TP53INP1 Research Articles

Causal Effects of Oxidative Stress on Diabetes Mellitus and Microvascular Complications: Insights Integrating Genome-Wide Mendelian Randomization, DNA Methylation, and Proteome.

Oxidative stress (OS) is involved in the development of diabetes, but the genetic mechanisms are not completely understood. We integrated multi-omics data in order to explore the genetic relations between OS-related genes, diabetes mellitus, and microvascular complications using Mendelian randomization and colocalization analysis. Summary-level data related to OS were acquired from respective studies of methylation, expression, and protein abundance quantitative trait loci. Genetic associations concerning diabetes, diabetic nephropathy (DN), and diabetic retinopathy (DR) were derived from the FinnGen study. Summary-data-based Mendelian randomization (SMR) analysis was conducted to evaluate the correlations between molecular features concerned with OS-related genes and diabetes mellitus, along with its microvascular complications. Additionally, we performed colocalization analysis to determine if the detected signal pairs shared a causal genetic variant. At the genetic level, we identified ten potential causal associations of oxidative stress genes with diabetes, along with microvascular complications, through SMR and colocalization analysis. After integrating the DNA methylation quantitative trait loci (mQTL) and expression QTL (eQTL) data, our analyses revealed a correlation between the methylation site cg26343298 and reduced expression of TP53INP1, supporting the protective role of cg26343298 methylation on type 2 diabetes (T2D) and diabetic nephropathy. Similarly, an inverse association was observed between gene methylation and expression in CHEK1 (cg07110182), confirming the beneficial effect of modification of CHEK1 by cg07110182 in diabetic retinopathy. In addition, upregulation of SUOX expression by cg22580629 was linked to a reduced risk of diabetic retinopathy. At circulating protein levels, genetically predicted a higher level of ICAM1 (OR 1.05, 95%CI 1.03-1.08) was positively connected with the risk of diabetic retinopathy. This SMR study elucidated that the TP53INP1 gene was putatively associated with T2D and DN risk, while the SUOX and CHEK1 genes were associated with DR risk through oxidative stress mechanisms. Additionally, our study showed a positive correlation between the ICAM-1 protein and DR. These findings may enhance our understanding of their pathogenesis and suggest new therapeutic targets for clinical practice.

Designed miR-19a/b sponge induces apoptosis in lung cancer cells through the PI3K-PTEN-Akt pathway regulation.

MicroRNAs (miRNAs) are one of the main factors in cancer development and can alter the activity of proto-oncogenic or tumor suppressor genes. The miR-17-92 cluster, which comprises miR-17, miR-18a, miR-19a/b, miR-20a, and miR-92a, has been identified as a biomarker in a variety of cancer types. Among them, miR-19a/b exerts an oncogenic effect by suppressing tumor suppressor genes, including PTEN and TP53INP1in numerous types of cancers, including NSCLC. An miRNA sponge is an mRNA with multiple repetitive sequences that prevents miRNAs from interacting with their targets, thereby inhibiting their action. In this study, we designed an miR-19a/b sponge plasmid and transfected it into A549 lung cancer cell lines and analyzed its effects on PTEN and TP53INP1 gene expression as the main miR-19a/b target and apoptosis rate in these cell lines. The findings revealed that miR-19a/b sponge significantly increased PTEN and TP53INP1 mRNA expression. The effect of the sponge on TP53INP1 was much greater than that on PTEN. This is because TP53INP1 is directly (sponge effect) and indirectly (AKT pathway is affected by the P53 gene) affected by this sponge. In addition, compared with the control group, the percentage of primary and secondary apoptosis increased significantly (P value < 0.0001).

Tempo Induces Switch Between Apoptosis and Autophagy in Combination with Hyperthermia via Modulation of TP53INP1 Gene

Tempo Induces Switch Between Apoptosis and Autophagy in Combination with Hyperthermia via Modulation of TP53INP1 Gene

EZH2 Gene Silencing Can Affect the Expression of miR-155 and TP53INP1

Many genetic, epigenetic, and cellular studies on cancer are underway today, and the completion of the genetic and epigenetic library of cancer could be the way to treat the disease in the future. In this study, we have investigated the parallel gene expression changes of EZH2 and miR-155. So far no study has examined the role of these two factors simultaneously and the results of this study could be useful for further studies. For this purpose, using specific shRNA, the EZH2 gene of HCT116 cells was downregulated and then the changes in expression of the miR-155 were investigated. For gene expression study, Real-time PCR as a standard quantitative method was used. The findings of this study showed that in HCT116 human colon cancer cells, downregulation of miR-155 using shRNA can reduce EZH2 expression and also can promote a significant increase in the expression of TP53INP1 gene. Based on the results, we can emphasize the interaction between these two genes. Importantly, EZH2 downregulation has been able to decrease the amount of miR-155 that has also increased expression in many types of cancers. It may be of interest in epigenetic treatments of colon cancer, because miR-155 can control a very important tumor suppressor gene, TP53INP1.

MicroRNA-125b mimic inhibits ischemia reperfusion-induced neuroinflammation and aberrant p53 apoptotic signalling activation through targeting TP53INP1

BackgroundIschemia reperfusion (IR) injury affects neuronal function through multiple pathogeneses that induce neuroinflammation and cellular apoptosis. The important roles of microRNAs (miRs) in the regulation of spinal cord IR have been recently reported. Among these roles, we investigated whether miR-125b and its downstream targets regulated the p53 signalling network and participated in both inflammation and apoptosis. MethodsAn IR model was established via 12-min occlusion of the aortic arch. The direct interaction between miR-125b and TP53INP1 was demonstrated by Western blotting and luciferase assays. The cellular distributions of TP53INP1 were visualised by double immunofluorescence labelling. The effects of miR-125b on the expression of TP53INP1, p53 and release of proinflammatory cytokines were evaluated by synthetic miRs. Additionally, the detection of hind-limb motor function in vivo and motor neuronal apoptosis in vitro were evaluated to explore the potential mechanisms. ResultsIR-induced alterations in hind-limb motor function were closely related to the temporal changes in miR-125b and TP53INP1 expression. The miR-125b/TP53INP1 gene pair was confirmed by luciferase assay. Compared with Sham controls, IR treatment resulted in increased TP53INP1 immunoreactivity that was primarily distributed in neurons. Treatment with miR-125b mimic markedly decreased the protein levels of TP53INP1, p53 and cytokines interleukin (IL)-1β and tumour necrosis factor (TNF)-α, whereas miR-125b control or inhibitor did not have the above-mentioned effects. Moreover, miR-125b mimic improved motor function in vivo and attenuated neuronal apoptosis in vitro, as demonstrated by the increased average Tarlov scores in lower limbs and lower percentages of neurons in the A4 and A2 quadrants of flow cytometry. Fluorescent staining and quantification further indicated that miR-125b mimic decreased the immunoreactivities of p53 and cleaved caspase 3 in neurons and simultaneously reduced the number of double-labelled cells with TP53INP1. ConclusionsmiR-125b mimic partially protected neurons against neuroinflammation and aberrant p53 network activation-induced apoptosis during IR injury through downregulation of TP53INP1.

Association of 48 type 2 diabetes susceptibility loci with fasting plasma glucose and lipid levels in Chinese Hans

AimDozens of susceptibility loci have been identified by type 2 diabetes (T2D) genome wide association study (GWAS) in Europeans. In our previous studies, we systematically evaluated the association of 48 susceptibility loci with T2D risk in Chinese Hans. Because dyslipidemia and hyperglycemia are implicated in the pathogenic process of T2D, we further evaluated whether these 48 single nucleotide polymorphisms (SNPs) were related to fasting plasma glucose (FPG) or lipid levels in Chinese Hans. MethodsThe 48 SNPs were genotyped by using the Taqman OpenArray Genotyping System and iPLEX Sequenom MassARRAY platform. Multiple linear regression was used to assess the relationship between genetic variants and FPG and lipid levels among 3281 non-diabetic, healthy Chinese Hans. ResultsAfter adjusting for age, gender, body mass index (BMI), smoking status and drinking status, the T allele of rs13266634 in the SLC30A8 gene was significantly associated with decreased glucose level (β = −0.0119, P = 8.05 × 10−5), whereas the T allele of rs896854 in the TP53INP1 gene was associated with increased triglyceride (TG) level (β = 0.0342, P = 9.61 × 10−4) and decreased high-density lipoprotein cholesterol (HDL-C) level (β = −0.015, P = 3.24 × 10−3) after Bonferroni correction. We also conducted a meta-analysis consisted of 11 studies and confirmed that SNP rs896854 in the TP53INP1 gene was associated with T2D risk. ConclusionOur findings indicated that SNP rs13266634 in SLC30A8 was associated with glucose level and SNP rs896854 in TP53INP1 was associated with lipid level.

MiR-524-5p of the primate-specific C19MC miRNA cluster targets TP53IPN1- and EMT-associated genes to regulate cellular reprogramming

BackgroundIntroduction of the transcription factors Oct4, Sox2, Klf4, and c-Myc (OSKM) is able to ‘reprogram’ somatic cells to become induced pluripotent stem cells (iPSCs). Several microRNAs (miRNAs) are known to enhance reprogramming efficiency when co-expressed with the OSKM factors. The primate-specific chromosome 19 miRNA cluster (C19MC) is essential in primate reproduction, development, and differentiation. miR-524-5p, a C19MC member, is highly homologous to the reprogramming miR-520d-5p; we also reported that miR-524-5p was expressed in iPSCs but not mesenchymal stem cells (MSCs). This study aimed to elucidate possible contributions of miR-524-5p to the reprogramming process.MethodsA miR-524-5p precursor was introduced into human fibroblast HFF-1 in the presence of OSKM, and the relative number of embryonic stem cell (ESC)-like colonies that stained positively with alkaline phosphatase (AP) and Nanog were quantified to determine reprogramming efficiency. A miR-524-5p mimic was transfected to MSCs to investigate the effects of miR-524-5p on TP53INP1, ZEB2, and SMAD4 expression by real-time polymerase chain reaction (PCR) and Western blot. Direct gene targeting was confirmed by luciferase activity. A phylogenetic tree of TP53INP1 was constructed by the Clustal method. Contribution of miR-524-5p to cell proliferation and apoptosis was examined by cell counts, BrdU, MTT, and cell death assays, and pluripotency gene expression by real-time PCR.ResultsCo-expressing the miR-524 precursor with OSKM resulted in a two-fold significant increase in the number of AP- and Nanog-positive ESC-like colonies, indicating a role for miR-524-5p in reprogramming. The putative target, TP53INP1, showed an inverse expression relationship with miR-524-5p; direct TP53INP1 targeting was confirmed in luciferase assays. miR-524-5p-induced TP53INP1 downregulation enhanced cell proliferation, suppressed apoptosis, and upregulated the expression of pluripotency genes, all of which are critical early events of the reprogramming process. Interestingly, the TP53INP1 gene may have co-evolved late with the primate-specific miR-524-5p. miR-524-5p also promoted mesenchymal-to-epithelial transition (MET), a required initial event of reprogramming, by directly targeting the epithelial-to-mesenchymal transition (EMT)-related genes, ZEB2 and SMAD4.ConclusionsVia targeting TP53INP1, ZEB2, and SMAD4, miR-524-5p contributes to the early stage of inducing pluripotency by promoting cell proliferation, inhibiting apoptosis, upregulating expression of pluripotency genes, and enhancing MET. Other C19MC miRNAs may have similar reprogramming functions.

Anti-oxydative effects of miR-204 on LECs in age-related cataract

Background miRNAs are a group of non-coding small RNA molecules, and they play an important role in regulating the apoptosis of LECs.The biological effects and mechanisams of miRNAs on LECs in age-related cataract still need to be further elucidated. Objective This study was to investigate the anti-oxidative effects of miR-204 on age-related cataract in vitro. Methods The specimens of anterior lens capsules from age-related cataract patients and normal donors were collected in Shandong Eye Institution and 20 subjects for each.The expression level of miR-204 was detected and compared between cataractous eyes and normol eyes by real-time fluorescence quantitative PCR (RT-qPCR). HLE-B3 cells, a human LEC line, were cultured, and the oxydative stress models of LECs were established by adding 200 μmol/L H2O2 in the medium.The models were divided into model control group, miR-204 agomir group, agomir negative control group, miR-204 antgomir group and antagomir negative control group according to the difference of tranfected agents, and normal cells served as the control group.Twenty-four hours after transfection, the expression levels of miR-204 mRNA in the cells of various groups were detected by RT-qPCR to varify the transfection rate.Apoptosis rate of the cells was assayed by flow cytometry.The relative expression levels of TP53INP1 mRNA and p53 mRNA as well as their proteins were detected by RT-qPCR and Western blot, respectively. Results The mean expression level of miR-204 was significantly lower in the anterior lens capsules of cataractous eyes than that in the normal eyes (t=14.21, P<0.05). The mean apoptosis rate of cells was 1.31±0.12, 4.90±0.28, 2.60±0.15, 4.39±0.20, 5.74±0.13 and 4.34±0.63 in the normal control group, model control group, miR-204 agomir group, agomir negative control group, miR-204 antgomir group and antagomir negative control group, respectively.The apoptosis rate was significantly increased in the model control group compared with the normal control group (t=-20.69, P<0.01) and the apoptosis rate was also increased in the miR-204 antagomir group compared with antagomir negative control group (t=3.79, P<0.05); while the apoptosis rate in the miR-204 agomir group was significantly declined in comparison with agomir negative control group (t=-12.20, P<0.01). The relative expression levels of TP53INP1 and p53 mRNA and proteins in the cells were significantly higher in the model control group than those in normal control group (mRNA: t=6.44, 11.71, both at P<0.01; protein: t=10.72, 19.40, both at P<0.01), and so were between the miR-204 antagomir group and antagomir negative control group (mRNA: t=4.07, 3.74, both at P<0.05; protein: t=7.18, 10.58, both at P<0.05). However, the expression levels of TP53INP1 and p53 mRNA and protein were significantly reduced in the miR-204 agomir group in comparison with the agomir negative control group (mRNA: t=-19.28, -10.58, both at P<0.05; protein: t=-6.50, -6.36, both at P<0.05). Conclusions miR-204 induces oxidative damage of age-related cataract via targeting TP53INP1, which suggests that the activation of TP53INP1-p53 signaling may be involved in the development of age-related cataract. Key words: MicroRNA; TP53INP1; p53; Age-related cataract; Oxidative damage

TP53inp1 Gene Is Implicated in Early Radiation Response in Human Fibroblast Cells.

Tumor protein 53-induced nuclear protein-1 (TP53inp1) is expressed by activation via p53 and p73. The purpose of our study was to investigate the role of TP53inp1 in response of fibroblasts to ionizing radiation. γ-Ray radiation dose-dependently induces the expression of TP53inp1 in human immortalized fibroblast (F11hT) cells. Stable silencing of TP53inp1 was done via lentiviral transfection of shRNA in F11hT cells. After irradiation the clonogenic survival of TP53inp1 knockdown (F11hT-shTP) cells was compared to cells transfected with non-targeting (NT) shRNA. Radiation-induced senescence was measured by SA-β-Gal staining and autophagy was detected by Acridine Orange dye and microtubule-associated protein-1 light chain 3 (LC3B) immunostaining. The expression of TP53inp1, GDF-15, and CDKN1A and alterations in radiation induced mitochondrial DNA deletions were evaluated by qPCR. TP53inp1 was required for radiation (IR) induced maximal elevation of CDKN1A and GDF-15 expressions. Mitochondrial DNA deletions were increased and autophagy was deregulated following irradiation in the absence of TP53inp1. Finally, we showed that silencing of TP53inp1 enhances the radiation sensitivity of fibroblast cells. These data suggest functional roles for TP53inp1 in radiation-induced autophagy and survival. Taken together, we suppose that silencing of TP53inp1 leads radiation induced autophagy impairment and induces accumulation of damaged mitochondria in primary human fibroblasts.

Curcumin alters gene expression-associated DNA damage, cell cycle, cell survival and cell migration and invasion in NCI-H460 human lung cancer cells in vitro.

Lung cancer is the most common cause of cancer mortality and new cases are on the increase worldwide. However, the treatment of lung cancer remains unsatisfactory. Curcumin has been shown to induce cell death in many human cancer cells, including human lung cancer cells. However, the effects of curcumin on genetic mechanisms associated with these actions remain unclear. Curcumin (2 µM) was added to NCI-H460 human lung cancer cells and the cells were incubated for 24 h. Total RNA was extracted from isolated cells for cDNA synthesis, labeling, microarray hybridization and flour‑labeled cDNA hybridized on chip. Localized concentrations of fluorescent molecules were detected and quantified using Expression Console software (Affymetrix) with default RMA parameters. GeneGo software was used for the key genes involved and their possible interaction pathways. The results showed that ~170 genes were significantly upregulated and 577 genes were significantly downregulated in curcumin‑treated cells. Specifically, the up‑ and downregulated genes included CCNE2, associated with DNA damage; ID3, associated with cell survival and 146 genes with a >2- to 3-fold change including the TP53INP1 gene, associated with DNA damage; CDC6, CDCA5, TAKMIP2, CDK14, CDK5, CDCA76, CDC25A, CDC5L and SKP2, associated with cell cycle; the CARD6, ID1 and ID2 genes, associated with cell survival and the BRMS1L, associated with cell migration and invasion. Additionally, 59 downregulated genes exhibited a >4-fold change, including the DDIT3 gene, associated with DNA damage; while 97 genes had a >3- to 4-fold change including the DDIT4 gene, associated with DNA damage; the CCPG1 gene, associated with cell cycle and 321 genes with a >2- to 3-fold including the GADD45A and CGREF1 genes, associated with DNA damage; the CCPG1 gene, associated with cell cycle, the TNFRSF10B, GAS5, TSSC1 and TNFRSF11B gene, associated with cell survival and the ARHAP29 and CADM2 genes, associated with cell migration and invasion. In conclusion, gene alterations provide information regarding the cytotoxic mechanism of curcumin at the genetic level and provide additional biomarkers or targets for the treatment of human lung cancer.

Defects in mitophagy promote redox-driven metabolic syndrome in the absence of TP53INP1.

The metabolic syndrome covers metabolic abnormalities including obesity and type 2 diabetes (T2D). T2D is characterized by insulin resistance resulting from both environmental and genetic factors. A genome-wide association study (GWAS) published in 2010 identified TP53INP1 as a new T2D susceptibility locus, but a pathological mechanism was not identified. In this work, we show that mice lacking TP53INP1 are prone to redox-driven obesity and insulin resistance. Furthermore, we demonstrate that the reactive oxygen species increase in TP53INP1-deficient cells results from accumulation of defective mitochondria associated with impaired PINK/PARKIN mitophagy. This chronic oxidative stress also favors accumulation of lipid droplets. Taken together, our data provide evidence that the GWAS-identified TP53INP1 gene prevents metabolic syndrome, through a mechanism involving prevention of oxidative stress by mitochondrial homeostasis regulation. In conclusion, this study highlights TP53INP1 as a molecular regulator of redox-driven metabolic syndrome and provides a new preclinical mouse model for metabolic syndrome clinical research.

Advances in research of miR-15's role in malignant tumors

MicroRNA-155 (miR-155) is over-expressed in many human malignant tumors. Recent studies have shown that miR-155 plays a crucial role in tumor development and is closely related to disease prognosis. It is found that miR-155 promotes tumor growth,invasion and metastasis through inhibition of downstream targets such as SHIP1、C/EBPβ、SCOS1 、TP53INP1. The expression level of miR-155 can be used as a marker for the early diagnosis of malignant tumors, and inhibition of miR-155 expression may become a novel approach for cancer treatment. Key words: MicroRNAs; Neoplasms; Gene expression regulation

TP53INP1s and Homeodomain-interacting Protein Kinase-2 (HIPK2) Are Partners in Regulating p53 Activity

The TP53INP1 gene encodes two protein isoforms, TP53INP1alpha and TP53INP1beta, located into the nucleus. Their synthesis is increased during cellular stress by p53-mediated activation of transcription. Overexpression of these isoforms induces apoptosis, suggesting an involvement of TP53INP1s in p53-mediated cell death. It was recently shown that p53-dependent apoptosis is promoted by homeodomain-interacting protein kinase-2 (HIPK2), which is known to bind p53 and induce its phosphorylation in promyelocytic leukemia protein nuclear bodies (PML-NBs). In this work we show that TP53INP1s localize with p53, PML-IV, and HIPK2 into the PML-NBs. In addition, we show that TP53INP1s interact physically with HIPK2 and p53. In agreement with these results we demonstrate that TP53INP1s, in association with HIPK2, regulate p53 transcriptional activity on p21, mdm2, pig3, and bax promoters. Furthermore, TP53INP1s overexpression induces G1 arrest and increases p53-mediated apoptosis. Although a TP53INP1s and HIPK2 additive effect was observed on apoptosis, G1 arrest was weaker when HIPK2 was transfected together with TP53INP1. These results indicate that TP53INP1s and HIPK2 could be partners in regulating p53 activity.