Dehydroepiandrosterone modulates the PTEN/PI3K/AKT signaling pathway to alleviate 4-vinylcyclohexene diepoxide-induced premature ovarian insufficiency in rats.

Dehydroepiandrosterone (DHEA) is frequently integrated as an adjuvant in over a quarter of controlled ovarian hyperstimulation (COH) protocols, despite the ongoing debate regarding its impact. This study aimed to evaluate the efficacy and mechanism of action of DHEA on ovarian follicular development and ovarian response in rats with varying ovarian reserves. The study involved 75 rats categorized into 15 distinct groups. The ovarian tissues of rats in both the normal ovarian reserve group and the premature ovarian insufficiency (POI) group, induced by 4-vinylcyclohexene diepoxide (VCD) injection, were subjected to histomorphological and biochemical analyses following the administration of DHEA, either alone or in combination with COH. Follicle counting was performed on histological sections obtained from various tissues. Serum concentrations of anti-Müllerian hormone (AMH) and the quantification of specific proteins in ovarian tissue, including phosphatase and tensin homolog of chromosome 10 (PTEN), phosphoinositide 3-kinase (PI3K), phosphorylated protein kinase B (pAKT), cyclooxygenase 2 (COX-2), caspase-3, as well as assessments of total antioxidant status and total oxidant status, were conducted employing the ELISA method. The impact of DHEA exhibited variability based on ovarian reserve. In the POI model, DHEA augmented follicular development and ovarian response to the COH protocol by upregulating the PTEN/PI3K/AKT signaling pathway, mitigating apoptosis, inflammation, and oxidative stress, contrary to its effects in the normal ovarian reserve group. In conclusion, it has been determined that DHEA may exert beneficial effects on ovarian stimulation response by enhancing the initiation of primordial follicles and supporting antral follicle populations.

Qinggan Huoxue Recipe attenuates Alcoholic Liver Disease by suppressing PI3K/AKT signaling pathway based on network pharmacology.

Qinggan Huoxue Recipe (QGHXR) is originated from Xiao Chaihu Decotion. Many experimental studies have confirmed that QGHXR can significantly alleviate the symptoms of alcoholic liver disease (ALD), but the detailed mechanism is still unclear. Using traditional Chinese medicine network pharmacology analysis system database and animal experiments, we found that 180 potentially chemical compositions and 618 potential targets were screened from the prescription, which shared 133 signal pathways with ALD. Through animal experiments, it was found that QGHXR could reduce the liver total cholesterol (TC), serum TC, alanine aminotransferase, aspartate aminotransferase of ALD mice, reduce the lipid droplets and inflammatory injury of liver tissue. Meanwhile, it can also increase PTEN, decrease PI3K and AKT mRNA levels. In this study, we obtained the targets and pathways of QGHXR in the treatment of ALD, and preliminatively verified that QGHXR may improve ALD through PTEN/PI3K/AKT signaling pathway.

Protective effect of total saponins of ginseng stems and leaves (GSLS) on chlorpyrifos-induced brain toxicity in mice through the PTEN/PI3K/AKT axis.

Chlorpyrifos (CPF) is a class of toxic compounds which has been widely used in agriculture that can cause multi-organ damage to the liver, kidneys, testes, and nervous system. Currently, most studies on ginseng have concentrated on the roots and rhizomes, and less research has been conducted on the above-ground parts. Our laboratory found that ginseng stem and leaf total saponin (GSLS) features strong antioxidant activity. In this experiment, we selected different concentrations of CPF to induce hippocampal neuronal cell injury model in mice, conducted a cell survival screening test, and also selected appropriate concentrations of CPF to induce brain injury model in mice. CCK-8, flow cytometry, Elisa, Hoechst 33258 staining, Annexin V-FITC/PI staining, HE staining, Morris water maze, and qRT-PCR were adopted for detecting the effects of GSLS treatment on CPF-induced cell viability, mitochondrial membrane potential, reactive oxygen species (ROS) levels, Ca2+ concentration and GSLS treatment on CPF-induced brain injury and related signaling in mice, respectively. The effects of GSLS treatment on CPF-induced brain injury and the related signaling pathways in mice were examined. The results showed that GSLS at 60 µg/ml and 125 µg/ml concentrations elevated the viability of CPF-induced HT22 cells, increased mitochondrial membrane potential, depleted ROS, decreased Ca2+ concentration, and decreased apoptosis rate. Meanwhile, GSLS treatment significantly reduced CPF-induced escape latency in mice, elevated the number of entries into the plateau and effective area, increased the effective area and target quadrant residence time, as well as improved the pathological damage of mouse hippocampal neurons. The results of mouse brain sections demonstrated that GSLS treatment significantly increased SOD and CAT activities and lowered MDA accumulation in CPF-induced mice. qRT-PCR revealed that PTEN mRNA expression was significantly decreased with PI3K and AKT expression being significantly increased in GSLS-treated CPF-induced mice. Thus, the obtained results indicate that GSLS can effectively antagonize CPF-induced brain toxicity in mice through regulating PTEN/PI3K/AKT pathway.

[Shenbai Jiedu Fang inhibits AOM/DSS-induced colorectal adenoma formation and carcinogenesis in mice via miRNA-22-mediated regulation of the PTEN/PI3K/AKT signaling pathway].

OBJECTIVE: To observe the inhibitory effect of Shenbai Jiedu Fang (SBJDF, a compound recipe of traditional Chinese herbal drugs) on chemically induced carcinogenesis of colorectal adenoma in mice and explore the role of PTEN/PI3K/AKT signaling pathway in mediating this effect. METHODS: Four-week-old male C57BL/6 mice were randomly divided into control group (n=10), AOM/DSS model group (n=20), low-dose (14 g/kg) SBJDF group (n=10) and high-dose (42 g/kg) SBJDF group (n= 10). In the latter 3 groups, the mice were treated with azoxymethane (AOM) and dextran sodium sulphate (DSS) to induce carcinogenesis of colorectal adenoma. In the two SBJDF treatment groups, SBJDF was administered daily by gavage during the modeling. The survival rate, body weight, general condition of the mice, and intestinal adenoma formation and carcinogenesis were observed. The expressions of proteins associated with the PTEN/PI3K/AKT signaling pathway in the intestinal tissue were detected using immunohistochemistry. RESULTS: Compared with those in the model group, the mice treated with SBJDF, especially at the high dose, showed a significantly lower incidence of intestinal carcinogenesis and had fewer intestinal tumors with smaller tumor volume. Pathological examination showed the occurrence of adenocarcinoma in the model group, while only low-grade and high-grade neoplasia were found in low-dose SBJDF group; the mice treated with high-dose SBJDF showed mainly normal mucosal tissues in the intestines with only a few lesions of low-grade neoplasia of adenoma. Compared with those in the control group, the mice in the model group had significantly elevated plasma miRNA-222 level (P < 0.05), which was obviously lowered in the two SBJDF groups (P < 0.01). The results of immunohistochemistry revealed that compared with the model group, the two SBJDF groups, especially the high-dose group, had significantly up-regulated expressions of PTEN, P-PTEN and GSK-3ß and down-regulated expressions of p-GSK-3 ß, PI3K, AKT, P-AKT, ß-catenin, c-myc, cyclinD1 and survivin in the intestinal tissues. CONCLUSION: SBJDF can significantly inhibit colorectal adenoma formation and carcino-genesis in mice possibly through regulating miRNA-222 and affecting PTEN/PI3K/AKT signaling pathway.

miR-21 antagonist alleviates colitis and angiogenesis via the PTEN/PI3K/AKT pathway in colitis mice induced by TNBS.

Background: The pathogenesis of Crohn's disease (CD) is unknown; however, angiogenesis is known to play an important role in the disease. The present research suggests that microRNA-21 (miR-21) may play a positive regulatory role in disordered angiogenesis in CD. Methods: C57 wild-type mice were divided into 6 groups. On day 0, all mice in the 2,4,6-trinitrobenzenesulfonic acid (TNBS) group were given an enema at the concentration of TNBS 100 mg/kg mouse body weight (solvent 50% alcohol). In the control group, the enema was performed with 50% alcohol. On day 0, 2, 4, and 6, the mice of the agomir-21 + TNBS group and the agomir control + TNBS group were injected with 200 µL, 5 nmol agomir-21 or agomir control [dissolved in ribonuclease (RNase)-free water] by tail vein injection, while the antagomir-21 + TNBS group and the antagomir control + TNBS group were injected with 200 µL, 20 nmol antagomir-21 or antagomir control (dissolved in RNase-free water). The body weight and disease activity index (DAI) score were recorded daily. The colons were obtained to assess macro and microscopic colon damage. The inferior vena cava and the accompanying abdominal aorta were chosen to detect the protein expression of the phosphatase and tensin homolog (PTEN)/phosphoinositide 3-kinase (PI3K)/serine/threonine kinase (AKT)/vascular endothelial growth factor (VEGF) axis through western blotting. Serum interleukin-1 beta (IL-1ß) and tumor necrosis factor alpha (TNF-α) were detected by enzyme-linked immunosorbent assay (ELISA). The distribution and expression of neovascularization were demonstrated by cluster of differentiation 31 (CD31) immunohistochemistry. Results: Compared with the only-TNBS group, the agomir-21 + TNBS group showed significantly severer colitis symptoms and more abnormal vascular hyperplasia, while the antagomir-21 + TNBS group showed symptom relief and reduced vascular hyperplasia. In addition, agomir-21 obviously inhibited the expression of PTEN and activated the PI3K/AKT/VEGF pathway in mice induced by TNBS, while antagomir-21 effectively antagonized this effect. Conclusions: miR-21 can promote the progression of colitis in mice induced by TNBS and aggravate the disordered angiogenesis by regulating the PTEN/PI3K/AKT axis. Intravenous injection of miR-21 antagonists can effectively relieve the symptoms of colitis and inhibit colonic angiogenesis.

Effect of Fushengong Decoction on PTEN/PI3K/AKT/NF-κB Pathway in Rats With Chronic Renal Failure via Dual-Dimension Network Pharmacology Strategy.

Overview: The treatment of chronic renal failure (CRF) with traditional Chinese medicine has attracted much attention, but its mechanism is not clear. Network pharmacology is an effective strategy for exploring the interaction mechanisms between Chinese herbs and diseases, however, it still needs to be validated in cell and/or animal experiments due to its virtual screening characteristics. Herein, the anti-CRF mechanism of the Fushengong decoction (FSGD) was investigated using a dual-dimension network pharmacological strategy combined with in vivo experiment. Methods: The traditional Chinese medicine systems pharmacology (TCMSP) database (https://tcmspw.com) and UHPLC-MS/MS technology were used to identify the effective compounds of FSGD in theory and practice, such as quercetin, formononetin, and pachymic acid. The putative targets of FSGD and CRF were obtained from the Swisstarget prediction platform and the Genecards database, respectively. The common target pathways between FSGD and CRF were got from the dual-dimension network pharmacology analysis, which integrated the cross-common targets from the TCMSP components-Swisstarget-Genecards-Venn platform analysis in theory, and the UHPLC-MS/MS identified effective ingredients-Swisstarget screening, such as TNF and PI3K/AKT. Furthermore, system molecular determinations were used to prove the dual-dimension network pharmacology study through CRF rat models, which were constructed using adenine and treated with FSGD for 4 weeks. Results: A total of 121 and 9 effective compounds were obtained from the TCMSP database and UHPLC-MS/MS, respectively. After dual-dimension network pharmacology analysis, the possible mechanism of PTEN/PI3K/AKT/NF-κB pathway was found for FSGD in CRF. In vivo experiments indicated that FSGD can play a role in protecting renal function and reducing fibrosis by regulating the PTEN/PI3K/AKT/NF-κB pathway. These findings provide a reference for FSGD in CRF. Conclusion: Based on the theoretical and practical dual-dimension network pharmacology analysis for FSGD in CRF, the possible molecular mechanism of PTEN/PI3K/AKT/NF-κB was successfully predicted, and these results were verified by in vivo experiments. In this study, the dual-dimension network pharmacology was used to interpret the key signal pathway for FSGD in CRF, which also proved to be a smart strategy for the study of effective substances and pharmacology in FSGD.

The Role of PTEN/PI3K/AKT Signaling Pathway in Apoptosis of Liver Cells in Cocks with Manganese Toxicity.

PTEN/PI3K/AKT signaling pathway is an important pathway for cell proliferation and apoptosis. Exposure to excess manganese (Mn) can cause damage in organisms. However, whether Mn toxicity can cause apoptosis is still not clear. In order to explore the mechanism of PTEN/PI3K/AKT signaling pathway responsible for Mn-induced apoptotic injury, 160 Hyline cocks were divided into four groups; there were the control group (Con group), the low-dose Mn group (L group), the medium-dose Mn group (M group), and the high-dose Mn group (H group). The cocks in Con group, L group, M group, and H group were fed with MnCl2 diet containing 100, 600, 900, and 1800 mg/kg, respectively. The growth status of cocks in each group was observed on days 30, 60, and 90. Thirty cocks were randomly selected from each group and sacrificed on day 90 for optical microscope observation and fluorescence microscopic observation, as well as for transcription-level expression of apoptosis-related genes and heat shock proteins (HSPs) in the liver. The results showed that the growth status of cocks was gradually depressed with the extension of feeding time and with the increase of Mn dose. On day 90, the results of optical microscope observation and fluorescence microscope observation showed that damage and apoptosis appeared in the cock liver cells under Mn exposure groups. The results of transcription-level detection of apoptosis-related genes and HSPs indicated that Mn exposure upregulated eleven pro-apoptotic genes (including RIP1, RIP3, MLKL, Bax, Caspase-3, FADD, Cyt-C, ERK, JNK, Caspase-8, and P38) and downregulated one anti-apoptotic gene Bcl-2, further meaning that exposure to Mn-induced apoptosis in cock liver cells and PTEN/PI3K/AKT signaling pathway took part in molecular mechanism of apoptosis caused by excess Mn. Moreover, in our experiment, the increase of four HSPs (including HSP27, HSP40, HSP60, and HSP70) was found after Mn treatment for 90 days, which indicated that Mn stress triggered HSPs and HSPs were involved in molecular mechanism of Mn poisoning in cock livers. In addition, we also found there was upregulated dose-dependent manner in fifteen detected genes and there was downregulated dose-dependent manner in Bcl-2, indicating that the apoptosis caused by Mn poisoning in cock liver cells was dose-dependent.

[Xinfeng capsule inhibits immune inflammation in osteoarthritis by inhibiting the miR- 23a-3p/PETN/PI3K/AKT/mTOR pathway].

OBJECTIVE: To observe the inhibitory effect of Xinfeng capsule (XFC) on immune inflammatory response in the coculture system of chondrocytes and CD4+ T cells from patients with osteoarthritis (OA). OBJECTIVE: Thirty OA patients and 30 healthy subjects were examined for the expression of miR-23a-3p/PTEN/PI3K/AKT/mTOR. The changes in the miR-23a-3p/ PTEN/PI3K/AKT/mTOR pathway and the clinical indicators of the OA patients after XFC treatment were observed. OA-CD4+ T cells and OA-Chondrocytes cells were cultured in Transwel chambers, and the expressions of miR-23a-3p, PTEN, PI3K, AKT and mTOR mRNA were detected by RT-PCR; the levels of IL-1ß, IL-6, IL-10 and TNF-α were detected by ELISA. The protein expressions of PTEN, PI3K, AKT, p-AKT, mTOR and p-mTOR were detected by Western blotting. OBJECTIVE: Compared with healthy individuals, OA patients showed significantly increased expressions of miR-23a-3p, PTEN, PI3K, AKT, IL-1ß, IL-6 and TNF-α and lowered expressions of PTEN and IL-10 (P < 0.01). Positive correlations were found between miR-23a-3p and IL-6 and between PI3K and IL-10; mTOR was positively correlated with IL-6, IL-10, complement C3 and C4 (P < 0.01 or 0.05). Intervention with XFC obviously down-regulated the expressions of IL-1ß, IL-6, and TNF-α and up-regulated the expression of IL-10 (P < 0.01). In the cell co-culture systems, the expressions of miR-23a-3p, PI3K, Akt, mTOR, IL-1ß, IL-6 and TNF-α were up-regulated while the expressions of PTEN and IL-10 were down-regulated in the model group (P < 0.01). Overexpression of miR-23a-3p significantly up-regulated the expressions of IL-1ß, IL-6, TNF-α, miR-23a-3p, PI3K, AKT and mTOR and lowered the expressions of PTEN and IL-10 (P < 0.01 or 0.05). The expressions of IL-1ß, IL-6, TNF-α, miR-23a-3p, PI3K, AKT and mTOR were down-regulated and the expressions of IL-10 and PTEN were up-regulated in XFC-treated cells (P < 0.01 or 0.05). OBJECTIVE: XFC can reduce the inflammatory response in patients with OA by down-regulating the expression of miR-23a-3p, inhibiting the activation of the PTEN/PI3K/AKT/ mTOR pathway, and regulating the expression of IL-1ß, IL-6, IL-10 and TNF-α.

Rosmarinic acid exerts an anticancer effect on osteosarcoma cells by inhibiting DJ-1 via regulation of the PTEN-PI3K-Akt signaling pathway.

BACKGROUND: Osteosarcoma is the most common type of primary malignant bone tumor. This disease has exhibited a progressively lower survival rate over the past several decades, which has resulted in it becoming a main cause of death in humans. Rosmarinic acid (RA), a water-soluble polyphenolic phytochemical, exerts powerful anticancer effects against multiple types of cancer; however, its potential effects on osteosarcoma remain unknown. Hence, the present study investigated the efficacy of RA against osteosarcoma and aimed to clarify the mechanisms underlying this process. METHODS: The effects of RA on cell viability, apoptosis, cell cycle distribution, migration, invasion, and signaling molecules were analyzed by CCK-8 assay, flowcytometric analysis, wound healing assay, Transwell assay, proteomic analysis, and use of shRNAs. RESULTS: RA exerted anti-proliferation and pro-apoptotic effects on U2OS and MG63 osteosarcoma cells. Apoptosis was induced via extrinsic and intrinsic pathways by increasing the Bax/Bcl-2 ratio, triggering the intracellular production of reactive oxygen species (ROS), reducing the mitochondrial membrane potential (MMP), and upregulating the cleavage rates of caspase-8, caspase-9, and caspase-3. Additionally, RA suppressed the migration and invasion of osteosarcoma cells by inhibiting the expression levels of matrix metalloproteinase-2 and -9 (MMP-2 and -9), which are associated with a weakening of the epithelial-mesenchymal transition (EMT). Moreover, proteomic analyses identified DJ-1 as a potential target for RA. Several studies have indicated an oncogenic role for DJ-1 using knockdowns via the lentiviral-mediated transfection of shRNA, which caused the conspicuous suppression of cell proliferation, migration, and invasion as well as the arrest of cell cycle progression. At the molecular level, the expression levels of DJ-1, p-PI3K, and p-Akt were reduced, whereas the protein levels of phosphatase and tensin homologue (PTEN) were increased. CONCLUSION: In conjunction with the high levels of DJ-1 expression in osteosarcoma tissues and cell lines, the present results suggested that RA exhibited anticancer effects in osteosarcoma cells by inhibiting DJ-1 via regulation of the PTEN-PI3K-Akt signaling pathway. Therefore, DJ-1 might be a biological target for RA in osteosarcoma cells.

Celastrol inhibits growth and metastasis of human gastric cancer cell MKN45 by down-regulating microRNA-21.

Celastrol could inhibit cancer cell growth in vitro. However, effect(s) of celastrol on gastric cancer is not well studied. Therefore, we investigated the effects of celastrol on human gastric cancer cell line MKN45 and the underlying mechanisms. We found that celastrol inhibited cell proliferation, migration, and invasion and induced cell apoptosis and G2/M cell cycle arrest (p < .05, p < .01, or p < .001). Under celastrol treatment, overexpression of microRNA-21 (miR-21) increased cell viability, migration, and invasion and inhibited cell apoptosis compared with negative control (p < .05, p < .01, or p < .001). In addition, the phosphorylation of PTEN was significantly up-regulated, whereas PI3K, AKT, p65, and IκBα phosphorylation was statistically decreased by celastrol (p < .05 or p < .01) and then further reversed by miR-21 overexpression (p < .05 or p < .01). On the other side, miR-21 silence showed contrary results (p < .05) as relative to miR-21 overexpression. In conclusion, celastrol inhibits proliferation, migration, and invasion and inactivates PTEN/PI3K/AKT and nuclear factor κB signaling pathways in MKN45 cells by down-regulating miR-21.

RETRACTED: Lycium barbarum polysaccharides promote osteoblasts viability by regulating microRNA-17/PTEN.

This article has been retracted: please see Elsevier Policy on Article Withdrawal (https://www.elsevier.com/about/our-business/policies/article-withdrawal). This article has been retracted at the request of the Editor-in-Chief. Given the comments of Dr Elisabeth Bik regarding this article "… the Western blot bands in all 400+ papers are all very regularly spaced and have a smooth appearance in the shape of a dumbbell or tadpole, without any of the usual smudges or stains. All bands are placed on similar looking backgrounds, suggesting they were copy/pasted from other sources, or computer generated", the journal requested the authors to provide the raw data. However, the authors were not able to fulfil this request and therefore the Editor-in-Chief decided to retract the article.

PI3K/Akt Cooperates with Oncogenic Notch by Inducing Nitric Oxide-Dependent Inflammation.

The PI3K/Akt signaling pathway, Notch, and other oncogenes cooperate in the induction of aggressive cancers. Elucidating how the PI3K/Akt pathway facilitates tumorigenesis by other oncogenes may offer opportunities to develop drugs with fewer side effects than those currently available. Here, using an unbiased in vivo chemical genetic screen in Drosophila, we identified compounds that inhibit the activity of proinflammatory enzymes nitric oxide synthase (NOS) and lipoxygenase (LOX) as selective suppressors of Notch-PI3K/Akt cooperative oncogenesis. Tumor silencing of NOS and LOX signaling mirrored the antitumor effect of the hit compounds, demonstrating their participation in Notch-PI3K/Akt-induced tumorigenesis. Oncogenic PI3K/Akt signaling triggered inflammation and immunosuppression via aberrant NOS expression. Accordingly, activated Notch tumorigenesis was fueled by hampering the immune response or by NOS overexpression to mimic a protumorigenic environment. Our lead compound, the LOX inhibitor BW B70C, also selectively killed human leukemic cells by dampening the NOTCH1-PI3K/AKT-eNOS axis.