Basic And Clinical Research On Familial Hypercholesterolemia And The Application Of Liver-targeted Nanolipid-lowering Material

Background:Familial hypercholesterolemia(FH),caused primarily by loss-of-function mutations in the low-density lipoprotein receptor(LDLR),is a life-threatening inherited metabolic disorder.More novel mutation sites causing FH were identified in the non-coding region of LDLR.Method:Based on a familial hypercholesterolemic individual(LDLR c.1988-8T>A),we constructed a novel humanized mouse model used to study the pathology and therapeutic evaluation of FH.However,since this locus is located at the intron-exon junction and the base sequences of human and mouse origin are not conserved,we performed partial humanized gene sequence replacement in the region of this locus(exon 13-exon 14)and introduced humanized point mutations,and finally constructed LDLR gene partially humanized wild-type as well as point mutant mice model.Results:Only part of the LDLR fragment sequence was humanized without changing the normal promoter region of LDLR in mice.Therefore,the normal function of LDLR was not affected in LDLR gene partially humanized wild-type mice model.Fortunately,LDLR gene partially humanized point mutant mice model developed hypercholesterolemia with an FH phenotype and did not respond to PCSK9 inhibitors treatment.Further analysis of the humanized point mutant mice model revealed that the point mutation resulted in a class Ⅱ mutation in the LDLR gene,impaired formation of LDLR mature protein,and ultimately inability of the receptor to translocate to the cell membrane.Conclusion:LDLR c.1988-8 T>A is a splice site mutation,resulting in a class Ⅱ mutation in the LDLR gene,which prevents the receptor from translocating to the cell membrane.In addition,the LDLR gene partially humanized mice model is a promising and breakthrough humanized mice as a favorable tool to study the unconserved FH mutation sites.Background:Familial hypercholesterolemia(FH)is an inherited metabolic disorder wi th a high level of low-density lipoprotein cholesterol and the worse prognosis.The triglyceride-glucose(TyG)index,an emerging tool to reflect insulin resistance(IR),is positively associated with a higher risk of atherosclerotic cardiovascular disease(ASCVD)in healthy individuals,but the value of TyG index has never been evaluated in FH patients.This study aimed to determine the association between the TyG index and glucose metabolic indicators,IR status,the risk of ASCVD and mortality among FH patients.Methods:Data from National Health and Nutrition Examination Survey(NHANES)1999-2018 were utilized.941 FH individuals with TyG index information were included and categorized into three groups:<8.5,8.59.0,and>9.0.Spearman correlation analysis was used to test the association of TyG index and various established glucose metabolismrelated indicators.Logistic and Cox regression analysis were used to assess the association of TyG index with ASCVD and mortality.The possible nonlinear relationships between TyG index and the all-cause or cardiovascular death were further evaluated on a continuous scale with restricted cubic spline(RCS)curves.Results:TyG index was positively associated with fasting glucose,HbAlc,fasting insulin and the homeostatic model assessment of insulin resistance(HOMA-IR)index(all p<0.001).The risk of ASCVD increased by 74%with every 1 unit increase of TyG index(95%CI:1.15-2.63,p=0.01).During a median follow-up of 114 months,151 all-cause deaths and 57 cardiovascular deaths were recorded.Strong U/J-shaped relations were observed according to the RCS results(p=0.0083 and 0.0046 for all-cause and cardiovascular death).A higher TyG index was independently associated with both all-cause death and cardiovascular death.Results remained similar among FH patients with IR(HOMA-IR ≥2.69).Moreover,addition of TyG index showed helpful discrimination of both survival from all-cause death and cardiovascular death(p<0.05).Conclusion:TyG index was applicable to reflect glucose metabolism status in FH adults,and a high TyG index was an independent risk factor of both ASCVD and mortality.Background:Metabolic diseases such as obesity,fatty liver,dyslipidemia and insulin resistance are the major risk factors for cardiovascular disease.Remarkable progress has been achieved in siRNAbased therapeutic approaches and their use for gene knockdown in mammalian cells.Hepatic inactivation of Surf4 in mice selectively lowers plasma lipids and protects mice from atherosclerosis.Method:Here,we constructed a liver-targeted siRNA delivery system of apolipoprotein E(apoE)peptide and poly(amidoamine)PAMAM/siRNA surf4 complex for metabolic diseases treatment,which was able to overcome the shortcomings such as low cellular uptake,short half-life,and rapid degradation of siRNA-based strategy.After four weeks of high-fat feeding,apoE-/-mice received apoE PAMAM/siRNA surf4 treatment via tail vein injection once a week for a total of six times.Results:ApoE-PAMAM/siRNA surf4 significantly reduced lipid levels and alleviated atherosclerosis.Additionally,body weight,hepatic steatosis,inflammation and fibrosis,and insulin resistance were alleviated in apoE-/-mice treated with apoE-PAMAM/siRNA surf4.RNA sequencing revealed that down-regulated MAPK pathway(ERK1/2 signal)and up-regulated bile acid secretion(Cyp7a1、ABCG5 and ABCG8)were involved in the mechanism.Moreover,no obvious toxic effects on heart,liver,spleen,lung and kidney were observed.Conclusion:Our study demonstrated that apoE-PAMAM/siRNA surf4 could serve as a promising siRNA-based therapy for metabolic diseases.