Research paperUpregulation of JHDM1D-AS1 protects PDLSCs from H2O2-induced apoptosis by decreasing DNAJC10 via phosphorylation of eIF2α
Introduction
Almost 90% people worldwide are suffering from periodontal related diseases [1]. Recently, stem cell based regeneration of periodontal tissues has showed promising potential [2]. Periodontal ligament stem cells (PDLSCs) are multipotent stem cells which can be differentiated into cementum, peripheral nerves, and other type of cells [[3], [4], [5], [6], [7]]. The in situ PDLSCs are limited and in vitro cultured PDLSCs endure apoptotic death which restricts the amount of PDLSCs that can be utilized in regenerative medicine. Previous studies have shown that genetic background affects the quality and function of PDLSCs [8,9]. External stimuli, such as hypoxia, are also well known factors that regulate PDLSC apoptosis [[10], [11], [12], [13], [14]]. Accumulating evidence has indicated that non-coding RNA, especially long non-coding RNA (lncRNA) response to external stimuli regulates cellular processes [15,16]. Oxidative stress is characterized by dysregulated levels of reactive oxygen species (ROS) which are primarily generated from mitochondrial complexes I and III. It has been previously demonstrated that oxidative stress is critical for stem cell aging and survival [[17], [18], [19]]; however, the underlying mechanisms are still unclear.
LncRNAs are a class of non-protein coding RNAs consisting of more than 200 nucleotides [20]. Studies have illustrated that lncRNAs are critical in multiple biological processes, including the regulation of stem cell and osteogenic differentiation [21]. Moreover, previous studies have shown that lncRNA HIF1α-anti-sense 1 (HIF11α-AS1) regulates osteogenic differentiation of bone marrow stromal cells and lncRNA ANCR modulates Wnt signaling to regulate osteogenic differentiation of PDLSCs [9,22]. Recently, large scale screening for lncRNAs that are critical for PDLSC osteogenic differentiation has been performed [23]. Since non-coding RNAs exhibit huge potentials in stem cell regulation, and disease diagnosis and treatment [24,25], it is important to further analyze the role of lncRNAs in PDLSC homeostasis. Histone demethylase Jumonji C domain containing histone demethylase 1 homolog D (JHDM1D) was initially identified to harbor a plant homeodomain and a Jumonji domain that binds to specific histone modification sites [26]. In addition, JHDM1D inhibited tumor growth by suppressing angiogenesis [27]. Importantly, a previous study has shown that the lncRNA JHDM1D antisense 1 promotes pancreatic tumorigenesis by regulating angiogenesis in response to nutrient starvation. In addition, JHDM1D-AS1 increases blood vessel formation and macrophage infiltration [28]. However, whether JHDM1D-AS1 is active in stem cells is unknown.
Eukaryotic DNAJ family proteins are conserved with the first J protein identified in Escherichia coli [29]. It was proposed that DNAJ is required for the loading of Hsc70 onto a nascent polypeptide to form a proper folding complex [30]. Previous studies have demonstrated that DNAJ family proteins are responsive to environmental stimuli, such as reactive oxygen species (ROS) [31,32]. However, how DNAJ proteins regulate PDLSCs, especially under environmental stress, is largely unknown.
Eukaryotic Translation Initiation Factor 2 Subunit Alpha (EIF2α) is a key regulator of apoptosis in multiple cell types [33]. Recently, it has been reported that EIF2α is responsive to endoplasmic reticulum stress and regulates the osteoblast differentiation [34]. These reports suggested that eIF2α may function as a link between stress response and apoptosis.
Hence, we explored the expression level of JHDM1D-AS1 in PDLSCs treated with H2O2 and examined the apoptotic status of PDLSCs. In addition, we assessed the changes in apoptotic signaling in PDLSCs overexpressing JHDM1D-AS1. Interestingly, our results revealed a regulatory axis involving JHDM1D-AS1, DNAJC10, and apoptotic inhibitor Bcl-2. Besides, we also examined the involvement of phosphorylated-eIF2α in JHDM1D-AS1-dependent regulation of PDLSC apoptosis.
Section snippets
Periodontal ligament stem cell isolation and culture
The rat PDLSCs were isolated by modifying the method described previously [35]. Briefly, PDLSCs were collected from the periodontal ligament by cutting periodontal ligament into small pieces and digesting in 0.3% collagenase (Sigma) at 37 °C for 4 h. The cells obtained were then seeded in culture dishes and cultured in DMEM (Dulbecco's modified Eagle's medium, Hyclone), containing 10% fetal bovine serum (Gibco), 100 IU/mL penicillin, and 100 IU/mL streptomycin (Gibco), in a humidified
H2O2 treatment transiently reduces JHDM1D-AS1 expression in human PDLSCs
To study the role of lncRNA JHDM1D-AS1 in human PDLSCs, we first established H2O2-induced apoptosis in human PDLSCs. We then treated the isolated PDLSCs with 100 μM, 200 μM, and 400 μM H2O2, and then, assessed the cell viability via MTT experiment. Our data showed that H2O2 treatment significantly affected the cell viability at the concentrations of 200 μM and 400 μM (Fig. 1A). Furthermore, we monitored apoptosis of PDLSCs treated with H2O2, and found that 200 μM H2O2 significantly induced
Discussion
PDLSCs are a promising tool for regenerative medicine [36]. Since PDLSCs potentially reside in extreme environment, it is of huge significance to monitor how PDLSCs response to external stimuli. Our data first revealed that under hydrogen peroxide treatment, a long non-coding RNA, JHDM1D-AS1, is rapidly downregulated. It has been reported that lncRNAs, MALAT1 and Sox2ot, were protective non-coding RNAs against hydrogen peroxide exposure [16,37]. These data suggested a promising role of these
Conclusion
Our data showed that lncRNA JHDM1D-AS1 is inhibited in PDLSCs by hydrogen peroxide treatment, which, in turn, leads to upregulation of DNAJC10. Upregulated DNAJC10 promoted the de-phosphorylation of phosphorylated eIF2α, which, in turn, caused the inhibition of anti-apoptotic gene, Bcl-2, and induction of pro-apoptotic gene, Bax, and thus, promoted apoptosis. Our data illustrated an alternative pathway of how hydrogen peroxide promoted PDLSC apoptosis, which provided insights into PDLSCs
Funding
This work was supported by grants from the Science and Technology Project of Yubei District, Chongqing (grant No. 2017 (agriculture society) 45), and the Administration of traditional Chinese Medicine of Zhejiang Province (2014ZA069), the Department of Education of Zhejiang Province(Y201534720).
Availability of supporting data
The datasets used and/or analyzed in the current study are available from the corresponding author based on a reasonable request.
Author contributions
BS and SB performed statistical analysis, evaluated the results, and drafted the manuscript. GN, BS, and SB participated in the conception and design of the study. YC, GY, XM, and FX contributed to laboratory measurements and data assurance. All authors have read and approved the final version of the manuscript.
Ethics approval and consent to participate
Not applicable.
Consent for publication
Not applicable.
Competing interests
The authors declare that they have no competing interests.
Acknowledgements
The authors thank all current and previous members of the laboratory for valuable discussions and support.
References (40)
- et al.
Periodontal diseases
Lancet
(2005) - et al.
Periodontal ligament stem cell-mediated treatment for periodontitis in miniature swine
Stem Cell.
(2008) - et al.
Investigation of multipotent postnatal stem cells from human periodontal ligament
Lancet
(2004) - et al.
Down-regulated non-coding RNA (lncRNA-ANCR) promotes osteogenic differentiation of periodontal ligament stem cells
Arch. Oral Biol.
(2015) - et al.
Protective effects of molecular hydrogen on steroid-induced osteonecrosis in rabbits via reducing oxidative stress and apoptosis
BMC Muscoskelet. Disord.
(2017) - et al.
The long non-coding RNA MALAT1 activates Nrf2 signaling to protect human umbilical vein endothelial cells from hydrogen peroxide
Biochem. Biophys. Res. Commun.
(2018) - et al.
DnaJ-like proteins: molecular chaperones and specific regulators of Hsp70
Trends Biochem. Sci.
(1994) - et al.
Redox-regulated cochaperone activity of the human DnaJ homolog Hdj2
Free Radic. Biol. Med.
(2006) - et al.
PERK-eIF2alpha-ATF4 pathway mediated by endoplasmic reticulum stress response is involved in osteodifferentiation of human periodontal ligament cells under cyclic mechanical force
Cell. Signal.
(2016) - et al.
Clinical utility of stem cells for periodontal regeneration
Periodontology
(2000)
Allogeneic stem cells from deciduous teeth in treatment for periodontitis in miniature swine
J. Periodontol.
Plasticity of stem cells derived from adult periodontal ligament
Regen. Med.
Efficacy of periodontal stem cell transplantation in the treatment of advanced periodontitis
Cell Transplant.
[The crosstalk between canonical and noncanonical Wnt signaling pathway in osteoblast differentiation of periodontal ligament stem cells in inflammatory microenvironments]
Zhonghua kou qiang yi xue za zhi = Zhonghua kouqiang yixue zazhi = Chinese journal of stomatology.
[Effect of trichostatin A on the osteogenic differentiation potential of periodontal ligament stem cells in inflammatory microenvironment induced by tumor necrosis factor-alpha stimulation]
Zhonghua kou qiang yi xue za zhi = Zhonghua kouqiang yixue zazhi = Chinese journal of stomatology
High glucose concentrations suppress the proliferation of human periodontal ligament stem cells and their differentiation into osteoblasts
J. Periodontol.
Estrogen enhances the bone regeneration potential of periodontal ligament stem cells derived from osteoporotic rats and seeded on nano-hydroxyapatite/collagen/poly(L-lactide)
Int. J. Mol. Med.
Hypoxia enhances periodontal ligament stem cell proliferation via the MAPK signaling pathway
Genet. Mol. Res.: GMR
Long non coding RNA-UCA1 contributes to cardiomyocyte apoptosis by suppression of p27 expression
Cell. Physiol. Biochem.: Int. J. Exper. Cell. Physiol. Biochem. Pharmacol.
Reprogramming to a pluripotent state modifies mesenchymal stem cell resistance to oxidative stress
J. Cell Mol. Med.
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2021, Life SciencesCitation Excerpt :Many studies have shown the protective function of JHDM1D-AS1 against external damage to the cells. Shi et al. recently reported that the expression of JHDM1D-AS1 is essential for the survival of periodontal ligament stem cells with hydrogen peroxide treatment [24]. Similarly, Liu et al. found that the upregulation of JHDM1D-AS1 suppresses neuronal apoptosis and microglial inflammation [25].
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2020, International ImmunopharmacologyCitation Excerpt :The gain of function experiments confirmed that JHDM1D-AS1 reduces the inflammation of microglia and also represses inflammation-induced neuronal damage. In fact, previous studies have shown that JHDM1D-AS1 attenuates H2O2-mediated apoptosis of Periodontal stem cells (PDLSCs) [12], further confirming that JHDM1D-AS1 exerts a protective role against cell damage. Additionally, this study found that miR-101-3p promoted the LPS-induced microglia cell inflammatory response and the IL-1β-treated neuronal apoptosis.
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2020, Biomedicine and PharmacotherapyCitation Excerpt :LncRNAs are also involved in the regulation of osteogenic differentiation of PDLSCs. JHDM1D-AS1 modulates PDLSC apoptosis through suppression of the heat shock protein DNAJC10 [39]. The lncRNA-TWIST1 has been shown to enhance osteogenic differentiation in periodontal mesenchymal stem cells from both periodontitis patients and healthy subjects through suppression of the TWIST1 expression [40].
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The first two authors contributed equally to this work.