Dietary silymarin improves performance by altering hepatic lipid metabolism and cecal microbiota function and its metabolites in late laying hens.

BACKGROUND: Liver lipid dysregulation is one of the major factors in the decline of production performance in late-stage laying hens. Silymarin (SIL), a natural flavonolignan extracted from milk thistle, is known for its hepatoprotective and lipid-lowering properties in humans. This study evaluates whether SIL can provide similar benefits to late-stage laying hens. A total of 480 68-week-old Lohmann Pink laying hens were randomly assigned into 5 groups, each group consisting of 6 replicates with 16 hens each. The birds received a basal diet either without silymarin (control) or supplemented with silymarin at concentrations of 250, 500, 750, or 1,000 mg/kg (SIL250, SIL500, SIL750, SIL1000) over a 12-week period. RESULTS: The CON group exhibited a significant decline in laying rates from weeks 9 to 12 compared to the initial 4 weeks (P = 0.042), while SIL supplementation maintained consistent laying rates throughout the study (P > 0.05). Notably, the SIL500 and SIL750 groups showed higher average egg weight than the CON group during weeks 5 to 8 (P = 0.049). The SIL750 group had a significantly higher average daily feed intake across the study period (P < 0.05), and the SIL500 group saw a marked decrease in the feed-to-egg ratio from weeks 5 to 8 (P = 0.003). Furthermore, the SIL500 group demonstrated significant reductions in serum ALT and AST levels (P < 0.05) and a significant decrease in serum triglycerides and total cholesterol at week 12 with increasing doses of SIL (P < 0.05). SIL also positively influenced liver enzyme expression (FASN, ACC, Apo-VLDL II, FXR, and CYP7A1; P < 0.05) and altered the cecal microbiota composition, enhancing species linked to secondary bile acid synthesis. Targeted metabolomics identified 9 metabolites predominantly involved in thiamin metabolism that were significantly different in the SIL groups (P < 0.05). CONCLUSIONS: Our study demonstrated that dietary SIL supplementation could ameliorate egg production rate in late stage laying hens, mechanistically, this effect was via improving hepatic lipid metabolism and cecal microbiota function to achieve. Revealed the potentially of SIL as a feed supplementation to regulate hepatic lipid metabolism dysregulation. Overall, dietary 500 mg/kg SIL had the best effects.

The role of TRPC6 in α1-AR activation-induced calcium signal changes in human podocytes.

BACKGROUND: An accumulating amount of evidence has suggested that there is a contributive role of sympathetic nervous hyperactivity in the pathogenesis of chronic kidney disease (CKD). α1-AR promotes an increase in calcium levels in podocytes and adjusts podocyte contraction. Changes in TRPC6 expression and function can directly affect the podocyte cytoskeleton, which is a key component in podocyte injury. This study proposed to clarify the correlation between α1-AR activation-induced signal cascade reaction and TRPC6 in human podocytes. METHODS: Human podocytes were incubated with the calcium probe Fluo-3/AM. Next, the effects of the α1-AR agonists or antagonists and nonselective TRPC6 blockers on intracellular calcium were observed under laser confocal microscopy. FITC-phalloidin was employed to stain podocytes, and the change of F-actin under the α1-AR activation condition was observed. RESULTS: The α1-AR agonist PE (phenylephrine hydrochloride) induced an increase in intracellular Ca2+ ([Ca2+]i) in human podocytes. Moreover, the downregulation of TRPC6 by siRNA or TRPC blocker could attenuate the PE-induced [Ca2+]i elevation in a phospholipase C (PLC)-dependent pattern. When podocytes were stimulated to the PE, their F-actin fiber cytoskeletal structure was lost. PE subsequently increased the expression of RhoA, and the TRPC6-dependent Ca2+ influx was involved in this process. The abnormal activation of RhoA could result in disturbance of the podocyte skeleton structure, thus leading to podocyte injury. CONCLUSIONS: We concluded that TRPC6 is involved in α1-AR activation-induced calcium signal changes in podocytes. Meanwhile, the α1-AR agonists can destroy the cell's cytoskeletal structure, which is mediated by TRPC6 via the RhoA/ROCK pathway.

Suppression of miR-30a-3p Attenuates Hepatic Steatosis in Non-alcoholic Fatty Liver Disease.

Non-alcoholic fatty liver disease (NAFLD) have a high prevalence in humans in the past two decades. Here, we elucidated the functions of miR-30a-3p in the development of NAFLD and identified its potential targets. HepG-2 cells and NAFLD patients' blood samples were used in our study. Bioinformatics analysis as well as luciferase reporter assays were employed to distinguish peroxisome proliferator-activated receptor alpha (PPAR-α) as a target gene. Western blotting showed the expressions of lipid metabolic proteins and the target gene PPAR-α. Oil red O staining and triglyceride activity tested the fatty deposits after treatment with miR-30a-3p. miR-30a-3p was substantially up-regulated in NAFLD. Bioinformatics analyses showed that PPAR-α was a possible target of miR-30a-3p, linked with signaling pathways in NAFLD. PPAR-α as a novel target of miR-30a-3p, and suppression of its levels. The lipid metabolic-related proteins ACC, p-GSK-3ß and FASN were up-regulated after transfecting with miR-30a-3p mimic, but the proteins CPT1, p-AMPK and UCP2 were down-regulated. miR-30a-3p inhibitor could diminish the protein manifestation of ACC, p-GSK-3ß and FASN; and augment the protein manifestation of CPT1, p-AMPK and UCP2. On the contrary, overexpression of miR-30a-3p had adverse impacts on the performance of hepatocellular lipid accumulation and Triglyceride (TG) activity. Co-treatment with miR-30a-3p mimic and overexpression PPAR-α could revise the hepatic steatosis and the TG level induced by fat milk. Our findings suggest that miR-30a-3p/PPAR-α may be developed as a potential agent in NAFLD, which is enough to attenuate triglyceride accumulation and hepatic steatosis.

Group I metabotropic glutamate receptor activation induces TRPC6-dependent calcium influx and RhoA activation in cultured human kidney podocytes.

Researches have shown that mice lacking the metabotropic glutamate receptor 1 (mGluR) showed albuminuria, remodeling of F-actin, with loss of stress fibers. Selective group I mGluRs agonist (S)-3,5-dihydroxyphenylglycine (DHPG) attenuated albuminuria in several rodent models of nephrotic syndrome. However, the molecular mechanism is obscure. Using a human podocyte cell line, we here investigated the molecular mechanisms of group I mGluRs-induced calcium influx and the formation of stress fibers. Our data showed that group I mGluRs activation by DHPG induced a significant calcium influx, and promoted cytoskeletal stress fiber formation and focal adhesions in podocytes. Pre-incubating podocytes with non-selective inhibitor of transient receptor potential channels (TRPC), or the knockdown of TRPC6 attenuated the calcium influx and the stress fiber formation induced by DHPG. Further, DHPG resulted in an increase of active RhoA expression. However, the knockdown of RhoA by siRNA abolished the DHPG-induced increase in stress fibers. Additionally, nonselective inhibitors of TRPC or TRPC6 knockdown clearly inhibited RhoA activation induced by DHPG, as assessed by Glutathione-S-transferase pull-down assay followed by Western blotting. Taken together, our findings suggest TRPC6 regulates actin stress fiber formation and focal adhesions via the RhoA pathway in response to group I mGluRs activation. Our data can potentially explain the mechanism of protective action of group I mGluRs in glomerular podocyte injury.