Investigation of the pathomechanism of uremic cardiomyopathy and the infarct
size-limiting effect of ischemic preconditioning in a rat model of chronic
kidney disease
Authors: Márványkövi Fanni Magdolna 
Published: July 2023

Chronic kidney disease (CKD) is a public health problem affecting 1 of 10
people worldwide. Interestingly, approximately 60% of patients are women in the
early stages of CKD. A common cardiovascular complication of CKD is uremic
cardiomyopathy, most characterized by left ventricular hypertrophy (LVH) and
fibrosis, ultimately leading to heart failure (HF). Moreover, uremic
cardiomyopathy enhances the susceptibility of the heart to acute myocardial
infarction (AMI). However, the precise molecular mechanisms and the role of
sex-based differences in the development of uremic cardiomyopathy and AMI in CKD
are still unclear. Therefore, novel therapeutic strategies that alleviate the
severity of uremic cardiomyopathy and AMI in CKD are urgently needed.
MicroRNA-212 (miR-212) has been demonstrated previously to be a crucial
regulator of pathologic LVH in pressure-overload-induced HF via regulating the
forkhead box O3 (FOXO3)/calcineurin/nuclear factor of activated T-cells (NFAT)
pathway. Here we aimed to investigate whether i) miR-212 and its selected
hypertrophy- and fibrosis-associated targets play a role in the development of
uremic cardiomyopathy, ii) the influence of sex on the severity of uremic
cardiomyopathy and AMI, as well as the infarct size-limiting effect of ischemic
preconditioning (IPRE) in experimental CKD. CKD was induced by 5/6 nephrectomy
in male and female Wistar rats. Serum and urine laboratory parameters were
measured to verify the development of CKD 8 or 9 weeks after the operations.
Transthoracic echocardiography was performed to assess cardiac function and
morphology. Cardiomyocyte hypertrophy and fibrosis were measured by histology.
Left ventricular (LV) samples were collected for RT-qPCR, Western blot, and
ELISA measurements. The LV expressions of miR-212 and its LVH and
fibrosis-associated selected targets, including FOXO3, AKT, and ERK1/2, were
measured only in males by RT-qPCR and/or Western blot. In a subgroup of animals,
hearts were perfused according to Langendorf and were subjected to 35 min global
ischemia and 120 min reperfusion with or without IPRE. Then the infarct size or
phosphorylated (p) and total forms of proteins related to the cardioprotective
RISK (AKT, ERK1/2) and SAFE (STAT3) pathways were measured in the myocardial
samples by Western blot. The severity of CKD was similar in males and females
based on serum urea and creatinine levels. In CKD, diastolic dysfunction
developed with preserved ejection fraction and increased A-type natriuretic
peptide (ANP) levels in both sexes; however, males developed more severe LVH
than females. Moreover, histology showed the development of marked cardiac
fibrosis only in CKD in males. The miR-212 was significantly overexpressed in
the LV samples in CKD in males. However, the LV expression of FOXO3, AMPK, and
ERK1/2 failed to change significantly at the mRNA or protein level.
Interestingly, only the LV pAKT/AKT ratio was significantly increased in males
in CKD. Females had significantly smaller infarct sizes both in the sham and CKD
groups compared to males. In both sexes, IPRE significantly decreased the
infarct size in both the sham-operated and CKD groups. IPRE significantly
increased the pSTAT3/STAT3 ratio in sham-operated but not in CKD animals in both
sexes. The groups had no significant differences in pAKT/AKT and pERK1/2 /
ERK1/2 ratios. In summary, cardiac overexpression of miR-212 in CKD failed to
affect its previously implicated hypertrophy- and fibrosis-associated downstream
targets in males. Thus, the molecular mechanism of the development of LVH in CKD
seems to be independent of the FOXO3, ERK1/2, and AMPK in our model. The infarct
size-limiting effect of IPRE was preserved in both sexes in CKD despite the more
severe uremic cardiomyopathy in male CKD rats and the smaller infarct size in
females. Further research is needed to identify crucial molecular mechanisms in
the development of uremic cardiomyopathy and the cardioprotective effects of
IPRE in CKD.