Molecular mechanism of N-acetylcysteine alleviating bladder fibrosis in mice with metabolic syndrome

【Objective】 To explore the molecular mechanism by which N-acetylcysteine （NAC） alleviated bladder fi brosis in metabolic syndrome mice,which could provide theoretical reference for a deeper understanding of the pathologi cal process of bladder fibrosis and the development of new treatment strategies.【Method】 C57BL/6 male mice and ob/ob（B6.V-Lepob/J） male mice （metabolic syndrome model） at 8 weeks of age were selected.The experiment lasted for 20weeks.After the end of the experiment,the mice were killed and the bladder tissues were collected.C57BL/6 male mice were used as the control group （Con）,and ob/ob （B6.V-Lepob/J） male mice were used as the metabolic syndrome model group （ob/ob）.The relative expression levels of TGF-β1,SMAD,p38,p-ERK,p-JNK,α-SMA,3-Nitrotyrosine （NT） and other proteins in the bladder tissues of mice were detected by Western blotting.Immunohistochemical staining was used to detect the expression and distribution characteristics of NF-κB and TGF-β1 in the bladder tissues of mice.Mouse bladder smooth muscle cells （BSMCs） were isolated,the glucose concentration was screened and the growth curve of BSMCs was drawn.The control group （Con）,high glucose group （HG）,high glucose+NAC group （HG+NAC） and hyperos‐motic group （Ho） were set up to detect the relative expression levels of CRP,IL-6,IL-1β,NLRP3,TGF-β1,BK-β1 and SK_Ca3 proteins.MCC950 and PDTC were used to inhibit the NLRP3 and NF-κB signaling pathways,and the relative ex‐pression levels of ROCK1,NLRP3,NF-κB and TGF-β1 proteins were detected. 【Result】 Compared with the Con group,the relative expression of NT,TGF-β1 and α-SMA proteins in the bladder tissue of the ob/ob group mice was signifi‐cantly increased （P＜0.05,the same below）,and there was no significant difference in the relative expression of SMAD,p38,p-ERK and p-JNK proteins （P＞0.05,the same below）.The results of glucose concentration screening showed that 45mmol/L glucose was the optimal intervention concentration.Western blotting results showed that compared with the Con group,the relative expression of CRP,IL-6,TGF-β1,IL-1β and NLRP3 proteins in BSMCs of the HG group was signifi‐cantly increased,and the relative expression of BK-β1 and SK_Ca3 proteins was significantly decreased.Compared with the HG group,the relative expression of CRP,IL-6,TGF-β1,IL-1β and NLRP3 proteins in BSMCs of the HG+NAC group was significantly decreased,and the relative expression of BK-β1 and SK_Ca3 proteins was significantly increased.After in‐hibiting the NLRP3 and NF-κB signaling pathways,compared with the HG group,the relative expression of TGF-β1 pro‐tein in BSMCs of HG+MCC950 and HG+PTCD groups was significantly decreased,and the relative expression of NF-κB protein was significantly increased;the relative expression of ROCK1 and NLRP3 proteins in BSMCs of HG+MCC950 group was significantly decreased,and there was no significant difference in the relative expression of ROCK1and NLRP3 proteins in BSMCs of HG+PTCD group.【Conclusion】 High glucose-induced bladder function damage in mice involves molecular mechanism pathways related to oxidative stress-inflammatory response-tissue fibrosis.NAC mainly blocks the development of oxidative stress,inflammatory response and fibrosis in functional damage by inhibiting the ROS/NLRP3/NF-κB/TGF-β1 signaling pathway.Compared with anti-inflammatory treatment,NAC antioxidant treat‐ment may have a more significant therapeutic effect on tissue fibrosis.