1.Study on the anti-pulmonary fibrosis effect of linarin in vivo and in vitro and its mechanism
Liting HUANG ; Zhuqiang WANG ; Yiting WANG ; Weifeng FAN ; Gengting DONG ; Weiwen PENG
China Pharmacy 2023;34(3):333-338
OBJECTIVE To investigate the anti-pulmonary fibrosis effect of linarin in vivo and in vitro, and investigate its mechanism preliminarily. METHODS C57BL/6J mice were randomly divided into normal group (carboxymethylcellulose sodium), model group (carboxymethylcellulose sodium), positive control group (pirfenidone, 200 mg/kg), linarin low-dose and high-dose groups (12.5, 25 mg/kg), with 8 mice in each group. Except for normal group, pulmonary fibrosis model was induced in other groups. After modeling, they were given relevant medicine intragastrically, once a day, for consecutive 14 d. The general situation of mice was observed, and their lung indexes were measured; the levels of tumor necrosis factor-α (TNF-α) and transforming growth factor-β1( TGF-β1) in serum and interleukin-6 (IL-6) in lung tissue were determined. Hematoxylin-eosin (HE) staining and Masson staining were used to observe the histopathological morphology of lung. The pulmonary fibrosis was scored according to Ashcroft score standard. The expressions of α-smooth muscle actin (α-SMA) and (type Ⅰ collagen, Collagen Ⅰ), phosphorylated extracellular signal-regulated kinase (p-ERK1/2) and TGF-β1 in lung tissues were detected. HFL1 cells were stimulated by TGF- β1 to form pulmonary fibrosis model in vitro, which were divided into normal group, model group and linarin low-, medium- and high-concentration groups (3.7, 7.4, 14.8 mg/L). After being cultured for 48 h, the protein expressions of α-SMA, Collagen Ⅰ and p-ERK1/2 in HFL1 cells were detected. RESULTS In vivo, compared with normal group, the lung index of model group and the levels of TNF- α, TGF- β1 and IL-6 were significantly increased (P<0.01). There were a large number of inflammatory infiltration and cellular fibrosis lesions in the alveoli, and a large number of collagen depositions. The scores of HE staining and Masson staining were significantly increased (P<0.01). The protein expressions of α-SMA, Collagen Ⅰ, p-ERK1/2 and TGF-β1 in lung tissue were up-regulated significantly (P<0.01). Compared with model group, above indexes of mice were improved significantly in linarin high-dose group (P<0.05 or P<0.01), and most of indexes (except for lung index) were improved significantly in linarin low-dose group (P<0.05 or P<0.01). In vitro, compared with blank group, the density of cells in the model group increased, and obvious proliferation and other changes occurred; protein expressions of α-SMA, Collagen Ⅰ and p-ERK1/2 were significantly up-regulated (P<0.05 or P<0.01). Compared with model group, the cell density of each concentration group was decreased and the morphology gradually returned to normal; the expressions of above proteins in linarin high-concentration group and the protein expression of p-ERK1/2 in linarin medium-concentration group were down-regulated significantly(P<0.05 or P<0.01). CONCLUSIONS Linarin may regulate ERK and inflammatory pathways to reduce the inflammatory response, thereby exerting anti-pulmonary fibrosis effect.
2.Study on the mechanism of berberine in improving diabetes mellitus type 2 combined with metabolic-associated fatty liver disease
Yi LI ; Shuyu KANG ; Qiwen WANG ; Manting HUANG ; Congyan ZENG ; Jun TONG ; Gengting DONG
China Pharmacy 2025;36(16):1975-1980
OBJECTIVE To investigate the potential mechanism of berberine improving diabetes mellitus type 2 (T2DM) combined with metabolic-associated fatty liver disease (MAFLD) by regulating ceramide. METHODS Thirty-two db/db mice with blood glucose levels>11.1 mmol/L (T2DM model) were divided into four groups: model group, berberine low- and high-dose groups [100, 200 mg/(kg·d)] and metformin group [300 mg/(kg·d)], with 8 mice in each group. Additionally, 8 wt/wt mice were selected as the normal control group. Mice in each group were administered the corresponding drug solution or water by gavage once daily for a continuous period of 6 weeks. During the experiment, the body weight of the mice was monitored, and the differences in final body weight were analyzed. After the last administration, the body shape of the mice in each group was observed, and their fasting blood glucose (FBG) and the lipid indicators [total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C)] were measured. Fasting serum insulin (FINS) levels were also measured, and the insulin resistance index HOMA-IR) and insulin sensitivity index (ISI) were calculated. Liver weight, liver index and serum liver function indicators [alanine transaminase (ALT), aspartate transaminase(AST)] were assessed, and hepatic histopathological changes were observed. Additionally, the expression of fatty acid synthesis-related proteins [sterol regulatory element-binding protein 1 (SREBP1), fatty acid synthase (FASN), acetyl-CoA carboxylase 1 (ACC1)] in liver tissue was examined. Serum samples from the normal control group, model group, and berberine high-dose group were collected for non-targeted lipidomics analysis and validation. RESULTS Compared with the model group, the pathological changes, including disordered liver tissue cell arrangement and lipid vacuoles, were significantly improved in the berberine low- and high-dose groups. The significant decreases or down-regulations were observed in body weight in the last week, as well as FBG, TC, TG, and LDL-C levels, HOMA-IR (except for the berberine low-dose group), liver weight, liver index, AST and ALT levels, and protein expressions of SREBP1, FASN and ACC1. Additionally, HDL-C levels, FINS (except for the berberine high-dose group), and ISI (except for the berberine low-dose group) were significantly increased (P<0.05). A total of 21 potential differential metabolites, including multiple types of ceramides, were identified; these metabolites were primarily enriched in sphingolipid metabolism and glycerophospholipid metabolism pathways. Verification experiments confirmed that high-dose berberine significantly reduced the serum content of ceramide in model mice (P<0.05). CONCLUSIONS Berberine reduces insulin resistance, improves liver damage and lipid accumulation in the T2DM combined with MAFLD mice, and these effects may be related to the reduction of ceramide content.