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Wogonin, a vital bioactive compound extracted from the medicinal plant, Scutellaria baicalensis, has been wildly used for its potential in mitigating the progression of chronic diseases. Chronic kidney disease (CKD) represents a significant global health challenge due to its high prevalence, morbidity and mortality rates, and associated complications. This study aimed to assess the potential of wogonin in attenuating renal fibrosis and to elucidate the underlying molecular mechanisms using a unilateral ureteral obstruction (UUO) mouse model as a CKD mimic. Male mice, 8 weeks old, underwent orally administrated of either 50 mg/kg/day of wogonin or positive control of 5 mg/kg/day candesartan following UUO surgery. NRK52E cells were exposed to tumor growth factors-beta (TGF-β) to evaluate the anti-fibrotic effects of wogonin. The results demonstrated that wogonin treatment effectively attenuated TGF-β-induced fibrosis markers in NRK-52E cells. Additionally, administration of wogonin significantly improved histopathological alterations and downregulated the expression of pro-fibrotic factors (Fibronectin, α-smooth muscle actin, Collagen IV, E-cadherin, and TGF-β), oxidative stress markers (Catalase, superoxide dismutase 2, NADPH oxidase 4, and thioredoxin reductase 1), inflammatory molecules (Cyclooxygenase-2 and TNF-α), and the infiltration of neutrophils and macrophages in UUO mice. Furthermore, wogonin treatment mitigated endoplasmic reticulum (ER) stress-associated molecular markers (GRP78, GRP94, ATF4, CHOP, and the caspase cascade) and suppressed apoptosis. The findings indicate that wogonin treatment ameliorates key fibrotic aspects of CKD by attenuating ER stress-related apoptosis, inflammation, and oxidative stress, suggesting its potential as a future therapeutic target., 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.)

Objective: The upregulation of long noncoding RNA intersectin 1-2 (lnc ITSN1-2) is associated with poor prognosis in acute ischemic stroke (AIS) patients, but the role and mechanism of lnc ITSN1-2 in AIS are rarely reported, which, thus, are highlighted in this study., Methods: AIS cell model was constructed by oxygen glucose deprivation and reoxygenation (OGD/R). The quantitative real-time PCR was used to detect the expression of lnc ITSN1-2 in HT22 cells. The effects of lnc ITSN1-2 overexpression or knockdown on viability, LDH release, apoptosis, inflammatory and apoptotic factor expressions in OGD/R-induced HT22 cells were measured by cell counting kit-8 assay, LDH release kit, flow cytometry, ELISA and western blot, respectively. Starbase was used to screen the target genes of lnc ITSN1-2. The targeting relationship between lnc ITSN1-2 and miR-195-5p was predicted by starbase and verified by dual-luciferase report assay. The above assays were conducted again to study the function of miR-195-5p. Lastly, the levels of activated mitogen-activated protein kinases (MAPK) pathway-related proteins were determined by western blot., Results: OGD/R treatment reduced the HT22 cell viability and enhanced LDH release rate and lnc ITSN1-2 expression. Lnc ITSN1-2 overexpression promoted the cell injury, apoptosis and inflammation in OGD/R-induced HT22 cells, while lnc ITSN1-2 knockdown generated the opposite effect and deactivated the MAPK pathways. However, the effect of lnc ITSN1-2 knockdown in OGD/R-induced HT22 cells was reversed by miR-195-5p inhibitor., Conclusion: Lnc ITSN1-2 knockdown suppressed the inflammation and apoptosis in OGD/R-induced HT22 cells by regulating the miR-195-5p-mediated MAPK pathways., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

Inorganic arsenic (As 3+ ), a well-known worldwide industrial and environmental pollutant, has been linked to neurodegenerative disorders (NDs). Autophagy plays an important role in controlling neuronal cell survival/death. However, limited information is available regarding the toxicological mechanism at the interplay between autophagy and As 3+ -induced neurotoxicity. The present study found that As 3+ exposure induced a concomitant activation of apoptosis and autophagy in Neuro-2a cells, which was accompanied with the increase of phosphatidylserine exposure on outer membrane leaflets and apoptotic cell population, and the activation of caspase-3, -7, and PARP as well as the elevation of protein expressions of LC3-II, Atg-5, and Beclin-1, and the accumulation of autophagosome. Pretreatment of cells with autophagy inhibitor 3-MA, but not that of Z-VAD-FMK (a pan-caspase inhibitor), effectively prevented the As 3+ -induced autophagic and apoptotic responses, indicating that As 3+ -triggered autophagy was contributing to neuronal cell apoptosis. Furthermore, As 3+ exposure evoked the dephosphorylation of Akt. Pretreatment with SC79, an Akt activator, could significantly attenuated As 3+ -induced Akt inactivation as well as autophagic and apoptotic events. Expectedly, inhibition of Akt signaling with LY294002 obviously enhanced As 3+ -triggered autophagy and apoptosis. Exposure to As 3+ also dramatically increased the phosphorylation level of AMPKα. Pretreatment of AMPK inhibitor (Compound C) could markedly abrogate the As 3+ -induced phosphorylated AMPKα expression, and autophagy and apoptosis activation. Taken together, these results indicated that As 3+ exerted its cytotoxicity in neuronal cells via the Akt inactivation/AMPK activation downstream-regulated autophagy-dependent apoptosis pathways, which ultimately lead to cell death. Our findings suggest that the regulation of Akt/AMPK signals may be a promising intervention to against As 3+ -induced neurotoxicity and NDs., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Background: Triple-negative breast cancer (TNBC) has a more aggressive phenotype and poorer prognosis than hormone receptor (HR+) and human epidermal growth factor receptor (HER2 -) subtypes. Inhibition of cyclin-dependent kinase (CDK)4 and CDK6 was successful in patients with advanced metastatic HR+/HER2- breast cancer, but those with TNBC exhibited low or no response to this therapeutic approach. This study investigated the dual therapeutic targeting of CDK2 and CDK4 by using 4-acetyl-antroquinonol B (4-AAQB) against TNBC cells., Methods: We examined the effects of CDK2, CDK4, and CDK6 inhibition through 4-AAQB treatment on TNBC cell lines and established an orthotropic xenograft mouse model to confirm the in vitro results of inhibiting CDK2, CDK4, and CDK6 by 4-AAQB treatment., Results: High expression and alteration of CDK2 and CDK4 but not CDK6 significantly correlated with poor overall survival of patients with breast cancer. CDK2 and CDK4 were positively correlated with damage in DNA replication and repair pathways. Docking results indicated that 4-AAQB was bound to CDK2 and CDK4 with high affinity. Treatment of TNBC cells with 4-AAQB suppressed the expression of CDK2 and CDK4 in vitro. Additionally, 4-AAQB induced cell cycle arrest, DNA damage, and apoptosis in TNBC cells. In vivo study results confirmed that the anticancer activity of 4-AAQB suppressed tumor growth through the inhibition of CDK2 and CDK4., Conclusion: The expression level of CDK2 and CDK4 and DNA damage response (DDR) signaling are prominent in TNBC cell cycle regulation. Thus, 4-AAQB is a potential agent for targeting CDK2/4 and DDR in TNBC cells., (Copyright © 2021. Published by Elsevier Inc.)

Bisphenol A (BPA) is recognized as a harmful pollutant in the worldwide. Growing studies have reported that BPA can cause adverse effects and diseases in human, and link to a potential risk factor for development of neurodegenerative diseases (NDs). 4-methyl-2,4-bis(4-hydroxyphenyl)pent-1-ene (MBP), which generated in the mammalian liver after BPA exposure, is a major active metabolite of BPA. MBP has been suggested to exert greater toxicity than BPA. However, the molecular mechanism of MBP on the neuronal cytotoxicity remains unclear. In this study, MBP exposure significantly reduced Neuro-2a cell viability and induced apoptotic events that MBP (5-15 μM) exhibited greater neuronal cytotoxicity than BPA (50-100 μM). The mitochondria-dependent apoptotic signals including the decrease in mitochondrial membrane potential (MMP) and the increase in cytosolic apoptosis-induced factor (AIF), cytochrome c release, and Bax protein expression were involved in MBP (10 μM)-induced Neuro-2a cell death. Exposure of Neuro-2a cells to MBP (10 μM) also triggered endoplasmic reticulum (ER) stress through the induction of several key molecules including glucose-regulated protein (GRP)78, C/EBP homologous protein (CHOP), X-box binding protein (XBP)-1, protein kinase R-like ER kinase (PERK), eukaryotic initiation factor 2α (eIF2α), inositol-requiring enzyme(IRE)-1, activation transcription factor(AFT)4 and ATF6, and caspase-12. Pretreatment with 4-PBA (an ER stress inhibitor) and specific siRNAs for GRP78, CHOP, and XBP-1 significantly suppressed the expression of these ER stress-related proteins and the activation of caspase-12/-3/-7 in MBP-exposed Neuro-2a cells. Furthermore, MBP (10 μM) exposure dramatically increased the activation of extracellular regulated protein (ERK)1/2 and decreased Akt phosphorylation. Pretreatment with PD98059 (an ERK1/2 inhibitor) and transfection with the overexpression of activation of Akt1 (myr-Akt1) effectively suppressed MBP-induced apoptotic and ER stress-related signals. Collectively, these results demonstrate that MBP exposure exerts neuronal cytotoxicity via the interplay of ERK activation and Akt inactivation-regulated mitochondria-dependent and ER stress-triggered apoptotic pathway, which ultimately leads to neuronal cell death., (Copyright © 2021 Elsevier B.V. All rights reserved.)

4-methyl-2,4-bis(4-hydroxyphenyl)pent-1-ene (MBP), a major active metabolite of bisphenol A (BPA), is generated in the mammalian liver. Some studies have suggested that MBP exerts greater toxicity than BPA. However, the mechanism underlying MBP-induced pancreatic β-cell cytotoxicity remains largely unclear. This study demonstrated the cytotoxicity of MBP in pancreatic β-cells and elucidated the cellular mechanism involved in MBP-induced β-cell death. Our results showed that MBP exposure significantly reduced cell viability, caused insulin secretion dysfunction, and induced apoptotic events including increased caspase-3 activity and the expression of active forms of caspase-3/-7/-9 and PARP protein. In addition, MBP triggered endoplasmic reticulum (ER) stress, as indicated by the upregulation of GRP 78, CHOP, and cleaved caspase-12 proteins. Pretreatment with 4-phenylbutyric acid (4-PBA; a pharmacological inhibitor of ER stress) markedly reversed MBP-induced ER stress and apoptosis-related signals. Furthermore, exposure to MBP significantly induced the protein phosphorylation of JNK and AMP-activated protein kinase (AMPK)α. Pretreatment of β-cells with pharmacological inhibitors for JNK (SP600125) and AMPK (compound C), respectively, effectively abrogated the MBP-induced apoptosis-related signals. Both JNK and AMPK inhibitors also suppressed the MBP-induced activation of JNK and AMPKα and of each other. In conclusion, these findings suggest that MBP exposure exerts cytotoxicity on β-cells via the interdependent activation of JNK and AMPKα, which regulates the downstream apoptotic signaling pathway.

Galectin-3 (Gal-3) is a 26-kDa lectin that regulates many aspects of inflammatory cell behavior. We assessed the hypothesis that increased levels of Gal-3 contribute to abdominal aortic aneurysm (AAA) progression by enhancing monocyte chemoattraction through macrophage activation. We analyzed the plasma levels of Gal-3 in 76 patients with AAA (AAA group) and 97 controls (CTL group) as well as in angiotensin II (Ang-II)-infused ApoE knockout mice. Additionally, conditioned media (CM) were used to polarize THP-1 monocyte to M1 macrophages with or without Gal-3 inhibition through small interfering RNA targeted deletion to investigate whether Gal-3 inhibition could attenuate macrophage-induced inflammation and smooth muscle cell (SMC) apoptosis. Our results showed a markedly increased expression of Gal-3 in the plasma and aorta in the AAA patients and experimental mice compared with the CTL group. An in vitro study demonstrated that the M1 cells exhibited increased Gal-3 expression. Gal-3 inhibition markedly decreased the quantity of macrophage-induced inflammatory regulators, including IL-8, TNF-α, and IL-1β, as well as messenger RNA expression and MMP-9 activity. Moreover, Gal-3-deficient CM weakened SMC apoptosis through Fas activation. These findings prove that Gal-3 may contribute to AAA progression by the activation of inflammatory macrophages, thereby promoting SMC apoptosis.

Cadmium (Cd) is known to be ranked the 7 th hazardous substance in the Substance Priority List by Agency for Toxic Substances and Disease Registry. The experimental and epidemiological data have suggested that Cd is linked to the development of diabetes mellitus (DM). The molecular mechanism of Cd on the pancreatic β-cell cytotoxicity still remains unclear. Evidence has pointed toward that Ca 2+ is an important regulator of toxic insult-induced β-cell cytotoxicity. The role of Ca 2+ in the Cd-induced β-cell cytotoxicity is still unknown. In this study, we found that Cd exposure significantly inhibited insulin secretion and cell viability in the pancreatic β-cell-derived RIN-m5F cells. Cd exposure induced apoptotic events, including the increased populations of apoptotic cells and sub-G1 hypodiploid cells, and caspase-3/-7/-9 and poly (ADP-ribose) polymerase (PARP) activation, which largely depended on the activation of c-Jun N-terminal kinase (JNK) and C/EBP homologous protein (CHOP). Transfection with siRNAs for JNK and CHOP or pretreatment with specific pharmacological inhibitor of JNK (SP600125) in β-cells effectively prevented the Cd-induced insulin secretion dysfunction and apoptosis. JNK-specific siRNA dramatically suppressed Cd-induced JNK phosphorylation and CHOP protein expression, but JNK phosphorylation could not be inhibited by CHOP-specific siRNA. Furthermore, Cd exposure significantly increased the intracellular calcium ([Ca 2+ ] i ) levels. Buffering the Ca 2+ response with BAPTA/AM effectively abrogated the Cd-induced [Ca 2+ ] i elevation, insulin secretion dysfunction, apoptosis, and protein expression of JNK phosphorylation and CHOP activation. Taken together, these findings demonstrated that Cd exposure exerts β-cell death via a [Ca 2+ ] i -dependent JNK activation-activated downstream CHOP-related apoptotic signaling pathway., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Most of the current cancer therapies may induce serious side effects and affect patient quality of life. Recently, a novel treatment using an alternating low-intensity and intermediate-frequency electric field was proposed and found to be a noninvasive and minimally toxic approach. However, additional fundamental studies and scientific evidence are required to further support the development of this treatment into a standard cancer therapy. In the current work, an in-house fabricated culture plate was developed to study the responses of hepatocellular carcinoma spheroids to treatment with an alternating electric field. From the results of the viability study, the electric field was confirmed to influence the dividing cells in the spheroids. Fluorescent staining of live and dead cells revealed that a fraction of the cells were damaged in the field-treated spheroids. Moreover, flow cytometry analyses were conducted and showed that a fraction of the cells in the spheroids underwent apoptosis and cell cycle arrest. Additionally, the apoptosis pathway (Bax/caspase) and cell cycle arrest pathway (p53/p21) were found to be activated after exposure to the electric field. In summary, the results further elucidated the cellular and molecular mechanism inducing apoptosis and cell cycle arrest in the field-treated hepatocellular carcinoma spheroids. This study provides more evidence to support the efficacy of electric-field-based cancer therapy., (© 2019 American Institute of Chemical Engineers.)

Tangeretin is one of the most abundant compounds in citrus peel, and studies have shown that it possesses anti-oxidant and anti-cancer properties. However, no study has been conducted on bladder cancer cells. Bladder cancer has the second highest mortality rate among urological cancers and is the fifth most common malignancy in the world. Currently, combination chemotherapy is the most common approach by which to treat patients with bladder cancer, and thus identifying more effective chemotherapeutic agents that can be safely administered to patients is a very important research issue. Therefore, this study investigated whether tangeretin can induce apoptosis and identified the signaling pathways of tangeretin-induced apoptosis in human bladder cancer cells using two-dimensional gel electrophoresis (2DGE). The results of the study demonstrated that 60 μM tangeretin reduced the cell survival of a BFTC-905 bladder carcinoma cell line by 42%, and induced early and late apoptosis in the cells. In this study 2DGE proteomics technology identified 41 proteins that were differentially-expressed in tangeretin-treated cells, and subsequently LC⁻MS/MS analysis was performed to identify the proteins. Based on the functions of the differentially-expressed proteins, the results suggested that tangeretin caused mitochondrial dysfunction and further induced apoptosis in bladder cancer cells. Moreover, western blotting analysis demonstrated that tangeretin treatment disturbed calcium homeostasis in the mitochondria, triggered cytochrome C release, and activated caspase-3 and caspase-9, which led to apoptosis. In conclusion, our results showed that tangeretin-induced apoptosis in human bladder cancer cells is mediated by mitochondrial inactivation, suggesting that tangeretin has the potential to be developed as a new drug for the treatment of bladder cancer.

Macrophages are the primary host target cells of Mycobacterium tuberculosis (M. tb). As a subunit of immunoregulatory cytokines IL-27 and IL-35, Epstein-Barr virus-induced gene 3 (EBI3) has typically been explored as the secreted form and assessed in terms of its effects triggered by extracellular EBI3. However, little is known about intracellular EBI3 function. In the current study, we report that EBI3 production by macrophages is elevated in TB patients. We further demonstrate that increased EBI3 accumulates in virulent M. tb-treated murine macrophages. Eukaryotic translation elongation factor 1-alpha 1 (eEF1A1) binds to intracellular EBI3 to reduce Lys48 (K48)-linked ubiquitination of EBI3, leading to EBI3 accumulation. Moreover, the intracellular EBI3 inhibits caspase-3-mediated apoptosis in M. tb-treated macrophages. Herein, we propose a novel mechanism for accumulating intracellular EBI3 and its regulation of macrophage apoptosis in response to virulent M. tb., (© FEMS 2019.)

Although it is well established that TNFα contributes to hepatitis, liver failure and associated hepatocarcinogenesis via the regulation of inflammation, its pro-apoptotic role in the liver has remained enigmatic. On its own, TNFα is unable to trigger apoptosis. However, when combined with the transcriptional inhibitor GaLN, it can cause hepatocyte apoptosis and liver failure in mice. Moreover, along with others, we have shown that TNFα is capable of sensitizing cells to FasL- or drug-induced cell death via c-Jun N-terminal kinase (JNK) activation and phosphorylation/activation of the BH3-only protein Bim. In this context, TNFα could exacerbate hepatocyte cell death during simultaneous inflammatory and T-cell-mediated immune responses in the liver. Here we show that TNFα sensitizes primary hepatocytes, established hepatocyte cell lines and mouse embryo fibroblasts to FasL-induced apoptosis by the transcriptional induction and higher surface expression of Fas via the NFκB pathway. Genetic deletion, diminished expression or dominant-negative inhibition of the NFκB subunit p65 resulted in lower Fas expression and inhibited TNFα-induced Fas upregulation and sensitization to FasL-induced cell death. By hydrodynamic injection of p65 shRNA into the tail vein of mice, we confirm that Fas upregulation by TNFα is also NFκB-mediated in the liver. In conclusion, TNFα sensitization of FasL-induced apoptosis in the liver proceeds via two parallel signaling pathways, activation of JNK and Bim phosphorylation and NFκB-mediated Fas upregulation.

Diabetic retinopathy (DR), one of the most common complications of late‑phase diabetes, is associated with the ectopic apoptosis of microvascular cells. Gastrodin, a phenolic glucoside derived from Gastrodia elata Blume, has been reported to have antioxidant and anti‑inflammation activities. The aim of the present study was to investigate the effects of gastrodin on high glucose (HG)‑induced human retinal endothelial cell (HREC) injury and its underlying mechanism. The results demonstrated that HG induced cell apoptosis in HRECs, which was accompanied by increased levels of reactive oxygen species production. Gastrodin treatment significantly alleviated HG‑induced apoptosis and oxidative stress. Furthermore, HG stimulation decreased the levels of SIRT1, which was accompanied by an increase in Toll‑like receptor 4 (TLR4) expression and the levels of phosphorylated nuclear factor (NF)‑κBp65. However, the administration of gastrodin significantly inhibited the activation of the sirtuin 1 (SIRT1)/TLR4/NF‑κBp65 signaling pathway in HRECs exposed to HG. Collectively, the present study demonstrated that gastrodin may be effective against HG‑induced apoptosis and its action may be exerted through the regulation of the SIRT1/TLR4/NF‑κBp65 signaling pathway.

Tributyltin (TBT), an endocrine disrupting chemical, can be found in food (particular in fish and seafood) and drinking water by contamination. Here, we elucidated the effects and possible mechanisms of low-dose TBT on the growth and function of pancreatic β-cells and glucose metabolism in mice. Submicromolar-concentration of TBT significantly induced β-cell cytotoxicity and apoptosis, which were accompanied by poly (ADP-ribose) polymerase cleavage and mitogen-activated protein kinases-JNK and ERK1/2 phosphorylation. TBT could also suppress the glucose-stimulated insulin secretion in β-cells and isolated mouse islets. TBT increased reactive oxygen species production. TBT-induced β-cell cytotoxicity and apoptosis were significantly prevented by antioxidant N-acetylcysteine (NAC) and JNK inhibitor SP600125, but not ERK1/2 inhibitor PD98059 and p38 inhibitor SB203580. Both NAC and SP600125 inhibited JNK phosphorylation and reduced cell viability in TBT-treated β-cells. Four-week exposure of TBT (0.25 mg/kg) to mice revealed the decreased plasma insulin, increased blood glucose and plasma malondialdehyde, suppressed islet insulin secretion, and increased islet caspase-3 activity, which could be reversed by NAC treatment. After removing the TBT exposure for 2 weeks, the TBT-induced glucose metabolism alteration was significantly reversed. These results suggest that low-dose TBT can induce β-cell apoptosis and interfere with glucose homeostasis via an oxidative stress-related pathway.

In this study, a compressional-puffing process (CPP) was used to pretreat Sargassum hemiphyllum (SH) and then fucoidan was extracted from SH by hot water. Three fucoidan extracts, namely SH1 (puffing at 0 kg/cm²); SH2 (puffing at 1.7 kg/cm²); and SH3 (puffing at 10.0 kg/cm²) were obtained, and their compositions and biological activities were evaluated. The results indicate that CPP increased the extraction yield, total sugar content, and molar ratios of sulfate/fucose of fucoidan and decreased molecular weight and impurities of fucoidan. The SH1-SH3 extracts exhibited characteristics of fucoidan as demonstrated by the analyses of composition, FTIR spectroscopy, NMR spectroscopy, and molecular weight. All SH1-SH3 extracts showed antioxidant activities. The SH1-SH3 extracts protected SH-SY5Y cells from 6-hydroxydopamine (6-OHDA)-induced apoptosis as illustrated by cell cycle distribution, cytochrome c release, activation of caspase-8, -9, and -3, and DNA fragmentation analyses. Additional experiments revealed that phosphorylation of Akt is involved in the opposing effects of SH1-SH3 on 6-OHDA-induced neurotoxicity. SH3 exhibited a relatively high extraction yield, the lowest levels of impurities, and was the most effective at reversing the 6-OHDA-induced neurotoxicity of SH-SY5Y cells among SH1-SH3, which taken together indicate that it may have potential as a candidate therapeutic agent for the preventive therapy of neurodegenerative diseases., Competing Interests: The authors declare no conflict of interest.

Background and Objective: Amarogentin has been reported to have a preventive effect on liver cancer via inducing cancer cell apoptosis. We attempted to elucidate the roles of p53-associated apoptosis pathways in the chemopreventive mechanism of amarogentin. The findings of this study will facilitate the development of a novel supplementary strategy for the treatment of liver cancer., Materials and Methods: The purity of amarogentin was assessed by high-performance liquid chromatography. The inhibitory ratios of the liver cell lines were determined using a Cell Counting Kit-8 following treatment with a gradient concentration of amarogentin. Cell apoptosis was detected by flow cytometry using annexin V-fluorescein isothiocyanate/propidium iodide kits. The gene and protein expression of p53-associated molecules, such as Akt, human telomerase reverse transcriptase, RelA, and p38, was detected by real-time quantitative polymerase chain reaction, Western blotting, and immunohistochemical staining in liver cancer cells and mouse tumor tissues after treatment with amarogentin., Results: The inhibitory effect of amarogentin on cell proliferation was more obvious in liver cancer cells, and amarogentin was more likely to induce the apoptosis of liver cancer cells than that of normal liver cells. The gene and protein expression levels of Akt, RelA, and human telomerase reverse transcriptase were markedly higher in the control group than in the preventive group and treatment groups. Only the expression of human telomerase reverse transcriptase was downregulated, accompanied by the upregulation of p53., Conclusion: The results of our study suggest that amarogentin promotes apoptosis of liver cancer cells by the upregulation of p53 and downregulation of human telomerase reverse transcriptase and prevents the malignant transformation of these cells.

Triple negative breast cancer (TNBC) is an aggressive cancer for which prognosis remains poor. Combination therapy is a promising strategy for enhancing treatment efficacy. Blockade of STAT3 signaling may enhance the response of cancer cells to conventional chemotherapeutic agents. Here we used a SHP-1 agonist SC-43 to dephosphorylate STAT3 thereby suppressing oncogenic STAT3 signaling and tested it in combination with docetaxel in TNBC cells. We first analyzed messenger RNA (mRNA) expression of SHP-1 gene (PTPN6) in a public TNBC dataset (TCGA) and found that higher SHP-1 mRNA expression is associated with better overall survival in TNBC patients. Sequential combination of docetaxel and SC-43 in vitro showed enhanced anti-proliferation and apoptosis associated with decreased p-STAT3 and decreased STAT3-downstream effector cyclin D1 in the TNBC cell lines MDA-MB-231, MDA-MB-468, and HCC-1937. Ectopic expression of STAT3 reduced the increased cytotoxicity induced by the combination therapy. In addition, this sequential combination showed enhanced SHP-1 activity compared to SC-43 alone. Furthermore, the combination treatment-induced apoptosis was attenuated by small interfering RNA (siRNA) against SHP-1 or by ectopic expression of SHP-1 mutants that caused SC-43 to lose its SHP-1 agonist capability. Moreover, combination of docetaxel and SC-43 showed enhanced tumor growth inhibition compared to single-agent therapy in mice bearing MDA-MB-231 tumor xenografts. Our results suggest that the novel SHP-1 agonist SC-43 enhanced docetaxel-induced cytotoxicity by SHP-1 dependent STAT3 inhibition in human triple negative breast cancer cells. TNBC patients with high SHP-1 expressions show better survival. Docetaxel combined with SC-43 enhances cell apoptosis and reduces p-STAT3. SHP-1 inhibition reduces the enhanced effect of docetaxel-SC-43 combination. Docetaxel-SC-43 combination suppresses xenograft tumor growth and reduces p-STAT3., Key Messages: TNBC patients with high SHP-1 expressions show better survival. Docetaxel combined with SC-43 enhances cell apoptosis and reduces p-STAT3. SHP-1 inhibition reduces the enhanced effect of docetaxel-SC-43 combination. Docetaxel-SC-43 combination suppresses xenograft tumor growth and reduces p-STAT3.

Triple-negative breast cancer (TNBC) remains difficult to treat and urgently needs new therapeutic options. Nintedanib, a multikinase inhibitor, has exhibited efficacy in early clinical trials for HER2-negative breast cancer. In this study, we examined a new molecular mechanism of nintedanib in TNBC. The results demonstrated that nintedanib enhanced TNBC cell apoptosis, which was accompanied by a reduction of p-STAT3 and its downstream proteins. STAT3 overexpression suppressed nintedanib-mediated apoptosis and further increased the activity of purified SHP-1 protein. Moreover, treatment with either a specific inhibitor of SHP-1 or SHP-1-targeted siRNA reduced the apoptotic effects of nintedanib, which validates the role of SHP-1 in nintedanib-mediated apoptosis. Furthermore, nintedanib-induced apoptosis was attenuated in TNBC cells expressing SHP-1 mutants with constantly open conformations, suggesting that the autoinhibitory mechanism of SHP-1 attenuated the effects of nintedanib. Importantly, nintedanib significantly inhibited tumor growth via the SHP-1/p-STAT3 pathway. Clinically, SHP-1 levels were downregulated, whereas p-STAT3 was upregulated in tumor tissues, and SHP-1 transcripts were associated with improved disease-free survival in TNBC patients. Our findings revealed that nintedanib induces TNBC apoptosis by acting as a SHP-1 agonist, suggesting that targeting STAT3 by enhancing SHP-1 expression could be a viable therapeutic strategy against TNBC.

Recurrent triple-negative breast cancer (TNBC) needs new therapeutic targets. Src homology region 2 domain-containing phosphatase-1 (SHP-1) can act as a tumor suppressor by dephosphorylating oncogenic kinases. One major target of SHP-1 is STAT3, which is highly activated in TNBC. In this study, we tested a sorafenib analogue SC-60, which lacks angiokinase inhibition activity, but acts as a SHP-1 agonist, in TNBC cells. SC-60 inhibited proliferation and induced apoptosis by dephosphorylating STAT3 in both a dose- and time-dependent manner in TNBC cells (MDA-MB-231, MDA-MB-468, and HCC1937). By contrast, ectopic expression of STAT3 rescued the anticancer effect induced by SC-60. SC-60 also increased the SHP-1 activity, but this effect was inhibited when the N-SH2 domain (DN1) was deleted or with SHP-1 point mutation (D61A), implying that SHP-1 is the major target of SC-60 in TNBC. The use of SC-60 in combination with docetaxel synergized the anticancer effect induced by SC-60 through the SHP-1/STAT3 pathway in TNBC cells. Importantly, SC-60 also displayed a significant antitumor effect in an MDA-MB-468 xenograft model by modulating the SHP-1/STAT3 axis, indicating the anticancer potential of SC-60 in TNBC treatment. Targeting SHP-1/p-STAT3 and the potential combination of SHP-1 agonist with chemotherapeutic docetaxel is a feasible therapeutic strategy for TNBC., (© 2017 The Authors. Published by FEBS Press and John Wiley & Sons Ltd.)

Objectives: Cancerous inhibitor of protein phosphatase 2A (CIP2A) has emerged as a therapeutic determinant mediating the anti-cancer effects of several new agents. We investigated the efficacy and mechanism of TD52, an erlotinib derivative with minimal p-EGFR inhibition but significant CIP2A downregulation, in triple-negative breast cancer (TNBC) cells., Methods: TNBC lines were used for in vitro studies. Cell apoptosis was examined by flow cytometry and Western blot. Signal transduction pathways in cells were assessed by Western blot. In vivo efficacy of TD52 was tested in xenograft nude mice., Results: We explored the CIP2A mRNA expression in a publically available database and found that higher levels of CIP2A mRNA is associated with worse recurrence-free survival in patients with TNBC. TD52-enhanced apoptosis accompanied with CIP2A downregulation and CIP2A overexpression protected cells from TD52-mediated apoptosis. The activity of protein phosphatase 2A (PP2A) was also increased in TD52-treated cells. TD52-induced apoptosis and p-Akt downregulation was attenuated by PP2A antagonist okadaic acid. Furthermore, TD52 indirectly downregulated CIP2A transcription via disturbing the binding of Elk1 to the CIP2A promoter. Importantly, TD52 showed anti-tumour activity in mice bearing TNBC xenograft tumours and downregulated CIP2A and p-Akt in these xenografted tumours. Interestingly, higher Elk1 mRNA expression was also associated with worse recurrence-free survival in TNBC patients by Kaplan-Meier survival analysis., Conclusion: Our findings indicated that EGFR-independent pharmacological modulation on Elk1/CIP2A signalling mediates the apoptotic effect of TD52 in TNBC cells, suggesting the potential therapeutic strategy., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Saikosaponin a (SSa) is one of the main active components of Bupleurum falcatum. It is commonly used to treat liver injury and fibrosis in traditional Chinese medicine. Our previous study showed that SSa induces apoptosis and inhibits the proliferation of rat hepatic stellate cell (HSC) line HSC-T6. The aim of the present study was to elucidate the mechanism of SSa-mediated apoptosis. Rat HSC cell line HSC-T6 and human HSC cell line LX-2 were used in this study. SSa triggered cell death mainly by apoptosis, as indicated by the typical morphological changes, sub-G1 phase of cell cycle increase, and activation of the caspase-9/caspase-3 cascade. In addition, SSa-induced apoptosis was partially inhibited by the caspase-3 inhibitor Z-DEVD-FMK, suggesting an involvement of caspase-3 dependent and independent pathways. Moreover, SSa upregulated pro-apoptotic proteins [BAK, Bcl-2-associated death promoter (BAD), and p53 upregulated modulator of apoptosis (PUMA)] and downregulated anti-apoptotic proteins (Bcl-2). In the mitochondria, SSa triggered the translocation of BAX and BAK from the cytosol to the outer membrane, resulting in a reduction of mitochondrial functions and membrane potential and subsequent release of apoptotic factors. Therefore, this study demonstrates that SSa induces apoptosis through the intrinsic mitochondrial-dependent pathway in HSCs.

Background: Testicular torsion/detorsion (T/D) can induce germ cells apoptosis, which may lead to impairment of spermatogenesis. FTY720, an agonist of the sphingosine-1-phosphate receptor 1 (S1PR1), inhibits apoptosis in ischemic stroke. We examined whether FTY720 could mitigate germ cell apoptosis in testicular T/D rats., Materials and Methods: Adult male Sprague-Dawley rats were allocated to receive testicular T/D (the T/D group), T/D plus FTY720 (the T/D-FTY group), or T/D plus FTY720 plus the potent S1PR1 antagonist VPC23019 (the T/D-FTY-VPC group; n = 6 in each group). Sham control groups were run simultaneously. At 24 h after detorsion, rats were euthanized., Results: Our data revealed that, in the ipsilateral twisted testes, sperm counts and expression of the S1PR1 of the T/D and the T/D-FTY-VPC groups were significantly lower than those of the T/D-FTY group (all P < 0.001). In contrast, signals of apoptotic cells stained by terminal deoxynucleotidyl transferase dUTP nick end labeling and the proapoptotic protein cleaved caspase-3 of the T/D, and the T/D-FTY-VPC groups were significantly stronger than those of the T/D-FTY group. Moreover, the terminal deoxynucleotidyl transferase dUTP nick end labeling signals mainly localized to germ cells., Conclusions: FTY720 could mitigate testicular T/D-induced germ cell apoptosis, and the mechanisms may involve the S1PR1., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Molybdenum (Mo), a well-known toxic environmental and industrial pollutant, causes adverse health effects and diseases in humans and has received attention as a potential risk factor for DM. However, the roles of Mo in the mechanisms of the toxicological effects in pancreatic β-cells are mostly unclear. In this study, the results revealed dysfunction of insulin secretion and apoptosis in the pancreatic β-cell-derived RIN-m5F cells and the isolated mouse islets in response to Mo. These effects were accompanied by a mitochondria-dependent apoptotic signals including a decreased in the MMP, an increase in cytochrome c release, and the activation of caspase cascades and PARP. In addition, ER stress was triggered as indicated by several key molecules of the UPR. Furthermore, exposure to Mo induced the activation of ERK1/2, JNK, AMPKα, and GSK3-α/β. Pretreatment with specific pharmacological inhibitors (in RIN-m5F cells and isolated mouse islets) of JNK (SP600125) and AMPK (Compound C) or transfection with si-RNAs (in RIN-m5F cells) specific to JNK and AMPKα effectively prevented the Mo-induced apoptosis and related signals, but inhibitors of ERK1/2 and GSK3-α/β (PD98059 and LiCl, respectively) did not reverse the Mo-induced effects. Additionally, both the inhibitors and specific si-RNAs could suppress the Mo-induced phosphorylation of JNK and AMPKα each other. Taken together, these results suggest that Mo exerts its cytotoxicity on pancreatic β-cells by inducing dysfunction and apoptosis via interdependent JNK and AMPK activation downstream-regulated mitochondrial-dependent and ER stress-triggered apoptosis pathways., (Copyright © 2016 Elsevier Inc. All rights reserved.)

We tested the efficacy of lapatinib, a dual tyrosine kinase inhibitor which interrupts the HER2 and epidermal growth factor receptor (EGFR) pathways, in a panel of triple-negative breast cancer (TNBC) cells, and examined the drug mechanism. Lapatinib showed an anti-proliferative effect in HCC 1937, MDA-MB-468, and MDA-MB-231 cell lines. Lapatinib induced significant apoptosis and inhibited CIP2A and p-Akt in a dose and time-dependent manner in the three TNBC cell lines. Overexpression of CIP2A reduced lapatinib-induced apoptosis in MDA-MB-468 cells. In addition, lapatinib increased PP2A activity (in relation to CIP2A inhibition). Moreover, lapatinib-induced apoptosis and p-Akt downregulation was attenuated by PP2A antagonist okadaic acid. Furthermore, lapatinib indirectly decreased CIP2A transcription by disturbing the binding of Elk1 to the CIP2A promoter. Importantly, lapatinib showed anti-tumor activity in mice bearing MDA-MB-468 xenograft tumors, and suppressed CIP2A as well as p-Akt in these xenografted tumors. In summary, inhibition of CIP2A determines the effects of lapatinib-induced apoptosis in TNBC cells. In addition to being a dual tyrosine kinase inhibitor of HER2 and EGFR, lapatinib also inhibits CIP2A/PP2A/p-Akt signaling in TNBC cells.

This study aimed to investigate the effects of harmine hydrochloride (HMH) on digestive tumor cells in vitro and its molecular mechanism. MTT assays showed that HMH inhibited the proliferation of some human cancer cell lines and had no obvious inhibitory effects on human LO2 cells. Flow cytometry assays showed that HMH trigged G2 phase arrest in MGC-803 cells and SMMC-7721 cells, while the expression of cyclin A, cyclin B, p21, Myt1, and p-cdc2 (Tyr15) was upregulated. Flow cytometry assays also showed that the percentages of apoptotic cells were increased, the mitochondrial transmembrane potential (ΔΨm) decreased, and the cleavage of caspase-9, caspase-3, and poly (Adenosine diphosphate ribose) polymerase (PARP) were observed, the expression of Bad increased, phospho-Bad (S112) decreased, pro-caspase-8 was cleaved, and Bid (22 kDa) was cleaved. The expression of p-ERK decreased in both cells. In conclusion, these results demonstrated that HMH upregulates the expression of p21, activates Myt1 and inhibits cdc2 by phospho-cdc2 (Y15), and triggers G2 phase arrest in both MGC-803 cells and SMMC-7721 cells. It can also activate the mitochondria-related cell apoptosis pathway through the caspase-8/Bid pathway, inhibiting the ERK/Bad pathway and promoting apoptosis in both of these two cell types., (Copyright © 2015 John Wiley & Sons, Ltd.)

Background/purpose: Monocytes play important roles in inflammatory responses and vascular remodeling after vascular stenting. This research focused on impacts of nickel (Ni) ions released from a corroded cardiovascular stent on cytotoxicity and monocyte activation., Methods: A human promonocytic (macrophage-like) cell line (U937) was exposed to graduated concentrations of Ni(2+)in vitro. Cells were observed and harvested at indicated times to determine the effects using histological and biochemical methods., Results: Ni caused U937 cell death in dose- and time-dependent manners. In vitro, high concentrations of Ni(2+) (>240 μM) significantly induced cell apoptosis and increased terminal deoxynucleotidyl transferase (TdT) dUTP nick end labeling (TUNEL)-positive cells according to flow cytometric surveillance and triggered apoptotic cell death. Although no significant changes in Bcl-2 or Bax expressions were detected after 24 hours of Ni(2+) treatment, increasing cleavage of caspase-3 and -8 was present. Results showed that cleavage of caspase-8 was inhibited by the presence of the inhibitor, Z-IETD-FMK, and this suggested the presence of Ni(2+)-induced U937 cell death through a death receptor-mediated pathway. Simultaneously, when treated with a high concentration of Ni(2+) ions, expressions of the vascular remodeling factors, matrix metalloproteinases (MMP)-9 and -2, were activated in dose- and time-dependent manners. Secretion of the proliferative factor, monocyte chemoattractant protein (MCP)-1, significantly increased during the first 6 hours of incubation with 480 μM Ni(2+)-treated medium., Conclusion: Our results demonstrated that a high concentration of Ni ions causes apoptotic cell death of circulating monocytes. They may also play different roles in vascular remodeling during the corrosion process following implantation of Ni alloy-containing devices., (Copyright © 2014. Published by Elsevier B.V.)

Early acute kidney injury (AKI) is a devastating complication in critical burn patients, and it is associated with severe morbidity and mortality. The mechanism of AKI is multifactorial. Astaxanthin (ATX) is a natural compound that is widely distributed in marine organisms; it is a strong antioxidant and exhibits other biological effects that have been well studied in various traumatic injuries and diseases. Hence, we attempted to explore the potential protection of ATX against early post burn AKI and its possible mechanisms of action. The classic severe burn rat model was utilized for the histological and biochemical assessments of the therapeutic value and mechanisms of action of ATX. Upon ATX treatment, renal tubular injury and the levels of serum creatinine and neutrophil gelatinase-associated lipocalin were improved. Furthermore, relief of oxidative stress and tubular apoptosis in rat kidneys post burn was also observed. Additionally, ATX administration increased Akt and Bad phosphorylation and further down-regulated the expression of other downstream pro-apoptotic proteins (cytochrome c and caspase-3/9); these effects were reversed by the PI3K inhibitor LY294002. Moreover, the protective effect of ATX presents a dose-dependent enhancement. The data above suggested that ATX protects against early AKI following severe burns in rats, which was attributed to its ability to ameliorate oxidative stress and inhibit apoptosis by modulating the mitochondrial-apoptotic pathway, regarded as the Akt/Bad/Caspases signalling cascade.

To investigate the association between high parity and triple-negative breast cancer (TNBC), and explore the etiologic mechanisms of TNBC in Tientsin Albinao 2 (TA2) mice model. After the TA2 mice model with high parity and TNBC had been established, the cell proliferation and apoptosis were detected by immunohistochemical and TUNEL staining in mammary epithelia from different conditions, which included non-pregnancy, low and high gravidity in pregnancy, and carcinogenesis. Apoptotic signaling was analyzed by measuring bcl-2, bax, caspase-3, and caspase-9 expression using immunohistochemistry (IHC), western blot, and real-time PCR technique. Estrogen receptor α (ERα) and progesterone receptor (PR) were determined by immunohistochemical staining and real-time PCR. Both proliferation and apoptosis in mammary epithelia changed with the increasing of parity. Immunohistochemistry revealed increased cell proliferation and apoptosis were related with upregulation of ERα, PR, bcl-2, bax, caspase-3, and caspase-9 expression, especially during the fourth pregnancy. Mammary gland in the fourth pregnancy stage was the closest to precancerous. In precancerous mammary gland, cell proliferation rate was much higher than apoptosis rate. High parity could increase the ovarian hormones level and alter ovarian hormone receptor levels in TA2 mice, and their sensitivity to ovarian hormones result in the imbalance between cell proliferation and apoptosis in precancerous mammary epithelial cells.

Through a high-fat diet, obesity leads to cardiomyocyte dysfunction and apoptosis. In addition, there is no evidence that probiotics have potential health effects associated with cardiac apoptosis in obese rats. The present study aimed to explore the effects of probiotics on obesity and cardiac apoptosis in rats fed a high-fat diet (HF). Eight‑week‑old male Wistar rats were separated randomly into five equally sized experimental groups: Normal diet (NC) and high-fat diet (HFC) groups, and high-fat diet supplemented with low (HFL), medium (HFM) or high (HFH) doses of multi‑strain probiotics groups. The rats were subsequently studied for 8 weeks. Food intake and body weights were recorded following sacrifice, and food utilization rates, body fat and serum cholesterol levels were analysed. The myocardial architecture of the left ventricle was evaluated by hematoxylin‑eosin staining, and key apoptotic‑related pathway molecules were analysed by western blotting. Rat weights and triglyceride levels were decreased with oral administration of high doses of probiotics (HFH) compared to the HFC group. Abnormal myocardial architecture and enlarged interstitial spaces were observed in HFC hearts, but were significantly decreased in groups that were provided multi‑strain probiotics compared with NC hearts. Western blot analysis demonstrated that key components of the Fas receptor‑ and mitochondrial‑dependent apoptotic pathways were significantly suppressed in multi‑strain probiotic treated groups compared to the HF group. Additionally, cardiac insulin, such as the insulin‑like growth factor I receptor (IGFIR)‑dependent survival signalling components, were highly induced in left ventricles from rats administered probiotics. Together, these findings strongly suggest that oral administration of probiotics may attenuate cardiomyocyte apoptosis by activation of the phosphatidylinositol‑3 kinase/AKT survival‑signalling pathway in obese rats.

Bladder cancer is a common malignancy worldwide. However, there is still no effective therapy for bladder cancer. In this study, we investigated the cytotoxic effects of cantharidin [a natural toxin produced (pure compound) from Chinese blister beetles (Mylabrisphalerata or Mylabriscichorii) and Spanish flies (Cantharis vesicatoria)] in human bladder cancer cell lines (including: T24 and RT4 cells). Treatment of human bladder cancer cells with cantharidin significantly decreased cell viability. The increase in the expressions of caspase-3 activity and cleaved form of caspase-9/-7/-3 were also increased in cantharidin-treated T24 cells. Furthermore, cantharidin increased the levels of phospho-eIF2α and Grp78 and decreased the protein expression of procaspase-12, which was accompanied by the increase in calpain activity in T24 cells. Cantharidin was capable of increasing the intracellular Ca (2+) and the phosphorylation of protein kinase C (PKC) in T24 cells. The addition of BAPTA/AM (a Ca (2+) chelator) and RO320432 (a selective cell-permeable PKC inhibitor) effectively reversed the increase in caspase-3 and calpain activity, the phosphorylation levels of PKC and eIF2α and Grp78 protein expression, and the decrease in procaspase-12 expression induced by cantharidin. Importantly, cantharidin significantly decreased the tumor volume (a dramatic 71% reduction after 21 days of treatment) in nude mice xenografted with T24 cells. Taken together, these results indicate cantharidin induced human bladder cancer cell apoptosis through a calcium/PKC-regulated ER stress pathway. These findings suggest that cantharidin may be a novel and potential anticancer agent targeting on bladder cancer cells.

Unlabelled: Abstract Purpose: The radioprotective effects of Antrodia cinnamomea (AC) were investigated for understanding the potential usefulness of AC as an adjunct treatment for reducing radiation side-effects., Materials and Methods: In this study, we determined the ability of AC extracts (AC539) to reduce radiation side-effects by analyzing cellular viability in normal mouse spleen immune cells and human cancer cells with different radiosensitivity. We further detected the effect of AC on radiation-induced changes in cytokine- and inflammatory-related gene expressions. Furthermore, apoptosis assay was performed to determine whether AC could inhibit radiation-induced cytotoxicity., Results: We found that an AC dose of 100-150 μg/ml in a time-dependent manner was the most effective in blocking radiation-induced cytotoxicity, in vitro. Radiation-induced cytotoxicity was inhibited in spleen immune cells by 37-56%; however, pretreatment of human colorectal cancer cell line HT-29 with AC did not have any effect on radiation-induced cytotoxicity, while pretreatment of radiosensitive human breast cancer cell lines BT-474 with AC caused a moderate enhancement of radiation-induced damage. Furthermore, AC pretreatment differentially regulated the mRNA expression of several important immunomodulatory genes in response to irradiation in normal and cancer cells., Conclusions: Our data indicate that AC may inhibit important immunoregulatory signaling which could be vital in the avoidance of an over-activated cytotoxic and inflammatory response of the immune system caused by radiation-induced tissue damage. Additionally, AC does not provide a radioprotective effect to tumor cells but instead enhances radiation-induced inflammation and cytotoxicity in cancer.

Introduction: Tamoxifen, a selective estrogen receptor (ER) modulator, may affect cancer cell survival through mechanisms other than ER antagonism. In the present study, we tested the efficacy of tamoxifen in a panel of ER-negative breast cancer cell lines and examined the drug mechanism., Methods: In total, five ER-negative breast cancer cell lines (HCC-1937, MDA-MB-231, MDA-MB-468, MDA-MB-453 and SK-BR-3) were used for in vitro studies. Cellular apoptosis was examined by flow cytometry and Western blot analysis. Signal transduction pathways in cells were assessed by Western blot analysis. The in vivo efficacy of tamoxifen was tested in xenograft nude mice., Results: Tamoxifen induced significant apoptosis in MDA-MB-231, MDA-MB-468, MDA-MB-453 and SK-BR-3 cells, but not in HCC-1937 cells. Tamoxifen-induced apoptosis was associated with inhibition of cancerous inhibitor of protein phosphatase 2A (CIP2A) and phospho-Akt (p-Akt) in a dose-dependent manner. Ectopic expression of either CIP2A or Akt protected MDA-MB-231 cells from tamoxifen-induced apoptosis. In addition, tamoxifen increased protein phosphatase 2A (PP2A) activity, and tamoxifen-induced apoptosis was attenuated by the PP2A antagonist okadaic acid in the sensitive cell lines, but not in resistant HCC-1937 cells. Moreover, silencing CIP2A by small interfering RNA sensitized HCC-1937 cells to tamoxifen-induced apoptosis. Furthermore, tamoxifen regulated CIP2A protein expression by downregulating CIP2A mRNA. Importantly, tamoxifen inhibited the in vivo growth of MDA-MB-468 xenograft tumors in association with CIP2A downregulation, whereas tamoxifen had no significant effect on CIP2A expression and anti-tumor growth in HCC-1937 tumors., Conclusions: Inhibition of CIP2A determines the effects of tamoxifen-induced apoptosis in ER-negative breast cancer cells. Our data suggest a novel "off-target" mechanism of tamoxifen and suggest that CIP2A/PP2A/p-Akt signaling may be a feasible anti-cancer pathway.

Osteosarcoma (OS) is the most common form of malignant bone tumor and is an aggressive malignant neoplasm exhibiting osteoblastic differentiation. Cisplatin is one of the most efficacious antitumor drugs for osteosarcoma patients. However, treatment failures are common due to the development of chemoresistance. CCN2 (also known as CTGF), is a secreted protein that binds to integrins, modulates the invasive behavior of certain human cancer cells. However, the effect of CCN2 in cisplatin-mediated chemotherapy is still unknown. Here, we found that CCN2 was upregulated in human osteosarcoma cells after treatment with cisplatin. Moreover, overexpression of CCN2 increased the resistance to cisplatin-mediated cell apoptosis. In contrast, reduction of CCN2 by CCN2 shRNA promoted the chemotherapeutic effect of cisplatin. We also found that CCN2 provided resistance to cisplatin-induced apoptosis through upregulation of Bcl-xL and survivin. Knockdown of Bcl-xL or survivin removed the CCN2-mediated resistance to apoptosis induced by cisplatin. On the other hand, CCN2 also promoted FAK, MEK, and ERK survival signaling pathways to enhance tumor survival during cisplatin treatment. In a mouse xenograft model, overexpression of CCN2 promoted resistance to cisplatin. However, knockdown of CCN2 increased the therapeutic effect of cisplatin. Therefore, our data suggest that CCN2 might be a critical oncogene of human osteosarcoma for cisplatin-resistance and supported osteosarcoma cell growth in vivo and in vitro.

Cervix cancer is the second most common cancer among women worldwide, whereas paclitaxel, the first line chemotherapeutic drug used to treat cervical cancer, shows low chemosensitivity on the advanced cervical cancer cell line. Tanshinone IIA (Tan IIA) exhibited strong growth inhibitory effect on CaSki cells (IC50 = 5.51 μM) through promoting caspase cascades with concomitant upregulating the phosphorylation of p38 and JNK signaling. Comprehensive proteomics revealed the global protein changes and the network analysis implied that Tan IIA treatment would activate ER stress pathways that finally lead to apoptotic cell death. Moreover, ER stress inhibitor could alleviate Tan IIA caused cell growth inhibition and ameliorate C/EBP-homologous protein as well as apoptosis signal-regulating kinase 1 mediated cell death. The therapeutic interventions targeting the mitochondrial-related apoptosis and ER stress responses might be promising strategies to conquer paclitaxel resistance., (© 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Purpose: Previous studies reported that patients with endometriosis had excess nitric oxide (NO) in the reproductive tract and poor embryo development in IVF cycles. This study aims to elucidate the effects of NO on early embryo development., Methods: Zygotes from superovulated B6CBF1 mice were cultured to blastocysts in a variety of media. Sodium nitroprusside (SNP) and N(G)-nitro-L-arginine (LNA) were added to the culture medium as a NO donor and a NO synthase inhibitor, respectively. The localization and fluorescence intensity of S-nitrosylated (SNO) proteins within 2-cell stage embryos were analyzed with confocal microscopy. Apoptosis and ATP production in the blastocysts were measured., Result(s): Subsequent to NO exposure, the SNO proteins mainly colocalized with the mitochondria and endoplasmic reticulum and the intensity of SNO proteins increased. The addition of a quanylate cyclase inhibitor and a cyclic GMP mimic agent induced nonsignificant changes in SNO proteins, whereas addition of a superoxide scavenger or a reduced form of glutathione rescued the embryos from the effects of NO. However, superoxide scavenger supplementation resulted in decreased blastocyst ATP production., Conclusion(s): Elevated NO exerts deleterious effects on embryo development, possibly through protein S-nitrosylation in the mitochondria and endoplasmic reticulum. Including glutathione as a component in the culture medium might counteract this effect.

Background: Apoptosis-associated biomarkers are rarely studied, especially their role in predicting the development of tuberculosis (TB) from latent TB infection and in prognostication., Methods: Patients with TB and interferon-gamma release assay (IGRA)-positive and IGRA-negative family contacts were evaluated to analyze changes in apoptosis-associated serum biomarkers, which included decoy receptor 3 (DcR3), prostaglandin 2 (PGE2), and lipoxin. The prognostic implications of these serum biomarkers were also analyzed., Results: One hundred TB patients and 92 IGRA-negative and 91 IGRA-positive family contacts were recruited. The DcR3 and PGE2 levels decreased from the IGRA-negative group to the IGRA-positive group, and peaked in the TB group. Lipoxin decreased to trough in the TB group. The three apoptosis serum markers and age were independent factors discriminating active TB from latent TB infection. In active TB, older age, co-morbidity, and higher serum DcR3 and monocyte chemotactic protein (MCP)-1 were independently associated with poorer six-month survival., Conclusion: Apoptosis-associated serum biomarkers change along with the status of Mycobacterium tuberculosis infection. In close contacts with positive IGRA, high DcR3 and PGE2 and low lipoxin may increase the probability of active TB. Older age, co-morbidity, and high DcR3 and MCP-1 levels might be important prognostic factors that warrant further investigation.

Chloroacetic acid (CA), a toxic chlorinated analog of acetic acid, is widely used in chemical industries as an herbicide, detergent, and disinfectant, and chemical intermediates that are formed during the synthesis of various products. In addition, CA has been found as a by-product of chlorination disinfection of drinking water. However, there is little known about neurotoxic injuries of CA on the mammalian, the toxic effects and molecular mechanisms of CA-induced neuronal cell injury are mostly unknown. In this study, we examined the cytotoxicity of CA on cultured Neuro-2a cells and investigated the possible mechanisms of CA-induced neurotoxicity. Treatment of Neuro-2a cells with CA significantly reduced the number of viable cells (in a dose-dependent manner with a range from 0.1 to 3mM), increased the generation of ROS, and reduced the intracellular levels of glutathione depletion. CA also increased the number of sub-G1 hypodiploid cells; increased mitochondrial dysfunction (loss of MMP, cytochrome c release, and accompanied by Bcl-2 and Mcl-1 down-regulation and Bax up-regulation), and activated the caspase cascades activations, which displayed features of mitochondria-dependent apoptosis pathway. These CA-induced apoptosis-related signals were markedly prevented by the antioxidant N-acetylcysteine (NAC). Moreover, CA activated the JNK and p38-MAPK pathways, but did not that ERK1/2 pathway, in treated Neuro-2a cells. Pretreatment with NAC and specific p38-MAPK inhibitor (SB203580), but not JNK inhibitor (SP600125) effectively abrogated the phosphorylation of p38-MAPK and attenuated the apoptotic signals (including: decrease in cytotoxicity, caspase-3/-7 activation, the cytosolic cytochrome c release, and the reversed alteration of Bcl-2 and Bax mRNA) in CA-treated Neuro-2a cells. Taken together, these data suggest that oxidative stress-induced p38-MAPK activated pathway-regulated mitochondria-dependent apoptosis plays an important role in CA-caused neuronal cell death., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Cadmium (Cd), one of well-known highly toxic environmental and industrial pollutants, causes a number of adverse health effects and diseases in humans. The growing epidemiological studies have suggested a possible link between Cd exposure and diabetes mellitus (DM). However, the toxicological effects and underlying mechanisms of Cd-induced pancreatic β-cell injury are still unknown. In this study, we found that Cd significantly decreased cell viability, and increased sub-G1 hypodiploid cells and annexin V-Cy3 binding in pancreatic β-cell-derived RIN-m5F cells. Cd also increased intracellular reactive oxygen species (ROS) generation and malondialdehyde (MDA) production and induced mitochondrial dysfunction (the loss of mitochondrial membrane potential (MMP) and the increase of cytosolic cytochrome c release), the decreased Bcl-2 expression, increased p53 expression, poly (ADP-ribose) polymerase (PARP) cleavage, and caspase cascades, which accompanied with intracellular Cd accumulation. Pretreatment with the antioxidant N-acetylcysteine (NAC) effectively reversed these Cd-induced events. Furthermore, exposure to Cd induced the phosphorylations of c-jun N-terminal kinases (JNK), extracellular signal-regulated kinases (ERK)1/2, and p38-mitogen-activated protein kinase (MAPK), which was prevented by NAC. Additionally, the specific JNK inhibitor SP600125 or JNK-specific small interference RNA (si-RNA) transfection suppressed Cd-induced β-cell apoptosis and related signals, but not ERK1/2 and p38-MAPK inhibitors (PD98059 and SB203580) did not. However, the JNK inhibitor or JNK-specific si-RNA did not suppress ROS generation in Cd-treated cells. These results indicate that Cd induces pancreatic β-cell death via an oxidative stress downstream-mediated JNK activation-triggered mitochondria-regulated apoptotic pathway.

Mercury is a toxic heavy metal that is an environmental and industrial pollutant throughout the world. Mercury exposure leads to many physiopathological injuries in mammals. However, the precise toxicological effects of mercury on pancreatic islets in vivo are still unclear. Here, we investigated whether mercuric compounds can induce dysfunction and damage in the pancreatic islets of mice, as well as the possible mechanisms involved in this process. Mice were treated with methyl mercuric chloride (MeHgCl, 2 mg/kg) and mercuric chloride (HgCl(2), 5 mg/kg) for more than 2 consecutive weeks. Our results showed that the blood glucose levels increased and plasma insulin secretions decreased in the mice as a consequence of their exposure. A significant number of TUNEL-positive cells were revealed in the islets of mice that were treated with mercury for 2 consecutive weeks, which was accompanied by changes in the expression of the mRNA of anti-apoptotic (Bcl-2, Mcl-1, and Mdm-2) and apoptotic (p53, caspase-3, and caspase-7) genes. Moreover, plasma malondialdehyde (MDA) levels increased significantly in the mice after treatment with mercuric compounds for 2 consecutive weeks, and the generation of reactive oxygen species (ROS) in the pancreatic islets also markedly increased. In addition, the mRNA expression of genes related to antioxidation, including Nrf2, GPx, and NQO1, were also significantly reduced in these islets. These results indicate that oxidative stress injuries that are induced by mercuric compounds can cause pancreatic islets dysfunction and apoptosis in vivo.

Unlabelled: What's known on the subject? and What does the study add? Protein kinase C inhibitor (PKCI) can decrease glomerular and tubular cell apoptosis and mitosis and attenuate collagen accumulation and fibronectin expression in a PUUO rat model. Although the role of PKC has been well studied in diabetic nephropathy, there is no report on its role in obstructive nephropathy. This investigation evaluated the processes that were associated with the activation of PKCα and PKCβ pathways and showed that PKCI played an important role in the protection of renal function during ureteric obstruction., Objectives: • To investigate the expression of the protein kinase C (PKC) pathway after partial unilateral ureteric obstruction (PUUO). • To evaluate the therapeutic potential of a PKC inhibitor (PKCI) in obstructive nephropathy., Materials and Methods: • Thirty-six rats were divided into three groups. One sham-operated group served as the control. The other two groups received PUUO surgery, after which one group received no treatment and the other group was treated with PKCI, chelerythrine. • The severity of hydronephrosis and renal morphology were assessed: tubular and glomerularcell apoptosis, mitosis and interstitial fibrosis were examined using immunohistochemistry. • Western immunoblots were performed to determine fibronectin, transforming growth factor-β (TGF-β), and PKC isoform levels., Results: • Two weeks after PUUO surgery, hydronephrosis progressively developed. Tubular-interstitial fibrosis, collagen deposition and fibronectin expression were increased. • PUUO also activated the expression of PKCα and PKCβ and the translocation of PKCs from cell cytosol to cell membranes. • Treatment with PKCI significantly decreased PKCα and PKCβ expression and translocation in the renal cortex. • Treatment with PKCI also reduced the severity of hydronephrosis, decreased both glomerular and tubular cell apoptosis and mitosis, and attenuated the collagen and fibronectin accumulation in renal interstitium., Conclusions: • Renal tubular apoptosis and interstitial fibrosis after obstructive nephropathy are associated with PKCα and PKCβ activation. • The PKCI, chelerythrine, is capable of decreasing PKC expression and translocation in the renal cortex, suggesting that this inhibitor may have therapeutic potential in the protection of renal function in the first few weeks after PUUO surgery., (© 2011 THE AUTHORS. BJU INTERNATIONAL © 2011 BJU INTERNATIONAL.)

A pool of myoblasts available for myogenesis is important for skeletal muscle size. The decreased number of skeletal muscle fibers could be due to the decreased myoblast proliferation or cytotoxicity. Identification of toxicants that regulate myoblast apoptosis is important in skeletal muscle development or regeneration. Here, we investigate the cytotoxic effect and its possible mechanisms of arsenic trioxide (As(2)O(3)) on myoblasts. C2C12 myoblasts underwent apoptosis in response to As(2)O(3) (1-10 μM), accompanied by increased Bax/Bcl-2 ratio, decreased mitochondria membrane potential, increased cytochrome c release, increased caspase-3/-9 activity, and increased poly (ADP-ribose) polymerase (PARP) cleavage. Moreover, As(2)O(3) triggered the endoplasmic reticulum (ER) stress indentified through several key molecules of the unfolded protein response, including glucose-regulated protein (GRP)-78, GRP-94, PERK, eIF2α, ATF6, and caspase-12. Pretreatment with antioxidant N-acetylcysteine (NAC, 0.5 mM) dramatically suppressed the increases in reactive oxygen species (ROS), lipid peroxidation, ER stress, caspase cascade activity, and apoptosis in As(2)O(3) (10 μM)-treated myoblasts. Furthermore, As(2)O(3) (10 μM) effectively decreased the phosphorylation of Akt, which could be reversed by NAC. Over-expression of constitutive activation of Akt (c.a. Akt) also significantly attenuated As(2)O(3)-induced myoblast apoptosis. Taken together, these results suggest that As(2)O(3) may exert its cytotoxicity on myoblasts by inducing apoptosis through a ROS-induced mitochondrial dysfunction, ER stress, and Akt inactivation signaling pathway.

Hypoxic-ischemia injury occurs after trauma causes consequential bone necrosis. Non-steroid anti-inflammatory drugs (NSAIDs) are frequently used in orthopedic clinics for pain relief. However, the underlying mechanism and outcome for usage of NSAIDs is poorly understood. To investigate the damage and loss of osteoblast function in hypoxia, two hypoxia mimetics, cobalt chloride (CoCl(2)) and desferrioxamine (DFO), were used to create an in vitro hypoxic microenvironment. The cell damage was observed by decreases of cell viability and increases in cyclooxygenase-2 and cleaved poly(ADP-ribose) polymerase (PARP). Cell apoptosis was confirmed by WST-1 cytotoxic assays and flow cytometry. The functional expression of osteoblast in alkaline phosphatase (ALP) activity was significantly decreased by CoCl(2) and inhibited when treated with DFO. To simulate the use of NSAID after hypoxic injury, four types of anti-inflammatory drugs, sulindac sulfide (SUL), indomethacin (IND), aspirin (Asp), and sodium salicylate (NaS), were applied to osteoblasts after 1 h of hypoxia mimetic treatment. SUL and IND further enhanced cell death after hypoxia. ALP activity was totally abolished in hypoxic osteoblasts under IND treatment. Facilitation of osteoblast apoptosis occurred regardless of IND dosage under hypoxic conditions. To investigate osteoblast in vivo, local hypoxia was created by fracture of tibia and then treated the injured mice with IND by oral feeding. IND-induced osteoblast apoptosis was confirmed by positive staining of TUNEL assay in fractured mice. Significant delay of fracture healing in bone tissue was also observed with the treatment of IND. These results provide information pertaining to choosing appropriate anti-inflammatory drugs for orthopedic patients., (Copyright © 2011 Wiley Periodicals, Inc.)

Programmed cell death (apoptosis) is essential for the development and homeostasis of metazoans. The central step in the execution of programmed cell death is the activation of caspases. In C. elegans, the core cell death regulators EGL-1(a BH3 domain-containing protein), CED-9 (Bcl-2), and CED-4 (Apaf-1) act in an inhibitory cascade to activate the CED-3 caspase. Here we have identified an additional component eif-3.K (eukaryotic translation initiation factor 3 subunit k) that acts upstream of ced-3 to promote programmed cell death. The loss of eif-3.K reduced cell deaths in both somatic and germ cells, whereas the overexpression of eif-3.K resulted in a slight but significant increase in cell death. Using a cell-specific promoter, we show that eif-3.K promotes cell death in a cell-autonomous manner. In addition, the loss of eif-3.K significantly suppressed cell death-induced through the overexpression of ced-4, but not ced-3, indicating a distinct requirement for eif-3.K in apoptosis. Reciprocally, a loss of ced-3 suppressed cell death induced by the overexpression of eif-3.K. These results indicate that eif-3.K requires ced-3 to promote programmed cell death and that eif-3.K acts upstream of ced-3 to promote this process. The EIF-3.K protein is ubiquitously expressed in embryos and larvae and localizes to the cytoplasm. A structure-function analysis revealed that the 61 amino acid long WH domain of EIF-3.K, potentially involved in protein-DNA/RNA interactions, is both necessary and sufficient for the cell death-promoting activity of EIF-3.K. Because human eIF3k was able to partially substitute for C. elegans eif-3.K in the promotion of cell death, this WH domain-dependent EIF-3.K-mediated cell death process has potentially been conserved throughout evolution.

Nickel (Ni), a well-known toxic metal, is widely used in electroplating and alloy production. It is also significantly implicated in industrial and environmental pollution caused by uncontrolled industrial and municipal discharges. In this study, we characterized and investigated the cytotoxic effects of Ni exposure and their probable toxicological mechanisms in the pancreatic β-cells. The results showed that it was significantly decreased cell viability after exposing pancreatic β-cell-derived RIN-m5F cells to NiCl(2) for 24h in a dose-dependent manner. NiCl(2) also increased sub-G1 hypodiploid cells and Annexin V-Cy3 binding population in RIN-m5F cells, indicating that it has apoptosis-inducing ability. Moreover, the exposure of RIN-m5F cells to NiCl(2) induced distinct signals of mitochondria-dependent apoptosis, including mitochondrial dysfunction (the disruption of mitochondrial membrane potential (MMP) and increase in mitochondrial cytochrome c release into the cytosol), Bak and Bid mRNA up-regulation, and activation of caspase-3, caspase-7, and caspase-9, and poly(ADP-ribose) polymerase (PARP) degradation. In addition, NiCl(2) also markedly induced the activation of c-Jun N-terminal kinases (JNK), but not of extracellular signal-regulated kinase (ERK)1/2 and p38. These NiCl(2)-induced apoptosis-related signaling responses could be effectively reversed by specific JNK inhibitor SP600125. To the best of our knowledge, this study is the first to show that Ni causes pancreatic β-cell death through a JNK activation-regulated mitochondria-dependent apoptosis-signaling pathway., (Crown Copyright © 2011. Published by Elsevier Ireland Ltd. All rights reserved.)

Methylmercury (MeHg) is well-known for causing irreversible damage in the central nervous system as well as a risk factor for inducing neuronal degeneration. However, the molecular mechanisms of MeHg-induced neurotoxicity remain unclear. Here, we investigated the effects and possible mechanisms of MeHg in the mouse cerebrum (in vivo) and in cultured Neuro-2a cells (in vitro). In vivo study showed that the levels of LPO in the plasma and cerebral cortex significantly increased after administration of MeHg (50μg/kg/day) for 7 consecutive weeks. MeHg could also decrease glutathione level and increase the expressions of caspase-3, -7, and -9, accompanied by Bcl-2 down-regulation and up-regulation of Bax, Bak, and p53. Moreover, treatment of Neuro-2a cells with MeHg significantly reduced cell viability, increased oxidative stress damage, and induced several features of mitochondria-dependent apoptotic signals, including increased sub-G1 hypodiploids, mitochondrial dysfunctions, and the activation of PARP, and caspase cascades. These MeHg-induced apoptotic-related signals could be remarkably reversed by antioxidant NAC. MeHg also increased the phosphorylation of ERK1/2 and p38, but not JNK. Pharmacological inhibitors NAC, PD98059, and SB203580 attenuated MeHg-induced cytotoxicity, ERK1/2 and p38 activation, MMP loss, and caspase-3 activation in Neuro-2a cells. Taken together, these results suggest that the signals of ROS-mediated ERK1/2 and p38 activation regulated mitochondria-dependent apoptotic pathways that are involved in MeHg-induced neurotoxicity., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Arsenic pollution is a major public health problem worldwide. Inorganic arsenic (iAs) is usually more harmful than organic ones. iAs pollution increases the risk of human diseases such as peripheral vascular disease and cancer. However, the toxicological effects of iAs in the brain are mostly unclear. Here, we investigated the toxic effects and possible mechanisms of iAs in the cerebrum of mice after exposure to iAs (0.5 and 5 ppm (mg/l) As(2)O(3), via the drinking water), which was the possible human exposed dose via the ingestion in iAs-contaminated areas, for 6 consecutive weeks. iAs dose-dependently caused an increase of LPO production in the plasma and cerebral cortex. iAs also decreased the reduced glutathione levels and the expressions of NQO1 and GPx mRNA in the cerebral cortex. These impairments in the cerebral cortex caused by iAs exposure were significantly correlated with the accumulation of As. Moreover, iAs induced the production of apoptotic cells and activation of caspase-3, up-regulation of Bax and Bak, and down-regulation of Mcl-1 in the cerebral cortex. Exposure to iAs also triggered the expression of ER stress-related genes, including GRP78, GRP94, and CHOP. Meanwhile, an increase of p38 activation and dephosphorylation of ERK1/2 were shown in the cerebral cortex as a result of iAs-exposed mice. These iAs-induced damages and apoptosis-related signals could be significantly reversed by NAC. Taken together, these results suggest that iAs-induced oxidative stress causes cellular apoptosis in the cerebrum, signaling of p38 and ERK1/2, and ER stress may be involved in iAs-induced cerebral toxicity.

Arsenic (As), a ubiquitous toxic metal, is an important environmental and industrial pollutant throughout the world. Inorganic As (iAs) is usually more harmful than organic ones and with a high risk of diabetes incidence by exposure. However, the toxicological effects of iAs on growth and function of pancreatic β-cells still remain unclear. Here, we found that iAs significantly decreased insulin secretion and cell viability, and increased ROS and MDA formation in pancreatic β-cell-derived RIN-m5F cells. iAs also induced the increases in sub-G1 hypodiploids, annexin V-Cy3 binding, and caspase-3 activity in RIN-m5F cells, indicating that iAs could induce β-cell apoptosis. Moreover, iAs induced MAPKs activation, mitochondria dysfunction, p53 up-regulation, Bcl-2 and Mdm-2 down-regulation, PARP, and caspase cascades, which displayed features of mitochondria-dependent apoptotic signals. In addition, exposure of RIN-m5F cells to iAs, could trigger ER stress as indicated by the enhancement in ER stress-related molecules induction (such as GRP78, GRP94, CHOP, and XBP1), procaspase-12 cleavage, and calpain activation. The iAs-induced apoptosis and its-related signalings could be effectively reversed by antioxidant N-acetylcysteine. We next observed that exposure of mice to iAs in drinking water for 6 consecutive weeks significantly decreased decreased the plasma insulin, elevated glucose intolerance and plasma lipid peroxidation, and induced islet cells apoptosis, which accompanied with arsenic accumulation in the whole blood and pancreas. N-acetylcysteine effectively antagonized the iAs-induced responses in mice. Taken together, these results suggest that iAs-induced oxidative stress causes pancreatic β-cells apoptosis via the mitochondria-dependent and ER stress-triggered signaling pathways., (Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.)

Fibronectin (FN) is an endogenous ligand of integrins, which plays a critical role in cell adhesion and growth. Here, we converted globular FN (G-FN) into a fibrillar form (F-FN) and found that, even though both G-FN and F-FN interacted with integrin alpha5beta1, G-FN induced cellular proliferation, whereas F-FN resulted in apoptosis that was associated with deactivation of Akt/GSK-3beta and phosphorylation of SHP-2. SHP-2 inhibitor and anti-sense oligodeoxynucleotide decreased SHP-2 level and reversed the F-FN mediated apoptosis. F-FN also induced stress fiber formation associated with activation of RhoA, Rho kinase (ROCK), and filamin. Inhibition of ROCK by ROCK inhibitor or dominant negative plasmid treatment modulated F-FN mediated apoptosis. Pharmacological studies revealed that F-FN was effective in inhibiting the survival of SKOV-3 and MCF-7 cancer cells. These findings thus demonstrate that unlike G-FN, F-FN exhibits fibrillar structure to induce cell apoptosis that is associated with phosphorylation of SHP-2, activation of RhoA/ROCK and formation of stress fibers as well as deactivation of Akt/GSK-3beta.

AAA domain-containing 3A (ATAD3A) is a member of the AAA-ATPase family. Three forms of ATAD3 have been identified: ATAD3A, ATAD3B and ATAD3C. In this study, we examined the type and expression of ATAD3 in lung adenocarcinoma (LADC). Expression of ATAD3A was detected by reverse transcription-polymerase chain reaction, immunoblotting, immunohistochemistry and confocal immunofluorescent microscopy. Our results show that ATAD3A is the major form expressed in LADC. Silencing of ATAD3A expression increased mitochondrial fragmentation and cisplatin sensitivity. Serum deprivation increased ATAD3A expression and drug resistance. These results suggest that ATAD3A could be an anti-apoptotic marker in LADC.

Apoptotic effects of oleanolic acid (OA) and ursolic acid (UA) on human liver cancer HepG2, Hep3B, Huh7 and HA22T cell lines were examined. OA or UA at 2, 4, 8 micromol/L were used and their effects on cell viability, DNA fragmentation, mitochondrial membrane potential (MMP), activity of Na(+)-K(+)-ATPase, caspase-3 and caspase-8, cell adhesion, level of intercellular adhesion molecule (ICAM)-1 and vascular endothelial growth factor (VEGF) in these cell lines were determined. OA or UA treatments concentration-dependently decreased cell viability and increased DNA fragmentation in HepG2 and Hep3B cell lines (P<0.05). However, these two compounds reduced viability and increased DNA fragmentation in Huh7 cell only at 4 and 8 micromol/L (P<0.05). OA or UA treatments concentration-dependently lowered MMP in HepG2, Hep3B and HA22T cell lines (P<0.05). These two compounds also concentration-dependently diminished Na(+)-K(+)-ATPase activity and VEGF level in four test cell lines (P<0.05). Besides Huh7 cell, OA or UA treatments concentration-dependently elevated caspase-3 and caspase-8 activities in other three cell lines (P<0.05). Besides HA22T cell, these two compounds concentration-dependently inhibited cell adhesion and decreased ICAM-1 level in other three cell lines (P<0.05). These findings support that OA and UA are potent anti-cancer agents to cause apoptosis in these liver cancer cell lines., (Copyright (c) 2009 Elsevier Ltd. All rights reserved.)