Improved production of class I phosphatidylinositol 4,5-bisphosphate 3-kinase.

Phosphatidylinositol 4,5-bisphosphate 3-kinases (PI3K) are a family of kinases whose activity affects pathways needed for basic cell functions. As a result, PI3K is one of the most mutated genes in all human cancers and serves as an ideal therapeutic target for cancer treatment. Expanding on work done by other groups we improved protein yield to produce stable and pure protein using a variety of modifications including improved solubility tag, novel expression modalities, and optimized purification protocol and buffer. By these means, we achieved a 40-fold increase in yield for p110α/p85α and a 3-fold increase in p110α. We also used these protocols to produce comparable constructs of the ß and δ isoforms of PI3K. Increased yield enhanced the efficiency of our downstream high throughput drug discovery efforts on the PIK3 family of kinases.

Mild therapeutic hypothermic protection activates the PI3K/AKT signaling pathway to inhibit TRPM7 and suppress ferroptosis induced by myocardial ischemia­reperfusion injury.

The present study aimed to investigate the role of PI3K­mediated ferroptosis signaling induced by mild therapeutic hypothermia (MTH), which was defined as a temperature of 34˚C, in protecting against myocardial ischemia-reperfusion (I/R) injury (MIRI). To meet this aim, H9C2 cells underwent hypoxia­reperfusion (H/R) and/or MTH. The MTT assay was used to assess cell viability, cytotoxicity was measured using a lactate dehydrogenase cytotoxicity assay, and Annexin V­FITC/PI flow cytometric analysis was used to analyze early and late cell apoptosis. In addition, 84 healthy adult male Sprague­Dawley rats were randomly divided into seven groups (n=12), and underwent I/R and various treatments. Hemodynamics were monitored, and the levels of myocardial injury marker enzymes and oxidative stress markers in myocardial tissue were measured using ELISA. The expression levels of PI3K, AKT, transient receptor potential cation channel subfamily M member 7 (TRPM7), glutathione peroxidase 4 (GPX4) and acyl­CoA synthetase long chain family member 4 (ACSL4) in animals and cells were measured using western blot analysis. These experiments revealed that MTH could effectively reduce myocardial infarct size, improve hemodynamic performance following MIRI and suppress myocardial apoptosis, thereby contributing to the recovery from H/R injury. Mechanistically, MTH was revealed to be able to activate the PI3K/AKT signaling pathway in cells, upregulating GPX4, and downregulating the expression levels of TRPM7 and ACSL4. Treatment with 2­aminoethoxydiphenyl borate (an inhibitor of TRPM7) could further strengthen the myocardial protective effects of MTH, whereas treatment with erastin (promoter of ferroptosis) and wortmannin (inhibitor of PI3K) led to the effective elimination of the myocardial protective effects of MTH. Compared with in the I/R group, the PI3K/AKT activation level and the expression levels of GPX4 were both significantly increased, whereas the expression levels of TRPM7 and ACSL4 were significantly decreased in the I/R + MTH group. Taken together, the results of the present study indicated that MTH may activate the PI3K/AKT signaling pathway to inhibit TRPM7 and suppress ferroptosis induced by MIRI.

Network-based pharmacology and experimental validation to explore the mechanism of action of the Jiawei Pentongling formula for the treatment of endometriosis-related pain.

OBJECTIVE: To investigate the effects and mechanisms of the Jiawei Pentongling formula (, JWPTL) in treating endometriosis-related pain using network pharmacology study and experimental validation. METHODS: Active ingredients and relevant targets of JWPTL, as well as genes for endometriosis-related pain, were collected from public databases. Prediction of core targets and pathways of JWPTL against pain associated with endometriosis by protein-protein interaction (PPI) network work, gene ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional enrichment analysis. The Sprague-Dawley rat endo-metriosis model was constructed using the autologous endosomal transplantation method, and the successfully modeled rats were randomly divided into the model group and the JWPTL group, with 8 rats in each group. Another 8 rats were set up in the sham group. Rats in the JWPTL group used the rectal delivery method with the addition of JWPTL (7.77 g·kg-1·d-1) once a day for 28 d. Rats in the model and sham-operated groups were given equal amounts of saline using the same administration method. The 50% paw withdrawal threshold (PWT) of the rats was measured at different time points. After the intervention, the expression of phosphatidylinositol 3-kinase (PI3K) and protein kinase B (Akt) proteins and mRNA in endometriotic tissues was detected by immune-ohistochemistry and reverse transcription-polymerase chain reaction. RESULTS: From GeneCards, Online Mendelian Inheritance in Man and other databases, a total of 964 endometriosis (EMs) -related pain targets were screened, 142 active ingredients of JWPTL, 605 targets, and 221 potential targets were obtained by intersection of Venn diagram; 44 core targets were identified by constructing PPI network. KEGG enrichment analysis showed that JWPTL mainly involves the PI3K-Akt signaling pathway, estrogen signaling pathway, hypoxia inducible factor-1 signaling pathway, tumour necrotizing factor signaling pathway, and other signaling pathways in the treatment of EMs-related pain. Animal experiments showed that JWPTL could up-regulate the mechanical pain threshold and reduce the expression of PI3K and Akt proteins and mRNA in ectopic endometrial tissues of model rats. CONCLUSIONS: The present study preliminarily analyzed the pharmacological mechanism of the formula, and molecular docking and animal experiments showed the feasibility of this study, suggesting that the formula may inhibit the release of inflammatory factors and reverse the pain associated with EMs by downregulating the PI3K/Akt signaling pathway.

Network pharmacology combined with experimental validation show that apigenin as the active ingredient of Campsis grandiflora flower against Parkinson's disease by inhibiting the PI3K/AKT/NF-κB pathway.

The exploration of novel natural products for Parkinson's disease (PD) is a focus of current research, as there are no definitive drugs to cure or stop the disease. Campsis grandiflora (Thunb.) K. Schum (Lingxiaohua) is a traditional Chinese medicine (TCM), and the exact active constituents and putative mechanisms for treating PD are unknown. Through data mining and network pharmacology, apigenin (APi) was identified as the main active ingredient of Lingxiaohua, and key targets (TNF, AKT1, INS, TP53, CASP3, JUN, BCL2, MMP9, FOS, and HIF1A) of Lingxiaohua for the treatment of PD were discovered. The primary routes implicated were identified as PI3K/AKT, Apoptosis, TNF, and NF-κB pathways. Subsequently, therapeutic potential of APi in PD and its underlying mechanism were experimentally evaluated. APi suppressed the release of mediators of inflammation and initiation of NF-κB pathways in MES23.5 cells induced by MPP+. APi suppressed caspase-3 activity and apoptosis and elevated p-AKT levels in MES23.5 cells. Pretreatment with LY294002, a PI3K inhibitor, resulted in APi treatment blocking the activation of NF-κB pathway and expression of inflammatory factors in MES23.5 cells by activating the PI3K/AKT pathway. In conclusion, APi protects dopaminergic neurons by controlling the PI3K/AKT/NF-κB pathway, giving novel insights into the pharmacological mechanism of Lingxiaohua in treating PD.

AEBP1 restores osteoblastic differentiation under dexamethasone treatment by activating PI3K/AKT signalling.

Adipocyte enhancer-binding protein 1 (AEBP1) is closely implicated in osteoblastic differentiation and bone fracture; this research aimed to investigate the effect of AEBP1 on restoring osteoblastic differentiation under dexamethasone (Dex) treatment, and its interaction with the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway. Pre-osteoblastic MC3T3-E1 cells were cultured in osteogenic medium and treated by Dex to mimic steroid-induced osteonecrosis cellular model. They were then further transfected with control or AEBP1-overexpressed lentiviral vectors. Finally, cells were treated with the PI3K inhibitor LY294002, with or without AEBP1-overexpressed lentiviral vectors. AEBP1 expression showed a downward trend in MC3T3-E1 cells under Dex treatment in a dose-dependent manner. AEBP1-overexpressed lentiviral vectors increased relative cell viability, alkaline phosphatase (ALP) staining, Alizarin red staining and osteoblastic differentiation markers including osteocalcin (OCN), osteopontin (OPN), collagen type I alpha 1 (COL1A1), runt-related transcription factor 2 (RUNX2) and bone morphogenetic protein 2 (BMP2), but decreased cell apoptosis rate in MC3T3-E1 cells under Dex treatment; besides, AEBP1-overexpressed lentiviral vectors positively regulated p-PI3K and p-AKT expressions. Furthermore, LY294002 treatment decreased relative cell viability, Alizarin red staining, osteoblastic differentiation markers including OCN, OPN, RUNX2 and BMP, increased cell apoptosis rate and did not affect ALP staining in MC3T3-E1 cells under Dex treatment; meanwhile, LY294002 treatment weakened the effect of AEBP1 overexpression vectors on the above cell functions. AEBP1 restores osteoblastic differentiation under Dex treatment by activating the PI3K/AKT pathway.

Polyphyllin II inhibits breast cancer cell proliferation via the PI3K/Akt signaling pathway.

Paridis Rhizoma saponins (PRS) are significant components of Rhizoma Paridis and have inhibitory effects on various tumors, such as bladder, breast, liver and colon cancer. Polyphyllin II (PPII), one of the PRS, has an unclear effect on breast cancer. The present study aimed to explore the effect and mechanism of PPII in breast cancer. A network pharmacology approach was employed to predict the core components and breast cancer­related targets of PRS. Moreover, a xenograft tumor model was established to determine the anti­breast cancer effect of PPII in vivo. The viability of MDA­MB­231 cells was determined by a Cell Counting Kit­8 assay. Apoptosis was analyzed using annexin V/PI double staining. Additionally, Transwell and scratch assays were performed to evaluate invasion and migration. The potential mechanism was predicted by Kyoto Encyclopedia of Genes and Genomes enrichment analysis and molecular docking analysis and verified by western blot analysis. The effect of PPII on aerobic glycolysis in breast cancer cells was detected by lactic acid and pyruvate kits and Western blotting of glycolytic rate­limiting enzymes. Network pharmacology analysis revealed 26 core targets involved in breast cancer and that PPII was the core active component of PRS. The in vivo studies showed that PPII could inhibit the growth of breast cancer in mice. In vitro experiments confirmed that PPII induced cancer cell apoptosis and inhibited invasion and migration. Furthermore, PPII was capable of suppressing the expression of key proteins in the PI3K/Akt signaling pathway, reducing the generation of aerobic glycolytic products, and diminishing the protein expression levels of hexokinase 2 and pyruvate kinase M2. The results indicated that PPII inhibited aerobic glycolysis in breast cancer cells through the PI3K/Akt signaling pathway, thereby inhibiting breast cancer growth.

MAGL blockade alleviates steroid-induced femoral head osteonecrosis by reprogramming BMSC fate in rat.

The leading cause of steroid-induced femoral head osteonecrosis (ONFH) is the imbalance of bone homeostasis. Bone marrow-derived mesenchymal stem cell (BMSC) differentiation and fate are closely associated with bone homeostasis imbalance. Blocking monoacylglycerol lipase (MAGL) could effectively ameliorate ONFH by mitigating oxidative stress and apoptosis in BMSCs induced by glucocorticoids (GC). Nevertheless, whether MAGL inhibition can modulate the balance during BMSC differentiation, and therefore improve ONFH, remains elusive. Our study indicates that MAGL inhibition can effectively rescue the enhanced BMSC adipogenic differentiation caused by GC and promote their differentiation toward osteogenic lineages. Cannabinoid receptor 2 (CB2) is the direct downstream target of MAGL in BMSCs, rather than cannabinoid receptor 1(CB1). Using RNA sequencing analyses and a series of in vitro experiments, we confirm that the MAGL blockade-induced enhancement of BMSC osteogenic differentiation is primarily mediated by the phosphoinositide 3-kinases (PI3K)/ the serine/threonine kinase (AKT)/ (glycogen synthase kinase-3 beta) GSK3ß pathway. Additionally, MAGL blockade can also reduce GC-induced bone resorption by directly suppressing osteoclastogenesis and indirectly reducing the expression of receptor activator of nuclear factor kappa-Β ligand (RANKL) in BMSCs. Thus, our study proposes that the therapeutic effect of MAGL blockade on ONFH is partly mediated by restoring the balance of bone homeostasis and MAGL may be an effective therapeutic target for ONFH.

SPP1 induces idiopathic pulmonary fibrosis and NSCLC progression via the PI3K/Akt/mTOR pathway.

BACKGROUND: The prevalence of non-small cell lung cancer (NSCLC) is notably elevated in individuals diagnosed with idiopathic pulmonary fibrosis (IPF). Secreted phosphoprotein 1 (SPP1), known for its involvement in diverse physiological processes, including oncogenesis and organ fibrosis, has an ambiguous role at the intersection of IPF and NSCLC. Our study sought to elucidate the function of SPP1 within the pathogenesis of IPF and its subsequent impact on NSCLC progression. METHODS: Four GEO datasets was analyzed for common differential genes and TCGA database was used to analyze the prognosis. The immune infiltration was analyzed by TIMER database. SPP1 expression was examined in human lung tissues, the IPF fibroblasts and the BLM-induced mouse lung fibrosis model. Combined with SPP1 gene gain- and loss-of-function, qRT-PCR, Western blot, EdU and CCK-8 experiments were performed to evaluate the effects and mechanisms of SPP1 in IPF progression. Effect of SPP1 on NSCLC was detected by co-cultured IPF fibroblasts and NSCLC cells. RESULTS: Through bioinformatics analysis, we observed a significant overexpression of SPP1 in both IPF and NSCLC patient datasets, correlating with enhanced immune infiltration of cancer-associated fibroblasts in NSCLC. Elevated levels of SPP1 were detected in lung tissue samples from IPF patients and bleomycin-induced mouse models, with partial colocalization observed with α-smooth muscle actin. Knockdown of SPP1 inhibits TGF-ß1-induced differentiation of fibroblasts to myofibroblasts and the proliferation of IPF fibroblasts. Conversely, SPP1 overexpression promoted IPF fibroblast proliferation via PI3K/Akt/mTOR pathway. Furthermore, IPF fibroblasts promoted NSCLC cell proliferation and activated the PI3K/Akt/mTOR pathway; these effects were attenuated by SPP1 knockdown in IPF fibroblasts. CONCLUSIONS: Our findings suggest that SPP1 functions as a molecule promoting both fibrosis and tumorigenesis, positioning it as a prospective therapeutic target for managing the co-occurrence of IPF and NSCLC.

Sirt6 ameliorates high glucose-induced podocyte cytoskeleton remodeling via the PI3K/AKT signaling pathway.

BACKGROUND: Podocyte injury plays an important role in the occurrence and progression of diabetic kidney disease (DKD), which leads to albuminuria. Cytoskeletal remodeling is an early manifestation of podocyte injury in DKD. However, the underlying mechanism of cytoskeletal remodeling has not been clarified. Histone deacetylase sirtuin6 (Sirt6) has been found to play a key role in DKD progression, and the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (PKB/AKT) pathway directly regulates the cytoskeletal structure of podocytes. Whereas, the relationship between Sirt6, the PI3K/AKT pathway and DKD progression remains unclear. METHODS: Renal injury of db/db mice was observed by PAS staining and transmission electron microscope. Expression of Sirt6 in the glomeruli of db/db mice was detected by immunofluorescence. UBCS039, a Sirt6 activator, was used to explore the renal effects of Sirt6 activation on diabetic mouse kidneys. We also downregulating Sirt6 expression in podocytes using the Sirt6 inhibitor, OSS_128167, and induced upregulation of Sirt6 using a recombinant plasmid, after which the effects of Sirt6 on high glucose (HG)-induced podocyte damage were assessed in vitro. Podocyte cytoskeletal structures were observed by phalloidin staining. The podocyte apoptotic rate was assessed by flow cytometry, and PI3K/AKT signaling activation was measured by Western blotting. RESULTS: Db/db mice exhibited renal damage including elevated urine albumin-to-creatinine ratio (ACR), increased mesangial matrix, fused podocyte foot processes, and thickened glomerular basement membrane. The expression of Sirt6 and PI3K/AKT pathway components was decreased in db/db mice. UBCS039 increased the expressions of Sirt6 and PI3K/AKT pathway components and ameliorated renal damage in db/db mice. We also observed consistent Sirt6 expression was in HG-induced podocytes in vitro. Activation of the PI3K/AKT pathway via a Sirt6 recombinant plasmid ameliorated podocyte cytoskeletal remodeling and apoptosis in HG-treated immortalized human podocytes in vitro, whereas Sirt6 inhibition by OSS_128167 accelerated HG-induced podocyte damage in vitro. CONCLUSIONS: Sirt6 protects podocytes against HG-induced cytoskeletal remodeling and apoptosis through activation of the PI3K/AKT signaling pathway. These findings provide evidence supporting the potential efficacy of Sirt6 activation as a promising therapeutic strategy for addressing podocyte injury in DKD.

Regulating the lncRNA DSCR9/RPLP2/PI3K/AKT axis: an important mechanism of Xinfeng capsules in improving rheumatoid arthritis.

Background: Rheumatoid arthritis (RA) is a systemic autoimmune disease characterized by chronic and symmetrical polyarthritis. RA patients often experience inflammatory reaction and hypercoagulable state, which together affect the self-perception of patient (SPP). Currently, inhibiting inflammation and hypercoagulable state are common treatment methods for alleviating RA symptoms. Xinfeng Capsules (XFC) has a long history of treating RA, and can effectively improve the inflammatory response and hypercoagulable state of RA. However, the potential mechanisms have not yet been determined. Purpose and study design: This study elucidated the action mechanism of XFC in RA inflammation and hypercoagulability through the lncDSCR9/RPLP2/PI3K/AKT axis. Results: Clinical observations indicated that there was a strong link between XFC therapy and improvements in inflammatory and coagulation biomarkers, as well as SPP among RA patients. The subsequent network pharmacology analysis results identified the PI3K/AKT signaling pathway as a potential mediator for XFC treatment of RA. Furthermore, clinical validation and sequencing results revealed that lncRNA DSCR9 expression (a gene implicated in inflammation and coagulation) was negatively correlated with clinical markers of inflammation and coagulation, while positively correlated with SF-36 indicators. Notably, XFC treatment remarkably upregulated lncRNA DSCR9 expression and downregulated PI3K and AKT expressions, showing opposite expression trends to the untreated cases.The regulatory effect of XFC on the lncRNA DSCR9/RPLP2/PI3K/AKT axis in RA was investigated using techniques such as RNA pull-down assay, Western blot analysis, RT-PCR, and EdU assay. Moreover, the administration of the PI3K/AKT agonist RMH can counteract the effects of XFC on p-PI3K, p-AKT, inflammation, and hypercoagulability, reinforcing the role of pathway. Finally, animal studies utilizing HE staining and transmission electron microscopy (TEM) demonstrated that XFC notably decreased PI3K and AKT expressions in adjuvant-induced arthritis (AA) rats, mitigated inflammation and hypercoagulability, and enhanced the ultrastructure of synovial cells. These findings underscored the potential mechanisms of XFC in the treatment of RA. Conclusion: Regulating the lncRNA DSCR9/RPLP2/PI3K/AKT axis may be an important mechanism by which XFC improved RA inflammatory response and hypercoagulable state.

Inhibition of HOXC11 by artesunate induces ferroptosis and suppresses ovarian cancer progression through transcriptional regulation of the PROM2/PI3K/AKT pathway.

BACKGROUND: Ferroptosis, a non-apoptotic form of regulated cell death, plays a critical role in the suppression of various tumor types, including ovarian cancer. Artesunate (ART), a derivative of artemisinin, exhibits extensive antitumor effects and is associated with ferroptosis. This study aimed to investigate the mechanisms through which ART induces ferroptosis to inhibit ovarian cancer. METHODS: RNA sequencing was conducted to identify differentially expressed genes associated with ART-induced ferroptosis. Dual-luciferase reporter assays and electrophoretic mobility shift assays were performed to confirm the interaction between Homeobox C11 (HOXC11) and the Prominin 2 (PROM2) promoter. Cell Counting Kit-8 (CCK-8) assays, flow cytometry, and wound healing assays were used to analyze the antitumor effects of ART. Western blot, biochemical assays and transmission electron microscope were utilized to further characterize ART-induced ferroptosis. In vivo, the effects of ART on ferroptosis were examined using a xenograft mouse model. RESULTS: RNA sequencing analysis revealed that the HOXC11, PROM2 and Phosphatidylinositol 3-Kinase/ Protein Kinase B (PI3K/AKT) pathways were downregulated by ART. HOXC11 was found to regulate PROM2 expression by binding to its promoter directly. HOXC11 overexpression reversed ART-induced effects on ovarian cancer cell proliferation, migration, apoptosis and ferroptosis by activating the PROM2/PI3K/AKT signaling axis. Conversely, silencing PROM2 in HOXC11-overexpressing cells restored ART-induced ferroptosis and its associated antitumor effects by inhibiting the PI3K/AKT pathway. Consistently, in vivo studies using a xenograft mouse model confirmed that ART-induced tumor inhibition was mediated by ferroptosis through the suppression of the HOXC11/PROM2/PI3K/AKT pathway. CONCLUSION: This study identifies the HOXC11/PROM2/PI3K/AKT axis as a novel regulatory mechanism underlying ART-induced ferroptosis in ovarian cancer. Targeting the HOXC11/PROM2 axis may represent a promising therapeutic strategy for enhancing ferroptosis, offering new insights for the treatment of ovarian cancer.

Erxian decoction ameliorates myocardial tissue damage through activating PI3K/AKT signaling pathway in ovariectomized rats.

BACKGROUND: Erxian decoction (EXD) is an empirical formula for treating cardiovascular disease, our previous work has shown that EXD could improve the cardiovascular structure and function in ovariectomized (OVX) rats, but its pharmacological mechanism is still unclear. MATERIALS AND METHODS: Network pharmacology was utilized to assess the key active components and central targets of EXD in treating postmenopausal cardiovascular disease. Then, an OVX rat model was established, HE staining and transmission electron microscope were utilized to observe myocardial tissue morphology, TUNEL staining was utilized to detect cardiomyocyte apoptosis, western blot, and ELISA were used to confirm efficacy and pathway of EXD. RESULTS: The network pharmacology prediction results showed that 129 common targets were identified by intersecting EXD targets and postmenopausal cardiovascular disease targets, including AKT1, TNF, IL-6, IL-1ß, PTGS2 and other core targets, apoptosis, PI3K/AKT, and other signaling pathways may be closely related to postmenopausal cardiovascular disease. After ovariectomy, the myocardial tissue of rats was damaged, the expression level of PI3K/AKT pathway-related molecules in the myocardial tissue were decreased, the apoptosis index of cardiomyocytes was increased, and the levels of inflammatory factors (TNF-α, IL-6, and IL-1ß) were enhanced. EXD intervention could improve myocardial tissue injury, EXD could up-regulate the protein expression of PI3K and p-AKT in myocardial tissue, and thereby prevent myocardial cell apoptosis. At the same time, EXD downregulated the levels of inflammatory factors in serum of ovariectomized rats. CONCLUSION: EXD may prevent myocardial tissue damage through induction of the PI3K/AKT signaling pathway, thereby reducing cardiomyocyte apoptosis and inflammation. EXD may be a potential drug for the treatment of postmenopausal cardiovascular disease.

Integrin αVß3 mediates porcine deltacoronavirus infection and inflammatory response through activation of the FAK-PI3K-AKT-nf-κB signalling pathway.

Porcine deltacoronavirus (PDCoV) is an emerging porcine enteropathogenic coronavirus that causes acute watery diarrhoea in piglets, resulting in significant economic losses to the global swine industry. However, the underlying mechanism of PDCoV infection is not well defined, which seriously hinders the development of effective drugs and vaccines. Integrins (ITG) are heterodimeric transmembrane glycoproteins that play important roles in the life cycle of many viruses. In the current study, the viral entry pathways of PDCoV were explored and the role of ITGαVß3 was investigated during PDCoV infection. Our results showed that the lysosomal acidification inhibitor bafilomycin-A1 (Baf-A1) significantly reduced PDCoV infection, while exogenous protease facilitated PDCoV infection and even allowed PDCoV entry to bypass the endosomal pathway, suggesting PDCoV entry into cells via the endocytic pathway and the exogenous protease-mediated pathway simultaneously. Furthermore, ITGαVß3 was identified to be involved in PDCoV infection, especially during viral entry stages. PDCoV infection triggers the activation of the focal adhesion kinase (FAK)-phosphatidylinositol 3-kinase (PI3K)-serine/threonine-specific protein kinase (AKT) signalling pathway, and this activation is ITGαVß3-dependent, suggesting that the activation of the FAK-PI3K-AKT signalling pathway during PDCoV infection is mediated by ITGαVß3. Our results further demonstrated that PDCoV infection induced the expression of inflammatory cytokines, which was mediated by activation of the ITGαVß3-FAK-PI3K-AKT-nuclear transcription factor-κB (NF-κB) signalling pathway. Overall, the results revealed that ITGαVß3 is an essential host factor for PDCoV infection and can serve as a supplementary receptor to facilitate PDCoV infection, which can help us to explore the molecular mechanism of PDCoV infection.

Identifying the host factors required for entry will be helpful in uncovering the pathogenesis mechanisms and developing antivirals against the emerging coronavirus porcine deltacoronavirus (PDCoV). Herein, we revealed that PDCoV enters cells via the endocytic and exogenous protease-mediated pathways simultaneously. Integrins (ITG) αVß3 is a host factor required for PDCoV infection, especially during virus adhesion, invasion, and release. Most importantly, PDCoV promotes viral infection by activating the ITGαVß3-focal adhesion kinase (FAK)-phosphatidylinositol 3-kinase (PI3K)-serine/threonine-specific protein kinase (AKT) signalling pathway and induces inflammation by activating the ITGαVß3-FAK-PI3K-AKT-NF-κB signalling pathway. Overall, this is the first study to identify ITGαVß3 as an essential factor for PDCoV infection, which can help us to confirm the molecular regulatory mechanism and provide a comprehensive resource for PDCoV infection.

Serum apolipoprotein H determines ferroptosis resistance by modulating cellular lipid composition.

Ferroptosis is a regulated cell death process dependent on iron, triggered by the accumulation of lipid peroxidation. The environmental context significantly impacts cellular sensitivities to ferroptosis. Serum, constituting the extracellular fluid composition in vivo, provides crucial environmental biomolecules. In this study, we investigated the influence of sera on ferroptosis induction, pinpointing the serum protein apolipoprotein H (APOH) as a pivotal inhibitor of ferroptosis. Moreover, we elucidated that APOH suppresses ferroptosis by activating the phosphoinositide 3-kinase (PI3K)-AKT-sterol regulatory element-binding proteins (SREBPs) pathway, thereby elevating stearoyl-CoA desaturase (SCD) levels and augmenting cellular monounsaturated fatty acid-containing phospholipids (MUFA-PLs). Furthermore, ApoHinfer, the peptide derivative of the active region of APOH, mimics its ferroptosis inhibitory activity. Our findings underscore the critical role of serum protein APOH in the inhibition of ferroptosis and indicates potential therapeutic applications in treating cancer and diseases associated with ferroptosis.

Effects and mechanisms of Salvia miltiorrhiza Bunge extract on myocardial cell apoptosis in rat heart failure model.

PURPOSE: This work aimed to investigate the effects of Tanshinone IIA (Tan IIA) on myocardial cell (MC) apoptosis in a rat model of heart failure (HF). METHODS: Tan IIA was extracted from Salvia miltiorrhiza Bunge (SMB) using an ethanol reflux method. Fifty rats were randomly divided into five groups: sham (no treatment), mod (HF model establishment), low dose (LD: 0.1 mL/kg Tan IIA), medium dose (MD: 0.3 mL/kg Tan IIA), and high dose (HD: 0.5 mL/kg Tan IIA), with 10 rats in each group. The effects of different doses of Tan IIA on cardiac function, MC apoptosis, and the levels of proteins associated with the PI3K/Akt/mTOR signaling pathway were compared. RESULTS: Mod group showed a significant decrease in systolic arterial pressure, mean arterial pressure, heart rate, left ventricular systolic pressure, left ventricular ejection fraction, left ventricular fractional shortening, and the levels of p-PI3K, p-Akt, and p-mTOR proteins versus sham group (p < 0.05). Additionally, the left ventricular end-diastolic diameter (LVIDd), end-systolic diameter, diastolic pressure, and MC apoptosis were significantly increased (p < 0.05). LD, MD, and HD groups exhibited significant improvements across various indicators of cardiac function and MC apoptosis versus mod group (p < 0.05). CONCLUSIONS: Tan IIA may improve cardiac function and inhibit MC apoptosis in rats with HF by modulating the PI3K/Akt/mTOR signaling pathway.

Diosgenin Alleviates Obesity-Induced Insulin Resistance by Modulating PI3K/Akt Signaling Pathway in Mice Fed a High-Fat Diet.

Obesity is a global medical issue that can be effectively treated by relieving adipose inflammation and subsequent insulin resistance. Diosgenin (DIOS) has various effects as a steroidal saponin in inflammatory disorders. This study explored the effects and mechanism of DIOS on adipose inflammation and insulin sensitivity, both in silico and in vivo. The high-fat diet-induced obesity model in C57BL/6 mice was divided into five groups: normal chow (NC), high-fat diet (HFD), HFD with atorvastatin 10 mg/kg (AT), HFD with DIOS 100 mg/kg (DIOS 100), and HFD with DIOS 200 mg/kg (DIOS 200). Each group underwent an oral intervention for seven weeks. DIOS significantly suppressed weight gain in the body, liver, and epididymal fat pads. Additionally, it significantly improved fasting glucose and insulin levels, homeostatic model assessment of insulin resistance (HOMA-IR), and oral glucose tolerance test results, and reduced the proportion of total and M1 adipose tissue macrophages. Significant changes were shown in mRNA expression of janus kinase 2 (JAK2), insulin receptor (INRS), insulin receptor substrate 1 (IRS-1), phosphatidylinositol 3-kinase (PI3K), and protein kinase B (Akt), all of which exhibited high binding affinity in the in silico. Safety indices, including aspartate aminotransferase (AST), alanine transaminase (ALT), and creatinine level indicated the preventive effects of DIOS. In conclusion, DIOS improves insulin resistance and obesity-associated inflammation via the PI3K/Akt signaling pathway.

A single center experience on PI3K/AKT/MTOR signaling defects: Towards pathogenicity assessment for four novel defects.

BACKGROUND: Phosphoinositide 3 kinases (PI3K) are lipid kinases expressed in lymphocytes/myeloid cells. PI3K/AKT/mTOR signaling defects present with recurrent infections, autoimmunity, lymphoproliferation, and agammaglobulinemia. OBJECTIVE: To characterize the PI3K/AKT/mTOR pathway defects and perform pathway analyses to assess novel variant pathogenicity. METHODS: We included 12 patients (heterozygous PIK3CD (n = 9) and PIK3R1 (n = 1) (activated PI3K delta syndrome (APDS) with gain-of-function mutations) and homozygous PIK3R1 variant (n = 2)), performed clinical/laboratory/genetic evaluation, and flow cytometric PI3K/AKT/mTOR pathway analyses. RESULTS: Median age at onset of complaints was 17.5 months (3 months to 12 years) and at diagnosis was 15.7 years (2.5-37) in APDS. Median diagnostic delay was 12.9 years (1.6-27). Recurrent respiratory tract infections (90%), lymphoproliferation (70%), autoimmune/inflammatory findings (60%), and allergy (40%) were common in APDS. Recurrent viral infections were present in 4/10 and malignancy (non-Hodgkin lymphoma and testicular yolk sac tumor) was present in 2/10 in APDS. Low CD4+ T cells(5/8) with increased CD4+ effector memory (8/8) and CD4+ TEMRA cells (6/8) were present in the given number of APDS patients. We diagnosed tubulointerstitial nephritis, Langerhans cell histiocytosis, and late-onset congenital adrenal hyperplasia in APDS. Allergic findings, lymphoproliferation/malignancy, and high IgM were present in the APDS but not in PIK3R1 deficiency. Low IgM/IgG/CD19+ B cell counts were characteristic in patients with PIK3R1 homozygous loss-of function mutations. CONCLUSION: Differential diagnosis with combined immunodeficiency and diseases of immune dysregulation make molecular genetic analysis crucial for diagnosing mTOR pathway defects. It is easy to differentiate APDS and homozygous PIK3R1 defects with specific laboratory features. Additionally, mTOR pathway functional analysis is a definitive diagnostic and pathogenicity assessment tool for novel APDS mutations.

ENAH transcriptionally activated by YY1 promotes growth and invasion of laryngocarcinoma cells through PI3K/AKT signaling.

BACKGROUND: Laryngocarcinoma is a common malignancy in the upper respiratory tract. Enabled homolog (ENAH) is an actin-binding protein that is associated with the development of various cancers. However, its role and mechanism in laryngocarcinoma remain unknown. METHODS: The ENAH level in laryngocarcinoma was examined in silico, in vitro and in vivo. The prognostic analysis of the ENAH level was assessed on laryngocarcinoma patients. Gain- and loss-of-function assays were conducted in AMC-HN-8 and TU686 cells. Sh-ENAH-containing AMC-HN-8 cells were implanted into naked mice. The role and mechanism of ENAH in laryngocarcinoma were investigated by CCK-8, transwell, immunofluorescence, dual luciferase, RT-qPCR, immunohistochemistry, and western blotting experiments. RESULTS: The ENAH level was upregulated in laryngocarcinoma, which predicted a poor prognosis in laryngocarcinoma patients. Gain- and loss-of-function results showed that ENAH promoted proliferation, invasion and EMT of laryngocarcinoma cells. Moreover, ENAH was transcriptionally activated by YY1, and YY1/ENAH axis enhanced these malignant progresses of laryngocarcinoma cells. Besides, ENAH activated the PI3K/AKT pathway, and 740Y-P abolished the accelerative role of ENAH in proliferation, invasion and EMT of laryngocarcinoma cells. Furthermore, knockdown of ENAH reduced tumor size and weight, and the expression level of vimentin and PI3K/AKT pathway in tumor-bearing mice. CONCLUSION: ENAH transcriptionally activated by YY1 promotes cell growth, invasion and EMT of laryngocarcinoma through the activation of PI3K/AKT signaling.

Modulatory effects of miRNAs in doxorubicin resistance: A mechanistic view.

MicroRNAs (miRNAs) are a group of small non-coding RNAs and play an important role in controlling vital biological processes, including cell cycle control, apoptosis, metabolism, and development and differentiation, which lead to various diseases such as neurological, metabolic disorders, and cancer. Chemotherapy consider as gold treatment approaches for cancer patients. However, chemotherapeutic is one of the main challenges in cancer management. Doxorubicin (DOX) is an anti-cancer drug that interferes with the growth and spread of cancer cells. DOX is used to treat various types of cancer, including breast, nervous tissue, bladder, stomach, ovary, thyroid, lung, bone, muscle, joint and soft tissue cancers. Also recently, miRNAs have been identified as master regulators of specific genes responsible for the mechanisms that initiate chemical resistance. miRNAs have a regulatory effect on chemotherapy resistance through the regulation of apoptosis process. Also, the effect of miRNAs p53 gene as a key tumor suppressor was confirmed via studies. miRNAs can affect main biological pathways include PI3K pathway. This review aimed to present the current understanding of the mechanisms and effects of miRNAs on apoptosis, p53 and PTEN/PI3K/Akt signaling pathway related to DOX resistance.