Mechanism of Shenshuai Xiezhuo Decoction in Intervention of Chronic Kidney Disease Combined with Atherosclerosis Based on Regulation of PI3K/Akt/mTOR Signaling Pathway by miRNA126 / 中国实验方剂学杂志

ObjectiveExploring the role of microRNA126 （miRNA126） in chronic kidney disease combined with atherosclerosis （CKD AS） by regulating the phosphatidylinositol-3-kinase （PI3K）/protein kinase B （Akt）/mammalian target of rapamycin （mTOR） signaling pathway and the mechanism of Shenshuai Xiezhuo decoction in the intervention of CKD AS rats with 5/6 nephrectomy combined with high-fat feeding. MethodA total of 60 SD rats were randomly divided into sham operation group， model group， losartan group， and low， medium， and high dose groups of Shenshuai Xiezhuo decoction. The CKD AS rat model was established by 5/6 nephrectomy combined with high-fat feeding for 10 weeks. The low， medium， and high dose groups （6.0， 12.0， 24.0 g·kg-1·d-1） of Shenshuai Xiezhuo decoction and the losartan group （20 mg·kg-1·d-1） were gavaged， and the corresponding intervention was carried out for eight weeks. Then， the rats were killed， and samples were collected for corresponding detection. Fully automated biochemical analyzers were used to detect kidney function and blood lipids in rats： blood creatinine （SCr）， blood urea nitrogen （BUN）， total cholesterol （TC）， triglyceride （TG）， and low-density lipoprotein cholesterol （LDL-C） levels. Hematoxylin-eosin （HE） and Masson staining of aortic tissue and pathological observation under a light microscope were carried out， and autophagosomes and autophagy lysosomes were observed by transmission electron microscopy. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was used to determine the mRNA levels of miRNA126， PI3K， Akt， and mTOR in rats， and Western blot was used to determine the protein expression levels of phosphorylated （p）-PI3K， PI3K， p-Akt， Akt， p -mTOR， mTOR， benzyl chloride 1 （Beclin-1）， and microtubule-associated protein light chain 3Ⅱ/Ⅰ （LC3Ⅱ/LC3Ⅰ）. ResultCompared with the sham operation group， the serum SCr， BUN， TC， TG， and LDL-C in the model group were significantly increased （P<0.01）. Compared with the model group， the SCr， BUN， TC， TG， and LDL-C were decreased in the losartan group and low， medium， and high dose groups of Shenshuai Xiezhuo decoction （P<0.05）. Compared with the sham operation group， thickening plaques， infiltration of mononuclear macrophages， a small number of foam cells， disordered arrangement of smooth muscle fibers in the tunica media， and increased collagen fibers were observed in the model group， and the lesions in the losartan group and Shenshuai Xiezhuo decoction groups were alleviated compared with those in the model group. Compared with the model group， the number of autophagosomes and autophagy lysosomes increased in the medium and high dose groups of Shenshuai Xiezhuo decoction. Compared with the sham operation group， the expression of miRNA126 in the aortic tissue of the model group was significantly decreased （P<0.01）， and the mRNA expressions of PI3K， Akt， and mTOR were significantly increased （P<0.01）. Compared with the model group， the expression of miRNA126 in the aortic tissue of rats in high， medium， and low dose groups of Shenshuai Xiezhuo decoction and losartan group was significantly increased （P<0.01）， while the mRNA expressions of PI3K， Akt， and mTOR were significantly decreased （P<0.01）. Compared with the sham operation group， the protein expressions of p-PI3K， PI3K， p-Akt， Akt， p-mTOR， and mTOR in the model group were significantly increased （P<0.01）， while the protein levels of Beclin-1， LC3Ⅰ， and LC3Ⅱ were significantly decreased （P<0.01）. Compared with the model group， the protein expressions of p-PI3K， PI3K， p-Akt， Akt， p-mTOR， and mTOR in the losartan group and low， medium， and high dose groups of Shenshuai Xiezhuo decoction were decreased （P<0.05）， while the protein levels of Beclin-1 and LC3Ⅱ/LC3Ⅰ were increased （P<0.05）. ConclusionThe expression of miRNA126 is decreased in the aortic tissue of CKD AS rats， and the PI3K/Akt/mTOR pathway is activated to inhibit autophagy flux. Shenshuai Xiezhuo decoction regulates the PI3K/Akt/mTOR signaling pathway through miRNA126， restores the autophagy of aortic endothelial cells， protects the damage of CKD vessels， reduces the formation of As plaques， and slows the development of cardiovascular complications.

Jiedu Huoxue Prescription Affects Plaque Stability in ApoE-/- Atherosclerosis Mice by Modulating PI3K/Akt/mTOR Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo investigate whether Jiedu Huoxue prescription can induce macrophage autophagy and inhibit inflammatory response to stabilize vulnerable plaques of atherosclerosis （AS） by regulating phosphatidylinositol 3-kinase （PI3K）/protein kinase B （Akt）/mammalian target of rapamycin （mTOR） signaling pathway. MethodThirty ApoE-/- mice fed with high-fat diet were randomly assigned into model， low-， medium-， and high-dose （5.35， 10.7， and 21.4 g·kg-1·d-1， respectively） Jiedu Huoxue prescription （Chinese medicine）， and rapamycin （2 mg·kg-1·d-1） groups. Six ApoE-/- mice fed with common diet were used as the control group， and 6 C57BL/6J mice fed with common diet as the blank group. The drugs or equal volume of normal saline were administrated by gavage after 7 weeks of modeling， and the treatment lasted for 4 weeks. The serum levels of lipids and inflammatory cytokines ［monocyte chemoattractant protein-1 （MCP-1） and interleukin-6 （IL-6）］ were measured. Hematoxylin-eosin （HE） staining was employed to observe the pathological changes of the vascular wall of the aortic root. Immunohistochemistry was employed to detect the expression of macrophages/monocytes monoclonal antibody （MOMA-2） and α-smooth muscle actin （α-SMA）. Transmission electron microscopy was employed to count the autophagosomes in the aorta， and Western blot to determine the protein levels of Beclin-1， LC3， PI3K， Akt， and mTOR. ResultCompared with the control group， the model group showed elevated serum levels of lipids， MCP-1， and IL-6 （P<0.05）， inhibited expression of MOMA-2 and α-SMA （P<0.05， P<0.01）， up-regulated protein level of Beclin-1 （P<0.05）， and down-regulated protein levels of PI3K， Akt， and mTOR （P<0.05， P<0.01）. The model group presented obvious atherosclerotic plaques on the inner wall of the aorta， infiltration of inflammatory cells in the plaque， thickened and disarranged vascular intima where the plaque was attached， decreased autophagosomes and mitochondria， and destroyed mitochondrial structure. Chinese medicine and rapamycin groups showed lower levels of total cholesterol， triglycerides， low-density lipoprotein cholesterol， MCP-1， and IL-6 （P<0.05）， higher level of high-density lipoprotein cholesterol （P<0.05）， inhibited expression of MOMA-2 and α-SMA （P<0.05， P<0.01）， higher protein levels of Beclin-1 and LC3Ⅱ （P<0.05， P<0.01）， and lower protein levels of PI3K， Akt， and mTOR （P<0.05， P<0.01） than the model group. Moreover， Chinese medicine and rapamycin groups showed only a small number of atherosclerotic plaques on the inner wall of the aorta， reduced infiltration of inflammatory cells and thickness of the blood vessel wall， and increased autophagosomes and autophagic lysosomes. ConclusionJiedu Huoxue prescription can improve lipid metabolism， enhance macrophage autophagy， and reduce AS-induced inflammation to improve the stability of vulnerable plaques in AS mice by inhibiting the PI3K/Akt/mTOR signaling pathway.

Danggui Sinitang Mitigates Gouty Arthritis in Rats by Regulating Autophagy via PI3K/Akt/mTOR Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo study the mechanism of Danggui Sinitang in mitigating gouty arthritis （GA） in rats by regulating autophagy via the phosphatidylinositol 3-kinase （PI3K）/protein kinase B （Akt）/mammalian target of rapamycin （mTOR） signaling pathway. MethodSixty male SD rats were randomly assigned into normal， model， colchicine （0.3 mg·kg-1）， and low-， medium-， and high-dose Danggui Sinitang （6.54， 13.08， and 26.16 g·kg-1） groups （n=10） and administrated with corresponding drugs by gavage. The rats in the normal group and model group were administrated with equal volume of normal saline by gavage for 7 days. One hour after administration on day 5， the GA model was established by injecting sodium urate suspension （50 g·L-1） into the right ankle joint of rats in other groups except the normal group， and the rats in the normal group were injected with sterile normal saline of the same volume. The swelling and pathological changes of the ankle joint were observed. The serum levels of tumor necrosis factor-α （TNF-α）， interleukin-6 （IL-6）， and IL-1β were determined. Western blot was employed to determine the protein levels of PI3K， phosphorylated PI3K （p-PI3K）， protein kinase B （Akt）， phosphorylated Akt （p-Akt）， mTOR， phosphorylated mTOR （p-mTOR）， microtubule-associated protein 1 light chain 3 Ⅱ/Ⅰ （LC3Ⅱ/Ⅰ）， autophagy effector Beclin-1， and ubiquitin-binding protein p62 in the synovial tissue. Real-time fluorescent quantitative PCR （Real-time PCR） was employed to determine the mRNA levels of PI3K， Akt， mTOR， LC3， Beclin-1 and p62. ResultCompared with the normal control， the model group showed increased joint swelling index （P<0.01）， elevated serum levels of TNF-α， IL-6， and IL-1β， inflammatory cell infiltration， and fibrous tissue hyperplasia. In addition， the model group showed up-regulated protein levels of PI3K， p-PI3K， Akt， p-Akt， mTOR， p-mTOR， and p62 and mRNA levels of PI3K， Akt， mTOR， and p62 in the synovial tissue， while it showed down-regulated protein levels of LC3Ⅱ/Ⅰ and Beclin-1 and mRNA levels of LC3 and Beclin-1 （P<0.01）. Compared with the model group， medium- and high-dose Danggui Sinitang alleviated the joint swelling （P<0.01）， lowered the serum levels of TNF-α， IL-6， and IL-1β （P<0.05）， and relieved the inflammatory cell infiltration in the synovial tissue of the ankle joint and the fibrous tissue hyperplasia. Moreover， they down-regulated the protein levels of PI3K， p-PI3K， Akt， p-Akt， mTOR， p-mTOR， and p62 and the mRNA levels of PI3K， Akt， mTOR， and p62 in the synovial tissue （P<0.05）， while they up-regulated the protein levels of LC3Ⅱ/Ⅰ and Beclin-1 and the mRNA levels of LC3 and Beclin-1 （P<0.05）. ConclusionDanggui Sinitang， especially at a high dose， can inhibit PI3K/Akt/mTOR signaling pathway to improve autophagy in the synovial tissue， thereby mitigating GA.

Mechanism of PI3K/Akt/mTOR Signaling Pathway Against Non-small Cell Lung Cancer and Chinese Medicine Intervention： A Review / 中国实验方剂学杂志

Non-small cell lung cancer （NSCLC） is a malignant tumor of the respiratory system with a high incidence. The early symptoms are not typical， and most patients are diagnosed at an advanced stage， which seriously threatens the lives and health of people. Surgery， chemotherapy， and targeted therapy are the main means of treatment at present， but the consequent drug resistance and adverse reactions restrict these treatment methods with certain limitations. In recent years， with the development of traditional Chinese medicine （TCM） in tumor resistance， TCM has attracted extensive attention for its obvious therapeutic effect and fewer adverse reactions. Numerous signaling pathways are involved in the formation and development of NSCLC， where phosphatidylinositol 3 kinase （PI3K）/protein kinase B （Akt）/mammalian target of rapamycin （mTOR） signaling pathway is one of the key regulatory pathways. The PI3K/Akt/mTOR signaling pathway affects the proliferation， invasion， and metastasis of NSCLC cells by affecting the cycle， inhibiting the apoptosis， inhibiting the autophagy of tumor cells， and promoting tumor neovascularization. As revealed by the recent literature， Chinese medicine plays an indispensable role in NSCLC cell autophagy， cell cycle， apoptosis， invasion and metastasis， neovascularization， and reversal of drug resistance by regulating the PI3K/Akt/mTOR signaling pathway. Although some Chinese medicinal extracts or compounds have made great breakthroughs in some mechanisms of action in the treatment of NSCLC， these studies only remain at the level of in vitro cell experiments and animal studies in vivo. Researchers are faced with the great challenge of "transforming the research results of Chinese medicines into clinical applications". Based on the current research status in China and abroad， this paper reviewed Chinese medicine in the intervention in NSCLC through the regulation of PI3K/Akt/mTOR signaling pathway in recent years， in order to open up new ideas for NSCLC drug therapy research and also provide a useful reference for further mechanism research.

Mechanism of Scutellariae Radix-Paeoniae Radix Rubra Combination of Different Proportions Against Hepatic Fibrosis in Rats / 中国实验方剂学杂志

ObjectiveTo observe the effects of Scutellariae Radix （SR）-Paeoniae Radix Rubra （PRR） combination of different proportions on the expression of the Toll-like receptor 4 （TLR4）/myeloid differentiation factor 88 （MyD88）/nuclear transcription factor κB （NF-κB） and phosphatidylinositol kinase （PI3K）/protein kinase B （Akt）/mammalian target of rapamycin （mTOR） signaling pathways in liver tissues of rats with hepatic fibrosis and explore the mechanism against hepatic fibrosis. MethodSixty male SD rats of SPF grade were randomly divided into a normal group， a model group， a positive control （silymarin） group， and SR-PRR 1∶1， SR-PRR 1∶2， and SR-PRR 1∶4 groups， with 10 rats in each group. The hepatic fibrosis model was induced in rats except for those in the normal group by intraperitoneal injection of 40% tetrachloromethane （CCl4）-olive oil solution at 3 mL·kg-1， 5 mL·kg-1 for the first time， for 8 weeks， twice per week. After 4 weeks， rats were treated correspondingly at 10 mL·kg-1 by intragastric administration， and the body weight of rats in each group was weighed for 8 weeks. After administration， histopathological changes in the liver were observed. Serum alanine aminotransferase （ALT）， aspartate aminotransferase （AST）， hyaluronic acid （HA）， laminin （LN）， albumin （ALB）， alkaline phosphatase （AKP）， and superoxide dismutase （SOD） activities， malondialdehyde （MDA）， and hydroxyproline （HYP） content in liver tissues were detected. The mRNA expression levels of TLR4， MyD88， NF-κB， PI3K， Akt， and mTOR in the liver of rats were detected by real-time fluorescence-based quantitative polymerase chain reaction （Real-time PCR）. ResultCompared with the model group， SR-PRR combination of different proportions could recover the body weight and improve the pathological injury of the liver. As revealed by enzyme linked immunosorbent assay （ELISA） results， compared with the normal group， the model group showed increased ALT， AST， HA， LN， AKP， MDA， and HYP levels to different degrees （P<0.05）. Compared with the model group， the groups with drug intervention showed decreased levels of ALT， AST， HA， LN， AKP， MDA， and HYP， potentiated SOD activity， and increased level of ALB （P<0.05）. As revealed by Real-time PCR results， compared with the normal group， the model group showed increased mRNA expression of TLR4， MyD88， NF-κB， PI3K， Akt， and mTOR （P<0.05）. Compared with the model group， the groups with drug intervention showed reduced mRNA expression of TLR4， MyD88， NF-κB， PI3K， Akt， and mTOR in the liver of rats （P<0.05）. ConclusionSR-PRR combination of different proportions can improve the histopathological injury in liver tissues caused by CCl4， with the optimal effect observed in the SR-PRR 1∶4 group. SR-PRR may inhibit the development of liver fibrosis by inhibiting the expression of TLR4/MyD88/NF-κB and PI3K/Akt/mTOR signaling pathways， thereby alleviating chemical-induced liver injury.

Effect of Bushen Tongluo Prescription on Synaptic Plasticity of Hippocampal Neurons of Vascular Dementia Model Rats Based on PI3K/Akt/mTOR Pathway / 中国实验方剂学杂志

Objective:To explore the effect of Bushen Tongluo prescription （BSTLP） on the synaptic plasticity of hippocampal neurons in vascular dementia （VD） model rats and its mechanism. Method:SD male rats of SPF grade were selected. The rat model of VD was established by permanent bilateral ligation of the common carotid artery several times. The model rats were randomly divided into a model group， an insulin-like growth factor-1 （IGF-1， 20 μg·kg^-1） group， high-dose （3 g·kg^-1）， medium-dose （1.5 g·kg^-1）， and low-dose （0.75 g·kg^-1） BSTLP groups. A sham operation group was also set. Drugs were administered to rats by gavage once a day for four weeks. The model group and the sham operation group received the same volume of normal saline. After the last administration， all the rats were detected for spatial learning and memory by the Morris water maze. The apoptosis of hippocampal neurons was detected by terminal deoxynucleotidyl transferase （TdT） dUTP nick-end labeling （TUNEL） assay. The changes in synaptic morphological structure and the number of dendritic spines in hippocampal neurons were detected by Golgi's method. The expression levels of phosphatidylinositol 3-kinase （PI3K）， protein kinase B （Akt）， mammalian target of rapamycin （mTOR）， synaptophysin （SYP）， and amyloid precursor protein （APP） in hippocampal neurons were detected by Western blot. Result:Compared with the sham operation group， the model group showed prolonged escape latency， lengthened swimming distance， dwindled the number of times for the platform crossing after platform removal （<italic>P</italic><0.05）， increased apoptotic cells （<italic>P</italic><0.05）， declining synaptic dendritic spines （<italic>P</italic><0.05）， down-regulated expression levels of PI3K， Akt， mTOR， and SYP proteins， and up-regulated expression level of APP protein in hippocampal neurons （<italic>P</italic><0.05）. Compared with the model group， the BSTLP groups and the IGF-1 group showed shortened escape latency and swimming distance， increased number of times for the platform crossing after platform removal （<italic>P</italic><0.05），declining apoptotic cells （<italic>P</italic><0.05）， up-regulated expression levels of PI3K， Akt， mTOR， and SYP proteins， and down-regulated expression level of APP protein in hippocampal neurons （<italic>P</italic><0.05）. Compared with the IGF-1 group， the high-dose BSTLP group showed no significant difference in the escape latency， swimming distance， the number of times for the platform crossing after platform removal， apoptotic cells， synaptic dendritic spines， and expression levels of PI3K， Akt， mTOR， SYP， and APP proteins in hippocampal neurons. However， the differences were significant in the medium-dose and low-dose BSTLP groups （<italic>P</italic><0.05）. Conclusion:BSTLP can improve the learning and memory of rats with VD. The mechanism is presumedly related to the activation of thePI3K/Akt/mTOR pathway and improvement of synaptic plasticity of hippocampal neurons.

Effect of Elian Granule on Autophagy and PI3K/Akt/mTOR Signaling Pathway in Gastric Tissue of Rats with Gastric Cancer / 中国实验方剂学杂志

Objective:To investigate the effect of Elian granule on autophagy and the phosphatidylinositol -3 kinase （PI3K）/protein kinase B （PKB/Akt）/mammalian target of rapamycin （mTOR） signaling pathway in gastric tissue of rats with gastric cancer. Method:SPF SD rats were randomly divided into the normal， model， Elian granule， and Weifuchun groups. In addition to the routine feeding in the normal group， the model， Elian granule， and Weifuchun groups received <italic>N</italic>-methyl-<italic>N</italic>'-nitro-<italic>N</italic>-nitrosoguanidine （MNNG） to induce gastric cancer in rats， and they were respectively given normal saline， Elian granule aqueous solution （3.240 g·kg^-1） and Weifuchun aqueous solution （0.390 g·kg^-1） by gavage （<italic>ig</italic>） for 48 weeks. The gross changes of the stomach taken by laparotomy were observed by naked eyes. Hematoxylin-eosin （HE） staining was performed to observe the histopathological changes of the gastric tissue in rats. Real-time quantitative polymerase chain reaction （Real-time PCR） and Western blot （WB） were used to detect the mRNA and protein expression of microtubule-associated protein 1 light chain 3 beta （LC3B）， Beclin1， p62， PI3K， Akt， mTOR in rat gastric tissue. Result:Compared with the normal group， the model group showed gastric distension， thinner gastric wall， pale gastric mucosa， atrophied and flat folds， disordered course， and visible nodules and vegetations. Compared with the model group， the Elian granule group demonstrated alleviated gastric distension， dark gastric mucosa， reduced folds， and regular course， with the thinned gastric wall improved and granular nodules observed occasionally. According to HE staining， compared with the normal group， the model group showed crowded and disordered rat gastric glands， diverse in shape， varied cell morphology， basophilic cytoplasm， large irregular hyperchromatic nuclei， visible mitosis， and infiltrated and destroyed muscularis mucosae. While compared with the model group， the arrangement of gastric glands was regular， and a few mildly atypical cells could be observed in rats of the Elian granule group. Compared with the normal group， the model group exhibited decreased expression of LC3B and Beclin1 mRNA and protein in gastric tissue （<italic>P</italic><0.05）， and increased expression of PI3K， p62， Akt， and mTOR mRNA and protein （<italic>P</italic><0.05）. Compared with the model group， the Elian granule group showed increased expression of LC3B and Beclin1 mRNA and protein in gastric tissue （<italic>P<</italic>0.05）， and decreased expression of PI3K mRNA and p62， Akt， and mTOR mRNA and protein （<italic>P</italic><0.05）. Conclusion:Elian granule can improve the cell atypia of gastric tissue in rats with gastric cancer， and the mechanism may be related to the inhibition of the PI3K/Akt/mTOR signaling pathway to promote autophagy.

Effect of Qigesan on Proliferation， Apoptosis and miR-133a/Akt/mTOR Signaling Pathway of Esophageal Cancer EC9706 Cells / 中国实验方剂学杂志

Objective:To observe the effect of Qigesan on the proliferation and apoptosis of the human esophageal cancer cell EC9706， and the effect on miR-133a/protein kinase B（Akt）/mammalian target of rapamycin （mTOR） signaling pathway. Method:The effective constituent of Qigesan was extracted by ethyl acetate. Thiazolyl blue tetrazolium bromide（MTT） colorimetric assay was used to determine the dosage of Qigesan on cells and to detect the effect of Qigesan on the proliferation of EC9706 cells. The effect of Qigesan on apoptosis of EC9706 cells was detected by flow cytometry. The effect of Qigesan on miR-133a and insulin-like growth factor 1 receptor（IGF-1R） mRNA expression was detected by Real-time quantitative polymerase chain reaction （Real-time PCR） . The protein expression of Akt and mTOR in EC9706 cells was detected by Western blot. Result:Qigesan can inhibit the proliferation of EC9706 cells in a dose-dependent manner（<italic>P</italic><0.01）. Inhibitory concentrations 30% inhibition concentration（IC₃₀） 40 mg·L^-1 and median inhibition concentration（IC₅₀） 80 mg·L^-1 were selected for follow-up experiments. Compared with the blank group， both the inhibitor group and the combination drug group can inhibit the proliferation of EC9706 cells （<italic>P</italic><0.01）. The inhibitor at 0.25 μmol·L^-1 was selected for subsequent experiments. Compared with the blank group， Qigesan 80 mg·L^-1 dose group could significantly promote the late apoptosis rate and total apoptosis rate of EC9706 cells（<italic>P</italic><0.05）， and the 40 mg·L^-1 dose group could significantly promote the late apoptosis rate of EC9706 cells（<italic>P</italic><0.05）， which shows synergistic effect after concomitant use with Akt/mTOR inhibitor（<italic>P</italic><0.05）. Compared with the blank control group， each group can effectively increase expression of miR-133a（<italic>P</italic><0.05）. The combination of inhibitor and traditional Chinese medicine（TCM） has obvious promotion effect. Compared with blank control group， the expressions of Akt and mTOR were significantly decreased in each group（<italic>P</italic><0.05）. Compared with single medication， the expressions of Akt and mTOR were decreased in combination of inhibitor and TCM group. Conclusion:Qigesan can inhibit the growth of EC9706 cells and promote apoptosis， and its inhibitory mechanism may be related to the Akt/mTOR signaling pathway by regulating the expression of miR-133a.

Effect of Jianpi Xiao' ai Prescription on Akt/mTOR Signaling Pathways in Colorectal Cells / 中国实验方剂学杂志

Objective::To investigate the effect of drug-containing serum of Jianpi Xiaoai prescription on protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathways in colorectal cells HCT116. Method::The HTC116 cells were treated by 15%concentration of drug-contained serum, and then the cell migration and invasion were detected by Transwell assay, the protein expression levels of Akt, phosphorylated protein kinase B (p-Akt), mTOR, phosphorylated mammalian target of rapamycin (p-mTOR), ribosomal protein S6 kinase, polypeptide1(S6K1), phosphorylated ribosomal protein S6 kinase, polypeptide1 (p-S6K1), 4E-binding protein1(4EBP1), and phosphorylated 4E-binding protein1(p-4EBP1) in HCT116 cells were detected by Western blot. The control group was treated by untreated serum (15%), and 10%fetal bovine serum(FBS). Result::As compared with the control group, the number of migration and invasion cells was significantly reduced in drug-contained serum group (P<0.01), the expression of Akt had no obvious decrease, p-Akt protein expression was significantly lowered in the drug-contained serum group (P<0.01), the expression of mTOR had no obvious decrease, but p-mTOR protein expression was significantly lowered in drug-contained serum group (P<0.01), the expression of S6K1 had no obvious decrease, but p-S6K1 protein expression was significantly lowered in the drug-contained serum group (P<0.01), the protein expression of 4EBP1 had no obvious decrease, but p-4EBP1 protein expression was significantly lowered in the drug-contained serum group (P<0.01). Conclusion::The anti-tumor mechanism and transfer of Jianpi Xiaoai prescription may be related to inhibiting the activation of Akt/mTOR signaling pathways in colorectal cancer.

MicroRNA-133a alleviates airway remodeling in asthtama through PI3K/AKT/mTOR signaling pathway by targeting IGF1R.

Asthma is characterized by chronic inflammation, and long-term chronic inflammation leads to airway remodeling. But the potential regulatory mechanism of airway remodeling is not clearly understood, and there is still no effective way to prevent airway remodeling. Present studies have confirmed the role of microRNAs (miRNAs) in the development of disease, which is known as suppressing translation or degradation of messenger RNA (mRNA) at the posttranscriptional stage. In this study, we described the role of miRNA-133a in asthma and demonstrated it in regulating airway remodeling of asthma through the phosphoinositide 3 kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathway by targeting IGF-1 receptor (IGF1R). IGF1R helps in mediating the intracellular signaling cascades. Asthmatic mice models were established by sensitization and Ovalbumin challenge. Adenovirus transfer vector carrying miR-133a or miR-133a sponge sequence was used to build the overexpression or downexpression of miR-133a modeling. Real-time polymerase chain reaction and Western blot were used to determine the alterations in the expression of miR-133a and mRNAs and their corresponding proteins. Results showed that miR-133a was downregulated in asthma. Upregulation of miR-133a expression in airway smooth muscle cells in vivo and in vitro could inhibit the activation of PI3K/AKT/mTOR pathway, and reduce the expression of α-smooth muscle actin (α-SMA), indicating that airway remodeling was inhibited. Functional studies based on luciferase reporter revealed miR-133a as a direct target of IGF1R mRNA. In conclusion, these data suggested that miR-133a regulated the expression of α-SMA through PI3K/AKT/mTOR signaling by targeting IGF1R. miR-133a plays a key role in airway remodeling of asthma and may serve as a potential therapeutic target for managing asthmatic airway remodeling.

Anticancer Effect of Isoliquiritigenin on Human Clear Cell Renal Cell Carcinoma 786-O Cells and Molecular Mechanism / 中国实验方剂学杂志

Objective: To investigate the anticancer effect of isoliquiritigenin (ISL) on human clear cell renal cell carcinoma 786-O cells, and explore its possible molecular mechanism. Method: Thiazolyl blue tetrazolium bromide (MTT) assay was used to detect effect of ISL (0, 10, 25,50, 75, 100 μmol·L-1) on proliferation of 786-O cells. The effect of ISL on migration and invasion of 786-O cells was detected by cell scratch test and Transwell assay. The autophagy was observed under the fluorescence microscope through acridine orange staining and Ad-GFP-LC3 transfection experiment. Western blot was used to detect the expression of autophagy related protein and analyze the changes of phosphatidylinositol-3-kinase (PI3K)/protein kinase B(Akt)/mammalian target of rapamycin (mTOR) signaling pathway to explore the possible mechanism. Result: MTT results showed that ISL could significantly inhibit the proliferation of 786-O cells in a time-dose dependent manner (PPPPPPPPConclusion: ISL can inhibit the proliferation, migration and invasion of clear cell renal carcinoma 786-O cells, and induce autophagy by inhibiting the PI3K/Akt/mTOR signaling pathway.