Pro-apoptotic Protein Research Articles

Neuregulin 4 attenuates pancreatic β-cell apoptosis induced by lipotoxicity via activating mTOR-mediated autophagy

ABSTRACT Neuregulin 4 (Nrg4) is a brown fat-enriched endocrine factor that ameliorates lipid metabolism disorders. Autophagy is critical for pancreatic β-cell to counteract lipotoxicity-induced apoptosis. This study aimed at exploring whether Nrg4 attenuates lipotoxicity-induced β-cell apoptosis by regulating autophagy. The mouse pancreatic β-cell line MIN6 was cultured in palmitic acid (PA) with or without Nrg4 administration. Apoptosis rate, together with anti-apoptotic and pro-apoptotic protein levels, was investigated. Autophagic flux and autophagy-related protein levels along with related signaling pathways that regulate autophagy were also evaluated. Results showed that Nrg4 decreased PA-induced MIN6 apoptosis, enhanced anti-apoptotic protein B-cell lymphoma 2 (Bcl-2) expression and reduced pro-apoptotic proteins Bcl-2-associated X protein (Bax) and cleaved-caspase 3 expressions. Autophagy levels in MIN6 also decreased with PA treatment and Nrg4 administration reactivated autophagy. Further, Nrg4 administration activated autophagy via the mammalian target of rapamycin (mTOR) signaling pathway. In addition, when the mTOR pathway was stimulated or autophagy was suppressed, the beneficial effects of Nrg4 administration on MIN6 apoptosis were diminished. These results imply that Nrg4 administration attenuates MIN6 apoptosis by promoting mTOR-dependent autophagy and thus may lead to a new therapeutic method for type 2 diabetes mellitus (T2DM).

Abstract A024: Assessing Mitondiral Priming in Leukemia Using BH3 Complex Specific Antibodies (PRIMABS): Prediciting Patient Response to Apoptosis Inducing Drugs

Abstract Diagnostic tools for cancer therapy that enable oncologists to inform patients about the likely benefits of their treatment versus toxicity will empower patients to make evidence-based decisions about the management of their disease, and in some cases, their last phase of life. Here we provide a novel approach that measures key drug response features of individual cancer cells with the potential for providing prognostic/predictive diagnostic utility for practicing precision medicine. Most cancer treatments work though activation of the cell suicide program of apoptosis. This process is tightly regulated by the BCL-2 family of proteins whose members either prevent or promote cell death. The relative levels of, and the specific interactions between the pro-survival and pro-apoptotic family members that dictates the fate of all cells and determines their propensity to undergo apoptosis. Cells that more readily die in response to various stimuli are referred to as “primed”. Hence, the degree of priming of a tumor sample can be predictive of responses to anti-cancer drugs and can be established using a technique called “BH3 profiling”. The relevance of this method has been demonstrated but it has had only limited application. To address this limitation a unique panel of antibody-based reagents that are mitochondrial PRIMing state detecting Antibodies (PRIMABS), have been developed by our lab, as a novel approach to implementing personalized medicine and developing predicative diagnostic tests. These antibodies enable the degree of tumor cell priming to be determined directly, unlike BH3 profiling which is an indirect measure. The key innovation lies in the approach taken to determine the presence (or absence) of the critical pro-survival : pro-apoptotic protein complexes , as these are the principal determinants of cell priming.. Such PRIMAB reagents have hitherto not been available. Here we provide data outlining the methodology development as well as evidence demonstrating feasibility of PRIMABTM Dx platform for predicting AML and CLL patient response in retrospective studies. Our platform provides a PD biomarker that will become a clinical predictor of cancer cell response to treatments tat are intended to induce apopstosis in tumor cells. The PRIMABTM -DX platform will enable heretofore hard to achieve clinical applicability of priming measurements. This will be significant due to the important prognostic implications of PRIMAB-DxTM readouts and will provide insights into drug resistance and sensitivity mechanisms and lead to new treatment options for cancer patients. Citation Format: Michael Cardone. Assessing Mitondiral Priming in Leukemia Using BH3 Complex Specific Antibodies (PRIMABS): Prediciting Patient Response to Apoptosis Inducing Drugs [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: RNAs as Drivers, Targets, and Therapeutics in Cancer; 2024 Nov 14-17; Bellevue, Washington. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(11_Suppl):Abstract nr A024.

XAF1 antagonizes TRIM28 activity through the assembly of a ZNF313-mediated destruction complex to suppress tumor malignancy.

X-linked inhibitor of apoptosis-associated factor 1 (XAF1) is a stress-inducible pro-apoptotic protein that is commonly inactivated in multiple human cancers. Nevertheless, the molecular basis for its tumor suppression function remains largely uncharacterized. Here we report that XAF1 antagonizes the oncogenic activity of tripartite motif containing 28 (TRIM28) ubiquitin E3 ligase through zinc finger protein 313 (ZNF313)-induced ubiquitination and proteasomal degradation. XAF1 exerts apoptosis-promoting effect more strongly in TRIM28+/+ versus XAF1-/- tumor cells and suppresses tumor cell growth, migration, invasion, and epithelial-to-mesenchymal transition and xenograft tumor growth in a highly TRIM28-dependent fashion. Mechanistically, XAF1 interacts directly with the RING domains of TRIM28 and ZNF313 through the ZF6 and ZF7 domain, respectively, thereby facilitating ZNF313 interaction with and ubiquitination of TRIM28. A mutant XAF1 lacking either ZF6 or ZF7 domain exhibits no activity to promote TRIM28 ubiquitination. By destabilizing TRIM28, XAF1 blocks TRIM28-driven ubiquitination of p53 and RLIM, p53-HDAC1 interaction, and TWIST1 stabilization. Intriguingly, TRIM28 destabilizes XAF1 through K48-linked polyubiquitination and proteasomal degradation to protect tumor cells from apoptotic stress, indicating its role as an intrinsic antagonist against XAF1 and the antagonistic interplay of XAF1 and TRIM28. XAF1 expression is inversely correlated with TRIM28 expression in cancer cell lines and tumor tissues and more tightly associated with the survival of TRIM28-high versus TRIM28-low patients. Together, this study uncovers a novel mechanism by which XAF1 suppresses tumor malignancy and an important role for XAF1-TRIM28 interplay in governing stress response, illuminating the mechanistic consequence of its alteration during tumorigenic process.

TMET-22. OVERCOMING RESISTANCE: PANOBINOSTAT SENSITIZES 2DG-RESISTANT PEDIATRIC HIGH-GRADE GLIOMAS WITH H3.3 K27M/G34R MUTATIONS TO APOPTOSIS

Abstract BACKGROUND Pediatric high-grade gliomas (pHGG) harboring H3.3 K27M/G34R mutations present formidable challenges in treatment due to their aggressiveness and resistance to standard chemotherapy. Our preliminary investigations have shown promising results, indicating that combining 2-deoxyglucose (2DG), a glycolytic inhibitor, with DNA-damaging agents synergistically inhibits pHGG growth. However, prolonged exposure to 2DG induces resistance, necessitating the exploration of alternative therapies. This study aims to investigate the potential of panobinostat, a histone deacetylase inhibitor, in sensitizing 2DG-resistant pHGG with H3.3 K27M/G34R mutations to apoptosis. METHODS Transcriptomic analysis via Illumina NovaSeq 6000 and metabolomic profiling employing Liquid Chromatography-High-Resolution Mass Spectrometry was used to delineate the molecular landscape of naive and 2DG-resistant pHGG cell lines. Functional assays, including the Agilent Seahorse Mito Stress Test and flow cytometry, assessed bioenergetics and mitochondrial dynamics. Intracellular NAD+ and NADH levels were quantified utilizing NAD+/NADH Glo assays, while apoptotic markers and metabolic stress were measured using the Annexin V-FITC Conjugates for Apoptosis Detection kit and western blot. RESULTS Comparative analysis revealed increased transketolase activity in 2DG-resistant cells, augmenting the interaction between the pentose phosphate pathway and glycolysis, resulting in elevated levels of NADPH and ATP. Additionally, these resistant cells displayed enhanced reliance on mitochondrial energy metabolism, as evidenced by substantial increases in mitochondrial membrane potential and mass, coupled with elevated NAD+ levels. Subsequent treatment with panobinostat induced a significant decrease in NAD+ biosynthesis and ATP generation. Furthermore, this intervention triggered the upregulation of pro-apoptotic BCL-2 family proteins, particularly BAX and BAK, concomitant with a notable reduction in mitochondrial membrane potential and the accumulation of reactive oxygen species. Consequently, heightened caspase-3 activity ensued, leading to significantly elevated cell death rates in 2DG-resistant gliomas. CONCLUSION Our findings highlight panobinostat as a promising therapy to resensitize 2DG-resistant pHGG cells to apoptosis, offering hope for treating these aggressive tumors with H3.3 K27M/G34R mutations.

Bio-mimetic strategies to re-activate apoptotic cell death for cancer treatments

Apoptosis is a crucial process to maintain the correct balance between healthy cells and committed-to-death cells in every tissue. The internal (or mitochondrial) and external (or death receptor) pathways are responsible for driving a series of molecular events that lead to apoptosis by releasing pro-apoptotic proteins, such as B-cell lymphoma-2 (BCL-2) homology 3 (BH3)-only proteins and second mitochondria-derived activator of caspases/diablo inhibitor of apoptosis protein-binding mitochondrial protein (SMAC/DIABLO), that in turn activate the caspase family of proteases. By counterbalancing the apoptogenic machinery, anti-apoptotic BCL-2 family members turn off pro-apoptotic signalling, favouring cell survival, a circumstance that is particularly pronounced in tumour cells in which apoptosis is deranged. Therefore, targeting the defective apoptotic process has become a viable therapeutic option for the treatment of several cancers and much effort is being made in the research and development of effective compounds. This review discussed and updated the most promising therapeutic strategies that target deranged apoptosis process in cancer by mimicking the pro-apoptotic effects of BH3-only and SMAC/DIABLO proteins.

Nanoconjugate Carrying pH-Responsive Transferrin Receptor-Targeted Hesperetin Triggers Triple-Negative Breast Cancer Cell Death through Oxidative Attack and Assemblage of Pro-Apoptotic Proteins.

Triple-negative breast cancer (TNBC) is recognized as a major aggressive subtype of breast cancer due to its expeditious worsening growth, extensive metastatic capability, and recalcitrance to standard current treatments. Hesperetin (HSP), a natural bioflavonoid from citrus fruits, demonstrates pronounced anticancer efficacy, but its hydrophobicity limits its clinical development. The present study reports the fabrication of a biocompatible and pH-responsive transferrin (TF) receptor-targeted HSP-loaded poly(lactic-co-glycolic acid) (PLGA) nanobioconjugate (PLGA-HSP-TF NPs) and the exploration of its in vitro and in vivo antineoplastic potential. PLGA nanoparticles (NPs), PLGA-HSP NPs, and PLGA-HSP-TF NPs were synthesized and characterized by DLS, FTIR, FE-SEM, and 1H NMR spectroscopy. The stability and in vitro release profile of nanoparticles were inspected, and anticancer efficacy was scrutinized in terms of in vitro cytotoxicity, oxidative stress and apoptosis biomarkers, and cell cycle arrest. In vivo tumor regression and host survival studies were executed in Ehrlich ascites carcinoma (EAC) cell-bearing Swiss albino mice. The drug uptake of highly stable PLGA-HSP-TF NPs was accomplished effectively in MDA-MB-231 cells and showed the pH-dependent intracellular release of HSP, which generated excessive intracellular reactive oxygen species (ROS) that led to oxidative assault to the TNBC cells. This elevated ROS dropped the mitochondrial membrane potential and triggered apoptosis-mediated cell death by arresting the cell cycle at the G0/G1 phase. Furthermore, PLGA-HSP-TF NPs unveiled significant in vivo Ehrlich ascites carcinoma regression and host survival compared to free HSP with minimum toxicity at a minimum dose of 20 mg/kg body weight. The study divulges that PLGA-HSP-TF NPs may be an astounding anticancer nanocandidate for aggressive triple-negative breast cancer therapy.

Design, in silico studies and biological evaluation of novel chalcones tethered triazolo[3,4-a]isoquinoline as EGFR inhibitors targeting resistance in non-small cell lung cancer

A novel series of six [1,2,4]triazolo[3,4-a]isoquinolin-3-yl)-3-(1,3-diphenyl-1H-pyrazol-4-yl)prop-2-en-1-ones (3a–3f) was designed and synthesized. They were characterized based on spectral and elemental analyses. In silico studies were also committed to provide insights and a better understanding of their structural features. The six compounds were screened for their antiproliferative activity using the MTT assay against five human cancer cell lines, namely, A549, HCT116, PC3, HT29, and MCF-7 in parallel with the non-cancerous human lung cell line WI-38. The results showed that 3e and 3f have potential cytotoxic activities, especially on A549 cells with IC50 = 2.3 µM and 1.15 µM, respectively. Meanwhile, they recorded a minimal cytotoxic effect on WI-38 cells. Concerning the molecular mechanism of action, the present study showed the inhibitory effect of the six compounds against total EGFR. The most potent EGFR inhibitors were 3e and 3f with IC50 = 0.031 µM and 0.023 µM, respectively. The selectivity index of 3f for EGFRT790M was 1.81 times more selective than that of lapatinib. In addition, 3e and 3f initiated cell cycle arrest at the G2/M and pre-G1 phases along with the downregulation of anti-apoptotic protein Bcl2 and the upregulation of pro-apoptotic proteins: p53, Bax, and caspases 3, 8, and 9. Further studies are recommended to evaluate animal models’ promising anticancer activity and molecular mechanism of triazolo[3,4-a]isoquinoline derivatives 3e and 3f.

MRPL41, as a target for acupuncture, promotes neuron apoptosis in models of ischemic stroke via activating p53 pathway

Neuronal death is the key cause of ischemic stroke. Acupuncture (Acu) is a recognized method for the treatment and amelioration of cerebral ischemia. However, the molecular mechanism of Acu for treating ischemic stroke has not yet been detailedly elucidated. Based our microarray analysis results, mitochondrial ribosomal protein L41 (MRPL41), which is related to apoptosis, was identified as the target of Acu. MRPL41 expression was increased in middle cerebral artery occlusion/reperfusion (MCAO/R) model and reduced after Acu treatment. Following, MCAO/R model and oxygen and glucose deprivation/reoxygenation (OGD/R) model were established to explore the effect of MRPL41. Knockdown of MRPL41 increased cell viability and ani-apoptotic protein (Bcl-2) expression, and reduced apoptosis intensity and pro-apoptotic protein (Bax and Cleaved caspase-3) of OGD/R neurons. In vivo, MRPL41 silencing decreased neurological severity score, shrank infarct area, reduced encephaledema and neuron apoptosis. In addition, reduction of MRPL41 caused loss of p53. Our data uncover that Acu targets MRPL41, following with inhibiting neuron apoptosis via p53 pathway, thereby ameliorating ischemic stroke.

Effect of Acacia concinna Extract on Apoptosis Induction Associated with Endoplasmic Reticulum Stress and Modulated Intracellular Signaling Pathway in Human Colon HCT116 Cancer Cells.

Colorectal cancer (CRC) stands as one of the most prevalent cancer types and among the most frequent causes of cancer-related death globally. Acacia concinna (AC) is a medicinal and edible plant that exhibits a multitude of biological properties, including anticancer properties. This study aimed to investigate the impact of the AC extract on apoptosis induction and the underlying mechanisms associated with this effect in KRAS-mutated human colon HCT116 cells. The effect of AC extract on cell cytotoxicity was evaluated using MTT assay. Nuclear morphological changes were visualized with Hoechst 33342 staining, while mitochondrial membrane potential (MMP) was assessed via JC-1 staining. Flow cytometry was employed for cell cycle analysis, and intracellular ROS levels were determined using DCFH-DA staining. The results showed that HCT116 cells exposed to AC extract showed reduced cell growth and prompted apoptosis, as indicated by an increase in chromatin condensation, apoptotic bodies, the sub-G1 apoptotic cell population, and disrupted MMP. Expression levels of apoptosis mediator proteins determined by Western blot analysis showed an increase in pro-apoptotic proteins (Bak and Bax) while decreasing anti-apoptotic proteins (Bcl-2, Bcl-xL, and Mcl-1), leading to caspase-7 activation and PARP inactivation. AC extract was also found to enhance intracellular reactive oxygen species (ROS) levels and stimulate endoplasmic reticulum (ER) stress. Furthermore, AC extract increases the phosphorylation of ERK1/2, p38, and c-Jun while downregulating PI3K, Akt, β-catenin, and their downstream target proteins. These results demonstrate that AC extract could inhibit cancer cell growth via ROS-induced ER stress associated with apoptosis and regulate the MAPK, PI3K/Akt, and Wnt/β-catenin signaling pathways in HCT116 cells. Therefore, AC extract may be a novel candidate for natural anticancer resources for colon cancer treatment.

Astaxanthin Protects Against H2O2- and Doxorubicin-Induced Cardiotoxicity in H9c2 Rat Myocardial Cells

Astaxanthin (AST) is a carotenoid that has positive effects on various organs and tissues. It also exhibits a cardioprotective action. In this study, the influence of AST on the survival of H9c2 cardiomyocytes under hydrogen peroxide (H2O2)- and doxorubicin (DOX)-induced cardiotoxicity was investigated. Under these conditions, the content of cytosolic Ca2+ was measured, and changes in the area of the mitochondrial mass, as well as in the content of the voltage-dependent anion channel 1 (VDAC1), the autophagy marker LC3A/B, and the pro-apoptotic transcription factor homologous protein (CHOP), were determined. It was found that AST removed the cytotoxic effect of H2O2 and DOX, while cell survival increased, and the mitochondrial mass did not differ from the control. At the same time, a decrease in the content of cytosolic Ca2+ and the restoration of the VDAC1 level to values close to the control were observed. The restoration of the CHOP level suggests a reduction in endoplasmic reticulum (ER) stress in cells. The results allow us to consider AST as a potential agent in the prevention and/or treatment of cardiac diseases associated with oxidative stress.

Early apoptosis detection in canine granulosa cells through the analysis of BCL-2 and BAX proteins during the follicular development associated with oocyte maturation

The objective of this study was to investigate the early follicular apoptosis in canine ovarian follicles by examining the expression of anti-apoptotic BCL-2 and pro-apoptotic BAX proteins throughout the estrous cycle associated with oocyte maturation. Follicular cells from preantral and antral follicles of varying sizes were isolated and grouped based on follicle type and estrous phase. Antral follicles underwent flow cytometry analysis, whereas preantral follicles were subjected to Western blotting. The meiotic capacity of oocytes from these different follicle types was evaluated through in vitro maturation. Statistical analysis included ANOVA and Duncan’s test. Results showed fluctuations in BCL-2 and BAX levels across different follicular stages and estrous phases. BCL-2 levels increased (P<0.05) with follicular development in antral follicles, particularly during estrus, while BAX exhibited variations peaking (P<0.05) during estrus. The BCL-2/BAX ratio in antral follicles was higher (P<0.05) in estrus and diestrus compared to anestrus and proestrus. Additionally, BCL-2 and BAX proteins were detected in preantral follicles, with varying expression levels (P<0.05) across estrous phases. The BCL-2/BAX ratio in preantral follicles was highest (P<0.05) during anestrus and estrus and decreased (P<0.05) in proestrus and diestrus. Oocytes from preantral follicles did not reach the MII stage, regardless of the levels of BCL-2/BAX. Conversely, oocytes obtained from large follicles during estrus showed the highest (P<0.05) maturation percentages, which were associated with the highest BCL-2/BAX ratio. These findings provide insights into the dynamic patterns of BCL-2 /BAX in canine follicles across the estrous cycle, shedding light on their potential roles in oocyte development.

Daidzein ameliorates experimental traumatic brain injury-induced neurological symptoms by suppressing oxidative stress and apoptosis.

Traumatic brain injury (TBI) causes deficits in neurological function, induces pathological changes, and increases oxidative stress. The current investigation aimed to determine Daidzein's neuroprotective potential in experimental TBI. Initially, the HT-22 cell line exposed to H2O2 underwent in vitro examination, and the results showed that Daidzein had a neuroprotective effect evident from enhanced cell viability and decreased NO generation. Using three different Daidzein doses-1 mg/kg, 5 mg/kg, and 10 mg/kg-in the in vivo experiment, the potential of Daidzein was evaluated against TBI. The neurological severity score (NSS), kondziela's screen test, and elevated plus maze showed improvements after treatment with Daidzein manifested by decreased score, enhanced motor coordination, and anti-anxiety effects. Additionally, Daidzein improved mechanical allodynia and restored the breakdown of the blood-brain barrier. The FTIR spectral analysis showed restoration of the biochemical compositional changes. Furthermore, H & E and Toluidine blue staining revealed an improvement in the histopathological alterations. The RT-qPCR revealed an increase in mRNA expression level of Nrf2, HO-1, and Bcl-2 and the downregulation of Keap-1, Bax and Cleaved caspase-3 expressions. Thus, exhibiting its antioxidant and antiapoptotic potential. The RT-qPCR also manifested a decrease in mRNA expression of GFAP and Iba-1. Further immunohistochemistry results indicated Daidzein's antioxidant and antiapoptotic properties by upregulating Nrf2 and downregulating cleaved caspase-3. Daidzein also lowered the apoptosis index and improved neuronal survival evidenced by flow cytometric analysis. In addition to this, Daidzein notably increased the antioxidant enzyme levels and decreased the oxidative stress markers. The current study's findings point to the neuroprotective potential of the phytoestrogen Daidzein as it lessened neurological abnormalities, decreased oxidative stress, and lowered proapoptotic protein expression.

Rational development of fingolimod nano-embedded microparticles as nose-to-brain neuroprotective therapy for ischemic stroke.

Ischemic stroke is one of the major diseases causing varying degrees of dysfunction and disability worldwide. The current management of ischemic stroke poses significant challenges due to short therapeutic windows and limited efficacy, highlighting the pressing need for novel neuroprotective treatment strategies. Previous studies have shown that fingolimod (FIN) is a promising neuroprotective drug. Here, we report the rational development of FIN nano-embedded nasal powders using full factorial design experiments, aiming to provide rapid neuroprotection after ischemic stroke. Flash nanoprecipitation was employed to produce FIN nanosuspensions with the aid of polyvinylpyrrolidone and cholesterol as stabilizers. The optimized nanosuspension (particle size = 134.0 ± 0.6nm, PDI = 0.179 ± 0.021, physical stability = 72 ± 0h, and encapsulation efficiency of FIN = 90.67 ± 0.08%) was subsequently spray-dried into a dry powder, which exhibited excellent redispersibility (RdI = 1.09 ± 0.04) and satisfactory drug deposition in the olfactory region using a customized 3D-printed nasal cast (45.4%) and an Alberta Idealized Nasal Inlet model (8.6%) at 15L/min. The safety of the optimized FIN nano-embedded dry powder was confirmed in cytotoxicity studies with nasal (RPMI 2650 and Calu-3 cells) and brain related cells (SH-SY5Y and PC 12 cells), while the neuroprotective effects were demonstrated by observed behavioral improvements and reduced cerebral infarct size in a middle cerebral artery occlusion mouse stroke model. The neuroprotective effect was further evidenced by increased expression of anti-apoptotic protein BCL-2 and decreased expression of pro-apoptotic proteins CC3 and BAX in brain peri-infarct tissues. Our findings highlight the potential of nasal delivery of FIN nano-embedded dry powder as a rapid neuroprotective treatment strategy for acute ischemic stroke.

Fatty acid composition and anti-cancer activity of essential oil from Tenebrio molitor larvae in combination with zoledronic acid on prostate cancer

The essential oil extracted from Tenebrio molitor larvae (EOTM) is a natural product containing trace elements with potential therapeutic properties. This study aimed to assess the anticancer effects of EOTM and its synergistic interactions with zoledronic acid, a bisphosphonate drug, on prostate cancer cell lines. The chemical composition of EOTM was analyzed using GC-MS revealing a high concentration of fatty acids. The cytotoxicity of EOTM, both as a standalone treatment and in combination with zoledronic acid, was evaluated on prostate cancer cell lines (LNCaP, PC3) and normal hSKM using MTT assays. Results demonstrated that EOTM exhibited selective toxicity, inhibiting the growth of cancer cells in a dose-dependent manner while sparing normal cells. Morphological assessments and gene expression analyses of BCL2 and BAX were conducted through microscopy, Western blotting, and real-time RT-qPCR. These analyses indicated that EOTM induced apoptosis in cancer cells, as evidenced by cellular shrinkage, membrane blebbing, and nuclear fragmentation. Western blot results showed that EOTM downregulated the anti-apoptotic protein BCL2 and upregulated the proapoptotic protein BAX, suggesting activation of apoptosis pathways. Additionally, the combination of EOTM with zoledronic acid amplified these effects. Hoechst 33258 staining further confirmed the purity of cells following treatment. In conclusion, EOTM exhibits strong anticancer properties by inducing apoptosis in prostate cancer cells and demonstrates synergistic potential when combined with zoledronic acid. These findings warrant further investigation of EOTM as a natural and effective cancer treatment option.

Adaptive Changes in Human Leukocytes in Response to a Long-Term Stay in Antarctica

Oxidative stress and aging are known to alter the copy number (CN) of satellite III repeat (1q12) (SatIII(1q)) and telomeric repeat (TR) in the DNA of human cells. The extreme conditions of Antarctica could potentially affect the CN of these repeats in human blood cells, which may be associated with inhibition of the antioxidant system and activation of apoptosis. In this work, we analyzed the CN of ribosomal DNA (rDNA), SatIII(1q), and TR repeats in the leukocytes of 11 male members of the expedition to Vostok station in 2019–2020. To observe dynamic changes in the number of repeating elements of the genome and the degree of their oxidation, six blood samples were taken: before arrival in Antarctica, after 27, 85, 160, 270, and 315 days of wintering. To analyze adaptive changes, the expression levels of the BAX, BCL2, NOX4, NRF2, SOD1, and HIF1 genes were measured. We detected a decrease in SatIII(1q) CN and an increase in TR CN against the background of a stable rDNA CN in human blood cells during wintering. These changes, along with a decrease in the 8-oxodG in DNA, are associated with an increase in the activity of the NOX4 gene, a decrease in the activity of the NRF2 gene, and an increase in the expression of the proapoptotic protein BAX. Thus, wintering in Antarctica stimulates an adaptive response in the human body, which includes increased elimination from the bloodstream of “ballast” cells with a high level of DNA oxidation, a high SatIII(1q) content, and a low TR content. An increase in ROS levels due to chronic activation of the NOX4 gene along with the blocked NRF2 gene may play a significant role in the development of the response.

Protective Role of Sphingosine-1-Phosphate During Radiation-Induced Testicular Injury

The impact of ionizing radiation on the male reproductive system is gaining increasing attention, particularly when it comes to testicular damage, which may result in decreased sperm quality and hormonal imbalances. Finding effective protective measures to mitigate testicular damage caused by radiation has become a focal point in the biomedical field. S1P, an essential biological signaling molecule, has garnered significant interest due to its multiple roles in regulating cellular functions and its protective effects against radiation-induced testicular injury. S1P not only effectively reduces the generation of ROS induced by radiation but also alleviates oxidative stress by enhancing the activity of antioxidant enzymes. Furthermore, S1P inhibits radiation-induced cell apoptosis by regulating the expression of anti-apoptotic and pro-apoptotic proteins. Additionally, S1P alleviates radiation-induced inflammation by inhibiting the production of inflammatory factors, thereby further protecting testicular tissue. In summary, S1P effectively reduces radiation-induced testicular damage through multiple mechanisms, offering a promising therapeutic approach to safeguard male reproductive health. Future research should explore the specific mechanisms of action and clinical application potential of S1P, aiming to contribute significantly to the prevention and treatment of radiation damage.

Synthesis of nanohybrid consisting of taurine derived carbon dots and nanoceria for anticancer applications

Here, we first synthesized carbon dots (CDs) from taurine and then a nanohybrid with ceria (CeO2) nanoparticles using thermal decomposition method for checking their antineoplastic efficacy against human triple negative mammary carcinoma cells, MDA-MB-231. The in vitro study demonstrated significant dose-dependent antineoplastic activity of the nanohybrids within a range of concentration from 10 to 50 μg/mL after 48 h of treatment. The cellular morphology analysis clearly depicted substantial amount of cell death which seems to stem from increased intracellular reactive oxygen species (ROS) production. However, the maximum anticancer activity of the nanohybrid as compared to bare CDs and CeO2 is supposed to be due to the combined anticancer effect of both CDs and CeO2 (a well-established antitumor agent). Further we have performed molecular docking study to reveal the anticancer mechanism of CDs which exhibited high binding capacity towards several proapoptotic and antiapoptotic protein molecules. The binding affinity values were found to be – 8.7 kcal/mol, – 7.9 kcal/mol, – 9.6 kcal/mol, – 9.5 kcal/mol, – 12 kcal/mol and – 11.1 kcal/mol for BAD, BCl-2, p53, Caspase-8, Caspase-9 and Caspase-3 respectively. Taken together, our synthesized CDs-CeO2 nanohybrid could be thought as a potential anticarcinogenic option in the field of breast cancer therapeutics.

Thyroid Hormone Supplementation Restores Cognitive Deficit, Insulin Signaling, and Neuroinflammation in the Hippocampus of a Sporadic Alzheimer's-like Disease Rat Model.

Thyroid hormones play a crucial role in the development of the central nervous system and are considered pivotal to cognitive functions in the adult brain. Recently, thyroid dysfunction has been associated with Alzheimer's disease. The aim of this study was to assess the neuroprotective effects of triiodothyronine (T3) on insulin signaling, neuroinflammation, apoptosis, and cognitive function in a streptozotocin (STZ)-induced sporadic Alzheimer's disease-like model. Male Wistar rats underwent stereotaxic surgery for intracerebroventricular injections of streptozotocin (STZ; 2 mg/kg) or vehicle in the lateral ventricles to induce an AD-like model. The animals received a daily dose of 1.5 μg of T3/100 g body weight or the same volume of vehicle for 30 days and were subdivided into four experimental groups: (1) animals receiving citrate treated with saline (Control = CTL); (2) animals receiving citrate treated with T3 (T3); (3) animals receiving STZ treated with saline (STZ); and (4) animals receiving STZ treated with T3 (STZ + T3). The novel object recognition test was used to measure cognitive function. Serum analysis, real-time RT-PCR, immunohistochemistry, and immunoblotting analyses were also carried out. Our results demonstrated that T3 treatment reversed cognitive impairment and increased Akt and GSK3 phosphorylation in the treated group, while also reducing microglial activation (Iba-1) and GFAP expression (reactive astrocytes), along with TNF-α, IL-6, and IL-1β levels in the hippocampus. Additionally, T3 treatment increased levels of the anti-apoptotic protein Bcl-2 and reduced the expression of the pro-apoptotic protein BAX in the hippocampus. Our study demonstrated that T3 could potentially protect neurons in an AD model induced by STZ.

Therapeutic potential of EVs loaded with CB2 receptor agonist in spinal cord injury via the Nrf2/HO-1 pathway

ABSTRACT Background Spinal cord injury (SCI) poses a challenge due to limited treatment options. Recently, the effect and mechanism of Exo-loaded cannabinoid receptor type 2 (CB2) agonist AM1241(Exo + AM1241) have been applied in other inflammatory diseases but not in SCI. Methods The SCI model was set up using C57BL/6 mice, followed by the treatment of Exo, AM1241, and Exo + AM1241. We assessed the effects of the following treatments on motor function recovery using BMS, and evaluated histological changes, apoptosis activity, inflammation, and oxidative stress in the SCI mice model. Additionally, the effect of following treatments on spinal cord neural stem cells (NSCs) was evaluated under lipopolysaccharides (LPS) induced inflammatory and oxidative models and, glutamate (Gluts) induced cell apoptosis models. Result Our results demonstrated that Exo + AM1241 treatment significantly improved motor function recovery, after SCI by decreasing proinflammatory cytokines, and suppressing astrocyte/microglia (GFAP/Iba1) activation in the injury zone. Additionally, this treatment reduces pro-apoptotic proteins (Bax and caspase 3), increases the levels of the anti-apoptotic protein Bcl-2, enhances antioxidant defenses by boosting SOD and GSH, and lowers oxidative stress markers such as MDA. It also activates the Nuclear factor erythroid-2 (Nrf2) related factor 2 signaling pathway, thereby enhancing tissue protection against damage and cell death.

Japanese encephalitis virus infection induces mitochondrial-mediated apoptosis through the proapoptotic protein BAX.

The Japanese encephalitis virus (JEV), a zoonotic flavivirus, is Asia's primary cause of viral encephalitis. JEV induces apoptosis in a variety of cells; however, the precise mechanisms underlying this apoptosis resulting from JEV infection remain to be elucidated. Our previous studies showed that the proapoptosis gene BAX may have a role in JEV proliferation. In this study, we constructed a PK-15 cell line (BAX.KO) with a knockout of the BAX gene using CRISPR/Cas9. The knockout of the BAX gene effectively inhibited the proliferation of JEV, resulting in a 39.9% decrease in viral protein levels, while BAX overexpression produced the opposite effect. We confirmed that JEV induces apoptosis of PK-15 using 4',6-diamidino-2-phenylindole (DAPI) staining and Annexin V-FITC/PI staining. Furthermore, we found that the phosphorylation of P53 and the expression levels of BAX, NOXA, PUMA, and cleaved-caspase-3/9 were significantly upregulated after JEV infection. Moreover, we found that JEV infection not only caused mitochondrial damage, the release of mitochondrial cytochrome C (Cyt C), and the downregulation of the apoptosis-inhibiting protein BCL-2 but also reduced the mitochondrial membrane potential (MOMP) and the accumulation of intracellular reactive oxygen species (ROS). These factors collectively encourage the activation of the mitochondrial apoptosis pathway. In contrast, BAX gene knockout significantly reduces the apoptotic changes caused by JEV infection. Treatment with the caspase3 inhibitor attenuated JEV-induced viral proliferation and release, leading to a decrease in viral protein levels of 46% in PK-15 cells and 30% in BAX.KO cells. In conclusion, this study clarified the molecular mechanisms of JEV-induced apoptosis and provided a theoretical basis for revealing the pathogenic mechanisms of JEV infection.