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Acute kidney injury (AKI) has become a disease of global concern due to its high morbidity and mortality. This has highlighted the need for renoprotective agents. Astragaloside IV (AS-IV) is a saponin isolated from Astragalus membranaceus with good antioxidant, anti-inflammatory and anti-tumor properties. In this study, HK2 cells and rat model were utilized to explore the protective effect of AS-IV against cadmium chloride-induced oxidative stress-induced apoptosis. CdCl 2 -induced apoptosis, ROS production, and mitochondrial membrane potential alterations were significantly inhibited in AS-IV -treated HK2 cells. Expression of the mitochondria-associated apoptotic proteins Cleaved-Caspase3, Cleaved-Caspase9, and Cleaved-PARP was significantly reduced after AS-IV intervention. In addition, AS-IV inhibited Rat weight loss and also alleviated the symptoms of CdCl 2 -induced nephrotoxicity in a rat model of CdCl 2 -induced kidney injury. Further experiments showed that AS-IV suppresses heavy metal Cd-induced mitochondria-mediated apoptosis by regulating the Nrf2/HO-1 pathway. In conclusion, AS-IV could protect the kidney from heavy metal-induced toxicity and could be used as a nephroprotective agent., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Colorectal adenocarcinoma (COAD) is a significant cause of cancer-related mortality worldwide, necessitating the identification of novel therapeutic targets and treatments. This research aimed to investigate the role of ARL3 in COAD progression and to explore the effects of Piperine on ARL3 expression, cell proliferation, epithelial-mesenchymal transition (EMT), and endoplasmic reticulum (ER) stress. Bioinformatics analysis of The Cancer Genome Atlas (TCGA)-COAD, GSE39582, and GSE44861 datasets assessed ARL3 expression levels. Immunohistochemical data from the Human Protein Atlas (HPA) database confirmed ARL3 overexpression in COAD. The association of ARL3 with COAD clinical parameters and prognosis was also examined. COAD cells were treated with Piperine, and in vitro assays evaluated cell proliferation, apoptosis, EMT marker expression, and ER stress (ERS) responses. ARL3 overexpression in COAD correlated with poor prognosis and varied across pathological stages. Piperine treatment inhibited COAD cell proliferation in a concentration- and time-dependent manner, as indicated by reduced Ki-67 levels and decreased colony-forming ability. Piperine induced S-phase cell cycle arrest and facilitated apoptosis in COAD cells, evidenced by changes in Bax, Bcl-2, cleaved caspase-3, and cleaved Poly (ADP-ribose) polymerase (PARP) levels. Moreover, Piperine downregulated ARL3 expression in COAD cells, thereby suppressing transforming growth factor beta (TGF-β)-induced EMT. Additionally, Piperine attenuated the ARL3-mediated ER stress response, significantly reducing binding immunoglobulin protein (BiP), inositol-requiring enzyme 1 alpha (p-IRE1α), activating transcription factor 6 (ATF6), and C/EBP homologous protein (CHOP) levels. Piperine exerts anti-cancer effects in COAD by modulating ARL3 expression, disrupting cell cycle progression, inhibiting the EMT pathway, and regulating ERS. These findings suggest that Piperine holds promise as a therapeutic agent for COAD through its targeting of ARL3.

Background: Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) are severe clinical conditions with limited treatment options. Toosendanin (TSN), a triterpenoid compound with anti-inflammatory effects, has unclear efficacy in ALI., Purpose: This study aimed to evaluate TSN's protective effects on ALI and the related mechanisms., Methods: Lipopolysaccharide (LPS)-induced ALI models were developed in vivo and in vitro. Endothelial permeability was measured using Evans Blue dye; lipid reactive oxygen species (ROS) and apoptosis were assessed using flow cytometry. Malondialdehyde (MDA) and superoxide dismutase (SOD) levels were determined, and cell viability was measured. mRNA and protein expression were quantified using qRT-PCR and Western blotting. Network pharmacology and surface plasmon resonance were used to identify and validate TSN's targets., Results: TSN reduced endothelial permeability and LPS-induced ALI. It lowered ROS levels, lipid peroxidation, endoplasmic reticulum (ER) stress, and apoptosis, both in vitro and in vivo. Network pharmacology identified mTOR as a key target of TSN, and surface plasmon resonance analysis confirmed TSN's direct binding to mTOR, underscoring mTOR's role in TSN's protective effects against ALI. Western blotting showed that TSN inhibits mTOR and its phosphorylation. In vitro, the mTOR activator MHY1485 reversed TSN's protective effects, increasing ER stress, apoptosis, and endothelial permeability. In vivo, TSN and rapamycin synergistically protected against ALI., Conclusion: This study is the first to demonstrate that TSN protects against ALI by targeting the mTOR pathway, regulating ER stress and apoptosis and mitigating endothelial damage. These findings suggest a novel approach for ALI treatment and underscore TSN's potential clinical value., Competing Interests: Declaration of competing interest The authors declare that the research was conducted independently of any commercial or financial relationships that could be interpreted as potential conflicts of interest. Furthermore, the authors confirm that there are no known financial or personal connections that could have appeared to influence the findings presented in this manuscript., (Copyright © 2024 Elsevier GmbH. All rights reserved.)

Spinal cord injury (SCI) can lead to permanent paralysis and various motor, sensory and autonomic nervous system dysfunction. The complex pathophysiological processes limit the effectiveness of many clinical treatments. Mitochondria has been reported to play a key role in the pathogenesis of SCI; while mitophagy is a protective mechanism against mitochondrial dysfunction. However, there is recently little drugs that may targeted activate mitophagy to treat SCI. In this study, we evaluated the role of 20-Deoxyingenol (20-DOI) in SCI and explored its potential mechanisms. We used a SCI rat model and evaluated the functional outcomes after the injury. Western blotting and immunofluorescence techniques were used to analyze the levels of mitophagy, apoptosis, and TFEB-related signaling pathways. Our research results show that 20-DOI significantly improves the apoptosis of neural cells after TBHP stimulation and functional recovery after spinal cord injury. In addition, mitophagy, TFEB levels, and apoptosis are related to the mechanism of 20-DOI treatment for spinal cord injury. Specifically, our research results indicate that 20-DOI restored the autophagic flux after injury, thereby inducing mitophagy, eliminating the accumulation of Cyto C, and inhibiting apoptosis. Further mechanism research suggests that 20-DOI may regulate mitophagy by promoting TFEB nuclear translocation. These results indicate that 20-DOI can significantly promote recovery after spinal cord injury, which may be a promising treatment method for spinal cord injury., Competing Interests: Declarations. Competing Interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

To explore role of NAALADL2-AS2 as ceRNA in DLBCL. Fluorescence in situ hybridization was used to determine location of NAALADL2-AS2 in cells and to verify its expression in DLBCL tissues. The miRNAs interacting with NAALADL2-AS2 and related regulatory genes were identified by small interfering RNA (siRNA) assay, luciferase reporter assay, fluorescent quantitative polymerase chain reaction, western blotting. DLBCL cells transfected with NAALADL2-AS2 siRNA or control siRNA were treated with doxorubicin, rituximab at different concentrations alone or in combination. The growth curves, drug sensitivity changes of cells before and after transfection were detected by MTT assay, ATP-TCA drug sensitivity test. Cell proliferation was detected by BrdU cell proliferation assay, and apoptosis was detected by Annexin V-fluorescein isothiocyanate/propidium iodide staining. The effects and mechanisms of NAALADL2-AS2 on proliferation, apoptosis, drug resistance of DLBCL cells were studied at cellular level. We confirmed expression of NAALADL2-AS2 in both cytoplasm and nuclei of DLBCL cells. Additionally, we observed elevated levels of NAALADL2-AS2 in DLBCL tissues. We discovered that NAALADL2-AS2 functions as ceRNA to inhibit expression of miR-34a, miR-125a, whereas overexpression of NAALADL2-AS2 indirectly upregulates expression of BCL-2. Interfering with NAALADL2-AS2 promoted apoptosis in DLBCL cells, resulting in approximately a 40% increase in sensitivity to doxorubicin and rituximab. In vivo experiments further confirmed that targeting NAALADL2-AS2 effectively suppressed tumor growth, leading to upregulation of miR-34a and miR-125a, downregulation of BCL-2, and enhanced apoptosis in DLBCL cells, which significantly improved their sensitivity to doxorubicin and rituximab by approximately 50%. These results indicate that NAALADL2-AS2/miR-34a, miR-125a/BCL-2 networks hold promise as therapeutic targets for treatment of DLBCL.

Phenylarsine oxide (PAO) is a known environmental pollutant and skin keratinocytes are most seriously affected. Baicalin (BCN) was reported to have antioxidant and anti-inflammatory effects, but its protective effect against PAO toxicity is unknown. This study aimed at exploring whether baicalin can reverse the toxicity of human epidermal keratinocytes that are subjected to PAO exposure and underlying mechanisms. In silico analysis from a publicly accessible HaCaT cell transcriptome dataset exposed to chronic Arsenic showed significant differential expression of several genes, including the genes related to DNA replication. Later, we performed in vitro experiments, in which HaCaT cells were exposed to PAO (500 nM) in the existence of BCN (10-50 µM). Treatment of PAO alone induces the JNK, p38 and caspase-3 activation, which were engaged in the apoptosis induction, while the activity of AKT was significantly inhibited, which was engaged in the suppression of apoptosis. PAO suppressed SIRT3 expression and induced intracellular reactive oxygen species (ROS), causing a marked reduce in cell viability and apoptosis. However, BCN treatment restored the PAO-induced suppression of SIRT3 and AKT expression, reduced intracellular ROS generation, and markedly suppressed both caspase-3 activation and apoptosis induction. However, the protective effect of BCN was significantly attenuated after pretreatment with nicotinamide, an inhibitor of SIRT3. These findings indicate that BCN protects against cell death induced by PAO via inhibiting excessive intracellular ROS generation via restoring SIRT3 activity and reactivating downstream AKT pathway. In this study, we firstly shown that BCN is an efficient drug to prevent PAO-induced skin cytotoxicity, and these findings need to be confirmed by in vivo and clinical investigations., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

Hepatocellular carcinoma (HCC) is a significant global health concern. However, there are limited effective treatments available for it. The use of natural products in the management and treatment of HCC is gaining more attention. Baicalein is a flavonoid compound that has been reported to have antitumor activities in HCC. However, the direct binding targets of baicalein are still unknown. Therefore, we used the DNA-programmed affinity labeling method to identify the target of baicalein and validated its function in HCC cells. We set blank and competitive DNA probes as negative controls. The results showed that baicalein had 136 binding targets, of which 13 targets were differently expressed in HCC tissues. The enriched cellular process of these targets was apoptosis, which involved MAPK9. We tested the binding affinity of baicalein with MAPK9 as 89.7 nM (Kd) by surface plasmon resonance and analyzed the binding sites by virtual docking. Notably, the binding of baicalein with MAPK9 increased the protein levels of MAPK9 itself and the related downstream apoptosis signaling, triggering the apoptosis of HCC cells. However, the inhibitor of MAPK9, SP600125, blocked the baicalein-induced apoptosis, and the amounts of MAPK9 and downstream molecules were also decreased, indicating that baicalein acted through MAPK9 to induce apoptosis of HCC cells. In conclusion, we used the DNA-programmed affinity labeling method to identify the direct-binding target MAPK9 of baicalein and validated its function in baicalein-induced apoptosis of HCC cells, which would be helpful to understand and use baicalein in HCC therapy., (© 2024 John Wiley & Sons Ltd.)

Objectives: To synthesize new bakuchiol aminoguanidine derivatives and test their effect on viability and apoptosis of human triple-negative breast cancer (TNBC) cells., Methods: Two bakuchiol derivatives 1 and 2 were obtained by formylation and Shiff base reaction of bakuchol. The structures of derivatives 1 and 2 were identified by 1 H-NMR, 13 C-NMR, and high-resolution electrospray ionization mass spectrometry (HR-ESI-MS) analysis. Human TNBC MDA-MB-231 cells were treated with bakuchiol and its derivatives and cell viability was measured by MTT assay. Apoptosis was detected by fluorescence microscopy and flow cytometry with Annexin V-FITC/PI staining. The expressions of apoptosis-related proteins were analyzed with Western blotting. The JC-1 and reactive oxygen species (ROS) assay kits were used to determine the effect of new bakuchiol derivatives on mitochondrial function., Results: Based on spectroscopic analysis, a new bakuchiol schiff base derivative was elucidated as 2-{( E )-5-[( S , E )-3, 7-dimethyl-3-vinylocta-1, 6-dien-1-yl]-2-hydroxylbenzylidene} hydrazine-1-carboximidamide (derivative 2). Bakuchiol and its derivatives 1 and 2 all showed cytotoxic activity against the MDA-MB-231 cells. Derivative 2 exhibited the most potent cytotoxic activity to MDA-MB-231 cell with IC 50 of (13.11±1.09), (6.91±1.78), and (2.23±1.32) μmol/L after 24, 48, and 72 h. It had low toxicity to normal mouse liver (AML-12) cells with IC 50 of (31.23±1.58) μmol/L at 72 h. Fluorescence microscopy and flow cytometry demonstrated apoptosis in breast cancer cells after treating with derivative 2 in a concentration dependent manner. Western blotting showed that after derivative 2 treatment, the expression of apoptosis-related proteins cytochrome C, cleaving caspase-3 and Bax/Bcl-2 radio in MDA-MB-231 cells increased; in addition, apoptosis was associated with the decreased mitochondrial membrane potential and increased reactive oxygen species accumulation., Conclusions: The novel bakuchiol aminoguanidine derivative (derivative 2) is capable of inducing apoptosis in MDA-MB-231 cells, but has low toxicity to normal liver cells, suggesting that it may be used as a lead compound for an anti-TNBC agent.

Objective: Multiple myeloma (MM) is caused by abnormal cloning of plasma cells. miR-184 is abnormally expressed in several types of tumors, but its expression and role in MM have not been reported., Methods: The bone marrow samples of healthy controls and MM patients were collected, and plasma cells were sorted. The multiple myeloma cell line OPM-2 was cultured and assigned into miR-NC+siRNA-NC group, miR-184 inhibitor+siRNA-NC group, and miR-184 inhibitor+siRNA-Notch1 group. Cell proliferation was assessed by MTT assay. Clone formation was evaluated by colony formation assay. Cell apoptosis activity was tested with flow cytometry. Notch1 and cleaved caspase3 protein expressions were detected., Results: MiR-184 expression was increased in myeloma plasma cells ( P <0.05). Transfection of miR-184 inhibitor can downregulate miR-184 expression, increase the levels of Notch1 and cleaved caspase3, inhibit OPM-2 cell proliferation, restrain colony formation, enhance caspase3 activity, and suppress tumor cell invasion ( P <0.05). However, administration of siRNA-Notch1 retarded the effect of miR-184 inhibitor by decreasing the expressions of Notch1 and cleaved caspase3, enhancing colony formation and tumor cell invasion, as well as inhibiting caspase3 activity and cell proliferation., Conclusion: Our data indicated that miR-184 expression is increased in myeloma plasma cells. Down-regulation of miR-184 promotes MM cell apoptosis and inhibits proliferation and colony formation by regulating Notch1 expression., (© 2024 by the Association of Clinical Scientists, Inc.)

In this study, we prepared a bionic nanosystem of trastuzumab-functionalized SK-BR-3 cell membrane hybrid liposome-coated pyrotinib (Ptb-M-Lip-Her) for the treatment of HER2-positive breast cancer. Transmission electron microscopy, dynamic light scattering, polyacrylamide gel electrophoresis (SDS-PAGE) and western blotting were used to verify the successful preparation of Ptb-M-Lip-Her. In vitro drug release experiments proved that Ptb-M-Lip-Her had a sustained release effect. Cell uptake experiments and in vivo imaging experiments proved that Ptb-M-Lip-Her had good targeting ability to homologous tumor cells (SK-BR-3). The results of cell experiments such as MTT, flow cytometry, immunofluorescence staining and in vivo antitumor experiments showed that Ptb-M-Lip-Her could significantly promote apoptosis and inhibit the proliferation of SK-BR-3 cells. These results clearly indicated that Ptb-M-Lip-Her may be a promising biomimetic nanosystem for targeted therapy of HER2-positive breast cancer., (© 2024. The Author(s).)

Vitamin D 3 is an important element in improving erectile function. However, the mechanisms of vitamin D 3 remain unknown. Thus, we explored the effect of vitamin D 3 on erectile function recovery after nerve injury in a rat model and investigated its possible molecular mechanisms. Eighteen male Sprague-Dawley rats were used in this study. The rats were randomly divided into three groups: the control, bilateral cavernous nerve crush (BCNC), and BCNC + vitamin D 3 groups. BCNC model was established in rats by surgery. The intracavernosal pressure and the ratio of intracavernosal pressure to mean arterial pressure were utilized to evaluate erectile function. Masson trichrome staining, immunohistochemistry, terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling and western blot analysis were performed on penile tissues to elucidate the molecular mechanism. The results indicated that vitamin D 3 alleviated hypoxia and suppressed the fibrosis signalling pathway by upregulating the expression of eNOS (p = 0.001), nNOS (p = 0.018) and α-SMA (p = 0.025) and downregulating the expression of HIF-1α (p = 0.048) and TGF-β1 (p = 0.034) in BCNC rats. Vitamin D 3 promoted erectile function restoration by enhancing the autophagy process through decreases in the p-mTOR/mTOR ratio (p = 0.02) and p62 (p = 0.001) expression and increases in Beclin1 expression (p = 0.001) and the LC3B/LC3A ratio (p = 0.041). Vitamin D 3 application improved erectile function rehabilitation by suppressing the apoptotic process through decreases in the expression of Bax (p = 0.002) and caspase-3 (p = 0.046) and an increase in the expression of Bcl2 (p = 0.004). Therefore, We concluded that vitamin D 3 improved the erectile function recovery in BCNC rats by alleviating hypoxia and fibrosis, enhancing autophagy and inhibiting apoptosis in the corpus cavernosum., (© 2023. The Author(s), under exclusive licence to Springer Nature Limited.)

ARID1A, a component of the SWI/SNF chromatin-remodeling complex, is frequently mutated in various cancer types and has emerged as a potential therapeutic target. In this study, we observed that ARID1A-deficient colorectal cancer (CRC) cells showed synthetic lethal effects with a p53 activator, RITA (reactivating p53 and inducing tumor apoptosis). RITA, an inhibitor of the p53-MDM2 interaction, exhibits increased sensitivity in ARID1A-deficient cells compared to ARID1A wild-type cells. Mechanistically, the observed synthetic lethality is dependent on both p53 activation and DNA damage accumulation, which are regulated by the interplay between ARID1A and RITA. ARID1A loss exhibits an opposing effect on p53 targets, leading to decreased p21 expression and increased levels of proapoptotic genes, PUMA and NOXA, which is further potentiated by RITA treatment, ultimately inducing cell apoptosis. Meanwhile, ARID1A loss aggravates RITA-induced DNA damage accumulation by downregulating Chk2 phosphorylation. Taken together, ARID1A loss significantly heightens sensitivity to RITA in CRC, revealing a novel synthetic lethal interaction between ARID1A and RITA. These findings present a promising therapeutic approach for colorectal cancer characterized by ARID1A loss-of-function mutations., (© 2024. The Author(s).)

Programed cell death (PCD) plays fundamental roles in plant development and responses to environmental stresses. Here, we report a protein, SICKLE (SIC), which represses PCD. In Arabidopsis (Arabidopsis thaliana), the loss-of-function mutant of SIC, sic-4, hyperaccumulated lariat intronic RNAs (lariRNAs) and exhibited PCD. The gene encoding an RNA debranching enzyme 1 (DBR1), a rate-limiting enzyme for lariRNAs decay, was overexpressed to reduce the level of lariRNAs in the sic-4 mutant, which led to suppression of PCD. Meanwhile, another lariRNAs hyper-accumulating mutant, dbr1-2, also exhibited PCD, further indicating that sic-4 PCD is caused by hyper-accumulation of lariRNAs. Transcriptional profiling analyses revealed that the sic-4 mutation disturbed alternative splicing and decay of mRNAs associated with salicylic acid (SA) homeostasis, a well-known molecule functioning in PCD regulation. Moreover, SA is dramatically increased in sic-4 and the disruption of SA biosynthesis and signaling suppressed PCD in the mutant, demonstrating that SA functions downstream of sic-4. Taken together, our results demonstrate that SIC is involved in regulating SA-triggered PCD., Competing Interests: Conflict of interest statement. The authors declare that the research was conducted without any known commercial or financial interests that would serve as a potential conflict of interest., (© American Society of Plant Biologists 2023. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Nonalcoholic fatty liver disease (NAFLD) is the most prevalent hepatic pathology worldwide. However, the precise molecular mechanisms for NAFLD are still not sufficiently explained. Recently, a new mode of cell death (cuproptosis) is found. However, the relationship between NAFLD and cuproptosis remains unclear. We analyzed three public datasets (GSE89632, GSE130970, and GSE135251) to identify cuproptosis-related genes stably expressed in NAFLD. Then, we performed a series of bioinformatics analyses to explore the relationship between NAFLD and cuproptosis-related genes. Finally, 6 high-fat diet- (HFD-) induced NAFLD C57BL/6J mouse models were established to carry out transcriptome analysis. The results of gene set variation analysis (GSVA) revealed that the cuproptosis pathway was abnormally activated to a certain degree ( p = 0.035 in GSE89632, p = 0.016 in GSE130970, p = 0.22 in GSE135251), and the principal component analysis (PCA) of the cuproptosis-related genes showed that the NAFLD group separated from the control group, with the first two principal components accounting for 58.63%-74.88% of the variation. Among three datasets, two cuproptosis-related genes ( DLD and PDHB , p < 0.01 or 0.001) were stably upregulated in NAFLD. Additionally, both DLD (AUC = 0.786-0.856) and PDHB (AUC = 0.771-0.836) had favorable diagnostic properties, and the multivariate logistics regression model further improved the diagnostic properties (AUC = 0.839-0.889). NADH, flavin adenine dinucleotide, and glycine targeted DLD , and pyruvic acid and NADH targeted PDHB in the DrugBank database. The DLD and PDHB were also associated with clinical pathology, especially with steatosis ( DLD , p = 0.0013-0.025; PDHB , p = 0.002-0.0026) and NAFLD activity score ( DLD , p = 0.004-0.02; PDHB , p = 0.003-0.031). What is more, DLD and PDHB were correlated with stromal score ( DLD , R = 0.38, p < 0.001; PDHB , R = 0.31, p < 0.001) and immune score ( DLD , R = 0.26, p < 0.001; PDHB , R = 0.27, p < 0.001) in NAFLD. Furthermore, Dld and Pdhb were also significantly upregulated in the NAFLD mouse model. In conclusion, cuproptosis pathways, especially DLD and PDHB , could be potential candidate genes for NAFLD diagnostic and therapeutic options., Competing Interests: The authors declare no conflict of interest., (Copyright © 2023 Chutian Wu et al.)

Context: Acacetin is a natural source of flavonoids with anti-inflammatory and antioxidant effects., Objective: This study determines acacetin's protective effect and mechanism on myocardial ischaemia/reperfusion (I/R) injury., Materials and Methods: Sprague-Dawley rats were divided into sham and I/R injury and treatment with acacetin. Acacetin (10 mg/kg) was subcutaneously injected for 7 days. ECG and echocardiography were conducted to determine arrhythmia and heart function. The pathological characters of the heart were determined with triphenyl tetrazolium chloride staining, Haematoxylin & Eosin staining, and Masson staining. Expression of proteins in infarct tissues was examined with western blots., Results: Administrated with acacetin in I/R rats significantly reduced the arrhythmia score from 4.90 to 2.50 and the reperfusion arrhythmia score from 3.79 to 1.82 in the vehicle or the acacetin group, respectively. LVEF was improved from 33.5% in the I/R group to 43.7% in the acacetin group, LVFS was increased from 16.4% to 24.5%, LVIDs was decreased from 6.5 to 5.3 mm. The inflammatory cell infiltration, myocardial fibrosis, and collagen 1 and 3 were reduced by acacetin. Acacetin promoted SOD and decreased MDA. In myocardial tissues, the expression level of TLR4 and IL-6 were restrained, and IL-10 was promoted. Apoptotic protein Bax was suppressed, and anti-apoptotic protein Bcl-2 was promoted in the acacetin group. Interestingly, the transcription factor Nrf-2/HO-1 pathway was also reversed by acacetin., Discussion and Conclusion: Our findings indicated that acacetin has a potential therapeutic effect in clinical application on treating I/R-induced heart injury.

Purpose: This study aimed to investigate the effect of Raf-1 kinase inhibitory protein (RKIP) on diabetic retinal neurodegeneration in streptozotocin-treated rat model and high glucose-treated rat Müller cells., Methods: Control and streptozotocin-treated rats were intravitreally injected with saline, RKIP gene overexpression lentivirus (oeRKIP) or negative control lentivirus (RKIP-vector). Normal or high glucose-treated Müller cells were transfected with saline, RKIP gene overexpression lentivirus or negative control lentivirus. Western blotting and immunofluorescence assay were utilized to evaluate the function of RKIP on the expression of RKIP, p38 mitogen-activated protein kinase (p38-MAPK), glutamate/aspartate transporter (GLAST), glutamine synthetase (GS), glial fibrillar acidic protein (GFAP) and cysteine-aspartic acid protease-3 (caspase-3). A glutamate assay kit was adopted to detect glutamate level in retina samples. Apoptosis of Müller cells was determined by Annexin-V/PI staining and flow cytometry., Results: High glucose-treated Müller cells exhibited promoted apoptosis, while RKIP overexpression in high glucose-treated Müller cells down-regulated the enhanced apoptosis. Compared with rats injected with saline, streptozotocin-treated hyperglycemic rats displayed enhancement in the immunoreactivities of p38-MAPK and GFAP as well as in the protein expression of p38-MAPK and caspase-3. Strikingly, intravitreal injection of RKIP gene overexpression lentivirus in the hyperglycemic rats reversed the augmented immunoreactivities and protein expression mentioned above. Meanwhile, RKIP overexpression in the hyperglycemic rats improved the immunoreactivities and protein expression of RKIP, GS and GLAST. Besides, RKIP down-regulated the increased level of retinal glutamate in the hyperglycemic rats., Conclusions: Intravitreal injection of RKIP gene overexpression lentivirus functioned in preventing diabetic retinal neurodegeneration in a rat model of diabetes presumably by inhibiting p38-MAPK pathway.

Long non-coding RNA (lncRNA) LINC00467 plays a proto-oncogenic role in non-small cell lung cancer. However, its effect and modulatory mechanism in gastric cancer (GC) are unknown. Thereby, we elucidated the mechanism of LINC00467 in GC. LINC00467 level in GC tissues was assessed by bioinformatic analysis, and clinicopathological parameters from GC patients were collected. The levels of LINC00467, integrin subunit beta 3 (ITGB3), proliferating cell nuclear antigen (PCNA), cleaved caspase-3 and cleaved poly (ADP-ribose) polymerase 1 (PARP1) in tissue samples or treated GC cells were assessed by quantitative real-time polymerase chain reaction (qRT-PCR), fluorescence in situ hybridization (FISH), or Western blot. The viability, proliferation and apoptosis of GC cells were detected by methyl thiazolyl tetrazolium assay, colony formation assay, and flow cytometry. Levels of LINC00467 and ITGB3 were up-regulated in GC, and highly expressed LINC00467 was positively associated with tumor size, differentiation, N stage, and T stage in GC patients. LINC00467 was enriched in cytoplasm of GC cells, and overexpressed LINC00467 promoted the viability and proliferation as well as levels of ITGB3 and PCNA, while suppressing the apoptosis and levels of cleaved caspase-3 and cleaved PARP1 in GC cells. Besides, the effects of shLINC00467 on inhibiting cell viability, proliferation of GC cells and PCNA level and promoting apoptosis as well as levels of cleaved caspase-3 and cleaved PARP1 were all partially reversed by overexpressed ITGB3. Overexpressed LINC00467 enhanced the viability and proliferation but inhibited apoptosis of GC cells via increasing ITGB3 level., (Copyright © 2021. Published by Elsevier Ltd.)

Cell apoptosis detection is vital for biological analysis and clinical application; some detection assays are already commercially available. However, it is still far from perfect and needs further improvement for less cost, time-consuming and operation demanding. TUNEL, a high market share cell apoptosis assay, depends on adulteration fluorescent labelling dUTP by terminal deoxynucleotidyl transferase(TdT) which randomly adds deoxyribonucleoside triphosphates (dNTPs) at the 3'-OH terminal of ssDNA with a template-free manner. Based on our previous work, we adopted a label-free strategy to reduce the cost and operation maintenance of TUNEL and developed a facile, rapid, convenient and in-situ assay for cell apoptosis., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Bacterial LPS is a heat-stable endotoxin and wall components of gram negative bacteria, which can exhibit a toxicological effect on physiology and biochemical activities of fish. In this study, we investigated the effect of LPS exposure on cell viability, oxidative stress, caspase activity and immune-related gene expressions in cultured fin cell lines of red crucian carp, white crucian carp and their hybrid offspring. LPS stimulation could reduce fish cell viability, whereas gene expression levels and promoter activities in inflammatory signals increased dramatically. Moreover, enhanced levels of intracellular oxidative stress and decreased levels of mitochondrial membrane potential (MMP) were observed in LPS-induced fish cells. N-Acetyl-L-cysteine (NAC) could alleviate LPS-stimulated reactive oxygen species (ROS) generation and caspase-3 activity in fish cells. These results suggested that ROS-mediated cytotoxic stress was involved in LPS-induced inflammation and mitochondrial damage in cultured fish cells., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Endoplasmic reticulum stress (ER stress) plays a critical role in neuronal apoptosis along with the aggravation of Alzheimer's disease (AD). Nuclear receptor peroxisome proliferator-activated receptor gamma (PPARγ) is a ligand-activated transcription factor that is involved in regulating ER stress in Alzheimer's disease (AD), therefore, this protein could be a promising therapeutic target for AD. Vanadium compounds, such as vanadyl acetylacetonate, sodium metavanadate and bis(maltolato)oxovanadium, are well-known as puissant PPARγ modulators. Thus, we are curious whether bis(ethylmaltolato)oxidovanadium (IV) (BEOV) can ameliorate ER stress and subsequent neuronal apoptosis by regulating PPARγ in AD models. To this end, we determined the effect of BEOV on behavioral performance, ER stress and neuronal apoptosis in the triple transgenic mouse AD model (3×Tg-AD). Our results showed that BEOV improved cognitive abilities and reduced the ER stress- and apoptosis-associated proteins in the brains of 3×Tg-AD mice. In vitro administration of BEOV in primary hippocampal neurons and N2asw cells achieved similar results in repressing ER stress. In addition, cotreatment with GW9662 (an antagonist of PPARγ) effectively blocked these neuroprotective effects of BEOV, which provided strong evidence that PPARγ-dependent signaling plays a key role in protecting against ER stress and neuronal apoptosis in AD. In conclusion, our data demonstrated that BEOV alleviated neuronal apoptosis triggered by ER stress by regulating PPARγ in a 3×Tg-AD model., (© 2021. Society for Biological Inorganic Chemistry (SBIC).)

Spinal cord injury (SCI) is an extremely destructive central nervous system lesion. Studies have shown that NGF can promote nerve regeneration after SCI. However, it cannot produce the desired effect due to its stability in the body and is difficulty in passing through the blood-brain barrier. In this study, we prepared nanovesicles derived from macrophage membrane encapsulating NGF (NGF-NVs) as a drug carrier for the treatment of SCI. Cell experiments showed that NGF-NVs were effectively taken up by PC12 cells and inhibited neuronal apoptosis. In vivo imaging experiments, a large quantity of NGF was delivered to the injured site with the aid of the good targeting of NVs. In animal experiments, NGF-NVs improved the survival of neurons by significantly activating the PI3K/AKT signaling pathway and had good behavioral and histological recovery effects after SCI. Therefore, NVs are a potential drug delivery vector for SCI therapy.

Glioblastoma multiforme (GBM) is an extremely aggressive brain tumor for which new therapeutic approaches are urgently required. Unfolded protein response (UPR) plays an important role in the progression of GBM and is a promising target for developing novel therapeutic interventions. We identified ubiquitin-activating enzyme 1 (UBA1) inhibitor TAK-243 that can strongly induce UPR in GBM cells. In this study, we evaluated the functional activity and mechanism of TAK-243 in preclinical models of GBM. TAK-243 significantly inhibited the survival, proliferation, and colony formation of GBM cell lines and primary GBM cells. It also revealed a significant anti-tumor effect on a GBM PDX animal model and prolonged the survival time of tumor-bearing mice. Notably, TAK-243 more effectively inhibited the survival and self-renewal ability of glioblastoma stem cells (GSCs) than GBM cells. Importantly, we found that the expression level of GRP78 is a key factor in determining the sensitivity of differentiated GBM cells or GSCs to TAK-243. Mechanistically, UBA1 inhibition disrupts global protein ubiquitination in GBM cells, thereby inducing ER stress and UPR. UPR activates the PERK/ATF4 and IRE1α/XBP signaling axes. These findings indicate that UBA1 inhibition could be an attractive strategy that may be potentially used in the treatment of patients with GBM, and GRP78 can be used as a molecular marker for personalized treatment by targeting UBA1., (© 2021. The Author(s).)

Atherosclerosis, a chronic multifactorial disease, is closely related to the development of cardiovascular diseases and is one of the predominant causes of death worldwide. Normal vascular endothelial cells play an important role in maintaining vascular homeostasis and inhibiting atherosclerosis by regulating vascular tension, preventing thrombosis and regulating inflammation. Currently, accumulating evidence has revealed that endothelial cell apoptosis is the first step of atherosclerosis. Excess apoptosis of endothelial cells induced by risk factors for atherosclerosis is a preliminary event in atherosclerosis development and might be a target for preventing and treating atherosclerosis. Interestingly, accumulating evidence shows that natural medicines have great potential to treat atherosclerosis by inhibiting endothelial cell apoptosis. Therefore, this paper reviewed current studies on the inhibitory effect of natural medicines on endothelial cell apoptosis and summarized the risk factors that may induce endothelial cell apoptosis, including oxidized low-density lipoprotein (ox-LDL), reactive oxygen species (ROS), angiotensin II (Ang II), tumor necrosis factor-α (TNF-α), homocysteine (Hcy) and lipopolysaccharide (LPS). We expect this review to highlight the importance of natural medicines, including extracts and monomers, in the treatment of atherosclerosis by inhibiting endothelial cell apoptosis and provide a foundation for the development of potential antiatherosclerotic drugs from natural medicines., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Ample empirical evidence suggests that mitochondrial dysfunction and endoplasmic reticulum (ER) stress play a crucial role in the pathogenesis of Parkinson's disease (PD). Prohibitin (PHB), a mitochondrial inner-membrane protein involved in mitochondrial homeostasis and function, may be involved in the pathogenesis of PD. We investigated the functional role of PHB in mitochondrial biogenesis and ER stress in methyl-4-phenylpyridinium (MPP +)-induced in vivo and in vitro models of PD. The overexpression of PHB in SH-SY5Y cells block ed cell death and the apoptosis induced by MPP + incubation. PHB also block ed the activation of ER stress markers, including glucose-regulated protein 78, while increasing the expression of Xbox- binding protein 1 and caspase-12. Moreover, the intracerebroventricular administration of the PHB overexpression vector greatly block ed motor dysfunction and the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-mediated neurodegeneration in the mouse model of PD. The production of reactive oxygen species, ER stress, and autophagic stress induced by MPTP were also significantly block ed in PD mice overexpressing PHB. Our results suggest that PHB blocks the dopaminergic-neuron depletion by preserving mitochondrial function and inhibiting ER stress. The genetic manipulation of PHB may feature potential as a treatment for PD., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Tumor necrosis factor receptor-associated factor 6 (TRAF6), a regulator of NF-κB signaling, has been discovered recently to be probably related to osteoarthritis, while the function of TRAF6 in lumbar facet joint osteoarthritis(FJOA)still remains unknown. The aim of this study was to probe the specific function of TRAF6 in chondrocytes and its connection with the pathophysiology of FJOA. We found upregulation of TRAF6 in FJOA cartilage by western blot analysis. In vitro, we stimulated immortalized human chondrocytes by LPS to establish the cells apoptosis model. Western blot analysis demonstrated that levels of TRAF6 and cleaved caspase-3/8 in the chondrocyte injury model increased significantly. Knockdown of TRAF6 suppressed the expression of matrix metallopeptidase-13 (MMP-13) and interleukin-6 (IL-6) induced by LPS, and alleviated cell apoptosis. Meanwhile, western blot and immunofluorescent staining demonstrated that IκBα degradation and p65 nuclear transportation were also inhibited, revealing that knockdown of TRAF6 suppressed activation of the NF-κB pathway in LPS-induced chondrocytes apoptosis model. Collectively, our findings suggest that TRAF6 plays a crucial role in FJOA development by regulating NF-κB signaling pathway. Knockdown of TRAF6 may supply a potential therapeutic strategy for FJOA.

Arsenic exposure could induce apoptosis and cause related cancer. It was reported that p38 signaling pathway played a key transcriptional regulatory factor in arsenic-induced apoptosis. However, there were certain disputable questions about this point of opinion. Therefore, the relationship between p38 signaling pathway and arsenic-induced apoptosis was systematically reviewed and analyzed by meta-analysis. Twelve essays were analyzed with StataSE15.0 and Review Manager 5.3. The regulatory variables, such as normal cells and cancer cells, arsenic exposure time and exposure dose were analyzed by the subgroup analysis. The comprehensive effects were compared and analyzed by SMD method. Publication bias, the monolithic impact and heterogeneity were inspected. Subgroup analysis showed, when arsenic exposure was ≥ 5 μmol/l, the expression of Bcl-2 and Bax was down-regulated and the expression of p38 and Caspase-3 was up-regulated. When arsenic exposure was < 5 μmol/l, the expression of Bcl-2, Bax, p38 and Caspase-3 was up-regulated. Arsenic exposure time (≥ 48 h) or arsenic exposure dose (≥ 5 μmol/l or < 5 μmol/l) can promote the expression of p38. Arsenic exposure time was ≥ 48 h or exposure dose was < 5 μmol/l in cancer cells, arsenic exposure dose was ≥ 5 μmol/l or exposure time was < 48 h in normal cells, and they are statistically significant in the expression of p38. This study evaluates the role of p38 signaling pathway in arsenic-induced apoptosis, which is helpful to provide theoretical basis for the differentiation of arsenic-induced injury and the therapeutic mechanism of arsenic-induced apoptosis.

In this study, we identified the multifaceted effects of atezolizumab, a specific monoclonal antibody against PD-L1, in tumor suppression except for restoring antitumor immunity, and investigated the promising ways to improve its efficacy. Atezolizumab could inhibit the proliferation and induce immune-independent apoptosis of osteosarcoma cells. With further exploration, we found that atezolizumab could impair mitochondria of osteosarcoma cells, resulting in increased release of reactive oxygen species and cytochrome-c, eventually leading to mitochondrial-related apoptosis via activating JNK pathway. Nevertheless, the excessive release of reactive oxygen species also activated the protective autophagy of osteosarcoma cells. Therefore, when we combined atezolizumab with autophagy inhibitors, the cytotoxic effect of atezolizumab on osteosarcoma cells was significantly enhanced in vitro. Further in vivo experiments also confirmed that atezolizumab combined with chloroquine achieved the most significant antitumor effect. Taken together, our study indicates that atezolizumab can induce mitochondrial-related apoptosis and protective autophagy independently of the immune system, and targeting autophagy is a promising combinatorial approach to amplify its cytotoxicity.

Epithelial ovarian cancer (EOC) is one of the leading causes of death from gynecologic cancers and peritoneal dissemination is the major cause of death in patients with EOC. Although the loss of 4.1N is associated with increased risk of malignancy, its association with EOC remains unclear. To explore the underlying mechanism of the loss of 4.1N in constitutive activation of epithelial-mesenchymal transition (EMT) and matrix-detached cell death resistance, we investigated samples from 268 formalin-fixed EOC tissues and performed various in vitro and in vivo assays. We report that the loss of 4.1N correlated with progress in clinical stage, as well as poor survival in EOC patients. The loss of 4.1N induces EMT in adherent EOC cells and its expression inhibits anoikis resistance and EMT by directly binding and accelerating the degradation of 14-3-3 in suspension EOC cells. Furthermore, the loss of 4.1N could increase the rate of entosis, which aggravates cell death resistance in suspension EOC cells. Moreover, xenograft tumors in nude mice also show that the loss of 4.1N can aggravate peritoneal dissemination of EOC cells. Single-agent and combination therapy with a ROCK inhibitor and a 14-3-3 antagonist can reduce tumor spread to varying degrees. Our results not only define the vital role of 4.1N loss in inducing EMT, anoikis resistance, and entosis-induced cell death resistance in EOC, but also suggest that individual or combined application of 4.1N, 14-3-3 antagonists, and entosis inhibitors may be a promising therapeutic approach for the treatment of EOC.

Photodynamic therapy (PDT) holds a number of advantages for tumor therapy. However, its therapeutic efficiency is limited by non-sustainable reactive oxygen species (ROS) generation and heterogeneous distribution of photosensitizer (PS) in tumor. Herein, a " S ustainable R OS G enerator" (SRG) is developed for efficient antitumor therapy. Methods: SRG was prepared by encapsulating small-sized Mn 3 O 4 -Ce6 nanoparticles (MC) into dendritic mesoporous silica nanoparticles (DMSNs) and then enveloped with hyaluronic acid (HA). Due to the high concentration of HAase in tumor tissue, the small-sized MC could be released from DMSNs and homogeneously distributed in whole tumor. Then, the released MC would be uptaken by tumor cells and degraded by high levels of intracellular glutathione (GSH), disrupting intracellular redox homeostasis. More importantly, the released Ce6 could efficiently generate singlet oxygen ( 1 O 2 ) under laser irradiation until the tissue oxygen was exhausted, and the manganese ion (Mn 2+ ) generated by degraded MC would then convert the low toxic by-product (H 2 O 2 ) of PDT to the most harmful ROS (·OH) for sustainable and recyclable ROS generation. Results: MC could be homogeneously distributed in whole tumor and significantly reduced the level of intracellular GSH. At 2 h after PDT, obvious intracellular ROS production was still observed. Moreover, during oxygen recovery in tumor tissue, ·OH could be continuously produced, and the nanosystem could induce 82% of cell death comparing with 30% of cell death induced by free Ce6. For in vivo PDT, SRG achieved a complete inhibition on tumor growth. Conclusion: Based on these findings, we conclude that the designed SRG could induce sustainable ROS generation, homogeneous intratumoral distribution and intracellular redox homeostasis disruption, presenting an efficient strategy for enhanced ROS-mediated anti-tumor therapy., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Muscle atrophy is a major character of cancer cachexia, whose mechanism remains enigmatic. During cancer cachexia, the function of endoplasmic reticulum stress (ERS), which ubiquitously exists in invasive cancer, remains unclear in muscle remodeling. In addition, ERS can be transmitted to surrounding and distant cells, terming transmissible ERS (TERS), by certain soluble factors, which have not been completely identified. In this study, tunicamycin-induced conditioned media from oral squamous cell carcinoma (OSCC) cell lines were proved to transmit ERS to muscle cells both in vivo and in vitro. We found for the first time that exosomes from the conditioned media were the key factors to mediate TERS signaling and induce muscle cell atrophy and apoptosis consequently. Next-generation RNA sequencing was applied to pinpoint exosome miR-181a-3p, which was then identified to play a critical role in regulating ERS, muscle atrophy and apoptosis pathways., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Zinc pyrithione (ZPT) is widely used as an antimicrobial. Zinc is a necessary trace element of the human whose homeostasis associated with several cancers. However, the anticancer effect of increased Zinc in ovarian cancer is still unclear. This study focussed on the anti-tumour effects of ZPT combined with Zinc in SKOV3 and SKOV3/DDP cells. The cell viability, apoptosis, migration, and invasion assays were detected by CCK-8, flow cytometry, wound healing and transwell assay, respectively. The distribution of Zinc in cells was monitored by staining of Zinc fluorescent dye and lysosome tracker. The changes in lysosomal membrane stability were reflected by acridine orange fluorescence and cathepsin D reposition. Expression of the proteins about invasion and apoptosis was evaluated by western blot. The results indicated that ZPT combined with Zinc could notably reduce cell viability, inhibit migration and invasion in SKOV3 and SKOV3/DDP cells. Besides, ZPT performed as a Zinc carrier targeted lysosomes, caused the increase of its membrane permeability and the release of cathepsin D accompanied by mitochondrial apoptosis in SKOV3/DDP cells. In conclusion, our work suggests that ZPT combined with Zinc could inhibit proliferation, migration, invasion, and promote apoptosis by trigger the lysosome-mitochondrial apoptosis pathway in ovarian carcinoma.

Selaginella moellendorffii Hieron. (SM), a perennial evergreen plant, has been used in the treatment of acute infectious hepatitis, thoracic and hypochondriac lumbar contusions, systemic oedema and thrombocytopaenia. However, the role of a biflavonoid-rich extract from SM (SM-BFRE) in anti-larynx cancer has rarely been reported. In this study, the in vitro and in vivo anti-laryngeal cancer activity and potential mechanisms of SM-BFRE were investigated. An off-line semipreparative liquid chromatography-nuclear magnetic resonance protocol was carried out to determine six biflavonoids from SM-BFRE. In vitro, MTT assay revealed that SM-BFRE inhibited the proliferation of laryngeal carcinoma cells. A wound healing assay indicated that SM-BFRE suppressed the migration of laryngeal cancer cells. Hoechst 33 258 and Annexin V-FITC/PI double staining assays were performed and verified that SM-BFRE induced apoptosis in laryngeal carcinoma cells. The Hep-2 bearing nude mouse model confirmed that SM-BFRE also exhibited anticancer effect in vivo. In addition, Western blot analysis demonstrated that SM-BFRE exerted its anti-laryngeal cancer effect by activating the mitochondrial apoptotic pathway and inhibiting STAT3 and Akt/NF-κB signalling pathways. All results suggested that SM-BFRE could be considered as a potential chemotherapeutic drug for laryngeal cancer., (© 2020 The Authors. Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.)

As a basic biological phenomenon of cells, regulated cell death (RCD) has irreplaceable influence on the occurrence and development of many processes of life and diseases. RCD plays an important role in the stability of the homeostasis, the development of multiple systems and the evolution of organisms. Thus comprehensively understanding of RCD is undoubtedly helpful in the innovation of disease treatment. Recently, research on the underlying mechanisms of the major forms of RCD, such as apoptosis, autophagy, necroptosis, pyroptosis, paraptosis and neutrophils NETosis has made significant breakthroughs. In addition, the interconnections among them have attracted increasing attention from global scholars in the field of life sciences. Here, recent advances in RCD research field are discussed.

The pathophysiological changes in cartilage are a crucial feature of lumbar facet joint (LFJ) degeneration and arthritis. However, the molecular mechanism of human LFJ degeneration remains largely defined. This study aimed to examine the changes in chondrocytes at different stages of degenerative LFJ using hematoxylin and eosin and Safranin O staining. The significant loss of chondrocytes in grades 2 and 3 of LFJs was observed. The expression levels of CCAAT enhancer binding protein β (C/EBPβ), Runt-related transcription factor 2 (RUNX2), and matrix metalloproteinase 13 (MMP13) also increased with the aggravation of degeneration (4.89, 5.77, and 6.3 times by Western blot). In vitro, chondrocytes scraped from the LFJs during surgery were stimulated by interleukin (IL)-1β to establish the injury model. The association of C/EBPβ and RUNX2 with active caspase-3 on chondrocytes was analyzed. The high expression level of C/EBPβ, RUNX2, and MMP13 was consistent with that of caspase-3, which reached a peak after 36 h of stimulation. Immunofluorescence suggested that C/EBPβ, RUNX2, and MMP13 co-labeled with active caspase-3. Moreover, immunoprecipitation data prompted that C/EBPβ was able to interact with RUNX2. The knockdown of C/EBPβ significantly decreased the expression levels of MMP13 and active caspase-3 (2.48 and 2.89 times as detected by Western blot analysis) and inhibited chondrocyte apoptosis, which was further demonstrated using flow cytometry. Taken together, the findings of this study uncovered that C/EBPβ could interact with RUNX2 to induce chondrocyte apoptosis in human LFJ degeneration by regulating the expression of MMP13.

Neuronal apoptosis caused by amyloid-beta (Aβ) overproduction is one of the most important pathological features in Alzheimer's disease (AD). Endoplasmic reticulum (ER) stress induced by Aβ overload plays a critical role in this process. Bis(ethylmaltolato)oxidovanadium (IV) (BEOV), a vanadium compound which had been regarded as peroxisome proliferator-activated receptor γ (PPARγ) agonist, was reported to exert an antagonistic effect on ER stress. In this study, we tested whether BEOV could ameliorate the Aβ-induced neuronal apoptosis by inhibiting ER stress. It was observed that BEOV treatment ameliorated both tunicamycin-induced and/or Aβ-induced ER stress and neurotoxicity in a dose-dependent manner through downgrading ER stress-associated and apoptosis-associated proteins in primary hippocampal neurons. Consistent with in vitro results, BEOV also reduced ER stress and inhibited neuronal apoptosis in hippocampi and cortexes of transgenic AD model mice. Moreover, by adopting GW9662 and salubrinal, the inhibitor of PPARγ and hyperphosphorylated eukaryotic translation initiation factor 2α, respectively, we further confirmed that BEOV alleviated Aβ-induced ER stress and neuronal apoptosis in primary hippocampal neurons by activating PPARγ. Taken together, these results provided scientific evidences to support the concept that BEOV ameliorates Aβ-induced ER stress and neuronal apoptosis through activating PPARγ., Competing Interests: Declaration of competing interest All authors declare that they have no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

As a top leading cause of cancer death in many countries, colorectal cancer (CRC) has drawn increasing attention to the study of the pathological mechanism. According to the "cancer stem cell hypothesis", malignancies originate from a small fraction of cancer cells that show self-renewal properties to initiate and sustain tumor growth and tumor metastasis. Therefore, these cancer stem cells (CSC) probably play important roles in tumor recurrence, metastasis, and drug resistance. Previous research reported that lysine-specific histone demethylase 1 (LSD1) maintains cancer stemness through up-regulating stemness markers SOX2 and OCT4. CD133 is believed to be the most robust surface marker for CRC stem cells, however the regulatory effect of LSD1 on stemness of CD133+ CRC has never been reported. In this study, our objectives included: 1) to isolate pure CD133+ and CD133- cells from SW620 cell line; 2) to investigate the effect of LSD1 on the characteristics of CD133+ stem cancer cells by knocking down the target gene. Results suggested that the SW620 cell line had both CD133+ and CD133- subsets. The CD133+ subset exhibited more CSC-like characteristics compared with the CD133- subset with higher viability, colony formation rate, migration and invasion rate, resistance to anti-cancer drugs, and apoptosis in vitro. The CD133+ also induced faster tumor formation and larger tumors in vivo. In the LSD1-knockdown CD133+ cells, the CSC-like characteristics had been all weakened. We conclude that LSD1 was important for CSCs to maintain their "stemness" features, which could be a potential therapeutic target of CRC.

Objective: The incidence of acute myocardial infarction (AMI) is increasing year by year, and it has become one of the diseases with the highest mortality in humans. MicroRNAs (miRNAs) are involved in the regulation of many diseases, including AMI. This study aims to investigate the function of miR-147 in myocardial infarction (MI) and its underlying mechanism of action., Materials and Methods: Quantitative Reverse Transcription-Polymerase Chain Reaction (qRT-PCR) was used to detect miR-147, Bcl-2 mRNA, and Bax mRNA expressions. Enzyme-linked immunosorbent assay (ELISA) kits were used to detect the levels of inflammatory factors (TNF-α, IL-6, IL-β) and lactate dehydrogenase (LDH). Besides, MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assay was performed to detect cell viability. Cell apoptosis was observed using terminal dexynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL). Moreover, echocardiography was utilized to measure cardiac function of rats. HIPK2 expression was detected by Western blot., Results: MiR-147 expression was significantly decreased in rat MI model and H2O2 treated H9c2 cells. H2O2 treatment increased the expression of inflammatory factors in H9c2 cells and induced apoptosis, while such effects were inhibited by overexpression of miR-147. Overexpression of miR-147 reversed the decrease in Bcl-2 expression and the increase in Bax expression in H9c2 cells caused by H2O2. In addition, overexpression of miR-147 could improve cardiac function and reduce serum LDH levels in myocardial infarction rats. Through TargetScan, we found that HIPK2 might have a binding site for miR-147. Moreover, overexpression of miR-147 could inhibit the expression of HIPK2., Conclusions: In MI, miR-147 expression is decreased in the myocardium and overexpression of miR-147 can inhibit myocardial inflammation and apoptosis and improve cardiac function in rats via targeting HIPK2.

C6-ceramide is an exogenous short-chain ceramide which can induce apoptosis of multiple cancer cells. Salivary adenoid cystic carcinoma (SACC) is a common salivary gland cancer, which possesses of high rate of local recurrence and distant metastasis. The mechanism of ceramide-induced SACC-83 and SACC-LM cell apoptosis has not been revealed. In our study, gene expression microarray was used to discover that the unfolded protein response (UPR) pathway, especially PRKR-like endoplasmic reticulum kinase (PERK) pathway, was the major activated pathway after treatment of c6-ceramide. D1ER, an endoplasmic-reticulum-targeted Ca2+ indicator, was used to measure Ca2+ release from endoplasmic reticulum (ER) dynamically. We found that inositol 1,4,5-trisphosphate receptor 3 (IP3R3) was activated, leading to Ca2+ release from ER, soon after c6-ceramide treatment. IP3R3 silencing could block UPR, although it could not prevent SACC-83 and SACC-LM cells from apoptosis. Moreover, we found that C/EBP-homologous protein could upregulate in a UPR-independent way. Mitochondria outer membrane permeabilization might play an important role in inducing SACC cell apoptosis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Acetylation is an important, highly conserved, and reversible post-translational modification of proteins. Previously, we showed by nano-HPLC/MS/MS that many nutrient storage proteins in the silkworm are acetylated. Among these proteins, most of the known 30K proteins were shown to be acetylated, including 23 acetylated 30K proteins containing 49 acetylated sites (Kac), indicating the importance of the acetylation of 30K proteins in silkworm. In this study, Bm30K-3, a 30K protein containing three Kac sites, was further assessed in functional studies of its acetylation. Increasing the level of Bm30K-3 acetylation by adding the deacetylase inhibitor trichostatin A (TSA) increased the levels of this protein and further inhibited cellular apoptosis induced by H 2 O 2 . In contrast, decreasing the level of acetylation by adding the acetylase inhibitor C646 could reduce the level of Bm30K-3 and increase H 2 O 2 -induced apoptosis. Subsequently, BmN cells were treated with CHX and MG132, and increasing the acetylation level using TSA was shown to inhibit protein degradation and improve the stability of Bm30K-3. Furthermore, the acetylation of Bm30K-3 could compete with its ability to be ubiquitinated, suggesting that acetylation could inhibit the ubiquitin-mediated proteasome degradation pathway, improving the stability and accumulation of proteins in cells. These results further indicate that acetylation might regulate nutrition storage and utilization in Bombyx mori, which requires further study., (© 2020 Wiley Periodicals, Inc.)

We herein develop a novel two-photon fluorescent probe termed L-pH for visualization of lysosomal pH within live cells. L-pH is composed of three moieties, including naphthalimide fluorophore as a fluorescence off-on response moiety, piperazine and morpholine groups as lysosomal targeting and pH responsive sites, as well as a reactive benzyl chloride segment for further lysosomal anchoring. The experimental results demonstrate that L-pH can instantaneously respond to various pH values with high sensitivity and selectivity, and has low cytotoxicity and excellent photostability. The one-photon and two-photon fluorescence imaging data indicate L-pH can preferably accumulate into lysosome and monitor the rise of lysosomal pH changes during myriad cell stress conditions, including heat shock, cell apoptosis and mitophagy. Moreover, L-pH was applied for imaging of pH difference in abdominal tissues of mice. L-pH will be a potential tool for monitoring lysosomal pH variation during lysosome-associated physiological and pathological states., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Background: Ovarian cancer is the most lethal gynecologic malignancy worldwide with poor prognosis owing to chemotherapy resistance and cancer relapse. Hence, there is an urgent need to develop novel anticancer agents against ovarian cancer., Objective: The aim of this research is to investigate the possible anticancer activity of aloperine, an active ingredient from a traditional Chinese medicine Sophora alopecuroides, and to explore the possible Reactive Oxygen Species (ROS)-related mechanism., Methods: Cell viability, cytotoxicity, apoptosis, ROS generation, and oxidant stress indicators were analyzed., Results: Our results demonstrated that aloperine significantly induced inhibition of cell viability, promoted cytotoxicity and mitochondrial-related apoptosis, and increased ROS generation in ovarian cancer cells. Furthermore, the antioxidant α-lipoic acid reversed apoptosis in aloperinetreated cells. In addition, we identified hydrogen peroxide as the main type of ROS, and the antioxidant catalase suppressed the apoptotic inducing effect of aloperine whereas hydrogen peroxide supplement exacerbated the effect of aloperine in ovarian cancer cells., Conclusion: Taken together, our results indicated that aloperine could exert anti-ovarian cancer cell activity through a reactive oxygen species activation mechanism and suggested aloperine as a potential agent against ovarian cancer., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Objective: This research was aimed to detect the functions of Lycium barbarum polysaccharides (LBPs) on oxygen and glucose deprivation (OGD) injury and potential mechanisms at PC-12 cells. Methods: CCK-8, flow cytometry and reactive oxygen species (ROS) assays were used to detect OGD, LBPs and miR-24 effects on cell viability, apoptosis, and oxidative stress. MiR-24 was transfected and texted by transfection and qRT-PCR. Moreover, the related-protein levels of apoptosis, autophagy and pathways were tested by Western blotting. Results: LBPs significantly enhanced cell viability , inhibited cell apoptosis, autophagy and ROS level in OGD injury. In addition, miR-24 expression was declined in OGD-treated cells, while it was elevated when added LBPs. The preventive effects of LBPs on PC-12 cell damage induced by OGD were reversed by down-regulating miR-24. Furthermore, miR-24 inhibitor declined LBPs-induced change in Wnt/β-catenin and JAK1/STAT3 pathways in OGD-injuried cells. Conclusions: LBPs exhibited preventive effects via up-regulating miR-122 and activating Wnt/β-catenin and JAK1/STAT3 pathways in OGD-induced PC-12 cells.

Gastric cancer is an aggressive disease and a common cause of cancer‑associated mortality worldwide. Recent studies have indicated that follistatin‑like protein 1 (FSTL‑1) is expressed and serves essential roles in tumorigenesis; however, the specific functional mechanism of FSTL‑1 in gastric cancer progression remains ambiguous. CellTiter‑Glo Luminescent Cell Viability and lactate dehydrogenase assays were used to measure cell survival and cell cytotoxicity, respectively. Cell apoptosis was ascertained using the Cell Death Detection ELISA assay and caspase‑3/9 activity kits. Reverse transcription‑quantitative polymerase chain reaction and western blotting were used to detect the expression levels of FSTL‑1. The present study confirmed that FSTL‑1 was highly expressed in gastric cancer cells compared with in control cells. Subsequently, FSTL‑1 inhibition by small interfering RNA significantly reduced cancer cell survival and induced cytotoxic effects. In addition, knockdown of FSTL‑1 in gastric cancer cells promoted apoptosis by increasing caspase‑3 and caspase‑9 expression. A decrease in signal transducer and activator of transcription 6 (STAT6) phosphorylation was observed in FSTL‑1 knockdown cells, and the results confirmed that STAT6 phosphorylation was essential for FSTL‑1 knockdown‑induced cell apoptosis of cancer cells. Taken together, these results demonstrated that FSTL‑1 knockdown may promote cell apoptosis via the STAT6 signaling pathway; therefore, FSTL1 may be considered a novel diagnostic and therapeutic target for gastric cancer.

It has been demonstrated that microRNAs (miRNAs) may contribute to tumorigenesis and tumor growth in osteosarcoma (OS), which is a primary malignant tumor of bone frequently diagnosed in adolescents and young people. The purpose of our investigation was to evaluate the functional relevance of miR-377 in OS and to investigate whether the mechanism was related to the histone acetyltransferase 1 (HAT1)-mediated Wnt signaling pathway. By screening differentially expressed genes in microarray GSE47572, HAT1 was found to be a candidate gene of interest. Besides, the regulatory miRNA (miR-377) of HAT1 was also selected. The interaction among miR-377, HAT1, and the Wnt signaling pathway was evaluated. In addition, the miR-377 expression was altered in OS cells (U-2OS and SOSP-9607) to assess the in vitro cell apoptosis and the in vivo tumor growth. OS tissues presented elevated HAT1 expression and decreased miR-377 expression. A putative miR-377 binding site in HAT1 3'-UTR HAT1 was verified. Cells with miR-377 overexpression or HAT1 silencing were observed to exhibit reduced HAT1 expression and promoted apoptosis, accompanied by blockade of Wnt signaling. Moreover, the in vivo experiment revealed that miR-377 overexpression or HAT1 silencing inhibited tumor growth and reduced tumor size in nude mice. Taken together, our results conclude that miR-377 may promote OS cell apoptosis through inactivation of the HAT1-mediated Wnt signaling pathway, highlighting the potential therapeutic effect of miR-377 on OS treatment., (© 2019 Wiley Periodicals, Inc.)

Cell death plays a pivotal role in animal development and tissue homeostasis. Dysregulation of this process is associated with a wide variety of human diseases, including developmental and immunological disorders, neurodegenerative diseases and tumors. While the fundamental role of JNK pathway in cell death has been extensively studied, its down-stream regulators and the underlying mechanisms remain largely elusive. From a Drosophila genetic screen, we identified Snail (Sna), a Zinc-finger transcription factor, as a novel modulator of ectopic Egr-induced JNK-mediated cell death. In addition, sna is essential for the physiological function of JNK signaling in development. Our genetic epistasis data suggest that Sna acts downstream of JNK to promote cell death. Mechanistically, JNK signaling triggers dFoxO-dependent transcriptional activation of sna. Thus, our findings not only reveal a novel function and the underlying mechanism of Sna in modulating JNK-mediated cell death, but also provide a potential drug target and therapeutic strategies for JNK signaling-related diseases.

The importance of vascular cell glycocalyx in mechanotransduction has been demonstrated by many studies. The simulated microgravity induced a region-dependent adaptation of arterial glycocalyx including its thickness, coverage, and gene expression in conduit arteries of tail-suspended rats has been reported in our previous studies. Herein, we extended this line of research by quantifying the mRNA levels of three nitric oxide synthase (NOSI, NOSII, and NOSIII) and evaluating the apoptotic rates of endothelial cells (ECs) and smooth muscle cells (SMCs) in the common carotid artery, abdominal aorta, and femoral artery of 3 week tail-suspended rats. Results indicated that the tail suspension of rats induced about 0.36, 0.22, and 0.33 fold down-regulation of NOSI, NOSII, and NOSIII in the abdominal aorta, while 3.21, and 3.48 fold up-regulation of NOSII and NOSIII in the carotid artery and no significant effects on three NOS isoforms in the femoral artery. Moreover, the apoptosis of ECs and SMCs were significantly inhibited in both carotid artery and abdominal aorta, while enhanced in the femoral artery of the tail-suspended rats. A linear positive correlation exists between the normalized coverage of the glycocalyx and the normalized NOSI and NOSIII mRNA levels. These results indicated that the redistribution of haemodynamics in the conduit arteries of 3 week tail-suspended rats regulated the glycocalyx, NOS expression, and vascular cell apoptosis in a region-dependent manner, contributing to the final vascular remodelling under simulated microgravity condition., (© 2019 John Wiley & Sons Australia, Ltd.)

CGA-N9, a peptide derived from human chromogranin A (CGA), was found to have antimicrobial activity in our previous investigation, but its mechanism of action remains unclear. Herein, the mechanism of action of CGA-N9 was investigated. We found that CGA-N9 induced the depolarization of the cell membrane and uptake of calcium ions into the cytosol and mitochondria. With the disruption of the mitochondrial membrane potential, the generation of intracellular reactive oxygen species (ROS) increased. Accordingly, we assessed apoptotic processes in Candida tropicalis cells post-treatment with CGA-N9 and found cytochrome c leakage, chromatin condensation and DNA degradation. The interaction of CGA-N9 with DNA in vitro showed that CGA-N9 did not degrade DNA but bound to DNA via an electrostatic interaction. In conclusion, CGA-N9 exhibits antifungal activity by inducing apoptosis in C. tropicalis., (© 2019 The Author(s).)

Objective: Atherosclerosis is one of the most important risk factors for coronary heart disease (CHD), and growing evidence has shown that long non-coding RNAs (lncRNAs) can serve as prospective markers for atherosclerosis. In this study, we mainly focused on the potential roles of linc00961 in CHD patients., Patients and Methods: qRT-PCR was used to detect the expressions of linc00961 and miR-367 in CHD patients and ApoE-/-mice, and the correlations were analyzed. Then, HA-VAMC was respectively treated with 5 inflammatory factors and hypoxia conditions to explore the factors that affect linc00961 levels. Furthermore, the linc00961 overexpression lentivirus (LV-linc00961) and linc00961 downregulation lentivirus (LV-sh linc00961) were purchased and transfected into human vascular smooth muscle cells (VSMCs). CCK8 assay was carried out to measure the cell proliferation of VSMC, and the levels of Cyclin D1, Bcl-2, Bax, and cleaved caspase-3 were detected by RT-PCR and Western blot. Moreover, the Luciferase assay was performed to explore the binding site of linc00961 and miR-367. Finally, the miR-367 inhibitor was transfected into LV-sh linc00961 VSMCs to confirm the linc00961 functions via miR-367., Results: We found that linc00961 was significantly decreased in patients with CHD and ApoE-/-mice. Additionally, linc00961 was reduced in VSMCs at the conditions of hypoxia and C-reactive protein (CRP). Most importantly, the overexpression of linc00961 significantly inhibited the VSMCs proliferation, repressed the levels of Cyclin D1 and Bcl-2, and increased the levels of Bax and cleaved caspase-3. However, the downregulation of linc00961 promoted VSMCs proliferation, increased the levels of Cyclin D1 and Bcl-2, and repressed the levels of Bax and cleaved caspase-3. We also found that miR-367 was downregulated following the upregulation of linc00961, while it was upregulated following the downregulation of linc00961. The Luciferase gene reporter assay indicated that linc00961 could directly bind with miR-367 in VSMCs. Finally, we found that linc00961 could inhibit proliferation and promote apoptosis of VSMCs via binding with miR-367., Conclusions: According to the results, our study revealed that linc00961 was significantly decreased in patients with CHD and ApoE-/-mice. Furthermore, our findings firstly uncovered that linc00961 was reduced by hypoxia and CRP in VSMCs. The downregulation of linc00961 contributed to promote proliferation and inhibit apoptosis of VSMCs by sponging miR-367 in CHD patients, which might provide a potential target for treating atherosclerosis.

Background: Rheumatoid arthritis (RA) is a known intractable chronic inflammatory disease of synovial joints characterized by hyperplasia and consecutive inflammation with a high prevalence.Guizhi-Shaoyao-Zhimu (GSZD) is the first choice for clinical treatment of RA in Chinese traditional medicine. This study is aimed to explore the possible pharmacological mechanisms of anti-arthritic effect of GSZD., Methods: Type II collagen-induced arthritis (CIA) rat model was used to study the anti-arthritic activity of GSZDin vivo, and toe swelling & arthritis score, serum levels of cytokines, and pathological examinations were carried out. In vitro, TNF-α induced MH7A cells were used to study the possible mechanisms of GSZD. The anti-proliferative effects of GSZD were determined by MMT assay, and pro-apoptotic activity of GSZD in MH7A cells was determined by flow cytometry analysis & DAPI staining. Furthermore, the adhesive and invasive abilities of MH7A cells were determined using cell adhesion and transwell assays. MMPs levels were determined by ELISA assays, and mRNA expressions of Caspase-3, -9, Bax, SOCS1, Bcl-2, JAK2, STAT-3 and -5 were determined using qRT-PCR analysis. Besides, the major chemical components in GSZD were analyzed by HPLC-QqQ-MS analysis., Results: Our results showed GSZD reduced the toe swelling & arthritis score, and serum levels of TNF-α, IL-1β, IL-6 & IL-17a in CIA rats; pathological examination results indicated GSZD improved ankle joint injury in CIA rats.In vitro, GSZD showed significant anti-proliferative and pro-apoptotic effects on TNF-α stimulated MH7A cells. After GSZD treatment, the adhesive and invasive abilities of MH7A cells were reduced, and secretions of MMPs, IL-6 and IL-8 were also reduced. GSZD decreased the releases of TNF-α and IL-1β in LPS stimulated RAW 264.7 cells. Further studies showed GSZD up-regulated mRNA expressions of Caspase-3, -9, Bax, and SOCS1, whereas down-regulated mRNA expressions of Bcl-2, JAK2, STAT3 and STAT5. Besides, 13 major chemical components were identified in GSZD extracts through HPLC-QqQ-MS analysis., Conclusion: Our results suggested GSZD possesses an anti-rheumatic effect on CIA rats, and the possible mechanism is related to inhibiting inflammatory response, inhibiting invasion and migration of synovial fibroblasts, and inducing apoptosis in synovial fibroblasts., (Copyright © 2019 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Objective Psoriasis is an immune-mediated inflammatory disease. Despite advances in the study of its pathogenesis, the exact development mechanism of psoriasis remains to be fully elucidated. Hyperproliferative epidermis plays a crucial role in psoriasis. This study aimed to investigate the effects of interleukin-36 β (IL-36 β ) on keratinocyte dysfunction in vitro . Methods Human keratinocyte cell lines, HaCaT cells, were treated with 0 (control), 50 or 100 ng/ml IL-36 β respectively for 24 h. Cell viability was determined with a cell counting kit-8 assay. Flow cytometry was used to assess the effects of IL-36 β on apoptosis and cell cycle distribution. Expressions of the differentiation markers, such as keratin 10 and involucrin, were evaluated by quantitative real-time polymerase chain reaction (RT-qPCR). Expressions of the inflammatory cytokines, IL-1 β and IL-6 were tested by ELISA. Results CCK8 assay showed the survival rate had no significant difference between the control and treated group ( P > 0.05). Flow cytometry analysis showed cell cycle arrest at S phase in the IL-36 β -treated groups compared with the control group ( P < 0.05). RT-qPCR verified the decreased mRNA expressions of keratin 10 and involucrin in the IL-36 β -treated groups compared with the negative control ( P < 0.01). ELISA showed 100 ng/ml IL-36 β enhanced levels of IL-1 β and IL-6 in culture supernatants of HaCaT cells compared with the negative control ( P < 0.05). Conclusion Taken together, these findings suggest that IL-36 β could induce cell cycle arrest at S phase, inhibit keratin 10 and involucrin expressions and promote inflammatory activity in HaCaT cell lines.