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Published on: May 10, 2024
MicroRNA-320-3p promotes the progression of acute pancreatitis by blocking DNMT3a-mediated MMP8 methylation in a
Huan Gu1, Jie Peng1, Meng Wang1
1Department of Gastroenterology, Xiangya Hospital, Central South University, Changsha, Hunan 410008, PR China.
Abstract:
In this research, we screened out two genes upregulated in mice with acute pancreatitis (AP) by gene sequencing: microRNA (miR)-320-3p and matrix metalloprotease 8 (MMP8). This study was designed to determine whether miR-320-3p and MMP8 participate in AP development and explore the mechanisms, with a new idea for clinical diagnosis and treatment of AP. Expression of miR-320-3p, DNA methyltransferase 3a (DNMT3a), and MMP8 in mouse pancreatic tissues and AR42J cells was tested by RT-qPCR and western blot assays. Pancreatic pathological changes, serum amylase and lipase, and inflammatory factors in mouse serum and cell supernatant were measured by hematoxylin-eosin staining, automation analyzer, and enzyme-linked immunosorbent assay, respectively. Cell proliferation and apoptosis were determined by CCK-8 assay and flow cytometry. The interaction between miR-320-3p, DNMT3a, and MMP8 was verified by luciferase activity assay, ChIP-qPCR, and MSP assay. High expression of miR-320-3p and MMP8, and low expression of DNMT3a were observed in pancreatic tissues of AP mice and caerulein-induced AP cellular model. Downregulation of miR-320-3p alleviated injury of mouse pancreas, reduced the levels of serum amylase and lipase, and blocked inflammatory factor levels in AP mice. In caerulein-induced AP cellular models, inhibiting miR-320-3p facilitated proliferation and inhibited apoptosis. Upregulation of MMP8 resulted in the opposite results, which could be reversed by simultaneous inhibition of miR-320-3p. miR-320-3p targeted DNMT3a, and downregulating miR-320-3p promoted DNMT3a expression. Moreover, DNMT3a promoted DNA methylation in MMP8 promoter region, thereby inhibiting MMP8 expression in AP mouse and cellular models. This research suggests that miR-320-3p inhibits DNMT3a to reduce MMP8 methylation and increase MMP8 expression, thereby promoting AP progression.
Insights
MicroRNA-320-3p and matrix metalloprotease 8 (MMP8) are upregulated in acute pancreatitis (AP). MiR-320-3p promotes AP by inhibiting DNMT3a, which reduces MMP8 methylation and increases MMP8 expression.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- Acute pancreatitis (AP) is a serious inflammatory condition with complex molecular mechanisms.
- Identifying key regulatory genes like microRNA (miR)-320-3p and matrix metalloprotease 8 (MMP8) is crucial for understanding AP.
- Gene sequencing identified miR-320-3p and MMP8 as upregulated in AP mouse models.
Purpose of the Study:
- To investigate the roles of miR-320-3p and MMP8 in the development of acute pancreatitis.
- To elucidate the underlying molecular mechanisms involving miR-320-3p, DNA methyltransferase 3a (DNMT3a), and MMP8.
- To explore potential new strategies for the clinical diagnosis and treatment of AP.
Main Methods:
- Gene expression analysis using RT-qPCR and western blot in mouse pancreatic tissues and AR42J cells.
- Assessment of pancreatic damage, serum biomarkers (amylase, lipase), and inflammatory factors.
- Cellular assays including CCK-8 for proliferation and flow cytometry for apoptosis.
- Molecular interaction studies using luciferase activity assay, ChIP-qPCR, and MSP assay.
Main Results:
- High expression of miR-320-3p and MMP8, with low DNMT3a, observed in AP models.
- Downregulation of miR-320-3p alleviated pancreatic injury and reduced inflammatory markers in vivo.
- Inhibition of miR-320-3p increased proliferation and decreased apoptosis in AP cellular models.
- miR-320-3p targets DNMT3a; DNMT3a promotes DNA methylation of the MMP8 promoter, inhibiting MMP8 expression.
Conclusions:
- miR-320-3p promotes AP progression by inhibiting DNMT3a, leading to reduced MMP8 methylation and increased MMP8 expression.
- The miR-320-3p/DNMT3a/MMP8 pathway represents a novel mechanism in AP pathogenesis.
- This pathway offers potential targets for future AP diagnosis and therapeutic interventions.
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