Serum circulating proteins from pediatric patients with dilated cardiomyopathy cause pathologic remodeling and cardiomyocyte stiffness

Abstract

Dilated cardiomyopathy (DCM) is the most common form of cardiomyopathy and main indication for heart transplantation in children. Therapies specific to pediatric DCM remains limited due to lack of a disease model. Our previous study showed that treatment of neonatal rat ventricular myocytes (NRVMs) with non-failing or DCM pediatric patient serum activates the fetal gene program (FGP). Here we show that serum treatment with Proteinase K prevents activation of the FGP, whereas RNase treatment exacerbates it, suggesting that circulating proteins, but not circulating microRNAs, promote these pathological changes. Evaluation of the protein secretome showed that midkine (MDK) is up-regulated in DCM serum, and NRVM treatment with MDK activates the FGP. Changes in gene expression in serum-treated NRVMs, evaluated by next-generation RNA sequencing (RNA-Seq), indicates extracellular matrix remodeling and focal adhesion pathways are upregulated in pediatric DCM serum and serum-treated NRVMs, suggesting alterations in cellular stiffness. Cellular stiffness was evaluated by Atomic Force Microscopy, which showed an increase in stiffness in DCM serum-treated NRVMs. Of the proteins increased in DCM sera, secreted frizzled related protein 1 (sFRP1) was a potential candidate for the increase in cellular stiffness, and sFRP1 treatment of NRVMs recapitulated the increase in cellular stiffness observed in response to DCM-serum treatment. Our results show that serum circulating proteins promote pathological changes in gene expression and cellular stiffness, and circulating miRNAs are protective against pathological changes.

Midkine—A novel player in cardiovascular diseases

“…MK hence drived cardiac remodeling in chronic kidney disease via EGFR signaling ( 143 ). Furthermore, a different report disclosed MK protein upregulation in the sera and cardiac tissue of pediatric patients with dilated cardiomyopathy (DCM) ( 121 ). Whereas, MK treatment of neonatal rat ventricular myocytes upregulated atrial natriuretic factor (ANF) and BNP expression, incubation with DCM serum, supplemented with anti-MK antibody, averted ANF and BNP upregulation in these cells ( 121 ).…”

“…Whereas, MK treatment of neonatal rat ventricular myocytes upregulated atrial natriuretic factor (ANF) and BNP expression, incubation with DCM serum, supplemented with anti-MK antibody, averted ANF and BNP upregulation in these cells ( 121 ). Here, MK participated in the activation of fetal gene program mediated by DCM serum ( 121 ). However, and in divergence to the above presented findings, MK had a protective influence on cardiac remodeling in tachycardia-induced congestive heart failure ( 144 ).…”

Hydroxychloroquine Mitigates Dilated Cardiomyopathy Phenotype in Transgenic D94A Mice

“…ANF and BNP peptides are closely related to LV function and are sensitive markers of the severity of heart injury [ 21 , 22 ] and sensitive markers of pathological DCM phenotype [ 23 ]. BNP and its N-terminal (NT)-prohormone (NT-proBNP) are released in response to changes in pressure inside the heart, and their levels were seen to increase in patients with heart failure [ 24 , 25 ].…”

Atomic Force Microscopy (AFM) Applications in Arrhythmogenic Cardiomyopathy

“…A genetic cardiocutaneous disorder associated with a desmoplakin mutation has also been recently studied by our group with AFM, and keratinocytes derived from a patient skin biopsy exhibited a higher deformability (lower stiffness) than controls [ 46 ]. Moreover, AFM has also been used to prove that serum circulating proteins from pediatric patients with DCM cause cardiomyocyte stiffness [ 47 ].…”