Introduction

We sought to quantify targeted metabolites (d-lactate, pyruvate, urea, ammonia) and the cytokine IL-8 produced by human cervicovaginal epithelial cells co-cultured with Ureaplasma urealyticum (a preterm birth-associated bacterium) or Lactobacillus crispatus (a healthy vaginal commensal associated with term birth).

Methods

Concentrations of d-lactate, pyruvate, urea and ammonia measured by enzyme-based spectrophotometry and IL-8 by ELISA were determined and compared between monolayer-cultured HeLa cells (ATCC 35241) infected with strains of U. urealyticum (ATCC 27618, 0.5 mL = 3640 CFU/mL, U. urealyticum) or L. crispatus (ATCC 33820, MOI = 10,000, 1000 and 100, L. crispatus) and incubated in 5% CO₂ at 37 °C for 24 h. Uninfected HeLa cells (Hc) were used as controls and cytotoxicity was determined by the amount (optical density) of lactate dehydrogenase (LDH) released by the dead HeLa cells.

Results

The amount of LDH released by untreated Hc (P = 0.002) and U. urealyticum-infected cells (P < 0.0001) was higher than those of L. crispatus-infected cells, with U. urealyticum-infected cells recording the highest % cytotoxicity and L. crispatus-infected cells MOI 10,000 (Lc10,000) the least (P < 0.0001). Though there was no significant difference in the concentration of urea between the samples, U. urealyticum-infected cells showed higher ammonia compared to other samples (p = 0.03). In contrast, all L. crispatus samples had higher d-lactate than untreated Hc (p = 0.01) and U. urealyticum-infected cells (P = 0.01). Also, Lc10,000 had the highest d-lactate (p = 0.001) and lowest pyruvate (P = 0.04, excluding UU) compared to other samples. Furthermore, U. urealyticum-infected cells produced the highest IL-8 (P = 0.01) compared to other samples, with Lc10,000 producing undetectable levels.

Conclusion

Infection of cervicovaginal epithelial cells by U. urealyticum stimulates production of ammonia from urea and induces elevated IL-8 production possibly leading to significantly higher cytotoxicity. In contrast, L. crispatus appeared protective against HeLa cell inflammation and death, producing more d-lactate and less IL-8, consistent with a role for L. crispatus in promoting vaginal floral health and reducing infection/inflammation-associated preterm birth.

中文翻译：

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曲霉乳杆菌和解脲脲原体感染的宫颈阴道上皮细胞产生的差异性细胞因子和代谢产物。

介绍

我们试图量化与解脲脲原体（一种与早产相关的细菌）或cristobacillus crispatus（一种健康的阴道细菌）共培养的人宫颈阴道上皮细胞产生的靶向代谢物（d-乳酸，丙酮酸，尿素，氨水）和细胞因子IL-8。与足月出生有关的共鸣）。

方法

确定了酶解分光光度法测定的d-乳酸，丙酮酸，尿素和氨的浓度，通过ELISA测定了IL-8的浓度，并比较了解脲脲原体（ATCC 27618，0.5 mL ）感染的单层培养HeLa细胞（ATCC 35241）= 3640 CFU /毫升，解脲支原体）或卷曲乳杆菌（ATCC 33820，MOI = 10,000，1000和100，卷曲乳杆菌），并温育，在5％CO ₂在37℃下24小时。将未感染的HeLa细胞（Hc）用作对照，并通过死亡的HeLa细胞释放的乳酸脱氢酶（LDH）的量（光学密度）确定细胞毒性。

结果

未经处理的Hc（P  = 0.002）和解脲支原体感染的细胞（P  <0.0001）释放的LDH量高于脆皮乳酸菌感染的细胞，其中解脲脲原体感染的细胞记录的％细胞毒性和L. crispatus-感染细胞MOI 10,000（Lc10,000）最少（P  <0.0001）。尽管样品之间的尿素浓度没有显着差异，但是与其他样品相比，解脲支原体感染的细胞显示出更高的氨（p  = 0.03）。相比之下，所有香菇样品的d均较高。-乳酸比未处理的Hc（p = 0.01）和解脲支原体感染的细胞（P  = 0.01）。同样， 与其他样品相比，Lc10,000具有最高的d-乳酸（p  = 0.001）和最低的丙酮酸（P = 0.04，不包括UU）。此外， 与其他样品相比，解脲支原体感染的细胞产生最高的IL-8（P = 0.01），Lc10,000产生不可检测的水平。

结论

解脲支原体感染宫颈阴道上皮细胞可刺激尿素产生氨并诱导升高的IL-8产生，可能导致明显更高的细胞毒性。相反，薄脆乳杆菌似乎对HeLa细胞炎症和死亡具有保护作用，产生更多的d-乳酸和更少的IL-8，这与薄脆乳杆菌在促进阴道花的健康和减少感染/炎症相关的早产中的作用一致。