Phosphorylation Of Phosphoinositide 3-kinase Research Articles

Recombinant human thrombopoietin combined with interleukin-2 improves the effects of chemosensitivity and thrombocytopenia on a basic gemcitabine and carboplatin combination therapy for non-small cell lung cancer in a nude mouse model.

To investigate the effects of recombinant human thrombopoietin (rhTPO) and interleukin-2 (IL-2) on a basic gemcitabine (GEM) plus carboplatin (GC) treatment regimen in a murine lung carcinoma model. Fifty nude mice with subcutaneous tumors derived from human lung cancer cells were divided into 5 groups, each comprised of 10 mice: A blank group (intraperitoneal injection of saline), a control group (GC) (intraperitoneal injections of GC), a rhTPO group (same as the control group plus subcutaneous injection of rhTPO), an IL-2 group (same as the control group plus subcutaneous injection of IL-2) and a rhTPO + IL-2 group (same as the rhTPO group plus subcutaneous injection of IL-2). Tumor development and histology as well as CD4+, phosphorylated-adenosine monophosphate-activated protein kinase (p-AMPK), phosphorylated-protein kinase B (p-AKT), phosphorylated-extracellular signal-regulated kinase (p-ERK), phosphorylated-phosphoinositide 3-kinase (p-pI3K) and GTPase RAS1 expression in tumor tissues were measured and blood analyses performed. Tumor sizes from all treated mice were significant smaller than the controls, as were the tumors of IL-2 plus GC treated mice compared to other treated groups. CD4+ expressing cells were increased in tumors after IL-2 and rhTPO treatment and the application of rhTPO significantly restored the blood platelet count. The expression of p-AMPK, p-AKT, p-ERK, p-pI3K and RAS1 in tumor cells were all significantly diminished after the addition of rhTPO and IL-2 to the GC regimen. The supplementation of rhTPO and IL-2 to a GC regime effectively reduced tumor sizes and restored the platelet count in a human lung cancer mouse model.

Peroxiredoxin 6 regulates the phosphoinositide 3-kinase/AKT pathway to maintain human sperm viability.

Peroxiredoxins (PRDXs) are antioxidant enzymes proven to control the levels of reactive oxygen species (ROS) and to avoid oxidative damage in the spermatozoon. Previously, we have shown that low amounts of PRDXs are associated with male infertility and that PRDX6 is the primary antioxidant defense in human spermatozoa, maintaining survival and DNA integrity (Gong et al., 2012, Fernandez and O'Flaherty, 2018). Oxidative stress can trigger different pathway cascades in the spermatozoa, including truncated apoptosis. It has been reported that the phosphorylation status of phosphoinositide 3-kinase (PI3K) and its target AKT (protein kinase B) prevent the spermatozoon from entering the truncated apoptotic cascade. Here, we aim to study the regulation of the PI3K/AKT pathway by PRDX6 and assess its role in maintaining sperm viability. Human semen samples were obtained over 1year from 20 healthy non-smoking volunteers aged 22-30years. Sperm viability, lipid peroxidation and apoptosis-like changes were determined by flow cytometry while phosphorylation of PI3K and AKT substrates were assessed by immunoblotting using anti-phospho-PI3K and anti-phospho-AKT substrates antibodies. We found that the addition of arachidonic acid and lysophosphatidic acid, products of PRDX6 calcium-independent phospholipase A2 (Ca2+-iPLA2), prevented loss of sperm viability and maintained the phosphorylation of PI3K. Antioxidant compounds such as D-penicillamine partially prevented the oxidative damage on spermatozoa that led to a reduction of their viability. Thus, other pathways can also participate in sperm survival and be regulated by PRDXs. In conclusion, PRDX6 contributes to the regulation of ROS production and the PI3K/AKT pathway for the maintenance of sperm survival.

Kinesin family member 18B: A contributor and facilitator in the proliferation and metastasis of cutaneous melanoma.

Melanoma is the most aggressive type of cutaneous tumor and the occurrence of metastasis makes it resistant to almost all available treatment and becomes incorrigible. Hence, identifying metastasis-related biomarkers and effective therapeutic targets will assist in preventing metastasis and ameliorating cutaneous melanoma. In our present study, we reported kinesin family member 18B (KIF18B) as a novel contributor in cutaneous melanoma proliferation and metastasis, and it was found to be of great significance in predicting the prognosis of cutaneous melanoma patients. Bioinformatics analysis based on ONCOMINE, The Cancer Genome Atlas, and Genotype-Tissue Expression database revealed that KIF18B was highly expressed in cutaneous melanoma and remarkably correlated with unfavorable clinical outcomes. Consistently, the results of the quantitative real-time polymerase chain reaction exhibited that the expression of KIF18B was significantly higher in cutaneous melanoma cell lines than that in normal cells. In vitro, biological assays found that knockdown of KIF18B in cutaneous melanoma cells noticeably repressed cell proliferation, migration, and invasion, while inducing cell apoptosis. Moreover, the protein expression of E-cadherin was enhanced while the expression of N-cadherin, vimentin, and Snail was decreased in M14 cells after knocking down KIF18B. In addition, the phosphorylation of phosphoinositide 3-kinase (PI3K) and extracellular-signal-regulated kinase (ERK) was significantly suppressed in M14 cells with silenced KIF18B. Above all, our results indicated that the repression of cutaneous melanoma cell migration and proliferation caused by KIF18B depletion suggested an oncogenic role of KIF18B in cutaneous melanoma, which acts through modulating epithelial-mesenchymal transition and ERK/PI3K pathway.

The up-regulated lncRNA DLX6-AS1 in colorectal cancer promotes cell proliferation, invasion and migration via modulating PI3K/AKT/mTOR pathway.

Colorectal cancer (CRC) is one of the leading causes of cancer-related deaths around the world. Recently, using the high-throughput techniques, long non-coding RNAs (lncRNAs) have been shown to play an important role in CRC progression. In the present study, we aimed to determine lncRNA DLX6 Antisense RNA 1 (DLX6-AS1) in CRC tissues and cell lines and to investigate the molecular mechanisms of DLX6-AS1 in CRC progression. Quantitative real-time PCR was performed to detect gene expression; cell counting kit-8, colony formation, cell invasion, and migration assays were performed to determine cell proliferation, invasion, and migration, respectively; caspase-3 activity assay kit was used to detect caspase-3 activity; in vivo tumor growth was evaluated in a nude mice xenograft model. DLX6-AS1 was up-regulated in 60 CRC tissues when compared to normal adjacent colorectal tissues, and high expression of DLX6-AS1 was correlated with advanced T stage and distant metastasis in CRC patients. The up-regulation of DLX6-AS1 was further confirmed in CRC cell lines. The gain-of-function assays showed that DLX6-AS1 overexpression promoted HCT116 cell proliferation, invasion, and migration, but inhibited cell apoptosis; while the loss-of-function assays showed that DLX6-AS1 knockdown exerted the opposite effects in SW480 cells. In vivo studies revealed that DLX6-AS1 knockdown suppressed tumor growth in the nude mice xenograft model. In addition, DLX6-AS1 overexpression caused an increase in the phosphorylated phosphoinositide 3-kinase (p-PI3K), p-AKT and p-mammalian target of rapamycin (mTOR) protein levels, and DLX6-AS1 knockdown had the opposite effects. Blockade of PI3K/AKT/mTOR signalling pathway by using mTOR inhibitor partially abolished the enhanced effects of DLX6-AS1 overexpression on CRC cell proliferation and metastasis. In summary, our data indicated that DLX6-AS1 promoted CRC cell proliferation, invasion, and migration but inhibited cell apoptosis via targeting PI3K/AKT/mTOR signalling pathway, suggesting the key role of DLX6-AS1 in CRC progression.

Downregulation of kynureninase restrains cutaneous squamous cell carcinoma proliferation and represses the PI3K/AKT pathway.

The protein kynureninase (KYNU) has recently been reported to participate in the pathological processes of various diseases. To explore the expression and the biological function of KYNU in cutaneous squamous cell carcinoma (cSCC). Expression of KYNU in cSCC cell lines and tissues was firstly evaluated based on the Gene Expression Omnibus and the Oncomine databases. Quantitative reverse transcription-PCR was performed to determine the mRNA expression of KYNU in cSCC cell lines. Small interfering RNA (siRNA) was used for silencing KYNU. The effect of KYNU on the growth and motility of cSCC cells was determined by cell counting kit-8, wound-healing and Transwell assays, and western blotting was used to determine the protein expression of KYNU, AKT, phosphoinositide 3-kinase (PI3K), phosphorylated (p)-AKT and p-PI3K. KYNU was significantly upregulated in cSCC tissues and cell lines. Knockdown of KYNU using siRNA noticeably suppressed the proliferation, migration and invasion ability of SCL-1 cells (P<0.01). Western blotting revealed that phosphorylation of AKT and PI3K was markedly inhibited after silencing KYNU. The ratios of p-AKT/AKT and p-PI3K/PI3K were significantly decreased in the si-KYNU group compared with the control group. Depletion of KYNU could inhibit the growth of cSCC cells, possibly through modulating PI3K/AKT pathway. These data indicate that KYNU takes a key part in the malignant progression of cSCC, and could be considered as a promising therapeutic target for cSCC treatment.

TWEAK promotes hepatic stellate cell migration through activating EGFR/Src and PI3K/AKT pathways.

Activated human hepatic stellate cells (HSCs) showed enhanced ability of migration compared with quiescent HSCs, which is pivotal in liver fibrogenesis. The aim of the present study was to investigate the effects of tumor necrosis factor-like weak inducer of apoptosis (TWEAK) on the migration of activated HSCs and to explore the relevant potential mechanisms. Human HSCs LX-2 cells were cultured with TWEAK. TNFRSF12A-downexpressing lentiviruses were used to infect LX-2 cells. The specific matrix metalloproteinases inhibitor BB94, the Src family kinase inhibitor, Dasatinib, and the specific inhibitor of phosphoinositide 3-kinase (PI3K), LY294002 were used to treat LX-2 cells combined with TWEAK. Cell migration and invasion was tested by the transwell assay. The expression of EGFR/Src, PI3K/AKT, and matrix metallopeptidase 9 (MMP9) was identified by real-time polymerase chain reaction or western blotting. The result showed TWEAK promoted HSC migration and collagen production. BB94 significantly attenuated the migration of LX-2 induced by TWEAK. Dasatinib inhibited the ability of cell migration stimulated by TWEAK. TWEAK upregulated the phosphorylation of epidermal growth factor receptor (EGFR) and Src. The phosphorylation of PI3K and AKT was significantly activated by TWEAK stimulation. Inhibition of PI3K/AKT reduced the expression of MMP9 induced by TWEAK. The present study, for the first time, demonstrated that TWEAK promoted HSC migration through the activation of EGFR/Src and PI3K/AKT pathways, and showed a novel potential mechanism of HSC migration regulated by TWEAK.

Electroacupuncture attenuates chronic fibromyalgia pain through the phosphorylated phosphoinositide 3-kinase signaling pathway in the mouse brain.

Objective(s):Fibromyalgia (FM) is a central nervous system disorder characterized by widespread mechanical hyperalgesia due to unknown mechanisms. Several inflammatory mediators, such as interleukin-1 (IL-1), IL-6, IL-8, and tumor necrosis factor, are increased in the serum of FM patients. Although medications including pregabalin, duloxetine, and milnacipran are used to treat FM, the results are unsatisfying. In the present study we assessed whether electroacupuncture (EA) can reduce chronic FM pain and then proposed an underlying mechanism for this effect. Materials and Methods:Chronic FM pain was induced in mice by dual acid saline injection lasting up to 4 weeks. Results:Chronic FM pain was treated by EA manipulation, but not in the sham operated group. Phosphorylated phosphatidylinositol 3-kinase (pPI3K), protein kinase B, mechanistic target of rapamycin, and nuclear factor kappa-light-chain-enhancer of activated B cells were unaltered in the mouse dorsal root ganglion (DRG) and spinal cord (SC) after inducing FM and administering EA treatment. The pPI3K-associated nociceptive signaling pathway was increased in the thalamus of FM mice, but reversed by EA. Similar results were observed in the mouse somatosensory cortex. Conclusion:These data suggest that EA has a significant effect on a signaling pathway in brain areas of FM mice. These findings suggest the value of EA for clinical practice.

ShRNA-interfering LSD1 inhibits proliferation and invasion of gastric cancer cells via VEGF-C/PI3K/AKT signaling pathway.

BACKGROUNDHistone Lysine Specific Demethylase 1 (LSD1) is the first histone demethylase to be discovered, which regulates various biological functions by making lysine of histone H3K4, H3K9 and non-histone substrates demethylated. Abnormal regulation of LSD1 is closely related to the occurrence and development of gastric cancer. The change of LSD1 expression level plays an important role in the proliferation and metastasis of gastric cancer cells. The study of its function and mechanism may provide a theoretical basis for early diagnosis and targeted therapy of gastric cancer.AIMTo investigate the effect of downregulation of lysine-specific demethylase 1 (LSD1) expression on proliferation and invasion of gastric cancer cells and the possible regulatory mechanisms of the VEGF-C/PI3K/AKT signaling pathway.METHODSThe LSD1-specific short hairpin RNA (shRNA) interference plasmid was transiently transfected, and expression of LSD1 was downregulated. The cell proliferation ability of LSD1 was observed by CCK-8 assay after downregulating expression of LSD1. Transwell invasion assay was used to observe the change of cell invasion ability after downregulating expression of LSD1. Expression of phosphorylated phosphoinositide 3-kinase (p-PI3K), PI3K, p-AKT, AKT, vascular endothelial growth factor receptor (VEGFR)-3, matrix metalloproteinase (MMP)-2 and MMP-9 in each group was detected by Western blotting.RESULTSThe cell proliferation ability of transiently transfected LSD1-shRNA interference plasmid group was significantly lower than that of the control group (P < 0.05). Transwell invasion assay showed that the number of cells across the membrane of the LSD1-shRNA transfection group (238.451 ± 5.216) was significantly lower than that of the control group (49.268 ± 6.984) (P < 0.01). Western blotting showed that expression level of VEGF-C, p-PI3K, PI3K, p-AKT, AKT, VEGFR-3, MMP-2 and MMP-9 in the LSD1-shRNA group was significantly lower than that in the control group (P < 0.05).CONCLUSIONDownregulation of LSD1 expression inhibits metastatic potential of gastric cancer cells, and VEGF-C-mediated activation of PI3K/AKT signaling pathway, which may be an important mechanism for inhibiting lymph node metastasis in gastric cancer cells.

EPO promotes axonal sprouting via upregulating GDF10

Erythropoietin (EPO) has an exact neuroprotective effect on stroke. However, it remains unknown whether it participates in axonal sprouting after neuron damage. Growth and differentiation factor 10 (GDF10) has been shown to be a trigger of axonal sprouting after stroke. Hence, it was hypothesized that EPO promotes axonal sprouting mainly through GDF10. In the present in vitro experiment, it was found that EPO could promote axonal sprouting and GDF10 expression in a dose–dependent manner. The knockdown of GDF10 using siRNA abolished the effect of EPO-mediated axonal sprouting, indicating that GDF10 is the executor of EPO-mediated axonal sprouting. The treatment of neurons with nuclear factor-kappaB (NF-κB) inhibitor JSH-23 could inhibit the accumulation of NF-κB phospho-p65 (p-p65) in the nucleus, the upregualtion of GDF10 and extending of axonal length. Furthermore, the addition of Janus kinase 2 (JAK2) inhibitor CEP-33779 or phosphoinositide 3-kinase (PI3K) inhibitor LY294002 to the culture medium also blocked the nuclear translocation of p-p65, the expression of GDF10, and axonal sprouting, suggesting that EPO induces axonal sprouting via activating cellular JAK2 and PI3K signaling. Impeding JAK2 signaling with CEP-33779 can suppress the phosphorylation of PI3K, and this confirms that the upstream of PI3K signaling is JAK2. These present results provide a novel insight into the role of EPO and the molecular mechanism of axonal sprouting, which is beneficial for the development of novel approaches for neurological recovery after brain injury, including stroke.

Oxymatrine reverses epithelial-mesenchymal transition in breast cancer cells by depressing αⅤβ3 integrin/FAK/PI3K/Akt signaling activation.

PurposeOxymatrine, an alkaloid extracted from the Chinese herb Sophora flavescens Aiton, possesses anti-inflammatory, anti-immune, anti-hepatic fibrosis, and anti-cancer properties. However, the effects of oxymatrine on epithelial-mesenchymal transition (EMT) of breast cancer cells are still unclear.AimThe present study was performed to investigate whether oxymatrine reverses EMT in breast cancer cells and to explore the underlying molecular mechanisms.Materials and methodsMTT assay was performed to evaluate cell viability. Wound-healing assay and transwell chamber assay were used to assess cell migration and invasion, respectively. Immunofluorescence and Western blot were used to study the expression of EMT-related molecules and αⅤβ3 integrin/focal adhesion kinase (FAK)/phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) signaling transduction. Fibronectin, a physiologic ligand of αⅤβ3 integrin, was used to stimulate αⅤβ3 integrin signaling.ResultsOur results demonstrated that oxymatrine effectively suppressed the viability of MDA-MB-231 and 4T1 breast cancer cells, and oxymatrine showed less cytotoxicity on normal breast mammary epithelial MCF-10A cells. In addition, oxymatrine reversed EMT in the MDA-MB-231 and 4T1 cells at nontoxic concentrations. Oxymatrine significantly inhibited cell migration and invasion, downregulated the expression of N-cadherin, vimentin, and Snail in MDA-MB-231 and 4T1 cells, but upregulated the expression of E-cadherin in 4T1 cells. The mechanism revealed that oxymatrine decreased the expression of αⅤ and β3 integrin and their co-localization. It also inhibited αⅤβ3 integrin downstream activation by suppressing the phosphorylation of FAK, PI3K, and Akt. Furthermore, oxymatrine prevented fibronectin-induced EMT and αⅤβ3 integrin/FAK/PI3K/Akt signaling activation.ConclusionOur results revealed that oxymatrine effectively reversed EMT in breast cancer cells by depressing αⅤβ3 integrin/FAK/PI3K/Akt signaling. Thus, oxymatrine could be a potential therapeutic candidate with anti-metastatic potential for the treatment of breast cancer.

Codelivery of Genistein and miRNA-29b to A549 Cells Using Aptamer-Hybrid Nanoparticle Bioconjugates.

This study aimed to evaluate the anti-cancer effect of a combination therapy of miRNA-29b and genistein loaded in mucin-1 (MUC 1)-aptamer functionalized hybrid nanoparticles in non-small cell lung cancer (NSCLC) A549 cell line. Genistein-miRNA-29b-loaded hybrid nanoparticles (GMLHN) was prepared and characterized. Particle size and zeta potential were measured using photon correlation spectroscopy (PCS). Encapsulation efficiency and loading efficiency were determined using HPLC. Preferential internalization of MUC 1-aptamer functionalized GMLHN by A549 cells was evaluated and compared to normal MRC-5 cells. The ability of GMLHN to downregulate targeted oncoproteins Phosphorylated protein kinase, strain AK, Thymoma (Phosphorylated protein kinase B) (pAKT), Phosphorylated phosphoinositide 3-kinase (p-PI3K), DNA (cytosine-5-)-methyltransferase 3 beta (DNMT3B) and Myeloid Cell Leukemia Sequence 1 (MCL 1) was evaluated using western blot, while antiproliferative effect and ability to initiate apoptosis was also assessed in A549 cells. MUC 1-aptamer functionalized GMLHN nanoparticles were prepared. These nanoparticles were preferentially internalized by A549 cells but less so, in MRC-5 cells. pAKT, p-PI3K, DNMT3B and MCL 1 were efficiently downregulated by these nanoparticles without affecting the levels of AKT and PI3K in A549 cells. GMLHN demonstrated a superior antiproliferative effect compared to individual genistein and miRNA-29b-loaded nanoparticles. Results generated were able to demonstrate that genistein-miRNA-29b-loaded hybrid nanoparticles (GMLHN) could be a potential treatment modality for NSCLC because of the ability of the payloads to attack multiple targets.

Alleviated Oxidative Damage by Taraxacum officinale through the Induction of Nrf2-MAPK/PI3K Mediated HO-1 Activation in Murine Macrophages RAW 264.7 Cell Line

Taraxacum officinale has been consumed as a folk remedy due to its diverse physiological activities. This study aimed to investigate the antioxidative potential of T. officinale water extract (TOWE) and ethanol extract (TOEE) against oxidative stress and compare their molecular mechanism via the induction of heme oxygenase-1 (HO-1) in RAW 264.7 cells. The antioxidative activity was evaluated through the radical scavenging assay, the cytoprotection assay against oxidative damage, and Western blot analysis. Both extracts dose-dependently induced HO-1 expression without any cytotoxicity in accordance with the activation of a transcription factor, nuclear factor-erythroid 2 p45-related factor 2 (Nrf2). In addition, TOWE induced HO-1 expression through the phosphorylation of phosphoinositide 3-kinase (PI3K)/Akt and c-Jun NH2-terminal kinase (JNK), while TOEE activated HO-1 by PI3K/Akt phosphorylation. In order to identify the antioxidative potential by HO-1 induction, oxidative damage-caused cell death by tert-butyl hydroperoxide (t-BHP) was significantly attenuated by both extracts. Their antioxidative potential was confirmed by HO-1 selective inducer and inhibitor, cobalt protoporphyrin (CoPP), and tin protoporphyrin (SnPP), respectively. These results indicate that TOWE and TOEE potently alleviated oxidative damage via the induction of Nrf2/MAPK/PI3K mediated HO-1 induction in RAW 264.7 cells.

MiR-125a-5p Inhibits Oxidized Low-Density Lipoprotein-Induced Proliferation and Migration of Vascular Smooth Muscle Cells Through PI3K/AKT Signaling

Certain microRNAs (miRNA/miRs) serve important roles in the progression of atherosclerosis (AS); however, the exact regulatory mechanisms of miRNAs in AS remain to be fully elucidated. The present study aimed to investigate the effects of miR-125a-5p in AS and the underlying mechanisms. Oxidized low-density lipoprotein (ox-LDL) was employed to stimulate human aortic vascular smooth muscle cells (HAVSMCs) to establish a cell model of AS. Reverse transcription-quantitative PCR was used to determine the expression levels of miR-125a-5p. Cell proliferation was evaluated using a Cell Counting Kit-8 (CCK-8) and migration was detected using a Transwell assay. In addition, the levels of the VSMC-specific marker gene α-smooth muscle actin, the cell cycleregulatory proteins cyclin-dependent kinase (CDK)2, cyclin D1, cyclin E and p27, as well as the migration-associated proteins matrix metalloproteinase-2 (MMP2) and MMP9, and phosphorylated phosphoinositide 3-kinase (p-PI3K) and p-AKT were determined by western blot analysis. The results revealed that transfection of miR-125a-5p mimics inhibited the ox-LDL-induced proliferation of HSVSMCs, accompanied by decreased expression of CDK-2, cyclin D1 and cyclin E, and increased expression of p27. Furthermore, miR-125a-5p mimics attenuated the ox-LDL-induced migration of HAVSMCs in parallel with downregulation of the expression of MMP2 and MMP9. Furthermore, the effect of ox-LDL to increase p-PI3K and p-AKT levels was significantly blunted by miR-125a-5p mimics. In conclusion, the present results suggested that miR-125a-5p mimics inhibited ox-LDL-induced proliferation and migration of HAVSMCs through inhibition of PI3K/AKT signaling, providing a potential novel therapeutic strategy for AS.

Long non-coding RNA H19 knockdown inhibits the cell viability and promotes apoptosis of thyroid cancer cells through regulating the PI3K/AKT pathway.

Certain long non-coding (lnc)RNAs have been reported to serve important roles in the genesis and progression of thyroid cancer (TC). Recent studies have demonstrated that the expression of lncRNA H19 is upregulated in TC tissues; however, knowledge of the associated molecular mechanisms is limited. Therefore, the present study aimed to clarify the roles of H19 in TC. The mRNA expression of lncRNA H19 in TC tissues was determined using reverse transcription-quantitative polymerase chain reaction analysis, and the effects of H19 knockdown on cell viability and apoptosis in vitro were assessed using MTT and flow cytometric assays, respectively. Finally, the signaling pathways involved in the effects of H19 were examined. The results indicated that H19 was upregulated in TC tissues. Silencing of H19 inhibited the cell viability and promoted apoptosis of FTC-133 and TPC-1 TC cells, accompanied by an increased expression of B-cell lymphoma 2 (Bcl-2)-associated X protein and caspase 3, and repressed expression of Bcl-2. The results of western blot analysis suggested that the levels of phosphorylated phosphoinositide-3 kinase (PI3K) and phosphorylated AKT were attenuated by H19 silencing. These results suggest that lncRNA H19 exerts an oncogenic function in TC, in part through the PI3K/AKT pathway.

MicroRNA‑214 suppresses the viability, migration and invasion of human colorectal carcinoma cells via targeting transglutaminase 2.

Colorectal carcinoma (CRC) is a common malignancy of the digestive tract. MicroRNA (miR)-214 is considered a key hub that controls tumor networks; therefore, the effects of miR-214 on CRC were examined and its target gene was investigated in this study. The expression levels of transglutaminase 2 (TGM2) and miR-214 were detected in CRC and adjacent normal tissues by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blotting, and luciferase activity was analyzed by dual luciferase reporter analysis. In addition, cell viability, invasion and migration were measured by Cell Counting kit-8 and Transwell assays, respectively. The expression levels of epithelial-mesenchymal transition-related proteins, and phosphoinositide-3 kinase (PI3K)/protein kinase B (Akt) signaling-associated factors were detected using RT-qPCR and western blotting. The results demonstrated that miR-214 expression was downregulated in CRC tissue, whereas TGM2 expression was upregulated. According to TargetScan prediction, miR-214 possesses a binding site to TGM2. In addition, transfection with miR-214 mimics markedly suppressed the viability of LoVo cells. miR-214 overexpression also inhibited cell invasion and migration by increasing E-cadherin and tissue inhibitor of metalloproteinases-2 expression, and decreasing matrix metalloproteinase (MMP)-2 and MMP-9 expression. Furthermore, miR-214 downregulated phosphorylation of PI3K and Akt; however, the expression levels of total PI3K and Akt were not affected by miR-214. In conclusion, this study indicated that TGM2 was a target gene of miR-214, and a negative correlation between miR-214 and TGM2 expression was determined in CRC. Notably, miR-214 markedly suppressed the viability, invasion and migration of CRC cells, which may be associated with a downregulation in PI3K/Akt signaling. These findings suggested that miR-214 may be considered a novel target for the treatment of CRC.

Tempol Protects Against Acetaminophen Induced Acute Hepatotoxicity by Inhibiting Oxidative Stress and Apoptosis.

Acetaminophen (APAP)-induced acute hepatotoxicity is the leading cause of drug-induced acute liver failure. The aim of this study was to evaluate the effects of 4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl (tempol) on the protection of APAP-induced hepatotoxicity in mice. Mice were pretreated with a single dose of tempol (20 mg/kg per day) orally for 7 days. On the seventh day, mice were injected with a single dose of APAP (300 mg/kg) to induce acute hepatotoxicity. Our results showed that tempol treatment markedly improved liver functions with alleviations of histopathological damage induced by APAP. Tempol treatment upregulated levels of antioxidant proteins, including superoxide dismutase, catalase, and glutathione. Also, phosphorylation of phosphoinositide 3-kinase (PI3K) and protein kinase B (Akt) and protein expression of nuclear factor erythroid 2-related factor (Nrf 2) and heme oxygense-1 (HO-1) were all increased by tempol, which indicated tempol protected against APAP-induced hepatotoxicity via the PI3K/Akt/Nrf2 pathway. Moreover, tempol treatment decreased pro-apoptotic protein expressions (cleaved caspase-3 and Bax) and increased anti-apoptotic Bcl-2 in liver, as well as reducing apoptotic cells of TUNEL staining, which suggested apoptotic effects of tempol treatment. Overall, we found that tempol normalizes liver function in APAP-induced acute hepatotoxicity mice via activating PI3K/Akt/Nrf2 pathway, thus enhancing antioxidant response and inhibiting hepatic apoptosis.

Knockdown of lncRNA-HOTAIR downregulates the drug-resistance of breast cancer cells to doxorubicin via the PI3K/AKT/mTOR signaling pathway.

The resistance to chemotherapeutic drugs is a critical feature of breast cancer recurrence and metastasis. Long non-coding RNAs (LncRNAs) serve key roles in tumor drug resistance. LncRNA-HOX transcript antisense RNA (HOTAIR) has been reported to be overexpressed in certain types of cancer and may be closely associated with tumor resistance. The current study aimed to investigate the role of lncRNA-HOTAIR in the regulation of breast cancer resistance to doxorubicin (DOX). A breast cancer cell line (MCF-7) and DOX-resistant breast cancer cell line (DOXR-MCF-7) were utilized in the current study. DOXR-MCF-7 cells were transfected with lncRNA-HOTAIR small interfering RNA (siRNA) and control siRNA. Subsequently, MTT and colony formation assays were performed to assess cell proliferation. Cell apoptosis was also evaluated via flow cytometry. In addition, western blotting and reverse transcription-quantitative polymerase chain reaction were performed to detect the expression of caspase-3, B-cell lymphoma 2, Bcl-2-associated X protein, phosphoinositide 3-kinase (PI3K), protein kinase B (AKT) and mechanistic target of rapamycin (mTOR), and the phosphorylation of PI3K, AKT, and mTOR. The data indicated that lncRNA-HOTAIR silencing decreased cell proliferation and increased apoptosis in MCF-7 and DOXR MCF-7 cells. Furthermore, lncRNA-HOTAIR silencing significantly decreased the phosphorylation of PI3K, AKT and mTOR, indicating that the knockdown of lncRNA-HOTAIR effectively attenuates the resistance of breast cancer cells to DOX by inhibiting the PI3K/AKT/mTOR pathway. In summary, the present study indicated that the knockdown of lncRNA-HOTAIR weakened the resistance of breast cancer cells to DOX via PI3K/AKT/mTOR signaling, suggesting that lncRNA-HOTAIR may be a novel intervention target to reverse DOX-resistance in breast cancer.

Effects of chrysin on the apoptosis in oral squamous carcinoma KB cell line and the underlying mechanisms

To investigate the effect of chrysin on apoptosis of oral squamous carcinoma KB cell line and the possible mechanisms, and to provide new ideas for the treatment of oral cancer. Methods: Oral cancer KB cells were treated with different concentrations of chrysin (1, 2, 4, 8, 16, and 32 μmol/L) for 24 h. Cell proliferation was detected by MMT assay; apoptosis was detected by flow cytometry; the activity of caspase-3/7 was detected by chemiluminescent assay; mitochondrial membrane potential in KB cells was determined by JC-1 assay; and Western blotting was used to determine the activation of protein kinase B (AKT) and phosphoinositide-3-kinase (PI3K). Results: Chrysin inhibited the proliferation of KB cells in a concentration-dependent manner, accompanied by increase in apoptosis of KB cells, activation of caspase-3/7, decrease in mitochondrial membrane potential, and suppression of the phosphorylation of AKT and PI3K. Conclusion: The effect of chrysin on KB cell apoptosis may be related to mitochondrial dysfunction and inhibition of PI3K/AKT pathway.

SIRT4 suppresses the PI3K/Akt/NF‑κB signaling pathway and attenuates HUVEC injury induced by oxLDL.

Atherosclerosis is a chronic and progressive disease. Its morbidity and mortality rates have demonstrated an increase in recent years. The present study aimed to explore the role of sirtuin (SIRT) 4 in the development of atherosclerosis. Alterations in SIRT4 expression in response to oxidized low density lipoprotein (oxLDL) were quantified in human umbilical vein endothelial cells (HUVECs) using western blotting. Cell counting kit‑8 and flow cytometry assays were used in order to explore the effects of SIRT4 on HUVEC proliferation and apoptosis. The effect of SIRT4 on the expression of inflammatory factors in HUVECs was analyzed using ELISA. The expression and phosphorylation of proteins in the phosphoinositide 3‑kinase (PI3K)/protein kinase B (Akt)/nuclear factor (NF)‑κB pathway were comparatively analyzed using western blotting. Nuclear translocation of p65 NF‑κB was examined using immunofluorescence. The present study indicated that oxLDL treatment decreased the expression of SIRT4 in HUVECs in a dose‑ and time‑dependent manner. SIRT4 overexpression promoted oxLDL‑induced HUVEC proliferation and inhibited cell apoptosis. Furthermore, SIRT4 overexpression suppressed the PI3K/Akt/NF‑κB pathway by inhibiting PI3K phosphorylation and phosphorylated (p)‑Akt, p‑nuclear factor of kappa light polypeptide gene enhancer in B‑cells inhibitor α and p‑p65 NF‑κB expression; blocking p65 NF‑κB nuclear translocation and decreasing interleukin (IL)‑1β, IL‑6, and tumor necrosis factor α expression in oxLDL‑induced HUVECs. In conclusion, SIRT4 overexpression enhanced HUVEC survival, suppressed the PI3K/Akt/NF‑κB signaling pathway and inhibited the expression of inflammatory cytokines in oxLDL‑induced HUVECs.

Tetramethylpyrazine exerts a protective effect against injury from acute myocardial ischemia by regulating the PI3K/Akt/GSK-3\u03b2 signaling pathway

ObjectiveWe investigated the protective effect of tetramethylpyrazine (TMP) on injury related to acute myocardial ischemia (AMI) induced by isoproterenol (ISO).Materials and methodsRats were randomly assigned to five groups: control, ISO, ISO + propranolol (10 mg/kg), ISO + TMP (10 mg/kg) and ISO + TMP (20 mg/kg). The rats in the three ISO + groups were pretreated with propranolol or TMP, while the rats in the control and ISO groups were pretreated with an equal volume of saline. Afterwards, the rats in the four administration groups were subcutaneously injected with ISO for two consecutive days. The levels of creatine kinase (CK), lactate dehydrogenase (LDH), superoxide dismutase (SOD), malondialdehyde (MDA), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and IL-1β in the serum were measured using ELISA. The expressions of B-cell lymphoma-associated X-2 (Bax-2), B-cell lymphoma-2 (Bcl-2), phosphoinositide-3-kinase (PI3K), protein kinase B (Akt), glycogen synthase kinase 3β (GSK-3β), MDA5 and SOD1 were determined using western blotting assay. The phosphorylation of PI3K, Akt and GSK-3β were also determined using western blotting assay. The left ventricles of the rats were extracted and stained using hematoxylin and eosin (H&E). The ST segment was recorded using electrocardiograms (ECGs).ResultsAdministration of TMP (10, 20 mg/kg) reduced the levels of MDA and CK and the activities of SOD and LDH in the serum. Pretreatment with TMP significantly reduced the levels of pro-inflammatory cytokines, including IL-1β, IL-6 and TNF-α. Treatment with TMP also improved the histopathological alteration and decreased the ST elevation. Furthermore, TMP ameliorated the expressions of Cu, SOD1, MDA5, Bax-2, Bcl-2, p-PI3K, p-Akt and p-GSK-3β in ISO-induced rats.ConclusionsTetramethylpyrazine protected against injury due to AMI by regulating the PI3K/Akt /GSK-3β signaling pathway.