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Objective: This study aimed to analyze the impact of HGF on cardiomyocyte injury, apoptosis, and inflammatory response induced by lipopolysaccharide (LPS)., Methods: Enzyme-linked immunosorbent assay (ELISA) was utilized to quantify the levels of HGF, interleukin (IL)-6, IL-10, creatine phosphokinase-isoenzyme-MB (CK-MB), and cardiac troponin I (cTnI) in the samples. qPCR and Western blotting (WB) were employed to assess the mRNA and protein expressions of HGF, IL-10, IL-6, PI3K, AKT, p-PI3K, and p-AKT., Results: The outcomes of the in vivo experiment revealed that serum levels of IL-6, IL-10, HGF and SOFA scores in the SC group were elevated in contrast to the non-SC group. The correlation analysis indicated a substantial and positive association among serum HGF, IL-6, and IL-10 levels and SOFA scores. Relative to IL-6, IL-10 levels, and SOFA scores, serum HGF demonstrated the highest diagnostic value for SC. Following LPS administration to stimulate H9c2 cells across various periods (0, 12, 24, 48, and 72 h), the levels of myocardial injury markers (CK-MB and cTnI) in the cell supernatants, intracellular inflammatory factors (mRNA and protein levels of IL-10 and IL-6), apoptosis and ROS levels, exhibited a gradual increase followed by a subsequent decline. Following the overexpression of HGF, there was an increase in cell viability, and a decrease in apoptosis, inflammation, oxidative stress injuries, and the protein phosphorylation expressions of PI3K and AKT. After knockdown of HGF expression, the activity of LPS-induced H9c2 cells was further reduced, leading to increased cell injury, apoptosis, inflammation, oxidative stress,and the expression levels of PI3K and Akt protein phosphorylation were further elevated., Conclusion: HGF was associated with decreased LPS-induced H9c2 apoptosis and inflammation in H9c2 cells, alongside an improvement in cell viability, indicating potential cytoprotective effects. The mechanism underlying these impacts may be ascribed to the suppression of the PI3K/AKT signaling pathway., 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 Elsevier B.V. All rights reserved.)

Cerebral ischemia-reperfusion injury is a complex pathophysiological process. Poly(ADP-ribose) (PAR) polymerase-1 (PARP-1)/apoptosis-inducing factor (AIF) signaling pathway-mediated apoptosis is one of the non-caspase-dependent cell death programs that are widely present in neurological diseases such as stroke. In our study, we aimed to conduct further research on the effects of Gualou Guizhi decoction (GLGZD) on the PARP-1/AIF signaling pathway in cell apoptosis after ischemia-reperfusion injury caused by middle cerebral artery occlusion (MCAO). The results showed that GLGZD administration for 7 days significantly ameliorated MCAO-induced neurological damage, limb paralysis and the pathological state of the ischemic cortex. GLGZD exerted its effects by significantly reducing the volume of ischemic cerebral infarction, increasing the number of Nissl-positive cells, and reducing neuronal apoptosis. Furthermore, Western blot analysis showed that GLGZD significantly inhibited the total protein expression of PARP-1, PAR, AIF and endonuclease G (Endo G) in the ischemic cortex and significantly increased the total protein expression of heat-shock protein 70 (Hsp70). On the one hand, the expression of PARP-1, AIF and Endo G protein in the nucleus significantly decreased while the expression of PAR nucleoprotein significantly upregulated. On the other hand, compared with the MCAO model group, the GLGZD-treated group showed a significantly reduced protein expression of PAR in mitochondria and significantly increased protein expression of mitochondrial AIF and Endo G. It was concluded that GLGZD had good therapeutic effects in MCAO model rats. These effects were closely related to GLGZD-mediated inhibition of ischemia-induced neuronal apoptosis by regulation of protein expression and translocation in the PARP-1/AIF signaling pathway.

Coronavirus membrane (M) protein is the most abundant structural protein playing a critical role in virion assembly. Previous studies show that the N-terminal ectodomain of M protein is modified by glycosylation, but its precise functions are yet to be thoroughly investigated. In this study, we confirm that N-linked glycosylation occurs at two predicted sites in the M protein ectodomain of infectious bronchitis coronavirus (IBV). Dual mutations at the two sites (N3D/N6D) did not affect particle assembly, virus-like particle formation and viral replication in culture cells. However, activation of the ER stress response was significantly reduced in cells infected with rN3D/N6D, correlated with a lower level of apoptosis and reduced production of pro-inflammatory cytokines. Taken together, this study demonstrates that although not essential for replication, glycosylation in the IBV M protein ectodomain plays important roles in activating ER stress, apoptosis and proinflammatory response, and may contribute to the pathogenesis of IBV., (Copyright © 2019. Published by Elsevier Inc.)

Epigallocatechin-3-gallate (EGCG), the major constituent of green tea, has been shown to induce cell death in cancer cells. Primary effusion lymphoma (PEL) is an aggressive neoplasm caused by human herpesvirus 8 (HHV8). In this study, we examined the role of EGCG on PEL cells in cell death and HHV8 replication. We performed trypan blue exclusion assay to assess the cell viability of PEL cells, flow cytometry analysis to examine the cell cycle distribution and reactive oxygen species (ROS) generation, caspase-3 activity to assay apoptosis, acridine orange staining to determine autophagy, and immunoblotting to detect the protein levels involved in apoptosis and autophagy as well as mitogen activated protein kinases (MAPKs) activation upon EGCG treatment. The expression of the HHV8 lytic gene was determined by luciferase reporter assay and reverse transcription-PCR, and viral progeny production was determined by PCR. Results revealed that EGCG induced cell death and ROS generation in PEL cells in a dose-dependent manner. N -acetylcysteine (NAC) inhibited the EGCG-induced ROS and rescued the cell from EGCG-induced cell death. Even though EGCG induced ROS generation in PEL cells, it reduced the production of progeny virus from PEL cells without causing HHV8 reactivation. These results suggest that EGCG may represent a novel strategy for the treatment of HHV8 infection and HHV8-associated lymphomas., Competing Interests: The authors declare no conflict of interest.

Pancreatic cancer (PC), as the leading cause of cancer death worldwide, is one of the deadliest tumors with a very low 5-year survival rate. Therefore, it is urgent to seek new biomarkers of PC for more accurate and reliable treatments. To identify the differentially expressed miRNAs (DEM) in PC tissues, we performed the systematic microarray and qRT-PCR analyses. We found miR-196b was the top dysregulated DEM in PC tissues as compared with the corresponding adjacent tissues, and positively correlated with poor differentiation, tumor size, lymphatic invasion and TNM stage. Furthermore, the late apoptosis rate was significantly reduced, while the cell proliferation was increased in PANC-1 and ASPC-1 cell-lines after treatment with miR-196b mimics. The qRT-PCR and Western blot analysis demonstrated that the level of CADM1 in PANC-1 cells response to the alteration of miR-196b. Moreover, blockade of CADM1 could decrease the late apoptosis in PANC-1 cells as up-regulated by inhibition of miR-196b. Finally, luciferase report assay confirmed that CADM1 was the direct target gene of miR-196b. Overexpression of miR-196b in PC tissues can increase the late apoptosis of pancreatic cancer cells by targeting CADM1. These findings suggested miR-196b is a potential target for diagnosis and therapeutics of human pancreatic cancer.

CPT-11 (Irinotecan) is a first-line chemotherapeutic agent in clinic, but it may induce side effects including diarrhea and enteritis in patients. The underlying mechanism of CPT-11's intestinal toxicity is unclear. Peritoneal resident macrophages have been reported to be important for the maintenance of intestinal homeostasis. In this study, we evaluated the cytotoxic effects of CPT-11 on mouse peritoneal resident macrophages. CPT-11 was administered intraperitoneally to mice and their peritoneal exudate cells were isolated for evaluation. CPT-11 treatment strikingly decreased the ratio of F4/80(hi)MHCII(low) large peritoneal macrophages (LPMs), which are regarded as prenatally-originated peritoneal resident macrophages. Consistent with this, the transcription factor GATA6 specifically expressed in LPMs was barely detectable in the macrophages from CPT-11-treated mice, indicative of elimination of LPMs. Such elimination of LPMs was at least partly due to CPT-induced apoptosis in macrophages, because inhibition of apoptosis by caspase-3 inhibitor z-DEVD-fmk significantly diminished the loss of GATA6(+) LPMs. As GATA6 is a transcription factor that controls expression of multiple genes regulating peritoneal B-1 cell development and translocation, elimination of GATA6(+) LPMs led to a great reduction in B-1 cells in the peritoneal cavity after CPT-11 treatment. These results indicated that CPT-11-induced apoptosis contributed to the elimination of peritoneal resident macrophages, which might in turn impair the function of peritoneal B-1 cells in maintaining intestinal homeostasis. Our findings may at least partly explain why CPT-11 treatment in cancer patients induces diarrhea and enteritis, which may provide a novel avenue to prevent such side effects.

Background: Adenomyosis is a specific subtype of endometriosis and recent evidences have indicated that Tanshinone IIA (TSIIA) might be a potential therapeutic option for endometriosis. Meanwhile, endometrial stromal cells (ESCs) of adenomyosis might play crucial roles in the progression of this disease, emphasizing the importance of targeting ESCs in the treatment of adenomyosis. Furthermore, previous evidences also implicated that deregulated 14-3-3ζ expression might be associated with therapeutic effects of certain drugs., Aim of the Study: The aim of this study is to evaluate the potential involvement of 14-3-3ζ in the process of TSIIA-treated adenomyosis., Materials and Methods: Ectopic endometrial stromal cells (EESCs) were isolated from a total of 3 patients with adenomyosis. Cells were treated with TSIIA and infected with 14-3-3ζ-overexpressing adenovirus, the expression level of 14-3-3ζ was determined by western blotting (WB), cell viability was detected by Cell Counting Kit-8 (CCK8), cell invasion and migration was evaluated by transwell assay, and cell apoptosis was detected by flow cytometry., Results: TSIIA could decrease cell viability, induce cell apoptosis, and inhibit cell migration and invasion in EESCs. Mechanistically, TSIIA markedly reduced the expression of 14-3-3ζ in EESCs, and overexpression of 14-3-3ζ could restore the ability of cell viability, migration and invasion, but has no effect on cell apoptosis., Conclusions: TSIIA could be a promising novel therapeutic agent for adenomyosis, via inducing cell apoptosis, inhibiting cell viability, migration and invasion in EESCs. Furthermore, the effects of cell viability, migration and invasion were mediated in 14-3-3ζ-dependent manner while that of cell apoptosis was mediated in 14-3-3ζ-independent manner.

Paeoniflorin (PF) is a principal bioactive component, which exhibits many pharmacological effects, including protection against ischemic injury. This paper aimed to investigate the protective effect of PF both in vivo and in vitro. Middle cerebral artery occlusion (MCAO) was performed on male Sprague-Dawley (SD) rat for 2 h, and different doses of PF or vehicle were administered 2 h after reperfusion. Rats were sacrificed after 7 days treatment of PF/vehicle. PF treatment for 7 days ameliorated MCAO-induced neurological deficit and decreased the infarct area. Further study demonstrated that PF inhibited the over-activation of astrocytes and apoptosis of neurons, and PF promoted up-regulation of neuronal specific marker neuron-specific nuclear (NeuN) and microtubule-associated protein 2 (MAP-2) in brain. Moreover, NMDA-induced neuron apoptosis was employed. The in vitro study revealed that PF treatment protected against NMDA-induced cell apoptosis and neuronal loss via up-regulation of neuronal specific marker NeuN, MAP-2 and Bcl-2 and the down-regulation Bax. Taken together, the present study demonstrates that PF produces its protective effect by inhibiting the over-activation of astrocytes, apoptosis of neurons and up-regulation of neuronal specific marker NeuN, MAP-2, and B-cell lymphoma-2 (Bcl-2), and down-regulation Bax. Our study reveals that PF may be a potential neuroprotective agent for stroke and can provide basic data for clinical use.

Induction of the unfolded protein response (UPR) is an adaptive cellular response to endoplasmic reticulum (ER) stress that allows a cell to reestablish ER homeostasis. However, under severe and persistent ER stress, prolonged UPR may activate unique pathways that lead to cell death. In this study, we investigated the activation of the protein kinase R-like ER kinase (PERK) pathway of UPR in cells infected with the coronavirus infectious bronchitis virus (IBV) and its relationship with IBV-induced apoptosis. The results showed moderate induction of PERK phosphorylation in IBV-infected cells. Meanwhile, activating transcription factor 4 (ATF4) was upregulated at the protein level in the infected cells, resulting in the induction in trans of the transcription factor ATF3 and the proapoptotic growth arrest and DNA damage-inducible protein GADD153. Knockdown of PERK by small interfering RNA (siRNA) suppressed the activation of GADD153 and the IBV-induced apoptosis. Interestingly, knockdown of protein kinase R (PKR) by siRNA and inhibition of the PKR kinase activity by 2-aminopurine (2-AP) also reduced the IBV-induced upregulation of GADD153 and apoptosis induction. In GADD153-knockdown cells, IBV-induced apoptosis was suppressed and virus replication inhibited, revealing a key role of GADD153 in IBV-induced cell death and virus replication. Analysis of the pathways downstream of GADD153 revealed much more activation of the extracellular signal-related kinase (ERK) pathway in GADD153-knockdown cells during IBV infection, indicating that GADD153 may modulate apoptosis through suppression of the pathway. This study provides solid evidence that induction of GADD153 by PERK and PKR plays an important regulatory role in the apoptotic process triggered by IBV infection.

Triptolide, a natural product derived from the Chinese plant Tripterygium wilfordii, is reported to exhibit antitumor effects in a broad range of cancers. The antitumor activity of triptolide is associated with its biologic activities, as it inhibits various proproliferative or antiapoptotic factors that are dominantly expressed in given types of cancer cells. Herein, we show that triptolide induced apoptosis in a subgroup of acute lymphoblastic leukemia (ALL) cells overexpressing the MDM2 oncoprotein by inhibiting MDM2 expression. More specifically, we found that triptolide inhibited MDM2 at the transcriptional level by suppressing its mRNA synthesis. This MDM2 inhibition led in turn to increased levels of p53 protein; however, p53 functionality was not activated due to the fact that triptolide-treated cells lacked induction of p21 and PUMA as well as in G(1) cell-cycle arrest. Triptolide-mediated downregulation of MDM2 increased inhibition of X-linked inhibitor of apoptosis protein (XIAP), its translational target, in a manner distinct from reactions to cellular stress and DNA-damaging agent ionizing radiation that induce XIAP due to p53-activated MDM2. These results suggest that increased inhibition of XIAP due to downregulation of MDM2 may play a critical role in triptolide-induced apoptosis in MDM2-overexpressing cancers., (©2012 AACR.)

Background: MicroRNAs are involved in various critical functions, including the regulation of cellular differentiation, proliferation, angiogenesis, and apoptosis. We hypothesize that microRNA-210 can rescue cardiac function after myocardial infarction by upregulation of angiogenesis and inhibition of cellular apoptosis in the heart., Methods and Results: Using microRNA microarrays, we first showed that microRNA-210 was highly expressed in live mouse HL-1 cardiomyocytes compared with apoptotic cells after 48 hours of hypoxia exposure. We confirmed by polymerase chain reaction that microRNA-210 was robustly induced in these cells. Gain-of-function and loss-of-function approaches were used to investigate microRNA-210 therapeutic potential in vitro. After transduction, microRNA-210 can upregulate several angiogenic factors, inhibit caspase activity, and prevent cell apoptosis compared with control. Afterward, adult FVB mice underwent intramyocardial injections with minicircle vector carrying microRNA-210 precursor, minicircle carrying microRNA-scramble, or sham surgery. At 8 weeks, echocardiography showed a significant improvement of left ventricular fractional shortening in the minicircle vector carrying microRNA-210 precursor group compared with the minicircle carrying microRNA-scramble control. Histological analysis confirmed decreased cellular apoptosis and increased neovascularization. Finally, 2 potential targets of microRNA-210, Efna3 and Ptp1b, involved in angiogenesis and apoptosis were confirmed through additional experimental validation., Conclusions: MicroRNA-210 can improve angiogenesis, inhibit apoptosis, and improve cardiac function in a murine model of myocardial infarction. It represents a potential novel therapeutic approach for treatment of ischemic heart disease.

Objective: To investigate the effect and mechanism of FTY720 on acute graft versus host disease (GVHD) in rat small bowel transplantation (SBTx)., Methods: Heterotopic SBTx was performed using a parent (WF)-into-F1 (WFxACI) rat combination. Recipient rats were divided into experimental group (n=6) and control group (n=6). Rats in the experimental group were administered with FTY720 at 0.5 mg/kg for 14 days. Lymphocyte apoptosis in the liver and the mucosa of intestine and graft was detected by TUNEL and flow cytometry 15 days after transplantation. Recipient survival and lymphocyte apoptosis were compared between the two groups., Results: Recipients in the control group died of GVHD after a mean survival time of (16+/-2.1) days. FTY720-treated recipients had a significantly longer survival (>100 days). After administration of FTY720, the percentage of apoptotic lymphocytes was significantly increased in the graft as compared to that in the control group by flow cytometry. The ratio of apoptotic lymphocyte in the liver and graft was also significantly higher in the experimental group by TUNEL., Conclusion: FTY720 effectively induces the lymphocyte apoptosis, inhibits the lesion of target tissues by GVHD, and prolongs the recipient survival.

Background: Many patients with ischemic heart disease have cardiovascular risk factors such as cigarette smoking. We tested the effect of nicotine (a key component of cigarette smoking) on the therapeutic effects of human embryonic stem cell-derived endothelial cells (hESC-ECs)., Methods and Results: To induce endothelial cell differentiation, undifferentiated hESCs (H9 line) underwent 4-day floating EB formation and 8-day outgrowth differentiation in EGM-2 media. After 12 days, CD31(+) cells (13.7+/-2.5%) were sorted by FACScan and maintained in EGM-2 media for further differentiation. After isolation, these hESC-ECs expressed endothelial specific markers such as vWF (96.3+/-1.4%), CD31 (97.2+/-2.5%), and VE-cadherin (93.7+/-2.8%), form vascular-like channels, and incorporated DiI-labeled acetylated low-density lipoprotein (DiI-Ac-LDL). Afterward, 5x10(6) hESC-ECs treated for 24 hours with nicotine (10(-8) M) or PBS (as control) were injected into the hearts of mice undergoing LAD ligation followed by administration for two weeks of vehicle or nicotine (100 microg/ml) in the drinking water. Surprisingly, bioluminescence imaging (BLI) showed significant improvement in the survival of transplanted hESC-ECs in the nicotine treated group at 6 weeks. Postmortem analysis confirmed increased presence of small capillaries in the infarcted zones. Finally, in vitro mechanistic analysis suggests activation of the MAPK and Akt pathways following activation of nicotinic acetylcholine receptors (nAChRs)., Conclusions: This study shows for the first time that short-term systemic administrations of low dose nicotine can improve the survival of transplanted hESC-ECs, and enhance their angiogenic effects in vivo. Furthermore, activation of nAChRs has anti-apoptotic, angiogenic, and proliferative effects through MAPK and Akt signaling pathways.

To study the effects of recombinant human tumor necrosis factor-alpha (rhTNF-alpha) on HL-60 cells in vitro and in vivo, MTT and colony forming assay were used to examine the effects of rhTNF-alpha on proliferation of HL-60 cells; AO/EB (acridine orange-ethidium bromide) staining, Annexin-V flow cytometry analysis and TUNEL assay were used to detect apoptotic cells. The effect of rhTNF-alpha on xenograft growth of HL-60 cells was evaluated by tumor inhibition rate, histology, ultrastructure and TUNEL assay. The results showed that rhTNF-alpha inhibited the proliferation of HL-60 cells in a dose-dependent manner. Staining of cells with AO/EB revealed that rhTNF-alpha induced nuclear chromatin condensation and fragmentation. Positive Annexin V-FITC on cell membrane showed that rhTNF-alpha induced apoptosis of HL-60 cells in a dose-dependent manner. TUNEL assay showed that the apoptotic percentage of HL-60 cells reached 37.5% when incubated with 3200 U/ml rhTNF-alpha for 48 hours. In vivo rhTNF-alpha inhibited xenograft growth of HL-60 cells with the highest inhibition rate of 60.33%. Pathologically it was found that there were necrotic areas in the tumors of groups treated with rhTNF-alpha. There were more apoptotic cells in treatment groups than in that control group by transmission electron microscopy (TEM) and TUNEL assay. It is concluded that rhTNF-alpha is able to inhibit the proliferation of HL-60 cells and to induce apoptosis of HL-60 cells in vitro and in vivo.

Purpose: Signal transducer and activator of transcription 3 (Stat3) protein is persistently activated in breast cancer and promotes tumor cell survival. To gain a better understanding of the role of constitutive Stat3 signaling in breast cancer progression, we evaluated the expression profile of potential Stat3-regulated genes that may confer resistance to apoptosis., Experimental Design: Stat3 signaling was blocked with antisense oligonucleotides in human MDA-MB-435s breast cancer cells and Affymetrix GeneChip microarray analysis was done. The candidate Stat3 target gene Survivin was further evaluated in molecular assays using cultured breast cancer cells and immunohistochemistry of breast tumor specimens., Results: Survivin, a member of the inhibitor of apoptosis protein family, was identified as a potential Stat3-regulated gene by microarray analysis. This was confirmed in Survivin gene promoter studies and chromatin immunoprecipitation assays showing that Stat3 directly binds to and regulates the Survivin promoter. Furthermore, direct inhibition of Stat3 signaling blocked the expression of Survivin protein and induced apoptosis in breast cancer cells. Direct inhibition of Survivin expression also induced apoptosis. Increased Survivin protein expression correlates significantly (P = 0.001) with elevated Stat3 activity in primary breast tumor specimens from high-risk patients who were resistant to chemotherapy treatment., Conclusions: We identify Survivin as a direct downstream target gene of Stat3 in human breast cancer cells that is critical for their survival in culture. Our findings suggest that activated Stat3 signaling contributes to breast cancer progression and resistance to chemotherapy by, at least in part, inducing expression of the antiapoptotic protein, Survivin.

Interactions between the histone deacetylase (HDAC) inhibitors suberanoylanilide hydroxamic acid (SAHA) and sodium butyrate (SB) and the heat shock protein (Hsp) 90 antagonist 17-allylamino 17-demethoxygeldanamycin (17-AAG) have been examined in Bcr-Abl(+) human leukemia cells (K562 and LAMA84), including those sensitive and resistant to STI571 (imatinib mesylate). Cotreatment with 17-AAG and SAHA or SB synergistically induced mitochondrial dysfunction (cytochrome c and apoptosis-inducing factor release), caspase-3 and -8 activation, apoptosis, and growth inhibition. Similar effects were observed in LAMA84 cells and K562 cells resistant to STI571, as well as in CD34(+) cells isolated from the bone marrows of three patients with chronic myelogenous leukemia. These events were associated with increased binding of Bcr-Abl, Raf-1, and Akt to Hsp70, and inactivation of extracellular signal-regulated kinase 1/2 and Akt. In addition, 17-AAG/SAHA abrogated the DNA binding and the transcriptional activities of signal transducer and activator of transcription (STAT) 5 in K562 cells, including those ectopically expressing a constitutively active STAT5A construct. Cotreatment with 17-AAG and SAHA also induced down-regulation of Mcl-1, Bcl-xL, and B-Raf; up-regulation of Bak; cleavage of 14-3-3 proteins; and a profound conformational change in Bax accompanied by translocation to the membrane fraction. Moreover, ectopic expression of Bcl-2 attenuated cell death induced by this regimen, implicating mitochondrial injury in the lethality observed. Together, these findings raise the possibility that combining HDAC inhibitors with the Hsp90 antagonist 17-AAG may represent a novel strategy against Bcr-Abl(+) leukemias, including those resistant to STI571.

DBM (dibenzoylmethane) is a minor constituent of licorice that has antimutagenic activity. However, its other biological activities are not well-known. The structurally related beta-diketones hydroxydibenzoylmethane (HDB) and hydroxymethyldibenzoylmethane (HMDB) were able to induce apoptosis in colorectal carcinoma COLO 205 cells. Thus, the effect of structurally related beta-diketones on cell viability, DNA fragmentation, and caspase activity was assessed. The potency of these compounds on these features of apoptosis were in the order of HDB > HMDB > DBM in colorectal carcinoma COLO 205 cells. Here, we found that HDB-induced apoptotic cell death was accompanied by upregulation of cyclin D3, Bax, and p21 and down-regulation of Bcl-X(L), while HDB had no effect on the levels of Bcl-2 and Bad protein. These results indicate that HDB allows caspase-activated deoxyribonuclease to enter the nucleus and degrade chromosomal DNA and induces DFF-45 degradation. It is suggested that HDB-induced apoptosis is triggered by the release of cytochrome c into cytosol, procaspase-9 processing, activation of caspase-3 and caspase-2, degradation of PARP, and DNA fragmentation caused by the caspase-activated deoxyribonuclease through the digestion of DFF-45. The induction of apoptosis by HDB may provide a pivotal mechanism for its cancer chemopreventive action.

Objective: To study the effect of endogenous TGF-beta(1) and TNF-alpha on As(2)O(3) inducing apoptosis of HL-60 cells., Methods: The expressions of endogenous TGF-beta(1) and TNF-alpha in apoptotic HL-60 cells induced by As(2)O(3) were assayed by RT-PCR, quantitative RT-PCR, ELISA, DNA fragmentation and TUNEL. The effect of TGF-beta(1) and TNF-alpha antisense phosphorothioate oligodeoxynucleotides (PSODNs) on As(2)O(3) inducing apoptotic HL-60 cells was further studied., Results: (1) Expressions of endogenous TGF-beta(1) and TNF-alpha were significantly up-regulated in As(2)O(3) inducing apoptotic HL-60 cells (from 13,546 +/- 124 and 497,216 +/- 187 before treatment to 23,273 +/- 229 and 674,217 +/- 189 after treatment, respectively), accompanied with down-regulated bcl-2 mRNA expression (from 10,424 +/- 274 before treatment to 3,361 +/- 89 after treatment). (2) TGF-beta(1) and TNF-alpha antisense PSODNs could rescue As(2)O(3) induced apoptosis of HL-60 cells, with a restoration of bcl-2 gene expression., Conclusions: Endogenous TGF-beta(1) and TNF-alpha played an important role in As(2)O(3) inducing HL-60 cells apoptosis through down-regulation of bcl-2 expression.

The molecular mechanisms underlying the cell cycle growth-inhibitory and apoptotic effects of flavopiridol (FP) were determined in human breast cancer cells. Treatment with FP caused accumulation in the G(1) phase of the cell cycle and induced apoptosis of SKBR-3 and MB-468 cells. This was associated with down-regulation of the levels of cyclins D1 and B1, as well as with inhibition of cyclin-dependent kinase (cdk) 1, cdk2, and cdk4. FP-induced apoptosis was accompanied by a conformational change and mitochondrial localization of Bax. This resulted in the accumulations of cytochrome c, Smac, and Omi/HtrA2 in the cytosol and induced the poly(ADP-ribose) polymerase cleavage activity of caspase-3. Treatment with FP also attenuated the mRNA and protein levels of XIAP, cIAP-2, Mcl-1, Bcl-x(L), and survivin. In MB-468 cells with overexpression of Bcl-2 (468/Bcl-2), FP-induced Bax conformational change and apoptosis were inhibited, whereas the FP-mediated decline in the levels of IAP proteins, Mcl-11 and Bcl-x(L) remained unaltered. The effects of cotreatment with FP and the nontaxane tubulin-polymerizing agent epothilone (Epo) B were also determined in MB-468 cells. Sequential treatment with Epo B followed by FP induced significantly more apoptosis of MB-468 cells than treatment with the reverse sequence of FP followed by Epo B or treatment with either agent alone (P < 0.05). Treatment with Epo B followed by FP induced more Bax conformational change and was associated with a greater decline in the levels of XIAP, cIAP-2, Mcl-1, and Bcl-x(L). However, MB-468/Bcl-2 cells remained relatively resistant to Epo B followed by FP. Taken together, these findings suggest that the superior sequence-dependent anti-breast cancer activity of Epo B followed by FP may be due to FP-induced Bax conformational change and down-regulation of the antiapoptotic IAP, Bcl-x(L), and Mcl-1 proteins, but this treatment may not overcome the resistance to apoptosis of breast cancer cells conferred by overexpression of Bcl-2.

MEK kinase 1 (MEKK1) induces apoptosis through the activation of caspases. The mechanism for MEKK1-induced apoptosis involves caspase-mediated cleavage of MEKK1, releasing a pro-apoptotic 91 kDa kinase fragment that serves to further amplify caspase activation in a feedback loop. Both cleavage of MEKK1 and increased expression of death receptor 4 (DR4, TRAILR1) and death receptor 5 (DR5, TRAILR2) occur following exposure of cells to genotoxins. Overexpression of kinase inactive MEKK1 inhibits MEKK1-mediated apoptosis and effectively blocks death receptor upregulation following etoposide treatment. Herein, we investigate the role of death receptor activation and the ability of AKT/PKB (AKT) to inhibit cell death in MEKK1-induced apoptosis. We show that by preventing DR4 and DR5 activation through expression of decoy receptor 1 (DcR1) and dominant negative FADD, we inhibit MEKK1-induced apoptosis. Furthermore, expression of 91 kDa MEKK1 increased DR4 and FAS mRNA and protein levels. MEKK1-induced apoptosis is amplified by blocking PI-3 kinase activation and overexpression of AKT blocked both MEKK1-induced apoptosis and caspase activation. AKT overexpression also prevented the cleavage of endogenous MEKK1 by genotoxins. AKT did not, however, block MEKK1-induced JNK activation, showing that regulation of the JNK pathway by MEKK1 is independent of its role in regulation of apoptosis. Thus, MEKK1-induced apoptosis requires TRAIL death receptor activation and is blocked by AKT through inhibition of MEKK1 cleavage.

Levan is an exopolysaccharide produced by Bacillus licheniformis (strain FRI MY-55) that shows promising pharmacological activity. Phosphorylation is a chemical modification that can increase the biological and antioxidant properties of levan. In this study, levan was phosphorylated by microwave-assisted synthesis to achieve a degree of substitution of 0.29. The hydroxyl radical scavenging activity of microwave-assisted phosphorylated levan (microwave P) increased significantly (6-fold) over native levan; this activity was only slightly lower than vitamin C. Other free radical scavenging and reducing power tests revealed that Microwave P activity was increased by 30–40%. Microwave P inhibited the proliferation of HCT-116 and A549 cancer cell lines more readily than native levan with an IC 50 of 1.03 mg/mL and 1.38 mg/mL for HCT-116 and A549 cells, respectively. Cells treated with native levan and its derivatives remained in the sub-G1 phase according to cell cycle analysis, whereas Microwave P treatment increased the proportion of cells undergoing apoptosis. Furthermore, Microwave P effectively upregulated pro-apoptosis marker Bax and downregulated anti-apoptosis marker Bcl-2, in addition to inducing the expression of caspase-9 and caspase-3. These findings show that levan phosphorylated via microwave-assisted synthesis showed increased antioxidant and antitumor activity over native levan or levan phosphorylated via traditional long-term heating. In particular, Microwave P possesses antiproliferative activity and can induce apoptosis through mitochondrial pathways in cancerous cells. [ABSTRACT FROM AUTHOR]

Epigallocatechin-3-gallate (EGCG), the major constituent of green tea, has been shown to induce cell death in cancer cells. Primary effusion lymphoma (PEL) is an aggressive neoplasm caused by human herpesvirus 8 (HHV8). In this study, we examined the role of EGCG on PEL cells in cell death and HHV8 replication. We performed trypan blue exclusion assay to assess the cell viability of PEL cells, flow cytometry analysis to examine the cell cycle distribution and reactive oxygen species (ROS) generation, caspase-3 activity to assay apoptosis, acridine orange staining to determine autophagy, and immunoblotting to detect the protein levels involved in apoptosis and autophagy as well as mitogen activated protein kinases (MAPKs) activation upon EGCG treatment. The expression of the HHV8 lytic gene was determined by luciferase reporter assay and reverse transcription-PCR, and viral progeny production was determined by PCR. Results revealed that EGCG induced cell death and ROS generation in PEL cells in a dose-dependent manner. N-acetylcysteine (NAC) inhibited the EGCG-induced ROS and rescued the cell from EGCG-induced cell death. Even though EGCG induced ROS generation in PEL cells, it reduced the production of progeny virus from PEL cells without causing HHV8 reactivation. These results suggest that EGCG may represent a novel strategy for the treatment of HHV8 infection and HHV8-associated lymphomas. [ABSTRACT FROM AUTHOR]

Coronavirus replication is structurally and functionally associated with the endoplasmic reticulum (ER), a major site of protein synthesis, folding, modification and sorting in the eukaryotic cells. Disturbance of ER homeostasis may occur under various physiological or pathological conditions. In response to the ER stress, signaling pathways of the unfolded protein response (UPR) are activated. UPR is mediated by three ER transmembrane sensors, namely the PKR-like ER protein kinase (PERK), the inositol-requiring protein 1 (IRE1) and the activating transcriptional factor 6 (ATF6). UPR facilitates adaptation to ER stress by reversible translation attenuation, enhancement of ER protein folding capacity and activation of ER-associated degradation (ERAD). In cells under prolonged and irremediable ER stress, UPR can also trigger apoptotic cell death. Accumulating evidence has shown that coronavirus infection causes ER stress and induces UPR in the infected cells. UPR is closely associated with a number of major signaling pathways, including autophagy, apoptosis, the mitogen-activated protein (MAP) kinase pathways, innate immunity and pro-inflammatory response. Therefore, studies on the UPR are pivotal in elucidating the complicated issue of coronavirus-host interaction. In this paper, we present the up-to-date knowledge on coronavirus-induced UPR and discuss its potential involvement in regulation of innate immunity and apoptosis. [ABSTRACT FROM AUTHOR]

Abstract: Lenalidomide, an analog of thalidomide, modified responses of stimulated T cells from healthy young (ages 21–40years) and old (≥age 65years) subjects. At 0.03μM to 1μM, lenalidomide enhanced generation of IL-2 and IFN-γ by T cell receptor-stimulated T cells of young subjects up to respective maximum increases of 17-fold and three-fold, but at 0.3μM and 1μM suppressed IL-17 generation. The same concentrations of lenalidomide enhanced IL-2 and IFN-γ generation by stimulated T cells of old subjects more, with greater respective maximal increases of up to 120-fold and six-fold, without suppressing IL-17 generation. Lenalidomide enhanced proliferation and suppressed apoptosis of stimulated T cells from old subjects, by IL-2-dependent mechanisms, and restored diminished T cell chemotactic responses to CCL21 and sphingosine 1-phosphate. The reversal of T cell abnormalities of immunosenescence by low concentrations of lenalidomide suggest a potential for improvement of immunity in the elderly. [Copyright &y& Elsevier]

Aspirin eugenol ester (AEE) is a new potential pharmaceutical compound possessing anti-inflammatory, anti-cardiovascular disease, and antioxidative stress activity. The pharmacological activities of AEE are partly dependent on its regulation of cell apoptosis. However, it is still unclear how AEE inhibits cell apoptosis on the basis of its antioxidative stress effect. This study aimed to reveal the vascular antioxidative mechanism of AEE in response to H2O2-induced oxidative stress in HUVECs and paraquat-induced oxidative stress in rats. In the different intervention groups of HUVECs and rats, the expression of ASK1, ERK1/2, SAPK/JNK, and p38 and the phosphorylation levels of ERK1/2, SAPK/JNK, and p38 were measured. The effects of ASK1 and ERK1/2 on the anti-apoptotic activity of AEE in the oxidative stress model were probed using the corresponding inhibitors ASK1 and ERK1/2. The results showed that in the HUVECs, 200 μM H2O2 treatment significantly increased the phosphorylation of SAPK/JNK and the level of ASK1 but decreased the phosphorylation of ERK1/2, while in the HUVECs pretreated with AEE, the H2O2-induced changes were significantly ameliorated. The findings were observed in vitro and in vivo. Moreover, inhibition of ASK1 and ERK1/2 showed that ASK1 plays a vital role in the protective effect of AEE on H2O2-induced apoptosis. All findings suggested that AEE protects the vascular endothelium from oxidative injury by mediating the ASK1 pathway. [ABSTRACT FROM AUTHOR]

Ethnopharmacological relevance Gualou Guizhi decoction (GLGZD) prescribed in traditional Chinese medicine has been reported to have protective effects on ischemic stroke. The present study is to investigate the therapeutic effect of GLGZD on ischemic stroke and explore its mode of action. Materials and methods GLGZD was studied on transient middle cerebral artery occlusion (MCAO) followed by reperfusion in viv o, as well as on hippocampal primary neuron c ultures in vitro . Results In viv o, it was shown that GLGZD treatment for 7 days could ameliorate transient middle cerebral artery occlusion (MCAO)-induced neurological deficit, histopathology changes and decrease infarct area. Further study demonstrated that GLGZD inhibited over-activation of astrocytes and apoptosis of neurons and GLGZD promoted up-regulation of neuronal specific marker neuron-specific nuclear (NeuN) and microtubule-associated protein 2 (MAP-2) in brain. Moreover, the in vitro study revealed that GLGZD treatment protected against NMDA-induced cell apoptosis and neuronal loss, and promoted up-regulation of neuronal specific marker NeuN. Conclusions Taken together, the present study demonstrates that GLGZD produces a protection in the MCAO model rats via inhibiting over-activation of astrocytes, apoptosis of neurons and up-regulation of neuronal specific marker NeuN and MAP-2. Our study reveals that GLGZD might be a potential neuroprotective agent for stroke and can provide basic data for clinical use. [ABSTRACT FROM AUTHOR]

Multiple myeloma (MM) is one of the most common tumors of the hematological system and remains incurable. Recent studies have shown that long noncoding RNA NORAD is a potential oncogene in a variety of tumors. However, the general biological role and clinical value of NORAD in MM remains unknown. In this study, we measured NORAD expression in bone marrow of 60 newly diagnosed MM, 30 post treatment MM and 17 healthy donors by real-time quantitative polymerase chain reaction (qPCR). The NORAD gene was knockdown by lentiviral transfection in MM cell lines, and the effects of NORAD on apoptosis, cell cycle and cell proliferation in MM cells were examined by flow cytometry, CCK8 assay, EDU assay and Western blot, and the differential genes after knockdown of NORAD were screened by mRNA sequencing, followed by in vivo experiments and immunohistochemical assays. We found that knockdown of NORAD promoted MM cell apoptosis, induced cell cycle G1 phase arrest, and inhibited MM cell apoptosis in in vivo and in vitro experiments. Mechanistically, NORAD plays these roles through the BMP6/P-ERK1/2 axis. We discuss a novel mechanism by which NORAD acts pro-tumorigenically in MM via the BMP6/P-ERK1/2 axis. [ABSTRACT FROM AUTHOR]