牛磺熊去氧胆酸抗炎和免疫调节作用的转录组学研究。
Transcriptome investigation of anti-inflammation and immuno-regulation mechanism of taurochenodeoxycholic acid.
机构信息
College of Veterinary Medicine, Inner Mongolia Agricultural University, Hohhot, China.
Key Laboratory of Clinical Diagnosis and Treatment Techniques for Animal Disease, Ministry of Agriculture, Hohhot, China.
出版信息
BMC Pharmacol Toxicol. 2021 Apr 29;22(1):23. doi: 10.1186/s40360-021-00491-0.
BACKGROUND
Taurochenodeoxycholic acid (TCDCA) is one of the major active components in bile acid. It was proven to have inhibitory activities on inflammation and also participate in host immuno-regulation. TCDCA exerts anti-inflammatory and immuno-regulatory effects through the glucocorticoid receptor (GR) mediated genomic signaling pathway and the G protein-coupled bile acid receptor 5 (TGR5) mediated AC-cAMP-PKA signaling pathway. However, it is unclear whether GR or TGR5 plays an important role in the regulatory effects of TCDCA. In order to further investigate this effects mechanism of TCDCA, the research use the transcriptome to identify the major genes and pathway in the anti-inflammatory and immuno-regulatory effects.
METHODS
After the Fibroblast-like synoviocytes (FLS) being treated by different concentrations (10, 10 and 10 M) of TCDCA for 12 h, the resulting mRNA was analyzed by RNA-seq. The differentially expressed genes were screened from sequencing results using bioinformatics techniques. In the next step, other published literature were referred in order to find out whether those genes mentioned above are related to inflammation. The final selected differentially expressed genes associated with inflammation were then validated by q-PCR and western blot assays.
RESULTS
Five genes associated with anti-inflammatory and immuno-regulatory effects, include Glyceraldehyde-3-phosphate dehydrogenase (GAPDH), Glutathione peroxidase 3 (GPX3), Serine/arginine-rich splicing factor-9 (SRSF9), Connective tissue growth factor (CTGF) and Cystatin B (CSTB) were identified. TCDCA at the concentrations of 10, 10 and 10 M significantly (p < 0.05) up-regulate the mRNA and protein expression of SRSF9 and GPX3 and also up-regulate the mRNA expression of CSTB, CTGF and GAPDH. RNA-seq results of GPX3 and SRSF9 expression were consistent with q-PCR results, while q-PCR results of CTGF, GAPDH showed inconsistent with their RNA-seq results. Q-PCR result of CSTB expression also showed inconsistent with the RNA-seq result.
CONCLUSIONS
The anti-inflammatory and immuno-regulatory activities of TCDCA are proven to be related to the up-regulation expression of GPX3, SRSF9 and CSTB.
背景
牛磺胆酸（TCDCA）是胆汁酸中的主要活性成分之一。已证实其具有抗炎活性，并且参与宿主免疫调节。TCDCA 通过糖皮质激素受体（GR）介导的基因组信号通路和 G 蛋白偶联胆汁酸受体 5（TGR5）介导的 AC-cAMP-PKA 信号通路发挥抗炎和免疫调节作用。然而，尚不清楚 GR 或 TGR5 在 TCDCA 的调节作用中是否发挥重要作用。为了进一步研究 TCDCA 的这种作用机制，研究人员使用转录组来鉴定抗炎和免疫调节作用的主要基因和途径。
方法
将成纤维样滑膜细胞（FLS）用不同浓度（10、10 和 10μM）的 TCDCA 处理 12 小时后，用 RNA-seq 分析其产生的 mRNA。使用生物信息学技术从测序结果中筛选差异表达基因。下一步，参考其他已发表的文献，以确定上述基因是否与炎症有关。最后，通过 q-PCR 和 Western blot 检测验证与炎症相关的差异表达基因。
结果
共鉴定出与抗炎和免疫调节作用相关的 5 个差异表达基因，包括甘油醛-3-磷酸脱氢酶（GAPDH）、谷胱甘肽过氧化物酶 3（GPX3）、丝氨酸/精氨酸丰富剪接因子 9（SRSF9）、结缔组织生长因子（CTGF）和胱抑素 B（CSTB）。浓度为 10、10 和 10μM 的 TCDCA 显著（p<0.05）上调 SRSF9 和 GPX3 的 mRNA 和蛋白表达，同时上调 CSTB、CTGF 和 GAPDH 的 mRNA 表达。GPX3 和 SRSF9 表达的 RNA-seq 结果与 q-PCR 结果一致，而 CTGF、GAPDH 的 q-PCR 结果与 RNA-seq 结果不一致。CSTB 表达的 q-PCR 结果也与 RNA-seq 结果不一致。
结论
TCDCA 的抗炎和免疫调节活性与 GPX3、SRSF9 和 CSTB 的上调表达有关。
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