Phosphatidylinositol 3-hydroxy Research Articles

Metformin reduces pancreatic cancer cell proliferation and increases apoptosis through MTOR signaling pathway and its dose-effect relationship.

To study the influences of metformin on the proliferation and apoptosis of pancreatic cancer cells and its dose-effect relationship and crucial molecular mechanism. With human pancreatic cancer cell line PANC-1 as the study object, different concentrations of metformin were added for intervention. Then, the proliferation of PANC-1 cells was detected via methyl thiazolyl tetrazolium (MTT) assay to determine the dose-effect relationship of metformin in PANC-1 cell proliferation. PANC-1 cells were treated with metformin at three appropriate concentrations as Metformin treatment groups, and an equal amount of dimethyl sulfoxide (DMSO) was added in Control group. Flow cytometry was performed to detect PANC-1 cell cycle and apoptosis, and the apoptosis of PANC-1 cells was also evaluated via terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay. Caspase-3 protein localization and expression in PANC-1 cells were detected using immunofluorescence assay. Besides, the expressions of the apoptosis-associated proteins Caspase-3, B-cell lymphoma 2 (Bcl-2), and Bcl-2 associated X protein (Bax) and phosphatidylinositol 3-hydroxy kinase (PI3K), phosphorylated protein kinase B (p-Akt), and p-mammalian target of rapamycin (mTOR) proteins related to the mTOR pathway were detected using Western blotting. Metformin repressed the proliferation of human pancreatic cancer PANC-1 cells in a concentration-dependent and time-dependent manner. Compared with Control group, Metformin treatment groups (0, 20 and 40 mM) exhibited a higher proportion of PANC-1 cells in G0/G1 phases, and a lower proportion of PANC-1 cells in S phase (p<0.05), and the change in the proportion of cells in G2/M phase was not statistically significant (p>0.05). Moreover, Metformin treatment groups (0, 20, and 40 mM) had more apoptotic PANC-1 cells, higher expression levels of pro-apoptosis proteins Caspase-3 and Bax and lower expression levels of anti-apoptosis protein Bcl-2 and the mTOR pathway-related proteins PI3K, p-Akt, and p-mTOR in cells than Control group (p<0.05). Metformin modulates the mTOR signaling pathway to reduce the proliferation of pancreatic cancer cell, but increase their apoptosis.

Rapid membrane effect of estrogens on stimulation of corticotropin-releasing hormone

BackgroundClassic nuclear-initiated estrogen signaling stimulates corticotropin-releasing hormone (CRH) gene expression as a transcription factor. However, the possible mechanism by which membrane-initiated estrogen signaling (MIES) influences CRH expression remains unclear. There are indications that MIES may upregulate nitric oxide (NO) production through the phosphatidylinositol 3-hydroxy kinase (PI3K) and potentially through the mitogen-activated protein kinase (MAPK) pathway. ObjectivesWe investigated the effect of MIES-mediated kinase pathways on CRH expression with or without NO synthesis. MethodIn SK-N-SH cell culture, estradiol-bovine serum albumin (E2-BSA) was used as the specific membrane estrogen receptor activator, with a specific NO donor, and/or inhibitors for NO synthase (NOS), PI3K, MAPK, protein kinase A (PKA), and protein kinase C (PKC). ResultsE2-BSA significantly increased NO and CRH levels in the medium and NOS1-mRNA levels in the cells. In addition, NO donor up-regulated CRH expression, while NOS-inhibitor down-regulated it. When the inhibitor of MAPK and/or the inhibitor of PI3K was added to the medium, only the latter appeared to significantly block the stimulating effect of E2-BSA on NO synthesis, and this was accompanied by an increased CRH expression in the medium. We further studied the effect of the MIES-PKC-mediated pathway on CRH expression, with or without NOS-inhibitor, while the MIES-PKA(-PI3K) pathway served as a control. We found that MIES-PKC upregulated CRH expression independent of NO synthesis. ConclusionMIES can efficiently upregulate CRH expression via various intracellular kinase pathways and may thus be a crucial component in the stress response.

FBXW7 inhibited cell proliferation and invasion regulated by miR-27a through PI3K/AKT signaling pathway and epithelial-to-mesenchymal transition in oral squamous cell carcinoma.

Our purpose was to detect the molecular mechanism of F-box and WD repeat domain containing 7 (FBXW7) in regulating cell growth and metastasis of oral squamous cell carcinoma (OSCC). Real Time-quantitative Polymerase Chain Reaction (RT-qPCR) and Western blot were applied to calculate the messenger ribonucleic acid (mRNA) and protein levels of genes and miR-27a. The proliferation and invasive abilities were measured by methyl thiazolyl tetrazolium (MTT) and transwell assays. The. Kaplan-Meier method was conducted to evaluate the 5-year overall survival of oral squamous cell carcinoma patients. FBXW7 was downregulated while miR-27a was upregulated in OSCC tissues and cells compared with the corresponding adjacent tissues. Downregulation of FBXW7 or upregulation of miR-27a in OSCC tissues predicted poor outcome of OSCC patients. FBXW7 suppressed the growth through the phosphatidylinositol 3-hydroxy kinase/protein kinase B (PI3K/AKT) signaling pathway in OSCC cell line HSC3. FBXW7 inhibited the invasion-mediated epithelial-mesenchymal transition (EMT) in HSC3 cells. The expression of FBXW7 was mediated by miR-27a by directly binding to the 3'-untranslated region (3'-UTR) of FBXW7 in HSC3 cells. MiR-27a reversed partial roles of FBXW7 on the proliferation and invasion in OSCC cells. FBXW7 was mediated by miR-27a and could inhibit the proliferation through the PI3K/AKT pathway and invasion-mediated EMT in OSCC cell line. The newly identified miR-27a/FBXW7/PI3K/AKT axis provides novel insights into the pathogenesis of osCC.

Effect of miR-26b on gestational diabetes mellitus in rats via PI3K/Akt signaling pathway.

The aim of this study was to explore the influence of micro ribonucleic acid (miR)-26b on gestational diabetes mellitus in rats via the phosphatidylinositol 3-hydroxy kinase/protein kinase B (PI3K/Akt) signaling pathway. A total of 60 healthy pregnant female rats were randomly divided into three groups, including group A (normal group), group B (model group), and group C (model + miR-26b group). The differences in fasting blood glucose (FBG), C-reactive protein (CRP), and phosphatidylinositol 3-hydroxy kinase/protein kinase B (PI3K/AKT) among the three groups were analyzed via serum CRP test, morphological observation, quantitative Reverse Transcription-Polymerase Chain Reaction (qRT-PCR), and Western blotting, respectively. The levels of FBG ad CRP were significantly up-regulated in group B when compared with group A (p<0.01). Meanwhile, they increased significantly in group C when compared with group B (p<0.01). Rats in group A exhibited smooth and flat thoracic aortic intimas, as well as neatly arranged smooth muscle cells at the media layer. However, rats in group B showed fractured intimas with enlarged junction gaps, as well as necrotic and detached endothelial cells. Compared with group B, group C exhibited extremely poorly arranged cells at all the layers, rough and rugged intimas, larger areas of necrotic and detached endothelial cells, and markedly worsened lesions. QRT-PCR results indicated that the expression of phosphorylated-PI3K (p-PI3K) was significantly lower in group B than that of group A (p=0.04). Meanwhile, it was markedly lower in group C than that in group B (p=0.04). The expression of p-Akt was remarkably lower in group B than group A (p=0.04), which was also significantly lower in group C than group B (p=0.04). Compared with group A, the expressions of p-PI3K and p-Akt in the thoracic aorta of group B were evidently down-regulated (p<0.01). Furthermore, they decreased markedly in group C when compared with group B (p<0.01). MiR-26b accelerates the progression of gestational diabetes by inhibiting the PI3K/Akt signaling pathway.

MiR-19 enhances pancreatic cancer progression by targeting PTEN through PI3K/AKT signaling pathway.

Abnormal expression of micro ribonucleic acids (miRNAs) has become an important marker of cancer. However, the exact molecular mechanisms of miRNAs were not very clear. Here, we decided to investigate the miR-19 effect and molecular mechanism on pancreatic cancer, which was blank until now. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis was applied for testing miR-19 and gene of phosphate and tensin homolog deleted on chromosome ten (PTEN) expression. Western blot was used for detecting the protein expression. Methyl thiazolyl tetrazolium (MTT) assay and transwell assay were carried out to measure cell proliferation, invasion, and migration. We showed that miR-19 expression was increased in cancerous tissues and was associated with the survival of patients, tumor node metastasis (TNM) stage, tumor size, and lymph node metastasis. MiR-19 mimic enhanced cell proliferation, invasion, and migration, while suppressing miR-19 cell progression was suppressed. With the help of TargetScanHuman and luciferase reporter assay, we verified PTEN as a specific target of miR-19. Moreover, PTEN expression was reduced by miR-19 mimic and was increased by miR-19 inhibitor. We next found that PTEN was elevated in cancerous tissues and its expression was negatively correlated with miR-19 expression. Furthermore, miR-19 regulated cell progression via activating phosphatidylinositol 3-hydroxy kinase/protein kinase B (PI3K/AKT) signaling pathway. We demonstrated that miR-19 facilitated cell progression through modulating PI3K/AKT signaling pathway by targeting PTEN, which provided a potential therapeutic target for pancreatic cancer patients.

Rapamycin inhibits proliferation and apoptosis of retinoblastoma cells through PI3K/AKT signaling pathway.

Effects of Rapamycin on the proliferation and apoptosis of retinoblastoma cells through the phosphatidylinositol 3-hydroxy kinase (PI3K)/protein kinase B (AKT) signaling pathway were studied. The retinoblastoma Y79 cells were selected and divided into negative control group (NC group), 0.2 µM Rapamycin group and 0.4 µM Rapamycin group. Then the proliferative activity of Y79 cells was detected using Cell Counting Kit-8 (CCK8) assay, the content of reactive oxygen species (ROS), malondialdehyde (MDA) and superoxide dismutase (SOD) in cells in each group was detected using enzyme-linked immunosorbent assay (ELISA), and the apoptosis of Y79 cells was detected via terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay. Moreover, the changes in Y79 cell cycle and apoptosis were determined through flow cytometry, and apoptosis and PI3K/AKT pathway were detected using reverse transcription-polymerase chain reaction (RT-PCR) and western blotting. It was found that the number of cells and the proliferative activity were significantly reduced in 0.2 µM Rapamycin group and 0.4 µM Rapamycin group. In 0.2 µM Rapamycin group and 0.4 µM Rapamycin group, the content of ROS and MDA was significantly decreased, while that of SOD was notably increased. TUNEL assay and flow cytometry showed that in 0.2 µM Rapamycin group and 0.4 µM Rapamycin group, the number of apoptotic cells was obviously increased, and the cell cycle was basically arrested in S phase. The expression levels of Bcl-2, PI3K and AKT declined in 0.2 µM Rapamycin group and 0.4 µM Rapamycin group, whereas the expression of Caspase 8 increased. Similar results were also obtained in the protein assay. The above results were significantly superior in 0.4 µM Rapamycin group to those in 0.2 µM Rapamycin group. Rapamycin inhibits proliferation and promotes apoptosis of retinoblastoma cells through inhibiting the PI3K/AKT signaling pathway.

Hyperhomocysteinemia inhibits tibial fracture healing in rats through PI3K/AKT signaling pathway.

To explore the influence of hyperhomocysteinemia (hHcys) on the tibial fracture healing in rats and its effect on the phosphatidylinositol 3-hydroxy kinase (PI3K)/protein kinase B (AKT) signaling pathway. A total of 36 Sprague-Dawley rats were randomly divided into sham group (n=12), tibial fracture group (n=12) and hHcys + fracture group (n=12). The rats in tibial fracture group underwent the tibial fracture surgery, while the model of tibial fracture and hHcys was established in hHcys + fracture group. The level of plasma homocysteine (Hcy) in each group was analyzed using the full-automatic biochemical analyzer, the fracture stress biomechanical measurement was performed, and the ultimate bending strength and torque were calculated. Moreover, the protein expressions of PI3K and phosphorylated (p)-AKT in tibial tissues were detected using western blotting, the messenger ribonucleic acid (mRNA) levels of Bcl-2 associated X protein (Bax) and caspase-3 were detected using quantitative polymerase chain reaction (qPCR), the apoptosis was detected via terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining, and the expressions of inflammatory factors were detected via immunohistochemistry. Compared with sham group, tibial fracture group and hHcys + fracture group had a significantly increased level of plasma Hcy, significantly decreased ultimate bending strength and torque, obviously decreased relative protein expressions of PI3K and p-AKT, increased mRNA levels of Bax and caspase-3 and an increased expression of pro-inflammatory factor tumor necrosis factor-α (TNF-α). Compared with tibial fracture group, hHcys + fracture group had a higher level of plasma Hcy, lower ultimate bending strength and torque, lower relative protein expressions of PI3K and p-AKT, higher mRNA levels of Bax and caspase-3, a higher apoptosis rate and a higher expression of TNF-α. hHcys blocks the downstream apoptotic signal transduction, promotes apoptosis and inflammatory response, and affects fracture healing through affecting the PI3K/AKT signaling pathway.

MiR-130a alleviates neuronal apoptosis and changes in expression of Bcl-2/Bax and caspase-3 in cerebral infarction rats through PTEN/PI3K/Akt signaling pathway.

Effect of micro ribonucleic acid (miR)-130a on neuronal apoptosis in rats with cerebral infarction (CI) was studied to explore whether phosphatase and tensin homolog deleted on chromosome ten (PTEN)/phosphatidylinositol 3-hydroxy kinase (PI3K)/protein kinase B (Akt) is involved in the regulation of neuronal apoptosis. Thirty-six Sprague-Dawley (SD) rats were randomly divided into blank control group, model group and miR-130a low-expression group. miR-130a was determined by quantitative polymerase chain reaction (qPCR), the content of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and IL-10 was detected using the enzyme-linked immunosorbent assay (ELISA) kits, and the neuronal apoptosis level in each group was determined through terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining. The neurobehavioral score was significantly lower in model group than that in blank control group (P<0.01), while it was significantly higher in miR-130a low-expression group than that in model group (P<0.01). Compared with blank control group, the model group had obviously increased content of TNF-α and IL-6 (P<0.01), decreased content of IL-10 (P<0.01), more apoptotic neurons (P<0.01), higher expression of caspase-3 (P<0.01), and obviously lower Bcl-2/Bax (P<0.01). Moreover, expression of phosphorylated (p)-PTEN, PI3K and p-Akt in brain tissues was remarkably lower in the model group than those in the blank control group (P<0.01). The expression level of miR-130a in brain tissues of CI rats is significantly increased. miR-130a promotes the release of inflammatory factors and facilitates neuronal apoptosis through suppressing the PTEN/PI3K/Akt signaling pathway.

Effect of acupuncture on neurovascular units after cerebral infarction in rats through PI3K/AKT signaling pathway.

To study the effect of acupuncture on neurovascular units after cerebral infarction (CI) in rats through the phosphatidylinositol 3-hydroxy kinase/protein kinase B (PI3K/AKT) signaling pathway. A total of 36 Sprague-Dawley rats were randomly divided into sham group (n = 12), model group (n = 12) and acupuncture group (n = 12). The external carotid artery was only exposed in model group, while the post-CI ischemia-reperfusion model was established using the suture method in the other 2 groups. After modeling, the rats in sham group and model group were fixed and sampled, while those in acupuncture group were treated with acupuncture intervention for 2 weeks and sampled. The neurological deficits of rats were evaluated using the Zea-Longa score, and the spatial learning and memory of rats were detected via water maze test. Moreover, the expressions of vascular endothelial growth factor (VEGF), growth associated protein-43 (GAP-43) and synuclein (SYN) in brain tissues were detected via immunohistochemistry, and the relative protein expressions of PI3K p85, PI3K p110 and p-AKT were detected via Western blotting. The messenger ribonucleic acid (mRNA) expressions of VEGF, GAP-43 and SYN were detected via quantitative polymerase chain reaction (qPCR). The Zea-Longa score was significantly increased in model group and acupuncture group compared with that in sham group (p < 0.05), while it significantly declined in acupuncture group compared with that in model group (p < 0.05). The escape latency was significantly prolonged and the times of crossing platform were significantly reduced in model group and acupuncture group compared with those in sham group (p < 0.05), while the escape latency was significantly shortened and the times of crossing platform were significantly increased in acupuncture group compared with those in model group (p < 0.05). The positive expressions of VEGF, GAP-43 and SYN were obviously increased in model group and acupuncture group compared with those in sham group (p < 0.05), while they were obviously increased in acupuncture group compared with those in model group (p < 0.05). Besides, model group and acupuncture group had significantly higher relative protein expressions of PI3K p85, PI3K p110 and p-AKT than sham group (p < 0.05), while acupuncture group also had significantly higher relative protein expressions of PI3K p85, PI3K p110 and p-AKT than model group (p < 0.05). The relative mRNA expressions of VEGF, GAP-43 and SYN were remarkably increased in model group and acupuncture group compared with those in sham group (p < 0.05), while they were remarkably increased in acupuncture group compared with those in model group (p < 0.05). Acupuncture promotes the repair of neurovascular units after CI in rats through activating the PI3K/AKT signaling pathway, thereby exerting a protective effect on neurovascular units.

Anti-tumor mechanism of eicosapentaenoic acid (EPA) on ovarian tumor model by improving the immunomodulatory activity in F344 rats

Eicosapentaenoic acid (EPA) has been known to induce human ovarian cancer cells apoptosis in vitro. However, the anti-tumor mechanisms of EPA in vivo have been rarely reported. In this study, the effect of EPA on the ovarian cancer rat model and its related mechanism were investigated. The results showed after EPA treatment, the index of spleens and thymus was significantly increased, and the proliferation of spleen cells, the natural killer (NK) cell activity and phagocytosis activity of macrophages were prompted. Besides, EPA could reverse the decrease of CD4+ and CD8+ T lymphocytes induced by ovarian cancer. It was found that EPA could inhibit the phosphorylation status of PI3K (phosphatidylinositol 3-hydroxy kinase)/Akt (serine-threonine kinase), ERK1/2 (extracellular signal-regulated kinase 1/2) and NF-κB p65, and prompt the expression of cytochrome C and caspase-3. These results suggested that EPA has a remarkable anti-tumor activity by improving the immunomodulatory on the ovarian cancer rat model.

Effect of atorvastatin on pulmonary arterial hypertension in rats through PI3K/AKT signaling pathway.

The aim of this study was to investigate the effect of atorvastatin on pulmonary arterial hypertension (PAH) in rats and to observe its specific regulatory mechanism through the phosphatidylinositol 3-hydroxy kinase/protein kinase B (PI3K/AKT) signaling pathway. The model of PAH was successfully established in rats via hypoxia feeding. All rats were divided into three groups, including Control group (n=15), PAH model group (Model group, n=15) and atorvastatin treatment group (Ator group, n=15). Tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and nitric oxide (NO) were detected via enzyme-linked immunosorbent assay (ELISA). Right ventricular systolic pressure (RVSP) and right ventricular hypertrophy index (RVHI) in each group were determined as well. Meanwhile, the pathological changes in lung tissues of rats were detected via hematoxylin-eosin (HE) staining. Furthermore, the apoptosis level of lung tissues in each group was detected via terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) staining. In addition, the expression levels of PI3K/AKT signaling pathway and apoptotic genes in lung tissues were detected via quantitative Polymerase Chain Reaction (qPCR). In Model group, the levels of TNF-α and IL-6 increased significantly, while the level of NO decreased. Both RVSP and RVHI in Model group were significantly higher than those of Control group and Ator group (p<0.05). The results of HE staining revealed that Model group showed significantly severe lung tissue injury (p<0.05). According to the results of TUNEL staining, the number of apoptotic cells in lung tissues in Model group was significantly smaller than that of Ator group (p<0.05). Meanwhile, the expression level of cysteinyl aspartate-specific proteinase-3 (Caspase-3) in Model group was markedly lower than that of Ator group (p<0.05). However, the expression level of B-cell lymphoma-2 (Bcl-2) in Model group was markedly higher than that of Ator group (p<0.05). In Ator group, the expression levels of PI3K and AKT in lung tissues were remarkably higher than those of Model group (p<0.05). All the above results indicated that atorvastatin could effectively up-regulate the expressions of PI3K and AKT (p<0.05). Atorvastatin regulates the symptoms of PAH in rats through activating the PI3K/AKT signaling pathway.

LncRNA MALAT1 knockdown alleviates myocardial apoptosis in rats with myocardial ischemia-reperfusion through activating PI3K/AKT signaling pathway.

To observe the effect of long non-coding ribonucleic acid metastasis-associated lung adenocarcinoma transcript 1 (lncRNA MALAT1) on the myocardial ischemia-reperfusion (I/R) injury in rats and to explore its potential mechanism, to provide certain references for clinical prevention and treatment of myocardial I/R injury. A total of 60 male Wistar rats were randomly divided into the Control group (n=20), I/R group (n=20) and I/R + MALAT1 small-interfering RNA (siRNA) group (n=20) using a random number table. The I/R model was established through recanalization after ligation of left anterior descending coronary artery (LAD), and the MALAT1 knockdown model was established via tail intravenous injection of MALAT1 siRNA in the I/R + MALAT1 siRNA group. The ejection fraction (EF%) and fractional shortening (FS%) of rats in each group were detected via echocardiography and the infarction area in each group was detected using 2,3,5-triphenyl tetrazolium chloride (TTC) assay. Moreover, the morphological changes in myocardial cells in each group were detected via hematoxylin-eosin (H&E) staining, and the myocardial apoptosis level was detected via terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining. At the same time, the expression levels of the anti-apoptotic protein B-cell lymphoma-2 (Bcl-2) and pro-apoptotic protein Bcl-2 associated X protein (Bax) in myocardial tissues in each group were determined via Western blotting. Finally, the effect of MALAT1 knockdown on the phosphatidylinositol 3-hydroxy kinase/protein kinase B (PI3K/AKT) protein expression was detected via Western blotting. The expression level of lncRNA MALAT1 in myocardial tissues was significantly higher in the I/R group than that in the Control group (p<0.05). The MALAT1 knockdown could significantly improve the cardiac insufficiency caused by I/R injury, and increase both EF% and FS% in rats (p<0.05). In addition, the MALAT1 knockdown could markedly inhibit myocardial infarction caused by I/R injury and reduce the infarction area from (62.12 ± 1.29) to (27.66 ± 3.58; p<0.05). The results of the H&E staining showed that the myofilaments were arranged more orderly, the degrees of degradation and necrosis were lower and the cellular edema was significantly alleviated in the I/R + MALAT1 siRNA group compared with those in the I/R group. According to the results of TUNEL staining, the rats in I/R + MALAT1 siRNA group had a markedly lower level of myocardial apoptosis than the I/R group (p<0.05), and the Bax/Bcl-2 ratio also remarkably declined in the I/R + MALAT1 siRNA group (p<0.05). Furthermore, the results of Western blotting revealed that MALAT1 siRNA could significantly reverse the I/R injury-induced inhibition on the AKT phosphorylation (p<0.05). The MALAT1 knockdown can markedly improve the I/R-induced myocardial injury and promote the cardiac function of rats, whose mechanism may be related to the activation of the AKT signaling pathway by MALAT1 siRNA. Therefore, lncRNA MALAT1 is expected to be a new therapeutic target for myocardial I/R injury.

Effect of bradykinin on rats with thromboangiitis obliterans through PI3K/Akt signaling pathway.

To explore the effect of bradykinin on rats with thromboangiitis obliterans (TAO) through the phosphatidylinositol 3-hydroxy kinase/protein kinase B (PI3K/Akt) signaling pathway. The female Wistar rats were injected with lauric acid via the femoral artery to establish the TAO model, and they were randomly divided into control group (healthy rats), model group (TAO rats) and bradykinin group (TAO rats injected with bradykinin B2 receptor-specific inhibitor). The control was set in each group before the operation. The level of serum bradykinin in each group was detected via enzyme-linked immunosorbent assay (ELISA), and the reactive oxygen species (ROS) level, Caspase-3 activity and PI3K/Akt protein concentration in vascular tissues were measured via ELISA, Western blotting, ROS assay, and Caspase-3 activity assay, respectively. Moreover, the specific therapeutic mechanism of bradykinin was analyzed. In control group, the intima of the lower extremity venous tissues was smooth, the extima had no evident changes, and there was no inflammatory cell invasion around the arteries and veins. In model group, there was massive inflammatory cell invasion into the lower extremity venous tissues. In bradykinin group, fibrosis and atrophy occurred in venous tissues, the extima was thickened without fibrosis, and there was phagocytosis of neutrophils and mononuclear macrophages around the arteries and veins, as well as massive inflammatory infiltration. The PI3K/Akt protein concentration in lower extremity venous tissues was the highest in control group and the lowest in bradykinin group, and there were statistically significant differences (p<0.01). At 24 h after administration of doxorubicin (DOX), the level of ROS in lower extremity venous tissues was higher in bradykinin group than that in model group (p<0.05), and it was also higher in model group than that in control group (p<0.05). Besides, the activity of Caspase-3 in lower extremity venous tissues was significantly increased in bradykinin group compared with that in model group and control group, while it was slightly higher in model group than that in control group (p<0.05). The low expression of bradykinin can promote TAO in rats by the mechanism that it inhibits the PI3K/Akt signaling pathway to raise the oxidative stress level, thereby aggravating TAO.

Influence of LncRNA UCA1 on glucose metabolism in rats with diabetic nephropathy through PI3K-Akt signaling pathway.

The aim of this study was to investigate the influence of long non-coding ribonucleic acid (lncRNA) urothelial carcinoma associated 1 (UCA1) on glucose metabolism in rats with diabetic nephropathy (DN), and to explore its regulatory mechanism. A total of 30 healthy Sprague-Dawley (SD) rats were selected in this study. All rats were randomly divided into three groups, including the control group, the model group, and the lncRNA UCA1 inhibitor group. The rat model of DN was successfully established via intraperitoneal injection of streptozotocin (STZ). The pathological changes in kidney tissues were detected via hematoxylin-eosin (HE) staining. The levels of blood urea nitrogen (BUN), serum creatinine (Scr), and urinary protein (UP) were detected using the biochemical method. Meanwhile, the content of serum tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) was detected via enzyme-linked immunosorbent assay (ELISA). In addition, the messenger RNA (mRNA) and protein levels of phosphatidylinositol 3-hydroxy kinase (PI3K) and protein kinase B (Akt) in kidney tissues were detected via reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting, respectively. The model group showed severe pathological damage to the kidney, compared with the control group. Meanwhile, the levels of BUN, Scr and UP, and the content of serum TNF-α and IL-6 increased significantly in the model group. The mRNA and the protein levels of PI3K and Akt in kidney tissues of the model group were significantly up-regulated as well. LncRNA UCA1 inhibitor group exhibited relieved pathological damage to the kidney, compared with the model group. The levels of BUN, Scr and UP, and the content of serum TNF-α and IL-6 remarkably decreased in UCA1 inhibitor group. Furthermore, the mRNA and the protein levels of PI3K and Akt in kidney tissues of UCA1 inhibitor groups were significantly down-regulated. LncRNA UCA1 can relieve the pathological damage to the kidney, improve renal function, and alleviate inflammatory response in DN rats. The underlying mechanism may be related to the inhibition of the PI3K-Akt signaling pathway.

Promethazine inhibits neuronal apoptosis via PI3K/Akt signaling pathway in rats with cerebral infarction.

To study the effect of promethazine on neuronal apoptosis in rats with cerebral infarction (CI) through the phosphatidylinositol 3-hydroxy kinase/protein kinase B (PI3K/Akt) signaling pathway. A total of 36 Sprague-Dawley rats were randomly divided into the sham group (n=12), model group (n=12), and promethazine group (n=12). The external carotid artery was only exposed in the model group, and the ischemia-reperfusion model after CI was established using the suture method in the other two groups. After modeling, the normal saline was intraperitoneally injected in the sham group and model group, while promethazine was intraperitoneally injected in the promethazine group. The rats were sampled after 1 week of intervention. The neurological deficits of rats were evaluated using the Zea-Longa score, and the cognitive function, the spatial learning, and memory of rats were detected via the water maze test. Moreover, the expressions of B-cell lymphoma-2 (Bcl-2) and Bcl-2 associated X protein (Bax) in brain tissues were detected via immunohistochemistry, and the relative protein expressions of PI3K p85, PI3K p110, and p-Akt were detected via Western blotting. The mRNA expressions of Bax and Bcl-2 were detected via quantitative Polymerase Chain Reaction (qPCR), and the apoptosis was detected via terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay. The Zea-Longa score was significantly increased in the model group and promethazine group compared with that in the sham group (p<0.05), while it significantly declined in the promethazine group compared with that in the model group (p<0.05). The escape latency was significantly prolonged and the times of crossing platform were significantly reduced in the model group and promethazine group compared with those in the sham group (p<0.05), while the escape latency was significantly shortened and the times of crossing platform were significantly increased in the promethazine group compared with those in the model group (p<0.05). Compared with those in the sham group, the positive expression of Bax was significantly increased, while the positive expression of Bcl-2 was remarkably decreased in the model group and promethazine group (p<0.05). Compared with those in the model group, the positive expression of Bax was significantly decreased, while the positive expression of Bcl-2 was remarkably increased in the promethazine group (p<0.05). Besides, the model group and promethazine group had evidently higher relative protein expressions of PI3K p85, PI3K p110, and p-Akt than the sham group (p<0.05), while the promethazine group also had evidently higher relative protein expressions of PI3K p85, PI3K p110, and p-Akt than the model group (p<0.05). Compared with the sham group, model group, and promethazine group had remarkably increased relative mRNA expression of Bax, and remarkably decreased relative mRNA expression of Bcl-2 (p<0.05). Compared with those in the model group, the relative mRNA expression of Bax was remarkably decreased, while the relative mRNA expression of Bcl-2 was remarkably increased in the promethazine group (p<0.05). Finally, the apoptosis rate was significantly higher in the model group and promethazine group than that in the sham group (p<0.05), while it was significantly lower in the promethazine group than that in the model group (p<0.05). Promethazine inhibits neuronal apoptosis in CI rats by upregulating the PI3K/Akt signaling pathway, thereby exerting a protective effect.

Let-7 participates in the regulation of inflammatory response in spinal cord injury through PI3K/Akt signaling pathway.

To study the potential mechanism of let-7 in participating in the regulation of inflammatory response in spinal cord injury (SCI). A total of 40 male Sprague-Dawley rats were randomly divided into four groups: group A (Sham, n=10), group B (SCI+NC, n=10), group C (SCI+antagomir, n=10), and group D (SCI+mimics, n=10). The SCI model was established via operation in all groups. After successful modeling, let-7-antagomir negative control (80 mg/kg) was intraperitoneally injected in SCI+NC group at 5 d, an equal amount of let-7-antagomir was intraperitoneally injected in SCI+antagomir group at 5 d, and an equal amount of let-7-mimics was intraperitoneally injected in SCI+mimics group at 5 d. The inflammatory cells in experimental groups and control group were observed via hematoxylin-eosin (HE) staining. At the same time, the expression of let-7 in the four groups was detected via Reverse Transcription-Polymerase Chain Reaction (RT-PCR), the expressions of phosphatidylinositol 3-hydroxy kinase (PI3K) and protein kinase B (Akt) in all groups were detected via Western blotting, and the inflammatory index levels in each group were detected via enzyme-linked immunosorbent assay (ELISA). In Sham group, it was observed via HE staining that there were only a few bleeding or inflammatory cells. In SCI+NC group, bleeding and inflammatory cells basically tended to be stable. There were a large number of inflammatory cells in SCI+mimics group, while there were some inflammatory cells in SCI+antagomir group, but showing a decreasing trend compared with SCI+NC group. It was found in the RT-PCR detection of let-7 expression level in all groups that the expression of let-7 significantly declined in SCI+antangomir group compared with that in Sham group and SCI+NC group, and there were significant differences (p<0.01). The expression of let-7 was significantly increased in SCI+mimics group compared with that in Sham group and SCI+NC group, and there were significant differences (p<0.01). The results of Western blotting revealed that the PI3K and Akt protein expressions were significantly decreased in SCI+mimics group compared with those in SCI+antagomir group, SCI+NC group, and Sham group (p<0.05). The ELISA results showed that the levels of inflammatory factors in SCI+mimics group, SCI+antagomir group, and SCI+NC group were significantly higher than those in Sham group. In SCI+mimics group, the levels of inflammatory factors were abnormally high and reached extremely significant levels (p<0.05), indicating that let-7 promotes the inflammatory response after SCI. Let-7 participates in the regulation of inflammatory response in SCI through the PI3K/Akt signaling pathway.

MiR-126 promotes endothelial cell apoptosis by targeting PI3K/Akt in rats with lower limb arteriosclerosis obliterans.

To investigate the influence of micro ribonucleic acid (miR)-126 on the rats with lower limb arteriosclerosis obliterans (ASO). Male Sprague- Dawley rats aged 3 months old were randomly divided into Sham operation group (Control group, n=10) and Model group (n=10), and the model of lower limb ASO was established. After modeling, the expression of miR-126 in arteries was detected using quantitative Real Time-Polymerase Chain Reaction (qRT-PCR), and the change in the downstream signaling pathway was examined via Western blotting. The human umbilical vein endothelial cells (HUVECs) were induced by oxidized low-density lipoprotein (Ox-LDL) to establish the model of endothelial injury, followed by detection of miR-126 expression. Then, the Luciferase assay was performed to verify the downstream target gene of miR-126. After being cultured, HUVECs were set as Control group, Ox-LDL induction group, and Ox-LDL + miR-126 inhibitor group, and the expressions of phosphorylated-protein kinase B (p-Akt) and cleaved cysteine-aspartic protease-3 (Caspase-3) were detected in the above groups. After the establishment of the model, the expression level of miR-126 was raised in vessels, but the phosphatidylinositol 3-hydroxy kinase (PI3K)/Akt signals were weakened (p<0.01). Ox-LDL-induced endothelial cell apoptosis promoted the expression of miR-126, and the difference was statistically significant. The bioinformatics analysis results showed that PI3KR2 was a direct target of miR-126, which was also proven via the Luciferase assay. Moreover, the transfection with miR-126 inhibitor into endothelial cells suppressed Ox-LDL-induced cell apoptosis, thereby persistently activating the PI3K/Akt signaling pathway (p<0.01). In rats with lower limb arteriosclerosis obliterans (ASO), miR-126 represses the PI3K/Akt signaling pathway to accelerate endothelial cell apoptosis.

Dexmedetomidine suppresses sevoflurane anesthesia-induced neuroinflammation through activation of the PI3K/Akt/mTOR pathway

BackgroundSevoflurane, an inhalational general anesthetic, has become one of the most widely used inhalational anesthetics in surgery. However, previous studies have found that sevoflurane anesthesia can trigger an inflammatory response, resulting in secondary damage. Dexmedetomidine (DEX), a highly-selective α adrenergic receptor agonist, is widely used as an anesthetic adjuvant in the clinic. In this study we investigated whether DEX was able to suppress sevoflurane-induced neuroinflammation.MethodsThe aim was to determine the mechanism of action of the suppressive effect of DEX using a rat model. Rats were randomly divided into a control group (n = 10), low-dose sevoflurane group (L-Sev; n = 10), high-dose sevoflurane group (H-Sev; n = 10), vehicle group (n = 10), DEX group (n = 10) and DEX + LY294002 (a specific inhibitor of PI3K) group (n = 10). The rats in vehicle, DEX and DEX + LY294002 groups were in the presence of high-dose sevoflurane exposure. Western blotting was used to measure the expression of proinflammatory cytokines (IL-6, IL-8, TNF-α) and the activity level of the phosphatidylinositol 3-hydroxy kinase/protein kinase B/mammalian target of rapamycin (PI3K/Akt/mTOR) pathway.ResultsWe found that sevoflurane anesthesia induced an increase in the levels of pro-inflammatory cytokines, while decreasing activation of the PI3K/Akt/mTOR pathway in both the cortex and hippocampus of rats. Treatment with DEX reduced pro-inflammatory cytokine levels and prevented inactivation of the PI3K/Akt/mTOR pathway. Moreover, LY294002, an inhibitor of the PI3K/Akt/mTOR pathway, reduced the anti-inflammatory activity of DEX.ConclusionsThese data suggest that the PI3K/Akt/mTOR pathway contributes to sevoflurane-induced neuroinflammation and that activation of PI3K/Akt/mTOR signaling by DEX could help reduce the neuroinflammatory effects of sevoflurane.

Low concentration of rutin treatment might alleviate the cardiotoxicity effect of pirarubicin on cardiomyocytes via activation of PI3K/AKT/mTOR signaling pathway.

Cancer is the leading cause of deaths around the world, especially in low- and middle- income countries. Pirarubicin (THP) is an effective drug for treatment of cancer, however, there still exists cardiotoxic effects of THP. Rutin is a kind of antioxidative compound extracted from plants, and might be a protective compound for cardiomyocytes. Phosphatidylinositol 3-hydroxy kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signaling pathway is critical for cellular survival, proliferation and metabolism, and thus we speculated rutin might perform a protective role in cardiomyocytes via PI3K/AKT/mTOR signaling pathway. And in this experiment, we first established a cardiotoxicity model of THP in mice model and cell models, and then found that rutin treatment could increase the proliferation of cells at low concentration. Then we explored the possible mechanism of the protective effect of rutin using Western blotting, quantitative polymerase chain reaction (qPCR) and ELISA methods, and found that the activation of PI3K/AKT/mTOR/nuclear factor-κB (NF-κB) signaling pathway was increased, and expression of downstream molecules involved in antioxidative stress were also increased. We further noticed that concentration of angiogenesis promoting factors were also increased in medium of cultured cells. Thus, we speculated that rutin could increase the activation of PI3K/AKT/mTOR signaling pathway, further decrease the oxidative stress level via increasing the expression of antioxidative stress enzymes with the increasing concentration of angiogenesis promoting factors, resulting in the protective role in cardiomyocytes and cardiac function.

Effect of miR-7 on resistance of breast cancer cells to adriamycin via regulating EGFR/PI3K signaling pathway.

To explore whether micro ribonucleic acid (miR)-7 affects the resistance of breast cancer cells to adriamycin (ADR) through regulating the epidermal growth factor receptor (EGFR)/phosphatidylinositol 3-hydroxy kinase (PI3K) signaling pathway. MiR-7 expression was compared among MCF-10A, MCF-7 and MCF-7/ADR cells. The MCF-7/ADR cells were divided into three groups, namely miR-7 control group (MCF-7/ADR drug-resistant strains), miR-7 inhibition group (miR-7-inhibited MCF-7/ADR drug-resistant strains) and miR-7 promotion group (MCF-7/ADR drug-resistant strains transfected with miR-7), and the messenger RNA (mRNA) and protein expression levels of MCF-7/ADR were evaluated via Western blotting. The expression level of miR-7 was substantially decreased in MCF-7 and MCF-7/ADR cells (p<0.05), and it was lowered more obviously in MCF-7/ADR cells than that in MCF-7 cells (p<0.05). Compared with that in miR-7 control group, miR-7 expression in miR-7 promotion group was notably raised (p<0.05), proving that the sensitivity of MCF-7/ADR cells to ADR was enhanced, while that in miR-7 inhibition group was significantly lowered (p<0.05). Compared with those in miR-7 control group, the mRNA and protein expression levels of EGFR and PI3K were elevated in miR-7 inhibition group (p<0.05), while they were lowered in miR-7 promotion group (p<0.05). Additionally, compared with those in miR-7 control group, the proliferation and apoptosis abilities of cells in miR-7 inhibition group were markedly enhanced (p<0.05) and weakened (p<0.05), respectively, while they were weakened (p<0.05) and significantly strengthened (p<0.05), respectively in miR-7 inhibition group. MiR-7 plays an important role in the resistance of breast cancer cells to ADR, and its over-expression can inhibit the EGFR/PI3K signaling pathway to raise their sensitivity to the chemotherapy drug ADR.