Cyclin L1 participates in Adriamycin resistance and progression of osteosarcoma via PI3K/AKT-mTOR pathway.

Chemoresistance is a common and thorny problem in the treatment of osteosarcoma (OS), which obstructs the response of relapse or metastasis of OS to chemotherapy and leads to the unfavorable prognosis of OS patients. Cyclin L1 (CCNL1) is a non-canonical cyclin that plays an important role in the regulation of tumor cell proliferation and lymph node metastasis. In this work, we explored the impact of CCNL1 expression levels on proliferation, migration, and Adriamycin (ADM) resistance in OS and related mechanisms. We found that CCNL1 expression levels were significantly associated with clinical prognosis of patients with OS and CCNL1 could promote OS proliferation and migration. In addition, we also revealed that cellular CCNL1 was significantly increased in ADM-resistant OS cells and promoted ADM resistance. The PI3K/AKT-mTOR pathway is involved in CCNL1-mediated ADM resistance in OS. In summary, CCNL1 is involved in the progression of ADM resistance and OS through the PI3K/AKT-mTOR pathway, which will provide a new clue to the mechanism of ADM resistance and a potential target for the treatment of ADM-resistant OS.

Nimbolide protects against diabetic cardiomyopathy by regulating endoplasmic reticulum stress and mitochondrial function via the Akt/mTOR pathway.

Nimbolide has been demonstrated to possess protective properties against gestational diabetes mellitus and diabetic retinopathy. However, the role and molecular mechanism of nimbolide in diabetic cardiomyopathy (DCM) remain unknown. Diabetes was induced in rats via a single injection of streptozotocin (STZ) and then the diabetic rats were administered nimbolide (5 mg/kg and 20 mg/kg) or dimethyl sulfoxide daily for 12 weeks. H9c2 cardiomyocytes were exposed to high glucose (25 mM glucose) to mimic DCM in vitro. The protective effects of nimbolide against DCM were evaluated in vivo and in vitro. The potential molecular mechanism of nimbolide in DCM was further explored. We found that nimbolide dose-dependently decreased blood glucose and improved body weight of diabetic rats. Additionally, nimbolide dose-dependently improved cardiac function, alleviated myocardial injury/fibrosis, and inhibited endoplasmic reticulum (ER) stress and apoptosis in diabetic rats. Moreover, nimbolide dose-dependently improved mitochondrial function and activated the Akt/mTOR signaling. We consistently demonstrated the cardioprotective effects of nimbolide in an in vitro model of DCM. The involvement of ER stress and mitochondrial pathways were further confirmed by using inhibitors of ER stress and mitochondrial division. By applying a specific Akt inhibitor SC66, the cardioprotective effects of nimbolide were partially blocked. Our study indicated that nimbolide alleviated DCM by activating Akt/mTOR pathway. Nimbolide may be a novel therapeutic agent for DCM treatment.

The effect and mechanism of palmar ginseng in type 2 diabetic cognitive impairment.

Objective: To investigate the therapeutic effect of palmar ginseng on cognitive impairment in rats with type 2 diabetes, evaluate its neuroprotective effects, and explore its underlying mechanism. Methods: A rat model of diabetic cognitive impairment (DCI) was established by feeding with homemade high-fat, high-sugar chow combined with intraperitoneal injection of streptozotocin (STZ). Rats were continually fed high-fat, high-sugar chow for 60 days after successful induction of the model. Palmar ginseng was administered via gavage. The Morris test was performed after 30 days of treatment. At the end of the test, blood samples were collected, and the activities of IL-6, IL-10, TNF-α, and IL-1ß in rat serum. Pathological changes in hippocampal tissues were observed by Haematoxylin-eosin (HE) staining of the brain, activation of microglia in hippocampal tissues was detected by immunofluorescence, and the expression of PI3K/Akt/mTOR and JAK2/STAT3 proteins in the hippocampal tissues by Western blot. Results: During the administration of palmar Ginseng, the body weight and blood glucose levels of DCI rats were measured weekly, with results showing that Palmar Ginseng effectively reduced blood glucose levels and body weight of DCI rats. Behavioural tests in the water maze indicated that palmar ginseng effectively improved the learning and memory ability of DCI rats. HE and immunofluorescence staining showed that palmar ginseng improved DCI in rats, ameliorated hippocampal neuronal damage, and improved microglial activation. ELISA showed that palmar ginseng significantly reduced the expression of pro-inflammatory factors in the serum of DCI rats. Increased expression of anti-inflammatory factors was observed, and Western blot analysis showed that Palmar Ginseng regulated PI3K/Akt/mTOR and JAK2/STAT3 protein expression, promoted the phosphorylation of PI3K/Akt/mTOR, and inhibited JAK2/STAT3 protein phosphorylation in rat hippocampal tissues as well as in BV2 cells. Conclusions: Palmar ginseng may improve the onset and development of DCI by upregulating the phosphorylation of proteins in the PI3K/Akt/mTOR pathway.

Protective effect of folic acid on MNNG-induced proliferation of esophageal epithelial cells via the PI3K/AKT/mTOR signaling pathway.

Recent research has revealed that N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) constitutes a significant risk factor in the development of esophageal cancer. Several investigations have elucidated the beneficial impact of folic acid (FA) in safeguarding esophageal epithelial cells against MNNG-induced damage. Therefore, we hypothesized that FA might prevent MNNG-induced proliferation of esophageal epithelial cells by interfering with the PI3K/AKT/mTOR signaling pathway. In vivo experiments, we found that FA antagonized MNNG-induced proliferation of rat esophageal mucosal epithelial echinocytes and activation of the PI3K/AKT/mTOR signaling pathway. In our in vitro experiments, it was observed that acute exposure to MNNG for 24 h led to a decrease in proliferative capacity and inhibition of the PI3K/AKT/mTOR signaling pathway in an immortalized human normal esophageal epithelial cell line (Het-1A), which was also ameliorated by supplementation with FA. We successfully established a Het-1A-T-cell line by inducing malignant transformation in Het-1A cells through exposure to MNNG. Notably, the PI3K/AKT2/mTOR pathway showed early suppression followed by activation during this transition. Next, we observed that FA inhibited cell proliferation and activation of the PI3K/AKT2/mTOR signaling pathway in Het-1A-T malignantly transformed cells. We further investigated the impact of 740Y-P, a PI3K agonist, and LY294002, a PI3K inhibitor, on Het-1A-T-cell proliferation. Overall, our findings show that FA supplementation may be beneficial in safeguarding normal esophageal epithelial cell proliferation and avoiding the development of esophageal cancer by decreasing the activation of the MNNG-induced PI3K/AKT2/mTOR signaling pathway.

Plumbagin ameliorates LPS-induced acute lung injury by regulating PI3K/AKT/mTOR and Keap1-Nrf2/HO-1 signalling pathways.

Acute lung injury (ALI) is a major pathophysiological problem characterized by severe inflammation, resulting in high morbidity and mortality. Plumbagin (PL), a major bioactive constituent extracted from the traditional Chinese herb Plumbago zeylanica, has been shown to possess anti-inflammatory and antioxidant pharmacological activities. However, its protective effect on ALI has not been extensively studied. The objective of this study was to investigate the protective effect of PL against ALI induced by LPS and to elucidate its possible mechanisms both in vivo and in vitro. PL treatment significantly inhibited pathological injury, MPO activity, and the wet/dry ratio in lung tissues, and decreased the levels of inflammatory cells and inflammatory cytokines TNF-α, IL-1ß, IL-6 in BALF induced by LPS. In addition, PL inhibited the activation of the PI3K/AKT/mTOR signalling pathway, increased the activity of antioxidant enzymes CAT, SOD, GSH and activated the Keap1/Nrf2/HO-1 signalling pathway during ALI induced by LPS. To further assess the association between the inhibitory effects of PL on ALI and the PI3K/AKT/mTOR and Keap1/Nrf2/HO-1 signalling, we pretreated RAW264.7 cells with 740Y-P and ML385. The results showed that the activation of PI3K/AKT/mTOR signalling reversed the protective effect of PL on inflammatory response induced by LPS. Moreover, the inhibitory effects of PL on the production of inflammatory cytokines induced by LPS also inhibited by downregulating Keap1/Nrf2/HO-1 signalling. In conclusion, the results indicate that the PL ameliorate LPS-induced ALI by regulating the PI3K/AKT/mTOR and Keap1-Nrf2/HO-1 signalling, which may provide a novel therapeutic perspective for PL in inhibiting ALI.

Hydrogen Sulfide Promotes Platelet Autophagy via PDGFR-α/PI3K/Akt Signaling in Cirrhotic Thrombocytopenia.

Background and Aims: The role of platelet autophagy in cirrhotic thrombocytopenia (CTP) remains unclear. This study aimed to investigate the impact of platelet autophagy in CTP and elucidate the regulatory mechanism of hydrogen sulfide (H2S) on platelet autophagy. Methods: Platelets from 56 cirrhotic patients and 56 healthy individuals were isolated for in vitro analyses. Autophagy markers (ATG7, BECN1, LC3, and SQSTM1) were quantified using enzyme-linked immunosorbent assay, while autophagosomes were visualized through electron microscopy. Western blotting was used to assess the autophagy-related proteins and the PDGFR/PI3K/Akt/mTOR pathway following treatment with NaHS (an H2S donor), hydroxocobalamin (an H2S scavenger), or AG 1295 (a selective PDGFR-α inhibitor). A carbon tetrachloride-induced cirrhotic BALB/c mouse model was established. Cirrhotic mice with thrombocytopenia were randomly treated with normal saline, NaHS, or hydroxocobalamin for 15 days. Changes in platelet count and aggregation rate were observed every three days. Results: Cirrhotic patients with thrombocytopenia exhibited significantly decreased platelet autophagy markers and endogenous H2S levels, alongside increased platelet aggregation, compared to healthy controls. In vitro, NaHS treatment of platelets from severe CTP patients elevated LC3-II levels, reduced SQSTM1 levels, and decreased platelet aggregation in a dose-dependent manner. H2S treatment inhibited PDGFR, PI3K, Akt, and mTOR phosphorylation. In vivo, NaHS significantly increased LC3-II and decreased SQSTM1 expressions in platelets of cirrhotic mice, reducing platelet aggregation without affecting the platelet count. Conclusions: Diminished platelet autophagy potentially contributes to thrombocytopenia in cirrhotic patients. H2S modulates platelet autophagy and functions possibly via the PDGFR-α/PI3K/Akt/mTOR signaling pathway.

Targeting the PI3K/AKT/mTOR pathway offer a promising therapeutic strategy for cholangiocarcinoma patients with high doublecortin-like kinase 1 expression.

BACKGROUND: Cholangiocarcinoma (CCA), characterized by high heterogeneity and extreme malignancy, has a poor prognosis. Doublecortin-like kinase 1 (DCLK1) promotes a variety of malignant cancers in their progression. Targeting DCLK1 or its associated regulatory pathways can prevent the generation and deterioration of several malignancies. However, the role of DCLK1 in CCA progression and its molecular mechanisms remain unknown. Therefore, we aimed to investigate whether and how DCLK1 contributes to CCA progression. METHODS: The expression of DCLK1 in CCA patients was detected using Immunohistochemistry (IHC). We established DCLK1 knockout and DCLK1 overexpression cell lines for Colony Formation Assay and Transwell experiments to explore the tumor-promoting role of DCLK1. RT-PCR, Western blot and multiple fluorescent staining were used to assess the association between DCLK1 and epithelial-mesenchymal transition (EMT) markers. RNA sequencing and bioinformatics analysis were performed to identify the underlying mechanisms by which DCLK1 regulates CCA progression and the EMT program. RESULTS: DCLK1 was overexpressed in CCA tissues and was associated with poor prognosis. DCLK1 overexpression facilitated CCA cell invasion, migration, and proliferation, whereas DCLK1 knockdown reversed the malignant tendencies of CCA cells, which had been confirmed both in vivo and in vitro. Furthermore, we demonstrated that DCLK1 was substantially linked to the advancement of the EMT program, which included the overexpression of mesenchymal markers and the downregulation of epithelial markers. For the underlying mechanism, we proposed that the PI3K/AKT/mTOR pathway is the key process for the role of DCLK1 in tumor progression and the occurrence of the EMT program. When administered with LY294002, an inhibitor of the PI3K/AKT/mTOR pathway, the tumor's ability to proliferate, migrate, and invade was greatly suppressed, and the EMT process was generally reversed. CONCLUSIONS: DCLK1 facilitates the malignant biological behavior of CCA cells through the PI3K/AKT/mTOR pathway. In individuals with cholangiocarcinoma who express DCLK1 at high levels, inhibitors of the PI3K/AKT/mTOR signaling pathway may be an effective therapeutic approach.

The Prognostic and Therapeutic Potential of Fragile X Mental Retardation 1 (FMR1) Gene Expression in Prostate Adenocarcinoma: Insights into Survival Outcomes and Oncogenic Pathway Modulation.

Prostate adenocarcinoma (PRAD) is the second most common tumor associated with death. The role and mechanisms of the fragile X mental retardation 1 (FMR1) gene in PRAD remain unknown. We conducted an analysis of FMR1 expression in PRAD to determine its prognostic importance and connection to carcinogenic pathways such as PI3K_AKT_mTOR. Survival analyses were utilized to establish a correlation between FMR1 expression and patient outcomes. We used the integration of genomic data with bioinformatic predictions to predict the regulatory factors of the FMR1 gene in PRAD. Our data revealed that individuals with higher levels of FMR1 expression experience worse survival outcomes compared to those with lower expression (hazard ratio [HR] = 5.08, 95% confidence interval [CI] = 1.07 - 24, p = 0.0412). FMR1 expression was significantly higher in patients with advanced pathological tumor stages, particularly in the pT3 and pT4 combined stages and the pN1 nodal stage. Furthermore, patients with high Gleason scores (GSs) (combined GSs 8 and 9) exhibited increased levels of FMR1 expression. Our results further identify a possible regulatory link between FMR1 and key oncogenic pathways, including PI3K_AKT_mTOR, and predict the possible mechanism by which FMR1 is regulated in PRAD. Our data suggest that the FMR1 gene could serve as a biomarker for PRAD progression. However, in-depth investigations, including those with large patient samples and in vitro studies, are needed to validate this finding and understand the mechanisms involved.

Bta-miR-330 promotes bovine intramuscular pre-adipocytes adipogenesis via targeting SESN3 to activate the Akt-mTOR signaling pathway.

Consumers are more inclined to choose beef with a high intramuscular fat content (IMF), which regulated by lots of factors. It is very significant to find a miRNA that plays a key role in the accumulation of IMF. In our study, we found that bta-miR-330 was highly expressed in Japanese black cattle and differentially expressed at intramuscular pre-adipocytes differentiation processes. Furthermore, we transfected the bta-miR-330 mimic & inhibitor in intramuscular pre-adipocytes. The results showed that bta-miR-330 inhibits the proliferation but promotes the adipogenesis of intramuscular pre-adipocytes. Subsequently, our study showed that bta-miR-330 binds to SESN3, which inhibits the adipogenesis of intramuscular pre-adipocytes. Moreover, we established the mechanism that bta-miR-330 promotes the adipogenesis of intramuscular pre-adipocytes by targeting SESN3 to activate the Akt-mTOR signaling pathway. Overall, our results revealed that bta-miR-330-SESN3-Akt-mTOR axis plays an important role in adipogenesis of intramuscular pre-adipocytes, which provides a molecular basis for increasing IMF content in beef cattle.

G-CSF promotes the development of hepatocellular carcinoma by activating the PI3K/AKT/mTOR pathway in TAM.

OBJECTIVE: This investigation seeks to elucidate the role of the Granulocyte Colony-Stimulating Factor (G-CSF) in the progression of hepatocellular carcinoma (HCC), as well as the impact of the substance on related signaling pathways within the disease matrix. METHODS: Nude mouse tumor-bearing assay was used to detect tumor progression. Levels of Mannose/CD68 and CD34/Mannose within these samples and the concentrations of Mannose and inducible Nitric Oxide Synthase (iNOS) in macrophages were quantified using immunofluorescence techniques. The angiogenic capability was assessed via tube formation assays, and protein expressions of G-CSF, Vascular Endothelial Growth Factor (VEGF), Transforming Growth Factor-beta (TGF-ß), Matrix Metalloproteinases 2 and 9 (MMP2/9), SH2-containing protein tyrosine phosphatase-2 (SHP-2), phosphorylated PI3K/total PI3K (P-PI3K/t-PI3K), phosphorylated AKT/total AKT (P-AKT/t-AKT), and phosphorylated mTOR/total mTOR (P-mTOR/t-mTOR) were measured through Western Blot analysis in both tumor tissues and macrophages. RESULTS: Administration of G-CSF resulted in a marked augmentation of tumor volume. Macrophage Mannose expression was significantly elevated upon G-CSF treatment, while iNOS levels were conspicuously diminished. G-CSF substantially enhanced the secretion of VEGF, TGF-ß, and MMPs in tumor tissues. Macrophage parameters, following incubation in G-CSF pre-treated conditioned medium, indicated enhanced tube-forming capabilities relative to the control, an effect mitigated by the introduction of specific inhibitors. Furthermore, the G-CSF group exhibited a notable reduction in SHP-2 expression, alongside a substantial elevation in the phosphorylation levels of the PI3K/AKT/mTOR pathway proteins across all tumor-bearing paradigms. CONCLUSION: G-CSF ostensibly facilitates the advancement of hepatocellular carcinoma by activating the PI3K/AKT/mTOR signaling cascade within Tumor-Associated Macrophages (TAM).

BI-7273, a BRD9 inhibitor, reduces lipid accumulation by downregulating the AKT/mTOR/SREBP1 signaling pathway.

Increases in de novo lipogenesis that disturbed lipid homeostasis and caused lipid accumulation are a major cause of NAFLD and obesity. SREBP1 is a crucial regulatory factor controlling the expression of rate-limiting enzymes of lipid synthesis. A reduction in SREBP1expression can reduce lipid accumulation. Thus, we utilized an SREBP1-luciferase-KI HEK293 cell line constructed by our lab to screen 200 kinds of epigenetic drugs for their ability to downregulate SREBP1expression. BI-7273, an inhibitor of bromodomain-containing protein 9 (BRD9), was screened and found to decrease SREBP1 expression. What is more, BI-7273 has been confirmed that it could reduce lipid accumulation in HepG2 cells by BODIPY staining, and significantly decrease the protein expression of SREBP1 and FASN. To explore the potential mechanism BI-7273 reducing lipid accumulation, RNA sequencing (RNA-seq) was performed and demonstrated that BI-7273 reduced lipid accumulation by downregulating the AKT/mTOR/SREBP1 pathway in vitro. Finally, these results were verified in NAFLD and obesity mouse model induced by high fat diet (HFD). The results indicated that BI-7273 could decrease mouse body weight and improve insulin sensitivity, but also exhibited a strong negative correlation with serum lipid levels, and also demonstrated that BI-7273 reduced lipid accumulation via AKT/mTOR/SREBP1 pathway in vivo. In conclusion, our results revealed that BI-7273 decreases lipid accumulation by downregulating the AKT/mTOR/SREBP1 pathway in vivo and in vitro. This is the first report demonstrating the protective effect of this BRD9 inhibitor against NAFLD and obesity. BRD9 may be a novel target for the discovery of effective drugs to treat lipid metabolism disorders.

[Astragali Radix-Salviae Miltiorrhizae Radix et Rhizoma alleviates acute lung injury in rats by regulating autophagy via PI3K/Akt/mTOR pathway].

This study aims to reveal the effects of the herb pair Astragali Radix-Salviae Miltiorrhizae Radix et Rhizoma(AR-SMRR) on phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin(PI3K/Akt/mTOR) pathway and autophagy in the lung tissue of the rat model of acute lung injury(ALI). Fifty adult male SD rats were randomized into sham, model, autophagy inhibition(intraperitoneal injection of chloroquine at 10 mg·kg~(-1)), autophagy induction(intraperitoneal injection of rapamycin at 15 mg·kg~(-1)), and AR-SMRR(5 g·kg~(-1), gavage) groups. The rats in the sham group received intratracheal instillation of normal saline, and those in other groups received intratracheal instillation of lipopolysaccharide(LPS, 5 mg·kg~(-1)) for the modeling of ALI. Seven days before the operation, the rats in the sham and model groups were administrated with normal saline, and those in other groups with corresponding drugs. Specimens were collected 24 h after modeling. The pathological changes of the lung tissue were observed under a light microscope. The lung wet/dry weight ratio and the lactate dehydrogenase(LDH) activity and total protein concentration in the bronchoalveolar lavage fluid(BALF) were measured. Western blot was employed to measure the protein levels of microtubule-associated protein 1-light chain 3(LC3), beclin-1, p62, PI3K, Akt, and mTOR. Compared with the sham group, the model group showed increased histopathological score of the lung tissue, lung wet/dry weight ratio, and LDH activity and protein concentration in BALF. Autophagy inhibition further increased these indicators compared with the model group, while autophagy induction and AR-SMRR lowered the levels. In addition, AR-SMRR up-regulated the protein levels of LC3-Ⅱ and beclin-1, down-regulated the expression of p62, and inhibited the expression of p-PI3K, p-Akt, and p-mTOR in the lung tissue of ALI rats. The findings suggest that AR-SMRR can alleviate the lung injury and edema in the rat model of ALI induced by LPS by enhancing autophagy via down-regulating PI3K/Akt/mTOR signaling pathway.

Anemoside B4 attenuates necrotic enteritis of laying hens induced by Clostridium perfringens via inhibiting NF-κB and PI3K/Akt/mTOR signalling pathways.

Poultry necrotic enteritis is an important enteric disease which might be controlled by antibiotics. However, with the excessive use of antibiotics, the phenomenon of drug resistance of Clostridium perfringens is becoming increasingly prominent. Anemoside B4 exhibits important anti-inflammatory, antioxidant and immunomodulatory effects. This study was performed to estimate the effect of Anemoside B4 on chicken necrotic enteritis induced by C. perfringens in vivo and in vitro. In the in vivo experiment we investigated the efficacy of Anemoside B4 on the growth curve, biofilm formation, haemolytic activity, virulence-related gene expression and NF-κB and PI3K/AKT/mTOR activation in Caco-2 cells induced by C. perfringens. The results showed that 12.5-50 µg/mL Anemoside B4 had no antibacterial activity but could inhibit biofilm formation, attenuate haemolytic activity and virulence-related gene expression of C. perfringens and weaken NF-κB and PI3K/Akt/mTOR activation triggered by C. perfringens in Caco-2 cells. In the in vivo experiment, 60 17-day-old healthy White Leghorns were randomly divided into six groups. The growing laying hens of the control group were fed a basic diet, and those of the five challenged groups were fed a basic diet (infection group), added 0.43 g/kg Anemoside B4 (0.43 g/kg Ane group), 0.86 g/kg Anemoside B4 (0.86 g/kg Ane group), 1.72 g/kg Anemoside B4 (1.72 g/kg Ane group) and 40 mg/kg lincomycin (lincomycin group), respectively. All challenged laying hens were infected with 1 × 109 CFU C. perfringens from day 17-20. Blood and intestinal samples were obtained, and the data demonstrated that Anemoside B4 improved the blood biochemical parameters, attenuated jejunum tissue injury, increased the spleen, thymus, bursa of fabricius index, and decreased lesion scores of the jejunum and the ileum. In the jejunum, Anemoside B4 and lincomycin downregulated the expression of IL-1ß, IL-6, IL-10, TNF-α and IFN-γ at mRNA levels. Moreover, Anemoside B4 significantly enhanced both mRNA and protein levels of tight junctions ZO-1, Claudin-1 and MUC-2 in the jejunum. Anemoside B4 weakened p-P65, p-PI3K, p-Akt and p-mTOR protein expression in the jejunum infected by C. perfringens. Diets supplemented with Anemoside B4 alleviated C. perfringens-induced necrotic enteritis in laying hens by inhibiting NF-κB and PI3K/Akt/mTOR signalling pathways and improving intestinal barrier functions.

The molecular genetics of PI3K/PTEN/AKT/mTOR pathway in the malformations of cortical development.

Malformations of cortical development (MCD) are a group of developmental disorders characterized by abnormal cortical structures caused by genetic or harmful environmental factors. Many kinds of MCD are caused by genetic variation. MCD is the common cause of intellectual disability and intractable epilepsy. With rapid advances in imaging and sequencing technologies, the diagnostic rate of MCD has been increasing, and many potential genes causing MCD have been successively identified. However, the high genetic heterogeneity of MCD makes it challenging to understand the molecular pathogenesis of MCD and to identify effective targeted drugs. Thus, in this review, we outline important events of cortical development. Then we illustrate the progress of molecular genetic studies about MCD focusing on the PI3K/PTEN/AKT/mTOR pathway. Finally, we briefly discuss the diagnostic methods, disease models, and therapeutic strategies for MCD. The information will facilitate further research on MCD. Understanding the role of the PI3K/PTEN/AKT/mTOR pathway in MCD could lead to a novel strategy for treating MCD-related diseases.

Integrating network pharmacology with experimental validation to investigate the mechanism of Wuwei Zishen formula in improving perimenopausal syndrome.

OBJECTIVES: To investigate the role of the Wuwei Zishen formula (WWZSF) in treating and preventing perimenopausal syndrome (PMS) and to understand its mechanism. METHODS: Network pharmacology and molecular docking was used to predict active compounds, potential targets, and pathways for PMS treatment using WWZSF. Female Sprague-Dawley (SD) rats were induced with D-galactose (D-gal) to establish a PMS model and treated with Kunbao pill (KBP) and WWZSF. Estrus cycles were observed using vaginal smears. Serum sex hormones were measured using the enzyme-linked immunosorbent assay (ELISA). Histological changes in the uterus and ovaries were evaluated using hematoxylin-eosin staining (HE). Western blot was used to assess the protein expression levels of Cleaved Caspase-3, p62, BAX/Bcl-2, p-PI3K/PI3K, p-AKT/AKT, and p-mTOR/mTOR in the uterus and ovaries. RESULTS: A total of 70 active compounds and 440 potential targets were screened out. Important targets and pathways, including AKT1, Bcl-2, Caspase-3, mTOR, and the PI3K/AKT/mTOR pathways, and molecular docking verified their high affinities to key WWZSF components. In vivo experiments showed that WWZSF can ameliorate the morphological abnormalities of the uterus and ovaries, increase sex hormone levels and organ index, and restore the estrus cycles in PMS rats. Moreover, the western blot results showed decreased Cleaved Caspase-3 and BAX/Bcl-2 protein levels in the ovarian and uterine tissues after WWZSF therapy. Concurrently, there was an increase in the expression of p62 and the ratios of p-AKT/AKT, p-mTOR/mTOR, and p-PI3K/PI3K. CONCLUSION: The PI3K/AKT/mTOR signaling pathway-mediated apoptosis and autophagy pathways may be how WWZSF efficiently reduces PMS.

Investigating the involvement of the NLRP3/ASC/caspase-1 and NF-κb/MAPK pathways in the pathogenesis of gouty arthritis: Insights from irradiated and non-irradiated Trifolium alexandrium L. extracts and some metabolites.

ETHNOPHARMACOLOGICAL RELEVANCE: Trifolium alexandrinum L. (TA), has traditionally been used in folk medicine for its anti-inflammatory properties against hyperuricemia and gout. However, the specific mechanisms of action of TA have not been thoroughly studied. AIM OF THE WORK: This study aimed to evaluate the protective effects of irradiated (TR25) and non-irradiated (TR0) Trifolium alexandrinum L. aqueous extract (TAAE), along with two isolated compounds, caffeine (CAF) and saponin (SAP), in a rat model of acute gouty arthritis (GA). MATERIALS AND METHODS: The GA model was established by injecting a monosodium urate (MSU) suspension into the knee joint. Synovial tissue pathology was assessed, and levels of TNF-α, IL-6, IL-1ß, NF-κB, mTOR, AKT1, PI3K, NLRP3, and ASC were measured by ELISA. mRNA expression of ERK1, JNK, and p-38 MAPK was detected using qRT-PCR, and Caspase-1 protein expression was assessed by immunohistochemical analysis. Knee swelling, uric acid levels, liver and kidney function, and oxidative stress markers were also evaluated. RESULTS: TAAE analysis identified 170 compounds, with 73 successfully identified using LC-HR-MS/MS, including caffeine citrate and theasapogenol B glycoside as the main constituents. The studied materials demonstrated significant protective effects against GA. TR25 administration significantly mitigated knee joint circumference compared to other treatments. It demonstrated potential in alleviating hyperuricemia, renal and hepatic impairments induced by MSU crystals. TR25 also alleviated oxidative stress and reduced levels of IL1ß, IL-6, TNF-α, and NF-κB. Weak Caspase-1 immune-positive staining was observed in the TR25 group. TR25 decreased NLRP3 and ASC expression, reducing inflammatory cytokine levels in GA. It effectively inhibited the PI3K, AKT, and mTOR signaling pathways, promoting autophagy. Additionally, TR25 suppressed ERK1, JNK, and p-38 MAPK gene expression in synovial tissue. These effects were attributed to various components in TAAE, such as flavonoids, phenolic acids, tannins, alkaloids, and triterpenes. CONCLUSION: Importantly, irradiation (25 KGy) enhanced the antioxidant effects and phtchemical contents of TAAE. Additionally, TR0, TR25, CAF, and SAP exhibited promising protective effects against GA, suggesting their therapeutic potential for managing this condition. These effects were likely mediated through modulation of the NLRP3/ASC/Caspase-1 and ERK/JNK/p-38 MAPK signaling pathways, as well as regulation of the PI3K/AKT/mTOR pathway. Further research is warranted to fully elucidate the underlying mechanisms and optimize their clinical applications.

Fermented Ginsenosides Alleviate Acute Liver Injury Induced by CCl4 in Mice by Regulating the AKT/mTOR Signaling Pathway.

This study investigated the alleviating effect of fermented ginsenosides obtained through yeast strain fermentation transformation on acute liver injury (ALI) induced by CCl4. Strains were screened for their ability to produce ß-glucosidase, the transformation ability of the strain was verified by high-performance liquid chromatography, and the Saccharomyces cerevisiae strain F6 was obtained by 26S rRNA sequencing. After fermentation by F6 strain, it was found that the content of ginsenosides Re, Rb1, and Rb2 was significantly decreased (P < 0.05), and rare ginsenosides were detected, with the content of Rh4 and Rg5 reaching 2.65 mg·g-1 and 2.56 mg·g-1. We also explored the preventive effect of fermented ginsenoside extract (FGE) on ALI. Mice were evenly divided into 9 groups as follows: control group, ALI model group, positive drug bifendate group, and treatment group, which included 3 ginsenoside extract (GE) groups and 3 FGE groups (dosage of 150, 300, and 450 mg·kg-1 b.w.). The results showed that compared with the ALI model group, FGE significantly increased the levels of glutathione peroxidase, hydroperoxidase, and superoxide dismutase and also decreased the malondialdehyde level. The levels of alanine aminotransferase, aspartate aminotransferase, and total bilirubin markers were significantly reduced, and the levels of inflammatory cytokines TNF-α, IL-6, and IL-1ß were significantly decreased. Bioinformatics analysis combined with Western blot validation explored the molecular mechanism of the effect of FGE. It was found that FGE could downregulate the expression of the p-AKT/AKT and the p-mTOR/mTOR ratios. These results suggested that FGE played an alleviative role in ALI by promoting autophagy to inhibit the AKT/mTOR signaling pathway.

Unilateral focal palmoplantar keratoderma associated with a postzygotic variant in PIK3CA and activation of the PI3K/AKT/mTOR pathway.

Palmoplantar keratoderma (PPK) is a group of -disorders with genetic and phenotypic heterogeneity featuring skin thickening of the palms and soles. More than 60 genes involved in various biological processes are implicated in PPK. PIK3CA is an oncogene encoding p110α, and its somatic variants contribute to a spectrum of congenital overgrowth disorders, including epidermal nevi (EN). To identify the genetic basis and elucidate the pathogenesis of a patient with unilateral focal PPK. Whole-exome sequencing and Sanger sequencing combined with laser capture microdissection (LCM) were performed on genomic DNA extracted from the patient's peripheral blood and skin lesion. Skin biopsies were taken from the lesion of the patient and normal controls for immunofluorescence. Molecular docking was performed using Alphafold2-multimer. A three-year-old girl presented with unilateral focal PPK with an identified missense -variant (c.3140A>G, p.His1047Arg) in PIK3CA from affected tissue. This variant only existed in the lesional epidermis. Elevated PI3K/AKT/mTOR signalling in the affected epidermis and an increased number of Ki67-positive keratinocytes were demonstrated. Molecular docking indicated instability of the p110α-p85α dimer caused by the PIK3CA His1047Arg variant. We describe the first PPK case associated with a variant in PIK3CA, which expands the spectrum of PIK3CA-related disorders. Our study further underscores the importance of the PI3K/AKT/mTOR pathway in the homeostasis of skin keratinization.

Downregulation of PRKCI inhibits osteosarcoma cell growth by inactivating the Akt/mTOR signaling pathway.

PRKCI is abnormally expressed in various cancers, but its role in osteosarcoma is unknown. This study aimed to explore the biological function of PRKCI in osteosarcoma and its potential molecular mechanism. PRKCI expression was evaluated in osteosarcoma cell lines using Western blot analysis and reverse transcription PCR. The CCK-8 assay, colony formation assay, flow cytometry, Transwell assay, and wound-healing assay were used to detect the proliferation, colony-forming capacity, cell cycle, migration, and invasion of osteosarcoma cells when PRKCI was overexpressed or knocked down. The interaction between PRKCI and SQSTM1 was explored using immunoprecipitation. Finally, the protein molecule expression of the Akt/mTOR signaling pathway in osteosarcoma was detected when PRKCI was knocked down. Our study found that PRKCI was overexpressed in osteosarcoma cell lines. The overexpression of PRKCI promoted the proliferation and colony-forming capacity of osteosarcoma cells, while silencing PRKCI inhibited the proliferation, colony-forming capacity, migration, and invasion of osteosarcoma cells and arrested the cell cycle at the G2/M phase. Both PRKCI and SQSTM1 were overexpressed in osteosarcoma. The expression of PRKCI was only related to histological type, while that of SQSTM1 was not related to clinical characteristics. The expression of PRKCI and SQSTM1 in osteosarcoma was higher than that in chondrosarcoma. Knockdown of PRKCI inhibited the proliferation of osteosarcoma cells by inactivating the Akt/mTOR signaling pathway, suggesting that PRKCI was a potential target for osteosarcoma therapy.

B. glomerulata promotes neuroprotection against ischemic stroke by inhibiting apoptosis through the activation of PI3K/AKT/mTOR pathway.

BACKGROUND: Brassaiopsis glomerulata (Blum) Regel (B.glomerulata) is recognized as a traditional Chinese medicine (TCM) primarily used for promoting blood circulation and removing stasis. It is frequently utilized in the treatment of injuries resulting from falls and bumps. PURPOSE: Despite its effective use in clinical treatment for ischemic stroke (IS), there are currently no reports on its composition and mechanism of action, which affects its promotion. The study investigated the chemical components and molecular mechanisms of B.glomerulata, with the following components: UPLC-Q-TOF-MS, network pharmacology Analysis and experimental verification in vivo and vitro. METHODS: The effect of B.glomerulata on interfering with ischemic stroke was assessed on MCAO/R rats and ORD cell model. Then the compositional analysis was conducted using UPLC-Q-TOF-MS. Furthermore, network pharmacology and molecular docking techniques were explored to identify potential targets and pathways. The predicted mechanisms of action were ultimately confirmed by immunohistochemistry and protein blotting. RESULTS: B. glomerulata exhibited neuroprotective effects in MCAO/R rats by reductions in hippocampal and cortical neuronal damage, brain infarction, and cerebral edema. Both in vivo and in vitro experiments demonstrated that it decreased ROS and MDA levels, increased SOD and GSH levels, thereby inhibiting oxidative stress. Moreover, the improvements in neuronal morphology and the modulation of Nissl bodies suggested a potential mechanism underlying its neuroprotective action. Additionally, B.glomerulata exhibited concentration-dependent reductions in Bax and Caspase-3 expressions, along with increases in GFAP, Bcl2/Bax ratio, p-PI3K, p-AKT, and p-mTOR levels. CONCLUSION: B.glomerulata exhibited neuroprotective effects against cerebral ischemia-reperfusion injury both in vivo and in vitro. It prevented oxidative stress damage and inhibited apoptosis of ischemic stroke through the PI3K/AKT/mTOR pathway.