Search

Summary Background/Objective Steroid-associated osteonecrosis (SAON) mostly occurs in large joints such as hip that finally results in joint collapse and replacement. An effective strategy for preventing SAON is inhibition of bone resorption and lipid formation, whereas promoting bone formation at osteonecrotic-sensitive skeletal sites. The objective of this study was to investigate whether gastrin administration could effectively prevent SAON in vivo and had a significant effect on osteogenesis, osteoclast differentiation and/or adipogenesis in vitro. Methods SAON was induced in rats by injection of both lipopolysaccharide and methylprednisolone. SAON rats were divided into three groups (untreated, low- ​and high-dose gastrin) and treated for two weeks. Lesions in SAON were identified, and efficacy of gastrin was evaluated in osteoclast activity, osteogenesis ​and adipogenesis. In vitro experiments evaluated the osteogenic and adipogenic potential of induced bone marrow stem cells, as well as osteoclast differentiation of stimulated bone marrow cells. Results Gastrin administration effectively prevented the development of SAON in vivo and significantly downregulated osteoclastogenesis, and upregulated osteogenesis in vitro. The underlying mechanism was potentially involved in the inhibited nuclear factor kappa-B (NF-κB) pathway leading to not only suppressed osteoclastogenesis but also facilitated osteoblastogenesis. Conclusion The present study demonstrated for the first time that gastrin could effectively prevent SAON in vivo and had a significant effect on osteogenesis, osteoclast differentiation and/or adipogenesis in vitro that should be an important link between gastrointestinal hormone and metabolic bone diseases. Translational potential of this article Gastrin prevents steroid-associated osteonecrosis with decreasing bone resorption and apoptosis, also enhancing bone formation. These findings provide in vivo evidence to support the use of gastrin to treat osteonecrosis.

Background Gastric cancer (GC) is a common form of malignant cancer in worldwide which has a poor prognosis. Despite recent improvements in the treatment of GC, the prognosis is not yet satisfactory for GC patients. CYT997, a novel microtubule-targeting agent, recently has been identified to be a promising anticancer candidate for the treatment of cancers; however, the effects of CYT997 in GC remain largely unknown. Methods Cell proliferation and apoptosis were detected by CCK8 assay and flow cytometry. The mitochondrial ROS were detected by confocal microscope and flow cytometry. Gastric cancer patient-derived xenograft (PDX) model was used to evaluate its antitumor activity of CYT997 in vivo. Results CYT997 inhibited gastric cancer cell proliferation and induced cell apoptosis and triggered autophagy. CYT997 induced apoptosis through triggering intracellular mitochondrial ROS generation in GC cells. ROS scavengers N-acetylcysteine (NAC) and Mitoquinone (MitoQ) distinctly weakened CYT997-induced cell cycle G2/M arrest and apoptosis in GC cells. Pretreatment with autophagy inhibitor 3-MA promoted the effect of CYT997 on cells apoptosis. Mechanistically, CYT997 performed its function through regulation of Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) signaling pathway in GC cells. In addition, CYT997 inhibited growth of gastric cancer patient-derived xenograft (PDX) tumors. Conclusions CYT997 induces autophagy and apoptosis in gastric cancer by triggering mitochondrial ROS accumulation to silence JAK2/STAT3 pathway. CYT997 might be a potential antitumor drug candidate to treat GC.

Objective: Integrin β3 is one of the main integrin heterodimer receptors on the surface of cardiac myocytes. Our previous studies showed that hypoxia induces apoptosis and increases integrin β3 expression in cardiomyocytes. However, the exact mechanism by which integrin β3 protects against apoptosis remains unclear. Hence, the present investigation aimed to explore the mechanism of integrin β3 in cardiomyocyte proliferation and hypoxia-induced cardiomyocyte apoptosis. Methods: Stable cells and in vivo acute and chronic heart failure rat models were generated to reveal the essential role of integrin β3 in cardiomyocyte proliferation and apoptosis. Western blotting and immunohistochemistry were employed to detect the expression of integrin β3 in the stable cells and rat cardiac tissue. Flow cytometer was used to investigate the role of integrin β3 in hypoxia-induced cardiomyocyte apoptosis. Confocal microscopy was used to detect the localization of integrin β3 and integrin αv in cardiomyocytes. Results: A cobaltous chloride-induced hypoxic microenvironment stimulated cardiomyocyte apoptosis and increased integrin β3 expression in H9C2 cells, AC16 cells, and cardiac tissue from acute and chronic heart failure rats. The overexpression of integrin β3 promoted cardiomyocyte proliferation, whereas silencing integrin β3 expression resulted in decreased cell proliferation in vitro. Furthermore, knocking down integrin β3 expression using shRNA or the integrin β3 inhibitor cilengitide exacerbated cobaltous chloride-induced cardiomyocyte apoptosis, whereas overexpression of integrin β3 weakened cobaltous chloride-induced cardiomyocytes apoptosis. We found that integrin β3 promoted cardiomyocytes proliferation through the regulation of the PTEN/Akt/mTOR and ERK1/2 signaling pathways. In addition, we found that knockdown of integrin αv or integrin β1 weakened the effect of integrin β3 in cardiomyocyte proliferation. Conclusion: Our findings revealed the molecular mechanism of the role of integrin β3 in cardiomyocyte proliferation and hypoxia-induced cardiomyocyte apoptosis, providing new insights into the mechanisms underlying myocardial protection.

Background To test the hypothesis that activated extracellular signal-regulated kinase (ERK) regulates P65-miR23a/27a/24 axis in gastric cancer (GC) and the ERK-P65-miR23a/27a/24 axis plays an important role in the development of GC, and to evaluate the role of gastrin in GC progression and ERK-P65-miR23a/27a/24 axis. Methods The component levels of the ERK-P65-miR23a/27a/24 axis in four fresh GC tissues, 101 paraffin-embedded GC tissues and four GC cell lines were determined by Western blotting, immunohistochemistry (IHC) or qRT-PCR. The effects of gastrin on GC were first evaluated by measuring gastrin serum levels in 30 healthy and 70 GC patients and performing a correlation analysis between gastrin levels and survival time in 27 GC patients after eight years of follow-up, then evaluated on GC cell lines, GC cell xenograft models, and patient-derived xenografts (PDX) mouse models. The roles of ERK-P65-miR23a/27a/24 axis in GC progression and in the effects of gastrin on GC were examined. Results ERK- P65-miR23a/27a/24 axis was proved to be present in GC cells. The levels of components of ERK-P65-miR23a/27a/24 axis were decreased in GC tissue samples and PGC cells. The decreased levels of components of ERK-P65-miR23a/27a/24 axis were associated with poor prognosis of GC, and ERK-P65-miR23a/27a/24 axis played a suppressive role in GC progression. Low blood gastrin was correlated with poor prognosis of the GC patients and decreased expression of p-ERK and p-P65 in GC tissues. Gastrin inhibited proliferation of poorly-differentiated GC (PGC) cells through activating the ERK-P65-miR23a/27a/24 axis. Gastrin inhibited GC growth and enhanced the suppression of GC by cisplatin in mice or PGC cell culture models through activating the ERK-P65-miR23a/27a/24 axis or its components. Conclusions ERK-P65-miR23a/27a/24 axis is down-regulated, leading to excess GC growth and poor prognosis of GC. Low gastrin promoted excess GC growth and contributed to the poor prognosis of the GC patients by down-regulating ERK-P65-miR23a/27a/24 axis. Gastrin inhibits gastric cancer growth through activating the ERK-P65-miR23a/27a/24 axis. Electronic supplementary material The online version of this article (10.1186/s13046-018-0782-7) contains supplementary material, which is available to authorized users.

Hypoxia-induced apoptosis plays an important role in cardiovascular diseases. Integrin β3 is one of the main integrin heterodimer receptors on the surface of cardiac myocytes. However, despite the important role that integrin β3 plays in the cardiovascular disease, its exact role in the hypoxia response remains unclear. Hence, in the present investigation we aimed to study the role of integrin β3 in hypoxia-induced apoptosis in H9C2 cells and primary rat myocardial cells. MTT assay, flow cytometry and TUNEL assay results showed that hypoxia inhibited cardiomyocyte proliferation and induced cardiomyocyte apoptosis. The expression levels of integrin β3 and HIF1α were upregulated in hypoxia-induced cardiomyocytes as revealed by real-time PCR and western blot analysis. Furthermore, knockdown of integrin β3 expression by siRNA increased hypoxia-induced cardiomyocyte apoptosis. In addition, integrin β3 overexpression weakened hypoxia-induced cardiomyocyte apoptosis. The protein expressions of integrin β3 and HIF1α were upregulated in acute myocardial infarction rat cardiac tissues compared with the control rat cardiac tissues. Our data suggest that integrin β3 plays a protective role in cardiomyocytes during hypoxia-induced apoptosis.

N-stearoylthrosine (NST), a synthesized anandamide (AEA) analogue, plays a neuroprotective role in neurodegenerative diseases and cerebrovascular diseases. Several studies have demonstrated that the endocannabinoids systems (ECS) are involved in the neuroprotective effects against cerebral ischemic injury. Oxygen-glucose deprivation (OGD)-induced neuronal injury elevated the levels of endocannabinoids and activated ECS. This research was conducted to investigate the neuroprotective effect of NST against OGD-induced neuronal injury in cultured primary cortical neurons and the potential mechanism involved. Cortical neurons were treated with NST at indicate concentrations for 30min prior to injury and OGD injured neurons were incubated with normal conditions for 0-24h. The best neuroprotective effect of NST against OGD-induced injury occurred at 10μM. All data indicated that the neuroprotective effect of NST against OGD-induced injury resulted from blocking anandamide membrane transporter (AMT) (IC50=11.74nM) and inhibiting fatty acid amide hydrolase activity (FAAH) (IC50=16.54nM). Our findings demonstrated that NST has an important role in cerebral ischemic injury pathological progression through activating cannabinoid receptors by inhibiting AEA inactivation system. These data suggested a potential role for NST in the therapeutic consideration of cerebral ischemic injury. However, inhibition of AEA inactivation system may provide a neuroprotective effect during cerebral ischemic injury.

p65 is a transcription factor that is involved in many physiological and pathologic processes. Here we report that p65 strongly binds to the miR-23a-27a-24 cluster promoter to up-regulate its expression. As bone marrow-derived cells differentiate into red blood cells in vitro, p65/miR-23a-27a-24 cluster expression increases sharply and then declines before the appearance of red blood cells, suggesting that this cluster is negatively related to erythroid terminal differentiation. Bioinformatic and molecular biology experiments confirmed that the miR-23a-27a-24 cluster inhibited the expression of the erythroid proteome and contributed to erythroleukemia progression. In addition, high level of the p65/miR-23a-27a-24 cluster was found in APL and AML cell lines and in nucleated peripheral blood cells from leukemia patients. Furthermore, anti-leukemia drugs significantly inhibited the expression of the p65/miR-23a-27a-24 cluster in leukemia cells. Administration of the p65 inhibitor parthenolide significantly improved hematology and myelogram indices while prolonging the life span of erythroleukemia mice. Meanwhile, stable overexpression of these three miRNAs in mouse erythroleukemia cells enhanced cell malignancy. Our findings thus connect a novel regulation pathway of the p65/miR-23a-27a-24 cluster with the erythroid proteome and provide an applicable approach for treating leukemia.

Anion exchanger 2 (AE2) mediates the exchange of C1-/HCO3- across the plasma membrane and plays a role in the regulation of intracellular pH. The present study showed that AE2 protein expression was upregulated immediately after exposure to either low (0.5 micromol/l) or high (1 and 2 micromol/l) concentrations of arsenic trioxide. This suggests that arsenic trioxide may act via regulation of intracellular pH. Changing the culture pH in NB4 cells modulated the degradation of promyelocytic leukaemia-retinoic acid receptor-alpha (PML-RARalpha), PML and RARalpha, which supported this hypothesis. DIDS (4,4'-diisothiocyanodihydrostilbene-2,2'-disulphonic acid) inhibited AE2 function, preventing the arsenic trioxide-induced degradation of RARalpha and low concentration showed synergistic effects on the expression of CD11c, which is related with cell differentiation. In addition, DIDS rescued the cells from 1 micromol/l arsenic trioxide-induced apoptosis. In conclusion, AE2 mediated the action of arsenic trioxide via regulation of intracellular pH and a novel pathway for the mechanism of action of arsenic trioxide is reported.

Successful application of arsenic trioxide (As2O3) in the treatment of acute promyelocytic leukemia (APL) has been attracting worldwide interest, but the exact mechanism for the action of As2O3 remains somewhat obscure. In the present work, we show for the first time that As2O3 facilitates the DIDS-sensitive anion transport activity of band 3 protein in red blood cells (RBCs) isolated from normal adults and APL patients. To elucidate the effect of As2O3 on band 3 protein, constructs encoding the full length of the band 3 transmembrane domain (mdb3) and its C-terminal deletion forms were transfected into yeast cells by a yeast display system. The results demonstrate that deletion of the C-terminal 16 residues of mdb3 (mdb3-d16) does not affect anion transport activity of mdb3 or its sensitivity to DIDS, but decreases its sensitivity to As2O3 in the yeast cell. More intriguingly, the forced expression of intact mdb3 by transfection significantly induces cell apoptosis in HeLa cells, to a higher degree than in cells transfected with mdb3-d16 or empty vector. Expression of activated caspase 3 in HeLa cells also indicates that the C-terminal 16 residues are important for mdb3-mediated apoptosis in cells treated with As2O3. Our results provide the first evidence that As2O3 enhances the anion transport activity of band 3 and the action is related with the C-terminal 16 residues of the protein.

Aims N-stearoyltyrosine (NsTyr) as an anandamide (AEA) analog has close relationships with AEA not only in structure but also in terms of biological actions of endocannabinoids. Since β-amyloid (Aβ)-induced primary neuronal injury involves the activation of the endocannabinoid systems (ECS), the protective effects of NsTyr against Aβ(1–40)-induced neuronal injury and the mechanism were studied systematically in this paper. Main methods Cortical neurons were incubated with Aβ(1–40) for 24 h. NsTyr was added to indicated concentrations 30 min prior to injury. Key findings The best effects of NsTyr on Aβ(1–40)-induced primary neuronal injury occurred at 10 μM. The mechanism of NsTyr on neuroprotective effects against Aβ(1–40)-induced cellular death first involved anti-apoptosis resulting from the activation of cannabinoid receptors, then the pre-receptor regulation of AEA by the inhibition of endocannabinoid inactivation. These data demonstrated that the protective effects of NsTyr on Aβ(1–40)-induced primary neuronal injury resulted from the inhibition of fatty acid amide hydrolase (FAAH) (IC 50 = 16.54 nM) and blocked AEA uptake mediated by anandamide membrane transporter (AMT) (IC 50 = 11.74 nM). Significance The activation of ECS by inhibiting the degradation of AEA is an effective pharmacological approach to suppress Aβ-induced neuropathic injury. Our research could result in a more realistic alternative for AD treatment.

Studies have shown that the expression of CD133, leucine-rich-repeat-containing G-protein-coupled receptor 5 (Lgr5), and ATP binding cassette (ABC)G2 proteins is associated with malignancy and poor prognosis in colon cancer. However, molecular regulation mechanism of the three proteins has not been elucidated. Here, we report that microRNA-142-3p (miR-142-3p) inhibits the expression of CD133, Lgr5, and ABCG2 in colon cancer cells by binding to both the 3′-untranslated region and the coding sequences of the three genes. The miR-142-3p was markedly decreased in colon cancer specimens, in which it was negatively correlated with the expression of CD133, Lgr5, and ABCG2. Reduction of miR-142-3p corresponds to poor differentiation and bigger tumor size in colon cancers. Moreover, miR-142-3p levels were reduced in cells that formed spheres compared to cells that were cultured in regular media. Transfection of miR-142-3p mimics in colon cancer cells downregulated cyclin D1 expression, induced G1 phase cell cycle arrest, and elevated the sensitivity of the cells to 5-fluorouracil. Furthermore, OCT4 suppressed miR-142-3p, and hypomethylation of the OCT4 promoter was associated with a reduction in miR-142-3p. Finally, the miR-142-3p inhibited the growth of colon cancer cells in vivo, which was accompanied by the downregulation of CD133, Lgr5, and ABCG2 in tumor tissues. Our results elucidate a novel regulation pathway in colon cancer cells and suggest a potential therapeutic approach for colon cancer therapy.

Infection with Helicobacter pylori (H. pylori) strains is linked to an increased risk of inflammation and gastric cancer. To investigate the effects of H. pylori on biological characteristics of gastric epithelial cells SGC-7901, derived from human adenocarcinoma, morphological appearances of both the pathogen and these cells, as well as features of attachment and internalization were observed by using transmission electron microscopy (TEM). We also investigated cell junctions and invasion by TEM and Transwell Invasion Assay. Cell proliferation and apoptosis were assessed by using chromogenic methylthiazol tetrazolium bromide (MTT) dye and flow cytometry. Three types of H. pylori were observed around, attaching to, or invading tumor cells. Cellular damage was characterized by vacuolar degeneration, dilated endoplasmic reticulum (ER), and reduction of organelles. Cell junctions and cell microvilli reduced or disappeared. H. pylori inhibited cell proliferation, whereas it had no effect on apoptosis. It also promoted gastric carcinoma cell invasion. H. pylori damages cell construction, destroys cell junctions, inhibits cell proliferation, promotes cell invasive ability, and, therefore, might accelerate the malignant progress and metastasis of gastric cancer.

Apoptosis is one of the major characteristics of delayed neuronal degeneration in neuronal injury following cerebral ischemia. Hypoxia-induced apoptosis may be co-regulated by HIF-1alpha as well as many other factors. In recent years, numerous studies concerning panaxynol (PNN) have been reported. However, whether PNN can show anti-hypoxia properties is still unknown. In this study, the protective effects of PNN on OGD-induced neuronal apoptosis and potential mechanisms were investigated. Pretreatment of the cells with PNN for 24h following exposure to OGD resulted in a significant elevation of cell survival determined by MTT assay, LDH assay, Hoechst staining and flow cytometric assessment. In addition to enhancing the expression of HIF-1alpha, PNN also normalized the caspase-3 expression/activation and increased the Bcl-2/Bax ratio. In our study, the increased level of HIF-1alpha with decreased cellular apoptosis suggested an important role for HIF-1alpha in hypoxic neurons. These results indicated that the neuroprotective effects of PNN on hypoxic neurons were at least partly due to up-regulation of HIF-1alpha and raised the possibility that PNN might reduce neurodegenerative disorders and ischemic brain diseases.