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The vulnerable plaque is a key distinguishing feature of atherosclerotic lesions that can cause acute atherothrombotic vascular disease. This study was designed to explore the effect of autophagy on mitochondria-mediated macrophage apoptosis and vulnerable plaques. Here, we generated the mouse model of vulnerable carotid plaque in ApoE -/- mice. Application of ApoE -/- mice with rapamycin (an autophagy inducer) inhibited necrotic core formation in vulnerable plaques by decreasing macrophage apoptosis. However, 3-methyladenine (an autophagy inhibitor) promoted plaque vulnerability through deteriorating these indexes. To further explore the mechanism of autophagy on macrophage apoptosis, we used macrophage apoptosis model in vitro and found that 7-ketocholesterol (7-KC, one of the primary oxysterols in oxLDL) caused macrophage apoptosis with concomitant impairment of mitochondria, characterized by the impairment of mitochondrial ultrastructure, cytochrome c release, mitochondrial potential dissipation, mitochondrial fragmentation, excessive ROS generation and both caspase-9 and caspase-3 activation. Interestingly, such mitochondrial apoptotic responses were ameliorated by autophagy activator, but exacerbated by autophagy inhibitor. Finally, we found that MAPK-NF-κB signalling pathway was involved in autophagy modulation of 7-KC-induced macrophage apoptosis. So, we provide strong evidence for the potential therapeutic benefit of macrophage autophagy in regulating mitochondria-mediated apoptosis and inhibiting necrotic core formation in vulnerable plaques., (© 2019 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

The present study aims to investigate the roles of steroidogenic acute regulatory protein (StAR) in Yangjing Capsule (YC) induced anti-apoptotic effects on Leydig cells and the related mechanism. Leydig tumor cells (MLTC-1) were cultured and treated with YC, and immunofluorescence assay was performed to examine the expression of StAR; furthermore, luciferase reporter assay was conducted to evaluate the impact of YC on StAR promoter; next, MLTC-1 cells were treated with StAR small interfering RNA (siRNA), and flow cytometry was carried out to examine the effect of StAR siRNA on the apoptosis of the cells; furthermore, quantitative (q)RT-PCR and Western blot methods was used to determine the expression of StAR and apoptosis related molecules Bcl-2, Bax and Caspase-3 on both mRNA and protein levels in different groups; finally the secretion of testosterone in different groups was examined by radioimmunoassay. We observed that the YC can increase the expression of StAR in a dose-dependent manner, and YC can activate the promoter of StAR; moreover, transfection of StAR siRNA can block YC induced anti-apoptotic effects and increased production of testosterone. In conclusion, our results suggested that YC might suppress the apoptosis of MLTC-1 cells and enhance the production of testosterone through regulating the expression of StAR.

Neuroglobin (Ngb) overexpression is considered as an intrinsic neuroprotective response. Therefore, exogenous Ngb increased in brain tissues has become a promising therapeutic strategy for neurological diseases. Previous studies demonstrated that transactivator of transcription (TAT) protein transduction domain was able to mediate synthetic Ngb entrance into neurons, and then protected brain from hypoxia-ischemic injury. However, the role of recombinant Ngb on early brain injury following subarachnoid hemorrhage (SAH) has not been elucidated. The objectives of this study were to investigate the expression of endogenous Ngb in brain using a rabbit model of SAH, and to verify whether TAT-Ngb fusion protein could be delivered into brain parenchyma, as well as to explore the neuroprotective effect of Ngb and its possible mechanisms. We found that Ngb expressions were up regulated in the transcript and protein levels in a similar time dependent manner after SAH as compared to the sham group. Moreover, TAT-Ngb fusion protein was successfully generated and transferred into brain neurons. Compared with the saline- and Ngb-treated group, neuronal viabilities and neurological outcomes were significantly improved 72 h post-SAH in the TAT-Ngb-treated group. Likewise, anti-apoptotic Bcl-2 protein was also elevated obviously. Conversely, pro-apoptotic factors including caspase 3, caspase 9 and Bax were greatly decreased after TAT-Ngb treatment. Our results suggest that Ngb plays a neuroprotective effect in rabbits suffering from SAH possibly through inhibiting the SAH-induced activation of mitochondria apoptotic pathway. Furthermore, TAT-mediated Ngb delivery into brain may be a promising therapeutic approach., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

The mechanisms by which oxidative stress induces spinal cord neuron death has not been completely understood. Investigation on the molecular signal pathways involved in oxidative stress-mediated neuronal death is important for development of new therapeutics for oxidative stress-associated spinal cord disorders. In current study we examined the role of heme oxygenase-1 (HO-1) in the modulation of MLK3/MKK7/JNK3 signaling, which is a pro-apoptotic pathway, after treating primary spinal cord neurons with H 2 O 2 . We found that MLK3/MKK7/JNK3 signaling was substantially activated by H 2 O 2 in a time-dependent manner, demonstrated by increase of activating phosphorylation of MLK3, MKK7 and JNK3. H 2 O 2 also induced expression of HO-1. Transduction of neurons with HO-1-expressing adeno-associated virus before H 2 O 2 treatment introduced expression of exogenous HO-1 in neurons. Exogenous HO-1 reduced phosphorylation of MLK3, MKK7 and JNK3. Consistent with its inhibitory effect on MLK3/MKK7/JNK3 signaling, exogenous HO-1 decreased H 2 O 2 -induced neuronal apoptosis and necrosis. Furthermore, we found that exogenous HO-1 inhibited expression of Cdc42, which is crucial for MLK3 activation. In addition, HO-1-induced down-regulation of MLK3/MKK7/JNK3 signaling might be related to up-regulation of microRNA-137 (mir-137). A mir-137 inhibitor alleviated the inhibitory effect of HO-1 on JNK3 activation. This inhibitor also increased neuronal death even when exogenous HO-1 was expressed. Therefore, our study suggests a novel mechanism by which HO-1 exerted its neuroprotective efficacy on oxidative stress.

Objectives: Neuroglobin (Ngb), an identified globin in vertebrate brain, is a potential novel protective protein against brain ischemia. In our previous study, the human immunodeficiency virus trans-activator of transcription (TAT) protein transduction domain successfully delivered exogenous Ngb into neurons in the mouse, and protected the brain from cerebral ischemia-induced apoptosis. The aim of this study is to investigate the role of TAT-Ngb in attenuating oxygen-glucose deprivation (OGD) induced apoptosis and to explore the possible mechanism., Methods: Nerve growth factor (NGF)-induced PC12 cells were divided into (1) the control group, (2) the OGD group (just OGD), (3) the Ngb treatment group (OGD and Ngb treatment), and (4) the TAT-Ngb treatment group (OGD and TAT-Ngb treatment). Cell viability and apoptosis were assessed by the MTT assay and the AnnexinV/propidium iodide (PI) staining, respectively. The mitochondrial transmembrane potential was measured by JC-1 staining. Caspase-3, Bcl-2, Bax, Stat3, Jak2, and Akt were determined by western blot analysis., Results: Trans-activator of transcription effectively delivered Ngb into NGF-induced PC12 cells. Neuroglobin-mediated neuroprotection rescued cultured cells from OGD. We also confirmed previous findings that TAT-Ngb inhibited mitochondrial apoptosis following OGD. Inhibition of apoptosis by Ac-DEVD-CHO showed that caspase-3 was a crucial factor in OGD-induced apoptosis cascades. AG490, a specific Jak2 inhibitor, attenuated the protective effects of TAT-Ngb. The TAT-Ngb promoted expression of the anti-apoptotic protein Bcl-2 through the Jak2/Stat3 signal pathway, and inhibited apoptosis by blocking caspase-3 activation, while the Jak-Akt-Stat3 signal network was not involved., Conclusion: Our results demonstrate that TAT-Ngb can protect neuron-like cells against OGD-induced apoptosis by activating the Jak2/Stat3 pathway.

Preeclampsia (PE) is a pregnancy-related syndrome characterized by new-onset hypertension and proteinuria after gestational 20 weeks. Oxidative stress, resulting from the imbalance between the production of oxidants and antioxidants in placentas, is recognized as a key pathology of PE. To date, the molecules that regulate antioxidants production remain unclear. CD151, a member of tetraspanins, is an important regulator of many physiological functions. However, the function of CD151 in oxidative stress and its association with pregnancy-related complications are currently unknown. In the present study, we have demonstrated that CD151 was a key regulator of antioxidants in placentas. Compared with the placentas of the controls, the placentas of PE patients exhibited decreased CD151 expression accompanying with decreased antioxidant gene expression (HO-1, NQO-1, GCLC and SOD-1). In vitro , overexpression of CD151 in trophoblast cells could enhance HO-1, NQO-1, GCLC and SOD-1 expression but downregulation of CD151 decreased those antioxidant genes expression, which indicates CD151 is the upstream of antioxidants. Importantly, the phenotype of PE (hypertension and proteinuria) was mimicked in the downregulating CD151 induced mouse model. Moreover, the beneficial effect of CD151 in trophoblast cells was hindered when ERK and Nrf2 signaling were blocked. Overall, our results revealed CD151 might be a new target for PE treatment. Image 1 • CD151 decreases in trophoblasts of placentas from PE patients. • CD151 exerts antioxidative ability via ERK/Nrf2 pathway. • Downregulation of CD151 in pregnant mice mimics the phenomenon of PE. [ABSTRACT FROM AUTHOR]

Aims: MicroRNA-186 (miR-186) has been shown to be involved in various types of cancer. The purpose of this study was to investigate the expression level and functional role of miR-186 in human cutaneous malignant melanoma cells.Subjects and Methods: Expression of miR-186 was analyzed in human cutaneous malignant melanoma (CMM) cell lines SK-MEL-1, G-361, A375 and A875, and human normal epidermal melanocytes cell line HEMn-LP by quantitative reverse transcription polymerase chain reaction (qRT-PCR). Additionally, the functional role of miR-186 on melanoma cells was investigated by transfection of miR-186 mimic followed by analyses of cell proliferation, apoptosis, and metastasis.Results: We found that the expression levels of miR-186 were decreased in CMM cell lines compared with normal epidermal melanocytes cell line. Moreover, overexpression of miR-186 inhibited cells proliferation through abrogating the G1-S transition, and reduced cells migration and invasion.Conclusions: Our findings suggested that miR-186 exhibit an inhibitory effect on CMM cell proliferation, migration, and invasion; thus, may serve as a potential therapeutic target for human CMM intervention. [ABSTRACT FROM AUTHOR]

MIR503HG is a 786 bp long lncRNA located on chromosome Xq26.3, and it can regulate diverse cellular processes. The pathogenesis of adenomyosis (AD) is associated with endometrial stromal cells (ESCs). The present study investigated the specific role of MIR503HG in AD pathogenesis and progression using ESCs derived from the endometrium of patients with AD as a model. Expression of MIR503HG and microRNA (miR)-191 were assessed using reverse transcription-quantitative PCR. An immunocytochemistry assay was used to detect cytokeratin- or vimentin-positive ESCs. Transfections of ESCs with MIR503HG overexpression plasmid, short hairpin-MIR503HG and miR-191 inhibitor were performed. ESC viability, migration, invasion and apoptosis were evaluated using Cell Counting Kit-8, Transwell and flow cytometry assays. The association between MIR503HG and miR-191 was predicted by StarBase and confirmed using a dual-luciferase reporter assay. Expression of epithelial-mesenchymal transition-related markers (E-cadherin and N-cadherin) and Wnt/β-catenin pathway-related molecules (β-catenin) in ESCs were analyzed by western blotting. The isolated ESCs were vimentin-positive and cytokeratin-negative. MIR503HG was lowly expressed in the endometrial tissues derived from patients with AD. MIR503HG overexpression hindered ESC viability, migration and invasion while enhancing the apoptosis and downregulating miR-191 expression. MIR503HG knockdown induced the opposite effects, accompanied by downregulation of the E-cadherin expression and upregulation of N-cadherin and β-catenin levels. MIR503HG directly targeted miR-191 that was highly expressed in endometrial tissues derived from patients with AD. In ESCs, downregulation of miR-191 inhibited the viability, migration and invasion and the expression of N-cadherin and β-catenin levels while enhancing the apoptosis and E-cadherin expression in ESCs. Moreover, downregulation of miR-191 partially reversed the effect of MIR503HG knockdown. Collectively, overexpressed MIR503HG impeded the proliferation and migration of ESCs derived from endometrium of patients with AD, while promoting apoptosis via inhibition of the Wnt/β-catenin pathway via targeting miR-191. [ABSTRACT FROM AUTHOR]

Abstract Taking sophoridine as a lead compound, 58 sophoridine derivatives were designed, synthesized and evaluated for their antiproliferative activity in the HepG2 cancer cell line. Among the 58 compounds, 33 compounds showed potent antiproliferative activity with IC 50 less than 10 μM. Compound 5w showed the most potent anti-proliferative activity in the HepG2 cancer cell line. Thus, we further extended our characterization of the antiproliferative activity of 5w in six cancer cell lines (HepG2, SMMC-7721, Hela, CNE1, CNE2 and MCF7). The representative compound 5w displayed robust anti-proliferative activities in all the tested cell lines with IC 50 values in range of 0.93–1.89 μM which were much lower than that of sophoridine. Here, we report the structure–activity relationships (SAR) in a sophoridine series of compounds, which indicated that introduction of N-benzyl indole group on the 14-carbon atom of sophoridine can significantly enhance the antiproliferative activity. By molecular docking and enzymatic assay, compound 5w was found to be able to inhibit the activity of DNA Topo I. Furthermore, apoptosis assay displayed that compound 5w could significantly induce the apoptosis of HepG2 cells in a dose-dependent manner by activating caspase-3, increasing expression of cleaved caspase-3 and reducing the ratio of Bcl-2/Bax. The in vivo antitumor assay demonstrated that 5w suppressed the growth of HepG2 xenografts in nude mice without any obvious side effects. Graphical abstract Image 1 Highlights • 58 sophoridine derivatives were designed, synthesized and evaluated. • 5w exhibited potent antitumor activity in vitro. • 5w inhibited the activity of DNA Topo I. • 5w arrested tumor growth in vivo. [ABSTRACT FROM AUTHOR]

Using matrine ( 1 ) as the lead compound, a series of new 14-( N -substituted-2-pyrrolemethylene) matrine and 14-( N -substituted-indolemethylene) matrine derivatives was designed and synthesized for their potential application as anticancer agents. The structure of these compounds was characterized by 1 H NMR, 13 C NMR and ESI-MS spectral analyses. The target compounds were evaluated for their in vitro cytotoxicity against three human cancer cell lines (SMMC-7721, A549 and CNE2). The results revealed that compound A6 and B21 displayed the most significant anticancer activity against three cancer cell lines with IC 50 values in range of 3.42–8.05 μM, which showed better activity than the parent compound (Matrine) and positive control Cisplatin. Furthermore, the Annexin V-FITC/PI dual staining assay revealed that compound A6 and B21 could significantly induce the apoptosis of SMMC-7721 and CNE2 cells in a dose-dependent manner. The cell cycle analysis also revealed that compound A6 could cause cell cycle arrest of SMMC-7721 and CNE2 cells at G2/M phase. [ABSTRACT FROM AUTHOR]

• Testicular inflammation induced by aging is identified as a potential risk factor for late-onset hypogonadism. • TNFAIP3, an anti-inflammatory factor, demonstrates close involvement in various aging and inflammatory diseases. • Our study reveals TNFAIP3 promotes testosterone production and suppresses apoptosis in Leydig cells within the aging inflammatory microenvironment. The aging inflammatory microenvironment surrounding Leydig cells is linked to reduced testosterone levels in males. Tumor necrosis factor alpha-induced protein 3 (TNFAIP3) acts as a critical anti-inflammatory factor in various aging-related diseases. This study aims to investigate the protective effect of TNFAIP3 on testosterone production in Leydig cells under an aging inflammatory microenvironment. Bioinformatics analysis examined TNFAIP3 expression differences in aging rat testes and validated the findings in aging mouse testes. In vitro models of inflammation were established using two Leydig cell lines, with tumor necrosis factor alpha (TNF-α) as the inflammatory factor. Lentiviral transduction was utilized to manipulate TNFAIP3 expression in these cell lines. Transcriptomic sequencing identified differentially expressed genes in TNFAIP3-overexpressing cells. Bioinformatics analysis and validation experiments revealed increased inflammatory signaling and elevated TNFAIP3 expression in aging rat and mouse testes. TNFAIP3 knockdown worsened testosterone synthesis inhibition and apoptosis in cells, while TNFAIP3 overexpression reversed these effects. Transcriptome analysis identified alterations in the P38MAPK pathway following TNFAIP3 overexpression. TNFAIP3 knockdown enhanced TNF-induced P38MAPK signaling, whereas its overexpression attenuated this effect. TNFAIP3 was found to regulate testosterone synthesis by upregulating CEBPB expression. TNFAIP3 exhibits inhibitory effects on apoptosis and promotes testosterone production in Leydig cells. The protective influence of TNFAIP3 on Leydig cells within an inflammatory microenvironment is likely mediated through by inhibiting the P38MAPK pathway and upregulating CEBPB expression. [ABSTRACT FROM AUTHOR]

The mechanism by which spinal cord injury (SCI) induces neuronal death has not been thoroughly understood. Investigation on the molecular signal pathways involved in SCI-mediated neuronal apoptosis is important for development of new therapeutics for SCI. In the current study, we explore the role of heme oxygenase-1 (HO-1) in the modulation of mixed lineage kinase 3/mitogen-activated protein kinase kinase/cJUN N-terminal kinase 3 (MLK3/MKK7/JNK3) signaling, which is a pro-apoptotic pathway, after SCI. We found that MLK3/MKK7/JNK3 signaling was activated by SCI in a time-dependent manner, demonstrated by increase in activating phosphorylation of MLK3, MKK7, and JNK3. SCI also induced HO-1 expression. Administration of HO-1-expressing adeno-associated virus before SCI introduced expression of exogenous HO-1 in injured spinal cords. Exogenous HO-1 reduced phosphorylation of MLK3, MKK7, and JNK3. Consistent with its inhibitory effect on MLK3/MKK7/JNK3 signaling, exogenous HO-1 decreased SCI-induced neuronal apoptosis and improved neurological score. Further, we found that exogenous HO-1 inhibited expression of cell division cycle 42 (Cdc42), which is crucial for MLK3 activation. In vitro experiments indicated that Cdc42 was essential for neuronal apoptosis, while transduction of neurons with HO-1-expressing adeno-associated virus significantly reduced neuronal apoptosis to enhance neuronal survival. Therefore, our study disclosed a novel mechanism by which HO-1 exerted its neuroprotective efficacy. Our discovery might be valuable for developing a new therapeutic approach for SCI. [ABSTRACT FROM AUTHOR]