Proteomic analysis of inducible pluripotent stem cell-derived cardiomyocytes (iPSC-CM) and permanent cardiomyocyte and monocytic cell lines with and without the rare variants observed in CCC under the effect of IFN-gamma and TNF-alpha

• Grant number: 25/27262-2
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)
• Start date: January 01, 2026
• End date: December 31, 2029
• Field of knowledge: Biological Sciences - Immunology - Applied Immunology
• Principal Investigator: Edecio Cunha Neto
• Grantee: Guilherme Fahl Zonzin
• Host Institution: Instituto do Coração Professor Euryclides de Jesus Zerbini (INCOR). Hospital das Clínicas da Faculdade de Medicina da USP (HCFMUSP). Secretaria da Saúde (São Paulo - Estado). São Paulo , SP, Brazil
• Associated research grant: 22/00758-0 - Mitochondria, Interferon-gamma and genetics in Chagas disease cardiomyopathy: Pathogenesis, therapeutic targets and prognostic markers, AP.TEM

Abstract

During Chagasic cardiomyopathy (CCC), IFN-gamma and TNF-alpha have been implicated in the pathogenesis and progression of heart disease. Mitochondrial dysfunction is a key finding in CCC. Recently, our group has shown that both cytokines impair mitochondrial function and changes mitochondrial protein expression in of AC-16 cardiomyocytes, an immortalized human cell line, similar with the findings in heart tissue from CCC patients. Pathogenic mitochondrial and inflammatory gene variants were found specifically in CCC patients but not indeterminate form siblings in families with multiple cases of Chagas disease. Inducible pluripotent stem cell-derived cardiomyocytes (iPSC-CM) derived from patients carrying or not the pathogenic gene variants offer the unique opportunity to assess the effect of gene variants in the cell type of interest. CRISPR-Cas9 enables the introduction of gene variants in multiple cell types. We thus propose to assess the effect of IFN-gamma and TNF-alpha on the whole cell and mitochondrial proteome of iPSC-CM from patients with CCC, indeterminate form or seronegative subjects from family 5 carrying or not the CCC-specific pathogenic gene variant DHODH R135C (C/T). In addition, we have also been able to use CRISPR-Cas9 to correct the gene variant DHODH R135C (C/T) in iPSC from patient 6135 to DHODH R135 C/C. We will also assess the whole cell and mitochondrial proteome of immortalized cardiomyocyte cell line AC16 gene edited with CRISPR-Cas9 to express the gene variants of interest. All cell lines have already been obtained and are available in the lab, and we have confirmed that cells carrying the variants are more susceptible to cytokine-induced mitochondrial dysfunction. (AU)