Anti-silencing function 1B knockdown suppresses the malignant phenotype of colorectal cancer by inactivating the phosphatidylinositol 3-kinase/AKT pathway

BACKGROUND Mounting studies have highlighted the pivotal influence of anti-silencing function 1B(ASF1B)on the malignancy of cancers.AIM To explore the influence and mechanism of ASF1B in colorectal cancer(CRC).METHODS Quantitative real-time polymerase chain reaction(qRT-PCR)was used to detect mRNA expression of ASF1B.Immunohistochemical staining was performed to detect protein expression of ASF1B and Ki67 in tumor tissues.Western blot analysis was used to determine levels of ASF1B and proliferation/epithelial mesenchymal transition(EMT)/stemness-related proteins.In addition,the proliferation of CRC cells was assessed using Cell Counting Kit-8 and 5-Ethynyl-2’-Deoxyuridine assays.The migration and invasion of CRC cells were evaluated using transwell assays.Stemness of CRC cells was tested using the sphere formation assay.To construct a xenograft tumor model,HCT116 cells were introduced into mouse flanks via subcutaneous injection.RESULTS ASF1B expression was markedly increased in CRC tissues and cells,and it was inversely correlated with overall survival of CRC patients and was positively associated with the tumor node metastasis(TNM)stage of CRC patients.Silencing of ASF1B suppressed proliferation,migration,invasion,stemness and EMT of CRC cells as well as tumorigenesis of xenograft mice.Furthermore,protein levels of Pphosphatidylinositol 3-kinase(p-PI3K)and p-AKT were decreased after silencing of ASF1B in CRC cells.The inhibitory effects of ASF1B knockdown on cell proliferation,stemness and EMT were partly abolished by PI3K activator in CRC cells.CONCLUSION Silencing of ASF1B inactivated the PI3K/AKT pathway to suppress CRC malignancy in vitro.

Phosphatidylinositol 3-kinase/Akt pathway regulates hepatic stellate cell apoptosis

AIM: To investigate the role of phosphatidylinositol 3-kinase (PI 3-K)/Akt signaling pathway in the balance of HSC activation and apoptosis in rat hepatic stellate cells (HSC). METHODS: An activated HSC cell line was used in this study. LY 294002, the PI 3-K/Akt signal pathway block-er was used to investigate the molecular events on apoptosis in HSC and to interpret the role of this path-way in HSC apoptosis. Immunocytochemistry, Western blot and reverse transcription polymerase chain reac-tion (RT-PCR) analysis were applied to detect the ex-pression of PI 3-K, and simultaneously phosphorylated-Akt (p-Akt) and total-Akt were determined by Western blot. The HSC apoptosis was examined by annexin-V/ propidium iodide double-labelled flow cytometry and transmission electron microscopy. RESULTS: The apoptosis rates in LY 294002 (30.82% ± 2.90%) and LY 294002 + PDGF-BB (28.16% ± 2.58%) groups were signif icantly increased compared with those of control (9.02% ± 1.81%) and PDGF-BB (4.35% ± 1.18%). PDGF-BB augmented PI 3-K and p-Akt expres-sion. LY 294002 signif icantly reduced the contents of PI 3-K and p-Akt. mRNA transcription evaluated by RT-PCR showed similar tendencies as protein expression. CONCLUSION: Inhibition of PI 3-K/Akt signaling path-way induces apoptosis in HSC.

Response of Subcutaneous Xenografts of Endometrial Cancer in Nude Mice to Inhibitors of Phosphatidylinositol 3-Kinase/Akt and Mitogen-Activated Protein Kinase (MAPK) Pathways: An Effective Therapeutic Strategy for Endometrial Cancer

Objective: This study was designed to explore whether inhibition of the extracellular-regulated kinase (ERK) and phosphatidylinositol-3-kinase (PI3K) signaling pathways can inhibit the growth of xenografts of endometrial cancer cell lines with different estrogen receptors (ER) profiles in vivo and to provide preliminary laboratory basis for the probability of endometrial adenocarcinoma treatment with blockage of the two pathways, especially to endometrial cancer with low ER status. Methods: Human endometrial cancer Ishikawa bearing ER and HEC-1Awith low ER status cells were subcutaneously injected into BALB/c nude mice to establish endometrial cancer xenograft tumor models. The effects of PI3K/Akt inhibitor LY294002, MAPK/ERK1/2 inhibitor PD-98059 and their combinations on the growth of the xenograft tumors and apoptotic state of Ishikawa and HEC-1Acells were tested in vivo using the inhibitory rate, the terminal deoxynucleotidyl transferase-mediated nick-end labeling assay, H/E-stain. Western blot analysis was used to detect the alterations of activated ERK (P-ERK) and AKT (P-AKT) during this process. Results: LY294002, a PI3K/Akt pathway inhibitor, induced significant suppression in the growth of both Ishikawa and HEC-1Acell xenograft tumors, concomitant with increased apoptosis in xenografts as evidenced by TUNEL. A similar effect was also observed when the MAPK/ERK1/2 signaling pathway was inhibited by PD98059. Concurrent inhibition of the PI3K/Akt and MAPK/ERK1/2 pathways showed enhanced anti-tumor effects in vivo as indicated by increased apoptosis. At the same time, the levels of P-ERK and P-AKT in both xenograft tumors decreased, and their levels in combination group was the lowest. Conclusions: PD98059, LY294002 and their combinations showed remarkable inhibitory effects on xenograft tumors of endometrial carcinoma cell lines with different expression status of ER in vivo through blockage of PI3K/Akt and MAPK/ERK1/2 signaling pathways. This suggests that targeting these pathways may be an effective therapeutic strategy against endometrial carcinomas, especially for ER-negative cancers which show poor response to endocrinal therapy.

Tumor-related factor complement Clq/TNF-related protein 6 affects the development of digestive system tumors through the phosphatidylinositol 3-kinase pathway

In this editorial,we review the work of Razali et al published in World J Gas-troenterology,with a particular focus on the effect of rs10889677 variation in the phosphatidylinositol 3-kinase(PI3K)pathway and buparlisib on colitis-associated cancer.The role of PI3K in promoting cancer progression has been widely recognized,as it is involved in regulating the survival,differentiation,and prolif-eration of cancer cells.The complement Clq/TNF-related protein 6(CTRP6)is a newer tumor-associated factor.Recent studies have revealed the pro-tumor effect of CTRP6 in gastric cancer,hepatocellular carcinoma,colorectal cancer,and other gastrointestinal tumors through the PI3K pathway.This article attempts to reveal the mechanism through which the CTRP6 affects the development of digestive system tumors through the PI3K pathway by summarizing recent research.

Alisol B acetate induces apoptosis of SGC7901 cells via mitochondrial and phosphatidylinositol 3-kinases/Akt signaling pathways

AIM： To examine the effect of alisol B acetate on the growth of human gastric cancer cell line SGC7901 and its possible mechanism of action.METHODS： The cytotoxic effect of alisol B acetate on SGC7901 cells was measured by 3-（4,5-dimethylthiazol- 2-yl）-2,5-diphenyltetrazolium bromide （MI-I-） assay. Phase-contrast and electron microscopy were used to observe the morphological changes. Cell cycle and mitochondrial transmembrane potential （A^Pm） were determined by flow cytometry. Western blotting was used to detect the expression of apoptosis-regulated gene Bcl-2, Bax, Apaf-1, caspase-3, caspase-9, Akt, P-Akt and phosphatidylinositol 3-kinases （PI3K）.RESULTS： Alisol B acetate inhibited the proliferation of SGC7901 cell line in a time- and dose-dependent manner. PI staining showed that alisol B acetate can change the cell cycle distribution of SGC7901, increase the proportion of cells in G0-G1 phase and decrease the proportion of S phase cells and G2-M phase cells. Alisol B acetate at a concentration of 30 pmol/L induced apoptosis after 24, 48 and 72 h incubation, with occurrence rates of apoptotic cells of 4.36%, 14.42% and 21.16%, respectively. Phase-contrast and electron microscopy revealed that the nuclear fragmentation and chromosomal condensed, cells shrank and attachment loss appeared in the SGC7901 treated with alisol B acetate. Apoptosis of SGC7901 cells was associated with cell cycle arrest, caspase-3 and caspase-9 activation, loss of mitochondrial membrane potential and up-regulation of the ratio of Bax/Bcl-2 and inhibition of the PI3K/Akt.CONCLUSION： Alisol B acetate exhibits an antiproliferative effect in SGC7901 cells by inducing apoptosis. Apoptosis of SGC7901 cells involves mitochondria-caspase and PI3K/Akt dependent pathways.

Alterations in the human epidermal growth factor receptor 2-phosphatidylinositol 3-kinase-v-Akt pathway in gastric cancer

AIM:To investigate human epidermal growth factor receptor 2(HER2)-phosphatidylinositol 3-kinase(PI3K)-vAkt murine thymoma viral oncogene homolog signaling pathway.METHODS:We analyzed 231 formalin-fixed,paraffinembedded gastric cancer tissue specimens from Japanese patients who had undergone surgical treatment.The patients' age,sex,tumor location,depth of invasion,pathological type,lymph node metastasis,and pathological stage were determined by a review of the medical records.Expression of HER2 was analyzed by immunohistochemistry(IHC) using the HercepTest TM kit.Standard criteria for HER2 positivity(0,1+,2+,and 3+) were used.Tumors that scored 3+ were considered HER2-positive.Expression of phospho Akt(pAkt) was also analyzed by IHC.Tumors were considered pAkt-positive when the percentage of positive tumor cells was 10% or more.PI3K,catalytic,alpha polypeptide(PIK3CA) mutations in exons 1,9 and 20 were analyzed by pyrosequencing.Epstein-Barr virus(EBV) infection was analyzed by in situ hybridization targeting EBV-encoded small RNA(EBER) with an EBER-RNA probe.Microsatellite instability(MSI) was analyzed by polymerase chain reaction using the mononucleotide markers BAT25 and BAT26.RESULTS:HER2 expression levels of 0,1+,2+ and 3+ were found in 167(72%),32(14%),12(5%) and 20(8.7%) samples,respectively.HER2 overexpression(IHC 3+) significantly correlated with intestinal histological type(15/20 vs 98 /205,P = 0.05).PIK3CA mutations were present in 20 cases(8.7%) and significantly correlated with MSI(10/20 vs 9/211,P < 0.01).The mutation frequency was high(21%) in T4 cancers and very low(6%) in T2 cancers.Mutations in exons 1,9 and 20 were detected in 5(2%),9(4%) and 7(3%) cases,respectively.Two new types of PIK3CA mutation,R88Q and R108H,were found in exon1.All PIK3CA mutations were heterozygous missense singlebase substitutions,the most common being H1047R(6/20,30%) in exon20.Eighteen cancers(8%) were EBV-positive and this positivity significantly correlated with a diffuse histological type(13/18 vs 93/198,P = 0.04).There were 7 cases of lymphoepithelioma-like carcinomas(LELC) and 6 of those cases were EBV-positive(percent/EBV:6/18,33%;percent/all LELC:6/7,86%).pAkt expression was positive in 119(53%) cases but showed no correlation with clinicopathological characteristics.pAkt expression was significantly correlated with HER2 overexpression(16/20 vs 103/211,P < 0.01) but not with PIK3CA mutations(12/20 vs 107/211,P = 0.37) or EBV infection(8/18 vs 103/211,P = 0.69).The frequency of pAkt expression was higher in cancers with exon20 mutations(100%) than in those with exon1(40%) or exon9(56%) mutations.One case showed both HER2 overexpression and EBV infection and 3 cases showed both PIK3CA mutations and EBV infection.However,no cases showed both PIK3CA mutations and HER2 overexpression.One EBVpositive cancer with PIK3CA mutation(H1047R) was MSI-positive.Three of these 4 cases were positive for pAkt expression.In survival analysis,pAkt expression significantly correlated with a poor prognosis(hazard ratio 1.75;95%CI:1.12-2.80,P = 0.02).CONCLUSION:HER2 expression,PIK3CA mutations and EBV infection in gastric cancer were characterized.pAkt expression significantly correlates with HER2 expression and with a poor prognosis.

PI3K/AKT signaling and neuroprotection in ischemic stroke:molecular mechanisms and therapeutic perspectives

It has been reported that the PI3K/AKT signaling pathway plays a key role in the pathogenesis of ischemic stroke.As a result,the development of drugs targeting the PI3K/AKT signaling pathway has attracted increasing attention from researchers.This article reviews the pathological mechanisms and advancements in research related to the signaling pathways in ischemic stroke,with a focus on the PI3K/AKT signaling pathway.The key findings include the following:(1)The complex pathological mechanisms of ischemic stroke can be categorized into five major types:excitatory amino acid toxicity,Ca^(2+)overload,inflammatory response,oxidative stress,and apoptosis.(2)The PI3K/AKT-mediated signaling pathway is closely associated with the occurrence and progression of ischemic stroke,which primarily involves the NF-κB,NRF2,BCL-2,mTOR,and endothelial NOS signaling pathways.(3)Natural products,including flavonoids,quinones,alkaloids,phenylpropanoids,phenols,terpenoids,and iridoids,show great potential as candidate substances for the development of innovative anti-stroke medications.(4)Recently,novel therapeutic techniques,such as electroacupuncture and mesenchymal stem cell therapy,have demonstrated the potential to improve stroke outcomes by activating the PI3K/AKT signaling pathway,providing new possibilities for the treatment and rehabilitation of patients with ischemic stroke.Future investigations should focus on the direct regulatory mechanisms of drugs targeting the PI3K/AKT signaling pathway and their clinical translation to develop innovative treatment strategies for ischemic stroke.

Human neural stem cell-derived extracellular vesicles protect against ischemic stroke by activating the PI3K/AKT/mTOR pathway

Human neural stem cell-derived extracellular vesicles exhibit analogous functions to their parental cells,and can thus be used as substitutes for stem cells in stem cell therapy,thereby mitigating the risks of stem cell therapy and advancing the frontiers of stem cell-derived treatments.This lays a foundation for the development of potentially potent new treatment modalities for ischemic stroke.However,the precise mechanisms underlying the efficacy and safety of human neural stem cell-derived extracellular vesicles remain unclear,presenting challenges for clinical translation.To promote the translation of therapy based on human neural stem cell-derived extracellular vesicles from the bench to the bedside,we conducted a comprehensive preclinical study to evaluate the efficacy and safety of human neural stem cell-derived extracellular vesicles in the treatment of ischemic stroke.We found that administration of human neural stem cell-derived extracellular vesicles to an ischemic stroke rat model reduced the volume of cerebral infarction and promoted functional recovery by alleviating neuronal apoptosis.The human neural stem cell-derived extracellular vesicles reduced neuronal apoptosis by enhancing phosphorylation of phosphoinositide 3-kinase,mammalian target of rapamycin,and protein kinase B,and these effects were reversed by treatment with a phosphoinositide 3-kinase inhibitor.These findings suggest that human neural stem cell-derived extracellular vesicles play a neuroprotective role in ischemic stroke through activation of phosphoinositide 3-kinase/protein kinase B/mammalian target of rapamycin signaling pathway.Finally,we showed that human neural stem cell-derived extracellular vesicles have a good in vivo safety profile.Therefore,human neural stem cell-derived extracellular vesicles are a promising potential agent for the treatment of ischemic stroke.

Herbal cake-partitioned moxibustion inhibits colonic autophagy in Crohn’s disease via signaling involving distinct classes of phosphatidylinositol 3-kinases

BACKGROUND Autophagy is an evolutionarily conserved biological process in eukaryotic cells that involves lysosomal-mediated degradation and recycling of related cellular components.Recent studies have shown that autophagy plays an important role in the pathogenesis of Crohn’s disease(CD).Herbal cake-partitioned moxibustion(HM)has been historically practiced to treat CD.However,the mechanism by which HM regulates colonic autophagy in CD remains unclear.AIM To observe whether HM can alleviate CD by regulating colonic autophagy and to elucidate the underlying mechanism.METHODS Rats were randomly divided into a normal control(NC)group,a CD group,an HM group,an insulin+CD(I+CD)group,an insulin+HM(I+HM)group,a rapamycin+CD(RA+CD)group,and a rapamycin+HM(RA+HM)group.2,4,6-trinitrobenzenesulfonic acid was administered to establish a CD model.The morphology of the colonic mucosa was observed by hematoxylin-eosin staining,and the formation of autophagosomes was observed by electron microscopy.The expression of autophagy marker microtubule-associated protein 1 light chain 3 beta(LC3B)was observed by immunofluorescence staining.Insulin and rapamycin were used to inhibit and activate colonic autophagy,respectively.The mRNA expression levels of phosphatidylinositol 3-kinase class I(PI3KC1),Akt1,LC3B,sequestosome 1(p62),and mammalian target of rapamycin(mTOR)were evaluated by RT-qPCR.The protein expression levels of interleukin 18(IL-18),tumor necrosis factor-α(TNF-α),nuclear factorκB/p65(NF-κB p65),LC3B,p62,coiled-coil myosin-like BCL2-interacting protein(Beclin-1),p-mTOR,PI3KC1,class III phosphatidylinositol 3-kinase(PI3KC3/Vps34),and p-Akt were evaluated by Western blot analysis.RESULTS Compared with the NC group,the CD group showed severe damage to colon tissues and higher expression levels of IL-18 and NF-κB p65 in colon tissues(P<0.01 for both).Compared with the CD group,the HM group showed significantly lower levels of these proteins(PIL-18<0.01 and Pp65<0.05).There were no significant differences in the expression of TNF-αprotein in colon tissue among the rat groups.Typical autophagic vesicles were found in both the CD and HM groups.The expression of the autophagy proteins LC3B and Beclin-1 was upregulated(P<0.01 for both)in the colon tissues of rats in the CD group compared with the NC group,while the protein expression of p62 and p-mTOR was downregulated(P<0.01 for both).However,these expression trends were significantly reversed in the HM group compared with the CD group(PLC3B<0.01,PBeclin-1<0.05,Pp62<0.05,and Pm-TOR<0.05).Compared with those in the RA+CD group,the mRNA expression levels of PI3KC1,Akt1,mTOR,and p62 in the RA+HM group were significantly higher(PPI3KC1<0.01 and PAkt1,mTOR,and p62<0.05),while those of LC3B were significantly lower(P<0.05).Compared with the RA+CD group,the RA+HM group exhibited significantly higher PI3KC1,p-Akt1,and pmTOR protein levels(PPI3KC1<0.01,Pp-Akt1<0.05,and Pp-mTOR<0.01),a higher p62 protein level(P=0.057),and significantly lower LC3B and Vps34 protein levels(P<0.01 for both)in colon tissue.CONCLUSION HM can activate PI3KC1/Akt1/mTOR signaling while inhibiting the PI3KC3(Vps34)-Beclin-1 protein complex in the colon tissues of CD rats,thereby inhibiting overactivated autophagy and thus exerting a therapeutic effect.

Phosphatidylinositol 3-kinase CB association with preoperative radiotherapy response in rectal adenocarcinoma

AIM: To examine the correlation of phosphatidylinositol 3-kinase (PIK3) CB expression with preoperative radiotherapy response in patients with stage II/III rectal adenocarcinoma.

Analysis of Phosphatidylinositol 3-kinase Activation in the Adipose Tissue of Gestational Diabetes Mellitus Patients and Insulin Resistance

The P85 regulatory subunit protein and gene expression and P110 catalylic subunit activity of phosphatidylinositol 3-kinase (PI-3K) were investigated in adipose tissue of patients with gestational diabetes mellitus (GDM) in order to explore the molecular mechanisms of insulin resistance (IR) of GDM. Samples from patients with GDM (n=50), and controls (n=50) were collected. Fasting insulin (FIN) was determined by radioimmunoassay. Fasting plasma glucose (FPG) was measured by oxidase assay. Western blot technique was used to detect the levels of PI-3K P85 subunit in adipose tissues of patients with GDM. The mRNA expression of PI-3K P85 subunit was detected by reverse transcription polymerase chain reaction (RT-PCR) method in the adipose tissue. PI-3K activity was examined by immunoprecipitation, thin-layer chromatography and gamma scintillation counting. The results were analyzed statistically. It was found that the levels of FPG, FIN and HOMA-IR in GDM group were significantly higher than those in control group (all P0.05). PI-3K activity was significantly decreased to 82.89% in GDM group as compared with control group (P<0.01) and negatively correlated with HOMA-IR (r=-0.75, P<0.01). It was concluded that PI-3K in GDM patients may be involved in the insulin signaling pathway, resulting in IR of GDM.

Glycine-extended gastrin activates two independent tyrosine-kinases in upstream of p85/p110 phosphatidylinositol 3-kinase in human colonic tumour cells

AIM： To investigate whether Src, JAK2 and phosphatidylinositol 3-kinase （PI3K） pathways are involved in the proliferation of human colonic tumour cells induced by glycine-extended gastrin （G-gly）, the precursor of the mature amidated gastrin and to elucidate the molecular interaction between these three kinases in response to this peptide. METHODS： Using the human colonic tumour cell line HCT116 as a model, we first measured the activation of PI3K, p60-Src and JAK2 in response to G-gly by in vitro kinase assays. Then we investigated the involvement of these kinases in G-gly-induced cell proliferation by MTT test. RESULTS： G-gly stimulation induced p60-Src, JAK2 and PI3K activation in HCT116. The different pathways were involved in proliferation of human colon cancer cells induced by G-gly. Furthermore, we found that both Src and JAK2 were necessary to PI3K regulation by this peptide. However, we did not find any cross-talk between the two tyrosine kinases. CONCLUSION： Our results suggest that the p60-Src/ PI3K and JAK2/PI3K pathways act independently to mediate G-gly proliferative effect on human colonic tumour cells.

Micro RNA-21 promotes phosphatase gene and protein kinase B/phosphatidylinositol 3-kinase expression in colorectal cancer

AIM: To explore the regulatory mechanism of the target gene of micro RNA-21(mi R-21), phosphatase gene(p TEN), and its downstream proteins, protein kinase B(AKT) and phosphatidylinositol 3-kinase(p I3K), in colorectal cancer(CRC) cells. METHODS: Quantitative real-time p CR(q RT-p CR) and Western blot were used to detect the expression levels of mi R-21 and p TEN in HCT116, HT29, Colo32 and SW480 CRC cell lines. Also, the expression levels of p TEN m RNA and its downstream proteins AKT and p I3 K in HCT116 cells after downregulating mi R-21 were investigated. RESULTS: Comparing the mi R-21 expression in CRC cells, the expression levels of mi R-21 were highest in HCT116 cells, and the expression levels of mi R-21 were lowest in SW480 cells. In comparing mi R-21 and p TEN expression in CRC cells, we found that the protein expression levels of mi R-21 and p TEN were inversely correlated(p < 0.05); when mi R-21 expression was reduced, m RNA expression levels of p TEN did not significantly change(p > 0.05), but the expression levels of its protein significantly increased(p < 0.05). In comparing the levels of p TEN protein and downstream AKT and p I3 K in HCT116 cells after downregulation of mi R-21 expression, the levels of AKT and p I3 K protein expression significantly decreased(p < 0.05). CONCLUSION: p TEN is one of the direct target genesof mi R-21. Thus, phosphatase gene and its downstream AKT and p I3 K expression levels can be regulated by regulating the expression levels of mi R-21, which in turn regulates the development of CRC.

Roles of phosphatidylinositol-3-kinases signaling pathway in inflammation-related cancer:Impact of rs10889677 variant and buparlisib in colitis-associated cancer

BACKGROUND Phosphatidylinositol-3-kinases(PI3K)is a well-known route in inflammationrelated cancer.Recent discovery on PI3K-related genes revealed a potential variant that links ulcerative colitis(UC)and colorectal cancer(CRC)with colitisassociated cancer(CAC).PI3K/AKT pathway has been recommended as a potential additional therapeutic option for CRC due to its substantial role in modifying cellular processes.Buparlisib is a pan-class I PI3K inhibitor previously shown to reduce tumor growth.AIM To investigate the regulation of rs10889677 and the role of buparlisib in the PI3K signaling pathway in CAC pathogenesis.METHODS Genomic DNA from 32 colonic samples,including CAC(n=7),UC(n=10)and CRC(n=15),was sequenced for the rs10889677 mutation.The mutant and wildtype fragments were amplified and cloned in the pmirGLO vector.The luciferase activity of cloned vectors was assessed after transfection into the HT29 cell line.CAC mice were induced by a mixture of a single azoxymethane injection and three cycles of dextran sulphate sodium,then buparlisib was administered after 14 d.The excised colon was subjected to immunohistochemistry for Ki67 and Cleaved-caspase-3 markers and quantitative real-time polymerase chain reaction analysis for Pdk1 and Sgk2.RESULTS Luciferase activity decreased by 2.07-fold in the rs10889677 mutant,confirming the hypothesis that the variant disrupted miRNA binding sites,which led to an increase in IL23R expression and the activation of the PI3K signaling pathway.Furthermore,CAC-induced mice had a significantly higher disease activity index(P<0.05).Buparlisib treatment significantly decreased mean weight loss in CAC-induced mice(P<0.05),reduced the percentage of proliferating cells by 5%,and increased the number of apoptotic cells.The treatment also caused a downward trend of Pdk1 expression and significantly decreased Sgk2 expression.CONCLUSION Our findings suggested that the rs10889677 variant as a critical initiator of the PI3K signaling pathway,and buparlisib had the ability to prevent PI3K-non-AKT activation in the pathophysiology of CAC.

The cardioprotection induced by lipopolysaccharide involves phosphoinositide 3-kinase/Akt and high mobility group box 1 pathways

Objective： The mechanisms by which lipopolysaccharide （LPS） pretreatment induces cardioprotection following ischaemia/reperfusion （I/R） have not been fully elucidated. We hypothesized that activation of phosphoinositide 3-kinase （PI3K）/Akt and high mobility group box 1 （HMGBxl） signaling plays an important role in LPS-induced cardioprotection. Methods： In in vivo experiments, age- and weight- matched male C57BL/10Sc wild type mice were pretreated with LPS before ligation of the left anterior descending coronary followed by reperfusion. Infarction size was examined by triphenyltetrazolium chloride （TTC） staining. Akt, phospho-Akt, and HMGBxl were assessed by immunoblotting with appropriate primary antibodies. In situ cardiac myocyte apop- tosis was examined by the TdT-mediated dUTP nick-end labeling （TUNEL） assay. In an in vitro study, rat cardiac myoblasts （H9c2） were subdivided into two groups, and only one was pretreated with LPS. After pretreatment, the cells were transferred into a hypoxic chamber under 0.5% 02. Levels of HMGBxl were assessed by immunoblot. Results： In the in vivo experiment, pretreatment with LPS reduced the at risk infarct size by 70.6% and the left ventricle infarct size by 64.93% respectively. Pretreatment with LPS also reduced cardiac myocytes apoptosis by 39.1% after ischemia and reperfusion. The mechanisms of LPS induced cardioprotection involved increasing PI3K/Akt activity and decreasing expression of HMGBxl. In the in vitro study, pretreatment with LPS reduced the level of HMGBxl in H9c2 cell cytoplasm following hypoxia. Conclusion： The results suggest that the cardioprotection following I/R induced by LPS pretreatment involves PI3K/Akt and HMGBxl pathways.

Yes相关蛋白(YAP)通过激活PI3K/AKT通路促进皮肤鳞状细胞癌细胞侵袭和迁移

目的探讨Yes相关蛋白(YAP)在皮肤鳞状细胞癌(cSCC)中表达及与cSCC侵袭和迁移中的作用。方法通过免疫组织化学染色法检测cSCC、鲍温病(BD)、癌旁正常皮肤组织中YAP的表达水平,并分析与临床病理参数之间的关系;利用慢病毒转染构建YAP基因敲低的A431稳定细胞株,利用四甲基罗丹明标记的鬼笔环肽检测A431细胞微丝分布和数量,Transwell TM实验检测细胞侵袭能力,划痕实验检测A431细胞的迁移能力;免疫荧光细胞化学染色法观察敲低YAP后上皮间质转化(EMT)相关标志物上皮钙黏素(E-cadherin)、锌指转录因子Snail的表达;Western blot法检测E-cadherin、Snail、β-catenin、磷脂酰肌醇3激酶(PI3K)、蛋白激酶B(AKT)、磷酸化的蛋白激酶B(p-AKT)、核糖体蛋白S6(S6)、磷酸化S6(p-S6)、4E结合蛋白1(4EBP1)、磷酸化的4EBP1(p-4EBP1)的表达。结果YAP在cSCC和BD中表达显著高于癌旁正常皮肤组织;cSCC中YAP高表达与肿瘤大小、分化程度、侵袭程度密切相关,与患者的性别、年龄、发病部位、形态类型、是否神经脉管侵犯不相关;敲低A431细胞中YAP后,肿瘤细胞的侵袭、迁移能力降低,细胞微丝变细、伪足变少;E-cadherin表达增加,Snail和β-catenin蛋白表达降低,p-AKT、p-S6及p-4EBP1蛋白表达降低。结论YAP在cSCC中高表达,YAP激活PI3K/AKT信号通路促进cSCC的侵袭、迁移及EMT过程。

栀子苷调节PI3K/AKT/mTOR信号通路在动脉粥样硬化形成过程中对Th17/Treg功能的影响

目的:观察栀子苷对载脂蛋白E缺乏(ApoE^(-/-))小鼠Th17/调节性T(Treg)细胞失衡的影响及其作用机制。方法:将50只纯合子ApoE^(-/-)雌性小鼠随机分为对照组、模型组和栀子苷低剂量组、栀子苷中剂量组、栀子苷高剂量组。对照组小鼠喂养普通饲料,模型组和栀子苷组小鼠喂养高脂饲料。从第8周开始,栀子苷各剂量组每日灌胃栀子苷(25、50、100 mg/kg),连续8周。试验结束时,采用油红O染色评估主动脉及其根部动脉粥样硬化(AS)病变面积比。采用定量逆转录聚合酶链式反应(RT-PCR)分析主动脉组织肿瘤坏死因子-α(TNF-α)、白细胞介素(IL)-6、IL-17A和IL-10 mRNA表达;采用流式细胞仪分析脾脏中Th17和Treg细胞百分比;蛋白免疫印迹法(Western Blot)检测主动脉组织磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B(AKT)/哺乳动物雷帕霉素靶蛋白(mTOR)信号通路相关蛋白表达。结果:油红O染色病变显示,栀子苷中剂量组、栀子苷高剂量组病变百分比低于模型组(P<0.05)。与对照组比较,模型组主动脉TNF-α、IL-6和IL-17A mRNA表达水平升高(P<0.05);栀子苷各剂量组主动脉TNF-α、IL-6和IL-17A mRNA表达水平降低(P<0.05)。与对照组比较,模型组主动脉抗炎细胞因子IL-10 mRNA表达水平降低(P<0.05);栀子苷各剂量组主动脉抗炎细胞因子IL-10 mRNA表达水平升高(P<0.05)。与对照组比较,模型组小鼠脾脏中Th17细胞百分比升高,Treg细胞百分比降低(P<0.05)。栀子苷处理恢复了AS小鼠Th17和Treg细胞的平衡。栀子苷抑制PI3K的表达及AKT和mTOR的磷酸化,MHY1485(mTOR活化剂)减弱了栀子苷对T细胞分化的影响。结论:栀子苷抗AS作用机制可能与抑制PI3K/AKT/mTOR信号引起的Treg细胞增多和Th17细胞减少有关。

基于PI3K/AKT/GSK-3β信号通路探讨EA改善APP/PS1双转基因小鼠认知功能障碍的内在机制

目的探讨鞣花酸(ellagicacid,EA)对APP/PS1双转基因小鼠认知功能的影响,并基于磷脂酰肌醇3-激酶/蛋白激酶B/糖原合成酶激酶-3(PI3K/AKT/GSK-3β)信号通路探讨鞣花酸对双转基因小鼠海马氧化应激水平的调节机制。方法将32只SPF级6月龄APP/PS1双转基因小鼠随机分为4组,即APP/PS1组、APP/PS1+EA组、APP/PS1+LY294002组、APP/PS1+EA+LY294002组,每组8只,另外选取8只SPF级C57BL/6J野生型小鼠(Wildtype)作为空白对照组,即WT组。APP/PS1+EA组给予50mg·kg^(-1)·d^(-1)灌胃EA;APP/PS1+LY294002组予以1.5mg·kg^(-1)·d^(-1)腹腔注射PI3K抑制剂LY294002;APP/PS1+EA+LY294002组予以50mg·kg^(-1)·d^(-1)灌胃EA,同时按1.5mg·kg^(-1)·d^(-1)腹腔注射LY294002;WT组和APP/PS1组于相同时间点灌胃等体积10%二甲基亚砜(DMSO)。每日给药1次,连续给药60天。Morris水迷宫检测小鼠学习和记忆能力,免疫组化、蛋白免疫印迹法检测PI3K、AKT、GSK-3β相关蛋白的表达,透射电镜观察小鼠海马组织超微结构变化。结果与WT组相比,其他四组的逃避潜伏期均增长(P<0.05),穿越平台次数明显减少(P<0.01);APP/PS1组、APP/PS1+LY294002组和APP/PS1+EA+LY294002组中的PI3K、AKT蛋白表达量显著降低(P<0.01),GSK-3β表达量显著升高(P<0.01);APP/PS1+EA组的PI3K表达量降低(P<0.05),AKT表达量显著降低(P<0.01),GSK-3β表达量升高(P<0.05);与WT组相比,APP/PS1组海马神经元细胞数目较少,线粒体结构破坏,大部分线粒体出现肿胀,线粒体的内膜和外模不完整,部分线粒体嵴消失,微管、微丝缠结,排列紊乱,而APP/PS1+EA组神经元细胞数较APP/PS1组增多,线粒体结构较清晰,可见清楚的线粒体嵴,线粒体轻度水肿。微管、微丝排列较整齐有序。结论鞣花酸改善AD模型小鼠的学习和记忆能力、减少海马神经元细胞损伤和凋亡,其作用机制可能是通过调节PI3K、AKT、GSK-3β等相关蛋白降低AD模型小鼠海马氧化应激水平。

SERPINH1 promoted the proliferation and metastasis of colorectal cancer by activating PI3K/Akt/mTOR signaling pathway

BACKGROUND Serpin peptidase inhibitor clade H member 1(SERPINH1)was initially recognized as an oncogene implicated in various human malignancies.Nevertheless,the clinical relevance and functional implications of SERPINH1 in colorectal cancer(CRC)remain largely elusive.AIM To investigate the effects of SERPINH1 on CRC cells and its specific mechanism.METHODS Quantitative real-time polymerase chain reaction,western blotting analysis,The Cancer Genome Atlas data mining and immunohistochemistry were employed to examine SERPINH1 expression in CRC cell lines and tissues.A series of in-vitro assays were performed to demonstrate the function of SERPINH1 and its possible mechanisms in CRC.RESULTS SERPINH1 demonstrated elevated expression levels in both CRC cells and tissues,manifested at both mRNA and protein tiers.Elevated SERPINH1 levels correlated closely with advanced T stage,lymph node involvement,and distant metastasis,exhibiting a significant association with poorer overall survival among CRC patients.Subsequent investigations unveiled that SERPINH1 overexpression notably bolstered CRC cell proliferation,invasion,and migration in vitro,while conversely,SERPINH1 knockdown elicited the opposite effects.Gene set enrichment analysis underscored a correlation between SERPINH1 upregulation and genes associated with cell cycle regulation.Our findings underscored the capacity of heightened SERPINH1 levels to expedite G1/S phase cell cycle progression via phosphatidylinositol 3-kinase/AKT/mechanistic target of rapamycin pathway activation,thereby facilitating CRC cell invasion and migration.CONCLUSION These findings imply a crucial involvement of SERPINH1 in the advancement and escalation of CRC,potentially positioning it as a novel candidate for prognostic assessment and therapeutic intervention in CRC management.

中医药调控PI3K/AKT信号通路干预溃疡性结肠炎研究进展

通过干预磷脂酰肌醇3-激酶(phosphatidylinositol 3-kinase,PI3K)/蛋白激酶B(protein kinase B,AKT)信号通路中的相关因子而发挥治疗作用的中医药疗法可分为清热解毒燥湿、活血化瘀止血、健脾温肾祛湿和攻补兼施等,能有效遏制UC的发生发展,其作用机制为:(1)通过抑制上游信号因子表皮生长因子受体,进而抑制PI3K/AKT通路,减少肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-1β(interleukin-1β,IL-1β)和白细胞介素-6(interleukin-6,IL-6)在内的促炎细胞因子的表达,降低结肠炎症反应,减轻炎症所引起的结肠组织损伤;(2)直接作用于PI3K-AKT通路,进而抑制核转录因子-κB(nuclear transcription factor-κB,NF-κB)的表达,减少细胞凋亡和促炎因子的表达,缓解肠黏膜组织损伤,修复肠黏膜屏障功能的完整性;(3)通过PI3K/AKT途径下调辅助性T细胞17(T helper cell 17,Th17)和辅助性T细胞3(T helper cell 3,Th3)的分化,调节免疫反应及细胞凋亡,改善肠黏膜屏障,缓解UC结肠炎症反应。中医药具有成分复杂,有效成分作用靶点多,治疗途径多样等特点,但其具体分子生物学作用机制尚未完全明确,今后,需对分子生物学作用机制进行深入研究,为临床治疗溃疡性结肠炎等易反复发作的消化性疾病和新药的开发提供新策略。