Inhibiting PTEN Expression Research Articles

MicroRNA-21 induces cisplatin resistance in head and neck squamous cell carcinoma.

Drug resistance, either intrinsic or acquired, can impair treatment effects and result in increased cell motility and death. MicroRNA-21 (miR-21), a proto-oncogene, may facilitate the development or maintenance of drug resistance in cancer cells. Restoring drug sensitivity can improve therapeutic strategies, a possibility that requires functional evaluation and mechanistic exploration. For miR-21 detection, matched tissue samples from 30 head and neck squamous cell carcinoma (HNSCC) patients and 8 head and neck cancer (HNC) cell lines were obtained. Reverse transcription-PCR to detect expression, MTT and clonogenic assays to evaluate cell proliferation, apoptosis assays, resazurin cell viability assays, western blot and luciferase reporter assays to detect protein expression, and flow cytometry to analyse the cell cycle were adopted. Compared to the corresponding normal control (NC) tissues, 25 cancer tissues had miR-21 upregulation among the 30 matched pair tissues (25/30, 83.8%); furthermore, among the 8 HNC cell lines, miR-21 expression that was notably upregulated in three: UPCI-4B, UMSCC-1, and UPCI-15B. In both the UMSCC-1 and UPCI-4B cell lines, the miR-21 mimic enhanced cell proliferation with reduced apoptosis and increased viability, whereas the miR-21 inhibitor resulted in the opposite effects (all P<0.001); additionally, miR-21 directly targeted the tumour suppressor phosphatase and tensin homologue (PTEN) and inhibited PTEN expression. Furthermore, the miR-21 mimic induced cisplatin resistance, while the miR-21 inhibitor restored cisplatin sensitivity. Overexpression of miR-21 can enhance cell proliferation, reduce apoptosis, and induce drug resistance by inhibiting PTEN expression. Targeting miR-21 may facilitate cancer diagnosis, restore drug sensitivity, and improve therapeutic effects.

Melatonin promotes apoptosis of thyroid cancer cells via regulating the signaling of microRNA-21 (miR-21) and microRNA-30e (miR-30e)

ABSTRACT Melatonin (MEL) is an effective therapeutic choice for thyroid cancer treatment. In this study, we aimed to explored the potential effect of MEL upon the drug sensitivity of cancer cells and the according underlying mechanisms. Thyroid cancer mice were established as a control group and a MEL group to observe the in vivo effect of MEL. Tumor size and weight in nude mice were detected to evaluate the effect of MEL on tumor growth. Immunohistochemistry assay (IHC) and Western blot were performed to analyze the expression of PTEN protein in tumor cells or tumor cells. After 32 days of cancer cell implantation, MEL was found to significantly repress tumor growth in nude mice approximately by half. Moreover, MEL also suppressed tumor cell proliferation, while apparently activating the apoptosis of tumor cells. In addition, hydrogen sulfide (H2S) production was obviously elevated by MEL treatment. Mechanistically, the expression of phosphatase and tensin homolog (PTEN) was remarkably activated by MEL treatment in tumor tissues of implanted TPC-1 and BCPaP cells in nude mice. Meanwhile, MEL inhibited the expression of miR-21 and miR-30e and promoted the expression of lncRNA-cancer susceptibility candidate 7 (CASC7). Both miR-21 and miR-30e could suppress PTEN expression, while miR-21 could also inhibit the expression of lncRNA-CASC7. In conclusion, the results demonstrated that the MEL administration could downregulate the expression of miR-21 and miR-30e, which resulted in increased expression of PTEN, a pro-apoptotic tumor suppressor, to promote the apoptosis of thyroid cancer cells.

Congenital microtia patients: the genetically engineered exosomes released from porous gelatin methacryloyl hydrogel for downstream small RNA profiling, functional modulation of microtia chondrocytes and tissue-engineered ear cartilage regeneration

BackgroundMesenchymal stem cells (MSCs) exosomes were previously shown to be effective in articular cartilage repairing. However, whether MSCs exosomes promote mature cartilage formation of microtia chondrocytes and the underlying mechanism of action remains unknown. Additionally, some hurdles, such as the low yield and unsatisfactory therapeutic effects of natural exosomes have emerged when considering the translation of exosomes-therapeutics to clinical practices or industrial production. Herein, we investigated the roles of human adipose-derived stem cells (ADSCs) exosomes in modulating microtia chondrocytes and the underlying mechanism of action. Special attention was also paid to the mass production and functional modification of ADSCs exosomes.ResultsWe firstly used porous gelatin methacryloyl (Porous Gelma) hydrogel with pores size of 100 to 200 μm for 3D culture of passage 2, 4 and 6 ADSCs (P2, P4 and P6 ADSCs, respectively), and obtained their corresponding exosomes (Exo 2, Exo 4 and Exo 6, respectively). In vitro results showed Exo 2 outperformed both Exo 4 and Exo 6 in enhancing cell proliferation and attenuating apoptosis. However, both Exo 4 and Exo 6 promoted chondrogenesis more than Exo 2 did. Small RNA sequencing results indicated Exo 4 was similar to Exo 6 in small RNA profiles and consistently upregulated PI3K/AKT/mTOR signaling pathway. Notably, we found hsa-miR-23a-3p was highly expressed in Exo 4 and Exo 6 compared to Exo 2, and they modulated microtia chondrocytes by transferring hsa-miR-23a-3p to suppress PTEN expression, and consequently to activate PI3K/AKT/mTOR signaling pathway. Then, we designed genetically engineered exosomes by directly transfecting agomir-23a-3p into parent P4 ADSCs and isolated hsa-miR-23a-3p-rich exosomes for optimizing favorable effects on cell viability and new cartilage formation. Subsequently, we applied the engineered exosomes to in vitro and in vivo tissue-engineered cartilage culture and consistently found that the engineered exosomes could enhance cell proliferation, attenuate apoptosis and promote cartilage regeneration.ConclusionsTaken together, the porous Gelma hydrogel could be applied to exosomes mass production, and functional modification could be achieved by selecting P4 ADSCs as parent cells and genetically modifying ADSCs. Our engineered exosomes are a promising candidate for tissue-engineered ear cartilage regeneration.Graphical

Resveratrol inhibits the expression of RYR2 and is a potential treatment for pancreatic cancer.

Resveratrol is a polyphonous natural compound that has cardioprotective, anticancer, and anti-inflammatory properties. Studies have proved that resveratrol (RES) inhibits cancer cell proliferation, migration, and invasion and promotes apoptosis. Elevated expression of ryanodine receptor type 2 (RYR2) may participate in the pathway responsible for calcium metabolism as well as anti-apoptosis and anti-autophagy events in malignant tumor cells. However, the underlying molecular mechanisms of RES anticancer effects with RYR2 are not completely understood in pancreatic cancer. The aim of the present study was tantamount to study the effect of RES in human pancreatic cancer and investigate the underlying mechanisms of RES. We found that RES inhibits proliferation, migration, and invasion and suppresses RYR2 expression in pancreatic cancer cells. In addition, RYR2 knockdown impedes the proliferation, migration, and invasiveness of pancreatic cancer cells. RYR2 knockdown can also increase PTEN expression, while increased RYR2 expression can inhibit PTEN expression. Moreover, RES can upregulate PTEN expression. Taken together, these results indicate that RES could play an antitumor role by decreasing RYR2 expression.

MiR-129-5p restores cardiac function in rats with chronic heart failure by targeting the E3 ubiquitin ligase Smurf1 and promoting PTEN expression

ABSTRACT Chronic heart failure (CHF) is a prevalent health concern with complex pathogenesis. This current study set out to estimate the function of the miR-129-5p/Smurf1/PTEN axis on cardiac function injury in CHF. The model of CHF in rats was established. The cardiac function indexes, myocardial tissue damage, and oxidative stress-related factors in CHF rats were evaluated after the interference of Smurf1/miR-129-5p/PTEN. The targeting relationships between miR-129-5p and Smurf1 and between PTEN and Smurf1 were verified. It was found that that after modeling, cardiac functions were impaired, heart/left ventricular/lung weight and the myocardial structure was destroyed, and the degree of fibrosis of myocardial tissue was increased. After Smurf1 knockdown, the cardiac function, myocardial structure, and oxidative stress were improved, and the fibrosis in myocardial tissue was decreased. Smurf1 was a target of miR-129-5p. miR-129-5p could annul the protective effect of Smurf1 silencing on CHF rats. Smurf1 inhibited PTEN expression by promoting PTEN ubiquitination, while miR-129-5p enhanced PTEN expression by inhibiting Smurf1. Meanwhile, overexpression of PTEN annulled the cardiac dysfunction in CHF rats induced by Smurf1. In conclusion, miR-129-5p targeted Smurf1 and repressed the ubiquitination of PTEN, and promoted PTEN expression, thus improving the cardiac function of CHF rats.

EVALUATION OF PTEN AND PI3K/AKT EXPRESSIONS IN STANOZOLOL-TREATED RAT KIDNEYS

Objective: Stanozolol is an anabolic androgenic steroid (AAS) that is widely used by teenagers and athletes in bodybuilding, sports, and athletics. The potential effects of stanozolol in kidney functions have not been defined. In this study we investigated the expression of tumor suppressor protein phosphatase and tensin homolog (Pten) and mRNA levels of phosphatidylinositol 4,5-bisphosphate 3 kinase (Pi3k) and the protein kinase B (Akt1) signaling pathway in rat kidneys treated by stanozolol. Material and Method: Rats were randomized to 5 groups as control, solvent control, steroid (stanozolol), solvent-exercise, and steroid-exercise. Subcutaneous injection of stanozolol (5 mg/kg) was applied for 28 days and swimming exercise was performed 20 min/day, 5 days/week in exercise groups. Expression of PTEN was evaluated by immunohistochemistry. Also, Pten, Pi3k, and Akt1 mRNA expressions were analyzed via RT-PCR. Results: Mesangial cells and renal tubules in the control, solvent control, and solvent exercise groups showed strong (+++) PTEN reactivity against weak PTEN reactivity in the steroid group. Moderate PTEN reactivity was detected in cells of the steroid exercise group. Pten mRNA expression was significantly decreased, and Pi3k and Akt1 mRNA expression were significantly increased in the steroid group versus other groups (p&lt;0.001). Pten expression showed increase while Pi3k and Akt1 expression showed decrease with exercise treatment (p&lt;0.05). Conclusion: Our findings suggest that AAS usage may inhibit PTEN expression in kidneys, which can be associated with increased Pi3k and Akt1 mRNA levels. Exercise performed with AAS usage can be protective on stanozolol-exposed kidneys due to increased levels of PTEN expression and decreased levels of Pi3k and Akt1.

EVALUATION OF PTEN AND PI3K/AKT EXPRESSIONS IN STANOZOLOL-TREATED RAT KIDNEYS

Objective: Stanozolol is an anabolic androgenic steroid (AAS) that is widely used by teenagers and athletes in bodybuilding, sports, and athletics. The potential effects of stanozolol in kidney functions have not been defined. In this study we investigated the expression of tumor suppressor protein phosphatase and tensin homolog (Pten) and mRNA levels of phosphatidylinositol 4,5-bisphosphate 3 kinase (Pi3k) and the protein kinase B (Akt1) signaling pathway in rat kidneys treated by stanozolol. Material and Method: Rats were randomized to 5 groups as control, solvent control, steroid (stanozolol), solvent-exercise, and steroid-exercise. Subcutaneous injection of stanozolol (5 mg/kg) was applied for 28 days and swimming exercise was performed 20 min/day, 5 days/week in exercise groups. Expression of PTEN was evaluated by immunohistochemistry. Also, Pten, Pi3k, and Akt1 mRNA expressions were analyzed via RT-PCR. Results: Mesangial cells and renal tubules in the control, solvent control, and solvent exercise groups showed strong (+++) PTEN reactivity against weak PTEN reactivity in the steroid group. Moderate PTEN reactivity was detected in cells of the steroid exercise group. Pten mRNA expression was significantly decreased, and Pi3k and Akt1 mRNA expression were significantly increased in the steroid group versus other groups (p&lt;0.001). Pten expression showed increase while Pi3k and Akt1 expression showed decrease with exercise treatment (p&lt;0.05). Conclusion: Our findings suggest that AAS usage may inhibit PTEN expression in kidneys, which can be associated with increased Pi3k and Akt1 mRNA levels. Exercise performed with AAS usage can be protective on stanozolol-exposed kidneys due to increased levels of PTEN expression and decreased levels of Pi3k and Akt1.

MicroRNA-195-3p promotes hepatic stellate cell activation and liver fibrosis by suppressing PTEN expression

Liver fibrosis is a reversible wound healing reaction characterized by abnormal accumulation of extracellular matrix (ECM) in response to liver injury. Recent studies have shown that it can be epigenetically regulated, especially by microRNAs (miRNAs). It has been acknowledged that activation of hepatic stellate cells (HSCs) is a pivotal step in the initiation and progression of liver fibrosis. Notably, our results showed that miR-195-3p was increased in HSCs isolated from CCl4-treated mice and that the increase was more pronounced as the degree of liver fibrosis increased. Moreover, treatment of LX-2 cells, a human immortalized hepatic stellate cell line, with TGF-β1 resulted remarkable upregulation of miR-195-3p. Gain-of-function and loss-of-function experiments have suggested that the increased levels of miR-195-3p inhibit the expression of phosphatase and tension homolog deleted on chromosome 10 (PTEN), a negative regulator of the PI3K/Akt/mTOR signaling pathway in liver fibrosis, thereby contributing to HSC activation and proliferation and promoting the expression of profibrotic genes, such as α-SMA and collagen I, in LX-2 cells, which accelerates the accumulation of fibrous extracellular matrix deposition in the liver, while knockdown of miR-195-3p induced the opposite effect. Taken together, these results provide evidence for the harmful role of miR-195-3p in CCl4-treated mouse liver fibrosis.

MiR-144-3p induced by SP1 promotes IL-1β-induced pyroptosis in chondrocytes via PTEN/PINK1/Parkin axis

Rheumatoid arthritis (RA) often leads to functional disabilities and deformities. MiRNA plays a vital role in cell pyroptosis. Nevertheless, the function and underlying mechanism of miR-144-3p in pyroptosis during the progression of RA remains unclear. In this study, N1511 cells were stimulated with IL-1β to construct a RA model. 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay was performed to assess the cell viability. Cell pyroptosis was detected by flow cytometry. The levels of inflammatory cytokines (TNF-α, IL-6, and IL-18) were assessed by enzyme-linked immunosorbent assay (ELISA). The relationship among specific protein 1 (SP1), microRNA-144-3p (miR-144-3p), and phosphatase and tensin homolog (PTEN) was explored by dual-luciferase reporter assay, RNA immunoprecipitation (RIP), and chromatin immunoprecipitation (ChIP), respectively. The level of miR-144-3p in N1511 cells was upregulated by IL-1β. MiR-144-3p knockdown inhibited IL-1β-induced pyroptosis in N1511 cells, and the expressions of NOD-like receptor family pyrin domain containing 3 (NLRP3), Cleaved caspase-1, Gasdermin D (GSDMD), and Cleaved caspase-3 in IL-1β-stimulated N1511 cells were increased. The levels of inflammatory cytokines in N1511 cells were increased by IL-1β, which were restored by miR-144-3p knockdown. MiR-144-3p knockdown abolished IL-1β-induced inactivation of putative kinase 1 (PINK1)/Parkin RBR E3 ubiquitin-protein (Parkin) signalling. Moreover, transcription factor SP1 could upregulate miR-144-3p expression and miR-144-3p negatively regulated PTEN expression. In summary, MiR-144-3p induced by SP1 could promote IL-1β-induced chondrocyte pyroptosis via inhibiting PTEN expression and suppressing the activation of PINK1/Parkin signalling, which provided a new strategy against RA.

Mechanism of IDH1-R132H mutation in T cell acute lymphoblastic leukemia mouse model via the Notch1 pathway

T-cell acute lymphoblastic leukemia (T-ALL) is a clonal malignant disease. Isocitrate Dehydrogenase 1-R123 (IDH1-R132 H) is related to T-ALL progression. This study explored the role of IDH1-R132H in T-ALL. Molt-4 cells with IDH1-R132H mutation were constructed by retroviral transfection of IDH1-R132H and T-ALL xenotransplantation mouse model was established by injection of Molt-4 cells through the tail vein. Infiltration of the liver, spleen, and bone marrow and the percentage of CD45-positive T-ALL cells in them were detected. Cell proliferation, apoptosis, and invasion were evaluated after the intervention of Notch1, PTEN, or PI3K expression. The leukocyte number was increased, the spleen was enlarged, infiltration in bone marrow, spleen, and liver tissue was worsened and the percentage of hCD45-positive T-ALL cells was increased by IDH1-R132H mutation, which promoted T-ALL deterioration. IDH1-R132H mutation promoted proliferation, invasion, and inhibited apoptosis of T-ALL cells, which were reversed by inhibition of Notch1. IDH1-R132H mutation upregulated HES1 expression and downregulated PTEN expression by activating the Notch1 pathway, while inhibition of Notch1 reversed these changes. PTEN inhibited the PI3K/AKT pathway activation. PTEN overexpression reversed IDH1-R132H mutation effect on promoting malignant behaviors of T-ALL cells. IDH1-R132H mutation inhibited PTEN expression by activating the Notch1/HES1 pathway, activated the PI3K/AKT pathway, thus promoting malignant behaviors of T-ALL cells.

Cooperative miRNA-dependent PTEN regulation drives resistance to BTK inhibition in B-cell lymphoid malignancies

Aberrant microRNA (miR) expression plays an important role in pathogenesis of different types of cancers, including B-cell lymphoid malignancies and in the development of chemo-sensitivity or -resistance in chronic lymphocytic leukemia (CLL) as well as diffuse large B-cell lymphoma (DLBCL). Ibrutinib is a first-in class, oral, covalent Bruton’s tyrosine kinase (BTK) inhibitor (BTKi) that has shown impressive clinical activity, yet many ibrutinib-treated patients relapse or develop resistance over time. We have reported that acquired resistance to ibrutinib is associated with downregulation of tumor suppressor protein PTEN and activation of the PI3K/AKT pathway. Yet how PTEN mediates chemoresistance in B-cell malignancies is not clear. We now show that the BTKi ibrutinib and a second-generation compound, acalabrutinib downregulate miRNAs located in the 14q32 miRNA cluster region, including miR-494, miR-495, and miR-543. BTKi-resistant CLL and DLBCL cells had striking overexpression of miR-494, miR-495, miR-543, and reduced PTEN expression, indicating further regulation of the PI3K/AKT/mTOR pathway in acquired BTKi resistance. Additionally, unlike ibrutinib-sensitive CLL patient samples, those with resistance to ibrutinib treatment, demonstrated upregulation of 14q32 cluster miRNAs, including miR-494, miR-495, and miR-543 and decreased pten mRNA expression. Luciferase reporter gene assay showed that miR-494 directly targeted and suppressed PTEN expression by recognizing two conserved binding sites in the PTEN 3′-UTR, and subsequently activated AKTSer473. Importantly, overexpression of a miR-494 mimic abrogated both PTEN mRNA and protein levels, further indicating regulation of apoptosis by PTEN/AKT/mTOR. Conversely, overexpression of a miR-494 inhibitor in BTKi-resistant cells restored PTEN mRNA and protein levels, thereby sensitizing cells to BTKi-induced apoptosis. Inhibition of miR-494 and miR-495 sensitized cells by cooperative targeting of pten, with additional miRNAs in the 14q32 cluster that target pten able to contribute to its regulation. Therefore, targeting 14q32 cluster miRNAs may have therapeutic value in acquired BTK-resistant patients via regulation of the PTEN/AKT/mTOR signaling axis.

LncRNA NEAT1 aggravates sepsis-induced lung injury by regulating the miR-27a/PTEN axis

Sepsis is an acute inflammatory reaction and a cause of acute respiratory distress syndrome (ARDS). In the present study, we explored the roles and underlying mechanism of the lncRNA Nuclear enriched abundant transcript 1 (NEAT1) in ARDS. The expression levels of genes, proteins and pro-inflammatory cytokines in patients with ARDS, LPS-stimulated cells and septic mouse models were quantified using qPCR, western blotting and ELISA assays, respectively. The molecular targeting relationship was validated by conducting a dual-luciferase reporter assay. Cell proliferation was assessed using the Cell Counting Kit-8 (CCK-8) assay. The cell cycle phase was determined by flow cytometry assay. The expression levels of NEAT1 and pro-inflammatory cytokines were higher in patients with ARDS and septic models than in controls. Knockdown of NEAT1 significantly increased cell proliferation and cycle progression and prolonged mouse survival in vitro and in vivo. Mechanistically, miR-27a was identified as a downstream target of NEAT1 and directly inhibited PTEN expression. Further rescue experiments revealed that inhibition of miR-27a impeded the promoting effects of NEAT1 silence on cell proliferation and cycle progression, whereas inhibition of PTEN markedly weakened the inhibitory effects of NEAT1 overexpression on cell proliferation and cycle progression. Altogether, our study revealed that NEAT1 plays a promoting role in the progression of ARDS via the NEAT1/miR-27a/PTEN regulatory network, providing new insight into the pathologic mechanism behind ARDS.Sepsis-induced acute lung injury has been clinically defined as acute respiratory distress syndrome (ARDS). In the present study, the authors reveal that NEAT1 plays a role in the inflammation and cell cycle progression of ARDS via the NEAT1/miR-27a/PTEN regulatory network, providing new insight into the pathologic mechanism behind ARDS.

MicroRNA-29b reduces myocardial ischemia-reperfusion injury in rats via down-regulating PTEN and activating the Akt/eNOS signaling pathway.

Reperfusion may cause injuries to the myocardium in ischemia situation, which is called ischemia/reperfusion (I/R) injury. The study aimed to explore the roles of microRNA-29b (miR-29b) in myocardial I/R injury. Myocardial I/R injury rat model was established. Differentially expressed miRNAs between the model rats and the sham-operated rats were analyzed. miR-29b expression in myocardial tissues was measured. Gain-of-function of miR-29b was performed, and then the morphological changes, infarct size, myocardial function, oxidative stress, and the cell apoptosis in myocardial tissues were detected. The target relation between miR-29b and PTEN was detected through bio-information prediction and dual luciferase reporter gene assay. Activation of Akt/eNOS signaling was detected. H9C2 cells were subjected to hypoxia/reoxygenation treatment to perform in vitro experiments. I/R rats presented severe inflammatory infiltration, increased infarct size and cell apoptosis, increased oxidative stress and decreased myocardial function. miR-29b was downregulated in I/R rats, and up-regulation of miR-29b reversed the above changes. miR-29b directly bound to PTEN, and overexpression of miR-29b reduced PTEN expression level and increased the protein levels of p-Akt/Akt and p-eNOS/eNOS. In vivo results were confirmed in in vitro experiments. This study provided evidence that miR-29b could alleviate the myocardial I/R injury in vivo and in vitro by inhibiting PTEN expression and activating the Akt/eNOS signaling pathway.

Bone marrow-mesenchymal stem cell-derived exosomal microRNA-141 targets PTEN and activates β-catenin to alleviate myocardial injury in septic mice

Background Mesenchymal stem cells (MSCs) and their derived exosomes have shown potentials in the control of myocardial dysfunction. This study aimed to reveal the function of bone marrow (BM)-MSC-derived exosomes in sepsis-induced myocardial injury and the molecular mechanism. Methods BM-MSC-derived exosomes were obtained and identified. A mouse model with sepsis was induced by cecalligation puncture (CLP) and treated with exosomes. The myocardial function of mice, the production of creatine kinase MB (CK-MB) and lactate dehydrogenase (LDH) in serum, the phosphorylation of a key myocardial contractility-related protein phospholamban (PLB), and the pathological changes in the myocardial tissues were examined. A microRNA (miRNA) microarray analysis was performed to examine the candidate miRNAs carried by the exosomes. Rescue experiments were conducted to validate the involvement of miR-141. Results CLP treatment led to sepsis and notably reduced the myocardial function in mice. Further treatment of BM-MSC-derived exosomes alleviated the CLP-induced myocardial impairment, production of CK-MB and LDH, and inflammatory infiltration and cell apoptosis in mouse myocardial tissues, and restored the PLB phosphorylation. miR-141 was the most upregulated miRNA in the myocardial tissues after exosome treatment. Downregulation of miR-141 blocked the myocardium-protective functions of the exosomes. miR-141 was found to bind to and suppress PTEN expression, which further enhanced the activity of β-catenin. Conclusion This study suggested that BM-MSC derived exosomes ameliorates myocardial injury in septic mice through conveying miRNA-141 and regulating the PTEN/β-catenin axis, and exosomes may serve as promising tools for the management of myocardial injury induced by sepsis or other factors.

Total Glucosides of Paeony Inhibited Autophagy and Improved Acute Kidney Injury Induced by Ischemia-Reperfusion via the lncRNA TUG1/miR-29a/PTEN Axis.

ObjectiveTotal glucosides of paeony (TGP) has been proven to affect anti-inflammatory, immunomodulatory and hypoxia tolerance. This study investigates the effect of TGP on autophagy in acute kidney injury (AKI) induced by ischemia-reperfusion (I/R).MethodsRat model of AKI induced by I/R was established. Rats were administered with TGP at different doses by oral gavage. The contents of BUN, creatinine, NGAL, Kim-1 and IL-18 were detected. The levels of inflammatory factors (TNF-α, IL-1β and IL-6) and autophagy were measured. The expressions of lncRNA TUG1, miR-29a and PTEN were detected and their binding relationships were verified. I/R rat model with overexpressed TUG1 was established to explore the effect of TGP on kidney injury and autophagy. The hypoxia/reoxygenation (HR) model of HK-2 cells and the HR model of HK-2 cells overexpressing TUG1 and low-expressing PTEN were established.ResultsTGP decreased the contents of BUN, creatinine, NGAL, Kim-1 and IL-18, and reduced the levels of inflammatory factors. LncRNA TUG1 and PTEN were downregulated, and miR-29a was upregulated in kidney tissues. The binding relationships between lncRNA TUG1 and miR-29a, and miR-29a and PTEN were confirmed. TGP suppressed PTEN expression via the lncRNA TUG1/miR-29a axis. Overexpressing lncRNA TUG1 attenuated the protective effect of TGP on AKI and autophagy in HK-2 cells. TGP improved cell viability and inhibited the autophagy in HR model of HK-2 cells via lncRNA TUG1/miR-29a/PTEN axis.ConclusionTGP inhibited autophagy and improved AKI induced by I/R via the lncRNA TUG1/miR-29a/PTEN axis.

Interference with lncRNA NEAT1 promotes the proliferation, migration, and invasion of trophoblasts by upregulating miR-411-5p and inhibiting PTEN expression

Background: Preeclampsia (PE) is an idiopathic hypertensive disorder of pregnancy, which is related to abnormal placental villi development. Our previous study has found that lncRNA NEAT1 promotes apoptosis of trophoblasts, but the role of NEAT1 in proliferation, migration, and invasion is still unclear. This study explores the role of NEAT1 in proliferation, migration, and invasion of trophoblasts. Methods: NEAT1 and miR-411-5p levels were detected by quantitative real-time PCR. Colony formation assay detected cell proliferation and transwell assay detected cell migration and invasion. Dual-luciferase reporter assay detected the binding between NEAT1 and miR-411-5p as well as the binding between miR-411-5p and PTEN. RNA pull-down assay detected the combination between NEAT1 and miR-411-5p. Result: NEAT1 was increased and miR-411-5p was reduced in PE patients and human trophoblasts (HTR8/SVneo cells) that were induced with H2O2. Interference with NEAT1 promoted cell proliferation, migration, and invasion, and the miR-411-5p inhibitor reversed the effect of siRNA-NEAT1. The expression of PTEN was promoted in PE patients and HTR8/SVneo cells that were induced with H2O2, while the miR-411-5p mimic inhibited PTEN expression, and the plasmid-mediated PTEN overexpression reversed the effect of the miR-411-5p mimic. Besides, under H2O2 induction, the miR-411-5p mimic promoted cell proliferation, migration, and invasion, and the plasmid-mediated PTEN overexpression reversed the effect of the miR-411-5p mimic. Conclusion: Interference with lncRNA NEAT1 promoted the proliferation, migration, and invasion of trophoblasts and alleviated the development of PE, which was partly mediated by upregulating miR-411-5p and inhibiting PTEN expression.

Targeting c-Jun in A549 Cancer Cells Exhibits Antiangiogenic Activity In Vitro and In Vivo Through Exosome/miRNA-494-3p/PTEN Signal Pathway.

The oncogene c-Jun is activated by Jun N-terminal kinase (JNK). Exosomes are nanometer-sized membrane vesicles released from a variety of cell types, and are essential for cell-to-cell communication. By using specific JNK inhibitor SP600125 or CRISPR/Cas9 to delete c-Jun, we found that exosomes from SP600125-treated A549 cancer cells (Exo-SP) or from c-Jun-KO-A549 cells (Exo-c-Jun-KO) dramatically inhibited tube formation of HUVECs. And the miR-494 levels in SP600125 treated or c-Jun-KO A549 cells, Exo-SP or Exo-c-Jun-KO, and HUVECs treated with Exo-SP or Exo-c-Jun-KO were significantly decreased. Meanwhile, Exo-SP and Exo-c-Jun-KO enhanced expression of phosphatase and tensin homolog deleted on chromosome ten (PTEN). Addition of miR-494 agomir in Exo-c-Jun-KO treated HUVECs inhibited PTEN expression and promoted tube formation, suggesting the target of miR-494 might be PTEN in HUVECs. Moreover, A549 tumor xenograft model and Matrigel plug assay demonstrated that Exo-c-Jun-KO attenuated tumor growth and angiogenesis through reducing miR-494. Taken together, inhibition of c-Jun in A549 cancer cells exhibited antiangiogenic activity in vitro and in vivo through exosome/miRNA-494-3p/PTEN signal pathway.

Effects of microRNA-103 on the Proliferation and Apoptosis of Pancreatic Cancer Cells via Targeting Phosphatase and Tensin Homolog Deleted on Chromosome Ten (PTEN) and Activating Phosphoinositide 3-Kinase/A Serine/Threonine Kinase (PI3K/Akt) Signaling Pathway

Objective: To investigate whether micro ribonucleic acid (miR)-103 affects pancreatic cancer (PaCa) cells via PTEN-activated PI3K/Akt signaling pathway. Methods: Differences in miR-103 expression in 35 pairs of PaCa tissues and cell lines (SW1990 and PATU8988S) were detected by RT-qPCR. miR-103 inhibitor was transfected into PaCa PATU8988S cell followed by analysis of proliferation and apoptosis of PaCa cells by MTT assay and flow cytometry, respectively. Results: MiR-103 exhibited a significantly high expression in PaCa tissues and cell lines (p &lt; 0.05). Besides, the exogenous inhibition of miR-103 expression in PATU8988S cells significantly inhibited cell proliferation and migration but increased apoptosis activity (p &lt; 0.05). According to the prediction of TargetScan biological database, miR-103 could bind PTEN 3′ untranslated region (3′UTR) and miR-103 was confirmed to suppress PTEN expression in a targeted way (p&lt;0.05). Furthermore, down-regulation of PTEN activated PI3K/Akt signaling to affect the proliferation and apoptosis of PaCa cells (p &lt; 0.05 or p &lt;0.01). Conclusion: MiR-103 displays a significantly increased expression in PaCa cells and targets PTEN to activate PI3K/Akt signaling pathway, thus promoting malignant phenotype formation.

Metformin attenuates synergic effect of diabetes mellitus and Helicobacter pylori infection on gastric cancer cells proliferation by suppressing PTEN expression.

It has been reported that CagA of Helicobacter pylori reduced PTEN expression by enhancing its promoter methylation. Furthermore, diabetes mellitus (DM) may also promote the methylation status of PTEN, a tumour suppressor gene in gastric cancer (GC). It is intriguing to explore whether DM may strengthen the tumorigenic effect of H pylori (HP) by promoting the methylation of PTEN promoter and whether the administration of metformin may reduce the risk of GC by suppressing the methylation of PTEN promoter. In this study, we enrolled 107 GC patients and grouped them as HP(−)DM(−) group, HP(+)DM(−) group and HP(+)DM(+) group. Bisulphite sequencing PCR evaluated methylation of PTEN promoter. Quantitative real‐time PCR, immunohistochemistry and Western blot, immunofluorescence, flow cytometry and MTT assay were performed accordingly. DNA methylation of PTEN promoter was synergistically enhanced in HP(+)DM(+) patients, and the expression of PTEN was suppressed in HP(+)DM(+) patients. Cell apoptosis was decreased in HP(+)DM(+) group. Metformin showed an apparent effect on restoring CagA‐induced elevation of PTEN promoter methylation, thus attenuating the PTEN expression. The reduced PTEN level led to increased proliferation and inhibited apoptosis of HGC‐27 cells. In this study, we collected GC tumour tissues from GC patients with or without DM/HP to compare their PTEN methylation and expression while testing the effect of metformin on the methylation of PTEN promoter. In summary, our study suggested that DM could strengthen the tumorigenic effect of HP by promoting the PTEN promoter methylation, while metformin reduces GC risk by suppressing PTEN promoter methylation.

MicroRNA-148a Regulates the Proliferation and Differentiation of Ovine Preadipocytes by Targeting PTEN.

Simple SummaryThe miR-148a has been shown to play an important role in preadipocyte differentiation. Herein, we explored the role of miR-148a in ovine adipocyte development, using Oil Red O staining, CCK-8, EdU, flow cytometry and RT-qPCR. The results showed that miR-148a suppressed proliferation and facilitated the differentiation of preadipocytes. The dual fluorescent reporter vector experiments showed that miR-148a directly targeted PTEN. Meanwhile, we demonstrated that PTEN significantly inhibited the differentiation of preadipocytes. In conclusion, our research provides new insights that miR-148a inhibits ovine preadipocyte proliferation and accelerates differentiation by the negative regulation of PTEN.MicroRNAs (miRNAs) have been found to be involved in lipid deposition and metabolism. However, there have been no reports on the roles of miR-148a in the proliferation and adipogenesis of preadipocytes in sheep. In this study, the expression of miR-148a was profiled in the eight tissues of Tibetan ewes and differentiated preadipocytes, and the role of miR-148a in differentiation and proliferation of ovine preadipocytes was investigated using Oil Red O staining, CCK-8, EdU staining, cell cycle detection, and RT-qPCR. The effect of PTEN on the differentiation of ovine preadipocytes was also investigated. The miR-148a was widely expressed in the eight tissues investigated and had significantly increased expression in liver, spleen and subcutaneous adipose tissues, and the heart. The expression of miR-148a continued to increase with the differentiation of ovine preadipocytes. The over-expression of miR-148a significantly promoted differentiation but inhibited the proliferation of ovine preadipocytes. The inhibition of miR-148a had the opposite effect on the differentiation and proliferation of ovine preadipocytes with over-expressed miR-148a. The results from the dual luciferase reporter assays showed that miR-148a mimic significantly decreased the luciferase activity of PTEN-3′UTR dual luciferase reporter vector, suggesting that PTEN is a target gene of miR-148a. In over-expressed-PTEN preadipocytes, the number of lipid droplets remarkably decreased, and the expression levels of adipogenesis marker genes PPARγ, FASN, FATP4, GLUT4, C/EBPβ and LPL were also significantly down-regulated. These results suggest that miR-148a accelerated the adipogenic differentiation of ovine preadipocytes by inhibiting PTEN expression, and also inhibited the proliferation of ovine preadipocytes.