N-myc Downstream-regulated Gene 2 Expression Research Articles

LncRNA XIST promotes insulin resistance in gestational diabetes mellitus via the microRNA-181b-5p/NDRG2 axis.

Many studies have explored the role of lncRNA X inactivation-specific transcript (XIST) in diabetes. This study was designed to unravel the regulatory mechanism of XIST on animal models of gestational diabetes mellitus (GDM) progression via the microRNA (miR)-181b-5p/N-myc downstream-regulated gene 2 (NDRG2) axis. XIST, miR-181b-5p, and NDRG2 expression levels in GDM mice were detected. The GDM mice were subjected to gain- and loss-of-function assays to examine the change of glucose metabolism indices (fasting blood glucose (FBG), fasting insulin (FINS) and homeostasis model assessment of insulin resistance (HOMA-IR)), serum oxidative stress factors (glutathione (GSH), superoxide dismutase (SOD), and malondialdehyde (MDA)), serum inflammatory factors (interleukin-1 β (IL-1β), IL-6, and tumor necrosis factor α (TNF-α)), pathological changes of pancreatic tissues, and apoptotic cells in pancreatic islets in GDM mice. XIST and NDRG2 expression were elevated while miR-181b-5p expression was depleted in GDM mice. Down-regulated XIST or NDRG2 or up-regulated miR-181b-5p reduced the FBG level, HOMA-IR, and serum IL-1β, IL-6, and TNF-α, and MDA contents, elevated the FINS, GSH, and SOD level, mitigated pathological changes in pancreatic tissues, and decelerated apoptotic cells in pancreatic islets in GDM mice. Silenced XIST dampens insulin resistance in GDM mice via the modulation of the miR-181b-5p/NDRG2 axis.

SPDR-2 DEVELOPMENT OF A NOVEL THERAPEUTIC STRATEGY AGAINST MEDULLOBLASTOMA TARGETING MYC

Abstract Medulloblastoma (MB) is the most common pediatric malignant brain tumor, with aggressive behavior and central nervous system dissemination. Standard of care for medulloblastoma include surgery, chemotherapy with platinum agent, and craniospinal radiation. If patients survived with these intensive treatments, frequently faced treatment-related adverse events. Therefore, development of novel therapeutic strategy is an important issue. Driver mutation underlying MB pathogenesis have been discovered, and molecular profiling led to the identification of subgroups; Wingless (WNT), Sonic Hedgehog(SHH), Group3, and Group4. Group 3, especially the group with MYC/MYCN amplification/overexpression confer a poor prognosis. Myc is attracting as a therapeutic target, however, to date, no drugs have been developed that target myc. We had found that N-myc downstream regulated gene-1 (NDRG1) protein expression is a good prognostic factor in glioblastoma, a representative of adult malignant brain tumors, and that Differentiation inducing factor-1 (DIF-1) increased NDRG1 expression. We reported that DIF-1 crossed the Blood-Brain Barrier and could be an effective treatment for GBM in vivo. In this research process, we found that DIF-1 markedly reduced Myc expression in patient-derived glioblastoma stem cells with MYC/MYCN amplification. Based on this result, we found that DIF-1 decreased Myc expression and inhibited cell survival/proliferation even in medulloblastoma cell lines with MYC/MYCN amplification. In this presentation, we will present the results obtained in the research of a novel Myc-targeted therapy using DIF-1 for MB with MYC/MYCN amplification.

MiR-96-5p induced NDRG1 deficiency promotes prostate cancer migration and invasion through regulating the NF-κB signaling pathway.

The N-myc downstream-regulated gene 1 (NDRG1) has been discovered as a significant gene in the progression of cancers. However, the regulatory mechanism of NDRG1 remained obscure in prostate cancer (PCa). The miR-96-5p and NDRG1 expression levels were evaluated in PCa cell lines, prostate tissues, and validated public databases by real-time PCR, western blot analysis, and immunohistochemistry. The function of miR-96-5p and NDRG1 were investigated by wound healing and transwell assays in vitro, and mouse xenograft assay in vivo. The candidate pathway regulated by NDRG1 was conducted by the next-generation gene sequencing technique. Immunofluorescence and luciferase assay was used to detect the relation between miR-96-5p, NDRG1, and NF-κB pathway. Overexpressing NDRG1 suppresses the migration, invasion, and epithelial-mesenchymal transition (EMT) in vitro, and inhibits metastasis in vivo. Moreover, miR-96-5p contributes to NDRG1 deficiency and promotes PCa cell migration and invasion. Furthermore, NDRG1 loss activates the NF-κB pathway, which stimulates p65 and IKBa phosphorylation and induces EMT in PCa. MiR-96-5p promotes the migration and invasion of PCa by targeting NDRG1 and regulating the NF-κB pathway.

The Function of N-Myc Downstream-Regulated Gene 2 (NDRG2) as a Negative Regulator in Tumor Cell Metastasis.

N-myc downstream-regulated gene 2 (NDRG2) is a tumor-suppressor gene that suppresses tumorigenesis and metastasis of tumors and increases sensitivity to anti-cancer drugs. In this review, we summarize information on the clinicopathological characteristics of tumor patients according to NDRG2 expression in various tumor tissues and provide information on the metastasis inhibition-related cell signaling modulation by NDRG2. Loss of NDRG2 expression is a prognostic factor that correlates with TNM grade and tumor metastasis and has an inverse relationship with patient survival in various tumor patients. NDRG2 inhibits cell signaling, such as AKT-, NF-κB-, STAT3-, and TGF-β-mediated signaling, to induce tumor metastasis, and induces activation of GSK-3β which has anti-tumor effects. Although NDRG2 operates as an adaptor protein to mediate the interaction between kinases and phosphatases, which is essential in regulating cell signaling related to tumor metastasis, the molecular mechanism of NDRG2 as an adapter protein does not seem to be fully elucidated. This review aims to assist the research design regarding NDRG2 function as an adaptor protein and suggests NDRG2 as a molecular target to inhibit tumor metastasis and improve the prognosis in tumor patients.

Expression and clinicopathological significance of glucocorticoid receptor, SGK1, and NDRG1 in hormone-naïve prostate carcinoma.

Glucocorticoid receptor (GR) has been implicated in prostate carcinoma growth and progression. Glucocorticoid receptor beta (GRβ) acts as an inhibitor of GR; however, its function is not well understood. Serum- and glucocorticoid-regulated kinase 1 (SGK1) is a GR-responsive gene that phosphorylates N-myc downstream-regulated gene 1 (NDRG1) and is involved in cancer growth and invasion. However, the expression of GR, GRβ, SGK1, and NDRG1 in prostate cancer and their relationship with clinicopathological and functional significance remain unknown. The association between the status of GR, GRβ, SGK1, and NDRG1 immunoreactivity and clinicopathological variables was analyzed in patients with prostate carcinoma to explore their clinical significance. In prostate carcinoma cases, the relative abundance of GR and NDRG1 immunoreactivity was inversely and significantly associated with the primary tumor stage (pT), while GR immunoreactivity was inversely and significantly associated with the Ki-67 score. The relative expression status of NDRG1 was significantly associated with that of GR. However, no significant correlation was observed between any of the clinicopathological parameters and GRβ and SGK1 expression. Our findings indicate that GR and NDRG1 expression status is correlated with clinicopathological features in patients with prostate cancer.

A Tumor Suppressor Gene, N-myc Downstream-Regulated Gene 1 (NDRG1), in Gliomas and Glioblastomas.

The development of potent and selective therapeutic approaches to glioblastoma (GBM) requires the identification of molecular pathways that critically regulate the survival and proliferation of GBM. Glioblastoma stem-like cells (GSCs) possess stem-cell-like properties, self-renewal, and differentiation into multiple neural cell lineages. From a clinical point of view, GSCs have been reported to resist radiation and chemotherapy. GSCs are influenced by the microenvironment, especially the hypoxic condition. N-myc downstream-regulated gene 1 (NDRG1) is a tumor suppressor with the potential to suppress the proliferation, invasion, and migration of cancer cells. Previous studies have reported that deregulated expression of NDRG1 affects tumor growth and clinical outcomes of patients with GBM. This literature review aimed to clarify the critical role of NDRG1 in tumorigenesis and acquirement of resistance for anti-GBM therapies, further to discussing the possibility and efficacy of NDRG1 as a novel target of treatment for GBM. The present review was conducted by searching the PubMed and Scopus databases. The search was conducted in February 2022. We review current knowledge on the regulation and signaling of NDRG1 in neuro-oncology. Finally, the role of NDRG1 in GBM and potential clinical applications are discussed.

Regulation of tumor-associated macrophage (TAM) differentiation by NDRG2 expression in breast cancer cells

Macrophages are a major cellular component of innate immunity and are mainly known to have phagocytic activity. In the tumor microenvironment (TME), they can be differentiated into tumor-associated macrophages (TAMs). As the most abundant immune cells in the TME, TAMs promote tumor progression by enhancing angiogenesis, suppressing T cells and increasing immunosuppressive cytokine production. N-myc downstream-regulated gene 2 (NDRG2) is a tumor suppressor gene, whose expression is down-regulated in various cancers. However, the effect of NDRG2 on the differentiation of macrophages into TAMs in breast cancer remains elusive. In this study, we investigated the effect of NDRG2 expression in breast cancer cells on the differentiation of macrophages into TAMs. Compared to tumor cell-conditioned medium (TCCM) from 4T1-mock cells, TCCM from NDRG2-overexpressing 4T1 mouse breast cancer cells did not significantly change the morphology of RAW 264.7 cells. However, TCCM from 4T1-NDRG2 cells reduced the mRNA levels of TAM-related genes, including MR1, IL-10, ARG1 and iNOS, in RAW 264.7 cells. In addition, TCCM from 4T1-NDRG2 cells reduced the expression of TAM-related surface markers, such as CD206, in peritoneal macrophages (PEM). The mRNA expression of TAM-related genes, including IL-10, YM1, FIZZ1, MR1, ARG1 and iNOS, was also downregulated by TCCM from 4T1-NDRG2 cells. Remarkably, TCCM from 4T1-NDRG2 cells reduced the expression of PD-L1 and Fra-1 as well as the production of GM-CSF, IL-10 and ROS, leading to the attenuation of T cell-inhibitory activity of PEM. These data showed that compared with TCCM from 4T1-mock cells, TCCM from 4T1-NDRG2 cells suppressed the TAM differentiation and activation. Collectively, these results suggest that NDRG2 expression in breast cancer may reduce the differentiation of macrophages into TAMs in the TME.

Abstract P5-08-13: NDRG1 expression is an independent prognostic factor in inflammatory breast cancer

Abstract Background: Inflammatory breast cancer (IBC) is a rare and highly aggressive form of breast cancer. Patients with IBC still have worse clinical outcomes compared to patients with non-IBC (40% versus 63% for 5 years overall survival, respectively) despite advances in breast cancer treatment. IBC remains a poorly characterized disease lacking specific therapeutic targets and prognostic biomarkers. Our group demonstrated that N-myc downstream regulated gene 1 (NDRG1) is crucial in promoting tumorigenesis and brain metastasis in mouse models of IBC. Our hypothesis is that NDRG1 is a prognostic marker associated with poor outcome in IBC patients. Methods: 64 IBC patients in a tissue microarray were evaluated by immunohistochemical staining with anti-NDRG1 primary antibody and NDRG1 levels were quantified. Using the median value, 32 patients were grouped as NDRG1-low (≤ median), and 32 as NDRG1-high (>median). Survival data were compared by Kaplan–Meier curves and log-rank test. Results: The average age of patients was 50 years. Sixty-two percent of patients had estrogen receptor (ER)-negative tumors, 83% were stage III, 80% high grade, and 67% of these patients received adjuvant radiation. The median follow-up time for the patients studied was 11.7 years, and the median overall survival (OS) time was 3.7 years. On univariate analysis, NDRG1 expression, tumor grade, disease stage, ER status, and adjuvant radiation therapy were associated with OS and disease specific survival (DSS). Patients with NDRG1-low tumors experienced better actuarial 10-year OS (p=0.0129) and DSS (p=0.0074), and showed significant higher 10-year OS and DSS rates than patients with NDRG1-high(OS, 45% vs. 19%, p=0.0278; DSS, 52% vs. 22%, p=0.0139). The median OS and DSS times were shorter for NDRG1-high patients (OS, 2.5 years; DSS, 3.1 years) than for NDRG1-low patients (OS, 5.9 years; DSS, 10.7 years). Multivariable analysis, NDRG1 was an independent predictor of OS (hazard ratio [HR]=2.449, p=0.0274) and DSS (HR=2.727, p=0.0039). ER status, disease stage and adjuvant radiation were also independent predictor for both OS and DSS. Furthermore, we observed that NDRG1-high expressing ER-negative tumors exhibit worse outcome in IBC patients (OS, p=0.0003; DSS, p=0.0003). NDRG1-high expression correlated with worse outcome in patients that received adjuvant radiation treatment (OS, p=0.0088; DSS, p=0.0093), and those with stage III (OS, p=0.0450; DSS, p=0.0239). Conclusions: Our findings demonstrated that NDRG1 is positively correlated with aggressive tumor characteristics in IBC and that it is an independent prognostic factor for OS and DSS in IBC patients, suggesting that targeting NDRG1 may provide a novel therapeutic strategy to improve outcomes for patients with IBC. Our data further suggests that NDRG1 warrants further investigation in radiation response. Citation Format: Emilly Schlee Villodre, Yun Gong, Xiaoding Hu, Lei Huo, Esther C Yoon, Naoto T Ueno, Wendy A Woodward, Debu Tripathy, Juhee Song, Bisrat G Debeb. NDRG1 expression is an independent prognostic factor in inflammatory breast cancer [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-08-13.

Expression and Significance of N-myc downstream regulated gene 2 in the process of Esophageal Squamous Cell Carcinogenesis

ABSTRACT It has been reported that the expression of tumor suppressor gene N-myc downstream-regulated gene 2 (NDRG2) was significantly reduced in human solid tumors, including esophageal squamous cell carcinoma (ESCC). This study aimed to explore whether the difference of NDRG2 expression exists in different stages of ESCC and provides a basis for the early diagnosis and prognosis of ESCC. Immunohistochemical staining was used to investigate the expression level of NDRG2 in samples from 91 patients with mild-to-moderate dysplasia, early ESCC, and advanced ESCC. The relationship between the expression of NDRG2 and clinicopathological characteristics of the patients was analyzed. The results showed that positive expression rates of NDRG2 in tissues adjacent to early ESCC (76.7%), or from mild-to-moderate dysplasia (74.1%), and early ESCC (83.3%) were significantly higher than in tissue from advanced ESCC (55.9%). The positive expression rate in advanced ESCC was significantly lower than in the other three tissue types (p < 0.05). There was a significant difference (p < 0.05) and correlation (Cramer’s V = 0.351, p = 0.019, <0.05) between the expression of NDRG2 and the clinical stage in the 64 patients with ESCC. In conclusion, this study found that the expression of NDRG2 gradually decreased with the progression of esophageal lesions into advanced ESCC. This difference in positive expression rate was more obvious in male patients and patients under 60 years of age. Therefore, the detection of NDRG2 plays an important role in differentiating early ESCC from advanced ESCC.

Targeting Wnt/tenascin C-mediated cross talk between pancreatic cancer cells and stellate cells via activation of the metastasis suppressor NDRG1

A major barrier to successful pancreatic cancer (PC) treatment is the surrounding stroma, which secretes growth factors/cytokines that promote PC progression. Wnt and tenascin C (TnC) are key ligands secreted by stromal pancreatic stellate cells (PSCs) that then act on PC cells in a paracrine manner to activate the oncogenic β-catenin and YAP/TAZ signaling pathways. Therefore, therapies targeting oncogenic Wnt/TnC cross talk between PC cells and PSCs constitute a promising new therapeutic approach for PC treatment. The metastasis suppressor N-myc downstream-regulated gene-1 (NDRG1) inhibits tumor progression and metastasis in numerous cancers, including PC. We demonstrate herein that targeting NDRG1 using the clinically trialed anticancer agent di-2-pyridylketone-4-cyclohexyl-4-methyl-3-thiosemicarbazone (DpC) inhibited Wnt/TnC-mediated interactions between PC cells and the surrounding PSCs. Mechanistically, NDRG1 and DpC markedly inhibit secretion of Wnt3a and TnC by PSCs, while also attenuating Wnt/β-catenin and YAP/TAZ activation and downstream signaling in PC cells. This antioncogenic activity was mediated by direct inhibition of β-catenin and YAP/TAZ nuclear localization and by increasing the Wnt inhibitor, DKK1. Expression of NDRG1 also inhibited transforming growth factor (TGF)-β secretion by PC cells, a key mechanism by which PC cells activate PSCs. Using an in vivo orthotopic PC mouse model, we show DpC downregulated β-catenin, TnC, and YAP/TAZ, while potently increasing NDRG1 expression in PC tumors. We conclude that NDRG1 and DpC inhibit Wnt/TnC-mediated interactions between PC cells and PSCs. These results further illuminate the antioncogenic mechanism of NDRG1 and the potential of targeting this metastasis suppressor to overcome the oncogenic effects of the PC-PSC interaction.

NDRG1 is expressed in human granulosa cells: An implicative role of NDRG1 in the ovary.

PurposeN‐myc downstream‐regulated gene 1 (NDRG1) is expressed in various human tissues and plays a role in regulating cellular proliferation, angiogenesis, and hypoxia sensing. However, the role of NDRG1 in the ovary remains poorly understood. Therefore, we investigated NDRG1 expression and the role of NDRG1 in the human ovary.MethodsFollicular fluid (FF) and luteinized granulosa cells were collected from follicles during oocyte retrieval. KGN cells were cultured with cobalt chloride (CoCl2, a hypoxia‐mimicking agent) and/or echinomycin. mRNA, protein levels and secretion, and localization were assessed by real‐time PCR, Western blotting, ELISA, and immunohistochemical analysis, respectively. KGN cells were also transfected with NDRG1 siRNA for 72 h.ResultsNDRG1 protein was expressed in luteinized granulosa cells. NDRG1 concentration was positively correlated with vascular endothelial growth factor (VEGF) and progesterone concentrations in FF. CoCl2‐induced hypoxic stress significantly increased NDRG1 and VEGF mRNA and protein and hypoxia‐inducible factor‐1α expression compared with those in the controls. The CoCl2‐induced overexpression of NDRG1 and VEGF was suppressed by echinomycin. Transfection with NDRG1 siRNA significantly suppressed the release of progesterone in the culture medium.ConclusionsThese results indicate that ovarian NDRG1 may play important roles in follicular development, especially in the early luteinization of pre‐ovulatory follicles.

Microrchidia family CW‑type zinc finger 2 promotes theproliferation, invasion, migration and epithelial‑mesenchymal transition of glioma by regulating PTEN/PI3K/AKT signaling via binding to N‑myc downstream regulated gene 1 promoter.

Glioma is a common malignant tumor of the central nervous system with high incidence and mortality. The present study aimed to investigate the role of Microrchidia family CW-type zinc finger 2 (MORC2) in the development of glioma. Firstly, MORC2 expression was detected in several glioma cell lines (U251, SHG44, LN229 and T98G). Following MORC2 silencing, cell proliferation was evaluated using the Cell Counting Kit-8 assay and the expression of proliferation-related proteins was assessed via immunofluorescence staining or western blotting. Cell invasion and migration were assessed using transwell and wound healing assays, respectively. Western blotting and immunofluorescence staining were employed to determine the expression of epithelial-mesenchymal transition (EMT)-associated proteins. The protein expression of N-myc downstream regulated gene 1 (NDRG1) and PTEN/PI3K/AKT signaling was determined with western blot analysis. Then, the luciferase reporter assay and chromatin immunoprecipitation (ChIP) assay were employed to evaluate the binding between MORC2 and NDRG1 promoter. Subsequently, cellular functional experiments were performed to assess the effects of NDRG1 on the progression of glioma after NDRG1 and MORC2 overexpression. In addition, tumor-bearing experiments were conducted using a U251 tumor-bearing nude mice model to detect tumor growth. The expression of proliferation (proliferating cell nuclear antigen, cyclin-dependent kinase 2 and cyclin E1), migration [matrix metalloproteinase (MMP)2 and MMP9], EMT (E-cadherin, N-cadherin and Vimentin) and PTEN/PI3K/AKT signaling proteins in tumor tissues was examined with immunohistochemistry assay or western blotting. Results revealed that MORC2 was notably unregulated in glioma cells compared with the normal human astrocyte. Loss-function of MORC2 inhibited the proliferation, invasion, migration and EMT of glioma cells. Importantly, MORC2 silencing upregulated NDRG1 expression and inactivated PTEN/PI3K/AKT signaling. Additionally, the luciferase reporter- and ChIP assays confirmed that MORC2 could bind to the NDRG1 promoter. NDRG1 upregulation suppressed the progression of glioma and these effects were partially reversed by MORC2 overexpression. Results of tumor-bearing experiments suggested that gain-function of NDRG1 inhibited tumor growth and downregulated the expression of proliferation, migration and EMT-related proteins in tumorous tissue in U251 tumor-bearing mice, which was partially counteracted after MORC2 overexpression. In addition, MORC2 overexpression abrogated the inhibitory effect of NDRG1 on PTEN/PI3K/AKT signaling. In summary, MORC2 promoted the progression of glioma by inactivation of PTEN/PI3K/AKT signaling via binding to NDRG1 promoter, providing a novel and potent target for the treatment of glioma.

Long noncoding RNA MIR22HG promotes Leydig cell apoptosis by acting as a competing endogenous RNA for microRNA-125a-5p that targets N-Myc downstream-regulated gene 2 in late-onset hypogonadism

Leydig cells (LCs) apoptosis is responsible for the deficiency of serum testosterone in Late-onset hypogonadism (LOH), while its specific mechanism is still unknown. This study focuses on the role of long noncoding RNA (lncRNA) MIR22HG in LC apoptosis and aims to elaborate its regulatory mechanism. MIR22HG was up-regulated in the testicular tissues of mice with LOH and H2O2-treated TM3 cells (mouse Leydig cell line). Interference of MIR22HG ameliorated cell apoptosis and upregulated miR-125a-5p expression in H2O2-treated TM3 cells. Then, the interaction between MIR22HG and miR-125a-5p was confirmed with RIP and RNA pull-down assay. Further study showed that miR-125a-5p downregulated N-Myc downstream-regulated gene 2 (NDRG2) expression by targeting its 3′-UTR of mRNA. What's more, MIR22HG overexpression aggravated cell apoptosis and reduced testosterone production in TM3 cells via miR-125a-5p/NDRG2 pathway. MIR22HG knockdown elevated testosterone levels in LOH mice. In conclusion, MIR22HG up-regulated NDRG2 expression through targeting miR-125a-5p, thus promoting LC apoptosis in LOH.Leydig cell apoptosis is responsible for the deficiency in serum testosterone in late-onset hypogonadism. This study reveals that lncRNA MIR22HG upregulates NDRG2 expression by targeting miR-125a-5p, thus promoting Leydig cell apoptosis in late-onset hypogonadism.

N-myc Downstream-Regulated Gene 2 (NDRG2) Function as a Positive Regulator of Apoptosis: A New Insight into NDRG2 as a Tumor Suppressor

N-myc downstream-regulated gene 2 (NDRG2) is a tumor suppressor gene that increases tumor sensitivity to anticancer drugs, slows tumor progression, and inhibits metastasis. NDRG2 is suppressed in various aggressive tumor positions, whereas NDRG2 expression is associated with patient prognosis, such as an improved survival rate. In this review, we summarize the tumor suppressor mechanism of NDRG2 and provide information on the function of NDRG2 concerning the susceptibility of cells to apoptosis. NDRG2 increases the susceptibility to apoptosis in various physiological environments of cells, such as development, hypoxia, nutrient deprivation, and cancer drug treatment. Although the molecular and cell biological mechanisms of NDRG2 have not been fully elucidated, we provide information on the mechanisms of NDRG2 in relation to apoptosis in various environments. This review can assist the design of research regarding NDRG2 function and suggests the potential of NDRG2 as a molecular target for cancer patients.

Upregulation of P53 promotes nucleus pulposus cell apoptosis in intervertebral disc degeneration through upregulating NDRG2.

P53 is an apoptosis marker which is involved in determining nucleus pulposus (NP) cell fate. Little is known about P53 interaction with N-Myc downstream-regulated gene 2 (NDRG2) in intervertebral disc degeneration (IVDD). Here, we studied the role of the P53-NDRG2 axis in IVDD. We found that NDRG2 was expressed in NP tissue obtained from patients with IVDD. The level of NDRG2 was positively related to the severity of IVDD, as determined by Pfirrmann grading. Subsequently, we overexpressed NDRG2 in human NP cells by adenoviral transfection and studied the effects of increased levels of NDRG2 on the viability and apoptosis of these cells. NDRG2 overexpression induced NP cell apoptosis and reduced viability in NP cells obtained from patient with IVDD. We also found that the level of P53 was elevated in NP cells from patients with IVDD and treatment with exogenous P53 upregulated NDRG2 in NP cells. Last, IVDD model was established in P53 knockout mice and the pathological changes in the intervertebral discs and NDRG2 expression were examined. P53 knockout can reduce the damage of NP tissues after IVDD surgery to some extent. Restoration of NDRG2 antagonized the effect of P53 knockout on IVDD. Collectively, this study suggests that elevated P53 in NP cells stimulates apoptosis of the cells by upregulating NDRG2 expression, thereby exacerbating IVDD.

TGF-β1 can be regulated by NDRG2 via the NF-κB pathway in hypoxia-induced liver fibrosis

BackgroundThe identification of the important elements that control hepatic stellate cell (HSC) activation will expand our understanding of the mechanism of liver fibrosis induced by hypoxia and affect the outcome of clinical treatment. A previous research demonstrated that N-Myc downstream-regulated gene 2 (NDRG2) is a potential regulator of fibrosis and a downstream target gene of hypoxia-inducible factor 1 (HIF-1). In this research, we studied the expression and function of NDRG2 in liver fibrosis induced by hypoxia.MethodsLX-2 cells/NF-κB-silenced LX-2 cells were exposed to hypoxic conditions (1% O2) to activate HSCs in vitro. The protein and mRNA expression levels of NDRG2, α-SMA and transforming growth factor beta 1 (TGF-β1) were evaluated by western blotting and real-time polymerase chain reaction (RT-PCR), respectively. Functional studies were performed using adenovirus-mediated gene upregulation.ResultsThe NDRG2 mRNA and protein levels were reduced under hypoxic conditions in LX-2 cells and overexpression of NDRG2 resulted in a decrease in the expression of TGF-β1 and α-SMA. Interestingly, no relationship was observed between NDRG2 and TGF-β1 when the NF-κB pathway was blocked, which indicates that NDRG2 can regulate the expression of TGF-β1 in LX-2 cells via the NF-κB pathway under hypoxic conditions.ConclusionsNDRG2 may regulate the expression of TGF-β1 via the NF-κB pathway and may be a novel therapeutic target for liver fibrosis induced by hypoxia.

N-myc downstream regulated gene 2 ameliorates myocardial remodeling and cardiac function in heart failure rats.

This study aims to explore the effect of NDRG2 (N-myc downstream regulated gene 2)-mediated Transforming growth factor-beta 1 (TGF-β1)/ Sma- and Mad-related protein (Smad) pathway in heart failure (HF) rats. HF rat models were established and treated with AdEGFP (adenovirus encoding enhanced green fluorescent protein) or AdNDRG2 (adenovirus encoding NDRG2). The echocardiography and hemodynamic parameters were detected, and the infarct size was calculated via 2,3,5-triphenyltetrazolium chloride (TTC) staining. Masson staining was performed to observe the collagen volume fraction (CVF), quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) to detect the expression of Collagen I (Col-I) and Collagen III (Col-III), and Transferase (TdT)-mediated D-UTP-biotin nick end labeling (TUNEL) staining to evaluate the apoptosis. Rats in the Model group presented with the decreases in left ventricular ejection fraction (LVEF), left ventricular shortening fraction (LVFS), left ventricular systolic pressure (LVSP) and maximal/minimum rate of left ventricular pressure (±dp/dt max), and significant increases in left ventricular end-diastolic pressure (LVEDP) and CVF. At the meantime, the expression of Col-I and Col-III as well as the apoptotic rate of myocardial cells was also elevated with increased infarct size in the Model group. The Model rats also had the significant reduction in the expression of NDRG2 and up-regulations of TGF-β1, p-Smad2/Smad2, p-Smad3/Smad3 and tissue inhibitor of metalloproteinases-2 (TIMP-2). However, model rats treated with AdNDRG2 had evident amelioration in aforementioned indicators. In conclusion, NDRG2 reduces the apoptosis of myocardial cells and improves the heart function and myocardial remodeling in HF rats via inhibiting the activity of TGF-β1/Smad.

Elemene Emulsion Injection Administration Reduces Neuropathic Pain by Inhibiting Astrocytic NDRG2 Expression within Spinal Dorsal Horn.

To investigate the mechanisms underlying elemene-induced analgesia in rats with spared nerve injury (SNI). Sixty-five rats were equally divided into 5 groups using a random number table: naive group, sham group, SNI group, SNI + elemene (40 mg·kg-1·d-1) group and naive + elemene (40 mg·kg-1·d-1) group. An SNI rat model was established and the intervention were given respectively for 14 consecutive days. Von Frey filament tests and elevated plus-maze (EPM) tests were used to evaluate the effect of elemene on the mechanical threshold and anxiety, respectively. Immunoblotting and immunostaining were used to measure the expression of glial fibrillary acidic protein (GFAP) and NMYC downstream-regulated gene 2 (NDRG2) within the lumbar spinal dorsal horn (SDH). The SNI rat model exhibited a significant decrease in paw withdrawal threshold and exploratory behaviour in the EPM (P<0.05). Consecutive administration of elemene alleviated SNI-induced mechanical allodynia and anxiety in rats (P<0.05). Immunohistochemical data showed that elemene decreased SNI-induced upregulation of NDRG2 within the SDH (P<0.05). Double immunofluorescent staining data further showed that elemene decreased SNI-induced upregulation of the number of GFAP immunoreactive (-ir), NDRG-ir, and GFAP/NDRG2 double-labelled cells within the SDH (P<0.05). Immunoblotting data showed that elemene decreased SNI-induced upregulation of GFAP and NDRG2 within the SDH (P<0.05). Elemene possibly alleviated neuropathic pain by downregulating the expression of NDRG2 in spinal astrocytes in a rat model of SNI.

Transcription factor Krüppel-like factor 5-regulated N-myc downstream-regulated gene 2 reduces IL-1β-induced chondrocyte inflammatory injury and extracellular matrix degradation

ABSTRACT Previous research has identified N-myc downstream-regulated gene 2 (NDRG2) as one of the differentially expressed genes common to rat models of osteoarthritis (OA) and human OA. The purpose of this study was to investigate the role of NDRG2 in OA. In this study, an in vitro OA model was constructed by challenging ATDC5 chondrocytes with 10 ng/ml IL-1β. After transfection of pcDNA3.1(+)/NDRG2, qPCR and western blot were performed to assay NDRG2 expression. The analyses of cell viability, apoptosis and inflammatory molecule expression were employed respectively by CCK-8, TUNEL and ELISA. The protein expression related to apoptosis, inflammation or extracellular matrix (ECM) degradation was detected by western blot. The binding of Krüppel-like factor 5 (KLF5) to NDRG2 promoter was verified by means of dual-luciferase reporter assay. After overexpression of both NDRG2 and KLF5 in IL-1β-stimulated ATDC5 chondrocytes, corresponding assays were performed to examine cell viability, apoptosis, inflammatory response and ECM degradation. In ATDC5 chondrocytes challenged with IL-1β, NDRG2 expression was much lower than that in the control group, whereas it’s overexpression helped restored cell viability and reduce cell apoptosis, inflammatory response and ECM degradation. It was also observed that KLF5 expression was decreased in IL-1β-stimulated ATDC5 chondrocytes, and that KLF5 bound to the NDRG2 promoter. Importantly, overexpressing KLF5 could reverse the protective effect of NDRG2 overexpression on IL-1β-stimulated ATDC5. Overall, NDRG2 could be transcriptionally regulated by transcription factor KLF5 and may play a protective role against chondrocyte the inflammatory response and ECM degradation in OA.

NDRG1 regulates Filopodia-induced Colorectal Cancer invasiveness via modulating CDC42 activity.

N-myc downstream regulated gene-1 (NDRG1) has been identified as a putative metastasis suppressor gene and proved to be a key player in cancer spreading and proliferation in our previous work. However, the effects of NDRG1 on tumor invasion and the mechanisms behind it are rarely understood. Here we provided in silico evidence that NDRG1 plays a crucial role in actin reorganization in colorectal cancer (CRC). Through in vitro experiments, we next observed filopodia formation was altered in NDRG1-modified cell lines, while cell division cycle-42 (CDC42) displayed excessive activation in NDRG1-silenced cells. Mechanistically, NDRG1 loss disrupts the binding between RhoGDIα and CDC42 and triggers the activation of CDC42 and the downstream cascades PAK1/Cofilin, thereby promotes the formation of filopodia and invasiveness of CRC. The knockdown of NDRG1 led to enhanced dissemination of CRC cells in vivo and correlates with active CDC42 expression. Using clinical sample analysis, we found an elevated level of active CDC42 in patients with advanced T stage, and it was negatively related to NDRG1 expression. In sum, these results uncover a mechanism utilized by NDRG1 to regulate CDC42 activity in coordinating cytoskeleton reorganization, which was crucial in cancer invasion.