| ObjectiveAbdominal obesity or overweight,abnormal lipid metabolism,hypertension,diabetes,insulin resistance and/or impaired glucose tolerance are important components of the metabolic syndrome.Obesity is an important risk factor for cardiovascular and cerebrovascular diseases.Lysine-specific demethylase 1(Lsdl)is the first histone demethylase identified in epigenetics of gene activation and inhibition.It plays an important role in regulation.In the first part of the experiment,Lsd1 gene-specific knockout mice were fed with high-fat diet to discuss the effect of cardiac cardiomyocyte Lsdl-specific knockout on obesity-related cardiomyopathy and its mechanism.Hypertension and obesity are closely linked and they share a common pathological basis.Hypertension can cause complications of target organs such as heart and kidney,especially in renal fibrosis.The second part of the experiment explored the intervention effect of Qian Yang Yu Yin Granules on renal fibrosis in spontaneously hypertensive rats by improving the fibrosis.MethodExperiment one1)Observing the the expression level of Lsdl in mice heart tissue which fed with high-fat diet;2)Determination of mitochondrial function in Lsdl gene-specific knockout(Lsdl-cKO)mice fed with high-fat diet;3)Heart tissue morphology(HE staining),degree of fibrosis(Masson staining),heart failure and fibrosis-related gene expression levels in Lsdl-cKO mice fed with high-fat diet;4)Doxorubicin stimulated the heart tissue morphology(HE staining),degree of fibrosis(Masson staining),mitochondrial function,heart failure and fibrosis-related gene expression levels in Lsdl-cKO mice fed with high-fat diet;5)Lsdl-shRNA transfection of neonatal primary cardiomyocytes mitochondrial function gene expression levels;Doxorubicin stimulated Lsdl-shRNA transfection of neonatal primary cardiomyocytes heart failure and fibrosis related gene expression levels.Experiment twoWKY rats were used as normal control group.SHR were randomly divided into 3 groups:model group(SHR),valsartan group(SHR+Valsartan)and QYYYG group(SHR+QYYY).Intragastric gavaged,once a day for 8 weeks.The following tests were performed on rat blood,urine,and kidney tissues.1)Determination of rat blood pressure test,rat early renal damage marker urine mALB,al-MG,β2-MG content;2)Determination of rat kidney weight,kidney index,rat kidney HE staining,Masson staining;3)Determination of urine neutrophil gelatinase-associated lipocalin(NGAL),kidney injury molecule-1(KIM-1)and interleukin-18(IL-1 8)in rat renal tubular injury markers;4)Determination of serum laminin,hyaluronidase,type Ⅲ procollagen and type IV collagen in rats with renal fibrosis;5)Network pharmacology predicts that the molecular mechanism of QYYYG improves hypertensive renal fibrosis;6)Real-time quantitative PCR verifies the prediction results;ResultsExperiment one1)The expression level of Lsd1 in heart tissue of mice led with high fat diet was decreased.2)The specific Lsdl gene knockout in mouse cardiomyocytes did not affect the body weight.heart weight and tibia length ratio,glucose and insulin tolerance.3)Lsd1-cKO mice were fed with high-fat diet、and the total RNA of heart tissue was extracted for RNA sequencing analysis.The differential gene was down-regulated in GO function clustering,which was mainly related to mitochondrial function and mitochondrial structure.Up-regulated genes are most involved in cell adhesion and intercellular adhesion.4)Decreased mitochondrial function gene expression level in heart tissue of Lsd1-cKO mice.5)The morphology of heart tissue of Lsdl-cKO mice fed with higlh-fat diet improved compared with the control group,the degree of cardiac fibrosis decreased,and cardiac function improved.The TGF-β1/Smads signaling pathway and downstream molecular activation were inhibited in Lsd1-cKO mouse heart tissue,and the expression level of heart failure-related genes was decreased.6)Doxorubicin stimulated Lsd-cKO mice fed with high-fat diet,the morphology of Lsdl-cKO mice heart tissue was improved compared with the control group,the degree of cardiac fibrosis was decreased,and the expression levels of genes related to fibrosis and heart failure were decreased.7)Lsd I-shRNA was transfected into primary cardiomyocytes of neonatal mice,and the expression lcvel of mitochondrial function-related genes and TGF-β1 was decreased.8)Doxorubicin stimulated Lsdl-shRNA transfection of primary cardiomyocytes in neonatal mice,and the expression levels of genes related to fibrosis and heart failure decreased compared with the control group.Experiment two1)QYYYG can reduce the levels of SBP,DBP and MAP in SI-HR.2)QYYYG can reduce the ccontent of urine mALB,al-MG andβ2-MG in SHR.3)There was no significant effect on the weight and body weight of rat kidney during the administration of QYYYG.4)QYYYG improves the pathological condition of the kidney by reducing the accumulation of inflammatory cells in the renal interstitium,reduces the deposition of collagen fibers,and reduces the degree of renal fibrosis.5)QYYYG can reduce the levels of urinary NGA L,KIM-1 and IL-18 in rats.6)QYYYG can reduce the serum Laminnin,HAase.Precollagen III and Collagen IV levels in SHR.7)Prediction of TGF-p signaling pathway by network pharmacology is a specific signaling path way associated with QYYYG in the treatment of hypertensive renal fibrosis.QYYYG can reduce the TGF-β1,a-Sma,Col IV and FN gene expression level in kidney tissue of SHR and increased the gene expression level of E-cadherin.ConclusionExperiment oneThe specific Lsdl gene knockout in mice cardiomyocytes can decrease expression of mitochondrial functional genes.However,Lsdl-cKO mice were paradoxically protected from high fat diet and doxorubicin-induced cardiomyopathy and fibrosis.Lsdl deletion in cardiomyocytes reduced the expression of TGF-β1 and the downstream fibrotic genes.These results suggest that Lsdl is a major factor reprograming cardiac gene expression and function.LSD1 may become a new target for intervention in obesity-related cardiomyopathy.Loss of Lsdl also shows protective effects on cardiomyocytes in the myocardial damage caused by doxorubicin,and proposes a new direction for the future use of drugs to guide the identification of myocardial damage.Experiment twoQYYYG can effectively reduce blood pressure,reduce microalbuminuria,repair renal tubular damage,and reduce the degree of renal fibrosis.It may inhibit the expression of TGF-β1,reduce the expression of stroimal cells,increase the phenotypic expression of epithelial cells,inhibit the activation of myofibroblasts,thereby reduce the secretion of collagen fibers and fibronectin,and reverse the EMT process to reduce the occurrence of hypertensive renal fibrosis. |
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