Study on the mechanism of hypericin improving acute pancreatitis in mice by regulating NLRP3 inflammasome

doi:10.11958/20251147

Abstract

Abstract:

Objective To investigate the therapeutic effect of hypericin on acute pancreatitis (AP) in mice and its effect on NLRP3 inflammasome signaling pathway. Methods The AP model in mice was established with caerulein (CER). The mice were divided into the normal control group, the model group (AP group), the low-dose HY group (CER + HY 5 mg/kg group), the medium-dose HY group (CER + HY 10 mg/kg group) and the high-dose HY group (CER + HY 20 mg/kg group), with 10 mice in each group. The 266-6 mouse pancreatic acinar cancer cells were treated with cholecystokinin (CCK) and divided into the control group, the AP group, the CCK + HY 1 μmol/L group, the CCK + HY 2 μmol/L group and the CCK + HY 4 μmol/L group. The activities of amylase (AMS), lipase, trypsin and myeloperoxidase (MPO) in the serum of each group of mice, and levels of inflammatory factors interleukin (IL)-1β and tumor necrosis factor (TNF)-α were detected by enzyme-linked immunosorbent assay (ELISA). The expression of NOD-like receptor family protein 3 (NLRP3) was detected by Western blot assay. The mRNA levels of NLRP3, caspase (Caspase)-1, IL-1β, TNF-α and IL-18 in pancreatic tissue of mice were detected by real-time quantitative PCR (q-PCR). The cell survival rate of cells in each group was detected by CCK8 method. The mRNA expression levels of NLRP3, Caspase-1 and IL-18 in each group of cells were detected by q-PCR. Results Compared with the normal control group, the levels of AMS, lipase, MPO, trypsin, IL-1β and TNF-α in serum of the model group, and the mRNA and protein expression levels of NLRP3, IL-1β, TNF-α, IL-18 and Caspase-1 in pancreatic tissue were increased (P<0.01). Compared with the model group, the levels of AMS, IL-1β and TNF-α, the enzymatic activity of trypsin in serum, and the mRNA levels of IL-1β, TNF-α, IL-18 and Caspase-1 in pancreatic tissue were decreased in the low-, medium- and high-dose HY groups. The serum levels of lipase and MPO and the mRNA expression levels of NLRP3 in pancreatic tissue were decreased in the medium- and high-dose HY groups (P<0.05). Compared with the AP group, the cell survival rates were increased in the CCK + HY 1 μmol/L group, the CCK + HY 2 μmol/L group and the CCK + HY 4 μmol/L group, and the mRNA levels of NLRP3, IL-18 and Caspase-1 were decreased in a dose-dependent manner (P<0.05). Conclusion Hypericin can effectively treat AP in vivo and in vitro, and its therapeutic effect may be related to the regulation of NLRP3 inflammasome signaling pathway.

Key words: pancreatitis, Hypericin, NLR family, pyrin domain-containing 3 protein, inflammation

CLC Number:

R576.1

Figures/Tables 7

References 27

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基因名称	引物序列（5'→3'）	产物大小/bp
β-actin	上游：AGAGGGAAATCGTGCGTGAC	236
β-actin	下游：CAATAGTGATGACCTGGCCGT	236
NLRP3	上游：TCCACAATTCTGACCCACAA	228
NLRP3	下游：ACCTCACAGAGGGTCACCAC	228
Caspase-1	上游：GGGCCCCAGGCAAGCCAAATC	225
Caspase-1	下游：AGGGCAAGACGTGTACGAGTGGT	225
IL-1β	上游：TCTTTGAAGTTGACGGACCC	235
IL-1β	下游：TGAGTGATACTG CCTGCCTG	235
TNF-α	上游：GTCGCTACCGTCGTGACTTC	216
TNF-α	下游：CAGACATGCACCTACCCAGC	216
IL-18	上游：ACACCAGCCTGGCTTCCATC	208
IL-18	下游：TTGGAGCTGGAGCTGCTTATAGTTG	208

基因名称	引物序列（5'→3'）	产物大小/bp
β-actin	上游：AGAGGGAAATCGTGCGTGAC	236
β-actin	下游：CAATAGTGATGACCTGGCCGT	236
NLRP3	上游：TCCACAATTCTGACCCACAA	228
NLRP3	下游：ACCTCACAGAGGGTCACCAC	228
Caspase-1	上游：GGGCCCCAGGCAAGCCAAATC	225
Caspase-1	下游：AGGGCAAGACGTGTACGAGTGGT	225
IL-1β	上游：TCTTTGAAGTTGACGGACCC	235
IL-1β	下游：TGAGTGATACTG CCTGCCTG	235
TNF-α	上游：GTCGCTACCGTCGTGACTTC	216
TNF-α	下游：CAGACATGCACCTACCCAGC	216
IL-18	上游：ACACCAGCCTGGCTTCCATC	208
IL-18	下游：TTGGAGCTGGAGCTGCTTATAGTTG	208

组别	AMS/（U/L）	脂肪酶/（U/L）
正常对照组	1 957.14±23.80	189.92±13.03
模型组	6 512.55±100.10^a	383.03±29.85^a
HY低剂量组	5 031.68±43.70^b	348.75±28.77
HY中剂量组	4 137.10±40.90^bc	310.43±22.43^bc
HY高剂量组	2 129.28±31.20^bcd	231.32±17.65^bcd
F	399.100^＊＊	82.500^＊＊

组别	AMS/（U/L）	脂肪酶/（U/L）
正常对照组	1 957.14±23.80	189.92±13.03
模型组	6 512.55±100.10^a	383.03±29.85^a
HY低剂量组	5 031.68±43.70^b	348.75±28.77
HY中剂量组	4 137.10±40.90^bc	310.43±22.43^bc
HY高剂量组	2 129.28±31.20^bcd	231.32±17.65^bcd
F	399.100^＊＊	82.500^＊＊

组别	胰蛋白酶/（U/mg）	MPO/（U/g）
正常对照组	4.59±0.33	1.25±0.08
模型组	15.22±1.05^a	2.77±0.36^a
HY低剂量组	11.28±0.75^b	2.61±0.39
HY中剂量组	8.38±0.66^b	2.33±0.26^bc
HY高剂量组	5.23±0.34^bcd	1.41±0.11^bcd
F	209.900^＊＊	45.550^＊＊