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  • Characterization and expression analysis of P5CS (Δ1-pyrroline-5-carboxylate synthase) gene in two distinct populations of the Atlantic Forest native species Eugenia uniflora L.
    Mol. Biol. Rep. (IF 2.107) Pub Date : null
    Débora Bublitz Anton,Frank Lino Guzman,Nicole Moreira Vetö,Felipe Augusto Krause,Franceli Rodrigues Kulcheski,Ana Paula Durand Coelho,Guilherme Leitão Duarte,Rogério Margis,Lúcia Rebello Dillenburg,Andreia Carina Turchetto-Zolet

    Eugenia uniflora is an Atlantic Forest native species, occurring in contrasting edaphoclimatic environments. The identification of genes involved in response to abiotic factors is very relevant to help in understanding the processes of local adaptation. 1-Pyrroline-5-carboxylate synthetase (P5CS) is one interesting gene to study in this species since it encodes a key enzyme of proline biosynthesis, which is an osmoprotectant during abiotic stress. Applying in silico analysis, we identified one P5CS gene sequence of E. uniflora (EuniP5CS). Phylogenetic analysis, as well as, gene and protein structure investigation, revealed that EuniP5CS is a member of P5CS gene family. Plants of E. uniflora from two distinct environments (restinga and riparian forest) presented differences in the proline accumulation and P5CS expression levels under growth-controlled conditions. Both proline accumulation and gene expression level of EuniP5CS were higher in the genotypes from riparian forest than those from restinga. When these plants were submitted to drought stress, EuniP5CS gene was up-regulated in the plants from restinga, but not in those from riparian forest. These results demonstrated that EuniP5CS is involved in proline biosynthesis in this species and suggest that P5CS gene may be an interesting candidate gene in future studies to understand the processes of local adaptation in E. uniflora.

    更新日期:2020-01-14
  • RNA: interactions drive functionalities
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-12-14
    Xiaofeng Dai, Shuo Zhang, Kathia Zaleta-Rivera

    Abstract RNA is produced from the majority of human genomic sequences, although only a relatively small portion of these transcripts has known functions. Diverse RNA species interact with RNA, DNA, proteins, lipids, and metabolites to form intricate molecular networks. In this review, we attempt to delineate diverse RNA functions by interaction types between RNA and other macromolecules. Through such interactions RNAs participate in essentially every major molecular function and process, including information flow and storage, environment sensing, signal transduction, and gene regulation at transcriptional and posttranscriptional levels. Through such interactions, RNAs promote or inhibit diverse biological processes, and act as catalyzer or quencher to modulate the pace of these progresses. Alterations and personal variations of these interactions are mechanistically coupled with disease etiology and phenotypical variations for clinical use.

    更新日期:2020-01-14
  • Ancestry informative DIP loci for dissecting genetic structure and ancestry proportions of Qinghai Tibetan and Tibet Tibetan groups.
    Mol. Biol. Rep. (IF 2.107) Pub Date : null
    Xiao-Ye Jin,Chun-Mei Shen,Chong Chen,Yu-Xin Guo,Wei Cui,Yi-Jie Wang,Wen-Qing Zhang,Ting-Ting Kong,Bo-Feng Zhu

    Tibetans living in the Qing-Tibet plateau show unique genetic features since they are exposed to the high altitude environment. Accordingly, it is necessary for us to analyze genetic components of the Tibetan groups. Here, genetic structure and ancestry proportions of Tibet Tibetan and Qinghai Tibetan groups are dissected by using a previously published ancestral deletion/insertion polymorphisms (DIPs) panel. Genetic distributions of the analyzed DIPs in both Tibetan groups reveal that some DIPs show relatively balanced frequency distributions with the values ranging from 0.4 to 0.6, implying that these DIPs could be used as individual identification loci for forensic applications in both groups. Besides, the cumulative power of discrimination of the panel also reflects that the panel could serve as a valuable tool for forensic individual identifications in Tibet Tibetan and Qinghai Tibetan groups. Population genetic analyses including principal component analysis, DA genetic distances, phylogenetic tree, and genetic structure reveal that two studied Tibetan groups have closer genetic affiliations with East Asian populations. Genetic differentiation analyses of two Han populations, Xinjiang Uyghur and two Tibetan groups reveal that some DIP loci might be informative for differentiating Uyghurs from the other populations.

    更新日期:2020-01-14
  • Evaluation of reference genes for quantitative real-time PCR in Wharton's Jelly-derived mesenchymal stem cells after lentiviral transduction and differentiation.
    Mol. Biol. Rep. (IF 2.107) Pub Date : null
    P Borkowska,A Zielińska,M Paul-Samojedny,R Stojko,J Kowalski

    Quantitative real time reverse transcription PCR, qRT-PCR, is one of the most important techniques for assessing the level of gene expression. Selecting the correct reference gene to normalize the results is a key step in this method. Inaccurate data can be generated if the correct reference gene is not selected. The level of the expression of reference genes is tissue-variable, and in the case of mesenchymal stem cells (MSC), it can be different depending on the source of their origin. The aim of this study was to select the reference gene for Wharton's Jelly-derived MSC (WJ- MSC) that were undergoing transduction and differentiation. In this work, the expression of 32 genes was analyzed, of which two (RPS17 and 18S rRNA), which had the most stable expression level, were selected. A comparative analysis of the expression stability of the selected genes was then performed with the genes that are most commonly used in the literature, i.e. β-actin and GAPDH. Next, it was determined that a false picture of the expression level of the studied genes can be obtained when a reference gene with variable expression level is used for normalization. RPS17 and 18S rRNA proved to be the most stable reference genes for the WJ-MSC that had been subjected to the lentiviral transfection procedure followed by differentiation. The expression of β-actin and GAPDH was highly unstable and therefore these genes are not suitable for use as reference genes in studies involving WJ- MSC.

    更新日期:2020-01-14
  • Endometrial cancer and its cell lines
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-12-17
    Kristijan Skok, Uroš Maver, Lidija Gradišnik, Nejc Kozar, Iztok Takač, Darja Arko

    Endometrial cancer is one of the most common gynaecological malignancies worldwide. One type of research in this field is the growing of cell lines (CLs) and cultures, which can be used to explore the biological mechanisms of cancer. The purpose of this review is to offer an overview of the current literature and highlight the importance of correct CL studies. We carried out a literature analysis of more than 60 articles from the Pubmed, Medline databases that were almost exclusively published in indexed journals in the last 10 years as well as the primary originating scientific studies of specific CLs. We then summarized the newest findings and recommendations. Cell lines are becoming widely used as in vitro tumour models. Recent work has shown inconsistencies in nomenclature and culturing of CLs. Their genomic evolution leads to a high degree of variation across CL strains therefore it is of the utmost importance to recognize the variability within laboratory cancer models. Laboratories must adapt, incorporate additional characterisation techniques and view this situation as an opportunity to improve the reproducibility of pre-clinical cancer research. The authors offer a comprehensive literature review about endometrial cancer CLs, a review of the current literature and advice on culturing CLs.

    更新日期:2020-01-14
  • GISH-based comparative genomic analysis in Urochloa P. Beauv.
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-11-18
    Caio T R Corrêa,Nathalia G Z Bonetti,Sanzio C L Barrios,Cacilda B do Valle,Giovana A Torres,Vânia H Techio

    The genus Urochloa P. Beauv. [syn. Brachiaria (Trin.) Griseb.] comprises species of great economic relevance as forages. The genomic constitution for the allotetraploid species Urochloa brizantha (cv. Marandu) and Urochloa decumbens (cv. Basilisk) and the diploid Urochloa ruziziensis was previously proposed as BBB1B1, B1B1B2B2 and B2B2, respectively. Evidence indicates U. ruziziensis as the ancestral donor of genome B2 in U. decumbens allotetraploidy, but the origin of the genomes B and B1 is still unknown. There are diploid genotypes of U. brizantha and U. decumbens that may be potential ancestors of the tetraploids. The aim of this study was to determine the genomic constitution and relationships between genotypes of U. brizantha (2x and 4x), U. decumbens (2x and 4x) and U. ruziziensis (2x) via genomic in situ hybridization (GISH). Additionally, chromosome number and genome size were verified for the diploid genotypes. The diploids U. brizantha and U. decumbens presented 2n = 2x = 18 chromosomes and DNA content of 1.79 and 1.44 pg, respectively. The GISH analysis revealed high homology between the diploids U. brizantha and U. decumbens, which suggests relatively short divergence time. The GISH using genomic probes from the diploid accessions on the tetraploid accessions' chromosomes presented similar patterns, highlighting the genome B1 present in both of the tetraploids. Based on GISH results, the genomic constitution was proposed for the diploid genotypes of U. brizantha (B1B1) and U. decumbens (B1'B1') and both were pointed as donors of genome B1 (or B1'), present in the allotetraploid genotypes.

    更新日期:2020-01-14
  • Establishment of a conditional Nomo1 mouse model by CRISPR/Cas9 technology
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-12-12
    Ignacio García-Tuñón, Elena Vuelta, Laura Lozano, María Herrero, Lucía Méndez, Javier Palomero-Hernandez, María Pérez-Caro, Jessica Pérez-García, Rogelio González-Sarmiento, Manuel Sánchez-Martín

    Abstract The Nomo1 gene mediates a wide range of biological processes of importance in embryonic development. Accordingly, constitutive perturbation of Nomo1 function may result in myriad developmental defects that trigger embryonic lethality. To extend our understanding of Nomo1 function in postnatal stages and in a tissue-specific manner, we generated a conditional knockout mouse model of Nomo1. To achieve this, we used clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technology in C57Bl/6J mouse zygotes to generate a new mouse model in which exon 3 of the Nomo1 gene is specifically flanked (or floxed) by LoxP sites (Nomo1f/f). Nomo1f/f mouse embryonic fibroblasts were transduced with a Cre adenovirus and efficiently recombined between LoxP sites. Genomic and expression studies in Nomo1-transduced MEFs demonstrated that the Nomo1 exon 3 is ablated. Western blot assay showed that no protein or early truncated protein is produced. In vivo assay crossing Nomo1f/f mouse with a Msi1-CRE transgenic mouse corroborated the previous findings and it showed Nomo1 exon 3 deletion at msi1+ cell compartment. This short technical report demonstrates that CRISPR/Cas9 technology is a simple and easy method for creating conditional mouse models. The Nomo1f/f mouse will be useful to researchers who wish to explore the role of Nomo1 in any developmental stage or in a tissue-specific manner.

    更新日期:2020-01-14
  • The dynamic responses of plant physiology and metabolism during environmental stress progression
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-12-10
    Amit Kumar Singh, Shanmuhapreya Dhanapal, Brijesh Singh Yadav

    Abstract At adverse environmental conditions, plants produce various kinds of primary and secondary metabolites to protect themselves. Both primary and secondary metabolites play a significant role during the heat, drought, salinity, genotoxic and cold conditions. A multigene response is activated during the progression of these stresses in the plants which stimulate changes in various signaling molecules, amino acids, proteins, primary and secondary metabolites. Plant metabolism is perturbed because of either the inhibition of metabolic enzymes, shortage of substrates, excess demand for specific compounds or a combination of these factors. In this review, we aim to present how plants synthesize different kinds of natural products during the perception of various abiotic stresses. We also discuss how time-scale variable stresses influence secondary metabolite profiles, could be used as a stress marker in plants. This article has the potential to get the attention of researchers working in the area of quantitative trait locus mapping using metabolites as well as metabolomics genome-wide association.

    更新日期:2020-01-14
  • Relationship between biofilm gene expression with antimicrobial resistance pattern and clinical specimen type based on sequence types (STs) of methicillin-resistant S. aureus
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-12-18
    Hamed Tahmasebi, Sanaz Dehbashi, Mojdeh Jahantigh, Mohammad Reza Arabestani

    The ica genes in methicillin-resistant Staphylococcus aureus (MRSA) play an important role in biofilm formation. The aim of this study is to define effect of antibiotic resistance and clinical specimens to the expression of ica genes based on their sequence types (STs) and clonal complex (CC). One-hundred (100) S. aureus strain were collected from two teaching therapeutic centers in Hamedan, Iran. Then, the PCR, qPCR, and MLST were used to characterize strains. The results indicated that 29 (29%), 15 (15%), and 5 (5%) strain were strong, mediate, weak biofilm producer, respectively, and the icaA (17%) and icaC (14%) genes were the most abundant. However, two unique STs (3667, 491) in Iran were reported and ST30 and ST11 were the most abundant STs and CC30 and CC5 were observed among MRSA and MSSA strains. High activity in ica locus was observed among strains collected from wound and catheter strains. Also, expression level of icaA gene increased in all strains except ST30 and ST491. Moreover, the highest expression level was observed in CC1, CC7, and CC11. Likewise, activity of the icaC gene was only observed in CC5. Furthermore, the expression of all ica genes in CC5 was significantly correlated with the type of biofilm and the clinical sample. In this study demonstrated that the frequency distribution of STs and CCs in different strains of MRSA was higher than methicillin-sensitive strains. Also, the type of clinical specimen and expression of ica genes played an important role in this abundance.

    更新日期:2020-01-14
  • Protein trap: a new Swiss army knife for geneticists?
    Mol. Biol. Rep. (IF 2.107) Pub Date : null
    Svetlana A Fedorova,Natalya V Dorogova

    The protein trap is a powerful tool for genetic and biochemical studies of gene function in the animal kingdom. Although the original protein trap was developed for flies, it can be easily adapted to other multicellular organisms, both known models and ones with an unsequenced genome. The protein trap has been successfully applied to the fruit fly, crustaceans Parhyale hawaiensis, zebrafish, and insect and animal cell cultures. This approach is based on the integration into genes of an artificial exon that carries DNA encoding a fluorescent marker, standardized immunoepitopes, an integrase docking site, and splice acceptor and donor sites. The protein trap for cell cultures additionally contains an antibiotic resistance gene, which facilitates the selection of trapped clones. Resulting chimeric tagged mRNAs can be interfered by dsRNA against GFP (iGFPi-in vivo GFP interference), or the chimeric proteins can be efficiently knocked down by deGradFP technology. Both RNA and protein knockdowns produce a strong loss of function phenotype in tagged cells. The fluorescent and protein affinity tags can be used for tagged protein localisation within the cell and for identifying their binding partners in their native complexes. Insertion into protein trap integrase docking sites allows the replacement of trap contents by any new constructs, including other markers, cell toxins, stop-codons, and binary expression systems such as GAL4/UAS, LexA/LexAop and QF/QUAS, that reliably reflect endogenous gene expression. A distinctive feature of the protein trap approach is that all manipulations with a gene or its product occur only in the endogenous locus, which cannot be achieved by any other method.

    更新日期:2020-01-14
  • The complete mitochondrial genomes of two octopods of the eastern Pacific Ocean: Octopus mimus and 'Octopus' fitchi (Cephalopoda: Octopodidae) and their phylogenetic position within Octopoda.
    Mol. Biol. Rep. (IF 2.107) Pub Date : null
    Erika Magallón-Gayón,Miguel Ángel Del Río-Portilla,Irene de Los Angeles Barriga-Sosa

    The complete mitochondrial genomes of two important octopus species from the eastern Pacific were sequenced, obtaining their complete nucleotide sequences. Octopus mimus is the most important commercially catched species along the eastern Pacific, from Mexico to Chile, whereas 'Octopus' fitchi is a pigmy species with uncertain taxonomic genus. The mitogenomes of Octopus mimus and 'Octopus' fitchi were 15,696 and 15,780 base pairs (bp) in length with an A + T composition of 75.5% and 75.8%, respectively. Each genome contains 13 protein-coding genes, 22 tRNA genes, and two rRNA genes, as well as a control region. Gene order is maintained as reported for other species of the Octopodidae. The phylogenetic analysis based on the concatenated thirteen protein-coding genes confirms that O. mimus belongs to the genus Octopus, which is supported by the genetic distance (11-16%) whereas the position of 'O'. fitchi within this group it is not supported. The analysis also indicated that the phylogenetic position of 'O'. fitchi is closer to Callistoctopus than to the Cistopus or the Amphioctopus clades. Based on the tree topology and the high genetic distance observed (24-25%), we suggest that 'O'. fitchi might represent a different genus.

    更新日期:2020-01-14
  • HINT1 gene pathogenic variants: the most common cause of recessive hereditary motor and sensory neuropathies in Russian patients
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-12-17
    O. A. Shchagina, T. B. Milovidova, A. F. Murtazina, G. E. Rudenskaya, S. S. Nikitin, E. L. Dadali, A. V. Polyakov

    Abstract Pathogenic variants in the HINT1 gene lead to hereditary axonopathy with neuromyotonia. However, many studies show that neuromyotonia may remain undiagnosed, while axonopathy is the major clinical finding. The most common cause of neuromyotonia and axonopathy, especially in patients of Slavic origin, is a c.110G>C (p.Arg37Pro) pathogenic variant in homozygous or compound heterozygous state. In this study, we analyzed a peripheral neuropathy caused by pathogenic variants in the HINT1 gene and evaluated its contribution to the hereditary neuropathy structure. The studied group included 1596 non-related families diagnosed with hereditary motor and sensory neuropathy (HMSN). The results show that HINT1 gene pathogenic variants make a significant contribution to the hereditary neuropathy epidemiology in Russian patients. They account for at least 1.9% of all HMSN cases and 9% of axonopathy cases. The most common HINT1 pathogenic variant in Russian patients is the c.110G>C (p.Arg37Pro) substitution. Its allelic frequency is 0.2% (95% CI 0.19–0.21%), carrier frequency is 1 in 250 people in Russian Federation, and the estimated disease incidence is 1 in 234,000 individuals. It was determined that the cause of this pathogenic variant’s prevalence is the founder effect.

    更新日期:2020-01-14
  • Thioredoxin, thioredoxin interacting protein and transducer and activator of transcription 3 in gestational diabetes
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-12-17
    Yael Pasternak, Meital Ohana, Tal Biron-Shental, Keren Cohen-Hagai, Sydney Benchetrit, Tali Zitman-Gal

    To evaluate changes in the inflammatory response of thioredoxin (TXN), thioredoxin interacting protein (TXNIP), transducer and activator of transcription 3, NFƙB-p50 and STAT3 at the level of maternal serum, placenta, and umbilical cord blood of women with gestational diabetes mellitus type 2 (GDMA2) compared to normal pregnancies (NP). Thirty pregnant women (20 with GDMA2 and 10 NP) were recruited during admission for delivery. Blood samples were obtained from the parturients and umbilical cords, as well as placental tissue for mRNA and protein extraction. TXNIP mRNA expression was significantly increased in maternal serum of women with GDMA2 compared to NP women. TXNIP mRNA was significantly decreased in GDMA2 placentas and cord blood compared to NP. TXN/TXNIP mRNA ratio showed significantly high absolute values in placental and cord blood (2.39 and 1.66) respectively, compared to maternal ratio (1.084) (P < 0.001). TXN/TXNIP placenta protein ratio showed similar values between GDMA2 and NP (0.98 and 0.86; P = 0.7). STAT3 and its target protein SOCS3, as well as NFƙB-p50 mRNA expression were significantly increased in placentas of GDMA2. NFƙB-p50 mRNA expression was significantly decreased in cord blood compared to both maternal and placental mRNA expression. Pro-inflammatory changes are expressed by low mRNA TXN/TXNIP ratio in maternal blood of GDMA2 patients, but not in placental and umbilical cord blood samples. This, as well as the feedback role of SOCS3 in STAT3 pathway and NFƙB-p50 expression, may indicate that the placenta has a role in protecting the fetus from damage due to inflammatory response, which is common in diabetes.

    更新日期:2020-01-14
  • Association between the transporters ABCA1/G1 and the expression of miR-33a/144 and the carotid intima media thickness in patients with arterial hypertension
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-12-18
    Claudia Huesca-Gómez, Yazmín Estela Torres-Paz, Rocío Martínez-Alvarado, Giovanny Fuentevilla-Álvarez, Leonardo Del Valle-Mondragón, Margarita Torres-Tamayo, Ma. Elena Soto, Ricardo Gamboa

    Abstract ATP-binding cassette membrane transporters (ABC), functions as an outflow facilitator of phospholipids and cellular cholesterol, playing an important role in the development of atherosclerosis and arterial hypertension. ABC’s transporters could post-transcriptionally regulated by miRs. Evaluate the association in the transporters ABCA1 and ABCG1 with the expression of miR-33a and miR-144 and the carotid intima media thickness (cIMT) in patients with essential arterial hypertension. The miR-33a-5p, miR-144-3p and mRNA ABCA1 and ABCG1 expression in monocytes from Mexican hypertensive patients were examined by RT-PCR. The miR-33a and miR-144 expression in monocytes and mRNA ABCA1 and ABCG1 from Mexican hypertensive patients were examined by RT-PCR. This study involved 84 subjects (42 normotensive subjects and 42 patients with essential hypertension). Our study revealed that miR-33a expression (p = 0.001) and miR-144 (p = 0.985) were up-regulated, meanwhile, ABCA1 and ABCG1 transporters were down-regulated (p = 0.007 and p = 0.550 respectively) in hypertensive patients compared with the control group. The trend remains for miR33a/ABCA1 in presence of cIMT. Moreover, an inverse correlation was found with the expression levels of ABCA1 and ABCG1 as well as in HDL-C with miR-33a and miR-144. Our results showed an increase in the expression of miR-33a and miR-144 and an inverse correlation in their target ABCA1 and ABCG1; it may be associated with essential arterial hypertension in patients with cIMT and as consequence for atheromatous plaque.

    更新日期:2020-01-14
  • Development and utilization of an InDel marker linked to the fertility restorer genes of CMS-D8 and CMS-D2 in cotton
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-01
    Juanjuan Feng, Haiyong Zhu, Meng Zhang, Xuexian Zhang, Liping Guo, Tingxiang Qi, Huini Tang, Hailin Wang, Xiuqin Qiao, Bingbing Zhang, Kashif Shahzad, Chaozhu Xing, Jianyong Wu

    Abstract The cytoplasmic male sterility (CMS) system is a useful tool for commercial hybrid cotton seed production. Two main CMS systems, CMS-D8 and CMS-D2, have been recognized with Rf2 and Rf1 as the restorer genes, respectively. The development of molecular markers tightly linked with restorer genes can facilitate the breeding of restorer lines. In this study, the InDel-1892 marker was developed to distinguish Rf2 and Rf1 simultaneously. Sequence alignment implied that CMS-D8-Rf2 has a 32 bp insertion and that CMS-D2-Rf1 has a 186 bp insertion at the InDel-1892 locus. The codominant marker was co-segregated with Rf1 and Rf2. Hence, this marker can be used for tracing Rf1 and Rf2 simultaneously and identifying the allele status at the restorer gene locus. The results of this study will facilitate efficient marker-assisted selection for restorer lines and hybrids of CMS systems.

    更新日期:2020-01-14
  • TNF-α G-308A genetic variants, serum CRP-hs concentration and DNA damage in obese women.
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-03-23
    Włodarczyk Marta,Ciebiera Michał,Nowicka Grażyna

    Obesity is associated with inflammation, which can disturb genome stability. Tumor necrosis factor (TNF-α) polymorphism was found to affect TNF-α protein production and inflammation. Therefore, the present study illustrates the relationship between TNF-α polymorphism, the degree of inflammation assessed by serum high sensitivity C-reactive protein concentration (CRP-hs) and basal DNA damage in patients with obesity (BMI 30-34.9 kg/m2) and control subjects with proper body mass (BMI < 25 kg/m2). A total of 115 participants (75 obese premenopausal women; and 40 age-, and gender-matched controls) were included. Biochemical parameters (serum concentrations of total-cholesterol, HDL-cholesterol, LDL- cholesterol, triglycerides, glucose, apolipoprotein AI, CRP-hs) and endogenous DNA damage (determined by comet assay) were measured. TNF-α G-308A polymorphism (rs1800629) was analyzed by PCR-RFLP (PCR-restriction fragments length polymorphism). An effect of TNF-α genotype on serum CRP-hs concentration was noted (p = 0.031). In general, carriers of the rare A allele of the TNF-α G-308A polymorphism had significantly lower endogenous DNA damage and serum CRP-hs concentrations than GG homozygotes, however, the protective effect of the A allele was especially visible in non-obese women. Serum CRP-hs concentrations and levels of DNA damage (% DNA in tail) were significantly higher in obese than in controls (p = 0.001 and p < 0.0001, respectively). The adjusted multiple linear regression analyses revealed a significant, independent impact of obesity on DNA damage (p = 0.00000) and no effect of other covariates i.e. age, TNF-α genotype and serum CRP-hs concentration. Our study showed that obesity has a significant impact on the levels of endogenous DNA damage. Obesity abolished the protective effect of A allele of the TNF-α G-308A polymorphism on DNA damage and on inflammation development observed in non-obese A allele carriers.

    更新日期:2020-01-14
  • Identification and characterization of microRNAs in phloem and xylem from ramie ( Boehmeria nivea )
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-12-09
    Fang Liu, Yinghong Tang, Qingquan Guo, Jianrong Chen

    Ramie (Boehmeria nivea) is a widely cropped species in southern China due to its high economic value of natural fiber for industry. Development of phloem and xylem is key evidence for generating fiber. However, the MicroRNA (miRNA) profiles of phloem and xylem in ramie have not been reported yet. miRNA belong to a small RNA family which has been recognized as an important regulator for various biological processes. In the present study, we aimed to identify differently expressed miRNAs between phloem and xylem in adult ramie. The results showed that 137 and 122 unique conserved miRNAs were identified from phloem and xylem libraries, respectively. Meanwhile, 4 novel miRNAs were identified from ramie by miRDeep2. Of these miRNAs, 77 conserved miRNAs in ramie were differentially expressed. Among the differentially expressed miRNAs, 44 miRNAs and 33 miRNAs were up-regulated and down-regulated in phloem compared to that in xylem, respectively. The functions of differentially expressed miRNAs were associated with regulating the development and differentiation of phloem and xylem. The present study provides a glance of miRNA profiles for further understanding of miRNA role in ramie development.

    更新日期:2020-01-14
  • Chlorogenic acid prevents hepatotoxicity in arsenic-treated mice: role of oxidative stress and apoptosis
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-12-09
    Mohamed A. Dkhil, Ahmed E. Abdel Moneim, Amira A. Bauomy, Mona Khalil, Esam M. Al-Shaebi, Saleh Al-Quraishy

    Arsenic is a potent and toxic heavy metal found in the environment that causes health problems, including liver disease, in humans and animals. Chlorogenic acid (CA) is the most abundant caffeoylquinic acid isomer present in plants. This study aims to assess how CA protects the liver tissue following sodium arsenite (NaAsO2)-induced toxicity in mice. Male Swiss mice were allocated into 5 groups: Control, intragastrically administered CA (200 mg/kg), intragastrically administered NaAsO2 (5 mg/kg), and two groups administered with CA (100 and 200 mg/kg) and NaAsO2. CA was administered 30 min before NaAsO2 and all the mice were treated daily for 28 days. To investigate the biochemical, histopathological, immunohistochemical, and molecular changes, blood and liver samples were collected. NaAsO2 treatment increased the liver function biomarkers such as alanine transaminase, aspartate transaminase, alkaline phosphatase, and total bilirubin. Lipid and nitric oxide production was elevated. Glutathione content and the activities of superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase decreased, indicating a disturbance in redox homeostasis. Histopathological examination revealed a granular degeneration of hepatocytes, infiltration of inflammatory cells, and centrilobular hepatocyte necrosis. Furthermore, tumor necrosis factor-α and interleukin-1β were upregulated upon NaAsO2 treatment, suggesting the induction of inflammation. Moreover, NaAsO2 triggered apoptosis in the liver by upregulating Bax and caspase-3 and downregulating Bcl-2. However, CA abrogated the biochemical, molecular, and histological changes, reflecting its hepatoprotective role in response to NaAsO2 treatment. Our findings demonstrate that CA could be a potential therapeutic to minimize NaAsO2-induced hepatic injury.

    更新日期:2020-01-14
  • Association of genetic variations in the vitamin D pathway with susceptibility to tuberculosis in Kazakhstan
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-13
    Mukhtar Sadykov, Azliyati Azizan, Ulan Kozhamkulov, Ainur Akilzhanova, Dauren Yerezhepov, Max Salfinger, Chee Kai Chan

    Tuberculosis (TB) poses an important health challenge and a significant economic burden for Kazakhstan and in Central Asia. Recent findings show a number of immunological related processes and host Mycobacterium tuberculosis defense are impacted by a variety of genes of the human host including those that play a part in the vitamin D metabolism. We investigated the genetic variation of genes in the vitamin D metabolic pathway of a cohort 50 TB cases in Kazakhstan and compared them to 34 controls living in the same household with someone infected with TB. We specifically analyzed 11 SNPs belonging to the following genes: DHCR7, CYP2R1, GC-1, CYP24A1, CYP27A1, CYP27B1, VDR and TNFα. These genes play a number of different roles including synthesis, activation, delivery and binding of the activated vitamin D. Our preliminary results indicate significant association of VDR (vitamin D receptor) SNPs (rs1544410, BsmI, with OR = 0.425, CI 0.221–0.816, p = 0.009 and rs731236, TaqI with OR = 0.443, CI 0.228–0.859, p = 0.015) and CYP24A1 (rs6013897 with OR = 0.436, CI 0.191–0.996, p = 0.045) with TB. Interaction of genetic variation of VDR and CYP24A1 may impact susceptibility to TB. The findings provided initial clues to understand individual genetic differences in relation to susceptibility and protection to TB.

    更新日期:2020-01-13
  • d -ribose and pathogenesis of Alzheimer’s disease
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-13
    Mehjbeen Javed, Md. Irshad Ahmad, Hina Javed, Sufia Naseem

    It is estimated that the global prevalence of dementia will rise as high as 24 million and predicted to be double in every 20 years which is attributed to the fact that the ageing population is increasing and so more individuals are at risk of developing neurodegenerative diseases like Alzheimer’s. Many scientists favored glycation of proteins such as tau, amyloid beta (Aβ) etc. as one of the important risk factor in Alzheimer’s disease (AD). Since, d-ribose shows highest glycation ability among other sugars hence, produces advanced glycation end products (AGEs) rapidly. However, there are several other mechanisms suggested by researchers through which d-ribose may cause cognitive impairments. There is a concern related to diabetic patients since they also suffer from d-ribose metabolism, may be more prone to AD risk. Thus, it is imperative that the pathogenesis and the pathways involved in AD progression are explored in the light of ribosylation and AGEs formation for identifying suitable diagnostics marker for early diagnosis or finding promising therapeutic outcomes.

    更新日期:2020-01-13
  • Adrenomedullin has a role in angiogenic effects of resveratrol in adipose tissues of obese female rats
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-13
    Ayse Asiye Culum, Muhittin Yurekli

    Abstract Obesity is a complex, chronic disease that arises according to the interaction between genetic and environmental factors. The expansion and growth of white adipose tissue (WAT) could be related to angiogenesis. Resveratrol and adrenomedullin (AdM) were used for the inhibition of angiogenesis in metabolically passive WAT for inhibiting the expansion of this tissue, and the activation of angiogenesis in metabolically active brown adipose tissue (BAT) for increasing daily energy consumption as a way of reducing obesity. Rats were divided into eight groups. Four obese groups were fed with a high-fat diet containing 60% fat as energy for three months. After obtaining obesity, 2.5 nmol/kg AdM and 10 mg/kg resveratrol were treated to experiment groups intraperitoneally (i.p.) every other day for four weeks. AdM and vascular endothelial growth factor A (VEGF-A) mRNA levels were detected with semi-quantitative PCR; protein levels were detected with Western Blotting. AdM and resveratrol are multifactorial molecules, thus, this study has revealed a few novel evidence. The results were distinct in the group and treatment levels. The results showed that resveratrol has a role in angiogenesis in obesity and contributed to AdM production. It is observed that AdM has regulated its expression and increased the effect of resveratrol in WAT. AdM and VEGF-A gene expressions could not be detected in BAT; however, it is suggested that resveratrol may have a pro-angiogenic effect in BAT of obese rats according to the protein levels. AdM also has regulated VEGF-A level according to the metabolic situation of the organism.

    更新日期:2020-01-13
  • Characterization of a novel cotton MYB gene, GhMYB108 - like responsive to abiotic stresses
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-13
    Abid Ullah, Muhammad Tahir Ul Qamar, Mohammad Nisar, Ali Hazrat, Gul Rahim, Aamir Hamid Khan, Kashif Hayat, Saeed Ahmed, Waqar Ali, Aziz khan, Xiyan Yang

    Abstract Transcriptional factors are the major regulators of plant signaling pathways in response to environmental stresses i.e., drought, salinity and cold. Hereby, the GhMYB108-like was characterized to determine whether it regulate these stresses. The GhMYB108-like cDNA consisted of 1107 base pairs (bp) with 807 open reading frame encoded a protein of 268 amino acids. Its isoelectric point and molecular weight are 5.51 and 30.3 kDa respectively. Phylogenetic analysis and online databases revealed that GhMYB108-like proteins are closely related with the Arabidopsis thaliana MYB2. Important cis-elements were detected in the promotor region of GhMYB108-like responding to stresses and phytohormones. The 3D structure of GhMYB108-like protein has been predicted. In addition, various physico-chemical properties of GhMYB108-like have been determined. Subcellular localization confirmed that GhMYB108-like are nuclear localized protein. Quantitative expression analysis showed that polyethylene glycol and salt treatments significantly induced the expression of GhMYB108-like. Overall, our findings suggest that GhMYB108-like is an important gene that would plays important regulatory role in response to drought and salt stresses.

    更新日期:2020-01-13
  • A novel bioactive nanoparticle synthesized by conjugation of 3-chloropropyl trimethoxy silane functionalized Fe 3 O 4 and 1-((3-(4-chlorophenyl)-1-phenyl-1H-pyrazol-4-yl)methylene)-2-(4-phenylthiazol-2-yl) hydrazine: assessment on anti-cancer against gastric AGS cancer cells
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-13
    Seyyedeh Zahra Habibzadeh, Ali Salehzadeh, Zeinab Moradi-Shoeili, Seyed Ataollah Sadat Shandiz

    Gastric cancer is one of the common types of cancer around the world which has few therapeutic options. Nitrogen heterocyclic derivatives such as thiazoles are used as the basis for the progression of the drugs. The objective of this study was to synthesize the 1-((3-(4-chlorophenyl)-1-phenyl-1H-pyrazol-4-yl) methylene)-2-(4-phenylthiazol-2-yl) hydrazine (TP) conjugating with (3-Chloropropyl) trimethoxysilane (CPTMOS)-coated Fe3O4 nanoparticles (NPs) for anti-cancer activities against gastric AGS cancer cell line. The synthesized Fe3O4@CPTMOS/TP NPs were characterized by FT-IR, XRD, EDX, SEM, TEM and Zeta potential analyses. To evaluate the toxicity of the above compound after AGS cell culture in RPMI1640 medium, the cells were treated at different concentrations for 24 h. The viability of the cells was investigated by MTT assay. Moreover, apoptosis induced by Fe3O4@CPTMOS/TP NPs was assessed by Hoechst 33432 staining, oxygen activity specification evaluation, caspase-3 activity assay, cell cycle analysis and annexin V/PI staining followed by flow cytometry analysis. The IC50 value in AGS cells was estimated to be 95.65 µg/ml. The flow cytometry results of Fe3O4@CPTMOS/TP NPs revealed a large number of cells in the apoptotic regions compared to the control cells and the cells treated with TP. In addition, the amount of ROS production and caspase-3 activity increased in the treated cells with Fe3O4@CPTMOS/TP NPs. The percentage of inhibited cancer cells in the G0/G1 phase increased under the treatment in the binding state to the nonionic iron oxide nanoparticles. Overall, this study showed that Fe3O4@CPTMOS/TP NP had effect on induction of apoptosis and inhibiting the growth of AGS cancer cells. Thus, Fe3O4@CPTMOS/TP NP can be considered as a new anti-cancer candid for next phase of studies on mouse models.

    更新日期:2020-01-13
  • Fenretinide reduces angiogenesis by downregulating CDH5, FOXM1 and eNOS genes and suppressing microRNA-10b
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-10
    Elif Isil Yücel, Mehmet Sahin

    Abstract Angiogenesis is a new vessel formation process that plays a role in various physiological and pathological conditions. This process is controlled by the balance between pro-angiogenic and anti-angiogenic mediators in the organism. Angiogenesis is needed for the growth and metastasis of solid tumors. Therefore, the anti-angiogenic treatment approach is seen as an interesting option in cancers. Fenretinide, a synthetic retinoic acid analog, is an effective agent on angiogenesis. In this study, we aimed to investigate the effects of the fenretinide on some miRNAs involving in angiogenesis process and on the expression of CDH5, FOXM1 and eNOS genes upregulated in angiogenesis. In addition, it was shown the effects of this agent on cell proliferation, cell migration and capillary-like tube formation. In our study, the data were analyzed using Kruskal–Wallis and Dunn’s test. Fenretinide applied to the cells for 24 and 48 h periods reduced cell proliferation (P < 0.001) and cell migration, and suppressed tube formation (P < 0.001) as a dose dependent manner. Endothelial cells were cultured in growth-inducing media containing a variety of growth factors such as VEGF, FGF, IGF and EGF. As a result of simultaneous PCR analysis, we found that angiogenesis-promoting miR-10b was effectively suppressed (P < 0.001) and interestingly angiogenesis-modulating miR-126 was slightly increased (P < 0.05), but other miRNAs, including miR-31, miR-21, miR-101, miR-340, miR-29c, miR-206 and miR-146a were not affected. Besides, a significant decrease was observed in the levels of some angiogenesis-inducing genes, CDH5 (P < 0.001), FOXM1 (P < 0.001) and eNOS (P < 0.01 and P < 0.001) in endothelial cells treated with fenretinide. Our results have shown that fenretinide exhibited anti-angiogenic activity through the down-regulation of CDH5, FOXM1 and eNOS genes, and suppression of miR-10b.

    更新日期:2020-01-11
  • NLRC5: new cancer buster?
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-10
    Feng Tang, Yadi Xu, Bing Zhao

    Recent decades, there is significant progress in understanding the mechanisms of tumor progression and immune evasion. The newly discovered protein NLRC5 is demonstrated to participate in regulating cancer immune escape through enhancing MHC class I genes expression in certain tumors. Nevertheless, increasing evidence has revealed that NLRC5 is up-regulated in some other tumors and promote tumor development and progression. The purpose of this review is to describe the role of NLRC5 in tumors and discuss whether NLRC5 can be a potential target in cancer treatment.

    更新日期:2020-01-11
  • NIT2 overexpression predicts poor prognosis in tongue squamous cell carcinoma patients
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-10
    Shan Chen, Zengyan Wang, Chongjin Feng

    Abstract There is disputable on the role of nitrilase-like 2 (NIT2) in cancer. Its expression and its relationship with clinicopathological features in tongue squamous cell carcinoma (TSCC) are not yet clear. The purpose of this study is to investigate the expression of NIT2 in TSCC and its correlation with clinicopathological characteristics in TSCC patients. Through proteomic identification, we found that the protein NIT2 was related to the development of TSCC. q-PCR, western blot and immunohistochemistry techniques were applied to detect the expression of NIT2 in TSCC. The relationship between the expression of NIT2 and clinicopathological features was analyzed by Chi square tests. The results showed the expression of NIT2 in TSCC was significantly higher than that in normal tongue tissues (p < 0.05). Univariate and multivariate analysis showed that the positive expression of NIT2 and N classification were associated with decreased disease-free survival rate (DFS) and overall survival (OS) (p < 0.05). The results suggested that NIT2 is overexpressed in TSCC and NIT2 may be a potential therapeutic target for TSCC.

    更新日期:2020-01-11
  • High-throughput retrotransposon-based genetic diversity of maize germplasm assessment and analysis
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-09
    Marwa Ghonaim, Ruslan Kalendar, Hoda Barakat, Nahla Elsherif, Naglaa Ashry, Alan H. Schulman

    Maize is one of the world’s most important crops and a model for grass genome research. Long terminal repeat (LTR) retrotransposons comprise most of the maize genome; their ability to produce new copies makes them efficient high-throughput genetic markers. Inter-retrotransposon-amplified polymorphisms (IRAPs) were used to study the genetic diversity of maize germplasm. Five LTR retrotransposons (Huck, Tekay, Opie, Ji, and Grande) were chosen, based on their large number of copies in the maize genome, whereas polymerase chain reaction primers were designed based on consensus LTR sequences. The LTR primers showed high quality and reproducible DNA fingerprints, with a total of 677 bands including 392 polymorphic bands showing 58% polymorphism between maize hybrid lines. These markers were used to identify genetic similarities among all lines of maize. Analysis of genetic similarity was carried out based on polymorphic amplicon profiles and genetic similarity phylogeny analysis. This diversity was expected to display ecogeographical patterns of variation and local adaptation. The clustering method showed that the varieties were grouped into three clusters differing in ecogeographical origin. Each of these clusters comprised divergent hybrids with convergent characters. The clusters reflected the differences among maize hybrids and were in accordance with their pedigree. The IRAP technique is an efficient high-throughput genetic marker-generating method.

    更新日期:2020-01-09
  • Investigating molecular evolutionary forces and phylogenetic relationships among melatonin precursor-encoding genes of different plant species
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-09
    Moncef Boulila, Abdelaleim Ismail ElSayed, Mohammed Suhail Rafudeen, Ahmad Alsayed Omar

    Abstract A total of 53 plant species accessions from different geographic regions, including four melatonin precursor-coding genes obtained from Arachis hypogaea (ASMT1, 2, 3 and T5H) underwent extensive molecular evolutionary analyses. Evolutionary relationships were inferred and showed that dichotomous bifurcating trees did not reflect the true phylogeny since reticulate events took place due likely to recombination. Thus, a phylogenetic network was reconstructed for each type of enzyme and highlighted the presence of such incompatibilities. GARD algorithm pointed out that ASMT1, 2, and 3-coding gene sequences contained recombination sites with significant topological incongruence on both sides of the breakpoints (for ASMT1, and 2), while only on one side of the breakpoints for ASMT3. In contrast, no statistically recombination signal was recorded in T5H-coding gene. Furthermore, gene duplication was localized in the ancestor of a monophyletic group of Populus accessions. Selection pressure was assessed using several statistical models incorporated in HyPhy package through the datamonkey web server. It was demonstrated that numerous individual sites and tree branches experienced predominantly purifying selection. In contrast, the BUSTED model evidenced a gene-wide episodic diversifying selection in the phylogeny of only three enzyme-coding genes (ASMT, and 2, and T5H). Likewise, it was shown that Mixed Effects Model of Episodic Selection (MEME) model detected only episodic positively selected sites in all four melatonin enzymes-coding genes; whereas, REL model failed to detect neither positive nor negative selection in tested individual sites of ASMT3-coding gene.

    更新日期:2020-01-09
  • Analysis of the DOK1 gene in breast cancer
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-09
    Esin Tuna, Yeliz Emine Ersoy, Pelin Bulut, Filiz Ozdemir, Nur Buyru

    Abstract Breast cancer, which is the most common type of cancer among women, is a heterogenous disease. It results from progressive accumulation of genetic and epigenetic alterations in different genes. The Dok1 protein has been identified as the major substrate of protein tyrosine kinases in hematopoietic cells. It is considered as a tumor suppressor due to the reports which describe its inhibitory effect on major oncogenic signaling pathways such as Mek/Erk/PI3k/Akt and Wnt/β-catenin. In this study, we investigated the mutation frequency of the DOK1 gene in 118 breast tumors using Sanger sequencing and DOK1 mRNA expression level in 63 breast cancer samples using qRT-PCR methods. Although the mutation frequency was low DOK1 mRNA expression levels were significantly reduced (63.5%) in the tumors compared to adjacent non-cancerous tissue. We also correlated expression changes with clinicopathological characteristics. Low mRNA levels correlated with age (p = 0.01) and c-erbB-2 (p = 0.05). In most of the previous reports, down-regulation of DOK1 mRNA expression has been associated with promoter methylation. We identified four different coding sequence alterations in 5.1% (6/118) of the tumor samples. However, all of these alterations were located in the functional domains of the protein. Therefore, these mutations may affect the function and/or cellular localization of the protein and contribute to cancer progression by this way. In conclusion our data indicate that DOK1 acts as a tumor suppressor in breast cancer and association of Dok1 with the c-erbB-2 mediated mechanism of action in breast cancer needs to be investigated.

    更新日期:2020-01-09
  • Probiotic mediated NF-κB regulation for prospective management of type 2 diabetes
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-09
    Rabia Bhardwaj, Brij Pal Singh, Nitika Sandhu, Niharika Singh, Ravinder Kaur, Namita Rokana, Kumar Siddharth Singh, Vishu Chaudhary, Harsh Panwar

    Diabetes and other lifestyle disorders have been recognized as the leading cause of morbidity and mortality globally. Nuclear factor kappa B (NF-κB) is a major factor involved in the early pathobiology of diabetes and studies reveal that hyperglycemic conditions in body leads to NF-κB mediated activation of several cytokines, chemokines and inflammatory molecules. NF-κB family comprises of certain DNA-binding protein factors that elicit the transcription of pro-inflammatory molecules. Various studies have identified NF-κB as a promising target for diabetic management. Probiotics have been proposed as bio-therapeutic agents for treatment of inflammatory disorders and many other chronic clinical stages. The precise mechanisms by which probiotics acts is yet to be fully understood, however research findings have indicated their role in NF-κB modulation. The current review highlights NF-κB as a bio-therapeutic target for probable management of type 2 diabetes through probiotic intervention.

    更新日期:2020-01-09
  • CA19-9 capability as predictor of pancreatic cancer resectability in a Spanish cohort
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-08
    Marta Herreros-Villanueva, Lourdes Ruiz-Rebollo, Mario Montes, Mario Rodriguez-Lopez, María Francisco, Joaquín Cubiella, Eduardo Iyo, Emilio Garabitos, Emma Martínez Moneo, Maider Martos, Enrique de Madaria, Ibon Martínez-Arránz, Marta García-Cougil, Agueda Iglesias-Gómez, Luis Bujanda

    Abstract CA19-9 serum has been suggested as a marker of unresectability but different cut-off levels have been published. A cut-off of 500 U/ml is currently considered in an international consensus as biological criteria of borderline resectable pancreatic adenocarcinoma. To evaluate whether serum CA19-9 threshold of 500 U/ml could be adequate predictor of resectability in pancreatic adenocarcinoma. Multicenter, observational, prospective study performed in Spain including 203 patients diagnosed with pancreatic adenocarcinoma. 43 (21.2%) cases were resectable and 160 (78.8%) unresectable. Among the 176 preoperative CA19-9 available values, 98 (58.3%) were ≤ 500 U/ml and 73 (42.7%) > 500 U/ml. Resectability rate in those patients with CA19-9 ≤ 500 U/ml was 60% while it was found to be 18% when CA19-9 > 500 U/ml. Statistical model to predict resectability based on CA19-9 provide an AUC of 0.6618 (95% CI 0.53–0.83) when only CA19-9 values > 500 U/ml are studied. Serum levels of CA19-9 higher than 500 U/ml are indicative of unresectable disease, however reduced sensitivity and specificity lead to a limited clinical applicability for resectability.

    更新日期:2020-01-09
  • Isolation and characterization of kelch repeat-containing F-box proteins from colored wheat
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-06
    Min Jeong Hong, Dae Yeon Kim, Hong-Il Choi, Yong Weon Seo, Jin-Baek Kim

    Abstract F-box proteins play important roles in the regulation of various developmental processes in plants. Approximately 1796 F-box genes have been identified in the wheat genome, but details of their functions remain unknown. Moreover, not much was known about the roles of kelch repeat domain-containing F-box genes (TaKFBs) in wheat. In the present study, we isolated five TaKFBs to investigate the roles of KFBs at different stages of colored wheat grain development. The cDNAs encoding TaKFB1, TaKFB2, TaKFB3, TaKFB4, and TaKFB5 contained 363, 449, 353, 382, and 456 bp open reading frames, respectively. All deduced TaKFBs contained an F-box domain (IPR001810) and a kelch repeat type 1 domain (IPR006652), except TaKFB2. Expression of TaKFBs was elevated during the pigmentation stages of grain development. To clarify how TaKFB and SKP interact in wheat, we investigated whether five TaKFB proteins showed specificity for six SKP proteins using a yeast two-hybrid (Y2H) assay. An Y2H screen was performed to search for proteins capable of binding the TaKFBs and interaction was identified between TaKFB1 and aquaporin PIP1. To examine the subcellular localization of TaKFBs, we transiently expressed TaKFB-green fluorescent protein (GFP) fusions in tobacco leaves; the TaKFB-GFP fusions were detected in the nucleus and the cytoplasm. Y2H and bimolecular fluorescence complementation (BiFC) assays revealed that TaKFB1 specifically interacts with aquaporin PIP1. These results will provide useful information for further functional studies on wheat F-box proteins and their possible roles.

    更新日期:2020-01-06
  • Rice momilactone gene cluster: transcriptional response to barnyard grass ( Echinochloa crus - galli )
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-04
    J. Bajsa-Hirschel, Z. Pan, S. O. Duke

    Expression of genes involved in diterpene biosynthesis, especially momilactone and gibberellins (GAs), in rice plants (Oryza sativa L.) in response to barnyard grass (Echinochloa crus-galli) stress was examined. The three analyzed class II diterpene synthases had the highest fold change expression. Transcription patterns of genes for two homologs of momilactone synthases, OsMAS and OsMAS2, suggests their distinct roles in response to the presence of barnyard grass.

    更新日期:2020-01-04
  • Effects of temozolomide on U87MG glioblastoma cell expression of CXCR4, MMP2, MMP9, VEGF, anti-proliferatory cytotoxic and apoptotic properties
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2020-01-02
    Seyedsaber Mirabdaly, Daniel Elieh Ali Komi, Yadollah Shakiba, Ali Moini, Amir Kiani

    Abstract Temozolomide is an alkylating agent which is used in glioblastoma treatment. We aimed to investigate the effects of different concentrations of temozolomide and exposure time on U87MG glioblastoma cell expression of CXCR4, MMP2, MMP9 and VEGF. U87MG cells were cultured in different temozolomide concentrations and incubation time and the effects of temozolomide on inducing apoptosis was investigated. The levels of VEGF and CXCR4 expression were measured by RT-PCR and flowcytometry. Moreover, MMP2 and MMP9 activity and expression were assessed by ELISA and zymography. CXCR4 and VEGF expression levels decreased upon applying higher concentration of temozolomide. MMP2 and MMP-9 had lower activity in cells with longer exposure time or higher doses of temozolomide. Temozolomide induces the apoptosis in U87MG glioblastoma cells at therapeutic or higher dose. It is capable of decreasing their expression levels of VEGF and CXCR4.

    更新日期:2020-01-04
  • Development of genomic microsatellite markers for Aconitum gymnandrum (Ranunculaceae) by next generation sequencing (NGS)
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-11-01
    Meng Hou, Guo-zhen Du, Zhi-gang Zhao

    Abstract Mating plays key roles in the demographic and genetic dynamics of populations. Estimates of mating portfolios and system based on progeny array (PA) method required highly polymorphic genetic markers, of which microsatellite is a good choice. In this study, we reported 19 polymorphic microsatellite loci for Aconitum gymnandrum. The number of alleles per locus ranged from 2 to 12. Observed and expected heterozygosity ranged from 0.000 to 1.000 and from 0.219 to 0.842, respectively. Seven loci showed significant deviation from Hardy–Weinberg equilibrium. These markers will provide a useful tool for pollination ecology and population genetic studies of A. gymnandrum in Qinghai-Tibet plateau.

    更新日期:2020-01-04
  • The testis-specific expressed gene Spata34 is not required for fertility in mice
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-10-16
    DongSong Nie, YanFa Dai, ZhongQin Luo

    Abstract It is estimated that more than two thousand genes exhibit testis-predominant expression pattern. The functions of hundreds of these genes have been explored during mouse spermatogenesis. However, there are still many genes whose relevance to reproduction in vivo remains unexplored. Our previous studies, as well as the other documented study, have indicated that Spata34, an evolutionarily conserved gene in metazoan species, was exclusively expressed in mouse testes and involved in spermatogenesis by regulating cell cycle progression. The present study aims to determine the effect of Spata34 gene knockout on mouse reproduction in vivo by generating a Spata34 gene knockout model using CRISPR/Cas9-mediated genome editing technology. We found that the Spata34 gene KO mice had normal fertility compared with wild type mice, and no overt detectable difference was found in testis/body weight ratios, testicular histology, sperm counts and spermatozoa motility parameters between WT and Spata34 KO mice. Our report indicated that the testis-specific-expressed gene Spata34 was not required for male mouse fertility, which will help to avoid unnecessary expenditures and effort by other researchers.

    更新日期:2020-01-04
  • Overview of Staphylococcus epidermidis cell wall-anchored proteins: potential targets to inhibit biofilm formation
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-10-22
    Silvestre Ortega-Peña, Sergio Martínez-García, Sandra Rodríguez-Martínez, Mario E. Cancino-Diaz, Juan C. Cancino-Diaz

    Abstract Currently, the treatment of infections by Staphylococcus epidermidis (S. epidermidis) represents a challenge because some strains have multidrug-resistance to antimicrobial products (antibiotic and biocides) and can produce biofilms. These biofilms protect bacterial cells from both antimicrobials and the host immune response. Therefore, it is crucial to encourage research on the development of new treatments. One method is immunotherapy, targeting components of S. epidermidis, such as S. epidermidis surface (Ses) proteins. Ses is expressed constitutively in most strains, and they participate in biofilm formation. This review is an update on Ses, regarding their structure, biological function, their relationship with S. epidermidis biofilm formation, and its possible role as therapeutic targets to develop immunotherapeutic treatments to prevent infections by S. epidermidis.

    更新日期:2020-01-04
  • Bacterial staphylokinase as a promising third-generation drug in the treatment for vascular occlusion
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-11-01
    Reza Nedaeinia, Habibollah Faraji, Shaghayegh Haghjooye Javanmard, Gordon A. Ferns, Majid Ghayour-Mobarhan, Mohammad Goli, Baratali Mashkani, Mozhdeh Nedaeinia, Mohammad Hossein Hayavi Haghighi, Maryam Ranjbar

    Abstract Vascular occlusion is one of the major causes of mortality and morbidity. Blood vessel blockage can lead to thrombotic complications such as myocardial infarction, stroke, deep venous thrombosis, peripheral occlusive disease, and pulmonary embolism. Thrombolytic therapy currently aims to rectify this through the administration of recombinant tissue plasminogen activator. Research is underway to design an ideal thrombolytic drug with the lowest risk. Despite the potent clot lysis achievable using approved thrombolytic drugs such as alteplase, reteplase, streptokinase, tenecteplase, and some other fibrinolytic agents, there are some drawbacks, such as high production cost, systemic bleeding, intracranial hemorrhage, vessel re-occlusion by platelet-rich and retracted secondary clots, and non-fibrin specificity. In comparison, bacterial staphylokinase, is a new, small-size plasminogen activator, unlike bacterial streptokinase, it hinders the systemic degradation of fibrinogen and reduces the risk of severe hemorrhage. A fibrin-bound plasmin–staphylokinase complex shows high resistance to a2-antiplasmin-related inhibition. Staphylokinase has the potential to be considered as a promising thrombolytic agent with properties of cost-effective production and the least side effects.

    更新日期:2020-01-04
  • DNA damage response manages cell cycle restriction of senile multipotent mesenchymal stromal cells
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-10-29
    Lin Yao, Fanyuan Yu, Yining Xu, Yitian Wang, Yanqin Zuo, Chenglin Wang, Ling Ye

    Abstract Multipotent mesenchymal stromal cells (MMSCs) are promising to treat a variety of traumatic and degenerative diseases. However, in vitro-passage aging induces cell cycle arrest and a series of genetic and biological changes, which greatly limits ex vivo cell number expansion and further clinical application of MMSCs. In most cases, DNA damage and DNA damage response (DDR) act as the main cause and executor of cellular senescence respectively. Mechanistically, DNA damage signals induce cell cycle arrest and DNA damage repair via DDR. If the DNA damage is indelible, MMSCs would entry into a permanent cell cycle arrest. It should be noted that apart from DDR signaling, certain proliferation or metabolism pathways are also occupied in DNA damage related cell cycle arrest. New findings of these aspects will also be summarized in this study. In summary, we aim to provide a comprehensive review of DDR associated cell cycle regulation and other major molecular signaling in the senescence of MMSCs. Above knowledge could contribute to improve the limited capacity of in vitro expansion of MMSCs, and then promote their clinical applications.

    更新日期:2020-01-04
  • In vivo flow cytometry combined with intravital microscopy to monitor kinetics of transplanted bone marrow mononuclear cells in peripheral blood and bone marrow
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-12-07
    Fen Wang, Dan Wei, Yuanzhen Suo, Xi Zhu, Yan Yuan, Wenyuan Gao, Hua Jiang, Xunbin Wei, Tong Chen

    Bone marrow mononuclear cells (BM-MNCs) transplantation has evolved as a promising experimental treatment in various regenerative therapy fields, especially in clinical hematopoietic stem cells transplantation (HSCT). In vitro methods have mainly been used to study the pre-clinical kinetics of BM-MNCs in mice after transplantation. And it is difficult to monitor the dynamic homing of BM-MNCs in living mice. The present study obtained the kinetics of transplanted BM-MNCs in the peripheral blood (PB) and the dynamic homing of BM-MNCs in the BM in living mice by a combination of in vivo flow cytometry (IVFC) and calvarium intravital microscopy. We found out that BM-MNCs were cleared rapidly from the PB and mainly localized to various hematopoietic tissues after transplantation. The number of BM-MNCs in the PB decreased over time accompanied by an increase in the BM indeed after transplantation. In addition, a lower number of BM-MNCs were found home to calvaria than long bone, probably indicating long bone marrow might also be an important hematopoietic organ. Clinical studies will benefit from non-invasive measurements to monitor the dynamic homing of transplanted cells. Our pre-clinical kinetics of BM-MNCs in living mice will have important clinical guiding significance in HSCT and other regenerative therapy fields.

    更新日期:2020-01-04
  • Stable reference genes for expression studies in breast muscle of normal and white striping-affected chickens
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-10-03
    Caroline Michele Marinho Marciano, Adriana Mércia Guaratini Ibelli, Jane de Oliveira Peixoto, Igor Ricardo Savoldi, Kamilla Bleil do Carmo, Lana Teixeira Fernandes, Mônica Corrêa Ledur

    Abstract The normalization with proper reference genes is a crucial step to obtain accurate mRNA expression levels in quantitative PCR (qPCR) studies. Therefore, in this study, 10 reference candidate genes were evaluated to determine their stability in normal pectoralis major muscle of broilers and those counterparts affected with White Striping (WS) myopathy at 42 days age. Four different tools were used for ranking the most stable genes: GeNorm, NormFinder, BestKeeper and Comparative Ct (ΔCt), and a general ranking was performed using the RankAggreg tool to select the best reference genes among all tools. From the 10 genes evaluated in the breast muscle of broilers, 8 were amplified. Most of the algorithms/tools indicated the same two genes, RPL30 and RPL5, as the most stable in the broilers breast muscle. In addition, there was agreement among the tools for the least stable genes: MRPS27, GAPDH and RPLP1 in the broilers breast muscle. Therefore, it is interesting to note that even with different tools for evaluating gene expression, there was consensus on the most and least stable genes. These results indicate that the Ribosomal protein L30 (RPL30) and Ribosomal protein L5 (RPL5) can be recommended for accurate normalization in qPCR studies with chicken pectoralis major muscle affected with White Striping and other myopathies.

    更新日期:2020-01-04
  • Development of an anti-CD45RA-quantum dots conjugated scFv to detect leukemic cancer stem cells
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-10-22
    Shima Moradi-Kalbolandi, Fariba Dashtestani, Malihe Salehi, Neda Jalili, Keivan Majidzadeh-A, Reza Rahighi, Amir Yadegari, Leila Farahmand

    Abstract Leukemic cancer stem cells (LSCs), aberrantly overexpressing CD45RA are among the major causes of relapse following chemotherapy in patients with acute myeloid leukemia and serve as a highly sensitive marker for predicting relapse occurrence following chemotherapy. The main purpose of current study was to develop a sensitive approach for detecting LSCs based on a conjugate of an anti-CD45 scFv and quantum dot. The variable light and heavy chain sequences of a recently developed anti-CD45RA monoclonal antibody were derived from hybridoma cells and PCR amplified to construct scFv. Following insertion of scFv gene into a pET32a-lic vector and expression in Escherichia coli and purification, the purified scFv, was conjugated with carbon dots (C dots) and used for the detection of CD45RA +cells while CD45RA-cells served as negative control. Subsequently, Functional activity of the conjugate was analyzed by flow cytometry and ICC to detect the cell surface antigen binding and detection ability. Based on results, purified CD45RA scFv conjugated C dots could specifically recognize CD45RA positive cells, but not any CD45RA negative ones. In conclusion, here we developed a low-cost but very efficient approach for detection of CD45RA positive cells including LSCs.

    更新日期:2020-01-04
  • Identification of a transforming growth factor-β type I receptor transcript in Eriocheir sinensis and its molting-related expression in muscle tissues
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-09-30
    Zhihuan Tian, Hongyuan Peng, Weide Deng, Chuanzhen Jiao

    The transforming growth factor-β (TGF-β) signaling pathway is conserved across animals, and knowledge of its roles during the molt cycle in crustaceans is presently very limited. This study investigates the roles of the TGF-β receptor in molting-related muscle growth in Eriocheir sinensis. Using the RT-PCR and RACE techniques, we obtained a 1722 bp cDNA sequence encoding a transforming growth factor-β type I receptor in Eriocheir sinensis, designated EsTGFBRI, which contains a 124 bp 5′-untranslated region, a 20 bp partial 3′-untranslated region and a 1578 bp open reading frame encoding 525 amino acids. The deduced EsTGFBRI contains an N-terminal 24 amino acid signal peptide, an activin type I and II receptor domain, a transmembrane helix region, a glycine-serine-rich motif, and a conserved serine/threonine kinase catalytic domain including an activation loop. The qRT-PCR results showed that EsTGFBRI gene was highly expressed in the intermolt testis and ovary in mature crabs. In juvenile crabs, the mRNA levels of EsTGFBRI in claw and abdominal muscles in the later premolt D3–4 stage were significantly higher than those in the intermolt C and postmolt A–B stages. There was no significant change in EsTGFBRI mRNA levels in walking leg muscles during the molt cycle. The results suggest that EsTGFBRI is probably play roles in molting-related muscle growth in E. sinensis. This study provides a necessary basis for elucidating the functions of TGF-β-like signaling mediated by TGFBRI in molting-related muscle growth in crustaceans.

    更新日期:2020-01-04
  • Induction of p73, Δ133p53, Δ160p53, pAKT lead to neuroprotection via DNA repair by 5-LOX inhibition
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-10-28
    Shashank Shekhar, Sharmistha Dey

    Abstract The inflammatory process plays a key role in neurodegenerative disorder. The inflammatory molecule, 5-lipooxygenase (5-LOX), protein is involved in the pathologic phenotype of AD which includes Aβ amyloid deposition and tau hyperphosphorylation. This study aims to identify the mechanistic role in neuroprotection by 5-LOX inhibitor in neurotoxic SH-SY5Y cell line model by evaluating different cell survival pathway. The neurotoxic SH-SY5Y cells were developed by the treatment of Aβ25–35. The cells were then treated with 5-LOX peptide inhibitor, YWCS to prevent neurotoxicity reported earlier from our lab. The effect of 5-LOX inhibition on cell survival pathways were determined by western blot experiment with different doses of peptide by using polyclonal anti body of p53, anti-Akt and anti-phosphorylated Akt. Immunoprecipitation and mass spectroscopic studies were done to identify the altered proteins appeared on the blot. Over expression of phosphorylated Akt and 3 bands on p53 lane blot other than p53 were observed. Three bands were identified as isoforms of p53 which correspond to p73, Δ133p53 and Δ160p53 in the cells treated only with 80 µM of YWCS compare to untreated cells. However, no alteration of total p53 and Akt were observed in treated cells. The results exposed the novel mechanistic pathway of neuroprotection by 5-LOX inhibition is likely to be mediated by DNA DSB repair through p53 isoforms and PI3K/Akt pathway. Our finding has opened a new window in the therapeutic approach for the prevention of AD.

    更新日期:2020-01-04
  • IFITM3/STAT3 axis promotes glioma cells invasion and is modulated by TGF-β
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-10-21
    Hongliang Wang, Feng Tang, Erbao Bian, Yile Zhang, Xinghu Ji, Zhihao Yang, Bing Zhao

    Abstract Glioma is the most aggressive primary brain tumor. We have previously provided evidence that IFITM3 promoted glioma cells migration. However, the mechanism of how IFITM3 regulates glioma cells invasion and whether IFITM3 participates in TGF-β-mediated glioma invasion are still unknown. In this paper, we proved that IFITM3 was notably up-regulated in glioma tissues. Knockdown of IFITM3 suppressed STAT3 phosphorylation in vitro, and a specific STAT3 inhibitor AG490 reversed IFITM3-induced invasion of glioma cells. Furthermore, IFITM3 expression was induced by TGF-β in glioma and IFITM3 knockdown abolished TGF-β-mediated glioma cells invasion. Collectively, the results indicate that IFITM3/STAT3 axis may promote TGF-β-induced glioma cells invasion. This study provided some suggestions for the clinical treatment of the brain tumor.

    更新日期:2020-01-04
  • Campylobacter jejuni isolated from poultry meat in Brazil: in silico analysis and genomic features of two strains with different phenotypes of antimicrobial susceptibility
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-11-20
    Simone de Fátima Rauber Würfel, Sérgio Jorge, Natasha Rodrigues de Oliveira, Frederico Schmitt Kremer, Christian Domingues Sanchez, Vinícius Farias Campos, Luciano da Silva Pinto, Wladimir Padilha da Silva, Odir Antônio Dellagostin

    Campylobacter jejuni is the most common bacterial cause of foodborne diarrheal disease worldwide and is among the antimicrobial resistant “priority pathogens” that pose greatest threat to public health. The genomes of two C. jejuni isolated from poultry meat sold on the retail market in Southern Brazil phenotypically characterized as multidrug-resistant (CJ100) and susceptible (CJ104) were sequenced and analyzed by bioinformatic tools. The isolates CJ100 and CJ104 showed distinct multilocus sequence types (MLST). Comparative genomic analysis revealed a large number of single nucleotide polymorphisms, rearrangements, and inversions in both genomes, in addition to virulence factors, genomic islands, prophage sequences, and insertion sequences. A circular 103-kilobase megaplasmid carrying virulence factors was identified in the genome of CJ100, in addition to resistance mechanisms to aminoglycosides, beta-lactams, macrolides, quinolones, and tetracyclines. The molecular characterization of distinct phenotypes of foodborne C. jejuni and the discovery of a novel virulence megaplasmid provide useful data for pan-genome and large-scale studies to monitor the virulent C. jejuni in poultry meat is warranted.

    更新日期:2020-01-04
  • Inheritance of faba bean resistance to Broomrape, genetic diversity and QTL mapping analysis
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-10-04
    Bahaa E. S. Abd El-Fatah, Dalia M. T. Nassef

    Abstract Six faba bean parents and their F1 and F2 generations were used in this investigation to study the genetic system controlling resistance of faba bean (Vicia faba L.) to broomrape (Orobanche crenata). Most of the F1 hybrids were tolerant to broomrape. In the F2 generation, the population P5 × P6 (Assiut 125 × Romy 12) gave the highest value of relative yield and tolerance index. Heterosis and inbreeding depression were only positive in number of tillers/plant and seed yield/plant characters. The results indicated that the additive effect was more important than the dominance one (D > H1) only for No. of pods/plant in the F1 generation. Moreover, the narrow-sense heritability was low for most of the studied traits. Three molecular marker systems, namely RAPD, ISSR and SRAP were used for identification and estimation of the genetic diversity among the six faba bean genotypes. The three molecular markers generated DNA unique bands for all genotypes. Only, eight DNA fragments were related to Orobanche tolerance. Clearly and reproducible polymorphic markers were subjected to QTL analysis. The linkage analysis showed that, out of 34 marker loci segregated in the F2 population, 29 (85.29%) were mapped on three linkage groups. QTL analysis using SIM method performed for the 29 markers assigned to LG-1, LG-2 and LG-3 with the eight traits, number of tillers/plant, plant height, number of pods/plant, seed yield/plant, number of broomrape spikes per plant, height of broomrape spikes, relative yield and tolerance index, showing 12 putative QTLs for all traits except number of tillers/plant. From this study, it is clear that P5 × P6 (Assiut 125 × Romy12) population could be considered promising for selection for resistance to broomrape infestation.

    更新日期:2020-01-04
  • Mechanism of CNP-mediated DG-PKC and IP4 signaling pathway in diabetic rats with gastric motility disorder
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-10-03
    Hui-Ming Lian, Jun-Yu Guo, Yan Sun, Mo-Han Zhang, Li-Hua Piao, Zheng Jin, Ying-Lan Cai

    In the precedent research conducted by the same team, it concluded that the activities in C-type natriuretic peptide (CNP)/cyclic guanosine monophosphate (cGMP)/cyclic adenosine monophosphate (cAMP)/β-type phospholipase C (PLCβ) pathways of rat antral smooth muscle were changed due to diabetes, which was the key pathogenetic mechanism for diabetic gastric dysmotility. As the follow-on step, this study was designed to probe into the downstream signaling pathway of CNP/PLCβ. The results showed that level of α-type protein kinase C (PKCα),cell membrane to cytoplasm ratio of PKCα, cell membrane to cytoplasmic ratio of βI-type protein kinase C (PKCβI) and level of Phosphor-PKCα (P-PKCα) were significantly reduced in diabetes rat antral smooth muscle samples. The content of tetraphosphate inositol (IP4) in gastric antral smooth muscle of diabetic rats reduced, and the content of diacyl-glycerol (DG) was unchanged. CNP significantly decreased the content of IP4 and DG, this effect was more obvious in diabetic rats. Subsequent to the addition of protein kinase A (PKA) blocker N-[2- (p-Bromocin-namylamino)ethyl]-5 -isoquinolinesulfonamide dihydrochloride (H-89) before CNP treatment, the inhibitory effect of CNP was reduced; subsequent to the addition of protein kinase G (PKG) blocker KT5823 before CNP treatment, the inhibitory effect of CNP was also reduced. With the addition of the combination of H-89 and KT5823 before CNP treatment, the inhibition by CNP could be offset. These results were concluded that CNP inhibited the activity of PKC family in rat smooth muscle and reduced the levels of IP4 and DG through the PKG/PKA-PLCβ pathways, causing inhibited muscular contractions, which may be a key pathogenetic factor for diabetic gastroparesis.

    更新日期:2020-01-04
  • Melatonin ameliorates sodium valproate-induced hepatotoxicity in rats
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-10-17
    Ozlem Oztopuz, Hakan Turkon, Basak Buyuk, Ozlem Coskun, Muserref Hilal Sehitoglu, Mehmet Akif Ovali, Metehan Uzun

    Valproic acid (VPA) is a anticonvulsant and mood-stabilizing agent used to treat epilepsy in patients of all ages. However, it can cause hepatotoxicity with increased oxidative stress. Melatonin (MEL) is known as antioxidant and antiinflammatory agent. Therefore, the present study designed to investigate the probable protective role of melatonin against VPA-induced liver toxicity. For that purpose, 28 Wistar rats were randomly selected and divided into four groups, namely the Group C (vehicle), VPA (500 mg/kg/day VPA), MEL + VPA (10 mg/kg/day melatonin + 500 mg/kg/day VPA) and MEL (10 mg/kg/day melatonin). The agents were given by oral gavage for 14 days. Blood and liver tissue samples from all the rats were harvested on the 15th day of experiment. Biochemical analyses were conducted on the blood samples. The levels of malondialdehyde (MDA), superoxide dismutase (SOD), alpha glutathione S-transferases (α-GST), nuclear factor-κB (NF-κB), myeloperoxidase (MPO) and changes in gene expression were examined in the liver tissues. Also, liver histopathological analyses were conducted. VPA administration significantly increased the levels of α-GST, MDA, NF-κB and of IL-1β, TNF-α gene expression in the liver compared to Group C. Moreover, vacuolization, hydropic degeneration, inflammatory cell infiltration, and sinusoidal congestion were commonly detected in the VPA-treated group along with the highest apoptotic index (TUNEL staining) values. Melatonin administration was revealed to exhibit powerful protective properties at cellular, inflammatory and oxidative level activities against VPA-induced liver toxicity. Therefore, melatonin administration may be used as an adjuvant therapy against to VPA-induced liver toxicity.

    更新日期:2020-01-04
  • Effects of different 2A peptides on transgene expression mediated by tricistronic vectors in transfected CHO cells
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-10-28
    Yan-mei Li, Meng Wang, Tian-yun Wang, Yong-ge Wei, Xiao Guo, Chun-liu Mi, Chun-peng Zhao, Xiang-xiang Cao, Yuan-yuan Dou

    Abstract Multicistronic vectors can increase transgene expression and decrease the imbalance of gene expression in the Chinese hamster ovary (CHO) cell expression system. Small, self-cleaving 2A peptides have a high cleavage efficiency and are essential for constructing high-expression multicistronic vectors. In this study, we investigated the effects of two different 2A peptides on transgene expression in CHO cells via their mediating action on tricistronic vectors. The enhanced green fluorescent protein (eGFP) and red fluorescent protein (RFP) genes were linked by the porcine teschovirus-1 (P2A) and Thosea asigna virus (T2A) peptides in a multicistronic vector. We transfected CHO cells with these vectors and screened for the presence of blasticidin-resistant colonies. Flow cytometry and real-time quantitative PCR (qPCR) were used to detect the expression levels of eGFP and RFP and the copy numbers of stably transfected cells. The results showed that P2A could enhance eGFP and RFP expression by 1.48- and 1.47-fold, respectively, compared to T2A. The expression levels of the genes were not proportional to their copy numbers. In conclusion, we found that P2A can effectively drive transgene expression in CHO cells and a potent 2A peptide can be used for recombinant protein production in the CHO cell system.

    更新日期:2020-01-04
  • Isolation, characterization and in silico analysis of 3-Hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR) gene from Andrographis paniculata (Burm. f) Nees
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-11-28
    Mote Srinath, Byreddi Bhavani Venkata Bindu, Ayeti Shailaja, Charu Chandra Giri

    3-Hydroxy-3-methylglutaryl-coenzymeA reductase (HMGR), the first rate-limiting enzyme of Mevalonate (MVA) pathway was isolated from Andrographis paniculata (ApHMGR) and expressed in bacterial cells. Full length ApHMGR (1937 bp) was submitted to NCBI with accession number MG271748.1. The open reading frame (ORF) was flanked by a 31-bp 5′-UTR, 118-bp 3′-UTR and ApHMGR contained a 1787 bp ORF encoding protein of 595 amino acids. ApHMGR protein was approximately 64 kDa, with isoelectric point of 5.75. Isolated ApHMGR was cloned into pET102 vector and expressed in E. coli BL21 (DE 3) cells, and characterized by SDS-PAGE. HPLC analysis for andrographolide content in leaf, stem and root of A. paniculata revealed highest in leaf tissue. The expression patterns of ApHMGR in different plant tissues using qRT-PCR revealed high in root tissue correlating with HPLC data. Three dimensional (3D) structural model of ApHMGR displayed 90% of the amino acids in most favored regions of the Ramachandran plot with 93% overall quality factor. ApHMGR was highly conserved with plant specific N-terminal membrane domains and C-terminal catalytic regions. Phylogenetic analysis showed A. paniculata sharing common ancestor with Handroanthus impetiginosus. 3D model of ApHMGR was screened for the interaction with substrates NADPH, HMG CoA and inhibitor using Auto Dock Vina. In silico analysis revealed that full length ApHMGR had extensive similarities to other plant HMGRs. The present communication reports the isolation of full length HMGR from A. paniculata, its heterologous expression in bacterial cells and in silico structural and functional characterization providing valuable genomic information for future molecular interventions.

    更新日期:2020-01-04
  • Comparative study of different glycating agents on human plasma and vascular cells
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-11-12
    Rashmi S. Tupe, Nilima Bangar, Arundhati Diwan, Dhanashri Changale, Shivani Choudhary, Shubhangi Chaware

    Diabetic complications are associated with the glycation and formation of advanced glycation end products (AGEs) which leads to structural modifications of biomolecules further affecting cells. Carbonyl compounds such as methylglyoxal and glyceraldehyde-3-phosphate are highly reactive and form an elevated amount of AGEs as compared to glucose and fructose. The investigation of glycation modifications by different compounds may be important to assess the specific pattern of biomolecular and cellular modifications and compare their glycation potential. The present work aims to comprehensively and comparatively examine the effect of glycating agents (glucose, fructose, ribose, methylglyoxal, and glyceraldehyde) on plasma, erythrocytes, platelets, and blood DNA. Glycation of plasma, cells, and DNA was initiated by incubating them with glycating agents for 24–48 h at 37 °C. Negative control samples (without glycating agents) were maintained simultaneously. After treatment, plasma and DNA samples were dialyzed and cell lysate was prepared. Markers of glycation (fructosamine), structural modifications (free amino, β-amyloid, absorption spectra), antioxidant indices (catalase activity, glutathione) and erythrocyte hemolysis were estimated. In the presence of glycating agents, there was a significant increase in the formation of fructosamine, structural modification markers and depletion in antioxidant indices. Overall results suggest that among all glycating agents; methylglyoxal and glyceraldehyde have more potency of glycation induced structural modifications in plasma and vascular cells. This indicates the specific glycation modifications in plasma and vascular cells by various glycating agents may be investigated further for controlling diabetic pathological changes.

    更新日期:2020-01-04
  • Tracking KPC-3-producing ST-258 Klebsiella pneumoniae outbreak in a third-level hospital in Granada (Andalusia, Spain) by risk factors and molecular characteristics.
    Mol. Biol. Rep. (IF 2.107) Pub Date : null
    Carmen Soria-Segarra,Pablo González-Bustos,Lorena López-Cerero,Felipe Fernández-Cuenca,María Dolores Rojo-Martín,María Amelia Fernández-Sierra,José Gutiérrez-Fernández

    The objective of this study was to determine clinical-epidemiological characteristics of the patients and the genetic characteristics of carbapenemase KPC-3-producing Klebsiella pneumoniae isolates belonging to sequence type ST258. The eligible study population was all patients with isolates detected between October 2015 and March 2017. Clinical-epidemiological and microbiological data were gathered on risk factors associated with infection by this clone. Antimicrobial susceptibility was determined using MicroScan system and diffusion in agar. Genes encoding carbapenemases were detected using PCR and Sanger sequencing. The sequence type was assigned by MLST, and the genetic relationship among clinical isolates was determined by pulsed field electrophoresis and by analysis of the genetic environment. The study included 23 individuals with isolates of KPC-3/ST258; the mean age was 77 year, and mean stay pre-isolation was 32 days; 81% received empirical antimicrobial treatment. Isolates were only susceptible to gentamicin (CIM ≤ 2 mg/L), tigecycline (CIM ≤ 1 mg/L), and colistin (CIM ≤ 2 mg/L). The isolates belonged to ST258, with five pulse types or subgroups. All isolates showed amplification of KPC, which was identified as KPC-3 variant. Gene blaKPC-3 was flanked by insertion sequences Kpn6 and Kpn7 within Tn4401 transposon isoform a. We report, for the first time in Spain, an 18-month outbreak by KPC-3-producing ST258 K. pneumoniae. Its acquisition was associated with a history of antimicrobial therapy, with three treatment options, and with high mortality. The detection of different pulse types is attributable to different introductions of the clone in our setting, supporting the need for multi-resistant isolate surveillance studies.

    更新日期:2019-11-01
  • Mutation-specific differences in arrhythmias and drug responses in CPVT patients: simultaneous patch clamp and video imaging of iPSC derived cardiomyocytes.
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-12-02
    R P Pölönen,H Swan,K Aalto-Setälä

    Catecholaminergic polymorphic ventricular tachycardia (CPVT) is an inherited cardiac disease characterized by arrhythmias under adrenergic stress. Mutations in the cardiac ryanodine receptor (RYR2) are the leading cause for CPVT. We characterized electrophysiological properties of CPVT patient-specific induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) carrying different mutations in RYR2 and evaluated effects of carvedilol and flecainide on action potential (AP) and contractile properties of hiPSC-CMs. iPSC-CMs were generated from skin biopsies of CPVT patients carrying exon 3 deletion (E3D) and L4115F mutation in RYR2. APs and contractile movement were recorded simultaneously from the same hiPSC-CMs. Differences in AP properties of ventricular like CMs were seen in CPVT and control CMs: APD90 of both E3D (n = 20) and L4115F (n = 25) CPVT CMs was shorter than in control CMs (n = 15). E3D-CPVT CMs had shortest AP duration, lowest AP amplitude, upstroke velocity and more depolarized diastolic potential than controls. Adrenaline had positive and carvedilol and flecainide negative chronotropic effect in all hiPSC CMs. CPVT CMs had increased amount of delayed after depolarizations (DADs) and early after depolarizations (EADs) after adrenaline exposure. E3D CPVT CMs had the most DADs, EADs, and tachyarrhythmia. Discordant negatively coupled alternans was seen in L4115F CPVT CMs. Carvedilol cured almost all arrhythmias in L4115F CPVT CMs. Both drugs decreased contraction amplitude in all hiPSC CMs. E3D CPVT CMs have electrophysiological properties, which render them more prone to arrhythmias. iPSC-CMs provide a unique platform for disease modeling and drug screening for CPVT. Combining electrophysiological measurements, we can gain deeper insight into mechanisms of arrhythmias.

    更新日期:2019-11-01
  • Upregulation of the clpB gene in response to heat shock and beta-lactam antibiotics in Acinetobacter baumannii.
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-12-02
    Waleska Yana Lazaretti,Elaine Luzia Dos Santos,José Luis da-Conceição Silva,Marina Kimiko Kadowaki,Rinaldo Ferreira Gandra,Alexandre Maller,Rita de Cássia Garcia Simão

    The role of the clpB gene encoding HSP/chaperone ClpB was evaluated in the multiresistant antibiotic cells of Acinetobacter baumannii (RS4 strain) under stress-induced heat shock and different beta-lactams. The expression of the clpB gene was assessed by qPCR during heat shock at 45 °C and subinhibitory concentrations of ampicillin (30 μg mL-1), amoxicillin + sulbactam (8/12 μg mL-1), cefepime (30 μg mL-1), sulfamethoxazole + trimethoprim (120/8 μg mL-1) and meropenem (18 μg mL-1). The results indicated a transient increase in clpB transcription in all treatments except cefepime. Both in the presence of ampicillin and amoxicillin/sulbactam for 20 min, the mRNA-clpB synthesis was 1.4 times higher than that of the control at time zero. Surprisingly, the mRNA-clpB levels were more than 30-fold higher after 10 min of incubation with meropenem and more than eightfold higher in the presence of trimethoprim/sulfamethoxazole. In addition, western blot assays showed that the RS4 strain treated with meropenem showed a marked increase in ClpB protein expression. Our data indicate that during exposure to beta-lactams, A. baumannii adjusts the transcription levels of the clpB mRNA and protein to respond to stress, suggesting that the chaperone may act as a key cellular component in the presence of antibiotics in this bacterium.

    更新日期:2019-11-01
  • A practicable method to prepare nitrated proteins with peroxynitrite and low concentration of sodium hydroxide.
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-12-02
    Nan Zhang,Xiaochen Gao,Weijia Zhang,Doaa Higazy,Ke Wang,Zhenfang Fu,Min Cui

    Peroxynitrite is an ion acting as a powerful oxidant and nucleophile, which plays a key role in the inflammation and aging process by nitrating tyrosine or tryptophan residues of the proteins. Nitration of a target protein is considered to be a proper method to study the behavioral change of the proteins being nitrated. The commonly used methods for peroxynitrite preparation in vitro usually contain high concentration of sodium hydroxide, which easily induces hydrolysis of target proteins. Accordingly, the method for peroxynitrite preparation was optimized in vitro by changing the sequence of hydrochloric acid and sodium hydroxide added. After different amount of hydrochloric acid added to the system following sodium nitrite, peroxynitrite can be yielded in a concentration up to 60 mM with sodium hydroxide as low as 17 mM. More importantly, biological activity of the target protein was well maintained after protein nitration since low sodium hydroxide was used.

    更新日期:2019-11-01
  • Testis-specific Arf promoter expression in a transposase-aided BAC transgenic mouse model.
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-10-05
    Caroline Y Sung,Yen-Ting Liu,Lynda B Bennett,Caitlin C Devitt,Stephen X Skapek

    CDKN2A is an evolutionarily conserved gene encoding proteins implicated in tumor suppression, ocular development, aging, and metabolic diseases. Like the human form, mouse Cdkn2a encodes two distinct proteins-p16Ink4a, which blocks cyclin-dependent kinase activity, and p19Arf, which is best known as a positive regulator of the p53 tumor suppressor-and their functions have been well-studied in genetically engineered mouse models. Relatively little is known about how expression of the two transcripts is controlled in normal development and in certain disease states. To better understand their coordinate and transcript-specific expression in situ, we used a transposase-aided approach to generate a new BAC transgenic mouse model in which the first exons encoding Arf and Ink4a are replaced by fluorescent reporters. We show that mouse embryo fibroblasts generated from the transgenic lines faithfully display induction of each transgenic reporter in cell culture models, and we demonstrate the expected expression of the Arf reporter in the normal testis, one of the few places where that promoter is normally expressed. Interestingly, the TGFβ-2-dependent induction of the Arf reporter in the eye-a process essential for normal eye development-does not occur. Our findings illustrate the value of BAC transgenesis in mapping key regulatory elements in the mouse by revealing the genomic DNA required for Cdkn2a induction in cultured cells and the developing testis, and the apparent lack of elements driving expression in the developing eye.

    更新日期:2019-11-01
  • Correction to: Potential role of lncRNA-TSIX, miR-548-a-3p, and SOGA1 mRNA in the diagnosis of hepatocellular carcinoma.
    Mol. Biol. Rep. (IF 2.107) Pub Date : 2019-05-03
    Alaa Habieb,Marwa Matboli,Hanaa El-Tayeb,Farid El-Asmar

    The original publication has been updated. The family name of Alaa Habieb contained a typo that has now been corrected.

    更新日期:2019-11-01
  • Molecular characterization of the Krüppel-homolog 1 and its role in ovarian development in Sogatella furcifera (Hemiptera: Delphacidae).
    Mol. Biol. Rep. (IF 2.107) Pub Date : null
    Kui Hu,Ping Tian,Lu Yang,Yan Tang,Lin Qiu,Hualiang He,Wenbing Ding,Youzhi Li

    Juvenile hormone (JH) plays a pivotal role in insect reproduction. The Krüppel-homolog 1 (Kr-h1) is a JH-inducible zinc finger transcription factor that has also been found to play a role in insect reproduction, however, its function varies across species. In this study, we cloned SfKr-h1 from Sogatella furcifera and investigated its role in ovarian development. The open reading frame of SfKr-h1 is 1 800 bp encoding 599 amino acids. The putative amino acid sequence of SfKr-h1 contains eight putative C2H2-type zinc finger domains and is highly homologous with the Kr-h1s of other hemipteran species. Expression of SfKr-h1 peaked 96 h after adult emergence and was highest in the ovary. RNA interference (RNAi) knockdown of SfKr-h1 substantially reduced the transcription of SfVg, and arrested ovarian development. These results suggest that SfKr-h1 is critical for normal ovarian development in S. furcifera.

    更新日期:2019-11-01
  • Isolation, characterization and in silico analysis of 3-Hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR) gene from Andrographis paniculata (Burm. f) Nees.
    Mol. Biol. Rep. (IF 2.107) Pub Date : null
    Mote Srinath,Byreddi Bhavani Venkata Bindu,Ayeti Shailaja,Charu Chandra Giri

    3-Hydroxy-3-methylglutaryl-coenzymeA reductase (HMGR), the first rate-limiting enzyme of Mevalonate (MVA) pathway was isolated from Andrographis paniculata (ApHMGR) and expressed in bacterial cells. Full length ApHMGR (1937 bp) was submitted to NCBI with accession number MG271748.1. The open reading frame (ORF) was flanked by a 31-bp 5'-UTR, 118-bp 3'-UTR and ApHMGR contained a 1787 bp ORF encoding protein of 595 amino acids. ApHMGR protein was approximately 64 kDa, with isoelectric point of 5.75. Isolated ApHMGR was cloned into pET102 vector and expressed in E. coli BL21 (DE 3) cells, and characterized by SDS-PAGE. HPLC analysis for andrographolide content in leaf, stem and root of A. paniculata revealed highest in leaf tissue. The expression patterns of ApHMGR in different plant tissues using qRT-PCR revealed high in root tissue correlating with HPLC data. Three dimensional (3D) structural model of ApHMGR displayed 90% of the amino acids in most favored regions of the Ramachandran plot with 93% overall quality factor. ApHMGR was highly conserved with plant specific N-terminal membrane domains and C-terminal catalytic regions. Phylogenetic analysis showed A. paniculata sharing common ancestor with Handroanthus impetiginosus. 3D model of ApHMGR was screened for the interaction with substrates NADPH, HMG CoA and inhibitor using Auto Dock Vina. In silico analysis revealed that full length ApHMGR had extensive similarities to other plant HMGRs. The present communication reports the isolation of full length HMGR from A. paniculata, its heterologous expression in bacterial cells and in silico structural and functional characterization providing valuable genomic information for future molecular interventions.

    更新日期:2019-11-01
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