基于生物信息学对扩张型心肌病与慢性心力衰竭共同生物标志物的预测*

目的：通过生物信息学的方法预测扩张型心肌病（DCM）与慢性心力衰竭（CHF）发病的共同生物标志物, 为临床上2 种疾病的发病及相关性奠定理论基础。方法：从Gene Expression Omnibus（GEO）数据库下载芯片数 据GSE3585,此为DCM和正常对照组原始数据,同时下载芯片数据GSE76701,此为CHF 和对照组原始数据。 通过R软件分析获得DCM和CHF 发病的差异表达基因,并获得2 种疾病发病的共同差异表达基因,进一步对共 同差异表达基因进行GO 和KEGG富集分析,构建差异表达基因的PPI 相互作用网络图,获得扩张型心肌病和心 衰发病的共同关键基因。结果：DCM的差异表达基因有240 个,其中141 个上调基因,99 个下调基因,CHF 的 差异表达基因有654 个,其中355 个上调基因,299 个下调基因。DCM和CHF 共同的差异表达基因有36 个,其中 19 个上调基因,17 个下调基因。GO 分析显示,差异表达基因主要集中在12 种不同的生理、病理过程中,KEGG 分析获得差异表达基因参与的主要信号通路为5 条,预测7 个关键差异表达基因,分别为：CD163、KYVE1、 MRC1、VSIG4、FCER1G、S100A9、F13A1。结论：该研究初步探讨了DCM与CHF 两种疾病发病分子机制, 获得了两种疾病发病的共同差异表达基因,仍需进一步的实验研究对基因的表达和临床病理特征的相关性进行验 证。     

Spock1参与Adipogenesis通路在肥大型与扩张型心肌病中的机制探讨

目的 探讨SPARC(骨素)、cwcv和Kazal样结构域蛋白多糖1(Spock1)在肥大型与扩张型心肌病发生中的作用与分子机制。方法 基于CNKI数据库明确Spock1在机体各组织中的表达情况,并获得Spock1参与信号通路的关联基因,对此关联基因进行GO和KEGG富集分析,对比同类基因与心脏疾病的关系,从而分析获得Spock1参与肥大型与扩张型心肌病的具体关联信息。结果 Spock1在心脏中较高表达,并且其可能通过Adipogenesis信号通路参与肥大型心肌病(HCM)和扩张型心肌病(DCM)的发生与发展。结论 Spock1可能与肥大型和扩张型心肌病等相关心血管疾病的发生密切相关,并可能成为心脏疾病发生发展的生物标记物,甚至可成为治疗心脏疾病的新靶点基因。     

基于生物信息学和机器学习识别扩张性心肌病特征基因与免疫细胞浸润

目的 通过生物信息学方法确定扩张性心肌病(DCM)特征基因及免疫细胞浸润。方法 在两个DCM基因表达数据集上鉴定差异表达基因(DEG)并进行基因本体论(GO)、疾病本体论(DO)和基因集富集分析(GSEA)功能富集以获得潜在途径。两种机器学习算法，包括支持向量机递归特征消除(SVM-RFE)以及最小绝对收缩和选择算子(LASSO)用于确定DCM特征基因。最后运用细胞类型分析工具CIBERSORT进行免疫细胞浸润分析。结果 共鉴定出51个DEG,其中机器学习算法识别硫氧还蛋白相互作用蛋白(TXNIP)、晶状体蛋白Mu(CRYM)、类热休克蛋白70蛋白A1(HSPA1L)、真核翻译延伸因子1A1(EEF1A1)为特征基因。富集分析集中在心脏过程、线粒体和细胞器的外膜、泛素样蛋白连接酶、自然杀伤细胞介导的细胞毒性，1型辅助T(Th1)细胞和Th2细胞分化，T细胞受体信号转导途径和Th17细胞分化方面。免疫细胞浸润发现幼稚B细胞，中性粒细胞和γT细胞可能参与DCM的发病过程。此外，中性粒细胞、Th细胞和M1巨噬细胞与4个特征基因高度相关。结论机器学习识别的4个特征基因TXNIP、 CRYM、 ...     

相关转录组分析鉴定色素沉着绒毛结节性滑膜炎的潜在生物标志物

背景：色素沉着绒毛结节性滑膜炎是一种罕见的滑膜炎性病变,早期诊断缺乏相关生物标志物,m RNA已被证实参与了疾病的发生发展,但其作用机制尚不清楚。目的：通过生物信息学及相关转录组分析鉴定色素沉着绒毛结节性滑膜炎的潜在生物标志物和发病机制,以用于疾病鉴别诊断。方法：通过GEO数据库检索与色素沉着绒毛结节性滑膜炎相关的滑膜组织微阵列数据集,并用Network Analyst分析鉴定差异表达基因(P <0.05)。使用疾病数据库检索与色素沉着绒毛结节性滑膜炎相关的基因,并与差异表达m RNA取交集得到最终差异表达基因。利用Bio GPS对特异性基因进行组织/器官定位。通过STRING数据库构建蛋白质-蛋白质相互作用网络,KOBAS 3.0和GSEA 4.1.0对差异表达基因进行富集分析,并运用多重计算方法鉴定核心基因。利用Cytoscape构建竞争性内源性RNA(Ce RNA)网络。GEO数据集验证了具有较高诊断价值的生物标志物。此外,通过Xcell网站分析64种免疫细胞和基质细胞的浓度,并计算丰度分数。结果与结论：在GSE175626数据集中共鉴别出2 546个差异表达基因,包括2 ...     

MicroRNA-146在糖尿病心肌病中的调控作用

目的:探讨微小RNA-146 (miR-146)在糖尿病心肌病(DCM)发病机制中的作用,观察miR-146及其下游靶基因表达水平的变化。方法:60只雄性C57BL/6小鼠随机分为实验(DCM)组和对照(control)组,每组30只,实验组采用低剂量(50 mg/kg链脲佐菌素(STZ)腹腔注射诱导建立糖尿病心肌病模型,对照组给予等量枸橼酸钠缓冲液腹腔注射,建模12周后取心脏做HE和Masson染色观察心脏病理改变,RT-qPCR检测miR-146 a和miR-146b及其下游靶基因白细胞介素-1受体相关激酶1(IRAK1)和TNF受体相关因子6(TRAF6)的mRNA表达,Western blot检测IRAK1和TRAF6的蛋白表达。结果:12周末HE染色显示,DCM组心肌肥大,结构紊乱;Masson染色显示,DCM组心肌内胶原纤维增多;RT-qPCR检测结果显示,DCM组中miR-146a及miR-146b较control组表达明显减少(P 0. 01),IRAK1的mRNA水平明显增高(P 0. 01),而TRAF6的mRNA水平下降(P 0. 01);Western blot检测结果显示DCM组的IRAK1蛋白表达增加,TRAF6的蛋白表达降低(P 0. 01)。结论:miR-146可能通过调节IRAK1介导的炎症反应参与糖尿病心肌损伤的发生发展。     

3-硝基酪氨酸与糖尿病心肌病大鼠心肌细胞凋亡的关系研究

目的：探讨2型糖尿病大鼠心肌组织及血中3-硝基酪氨酸（3-NT）与糖尿病心肌病（DCM）心肌细胞凋亡的关系。方法：选取8周龄雄性SD大鼠60只,将其随机分为4组,空白对照组、DCM组、糖尿病+缬沙坦处理组和DCM+缬沙坦处理组,每组15只。用TUNEL法检测DCM心肌细胞的凋亡指数（AI）;用免疫组化的方法检测心肌组织中3-NT的表达指数（EI）;用ELISA法检测血清中3-NT的浓度。结果：(1) 4组大鼠的心脏重量指数有显著差异（P<0.01）,DCM组和DCM+缬沙坦处理组心脏重量指数大于空白对照组和糖尿病+缬沙坦处理组。(2) 心肌组织免疫组化方法检测表明,心肌组织中3-NT的EI与心肌细胞的AI呈正相关（P<0.01）,而血中3-NT的含量与心肌细胞的AI无相关关系（P>0.05）。(3) 4组大鼠间的AI比较有显著差异（P<0.01）。两两比较各组AI,DCM组 > 糖尿病+缬沙坦预处理组、DCM+缬沙坦处理组 > 空白对照组。(4) 4组大鼠心肌组织3-NT表达指数比较有显著差异（P<0.01）;两两比较,DCM组显著大于其它3组。(5) 4组大鼠血中3-NT的浓度比较无显著差异（P>0.05）。结论：(1) DCM大鼠心肌组织中3-NT的表达显著增多并与心肌细胞凋亡密切相关,缬沙坦能够抑制DCM大鼠心肌细胞中3-NT的表达,并由此抑制DCM大鼠心肌细胞凋亡。(2) 循环血中的3-NT水平不能真实反映DCM大鼠心肌组织中3-NT表达水平及其对心肌细胞凋亡的影响。     

基于通路及网络分析方法探究肝细胞肝癌差异表达基因

目的基于通路及网络分析方法,对肝细胞肝癌(HCC)相关差异表达基因进行系统分析,旨在由分子、通路及网络层面理解HCC发生、发展机制,并对HCC的防治靶点进行预测,为后续实验提供一些有意义的信息。方法由公共基因表达数据库(GEO)下载4组与HCC相关的生物基因芯片(GSE62232、GSE49515、GSE19665和GSE29721),利用相应R包对基因表达阵列的差异表达基因进行计算与比较,确定HCC相关差异表达基因。利用功能富集分析方法 ,对HCC相关差异表达基因进行生物过程及通路注释。利用网络分析方法,对HCC相关差异表达基因进行生物调控网络构建,并对关键基因进行筛选,同时利用分子复合物检测(MCODE)方法对HCC相关疾病模块进行搜索。结果确定442个HCC相关差异表达基因,这些差异表达基因主要调控细胞周期、核分裂、染色体分离及代谢相关生物过程;参与影响细胞周期通路、有丝分裂过程通路、生物氧化通路、金属离子反应通路、生物代谢通路等多条信号通路的正常信号转导。网络分析提示,HCC相关差异表达基因之间互作关系紧密,进一步分析鉴定出20个处于HCC调控网络中心的关键Hub基因,推测Hub基因是HCC防治靶点。同时,鉴定出5个与HCC紧密相关的疾病模块,分别为细胞周期处理模块、趋化因子介导的信号处理模块、血管生成调控模块、与氧化应激和代谢过程相关功能模块及细胞核分裂模块。结论利用生物信息学方法,可以系统分析HCC的发生、发展机制,并对其防治靶点进行预测,为临床诊疗及后续基础实验提供有价值的信息。     

扩张型心肌病与缺血性心肌病二维彩色多普勒的对比研究

利用二维彩色多普勒在心脏形态学、血液动力学及心室功能方面对扩张型心肌病 (DCM )及缺血性心肌病 (ICM )进行诊断及鉴别诊断。利用二维彩色多普勒技术分别测定 5 2例DCM组、ICM组及正常对照组的数据 ,利用定量和半定量的方法对数据进行测定 ,用统计学对数据进行处理。结果表明正常组分别与DCM组及ICM组比较有显著的差异 (P <0 0 5或0 0 1) ,ICM组与DCM组有比较显著的差异 (P <0 0 1)。DCM组主要是心肌弥漫性改变 ,心腔普遍扩大 ,心脏收缩功能明显减退 ,舒张功能为“正常”。多瓣膜返流发生率高且严重。ICM组以心肌局部病变为主 ,主要是左心腔扩大 ,左室收缩功能正常或轻度减低 ,舒张功能明显减低。瓣膜多为单瓣膜返流且较轻。结论 :二维彩色多普勒是临床鉴别诊断DCM和ICM最可靠、最简单及无创伤的首选方法。     

肠道病毒感染致扩张型心肌病发生的机制

肠道病毒持续感染是扩张型心肌病(dilated cardiomyopathy,DCM)发生的重要病因,然而病毒感染后导致DCM发生的机制至今尚不明了.研究表明:肠道病毒一方面可以在心肌组织中持续复制,进而通过自身蛋白酶直接损伤心肌细胞,或是通过机体的免疫系统对心肌产生间接的损伤;另一方面,病毒也可以一种自然变异的形式在心肌中长期潜伏.     

家族性肥厚型心肌病与肌钙蛋白T

家族性肥厚型心肌病(FHCM)是一种常染色体显性遗传病, 由于编码心肌蛋白的基因突变引起,目前已识别出至少13个不同致病基因的200余种突变。目前,阐明FHCM的分子遗传学机制已经成为当前研究的热点之一。肌钙蛋白T通过与原肌宁蛋白结合,在将肌钙蛋白复合体锚钉到细肌丝上起重要作用。肌钙蛋白T基因突变是导致家族性肥厚型心肌病的主要原因,至今已经发现了大约30个突变,约占所有突变的15~20%。肌钙蛋白T基因突变所致FHCM有两个主要特征：（1）心肌肥厚程度较轻, 疾病外显率差别较大,（2）猝死率高。目前所发现的致FHCM突变,主要集中在肌钙蛋白T的T1和T2结构域。对肌钙蛋白T突变致FHCM分子机制的研究将有助于肥厚型心肌病的基因诊断和临床治疗。     

Cardiac troponin T mutation in familial cardiomyopathy with variable remodeling and restrictive physiology

We identified a unique family with autosomal dominant heart disease variably expressed as restrictive cardiomyopathy (RCM), hypertrophic cardiomyopathy (HCM), and dilated cardiomyopathy (DCM), and sought to identify the molecular defect that triggered divergent remodeling pathways. Polymorphic DNA markers for nine sarcomeric genes for DCM and/or HCM were tested for segregation with disease. Linkage to eight genes was excluded, but a cardiac troponin T (TNNT2) marker cosegregated with the disease phenotype. Sequencing of TNNT2 identified a heterozygous missense mutation resulting in an I79N substitution, inherited by all nine affected family members but by none of the six unaffected relatives. Mutation carriers were diagnosed with RCM (n = 2), non-obstructive HCM (n = 3), DCM (n = 2), mixed cardiomyopathy (n = 1), and mild concentric left ventricular hypertrophy (n = 1). Endomyocardial biopsy in the proband revealed non-specific fibrosis, myocyte hypertrophy, and no myofibrillar disarray. Restrictive Doppler filling patterns, atrial enlargement, and pulmonary hypertension were observed among family members regardless of cardiomyopathy subtype. Mutation of a sarcomeric protein gene can cause RCM, HCM, and DCM within the same family, underscoring the necessity of comprehensive morphological and physiological cardiac assessment in familial cardiomyopathy screening.     

Investigation of de novo variation in pediatric cardiomyopathy

Pediatric cardiomyopathies can be caused by variants in genes encoding the sarcomere and cytoskeleton in cardiomyocytes. Variants are typically inherited in an autosomal dominant manner with variable expressivity. De novo variants have been reported, however their overall frequency is largely unknown. We sought to determine the rate of de novo, pathogenic and likely pathogenic (P/LP) variants in children with a diagnosis of hypertrophic, dilated, or restrictive cardiomyopathy (HCM, DCM, or RCM), and to compare disease outcomes between individuals with and without a de novo variant. A retrospective record review identified 126 individuals with HCM (55%), DCM (37%), or RCM (8%) ≤18 years of age who had genetic testing. Overall, 50 (40%) had positive genetic testing and 18% of P/LP variants occurred de novo. The rate of de novo variation in those with RCM (80%) was higher than in those with HCM (9%) or DCM (20%). There was evidence of germline mosaicism in one family with RCM. Individuals with de novo variants were more likely than those without to have a history of arrhythmia (p = .049), sudden cardiac arrest (p = .024), hospitalization (p = .041), and cardiac transplantation (p = .030). The likelihood of de novo variation and impact on family risk and screening should be integrated into genetic counseling.     

Evolution of dilated cardiomyopathy (DCM) from idiopathic hypertrophic cardiomyopathy (IHCM) vs. inflammatory dilated cardiomyopathy (DCMi): a rare case of sudden death in an 8-year-old boy

In rare cases, the diagnosis of hypertrophic and dilated cardiomyopathy (DCM) in children was established postmortem. Our case report deals with the sudden and unexpected death of an 8-year-old boy. The postmortem examination revealed non-obstructive hypertrophy with irregular arrangement of muscular fibers, dilatation of the ventricles, endocardial fibrosis, microfocal vacuolization with enlarged hyperchromatic nuclei, and signs of inflammation with interstitial fibrosis. We present an evolution from idiopathic cardiomyopathy to DCM. To some extent, there were morphologic signs of an inflammatory process that first led us to suspect a specific inflammatory DCM.     

The haplotype block, NFKBIL1-ATP6V1G2-BAT1-MICB-MICA, within the class III-class I boundary region of the human major histocompatibility complex may control susceptibility to hepatitis C virus-associated dilated cardiomyopathy

Cardiomyopathy is a heart muscle disease with impaired stretch response that can result in severe heart failure and sudden death. A small proportion of hepatitis C virus (HCV)-infected patients may be predisposed to develop dilated cardiomyopathy (DCM) and hypertrophic cardiomyopathy (HCM). The molecular mechanisms involved in the predisposition remain unknown due in part to the lack of information on their genetic background. Because the human leukocyte antigen (HLA) region has a pivotal role in controlling the susceptibility to HCV-induced liver disease, we hypothesized that particular HLA alleles and/or non-HLA gene alleles within the human major histocompatibility complex (MHC) genomic region might control the predisposition to HCV-associated DCM (HCV-DCM) and/or HCV-associated HCM (HCV-HCM). Here, we present mapping results of the MHC-related susceptibility gene locus for HCV-associated cardiomyopathy by analyzing microsatellite and single nucleotide polymorphism markers. To delineate the susceptibility locus, we genotyped 44 polymorphic markers scattered across the entire MHC region in a total of 59 patients (21 HCV-DCM and 38 HCV-HCM) and 120 controls. We mapped HCV-DCM susceptibility to a non-HLA gene locus spanning from NFKBIL1 to MICA gene loci within the MHC class III-class I boundary region. Our results showed that HCV-DCM was more strongly associated with alleles of the non-HLA genes rather than the HLA genes themselves. In addition, no significant association was found between the MHC markers and HCV-HCM. This marked difference in the MHC-related disease susceptibility for HCV- associated cardiomyopathy strongly suggests that the development of HCV- DCM and HCV-HCM is under the control of different pathogenic mechanisms.     

The novel αB‐crystallin (CRYAB) mutation p.D109G causes restrictive cardiomyopathy

Restrictive cardiomyopathy (RCM) is a rare heart disease characterized by diastolic dysfunction and atrial enlargement. The genetic etiology of RCM is not completely known. We identified by a next‐generation sequencing panel the novel CRYAB missense mutation c.326A>G, p.D109G in a small family with RCM in combination with skeletal myopathy with an early onset of the disease. CRYAB encodes αB‐crystallin, a member of the small heat shock protein family, which is highly expressed in cardiac and skeletal muscle. In addition to in silico prediction analysis, our structural analysis of explanted myocardial tissue of a mutation carrier as well as in vitro cell transfection experiments revealed abnormal protein aggregation of mutant αB‐crystallin and desmin, supporting the deleterious effect of this novel mutation. In conclusion, CRYAB appears to be a novel RCM gene, which might have relevance for the molecular diagnosis and the genetic counseling of further affected families in the future.     

Inflammation in myocardial disease: From myocarditis to dilated cardiomyopathy

Dilated cardiomyopathy (DCM) is a heterogeneous group of myocardial diseases clinically defined by the presence of left ventricular dilatation and contractile dysfunction. Among various causes of DCM, a progression from viral myocarditis to DCM has long been hypothesized. Supporting this possibility, studies by endomyocardial biopsy, the only method to obtain a definite diagnosis of myocarditis at present, have provided evidence of inflammation in the myocardium in DCM patients. A number of experimental studies have elucidated a cell‐mediated autoimmune mechanism triggered by viral infection in the progression of myocarditis to DCM. In addition, the important role of inflammation in the pathogenesis of heart failure has been recognized, and many terms including myocarditis, inflammatory cardiomyopathy, and inflammatory DCM have been used for myocardial diseases associated with inflammation. This review discusses the pathophysiology of inflammation in the myocardium, and refers to diagnosis and treatment based on these concepts.     

Mutations in PDLIM3 and MYOZ1 encoding myocyte Z line proteins are infrequently found in idiopathic dilated cardiomyopathy

Dilated cardiomyopathy (DCM), characterized by ventricular dilation and decreased systolic function, is estimated to be of genetic origin in up to 50% of cases. In the present study, we investigated the role of two genes, encoding the Z line proteins PDZ and LIM domain protein 3 (PDLIM3) and myozenin-1 (MYOZ1), in the etiology of DCM. The coding regions of PDLIM3 and MYOZ1 were first amplified from the genomic DNA of 185 unrelated DCM patients by polymerase chain reaction (PCR), followed by denaturing high-performance liquid chromatography (DHPLC) analysis. The samples that exhibited abnormal peaks on DHPLC were re-amplified, purified and sequenced using a Big-Dye Terminator cycle sequencing system. Interestingly, a 2-bp insertion (178insCA) in exon 2 of PDLIM3 was identified in one patient who presented with DCM during pregnancy and died a year later awaiting heart transplant. No other significant mutations were found in either PDLIM3 or MYOZ1. The mutation probably resulted in an unstable protein, since no exogenous protein could be detected in transfected murine myoblastoid cells by immunohistochemical or Western blot analyses. We conclude that mutations in PDLIM3 and MYOZ1, encoding myocyte Z line proteins, do not play any significant role in the genetic etiology of idiopathic DCM. The exact mechanism by which the mutation identified in the present study is linked to DCM phenotype remains unknown. The hemodynamic burden of pregnancy and/or other genetic or environmental factors could have precipitated heart failure symptoms in an individual with defective myocardial cytoarchitecture.     

NGS testing for cardiomyopathy: Utility of adding RASopathy‐associated genes

RASopathies include a group of syndromes caused by pathogenic germline variants in RAS‐MAPK pathway genes and typically present with facial dysmorphology, cardiovascular disease, and musculoskeletal anomalies. Recently, variants in RASopathy‐associated genes have been reported in individuals with apparently nonsyndromic cardiomyopathy, suggesting that subtle features may be overlooked. To determine the utility and burden of adding RASopathy‐associated genes to cardiomyopathy panels, we tested 11 RASopathy‐associated genes by next‐generation sequencing (NGS), including NGS‐based copy number variant assessment, in 1,111 individuals referred for genetic testing for hypertrophic cardiomyopathy (HCM) or dilated cardiomyopathy (DCM). Disease‐causing variants were identified in 0.6% (four of 692) of individuals with HCM, including three missense variants in the PTPN11, SOS1, and BRAF genes. Overall, 36 variants of uncertain significance (VUSs) were identified, averaging ～3VUSs/100 cases. This study demonstrates that adding a subset of the RASopathy‐associated genes to cardiomyopathy panels will increase clinical diagnoses without significantly increasing the number of VUSs/case.