evaluating-cell-death-signaling-immunofluorescence-rat-model-ischemic

Evaluating Cell Death Signaling by Immunofluorescence in a Rat Model of Ischemic Stroke

Stroke is a leading cause of death and disability worldwide. Most cases of stroke are ischemic and result from the occlusion of the middle cerebral artery ...

Sichuan University

Angiogenesis plays an important role in tumor growth, invasion, and eventually metastasis. Antiangiogenic strategies have been proven to be a promising approach for clinical therapy for a variety of tumors. As a potent inhibitor of tumor angiogenesis, pigment epithelium-derived factor (PEDF) has recently been studied and used as an anticancer agent in several tumor models.

Anti-tumor effect of adenovirus-mediated gene transfer of pigment epithelium-derived factor on mouse B16-F10 melanoma.

Myocardial ischemia is a primary cause for the loss of vital components such as cardiomyocytes in the heart, leading to myocardial infarction and eventual cardiac dysfunction or heart failure. Suppressed angiogenesis plays a determinant role in the pathogenesis of myocardial infarction. In response to myocardial ischemia, hypoxia-inducible factor-1α and 2α (HIF-1α and HIF-2α) accumulate in cardiomyocytes and other cell types. This would up-regulate the expression of genes involved in angiogenesis such as vascular endothelial growth factor (VEGF); however, it is often observed that the angiogenic capacity is suppressed rather than enhanced. Ischemic toxicity, which has not been fully recognized, is highly responsible for the compromised angiogenic capacity. One of the toxic effects resulting from myocardial ischemia is the loss of copper content in the heart. Although the reason for this loss has not been elucidated, the essential role of copper in the regulation of HIF-1 transcriptional activity has been described. Copper does not affect the accumulation of HIF-1α in the cell, but is required for the HIF-1 transcriptional complex formation and its interaction with the hypoxia-responsive element in target genes. Copper supplementation can stimulate the transcriptional activity of HIF-1 and restore angiogenic capacity, leading to increased capillary density in the heart. The recognition of ischemic toxicity and the effort to overcome the toxic effect would help develop alternative approaches in the treatment of ischemic heart disease.

Ischemia-induced copper loss and suppression of angiogenesis in the pathogenesis of myocardial infarction.

We aimed to explore the association of serum level of the nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3) and its related inflammatory biomarkers (hypoxia inducible factor-1α, cathepsin B, caspase-1 and matrix metalloproteinase-9) with malignant brain edema (MBE) in patients with acute ischemic stroke.

Association between serum NLRP3 and malignant brain edema in patients with acute ischemic stroke.

It aims to analyze the influential mechanism of microRNA-9 (miR-9) and long non-coding RNA XIST (lncRNA XIST) expression on the proliferation and apoptosis of macrophages induced by oxidized-low density lipoprotein (ox-LDL). Firstly, lncRNA XIST overexpression vector was constructed, and then RAW264.7 cells were used as the research object. Methylthiazolyl tetrazolium (MTT) method, flow cytometry, and Western blot were used to detect and compare the differences of cell proliferation, apoptosis, and the expression levels of apoptosis signal-regulating kinase 1 (ASK1), c-Jun N-terminal kinase (JNK), matrix metalloproteinase-9 (MMP-9), and B-cell lymphoma-2 (Bcl-2) after ox-LDL induction and transfection of miR-9 mimic, miR-9 inhibitor and XIST expression vector, respectively. The results showed that lncRNA XIST overexpression vector was successfully constructed and transfected into cells, wh5ich can inhibit the expression level of miR-9. Compared with the normal control group, ox-LDL can inhibit cell proliferation, promote cell apoptosis, and increase the expression level of target protein. Moreover, transfection of XIST expression vector based on ox-LDL induction can significantly enhance the inhibition of cell proliferation, and promote cell apoptosis and the expression of target protein. Transfection of miR-9 mimic can improve the biological changes induced by ox-LDL. After co-transfection of miR-9 mimic and XIST expression vector based on ox-LDL induction, cell proliferation, apoptosis, and target protein expression level were not significantly different from those induced by ox-LDL alone. In summary, the increased expression level of miR-9 can inhibit the apoptosis of macrophages induced by ox-LDL. lncRNA XIST can positively regulate the apoptosis of macrophages induced by ox-LDL.

Regulatory Mechanism of MicroRNA-9 / Long Non-Coding RNA XIST Expression on Mouse Macrophage RAW264.7 Apoptosis Induced by Oxidized Low Density Lipoprotein.

To investigate whether serum calcium and magnesium on the day of symptom onset contribute to prognosis at 1 year after intracerebral hemorrhage (ICH).

Prognostic value of day-of-event serum calcium and magnesium for predicting 1-year prognosis after intracerebral hemorrhage.

Runt-related transcription factor-1 (RUNX1), a key member of the core-binding factor family of transcription factors, has emerged as a novel therapeutic target for cardiovascular disease. There is an urgent need to fully understand the expression pattern of Runx1 in the heart and the mechanisms by which it is controlled under normal conditions and in response to disease. The expression of Runx1 is regulated at the transcriptional level by two promoters designated P1 and P2. Alternative usage of these two promoters creates differential mRNA transcripts diversified in distribution and translational potential. While the significance of P1/P2 promoter-switch in the transcriptional control of Runx1 has been highlighted in the embryogenic process, very little is known about the level of P1- and P2-specific transcripts in adult hearts, and the underlying mechanisms controlling the promoter-switch.

Age-related promoter-switch regulates Runx1 expression in adult rat hearts.

Medial arterial calcification (MAC) accompanying chronic kidney disease (CKD) leads to increased vessel wall stiffness, myocardial ischemia, heart failure, and increased cardiovascular morbidity and mortality. Unfortunately, there are currently no drugs available to treat MAC. The natural polyphenol epigallocatechin-3-gallate (EGCG) has been demonstrated to protect against cardiovascular disease; however, whether EGCG supplementation inhibits MAC in CKD remains unclear. In this study, we utilize a CKD-associated MAC model to investigate the effects of EGCG on vascular calcification and elucidate the underlying mechanisms involved. Our findings demonstrate that EGCG treatment significantly reduces calcium phosphate deposition and osteogenic differentiation of VSMCs  and  in a dose-dependent manner. In addition, through RNA sequencing (RNA-seq) analysis, we show a significant activation of the transcription factor JunB both in CKD mouse arteries and in osteoblast-like VSMCs. Notably, EGCG effectively suppresses CKD-associated MAC by inhibiting the activity of JunB. In addition, overexpression of JunB can abolish while knockdown of  can enhance the inhibitory effect of EGCG on the osteogenic differentiation of VSMCs. Furthermore, EGCG supplementation inhibits MAC in CKD via modulation of the JunB-dependent Ras/Raf/MEK/ERK signaling pathway. In conclusion, our study highlights the potential therapeutic value of EGCG for managing CKD-associated MAC, as it mitigates this pathological process through targeted inactivation of JunB.

Epigallocatechin-3-gallate inhibits osteogenic differentiation of vascular smooth muscle cells through the transcription factor JunB.

Myocardial infarction (MI) results in prolonged ischemia and the subsequent cell death leads to heart failure which is linked to increased deaths or hospitalizations. New therapeutic targets are urgently needed to prevent cell death and reduce infarct size among patients with MI. Runt-related transcription factor-1 (RUNX1) is a master-regulator transcription factor intensively studied in the hematopoietic field. Recent evidence showed that RUNX1 has a critical role in cardiomyocytes post-MI. The increased RUNX1 expression in the border zone of the infarct heart contributes to decreased cardiac contractile function and can be therapeutically targeted to protect against adverse cardiac remodelling. This study sought to investigate whether pharmacological inhibition of RUNX1 function has an impact on infarct size following MI. In this work we demonstrate that inhibiting RUNX1 with a small molecule inhibitor (Ro5-3335) reduces infarct size in an in vivo rat model of acute MI. Proteomics study using data-independent acquisition method identified increased cathepsin levels in the border zone myocardium following MI, whereas heart samples treated by RUNX1 inhibitor present decreased cathepsin levels. Cathepsins are lysosomal proteases which have been shown to orchestrate multiple cell death pathways. Our data illustrate that inhibition of RUNX1 leads to reduced infarct size which is associated with the suppression of cathepsin expression. This study demonstrates that pharmacologically antagonizing RUNX1 reduces infarct size in a rat model of acute MI and unveils a link between RUNX1 and cathepsin-mediated cell death, suggesting that RUNX1 is a novel therapeutic target that could be exploited clinically to limit infarct size after an acute MI.

Pharmacological inhibition of RUNX1 reduces infarct size after acute myocardial infarction in rats and underlying mechanism revealed by proteomics implicates repressed cathepsin levels.

Atherosclerotic calcification often coincides with osteoporosis, suggesting a potential interplay between bone and vascular mineralization. Osteoblast-derived matrix vesicles (Ost-MVs), pivotal in bone mineralization, have emerged as potential contributors to ectopic vascular calcification. However, the precise role of Ost-MVs in vascular calcification and the underlying mechanisms remain elusive. In this study, we observed a concomitant increase in atherosclerotic calcification and bone loss, accompanied by elevated release of Ost-MVs into circulation. We demonstrate that circulating Ost-MVs target plaque lesions in the setting of atherosclerosis. Mechanistically, vascular injury facilitates transendothelial transport of Ost-MVs, collagen І remodeling promotes Ost-MVs aggregation, and vascular smooth muscle cell (VSMC) phenotypic switching enhances MV uptake. These pathological changes during atherosclerosis collectively contribute to Ost-MVs recruitment into the vasculature. Furthermore, Ost-MVs and VSMC-derived matrix vesicles (VSMC-MVs) exacerbate calcification via the Ras-Raf-ERK pathway. Our findings unveil a novel Ost-MVs-mediated mechanism participating in vascular calcification and enriching our understanding of bone-vascular crosstalk.

Weihong He

Department of Physiology,
West China School of Basic Medical Sciences and Forensic Medicine,
Department of Physiology, West China School of Basic Medical Sciences and Forensic Medicine

Sichuan University

Evaluating Cell Death Signaling by Immunofluorescence in a Rat Model of Ischemic Stroke

Weihong He

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Department of Physiology,West China School of Basic Medical Sciences and Forensic Medicine,Department of Physiology, West China School of Basic Medical Sciences and Forensic Medicine

Sichuan University

Evaluating Cell Death Signaling by Immunofluorescence in a Rat Model of Ischemic Stroke

Department of Physiology,
West China School of Basic Medical Sciences and Forensic Medicine,
Department of Physiology, West China School of Basic Medical Sciences and Forensic Medicine