OBJECTIVE: To investigate the influence of electroacupuncture (EA) on ghrelin and the phosphoinositide 3-kinase/protein kinase B/endothelial nitric oxide synthase (PI3K/Akt/eNOS) signaling pathway in spontaneously hypertensive rats (SHRs). METHODS: Eight Wistar-Kyoto rats were used as the healthy blood pressure (BP) control (normal group), and 32 SHRs were randomized into model group, EA group, EA plus ghrelin group (EA + G group), and EA plus PF04628935 group (a potent ghrelin receptor blocker; EA + P group) using a random number table. Rats in the normal group and model group did not receive treatment, but were immobilized for 20 min per day, 5 times a week, for 4 continuous weeks. SHRs in the EA group, EA + G group and EA + P group were immobilized and given EA treatment in 20 min sessions, 5 times per week, for 4 weeks. Additionally, 1 h before EA, SHRs in the EA + G group and EA + P group were intraperitoneally injected with ghrelin or PF04628935, respectively, for 4 weeks. The tail-cuff method was used to measure BP. After the 4-week intervention, the rats were sacrificed by cervical dislocation, and pathological morphology of the abdominal aorta was observed using hematoxylin-eosin (HE) staining. Enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of ghrelin, nitric oxide (NO), endothelin-1 (ET-1) and thromboxane A2 (TXA2) in the serum. Isolated thoracic aortic ring experiment was performed to evaluate vasorelaxation. Western blot was used to measure the expression of PI3K, Akt, phosphorylated Akt (p-Akt) and eNOS proteins in the abdominal aorta. Further, quantitative real-time polymerase chain reaction (qRT-PCR) was conducted to measure the relative levels of mRNA expression for PI3K, Akt and eNOS in the abdominal aorta. RESULTS: EA significantly reduced the systolic BP (SBP) and diastolic BP (DBP) (P < 0.05). HE staining showed that EA improved the morphology of the vascular endothelium to some extent. Results of ELISA indicated that higher concentrations of ghrelin and NO, and lower concentrations of ET-1 and TXA2 were presented in the EA group (P < 0.05). The isolated thoracic aortic ring experiment demonstrated that the vasodilation capacity of the thoracic aorta increased in the EA group. Results of Western blot and qRT-PCR showed that EA increased the abundance of PI3K, p-Akt/Akt and eNOS proteins, as well as expression levels of PI3K, Akt and eNOS mRNAs (P < 0.05). In the EA + G group, SBP and DBP decreased (P < 0.05), ghrelin concentrations increased (P < 0.05), and the concentrations of ET-1 and TXA2 decreased (P < 0.05), relative to the EA group. In addition, the levels of PI3K and eNOS proteins, the p-Akt/Akt ratio, and the expression of PI3K, Akt and eNOS mRNAs increased significantly in the EA + G group (P < 0.05), while PF04628935 reversed these effects. CONCLUSION EA effectively reduced BP and protected the vascular endothelium, and these effects may be linked to promoting the release of ghrelin and activation of the PI3K/Akt/eNOS signaling pathway.
Animals ; Electroacupuncture ; Ghrelin/pharmacology* ; Nitric Oxide/metabolism* ; Nitric Oxide Synthase Type III/pharmacology* ; Phosphatidylinositol 3-Kinase/pharmacology* ; Phosphatidylinositol 3-Kinases/metabolism* ; Proto-Oncogene Proteins c-akt/pharmacology* ; Rats ; Rats, Inbred SHR ; Rats, Inbred WKY ; Signal Transduction

Objective: To investigate the therapeutic effect and mechanism of Liangge Powder against sepsis-induced acute lung injury (ALI) . Methods: From April to December 2021, the key components of Liangge Powder and its targets against sepsis-induced ALI were analyzed by network pharmacology, and to enrich for relevant signaling pathways. A total of 90 male Sprague-Dawley rats were randomly assigned to sham-operated group, sepsis-induced ALI model group (model group), Liangge Powder low, medium and high dose group, ten rats in the sham-operated group and 20 rats in each of the remaining four groups. Sepsis-induced ALI model was established by cecal ligation and puncture. Sham-operated group: gavage with 2 ml saline and no surgical treatment. Model group: surgery was performed and 2 ml saline was gavaged. Liangge Powder low, medium and high dose groups: surgery and gavage of Liangge Powder 3.9, 7.8 and 15.6 g/kg, respectively. To measure the wet/dry mass ratio of rats lung tissue and evaluate the permeability of alveolar capillary barrier. Lung tissue were stained with hematoxylin and eosin for histomorphological analysis. The levels of tumor necrosis factor-alpha (TNF-α), interleukin (IL) -6 and IL-1β in bronchoalveolar lavage fluid (BALF) were measured by enzyme-linked immunosorbent assay. The relative protein expression levels of p-phosphatidylinositol 3-kinase (PI3K), p-protein kinase B (AKT), and p-ertracellular regulated protein kinases (ERK) were detected via Western blot analysis. Results: Network pharmacology analysis indicated that 177 active compounds of Liangge Powder were selected. A total of 88 potential targets of Liangge Powder on sepsis-induced ALI were identified. 354 GO terms of Liangge Powder on sepsis-induced ALI and 108 pathways were identified using GO and KEGG analysis. PI3K/AKT signaling pathway was recognized to play an important role for Liangge Powder against sepsis-induced ALI. Compared with the sham-operated group, the lung tissue wet/dry weight ratio of rats in the model group (6.35±0.95) was increased (P<0.001). HE staining showed the destruction of normal structure of lung tissue. The levels of IL-6 [ (392.36±66.83) pg/ml], IL-1β [ (137.11±26.83) pg/ml] and TNF-α [ (238.34±59.36) pg/ml] were increased in the BALF (P<0.001, =0.001, <0.001), and the expression levels of p-PI3K, p-AKT and p-ERK1/2 proteins (1.04±0.15, 0.51±0.04, 2.31±0.41) were increased in lung tissue (P=0.002, 0.003, 0.005). The lung histopathological changes were reduced in each dose group of Liangge Powder compared with the model group. Compared with the model group, the wet/dry weight ratio of lung tissue (4.29±1.26) was reduced in the Liangge Powder medium dose group (P=0.019). TNF-α level [ (147.85±39.05) pg/ml] was reduced (P=0.022), and the relative protein expression levels of p-PI3K (0.37±0.18) and p-ERK1/2 (1.36±0.07) were reduced (P=0.008, 0.017). The wet/dry weight ratio of lung tissue (4.16±0.66) was reduced in the high-dose group (P=0.003). Levels of IL-6, IL-1β and TNF-α[ (187.98±53.28) pg/ml, (92.45±25.39) pg/ml, (129.77±55.94) pg/ml] were reduced (P=0.001, 0.027, 0.018), and relative protein expression levels of p-PI3K, p-AKT and p-ERK1/2 (0.65±0.05, 0.31±0.08, 1.30±0.12) were reduced (P=0.013, 0.018, 0.015) . Conclusion: Liangge Powder has therapeutic effects in rats with sepsis-induced ALI, and the mechanism may be related to the inhibition of ERK1/2 and PI3K/AKT pathway activation in lung tissue.
Male ; Animals ; Rats ; Rats, Sprague-Dawley ; Proto-Oncogene Proteins c-akt ; Phosphatidylinositol 3-Kinase ; Phosphatidylinositol 3-Kinases ; Powders ; Animal Experimentation ; Interleukin-6 ; MAP Kinase Signaling System ; Network Pharmacology ; Tumor Necrosis Factor-alpha ; Acute Lung Injury/drug therapy* ; Sepsis/drug therapy*

OBJECTIVETo study the molecular mechanism of curcumin in human esophageal carcinoma cell line (EC109).

METHODSEC109 cells were cultivated in vitro. When 80%-90% confluence was reached, they were treated with curcumin in different concentrations (15-120 µmol/L). The effects on cell proliferation were examined by CCK-8 colorimetry. The ultrastructure of EC109 cells were detected with transmission electron microscope(TEM). The cells apoptosis was observed with laser confocal microscope(LCM) by AnnexinV-FITC/PI double staining. The proteins level of PTEN, AKT, GSK3β and Caspase 3 were tested by flow cytometry(FCM) .

RESULTSCCK-8 test showed that curcumin could inhibit the proliferation of EC109 cells in a time- and concentration-dependent manner. TEM and LCM examinations indicated that curcumin could make EC109 cells apoptosis. The data of FCM showed that curcumin could increase the expression of PTEN, GSK3β and Caspase 3, decreased the expression of AKT.

CONCLUSIONThe effects of curcumin on inhibiting proliferation and promoting apoptosis of EC109 cells were related with increased expression of PTEN and inhibition of PI3K/AKT signaling pathway.

Apoptosis ; Caspase 3 ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; Curcumin ; pharmacology ; Esophageal Neoplasms ; metabolism ; Glycogen Synthase Kinase 3 ; metabolism ; Glycogen Synthase Kinase 3 beta ; Humans ; PTEN Phosphohydrolase ; metabolism ; Phosphatidylinositol 3-Kinases ; metabolism ; Proto-Oncogene Proteins c-akt ; metabolism ; Signal Transduction

The objective of study was to compare the influences of wortmannin on platelet aggregation and platelet membrane surface glycoproteins GPIb expression after thrombin receptor activation, and to investigate the role of phosphatidylinositol 3-kinase (PI3-K) and myosin light chain kinase (MLCK) in the course of thrombin receptor activation. Peptide SFLLRN (PAR1-AP) and AYPGKF (PAR4-AP) were used for stimulating platelet, and the changes of platelet aggregation and GPIb were analyzed with 100 nmol/L wortmannin (inhibitor of PI3-K) and 10 micromol/L wortmannin (inhibitor of MLCK). The results indicated that the platelet activation was influenced by either concentration of wortmannin in response to PAR stimulation. Platelet aggregation was apparently inhibited by 10 micromol/L wortmannin through both PAR peptides, and was slightly inhibited by 100 nmol/L wortmannin only under PAR1-AP activation. In addition, GPIbalpha internalization was partly inhibited by 100 nmol/L wortmannin in response to PAR1 (p < 0.05 at 1, 2, 5 min) and PAR4 (p < 0.05 at 2, 5, 10 min) activation. Meanwhile, 10 micromol/L wortmannin induced little change for GPIbalpha centralisation in the course of PAR activation, with a delayed restoration of surface GPIbalpha observed under PAR1-AP activation, and no change of GPIbalpha redistribution existed under PAR4-AP activation. It is concluded that the different roles of PI3-K and MLCK exist in the course of thrombin receptor activation. PI3-K accelerates the short course of GPIb centralisation for two PAR signal pathways, while MLCK inhibits the restoration of GPIbalpha in PAR1 pathway.
Adult ; Androstadienes ; pharmacology ; Female ; Humans ; Male ; Myosin-Light-Chain Kinase ; metabolism ; Phosphatidylinositol 3-Kinase ; metabolism ; Platelet Activation ; drug effects ; Platelet Aggregation ; Receptors, Thrombin ; metabolism ; physiology ; Signal Transduction

AIMTo explore the effect of Triptolide on airway remodeling and the expression of Phosphoinositide 3-Kinases in asthmatic rats.

METHODS40 rats were randomly divided into 5 groups (n = 8): (1) Control group; (2) Asthmatic 4 weeks group; (3) Asthmatic 6 weeks group; (4) Therapeutic 4 weeks group; (5) Therapeutic 6 weeks group. The airway resistance and eosinophilic inflammation of airway wall were observed. The airway wall thickness (WA/Pi), the bronchial smooth muscle thickness (smooth muscle area/Pi) and the number of bronchial smooth muscle nucleus (N/Pi) were measured by image analysis system. The expression of PI3K protein and mRNA were determined by immunohistochemical staining and reverse transcription-polymerase chain reaction (RT-PCR).

RESULTS(1) The expression of PI3K p85alpha protein and mRNA in asthmatic 4 weeks group and asthmatic 6 weeks group were significantly higher than control group, respectively (P < 0.01). The above-mentioned parameters of therapeutic 6 weeks group were significantly lower than those of asthmatic 4 weeks group, asthmatic 6 weeks group and therapeutic 4 weeks group, respectively (P < 0.01, P < 0.01 P < 0.05). (2) The WA/Pi, the smooth muscle area/Pi and the N/Pi of asthmatic 4 weeks group and asthmatic 6 weeks group were significantly higher than control group, respectively (P < 0.01). The above-mentioned parameters of therapeutic 6 weeks group were significantly lower than those of asthmatic 4 weeks group, asthmatic 6 weeks group and therapeutic 4 weeks group, respectively (P < 0.01). (3) The airway resistance of asthmatic 4 weeks group and asthmatic 6 weeks group were significantly higher than the control group, respectively (P < 0.01). The above-mentioned parameters of therapeutic 6 weeks group were significantly lower than those of asthmatic 4 weeks group, asthmatic 6 weeks group and therapeutic 4 weeks group, respectively (P < 0.01, P < 0.01, P < 0.05).

CONCLUSIONThe proliferation of airway smooth muscle is a remarkable character of airway remodeling in asthma. The PI3K signal pathway may be involved in the process. Triptolide may reduce AHR and decrease the proliferation of ASMCs by inhibiting the expression of PI3K. It may have potential therapeutic effects in the asthmatic airway remodeling.

Airway Remodeling ; Animals ; Asthma ; metabolism ; physiopathology ; Diterpenes ; pharmacology ; Epoxy Compounds ; pharmacology ; Male ; Phenanthrenes ; pharmacology ; Phosphatidylinositol 3-Kinase ; metabolism ; Rats ; Rats, Sprague-Dawley ; Signal Transduction

OBJECTIVESTo explore the effects of insulin on the expression and the regulatory pathway of AQP9 in normal human liver cells.

METHODSNormal human liver cells L02 were cultured and treated with PI3K inhibitor LY294002, AKT inhibitor A-443654, MAPK inhibitors SB2030580 and insulin at different concentrations respectively. The AQP9 mRNA and protein expressions were detected with semi-quantitative RT-PCR and Western blot respectively.

RESULTSThe insulin (100 nmol/L approximately 500 nmol/L) treatment decreased the expression of AQP9 in normal human liver cells (P less than 0.05) concentration dependently, and the expression of AQP9 began to reduce from 3 hours of insulin stimulation (P less than 0.05), especially at insulin treatment for 12 hours (P less than 0.05); Incubated with the selective inhibitor of PI3K (LY294002) and AKT (A-443654), the inhibitory effects of insulin on AQP9 expression decreased (P less than 0.05); but it did not change significantly by blocking the MAPK signaling pathway.

CONCLUSIONThe insulin treatment inhibited the expression of AQP9 and the PI3K/akt signal transduction pathway was involved in the mechanism.

Aquaporins ; metabolism ; Cells, Cultured ; Chromones ; pharmacology ; Hepatocytes ; drug effects ; metabolism ; Humans ; Insulin ; pharmacology ; Morpholines ; pharmacology ; Phosphatidylinositol 3-Kinase ; antagonists & inhibitors ; metabolism ; Protein-Serine-Threonine Kinases ; metabolism ; Signal Transduction

OBJECTIVE: Moxibustion, a common therapy in traditional Chinese medicine, has potential benefits for treating decreased ovarian reserve (DOR). The present study investigates the protective effect of moxibustion in a rat model of DOR and explores the possible mechanisms. METHODS: Sixty-four female Sprague-Dawley rats were randomly divided into four groups: control, DOR, moxibustion (MOX), and hormone replacement therapy (HRT). The DOR rat model was established by intragastric administration of 50 mg/kg Tripterygium glycoside suspension (TGS), once daily for 14 days. MOX and HRT treatments were given from the day TGS administration was initiated. The ovarian reserve function was evaluated by monitoring the estrus cycle, morphological changes in ovaries, levels of serum estradiol (E₂), follicle-stimulating hormone (FSH), luteinizing hormone (LH), and anti-Mullerian hormone (AMH), pregnancy rate and embryo numbers. Terminal-deoxynucleotidyl transferase-mediated nick-end-labeling staining was used to identify ovarian granulosa cell apoptosis, while the protein and mRNA expressions of Bax, B-cell lymphoma-2 (Bcl-2), phosphatidylinositol 3-kinase (PI3K) and protein kinase B (AKT) in ovarian tissues were examined by immunohistochemistry, Western blot and quantitative reverse transcription-polymerase chain reaction. RESULTS: Compared with the DOR group, MOX improved the disordered estrous cycle, promoted follicular growth, reduced the number of atresia follicles, increased the concentrations of serum E₂ and AMH, and decreased serum FSH and LH concentrations. More importantly, the pregnancy rate and embryo numbers in DOR rats were both upregulated in the MOX treatment group, compared to the untreated DOR model. Further, we found that the MOX group had reduced apoptosis of ovarian granulosa cells, increased Bcl-2 expression and reduced expression of Bax. Furthermore, the PI3K/AKT signaling pathway was triggered by the moxibustion treatment. CONCLUSION Moxibustion improved ovarian function and suppressed apoptosis of ovarian granulosa cells in a rat model of DOR induced by TGS, and the mechanism may involve the PI3K/AKT signaling pathway.
Animals ; Female ; Follicle Stimulating Hormone ; Luteinizing Hormone ; Moxibustion ; Ovarian Reserve ; Phosphatidylinositol 3-Kinase/pharmacology* ; Phosphatidylinositol 3-Kinases/metabolism* ; Pregnancy ; Proto-Oncogene Proteins c-akt/pharmacology* ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; bcl-2-Associated X Protein/genetics*

BACKGROUND AND OBJECTIVEInvasion and metastasis are the primary causes of death in patients with pulmonary carcinoma. The epidermal growth factor (EGF) stimulates A549 cells invasion greatly through activating ERK and PI3K-Akt signaling pathway. The aim of this study is to elucidate the inhibitory effect of Resveratrol on EGF-induced invasive ability of A549 cells in vitro and explore the molecular mechanism.

METHODSThe cytotoxicity of Resveratrol was evaluated by methyl thiazolyltetrazolium (MTT) assay. Then, the A549 cells were treated with EGF and non-cytotoxic concentration of Resveratrol. The cells' invasion were detected by Boyden chamber assay; MMP-2 activity was determined by gelatine zymography assay; the changes of the related proteins were detected by Western blot.

RESULTSResveratrol was not toxic to A549 cells at the concentration between 0 to 30 microM. The invasion ability of EGF-induced A549 cells was decreased after treatment with 20 microM resveratrol for 24 h, accompanied by the inhibition of MMP-2 secretion. And the levels of p-ERK1/2, PI3K (within 6 h) were suppressed too.

CONCLUSION20 microM Resveratrol inhibits A549 cells' invasion possibly through the suppression of the activation of ERK and PI3K-Akt signaling pathways, subsequently exerting inhibitory effect on MMP-2.

Adenocarcinoma ; metabolism ; Anticarcinogenic Agents ; pharmacology ; Blotting, Western ; Cell Line, Tumor ; Epidermal Growth Factor ; pharmacology ; Humans ; Lung Neoplasms ; metabolism ; Mitogen-Activated Protein Kinase 1 ; metabolism ; Mitogen-Activated Protein Kinase 3 ; metabolism ; Phosphatidylinositol 3-Kinases ; metabolism ; Signal Transduction ; drug effects ; Stilbenes ; pharmacology

CAL-101 is a selective inhibitor of the phosphatidylinositol-3 kinase (PI3K), it inhibits the survival, proliferation and migration of tumor cells by directly inducing apoptosis and inhibiting micro-environmental interactions. It has been determined that the P110δ isoforms of PI3K expressed primarily in cells of hematopoietic lineage, such as B and T cells. This review focuses on the target, mechanism of action, the use and prospect of CAL-101 in tumors of blood and lymph systems.
Animals ; Class Ia Phosphatidylinositol 3-Kinase ; antagonists & inhibitors ; Hematologic Neoplasms ; drug therapy ; Humans ; Purines ; pharmacology ; therapeutic use ; Quinazolinones ; pharmacology ; therapeutic use ; Signal Transduction ; drug effects

OBJECTIVETo investigate the effect of Jiangtang Yishen Recipe (JTYSR) on high insulin induced cell proliferation of human glomerular mesangial cells (HMCs) and the expression of insulin receptor substrate 1 (IRS-1) and phosphatidylinositol-3-kinase (PI-3K).

METHODSHMCs were divided into 4 groups, i.e., the negative control group, the high insulin model group, the JTYSR group, and the LY294002 group. The concentration of insulin, JTYSR, and LY294002 was respectively confirmed by pre-experiment. Different culture solution was respectively added for different groups. RPMI1640 culture solution was added for HMCs in the negative control group, while HMCs in the rest 3 groups were cultured by 100 nmol/L insulin for 24 h. Meanwhile, HMCs from the JTYSR group and the LY294002 group were exposed to 125 mg/L JTYSR and 80 micromol/L LY294002 respectively for further 48 h. The proliferation of HMCs was detected by MTT and flow cytometry. The protein expression of IRS-1 and PI-3K in HMC was detected by immunohistochemical assay and Western blot. Results The proliferation of HMCs induced by high insulin could be significantly lowered, and the protein expression of IRS-1 and PI-3K could be down-regulated in the JTYSR group and the LY294002 group (P <0.01). Compared with the LY294002 group, the protein expression of IRS-1 and PI-3K could be slightly down-regulated in the JTYSR group (P <0.05).

CONCLUSIONJTYSR could lower high insulin induced proliferation of HMCs, and its mechanism might be related to insulin signaling pathway.

Cell Proliferation ; drug effects ; Chromones ; Drugs, Chinese Herbal ; pharmacology ; Humans ; Insulin Receptor Substrate Proteins ; metabolism ; Mesangial Cells ; physiology ; Morpholines ; Phosphatidylinositol 3-Kinase ; metabolism ; Phosphatidylinositol 3-Kinases ; metabolism ; Signal Transduction