Post-myocardial infarction heart failure and long-term high-fat diet: Cardiac endoplasmic reticulum stress and unfolded protein response in Sprague Dawley rat model

Background Myocardial infarction (MI) significantly contributes to the global mortality rate, often leading to heart failure (HF) due to left ventricular remodeling. Key factors in the pathomechanism of HF include nitrosative/oxidative stress, inflammation, and endoplasmic reticulum (ER) stress. Furthermore, while a high-fat diet (HFD) is known to exacerbate post-MI cardiac remodeling, its impact on these critical factors in the context of HF is not as well understood. Aims This study aimed to assess the impact of post-MI HF and HFD on inflammation, nitro-oxidative stress, ER stress, and unfolded protein response (UPR). Methods The study was performed on fragments of the left ventricle harvested from 30 male adult Sprague Dawley rats, which were divided into four groups based on diet (normal-fat vs. high-fat) and surgical procedure (sham operation vs. coronary artery ligation to induce MI). We assessed body weight, NT-proBNP levels, protein levels related to nitrosative/oxidative stress, ER stress, UPR, apoptosis, and nitric oxide synthases, through Western Blot and ELISA. Results HFD and MI significantly influenced body weight and NT-proBNP concentrations. HFD elevated 3-nitrotyrosine and myeloperoxidase levels and altered nitric oxide synthase levels. HFD and MI significantly affected ER stress markers and activated or inhibited UPR pathways. Conclusions The study demonstrates significant impacts of post-MI HF and dietary fat content on cardiac function and stress markers in a rat model. The interaction between HFD and MI on UPR activation suggests the importance of dietary management in post-MI recovery and HF prevention.

The expected size of the detected IRE1α protein was around 110 kDa.However, in our results, a band was observed at approximately 70 kDa.This discrepancy might be due to differences in protein migration during electrophoresis, post-translational modifications, or protein degradation during sample preparation.We also observed additional bands at sizes other than 70 kDa, which could represent degradation products of IRE1α or non-specific antibody binding.Each sample was normalized to βactin to minimize these differences, allowing for relative protein level comparisons between samples.
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C .The expected size of the detected phospho-IRE1α protein was around 110 kDa.However, in our results, a band was observed at approximately 75 kDa.This discrepancy might be due to differences in protein migration during electrophoresis, post-translational modifications, or protein degradation during sample preparation.We also observed additional bands at sizes other than 75 kDa, which could represent degradation products of phospho-IRE1α or non-specific antibody binding.Each sample was normalized to β-actin to minimize these differences, allowing for relative protein level comparisons between samples.The expected size of the detected eNOS protein was around 140 kDa.However, in our results, a band was observed at approximately 75 kDa.This discrepancy might be due to differences in protein migration during electrophoresis, post-translational modifications, or protein degradation during sample preparation.We also observed additional bands at sizes other than 75 kDa, which could represent degradation products of phospho-IRE1α or non-specific antibody binding.Each sample was normalized to β-actin to minimize these differences, allowing for relative protein level comparisons between samples.

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C .The expected size of the detected nNOS protein was around 160 kDa.However, in our results, a band was observed at approximately 75 kDa.This discrepancy might be due to differences in protein migration during electrophoresis, post-translational modifications, or protein degradation during sample preparation.We also observed additional bands at sizes other than 75 kDa, which could represent degradation products of phospho-IRE1α or non-specific antibody binding.Each sample was normalized to β-actin to minimize these differences, allowing for relative protein level comparisons between samples.The expected size of the detected PERK protein was around 125 kDa.However, in our results, a band was observed at approximately 70 kDa.This discrepancy might be due to differences in protein migration during electrophoresis, post-translational modifications, or protein degradation during sample preparation.We also observed additional bands at sizes other than 70 kDa, which could represent degradation products of phospho-IRE1α or non-specific antibody binding.Each sample was normalized to β-actin to minimize these differences, allowing for relative protein level comparisons between samples.

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ATF6 and ATF6-cleaved The expected size of the detected ATF6 (non-cleaved) protein was around 90 kDa.However, in our results, a band was observed at approximately 70 kDa.This discrepancy might be due to differences in protein migration during electrophoresis, post-translational modifications, or protein degradation during sample preparation.We also observed additional bands at sizes other than 70 kDa, which could represent degradation products of phospho-IRE1α or non-specific antibody binding.Each sample was normalized to β-actin to minimize these differences, allowing for relative protein level comparisons between samples.

Figure 1 .
Figure 1.Western Blot results for 3-nitrosine.A. PVDF membrane with left ventricular samples from sham operated rats fed with the normal fat diet (NFD SO).B. PVDF membrane with left ventricular samples from rats with ligation of the coronary artery fed with the normal fat diet (NFD MI). C. PVDF membrane with left ventricular samples from sham operated rats fed with the normal fat diet (HFD SO).D. PVDF membranę with left ventricular samples from rats with ligation of the coronary artery fed with the normal fat diet.St -protein standard.

Figure 2 .
Figure 2. Western Blot results for 78-kDa glucose-regulated protein (GRP78).A. PVDF membrane with left ventricular samples from sham operated rats fed with the normal fat diet (NFD SO).B. PVDF membrane with left ventricular samples from rats with ligation of the coronary artery fed with the normal fat diet (NFD MI). C. PVDF membrane with left ventricular samples from sham operated rats fed with the normal fat diet (HFD SO).D. PVDF membranę with left ventricular samples from rats with ligation of the coronary artery fed with the normal fat diet.St -protein standard.

Figure 3 .Figure 4 .
Figure 3.Western Blot results for myeloperoxidase (MPO).A. PVDF membrane with left ventricular samples from sham operated rats fed with the normal fat diet (NFD SO).B. PVDF membrane with left ventricular samples from rats with ligation of the coronary artery fed with the normal fat diet (NFD MI). C. PVDF membrane with left ventricular samples from sham operated rats fed with the normal fat diet (HFD SO).D. PVDF membranę with left ventricular samples from rats with ligation of the coronary artery fed with the normal fat diet.St -protein standard.

Figure 5 .
Figure 5.Western Blot results for phospho-IRE1α. A. PVDF membrane with left ventricular samples from sham operated rats fed with the normal fat diet (NFD SO).B. PVDF membrane with left ventricular samples from rats with ligation of the coronary artery fed with the normal fat diet (NFD MI). C. PVDF membrane with left ventricular samples from sham operated rats fed with the normal fat diet (HFD SO).D. PVDF membranę with left ventricular samples from rats with ligation of the coronary artery fed with the normal fat diet.St -protein standard.The expected size of the detected phospho-IRE1α protein was around 110 kDa.However, in our results, a band was observed at approximately 75 kDa.This discrepancy might be due to differences in protein migration during electrophoresis, post-translational modifications, or protein degradation during sample preparation.We also observed additional bands at sizes other than 75 kDa, which could represent degradation products of phospho-IRE1α or non-specific antibody binding.Each sample was normalized to β-actin to minimize these differences, allowing for relative protein level comparisons between samples.

Figure 6 .
Figure 6.Western Blot results for endothelial nitric oxide syntase (eNOS).A. PVDF membrane with left ventricular samples from sham operated rats fed with the normal fat diet (NFD SO).B. PVDF membrane with left ventricular samples from rats with ligation of the coronary artery fed with the normal fat diet (NFD MI). C. PVDF membrane with left ventricular samples from sham operated rats fed with the normal fat diet (HFD SO).D. PVDF membranę with left ventricular samples from rats with ligation of the coronary artery fed with the normal fat diet.St -protein standard.The expected size of the detected eNOS protein was around 140 kDa.However, in our results, a band was observed at approximately 75 kDa.This discrepancy might be due to differences in protein migration during electrophoresis, post-translational modifications, or protein degradation during sample preparation.We also observed additional bands at sizes other than 75 kDa, which could represent degradation products of phospho-IRE1α or non-specific antibody binding.Each sample was normalized to β-actin to minimize these differences, allowing for relative protein level comparisons between samples.

Figure 7 .Figure 8 .
Figure 7.Western Blot results for inducible nitric oxide syntase (iNOS).A. PVDF membrane with left ventricular samples from sham operated rats fed with the normal fat diet (NFD SO).B. PVDF membrane with left ventricular samples from rats with ligation of the coronary artery fed with the normal fat diet (NFD MI). C. PVDF membrane with left ventricular samples from sham operated rats fed with the normal fat diet (HFD SO).D. PVDF membranę with left ventricular samples from rats with ligation of the coronary artery fed with the normal fat diet.St -protein standard.The expected size of the detected iNOS protein was around 130 kDa.However, in our results, a band was observed at approximately 75 kDa.This discrepancy might be due to differences in protein migration during electrophoresis, post-translational modifications, or protein degradation during sample preparation.We also observed additional bands at sizes other than 75 kDa, which could represent degradation products of phospho-IRE1α or non-specific antibody binding.Each sample was normalized to β-actin to minimize these differences, allowing for relative protein level comparisons between samples.

Figure 9 .
Figure 9.Western Blot results for protein kinase R-like endoplasmic reticulum kinase (PERK).A. PVDF membrane with left ventricular samples from sham operated rats fed with the normal fat diet (NFD SO).B. PVDF membrane with left ventricular samples from rats with ligation of the coronary artery fed with the normal fat diet (NFD MI). C. PVDF membrane with left ventricular samples from sham operated rats fed with the normal fat diet (HFD SO).D. PVDF membranę with left ventricular samples from rats with ligation of the coronary artery fed with the normal fat diet.St -protein standard.The expected size of the detected PERK protein was around 125 kDa.However, in our results, a band was observed at approximately 70 kDa.This discrepancy might be due to differences in protein migration during electrophoresis, post-translational modifications, or protein degradation during sample preparation.We also observed additional bands at sizes other than 70 kDa, which could represent degradation products of phospho-IRE1α or non-specific antibody binding.Each sample was normalized to β-actin to minimize these differences, allowing for relative protein level comparisons between samples.

Figure 10 .
Figure 10.Western Blot results for activating transcription factor 6 (ATF6).A. PVDF membrane with left ventricular samples from sham operated rats fed with the normal fat diet (NFD SO).B. PVDF membrane with left ventricular samples from rats with ligation of the coronary artery fed with the normal fat diet (NFD MI). C. PVDF membrane with left ventricular samples from sham operated rats fed with the normal fat diet (HFD SO).D. PVDF membranę with left ventricular samples from rats with ligation of the coronary artery fed with the normal fat diet.St -protein standard.The expected size of the detected ATF6 (non-cleaved) protein was around 90 kDa.However, in our results, a band was observed at approximately 70 kDa.This discrepancy might be due to differences in protein migration during electrophoresis, post-translational modifications, or protein degradation during sample preparation.We also observed additional bands at sizes other than 70 kDa, which could represent degradation products of phospho-IRE1α or non-specific antibody binding.Each sample was normalized to β-actin to minimize these differences, allowing for relative protein level comparisons between samples.