Expression Of Apoptosis-related Proteins Research Articles

MiR-146a-5p downregulated TRAF6/NF-κB p65 pathway to attenuate the injury of HT-22 cells induced by oxygen-glucose deprivation/reoxygenation.

MicroRNA-146a-5p (miR-146a-5p) actively participates in the process of cerebral ischemia-reperfusion (CI/R) injury. Dysregulation of the tumor necrosis factor receptor-associated factor 6 (TRAF6)/nuclear factor kappa-B (NF-κB) p65 axis is closely associated with inflammatory response. This study aimed to investigate the potential involvement of miR-146a-5p and TRAF6/NF-κB p65 in mediating CI/R progression in vitro. HT-22 cells were challenged with oxygen-glucose deprivation/reoxygenation (OGD/R) to simulate CI/R in vitro. HT-22 cells were transfected with miR-146a-5p mimics or TRAF6 overexpression constructs. The impact of miR-146a-5p on apoptosis, inflammation, and TRAF6/NF-κB p65 activation were investigated. OGD/R inhibited HT-22 cell viability, induced apoptosis, reduced miR-146a-5p levels and activated the TRAF6/NF-κB p65 pathway. MiR-146a-5p mimics reduced pro-inflammatory factor release, limited apoptosis-related protein expression, and inactivated the TRAF6/NF-κB p65 pathway in OGD/R-challenged HT-22 cells. Mechanistically, miR-146a-5p was verified to bind to TRAF6 3'UTR. TRAF6 overexpression reversed the beneficial effects of miR-146a-5p mimics on apoptosis, inflammation, and TRAF6/NF-κB p65 activation. This work revealed that miR-146a-5p targeted TRAF6 and suppressed the TRAF6/NF-κB p65 pathway, thereby reducing OGD/R-induced inflammation and apoptosis in HT-22 cells. These findings suggest the potential of the miR-146a-5p/TRAF6/NF-κB p65 axis in the treatment of CI/R.

Lead (Pb) Induces Osteotoxicity Through the Activation of Mutually Reinforced ER Stress and ROS in MC3T3-E1 Cells.

Lead (Pb) is the most common contaminant of heavy metals and is widely present in the environment. Destruction of bone structure, malformation of bone development, and loss of bone mass are important pathological features of lead-exposed individuals. However, the exact molecular mechanisms associated with lead exposure and osteogenic injury are still not fully understood. MC3T3-E1 mouse embryonic osteoblast is a cell line widely used in osteoblast cytology. It can differentiate into mature osteoblasts and express bone-specific genes in cell culture. The doses of 1, 2, and 4 mM Pb were adopted to study the toxicity of Pb on MC3T3-E1 proliferation and differentiation. In this study, the results show that Pb increases the expression of apoptosis-related proteins, including PARP1, cleaved caspase-3, Bax, and cleaved caspase-9. More importantly, Pb activated endoplasmic reticulum stress and oxidative stress, as evident by elevated PERK/ATF4/CHOP and ROS/NRF2 signaling pathway. Pb induced ROS production in MC3T3-E1 cells through endoplasmic reticulum stress and produced a lethal effect. NAC mitigated these effects. Endoplasmic reticulum stress inhibitor 4-PBA can block the ER stress pathway, reduce ROS production, and enhance cell viability. In addition, studies have shown that ERO1 activation in the ER stress pathway is responsible for inducing ROS production. ROS produced by the mitochondrial pathway also aggravates ER stress. This study suggests that Pb induces MC3T3-E1 cell apoptosis by inducing PERK-mediated ER stress and NRF2-mediated oxidative stress via mutual enhancement, which may be an important mechanism leading to skeletal toxicity.

Integrating Metabolomics, Histopathology, and Cardiac Marker Analysis to Assess Valsartan's Efficacy in Mitigating Dasatinib-Induced Cardiac Toxicity in Sprague-Dawley Rats.

Dasatinib (DASA) is associated with cardiotoxic effects, posing risks to patients. Valsartan (VAL) may offer protective benefits against these effects. This study evaluates the impact of DASA, VAL, and their combination on cardiac health. Wistar rats were treated with DASA, VAL, and a combination of VAL and DASA intraperitoneally every other day for 14 days. Body weight and survival rates were monitored. Serum levels of cardiac biomarkers (CPK, LDH, AST) were analyzed. Histopathological and immunohistochemical analyses assessed myocardial architecture and apoptosis-related protein expression. Metabolomic profiling was conducted using GC-MS to identify metabolic changes across treatment groups. The DASA group experienced significant weight loss and a 50% mortality rate, while the combination group had no mortality. Cardiac biomarkers like CPK, LDH, and AST were elevated in the DASA group but significantly reduced in the VAL + DASA group. Histopathological examination showed significant myocardial injury in the DASA group, with improved cardiac tissue morphology in the combination group. Immunohistochemical analysis revealed altered expression of apoptosis-related proteins, including caspase-3 and BCL-2, with improved levels in the combination group compared to DASA alone. Metabolomic profiling identified significant metabolic shifts, with 15 metabolites differentiating the treatment groups, and the VAL + DASA group mitigated the metabolic disturbances caused by DASA. The study suggesting VAL's potential therapeutic role in managing DASA-induced cardiac toxicity. The combination of VAL with DASA not only improved survival rates and reduced cardiac biomarker levels but also preserved myocardial architecture and normalized metabolic profiles. These findings highlight the importance of integrated approaches in evaluating drug efficacy and suggest VAL as a promising candidate for protecting cardiac function in preclinical models of DASA therapy.

CircKRT75 augments the cisplatin chemoresistance of nasopharyngeal carcinoma via targeting miR-659/CCAR2 axis.

Cisplatin resistance is a clinical challenge limiting the treatment of nasopharyngeal carcinoma (NPC). CircRNAs have been evidenced as key molecules involved in tumor advancement and drug resistance. The present study aimed to elucidate the potential biological value of circKRT75 in NPC cisplatin resistance. CircKRT75 levels in NPC clinical samples and parental/resistant cell lines were analyzed based on qRT-PCR. CCK-8 and flow cytometry were adopted to assess the impacts of circKRT75 on the growth viability and apoptotic ability of NPC resistant cells. Meanwhile, western blot was performed to detect changes in the expression of apoptosis-related proteins. Bioinformatics analysis predicted miRNAs and mRNAs downstream of circKRT75, and the interaction between circKRT75 and downstream targets was validated by RNA pull-down, dual-luciferase reporter and rescue experiments. CircKRT75 was notably enhanced in NPC tissues and NPC cisplatin-resistant cells. Functional experiments disclosed that circKRT75 silencing repressed NPC-resistant cell growth and promoted apoptosis. Bioinformatics screening identified that circKRT75 performed as a molecular sponge for miR-659, and CCAR2 was a direct target of miR-659. Further rescue assays confirmed that miR-659 inhibitor restored the inhibitory effect of circKRT75 knockdown on the growth of drug-resistant cells, while CCAR2 silencing could reverse the promotion of NPC cisplatin resistance by circKRT75 upregulation. Additionally, animal experiments revealed that circKRT75 knockdown restrained NPC cisplatin resistance in vivo. CircKRT75 contributed to cisplatin resistance in NPC through miR-659/CCAR2 signaling, which provided a novel perspective and direction to solve the problem of chemoresistance in NPC.

Mechanism of jianxin granules in the treatment of heart failure based on proteomics and metabolomics

BackgroundHeart failure (HF) is associated with high mortality and rehospitalization rates, highlighting the need for novel therapeutic approaches. Jianxin (JX) granules, a Traditional Chinese Medicine formulation, have been patented for the treatment of HF. However, the specific therapeutic effects and underlying mechanisms of JX granules have not been fully elucidated. This study aimed at investigating the effects and mechanism of JX granules in the treatment of HF based on proteomics and metabolomic profiling.MethodsHF model was established in rats by ligation of left coronary artery. The successfully modeled rats were randomly divided into three groups: the model group, the JX granules group, and Sacubitril/Valsartan (S/V) group. Four weeks after treatment, left ventricular (LV) function was evaluated via echocardiography. LV fibrosis and apoptosis were examined through histological analyses, while mitochondrial morphology was assessed using transmission electron microscopy. Quantitative assessment of oxidative stress was also conducted. Proteomics was used to identify the differentially expressed proteins and potential pathways. Metabolomics was utilized to elucidate the variations in metabolism. Then western blotting and in vitro analyses were performed.ResultsA rat model of HF was established, evidenced by a decrease in left ventricular ejection fraction (LVEF), stroke volume (SV), and left ventricular fractional shortening (LVFS), alongside diminished adenosine triphosphate (ATP) content, elevated oxidative stress, augmented apoptosis, and disrupted pyruvate metabolism. Treatment with JX granules ameliorated these effects, improving systolic function, reducing ventricular chamber size, and increasing LVEF, SV, and LVFS, as assessed by echocardiography. Additionally, JX granules attenuated cardiac fibrosis and improved mitochondrial structure, as evidenced by less vacuolation and clearer mitochondrial cristae, when compared to the model group. The treatment also regulated apoptosis-related protein expression, partially reversing the increase in cleaved Caspase-9, cleaved Caspase-3, and Bax and the suppression of Bcl-2 observed in the heart failure rats. All of these effects were similar to S/V. Proteomic and metabolomic analyses identified key differential genes, such as triosephosphate isomerase 1 (TPI1), lactate dehydrogenase B (LDHB), pyruvate kinase M (PKM), protein kinase B (Akt), Pyruvate Dehydrogenase Beta (PDHB) and lactate dehydrogenase A (LDHA), as well as vital pathways including carbon metabolism, the PI3K-Akt signaling pathway, pyruvate metabolism, and HIF-1α signaling pathway. Moreover, JX granules mitigated oxidative stress, inhibited apoptosis, and activated Akt in H9c2 cells exposed to angiotensin II, which could be reversed by the PI3K inhibitor LY294002.ConclusionJX granules improve HF in parallel to the efficacy of S/V, at least in part, through enhancing pyruvate metabolism, inhibiting oxidative stress and activating PI3K/Akt pathway.

Bushen Jianpi Tiaoxue Decoction (BJTD) ameliorates oxidative stress and apoptosis induced by uterus ageing through activation of the SIRT1/NRF2 pathway

BackgroundUterus ageing is a crucial factor contributing to decreased fertility in older women and is also implicated in menstrual disorders, endometritis, and adenomyosis. Bushen Jianpi Tiaoxue Decoction (BJTD) is a traditional Chinese medicine formulation used to ameliorate endocrine disorders in the female reproductive system and finds extensive application in ageing-related endometrial diseases. However, the mechanisms underlying its improvement of uterus ageing have not been thoroughly investigated. PurposeTo explore the potential components and mechanisms of BJTD in ameliorating uterus ageing through network pharmacology, in vivo, and in vitro experiments. MethodsMorphological changes were observed using hematoxylin and eosin staining, collagen deposition was assessed using Masson staining, and apoptotic-related molecules were detected using Western blot. After determining the modeling doses, BJTD intervention was administered at two doses, and the expression of oxidative stress and apoptosis-related genes and proteins was measured. The levels of cellular apoptosis were evaluated using the TUNEL assay kit and Annexin V/FITC-PI assay kit. The main components of BJTD were determined by UPLC-MS, and the potential targets and mechanisms of BJTD action were explored using network pharmacology and molecular docking. BJTD-Containing Serum (BJTD-S) was extracted and applied in vitro experiments using human endometrial stroma cells (hESC) to preliminarily identify the pathways affected. ResultsWe demonstrated that modeling with 600 mg/kg/day D-Gal for 5 weeks significantly increased collagen deposition in uterine tissues, particularly in the glands and stroma. Additionally, it significantly elevated the levels of TNF-α and IL-1β and increased the expression of p53 and BAX while decreasing BCL-2 expression. BJTD significantly reduced the increased levels of TNF-α and IL-1β induced by D-Gal, and modulated oxidative stress markers such as SOD, MDA, GSH-Px, and T-AOC. BJTD also inhibited the cascade activation of apoptosis induced by D-Gal, suppressing the expression of cleaved-Caspase 8, cleaved-Caspase 3, and BAX. SIRT1 is a potential target of BJTD action. In vitro experiments showed that BJTD-S significantly improved D-Gal-induced apoptosis in hESC cells, and the expression levels of SIRT1, NRF2, and HO-1 were significantly decreased in D-Gal-induced hESC, and BJTD-S significantly increased their expression. ConclusionBJTD can ameliorate oxidative stress and cell apoptosis levels in D-Gal-induced uterine aging, and its active ingredients can activate the SIRT1/NRF2 pathway to exert its effects. Importantly, our study provides novel insights into the molecular mechanisms by which traditional Chinese medicine influence uterus ageing. By specifically targeting the SIRT1/NRF2 pathway, BJTD presents a unique therapeutic approach that has not been extensively explored in previous studies, marking a significant advancement in the treatment of uterus ageing.

Saikosaponins from Bupleurum scorzonerifolium Willd. alleviates microglial pyroptosis in depression by binding and inhibiting P2X7 expression

BackgroundThe major depressive disorder (MDD) is a heterogeneous condition and characterized by a high recurrence rate, with neuroinflammation playing a significant role in its various potential pathologies. In recent years, neuronal pyroptosis induced by neuroinflammation has garnered increasing attention in depression research. Bupleurum scorzonerifolium Willd. is depression research in traditional Chinese medicine prescriptions to treat depression and exhibits notable anti-inflammatory properties. Saikosaponins (SSs) are the primary active components of Bupleurum scorzonerifolium Willd., although their mechanism of action in relation to anti-depressive effects remains unclear. PurposeThe purpose is to evaluate the efficacy of Chinese herbal medicine in treating depression through the lens of modern pharmacology. Additionally, it aims to explore the potential value of new mechanisms in the treatment of depression. Study design and methodsAn in vivo model of depression was established by intraperitoneal injection of LPS. HE staining, Nissl staining, and Golgi staining were used to evaluate the effectiveness of depression treatments in each group of mice. The activation of microglia was analyzed using immunofluorescence to evaluate the anti-inflammatory effects of drugs. In vitro models, N9 cells induced by LPS and ATP used CCK-8 assay, and lactate dehydrogenase assay were conducted to explore the therapeutic effect of the SSs. Western blot and immunohistochemical methods were employed to investigate the expression of P2X7 and apoptosis-related proteins. BzATP was introduced as a P2X7 agonist, and CETSA and CHX pulse-chase assay were utilized to verify the combined action and stability of the SSs with P2X7. Results57 triterpenoid saponin compounds were identified by Unifi in Bupleurum scorzonerifolium Willd. In both in vivo and in vitro models, the SSs has been shown to significantly improve depression-like behavior, reduce central inflammation, and significantly inhibit neuronal pyroptosis in mice. The SSs can significantly decrease the expression of P2X7 in the brains of depressed mice and enhance the stability of P2X7 protein in N9 cells. ConclusionSSs in Bupleurum scorzonerifolium Willd. bind and inhibit the P2X7 receptor to alleviate neuronal pyroptosis and improve symptoms of depression.

Effect and underlying mechanism of a photochemotherapy dual-function nanodrug delivery system for head and neck squamous cell carcinoma

BackgroundThe novel nanomaterials PNA-TN (PN) and PNA-TN-Dox (PND) have been shown to have strong inhibitory effects on breast cancer; however, it is unclear whether PN and PND have anti-head and neck squamous cell carcinoma (HNSCC) activity, and their potential mechanisms of activity are unknown. So, our study aims to explore the therapeutic effects of PN and PND on HNSCC and their possible mechanisms.MethodsWe used a series of phenotypic research to evaluate the effects of PN + Laser (L) and PND + L on the biological function of HNSCC cells in vitro and in vivo. We subsequently used mechanism research to examine changes in mRNA and protein expression related to apoptosis, epithelial‒mesenchymal transition (EMT), and the JNK signalling pathway.ResultsOur study revealed that PN and PND have strong inhibitory effects on HNSCC cells both in vitro and in vivo. In vitro, PN and PND significantly inhibited the proliferation, migration, invasion and EMT ability of HNSCC cells and promoted apoptosis; the inhibitory effect in the PND + L group was significantly greater than that in the PN + L group. In vivo, both treatments led to significant reductions in tumour volume and weight. Notably, the tumour volume and weight in the PND + L group were significantly lower than those in the PN + L group. Mechanism research confirmed that PN + L activated the expression of apoptosis-related proteins and inhibited the expression of EMT-related proteins via the JNK pathway. Furthermore, the anti-HNSCC effect of PN + L was blocked after the use of a JNK pathway inhibitor.ConclusionTreatment with PN + L or PND + L significantly inhibited the malignant progress of HNSCC cells, and the therapeutic effect of PND + L was significantly stronger than that of PN + L. The JNK signalling pathway is a key mechanism by which PN exerts its anti-HNSCC activity.

FGF6 inhibits oral squamous cell carcinoma progression by regulating PI3K/AKT and MAPK pathways

To explore diagnostic and prognostic biomarkers in the progression of oral squamous cell carcinoma (OSCC) and to reveal their regulatory mechanisms in key pathways. A RayBiotech protein chip was used to screen differentially expressed serum proteins in OSCC, oral leukoplakia (OLK), and healthy participants. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis were used to determine the pathways enriched by characteristic differential proteins. Immunohistochemical analysis and western blotting were used to verify the expression of characteristic differential proteins and key regulatory factors in human tissues and in a nude mouse model. Fibroblast growth factor 6 (FGF6) was identified as a key differential protein and was weakly expressed in OSCC tissues. The mitogen-activated protein kinases (MAPK) and PI3K-AKT pathways were identified as key signaling pathways. The results showed that pERK, Cyclin D1, pAKT, and BCL2 were highly expressed in OSCC, Caspase9 was lowly expressed in OSCC. With an increase in FGF6 expression in nude mice, the expression of FGFR4, pERK, Cyclin D1, pAKT, BCL2, GPX4, and ACSL4 increased, and the expression of Caspase9 decreased. FGF6 may change the expression of apoptosis-related proteins and proliferation factors by binding to FGFR4 in the PI3K-AKT/MAPK pathway and may inhibit the ferroptosis of OSCC, thereby possibly participating in the process of inhibiting OSCC.

Overcoming Stress Response-Induced Drug Resistance through Targeting CDK9 in Ph+ Leukemia

Introduction: The toxicity and quality of life burden following polychemotherapy protocols are significant, even among pediatric ALL patients classified as “low risk.” Therefore, it is essential to look for alternative therapies for patients with high-risk features, such as Philadelphia chromosome-positive (Ph+) leukemia, where harsh treatments are usually required. Heat shock protein 90 (HSP90), involved in stress response, is a critical target in leukemia due to its significant upregulation and essential role in stabilizing numerous oncogenic proteins, such as BCR::ABL1 kinase. However clinical approval of HSP90 inhibitors (HSP90i) is hampered due to resistance mechanisms, such as the heat shock response (HSR). Our recent findings identified HSP90α as the principal contributor to resistance in Ph+ leukemia cells. The combination of CDK7i and HSP90i synergistically inhibited the HSR via RNA Polymerase II (RNAPII) in Ph+ leukemia cells resistant to multiple TKIs.1 Methods: The present study investigated the effects of HSP90 and CDK9 inhibitors on Ph+ leukemia cells. Conventional and capillary-based (JESS) Western blotting assessed HSR- and apoptosis-related protein expression following HSP90i and CDK9i treatment. Isogenic cell line models overexpressing anti-apoptotic proteins MCL-1, BCL-2, and BCL-xL were generated. Time-dependent qPCR validated transcriptional changes in HSR- and apoptosis-related genes post-treatment. Matrix synergy drug screening was performed to identify synergistic drug interactions in Ph+ leukemia cell lines and patient-derived xenograft (PDX) leukemia cells. Results: We investigated the pivotal role of CDK9, a crucial mediator in the transcriptional elongation of stress response genes and downstream of CDK7 in regulating RNAPII mediated transcription. Concurrent inhibition of CDK9 and HSP90 effectively diminishes HSP90AA1 expression, a key chaperone in cellular stress responses. The co-targeting strategy disrupted the induction of the HSR by specifically downregulating HSP90α (p = 0.0003) and HSP70 (p < 0.0001) at the protein level, along with downregulating HSP90α/β, HSP70, HSP40, and HSP27 at the transcriptional level. A notable consequence of this dual inhibition is the downregulation of short-lived oncogenic proteins such as c-Myc and Mcl-1 over a time period of 1-24 hours. Additionally, other anti-apoptotic members of the BCL-2 family, including BCL-2 and BCL-xL are also downregulated within 6-24 hours after the treatment. This reduction in anti-apoptotic proteins correlates with an increase in cleaved Poly (ADP-ribose) polymerase (cl-PARP) and apoptotic cells (% sub-G1; pHSP90i:Combi < 0.0001; pCDK9i:Combi = 0.0001). Notably the combination of CDK9 and HSP90 inhibition is equally effective in isogenic leukemia cell line models, overexpressing BCL2 family members (BCL2, BCL-xl, and MCL-1). The synergistic efficacy of CDK9i and HSP90i has been validated across various leukemia cell lines (maximum ZIP score: >15; N=12) and patient-derived leukemia cells (N=3), highlighting its potential therapeutic significance in combating resistance. Summary: Dual targeting of HSP90 and CDK9 achieves HSR abrogation through RNAPII inhibition, leading to disruption of the transcription machinery responsible for activating stress response genes. Further, the combination will be tested in preclinical leukemia models, both alone and in conjunction with standard chemotherapy, with the goal of reducing doses and achieving stable remissions. 1. Vogt, M.; Dienstbier, N. et al. Co-targeting HSP90 alpha and CDK7 overcomes resistance against HSP90 inhibitors in BCR-ABL1+ leukemia cells. Cell Death & Disease 2023, 14 (12), 799.

Near-infrared modulated photothermal/nanozymatic dual-modal strategy for combating cancer via molybdenum selenide nanourchins

BackgroundBreast cancer, a major health concern globally, often faces challenges in the non-invasive elimination of cancer cells. This has prompted researchers to explore alternative strategies, such as photothermal therapy (PTT), which takes advantage of the location of breast cancer lesions. Additionally, the tumor microenvironment with high hydrogen peroxide (H2O2) provides a key prerequisite for treating tumors with nanozymes, especially those with peroxidase (POD)-like activity. However, the overexpression of TrxR1, which maintains redox homeostasis in tumor cells, limits reactive oxygen species (ROS)-based cytotoxic therapy. This study aims to present molybdenum selenide nano urchins (MoSe2 NUs) with excellent photothermal conversion efficiency, POD-like activity and selective inhibition of TrxR1 to achieve photothermal enhanced nanocatalytic therapy for combating breast cancer.MethodsThe characterization of MoSe2 NUs synthesized by the hydrothermal method was identified by TEM, XRD, Zeta potential and DLS. Whether MoSe2 NUs in conjunction with NIR possessed POD-like activities or not was identified via a TMB colorimetric method. The photothermal characteristics of MoSe2 NUs excited by near-infrared were recorded by an infrared thermal imaging camera. The antitumor effect of MoSe2 NUs was detected by cell death staining, apoptosis assay and animal experiments. TrxR1 and apoptosis-related protein expression were identified by Western blot and Immunohistochemistry.ResultsMoSe2 NUs possessed excellent photothermal conversion efficiency, peroxidase (POD)-like activity, and selective inhibition of TrxR1. Furthermore, the photothermal effect of MoSe2 NUs can enhance their POD-like activity, allowing for accurate cancer treatment under near-infrared (NIR) light. MoSe2 NUs plus NIR and MoSe2 NUs alone exhibited in vivo tumor inhibition rates of 69.7% and 35.1%, respectively. Mechanistically, NIR-regulated MoSe2 NUs induced potent apoptosis of cancer cells by downregulating the TrxR1 and elevating intracellular ROS, thereby leading to caspase-3 cleavage.ConclusionsThis study demonstrated that MoSe2 NUs under NIR irradiation can precisely and efficiently treat breast cancer and have great potential for clinical application.Graphical

Diagnostic and Prognostic Potential of FBXO8 Expression in Kidney Renal Clear Cell Carcinoma and Its Regulation of Renal Adenocarcinoma Cells

BackgroundThe F-box protein 8 Gene (FBXO8) has been shown to suppress invasion and metastasis in various cancer types. Recurrence and drug resistance pose significant challenges in renal cell carcinoma (RCC). Identifying novel biomarkers is crucial for addressing these issues. MethodsData on RNA sequencing and patient survival for KIRC was obtained from The Cancer Genome Atlas (TCGA), UALCAN, and Gene Expression Omnibus (GEO) databases. We confirmed FBXO8 gene expression and its impact on survival. Clinical characteristics were classified, and FBXO8 expression differences among various categories were observed. We conducted biofunctional predictions and analyzed the tumor microenvironment (TME), immune cell infiltration, and immune checkpoints in relation to FBXO8 expression. FBXO8 was overexpressed using a plasmid, and we assessed Kidney renal clear cell carcinoma (KIRC) cell proliferation, migration, and apoptosis through CCK8, wound healing tests, and western blot analysis. ResultsOur findings revealed decreased FBXO8 expression in KIRC, with patients exhibiting low FBXO8 expression experiencing shorter survival times. The low expression group showed elevated TME immune and estimate scores. Biofunctional analyses indicated that FBXO8 expression was notably linked to drug metabolism cytochrome P450, nutrition disease, receptor-ligand activity, and neuroactive ligand-receptor interaction. Furthermore, we discovered significant correlations between FBXO8 expression and immune cell infiltration, as well as checkpoints such as CD274. Overexpression (OE) of FBXO8 led to a marked reduction in cell proliferation and migration, along with increased apoptosis, as evidenced by apoptosis-related protein expression. ConclusionThis study demonstrates that FBXO8 serves as a biomarker for KIRC and plays a role in regulating cell proliferation, migration, and apoptosis.

Folic-acid-targeted drug delivery system implementing Angelica gigas polysaccharide: A potential strategy for colorectal cancer treatment.

The study focuses on the development of folate-targeted conjugates utilizing Angelica gigas polysaccharide (F2) as a drug carrier for colorectal cancer therapy. We synthesized F2-C-5-FU conjugates by linking carboxymethyl-5-fluorouracil (C-5-FU) with folic acid (FA) through ester bonding. The drug release behavior of F2-C-5-FU-FA was pH-dependent, favoring release under alkaline conditions. After 96 h in phosphate buffer (pH 7.4), the conjugate exhibited a cumulative release of 54.7 %, which was higher compared to other pH environments. In vitro, F2-C-5-FU-FA showed enhanced cytotoxicity and increased cellular uptake in folate receptor-positive HCT-116 cells compared to A549 cells. The conjugate also induced G2/M cell cycle arrest and modulated the BAX/BCL-2 mRNA expression ratio through the MAPK and NF-κB signaling pathways. In vivo, F2-C-5-FU-FA increased tumor fluorescence intensity, prolonged drug circulation, and reduced organ toxicity to non-target organs. The treatment promoted cancer cell apoptosis by inhibiting the expression of apoptosis-related proteins. Overall, F2-C-5-FU-FA conjugates demonstrate potential as an effective drug delivery system for targeted colorectal cancer therapy.

Intrauterine inoculation of pseudorabies virus impairs mouse embryo implantation via inducing inflammation and apoptosis in endometrium.

Pseudorabies virus (PRV) is the pathogenic agent of pseudorabies, causing serious reproductive failure in swine. However, it is still unknown whether PRV uterine inoculation impairs blastocyst implantation. In the present study, a PRV infection mouse model was developed. Pregnant mice were inoculated with either 104 or 105 50% tissue culture infectious dose (TCID50) units of PRV on gestation day 2 (GD2). Viral DNA was detected in tissues by PCR and/or in situ hybridization. Histopathological change and expression of proinflammatory cytokines in uterus were analyzed by H.E. staining and qPCR, respectively. Apoptosis was also investigated by TUNEL assay, and the expression of apoptosis-related proteins including Bax and Bcl-2 was detected by Western blot. The results showed that intrauterine exposure of PRV on GD2 reduced the number of embryo implantation site. Abundant viral DNA was detected in spinal marrow and brain, and small amounts of PRV genomes were detected in embryo implantation site, ovary as well as thymus. Considerable inflammatory cells infiltrating in the endometrium, with high levels of pro-inflammatory cytokines of interleukin (IL)-6, IL-1β and tumor necrosis factor-α mRNA after infection. In addition, PRV infection promoted apoptosis in stroma and endothelium of the mouse endometrium. Collectively, intrauterine inoculation of PRV during early pregnancy causes cytokine release syndrome and apoptosis in endometrium, which impairs mouse embryo implantation.

Experimental study on the treatment of diabetic cardiomyopathy in rats by using ultrasound-targeted microbubble destruction combined with coenzyme Q10 loaded long-circulating nanoliposomes

Objective: To investigate the therapeutic effects and mechanisms of Ultrasound-targeted microbubble destruction (UTMD) technology combined with CoQ10 loaded PEGylated nanoliposomes (CoQ10-PEG-lips) on diabetic cardiomyopathy (DCM) in rats. Methods: CoQ10-PEG-lips were prepared using the thin-film dispersion method combined with ultrasonic hydration, followed by quality assessment. Sixty healthy and clean male SD rats were selected, and 50 were randomly chosen using a random number table to establish a type 1 diabetes mellitus (DM) model via a single intraperitoneal injection of streptozotocin. The remaining 10 rats were assigned as the normal control group. A total of 47 rats successfully developed the DM model, and 40 were selected using the random number table method. Based on different intervention methods, these rats were then randomly divided into DM model group, CoQ10 solution group, CoQ10-PEG-Lips group, and CoQ10-PEG-Lips+UTMD group (n=10 per group). Normal control group and DM model group rats were injected with 1 ml of normal saline through caudal vein. CoQ10 solution group and CoQ10-PEG-Lips group were injected with 1 ml of CoQ10 solution or CoQ10-PEG-Lips solution containing 10 mg/kg of CoQ10 through caudal vein, respectively. Rats in the CoQ10-PEG-Lips+UTMD group were injected with 1 ml of CoQ10-PEG-Lips solution containing 10 mg/kg of CoQ10+100 mg of freeze-dried ultrasound microbubble powder through caudal vein and were given UTMD treatment at the same time, with the intervention given twice a week. After 12 weeks of intervention, cardiac function indexes [left ventricular end-systolic diameter (LVEDs), left ventricular end-diastolic diameter (LVEDd), and left ventricular ejection fraction (LVEF)]of each group were measured by ultrasonic cardiac function detection in vivo. Simultaneously, ex-vivo histopathological examination was conducted to assess myocardial cell morphology, cross-sectional area, collagen volume fraction (CVF), and apoptosis index (AI) in each group. Additionally, molecular biology techniques were employed to measure oxidative stress-related indicators and the expression of apoptosis-related pathway proteins. Results: The prepared CoQ10-PEG-lips had a well-rounded morphology, good dispersibility, and a high encapsulation efficiency of 87.45%±3.23%. After 12 weeks of intervention, the myocardial cell morphology in the CoQ10-PEG-Lips+UTMD group was intact, with orderly arrangement, closely resembling that of the normal control group. There were no statistically significant differences between the two groups in terms of LVEDs [(1.53±0.07) mm vs (1.42±0.04) mm], LVEDd [(2.93±0.15) mm vs (2.81±0.05) mm], or LVEF (80.76%±3.42% vs 84.60%±2.10%) (all P>0.05). Similarly, there were no significant differences between the two groups in myocardial cell cross-sectional area, CVF, or AI (all P>0.05). The CoQ10-PEG-Lips+UTMD group showed statistically significant differences in the above-mentioned indicators compared to the DM group, CoQ10 solution group, and CoQ10-PEG-Lips group (all P<0.05). In the CoQ10-PEG-Lips+UTMD group, the levels of myocardial superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and the expression of the anti-apoptotic protein B-cell lymphoma-2 (Bcl-2) were significantly higher compared to the DM model group, CoQ10 solution group, and CoQ10-PEG-Lips group (all P<0.05), while malondialdehyde levels and the expression of Bcl-2-associated X protein (Bax) and caspase-3 were lower (all P<0.05). Conclusions: Using PEG-lips to encapsulate the poorly soluble drug CoQ10 in combination with UTMD technology enables targeted delivery of the drug to the myocardium, which can help reduce myocardial cell damage, fibrosis, and apoptosis caused by diabetes mellitus (DM) by inhibiting oxidative stress damage and the Bcl-2/Bax/caspase-3 apoptosis signaling pathway, ultimately improving cardiac function in DCM rats.

Intranasal administration of Clostridium butyricum and its derived extracellular vesicles alleviate LPS-induced acute lung injury.

Acute lung injury (ALI) is associated with high morbidity and mortality rates. However, its clinical treatment is limited. Currently, the treatment of lung diseases by regulating the lung microbiota has become a research hotspot. In this study, we investigated the protective effects of the intranasal administration of Clostridium butyricum and its derived extracellular vesicles (EVs) against lipopolysaccharide (LPS)-induced ALI. The results demonstrated that compared with the LPS group, the pre-treatment group with C. butyricum and its EVs reduced the expression of pro-inflammatory cytokines and alleviated the symptoms in ALI mice by inhibiting the TLR4/MyD88 signaling pathway. Moreover, C. butyricum and its derived EVs inhibited the expression of apoptosis-related proteins and increased the expression of lung barrier proteins. Additionally, the intervention of C. butyricum changed the composition of the pulmonary microbiota. At the species level, LPS significantly increased the relative abundance of Acinetobacter johnsonii, while C. butyricum reversed this effect. In conclusion, these data demonstrate that intranasal administration of C. butyricum and its EVs can prevent LPS-induced ALI by reducing inflammation, inhibiting apoptosis, and improving lung barrier function. Additionally, C. butyricum regulated the pulmonary microbiota of mice to alleviate LPS-induced ALI.IMPORTANCEThe disorder of pulmonary microbiota plays an important role in the progression of acute lung injury (ALI). However, very few studies have been conducted to treat ALI by modulating pulmonary microbiota. In this study, the diversity and composition of pulmonary microbiota were altered in lipopolysaccharide (LPS)-induced ALI mice, but the ecological balance of the pulmonary microbiota was restored by intranasal administration of Clostridium butyricum. Moreover, the study reported the mechanism of C. butyricum and its derived extracellular vesicles for the treatment of LPS-induced ALI. These results reveal the importance of pulmonary microbiota in ALI disease. It provides a new approach for the treatment of ALI with new-generation probiotics.

Nimbolide Induces Cell Apoptosis via Mediating ER Stress-Regulated Apoptotic Signaling in Human Oral Squamous Cell Carcinoma.

Human oral squamous cell carcinoma (OSCC) poses a significant health challenge in Asia, with current therapeutic strategies failing to improve the survival rates for OSCC patients sufficiently. To elucidate the effects of Nimbolide on OSCC cell proliferation and apoptosis, we performed a series of experiments, including cell proliferation assays, annexin V/PI assays, and cell cycle analysis. We further investigated nimbolide's role in modulating endoplasmic reticulum (ER) stress, reactive oxygen species (ROS) production, and mitochondrial dysfunction using flow cytometry. Additionally, Western blotting was used to detect apoptosis-related protein expression. Our findings reveal that nimbolide exerts its anti-proliferative effects on OSCC cells by inducing apoptosis. The nimbolide increased intracellular ROS levels and acceleration of cellular calcium accumulation, respectively promoting endoplasmic reticulum stress and cancer cell apoptosis. Furthermore, nimbolide activates the caspase cascade by altering the mitochondrial membrane potential and apoptotic protein expression, thereby inhibiting the viability of tumor cells. Our data show that Nimbolide suppresses tumor growth through the induction of ROS production, ER stress, and mitochondrial dysfunction, resulting in apoptosis in OSCC cells. Overall, our study highlights nimbolide as a potential natural compound for OSCC therapy.

Green synthesis of Au nanoparticles by Scutellaria barbata extract for chemo-photothermal anticancer therapy

IntroductionThe combination of photothermal therapy and chemotherapy has emerged as a promising strategy for cancer treatment. The green synthesis of gold nanoparticles (Au NPs) using extracts from traditional Chinese medicine Scutellaria barbata D. Don (Scu) has revealed an economical, sustainable, and eco-friendly approach for the outstanding anticancer activities. MethodsScu extract was mixed with HAuCl4 solution to fabricate Scu-Au NPs. The synthesis process was optimized by varying the reaction temperature, reaction time, and HAuCl4 concentration. The changes in the components of Scu before and after synthesis were analyzed using the 3,5-dinitrosalicylic acid (DNS) reduction test, the rutin colorimetric method, and the Folin–Ciocalteu method. The morphology, size, and structure of Scu-Au NPs were characterized by TEM, DLS, XRD, and XPS. The photothermal characteristics induced by near-infrared (NIR) irradiation were assessed by continuously monitoring the temperature elevation. The anticancer activity and mechanisms of Scu-Au NPs were investigated using MTT assay, cell uptake studies, ROS level detection, cell apoptosis and necrosis assays, flow cytometry analysis, intracellular Caspase-3 protein activity fluorescence detection, and Western blot analysis on A549, 4T1, and RAW264.7 cells. ResultsScu-Au NPs were successfully fabricated using Scu extract as a reducing agent. The Scu-Au NPs were spherical with a size of 50 nm. During the synthesis process, the levels of reducing sugars, flavonoids, and polyphenols in the Scu extract decreased by 80.8%, 54.3%, and 70.1%, respectively. Scu-Au NPs demonstrated excellent photothermal responsiveness, thermostability, and biocompatibility. For chemo-photothermal anticancer therapy, the cytotoxicity of Scu-Au NPs on 4T1 cells reached approximately 85% upon exposure to 808 nm NIR irradiation. The anticancer activity was attributed to the induction of apoptosis and necrosis, achieved by increasing intracellular ROS levels, causing cell cycle arrest at the S phase, and modulating the expression of apoptosis-related proteins. DiscussionOur synthesized Scu-Au NPs exhibit favorable photothermal conversion characteristics, demonstrating excellent therapeutic efficacy and safety for cancer treatment.

Photobiomodulation and Physical Exercise Modulate of Cell Survival Proteins in the Skeletal Muscle of Rats with Heart Failure and Diabetes Mellitus.

Introduction: Heart failure (HF) and type 2 diabetes mellitus (DM2) are global health problems that often lead to muscle atrophy. These conditions are associated with increased autophagy and apoptosis in the muscle cells, resulting in decreased muscle mass. Physical exercise associated with photobiomodulation (PBM) seems promising to attenuate the skeletal muscle changes caused by HF and DM2, due to its direct effects on mitochondria, which may result in an increase in antioxidant capacity. Objective: To verify the influence of physical exercise and the association with PBM on autophagy, apoptosis, and cell survival signaling pathways in myocytes from rats with HF and DM2. Materials and Methods: Male rats were assigned to one of four groups: control (CT), HF+DM (disease model), exercise+HF+DM (EX+HF+DM), and EX+HF+DM+PBM (EX+HF+DM+PBM). To induce DM2, we administered streptozotocin (STZ) (0.25 mL/kg, intraperitoneally). HF was induced by coronary ligation. One week post-induction, an 8-week aerobic exercise and PBM protocol was initiated. Western blot analysis was used to measure the expression of apoptosis-related proteins and autophagy. Results: The EX+HF+DM+PBM group showed a substantial increase in Nrf2, p-AKT, and LC3-I levels compared to the HF+DM group. Conclusions: These findings suggest that physical exercise combined with PBM can upregulate proteins that promote myocyte survival in rats with HF and DM2.

Honokiol protects against diabetic retinal microvascular injury via sirtuin 3-mediated mitochondrial fusion.

Mitochondrial dysfunction and oxidative stress play important roles in diabetic retinal vascular injuries. Honokiol (HKL) is a small-molecule polyphenol that exhibits antioxidant effects and has a beneficial effect in diabetes. This study aimed to explore the potential ability of HKL to ameliorate vascular injury in diabetic retinopathy (DR) and its possible mechanisms of action. The effect of HKL was evaluated in vascular injury in an in vivo type 2 diabetic (db/db) mouse model. In vitro, retinal microvascular endothelial cells were treated with high glucose (HG) to simulate the pathological diabetic environment. Cell viability, expression of apoptosis-related proteins, cellular reactive oxygen species, mitochondrial membrane potential, and morphological changes in the mitochondria were examined. The diabetic mice exhibited severe retinal vascular damage, including vascular leakage in vivo and capillary endothelial cell apoptosis in vitro. HKL reversed the retinal vascular leakage in the diabetic mice. In vitro, HKL improved retinal capillary endothelial cell viability, decreased apoptosis, and reversed the HG-induced increased cellular oxidative stress and mitochondrial fragmentation. The sirtuin 3 (SIRT3) inhibitor 3-TYP blocked all the in vivo and in vitro protective effects of HKL against diabetic retinal vascular leakage and capillary endothelium and eliminated the decrease in oxidative stress levels and reduction of mitochondrial fragmentation. In conclusion, these findings suggest that HKL inhibits vascular injury in DR, which was likely achieved through SIRT3-mediated mitochondrial fusion. This study provides a potential new strategy for the treatment of DR.