Annexin V, a protein with high affinity to phosphatidylserine (PS) in a calcium dependent manner, has been widely used to probe apoptosis. Annexin V in inhibiting engulfment of apoptotic cells by macrophages had been reported to increase the immunogenicity of tumor cells undergoing apoptosis. However, far less is known about its multiple properties, especially in cancer therapies. Here we found that Annexin V had a good anti-tumor activity in murine melanomaxenograft model. Treatment with Annexin V showed significant reduction in tumor size and remarkable tumor necrosis areas. The serum level of VEGF was downregualted by Annexin V both in normal mice and mice bearing tumor, suggesting that its new role on impeding tumor angiogenesis. In Silico analysis using Oncomine database, we also found the negative correlation of AnnexinV and VEGF both in skin and melanoma. The decreased Annexin V expression shows linearity relation with the elevated VEGF expression. These data provided a possibility that Annexin V can be used as a novel angiogenesis inhibitor in tumor therapy.

This study aims to determine the effects and mechanism of action of muscone on the biological activity of the gastric cancer cell lines SGC-7901 and MGC-803 (proliferation, apoptosis, invasion, and migration) in vitro. An optimal muscone concentration was determined using MTT and cell apoptosis tests. The SGC-7901 and MGC-803 cells were divided into five groups: normal control, muscone, miRNA, muscone + miRNA, and muscone + miRNA inhibitor. Cell proliferation rate, apoptosis rate, cell cycle phase distribution, number of invading cells, and wound healing rate were compared among the five groups using MTT, flow cytometry, transwell, and wound healing assays. Relative expression levels of the proteins PI3K, AKT, P21, c-Myc, MMP-2, and MMP-9 were measured by Western blot. Compared with the control group, the groups treated with muscone and miRNA showed significantly lower cell proliferation rate, number of invading cells, and wound healing rate (p < .05 for all), but significantly higher rates of cell apoptosis rate and numbers of cells in the G1 phase (p < .05 for all). These groups also showed significantly lower expression of the proteins PI3K, AKT, c-Myc, MMP-2, and MMP-9 but significantly increased expression of the protein P21 (p < .05). Transfecting muscone-treated SGC-7901 and MGC-803 cells with miRNA-145 inhibitor resulted in a significant recovery of biological activity (p < .05). Muscone suppresses the biological activity of SGC-7901 and MGC-803 gastric cancer cells in vitro via regulation of miRNA-145.

Transforming growth factor‑β1 (TGF‑β1)‑induced epithelial‑mesenchymal transition (EMT) serves a significant role in pulmonary fibrosis (PF). Increasing evidence indicates that microRNAs (miRNAs or miRs) contribute to PF pathogenesis via EMT regulation. However, the role of miR‑483‑5p in PF remains unclear. Therefore, the present study investigated the potential effect of miR‑483‑5p on TGF‑β1‑induced EMT in PF. It was found that the expression of miR‑483‑5p was upregulated in both PF tissue and A549 cells treated with TGF‑β1, whereas expression of Rho GDP dissociation inhibitor 1 (RhoGDI1) was downregulated. miR‑483‑5p mimic transfection promoted TGF‑β1‑induced EMT; by contrast, miR‑483‑5p inhibitor inhibited TGF‑β1‑induced EMT. Also, miR‑483‑5p mimic decreased RhoGDI1 expression, whereas miR‑483‑5p inhibitor increased RhoGDI1 expression. Furthermore, dual‑luciferase reporter gene assay indicated that miR‑483‑5p directly regulated RhoGDI1. Moreover, RhoGDI1 knockdown eliminated the inhibitory effect of the miR‑483‑5p inhibitor on TGF‑β1‑induced EMT via the Rac family small GTPase (Rac)1/PI3K/AKT pathway. In conclusion, these data indicated that miR‑483‑5p inhibition ameliorated TGF‑β1‑induced EMT by targeting RhoGDI1 via the Rac1/PI3K/Akt signaling pathway in PF, suggesting a potential role of miR‑483‑5p in the prevention and treatment of PF.

Gastric cancer is an important cancer type worldwide, the anti‑angiogenic agent BC001 can target the vascular endothelial growth factor receptor 2 (VEGFR2), and significantly suppresses the growth of gastric cancer BGC823 cells in vitro and in vivo. However, numerous results indicated that antiangiogenic drugs could induce autophagy, and the inhibition of autophagy enhanced the anticancer effects of antiangiogenic agents. In the present study, hydroxychloroquine (HCQ), an inhibitor of autophagy, enhanced the antiproliferative and proapoptotic effects of BC001 in vitro. Furthermore, HCQ enhanced the antitumor effects of BC001 on BGC823 xenograft tumors in vivo. Of note, BC001 neither induced nor inhibited autophagy. RNA‑sequencing results revealed that HCQ regulated autophagy or lysosomal‑associated genes, such as tumor protein p53‑inducible nuclear protein 1, interleukin (IL)1B, tumor necrosis factor (TNF), Mediterranean fever, ubiquitin specific peptidase 36, IL6, neuraminidase (NEU)1, ATP‑binding cassette subfamily A member 1, proprotein convertase subtilisin/kexin type 9, myelin basic protein and NEU3. Importantly, HCQ was determined to affect multiple pathways, including 'negative regulation of endothelial cell proliferation', 'blood vessel remodeling', 'cell surface receptor signaling pathways' and 'notch receptor processing' associated with 'signal transduction', 'cancers' and 'immune system', through regulating C‑X‑C motif chemokine ligand 8, TNF, IL6, intercellular adhesion molecule 1 and FOS genes. In summary, HCQ was proposed to enhance the anticancer effects of BC001 in gastric cancer via complex mechanisms.

The aim of the present study was to explore the effect of baicalin on liver hypoxia/reoxygenation (H/R) injury and the possible mechanism involved. A cellular H/R model was established and cells were treated with 50, 100 and 200 µmol/l baicalin. Following reoxygenation for 6 h, cell viability, lactate dehydrogenase (LDH), B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax), caspase 3 and cleaved caspase 3 were assessed. Furthermore, levels of endoplasmic reticulum stress markers binding of immunoglobulin protein (BIP) and CCAAT/enhancer-binding protein homologous protein (CHOP) and autophagy markers microtubule-associated proteins 1A/1B light chain 3B (LC3) and beclin 1 were measured. To confirm the involvement of autophagy in baicalin-mediated attenuation of H/R injury, the autophagy inhibitor 3-methyladenine (3-MA) was administered. The results revealed that baicalin administration increased cell viability and decreased LDH levels, most notably at a dosage of 100 µmol/l. Baicalin pretreatment also downregulated the expression of caspase 3, cleaved caspase 3 and Bax, while upregulating the expression of Bcl-2. Furthermore, BIP and CHOP were decreased while LC3 and beclin-1 were significantly increased by baicalin pretreatment. Inhibiting autophagy using 3-MA, resulted in a significant decrease in LC3-II, beclin-1 and LDH, as well as increase in the expression of BIP, CHOP, caspase 3, cleaved caspase 3 and Bax. Bcl-2 and cell viability were also decreased. In conclusion, the results of the present study indicate that baicalin exerts a protective effect on liver H/R injury and this may be achieved via the induction of autophagy.

Diabetic kidney disease (DKD) can lead to end-stage kidney disease (ESKD) and mortality; however, few mechanistic biomarkers are available for high-risk patients, especially those without macroalbuminuria. Urine from participants with diabetes from the Chronic Renal Insufficiency Cohort (CRIC) study, the Singapore Study of Macro-angiopathy and Micro-vascular Reactivity in Type 2 Diabetes (SMART2D), and the American Indian Study determined whether urine adenine/creatinine ratio (UAdCR) could be a mechanistic biomarker for ESKD. ESKD and mortality were associated with the highest UAdCR tertile in the CRIC study and SMART2D. ESKD was associated with the highest UAdCR tertile in patients without macroalbuminuria in the CRIC study, SMART2D, and the American Indian study. Empagliflozin lowered UAdCR in nonmacroalbuminuric participants. Spatial metabolomics localized adenine to kidney pathology, and single-cell transcriptomics identified ribonucleoprotein biogenesis as a top pathway in proximal tubules of patients without macroalbuminuria, implicating mTOR. Adenine stimulated matrix in tubular cells via mTOR and stimulated mTOR in mouse kidneys. A specific inhibitor of adenine production was found to reduce kidney hypertrophy and kidney injury in diabetic mice. We propose that endogenous adenine may be a causative factor in DKD.

Our previous studies suggested that both hydrogen gas (H₂) and nitric oxide (NO) could enhance the postharvest freshness of cut flowers. However, the crosstalk of H₂ and NO during that process is unknown. Here, cut lilies (Lilium "Manissa") were used to investigate the relationship between H₂ and NO and to identify differentially accumulated proteins during postharvest freshness. The results revealed that 1% hydrogen-rich water (HRW) and 150 μM sodium nitroprusside (SNP) significantly extended the vase life and quality, while NO inhibitors suppressed the positive effects of HRW. Proteomics analysis found 50 differentially accumulated proteins in lilies leaves which were classified into seven functional categories. Among them, ATP synthase CF1 alpha subunit (chloroplast) (AtpA) was up-regulated by HRW and down-regulated by NO inhibitor. The expression level of LlatpA gene was consistent with the result of proteomics analysis. The positive effect of HRW and SNP on ATP synthase activity was inhibited by NO inhibitor. Meanwhile, the physiological-level analysis of chlorophyll fluorescence and photosynthetic parameters also agreed with the expression of AtpA regulated by HRW and SNP. Altogether, our results suggested that NO might be involved in H₂-improved freshness of cut lilies, and AtpA protein may play important roles during that process.

Human periodontal ligament cells (PDLCs) play an important role in periodontal tissue stabilization and function. In the process of osteogenic differentiation of PDLSCs, the regulation of molecular signal pathways are complicated. In this study, the sequencing results of three datasets on GEO were used to comprehensively analyze the miRNA-mRNA network during the osteogenic differentiation of PDLSCs. Using the GSE99958 and GSE159507, a total of 114 common differentially expressed genes (DEGs) were identified, including 62 up-regulated genes and 52 down-regulated genes. GO enrichment analysis was performed. The up-regulated 10 hub genes and down-regulated 10 hub genes were screened out by protein-protein interaction network (PPI) analysis and STRING in Cytoscape. Similarly, differentially expressed miRNAs (DEMs) were selected by limma package from GSE159508. Then, using the miRwalk website, we further selected 11 miRNAs from 16 DEMs that may have a negative regulatory relationship with hub genes. In vitro RT-PCR verification revealed that nine DEMs and 18 hub genes showed the same trend as the RNA-seq results during the osteogenic differentiation of PDLSCs. Finally, using miR-584-5p inhibitor and mimics, it was found that miR-584-5p negatively regulates the osteogenic differentiation of PDLSCs in vitro. In summary, the present results found several potential osteogenic-related genes and identified candidate miRNA-mRNA networks for the further study of osteogenic differentiation of PDLSCs.

Endothelial cells are an important component of the heart and vasculature and form a crucial link between the cardiovascular system and the immune system. Sestrin 1 (SESN1) has an important role in atherosclerosis by inhibiting NOD‑like receptor family pyrin domain containing 3 inflammasome activation. However, whether SESN1 is involved in human umbilical vein endothelial cell (HUVEC) injury caused by atherosclerosis has remained to be elucidated. The present study aimed to investigate the functions of SESN1 in the inflammatory response, apoptosis and endothelial‑mesenchymal transition (EndMT) of HUVECs following stimulation with oxidized low‑density lipoprotein (Ox‑LDL). SESN1 expression at the mRNA and protein levels was detected using reverse transcription‑quantitative PCR (RT‑qPCR) and western blot analysis. Following SESN1 overexpression in Ox‑LDL‑stimulated HUVECs, cell viability was determined using a Cell Counting Kit‑8 assay. Terminal deoxynucleotidyl transferase‑mediated nick‑end labeling staining was employed to detect cell apoptosis and western blot analysis was used to determine the levels of apoptosis‑related proteins. RT‑qPCR, ELISA and western blot were utilized to determine the levels of inflammatory factors. Immunofluorescence staining, RT‑qPCR and western blot analysis were employed to assess the EndMT of Ox‑LDL‑stimulated HUVECs. The results revealed that SESN1 exhibited a low expression in HUVECs following Ox‑LDL stimulation. SESN1 overexpression suppressed inflammation, apoptosis and EndMT in Ox‑LDL‑induced HUVECs. In addition, SESN1 stimulated adenosine monophosphate‑activated protein kinase catalytic subunit α1/sirtuin 1 signaling to suppress Ox‑LDL receptor‑1 expression. An AMPK and SIRT1 inhibitor reversed the effects of SESN1 overexpression on the inflammatory response, apoptosis and EndMT of HUVECs exposed to Ox‑LDL. Taken together, the present study demonstrated that SENS1 exerts a suppressive effect on Ox‑LDL‑induced inflammation, apoptosis and EndMT of HUVECs, suggesting that SENS1 may be used as a novel biomarker for endothelial injury‑related disorders.

This study is aimed to elucidate the mechanisms underlying the role of miR-485-5p in small cell lung cancer (SCLC). The expression of miR-485-5p were quantified with real time quantitative PCR and it was found that the level of miR-485-5p was lower in SCLC tissues than normal tissues. In cultured SCLC cell lines, overexpression of miR-485-5p reduced cell proliferation, migration, and invasion in vitro, whereas knockdown of miR-485-5p performed contrary. FLOT2 expression was obviously upregulated and negatively correlated with miR-485-5p expression level in SCLC tissues. Overexpression of miR-485-5p significantly inhibited the protein expression of flotillin-2 (FLOT2) in cultured SCLC cells. Luciferase reporter assay confirmed that FLOT2 was a direct target of miR-485-5p in SCLC cells. It is concluded that miR-485-5p, as a tumor suppressor, inhibits the growth and metastasis in SCLC by targeting FLOT2. Upregulation of miR-485-5p expression may be an attractive strategy for SCLC therapy.

Candida glabrata is an emerging human fungal pathogen that is frequently drug tolerant, resulting in difficulties in treatment and a higher mortality in immunocompromised patients. The calcium-activated protein phosphatase calcineurin plays critical roles in controlling drug tolerance, hyphal growth, and virulence in diverse fungal pathogens via distinct mechanisms involving survival in serum or growth at host temperature (37° and higher). Here, we comprehensively studied the calcineurin signaling cascade in C. glabrata and found novel and uncharacterized functions of calcineurin and its downstream target Crz1 in governing thermotolerance, intracellular architecture, and pathogenesis in murine ocular, urinary tract, and systemic infections. This represents a second independent origin of a role for calcineurin in thermotolerant growth of a major human fungal pathogen, distinct from that which arose independently in Cryptococcus neoformans. Calcineurin also promotes survival of C. glabrata in serum via mechanisms distinct from C. albicans and thereby enables establishment of tissue colonization in a murine systemic infection model. To understand calcineurin signaling in detail, we performed global transcript profiling analysis and identified calcineurin- and Crz1-dependent genes in C. glabrata involved in cell wall biosynthesis, heat shock responses, and calcineurin function. Regulators of calcineurin (RCN) are a novel family of calcineurin modifiers, and two members of this family were identified in C. glabrata: Rcn1 and Rcn2. Our studies demonstrate that Rcn2 expression is controlled by calcineurin and Crz1 to function as a feedback inhibitor of calcineurin in a circuit required for calcium tolerance in C. glabrata. In contrast, the calcineurin regulator Rcn1 activates calcineurin signaling. Interestingly, neither Rcn1 nor Rcn2 is required for virulence in a murine systemic infection model. Taken together, our findings show that calcineurin signaling plays critical roles in thermotolerance and virulence, and that Rcn1 and Rcn2 have opposing functions in controlling calcineurin signaling in C. glabrata.

BACKGROUND Microglia reside in the spinal cord plays a key role in the onset, progression of post-spinal cord injury (SCI) neuroinflammation. Curcumin has been shown to exhibit diverse anti-inflammatory and anti-tumor activities. The aim of this study was to explore the effect of curcumin on the inflammatory response in lipopolysaccharide (LPS)-activated microglia and its mechanism. MATERIAL AND METHODS The expression levels of phosphorylated-p65 (p-p65), tumor necrosis factor (TNF)-alpha, interleukin (IL)-1ß, and IkappaB kinase ß (IKKß) were examined by western blot assay. MiR-199b-5p expression was detected by quantitative real-time polymerase chain reaction assay. The putative binding sites of miR-199b-5p in IKKß 3'UTR were predicted by bioinformatics, and direct interaction between miR-199b-5p and IKKß was verified by dual-luciferase reporter assay and RNA-immunoprecipitation assay. RESULTS Curcumin significantly suppressed inflammatory response induced by LPS by inactivation of nuclear factor kappa B (NF-kappaB) in microglial cells, as reflected by the decreased levels of p-p65, as well as the pro-inflammatory mediators, including inducible nitric oxide synthase (iNOS), TNF-alpha, and IL-1ß. Moreover, curcumin increased the level of miR-199b-5p and decreased IKKß expression in activated microglial cells. Knockdown of miR-199b-5p or overexpression of IKKß reversed the inhibitory effect of curcumin on inflammatory response and NF-kappaB activation. MiR-199b-5p directly targeted IKKß and suppressed its expression. Silencing of IKKß abolished miR-199b-5p-stimulated inflammatory cytokines production and NF-kappaB activation. CONCLUSIONS Curcumin attenuated neuroinflammation induced by LPS through regulating miR-199b-5p/IKKß/NF-kappaB axis in microglia.

The Prp8 intein is one of the most widespread eukaryotic inteins, present in important pathogenic fungi, including Cryptococcus and Aspergillus species. Because the processed Prp8 carries out essential and non-redundant cellular functions, a Prp8 intein inhibitor is a mechanistically novel antifungal agent. In this report, we demonstrated that cisplatin, an FDA-approved cancer drug, significantly arrested growth of Prp8 intein-containing fungi C. neoformans and C. gattii, but only poorly inhibited growth of intein-free Candida species. These results suggest that cisplatin arrests fungal growth through specific inhibition of the Prp8 intein. Cisplatin was also found to significantly inhibit growth of C. neoformans in a mouse model. Our results further showed that cisplatin inhibited Prp8 intein splicing in vitro in a dose-dependent manner by direct binding to the Prp8 intein. Crystal structures of the apo- and cisplatin-bound Prp8 inteins revealed that two degenerate cisplatin molecules bind at the intein active site. Mutation of the splicing-site residues led to loss of cisplatin binding, as well as impairment of intein splicing. Finally, we found that overexpression of the Prp8 intein in cryptococcal species conferred cisplatin resistance. Overall, these results indicate that the Prp8 intein is a novel antifungal target worth further investigation.

SHH subgroup medulloblastoma (SHH-MB) is one of the most common malignant pediatric tumors that arises in the cerebellum. Previously, we showed that RNA m6A methylation participates in regulation of cerebellar development. Here we investigate whether dysregulated m6A methylation contributes to tumorigenesis of SHH-MB. We show that high expression of m6A methyltransferase METTL3 associates with worse survival in the patients with SHH-MB. A large number of hypermethylated transcripts are identified in SHH-MB tumor cells by m6A-seq. We find that METTL3 promotes tumor progression via activating Sonic hedgehog signaling. Mechanistically, METTL3 methylates PTCH1 and GLI2 RNAs and further regulates their RNA stability and translation. Importantly, targeting METTL3 by depleting METTL3 expression or treatment with its catalytic inhibitor STM2457 restrains tumor progression. Collectively, this study shows a critical function for METTL3 and m6A methylation in SHH-MB, indicative of a potential role of METTL3 as therapeutic target in SHH-MB.

The flavivirus genome encodes a single polyprotein precursor requiring multiple cleavages by host and viral proteases in order to produce the individual proteins that constitute an infectious virion. Previous studies have revealed that the NS2B cofactor of the viral NS2B-NS3 heterocomplex protease displays a conformational dynamic between active and inactive states. Here, we developed a conformational switch assay based on split luciferase complementation (SLC) to monitor the conformational change of NS2B and to characterize candidate allosteric inhibitors. Binding of an active-site inhibitor to the protease resulted in a conformational change of NS2B and led to significant SLC enhancement. Mutagenesis of key residues at an allosteric site abolished this induced conformational change and SLC enhancement. We also performed a virtual screen of NCI library compounds to identify allosteric inhibitors, followed by in vitro biochemical screening of the resultant candidates. Only three of these compounds, NSC135618, 260594, and 146771, significantly inhibited the protease of Dengue virus 2 (DENV2) in vitro, with IC50 values of 1.8 μM, 11.4 μM, and 4.8 μM, respectively. Among the three compounds, only NSC135618 significantly suppressed the SLC enhancement triggered by binding of active-site inhibitor in a dose-dependent manner, indicating that it inhibits the conformational change of NS2B. Results from virus titer reduction assays revealed that NSC135618 is a broad spectrum flavivirus protease inhibitor, and can significantly reduce titers of DENV2, Zika virus (ZIKV), West Nile virus (WNV), and Yellow fever virus (YFV) on A549 cells in vivo, with EC50 values in low micromolar range. In contrast, the cytotoxicity of NSC135618 is only moderate with CC50 of 48.8 μM on A549 cells. Moreover, NSC135618 inhibited ZIKV in human placental and neural progenitor cells relevant to ZIKV pathogenesis. Results from binding, kinetics, Western blot, mass spectrometry and mutagenesis experiments unambiguously demonstrated an allosteric mechanism for inhibition of the viral protease by NSC135618.

It has been reported that skin/muscle incision and retraction (SMIR) in the thigh, produces mechanical allodynia in the hind paw, far from the site of incision/retraction. The mechanical allodynia lasts about 22 days, indicating chronic post-operative pain develops. The precise mechanisms, however, are largely unclear. In the current study, we further found that SMIR surgery induced LTP of c-fiber evoked field potentials that lasted at least 4 h. The mRNA and protein level of tumor necrosis factor-alpha (TNFα) and acetylated nuclear factor-kappaB p65 (ac-NF-κB p65) in the lumbar spinal dorsal horn was gradually increased during LTP development, while pretreatment with either TNFα neutralization antibody or NF-κB inhibitor PDTC completely prevented the induction of LTP. Moreover, the expression of Silent information regulator 1 (SIRT1) in the lumbar spinal dorsal horn was decreased and activation of SIRT1 by SRT1720 also prevented the induction of LTP. Importantly, the spinal expression of Liver X receptors (LXRs) was increased, both at mRNA and protein level following SMIR. Application of LXRs agonist T0901317 to the spinal dorsal horn prevented LTP induction following SMIR. Mechanistically, T0901317 enhanced the expression of SIRT1 and decreased the expression of ac-NF-κB p65 and TNFα. Spinal application of SIRT1 antagonist EX-527, 30 min before T0901317 administration, completely blocked the inhibiting effect of T0901317 on LTP, and on expression of ac-NF-κB p65 and TNFα. These results indicated that activation of LXRs prevented SMIR-induced LTP by inhibiting NF-κB/TNFα pathway via increasing SIRT1 expression.

AG36 is the biotransformation product of triterpenoid saponin from Ardisia gigantifolia stapf. In this study, the antitumor activity and underlying molecular mechanisms of AG36 against human breast MCF-7, MDA-MB-231, and SK-BR-3 cancer cells were investigated. AG36 inhibited the viability of MCF-7, MDA-MB-231, and SK-BR-3 cells in a dose and time-dependent manner, with an IC50 of approximately 0.73, 18.1, and 23.4 μM at 48 h, respectively. AG36 obviously induced apoptosis and G2/M arrest of all the three breast cancer cells. Moreover, AG36 decreased the protein expression of cycle regulatory proteins cyclin B1 or cyclin D1. In MCF-7 and MDA-MB-231 cells, AG36 strongly increased the cleaved caspase-3 and -8 protein expressions, while in SK-BR-3 cells, AG36 only increased the protein expression of cleaved caspase-3. In all the three breast cancer cells, the ratio of Bax/Bcl-2 and cytosolic cytochrome c content increased significantly compared with control group. The death receptor-related proteins Fas/FasL, TNFR1, and DR5 were detected by Western blot, it showed that different breast cancer cells activated the death receptor-mediated extrinsic caspase-8 pathway through different receptors. In addition, the caspase-8 inhibitor z-IETD-fmk could significantly block AG36-triggered MCF-7 cells apoptosis. The in vivo studies showed that AG36 significantly inhibited the growth of MCF-7 xenograft tumors in BALB/c nude mice comparing with control. In conclusion, AG36 inhibited MCF-7, MDA-MB-231, and SK-BR-3 cells proliferation by the intrinsic mitochondrial and the extrinsic death receptor pathways and AG36 might be a potential breast cancer therapeutic agent.

Background and Objective: Diabetes mellitus (DM) is reportedly a significant risk factor for intervertebral disc degeneration (IDD). Incretin system and particularly glucagon-like peptide 1 (GLP-1) because of its glucose-lowering effects has become an important target in therapeutic strategies of type 2 diabetes (T2D). Liraglutide is a GLP-1 receptor (GLP-1R) agonist with glucoregulatory and insulinotropic functions as well as regulatory functions on cell proliferation, differentiation, and apoptosis. However, little is known on the roles and signaling pathways of apoptosis protecting effects of liraglutide in IDD. This study aimed to investigate the potential protective effects of liraglutide against high glucose-induced apoptosis of nucleus pulposus cells (NPCs) and the possible involved signaling pathways. Methods: The human NPCs were incubated with 100 nM liraglutide alone or in combination with LY294002 (PI3K inhibitor), rapamycin (mTOR inhibitor), and SB216763 (GSK3β inhibitor) in a high glucose culture for 48 h. The four groups were assessed further for apoptosis and genes expressions. The apoptotic effect was evaluated by flow cytometry and further confirmed by cell death detection enzyme-linked immunoassay plus (ELISAPLUS). The gene and protein expression levels were assessed by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting techniques. The results were comparatively assessed between the four groups. Results: The results confirmed the presence of GLP-1R in the NPCs indicating that liraglutide inhibited the high glucose-induced apoptosis, which was blocked by silencing GLP-1R with siRNA. Moreover, liraglutide stimulated the phosphorylation of Akt, mTOR and GSK3β. Treatment with LY294002 significantly increased the apoptosis of NPCs and reduced the levels of their downstream substrates (p-AKT, p-mTOR, and p-GSK3β). Further assessments revealed that activation of mTOR and GSK3β was almost completely inhibited by rapamycin and SB216763, respectively, which significantly increased the caspase-3 levels. Conclusion: Liraglutide could protect NPCs against high glucose-induced apoptosis by activating the PI3K/AKT/mTOR/caspase-3 and PI3K/AKT/GSK3β/caspase-3 signaling pathways.

Platinum‑containing doublet chemotherapy is the cornerstone of lung cancer treatment; however, cisplatin resistance is a major obstacle in the treatment of lung cancer. However, the mechanism underlying this resistance has not been fully elucidated. Previous studies have shown that serum apurinic/apyrimidinic endonuclease 1 (APE1) levels in patients with NSCLC are inversely associated with progression‑free survival after platinum‑containing doublet chemotherapy, and can serve as a biomarker for predicting disease prognosis and treatment efficacy. The present study was designed to investigate the role played by APE1 in the resistance of lung cancer to cisplatin. The levels of mitochondrial apurinic endonuclease 1 (m‑APE1) and total APE1 (t‑APE1) protein in a cisplatin‑resistant A549 cell line (A549/DDP) and cisplatin‑sensitive A549 cells were analyzed by western blotting. Mitochondrial membrane potential was detected by using the JC‑1 staining method. The cisplatin‑resistance of APE1‑overexpressing A549 cells and APE1‑silenced A549/DDP cells was assessed by cell apoptosis and colony formation assays. The results revealed that cisplatin‑resistant A549 cells contained high levels of APE1, and exhibited elevated levels of autophagy. The levels of m‑APE1 and t‑APE1 protein were increased in the A549/DDP cells when compared with these levels in the A549 cells. Overexpression of APE1 and Mia40 enhanced the cisplatin resistance and autophagy of the A549 cells. APE1 knockdown restored the cisplatin sensitivity and reduced the levels of LC3II and Parkin in the A549/DDP cells, but promoted the release of cytochrome c. Furthermore, Parkin silencing or treatment with 3‑methyladenine (3‑MA, an autophagy inhibitor) promoted the apoptosis of APE1‑overexpressing A549 cells, indicating that Parkin‑mediated mitophagy plays an important role in the APE1‑induced cisplatin resistance of A549 cells. In conclusion, APE1 promotes the cisplatin resistance of lung cancer cells by inducing Parkin‑mediated mitophagy.

Increased mechanical stress after spinal cord injury (SCI) expands the scope of nerve tissue damage and exacerbates nerve function defects. Surgical decompression after SCI is a conventional therapeutic strategy and has been proven to have neuroprotective effects. However, the mechanisms of the interaction between mechanical stress and neurons are currently unknown. In this study, we monitored intramedullary pressure (IMP) and investigated the therapeutic benefit of decompression (including durotomy and piotomy) after injury and its underlying mechanisms in SCI. We found that decreased IMP promotes the generation and degradation of LC3 II, promotes the degradation of p62 and enhances autophagic flux to alleviate apoptosis. The lysosomal dysfunction was reduced after decompression. Piotomy was better than durotomy for the histological repair of spinal cord tissue after SCI. However, the autophagy-lysosomal pathway inhibitor chloroquine (CQ) partially reversed the apoptosis inhibition caused by piotomy after SCI, and the structural damage was also aggravated after CQ administration. An antibody microarray analysis showed that decompression may reverse the up-regulated abundance of p-PI3K, p-AKT and p-mTOR caused by SCI. Our findings may contribute to a better understanding of the mechanism of decompression and the effects of mechanical stress on autophagy after SCI.