CASP8 Genes Research Articles

Antioxidant, cytotoxic, anti-migratory, and pro-apoptotic effects of Bolanthus turcicus extracts on head and neck cancer cells.

Investigation of various plant extracts using in-vitro/in-vivo assays has emerged as a promising avenue for identifying potential pharmacophores that can be developed into therapeutic drugs. This study aims to assess the bioactive compounds and antioxidant capacity of the Bolanthus turcicus (B. turcicus) and to investigate the effects on head and neck cancer (HNC) cell lines. Methanol (MeOH), ethyl acetate (EA) and aqueous (Aq) extracts were prepared from B. turcicus, and the amount of total phenolic content (TPC) and total flavonoid content (TFC) in the extracts were analyzed by the Folin-Ciocalteu and Aluminum chloride method, respectively. In addition, the total antioxidant capacity and iron reducing potential of B. turcicus extracts were determined by the Phosphomolybdenum and Ferric ion reducing antioxidant power (FRAP) method. The effect of B. turcicus on HEp-2, SCC-90, SCC-9, FaDu HNC cell viability, motility, and cell-nuclear morphology was evaluated by MTT, scratch-wound healing assay, and Pllalloidin-DAPI staining, respectively. The effect of B. turcicus on the expression of CASP-3, BAX, and BCL-2 genes at the mRNA, protein, and intracellular level was evaluated by quantitative PCR (qPCR), western blot, and immunofluorescence staining. Moreover, Annexin V-FITC/PI, was used in flow cytometry to investigate the effect of B. turcicus on apoptosis. The MeOH extract exhibited the highest phenolic content, flavonoid content and antioxidant activity (p < 0.05 for all). HNC cells treated with extracts indicated delayed wound healing and decreased motility (p < 0.05 for all). Analysis of annexin V-PI staining indicated that the B. turcicus extracts induced apoptosis but not viability and necrosis in the HNC cell (p < 0.05 for all). Moreover, qPCR data regarding the apoptotic mechanism showed that the extracts could induce apoptosis by upregulation of pro-apoptotic CASP-3 and BAX genes and downregulation of anti-apoptotic BCL-2 gene (p < 0.05 for all). The expression of protein and intracellular levels of CASP-3 and BAX were increased, while the BCL-2 was decreased in cells treated with the extracts (p < 0.05 for all). In addition, diffuse pycnosis and DNA condensation in HNC cell nuclei, confirming apoptotic cell death (p < 0.05 for all). This study data indicated that B. turcicus extracts have antioxidant, cytotoxic, anti-migratory and pro-apoptotic activity. In conclusion, it has been shown that B. turcicus can be used as a potential therapeutic agent against HNC.

Cardioprotective effects of cinnamoyl imidazole on apoptosis and oxidative stress in hypoxia/reoxygenation-induced H9C2 cell lines

BackgroundThis study explored the effects of cinnamoyl imidazole on alleviating oxidative stress and apoptosis in hypoxia/reoxygenation (H/R)-induced H9C2 cells, using computational analysis with in-vitro validation. MethodsComputational techniques, including SwissADME and Swiss Target Prediction, were employed to predict the ADME properties and to identify targets of cinnamoyl imidazole. Differential gene expression (DEG) analysis was conducted on myocardial infarction (MI) datasets obtained from the Gene Expression Omnibus. Gene enrichment and molecular simulation studies were done to focus on apoptotic pathways. The computational findings were validated through In vitro experiments on H9C2 cardiomyocytes subjected to 8 h of hypoxia followed by 24 h of reoxygenation. Antioxidant enzyme levels (catalase, GST, GSH-Px, and SOD), mitochondrial membrane potential (ΔΨm), caspase-3 activity, and the expression of CASP3, MAPK8, JAK2, and BCL2L1 were assessed. ResultsCinnamoyl imidazole has demonstrated favourable pharmacokinetic properties, characterized by high gastrointestinal absorption and low toxicity with negative toxicity for organ endpoints. Molecular docking studies revealed the strong binding affinities for CASP3, MAPK8, and JAK2. In vitro results showed a significant increase in cell viability (94.7 % at 10 μM, p < 0.001) and antioxidant enzyme activity, along with a 64.3 % reduction in caspase-3 activity at 1000 μM (p < 0.01). Cinnamoyl imidazole treatment preserved mitochondrial membrane potential, downregulated pro-apoptotic genes CASP3 and MAPK8, and upregulated the anti-apoptotic gene BCL2L1. ConclusionCinnamoyl imidazole effectively mitigates oxidative stress and apoptosis in H/R-induced H9C2 cells, enhancing cell viability and antioxidant defenses while maintaining mitochondrial integrity.

Effects of Nitrite Stress on the Antioxidant, Immunity, Energy Metabolism, and Microbial Community Status in the Intestine of Litopenaeus vannamei

Nitrite is the main environmental pollutant that endangers shrimp culture. Intestinal health is essential for the disease resistance of shrimp. In this study, Litopenaeus vannamei shrimps were separately exposed to 1 and 5 mg/L of nitrite stress for 48 h, and then the variations in intestinal health were investigated from the aspects of histology, antioxidant, immunity, energy metabolism, and microbial community status. The results showed that nitrite stress damaged intestinal mucosa, and 5 mg/L of nitrite induced more obvious physiological changes than 1 mg/L. Specifically, the relative expression levels of antioxidant (ROMO1, Nrf2, SOD, GPx, and HSP70), ER stress (Bip and XBP1), immunity (proPO, Crus, ALF, and Lys), inflammation (JNK and TNF-α), and apoptosis (Casp-3 and Casp-9) genes were increased. Additionally, intestinal energy metabolism was activated by inducing glucose metabolism (HK, PK, PDH, and LDH), lipid metabolism (AMPK and FAS), tricarboxylic acid cycle (MDH, CS, IDH, SDH, and FH), and electron transfer chain (NDH, CytC, COI, CCO, and AtpH) gene transcription. Further, the homeostasis of intestinal microbiota composition was also disturbed, especially the abundance of some beneficial genera (Clostridium sensu stricto 1, Faecalibacterium, Romboutsia, and Ruminococcaceae UCG-010). These results reveal that nitrite stress could damage the intestinal health of L. vannamei by destroying mucosal integrity, inducing oxidation and ER stress, interfering with physiological homeostasis and energy metabolism, and disrupting the microbial community.

The Impact of Acute Ammonia Nitrogen Stress on Serum Biochemical Indicators and Spleen Gene Expression in Juvenile Yellowfin Tuna (Thunnus albacares)

The presence of ammonia nitrogen in water has a significant impact on the serum and spleen of fish, potentially leading to changes in substances such as proteins in the serum while also causing damage to the immune function of the spleen. To investigate the effects of ammonia nitrogen (NH3-N) stress on juvenile yellowfin tuna (Thunnus albacares), this study established three NH3-N concentrations, 0, 5, and 10 mg/L, denoted as L0, L1, and L2, respectively. Serum and spleen samples were collected at 6, 24, and 36 h. The effects of different NH3-N concentrations and exposure times on the physiological status of juvenile fish were investigated by analyzing serum biochemical indices and splenic gene expression. The results indicate that in the L1 group, the serum high-density lipoprotein cholesterol (HDL-C), triglycerides (TG), complement 3 (C3) and complement 4 (C4) levels, and acid phosphatase (ACP) activity showed a trend of initially increasing and then decreasing. In the L2 group, the serum low-density lipoprotein cholesterol (LDL-C), HDL-C, and C4 levels and ACP activity also displayed an initially rising and then declining trend, while TG, C3, and creatinine (CRE) levels and alkaline phosphatase (AKP) activity showed an upward trend. In the L1 group, glutathione peroxidase 1b (GPX1b), interleukin 10 (IL-10), interleukin 6 receptor (IL-6r), and tumor necrosis factor alpha (TNF-α) gene expression levels in the spleen exhibited a trend of first increasing and then decreasing. In the L2 group, IL-10, IL-6r, tumor necrosis factor beta (TNF-β), caspase 2 (casp2), and caspase 9 (casp9) gene expression levels in the spleen also showed an initial increase followed by a decrease. When NH3-N levels are below 5 mg/L, it is recommended to limit stress exposure to no more than 36 h for the juvenile fish. For concentrations ranging from 5 to 10 mg/L, stress should be strictly controlled to within 24 h. Exposure to high NH3-N levels may affect biochemical indicators such as serum lipid metabolism, immunity, and metabolism in juvenile fish, and may damage the expression of antioxidant, immune gene, and apoptosis factors in the spleen. This study aims to deepen our understanding of the effects of NH3-N on juvenile tuna, with the goal of establishing effective water quality monitoring and management strategies. This will ensure the quality of aquaculture water, reduce the harm caused by NH3-N to juvenile yellowfin tuna, and enhance aquaculture efficiency and product quality.

Pterostilbene suppresses head and neck cancer cell proliferation via induction of apoptosis.

Head and neck cancer (HNC) is one of the most prevalent causes of death worldwide, and so discovering anticancer agents for its treatment is very important. Pterostilbene (PS) is a trans-stilbene reported to be beneficial in managing various cancers. The objective of the study was to evaluate the cytotoxic, antiproliferative, proapoptotic, and antimigrative effect of PS on HEp-2, SCC-90, SCC-9, FaDu, and Detroit-551 cell lines. MTT and live/dead assays were employed to assess cell viability, while a cell migration test was performed to evaluate wound healing capacity. The mRNA, protein, and intracellular expression levels of CASP-3, BAX, and BCL-2 genes were evaluated by real-time PCR, western blotting, and immunofluorescence staining. Annexin V-PI staining was conducted to assess the amounts of viable, apoptotic, and necrotic cells. The results revealed that PS displayed cytotoxic, antiproliferative activity in a dose-dependent manner in HNC cells by upregulating CASP-3 and BCL-2 while downregulating BCL-2 in the apoptotic pathway. The proapoptotic properties were confirmed by the annexin-V-IP results. Moreover, PS displayed a significant suppressive efficacy on the migration capacity of HNC cells. The present study provides proof that PS has the prospective to be improved as an attractive anticancer agent against HNC following advanced studies.

A Machine Learning Approach to Gene Expression in Hypertrophic Cardiomyopathy

Background/Objectives: Hypertrophic cardiomyopathy (HCM) is a common heart disorder characterized by the thickening of the heart muscle, particularly in the left ventricle, which increases the risk of cardiac complications. This study aims to analyze the expression of apoptosis-regulating genes (CASP8, CASP9, CASP3, BAX, and BCL2) in blood samples from HCM patients, to better understand their potential as biomarkers for disease progression. Methods: Quantitative real-time PCR (qPCR) was used to evaluate gene expression in blood samples from 93 HCM patients. The correlation between apoptosis-regulating genes was conducted and clinical parameters were integrated for feature importance and clustering analysis. Results: Most patients exhibited significant downregulation of CASP8, CASP9, and CASP3. In contrast, BAX expression was elevated in 71 out of 93 patients, while BCL2 was increased in 55 out of 93 patients. Correlation analysis revealed weak negative correlations between the BAX/BCL2 ratio and CASP gene expression. Conclusions: These findings suggest that reduced expression of apoptotic genes may indicate a protective cellular mechanism, which could serve as a biomarker for disease progression. Further studies are needed to investigate the potential for therapeutic modulation of these pathways to improve patient outcomes.

Expression Analysis of Caspase-3 (CASP3) Gene in Leukemia Patients Using Quantitative Polymerase Chain Reaction (qPCR) and Western Blot Techniques

CASP3 belongs to the caspases family and accounts for several critical activities, including apoptosis and other pathological and physiological processes. CASP3 also influences the multiple functions in cellular development such as homeostasis, hematopoietic growth and embryonic growth. It has significant involvement in the initiation and development of cancers including breast, colon and uterine cancers. However, molecular studies on the CASP3 gene in leukemia have not been widely conducted. The present study was designed to analyse the expression level of the CASP3 gene in leukemia patients. The results were further compared between different parameters such as types of leukemia, age, gender, occupation, WBC count and location-based. About 112 leukemia blood samples along adjacent controls were collected to analyse the CASP3 expression in this study cohort. The mRNA of each patient and control samples were isolated, quantified and converted to complementary DNA (cDNA). Relative expression of the CASP3 gene was measured using the quantitative PCR (qPCR) and Western blot analyses. Results showed that the relative expression of CASP3 was found to be significantly downregulated at both mRNA and protein levels in leukemia patients compared to normal controls. Further analysis showed that expression of CASP3 was downregulated in patients receiving the combination of chemotherapy as well as radiotherapy compared with patients receiving chemotherapy. These findings from our research showed the lower expression of CASP3 is associated with an increased risk of development and proliferation of leukemia. It can, therefore, be concluded that lower expression of the CASP3 gene may contribute to the initiation and development of leukemia.

Analysis of the Dynamics of Cognitive Impairments and Expression of Caspase Cascade Genes in Preclinical Stages of Parkinsonism Modeled Using α-Synuclein Oligomers.

The behavioral effects of α-synuclein oligomers were studied at various times after its chronic intranasal administration to 75-day-old C57BL/6J mice in comparison with the dynamics of changes in the transcriptional activity of caspases genes (Casp9, Casp8, and Casp3) in the hippocampus, frontal cortex, and cerebellum. The negative effects of α-synuclein oligomers on exploratory activity and short-term memory in the novel object recognition test were most pronounced after 90 days from the end of administration, while after 1 and 270 days, partial compensation of the studied cognitive functions was observed. Analysis of the expression of caspase genes suggests that early compensatory mechanisms are associated with suppression of the effector caspase-3 gene expression along with increased activity of the genes encoding initiator caspases-9 and -8. Late compensation processes are associated with a decrease in the activity of initiator caspases in the frontal cortex and cerebellum.

An Organic Fraction of Oenothera rosea L'Her Ex. Aiton Prevents Neuroinflammation in a Rat Ischemic Model.

Oenothera rosea L'Her Ex. Aiton, presenting antioxidant and anti-inflammatory activities, is traditionally used to treat bruises and headaches and as a healing agent. This study aimed to investigate whether its organic fraction (EAOr) has neuroprotective properties against neuroinflammation in the context of ischemia/reperfusion. The chemical composition of EAOr was determined using HPLC techniques, and its neuroprotective activities were evaluated in a common carotid-artery ligation model for the induction of ischemia/reperfusion (I/R). The animals were supplemented with EAOR for 15 days. On the last day, the animals were rested for one hour, following which the common carotid-artery ligation procedure was performed to induce I/R. The neurological deficit was evaluated at 24 h after I/R using Bederson's scale, and the relative expression of inflammatory genes and structure of hippocampal neurons were analyzed at 48 h. The chemical analysis revealed five major compounds in EAOr: gallic acid, rutin, ellagic acid, and glucoside and rhamnoside quercetin. EAOr prevented neurological deficit 24 h after I/R; led to the early activation of the AIF and GFAP genes; reduced Nfkb1, IL-1beta, Il-6 and Casp3 gene expression; and protected hippocampal neurons. Our findings demonstrate that EAOr contains polyphenol-type compounds, which could exert a therapeutic effect through the inhibition of neuroinflammation and neuronal death genes, thus maintaining hippocampal neurons.

Multi-regional genomic and transcriptomic characterization of a melanoma-associated oral cavity cancer provide evidence for CASP8 alteration-mediated field cancerization

BackgroundPrecancerous and malignant tumours arise within the oral cavity from a predisposed “field” of epithelial cells upon exposure to carcinogenic stimulus. This phenomenon is known as “Field Cancerization”. The molecular genomic and transcriptomic alterations that lead to field cancerization and tumour progression is unknown in Indian Oral squamous cell carcinoma (OSCC) patients.MethodsWe have performed whole exome sequencing, copy-number variation array and whole transcriptome sequencing from five tumours and dysplastic lesions (sampled from distinct anatomical subsites - one each from buccal anterior and posterior alveolus, dorsum of tongue–mucosal melanoma, lip and left buccal mucosa) and blood from a rare OSCC patient with field cancerization.ResultsA missense CASP8 gene mutation (p.S375F) was observed to be the initiating event in oral tumour field development. APOBEC mutation signatures, arm-level copy number alterations, depletion of CD8 + T cells and activated NK cells and enrichment of pro-inflammatory mast cells were features of early-originating tumours. Pharmacological inhibition of CASP8 protein in a CASP8-wild type OSCC cell line showed enhanced levels of cellular migration and viability.ConclusionCASP8 alterations are the earliest driving events in oral field carcinogenesis, whereas additional somatic mutational, copy number and transcriptomic alterations ultimately lead to OSCC tumour formation and progression.

Unraveling the immune response of the spleen in sepsis using green-synthesized silver nanoparticles from pomegranate peel extracts.

Sepsis is a serious disease characterized by an inappropriate host response to infection, resulting in widespread inflammation and systemic organ failure. The aim of this research is to investigate the possibility of pomegranate peel-derived silver nanoparticles (PGNP) as a potential alternative therapy for sepsis. Characterization using transmission electron microscopy revealed 10-30 nm spherical nanoparticles. In a rat model of sepsis, PGNP treatment improved spleen health, histology, and immune response as compared with septic rats. In rats treated with PGNP during sepsis, significant alterations in oxidative stress markers (p < .01) were observed. These included elevated levels of glutathione (0.63 ± 0.08 mmol/mg protein), reduced concentrations of nitric oxide (8.7 ± 0.8 μ mol/mg protein) and malondialdehyde (2.2 ± 0.3 nmol/mg protein), as well as increased activity of superoxide dismutase (159 ± 33 U/mg protein). Following PGNP administration, gene expression analysis revealed a decrease in spleen IL-1β, IL-6, and TNF-α, highlighting its anti-inflammatory potential. Furthermore, PGNP effectively controlled apoptosis-related genes (Bax, Bcl-2, and Casp3), indicating its role in cellular survival pathways. This study sheds light on the immunological regulation of the spleen during sepsis using PGNP, demonstrating its potential as a new effective treatment approach. The study emphasizes the necessity of continuing to investigate and develop alternative medicines, particularly in light of antibiotic resistance and the global impact of sepsis. RESEARCH HIGHLIGHTS: The study explored the potential medicinal benefits of pomegranate peel-derived silver nanoparticles (PGNP) in the treatment of sepsis. PGNP suppressed pro-inflammatory cytokines and enhanced the immune response. The study recommends PGNP as a viable substitute treatment.

Tocotrienol-rich fraction (TRF) protects against retinal cell apoptosis and preserves visual behavior in rats with streptozotocin-induced diabetic retinopathy

BackgroundTocotrienol is a vitamin E analogue that is known to exert anti-inflammatory and antioxidant effects. Hence, in the current study, the effects of TRF on the expression of pro- and anti-apoptotic proteins in the streptozotocin-induced diabetic rat retinas were investigated. The effect of TRF on the visual behaviour of rats was also studied.MethodsDiabetes was induced in rats by intraperitoneal injection of streptozotocin and was confirmed by a blood sugar level of at least 20 mmol/L, 48 h, post-injection. Diabetic rats were divided into a group treated with vehicle (DV) and the other treated with TRF (100 mg/kg; DT). A group of non-diabetic rats treated with vehicle (N) served as the control group. All treatments were administered orally for 12 weeks. Rats were then subjected to an assessment of general behaviour in an open field arena and a two-chamber mirror test to assess their visual behaviour. At the end of the experimental period, rats were sacrificed, and their retinas were isolated to measure the expression of pro- (Casp3, Bax) and anti-apoptotic (Bcl2) markers using RT-qPCR and ELISA. TUNEL staining was used to detect the apoptotic retinal cells.ResultsTreatment with TRF lowered the retinal expression of Casp3 protein by 2.26-folds (p < 0.001) and Bax protein by 2.18-fold (p < 0.001) compared to vehicle-treated rats. The retinal anti-apoptotic protein Bcl2 expression was 1.87-fold higher in DT compared to DV rats (p < 0.001). Accordingly, the Bax/Bcl2 ratio in the TRF-treated group was significantly greater in DT compared to DV rats. Retinal Casp3, Bax, and Bcl2 gene expression, as determined by RT-qPCR, also showed changes corresponding to protein expression. In the open field test, DV rats showed greater anxiety-related behaviour than group N, while the behaviour of DT rats was similar to the N group of rats. DT rats and group N rats preferred the inverse mirror chamber over the mirror-containing chamber in the two-mirror chamber test (p < 0.01).ConclusionOral TRF therapy for 12 weeks lowers retinal cell apoptosis by decreasing pro- and increasing anti-apoptotic markers. The preservation of visual behaviour in a two-chamber mirror test supported these retinal molecular alterations in diabetic rats.

Novel strategies in HPV‑16‑related cervical cancer treatment: An in vitro study of combined siRNA-E5 with oxaliplatin and ifosfamide chemotherapy

BackgroundCervical cancer, primarily caused by HPV infection, remains a global health concern. Current treatments face challenges including drug resistance and toxicity. This study investigates combining E5-siRNA with chemotherapy drugs, Oxaliplatin and Ifosfamide, to enhance treatment efficacy in HPV-16 positive cervical cancer cells, targeting E5 oncoprotein to overcome limitations of existing therapies. MethodsThe CaSki cervical cancer cell line was transfected with E5-siRNA, and subsequently treated with Oxaliplatin/Ifosfamide. Quantitative real-time PCR was employed to assess the expression of related genes including p53, MMP2, Nanog, and Caspases. Cell apoptosis, cell cycle progression, and cell viability were evaluated using Annexin V/PI staining, DAPI staining, and MTT test, respectively. Furthermore, stemness ability was determined through a colony formation assay, and cell motility was assessed by wound healing assay. ResultsE5-siRNA transfection significantly reduced E5 mRNA expression in CaSki cells compared to the control group. The MTT assay revealed that monotherapy with E5-siRNA, Oxaliplatin, or Ifosfamide had moderate effects on cell viability. However, combination therapy showed synergistic effects, reducing the IC50 of Oxaliplatin from 11.42 × 10−8 M (45.36 μg/ml) to 6.71 × 10−8 M (26.66 μg/ml) and Ifosfamide from 12.52 × 10−5 M (32.7 μg/ml) to 8.206 × 10−5 M (21.43 μg/ml). Flow cytometry analysis demonstrated a significant increase in apoptosis for combination treatments, with apoptosis rates rising from 11.02 % (Oxaliplatin alone) and 16.98 % (Ifosfamide alone) to 24.8 % (Oxaliplatin + E5-siRNA) and 34.9 % (Ifosfamide + E5-siRNA). The sub-G1 cell population increased from 15.7 % (Oxaliplatin alone) and 18 % (Ifosfamide alone) to 21.9 % (Oxaliplatin + E5-siRNA) and 27.1 % (Ifosfamide + E5-siRNA), indicating cell cycle arrest. The colony formation assay revealed a substantial decrease in the number of colonies following combination treatment. qRT-PCR analysis showed decreased expression of stemness-related genes CD44 and Nanog, and migration-related genes MMP2 and CXCL8 in the combination groups. Apoptosis-related genes Casp-3, Casp-9, and pP53 showed increased expression following combination therapy, while BAX expression increased and BCL2 expression decreased relative to the control. ConclusionThe study demonstrates that combining E5-siRNA with Oxaliplatin or Ifosfamide enhances the efficacy of chemotherapy in HPV-16 positive cervical cancer cells. This synergistic approach effectively targets multiple aspects of cancer cell behavior, including proliferation, apoptosis, migration, and stemness. The findings suggest that this combination strategy could potentially allow for lower chemotherapy doses, thereby reducing toxicity while maintaining therapeutic efficacy. This research provides valuable insights into targeting HPV E5 as a complementary approach to existing therapies focused on E6 and E7 oncoproteins, opening new avenues for combination therapies in cervical cancer treatment.

Exploring the mechanism of intestinal injury induced by Bisphenol S in freshwater crayfish (Procambarus clarkii): Molecular and biochemical approaches

Bisphenol S (BPS) is extensively utilized in various industries such as plastic manufacturing, food packaging, and electronics. The release of BPS into aquatic environments has been observed to have negative impacts on aquatic ecosystems. Research has shown that exposure to BPS can have adverse effects on the health of aquatic animals. This study aimed to explore the mechanism of oxidative stress and endoplasmic reticulum stress induced in freshwater crayfish (Procambarus clarkii) by exposure to BPS (0 µg/L, 1 µg/L, 10 µg/L, and 100 µg/L) for 14 days. The results showed that BPS exposure resulted in elevated levels of reactive oxygen species (ROS) and malondialdehyde (MDA) and severe intestinal histological damage. In addition, oxidative stress can occur in the body by inhibiting the activity of antioxidant enzymes and the expression of related genes. BPS exposure induced a significant increase in the relative mRNA expression levels of inflammatory cytokines (NF-κB and TNF-α) and key unfolded protein response (UPR) related genes (Bip, Ire1, and Xbp1). At the same time, BPS exposure also induced up-regulation of apoptosis genes (Cytc and Casp3), suggesting that UPR and Nrf2-Keap1 signaling pathways may play a protective role in the process of apoptosis and oxidative stress. In conclusion, Our findings present the initial evidence that exposure to environmentally relevant levels of BPS can lead to intestinal injury through various pathways, highlighting concerns about the potential harm at a population level from BPS and other bisphenol analogs.

The Effect of ZnO Nanoparticles Functionalized with Glutamine and Conjugated with Thiosemicarbazide on Triggering of Apoptosis in the Adenocarcinoma Gastric Cell Line.

Gastric carcinoma is the fourth most common malignancy worldwide. Conjugation of metal nanoparticles with thiosemicarbazones has shown considerable anti-cancer potential. Zinc oxide nanoparticles (ZnO NPs) were synthesized, functionalized by glutamine, and conjugated with thiosemicarbazide (ZnO@Gln-TSC). Fourier transform infrared spectroscopy, X-ray diffraction, scanning electron microscopy and transmission electron microscopy imaging, energy-dispersive X-ray, DLS, and zeta potential were used to characterize the NPs. The toxicity of ZnO NPs, TSC, ZnO@Gln-TSC NPs, and oxaliplatin in AGS cells and ZnO NPs and ZnO@Gln-TSC NPs in HEK293 cells was investigated by MTT assay. Cell apoptosis was evaluated by flow cytometry, caspase-3 activity, and Hoechst staining assays. The intra-cellular reactive oxygen species level and expression level of the CASP3 gene in AGS cells treated with ZnO@Gln-TSC NPs were evaluated. The NPs were in the size range of 20 to 70 nm. The DLS and zeta potential were 374 nm and -31.7 mV, respectively. In MTT, the IC50 of ZnO, TSC, oxaliplatin, and ZnO@Gln-TSC NPs for AGS cells were 130, 80.5, 67.7, and 9.8 μg/mL, respectively, and the IC50 of ZnO and ZnO@Gln-TSC NPs for HEK293 cells were 215 and 150.5 μg/mL, respectively. Flow cytometry showed higher apoptosis in the cell treated with the NPs and TSC. Apoptotic features, including cell shrinkage, were recognized. A significant increase of 5.9 folds in the level of ROS was noticed. The activity of caspase-3 and the expression level of the CASP3 gene were increased by1.83 and 1.6 folds after exposure to ZnO@Gln-TSC NPs, respectively. This study revealed the anti-cancer potential of ZnO@Gln-TSC NPs to be used for gastric cancer treatment after further in vitro and in vivo assays.

Yinchenhao Decoction mitigates intestinal impairment induced by high carbohydrate diet in largemouth bass (Micropterus salmoides): insights from inflammation, apoptosis, oxidative stress, tight junctions, and microbiota homeostasis.

As a major source of energy, carbohydrates have a protein-saving effect. However, excessive consumption of carbohydrates can lead to the disruption of the intestinal barrier in fish, especially for carnivorous fish. Therefore, traditional Chinese medicine component Yinchenhao Decoction (YD), was used to detect the effect on intestinal barriers and microbial community equilibrium for largemouth bass in current research. In this research, a series of NC (normal carbohydrate diet) and HC (high carbohydrate diet) with graded YD treatments during 10 weeks feeding trial. Results suggested that 2% and 4% YD treatments significantly reduced gut inflammation and mucosal loss caused by HC. Compared with NC, HC significantly decreased the relative expression of intestinal tight junction-related genes (zo1, claudin1, claudin7, and occludin). However, with the application of YD, the expression of tight junction-related genes (zo1, claudin1, and claudin7) increased significantly (p < 0.05). Likewise, administration of YD significantly reduced elevated plasma diamine oxidase (DAO) activity caused by HC (p < 0.05). Additionally, YD significantly downregulated the mRNA expression of endoplasmic reticulum stress (ERS)-related genes (grp78, atf6, chopα, ire1, xbp1, and eifα) and pro-apoptosis genes (casp3, casp8, and bax) (p < 0.05), while upregulating the anti-apoptosis gene bcl2 (p < 0.05). Moreover, YD significantly increased the mRNA expression of antioxidant genes and the enzyme activities of CAT and GPX, while decreased MDA concentration significantly (p < 0.05). Whereas, YD markedly decreased the expression of pro-inflammatory genes (il1β, tnfα, il8, and nf-κB) and the immune enzymes activity (ACP and AKP) (p < 0.05) by up-regulating the expression of anti-inflammatory genes (ikb and il10). Notably, YD modulated the largemouth bass intestinal microbial community, enhanced the diversity and increased the abundance of probiotic microorganisms in the intestinal microbiota. In summary, YD supplementation in HC alleviated inflammation, apoptosis, oxidative stress, tight-junction injury, and microbiota disequilibrium in the intestine, which suggested that YD could be a valuable functional additive in aquaculture.

Piceatannol induces caspase-dependent apoptosis by modulating intracellular reactive oxygen species/mitochondrial membrane potential and enhances autophagy in neuroblastoma cells.

The aim of this study was to evaluate the anticancer effects of piceatannol, a natural stilbenoid, on human neuroblastoma cells. In order to accomplish this goal, we performed various cellular assays, including the XTT cell proliferation assay for cell viability, colony formation assay for colony formation capacity, FITC Annexin V and cell death detection kit for apoptosis, matrigel invasion assay for invasion capacity, intracellular reactive oxygen species (ROS) red dye for intracellular ROS levels, TMRM staining method for mitochondrial membrane potential (MMP), and the CYTO-ID autophagy detection kit for autophagy. Furthermore, we analyzed the expression levels of genes associated with apoptosis and autophagy using RT-qPCR. Based on our findings, piceatannol exhibited cytotoxic effects on neuroblastoma cells. Besides, treatment with piceatannol at both 50 and 100 μM concentrations for 72 h decreased colony formation, induced apoptosis and autophagy, inhibited cell invasion, decreased MMP, and increased ROS levels in SH-SY5Y cells. In addition, we observed significant upregulation in the expression levels of CASP8, BECLIN, ATG5, ATG7, and MAPILC3A genes between the two doses. These results suggest that piceatannol enhances autophagic activity and induces caspase-dependent apoptosis, indicating its potential as a therapeutic agent against neuroblastoma cells.

The Effects of Seleno-Methionine in Cadmium-Challenged Human Primary Chondrocytes.

Cadmium (Cd) is a potentially toxic element able to interfere with cellular functions and lead to disease or even death. Cd accumulation has been demonstrated in cartilage, where it can induce damage in joints. The aim of this study was to evaluate the effect of CdCl2 on primary cultures of human chondrocytes and the possible protective effect of seleno-methionine (Se-Met). Human primary articular chondrocytes were cultured and treated as follows: control groups, cells challenged with 7.5 μM and 10 μM CdCl2 alone, and cells pretreated with 10 and 20 μM Se-Met and then challenged with 7.5 μM and 10 μM CdCl2. Twenty-four hours after incubation, cell viability, histological evaluation with hematoxylin-eosin stain, and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay were performed. Furthermore, reverse transcription-PCR was carried out to evaluate mRNA levels of BAX, BAK1, CASP-3, and CASP-9. After CdCl2 challenge at both doses, a reduced cell viability and an overexpression of BAX, BAK1, CASP-3, and CASP-9 genes, as well as a high number of TUNEL-positive cells, were demonstrated, all parameters becoming higher as the dose of CdCl2 was increased. The pretreatment with Se-Met lowered the expression of all considered genes, improved cell viability and morphological changes, and reduced the number of TUNEL-positive cells. It was concluded that Se-Met plays a protective role against CdCl2-induced structural and functional changes in chondrocytes in vitro, as it improved cell viability and showed a positive role in the context of the apoptotic pathways. It is therefore suggested that a translational, multifaceted approach, with plant-based diets, bioactive functional foods, nutraceuticals, micronutrients, and drugs, is possibly advisable in situations of environmental pollution caused by potentially toxic elements.

Curcumin ameliorates pyroptosis in diabetic seminal vesicles by upregulating TRPV6.

Diabetes damages the seminal vesicle tissues leading to a decrease in seminal fluid secretion, so investigations are ongoing to identify specific therapeutic approaches to address diabetes-induced damage to seminal vesicles. This study investigated the secretory dysfunction of seminal vesicles and how curcumin can ameliorate this dysfunction. First, 40 diabetic males (DM group) and 40 nondiabetic males (control group) underwent seminal vesicle ultrasound evaluation and ejaculate volume measurements. Then, the effects of curcumin on seminal vesicle function were investigated in a diabetic rat model. Fifty 8-week-old SPF-grade SD rats were categorized into five groups: control, DM (diabetes mellitus), low-dose CUR (curcumin 50mg/kg/d), medium-dose CUR (curcumin 100mg/kg/d), and high-dose CUR (curcumin 150mg/kg/d). After a month-long diet with varying curcumin doses, key parameters such as body weight, blood glucose levels, seminal vesicle volume, and seminal fluid secretion were measured. Transcriptome sequencingwas performed to assess differences in gene expression and structural changes in rat seminal vesicle tissues were examined by HE staining. Finally, human seminal vesicle cell lines were cultured and divided into five groups (HG-CON, HG-CUR-5µM, HG-CUR-10µM, HG-CUR-20µM, and HG-CUR-50µM) to measure the fructose levels in the seminal vesicle cell culture fluids and evaluate the expression of CASP1, GSDMD, and TRPV6. Post TRPV6 interference, variations in the gene expression of CASP1, GSDMD, and TRPV6 were monitored. Diabetic patients exhibited a notable reduction in seminal vesicle volume and ejaculate volume compared with the control group, with a direct correlation between the decrease in ejaculate and seminal vesicle volume. Animal studies demonstrated that curcumin supplementation significantly augmented seminal vesicle volume in diabetic rats and notably improved their seminal vesicle secretory dysfunction, particularly in the high-dose curcumin group. Transcriptome sequencing and experimental verification pinpointed the differential expression of TPRV6 and pyroptosis-associated genes (CASP1, GSDMD), with reduced TRPV6 expression but increased markers of pyroptosis (CASP1 and GSDMD) in diabetic rats. Curcumin treatment reversed these effects with an increase in TRPV6 and a decrease in GSDMD and CASP1. Cell transfection experiments indicated that TRPV6 downregulation increased GSDMD and CASP1 gene expression. Curcumin effectively activates TRPV6, thereby diminishing pyroptosis in the seminal vesicle tissues of diabetic rats. This activation not only leads to an increase in the seminal vesicle volume but also significantly ameliorates the seminal vesicle secretory dysfunction in diabetic rats.

P-368 In vitro culture with rutin and VEGF-A before xenotransplantation protects ovarian tissue from ischemia and oxidative damage but not from growing follicle depletion

Abstract Study question Can pre-treatment with antioxidant and vascular endothelial factor (VEGF-A) help to maintain healthy tissue structure and follicle pool in xenotransplanted human ovarian tissue? Summary answer Rutin and VEGF-A help to prevent fibrosis and oxidative stress damage in grafted ovarian tissue but cannot preserve growing follicles. What is known already Nowadays, ovarian tissue (OT) cryopreservation and autotransplantation become established fertility preservation methods in numerous countries. However, even if hundreds of children has been born thanks to this procedure, most crucial process during which 70-80% of the follicles are lost in the graft is the transplantation itself. As the graft restores vascularisation and regains its function the tissue undergoes ischemia and oxidative stress. There have been attempts made to improve this critical postoperative period by administration of different growth factors, hormones, antoxidants or mechanical tissue damage. Study design, size, duration Pieces of frozen-tharwed OT (2,5 × 2,5 mm) from 10 women were transplanted into immunodeficient mice for 4 weeks. Before xenotransplantation OT fragments were in vitro cultured for 36h in culture media supplemented with 1 μg/ml rutin and 50 ng/ml VEGF-A (n = 10 OT, pretreated grafted) or without (n = 10 OT, non-pretreated grafted). Frozen-thawed OTs were used as non-grafted controls for ischemia-reperfusion injury, apoptosis and follicle loss evaluation. Participants/materials, setting, methods OTs were received from cancer patients (n = 10) undergoing cryopreservation at National Cancer Institute. Xenotransplantation was performed at Life Sciences Centre, Vilnius University. Retrieved grafts were devided into one small piece for immunochistochemical evaluation of α-SMA and apoptosis by TUNEL, and one bigger piece for qPCR analysis of genes related to tissue integrity (PRDX1, SOD1, FN1, COL6A1, CASP3, ANGPT2, mouse CD31) and follicle development (FOXO3, FSHR, GDF9), house keeping gene being GAPDH/Gapdh. Main results and the role of chance After 4 weeks of grafting, multiple genes and protein expressions were evaluated. Vascularisation evaluation: compared to non-grafted group, in grafted OTs we observed significantly more vessels (α-SMA staining, p &amp;lt; 0.0001 and ANGPT2 expression, p &amp;lt; 0.05), as well 1,5-fold higher relative expression of mouse CD31 in pretreated grafted group vs non-pretreated grafted group. Qxidative stress and apoptosis evaluation: expression of SOD2, PRDX1 and CASP3 genes was lower in both grafted groups compared to non-grafted control, significant alterations between xenotransplanted groups were found. Structural gene evaluation: in non-grafted control we could find higher expression of COL6A1 and FN1 genes compared to grafted groups and statistical difference of COL6A1 expression in between grafted groups (p &amp;lt; 0.05). Follicle pool evaluation: primordial follicle activation was measured by FOXO3 gene expression change, though we did not find significant differences between any of three groups, p = 0.06 was observed between grafted OT groups. Maturing follicles were evaluated by expression changes in GDF9 and FSHR genes. We observed significant decrese in expression of these genes in grafted groups (p &amp;lt; 0.05). Results from TUNEL are awaited, thus are the quantification of apoptotic cells, which will provide valuable insights for correlation with vascularisation and oxidative stress. Limitations, reasons for caution This study was performed only in a long-term xenotransplantation setting, we cannot know if rutin and VEGF-A had a significant influence during the first days of OT transplantation. Due to the small size of OT grafted, counting and distribution of follicles was not performed, Wider implications of the findings This study shows that pretreatment of OTs with rutin and VEGF-A could be potentially used in human setting due to protection of ischemia and oxidative stress, especially if the survival of growing follicles is not required, as it is in cases of prepubertal girls. Trial registration number not applicable