Expression Of Phosphoinositide 3-kinase Research Articles

Disrupted Phosphorylated Phosphoinositide 3-kinase in the NTS of Type 2 Diabetic Rats

The hallmark of type 2 diabetes mellitus (T2DM) is systemic insulin resistance, where insulin signaling is impaired, blood glucose is chronically elevated, and various pathophysiological processes ensue. Much is known about the cardiovascular and metabolic consequences of T2DM. However, the impact of T2DM on autonomic nervous system function remains to be fully elucidated. Increasing evidence suggests that in both human and animal models of T2DM, the exercise pressor reflex (EPR) is abnormally augmented, characterized by heightened cardiovascular and sympathetic responses to physical activity. The underlying mechanisms are still not completely understood. Our recent studies suggest that T2DM induces alterations in peripheral somatosensory nerve activity that could explain the augmented EPR function. However, it is possible that neurons within the cardiovascular control areas of the brainstem, such as the nucleus tractus solitarius (NTS), could also be impaired by this disease. In the central nervous system, insulin has been shown to hyperpolarize neurons through activation of the phosphoinositide 3-kinase (PI3K) pathway, resulting in reduced neuronal excitability. Therefore, we hypothesized that decreased insulin transport into the brain in T2DM rats, results in increased cellular excitability as evidenced by decreased activity of the PI3K pathway in the NTS. Purpose: The aim of this study was to examine alterations in brain insulin levels and the PI3K pathway in T2DM rats as compared to control rats. Methods: Control rats were fed a low fat control diet (13% fat, 67.8% carbohydrate, and 19.2% protein) for 12-16 wks, while T2DM were generated by two low dose bolus injections of i.p. streptozotocin (35mg/kg, week 1; 25mg/kg week 2) and then fed a high fat diet (42% fat, 42.8% carbohydrate, and 15.2% protein) for 12-16wks. Cerebrospinal fluid (CSF) were collected from control and T2DM rats after an overnight fast. NTS micro-punches were performed, and then total and phosphorylated PI3K protein expressions were examined by the western blot technique. Results: CSF insulin was significantly decreased in T2DM rats as compared to control rats (0.41 ± 0.19 vs 0.11 ± 0.05 (ng/mL), Control (n=14) vs T2DM (n=4), p<0.01). Total PI3K expression in the NTS did not differ between control and T2DM (1.0 ± 0.2 vs 1.1 ± 0.2 (au), Control (n=9) vs T2DM (n=9), P=0.172). However, phosphorylated PI3K expression in the NTS of T2DM was significantly decreased as compared with control animals (1.0 ± 0.2 vs 0.8 ± 0.1 (au), Control (n=9) vs T2DM (n=9), p<0.05). Conclusion: These results suggest that 1) in consistent with earlier studies, insulin transport into the brain is impaired in high fat diet/streptozotocin-induced T2DM animals, and 2) decreased phospho-PI3K signaling in the NTS of diabetic animals is associated with lower brain insulin. Taken together, these alterations may increase neuronal excitability in the NTS, thereby contributing, at least in part, to the exaggerated EPR response to exercise in this disease. Supported by NIH HL-151632 and NIH HL-151632-01S1. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Network pharmacology prediction and experiment validation of anti liver cancer activity of curcumae rhizoma and hedyotis diffusa willd

Objective: This study aims to elucidate anti liver cancer components and potential mechanisms of Curcumae Rhizoma and Hedyotis diffusa Willd (CR-HDW). Methods: Effective components and targets of CR-HDW were identified from the Traditional Chinese Medicine Systems Pharmacology (TCMSP) database. Liver cancer-related genes were collected from GeneCards, Gene-Disease Association (DisGeNET), and National Center for Biotechnology Information (NCBI). Protein-protein interaction networks, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment were conducted to analyze the identified genes. Molecular docking was used to simulate binding of the active components and their target proteins. Cell activity assay, Western blot, and Senescence-associated β-galactosidase (SA-β-gal) experiments were conducted to validate core targets identified from molecular docking. Results: Ten active compounds of CR-HDW were identified including quercetin, 3-epioleanic acid and hederagenin. The primary core proteins comprised Glyceraldehyde-3-phosphate dehydrogenase (GAPDH), Protein Kinase B（AKT1）, etc. The pathways for Phosphoinositide 3-kinase (PI3K)/ AKT, cellular senescence, Fork head boxO (FOXO) were revealed as important for anti-cancer activity of CR-HDW. Molecular docking demonstrated strong binding between liver cancer target proteins and major active components of CR-HDW. In vitro experiments confirmed that hederagenin and 3-epioleolic acid inhibited HuH-7 cell growth, reduced expression of PI3K, AKT, and mechanistic target of rapamycin (mTOR) proteins. Hederagenin also induced HuH-7 senescence. Conclusion: In summary, our results suggest that the CR-HDW component (Hederagenin, 3-epoxy-olanolic acid) can inhibit the proliferation of HuH-7 cells by decreasing PI3K, AKT, and mTOR. Hederagenin also induced HuH-7 senescence.

Potential mitigating impact of a dipeptidyl peptidase-IV inhibitor, vildagliptin, on oxazolone-induced ulcerative colitis: Targeting the role of PI3K/AKT/mTOR and AMPK/Nrf2 signaling pathways

Growing evidence suggests that phosphoinositide 3-kinase (PI3K) and adenosine monophosphate-activated protein kinase (AMPK) signaling cascades are critical in ulcerative colitis (UC) pathophysiology by influencing gut mucosal inflammation. Recently, the coloprotective properties of dipeptidyl peptidase-IV (DPP-IV) inhibitors have emerged. Thus, this study assessed for the first time the potential mitigating impact of a DPP-IV inhibitor, vildagliptin (Vilda), on oxazolone (OXZ)-induced colitis in rats, targeting the role of PI3K/AKT/mTOR and AMPK/Nrf2 pathways. Thirty-two adult Albino rats were divided into four groups: control, Vilda (10 mg/kg/day orally), OXZ (300 µL of 5 % OXZ in 50 % aqueous ethanol solution introduced once into the colon via catheter), and Vilda+OXZ. Inflammatory cytokines (interleukin 13, tumor necrosis factor-α, interleukin 10), oxidative/endoplasmic reticulum stress markers (myeloperoxidase, reduced glutathione, catalase, CHOP), mitochondrial reactive oxygen species, adenosine triphosphate levels, and mitochondrial transmembrane potential were estimated. p-AMPK, p-AKT, beclin-1, and SQSTM1 levels were immunoassayed. Nrf2, PI3K, and mTOR expression levels were quantified using the real-time polymerase chain reaction. Furthermore, p-NF-ĸBp65 and LC3II immunoreactivity were evaluated. Vilda administration effectively ameliorated OXZ-induced colitis, as evidenced by the reduced Disease Activity Index, macroscopic colon damage score, colon weight/length ratio, ulcer index, and histopathological and electron microscopic changes in the colon tissues. Vilda treatment also counteracted OXZ-triggered inflammation, oxidative/endoplasmic reticulum stress, mitochondrial dysfunction, and enhanced autophagy in the colon. Vilda substantially suppressed PI3K/AKT/mTOR and activated the AMPK/Nrf2 pathway. Vilda has potent coloprotective and anti-ulcerogenic properties, primarily attributed to its antiinflammatory, antioxidant, and modulatory impact on mitochondrial dysfunction and autophagy activity. These effects were mostly mediated by suppressing PI3K/AKT/mTOR and activating AMPK/Nrf2 signaling cascades, suggesting a potential role of Vilda in UC therapy.

Cytotoxic activity of bimetallic Ag@Se green synthesized nanoparticles using Jerusalem Thorn (Parkinsonia aculeata).

Introduction: The process of green synthesis of metal nanoparticles is considered to be eco-friendly and cost-effective. Methods: In this study, bimetallic Ag@Se-P and Ag@Se-S nanoparticles were synthesized successfully using Parkinsonia aculeata aerial parts and seed extracts. The phytochemical contents in P. aculeata aerial parts and seed aqueous extract serve as reducing and stabilizing capping agents without the need for any chemical stabilization additive in the synthesis of bimetallic nanoparticles. Result and Discussion: The obtained results from UV-vis spectrophotometry, scanning electron microscopy (SEM), X-ray powder diffraction (XRD), energy-dispersive X-ray spectroscopy (EDS), transmission electron microscopy (TEM), and Fourier-transform infrared spectroscopy (FT-IR) confirmed the successful synthesis of bimetallic nanoparticles with cluster irregular spherical morphology, crystalline nature, and average particle sizes of 17.65 and 24.36nm for Ag@Se-S and Ag@Se-P, respectively. The cytotoxicity assessment of greenly synthesized nanomaterials using seed and plant extracts showed cell inhibition >50μg/mL. Ag@Se-S and Ag@Se-P seed and plant extracts significantly reduced LPS-induced inflammation, which was assessed by NO and cytokines IL-1β, IL-6, and TNF-α. The mRNA and protein expression levels of phosphoinositide 3 kinase (PI3K) and nuclear factor kappa B (NFkB) were significantly overexpressed in LPS-induced RAW 264.7 cell lines. Ag@Se-S and Ag@Se-P downregulated the expression of PI3K and NFkB in LPS-induced cell models.

Methanolic extract of Cleome droserifolia mitigates epinephrine-induced cardiac injury

Ethnopharmacological relevanceCleome droserifolia treatment has potent antioxidant, antimicrobial, and immunomodulatory potentials, which can help recover the general health status. Aim of the studyThis research aims to estimate the potential cardioprotective properties of the methanolic extract of Cleome droserifolia (MEC) against epinephrine-induced cardiac injury (MI) in rats. Materials and methodsThirty-six male Wistar rats were separated equally into 6 groups. The control group (Cont) received oral distilled water for 30 uninterrupted days and was administered subcutaneous saline on the 31st and 32nd days. The MEC 100 group received MEC 100 mg/kg, P.O., for 30 uninterrupted days and administered subcutaneous saline on the 31st and 32nd days. The MEC 200 group received MEC 200 mg/kg, P.O. for 30 uninterrupted days and subcutaneous saline was administered on the 31st and 32nd days. The epinephrine group (EP) received distilled water orally for 30 uninterrupted days and administered subcutaneous epinephrine (2 mg/kg, s.c.) divided into two doses (1 mg/kg, s.c) each on the 31st and 32nd days. MEC 100+Ep group received MEC 100 mg/kg, P.O., for 30 uninterrupted days and administered subcutaneous epinephrine (2 mg/kg, s.c.) divided into two doses (1 mg/kg, s.c) each on the 31st and 32nd days. The MEC 200+EP received MEC 200 mg/kg, P.O. for 30 uninterrupted days and administered subcutaneous epinephrine (2 mg/kg, s.c.) divided into two doses (1 mg/kg, s.c) each on the 31st and 32nd days. Electrocardiography (ECG), biochemical, oxidative stress, inflammatory mediators, histopathological, immunohistochemical analysis and the expression of phosphoinositide 3-kinases (PI3K) and protein kinase B (AKT) were assessed. ResultsMEC reversed epinephrine-induced lessening of heart rate, prolonged QT interval and ST displacement. Pretreatment with MEC significantly reduced serum aspartate aminotransferase (AST), lactate dehydrogenase (LDH) and creatine kinase-MB (CK-MB) levels in MI rats. The MEC pretreatment promoted total antioxidant capacity (TAC), glutathione (GSH), and total oxidant status (TOS). In addition, it reduced interleukin-1β (IL-1β), interleukin-6 (IL-6), malondialdehyde (MDA), and tumour necrosis factor (TNF-α) in MI rats. Furthermore, MEC statistically reduced immunohistochemical promotion of nuclear factor–κB (NF-κB) and expression of AKT and PI3K in cardiac tissue and perfected histopathological changes. ConclusionThe MEC, rich in phenolic and flavonoid contents, had a cardioprotective effect on rats suffering from epinephrine-induced MI. Such effect was achieved via improving cardiac function, antioxidant status, attenuating ECG pattern and histological picture, as well as reducing IL-6, IL-1β, TNF-α and NF-κB, which its antioxidant and anti-inflammatory potential may partly arbitrate.

Echinacoside ameliorates hepatic fibrosis and tumor invasion in rats with thioacetamide-induced hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) affects approximately 800,000 individuals globally each year. Despite advancements in HCC treatments, there is still a pressing need to identify new drugs that can combat resistance. One potential option is echinacoside, a natural caffeic acid glycoside with antioxidant, anti-inflammatory, antidepressant, and antidiabetic properties. Therefore, we aimed to investigate the ability of echinacoside to exhibit antitumor activity against HCC in rats through ameliorating hepatic fibrosis and tumor invasion. Rats were given thioacetamide to induce HCC, and some were given 30 mg/kg of echinacoside twice a week for 16 weeks. The liver impairment was assessed by measuring serum α-fetoprotein (AFP) and examining liver sections stained with Masson trichrome or anti-transforming growth factor (TGF)-β1 antibodies. The hepatic expression of mRNA and protein levels of TGF-β1, β-catenin, SMAD4, matrix metalloproteinase-9 (MMP9), phosphoinositide 3-kinases (PI3K), mammalian target of rapamycin (mTOR), connective tissue growth factor 2 (CCN2), E-Cadherin, platelets derived growth factor (PDGF)-B and fascin were also analyzed. Echinacoside improved the survival rate of rats by decreasing serum AFP and the number of hepatic nodules. Examination of micro-images indicated that echinacoside can reduce fibrosis. It also significantly decreased the expression of TGF-β1, β-catenin, SMAD4, MMP9, PI3K, mTOR, CCN2, PDGF-B, and fascin while enhancing the expression of E-Cadherin. In conclusion, echinacoside exhibits a protective effect against HCC by increasing survival rates and decreasing tumor growth. It also acts as an inhibitor of the hepatic tissue fibrosis pathway by reducing the expression of TGF-β1, β-catenin, SMAD4, PI3K, CCN2, PDGF-B and mTOR. Additionally, it prevents tumor invasion by suppressing MMP9 and fascin, and increasing the expression of E-Cadherin.

Cilostazol Stimulates Angiogenesis and Accelerates Fracture Healing in Aged Male and Female Mice by Increasing the Expression of PI3K and RUNX2.

Fracture healing in the aged is associated with a reduced healing capacity, which often results in delayed healing or non-union formation. Many factors may contribute to this deterioration of bone regeneration, including a reduced 'angiogenic trauma response'. The phosphodiesterase-3 (PDE-3) inhibitor cilostazol has been shown to exert pro-angiogenic and pro-osteogenic effects in preclinical studies. Therefore, we herein analyzed in a stable closed femoral fracture model whether this compound also promotes fracture healing in aged mice. Forty-two aged CD-1 mice (age: 16-18 months) were daily treated with 30 mg/kg body weight cilostazol (n = 21) or vehicle (control, n = 21) by oral gavage. At 2 and 5 weeks after fracture, the femora were analyzed by X-ray, biomechanics, micro-computed tomography (µCT), histology, immunohistochemistry, and Western blotting. These analyses revealed a significantly increased bending stiffness at 2 weeks (2.2 ± 0.4 vs. 4.3 ± 0.7 N/mm) and an enhanced bone formation at 5 weeks (4.4 ± 0.7 vs. 9.1 ± 0.7 mm3) in cilostazol-treated mice when compared to controls. This was associated with a higher number of newly formed CD31-positive microvessels (3.3 ± 0.9 vs. 5.5 ± 0.7 microvessels/HPF) as well as an elevated expression of phosphoinositide-3-kinase (PI3K) (3.6 ± 0.8 vs. 17.4 ± 5.5-pixel intensity × 104) and runt-related transcription factor (RUNX)2 (6.4 ± 1.2 vs. 18.2 ± 2.7-pixel intensity × 104) within the callus tissue. These findings indicate that cilostazol accelerates fracture healing in aged mice by stimulating angiogenesis and the expression of PI3K and RUNX2. Hence, cilostazol may represent a promising compound to promote bone regeneration in geriatric patients.

The Effect of Aerobic Training and Eugenol Supplementation on the Brain-Derived Neurotrophic Factor and Phosphoinositide 3-Kinase Expression in the Skeletal Muscle of Male Rats Poisoned With Chlorpyrifos

Background: Chlorpyrifos (CPF) is an insecticide and has a wide range of applications in the world. The purpose of this research was to investigate the effect of 4-week aerobic exercise and eugenol supplementation on the brain-derived neurotrophic factor/phosphoinositide 3-kinase (BDNF/PI3K) pathway on the skeletal muscle of male rats poisoned with CPF. Materials and Methods: Overall, 12-week-old female rats were used in this experimental research. The rats were randomly divided into 7 groups (8 rats in each group), including healthy control, toxic control, poison solvent, corn oil solvent, poisoned+eugenol, poisoned+aerobic exercise, and poisoned+aerobic exercise+eugenol. Moderate training in the range of 50%-60% VO2max, including 5 training sessions per week (treadmill). Poisoning was performed with CPF poison with a dose of 3 mg/kg. The dose of eugenol was determined to be 250 mg/kg. Finally, a one-way analysis of variance and Tukey’s post hoc test were employed to check the difference between groups. Results: The expression of PI3K and BDNF in the poisoned control group was lower than that in the healthy control group (P=0.049 and P=0.001, respectively). In addition, the expression of PI3K and BDNF was higher in the poisoned+eugenol+exercise group compared to the poisoned control group (P=0.009 and P=0.03, respectively). The corn solvent group also had a higher PI3K and BDNF expression in comparison to the poisoned control group (P=0.025 and P=0.01, respectively). Eventually, there was no significant difference among other groups. Conclusion: Exercise and eugenol increased PI3K and BDNF expression. It is thought that exercise and consumption of eugenol in poisoned rats reduces damage and improves muscle function and the BDNF/PI3K signaling pathway.

Effects of Echinacoside on Ehrlich Carcinoma in Rats by Targeting Proliferation, Hypoxia and Inflammation.

Background and objectives Ehrlich solid carcinoma (ESC) is a type of tumor originating from a spontaneous mammary adenocarcinoma in mice. It is highly aggressive and fast-growing and can create a solid undifferentiated mass when inserted under the skin. This makes it an ideal model for assessing cancer biology and tumor immunology. Echinacoside is a natural phenylethanoid glycoside with anti-inflammatory, anti-endoplasmic reticulum stress, anti-oxidative stress, and other beneficial properties. This study explored the potential anti-cancer benefits of echinacoside in rats with ESC. The study also analyzed its effects on tumor cell proliferation, differentiation, motility, and inflammation. Methods The study involved injecting rats with tumors in their left hind limb using an intramuscular injection of 2×106cells. After 14 days, some rats were given a daily intraperitoneal dose of 30 mg/kg echinacoside for three weeks. Muscle samples were then analyzed under an electron microscope. In addition, gene expression and protein levels of various factors such as phosphoinositide 3-kinases (PI3K), mammalian target of rapamycin (mTOR), hypoxia-inducible factor(HIF)-1α, cyclin D1, cyclin-dependent kinase 2 (CDK2), tumor necrosis factor (TNF)-α, and nuclear factor (NF)κB were evaluated in another part of the muscle samples. Results After being treated with echinacoside, theESC rats experienced a significant increase in their mean survival time from 27 days to 48 days. This treatment also resulted in a decrease in the volume and weight of the tumor. Upon examining the tumor tissue under an electron microscope, signs of damage such as pleomorphic cells, necrosis, nuclear fragmentation, membrane damage with cytoplasmic content spilling, and loss of cellular junction were observed. However, the treatment with echinacoside was effective in improving these effects. Furthermore, the expression of PI3K, mTOR, HIF-1α, cyclin D1, CDK2, TNF-α, and NFκB was significantly reduced due to the echinacoside treatment. Conclusions Our research found that echinacoside has antitumor properties that resulted in a substantial decrease in tumor size and weight, leading to an increase in the average survival time of rats and an improvement in muscle structure. Additionally, echinacoside was shown to ameliorate hypoxia by suppressing HIF-1α, reduce inflammation by decreasing NFκB and TNF-α, decrease proliferation by reducing PI3K, and block cyclin D1 and CDK2 to inhibit differentiation.

Cardioprotective effects of plant-based silver nanoparticles: Describing a modern drug

Regarding applicative, facile, green chemical research, a bio-inspired approach is being reported for the synthesis of Ag nanoparticles by Rumex alpinus L as a natural reducing and stabilizing agent without using any toxic and harmful reagent. Our study examined the ability of Ag NPs to make the cardioprotective effects. The biosynthesized Ag NPs@Rumex alpinus L were characterized by advanced physicochemical techniques like ultraviolet–visible (UV–Vis), Fourier Transformed Infrared spectroscopy (FT-IR), and Scanning Electron Microscopy (SEM) study. It has been established that Rumex alpinus L-stabilized silver nanoparticles have a spherical shape with a mean diameter from 12 to 55 nm. In the in medicinal part of this research, four groups of Wister rats were used: a control group; a group treated only with isoproterenol, to induce myocardial infarction; and two groups pretreated with silver nanoparticles containing Rumex alpinus (15 or 45 mg/kg, respectively) for 14 days and challenged with isoproterenol on the 13th and 14th days. Several parameters were measured including cardiac markers, electrocardiogram (ECG), nuclear factor erythroid 2-related factor 2 (Nrf2), the expression of Kelch-like ECH-associated protein 1 (Keap1), and other downstream antioxidant enzymes, as well as the expression of phosphoinositide 3-kinases (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) and other downstream apoptotic and inflammatory mediators. Silver nanoparticles containing Rumex alpinus may possess a protective effect against myocardial infarction through moderating myocardial infarction-induced myocardial oxidative stress (GSH, SOD, GPx, GST, and Keap1/Nrf2 pathway), inflammation (IL-1β, IL-6, TNF-α, and NF-κB), apoptosis (caspase-3, caspase-9, Bcl2, and Bax), and autophagy (PI3K/Akt/mTOR pathway). Silver nanoparticles containing Rumex alpinus treatment activated the Keap1/Nrf2/heme oxygenase-1 (HO-1) pathway, increased antioxidant enzyme activities, modulated the PI3K/Akt/mTOR pathway, and ameliorated myocardial autophagy, inflammation, and apoptosis. Silver nanoparticles containing Rumex alpinus treatment reduced the size of the infarct region, attenuated the levels of cardiac indicators, and diminished myocardial necrosis and immune cell infiltration.

Therapeutic effect of Hosta plantaginea (Lam.) Aschers flowers on acute pharyngitis through inhibition of multi-inflammatory pathways in rats

BackgroundHosta plantaginea (Lam.) Aschers flower is a famous Mongolian folk medicine in China and has a therapeutic effect on acute pharyngitis (AP). However, the effect and potential mechanism of H. plantaginea flower on AP have not been fully elucidated. Aim of the studyThe present work aimed to evaluate the effects and mechanisms of the crude extract of H. plantaginea flowers (HP) and its four fractions of petroleum ether fraction (HPA), ethyl acetate fraction (HPB), n-butanol fraction (HPC), and water residue (HPD) against AP in rats. Materials and methodsA 15% ammonia-induced AP rat model in rats was established. Therapeutic effects of HP and HPA∼D in model rats were evaluated based on body weight, histopathological analysis, and inflammatory parameters, including tumor necrosis factor α (TNF-α), prostaglandin E2 (PGE2), interleukin 1β (IL-1β), and IL-6. The protein expression of nuclear factor kappa-B p65 (NF-κB p65), inhibitor of NF-κB alpha (IκBα), c-Jun N-terminal kinases (JNK), mitogen-activated protein kinase (MAPK) p38, extracellular signal-regulated kinase (Erk), just another kinase 1 (JAK1), signal transducer and activator of transcription 3 (STAT3), phosphoinositide 3-kinase (PI3K), and protein kinase B (Akt) were detected by a Western blotting assay. ResultsHP, HPB, and HPC treatments markedly alleviated AP in rats by increasing body weight and improving pathological damages in pharyngeal tissues. In addition, HP, HPB, and HPC treatments significantly inhibited inflammation, including decreasing the levels of TNF-α, PGE2, IL-1β, and IL-6, and suppressing phosphorylated protein expression of p65, IκBα, JNK, p38, Erk, JAK1, STAT3, PI3K, and Akt in pharyngeal tissues of rats. ConclusionCollectively, HP, HPB, and HPC can attenuate pharynx injury in rats by suppressing inflammation via inhibition of NF-κB, MAPKs, JAK-STAT, and PI3K-Akt pathways, which supports the traditional use of H. plantaginea flowers.

Anti-fatigue effect of Lubian on kidney Yin deficiency and kidney Yang deficiency mice and mechanism based on PI3K-Akt pathway

This study aimed to investigate the anti-fatigue effect and mechanism of Lubian(Cervi Penis et Testis) on kidney Yin deficiency and kidney Yang deficiency mice. After one week of adaptive feeding, 88 healthy male Kunming mice were randomly divided into a blank group, a kidney Yin deficiency model group, a kidney Yin deficiency-Panacis Quinquefolii Radix(PQR) group, kidney Yin deficiency-Lubian treatment groups, a kidney Yang deficiency model group, a kidney Yang deficiency-Ginseng Radix et Rhizoma(GR) group, and kidney Yang deficiency-Lubian treatment groups, with eight mice in each group. The kidney Yin deficiency model and kidney Yang deficiency model were prepared by daily regular oral administration of dexamethasone acetate and hydrocortisone, respectively, and meanwhile, corresponding drugs were provided. The mice in the blank group received blank reagent. The treatment lasted 14 days. The exhaustive swimming time was measured 30 min after drug administration on the 14th day. On the 15th day, blood was collected from eyeballs and the serum was separated to determine the content of lactic acid(LD), blood urea nitrogen(BUN), lactate dehydrogenase(LDH), cyclic adenosine monophosphate(cAMP), and cyclic guanosine monophosphate(cGMP). The liver was dissected to determine the content of liver glycogen and the protein expression of phosphoinositide 3-kinase(PI3K) and protein kinase B(Akt). Compared with the kidney Yang deficiency model group, the kidney Yang deficiency-Lubian treatment groups showed increased body weight(P<0.05), relieved symptoms of Yang deficiency, decreased cGMP content(P<0.01), increased cAMP/cGMP(P<0.01), prolonged exhausted swimming time(P<0.01), reduced LD(P<0.01), elevated BUN content(P<0.01), increased liver glycogen content(P<0.01), and increased protein expression of PI3K and Akt in the liver(P<0.05). Compared with the kidney Yin deficiency model group, the kidney Yin deficiency-Lubian treatment groups showed increased body weight(P<0.01), relieved symptoms of Yin deficiency, increased content of cGMP(P<0.01), decreased cAMP/cGMP(P<0.01), prolonged exhausted swimming time(P<0.01), decreased LD(P<0.01), decreased BUN content(P<0.01), increased liver glycogen content(P<0.01), and increased protein expression of PI3K(P<0.05) and Akt in the liver(P<0.05). To sum up, Lubian can regulate Yin deficiency and Yang deficiency and increase glycogen synthesis by affecting the PI3K-Akt pathway, thereby exerting an anti-fatigue role.

Cilostazol novel neuroprotective mechanism against rotenone-induced Parkinson's disease in rats: Correlation between Nrf2 and HMGB1/TLR4/PI3K/Akt/mTOR signaling.

This study investigated the novel neuroprotective effect of cilostazol on rotenone-intoxicated rats focusing on the HMGB1/TLR4 axis, erythroid-related factor 2 (Nrf2)/hemeoxygenase-1 (HO-1), and phosphoinositide 3-kinase (PI3K)/Protein kinase B (Akt)/the mammalian target of rapamycin (mTOR) pathway. The aim is extended to correlate the Nrf2 expression with all assessed parameters as promising therapeutic targets for neuroprotection. Our experiment was designed as follows: vehicle group, cilostazol group, rotenone group (1.5mg/kg, s.c), and the rotenone pretreated with cilostazol (50mg/kg, p.o.) group. Eleven rotenone injections were injected day after day, while cilostazol was administered daily for 21days. Cilostazol significantly improved the neurobehavioral analysis, the histopathological examination, and dopamine levels. Moreover, the immunoreactivity of tyrosine hydroxylase (TH) in substantia nigra pars compacta (SNpc) enhanced. These effects were associated with enhancement of the antioxidant expression of Nrf2 and HO-1 by 1.01 and 1.08-fold, respectively, and repression of HMGB1/TLR4 pathway by 50.2% and 39.3%, respectively. Upregulation of the neuro-survival PI3K and Akt expression by 2.26 and 2.69-fold, respectively, and readjusting mTOR overexpression. Cilostazol exerts a novel neuroprotective strategy against rotenone-induced neurodegeneration via activation of Nrf2/HO-1, suppression of HMGB1/TLR4 axis, upregulation of PI3K/Akt besides mTOR inhibition that compels more investigations with different PD models to clarify its precise role.

Ethyl Acetate Extract of Halymenia durvillei Induced Apoptosis, Autophagy, and Cell Cycle Arrest in Colorectal Cancer Cells.

Colorectal cancer is one of the most death-dealing cancers. However, conventional cancer treatments still have side effects. Therefore, novel chemotherapeutic agents with less side effects are still in search. A marine red seaweed, Halymenia durvillei, is recently interested in its anticancer effects. This study investigated the anticancer effect of ethyl acetate extract of H. durvillei (HDEA) on HT-29 colorectal cancer cells in association with the phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signaling pathway. HDEA-treated HT-29 and OUMS-36 cells were used for cell viability tests by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyltetrazolium bromide assay. The effects of HDEA on apoptosis and cell cycle were evaluated. The nuclear morphology and mitochondrial membrane potential (ΔΨm) were observed by Hoechst 33342 and JC-1 staining, respectively. The gene expression of PI3K, AKT, and mTOR genes was evaluated using a real-time semiquantitative reverse transcription-polymerase chain reaction. The corresponding protein expressions were assessed by western blot analysis. The result revealed that the cell viability of treated HT-29 cells diminished while that of OUMS-36 cells was non-significant. By the down-regulation of cyclin-dependent ki-nase 4 and cyclin D1, HDEA-treated HT-29 cells were arrested in the G0/G1 phase. By the up-regulation of cleaved poly(adenosine diphosphate-ribose) polymerase, caspase-9, caspase-8, caspase-3, and Bax, HDEA-treated HT-29 cells underwent apoptosis, but suppressed Bcl-2, disrupted nuclear morphology and ΔΨm. Furthermore, treated HT-29 cells underwent autophagy by up-regulation of light chain 3-II and beclin-1. Lastly, HDEA suppressed the expression of PI3K, AKT, and mTOR. Therefore, HDEA exerts anticancer effects against HT-29 cells, confirmed by apoptosis, autophagy, and cell cycle arrest induction via regulation of the PI3K/AKT/mTOR signaling pathway.

The Attenuation of Insulin/IGF-1 Signaling Pathway Plays a Crucial Role in the Myo-Inositol-Alleviated Aging in Caenorhabditis elegans.

Myo-Inositol (MI) has been shown to alleviate aging in Caenorhabditis (C). elegans. However, the mechanism by which MI alleviates aging remains unclear. In this study, we investigate whether MI can modulate the PI3K so as to attenuate the insulin/IGF-1 signaling (IIS) pathway and exert the longevity effect. The wild-type C. elegans and two mutants of AKT-1 and DAF-16 were used to explore the mechanism of MI so as to extend the lifespan, as well as to improve the health indexes of pharyngeal pumping and body bend, and an aging marker of autofluorescence in the C. elegans. We confirmed that MI could significantly extend the lifespan of C. elegans. MI also ameliorated the pharyngeal pumping and body bend and decreased autofluorescence. We further adopted the approach to reveal the loss-of-function mutants to find the signaling mechanism of MI. The functions of the lifespan-extending, health-improving, and autofluorescence-decreasing effects of MI disappeared in the AKT-1 and DAF-16 mutants. MI could also induce the nuclear localization of the DAF-16. Importantly, we found that MI could dramatically inhibit the phosphoinositide 3-kinase (PI3K) activity in a dose-dependent manner with an IC50 of 90.2 μM for the p110α isoform of the PI3K and 21.7 μM for the p110β. In addition, the downregulation of the PI3K expression and the inhibition of the AKT phosphorylation by MI was also obtained. All these results demonstrate that MI can inhibit the PI3K activity and downregulate the PI3K expression, and the attenuation of the IIS pathway plays a crucial role for MI in alleviating aging in C. elegans.

Vitamin D ameliorates insulin resistance-induced osteopenia by inactivating the nucleotide-binding oligomerization domain-like receptor protein 3 inflammasome

ObjectiveOsteoporosis (OP) can be considered a chronic complication of type 2 diabetes mellitus (T2DM). Aberrant activation of the nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3) inflammasome is associated with the pathogenesis of various inflammation-related diseases, e.g., T2DM and OP. Vitamin D affects the inflammatory pathway and inhibits an excessive inflammatory response. The current study investigated the inter-relationship between vitamin D and inflammasome activation in T2DM. MethodHepatocellular carcinoma (HepG2) cells and bone marrow stromal cells (BMSCs) were treated with Conditioned Medium of bone marrow mesenchymal stem cells after VitD treatment (CM-VitD), as well as phosphoinositide 3-kinase (PI3K) specific agonist, 740Y-P, or the PI3K specific inhibitor, LY294002, respectively, or both. 40 Eight-week-old female Sprague Dawley rats were selected and established as a DM model. The rats were injected with CM-VitD, as well as the 740Y-P specific agonist, or the LY294002 inhibitor, respectively, or both. A quantitative reverse transcription polymerase chain reaction and western blotting were conducted to evaluate the expression of messenger ribonucleic acid and protein in the RUX2 gene, alkaline phosphatase (ALP), OsteoPontiN (OPN), peroxisome proliferator-activated receptor gamma (PPARγ), fatty acid-binding protein 4 (FABP4), protein kinase B (AKT), PI3K, NLRP3, apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), caspase-1, interleukin (IL)-1 beta (β), IL-18, and tumor necrosis factor alpha (TNF-α) in the BMSCs and liver tissue of rats. Enzyme-linked immunosorbent assay was used to detect the concentration of inflammatory factors in the cell supernatant and serum of rats. ResultsAn isolated co-culture of HepG2/insulin-resistance cells and BMSCs promoted the adipogenic transformation of the latter and inhibited the transformation of BMSCs into osteogenesis. The PI3K specific agonist, 740Y-P, significantly increased the expression of PI3K, AKT, NLRP3, ASC and Caspase-1 while the PI3K specific inhibitor, LY294002, does the opposite. Additionally, CM-VitD reduced the expression of NLRP3, ASC, caspase-1, IL-1β, and IL-18 in BMSCs and rat liver via the PI3K/AKT pathway. ConclusionVitamin D can inhibit the inflammatory response induced by T2DM and promote the osteogenesis of BMSCs, which may play a key role in the treatment of type 2 diabetes patients with OP.

Kunxian Capsule Extract Inhibits Angiogenesis in Zebrafish Embryos via PI3K/AKT-MAPK-VEGF Pathway.

To investigate the anti-angiogenic activity of Kunxian Capsule (KX) extract and explore the underlying molecular mechanism using zebrafish. The KX extract was prepared with 5.0 g in 100 mL of 40% methanol followed by ultrasonication and freeze drying. Freeze dried KX extract of 10.00 mg was used as test stock solution. Triptolide and icariin, the key bioactive compounds of KX were analyzed using ultra-high performance liquid chromatography. The transgenic zebrafish Tg(flk1:GFP) embryos were dechorionated at 20-h post fertilization (hpf) and treated with PTK 787, and 3.5, 7, 14 and 21 µg/mL of KX extract, respectively. After 24-h post exposure (hpe), mortality and malformation (%), intersegmental vessels (ISV) formation, and mRNA expression level of angiogenic pathway genes including phosphoinositide 3-kinase (PI3K), protein kinase B (AKT), extracellular signal-regulated kinases (ERKs), mitogen-activated protein kinase (MAPK), vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF-2) were determined. Further, the embryos at 72 hpf were treated with KX extract to observe the development of sub-intestinal vein (SIV) after 24 hpe. The chromatographic analysis of test stock solution of KX extract showed that triptolide and icariin was found as 0.089 mg/g and 48.74 mg/g, respectively, which met the requirements of the national drug standards. In zebrafish larvae experiment, KX extract significantly inhibited the ISV (P<0.01) and SIV formation (P<0.05). Besides, the mRNA expression analysis showed that KX extract could significantly suppress the expressions of PI3K and AKT, thereby inhibiting the mRNA levels of ERKs and MAPK. Moreover, the downstream signaling cascade affected the expression of VEGF and its receptors (VEGFR and VEGFR-2). FGF-2, a strong angiogenic factor, was also down-regulated by KX treatment in zebrafish larvae. KX extract exhibited anti-angiogenic effects in zebrafish embryos by regulating PI3K/AKT-MAPK-VEGF pathway and showed promising potential for RA treatment.

Glycoprotein M6A Suppresses Lung Adenocarcinoma Progression via Inhibition of the PI3K/AKT Pathway.

Lung adenocarcinoma is the most common subtype of lung cancer and has high morbidity and mortality. Glycoprotein M6A (GPM6A) is a neuronal membrane glycoprotein reported to be related with cancer. However, studies on GPM6A in lung adenocarcinoma are rare. This study aimed to investigate the role of GPM6A in lung adenocarcinoma and its potential mechanism. GPM6A mRNA expression was analysed in 33 types of cancers using The Cancer Genome Atlas (TCGA) datasets. It was compared among normal lung tissues, lung adenocarcinoma tissues, and adjacent tissues using the Oncomine database. Real-time quantitative polymerase chain reaction (RT-qPCR) was performed to detect GPM6A expression in human lung adenocarcinoma cell lines (A549 and H1299) and normal pulmonary epithelial cells (BEAS-2B). When GPM6A was inhibited, cell proliferative capacity was detected by Cell Counting Kit 8 (CCK8), EdU, and colony formation assays. Cell migration ability was detected by wound healing and transwell assays. The expression of epithelial-mesenchymal transition (EMT) markers was detected by Western blotting (WB) and RT-qPCR. When GPM6A was overexpressed, cell proliferation and migration were detected again. Ten nude mice were subcutaneously injected with cells overexpressing GPM6A or empty vector, and the tumor size was recorded on day 14 and then measured every 3 days thereafter. The final tumor weight was measured on day 36. Furthermore, the expressions of phosphoinositide 3-kinase (PI3K), phosphorylated PI3K, AKT, and phosphorylated AKT were detected by WB. Results showed that GPM6A mRNA expression decreased in 15 types of tumors in TCGA dataset. GPM6A expression was lower in lung adenocarcinoma than in normal lung tissues or adjacent tissues in the Oncomine dataset. Similar results were found in lung adenocarcinoma cells. The function study showed that GPM6A downregulation enhanced the proliferation, migration, and EMT of lung adenocarcinoma cells, while GPM6A upregulation inhibited their development. The xenograft results suggested that GPM6A upregulation delayed tumor growth and reduced tumor weight. Moreover, WB showed that GPM6A knockdown activated the PI3K/AKT pathway, while GPM6A upregulation inhibited the activation of the PI3K/AKT pathway. In conclusion, GPM6A suppresses lung adenocarcinoma progression via inhibition of the PI3K/AKT pathway. Thus, GPM6A could be a possible treatment target for lung cancer therapy.

Advanced glycation end products (AGEs) downregulate the miR-4429/PTEN axis to promote apoptosis of fibroblasts in pelvic organ prolapse.

BackgroundPelvic organ prolapse (POP) is a common degenerative disease among females. We previously reported that advanced glycation end products (AGEs), compounds derived from nonenzymatic glycoxidation reactions, accumulated in the human vaginal wall and impaired the function of fibroblasts in the pathogenesis of POP. This study investigated the apoptosis induced by AGEs in human uterosacral ligament fibroblasts and the underlying mechanism.MethodsHuman uterosacral ligament fibroblasts were cultured and identified. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis was performed to identify the expression of miR-4429, phosphatase and tensin homolog (PTEN), and caspase-3. Flow cytometric analysis was applied to detect the apoptosis rate of fibroblasts. Dual-luciferase reporter assay was performed to verify the relationship between miR-4429 and PTEN. The overexpression of miR-4429 and the inhibition of PTEN were achieved by cell transfections. Western blot analysis was used to detect the protein levels of PTEN, phosphoinositide 3-kinase (PI3K), and protein kinase B (Akt).ResultsThe AGEs promoted fibroblast apoptosis both in the POP and the non-POP groups. The expression of PTEN increased in fibroblasts from the POP group or fibroblasts treated with AGEs. It was confirmed that miR-4429 interacted with PTEN messenger RNA (mRNA), and the expression level of miR-4429 was reduced in fibroblasts from the POP group or fibroblasts treated with AGEs. Further, overexpression of miR-4429 alleviated increased PTEN expression and fibroblast apoptosis induced by AGEs. Similarly, inhibition of PTEN expression alleviated increased fibroblast apoptosis induced by AGEs. In addition, the protein expressions of PI3K and phosphorylated Akt were reduced in fibroblasts exposed to AGEs.ConclusionsWe proposed that AGEs induced fibroblast apoptosis by regulating the miR-4429/PTEN/PI3K/Akt pathway in POP. Our results revealed a novel mechanism by which AGEs contributed to the molecular pathological alteration in POP.

Discovery of Novel Phosphoinositide-3-Kinase α Inhibitors with High Selectivity, Excellent Bioavailability, and Long-Acting Efficacy for Gastric Cancer.

Phosphoinositide-3-kinase (PI3K) overexpressed in many tumors is a promising target for cancer therapy. However, due to toxicity from the ubiquitous expression of PI3K in many tissues, the development of PI3K inhibitors with high selectivity and low toxicity has become an urgent need for tumor treatment. Herein, based on the HipHop, we designed and synthesized a series of 6-(4,6-dimorpholino-1,3,5-triazin-2-yl)benzo[d]oxazol-2-amine derivatives as potent, selective, and long-acting PI3Kα inhibitors. Compound 27 was determined with potent PI3Kα inhibitory activity (IC50 = 4.4 nM), which exhibited excellent selectivity for homologous PI3K enzymes and a 370 kinome panel. Meanwhile, 27 featured favorable stability (T1/2 > 10 h) and high bioavailability (130%). Importantly, compound 27 exerted great antigastric cancer activity in vivo when combined with taxol. Collectively, these characteristics suggested 27 to be a promising PI3K agent for cancer treatment.