Gentianella acuta improves TAC-induced cardiac remodelling by regulating the Notch and PI3K/Akt/FOXO1/3 pathways.

Cardiac remodelling mainly manifests as excessive myocardial hypertrophy and fibrosis, which are associated with heart failure. Gentianella acuta (G. acuta) is reportedly effective in cardiac protection; however, the mechanism by which it protects against cardiac remodelling is not fully understood. Here, we discuss the effects and mechanisms of G. acuta in transverse aortic constriction (TAC)-induced cardiac remodelling in rats. Cardiac function was analysed using echocardiography and electrocardiography. Haematoxylin and eosin, Masson's trichrome, and wheat germ agglutinin staining were used to observe pathophysiological changes. Additionally, real-time quantitative reverse transcription polymerase chain reaction and western blotting were used to measure protein levels and mRNA levels of genes related to myocardial hypertrophy and fibrosis. Immunofluorescence double staining was used to investigate the co-expression of endothelial and interstitial markers. Western blotting was used to estimate the expression and phosphorylation levels of the regulatory proteins involved in autophagy and endothelial-mesenchymal transition (EndMT). The results showed that G. acuta alleviated cardiac dysfunction and remodelling. The elevated levels of myocardial hypertrophy and fibrosis markers, induced by TAC, decreased significantly after G. acuta intervention. G. acuta decreased the expression of LC3 II and Beclin1, and increased p62 expression. G. acuta upregulated the expression of CD31 and vascular endothelial-cadherin, and prevented the expression of α-smooth muscle actin and vimentin. Furthermore, G. acuta inhibited the PI3K/Akt/FOXO1/3a pathway and activated the Notch signalling. These findings demonstrated that G. acuta has cardioprotective effects, such as alleviating myocardial fibrosis, inhibiting hypertrophy, reducing autophagy, and blocking EndMT by regulating the PI3K/Akt/FOXO1/3a and Notch signalling pathways.

Gentianella acuta prevents acute myocardial infarction induced by isoproterenol in rats via inhibition of galectin-3/TLR4/MyD88/NF-кB inflammatory signalling.

Gentianella acuta (G. acuta), as a folk medicine, was used to treat heart disease by the Ewenki people in Inner Mongolia. However, the effect of G. acuta on acute myocardial infarction (AMI) is not clear. To explore the mechanisms of G. acuta on isoproterenol (ISO)-induced AMI, rats were administered G. acuta for 28 days, then injected intraperitoneally with ISO (85 mg/kg) on days 29 and 30. An electrocardiogram helped to evaluate the myocardial injury. Serum lactate dehydrogenase (LDH), creatinine kinase (CK) and aspartate aminotransferase (AST) levels were evaluated, and haematoxylin eosin, Masson's trichrome staining and terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) staining were used to detect myocardial histological changes. Radioimmunoassay was used to measure serum tumour necrosis factor alpha (TNFα) and interleukin (IL)-6. An enzyme-linked immunosorbent assay kit was used to analyse serum galectin-3 (Gal-3) levels. Immunohistochemistry, Western blotting and reverse transcription polymerase chain reaction were used to examine relevant molecular events. The results revealed that pre-treatment with G. acuta decreased the elevation in the ST segment; reduced serum LDH, CK and AST levels; alleviated cardiac structure disorder; and reduced inflammatory infiltration, abnormal collagen deposition and cardiomyocyte apoptosis that were induced by ISO. Furthermore, pre-treatment with G. acuta inhibited serum Gal-3 levels and Gal-3 expression in heart tissue, and also impeded TLR4/MyD88/NF-кB signalling activation, which ultimately prevented the expression of inflammatory cytokines. The study indicated that pre-treatment with G. acuta protects against ISO-induced AMI, and the protective role may be related to inhibiting Gal-3/TLR4/MyD88/NF-кB inflammatory signalling.

Modified citrus pectin ameliorates myocardial fibrosis and inflammation via suppressing galectin-3 and TLR4/MyD88/NF-κB signaling pathway.

Myocardial fibrosis (MF) plays a key role in the development and progression of heart failure (HF) with limited effective therapies. Galectin-3 (Gal-3) is a biomarker associated with fibrosis and inflammation in patients with HF. The Gal-3 inhibitor modified citrus pectin (MCP) protects against cardiac dysfunction, though the underlying mechanism remains unclear. The aim of this study was to investigate the effect and mechanism of MCP on MF using an isoproterenol (ISO)-induced rat model of HF. Cardiac function was analyzed by echocardiography and electrocardiography. Histopathological changes in the heart tissue were assessed by hematoxylin-eosin and Masson trichrome staining. The mRNA and protein expression levels of signaling molecules and pro-inflammatory cytokines were monitored by immunohistochemistry, western blot, qRT-PCR and ELISA analyses. The results demonstrated that MCP ameliorated cardiac dysfunction, decreased myocardial injury and reduced collagen deposition. Furthermore, MCP downregulated the expression of Gal-3, TLR4 and MyD88, thereby inhibiting NF-κB-p65 activation. MCP also decreased the expression of IL-1ß, IL-18 and TNF-α, which have been implicated in the pathogenesis of HF. These inhibitory effects were observed on day 15 and continued until day 22. Taken together, these results suggest that MCP ameliorates cardiac dysfunction through inhibiting inflammation and MF. These effects may be through downregulating Gal-3 expression and suppressing activation of the TLR4/MyD88/NF-κB signaling pathway. The present study supports the use of Gal-3 as a therapeutic target for the treatment of MF after myocardial infarction.

Inhibitory effects of CP on the growth of human gastric adenocarcinoma BGC-823 tumours in nude mice.

Objective To investigate the potential antitumour effects of [2-(6-amino-purine-9-yl)-1-hydroxy-phosphine acyl ethyl] phosphonic acid (CP) against gastric adenocarcinoma. Methods Human BGC-823 xenotransplants were established in nude mice. Animals were randomly divided into control and CP groups, which were administered NaHCO3 vehicle alone or CP dissolved in NaHCO3 (200 µg/kg body weight) daily, respectively. Tumour volume was measured weekly for 6 weeks. Resected tumours were assayed for proliferative activity with anti-Ki-67 or anti-proliferating cell nuclear antigen (PCNA) antibodies. Cell apoptosis was examined using terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling (TUNEL) assays and with caspase-3 immunostaining. Proteins were measured by Western blotting. Results There was a significant reduction in tumour volume and a reduced percentage of Ki-67-positive or PCNA-positive cells in the CP group compared with the control group. The percentage of TUNEL-positive or caspase 3-positive cells significantly increased following CP treatment compared with the control group. Tumours from the CP group had higher levels of phosphorylated-extracellular signal-regulated kinase (p-ERK) and phosphorylated-AKT (p-AKT) compared with control tumours. Conclusion CP treatment inhibited tumour growth and induced tumour cell apoptosis in a nude mouse model of BGC-823 gastric adenocarcinoma. Activation of the AKT and ERK signalling pathways may mediate this antitumour activity.

Roscovitine attenuates intimal hyperplasia via inhibiting NF-κB and STAT3 activation induced by TNF-α in vascular smooth muscle cells.

Roscovitine is a selective CDK inhibitor originally designed as anti-cancer agent, which has also been shown to inhibit proliferation in vascular smooth muscle cells (VSMCs). However, its effect on vascular remodeling and its mechanism of action remain unknown. In our study, we created a new intimal hyperplasia model in male Sprague-Dawley rats by trypsin digestion method, which cause to vascular injury as well as the model of rat carotid balloon angioplasty. Roscovitine administration led to a significant reduction in neointimal formation and VSMCs proliferation after injury in rats. Western blot analysis revealed that, in response to vascular injury, TNF-α stimulation induced p65 and STAT3 phosphorylation and promoted translocation of these molecules into the nucleus. p65 can physically associate with STAT3 and bind to TNF-α-regulated target promoters, such as MCP-1 and ICAM-1, to initiate gene transcription. Roscovitine can interrupt activation of NF-κB and reduce expression of TNF-α-induced proinflammatory gene, thus inhibiting intimal hyperplasia. These findings provide a novel mechanism to explain the roscovitine-mediated inhibition of intimal hyperplasia induced by proinflammatory pathways.

Roscovitine Protects From Arterial Injury by Regulating the Expressions of c-Jun and p27 and Inhibiting Vascular Smooth Muscle Cell Proliferation.

PURPOSE: Roscovitine (Rosc) is a selective inhibitor of cyclin-dependent kinases (CDKs) and a promising therapy for various cancers. However, limited information is available on the biological significance of Rosc in vascular smooth muscle cells (VSMCs), the cell type critical for the development of proliferative vascular diseases. In this study, we address the effects of Rosc in regulating VSMC proliferation, both in vitro and in vivo, exploring the underlying molecular mechanisms. METHODS: The proliferations and cell-cycle distributions of in vitro cultured VSMCs, as well as several other cancer cell lines, were examined by cell-counting assay and flow cytometry, respectively. Molecular changes in various CDKs, cyclins, and other regulatory molecules were examined by reverse transcription polymerase chain reaction, Western blot, or immunocytochemistry. The in vivo effects of Rosc were examined on a carotid arterial balloon-injury model. RESULTS: Rosc significantly inhibited VSMC proliferation in response to serum or angiotensin II and arrested these cells at the G0/G1 phase. These changes were associated with a specific and robust decrease in CDK4, cyclin E, c-Jun, and a dramatic increase in p27kip1 in VSMCs, which was also translated in vivo and correlated with the protection of Rosc on injury-induced neointimal hyperplasia. CONCLUSIONS: Acting on distinct molecular targets in VSMCs versus cancer cells, Rosc inhibits VSMC proliferation and protects from proliferative vascular diseases.

Endometrial breakdown with sustained progesterone release involves NF-κB-mediated functional progesterone withdrawal in a mouse implant model.

Irregular uterine bleeding is a major side effect of long-acting progestogen-only contraceptives in women, and is the primary reason women discontinue their use. In this study, a mouse model of endometrial breakdown was established using a subcutaneous progesterone implant to understand how irregular bleeding begins. Although progestogens sustained decidualization, endometrial breakdown was still observed in this model. We, therefore, hypothesized that endometrial breakdown might involve functional progesterone withdrawal. Using co-immunoprecipitation assays, we observed the constitutive activation of nuclear factor kappa-b (NF-κB) p65 and its interaction with the progesterone receptor (PGR); moreover, transcriptional activity of the PGR was also repressed by NF-κB activity in primary mouse and human decidual stromal cells that mimic progesterone maintenance. Yet the ratio of PGR-B to PGR-A was not increased in the mouse model. In vivo comparison of endometrial breakdown induced by progesterone withdrawal to that seen during sustained progesterone exposure, in the presence of NF-κB inhibitors, revealed that NF-κB-mediated functional progesterone withdrawal is involved in endometrial breakdown in this implant model. These data prompt further studies to determine the homology of this functional progesterone withdrawal mechanism in human endometrium. Mol. Reprod. Dev. 83: 780-791, 2016 © 2016 Wiley Periodicals, Inc.