Background: Anthracycline-induced cardiotoxicity (ACT) occurs in 57% of treated cancer patients. Effects of genetic variations to individual susceptibility to ACT is emerging. However, diverse lifestyle and environmental factors combined with small sample sizes in human cohorts remain as limitations to effectively clarify the genetic susceptibility to ACT. This study employed a murine genetic reference population (GRP) of BXD mice derived from crosses between DBA/2J and C57BL/6J strains for mapping of quantitative trait loci (QTLs) associated with doxorubicin (DOX)-induced traits using systems genetics methods. Methods: To model the variability in ACT to DOX, 56 BXD strains (N≥3 mice/sex, 3-4 month-old) were injected intraperitonially once with DOX (20 mg/kg). Survival and body weight (BW) were monitored daily for 10 Days (D). Echocardiography was performed before and on D5 after DOX. Genetic mapping was performed using WebQTL (www.genenetwork.org) and multi-criteria scoring was used for filtering candidate genes associated with DOX-induced traits. Results: Varied body weight loss, proportions and median survival rates were found among DOX-treated BXDs. Parental C57BL/6J strain had 100% survival, while 75% of DBA/2J mice survived on D10. Among BXDs tested, 47 strains had median survival of 6±1 D, with the lowest seen in BXD77 (4 D). Echocardiography revealed abnormal small heart and restrictive dysfunction in BXDs, resembling Grinch syndrome seen in ACT patients. A significant QTL (LRS = 17.11) at 86~94 Mb of Chromosome (Chr) 10 was identified (genome wide P < 0.05). Further filtering identified Gnptab , Slc25a3 , Uhrf1bp1l, and Chpt1 as top candidate genes associated with the survival of DOX-treated BXDs. Mapping of QTLs associated with echocardiography values and weight loss are in a progress. Conclusions: Survival, BW loss, and echocardiography parameters significantly varied among DOX-treated BXDs suggesting an influence of genetic background on expression of those traits. Gnptab , Slc25a3, Uhrf1bp1l, and Chpt1 (Chr 10) are associated with DOX-induced survival traits in BXDs of GRP, suggesting that these genes are involved in modulating ACT severity in humans and require further functional and biological validation.

Acute rheumatic fever (ARF) is an inflammatory disease that produces cardiac, joint, neurological, and dermatological manifestations. It is caused by an abnormal immune response to Group A streptococcus (GAS) infection, most commonly after tonsillopharyngitis. ARF can affect anyone but commonly occurs in children. Severe or recurrent disease can cause rheumatic heart disease (RHD), which results in severe morbidity and mortality. Management of ARF includes eradicating GAS colonization, controlling symptoms, and secondary prophylaxis. Although the prevalence of ARF was high across the globe in the 1900s, its incidence has declined dramatically in industrialized countries after the development of antibiotics, accessible healthcare, and improved housing conditions and hygiene. However, this disease continues to affect people in developing nations. Improved international awareness of ARF and RHD is required for its control. This chapter will focus on the epidemiology, etiology, and pathogenesis of ARF and RHD along with a thorough description of clinical manifestations with their underlying mechanism. Diagnostic criteria, differentials, management, and prevention are also described in this chapter.

BACKGROUND:There is growing evidence of a strong correlation between pain sensitivity and cognitive function under both physiological and pathological conditions. However, the detailed mechanisms remain largely unknown. In the current study, we sought to explore candidate genes and common molecular mechanisms underlying pain sensitivity and cognitive function with a transcriptome-wide association study using recombinant inbred mice from the BXD family. METHODS:The pain sensitivity determined by Hargreaves' paw withdrawal test and cognition-related phenotypes were systematically analyzed in 60 strains of BXD mice and correlated with hippocampus transcriptomes, followed by quantitative trait locus (QTL) mapping and systems genetics analysis. RESULTS:The pain sensitivity showed significant variability across the BXD strains and co-varies with cognitive traits. Pain sensitivity correlated hippocampual genes showed a significant involvement in cognition-related pathways, including glutamatergic synapse, and PI3K-Akt signaling pathway. Moreover, QTL mapping identified a genomic region on chromosome 4, potentially regulating the variation of pain sensitivity. Integrative analysis of expression QTL mapping, correlation analysis, and Bayesian network modeling identified Ring finger protein 20 (Rnf20) as the best candidate. Further pathway analysis indicated that Rnf20 may regulate the expression of pain sensitivity and cognitive function through the PI3K-Akt signaling pathway, particularly through interactions with genes Ppp2r2b, Ppp2r5c, Col9a3, Met, Rps6, Tnc, and Kras. CONCLUSIONS:Our study demonstrated that pain sensitivity is associated with genetic background and Rnf20-mediated PI3K-Akt signaling may involve in the regulation of pain sensitivity and cognitive functions.

Heart failure is a clinical syndrome that has become a leading public health problem worldwide. Globally, nearly 64 million individuals are currently affected by heart failure, causing considerable medical, financial, and social challenges. One therapeutic option for patients with advanced heart failure is mechanical circulatory support (MCS) which is widely used for short-term or long-term management. MCS with various ventricular assist devices (VADs) has gained traction in end-stage heart failure treatment as a bridge-to-recovery, -decision, -transplant or -destination therapy. Due to limitations in studying VADs in humans, animal studies have substantially contributed to the development and advancement of MCS devices. Large animals have provided an avenue for developing and testing new VADs and improving surgical strategies for VAD implantation and for evaluating the effects and complications of MCS on hemodynamics and organ function. VAD modeling by utilizing rodents and small animals has been successfully implemented for investigating molecular mechanisms of cardiac unloading after the implantation of MCS. This review will cover the animal research that has resulted in significant advances in the development of MCS devices and the therapeutic care of advanced heart failure.

The genetic reference population of recombinant inbred BXD mice has been derived from crosses between C57BL/6J and DBA/ 2J strains. The DBA/2J parent exhibits cardiomyopathy phenotypes, whereas C57BL/6J has normal heart. BXD mice are sequenced for studying genetic interactions in cardiomyopathies. The study aimed to assess cardiomyopathy traits in BXDs and investigate the quantitative genetic architecture of those traits. Echocardiography, blood pressure, and cardiomyocyte size parameters obtained from 44 strains of BXD family (n > 5/sex) at 4-5 mo of age were associated with heart transcriptomes and expression quantitative trait loci (eQTL) mapping was performed. More than twofold variance in ejection fraction (EF%), fractional shortening (FS%), left ventricular volumes (LVVols), internal dimensions (LVIDs), mass (LVM), and posterior wall (LVPW) thickness was found among BXDs. In male BXDs, eQTL mapping identified Ndrg4 on chromosome 8 QTL to be positively correlated with LVVol and LVID and negatively associated with cardiomyocyte diameter. In female BXDs, significant QTLs were found on chromosomes 7 and 3 to be associated with LVPW and EF% and FS%, respectively, and Josd2, Dap3, and Tpm3 were predicted as strong candidate genes. Our study found variable cardiovascular traits among BXD strains and identified multiple associated QTLs, suggesting an influence of genetic background on expression of echocardiographic and cardiomyocyte diameter traits. Increased LVVol and reduced EF% and FS% represented dilated cardiomyopathy, whereas increased LV mass and wall thickness indicated hypertrophic cardiomyopathy traits. The BXD family is ideal for identifying candidate genes, causal and modifier, that influence cardiovascular phenotypes. NEW & NOTEWORTHY This study aimed to establish a cardiac phenotype-genotype correlation in murine genetic reference population of BXD RI strains by phenotyping the echocardiography, blood pressure, and cardiomyocyte diameter traits and associating each collected phenotype with genetic background. Our study identified several QTLs and candidate genes that have significant association with cardiac hypertrophy, ventricular dilation, and function including systolic hyperfunction and dysfunction.

Background:Copper (Cu) is essential for the functioning of various enzymes involved in important cellular and physiological processes. Although critical for normal cardiac function, excessive accumulation, or deficiency of Cu in the myocardium is detrimental to the heart. Fluctuations in cardiac Cu content have been shown to cause cardiac pathologies and imbalance in systemic Cu metabolism. However, the genetic basis underlying cardiac Cu levels and their effects on heart traits remain to be understood. Representing the largest murine genetic reference population, BXD strains have been widely used to explore genotype-phenotype associations and identify quantitative trait loci (QTL) and candidate genes.Methods:Cardiac Cu concentration and heart function in BXD strains were measured, followed by QTL mapping. The candidate genes modulating Cu homeostasis in mice hearts were identified using a multi-criteria scoring/filtering approach.Results:Significant correlations were identified between cardiac Cu concentration and left ventricular (LV) internal diameter and volumes at end-diastole and end-systole, demonstrating that the BXDs with higher cardiac Cu levels have larger LV chamber. Conversely, cardiac Cu levels negatively correlated with LV posterior wall thickness, suggesting that lower Cu concentration in the heart is associated with LV hypertrophy. Genetic mapping identified six QTLs containing a total of 217 genes, which were further narrowed down to 21 genes that showed a significant association with cardiac Cu content in mice. Among those, Prex1 and Irx3 are the strongest candidates involved in cardiac Cu modulation.Conclusion:Cardiac Cu level is significantly correlated with heart chamber size and hypertrophy phenotypes in BXD mice, while being regulated by multiple genes in several QTLs. Prex1 and Irx3 may be involved in modulating Cu metabolism and its downstream effects and warrant further experimental and functional validations.

Introduction: Heart rhythm disorders or cardiac arrhythmias can cause sudden death and heart failure. Risk factors for cardiac arrhythmias vary including genetics, age, and other environmental and lifestyle factors. To sought how genetic background affects an expression of cardiac rhythm phenotypes, we assessed electrocardiography (ECG) traits in BXD family of mice derived from crosses between DBA/2J (D2) and C57BL/6J (B6) strains and explored the quantitative genetic architecture of those traits. Methods: ECG tracings were recorded in 44 BXD male (M) and female (F) strains (N >5 mice/sex) at 4-5 months of age anesthetized with 2% isoflurane. Heart rate (HR), ECG parameters, and frequency of arrhythmias in percentile (‰) were associated with blood pressure, echocardiography, and heart transcriptome values followed by quantitative trait loci (QTLs) mapping. Results: Parental D2 strains had the lowest HR and significantly prolonged QT intervals (62.82 ± 9.42 ms in D2F and 53.42 ± 1.73 ms in D2M) compared to sex matched B6 parental strains. Significantly widened QRS complexes were seen in BXD65F (14.27±2.97 ms) compared to 11.67±1.03 ms in B6F controls. In males, BXD83M had the widest QRS (13.48±1.74 ms) vs 10.89±0.41 ms seen in B6M controls. We found a significant association between QRS duration and increased left ventricular internal diameter (LVID) and reduced ejection fraction and fractional shortening. Further, varied cardiac arrhythmias including bradycardia, atrioventricular block (AVB), premature atrial or ventricular complexes (PAC and PVC, respectively), ventricular tachycardia (VT) and sick sinus syndrome (SSS) were seen among BXD strains. PACs were recorded in BXD73F (14.57 ‰), 40M, 101F, 51M, and 101M strains. Strains, BXD79M (14.97 ‰), 83M, 78M, 69F, and 171F, had frequent PVCs compared to B6 controls. AVB II was recorded in BXD48M and BXD66M had SSS. Both PAC and PVC were positively associated with cardiac phenotype burden. Specifically, PVCs were significantly correlated with echocardiographic pulmonary vein peak pressure, LVID and volumes at end-diastole (P = 0.04) among male BXDs. In females, right ventricular internal diameters (RVID) and cardiac output were significantly correlated with PVC and PAC frequencies. Moreover, PVCs were significantly correlated with systolic blood pressure in both male and female mice. QTL mapping identified a significant locus on Chromosome 3 associated with QTC and JT durations, while the same locus was suggestive for association with QT interval, suggesting that Chromosome 3 loci may be associated with repolarization abnormalities such as long QT syndromes. Conclusions: ECG parameters, heart rate, type and frequency of arrhythmias significantly varied between mouse strains of the BXD family suggesting an influence of genetic background on expression of those traits. Abnormal heart rhythms detected in BXD strains mimic cardiac rhythm and conduction disorders in humans. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

The transmembrane protein 43 (TMEM43/LUMA) p.S358L mutation causes arrhythmogenic cardiomyopathy named as ARVC5, a fully penetrant disease with high risk of ventricular arrhythmias, sudden death, and heart failure. Male gender and vigorous exercise independently predicted deleterious outcome. Our systems genetics analysis revealed the importance of Tmem43 for cardiac and metabolic pathways associated with elevated lipid absorption from small intestine. This study sought to delineate gender-specific cardiac, intestinal, and metabolic phenotypes in vivo and investigate underlying pathophysiological mechanisms of S358L mutation. Serial echocardiography, surface electrocardiography (ECG), treadmill running, and body EchoMRI have been used in knock-in heterozygous (Tmem43WT/S358L), homozygous (Tmem43S358L), and wildtype (Tmem43WT) littermate mice. Electron microscopy, histology, immunohistochemistry, transcriptome, and protein analysis have been performed in cardiac and intestinal tissues. Systolic dysfunction was apparent in 3-mo-old Tmem43S358L and 6-mo-old Tmem43WT/S358L mutants. Both mutant lines displayed intolerance to acute stress at 6 mo of age, arrhythmias, fibro-fatty infiltration, and subcellular abnormalities in the myocardium. Microarray analysis found significantly differentially expressed genes between left ventricular (LV) and right ventricular (RV) myocardium. Mutants displayed diminished PPARG activities and significantly reduced TMEM43 and β-catenin expression in the heart, whereas junctional plakoglobin (JUP) translocated into nuclei of mutant cardiomyocytes. Conversely, elongated villi, fatty infiltration, and overexpression of gut epithelial proliferation markers, β-catenin and Ki-67, were evident in small intestine of mutants. We defined Tmem43 S358L-induced pathological effects on cardiac and intestinal homeostasis via distinctly disturbed WNT-β-catenin and PPARG signaling thereby contributing to ARVC5 pathophysiology. Results suggest that cardiometabolic assessment in mutation carriers may be important for predictive and personalized care.NEW & NOTEWORTHY This manuscript describes the findings of our investigation of cardiac, small intestine, and metabolic features of Tmem43-S358L mouse model. By investigating interorgan pathologies, we uncovered multiple mechanisms of the S358L-induced disease, and these unique mechanisms likely appear to contribute to the disease pathogenesis. We hope our findings are important and novel and open new avenues in the hunting for additional diagnostic and therapeutic targets in subjects carrying TMEM43 mutation.

Most prominent functional abnormalities seen in the failing human heart are impaired contraction and slowed rates of relaxation of cardiac cells in the face of increased neurohormonal activation, sustained inflammation, mechanical and volume overload, and progressive maladaptive remodeling of the myocardium. Mechanical circulatory support devices (MCS) improve cardiac function and outcomes of patients with end-stage heart failure, allowing to bridge to heart transplantation and permitting the removal of MCS device as a bridge to recovery, in some patients with the sufficient recovery of heart function. Numerous reports have demonstrated favorable myocardial recovery and reverse remodeling after prolonged ventricular unloading by MCS. Ventricular unloading by MCS leads to a decreased concentration of peripheral natriuretic peptides in plasma, reduction in cardiac cytokines, kinases, collagens, and proteins involved in hypertrophy, fibrosis, programmed cell death, and necrosis in the heart. This chapter will summarize and review the effects and underlying mechanisms of myocardial remodeling during prolonged MCS in patients with end-stage heart failure. The mechanisms of myocardial recovery are multifactorial and remain to be further explored on cellular, organ, and systems levels.

Background: Cardiomyopathies are responsible for nearly 10% of cardiac deaths in neonates. Recently, biallelic variants in ribosomal protein L3-like ( RPL3L , NM_005061.2) have been reported in neonates with dilated cardiomyopathy (DCM) and heart failure (HF). This study is aimed to study RPL3L induced genetic pathways and mechanisms involved in the development of DCM. Methods: Whole exome and RNA sequencing, histology, immunohistochemistry, and Western blotting were performed in a human explanted heart. Genetic correlation, expression quantitative trait loci (eQTL) mapping and functional analysis were performed using cardiac gene expression data from the patient as well as human and murine genetic reference populations (GRPs). Results: Two-month-old patient with DCM and HF carrying compound heterozygous RPL3L variants, p.A51T and p.V231F, has been managed on an INTERMACS profile 2 with continuous-flow ventricular assist device and successfully bridged to heart transplantation at the age of 4 months. Morphology of the explanted heart displayed apical deep intertrabecular recesses consistent with left ventricular noncompaction (LVNC). Histology revealed interstitial fibrosis and myocardial hypertrophy. No signs of myocarditis and glycogen or mucopolysaccharides deposits were detected. Immunohistochemistry revealed abnormal clusters of RPL3L protein in the cytosol of cardiomyocytes and disruption of F-actin, Z-disk proteins, MYL4 and dystrophin was evident. In a mouse heart, the Rpl3l level was of 33.45 RPKM, while levels of RPL3L appeared to be 32 RPKM in a human heart compared to its mean expression in other adult tissues of 2.54 (mouse) and 2.36 (human). Systems genetics analysis identified high expression values ranged from 11.31 to 12.16 across murine GRPs of BXD mice with the ~1.8-fold difference. Pathways such as “thermogenesis”, “diabetic cardiomyopathy” and “DCM” significantly associated with RPL3L. e QTL mapping suggested Myl4 (Chr 11) and Sdha (Chr 13) as the upstream regulators of Rpl3l . Conclusions: Compound RPL3L heterozygosity is causal for neonatal DCM and LVNC. The RPL3L is highly expressed in the heart tissue of humans and mice. Genes, Myl4 and Sdha, are strong candidates that regulate expression of Rpl3l in murine heart.

Introduction: Mutations in myopalladin ( MYPN) gene are associated with cardiomyopathies (CM) and skeletal myopathies (SM). This study is aimed to identify Mypn -induced cardioskeletal phenotypes in a murine genetic reference population (GRP) and uncover common genetic mechanisms underlying the development of CM and SM. Hypothesis: Mypn induces common molecular signaling in cardiac and skeletal muscles. Methods: Pearson correlation between cardiac function and expression of Mypn in gastrocnemius muscles was performed in 40 mouse BXD recombinant inbred strains derived from C57BL/6J and DBA/2J parental mice. A causal Mypn-Q526X mutation was introduced to BXDs by crossing with MypnWT/Q526X knock-in mice and this generated F1 hybrid mutant (BXDQ526X) and wild type (BXDWT) strains. Cardiac (morphology, function by echo) and skeletal muscle (muscle mass, strength) phenotypes and transcriptomics (RNA-seq) were assessed in 3- and 6-month-old hybrid mice (N=5). Results: Skeletal muscle expression of Mypn was significantly associated with heart rate (HR) and heart mass (HM) in BXDs, while it was negatively associated with interventricular septum (IVS) thickness, indicating BXD strains with higher skeletal muscle Mypn expression had faster HR, higher HM but thinner IVS. Varied phenotypes in ejection fraction (EF%), ventricular volume and hypertrophy were found among BXD Q526X mice; BXD51 Q526X , BXD48 Q526X and BXD69 Q526X strains had significantly higher EF%, while some mutant mice showed EF% decrease compared to their respective BXD WT littermates. Many mutant lines displayed significantly reduced skeletal muscle strength or mass. Overt SM-related signs (rear limb deformity, restricted movement, waddling) were seen in BXD78 Q526X , while we found overlapping CM/SM traits in BXD51 Q526X mice. Conclusions: In this study, common Mypn -induced molecular signaling in cardiac and skeletal muscles is identified. Introduction of the Mypn -Q526X mutation into the diverse genetic background of BXD GRP exposed CM- and SM-related phenotypes in hybrid lines. The most susceptible lines with overt and overlapping CM/SM phenotypes will be useful to identify common genes, networks and pathways involved in CM and SM using systems genetics methods.

BACKGROUND:Troponin-I interacting kinase encoded by the TNNI3K gene is expressed in nuclei and Z-discs of cardiomyocytes. Mutations in TNNI3K were identified in patients with cardiac conduction diseases, arrhythmias, and cardiomyopathy. METHODS:We performed cardiac gene expression, whole genome sequencing (WGS), and cardiac function analysis in 40 strains of BXD recombinant inbred mice derived from C57BL/6J (B6) and DBA/2J (D2) strains. Expression quantitative trait loci (eQTLs) mapping and gene enrichment analysis was performed, followed by validation of candidate Tnni3k-regulatory genes. RESULTS:WGS identified compound splicing and missense T659I Tnni3k variants in the D2 parent and some BXD strains (D allele) and these strains had significantly lower Tnni3k expression than those carrying wild-type Tnni3k (B allele). Expression levels of Tnni3k significantly correlated with multiple cardiac (heart rate, wall thickness, PR duration, and T amplitude) and metabolic (glucose levels and insulin resistance) phenotypes in BXDs. A significant cis-eQTL on chromosome 3 was identified for the regulation of Tnni3k expression. Furthermore, Tnni3k-correlated genes were primarily involved in cardiac and glucose metabolism-related functions and pathways. Genes Nodal, Gnas, Nfkb1, Bmpr2, Bmp7, Smad7, Acvr1b, Acvr2b, Chrd, Tgfb3, Irs1, and Ppp1cb were differentially expressed between the B and D alleles. CONCLUSIONS:Compound splicing and T659I Tnni3k variants reduce cardiac Tnni3k expression and Tnni3k levels are associated with cardiac and glucose metabolism-related phenotypes.

Introduction: Desmoplakin (DSP) is a key protein for cardiac myocyte cell-to-cell adhesion that connects the desmosomes present in intercalated discs to the cytoplasmic network of desmin intermediate filaments. Pathogenic variants of the human DSP gene often lead to left ventricular (LV)-dominant and biventricular forms of arrhythmogenic cardiomyopathy characterized by a dilated LV, arrhythmias, and an elevated risk of sudden cardiac death. Nonetheless, the pathophysiological pathways triggered by DSP variants remain unknown. Hypothesis: We hypothesized that DSP pathogenic variants modify cardiomyocyte excitation-contraction coupling leading to life-threatening ventricular arrhythmias. Methods: We created a novel knock-in mouse model based on the human DSP p.S2859LfsX5 variant that leads to a murine protein truncated from its last 9 amino-acids. We compared one-year-old sedentary heterozygous (Dsp +/- ) animals to WT littermates and defined histologic, cellular, and molecular outcomes of the variant. Results: Western blotting revealed 50% Dsp reduction (N= 5 hearts/group; P < 0.05) in mutant right ventricle (RV) indicating mutant protein instability. Histologic assessments of Dsp +/- hearts revealed LV chamber enlargement and RV wall thinning in the absence of fibrosis and lipid infiltration. Dsp +/- myocytes isolated from LV and RV free walls and field stimulated at 3 Hz exhibited 30-40% reduction in sarcomere shortening (n= 17-25 myocytes/ventricle/3-4 hearts/group; P < 0.05), while their Ca 2+ handling properties remained normal. Pacing of Dsp +/- isolated myocytes (LV and RV) in the presence of (100 nmol/L) isoproterenol provoked a 20-30% increase in Ca 2+ release due to sarcoplasmic reticulum Ca 2+ overload (n= 15-20 myocytes/ventricle/3-4 hearts/group; P < 0.05). Finally, surface ECGs showed that activation of the sympathetic pathway (isoproterenol + caffeine) in 6-month-old Dsp +/- mice increased mutant heart susceptibility to ventricular arrhythmias. Conclusions: The pathogenic variant p.S2859LfsX5 promotes biventricular dysfunction due to the loss in myocyte contraction capacity and catecholaminergic-mediated ventricular arrhythmias from cardiomyocyte Ca 2+ overload.

Background: Cardiomyopathies, diseases affecting the myocardium, are common causes of congestive heart failure (CHF) and sudden cardiac death. Recently, biallelic variants in ribosomal protein L3-like (RPL3L) have been reported to be associated with severe neonatal dilated cardiomyopathy (DCM) and CHF. This study employs a systems genetics approach to gain understanding of the regulatory mechanisms underlying the role of RPL3L in DCM. Methods: Genetic correlation, expression quantitative trait loci (eQTL) mapping, differential expression analysis and comparative functional analysis were performed using cardiac gene expression data from the patients and murine genetic reference populations (GRPs) of BXD mice (recombinant inbred strains from a cross of C57BL/6J and DBA/2J mice). Additionally, immune infiltration analysis was performed to understand the relationship between DCM, immune cells and RPL3L expression. Results: Systems genetics analysis identified high expression of Rpl3l mRNA, which ranged from 11.31 to 12.16 across murine GRPs of BXD mice, with an ~1.8-fold difference. Pathways such as “diabetic cardiomyopathy”, “focal adhesion”, “oxidative phosphorylation” and “DCM” were significantly associated with Rpl3l. eQTL mapping suggested Myl4 (Chr 11) and Sdha (Chr 13) as the upstream regulators of Rpl3l. The mRNA expression of Rpl3l, Myl4 and Sdha was significantly correlated with multiple echocardiography traits in BXD mice. Immune infiltration analysis revealed a significant association of RPL3L and SDHA with seven immune cells (CD4, CD8-naive T cell, CD8 T cell, macrophages, cytotoxic T cell, gamma delta T cell and exhausted T cell) that were also differentially infiltrated between heart samples obtained from DCM patients and normal individuals. Conclusions: RPL3L is highly expressed in the heart tissue of humans and mice. Expression of Rpl3l and its upstream regulators, Myl4 and Sdha, correlate with multiple cardiac function traits in murine GRPs of BXD mice, while RPL3L and SDHA correlate with immune cell infiltration in DCM patient hearts, suggesting important roles for RPL3L in DCM and CHF pathogenesis via immune inflammation, necessitating experimental validations of Myl4 and Sdha in Rpl3l regulation.

The murine recombinant inbred (RI) family of BXD strains derived from crosses between C57BL/6J (B6) and DBA/2J (D2) mice are the largest genetic reference population (GRP). The parental D2 mouse is a natural model of hypertrophic cardiomyopathy (HCM). Thus, the BXD family represents an applicable rodent model for phenotype‐genotype studies of complex cardiomyopathy traits.

Introduction: Prominent arrhythmias are features of arrhythmogenic cardiomyopathy (ACM) and arrhythmogenic dilated cardiomyopathy (aDCM). Despite clinicopathological differences, these diseases share common genetic causes. Hypothesis: This study aimed to identify variants in ACM and aDCM and map the candidate genetic regulators using a mouse genetic reference population (GRP). Methods: Whole exome sequencing was performed on 226 individuals with ACM and 102 patients with aDCM. Bioinformatics analysis was applied to identify the pathogenic or likely pathogenic variants followed by direct sequencing confirmation. Expression quantitative trait loci (eQTLs) mapping to heart transcriptomes of GRP of BXD recombinant inbred (RI) mice was conducted in GeneNetwork (http://www.genenetwork.org). Results: In the ACM cohort, a total of 82 variants (MAF<0.01) in 24 genes have been confirmed being positive in 42 probands. Depending on the frequency of the identified variants, the top 10 genes (excluding TTN) with the most pathogenic variants were PKP2, MYH6, FLNC, RYR2, MYH7B, PLEC, JPH2, SYNE1, NRAP and POSTN. In the aDCM cohort, 141 pathogenic variants have been identified and 91 patients harbored at least 1 deleterious variant. The top 10 genes were RYR2, FLNC, MYH7, DSP, SCN5A,MYPN, NEBL, CTNNA3, ACAP6 and SLC22A5. We analyzed the distribution of the first principal component (PC1) of mRNA expression levels of those 10 genes (eigentraits) from each cohort in the heart across BXD strains. Interval mapping for PC1 eigentraits for ACM identified a significant eQTL (P<0.05) in the mouse genome at 24-36 Mb of chromosome (Chr) 6. In aDCM, interval mapping for PC1 eigentraits identified 3 suggestive eQTLs at Chr 5 (44-53 Mb), Chr 6 (32-34 Mb) and Chr 15 (82-84 Mb). Further gene exploration of these eQTLs suggested that Mtpn, Lep, Cyb5r3, Tspo, Slit2, and Ppargc1a could be common regulators that interact with those top ACM and aDCM genes. Conclusions: This study identified diverse genes in ACM and aDCM cases. Our joint human and mouse analysis using a systems genetics approach identified genetic regulators that could affected expression of genes with pathogenic variants in ACM and aDCM patients.

Broad cellular functions and diseases including muscular dystrophy, arrhythmogenic right ventricular cardiomyopathy (ARVC5) and cancer are associated with transmembrane protein43 (TMEM43/ LUMA). The study aimed to investigate biological roles of TMEM43 through genetic regulation, gene pathways and gene networks, candidate interacting genes, and up- or downstream regulators. Cardiac transcriptomes from 40 strains of recombinant inbred BXD mice and two parental strains representing murine genetic reference population (GRP) were applied for genetic correlation, functional enrichment, and coexpression network analysis using systems genetics approach. The results were validated in a newly created knock-in Tmem43-S358L mutation mouse model (Tmem43S358L) that displayed signs of cardiac dysfunction, resembling ARVC5 phenotype seen in humans. We found high Tmem43 levels among BXDs with broad variability in expression. Expression of Tmem43 highly negatively correlated with heart mass and heart rate among BXDs, whereas levels of Tmem43 highly positively correlated with plasma high-density lipoproteins (HDL). Through finding differentially expressed genes (DEGs) between Tmem43S358L mutant and wild-type (Tmem43WT) lines, 18 pathways (out of 42 found in BXDs GRP) that are involved in ARVC, hypertrophic cardiomyopathy, dilated cardiomyopathy, nonalcoholic fatty liver disease, Alzheimer’s disease, Parkinson’s disease, and Huntington’s disease were verified. We further constructed Tmem43-mediated gene network, in which Ctnna1, Adcy6, Gnas, Ndufs6, and Uqcrc2 were significantly altered in Tmem43S358L mice versus Tmem43WT controls. Our study defined the importance of Tmem43 for cardiac- and metabolism-related pathways, suggesting that cardiovascular disease-relevant risk factors may also increase risk of metabolic and neurodegenerative diseases via TMEM43-mediated pathways.

Restrictive cardiomyopathy (RCM), a potentially devastating heart muscle disorder, is characterized by diastolic dysfunction due to abnormal muscle relaxation and myocardial stiffness resulting in restrictive filling of the ventricles. Diastolic dysfunction is often accompanied by left atrial or bi-atrial enlargement and normal ventricular size and systolic function. RCM is the rarest form of cardiomyopathy, accounting for 2-5% of pediatric cardiomyopathy cases, however, survival rates have been reported to be 82%, 80%, and 68% at 1-, 2-, and 5-years after diagnosis, respectively. RCM can be idiopathic, familial, or secondary to a systemic disorder, such as amyloidosis, sarcoidosis, and hereditary hemochromatosis. Approximately 30% of cases are familial RCM, and the genes that have been linked to RCM are cTnT, cTnI, MyBP-C, MYH7, MYL2, MYL3, DES, MYPN, TTN, BAG3, DCBLD2, LNMA, and FLNC. Increased Ca2+ sensitivity, sarcomere disruption, and protein aggregates are some of the few mechanisms of pathogenesis that have been revealed by studies utilizing cell lines and animal models. Additional exploration into the pathogenesis of RCM is necessary to create novel therapeutic strategies to reverse restrictive cardiomyopathic phenotypes.