Renal protective effects of Alpinate Oxyphyllae Fructus and mesenchymal stem cells co-treatment against D- galactose induced renal deterioration.

Age-related structural and functional changes in the kidney can eventually lead to development of chronic kidney disease, which is one of the leading causes of mortality among elderly people. For effective management of age-related kidney complications, it is important to identify new therapeutic interventions with minimal side-effects. The present study was designed to evaluate the synergistic effect of a traditional Chinese herb, Alpinate Oxyphyllae Fructus (AOF), and adipose-derived mesenchymal stem cells (ADMSCs) in ameliorating D-galactose (D-gal)-induced renal aging phenotypes in WKY rats. The study findings showed that D-gal-induced alteration in the kidney morphology was partly recovered by the AOF and ADMSC co-treatment. Moreover, the AOF and ADMSC co-treatment reduced the expression of proinflammatory mediators (NFkB, IL-6, and Cox2) and increased the expression of redox regulators (Nrf2 and HO-1) in the kidney, which were otherwise augmented by the D-gal treatment. Regarding kidney cell death, the AOF and ADMSC co-treatment was found to abolish the proapoptotic effects of D-gal by downregulating Bax and Bad expressions and inhibiting caspase 3 activation. Taken together, the study findings indicate that the AOF and ADMSC co-treatment protect the kidney from D-gal-induced aging by reducing cellular inflammation and oxidative stress and inhibiting renal cell death. This study can open up a new path toward developing novel therapeutic interventions using both AOF and ADMSC to effectively manage age-related renal deterioration.

Anethole mitigates H2 O2 -induced inflammation in HIG-82 synoviocytes by suppressing the aquaporin 1 expression and activating the protein kinase A pathway.

Rheumatoid arthritis (RA) is an autoimmune inflammatory disease affecting approximately 1% of the global population, with a higher prevalence in women than in men. Chronic inflammation and oxidative stress play pivotal roles in the pathogenesis of RA. Anethole, a prominent compound derived from fennel (Foeniculum vulgare), possesses a spectrum of therapeutic properties, including anti-arthritic, anti-inflammatory, antioxidant, and tumor-suppressive effects. However, its specific impact on RA remains underexplored. This study sought to uncover the potential therapeutic value of anethole in treating RA by employing an H2 O2 -induced inflammation model with HIG-82 synovial cells. Our results demonstrated that exposure to H2 O2 induced the inflammation and apoptosis in these cells. Remarkably, anethole treatment effectively countered these inflammatory and apoptotic processes triggered by H2 O2 . Moreover, we identified the aquaporin 1 (AQP1) and protein kinase A (PKA) pathway as critical regulators of inflammation and apoptosis. H2 O2 stimulation led to an increase in the AQP1 expression and a decrease in p-PKA-C, contributing to cartilage degradation. Conversely, anethole not only downregulated the AQP1 expression but also activated the PKA pathway, effectively suppressing cell inflammation and apoptosis. Furthermore, anethole also inhibited the enzymes responsible for cartilage degradation. In summary, our findings highlight the potential of anethole as a therapeutic agent for mitigating H2 O2 -induced inflammation and apoptosis in synovial cells, offering promising prospects for future RA treatments.

Platycodi radix aqueous extract salvages doxorubicin-induced senescence by mitochondrial reactive oxygen species reduction in umbilical cord matrix stem cells.

Platycodi radix is a widely used herbal medicine that contains numerous phytochemicals beneficial to health. The health and biological benefits of P. radix have been found across various diseases. The utilization of umbilical cord stromal stem cells, derived from Wharton's jelly of the human umbilical cord, has emerged as a promising approach for treating degenerative diseases. Nevertheless, growing evidence indicates that the function of stem cells declines with age, thereby limiting their regenerative capacity. The primary objective in this study is to investigate the beneficial effects of P. radix in senescent stem cells. We conducted experiments to showcase that diminished levels of Lamin B1 and Sox-2, along with an elevation in p21, which serve as indicative markers for the senescent stem cells. Our findings revealed the loss of Lamin B1 and Sox-2, coupled with an increase in p21, in umbilical cord stromal stem cells subjected to a low-dose (0.1 µM) doxorubicin (Dox) stimulation. However, P. radix restored the Dox-damage in the umbilical cord stromal stem cells. P. radix reversed the senescent conditions when the umbilical cord stromal stem cells exposed to Dox-induced reactive oxygen species (ROS) and mitochondrial membrane potential are significantly changed. In Dox-challenged aged umbilical cord stromal stem cells, P. radix reduced senescence, increased longevity, prevented mitochondrial dysfunction and ROS and protected against senescence-associated apoptosis. This study suggests that P. radix might be as a therapeutic and rescue agent for the aging effect in stem cells. Inhibition of cell death, mitochondrial dysfunction and aging-associated ROS with P. radix provides additional insights into the underlying molecular mechanisms.

Cardiac-specific overexpression of insulin-like growth factor II receptor-α interferes with the regulation of calcium homeostasis in the heart under hyperglycemic conditions.

BACKGROUND: Diabetic cardiomyopathy is a progressive disease caused by inexplicit mechanisms, and a novel factor, insulin-like growth factor II receptor-α (IGF-IIRα), may contribute to aggravating its pathogenesis. We hypothesized that IGF-IIRα could intensify diabetic heart injury. METHODS AND RESULTS: To demonstrate the potential role of IGF-IIRα in the diabetic heart, we used (SD-TG [IGF-IIRα]) transgenic rat model with cardiac-specific overexpression of IGF-IIRα, along with H9c2 cells, to study the effects of IGF-IIRα in the heart under hyperglycemic conditions. IGF-IIRα was found to remodel calcium homeostasis and intracellular Ca2+ overload-induced autophagy disturbance in the heart during diabetes. IGF-IIRα overexpression induced intracellular Ca2+ alteration by downregulating phosphorylated phospholamban/sarcoplasmic/endoplasmic reticulum calcium-ATPase 2a (PLB/SERCA2a), resulting in the suppression of Ca2+ uptake into the endoplasmic reticulum. Additionally, IGF-IIRα itself contributed to Ca2+ withdrawal from the endoplasmic reticulum by increasing the expression of CaMKIIδ in the active form. Furthermore, alterations in Ca2+ homeostasis significantly dysregulated autophagy in the heart during diabetes. CONCLUSIONS: Our study reveals the novel role of IGF-IIRα in regulating cardiac intracellular Ca2+ homeostasis and its related autophagy interference, which contribute to the development of diabetic cardiomyopathy. In future, the present study findings have implications in the development of appropriate therapy to reduce diabetic cardiomyopathy.

Fisetin attenuates doxorubicin-induced cardiotoxicity by inhibiting the insulin-like growth factor II receptor apoptotic pathway through estrogen receptor-α/-ß activation.

Doxorubicin (DOX), an effective chemotherapeutic drug, has been used to treat various cancers; however, its cardiotoxic side effects restrict its therapeutic efficacy. Fisetin, a flavonoid phytoestrogen derived from a range of fruits and vegetables, has been reported to exert cardioprotective effects against DOX-induced cardiotoxicity; however, the underlying mechanisms remain unclear. This study investigated fisetin's cardioprotective role and mechanism against DOX-induced cardiotoxicity in H9c2 cardiomyoblasts and ovariectomized (OVX) rat models. MTT assay revealed that fisetin treatment noticeably rescued DOX-induced cell death in a dose-dependent manner. Moreover, western blotting and TUNEL-DAPI staining showed that fisetin significantly attenuated DOX-induced cardiotoxicity in vitro and in vivo by inhibiting the insulin-like growth factor II receptor (IGF-IIR) apoptotic pathway through estrogen receptor (ER)-α/-ß activation. The echocardiography, biochemical assay, and H&E staining results demonstrated that fisetin reduced DOX-induced cardiotoxicity by alleviating cardiac dysfunction, myocardial injury, oxidative stress, and histopathological damage. These findings imply that fisetin has a significant therapeutic potential against DOX-induced cardiotoxicity.

Insulin-like growth factor II receptor alpha overexpression in heart aggravates hyperglycemia-induced cardiac inflammation and myocardial necrosis.

Diabetes-induced cardiovascular complications are mainly associated with high morbidity and mortality in patients with diabetes. Insulin-like growth factor II receptor α (IGF-IIRα) is a cardiac risk factor. In this study, we hypothesized IGF-IIRα could also deteriorate diabetic heart injury. The results presented that both in vivo transgenic Sprague-Dawley rat model with specific IGF-IIRα overexpression in the heart and in vitro myocardium H9c2 cells were used to investigate the negative function of IGF-IIRα in diabetic hearts. The results showed that IGF-IIRα overexpression aided hyperglycemia in creating more myocardial injury. Pro-inflammatory factors, such as Tumor necrosis factor-alpha, Interleukin-6, Cyclooxygenase-2, Inducible nitric oxide synthase, and Nuclear factor-kappaB inflammatory cascade, are enhanced in the diabetic myocardium with cardiac-specific IGF-IIRα overexpression. Correspondingly, IGF-IIRα overexpression in the diabetic myocardium also reduced the PI3K-AKT survival axis and activated mitochondrial-dependent apoptosis. Finally, both ejection fraction and fractional shortening were be significantly decrease in diabetic rats with cardiac-specific IGF-IIRα overexpression. Overall, all results provid clear evidence that IGF-IIRα can enhance cardiac damage and is a harmful factor to the heart under high-blood glucose conditions. However, the pathophysiology of IGF-IIRα under different stresses and its downstream regulation in the heart still require further research.

Carboxyl terminus of HSP70-interacting protein attenuates advanced glycation end products-induced cardiac injuries by promoting NFκB proteasomal degradation.

Advanced glycation end products (AGEs), which are highly reactive molecules resulting from persistent high-glucose levels, can lead to the generation of oxidative stress and cardiac complications. The carboxyl terminus of HSP70 interacting protein (CHIP) has been demonstrated to have a protective role in several diseases, including cardiac complications; however, the role in preventing AGE-induced cardiac damages remains poorly understood. Here, we found that elevated AGE levels impaired cardiac CHIP expression in streptozotocin-induced diabetes and high-fat diet-administered animals, representing AGE exposure models. We used the TUNEL assay, hematoxylin and eosin, Masson's trichrome staining, and western blotting to prove that cardiac injuries were induced in diabetic animals and AGE-treated cardiac cells. Interestingly, our results collectively indicated that CHIP overexpression significantly rescued the AGE-induced cardiac injuries and promoted cell survival. Moreover, CHIP knockdown-mediated stabilization of nuclear factor κB (NFκB) was attenuated by overexpressing CHIP in the cells. Furthermore, co-immunoprecipitation and immunoblot assay revealed that CHIP promotes the ubiquitination and proteasomal degradation of AGE-induced NFκB. Importantly, fluorescence microscopy, a luciferase reporter assay, electrophoretic mobility shift assay, and subcellular fractionation further demonstrated that CHIP overexpression inhibits AGE-induced NFκB nuclear translocation, reduced its binding ability with the promoter sequences of the receptor of AGE, consequently inhibiting the translocation of the receptor AGE to the cell membrane for its proper function. Overall, our current study findings suggest that CHIP can target NFκB for ubiquitin-mediated proteasomal degradation, and thereby potentially rescue AGE-induced cardiac damages.

MicroRNA-210 repression facilitates advanced glycation end-product (AGE)-induced cardiac mitochondrial dysfunction and apoptosis via JNK activation.

Hyperglycemia results in the formation of reactive oxygen species which in turn causes advanced glycation end products (AGEs) formation, leading to diabetic cardiomyopathy. Our previous study showed that AGE-induced reactive oxygen species-dependent apoptosis is mediated via protein kinase C delta (PKCδ)-enhanced mitochondrial damage in cardiomyocytes. By using microRNA (miRNA) database, miRNA-210 was predicted to target c-Jun N-terminal kinase (JNK), which were previously identified as downstream of PKCδ in regulating mitochondrial function. Therefore, we hypothesized that miR-210 mediates PKCδ-dependent upregulation of JNK to cause cardiac mitochondrial damage and apoptosis following AGE exposure. AGE-exposed cells showed activated cardiac JNK, PKCδ, and apoptosis, which were reversed by treatment with a JNK inhibitor and PKCδ-KD (deficient kinase). Cardiac miR-210 and mitochondrial function were downregulated following AGE exposure. Furthermore, JNK was upregulated and involved in AGE-induced mitochondrial damage. Interestingly, luciferase activity of the miR-210 mimic plus JNK WT-3'-untranslated region overexpressed group was significantly lower than that of miR-210 mimic plus JNK MT-3'UTR group, indicating that JNK is a target of miR-210. Moreover, JNK activation induced by AGEs was reduced by treatment with the miR-210 mimic and reversed by treatment with the miR-210 inhibitor, indicating the regulatory function of miR-210 in JNK activation following AGE exposure. Additionally, JNK-dependent mitochondrial dysfunction and apoptosis were reversed following treatment with the miR-210 mimic, while the miR-210 inhibitor showed no effect on JNK-induced mitochondrial dysfunction and apoptosis in AGE-exposed cardiac cells. Taken together, our study showed that PKCδ-enhanced JNK-dependent mitochondrial damage is mediated through the reduction of miR-210 in cardiomyocytes following AGE exposure.

miR-145-5p targets paxillin to attenuate angiotensin II-induced pathological cardiac hypertrophy via downregulation of Rac 1, pJNK, p-c-Jun, NFATc3, ANP and by Sirt-1 upregulation.

Pathological cardiac hypertrophy is associated with many diseases including hypertension. Recent studies have identified important roles for microRNAs (miRNAs) in many cardiac pathophysiological processes, including the regulation of cardiomyocyte hypertrophy. However, the role of miR-145-5p in the cardiac setting is still unclear. In this study, H9C2 cells were overexpressed with microRNA-145-5p, and then treated with Ang-II for 24 h, to study the effect of miR-145-5p on Ang-II-induced myocardial hypertrophy in vitro. Results showed that Ang-II treatment down-regulated miR-145-5p expression were revered after miR-145-5p overexpression. Based on results of bioinformatics algorithms, paxillin was predicted as a candidate target gene of miR-145-5p, luciferase activity assay revealed that the luciferase activity of cells was substantial downregulated the following co-transfection with wild paxillin 3'UTR and miR-145-5p compared to that in scramble control, while the inhibitory effect of miR-145-5p was abolished after transfection of mutant paxillin 3'UTR. Additionally, overexpression of miR-145-5p markedly inhibited activation of Rac-1/ JNK /c-jun/ NFATc3 and ANP expression and induced SIRT1 expression in Ang-II treated H9c2 cells. Jointly, our study suggested that miR-145-5p inhibited cardiac hypertrophy by targeting paxillin and through modulating Rac-1/ JNK /c-jun/ NFATc3/ ANP / Sirt1 signaling, therefore proving novel downstream molecular pathway of miR-145-5p in cardiac hypertrophy.

Protective effects of luteolin against oxidative stress and mitochondrial dysfunction in endothelial cells.

BACKGROUND AND AIMS: Luteolin is a common flavonoid that is abundantly present in various edible plants, it is known to exhibit beneficial effects on cardiovascular system. However, the mechanisms which underlie the protective effects of luteolin on endothelial cell damage caused by oxidative stress remains unclear. The purpose of this study is to test the hypothesis which states that luteolin protects against H2O2-induced oxidative stress via modulating ROS-mediated P38 MAPK/NF-κB and calcium-evoked mitochondrial apoptotic signalling pathways. METHODS AND RESULTS: Human umbilical vein endothelial cells (HUVECs) were pretreated with luteolin prior to being stimulated by 600 µM H2O2 for another 24 h. The expression of native and phosphorylated-P38, IκB, NF-κB, native eNOS, phosphorylated-eNOS, iNOS and several apoptosis-related proteins were analyzed by Western blot. In addition, intracellular calcium was determined by fura-2 AM and mitochondrial membrane potential was examined by using JC1. Using the data gathered, we found indications that H2O2 induced P38 MAPK/NF-κB activation. H2O2 downregulated the expression of eNOS and upregulated iNOS, which in turn contribute to an elevated NO generation and protein nitrosylation. However, pretreatment with luteolin markedly reversed all of these alterations dose-dependently. Additionally, an intracellular calcium rise and subsequent mitochondrial membrane potential collapse, P53 phosphorylation, reduced BcL-2/Bax ratio in the mitochondrial membrane, release cytochrome c from mitochondria, leading to the subsequent activation of caspase 3 activation by H2O2 were all markedly suppressed in the presence of luteolin. CONCLUSION: Results from this study may provide the possible molecular mechanisms underlying cardiovascular protective effects of luteolin.

Andrographolide mitigates cardiac apoptosis to provide cardio-protection in high-fat-diet-induced obese mice.

Excessive intake of high fat diet (HFD) and associated obese conditions are critical contributors of cardiac diseases. In this study, an active metabolite andrographolide from Andrographis paniculata was found to ameliorate HFD-induced cardiac apoptosis. C57/BL6 mouse were grouped as control (n = 9), obese (n = 8), low dose (25 mg/kg/d) andrographolide treatment (n = 9), and high dose (50 mg/kg/d) andrographolide treatment (n = 9). The control group was provided with standard laboratory chow and the other groups were fed with HFD. Andrographolide was administered through oral gavage for 1 week. Histopathological analysis showed increase in apoptotic nuclei and considerable cardiac-damages in the obese group signifying cardiac remodeling effects. Further, Western blot results showed increase in pro-apoptotic proteins and decrease in the proteins of IGF-1R-survival signaling. However, feeding of andrographolide significantly reduced the cardiac effects of HFD. The results strongly suggest that andrographolide supplementation can be used for prevention and treatment of cardiovascular disease in obese patients.

HSF1 phosphorylation by ERK/GSK3 suppresses RNF126 to sustain IGF-IIR expression for hypertension-induced cardiomyocyte hypertrophy.

Hypertension-induced cardiac hypertrophy and apoptosis are major characteristics of early-stage heart failure (HF). Inhibition of extracellular signal-regulated kinases (ERK) efficaciously suppressed angiotensin II (ANG II)-induced cardiomyocyte hypertrophy and apoptosis by blocking insulin-like growth factor II receptor (IGF-IIR) signaling. However, the detailed mechanism by which ANG II induces ERK-mediated IGF-IIR signaling remains elusive. Here, we found that ANG II activated ERK to upregulate IGF-IIR expression via the angiotensin II type I receptor (AT1 R). ERK activation subsequently phosphorylates HSF1 at serine 307, leading to a secondary phosphorylation by glycogen synthase kinase III (GSK3) at serine 303. Moreover, we found that ANG II mediated ERK/GSK3-induced IGF-IIR protein stability by downregulating the E3 ubiquitin ligase of IGF-IIR RING finger protein CXXVI (RNF126). The expression of RNF126 decreased following ANG II-induced HSF1S303 phosphorylation, resulting in IGF-IIR protein stability and increased cardiomyocyte injury. Inhibition of GSK3 significantly alleviated ANG II-induced cardiac hypertrophy in vivo and in vitro. Taken together, these results suggest that HSF1 phosphorylation stabilizes IGF-IIR protein stability by downregulating RNF126 during cardiac hypertrophy. ANG II activates ERK/GSK3 to phosphorylate HSF1, resulting in RNF126 degradation, which stabilizes IGF-IIR protein expression and eventually results in cardiac hypertrophy. HSF1 could be a valuable therapeutic target for cardiac diseases among hypertensive patients.

GABA tea attenuates cardiac apoptosis in spontaneously hypertensive rats (SHR) by enhancing PI3K/Akt-mediated survival pathway and suppressing Bax/Bak dependent apoptotic pathway.

Cardiomyocyte apoptosis is the major risk factor for the development of heart failure (HF). The purpose of this study was to evaluate the effects of Gamma-aminobutyric acid (GABA) tea on hypertension-induced cardiac apoptotic pathways in spontaneously hypertensive rats (SHR). In order to reveal the mechanisms, 36 male SHR at eight weeks of age, 200 g were divided into six groups. One group was fed water as a control group. Other rats were administered one of the following treatments: GABA tea at dose 150 and 300 mg/kg/day as low GABA tea (LGT) and high GABA tea (HGT) groups, respectively, pure GABA at dose 150 and 300 mg/kg/day as LG and HG groups, respectively, green tea (GT) as control of LGT and HGT groups. After 12 weeks, cardiac tissues were analyzed by histological analysis, western blotting, and TUNEL assays. GABA tea, GT, and pure GABA decreased hypertension-induced cardiac abnormalities, including abnormal myocardial architecture. In addition, GABA tea, GT, and pure GABA dramatically increased anti-apoptotic protein, Bcl2. Furthermore, GABA tea, GT, and pure GABA also decreased activated-caspase 9 and activated-caspase 3. Additionally, the survival associated protein IGF-I and PI3K/Akt were enhanced in cardiac tissues upon treatment. Our results showed an optimistic anti-apoptotic and pro-survival effects of GABA tea treatment against hypertensive rat hearts.

The preventive effects of edible folic acid on cardiomyocyte apoptosis and survival in early onset triple-transgenic Alzheimer's disease model mice.

In recent years, neuropathological and epidemiological studies have indicated an association between Alzheimer's disease (AD) and several cardiovascular risk factors. In this study, the cardio-protective effects of folic acid (FA) in early stage AD was elucidated using a triple-transgenic (3xTg) Alzheimer's mouse model. Eleven-month-old C57BL/6 mice and 3xTg mice were assigned to five groups. During the four-month treatment period, the low-FA treatment group received FA through their diet, and the high-FA treatment groups received 3 mg/dl folate in drinking water and were also gastric-fed 1.2 mg/kg folate every day. In the C57B1/6J mice, treatment with high doses of FA (HFA) did not show any considerable effect compared to the control group or the low-dose dietary FA treatment group. However, Alzheimer's mice treated with HFA showed enhanced cardio-protection. Western blot analysis revealed that FA treatment restored SIRT1 expression, which was suppressed in 3xTg mice, through enhanced AMPK expression. FA significantly enhanced the IGF1 receptor survival mechanism in the hearts of the 3xTg mice and suppressed the expression-intrinsic and extrinsic apoptosis-associated proteins. The results suggest that FA intake may significantly alleviate cellular pathological events in the heart associated with AD.

Taiwanin C selectively inhibits arecoline and 4-NQO-induced oral cancer cell proliferation via ERK1/2 inactivation.

Arecoline, the most abundant alkaloid in betel nut is known to promote abnormal proliferation of epithelial cells by enhancing epidermal growth factor receptor (EGFR) activation and cyclooxygenase-2 (COX2) expression. Taiwanin C, a naturally occurring lignan extracted from Taiwania cryptomerioides, has been found to be a potential inhibitor of COX2 expression. Based on the MTT assay results, taiwanin C was found to be effective in inhibiting the tumorous T28 cell than the non-tumorous N28 cells. The modulations in the expression of relevant proteins were determined to understand the mechanism induced by taiwanin C to inhibit T28 cell proliferation. The levels of activated EGFR and COX2 were found to be abnormally high in the T28 oral cancer cells. However, taiwanin C was found to inhibit the activation of EGFR and regulated other related downstream proteins and thereby inhibited the T28 cell proliferation. In conclusion the results indicate that taiwanin C suppresses COX2-EGFR and enhances P27 pathways to suppress arecoline induced oral cancer cell proliferation via ERK1/2 inactivation. © 2015 Wiley Periodicals, Inc. Environ Toxicol 32: 62-69, 2017.

E2/ER ß Enhances Calcineurin Protein Degradation and PI3K/Akt/MDM2 Signal Transduction to Inhibit ISO-Induced Myocardial Cell Apoptosis.

Secretion of multifunctional estrogen and its receptor has been widely considered as the reason for markedly higher frequency of heart disease in men than in women. 17ß-Estradiol (E2), for instance, has been reported to prevent development of cardiac apoptosis via activation of estrogen receptors (ERs). In addition, protein phosphatase such as protein phosphatase 1 (PP1) and calcineurin (PP2B) are also involved in cardiac hypertrophy and cell apoptosis signaling. However, the mechanism by which E2/ERß suppresses apoptosis is not fully understood, and the role of protein phosphatase in E2/ERß action also needs further investigation. In this study, we observed that E2/ERß inhibited isoproterenol (ISO)-induced myocardial cell apoptosis, cytochrome c release and downstream apoptotic markers. Moreover, we found that E2/ERß blocks ISO-induced apoptosis in H9c2 cells through the enhancement of calcineurin protein degradation through PI3K/Akt/MDM2 signaling pathway. Our results suggest that supplementation with estrogen and/or overexpression of estrogen receptor ß gene may prove to be effective means to treat stress-induced myocardial damage.

Treatment with 17ß-Estradiol Reduced Body Weight and the Risk of Cardiovascular Disease in a High-Fat Diet-Induced Animal Model of Obesity.

Estrogen receptor α (ERα) and estrogen receptor ß (ERß) play important roles in cardiovascular disease (CVD) prevention. Recently, these estrogen receptors were reconsidered as an important treatment target of obesity leading to CVD. In this study, 17ß-estradiol (17ß-E) replacement therapy applied to high-fat diet-induced obese C57B male mice and ovariectomized (OVX) rats were evaluated, and the protective effects against high-fat diet-induced obesity were assessed in C57B mouse hearts. The results showed that 17ß-E treatment activated both ERα and ERß, and ERß levels increased in a dose-dependent manner in high-fat diet C57B mouse cardiomyocytes following 17ß-E treatment. Notably, an almost 16% reduction in body weight was observed in the 17ß-E-treated (12 µg/kg/day for 60 days) high-fat diet-induced obese C57B male mice. These results suggested that 17ß-E supplements may reduce CVD risk due to obesity.

Hypoxia suppresses myocardial survival pathway through HIF-1α-IGFBP-3-dependent signaling and enhances cardiomyocyte autophagic and apoptotic effects mainly via FoxO3a-induced BNIP3 expression.

The HIF-1α transcriptional factor and the BH-3 only protein BNIP3 are known to play fundamental roles in response to hypoxia. The objective of this research is to investigate the molecular mechanisms and the correlation of HIF-1α, BNIP3 and IGFBP-3 in hypoxia-induced cardiomyocytes injuries. Heart-derived H9c2 cells and neonatal rat ventricular myocytes (NRVMs) were incubated in normoxic or hypoxic conditions. Hypoxia increased HIF-1α expression and activated the downstream BNIP3 and IGFBP-3 thereby triggered mitochondria-dependent apoptosis. Moreover, IGF1R/PI3K/Akt signaling was attenuated by HIF-1α-dependent IGFBP-3 expression to enhance hypoxia-induced apoptosis. Autophagy suppression with 3-methyladenine or siATG5 or siBeclin-1 significantly decreased myocardial apoptosis under hypoxia. Knockdown of FoxO3a or BNIP3 significantly abrogated hypoxia-induced autophagy and mitochondria-dependent apoptosis. Moreover, prolonged-hypoxia induced HIF-1α stimulated BNIP3 and enhanced IGFBP-3 activation to inhibit IGF1R/PI3K/Akt survival pathway and mediate mitochondria-dependent cardiomyocyte apoptosis. HIF-1α and FoxO3a blockage are sufficient to annul the change of excessive hypoxia of hearts.

NFIL3 suppresses hypoxia-induced apoptotic cell death by targeting the insulin-like growth factor 2 receptor.

The insulin-like growth factor-II/mannose 6-phosphate receptor (IGF2R) over-expression correlates with heart disease progression. The IGF2R is not only an IGF2 clearance receptor, but it also triggers signal transduction, resulting in cardiac hypertrophy, apoptosis and fibrosis. The present study investigated the nuclear factor IL-3 (NFIL3), a transcription factor of the basic leucine zipper superfamily, and its potential pro-survival effects in cardiomyocytes. NFIL3 might play a key role in heart development and act as a survival factor in the heart, but the regulatory mechanisms are still unclear. IGF2 and IGF2R protein expression were highly increased in rat hearts subjected to hemorrhagic shock. IGF2R protein expression was also up-regulated in H9c2 cells exposed to hypoxia. Over-expression of NFIL3 in H9c2 cardiomyoblast cells inhibited the induction of hypoxia-induced apoptosis and down-regulated IGF2R expression levels. Gel shift assay, double-stranded DNA pull-down assay and chromatin immune-precipitation analyses indicated that NFIL3 binds directly to the IGF2R promoter region. Using a luciferase assay, we further observed NFIL3 repress IGF2R gene promoter activity. Our results demonstrate that NFIL3 is an important negative transcription factor, which through binding to the promoter of IGF2R, suppresses the apoptosis induced by IGF2R signaling in H9c2 cardiomyoblast cells under hypoxic conditions.

Tetramethylpyrazine Ameliorated Hypoxia-Induced Myocardial Cell Apoptosis via HIF-1α/JNK/p38 and IGFBP3/BNIP3 Inhibition to Upregulate PI3K/Akt Survival Signaling.

BACKGROUND: Hemorrhagic shock (HS) is the major cause of death from trauma. Hemorrhagic shock may lead to cellular hypoxia and organ damage. Our previous findings showed that HS induced a cardiac apoptosis pathway and synergistically caused myocardial cell damage in diabetic rats under trauma-induced HS. Tetramethylpyrazine (TMP) is a major biologically active ingredient purified from the rhizome of Ligusticum wallichii (called Chuang Xiong in Chinese). Chuan Xiong rescued cells from synergistic cardiomyoblast cell injury under high-glucose (HG) conditions plus hypoxia. TMP is one of the most important active ingredients that elevated the survival rate in ischemic brain injury and prevented inducible NO synthase expression to have anti-inflammatory effects against cell damage in different cell types. METHOD: Here, we further investigate whether TMP can protect against hypoxic (<1% oxygen) conditions in H9c2 cardiomyoblast cells for 24 hrs. RESULTS: Our results showed that hypoxia mediated through HIF-1α/JNK/p38 activation significantly elevated the levels of the hypoxia-related proteins HIF-1α, BNIP3 and IGFBP3, further enhanced the pro-apoptotic protein Bak and upregulated downstream Caspase 9 and 3, resulting in cell death. All of these phenomena were fully recovered under TMP treatment. We observed that TMP exerted this effect by activating the IGF1 receptor survival pathway, dependent primarily on PI3K/Akt. When PI3K (class I) was blocked by specific siRNA, the hypoxia-induced activated caspase 3 and cell apoptosis could not be reversed by TMP treatment. CONCLUSION: Our results strongly suggest that TMP could be used to restore hypoxia-induced myocardial cell apoptosis and cardiac hypoxic damage.