Showing papers in "Journal of Molecular Histology in 2021"

Exosomes with high level of miR-181c from bone marrow-derived mesenchymal stem cells inhibit inflammation and apoptosis to alleviate spinal cord injury

Abstract: Stem cell transplantation is a promising method in the treatment of spinal cord injury (SCI). Researches have shown that stem cell-derived exosomes as well as its contents such as microRNAs contribute to the protective effects of stem cell against SCI. However, the effects of exosomes derived from bone marrow stem cells on SCI and the underlying mechanisms remain unknown. In this study, we collected bone marrow stem cells derived exosomes (BMSCs-exo) to deal with SCI rats and LPS induced microglia to explore the possible mechanisms. We found that BMSCs-exo showed significant effects on decreasing pro-inflammatory cytokines as well as increasing Basso-Beattie-Bresnahan score after acute SCI. MicroRNA-181c levels in tissue were elevated with the use of BMSCs-exo. Then we verified the effect in vitro and found that in LPS induced microglia, the administration of BMSCs-exo could inhibit the expression of pro-inflammatory cytokines, and the phosphorylation of NF-κB signal was also suppressed. During which, the expression of microRNA-181c in microglia was elevated. When LPS induced microglia were treated with BMSCs-exo over-expressing microRNA-181c, the levels of pro-inflammatory cytokines decreased. Then bioinformatics techniques were used to detect the possible target gene of microRNA-181c and then PTEN was found as a candidate. Further experiments showed that the protection effects of BMSCs-exo over-expressing microRNA-181c could be antagonized by the elevation of PTEN expression both in vitro and in vivo. In conclusion, we verified that BMSCs-exo could protect against SCI through its content microRNA-181c which suppressed the inflammation in microglia and spinal cord. It was related to the inhibition of PTEN and the suppression of NF-κB signal, and finally decreasing inflammation and apoptosis in spinal cord and improved SCI.

LncRNA KCNQ1OT1 acts as miR-216b-5p sponge to promote colorectal cancer progression via up-regulating ZNF146.

Abstract: Long non-coding RNAs (lncRNAs) have shown to act as important regulators in cancer biology. The aim of this study was to investigate the role and mechanism of lncRNA KCNQ1 opposite strand/antisense transcript 1 (KCNQ1OT1) in colorectal cancer (CRC) progression. The abundance of KCNQ1OT1, microRNA-216b-5p (miR-216b-5p) and zinc finger protein 146 (ZNF146) messenger RNA (mRNA) was measured by quantitative real-time polymerase chain reaction (qRT-PCR). Cell proliferation was analyzed by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and colony formation assay. Cell migration and invasion abilities were assessed by transwell assays. Western blot assay was performed for determination of protein levels. LncBase v.2 of DIANA Tool and StarBase software were used to predict the targets of KCNQ1OT1 and miR-216b-5p, respectively. Dual-luciferase reporter assay was implemented to confirm the target interaction between miR-216b-5p and KCNQ1OT1 or ZNF146. KCNQ1OT1 expression was higher in CRC tissues and cell lines. KCNQ1OT1 interference restrained the proliferation, migration and invasion of CRC cells. MiR-216b-5p was a target of KCNQ1OT1 in CRC cells, and KCNQ1OT1 knockdown-induced effects in CRC cells were partly overturned by miR-216b-5p silencing. MiR-216b-5p bound to the 3' untranslated region (3'UTR) of ZNF146, and ZNF146 overexpression partly attenuated miR-216b-5p overexpression-mediated influences in CRC cells. KCNQ1OT1 up-regulated the abundance of ZNF146 through sequestering miR-216b-5p in CRC cells. KCNQ1OT1 accelerated the proliferation and motility of CRC cells through elevating ZNF146 expression via sponging miR-216b-5p. KCNQ1OT1/miR-216b-5p/ZNF146 axis might be underlying target for the diagnosis and treatment of CRC patients.

Circular RNA RBM33 contributes to cervical cancer progression via modulation of the miR-758-3p/PUM2 axis

Abstract: Cervical cancer (CC) is a gynecological malignant tumor. Circular RNA (hsa_circ_0001772) (circRBM33) is implicated in the tumorigenesis of cancers. Nevertheless, the role of circRBM33 in CC is indistinct. Quantitative real-time polymerase chain reaction (qRT-PCR) was employed to evaluate the levels of circRBM33, miR-758-3p, and pumilio RNA binding family member 2 (PUM2) mRNA in tissue samples and cells. Cell proliferation, apoptosis, migration, invasion, and glycolysis were assessed using 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay, flow cytometry assay, transwell assay, or special commercial kits. Relative protein levels were examined via western blotting. The targeting relationship between circRBM33 or PUM2 and miR-758-3p was verified via dual-luciferase reporter or RNA pull-down assays. The role of circRBM33 was confirmed via tumor formation experiments. CircRPPH1 and PUM2 were upregulated while miR-758-3p was downregulated in CC tissues and cells. Functionally, circRBM33 knockdown constrained tumor growth in vivo and cured CC cell proliferation, migration, invasion, glycolysis, and fostered CC cell apoptosis in in vitro. Mechanistically, circRBM33 sponged miR-758-3p to modulate PUM2 expression. MiR-758-3p inhibitor neutralized circRBM33 silencing-mediated effects on the malignant behaviors of CC cells. PUM2 elevation overturned the suppressive influence of miR-758-3p upregulation on the malignant behaviors of CC cells. CircRBM33 fostered CC advancement via absorbing miR-758-3p and upregulating PUM2, indicating that circRBM33 was a possible target for CC treatment.

Circ_0091702 serves as a sponge of miR-545-3p to attenuate sepsis-related acute kidney injury by upregulating THBS2

Abstract: Circular RNA (circRNA) has been shown to play an important function in the progression of human diseases, including sepsis with acute kidney injury (AKI). However, the role and mechanism of circ_0091702 in sepsis-induced AKI have yet to be confirmed. Lipopolysaccharide (LPS) was used to induce HK2 cells to construct AKI cell models in vitro. Quantitative real-time PCR was used to measure the expression of circ_0091702, inflammatory cytokines, microRNA (miR)-545-3p and thrombospondin 2 (THBS2). Cell counting kit 8 assay and flow cytometry were used to assess cell viability and apoptosis. Besides, the protein levels of apoptosis markers and THBS2 were evaluated by western blot analysis. In addition, the concentrations of inflammatory cytokines were detected by enzyme-linked immunosorbent assay (ELISA). Cell oxidative stress was determined by detecting the contents of oxidative stress markers. Dual-luciferase reporter assay and RIP assay were used to confirm the relationship between miR-545-3p and circ_0091702 or miR-545-3p and THBS2. Circ_0091702 was downregulated in septic AKI patients and LPS-induced HK2 cells. Circ_0091702 could attenuate LPS-induced HK2 cell injury, while its silencing had an opposite effect. In the terms of mechanism, circ_0091702 could act as a sponge of miR-545-3p, and miR-545-3p could directly target THBS2. Functional experiments revealed that miR-545-3p could reverse the alleviating effect of circ_0091702 on LPS-induced HK2 cell injury, and THBS2 knockdown also could overturn the suppressing effect of miR-545-3p inhibitor on LPS-induced HK2 cell injury. Additionally, we also suggested that circ_0091702 could sponge miR-545-3p to regulate THBS2 expression. In conclusion, our results showed that circ_0091702 could suppress LPS-induced HK2 cell injury via the miR-545-3p/THBS2 axis, indicating that circ_0091702 might be an important biomarker for relieving sepsis-related AKI.

Pregnant women with COVID-19: the placental involvement and consequences.

Abstract: SARS-CoV-2, the leading cause of COVID-19 pandemic, was detected for the first time in Wuhan. In this study, we investigated the potential undesirable maternal and feto-neonatal consequences of COVID-19, and the related pathophysiological alterations in mother, neonate, and especially in the placenta as a vital organ, were reviewed. Also, the possibility of vertical transmission of virus and placental abnormalities were evaluated. The pregnant women were a vulnerable population for COVID-19, and several obstetric consequences were reported following SARS-CoV-2 infection. The higher risk of abruption, preterm labor, maternal death, stillbirth, intrauterine growth restriction, and newborns with fetal distress were adverse pregnancy and perinatal outcomes of COVID-19. Despite the ACE2 expression on placental components was confirmed, there is no agreement on the mother-child vertical transmission of this virus. Therefore, feto-neonatal consequences might be associated with placental abnormalities. The placental abnormalities are characterized by feto-maternal vascular malperfusion. Additionally, these adverse consequences lead to early termination of pregnancy in some cases, mostly via cesarean section. The pregnant women screening, coordination between healthcare personnel and neonatal unit, and infected women quarantine may decrease the risk of maternal and neonatal death after delivery.

Altered BMP-Smad4 signaling causes complete cleft palate by disturbing osteogenesis in palatal mesenchyme

Abstract: As the major receptor mediated BMP signaling in craniofacial development, Bmpr1a expression was detected in the anterior palatal shelves from E13.5 and the posterior palatal shelves from E14.5. However, inactivating BMP receptor in the mesenchyme only leads to anterior cleft palate or submucous cleft palate. The role of BMP signaling in posterior palatal mesenchyme and palatal osteogenesis is still unknown. In this study, a secreted BMP antagonist, Noggin was over-expressed by Osr2-creKI to suppress BMP signaling intensively in mouse palatal mesenchyme, which made the newborn mouse displaying complete cleft palate, a phenotype much severer than the anterior or submucous cleft palate. Immunohistochemical analysis indicated that in the anterior and posterior palatal mesenchyme, the canonical BMP-Smad4 signaling was dramatically down-regulated, while the non-canonical BMP signaling pathways were altered little. Although cell proliferation was reduced only in the anterior palatal mesenchyme, the osteogenic condensation and Osterix distribution were remarkably repressed in the posterior palatal mesenchyme by Noggin over-expression. These findings suggested that BMP-Smad4 signaling was essential for the cell proliferation in the anterior palatal mesenchyme, and for the osteogenesis in the posterior palatal mesenchyme. Interestingly, the constitutive activation of Bmpr1a in palatal mesenchyme also caused the complete cleft palate, in which the enhanced BMP-Smad4 signaling resulted in the premature osteogenic differentiation in palatal mesenchyme. Moreover, neither the Noggin over-expression nor Bmpr1a activation disrupted the elevation of palatal shelves. Our study not only suggested that BMP signaling played the differential roles in the anterior and posterior palatal mesenchyme, but also indicated that BMP-Smad4 signaling was required to be finely tuned for the osteogenesis of palatal mesenchyme.

LINC00152 mediates CD8+ T-cell infiltration in gastric cancer through binding to EZH2 and regulating the CXCL9, 10/CXCR3 axis.

Abstract: This study aimed to annotate the role of long intergenic non-coding RNA 152 (LINC00152) in CD8+ T cells mediated immune responses in gastric cancer (GC) and the underlying mechanism. LINC00152 expression levels were detected through RT-PCR. For tumor engraftment, HGC-27 cells that received LINC00152 shRNA, LINC00152 overexpression vectors, enhancer of zeste homolog 2 (EZH2) shRNA or combination transfection were injected into mice. Chromatin immunoprecipitation (ChIP) assay was used to explore the interaction between LINC00152, Cys-X-cys ligand 9 (CXCL9) and Cys-X-cys ligand 10 (CXCL10). Flow cytometry was adopted to measure the CD8+ T-cell infiltration in tumor issue. In this study, we found increased LINC00152 expression levels are positively associated with the poor prognosis of GC patients and negatively associated with the CD8 levels. ChIP assay verified that LINC00152 recruits EZH2 to the promoters of CXCL9 and CXCL10, thus the silencing of LINC00152 promoted the production of CXCL9 and CXCL10. Knockdown of LINC00152 suppressed tumor cells growth in vivo and in vitro, increased tumor-infiltrating CD8+ T cells numbers and promoted the expression of CXCL9, CXCL10 and C-X-C Motif Chemokine Receptor 3 (CXCR3) in xenograft tumors. While CD8+ T cell depletion reversed the tumor suppression effect of LINC00152 silence. Besides, the silencing of EZH2 partly inhibited the promotion effect LINC00152 on tumor growth. Our study indicated that LINC00152 inhibition suppressed the tumor progress may through promoting CD8+ T-cell infiltration.

LncRNA SNHG1 promotes neuronal injury in Parkinson’s disease cell model by miR-181a-5p/CXCL12 axis

Abstract: Small molecule RNA host gene 1 (SNHG1) has been found to be an important regulator in the neurotoxicity of Parkinson's disease (PD). However, the underlying molecular mechanisms of SNHG1 in PD remains elusive. The expression of SNHG1, microRNA (miR)-181a-5p, and C-X-C motif chemokine 12 (CXCL12) mRNA was detected using quantitative real-time polymerase chain reaction. Cell viability and apoptosis were analyzed by cell counting kit-8 and Flow cytometry, respectively. Western blot was utilized to determine the levels of B-cell lymphoma-2 (Bcl-2), CyclinD1, Cleaved-caspase-3, and CXCL12 protein. The interaction between miR-181a-5p and SNHG1 or CXCL12 was confirmed by the dual-luciferase reporter assay. We discovered that SNHG1 was significantly elevated, while miR-181a-5p was decreased in N-methyl-4-phenylpyridinium (MPP+)-treated neuroblastoma cells in dose-dependent manners. MPP+ induced cell viability inhibition and apoptosis promotion, while these effects were reversed by SNHG1 knockdown or miR-181a-5p re-expression. SNHG1 directly bound to miR-181a-5p, and miR-181a-5p inhibition could block the action of SNHG1 knockdown on MPP+-induced neurotoxicity in neuroblastoma cells. CXCL12 was identified as a downstream target of miR-181a-5p, and the impact of miR-181a-5p on MPP+-induced neuronal damage could be attenuated by CXCL12 overexpression. Besides, SNHG1 could indirectly regulate CXCL12 expression via miR-181a-5p. We demonstrated that SNHG1 promoted MPP+ induced neuronal injury in neuroblastoma cells by regulating miR-181a-5p/CXCL12 axis, suggesting SNHG1 might contribute to the development of PD, which provided a novel insight into the pathogenesis and treatment of PD.

Polydatin and polydatin-loaded chitosan nanoparticles attenuate diabetic cardiomyopathy in rats

Abstract: Hyperglycemia is associated with impairment of heart function. The current study aimed to investigate the ameliorative effect of polydatin-loaded chitosan nanoparticles (PD-CSNPs), polydatin (PD) and metformin (MET) on diabetic cardiomyopathy in rats. Rats divided into six groups; normal-control, diabetic-control, diabetic + CSNPs (diabetic rats treated with 50 mg/kg blank chitosan nanoparticles), diabetic + PD-CSNPs (diabetic rats treated with PD-CSNPs equivalent to 50 mg/kg of polydatin), diabetic + PD (diabetic rats given 50 mg/kg polydatin), diabetic + MET (diabetic rats given 100 mg/kg metformin), orally and daily for 4 weeks. Treatment of diabetic rats with PD-CSNPs, PD and MET showed a significant reduction in the values of glucose and glycosylated hemoglobin with improvement in heart function biomarkers through decreasing serum creatine kinase and creatine kinase myocardial band activities compared to diabetic control. The treatment agents also suppressed the elevated lipid peroxidation product, increased values of glutathione content, superoxide dismutase, superoxide peroxidase, and catalase activities in the heart of diabetic treated rats. Furthermore, PD-CSNPs, PD and MET decreased heart tissue levels of a pro-inflammatory cytokine; tumor necrosis factor-alpha and nuclear factor-kappa β, upregulation of heart gene expressions; nuclear factor erythroid 2-related factor 2 and heme oxygenase-1. Histological and ultrastructural examinations revealed the ameliorative effect of PD-CSNPs, PD and MET against the harmful of diabetic cardiomyopathy by reducing the cardiac fibers, necrotic cardiac myocytes, inflammatory cell infiltration, and the arrangement of the myofibrils and intercalated discs. In conclusion, the new formula of PD-CSNPs was more effective than PD and MET in amelioration the diabetic cardiomyopathy through its antioxidant, anti-inflammatory and prolonged-release properties.

Coenzyme Q10 enhances remyelination and regulate inflammation effects of cuprizone in corpus callosum of chronic model of multiple sclerosis

Abstract: Multiple Sclerosis (MS) is a chronic, progressive demyelinating disease of the central nervous system that causes the most disability in young people, besides trauma. Coenzyme Q10 (CoQ10)—also known as ubiquinone—is an endogenous lipid-soluble antioxidant in the mitochondrial oxidative respiratory chain which can reduce oxidative stress and inflammation, the processes associated with demyelination in MS. Cuprizone (CPZ) intoxication is a well-established model of inducing MS, best for studying demyelination—remyelination. In this study, we examined for the first time the role of CoQ10 in preventing demyelination and induction of remyelination in the chronic CPZ model of MS. 40 male mice were divided into four groups. 3 group chewed CPZ-containing food for 12 weeks to induce MS. After 4 weeks, one group were treated with CoQ10 (150 mg/kg/day) by daily gavage until the end of the experiment, while CPZ poisoning continued. At the end of 12 weeks, tail suspension test (TST) and open field test (OFT) was taken and animals were sacrificed to assess myelin basic protein (MBP), oligodendrocyte transcription factor-1 (Olig1), tumor necrosis factor-α (TNF-α) and interleukin 6 (IL-6) by real-time polymerase chain reaction (real-time PCR) and total antioxidant capacity (TAC) and superoxide dismutase (SOD) by Elisa test. Luxol fast blue (LFB) staining was used to evaluate histological changes. CoQ10 administration promoted remyelination in histological findings. MBP and Olig-1 expression were increased significantly in CoQ10 treated group compare to the CPZ-intoxicated group. CoQ10 treatment alleviated stress oxidative status induced by CPZ and dramatically suppress inflammatory biomarkers. CPZ ingestion made no significant difference between normal control group and the CPZ-intoxicated group in TST and OFT. CoQ10 can enhance remyelination in the CPZ model and potentially might have same effects in MS patients.

Correlation between the immune checkpoints and EMT genes proposes potential prognostic and therapeutic targets in ESCC

Abstract: PD-1, PD-L1, CTLA-4, TIM-3, and LAG-3, crucial immune checkpoint molecules in the tumor microenvironment, identify as key targets for cancer immunotherapy. There is a correlation between immune cells and epithelial-mesenchymal transition (EMT)-related genes expression in varies human cancers. In this study, we aimed to investigate the probable association between expression of immune checkpoints and EMT in esophageal squamous cell carcinoma (ESCC) with clinical treats for providing the new therapeutic targets and prognostic value for the disease. Quantitative real-time PCR was used to investigate the gene expression profile of immune checkpoints (PD-1, PD-L1, CTLA-4, TIM-3, and LAG-3) and EMT (TWIST1 and MMP-13) genes based on the mRNA expression levels in 51 ESCC tissues. The upregulation of CTLA-4, PD-1, PD-L1, TIM-3, LAG-3, MMP-13, and TWIST1 were observed in 31.37%, 29.41%, 21.56%, 39.21%, 25.49%, 60.78%, and 56.86% of ESCC cases at the mRNA level, respectively. Dysregulation of immune checkpoints was related to lymph node involvement, stage of tumor progression, and depth of tumor invasion (P < 0.05). While overexpression of MMP-13 and TWIST1 was associated with lymph node involvement, stage of tumor progression, and grade of tumor differentiation (P < 0.05). The mRNA expression of immune checkpoint genes was significantly correlated to each other's (P = 0.000). Of importance, the data explored the significant association between the concomitant expression of immune checkpoints and EMT-related genes with each other in a variety of clinicopathological traits (P < 0.05). Consequently, immune checkpoints were positively correlated with EMT status in ESCC. The correlation between tumor immune microenvironment with the elevation of multiple immune checkpoints and EMT status may help to identify potential biomarkers for the simultaneous clinical use of multiple immune checkpoints blockade and other immunotherapies approaches for advanced ESCC patients.

COVID-19 and male reproductive system: pathogenic features and possible mechanisms.

Abstract: Multiorgan dysfunction is the main characteristic of severe COVID-19 patients and the involvement of male reproductive system may occur among these patients. Although there is a limited evidence to confirm the orchitis and virus presence in the semen of patients, there are concerns about the transmission of virus through the semen. In addition, reduced fertility or infertility can be seen as consequences of severe COVID-19 in recovered subjects. In this study, we aimed to review articles related to COVID-19 and male reproductive system to find the possible underlying mechanisms of SARS-CoV-2 in affecting male fertility. The following keywords of SARS-CoV-2, COVID-19, testis, orchitis, semen, angiotensin-converting enzyme 2 (ACE2), hypothalamic-pituitary-testicular (HPT) axis, Hypothalamus, etc., were defined to find the related publications from standard search engines, e.g., PUBMED, SCOPUS, Google Scholar. According to studies, COVID-19 occurs in severe patients as respiratory disease, along with multi-organ failure. The most important mechanisms are classified as direct and indirect pathogenesis of SARS-CoV-2. The presence of ACE2 on the cell surface of various cells in testis increases the risk of direct infection by this virus. SARS-CoV-2 also affects the testis through the cytokine storm. In addition, the important role of HPT axis dysregulation through impaired Leydig cells and hypothalamus should be considered. Using antiviral and immunomodulatory therapy can be harmful for testis function. Further investigations are required to investigate potential mechanisms of male infertility in survivals of COVID-19. Since involvement of testis is essential for fertility, increasing the knowledge of health system may improve the outcomes.

LncRNA TRHDE-AS1 inhibit the scar fibroblasts proliferation via miR-181a-5p/PTEN axis

Abstract: Hypertrophic scar (HS), a fibroproliferative disorder caused by abnormal wound healing after skin injury, which is characterized by excessive deposition of extracellular matrix and invasive growth of fibroblasts. Recent studies have shown that some non-coding RNA implicated the formation of HS, but the mechanism remains unclear. In this study, we found that lncRNA TRHDE-AS1 was downregulated in HS tissues and HSFs, and the level of lncRNA TRHDE-AS1 negatively correlated with the level of miR-181a-5p in HS tissue and HSFs. Overexpressed lncRNA TRHDE-AS1 significantly suppressed miR-181a-5p level, while promoted HSFs apoptosis and inhibited HSFs proliferation. Further study shown that PTEN was a direct target of miR-181a-5p, and lncRNA TRHDE-AS1 served as a molecular sponge for miR-181a-5p to regulate the expression of PTEN. Overexpression of PTEN could eliminate lncRNA TRHDE-AS1-mediated proliferation suppression of HSFs. In conclusion, our study suggested that lncRNA TRHDE-AS1/miR-181a-5p/PTEN axis plays an important role in promoting hypertrophic scar formation, which may be effectively used as a therapeutic target for hypertrophic scar treatment.

Long noncoding HOXA11-AS knockdown suppresses the progression of non-small cell lung cancer by regulating miR-3619-5p/SALL4 axis

Abstract: Accumulating evidence suggested that many long noncoding RNAs (lncRNAs) were widely involved in the development and progression of non-small cell lung cancer (NSCLC). However, the roles of lncRNA homeobox A11 antisense (HOXA11-AS) and its underlying mechanism in NSCLC remains largely unknown. The expression levels of HOXA11-AS, miR-3619-5p and sal-like protein 4 (SALL4) were detected by quantitative real-time polymerase chain reaction (qRT-PCR). Western blot analysis was used to measure the protein levels of hexokinase II (HK2) and SALL4. Cell proliferation, apoptosis, migration and invasion were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, flow cytometry and transwell assay, respectively. The glucose consumption and lactate production were measured using glucose assay kit and lactate assay kit, respectively. The potential binding sites between miR-3619-5p and HOXA11-AS or SALL4 were predicted by online software and verified by luciferase report assay. A xenograft tumor model was established to confirm the function of HOXA11-AS in NSCLC in vivo. HOXA11-AS and SALL4 were upregulated while miR-3619-5p was downregulated in NSCLC tissues and cells. HOXA11-AS knockdown suppressed cell proliferation, migration, invasion, and glycolysis but promoted apoptosis in NSCLC cells. Moreover, miR-3619-5p could directly bind to HOXA11-AS and its inhibition attenuated the inhibitory effect of HOXA11-AS knockdown on progression of NSCLC cells. Furthermore, SALL4 was a direct target of miR-3619-5p and its overexpression reversed the anti-tumor role of miR-3619-5p in NSCLC cells. Besides, HOXA11-AS modulated SALL4 expression via sponging miR-3619-5p. Additionally, silencing HOXA11-AS inhibited tumor growth though upregulating miR-3619-5p and downregulating SALL4. Collectively, HOXA11-AS knockdown inhibited the progression of NSCLC by regulating miR-3619-5p/SALL4 axis, which might offer a novel avenue for interpreting the mechanism of NSCLC development.

TRIP13 promotes the proliferation and invasion of lung cancer cells via the Wnt signaling pathway and epithelial–mesenchymal transition

Abstract: Thyroid hormone receptor interactor 13 (TRIP13) is an ATPase that has been found to be overexpressed in many tumors. The aim of this study was to investigate the role of TRIP13 and its mechanism of action in lung cancer. The expression of TRIP13 was examined in lung cancer tissues and corresponding normal lung tissues by western blotting. TRIP13 was overexpressed or knocked down by transient transfection or siRNA interference in lung cancer cells, respectively. The expression of key proteins associated with the Wnt signaling pathway and epithelial-mesenchymal transition (EMT) was assessed. The interaction between TRIP13 and low-density lipoprotein receptor-related protein 6 (LRP6) was examined by co-immunoprecipitation and laser confocal immunofluorescence. Moreover, this study determined the proliferative and invasive ability of cells through colony formation, cell proliferation, and Matrigel invasion assays. The expression of TRIP13 was higher in lung cancer tissues than in normal lung tissues (p = 0.002), and this correlated with poor patient prognosis (p < 0.001). In addition, overexpression of TRIP13 enhanced the levels of active β-catenin and target proteins of the Wnt signaling pathways (p < 0.05). This study found that TRIP13 can co-localize and bind with LRP6. Furthermore, overexpression of TRIP13 caused the upregulation of N-cadherin, Snail, and vimentin, and the downregulation of E-cadherin (p < 0.05). The aforementioned results were reversed after knocking down the expression of TRIP13 (p < 0.05). TRIP13 is highly expressed in lung cancers, indicating poor prognosis. overexpression of TRIP13 promotes the proliferative and invasive ability of lung cancer cells via the activation of Wnt signaling pathway and EMT.

SUMO modification in apoptosis.

Abstract: Apoptosis and clearance of dead cells is highly evolutionarily conserved from nematode to humans, which is crucial to the growth and development of multicellular organism. Fail to remove apoptotic cells often lead to homeostasis imbalance, fatal autoimmune diseases, and neurodegenerative diseases. Small ubiquitin-related modifiers (SUMOs) modification is a post-translational modification of ubiquitin proteins mediated by the sentrin-specific proteases (SENPs) family. SUMO modification is widely involved in many cellular biological process, and abnormal SUMO modification is also closely related to many major human diseases. Recent researches have revealed that SUMO modification event occurs during apoptosis and clearance of apoptotic cells, and plays an important role in the regulation of apoptotic signaling pathways. This review summarizes some recent progress in the revelation of regulatory mechanisms of these pathways and provides some potential researching hotpots of the SUMO modification regulation to apoptosis.

Transferrin improved the generation of cardiomyocyte from human pluripotent stem cells for myocardial infarction repair

Abstract: Human pluripotent stem cell (hPSC)-derived cardiomyocytes (CMs) hold great promise for the repair of the injured heart, but optimal cell production in a fully chemically defined and cost-effective system is essential for the efficacy and safety of cell transplantation therapies. In this study, we provided a simple and efficient strategy for cardiac differentiation from hPSCs and performed functional evaluation in a rat model of myocardial infarction. Using a chemically defined medium including four components, recombinant human albumin, ascorbic acid, human transferrin, and RPMI 1640, we developed a manageable and cost-effective protocol for robust generation of CMs from hPSCs. Interestingly, the addition of transferrin helped hPSCs to transit from TeSR-E8 medium to the simple cardiac differentiation medium and successfully initiated mesoderm differentiation without significant cell death. The CM generation efficiency was up to 85% based on cTnT expression. We performed transcriptome profiling from differentiation day 0 to 35, and characterized interesting dynamic change of cardiac genes. CMs derived from transferrin-supplemented simple medium have similar transcriptome and the maturation level compared to those generated in B27 minus insulin medium as well as their in vivo counterparts. Importantly, after transplantation, hPSC-derived CMs survived in the infarcted rat heart, significantly improved the physiological function and reduced fibrosis. Our study offers an easy-to-use and cost-effective method for cardiac differentiation and facilitates the translational application of hPSC-derived CMs for heart repair.

Nogo-A-Δ20/EphA4 interaction antagonizes apoptosis of neural stem cells by integrating p38 and JNK MAPK signaling.

Abstract: Nogo-A protein consists of two main extracellular domains: Nogo-66 (rat amino acid [aa] 1019-1083) and Nogo-A-Δ20 (extracellular, active 180 amino acid Nogo-A region), which serve as strong inhibitors of axon regeneration in the adult CNS (Central Nervous System). Although receptors S1PR2 and HSPGs have been identified as Nogo-A-Δ20 binding proteins, it remains at present elusive whether other receptors directly interacting with Nogo-A-Δ20 exist, and decrease cell death. On the other hand, the key roles of EphA4 in the regulation of glioblastoma, axon regeneration and NSCs (Neural Stem Cells) proliferation or differentiation are well understood, but little is known the relationship between EphA4 and Nogo-A-Δ20 in NSCs apoptosis. Thus, we aim to determine whether Nogo-A-Δ20 can bind to EphA4 and affect survival of NSCs. Here, we discover that EphA4, belonging to a member of erythropoietin-producing hepatocellular (Eph) receptors family, could be acting as a high affinity ligand for Nogo-A-Δ20. Trans-membrane protein of EphA4 is needed for Nogo-A-Δ20-triggered inhibition of NSCs apoptosis, which are mediated by balancing p38 inactivation and JNK MAPK pathway activation. Finally, we predict at the atomic level that essential residues Lys-205, Ile-190, Pro-194 in Nogo-A-Δ20 and EphA4 residues Gln-390, Asn-425, Pro-426 might play critical roles in Nogo-A-Δ20/EphA4 binding via molecular docking.

Histological and imunohistochemical alterations of hippocampus and prefrontal cortex in a rat model of Alzheimer like-disease with a preferential role of the flavonoid “hesperidin”

Abstract: Alzheimer’s disease (AD) is a chronic age-related neurodegenerative disease characterized by degeneration of the central cholinergic neurons, inflammation and oxidative stress in the basal forebrain, prefrontal cortex and hippocampus. Hesperidin (Hesp) is one of the flavonoids havinganti-inflammatory and anti-oxidative properties in some neurodegerative brain lesions. To investigate the possible neuroprotective role of Hespin an AD-like rat model induced experimentally by Scopolamine (Scop). Forty adult male Sprague Dawley rats were randomly allocated into four groups. Group I—(Control), group II—(Hesp) (supplemented orally with 100 mg/kg Hesp for 28 days), group III—(AD) (injected i.p with 1 mg/kg Scop for 9 days) and group IV—(Hesp/AD). At the end of the experiment, behavioral (Y-maze test) and biochemical analysis were carried out along with histological, immunohistochemical and morphometric studies of the hippocampus and prefrontal cortex. AD rats displayed memory impairment in the behavioural paradigm with a concomitant increase of serum TNF-α and IL-1β, while IL-10 decreased significantly. Also, there was a rise of amyloid beta-42 (Aβ-42), acetylcholinesterase (AChE) activity and malondialdehyde (MDA) together with a decrease of reduced glutathione (GSH) in hippocampal and prefrontal homogenate. In addition, sections of the hippocampus and prefrontal cortex revealed obvious histopathological changes, overexpression of p-Tau protein and glial fibrillary acidic protein (GFAP) with a decrease in the expression of synaptophysin (SYN). Contradictorily, pre-treatment with Hesp offset the spatial memory deficits, redox imbalance, Aβ-42 and AChE over activity as well as preserved the histological architecture and attenuated the raised p-Tau protein and GFAP while upregulated SYN immuoreactivity of AD rats. Collectively, our results highlight the potential mitigating role of Hesp in AD-like state in rats and this may presumably raise the possibility of its future implementation as a prophylactic remedy against AD in humans.

Circ_PRKDC knockdown promotes skin wound healing by enhancing keratinocyte migration via miR-31/FBN1 axis

Abstract: Circular RNA protein kinase, DNA-activated, catalytic subunit (circ_PRKDC) has been found to impede wound healing in diabetic foot ulcers via regulating keratinocyte proliferation and migration. However, the mechanisms underlying circ_PRKDC in skin wound healing remain unclear. The expression of circ_PRKDC, microRNA (miR)-31 and fibrillin 1 (FBN1) was detected using quantitative reverse transcription-polymerase chain reaction and Western blot assays. The migration ability and the changes of matrix metallopeptidase 9 (MMP-9) and MMP2 levels were determined using wound healing, transwell and Western blot assays. The interaction between miR-31 and circ_PRKDC or FBN1 was verified by dual-luciferase reporter assay. The expression of circ_PRKDC was gradually down-regulated in wound edge at 1 and 7 days after injury relative to the unwounded skin. In human epidermal keratinocytes (HEKa), knockdown of circ_PRKDC promoted cell migration partly through up-regulating MMP-2 and MMP9, while circ_PRKDC overexpression showed opposite effects. In a mechanical study, we confirmed that miR-31 was a target of circ_PRKDC, and inhibition of miR-31 reversed the promotive effect of circ_PRKDC knockdown on HEKa migration. Besides that, miR-31 was verified to target FBN1, and ectopic overexpression of miR-31 accelerated HEKa migration via FBN1. Importantly, we also demonstrated that FBN1 overexpression attenuated the effects of circ_PRKDC knockdown on HEKa migration. In all, circ_PRKDC knockdown promoted HEKa migration during wound healing through miR-31/FBN1 axis, suggesting the therapeutic potential for circ_PRKDC on skin wound healing.

Effect of endoplasmic reticulum stress-induced apoptosis in the role of periodontitis on vascular calcification in a rat model.

Abstract: The present study aimed to investigate the mechanism(s) through which endoplasmic reticulum stress (ERS)-induced apoptosis, in the role of periodontitis, affects vascular calcification. Rat models of periodontitis, vascular calcification, periodontitis-vascular calcification, and a normal group were established. Cardiovascular tissues were obtained, and hematoxylin–eosin staining was applied to demonstrate the morphological changes in vascular tissues. Immunohistochemical staining was applied to analyze apoptosis in cardiovascular tissues. The expression levels of apoptotic factor cysteinyl aspartate specific proteinase 3 (Caspase-3), ERS-induced apoptotic factors glucose-regulated protein 78 (GRP78), 94 (GRP94), and ERS-induced apoptosis pathways Caspase-12, C/EBP homologous protein (CHOP), and c-Jun N-terminal kinase (JNK) were analyzed and compared. Hematoxylin–eosin staining revealed that the arterial layers in the normal group were structurally intact. The structural damage to the aortic wall gradually aggravated from the periodontitis group to the vascular calcification group to the combined group. The immunohistochemistry results showed Caspase-3, GRP78, GRP94, and ERS-induced apoptosis pathways in the cardiovascular tissues cells in the periodontitis group, vascular calcification group, and combined group. The Caspase-3, GRP78, GRP94, and CHOP expression levels in the combined group were significantly higher than that in the normal group (P 0.05). Apoptosis induced by ERS is involved in the effect of periodontitis on vascular calcification and might be mainly achieved through the activation of the CHOP transcription pathway.

Salivary gland proteins alterations in the diabetic milieu

Abstract: Salivary glands are considered the chief exocrine glands of the mouth and physiologically contribute to the maintenance of the homeostasis of the oral cavity. They consist of the parotid, submandibular and sublingual glands, which come in pairs and are collectively called the major glands, and the minor glands, which are much smaller and are dispersed throughout the buccal cavity. Salivary glands are distinguished by their size, amount of saliva secretion and their location in the oral cavity. Salivary glands pathophysiology has been a subject of interest in various worldwide metabolic disorders, including diabetes mellitus. Diabetes mellitus (DM), a global health concern, with a pathological imprint involved in vasculature, promotes microvascular and macrovascular complications among which periodontitis ranks sixth. Indeed, DM has also been directly associated with oral health lesions. Specifically, salivary glands in the context of diabetes have been a focal point of study and emphasis in the research field. There is evidence that relates salivary secretion content and diabetes progression. In this review, we present all the reported evidence of the deregulation of specific salivary proteins associated with the progression of diabetes in parallel with changes in salivary gland morphology, cellular architecture, and salivary secretion and composition more generally.

Long non-coding RNA TDRG1 promotes hypoxia-induced glycolysis by targeting the miR-214-5p/SEMA4C axis in cervical cancer cells

Abstract: Long non-coding RNA (lncRNA) has been demonstrated as vital regulator in human cancer. However, the precise role of lnc-TDRG1 in cervical cancer (CC) remains unclear, so this study was aimed to clarify the role and underlying molecular mechanism of lnc-TDRG1 in CC. The real-time quantitative polymerase chain reaction (RT-qPCR) was conducted to assess the expression levels of lnc-TDRG1, miR-214-5p and Semaphorin 4C (SEMA4C). Under hypoxia condition, the biological behaviors of CC cell, including invasion and glycolysis were determined by transwell assay and Glucose Assay Kit and Lactate Assay Kit, respectively. The Western blot assay was employed to test the expression level of SEMA4C and hexokinase 2 (HK2) expression. The interaction relationship between miR-214-5p and lnc-TDRG1 or SEMA4C was analyzed bioinformatics database and confirmed by dual-luciferase reporter assay, respectively. A xenograft experiment in nude mice was established to clarify the functional role of lnc-TDRG1 in vivo. We found Lnc-TDRG1 was highly expressed in CC tissues and cells and it was upregulated in response to hypoxia. Loss-of-functional experiment suggested that knockdown of lnc-TDRG1 impede invasion, hypoxia-induced glycolysis in vitro and tumor growth in vivo, which was abolished by knockdown of miR-214-5p or overexpression of SEMA4C. Moreover, we confirmed that miR-214-5p specifically bound to SEMA4C and negatively correlated with SEMA4C expression. Collectively, lnc-TDRG1 regulated SEMA4C expression by sponging miR-214-5p in CC. Collectively, mechanistically, lnc-TDRG1 could act as a sponge of miR-214-5p to regulate the expression of SEMA4C, and further regulate invasion and hypoxia-glycolysis in CC cells.

Salivary exosomes as a new therapy to ameliorate diabetes mellitus and combat xerostomia and submandibular salivary glands dysfunction in diabetic rats

Abstract: Diabetes mellitus (DM) is one of the major metabolic diseases. Xerostomia and salivary gland dysfunction are of its common oral complications. Exosomes, as a new therapeutic potential containing nucleic acids, proteins and lipids, act as effective vehicles for target molecules delivery. Accordingly, their therapeutic use is gaining much interest. Therefore, this work aimed to assess the therapeutic efficacy of salivary exosomes in ameliorating DM and combating xerostomia as a complication of salivary gland dysfunction in diabetic rats. In the current study, salivary exosomes were injected intravenously to rats of group II (Salivary Exo-treated group) one week after diabetes induction. Group I (Diabetic group) was left untreated. Blood sugar level was checked weekly. Water intake, salivary flow rate, salivary amylase and serum nitric oxide were assessed before and after diabetes induction and at the end of the study. After 5 weeks from the beginning of the study, salivary gland tissues were dissected and examined histologically and ultrastructurally. Gene expression of the inflammatory markers NFκB/p65 and TNFα was assessed by polymerase chain reaction. The results showed that salivary exosomes reduced blood glucose levels and enhanced salivary glands’ function. This was indicated by a decrease in water intake, salivary amylase and serum nitric oxide in addition to an increase in salivary flow rate. This was confirmed histologically, ultrastructurally and via downregulation of NFκB/p65 and TNFα gene expression. Our results concluded that salivary exosomes could be considered as a novel cell free based therapy in treatment of xerostomia and salivary gland dysfunction in DM.

MiR-590-5p inhibits pathological hypertrophy mediated heart failure by targeting RTN4.

Abstract: Heart failure (HF) is a rising epidemic and public health burden in modern society. It is of great need to find new biomarkers to ensure a timely diagnosis and to improve treatment and prognosis of the disease. The mouse model of HF was established by thoracic aortic constriction. Color Doppler ultrasound was performed to detect left ventricular end-diastolic diameter. Hematoxylin and eosin staining was conducted to observe the pathological changes of mouse myocardium. The RT-qPCR analysis was performed to detect miR-590-5p and RTN4 expression levels. Western blot was conducted to detect protein levels of the indicated genes. We found that the expression of miR-590-5p was downregulated in cardiac tissues of HF mice. Injection of AAV-miR-590-5p attenuated myocardium hypertrophy and myocyte apoptosis. Additionally, miR-590-5p overexpression promoted viability, inhibited apoptosis, and decreased ANF, BNP and beta-MHC protein levels in H9c2 cell. Mechanistically, miR-590-5p binds to RTN4 3'-untranslated region, as predicted by starBase online database and evidenced by luciferase reporter assay. Furthermore, miR-590-5p negatively regulates RTN4 mRNA expression and suppresses its translation. The final rescue experiments revealed that miR-590-5p modulated cardiomyocyte phenotypes by binding to RTN4. In conclusion, miR-590-5p modulates myocardium hypertrophy and myocyte apoptosis in HF by downregulating RTN4.

SDF-1 secreted by mesenchymal stem cells promotes the migration of endothelial progenitor cells via CXCR4/PI3K/AKT pathway

Abstract: Cell-based therapeutics bring great hope in areas of unmet medical needs. Mesenchymal stem cells (MSCs) have been suggested to facilitate neovascularization mainly by paracrine action. Endothelial progenitor cells (EPCs) can migrate to ischemic sites and participate in angiogenesis. The combination cell therapy that includes MSCs and EPCs has a favorable effect on ischemic limbs. However, the mechanism of combination cell therapy remains unclear. Herein, we investigate whether stromal cell-derived factor (SDF)-1 secreted by MSCs contributes to EPC migration to ischemic sites via CXCR4/Phosphoinositide 3-Kinases (PI3K)/protein kinase B (termed as AKT) signaling pathway. First, by a “dual-administration” approach, intramuscular MSC injections were supplemented with intravenous Qdot® 525 labeled-EPC injections in the mouse model of hind limb ischemia. Then, the mechanism of MSC effect on EPC migration was detected by the transwell system, tube-like structure formation assays, western blot assays in vitro. Results showed that the combination delivery of MSCs and EPCs enhanced the incorporation of EPCs into the vasculature and increased the capillary density in mouse ischemic hind limb. The numbers of CXCR4-positive EPCs increased after incubation with MSC-conditioned medium (CM). MSCs contributed to EPC migration and tube-like structure formation, both of which were suppressed by AMD3100 and wortmannin. Phospho-AKT induced by MSC-CM was attenuated when EPCs were pretreated with AMD3100 and wortmannin. In conclusion, we confirmed that MSCs contributes to EPC migration, which is mediated via CXCR4/PI3K/AKT signaling pathway.

LINC01515 promotes nasopharyngeal carcinoma progression by serving as a sponge for miR-325 to up-regulate CDCA5.

Abstract: Long non-coding RNAs (LncRNAs) have gained widespread interest and attention as vital regulators in cancer occurrence and development. Nonetheless, the functions and mechanisms of lncRNAs involved in nasopharyngeal carcinoma (NPC) are largely unknown. By analysing the data from GSE61218, we identified a novel lncRNA LINC01515 which is altered in NPC. A series of experiments were performed to examine the exact roles of LINC01515 as well as the molecular mechanisms by which LINC01515 acted in NPC. LINC01515 expression was increased in NPC and that high LINC01515 expression was associated with a worse prognosis. Functionally, depletion of LINC01515 resulted in an inhibition of cell proliferation, migration and invasion, while apoptosis was promoted. Mechanistically, LINC01515 facilitated cell division cycle associated 5 (CDCA5) expression via serving as a sponge for miR-325. And more notably, miR-325 suppressed NPC progression in vitro by targeting CDCA5. Furthermore, the anti-tumor property induced by LINC01515 knockdown was partially reversed due to the overexpression of CDCA5. Taken together, LINC01515 exerted the carcinogenic effect in NPC through regulating miR-325/CDCA5 pathway. Our findings shed light on the possibility of exploiting LINC01515 as a prognostic biomarker or therapeutic target in NPC.

Semaphorin 3A promotes the osteogenic differentiation of rat bone marrow-derived mesenchymal stem cells in inflammatory environments by suppressing the Wnt/β-catenin signaling pathway

Abstract: After periodontal treatment, the local inflammatory environment surrounding periodontal tissues cannot be entirely eliminated The means by which alveolar bone repair and regeneration are promoted in inflammatory environments have important clinical significance As a powerful protein that promotes the differentiation of osteocytes, semaphorin 3A (Sema3A) shows potential for bone regeneration therapy However, the effect of Sema3A on osteogenic differentiation in an inflammatory environment, as well as the underlying mechanism, have not yet been explored We used lentivirus to transduce rat bone marrow-derived mesenchymal stem cells (rBMSCs) to stably overexpress Sema3A Lipopolysaccharide from Escherichia coli (E coli LPS) was used to stimulate rBMSCs to establish an inflammatory environment ALP staining, Alizarin red staining, ALP activity tests, quantitative RT-PCR (qRT-PCR), and Western blotting were used to elucidate the effect of Sema3A on the osteogenesis of rBMSCs in inflammatory environments XAV939 and LiCl were used to determine whether the Wnt/β-catenin signaling pathway was involved in attenuating the inhibition of Sema3A-induced osteogenic differentiation by LPS The qRT-PCR and Western blot results demonstrated that the lentiviral vector (LV-NC) and lentiviral-Sema3A (LV-Sema3A) were successfully transduced into rBMSCs An inflammatory environment could be established by stimulating rBMSCs with 1 μg/ml E coli LPS After Sema3A overexpression, mineral deposition was exacerbated, and the BSP and Runx2 gene and protein expression levels were increased Furthermore, E coli LPS activated the Wnt/β-catenin signaling pathway and decreased rBMSC osteogenesis, but these effects were attenuated by Sema3A In conclusion, Sema3A could protect BMSCs from LPS-mediated inhibition of osteogenic differentiation in inflammatory environments by suppressing the Wnt/β-catenin pathway

The role of MYB proto-oncogene like 2 in tamoxifen resistance in breast cancer.

Abstract: Despite the efficacy of tamoxifen in preventing disease relapse, a large portion of breast cancer patients show intrinsic or acquired resistance to tamoxifen, leading to treatment failure and unfavorable clinical outcome. MYB proto-oncogene like 2 (MYBL2) is a transcription factor implicated in the initiation and progression of various human cancers. However, its role in tamoxifen resistance in breast cancer remained largely unknown. In the present study, by analyzing public transcriptome dataset, we found that MYBL2 is overexpressed in breast cancer and is associated with the poor prognosis of breast cancer patients. By establishing tamoxifen-resistant breast cancer cell lines, we also provided evidence that MYBL2 overexpression contributes to tamoxifen resistance by up-regulating its downstream transcriptional effectors involved in cell proliferation (PLK1, PRC1), survival (BIRC5) and metastasis (HMMR). In contrast, inhibiting those genes via MYBL2 depletion suppresses cancer progression, restores tamoxifen and eventually reduces the risk of disease recurrence. All these findings revealed a critical role of MYBL2 in promoting tamoxifen resistance and exacerbating the progression of breast cancer, which may serve as a novel therapeutic target to overcome drug resistance and improve the prognosis of breast cancer patients.

LGR4 silence aggravates ischemic injury by modulating mitochondrial function and oxidative stress via ERK signaling pathway in H9c2 cells.

Abstract: It is reported that LGR4 (leucine-rich repeat domain containing G protein-coupled receptor 4) plays a crucial role in the physiological function of many organs. However, few data are available on the function and mechanism of LGR4 in myocardial ischemia-reperfusion (I/R) injury. The aim of this study was to explore the function and mechanism of LGR4 in I/R injury. We incubated H9c2 cells in simulating ischemia buffer and then re-incubated them in normal culture medium to establish a model of I/R injury in vitro. The expression of LGR4 was evaluated by RT-PCR and western blot. Besides, the cell apoptosis was evaluated by flow cytometric analysis and the content of ROS, SOD, MDA, LDH, CK, ATP, cyt c were detected by special commercial kits. The expression of mitochondrial function-related proteins were detected by western blot. Then, the roles of ERK signaling pathway was determined with TBHQ (ERK activator) treatment. Our data have demonstrated that I/R boosted the expression of LGR4 in H9c2 cells. Knockdown of LGR4 increased the apoptosis rate of H9c2 cells and led to excessed oxidant stress and impaired mitochondrial function by increasing the levels of ROS, MDA, LDH, CK and cyt c and inhibiting SOD activity, ATP production. In addition, LGR4 silence inhibited the activation of ERK pathway. And TBHQ partially reversed the effects of LGR4 knockdown on H9c2 cells. To conclude, our study indicated that LGR4 regulated mitochondrial dysfunction and oxidative stress by ERK signaling pathways, which provides a potential cardiac protective target against I/R.