Beneficial Role of Vitexin in Parkinson's Disease.

Today, Parkinson's disease (PD) is the foremost neurological disorder all across the globe. In the quest for a novel therapeutic agent for PD with a multimodal mechanism of action and relatively better safety profile, natural flavonoids are now receiving greater attention as a potential source of neuroprotection. Vitexin have been shown to exhibit diverse biological benefits in various disease conditions, including PD. It exerts its anti-oxidative property in PD patients by either directly scavenging reactive oxygen species (ROS) or by upregulating the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and enhancing the activities of antioxidant enzymes. Also, vitexin activates the ERK1/1 and phosphatidyl inositol-3 kinase/Akt (PI3K/Akt) pro-survival signalling pathway, which upregulates the release of anti-apoptotic proteins and downregulates the expression of pro-apoptotic proteins. It could be antagonistic to protein misfolding and aggregation. Studies have shown that it can also act as an inhibitor of monoamine oxidase B (MAO-B) enzyme, thereby increasing striatal dopamine levels, and hence, restoring the behavioural deficit in experimental PD models. Such promising pharmacological potential of vitexin could be a game-changer in devising novel therapeutic strategies against PD. This review discusses the chemistry, properties, sources, bioavailability and safety profile of vitexin. The possible molecular mechanisms underlying the neuroprotective action of vitexin in the pathogenesis of PD alongside its therapeutic potential is also discussed.

SDC4-rs1981429 and ATM-rs228590 may provide early biomarkers of breast cancer risk.

In Australia, 13% of women are diagnosed with breast cancer (BC) in their lifetime with approximately 20,000 women diagnosed with the disease in 2021. BC is characterised by complex histological and genomic influences with recent advances in cancer biology improving early diagnosis and personalised treatment interventions. The Phosphatidyl-inositol-3-kinase/Protein kinase B (PI3K/AKT) pathway is essential in apoptosis resistance, cell survival, activation of cellular responses to DNA damage and DNA repair. Heparan sulfate proteoglycans (HSPGs) are ubiquitous molecules found on the cell surface and in the extracellular matrix with essential functions in regulating cell survival, growth, adhesion and as mediators of cell differentiation and migration. HSPGs, particularly the syndecans (SDCs), have been linked to cancers, making them an exciting target for anticancer treatments. In the PI3K/AKT pathway, syndecan-4 (SDC4) has been shown to downregulate AKT Serine/Threonine Kinase (AKT1) gene expression, while the ATM Serine/Threonine Kinase (ATM) gene has been found to inhibit this pathway upstream of AKT. We investigated single-nucleotide polymorphisms (SNPs) in HSPG and related genes SDC4, AKT1 and ATM and their influence on the prevalence of BC. SNPs were genotyped in the Australian Caucasian Genomics Research Centre Breast Cancer (GRC-BC) population and in the Griffith University-Cancer Council Queensland Breast Cancer Biobank (GU-CCQ BB) population. We identified that SDC4-rs1981429 and ATM-rs228590 may influence the development and progression of BC, having the potential to become biomarkers in early BC diagnosis and personalised treatment.

Hydrophobic Bile Salts Induce Pro-Fibrogenic Proliferation of Hepatic Stellate Cells through PI3K p110 Alpha Signaling.

Bile salts accumulating during cholestatic liver disease are believed to promote liver fibrosis. We have recently shown that chenodeoxycholate (CDC) induces expansion of hepatic stellate cells (HSCs) in vivo, thereby promoting liver fibrosis. Mechanisms underlying bile salt-induced fibrogenesis remain elusive. We aimed to characterize the effects of different bile salts on HSC biology and investigated underlying signaling pathways. Murine HSCs (mHSCs) were stimulated with hydrophilic and hydrophobic bile salts. Proliferation, cell mass, collagen deposition, and activation of signaling pathways were determined. Activation of the human HSC cell line LX 2 was assessed by quantification of α-smooth muscle actin (αSMA) expression. Phosphatidyl-inositol-3-kinase (PI3K)-dependent signaling was inhibited both pharmacologically and by siRNA. CDC, the most abundant bile salt accumulating in human cholestasis, but no other bile salt tested, induced Protein kinase B (PKB) phosphorylation and promoted HSC proliferation and subsequent collagen deposition. Pharmacological inhibition of the upstream target PI3K-inhibited activation of PKB and pro-fibrogenic proliferation of HSCs. The PI3K p110α-specific inhibitor Alpelisib and siRNA-mediated knockdown of p110α ameliorated pro-fibrogenic activation of mHSC and LX 2 cells, respectively. In summary, pro-fibrogenic signaling in mHSCs is selectively induced by CDC. PI3K p110α may be a potential therapeutic target for the inhibition of bile salt-induced fibrogenesis in cholestasis.

Ticagrelor Protects against Sepsis-Induced Acute Kidney Injury through an Adenosine Receptor-Dependent Pathway.

OBJECTIVE: Ticagrelor is a widely used anti-platelet drug. However, the mechanisms by which ticagrelor protects against sepsis-induced acute kidney injury (AKI) have not been clearly demonstrated. We designed this study to explore the protective effect of ticagrelor on sepsis-induced AKI and to explore the underlying mechanisms. METHODS: C57BL6J mice received oral ticagrelor (20 mg/kg and 50 mg/kg) for 7 days, and then caecal ligation and puncture (CLP) were performed. An adenosine receptor antagonist, CGS15943, was administered (10 mg/kg, intraperitoneal injection) to block the adenosine pathway 2 h before CLP. After 24 h, serum creatinine levels were measured. Periodic acid-Schiff (PAS) staining and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining were employed to analyze pathological changes and cell apoptosis. Serum concentrations of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and mRNA expression of tissue TNF-α and IL-1ß were detected. Western blotting analysis was used to determine AKT and mammalian target of rapamycin (mTOR) protein expression in the kidney. RESULTS: PAS staining showed less swelling of renal tubules, and TUNEL staining revealed less cell apoptosis in the ticagrelor group than in the CLP group. Serum creatinine levels were significantly lower in the ticagrelor group than in the CLP group. Moreover, significantly lower serum and kidney levels of TNF-α and IL-1ß were observed in the ticagrelor group. CGS15943 blocked the effects of ticagrelor. Western blotting analysis showed increased phosphorylation of AKT and mTOR in the kidneys of the 50 mg/kg ticagrelor group. The adenosine receptor antagonist inhibited the activation of AKT and mTOR. CONCLUSION: This study demonstrates that the protective effect of ticagrelor on sepsis-induced AKI depends on adenosine receptor activation and the subsequent increase of AKT and mTOR phosphorylation.

The PI3K/Akt/mTOR pathway in lung cancer; oncogenic alterations, therapeutic opportunities, challenges, and a glance at the application of nanoparticles.

Lung cancer is the leading cause of cancer-related mortality worldwide. Although the PI3K/Akt/mTOR signaling pathway has recently been considered as one of the most altered molecular pathways in this malignancy, few articles reviewed the task. In this review, we aim to summarize the original data obtained from international research laboratories on the oncogenic alterations in each component of the PI3K/Akt/mTOR pathway in lung cancer. This review also responds to questions on how aberrant activation in this axis contributes to uncontrolled growth, drug resistance, sustained angiogenesis, as well as tissue invasion and metastatic spread. Besides, we provide a special focus on pharmacologic inhibitors of the PI3K/Akt/mTOR axis, either as monotherapy or in a combined-modal strategy, in the context of lung cancer. Despite promising outcomes achieved by using these agents, however, the presence of drug resistance as well as treatment-related adverse events is the other side of the coin. The last section allocates a general overview of the challenges associated with the inhibitors of the PI3K pathway in lung cancer patients. Finally, we comment on the future research aspects, especially in which nano-based drug delivery strategies might increase the efficacy of the therapy in this malignancy.

[Corrigendum] Panax notoginseng saponins prevent senescence and inhibit apoptosis by regulating the PI3K­AKT­mTOR pathway in osteoarthritic chondrocytes.

Following the publication of the above article (and a Corrigendum that has already been published with the intention of showing the corrected version of Fig. 6 (DOI:10.3892/ijmm.2020.4786; published online on November 11, 2020), an interested reader drew to the authors' attention that, in Fig. 5B on p. 1233, the 'OA' and 'OA+IGF­1+PNS' data panels appeared to show overlapping data. The authors have re­examined their original data, and realize that Fig. 5 was assembled incorrectly; essentially, the 'OA+IGF­1+PNS' data panel for Fig. 5B was selected in error. The corrected version of Fig. 5 is shown on the next page. Note that this inadvertent error did not affect the main conclusions reported in this study. The authors are grateful to the Editor of International Journal of Molecular Medicine for granting them the opportunity to publish this Corrigendum, and apologize to the readership for any inconvenience caused. [the original article was published on International Journal of Molecular Medicine 45: 1225­1236, 2020; DOI: 10.3892/ijmm.2020.4491].

miR­519d­3p/HIF­2α axis increases the chemosensitivity of human cervical cancer cells to cisplatin via inactivation of PI3K/AKT signaling.

Cisplatin (DDP)­based chemotherapy is a standard treatment for cervical cancer, although chemotherapy resistance remains a major concern. Hypoxia­inducible factor­2 α (HIF­2α) plays an important role in chemotherapy resistance. MicroRNAs (miRs) can inhibit gene expression by binding to the 3'­untranslated region of the target gene. The authors' previous study showed that miR­519d­3p plays an important role in the regulation of HIF­2α expression under hypoxic conditions in cervical cancer. However, the function and regulatory mechanisms of the miR­519d­3p/HIF­2α axis in DDP­resistance in cervical cancer are not fully understood. Therefore, the aim of the present study was to investigate whether the miR­519d­3p/HIF­2α axis increased DDP resistance by regulating the PI3K/AKT signaling pathway. It was found that the expression of miR­519d­3p was lower in DDP­resistant cervical cancer cells (CaSki/DDP and HeLa/DDP) compared with CaSki and HeLa cells under hypoxic conditions. Additionally, miR­519d­3p overexpression decreased the IC50 value in CaSki/DDP and HeLa/DDP cells, and inhibited HIF­2α protein expression and the PI3K/AKT signaling pathway under hypoxic conditions. Furthermore, it was demonstrated that HIF­2α overexpression reduced the effect of miR­519d­3p overexpression on HeLa/DDP and CaSki/DDP cells. Moreover, the present results suggested that HIF­2α overexpression increased the IC50 value in CaSki/DDP and HeLa/DDP cells. It was also found that HIF­2α overexpression reduced the effect of miR­519d­3p overexpression on the PI3K/AKT signaling pathway. Therefore, the present results indicated that the miR­519d­3p/HIF­2α axis increased DDP resistance of cervical cancer cells by suppressing the PI3K/AKT signaling pathway under hypoxic conditions.

An information theoretic approach to insulin sensing by human kidney podocytes.

Podocytes are key components of the glomerular filtration barrier (GFB). They are insulin-responsive but can become insulin-resistant, causing features of the leading global cause of kidney failure, diabetic nephropathy. Insulin acts via insulin receptors to control activities fundamental to GFB integrity, but the amount of information transferred is unknown. Here we measure this in human podocytes, using information theory-derived statistics that take into account cell-cell variability. High content imaging was used to measure insulin effects on Akt, FOXO and ERK. Mutual Information (MI) and Channel Capacity (CC) were calculated as measures of information transfer. We find that insulin acts via noisy communication channels with more information flow to Akt than to ERK. Information flow estimates were increased by consideration of joint sensing (ERK and Akt) and response trajectory (live cell imaging of FOXO1-clover translocation). Nevertheless, MI values were always <1Bit as most information was lost through signaling. Constitutive PI3K activity is a predominant feature of the system that restricts the proportion of CC engaged by insulin. Negative feedback from Akt supressed this activity and thereby improved insulin sensing, whereas sensing was robust to manipulation of feedforward signaling by inhibiting PI3K, PTEN or PTP1B. The decisions made by individual podocytes dictate GFB integrity, so we suggest that understanding the information on which the decisions are based will improve understanding of diabetic kidney disease and its treatment.

Aspartic Aminopeptidase Is a Novel Biomarker of Aggressive Chronic Lymphocytic Leukemia.

Treatment of chronic lymphocytic leukemia has advanced substantially as our understanding of the kinase signal transduction pathways driven by the B cell receptor (BcR) has developed. Particularly, understanding the role of Bruton tyrosine kinase and phosphatidyl inositol 3 kinase delta in driving prosurvival signal transduction in chronic lymphocytic leukemia (CLL) cells and their targeting with pharmacological inhibitors (ibrutinib and idelalisib, respectively) has improved patient outcomes significantly. The kinase signaling pathway induced by the BcR is highly complex and has multiple interconnecting branches mediated by tyrosine and serine/threonine kinases activated downstream of the BcR. There is a high level of redundancy in the biological responses, with several BcR-signaling kinases driving nuclear factor kappa B activation or inducing antiapoptotic Bcl-2 genes. Accordingly, common gene targets of BcR-signaling kinases may serve as biomarkers indicating enhanced BCR-signaling and aggressive disease progression. This study used a gene expression correlation analysis of malignant B cell lines and primary CLL cells to identify genes whose expression correlated with BCR-signaling kinases overexpressed and/or overactivated in CLL, namely: AKT1, AKT2, BTK, MAPK1, MAPK3, PI3KCD and ZAP70. The analysis identified a 32-gene signature with a strong prognostic potential and DNPEP, the gene coding for aspartic aminopeptidase, as a predictor of aggressive CLL. DNPEP gene expression correlated with MAPK3, PI3KCD, and ZAP70 expression and, in the primary CLL test dataset, showed a strong prognostic potential. The inhibition of DNPEP with a pharmacological inhibitor enhanced the cytotoxic potential of idelalisib and ibrutinib, indicating a biological functionality of DNPEP in CLL. DNPEP, as an aminopeptidase, contributes to the maintenance of the free amino acid pool in CLL cells found to be an essential process for the survival of many cancer cell types, and thus, these results warrant further research into the exploitation of aminopeptidase inhibitors in the treatment of drug-resistant CLL.

Beta carotene protects H9c2 cardiomyocytes from advanced glycation end product-induced endoplasmic reticulum stress, apoptosis, and autophagy via the PI3K/Akt/mTOR signaling pathway.

BACKGROUND: Diabetic cardiomyopathy (DCM), which is associated with many pathological processes, commonly occurs when advanced glycation end products (AGEs) are present. ß-carotene (BC) is a well-known vitamin A precursor that is found in many fruits and vegetables. BC can reduce the risk of cancer and cardiovascular disease. This study aimed to investigate the effect of BC on AGE-induced myocardial injury in vitro. METHODS: Cell viability test was used to select 40 µM concentrations of BC to treat AGE-induced H2c9 cells. The cell apoptosis was detected by flow cytometry. Western blotting was used to measure the protein expression levels of Bcl-2-associated X protein (Bax), B-cell lymphoma-2 (Bcl-2), cleaved caspase-3, activating transcription factor 4 (ATF4), glucose-regulated protein 78 (GRP78), CCAAT/enhancer-binding protein homologous protein (CHOP), beclin 1, p62,microtubule-associated protein 1 light chain 3 (LC3), phosphorylated PI3K (p-PI3K), phosphorylated Akt (p-AKT), and phosphorylated mTOR (p-mTOR). Enzyme-linked immunosorbent assay (ELISA) was performed to measure the levels of lactate dehydrogenase (LDH) and cardiac troponin-1 (cTn-I). Reactive oxygen species (ROS) was detected by flow cytometry. The levels of malondialdehyde (MDA), glutathione peroxidase (GSH-Px), and superoxide dismutase (SOD) were used to determine MDA kits, SOD assay kit and GSH-Px kit, respectively. RESULTS: BC significantly inhibited AGE-induced cell death and apoptosis in H9c2 cells. BC had a suppressive effect on intracellular ROS production and antioxidative enzyme reduction. Moreover, BC decreased hyperactive endoplasmic reticulum (ER) stress and autophagy in H9c2 cells. Furthermore, BC exerted a cardioprotective effect in AGE-induced H9c2 cells via the activation of the PI3K/Akt/mTOR signaling pathway. CONCLUSIONS: BC exhibited a cardioprotective effect AGE-induced apoptosis. Our study provides a foundation for further study into the potential value of BC for treating DCM or other heart diseases.

Ginkgolide B inhibits hydrogen peroxide­induced apoptosis and attenuates cytotoxicity via activating the PI3K/Akt/mTOR signaling pathway in H9c2 cells.

Ginkgolide B (GB) is a diterpene lactone found in the leaves of the traditional Chinese medicinal plant Ginkgo that has been shown to have various pharmacological effects. However, the anti­apoptotic properties of GB in cardiovascular disease remain poorly understood. The present study aimed to investigate the effect of GB on hydrogen peroxide­induced cell injury in cardiac H9c2 cells, and to further clarify its protective mechanism of action. An in vitro hydrogen peroxide­treated H9c2 cell model was used in order to mimic myocardial ischemia­reperfusion (I/R) injury. Cell viability was assessed by the Cell Counting Kit­8 assay. The induction of apoptosis was determined by flow cytometry and staining was performed using Hoechst 33342. In addition, the effect of GB on the expression levels of apoptosis­associated proteins was evaluated by western blot analysis. The present study demonstrated that GB protected against hydrogen peroxide­induced cytotoxicity and cell apoptosis in H9c2 cardiac cells. GB upregulated the expression level of the anti­apoptotic protein Bcl­2 and downregulated the expression levels of the pro­apoptotic proteins cleaved caspase­3 and Bax in hydrogen peroxide­treated H9c2 cells. The molecular mechanism underlying the anti­apoptotic effects of GB was subsequently detected. GB pretreatment activated the PI3K/Akt/mTOR signaling pathway and caused an increase in the phosphorylation levels of Akt and mTOR in hydrogen peroxide­treated H9c2 cells. These results revealed that GB inhibited hydrogen peroxide­induced apoptosis in H9c2 cells via activation of the PI3K/Akt/mTOR signaling pathway. These findings indicate the potential therapeutic benefits of GB in the treatment of myocardial I/R injury.

RRM2 elicits the metastatic potential of breast cancer cells by regulating cell invasion, migration and VEGF expression via the PI3K/AKT signaling.

Breast cancer is the second leading primary cause for cancer-related mortality among women and metastasis to the brain is a disastrous event for patients with increasing incidence. A previous study confirmed the critical function of RRM2 in breast cancer cell growth. Unfortunately, the role and fundamental molecular mechanism of RRM2 in breast cancer metastasis remains elusive. In the current study, higher RRM2 expression was validated in breast cancer tissues, especially in the brain metastasis group. Simultaneously, the expression of RRM2 was increased in breast cancer cells relative to the normal breast epithelial cell line MCF-10A, concomitant with higher levels of RRM2 in the highly metastatic MDA-MB-231 cell line relative to the weakly metastatic MCF-7 cell line. Knockdown of RRM2 by small interfering-RRM2 transfection notably suppressed the malignant metastatic behavior of breast cancer cells, including invasion and migration. Simultaneously, RRM2 downregulation also restrained the transcription and release of vascular endothelial growth factor (VEGF) in breast cancer cells. Moreover, inhibition of RRM2 dampened the activation of phosphatidylinositol 3 kinase (PI3K)/protein kinase B (AKT) signaling by decreasing phosphorylated-AKT and downstream matrix metalloproteinases-2 expression. Intriguingly, reactivation of the PI3K/AKT pathway with its agonist insulin-like growth factor-1 reversed the adverse effects of RRM2 suppression on cancer cell invasion, migration and VEGF expression. Together, these findings suggest that RRM2 may act as a pro-metastatic factor to facilitate breast cancer metastasis by evoking cell invasion, migration and VEGF expression through the PI3K/AKT signaling pathway. This study may provide an attractive target for metastatic intervention in breast cancer.

MicroRNA-19a-3p regulates cell growth through modulation of the PIK3IP1-AKT pathway in hepatocellular carcinoma.

There are some controversies about the involvement of microRNA (miR)-19a-3p in hepatocellular carcinoma (HCC) biology, even though many studies have shown that it plays an important role in cancer. In this study, we found that miR-19a-3p is usually overexpressed in HCC tissues compared with corresponding peritumorous tissues, and its expression was associated with tumor size and poor overall survival. MiR-19a-3p promoted cell proliferation significantly, and more cells were found in the S phase. In vivo, miR-19a-3p promoted liver tumor growth, and more HCC cells were found in the active cell cycle. Sequencing and bioinformatics analysis predicted that PIK3IP1 is a likely target gene of miR-19a-3p, and we next confirmed it by luciferase and rescue assays. Altogether, our data showed an important role of PIK3IP1 downregulation by miR-19a-3p in HCC progression, and the miR-19a-3p-PIK3IP1-AKT pathway may be a potential therapeutic target.

Network Reconstruction and Significant Pathway Extraction Using Phosphoproteomic Data from Cancer Cells.

Protein phosphorylation acts as an efficient switch controlling deregulated key signaling pathway in cancer. Computational biology aims to address the complexity of reconstructed networks but overrepresents well-known proteins and lacks information on less-studied proteins. A bioinformatic tool to reconstruct and select relatively small networks that connect signaling proteins to their targets in specific contexts is developed. It enables to propose and validate new signaling axes of the Syk kinase. To validate the potency of the tool, it is applied to two phosphoproteomic studies on oncogenic mutants of the well-known phosphatidyl-inositol 3-kinase (PIK3CA) and the unfamiliar Src-related tyrosine kinase lacking C-terminal regulatory tyrosine and N-terminal myristoylation sites (SRMS) kinase. By combining network reconstruction and signal propagation, comprehensive signaling networks from large-scale experimental data are built and multiple molecular paths from these kinases to their targets are extracted. Specific paths from two distinct PIK3CA mutants are retrieved, and their differential impact on the HER3 receptor kinase is explained. In addition, to address the missing connectivities of the SRMS kinase to its targets in interaction pathway databases, phospho-tyrosine and phospho-serine/threonine proteomic data are integrated. The resulting SRMS-signaling network comprises casein kinase 2, thereby validating its currently suggested role downstream of SRMS. The computational pipeline is publicly available, and contains a user-friendly graphical interface (http://doi.org/10.5281/zenodo.3333687).

Drosophila melanogaster as a Model System for Human Glioblastomas.

Glioblastoma multiforme (GBM) is the most common primary malignant adult brain tumor. Genomic amplifications, activating mutations, and overexpression of receptor tyrosine kinases (RTKs) such as EGFR, and genes in core RTK signaling transduction pathways such as PI3K are common in GBM. However, efforts to target these pathways have been largely unsuccessful in the clinic, and the median survival of GBM patients remains poor at 14-15 months. Therefore, to improve patient outcomes, there must be a concerted effort to elucidate the underlying biology involved in GBM tumorigenesis. Drosophila melanogaster has been a highly effective model for furthering our understanding of GBM tumorigenesis due to a number of experimental advantages it has over traditional mouse models. For example, there exists extensive cellular and genetic homology between humans and Drosophila, and 75% of genes associated with human disease have functional fly orthologs. To take advantage of these traits, we developed a Drosophila GBM model with constitutively active variants of EGFR and PI3K that effectively recapitulated key aspects of GBM disease. Researchers have utilized this model in forward genetic screens and have expanded on its functionality to make a number of important discoveries regarding requirements for key components in GBM tumorigenesis, including genes and pathways involved in extracellular matrix signaling, glycolytic metabolism, invasion/migration, stem cell fate and differentiation, and asymmetric cell division. Drosophila will continue to reveal novel biological pathways and mechanisms involved in gliomagenesis, and this knowledge may contribute to the development of effective treatment strategies to improve patient outcomes.

Targeting the PI3K/STAT3 axis modulates age-related differences in macrophage phenotype in rats with myocardial infarction.

Ageing is associated with impaired repair mechanisms in cardiovascular diseases. Macrophages contribute to cardiac fibrosis after myocardial infarction (MI). The phosphatidyl-inositol-3-kinase (PI3K) pathway has been shown to play a role in cardiac remodelling after MI. It remained unclear whether n-butylidenephthalide, a major component of Angelica sinensis, can attenuate cardiac fibrosis by regulating the PI3K/signal transducer and activator of transcription 3 (STAT3)-mediated macrophage phenotypes in ageing rats after MI. Twenty-four hours after ligation of the left anterior descending artery, young (2-month-old) and ageing (18-month-old) male Wistar rats were treated with either vehicle or n-butylidenephthalide for 4 weeks. There were similar infarct sizes in both age groups. Compared with young rats, ageing rats exhibited significant increased cardiac fibrosis after MI, which can be attenuated after administering n-butylidenephthalide. MI was associated with decreased activities of PI3K and STAT3 in ageing rats compared with young rats. In both age groups, n-butylidenephthalide effectively provided a significant increase of STAT3 phosphorylation, STAT3 activity, STAT3 nuclear translocation, myocardial IL-10 levels and the percentage of M2c macrophage and a decrease of myofibroblast infiltration. The effects of n-butylidenephthalide on increased IL-10 levels were reversed by LY294002 or S3I-201. Furthermore, LY294002 abolished the STAT3 phosphorylation, whereas PI3K activity was not affected following the inhibition of STAT3. In conclusions, the host environment is responsible for ageing-related myofibroblast dysregulation in response to MI which can be improved by administering n-butylidenephthalide via macrophage differentiation towards M2 phenotype by targeting the PI3K/STAT3 axis.

PEDF increases GLUT4-mediated glucose uptake in rat ischemic myocardium via PI3K/AKT pathway in a PEDFR-dependent manner.

BACKGROUND: Targeted increase in glucose uptake of ischemic myocardium is a potential therapeutic strategy for myocardial ischemia. PEDF presents a profound moderating effect on glucose metabolism of cells, but its role is still controversial. Here, we try to demonstrate the direct effect of PEDF on glucose uptake in ischemic myocyte and to elucidate its underlying mechanism. METHODS AND RESULTS: Lentivirus vectors carrying PEDF gene were delivered into the myocardium to locally overexpress PEDF in a myocardial ischemia/reperfusion rat model. PET imaging showed that PEDF local overexpression increased [18F]-FDG uptake of ischemic myocardium. In vitro, PEDF directly increased the glucose uptake in hypoxic cardiomyocytes. The expression of glucose transporter 4 (GLUT4) on plasma membrane of hypoxic cardiomyocytes was significantly upregulated by PEDF, but its total amount was not changed. The increased glucose uptake and cardioprotective effects induced by PEDF were blocked by the GLUT4 inhibitor indinavir. PEDF-mediated GLUT4 translocation and glucose uptake increase in hypoxic cardiomyocytes were prevented by phosphatidyl-inositol-3 kinase (PI3K) inhibitor or AKT inhibitor. The PEDF-mediated glucose uptake was also diminished when PEDF receptor (PEDFR) was downregulated or potent phospholipase A2 enzymatic activity was inhibited. CONCLUSIONS: PEDF can increase glucose uptake in ischemic myocardium through a PEDFR-dependent mechanism, involving PI3K/AKT signaling and GLUT4 translocation.

PI3k and Stat3: Oncogenes that are Required for Gap Junctional, Intercellular Communication.

Gap junctional, intercellular communication (GJIC) is interrupted in cells transformed by oncogenes such as activated Src. The Src effector, Ras, is required for this effect, so that Ras inhibition restores GJIC in Src-transformed cells. Interestingly, the inhibition of the Src effector phosphatidyl-inositol-3 kinase (PI3k) or Signal Transducer and Activator of Transcription-3 (Stat3) pathways does not restore GJIC. In the contrary, inhibition of PI3k or Stat3 in non-transformed rodent fibroblasts or epithelial cells or certain human lung carcinoma lines with extensive GJIC inhibits communication, while mutational activation of PI3k or Stat3 increases GJIC. Therefore, it appears that oncogenes such as activated Src have a dual role upon GJIC; acting as inhibitors of communication through the Ras pathway, and as activators through activation of PI3k or Stat3. In the presence of high Src activity the inhibitory functions prevail so that the net effect is gap junction closure. PI3k and Stat3 constitute potent survival signals, so that their inhibition in non-transformed cells triggers apoptosis which, in turn, has been independently demonstrated to suppress GJIC. The interruption of gap junctional communication would confine the apoptotic event to single cells and this might be essential for the maintenance of tissue integrity. We hypothesize that the GJIC activation by PI3k or Stat3 may be linked to their survival function.

Vanadate inhibits transcription of the rat insulin receptor gene via a proximal sequence of the 5'flanking region.

Vanadate, a protein tyrosine phosphatase inhibitor which elicits insulin-like effects, has previously been shown to inhibit expression of the insulin receptor gene at the transcriptional level in rat hepatoma cells. In an attempt to identify the DNA sequence and transcription factors potentially involved in this effect, a fragment of the proximal 5'flanking region of the IR gene (-1143/-252 upstream the ATG codon) has been cloned and functionally characterized. RNase protection allowed the identification of several transcription start sites in the conserved region of the gene, among which two major sites at -455 and -396. Upon fusion to the luciferase gene and transient transfection into hepatoma cells, the -1143/-252 fragment showed promoter activity. This was unaffected by deletion of the -1143/-761 sequence, but markedly decreased (90%) by additional deletion of the -760/-465 sequence. Treatment of hepatoma cells with vanadate led to a dose-dependent decrease in promoter activity of the 1143/-252, -760/-252 and -464/-252 constructs (change relative to untreated cells, 40, 55 and 23% at 125 µM, and 70, 85 and 62% at 250 µM, respectively). These data suggest that although the entire DNA sequence upstream the transcription start sites is probably involved in vanadate-induced inhibition, the short sequence downstream of position -464 and is sufficient for inhibition. Potential targets of vanadate are the transcription factors FoxO1 and HMGA1, two downstream targets of the insulin signaling pathway which have been shown to mediate the inhibitory effect of insulin on IR gene expression.

Pharmacological Inhibition of Casein Kinase 2 Enhances the Effectiveness of PI3K Inhibition in Colon Cancer Cells.

BACKGROUND/AIM: Serine/threonine kinase B-Raf proto-oncogene (BRAF) mutant colon cancer has a poor prognosis and there is an absence of targeted treatments for this subtype. Here, we investigated the effects of inhibition of casein kinase 2 (CK2) on the inhibitory effects of BRAF and phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K) inhibition in BRAF-mutant colon cancer cells. MATERIALS AND METHODS: Colon cancer cell lines with mutations in components of the mitogen-activated protein kinase (MAPK) and PI3K signalling pathway were used. Cell viability was determined after exposure to single agent and combinations of erlotinib (EGFR inhibitor), dabrafenib (MEK inhibitor), GDC0941 (PI3K inhibitor) and CX4945 (CK2 inhibitor). Western blots were used to examine MAPK and AKT serine/threonine kinase (AKT) pathway activation. RESULTS: Addition of CX4945 to dabrafenib did not enhance the antiproliferative effects of single-agent dabrafenib. Use of GDC0941 alone was highly effective in controlling growth of both BRAF-mutant and wild-type cells and this effect was enhanced by CK2 inhibition. CONCLUSION: Inhibition of the PI3K/AKT pathway is central to regulating growth of colon cancer cells and this can be enhanced by CK2 inhibition.