Caspase-3 Activity Levels Research Articles

Sensitivity of Primary Human Glioblastoma Cell Lines to Mumps Virus Vaccine Strain

The sensitivity of human glioblastoma cells to virus-mediated oncolysis was investigated on five patient-derived cell lines. Primary glioblastoma cells (Gbl13n, Gbl16n, Gbl17n, Gbl25n, and Gbl27n) were infected with 10-fold serial dilutions of the Leningrad-3 strain of mumps virus, virus reproduction and cytotoxicity were monitored for 96–120 hours. Immortalized human non-tumor NKE cells were used as controls to determine virus specificity. Four out of the five glioblastoma cell lines examined were susceptible to mumps virus infection, whereas no virus reproduction was observed in the non-tumor cell line. Moreover, the level of proapoptotic caspase-3 activity was increased in all infected cells 48 hours after infection. The kinetics of viral RNA accumulation in the studied glioblastoma cell lines was comparable with the rate of cell death. The data suggest that glioblastoma cell lines are permissive for mumps virus. Glioblastoma cell lines differed in type I IFN production in response to mumps virus infection. In addition, it was shown that MV infection was able to induce immunogenic death of glioblastoma cells.

Neuroprotective effect of red dragon fruit extract ameliorates oxidative stress and inflammation in D-galactose-induced aging rat model: biochemical, histological and immunohistochemical study.

Aging is a worldwide socioeconomic burden. Cerebellar aging is an enigma contributing to many behavioral aging disorders, hence is its hindering by prophylactic measurements is a crucial geriatric research target. Red dragon fruit (RDF) is a tropical fruit with antioxidant, anti-inflammatory and anti-apoptotic properties. This study aimed to determine the protective effect of RDF extract against cerebellar aging. Thirty-two male albino rats were randomly allocated into 4 groups: Control, RDF, aged and RDF-aged groups. Aged group revealed structural distortion affecting cerebellar layers including a significant (P < 0.05) decrease in Purkinje cells number and decrease in granular cell layer thickness by comparison to the control and RDF groups. Additionally, distorted capillary endothelium, and defective myelination were noticed. Interestingly, cerebellar active caspase-3, iNOS, MDA and 3-NT and serum TNF-α levels significantly increased with aging by comparison to the control and RDF groups (all P < 0.05). Biochemical analysis revealed a significant (P < 0.05) decrease in cerebellar SOD and serum GSH levels in aged rats. RDF extract remarkably ameliorated most of the neuronal degenerative changes with a significant (P < 0.05) increase in Purkinje cells numbers, and granular cell layer thickness by comparison to the aged group. Furthermore, it resulted in a significant (P < 0.05) decrease in cerebellum expression of active caspase-3, iNOS, MDA, 3-NT, and serum TNF-α levels associated with a significant (P < 0.05) increase in cerebellar SOD and serum GSH levels by comparison to the aged group. To the best of our knowledge this is the first study showing a neuroprotective effect for RDF against cerebellar aging. RDF might be effective in attenuation of age-induced cerebellar degenerative changes through its anti-apoptotic, antioxidant and anti-inflammatory effects.

Comparative study of cytotoxic Signaling pathways in H1299 cells exposed to alternative Bisphenols: BPA, BPF, and BPS.

Bisphenols are prevalent in food, plastics, consumer goods, and industrial products. Bisphenol A (BPA) and its substitutes, bisphenol F (BPF) and bisphenol S (BPS), are known to act as estrogen mimics, leading to reproductive disorders, disruptions in fat metabolism, and abnormalities in brain development. Despite numerous studies exploring the adverse effects of bisphenols both in vitro and in vivo, the molecular mechanisms by which these compounds affect lung cells remain poorly understood. This study aims to compare the effects of BPA, BPF, and BPS on the physiological behavior of human nonsmall cell lung cancer (NSCLC) cells. Human non-small cell lung cancer (NSCLC) H1299 cells were treated with various concentration of BPA, BPF and BPS during different exposure time. Cellular physiology for viability and cell cycle was assessed by the staining with apoptotic cell makers such as active Caspase-3 and cyclins antibodies. Toxicological effect was quantitatively counted by using flow-cytometry analysis. Our findings indicate that BPA induces apoptosis by increasing active Caspase-3 levels in H1299 cells, whereas BPF and BPS do not promote late apoptosis. Additionally, BPA was found to upregulate cyclin B1, causing cell cycle arrest at the G0/G1 phase and leading to apoptotic cell death through Caspase-3 activation. Conclusion: These results demonstrate that BPA, BPF, and BPS differentially impact cell viability, cell cycle progression, and cell death in human NSCLC cells.

Herbo-vitamin medicine Livogrit Vital ameliorates isoniazid induced liver injury (IILI) in human liver (HepG2) cells by decreasing isoniazid accumulation and oxidative stress driven hepatotoxicity

BackgroundTuberculosis (TB) is a leading cause of infection related mortality. Isoniazid is one of the frontline drugs for anti-TB therapy. Hepatotoxicity induced by isoniazid is a major cause of drug-discontinuation which may lead to development of resistant TB or increased mortality.PurposeTo characterize pharmacological properties of plant-based prescription medicine, Livogrit Vital (LVV) against isoniazid-induced liver injury (IILI) using HepG2 cells.MethodPhytometabolite characterization of LVV was performed by High-performance liquid chromatography (HPLC). The effects of LVV on cytosafety, IC50 shift, oxidative stress, ER stress, apoptosis, liver injury markers, and accumulation of isoniazid and hydrazine was performed on HepG2 cells induced with isoniazid. Silymarin was used as the positive control.ResultsHPLC based phytometabolite characterization of LVV revealed the presence of several anti-oxidant, anti-apoptotic, and hepatoprotective compounds. In isoniazid-induced HepG2 cells, LVV reduced cytotoxicity of isoniazid and shifted its IC50 value. Treatment with LVV reduced ROS generation and lipid peroxidation; enhanced GSH enzyme levels in isoniazid-induced HepG2 cells. As per the mechanistic evaluation, LVV modulated gene expression level of Caspase-3, FGF21, and IRE-1α. LVV treatment also normalized isoniazid-induced elevated Caspase-3 activity and cPARP1 protein levels, indicating its potentials to regulate liver cell apoptosis. Concomitantly, biomarkers of hepatotoxicity, ALT and GGT, also decreased by LVV treatment. Interestingly, LVV treatment reduced intracellular accumulation of isoniazid and its toxic metabolite hydrazine, in isoniazid-stimulated HepG2 cells.ConclusionTreatment of hepatic cells with the herbo-vitamin medicine, Livogrit Vital, regulates IILI by modulation of oxidative and ER stress, apoptosis, and bioaccumulation of isoniazid and hydrazine. Collectively, Livogrit Vital could well be explored as an adjuvant hepatoprotective agent alongwith anti-TB medicines.

Role of MMP-2 and MMP-9 in the Relationship between Inflammation, Fibrosis, and Apoptosis during Progression of Non-Alcoholic Fatty Liver Disease and Diagnostic Significance of Plasma Levels of Their Active Forms.

MMP-2 and MMP-9 play an important role in pathogenesis of chronic liver diseases, participating in the processes of inflammation and fibrosis. Their role in progression of non-alcoholic fatty liver disease (NAFLD) is poorly understood. Analysis of MMP-2, -9 levels in the blood plasma of patients with different forms of NAFLD [liver steatosis (LS) and non-alcoholic steatohepatitis (NASH) of weak (-WA), moderate (MA), high(-HA) activity without pronounced fibrosis] was performed. Correlations between the levels of MMP-2,-9 and mRNA of the genes MMP2, MMP9, ADAM17, NLRP3, caspase3 activity in peripheral blood leukocytes (PBL), TNFα, IL-6, sIL-6R, cytokeratin-18 fragments in plasma were assessed. In steatosis, the levels of MMP2 gene mRNA in PBL and MMP-2 in plasma are lower than in the control, and expression of the NLRP3 gene in PBL is increased relative to other groups. In the NASH-WA, the level of MMP-9 is higher than in the control, in LS, and in NASH-MA, which could be associated with activation of inflammation during transformation of LS into NASH. The plasma level of MMP-9 over 389.50pg/ml has been shown to be diagnostically significant for identification of NASH-WA among the patients with steatosis (AUCROC=0.818, 95% CI=0.689-0.948, p&lt;0.001). InNAFLD, the level of MMP-9 could be associated not only with inflammation, but also with apoptosis. ADAM17 probably plays a certain role in this regard. In the advanced NASH, hepatocyte apoptosis is increased, the level of caspase3 activity in PBL is increased, the level of MMP-9 in the blood is reduced to the level of the control and LS. In the NASH-HA, the level of mRNA of the ADAM17 gene in PBL is increased compared to the control, NASH-WA, and NASH-MA. Thus, MMP-2 and MMP-9 are involved in pathogenesis of NAFLD already at the early stages and their level in blood could be associated with the presence and severity of inflammation in the liver parenchyma.

Prolonged Survival of Neutrophils Induced by Tumor-Derived G-CSF/GM-CSF Promotes Immunosuppression and Progression in Laryngeal Squamous Cell Carcinoma.

Tumor-associated neutrophils (TANs) play a crucial role in tumor progression and exhibit prolonged survival. However, the mechanism underlying their extended lifespan and significance in laryngeal squamous cell carcinoma (LSCC) remains unclear. Herein, it is observed that apoptosis of TANs is significantly delayed owing to induction by tumor-derived G-CSF and GM-CSF through the activation of the PI3K-AKT signaling pathway, upregulation of anti-apoptotic Mcl-1 expression, and downregulation of activated Caspase-3 levels. It is found that prolonged survival of TANs leads to the accumulation of aged CXCR4+ neutrophils that exhibit potent immunosuppressive properties and are associated with poor patient prognosis. Furthermore, extended survival promotes the enhanced immunosuppressive function of CD8+ T cells by TANs, thereby facilitating the in vitro and in vivo progression and growth of human LSCC tumors. Importantly, this effect could be reversed by blocking G-CSF and GM-CSF stimulation of neutrophils. These findings elucidate the pivotal role of pathologically prolonged neutrophil survival in impairing CD8+ T cell immunity and suggest targeting it as a potential therapeutic strategy for tumors.

Design, synthesis and mechanistic study of N-4-Piperazinyl Butyryl Thiazolidinedione derivatives of ciprofloxacin with Anticancer Activity via Topoisomerase I/II inhibition

A new group of thiazolidine-2,4-dione derivatives of ciprofloxacin having butyryl linker 3a-l was synthesized via an alkylation of thiazolidine-2,4-diones with butyryl ciprofloxacin with yield range 48–77% andfully characterized by various spectroscopic and analytical tools. Anti-cancer screening outcomes indicated that 3a and 3i possess antiproliferative activities against human melanoma LOX IMVI cancer cell line with IC50 values of 26.7 ± 1.50 and 25.4 ± 1.43 µM, respectively, using doxorubicin and cisplatin as positive controls with an IC50 of 7.03 ± 0.40 and 5.07 ± 0.29 µM, respectively. Additionally, compound 3j showed promising anticancer activity against human renal cancer A498 cell line with IC50 value of 33.9 ± 1.91 µM while doxorubicin and cisplatin showed IC50 values of 3.59 ± 0.20 and 7.92 ± 0.45, respectively. On the other hand, compound 3i did not show considerable anti-bacterial activity against S. aureus, E. coli and P. aeruginosa, and only moderate activity against K. pneumoniae with only a tenth of the activity of ciprofloxacin, confirming the cytotoxicity observed. Mechanistically, compound 3i inhibited both topoisomerase I and II with IC50 of 4.77 ± 0.26 and 15 ± 0.81 µM. Furthermore, it induced cell cycle arrest at S phase in melanoma LOX IMVI cells. Moreover, 3i provoked substantial levels of early, late apoptosis and necrosis in melanoma LOX IMVI cell line comparable to that induced by doxorubicin. Furthermore, compound 3i increased the expression level of active caspase-3 by 49 folds higher in LOX IMVI cell, increased protein expression level of Bax more than the control by 3 folds and inhibited PARP-1by 33% in LOX IMVI. All results were supported by theoretical docking studies on both tested enzymes confirming potential cytotoxicity for the synthesized hybrids.

Effect of Tualang Honey-Mediated Silver Nanoparticles on TNF-α level, Caspase-3 Activity and Hippocampal Morphology in Kainic Acid-Induced Neurodegeneration in Male Rats

INTRODUCTION: Despite being common disorder, the curative treatment for degenerative diseases are not yet available. Although Tualang honey (TH) has been reported to protect against neurodegeneration, but the effect of TH-mediated silver nanoparticles (THSN) on neurodegeneration is poorly understood. Thus, we conducted this study aimed to determine the effects of THSN on the levels of tumour necrosis factor alpha (TNF-α), caspase-3 activity, and hippocampal morphology in Kainic Acid (KA) induced neurodegeneration in rats. MATERIALS AND METHODS: A total of 72 Male Sprague Dawley rats were randomized into six groups which were the control, THSN 10mg, THSN 50 mg, KA only, KA+THSN 10 mg, and KA+THSN 50 mg groups. Each group was pre-treated orally with either distilled water or THSN (10 mg/kg or 50 mg/kg), according to their respective group. Following the last pre-treatment, each rat was injected with KA (15 mg/kg) or saline. After 24 h and 5 days of KA induction, all rats were sacrificed, and the hippocampus from each rat was harvested. Cresyl Violet and Fluoro Jade C staining were carried out to examine the number of viable cells and degenerating neurons. TNF-α level and caspase-3 activity in the hippocampus were measured using commercially available ELISA kits. RESULTS: Rats with KA-induced neurodegeneration demonstrated a significant increase (p&lt;0.05) of TNF-α level and caspase-3 activity with a lower number of viable cells and increased number of degenerating neurons in the hippocampus. The pre-treatments of THSN groups improved these changes by lowering the TNF-α level and caspase-3 activity and decreasing the number of degenerating neurons. CONCLUSION: THSN could have potential neuroprotective effects in ameliorating TNF-α level, caspase-3 activity, and hippocampal damage in KA-induced male rats.

Randomized clinical trial of astaxanthin supplement on serum inflammatory markers and ER stress-apoptosis gene expression in PBMCs of women with PCOS.

Polycystic ovarian syndrome (PCOS) is related to pro-apoptotic and pro-inflammatory conditions generated by Endoplasmic reticulum (ER) stress. This study aimed to determine the effect of Astaxanthin (ASX), as carotenoid with potent antioxidant and anti-inflammatory properties, on serum inflammatory markers, apoptotic factors and ER stress-apoptotic genes in peripheral blood mononuclear cells (PBMCs) of women with PCOS. This randomized, double-blind clinical trial included 56 PCOS patients aged 18-40. For 8 weeks, subjects were randomly assigned to one of two groups: either 12 mg ASX (n = 28) or placebo (n = 28). Real-time PCR was used to quantify gene expression associated with ER stress-apoptosis in PCOS women's PBMCs. The levels of TNF-α, IL18, IL6 and CRP were determined by obtaining blood samples from all patients before and after the intervention using Enzyme-linked immunosorbent assay (ELISA). Also, the levels of active caspase-3 and caspase-8 were detected in the PBMC by ELISA kit. Furthermore, we evaluated the efficacy of ASX on disease symptoms. Following the 8-week intervention, ASX supplementation was able to reduce the expression of GRP78 (p = 0.051), CHOP (p = 0.008), XBP1 (p = 0.002), ATF4 (0.038), ATF6 (0.157) and DR5 (0.016) when compared to the placebo. However, this decrease was not statistically significant for ATF6 (p = 0.067) and marginally significant for GRP78 (p = 0.051). The levels of TNF-α (p = 0.009), IL-18 (p = 0.003), IL-6 (p = 0.013) and active caspase-3 (p = 0.012) were also statistically significant lower in the therapy group. However, there was no significant difference in CRP (p = 0.177) and caspase-8 (p = 0.491) levels between the treatment and control groups. In our study, ASX had no significant positive effect on BMI, hirsutism, hair loss and regularity of the menstrual cycle. It appears that ASX may benefit PCOS by changing the ER stress-apoptotic pathway and reducing serum inflammatory markers; however, additional research is required to determine this compound's potential relevance.

Morin attenuates arsenic-induced toxicity in 3T3 embryonic fibroblast cells by suppressing oxidative stress, inflammation, and apoptosis: In vitro and silico evaluations.

This study aims to investigate the curative effects of Morin, a flavonoid, against arsenic toxicity in 3T3 embryonic fibroblast cells and its effect on the molecular mechanisms of cells. The cytotoxicity and viability of the cells were measured by MTT and LDH tests. Arsenic (0.74μM) was used to trigger toxicity and Morin (50μM) was used for treatment. The levels of oxidative stress biomarkers and the activities of antioxidant enzymes were measured by spectrophotometric method, and inflammatory markers were measured by ELISA method. While mRNA expression levels of Bax, Bcl-2 levels, and Caspase-3 activity were measured by qRT-PCR technique, TUNEL staining was performed to detect DNA breaks and DAPI staining to visualize nuclear changes. Protein structures were retrieved from the protein data bank. OpenBabel and Autodock programs were used for the molecular docking study. Morin rescued the 3T3 embryonic fibroblast cells exposed to arsenic. However, Arsenic decreased the activities of antioxidant enzymes in cells and significantly increased oxidative stress, inflammation, and apoptosis. Morin treatment reduced oxidative damage and TNF-α and IL-1β levels. Arsenic-induced Caspase-3 mRNA expression level and Bax protein mRNA expression level were significantly increased, while Bcl-2 mRNA expression level was significantly decreased. While Caspase-3 mRNA expression level and Bax protein mRNA expression level decreased with morin treatment, Bcl-2 mRNA expression level increased significantly. Molecular docking study results showed good binding affinity of morin in SOD, GSH-Px, Bax, Bcl-2, Caspase-3, TNF-α, and IL-1β structures. Morin showed antioxidant, anti-inflammatory, and anti-apoptotic effects against Arsenic-induced cellular toxicity.

Emetine induces oxidative stress, cell differentiation and NF-κB inhibition, suppressing AML stem/progenitor cells

Acute myeloid leukemia (AML) is a fatal malignancy of the blood and bone marrow. Leukemic stem cells (LSCs) are a rare subset of leukemic cells that promote the development and progression of AML, and eradication of LSCs is critical for effective control of this disease. Emetine is an FDA-approved antiparasitic drug with antitumor properties; however, little is known about its potential against LSCs. Herein, we explored the antileukemic potential of emetine, focusing on its effects on AML stem/progenitor cells. Emetine exhibited potent cytotoxic activity both in hematologic and solid cancer cells and induced AML cell differentiation. Emetine also inhibited AML stem/progenitor cells, as evidenced by decreased expression of CD34, CD97, CD99, and CD123 in KG-1a cells, indicating anti-AML stem/progenitor cell activities. The administration of emetine at a dosage of 10 mg/kg for two weeks showed no significant toxicity and significantly reduced xenograft leukemic growth in vivo. NF-κB activation was reduced in emetine-treated KG-1a cells, as shown by reduced phospho-NF-κB p65 (S529) and nuclear NF-κB p65. DNA fragmentation, YO-PRO-1 staining, mitochondrial depolarization and increased levels of active caspase-3 and cleaved PARP (Asp214) were detected in emetine-treated KG-1a cells. Moreover, treatment with the pancaspase inhibitor Z-VAD(OMe)-FMK partially prevented the apoptotic cell death induced by emetine. Emetine treatment also increased cellular and mitochondrial reactive oxygen species, and emetine-induced apoptosis in KG-1a cells was partially prevented by the antioxidant N-acetylcysteine, indicating that emetine induces apoptosis, at least in part, by inducing oxidative stress. Overall, these studies indicate that emetine is a novel potential anti-AML agent with promising activity against stem/progenitor cells, encouraging the development of further studies aimed at its clinical application.

Comparative neuroprotective effects of royal jelly and its unique compound 10-hydroxy-2-decenoic acid on ischemia-induced inflammatory, apoptotic, epigenetic and genotoxic changes in a rat model of ischemic stroke

ABSTRACT Objectives This study aimed to compare the efficacy of royal jelly (RJ) and its major fatty acid 10-hydroxy-2-decenoic acid (10-HDA) on ischemic stroke-related pathologies using histological and molecular approaches. Methods Male rats were subjected to middle cerebral artery occlusion (MCAo) to induce ischemic stroke and were supplemented daily with either vehicle (control group), RJ or 10-HDA for 7 days starting on the day of surgery. On the eighth day, rats were sacrificed and brain tissue and blood samples were obtained to analyze brain infarct volume, DNA damage as well as apoptotic, inflammatory and epigenetic parameters. Results Both RJ and 10-HDA supplementation significantly reduced brain infarction and decreased weight loss when compared to control animals. These effects were associated with reduced levels of active caspase-3 and PARP-1 and increased levels of acetyl-histone H3 and H4. Although both RJ and 10-HDA treatments significantly increased acetyl-histone H3 levels, the effect of RJ was more potent than that of 10-HDA. RJ and 10-HDA supplementation also alleviated DNA damage by significantly reducing tail length, tail intensity and tail moment in brain tissue and peripheral lymphocytes, except for the RJ treatment which tended to reduce tail moment in lymphocytes without statistical significance. Conclusions Our findings suggest that neuroprotective effects of RJ in experimental stroke can mostly be attributed to 10-HDA.

Abstract 6002: Increased ferroptosis sensitivity and epithelial to mesenchymal transition of breast cancer cells overcoming chemotherapeutic mediated apoptotic caspase activation

Abstract The survival of cancer cells post chemotherapeutic treatment can lead to the presence of recurrent tumors and continues to be a barrier to effective cancer treatment. The majority of chemotherapeutics kill cells through the induction of executioner caspases and subsequent apoptotic death, and resistance to apoptosis can lead to the presence of anastatic cancer cells. Executioner caspase release was previously thought to be the point of no return from apoptotic cell death, however, it has been shown that removal of the reagent causing caspase release can lead to recovery of cells from apoptotic signaling, a phenomenon which has been termed ``Anastasis``. This presents a need for a better understanding of the underlying mechanisms behind cancer cell survival and the mechanisms that lead to cell recovery and recurrent tumors. We have hypothesized that when treated with chemotherapeutics, some cells will evade cell death even when executioner caspases have been activated. By identifying surviving anastatic cells and the pathways involved in their evasion of cell death, we hope to propose novel therapeutic strategies to prevent their survival. The novel CasExpress system, developed by the Denise Montell lab group, was used to identify and isolate a population of cells surviving caspase-3 activation as a result of chemotherapeutic treatment in Triple Negative Breast Cancer (TNBC) cell lines. The CasExpress system permanently labels these cells with GFP post caspase-3 activation. Analysis of this population of SUM159 anastatic cells, identified by their GFP expression, demonstrates that these cells have an increased resistance to further chemotherapeutic treatment, a decrease in levels of active caspase-3, an up-regulation of Cancer Stem Cell marker CD44, and a more mesenchymal phenotype. Furthermore, these “anastatic cells” express decreased levels of GPX-4, an enzyme that mitigates ferroptosis via lipid peroxide reduction, and are more sensitive to GPX-4 and xCT inhibitor mediated ferroptosis cell death.To investigate the link between Epithelial to Mesenchymal Transition (EMT) and ferroptosis mediated cell death, we used TGF-b to stimulate EMT in NMe mouse epithelial cells. We found that TGF-b induced EMT is associated with increased sensitivity to ferroptosis mediated by GPX-4 inhibition. Furthermore, the TGF-b induced EMT cells have a marked decrease in the expression of GPX4, similar to what was observed in the TNBC anastatic cells. These results indicate a potential link between EMT and an increased sensitivity to ferroptosis, and provide novel strategies for identifying and targeting chemotherapeutic resistant tumor cells. Citation Format: Rachel Hausman, Wells Brown, Paul McDonald, Shannon Awrey, Gongping Sun, Denise Montell, Shoukat Dedhar. Increased ferroptosis sensitivity and epithelial to mesenchymal transition of breast cancer cells overcoming chemotherapeutic mediated apoptotic caspase activation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6002.

DMC triggers MDA-MB-231 cells apoptosis via inhibiting protective autophagy and PI3K/AKT/mTOR pathway by enhancing ROS level

DMC, a kind of compound derived from the dry flower buds of Cleistocalyx operculatus, has been shown to inhibit the growth of various cancer cells, but research on triple-negative breast cancer cells remains scarce. To explore this issue, MDA-MB-231 cells were selected, and the results showed that DMC has strong proliferation inhibit effects on this kind of cells. The inhibit rate of 30 μM DMC incubated for 24 h was 56.25%, and 40.6% cells were arrested under the G2/M phase. The levels of pro-apoptosis protein Bax and active caspase-3, cleaved PARP and cell cycle related proteins, such as p21 and p27 increased, but apoptosis regulators, like Bcl-2, Cdc 2, Cyclin B1, and LC3 II decreased dramatically. In addition, DMC induced the accumulation of autophagosomes and autophagic substrates, and the combination of DMC with CQ promoted apoptosis of MDA-MB-231 cells, which suggested that DMC induced apoptosis partly by blocking autophagy flow.Moreover, the phosphorylation levels of phosphatidylinositol 3-kinase (PI3K), protein kinase B (AKT), and its mechanistic target of rapamycin kinase (mTOR) were also decreased after 30 μM DMC incubating for 24 h. The proteins play a critical role in cell proliferation, apoptosis, and autophagy modulation. The inhibition of autophagy flow and PI3K/AKT/mTOR pathway could be reversed after being treated with ROS scavenger NAC. Altogether, the results of the present study suggest that DMC effectively induces apoptosis and growth inhibition in MDA-MB-231 cells through blocking autophagy flow and regulating the PI3K/AKT/mTOR pathway by increasing ROS level.

A novel approach of using Maca root as a radioprotector in a rat testicular damage model focusing on GRP78/CHOP/Caspase-3 pathway

PurposeDespite the effectiveness of ionizing radiation in treating cancer, it can damage healthy tissues in the vicinity. Due to the high radio-sensitivity of testicular tissues, radiation therapy may affect spermatogenesis, which may result in infertility. Hence, in this study testicular damage model is constructed to investigate the mitigation effect of Maca root powder and its potential radioprotective activity through both oxidative and endoplasmic reticulum (ER) stresses, besides the apoptotic pathway. MethodsMale albino rats were exposed to 6Gy of whole-body gamma radiation single dose. Maca root powder (1 g/kg b.wt./day, by oral gavage) was administered for a week before irradiation, then d-galactose (300 mg/kg, by oral gavage) and Maca daily for another week. ResultsGamma radiation and d-galactose revealed a significant decrease in serum testosterone, sperm count, and motility and higher percentage of the sperm head abnormality, while Maca root treatment maintained all sperm morphology parameters. Maca root treatment demonstrated a notable defense against radiation-induced oxidative stress and ameliorated malonaldehyde (MDA), reactive oxygen species (ROS), nitric oxide (NO), glutathione-S-transferase (GST) levels, reduced glutathione (GSH), oxidized glutathione (GSSG) and the ratio of GSH/GSSG in testis tissues. Exposure to gamma rays and d-galactose displayed a significant elevation in GRP78, CHOP, total caspase-3 as well as active (cleaved) caspase-3 levels, whereas treatment with Maca significantly reduced the ER and apoptotic markers levels. Also, Maca improved the histological changes of the disorganized seminiferous tubules induced by irradiation. ConclusionOur findings show for the first time that Maca has a protective effect on male reproductive damage induced by radiotherapy. Maca root reveals anti-apoptotic effect and protection against testicular damage via GRP78/CHOP/caspase-3 pathway.

Sensitivity of Primary Human Glioblastoma Cell Lines to the Mumps Virus Vaccine Strain

The sensitivity of human glioblastoma cells to virus-mediated oncolysis was investigated on five patient-derived cell lines. Primary glioblastoma cells (Gbl13n, Gbl16n, Gbl17n, Gbl25n, and Gbl27n) were infected with tenfold serial dilutions of the Leningrad-3 strain of the mumps virus, and virus reproduction and cytotoxicity were monitored for 96-120 h. Immortalized human non-tumor NKE cells were used as controls to determine the virus specificity. Four out of the five glioblastoma cell lines examined were susceptible to mumps virus infection, whereas no virus reproduction was observed in the non-tumor cell line. Moreover, the level of proapoptotic caspase-3 activity was increased in all infected cells 48 h after infection. The kinetics of viral RNA accumulation in the studied glioblastoma cell lines was comparable with the rate of cell death. The data suggest that glioblastoma cell lines were permissive for the mumps virus. Glioblastoma cell lines differed in type I IFN production in response to the mumps virus infection. In addition, it was shown that MV infection was able to induce immunogenic death of glioblastoma cells.

Antiproliferative and Apoptotic Effects of Tempol, Methotrexate, and Their Combinations on the MCF7 Breast Cancer Cell Line.

Breast cancer holds the top position among the cancers occurring in women. Despite the utilization of surgical removal, chemotherapy, and radiation therapy, there is currently no conclusive treatment available to prevent breast cancer. New treatment approaches are being studied since traditional chemotherapeutics also damage healthy cells. Tempol (TPL) is a potent antioxidant agent that has been shown to exhibit anticancer activity. The objective of this research was to examine the impacts on cell proliferation and apoptosis by using methotrexate (MTX) and TPL individually and in combination on MCF7 breast cancer cells. MCF7 cells were exposed to TPL, MTX, and MTX + TPL for 48 h. The effects of the administered drugs on cell viability were determined using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Enzyme-linked immunosorbent assay analysis was conducted to assess the levels of the antiapoptotic protein Bcl-2, the pro-apoptotic protein Bax, and the activity of caspase-3 in MCF7 cells. Increasing concentrations of TPL and MTX significantly decreased the proliferation in MCF7 cells in both solo and combined use. Solo and combined use of TPL and MTX significantly increased caspase-3 activity and Bax levels and significantly decreased Bcl-2 levels in the cells. This study revealed that the solo use of TPL and MTX inhibited proliferation and increased apoptotic activity in the cells. In addition, TPL increased the antiproliferative and apoptosis efficiency of MTX on cancer cells as a result of the combined use of these drugs.

Diosgenin potentiates the anticancer effect of doxorubicin and volasertib via regulating polo-like kinase 1 and triggering apoptosis in hepatocellular carcinoma cells.

A common approach to cancer therapy is the combination of a natural product with chemotherapy to overcome sustained cell proliferation and chemotherapy resistance obstacles. Diosgenin (DG) is a phytosteroidal saponin that is naturally present in a vast number of plants and has been shown to exert anti-cancer activities against several tumor cells. Herein, we assessed the chemo-modulatory effects of DG on volasertib (Vola) as apolo-like kinase 1 (PLK1) inhibitor and doxorubicin (DOX) in hepatocellular carcinoma (HCC) cell lines. DOX and Vola were applied to two human HCC cell lines (HepG2 and Huh-7) alone or in combination with DG. The cell viability was determined, and gene expressions of PLK1, PCNA, P53, caspase-3, and PARP1 were evaluated by RT-qPCR. Moreover, apoptosis induction was determined by measuring active caspase-3 level using ELISA method. DG enhanced the anticancer effects of Vola and DOX. Moreover, DG enhanced Vola- and DOX-induced cell death by downregulating the expressions of PLK1 and PCNA, elevating the expressions of P53 and active caspase-3. DG showed promising chemo-modulatory effects to Vola and DOX against HCC that may be attributed partly to the downregulation of PLK1 and PCNA, upregulation of tumor suppressor protein P53, and apoptosis induction. Thus,DG combination with chemotherapy may be a promising treatment approach for HCC.

Chemopreventive effects of α-tocopherol and its long-chain metabolites α-13'-hydroxy- and α-13'-carboxychromanol in LT97 colon adenoma cells.

Anticancer effects of vitamin E (tocopherols) have been studied extensively. While in vitro and animal studies showed promising results regarding anticancer effects of tocopherols, human intervention studies failed to reproduce these results. In vivo, α-tocopherol (α-TOH) is metabolized to the long-chain metabolites (LCM) 13'-hydroxychromanol (α-13'-OH) and 13'-carboxychromanol (α-13'-COOH), which likely reach the large intestine. The LCM showed antiproliferative effects in different colon cancer cell lines, but the exact mechanism of action remains unclear. To further clarify the chemopreventive action of the LCM, premalignant LT97 colon adenoma cells were treated with α-TOH, α-13'-OH and α-13'-COOH to study their impact on growth, apoptosis, antigenotoxicity, and ROS-scavenging capacity as well as expression of selected genes involved in detoxification and the cell cycle. Growth inhibitory potential was observed for α-13'-OH (IC50: 37.4 μM) and α-13'-COOH (IC50: 5.8 μM) but not for α-TOH in the tested concentrations. Levels of caspase-3 activity and expression of genes regulating the cell cycle and detoxification remained unchanged. However, α-TOH, α-13'-OH and α-13'-COOH exhibited antigenotoxic and partly ROS-scavenging capacity. The results indicate that the LCM exert chemopreventive effects via ROS-scavenging capacity, the protection against DNA damage and the induction of cell death via caspase-independent mechanisms in premalignant colon cells.

Levofloxacin reposition-based design: synthesis, biological evaluation of new levofloxacin derivatives targeting topoisomerase II beta polymerase as promising anticancer agents, molecular docking, and physicochemical characterization.

Repositioning of already approved medications through repurposing or re-profiling for new medical uses after certain structural modifications is a novel approach in drug discovery. Fluoroquinolone antibiotics are one of the cardinal agents investigated for potential anticancer activities. In this work, levofloxacin was repositioned for anticancer activities. A series of levofloxacin-based compounds were designed and synthesized through the derivatization of levofloxacin's carboxylic acid functionality. The newly synthesized compounds were screened for cytotoxic activities against breast, liver, and leukemia cancer cell lines. Their effect on normal cells was also investigated. The target compounds were evaluated for their proliferative inhibitory activity toward topoisomerase II beta polymerization. Compound 5 showed higher inhibitory activity against a breast cancer cell line (MCF-7) with IC50 = 1.4 μM and lesser side effects on a normal breast cell line (MCF-10a) with IC50 = 30.40 μM than reference drugs. The best activity against a liver cancer cell line (Hep3B) was exhibited by compounds 3c, 4b, 5, 7, 8, 13a and 13c with IC50 values ranging from 0.43 to 8.79 μM. Regarding the effect of compounds 5 and 13a on a leukemia cancer cell line (L-SR), their IC50 values were 0.96 and 3.12 μM, respectively. Compounds 3c and 5 showed Topo2β inhibitory effects on Hep3B cells (81.33% and 83.73%, respectively), which was better than levofloxacin and etoposide as reference drugs. Cytometry cell cycle analysis revealed that compounds 3c and 5 arrested the cell cycle at the S phase (37.56% and 39.09%, respectively). Compounds 3c and 5 exhibited an elevation in active caspase-3 levels by 4.9 and 4.5 folds, respectively. Molecular modeling simulation of compounds 3c and 5 demonstrated energy scores (-29.77 and -20.46 kcal mol-1, respectively) more than those of the reference drugs as they interact with the most essential amino acids required for good affinity towards human topoisomerase II beta enzyme (PDB ID: 3QX3). Physicochemical characteristics of the most promising cytotoxic compounds 3c and 5 were investigated and compared to etoposide and levofloxacin as reference drugs. However, they showed high gastrointestinal absorption and could not penetrate the blood-brain barrier.