Micheliolide ameliorates renal lesion of unilateral ureteral obstruction mice by inhibiting NF-κB/NLRP3 axis

doi:10.11958/20230150

Abstract

Abstract:

Objective To investigate the effect and mechanism of micheliolide on renal lesions of unilateral ureteral obstruction (UUO) mice. Methods Male C57BL/6J mice were randomly divided into the sham group, the UUO (model) group and the UUO+micheliolide (administration) group. Mice in the UUO+ micheliolide group were given micheliolide by gavage at 25 mg/kg body weight after the establishment of the animal model. Renal tissue samples were collected 8 days after operation, and the renal tissue was stained with HE, Masson and TUNEL staining. The protein expressions of nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3), caspase-1, interleukin (IL)-1β, tumor necrosis factor (TNF)-α and (phosphorylation)- nuclear transcription factor (NF)-κB p65 were detected by immunohistochemistry and Western blot assay. Results HE staining showed that there were no pathological changes in renal tissue in the sham group. Mice in the UUO group had progressive renal tubule dilation and interstitial edema with a small amount of inflammatory cell infiltration. The renal lesions were significantly improved in the UUO+MCL group compared with those of the UUO group. Masson staining showed that compared with the sham group, the collagen volume fraction (CVF) was significantly increased in the UUO group, while CVF was significantly reduced in the UUO+MCL group than that in the UUO group. TUNEL staining showed that compared with the sham group, the apoptosis rate was increased in the UUO group, and the apoptosis rate was significantly decreased in the UUO+MCL group than that of the UUO group. Western blot assay and immunohistochemistry result showed that the expression levels of NLRP3, caspase-1, IL-1β, TNF-α and p-NF-κB p65 were increased in kidney tissue of the UUO group compared with those of the sham group. Compared with the UUO group, NLRP3, caspase-1, IL-1β, TNF-α and p-NF-κB p65 were significantly decreased in the UUO+MCL group. Conclusion The expression of NLRP3 inflammasome and related inflammatory factors in renal tissue of mice with unilateral ureteral obstruction are increased. Micheliolide can reduce the inflammatory response of renal tubule interstitium by inhibiting NF-κB pathway and NLRP3 inflammasome activation, thereby improving renal fibrosis.

Key words: ureteral obstruction, fibrosis, NLR family, pyrin domain-containing 3 protein, NF-kappa B, micheliolide

CLC Number:

R692.5

Figures/Tables 10

Fig.1 Schematic diagram of animal experiment design

Fig.2 HE staining results of renal tissue in each group (×400)

Fig.3 Masson staining results of kidney tissue in each group (×400)

Fig.4 TUNEL staining results of kidney tissue in each group (×400)

Fig.5 Immunohistochemical results of of NLRP3, caspase-1, IL-1β and TNF-ɑ in renal tissue (×400)

Tab.1 Comparison of the expression levels of NLRP3, caspase-1, IL-1β and TNF-α proteins in renal tissue between the three groups

组别	NLRP3	caspase-1	IL-1β	TNF-α
Sham组	0.356±0.031	0.166±0.018	0.226±0.040	0.205±0.034
UUO组	1.284±0.060^a	0.917±0.069^a	0.902±0.078^a	1.007±0.077^a
UUO+MCL组	0.454±0.021^ab	0.281±0.046^ab	0.491±0.056^ab	0.201±0.038^ab
F	947.521^＊＊	410.538^＊＊	193.414^＊＊	454.035^＊＊

Fig.6 The expression levels of NLRP3, caspase-1, IL-1β and TNF-α proteins in renal tissue of each group detected by Western blot assay

Tab.2 Comparison of expression levels of NLRP3, caspase-1, IL-1β and TNF-α proteins in renal tissue between the three groups

组别	NLRP3	caspase-1	IL-1β	TNF-α
Sham组	0.422±0.091	0.155±0.044	0.367±0.095	0.401±0.118
UUO组	0.840±0.138^a	0.722±0.123^a	0.792±0.107^a	1.012±0.152^a
UUO+MCL组	0.470±0.110^b	0.462±0.093^ab	0.573±0.066^ab	0.646±0.156^ab
F	23.891^＊＊	56.410^＊＊	32.841^＊＊	27.745^＊＊

Fig.7 Immunohistochemical results of NF-κB p65 in renal tissue(×400)

Fig.8 The expression of p-NF-κB p65 protein in kidney tissue of each group detected by Western blot assay

References 17

[1]	CHEN T K, KNICELY D H, GRAMS M E. Chronic kidney disease diagnosis and management:A review[J]. JAMA, 2019, 322(13):1294-1304. doi:10.1001/jama.2019.14745.
[2]	GBD Chronic Kidney Disease Collaboration. Global,regional,and national burden of chronic kidney disease,1990-2017:A systematic analysis for the Global Burden of Disease Study 2017[J]. Lancet, 2020, 395(10225):709-733. doi:10.1016/S0140-6736(20)30045-3.
[3]	YUAN Q, TANG B, ZHANG C. Signaling pathways of chronic kidney diseases, implications for therapeutics[J]. Signal Transduct Target Ther, 2022, 7(1):182. doi:10.1038/s41392-022-01036-5.
[4]	LEI X, LI S, LUO C, et al. Micheliolide attenuates lipopolysaccharide-induced inflammation by modulating the mROS/NF-kappaB/NLRP3 axis in renal tubular epithelial cells[J]. Mediators Inflammation, 2020, 2020:3934769. doi:10.1155/2020/3934769.
[5]	TANAKA S, ZHENG S, KHAREL Y, et al. Sphingosine 1-phosphate signaling in perivascular cells enhances inflammation and fibrosis in the kidney[J]. Sci Transl Med, 2022, 14(658):eabj2681. doi:10.1126/scitranslmed.abj2681.
[6]	ZHOU R, LIAO J, CAI D, et al. Nupr1 mediates renal fibrosis via activating fibroblast and promoting epithelial-mesenchymal transition[J]. FASEB J, 2021, 35(3):e21381. doi:10.1096/fj.202000926RR.
[7]	PUTHUMANA J, THIESSEN-PHILBROOK H, XU L, et al. Biomarkers of inflammation and repair in kidney disease progression[J]. J Clin Invest, 2021, 131(3):e139927. doi:10.1172/JCI139927.
[8]	YIU W H, LIN M, TANG S C. Toll-like receptor activation:from renal inflammation to fibrosis[J]. Kidney Int Suppl(2011),2014, 4(1):20-25. doi:10.1038/kisup.2014.5.
[9]	JIN J, LI W, WANG T, et al. Loss of proximal tubular sirtuin 6 aggravates unilateral ureteral obstruction-induced tubulointerstitial inflammation and fibrosis by regulation of β-catenin acetylation[J]. Cells, 2022, 11(9):1477. doi:10.3390/cells11091477.
[10]	FU H, GU Y H, TAN J, et al. CircACTR2 in macrophages promotes renal fibrosis by activating macrophage inflammation and epithelial-mesenchymal transition of renal tubular epithelial cells[J]. Cell Mol Life Sci, 2022, 79(5):253. doi:10.1007/s00018-022-04247-9.
[11]	BAUERNFEIND F G, HORVATH G, STUTZ A, et al. Cutting edge:NF-kappaB activating pattern recognition and cytokine receptors license NLRP3 inflammasome activation by regulating NLRP3 expression[J]. J Immunol, 2009, 183(2):787-791. doi:10.4049/jimmunol.0901363.
[12]	VILAYSANE A, CHUN J, AMONE M E, et al. The NLRP3 inflammasome promotes renal inflammation and contributes to CKD[J]. J Am Soc Nephrol, 2010, 21(10):1732-1744. doi:10.1681/ASN.2010020143.
[13]	SCHRODER K, ZHOU R, TSCHOPP J. The NLRP3 inflammasome:A sensor for metabolic danger?[J]. Science, 2010, 327(5963):296-300. doi:10.1126/science.1184003.
[14]	LI J, LI S, GUO J, et al. Natural product micheliolide (MCL) irreversibly activates pyruvate kinase M2 and suppresses leukemia[J]. J Med Chem, 2018, 61(9):4155-4164. doi:10.1021/acs.jmedchem.8b00241.
[15]	TONG L, LI J, LI Q, et al. ACT001 reduces the expression of PD-L1 by inhibiting the phosphorylation of STAT3 in glioblastoma[J]. Theranostics, 2020, 10(13):5943-5956. doi:10.7150/thno.41498.
[16]	LI S, PENG F, GONG W, et al. Dimethylaminomicheliolide ameliorates peritoneal fibrosis through the activation of autophagy[J]. J Mol Med(Berl), 2019, 97(5):659-674. doi:10.1007/s00109-019-01757-1.
[17]	LIU W, CHEN X, WANG Y, et al. Micheliolide ameliorates diabetic kidney disease by inhibiting Mtdh-mediated renal inflammation in type 2 diabetic db/db mice[J]. Pharmacol Res, 2019, 150:104506. doi:10.1016/j.phrs.2019.104506.