LY294002 Group Research Articles

Sevoflurane ameliorates adriamycin-induced myocardial injury in rats through the PI3K/Akt/GSK-3β pathway.

The aim of this study was to explore the effects of sevoflurane (SEV) pretreatment on Adriamycin (ADR)-induced myocardial injury through the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/glycogen synthase kinase-3β (GSK-3β) pathway. A total of 24 rats weighing 200-250 g were divided into four groups, including: control group (C group), ADR injection group (ADR group), SEV pretreatment group (ADR + SEV group) and inhibitor group (ADR + SEV + LY group). H9c2 cells were cultured in vitro and were divided into control group (C group), ADR treatment group (ADR group), and SEV pretreatment group (ADR + SEV group) and inhibitor group (ADR + SEV + LY group) as well. Next, the content of aspartate aminotransferase (AST), lactate dehydrogenase (LDH) and creatine kinase (CK) in the serum was detected via Enzyme-Linked Immunosorbent Assay (ELISA). Hematoxylin-eosin (HE) staining assay was performed to determine the severity of myocardial injury. Meanwhile, the apoptosis rate of cells was detected through terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL) assay. Additionally, Western blotting (WB) was employed to measure the protein expression levels of phosphorylated (p)-GSK-3β, p-PI3K, Akt and p-Akt. Compared with C group, ADR significantly increased the content of AST, LDH and CK in the serum (p<0.01), reduced protein expression levels of p-GSK-3β, p-PI3K and p-Akt (p<0.01), increased apoptosis rate (p<0.01), and induced myocardial injury. SEV pretreatment significantly alleviated the effect of ADR, manifested as significantly lowered content of AST, LDH and CK in the serum (p<0.01), distinctly elevated protein expression levels of p-GSK-3β, p-PI3K and p-Akt (p<0.01), notably reduced apoptosis rate (p<0.01), and relieved myocardial injury. LY294002 remarkably inhibited the protective effect of SEV against myocardial injury (p<0.01) CONCLUSIONS: SEV is able to prominently ameliorate ADR-induced myocardial injury by regulating the phosphorylation level of the PI3K/Akt/GSK-3β signaling pathway.

AGEs promote calcification of HASMCs by mediating Pi3k/AKT-GSK3β signaling.

This study aimed to investigate the effects of advanced glycation end products (AGEs) on the calcification of human arterial smooth muscle cells (HASMCs) and to explore whether AGEs can promote the calcification of HASMCs by activating the phosphoinositide 3-kinase (PI3K)/AKT-glycogen synthase kinase 3 beta (GSK3-β) axis. Cultured HASMCs were divided into five groups: blank control group, dimethyl sulfoxide (vehicle) group, AGEs group, LY294002 (AKT inhibitor) group, and TWS119 (GSK3-β inhibitor) group. Cells were pretreated with either vehicle, LY294002, or TWS119 for 2 hours followed by incubation with AGEs (25 μg/mL) for 5 days, and the expression levels of proteins in each group were analyzed by western blotting. AGE treatment promoted HASMC calcification, which coincided with increased expression of p-AKT and p-GSK3-β (serine 9). Also, AGEs upregulated the expression of osteoprotegerin and bone morphogenetic protein, and these effects were suppressed by LY294002 but enhanced by TWS119. In conclusion, AGEs promote calcification of HASMCs, and this effect is ameliorated by inhibition of AKT activity but potentiated by inhibition of GSK3-β activity. Hence, AGEs trigger HASMC calcification by regulating PI3K/AKT-GSK3-β signaling.

Oleanolic Acid Improves the Symptom of Renal Ischemia Reperfusion Injury via the PI3K/AKT Pathway

Purpose: The aim of this study was to investigate the therapeutic effect of oleanolic acid (OA) on the renal ischemia reperfusion injury (RIRI) and the possible mechanism. Methods: The RIRI model was successfully established in rats. OA, LY294002 (a PI3K inhibitor), and OA combined with LY294002 were dosed to rats in 3 therapeutic groups, respectively. The blood was collected to detect the concentration of Cr and BUN by ELISA. The kidney of each rat was collected to detect the concentration of renal injury factor (Kim-1) and the HE staining was performed. Western blot was used to detect the expression level of PI3K, p-AKT, AKT, PDK1, Skp2, and p27 in the renal tissue homogenate. Results: The symptom of vacuolar degeneration and interstitial edema was greatly improved in the rat kidney from the 3 therapeutic groups, compared with that from the RIRI model group. No significant difference was observed among the 3 therapeutic groups. The concentration of Cr in the 3 therapeutic groups was greatly lower than that in the RIRI model group. The expression level of p-AKT/AKT, PI3K, PDK1, Skp2, and p27 in OA group, LY294002 group, and OA combined with LY294002 group was significantly lower than that in the RIRI model group, respectively. Conclusion: OA could improve the symptom of RIRI, possibly by inhibiting PI3K/AKT signal pathway.

153 Effect of dietary live yeast supplementation on growth performance, diarrhea severity, antioxidant status and immunological parameters of weaned piglets

Abstract The objective of this study was to determine the effects of a Saccharomyces cerevisiae yeast as a substitute to AGP on intestinal health and performance of piglets. A total of 192 piglets weaned at 28 d of age were randomly assigned into one of four treatment groups with 8 pens/ treatment and 6 pigs/pen. Piglets were supplemented with either a regular nursery diet with two phase feeding (Phase 1: d1-d14 and Phase 2: d15-d28) (CON); CON diet supplemented with 75mg/kg Chlortetracycline plus ZnO 2000 mg/kg in phase 1 and 160 mg/kg in phase 2 (AGP); Con diet supplemented with 2kg/ton live yeast in both phases (LY); Con diet plus live yeast 1kg/ton in both phases plus ZnO 2000 mg/kg in phase 1 and 160 mg/kg in phase 2 (LY-ZnO). During Phase 1, LY and LY-ZnO group had significantly higher ADG (p&amp;lt; 0.01). In Phase 2 only the LY group has a significantly higher ADG (P=0.02). The LY and LY-ZnO group had also better feed efficiency (P&amp;lt; 0.05) during phase 1. On the other hand, the CON group had significantly higher (P&amp;lt; 0.01) diarrhea score. Test on antioxidant status has not shown any significant difference between the groups for most parameters measured; however, the concentration of Malondialdehyde (MDA, a marker for oxidative stress) was significantly decreased (P&amp;lt; 0.01) in the LY and LY-ZnO group on day 28. Furthermore, superoxide dismutase (SOD, the most powerful antioxidant) was significantly increased (P&amp;lt; 0.05) in these two treatment groups at different time points. Live yeast and Live yeast plus ZnO treatment has also significantly increased (P&amp;lt; 0.01) concentration of IgA and IgG in the serum of piglets on d7 and d14. In conclusion, supplementation of nursery piglet diets with either live yeast or live yeast plus ZnO improved zootechnical performance of animals by improving gut health through reduction of oxidative stress and enhancement of immune response, suggesting that yeast probiotics could be a suitable alternative to substitute AGP.

Avenanthramide C Prevents Neuronal Apoptosis via PI3K/Akt/GSK3β Signaling Pathway Following Middle Cerebral Artery Occlusion.

Avenanthramides are a group of phenolic alkaloids that have been shown to have anti-inflammatory, anti-oxidant, anti-atherogenic, and vasodilation effects. The aim of the present study was to investigate the neuroprotective effect of avenanthramide-c (Avn-c) in focal brain ischemia and reperfusion injury using middle cerebral artery occlusion (MCAo) model with mice. Male C57BL/6 mice were divided into 4 groups: sham, control (MCAo), Avn-c, and Avn-c + LY294002 (phosphoinositide 3-kinase inhibitor) group. They were subjected to 60 min MCAo followed by reperfusion. Brain infarct volume and neurological deficit scores were measured after 24 h of reperfusion. We evaluated the blood brain barrier (BBB) integrity (ZO-1, VE-cadherin and occludin) and apoptosis (Bax, Bcl2, caspase3, Cytochrome C, and poly ADP ribose polymerase(PARP)-1). We also measured GSK3β for evaluation of the downstream mechanism of Akt. We examined the effect of the Avn-c in the phosphoinositide 3-kinase pathway. Avn-c reduced neurological score and infarction size. Avn-c inhibited the MCAo-induced disruption of tight junction proteins. Avn-c decreased apoptotic protein expression (Bax, Cytochrome C, and cleaved PARP-1) and increased anti-apoptotic protein expression (Bcl2) after MCAo. Akt and GSK3β were decreased in MCAo group and were restored in Avn-c group. This effect of Avn-c was abolished by PI3K inhibitor. In summary, Avn-c showed neuroprotective effects through PI3K-Akt-GSK3β signaling pathway.

Klotho gene improves oxidative stress injury after myocardial infarction

The aim of the present study was to investigate the effects and mechanisms of the Klotho gene in oxidative stress injury after myocardial infarction. Sprague-Dawley rats were divided into five groups (sham, model, pDC316, LY294002, and pDC316-Klotho). Subsequently, the superoxide dismutase (SOD), glutathione (GSH), and malondialdehyde (MDA) concentrations were measured in myocardial tissues. Additionally, pathological differences among the groups were evaluated using hematoxylin and eosin and Masson's trichrome staining. Apoptosis was assayed by terminal deoxynucleotidyl transferase 2'-deoxyuridine-5'-triphosphate nick end-labeling assay, evaluated Klotho protein expression by immunohistochemical assay, and assessed Nrf 2 and ARE protein expressions using western blotting assay. As compared with in the sham group, the SOD, MDA, and GSH concentrations were significantly deteriorated (P<0.001, respectively); cardiomyocyte apoptosis index values were significantly increased (P<0.001); Klotho protein expression was significantly depressed; and Nrf-2 and ARE protein expressions were significantly (P<0.001, respectively) in the model and pDC316 groups. However, with Klotho supplementation by pDC316 transfection, as compared with in the model group, the SOD, MDA, and GSH concentrations were significantly improved (P<0.001, respectively); the cardiomyocyte apoptosis index values were significantly suppressed (P<0.001); and the pathology was improved. Further, the Klotho protein expression of the pDC316-Klotho group was significantly upregulated and the Nrf-2 and ARE proteins expressions of the LY294002 and pDC316-Klotho groups were significantly suppressed. Klotho overexpression improved findings of oxidative stress injury after myocardial infarction.

Abstract 15183: Thrombopoietin Has a Protective Effect Against Apoptosis of Endothelial Cells Through Activating Pi3k/akt Pathway

Introduction: Thrombopoietin (TPO) is a hematopoietic growth factor for platelet lineage. TPO was found to be neuroprotective in hypoxic-ischemic neonatal rat brain models and treatment with TPO reduced brain damage and improved sensorimotor functions (Li L et al, Aging-US, 2020). However, the underlying mechanism of TPO in this model, and its role in neural cells and endothelial cells were still unclear. Methods: C17.2 cells were divided into control (0.5% FCS), Normal (10% FCS), TPO and TPO + LY294002 groups. Human umbilical vein endothelial (HUVEC) cells were divided in to normal, CoCl 2 , TPO and TPO + CoCl2 groups. The expressed of TPO and c-mpl was tested by RT-PCR. The cell viability and apoptosis of each group were tested by Cell Counter Kit 8 (CCK-8) assay and flow cytometry. The expression of Caspase-3 and mitochondrial membrane potential (MMP) were then determined by flow cytometry with Caspase-3-PE and JC-1. The effect of TPO in PI3K/AKT pathway was detected by using Western blot. Results: Both TPO and c-mpl are expressed in the neurons of the human CNS. TPO was also detected in human cerebrospinal fluid. TPO promoted C17.2 cell proliferation through activation of the PI3K/Akt signaling pathway. Via the Bcl-2/BAX signaling pathway, TPO exerted an anti-apoptotic effect by suppressing mitochondria membrane potentials. We also investigated the protective effect of TPO on human endothelial cells. CoCl 2 significantly inhibited the growth of HUVECs. The cell viability of HUVECs decreased gradually with the enhancement of CoCl2 at a gradient of chemical concentrations (r= -0.997). CoCl 2 dramatically increased apoptosis of HUVECs, whereas pre-treatment with TPO rescued cell apoptosis induced by CoCl 2 (P&lt;0.001). Further investigation found that TPO decreased the expression of Caspase-3 and inhibited the reduction of MMP induced by CoCl 2 (P&lt;0.05). TPO increased the activation of PI3K/AKT pathway in HUVECs. Conclusions: TPO has a protective effect against apoptosis of neural cells and endothelial cells through activating the PI3K/AKT pathway, thus decreasing the expression of apoptosis protease Caspase-3 and inhibiting the reduction of MMP.

Effect of moxa-cone moxibustion at lung's back-shu points and front-mu points on Th17/Treg balance in mice with asthma

To observe the effect of moxa-cone moxibustion at lung's back-shu points and front-mu points on the expression of helper T lymphocyte 17 (Th 17)/regulatory T lymphocyte (Treg) in mice with asthma, and to explore the possible mechanism of moxa-cone moxibustion on asthma. Sixty SPF-grade healthy male Balb/c mice were randomly divided into a normal group, a model group, an LY294002 group (LY group), an electroacupuncture (EA) group and a moxibustion group, 12 mice in each group. Asthma model was replicated by using ovalbumin (OVA) sensitization. Except the mice in the normal group, all the mice were intraperitoneally injected with sensitization solution (containing 15 μg of OVA and 30 mg of aluminum hydroxide) on the 1st day, 7th day and 14th day, 0.5 mL per mice; from the 15th day, 1% OVA solution was atomized for 20 min, once a day for 2 weeks; the mice in the normal group was treated with identical operations but with 0.9% sodium chloride solution. The mice in the LY group were treated with injection of LY294002 at tail vein on the 13th day, 14th day and 15th day. At the beginning of the 15th day, The mice in the EA group were treated with EA at "Feishu" (BL 13) and "Zhongfu" (LU 1) with disperse-dense wave, frequency of 2 Hz/20 Hz, intensity of 1 mA, 15 min each time, once a day for 2 weeks. The mice in the moxibustion group was treated with moxa-cone moxibustion at "Feishu" (BL 13) and "Zhongfu" (LU 1) from the 15th day, three moxa-cones per acupoint, once a day for 2 weeks. On the 16th day, 18th day and 22nd day, the incubation period of asthma was recorded. On the 29th day, all the samples were collected. The expressions of IL-17 and IL-10 in serum and bronchoalveolar lavage fluid (BALF) were detected by ELISA method. The pathological changes of lung tissue were observed by HE staining. The percentage of Th17, Treg and Th17/Treg ratio in spleen tissue were detected by flow cytometry method. Compared with the normal group, the incubation period of asthma in the model group was significantly shortened (P<0.01), while that in the LY group, the EA group and the moxibustion group was longer than that in the model group (P<0.01). After the intervention, bleeding and inflammatory exudation in the lung tissue were observed in the model group, and a large number of inflammatory cell infiltration around the bronchus and alveoli was found, and extensive consolidation appeared; the pathological changes of lung tissue in the LY group, the EA group and the moxibustion group were significantly reduced, and the structure of bronchus and alveoli was basically normal, and the inflammatory exudation and inflammatory cell infiltration were significantly reduced compared with the model group; the inflammatory reaction in the moxibustion group was slightly less than that in the LY group and the EA group. Compared with the normal group, the content of IL-17 in serum and BALF, the percentage of Th17 and Th17/Treg ratio in spleen tissue in the model group after intervention were increased (P<0.01), while the content of IL-10 in serum and BALF and the percentage of Treg in spleen tissue were decreased (P<0.01, P<0.05). Compared with the model group, the contents of IL-17 in serum and BALF, Th17 percentage and Th17/Treg ratio in spleen tissue in the LY group, the EA group and the moxibustion group were significantly reduced (P<0.01, P<0.05), while the content of IL-10 in serum and BALF and the percentage of Treg in spleen tissue were increased (P<0.01, P<0.05). The above indice had no significant difference among all the intervention groups (P>0.05). The Th17/Treg is imbalanced in asthmatic body. The moxibustion at lung's back-shu points and front-mu points can control asthma by regulating Th17/Treg imbalance and restoring immune homeostasis.

Effects of Rearing Conditions and Sex on Cecal Microbiota in Ducks.

The intestinal microbiome influences the health of animals. However, little is known about the impact of indoor conditions and sex on intestinal microbiome diversity and composition in ducks. The present study aimed to investigate differences in the cecal microbiome between male and female ducks reared on the floor (PY group) or in cages (LY group). We also determined the relationships between cecal microbiota composition and slaughter traits, and the expression levels of mucosal and intestinal structural genes in ducks. There was a slight difference in slaughter traits among the groups, with cecum weight being significantly lighter in the LY compared with the PY group, especially in females (p < 0.05). Analysis of the alpha diversity of the cecal microbiota between males and females in the LY and PY groups showed that LY males had significantly lower diversity and richness. Beta diversity analysis demonstrated differences in the microbiota composition in relation to rearing conditions, and a significant difference between the sexes in the PY groups. The dominant bacterial phyla in duck cecum were Bacteroidetes, Firmicutes, Proteobacteria, and Fusobacteria. The relative abundances of the most common bacteria revealed that the intestinal microbiota diversity and composition were affected by both feeding conditions and sex. Several bacterial genera were detected differentially among the groups. These genera were correlated with slaughter traits and expression levels of mucosal and cecal structural genes in ducks. In conclusion, rearing conditions, sex, and associated changes in the cecal microbiota are thus associated with gut barrier functions in ducks.

Qishen Huanwu capsule reduces pirarubicin-induced cardiotoxicity in rats by activating the PI3K/Akt/mTOR pathway

One of the common adverse reactions to anthracyclines, a group of chemotherapeutics, is cardiotoxicity. Cancer patients receiving anthracycline-based chemotherapeutic regimens often discontinue treatment due to cardiotoxicity. How to prevent and reduce the cardiotoxicity of anthracyclines is one of the hot topics in the field of onco-cardiology. Traditional Chinese medicine can reduce the toxic side effects of chemotherapeutics. The present study aimed to investigate the protective effect of Qishen Huanwu capsule (QSHWC) on pirarubicin (THP)-induced myocardial injury in rats and the underlying mechanisms. Forty-eight male Sprague-Dawley (SD) rats were randomly divided into six groups: control group, THP, low-dose QSHWC, moderate-dose QSHWC, high-dose QSHWC, and LY294002 [phosphatidylinositol 3-kinase (PI3K) inhibitor] (n=8 each). Echocardiographic examination was performed to examine heart structure and function. Hematoxylin and eosin (HE) staining was conducted to examine histopathological changes in myocardial tissue. Immunofluorescence staining was carried out to examine the expression of the autophagosome-specific marker protein microtubule-associated protein 1 light chain 3 (LC3). Western blot was performed to analyze the expression of LC3-I, LC3-II, PI3K, phosphorylated (p)-PI3K, protein kinase B (Akt), p-Akt, mammalian target of rapamycin (mTOR), and p-mTOR. Compared with the control group, the THP group had a higher left ventricular end-systolic diameter (LVESD), lower left ventricular ejection fraction (LVEF), lower left ventricular fractional shortening (LVFS), and inferior heart function. In addition, compared with the control group, the THP group had significantly higher LC3 protein expression, a significantly higher LC3-II/LC3-I ratio (P<0.05), and significantly lower p-PI3K, p-Akt, and p-mTOR (P<0.05). QSHWC attenuated the THP-induced decline in heart function, downregulated LC3 protein in rat myocardial tissue, decreased the LC3-II/LC3-I ratio, and increased p-PI3K, p-Akt, and p-mTOR. In the LY294002 group, the above effects of QSHWC were reversed. QSHWC alleviated THP-induced myocardial injury. The underlying mechanism was related to the activation of the PI3K/Akt/mTOR pathway and the mitigation of the excessive autophagy of cardiomyocytes caused by THP.

Glutamine relieves oxidative stress through PI3K/Akt signaling pathway in DSS-induced ulcerative colitis mice.

Objective(s):Ulcerative colitis (UC) is a kind of complex immune disease, and a major cause of destruction of intestinal barrier and oxidative stress in this field. In this paper, glutamine (Gln) was believed to offer protection against oxidative stress injury in colitis mice.Materials and Methods:Thirty mice were randomly assigned into control, model, LY294002 (PI3K/Akt inhibitor), Gln, Gln+LY294002 and 5-Aminosalicylic acid (5-ASA) groups. The mice in the experimental group drank 4% dextran sulfate sodium salt (DSS) for 7 consecutive days. The protective effect of Gln on oxidative stress was quantified by keeping colitis mice, involving Phosphatidylinositol-3-kinase (PI3K)/Protein kinase B (Akt)/mammalian target of Rapamycin (mTOR) signaling pathway, with different medications or distilled water through intragastric administration for 10 consecutive days.Results: In vivo administration of Gln, LY294002 or 5-ASA was found to ameliorate the symptoms of colitis in mice, such as reduced growth, loose stools and stool bleeding; protected DSS-induced colitis mice from goblet cell loss, lymphocytosis, mucosal erosion, loss of crypts, and neutrophil infiltration; improved the activity of superoxide dismutase (SOD) and glutathione peroxidase (GSH-XP); decreased the content of malondialdehyde (MDA); and inhibited the activation of PI3K/Akt signaling pathway.Conclusion:Administration of Gln to the DSS-induced colitis mice led to a clearly reduction in oxidative stress-induced injury. The Gln is confirmed as inhibiting the PI3K/Akt signaling pathway activity.

Ischaemic preconditioning-induced serum exosomes protect against myocardial ischaemia/reperfusion injury in rats by activating the PI3K/AKT signalling pathway.

Ischaemia/reperfusion (I/R) injury can lead to severe arrhythmia and aggravate myocardial damage. Exosomes are small-membrane vesicles that play a protective role in myocardial I/R injury. This study aimed to explore the protective effects of ischaemic preconditioning (IPC)-induced serum exosomes (IPC-Exo) on myocardial I/R injury in rats and its underlying mechanism. Serum exosomes were extracted from IPC rats and quantified using a bicinchoninic acid assay kit. IPC-Exo (50 μg) was injected into the infarcted myocardium immediately after ligation. Rats were randomly divided into Sham, I/R, IPC-Exo + I/R, I/R + LY294002, and I/R + IPC-Exo + LY294002 groups. Haemodynamic parameters were measured by physiological recording. Transthoracic echocardiography was used to detect cardiac function. The serum levels of creatine kinase isomer-MB, lactate dehydrogenase, aspartate transaminase, tumour necrosis factor-alpha, interleukin (IL)-1β, and IL-10 were detected by enzyme-linked immunosorbent assay. Triphenyl tetrazolium chloride staining was used to measure the myocardial infarct size. Apoptosis in myocardial tissues was detected by TUNEL staining. Western blotting was used to detect the levels of PI3K/AKT and apoptosis-related proteins. Our results showed that treatment with IPC-Exo ameliorated cardiac function and reduced inflammatory factor production, cardiomyocyte apoptosis, and myocardial infarct size. Moreover, IPC-Exo treatment promoted the protein expression of Bcl-2, p-PI3K, and p-AKT but inhibited that of caspase-3 and Bax. However, treatment with LY294002 significantly reversed that IPC-Exo-induced increase in p-PI3K and p-AKT levels, improvement of haemodynamics, and decrease of inflammatory factor production and apoptosis in the I/R + IPC-Exo group. Taken together, our results suggest that IPC-Exo may alleviate I/R injury via activating the PI3K/AKT signalling pathway.

Study on mechanism of curcumin in treatment of acute pancreatitis based on regulation of PI3K-Akt signaling pathway by miR-198

Curcumin was used to interfere with acute pancreatitis model rats to explore its possible mechanism. One hundred and twenty rats were randomly divided into blank group, model group, model+curcumin group, model+mock+curcumin group, model+antagonist+curcumin group and model+curcumin+LY294002 group, with 20 rats in each group. The wet/dry weight ratio of pancreatic tissue was measured and the pathological changes of pancreas were observed by HE staining. The apoptosis was detected by TUNEL staining; the levels of serum amylase, lipase, Bcl-2 and Bax were detected by ELISA, and the levels of PI3 K, Akt and p-Akt in pancreatic tissue were measured by Western blot. HE staining showed that curcumin could improve the pathological changes of pancreas and reduce the pathological score of pancreas, while ELISA results showed that curcumin could decrease the levels of amylase, lipase and Bax in peripheral serum and increase the concentration of Bcl-2. Western blot results showed that the expression levels of PI3 K and p-Akt in pancreatic tissue of model rats were up-regulated after the intervention of curcumin, and the apoptosis rate of pancreatic cells decreased in TUNEL staining. The above effects could be weakened by miR-198 antagonist and PI3 K-Akt signal pathway inhibitor LY294002. In conclusion, curcumin has an ideal effect on acute pancreatitis, and its mechanism may be mediated by miR-198-PI3 K-Akt axis.

Proliferation and invasion of ovarian cancer SK-OV-3 cells inhibited by Tectoridin

Objective: To study the effects of Tectoridin on the proliferation, migration and invasion of ovarian cancer SK-OV-3 cells and its mechanism. Methods: SK-OV-3 cells were treated with 0, 5, 10, 50 and 100 μg/L Tectoridin for 24 hours. The proliferation of SK-OV-3 cells treated with different concentrations of Tectoridin was detected by cell counting kit-8 (CCK-8). The migration was analyzed by wound healing test and the invasion was observed by Transwell. The effects of different concentrations of Tectoridin on the protein levels of phosphoinositide 3-kinase (PI3K)/serine-threonine kinase (AKT) signaling pathway were detected by western blot assay. Results: The A values of 0, 5, 10, 50, 100 μg/L Tectoridin groups were 0.857±0.043, 0.725±0.036, 0.611±0.032, 0.410±0.027, and 0.321±0.023, respectively. The relative migration distances of the cells were 1.000±0.083, 0.896±0.092, 0.644±0.075, 0.432±0.056, and 0.215±0.043, respectively. The numbers of cell invasion were (106.258±11.785), (93.241±10.251), (74.258±7.963), (40.236±5.210), and (32.147±3.458), respectively. The phosphorylated PI3K (p-PI3K) protein levels were 1.000±0.079, 0.875±.089, 0.727±0.075, 0.452±0.053, 0.365±0.052, respectively. The phosphorylated AKT (p-AKT) protein levels were 1.000±0.070, 0.816±0.072, 0.635±0.079, 0.463±0.065, 0.305±0.047, respectively. Compared with the 0 μg/L Tectoridin group, the differences of 5, 10, 50, 100 μg/L Tectoridin group were statistically significant (all P<0.05). The A values of the control group and the LY294002 group were 0.873±0.081 and 0.494±0.038, respectively. The relative cell migration distances were 1.000±0.081 and 0.523±0.051, respectively. The numbers of cell invasion were (106.228±11.783) and (61.243±9.251), respectively. The protein levels of p-PI3K were 1.000±0.075, 0.521±0.046, respectively. The protein levels of p-AKT were 1.000±0.070, 0.465±0.051, respectively. Compared with the control group, the difference in the LY294002 group was statistically significant (P<0.05). Conclusion: Tectoridin may inhibit proliferation, migration and invasion of SK-OV-3 cells by regulating PI3K/AKT signaling pathway.

Effect of PI3K/Akt Signaling Pathway on PRAS40Thr246 Phosphorylation in Gastric Cancer Cells

We aimed to investigate the effect of PI3K/Akt signaling pathway on PRAS40Thr246 phosphorylation in gastric cancer cells. The study was conducted from April 2017 to January 2018 in Zhongshan Hospital, Xiamen University, Xiamen, China. Gastric cancer cells were divided into three groups: gastric cancer cell group, LY294002 group and MK-2206 group. Specific tests were conducted accordingly. Inhibition of PI3K/Akt signaling pathway activation and PRAS40Thr246 phosphorylation could inhibit proliferation and invasion and promote apoptosis of gastric cancer cells, and PRAS40Thr246 phosphorylation could activate PI3K/Akt signaling pathway. The levels of PI3K/Akt signaling pathway related proteins and p-PRAS40Thr246 were significantly increased in gastric cancer cells. p-PRAS40-Thr246 was able to reflect the activation of the PI3K/Akt signaling pathway, reflecting the sensitivity of the PI3K/AKT signaling pathway to inhibitors.

Protective effect of insulin on burn serum-challenged cardiomyocytes in vitro

To investigate the protective effect of insulin on burn serum-challenged cardiomyocytes in vitro. Primary culture of cardiomyocytes from Sprague-Dawley (SD) 2-day-old neonate rats were divided into sham group, burn group, insulin group, and insulin activation inhibitor LY294002 pretreatment group (LY group). The model of cardiomyocytes injury induced by burn serum of 3-month-old SD rats [the serum of abdominal aortic was collected at 6 hours after modelling 30% total surface area (TBSA) III degree scald rat] was reproduced. In the insulin group, 10% burn serum and insulin (10 U/L) were added into cell culture medium, and in the LY group, LY294002 (50 μmol/L) was pretreated for 30 minutes before the addition of burn serum and insulin. Sham group was only given 10% serum of sham injured rats (sham rats were only placed in 37 centigrade warm water). After the cells were cultured for 12 hours, the release of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and creatine kinase (CK) were determined by enzyme-linked immunosorbent assay (ELISA). The cardiac troponin T (cTnT) protein expression was examined by Western Blot. Apoptosis of cardiomyocytes was observed after Hoechst 33258 staining. Compared with the sham group, the cardiomyocytes were damaged and released inflammatory cytokines after burn serum-challenged. The levels of TNF-α, IL-6 and CK increased [TNF-α (ng/L): 273±48 vs. 21±6, IL-6 (ng/L): 416±83 vs. 44±11, CK (U/L): 1.44±0.24 vs. 0.14±0.08, all P < 0.01], while the expression of cTnT protein decreased (cTnT/β-actin: 0.12±0.04 vs. 0.86±0.34, P < 0.01), and the cardiomyocyte apoptosis increased [(19.1±5.6)% vs. (5.2±1.3)%, P < 0.01]. Insulin could significantly reduce the damage of cardiomyocytes, decrease the release of TNF-α, IL-6 and CK induced by burn serum [TNF-α (ng/L): 105±37 vs. 273±48, IL-6 (ng/L): 176±77 vs. 416±83, CK (U/L): 0.82±0.26 vs. 1.44±0.24, all P < 0.05], the expression of cTnT protein significantly increased (cTnT/β-actin: 0.41±0.16 vs. 0.12±0.04, P < 0.05), and the cells apoptosis rate significantly decreased [(10.7±3.2)% vs. (19.1±5.6)%, P < 0.05]. Further blocking experiments showed that LY294002 could mitigate the protective effects of insulin. For cardiomyocytes challenged by burn serum, insulin may decrease inflammation, apoptosis and then protect the cardiomyocytes.

The Effect and Mechanism of Emodin on the NO Secretion of Human Umbilical Vein Endothelial Cells (HUVECs) Induced by High Glucose

Objective. To investigate the effects of emodin on nitric oxide (NO) secretion induced by high glucose in human umbilical vein endothelial cells (HUVECs) through the p-Akt signaling pathway. Methods. Sensitivity of cells to emodin was determined by MTT assay to establish the experimental concentrations; then, HUVECs were treated with high-dose (33.3 mmol/L) glucose (HG), HG + emodin (HG + E), HG + the Akt phosphorylation inhibitor LY294002 (HG + LY), or HG + E + LY. The p-Akt (Ser 473) expression in 48 h was analyzed using Western blot. NO effect on the secretion of HUVECs was analyzed using nitrate reductase assay. Results. The sensitive emodin concentration for HUVECs growth was 10 mol/L (P&lt;0.05). Compared with the HG group, NO secretion was significantly higher in the HG + E group (P&lt;0.05), whereas it was lowest in the HG + LY group (P&lt;0.05). Compared with the HG + LY group, NO secretion was increased in the HG + E + LY group (P&lt;0.05). The p-Akt protein expression was decreased in the HG + LY group when compared to the HG group (P&lt;0.05), while it significantly increased in the HG + E group (P&lt;0.05). Compared with HG + LY group, p-Akt protein expression was significantly higher in the HG + E + LY group (P&lt;0.05). Conclusion. Emodin could improve the NO secretion of HUVECs by high glucose through the p-Akt signaling pathway.

Effect of Methylene Blue and PI3K-Akt Pathway Inhibitors on the Neurovascular System after Chronic Cerebral Hypoperfusion in Rats.

Methylene blue (MB) has a protective effect on cognitive decline caused by chronic hypoperfusion, but the specific mechanism is not clear. This article aims to determine whether MB protects vascular neurons through PI3K/Akt and plays a role in improving cognitive impairment. Molecular biological methods, the hippocampal neuronal density test, the hippocampal vascular network density test, and dynamic enhanced magnetic resonance imaging (MRI) were used to detect the blood-brain barrier permeability and Evans blue leakage rate in the hippocampus. We also observed and evaluated the changes in the above results after administration of the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) pathway protein inhibitor LY294002. There were significant differences for cerebral blood flow (CBF) between the chronic cerebral hypoperfusion (CCH) + MB group (100ml/100g/min) and the CCH group (60ml/100g/min, P < 0.05). After using LY294002, the CBF of the CCH + MB + LY294002 group dropped to 82ml/100g/min. The vascular density in the CCH + MB group was 23%, which is significantly higher than that in the CCH group (15.1%) (P < 0.05). The vascular density (17.5%) in the CCH + MB + LY294002 group was significantly higher than that in the CCH group but lower than that in the CCH + MB group. Western blotting results showed that one week after intraperitoneal injection of MB, the expression of t-Akt and p-Akt in the CCH + MB group was increased after CCH, and LY294002 partially blocked this up-regulation effect (CCH + MB + LY294002 group). MB is a potential therapy for the relief of mild cognitive impairment associated with CCH, vascular dementia, and Alzheimer's disease.

The multidrug resistance can be reversed for the decrease of P-gp and LRP by inhibiting PI3K/Akt/NF-κB signal pathway in nasopharynx carcinoma.

Aim: To investigate the relationship between PI3K/Akt/NF-κB cellular signal pathway and the expression of P-gp and LRP in multidrug resistance (MDR) cell of nasopharyngeal carcinoma.Method: The PI3K, p-Akt and NF-κB/p65 as the activity of PI3K/Akt/NF-κB were detected by Western blot. The expressions of LRP and P-gp were detected by Western blot and real-time PCR.Result: The RIs of CNE/DDP group to DDP, 5-Fu, VCR, ADR and PTX were 35.04, 18.14, 24.13, 12.00 and 10.18, respectively. The RIs of LY-294002 group were 11.77, 5.83, 3.07, 3.86 and 3.34, and PDTC group were 11.08, 6.55, 7.66, 2.18 and 4.05. The expressions of PI3K, p-Akt and NF-κBp65, LRP and P-gp were increased and mRNA of LRP and P-gp were up-regulated in CNE/DDP. The expression of p-Akt in LY-294002 group was down-regulated. The expression of NF-κB p65 in PDTC group was decreased. The mRNA of LRP and P-gp in LY-294002 group and PDTC group were decreased.Conclusion: MDR of nasopharyngeal carcinoma cell can be regulated by activating PI3K/Akt/NF-κB signal pathway and then increase the expression of P-gp and LRP. The MDR of nasopharyngeal carcinoma cell can be reversed by inhibiting PI3K/Akt/NF-κB signal pathway.

Effect of miR-124 on PI3K/Akt signal pathway in refractory epilepsy rats

This research aimed to investigate the effect of miR-124 on the PI3K/Akt signaling pathway in refractory epilepsy rats. Ten healthy SD rats were selected as a control group, thirty-six successfully established model rats were randomly divided into the model group, mir-124-agomir, mir-124-antagomir, mir-124-agomir+ LY294002 group (combined group), with 9 rats in each group. PI3K or (and) Akt expressions were intervened respectively. The changes in attack latency and cognitive function were observed, and the correlation among miR-124, PI3K, Akt, and attack latency was analyzed. Up-regulation of the level of miR-124 could increase the seizure interval of rats (P&lt; 0.05), improve the cognitive function of rats (P&lt; 0.05), and promote the expression of PI3K, AKT. The level of miR-124, PI3K, Akt was positively correlated with the latent time of seizures in rats, and the level of miR-124 was positively correlated with the level of PI3K, Akt (P&lt; 0.05). In conclusion, miR-124 can play a protective role in temporal lobe epilepsy by promoting PI3K/Akt signaling pathway, including the protection of cognitive function, which may be a potential target for clinical treatment of intractable epilepsy in the future.