Study on the Synergistic Effect of Klotho and KRAS on Reducing Ferroptosis After Myocardial Infarction by Regulating...

Myocardial infarction (MI) is a coronary artery-related disease that seriously threatens human life and is the leading cause of sudden death worldwide, where a lack of nutrients and oxygen leads to an inflammatory response and death of cardiomyocytes. Ferroptosis is a form of non-apoptotic cell death associated with metabolic dysfunction, resulting in abnormal breakdown of glutamine and iron-dependent accumulation of reactive oxygen species (ROS) during metabolism. However, the molecular mechanism of ferroptosis in the pathogenesis of MI and the function of Klotho and KRAS on ferroptosis during MI remain unclear. The MI rat model was established by LAD ligation with a 6-0 suture. H9c2 cells were placed in glucose-deficient DMEM (Thermo) and cultured in an anaerobic environment (1% CO₂ and 5% CO) to establish an in vitro OGD cell model. The damage to rat heart tissue was detected by HE staining, and Klotho and KRAS were determined by RT-qPCR, Western Blot, and IHC. TUNEL staining was used to determine apoptosis in rat heart tissue samples. The interaction between Klotho and KRAS was verified by co-immunoprecipitation and Western Blot. The cardiomyocyte activity was measured by CCK-8 assay. LDH, CK-MB, cTnT, and Fe²⁺ markers were detected by the kits. For the assessment of ferroptosis, GSH and ROS in cardiomyocytes and serum were detected by kits, and PTSG was detected by Western Blot. IL-1β and IL-6 in cardiomyocytes and serum were determined by ELISA. Klotho was downregulated in MI. Downregulation of Klotho promoted myocardial injury; increased apoptosis of cardiomyocytes; promoted LDH, CK-MB, and cTnT concentrations; inhibited GSH; and promoted ROS levels, PTGS2 expression, and ferroptosis in rats. The same results were obtained in vitro. Klotho and KRAS had endogenous interactions. KRAS knockdown can reverse Klotho knockdown-mediated MI and ferroptosis. RAP1/ERK pathway was highly expressed in MI, and inhibiting RAP1/ERK pathway activation can reverse the promoting effect of overexpressed KRAS on MI progression and ferroptosis. Klotho interacts with KRAS and inhibits ferroptosis after MI by regulating the RAP1/ERK pathway.