Rho GDP Dissociation Inhibitor 2 Expression Research Articles

Complement component C3 and C5b-9 deposition on hypoxia reperfused endothelial cells by non-HLA antibodies against RhoGDI2: A player involved in graft failure?

Antibodies against Rho GDP-dissociation inhibitor 2 (RhoGDI2) are associated with inferior graft survival in transplant patients receiving a kidney from deceased donors. Although this suggests that these antibodies contribute to graft injury because of ischemia, it remains unknown whether they are also pathogenically involved in the process of graft loss. To study this, we firstly analyzed the IgG subclass profile of anti-RhoGDI2 antibodies in kidney transplant recipients, and whether antibody titers change over time or because of acute rejection. Next, we investigated the expression of RhoGDI2 on primary kidney and lung endothelial cells (ECs) upon hypoxia reperfusion. In addition, the complement-fixing properties of anti-RhoGDI2 antibodies were studied using imaging flow cytometry. Anti-RhoGDI2 antibodies in patients are mainly IgG1, and titers remained stable and seemed not be changed because of rejection. Antibodies against RhoGDI2, which surface expression seemed to increase upon hypoxia reperfusion, co-localized with C3 on ECs. Binding of human IgG1 monoclonal anti-RhoGDI2 antibodies as well as patient derived antibodies, resulted in complement activation, suggesting that these antibodies are complement fixing. This study suggested a potential pathogenic role of anti-RhoGDI2 antibodies in kidney graft loss. During ischemia reperfusion, the ability of these antibodies to fix complement could be one of the mechanisms resulting in tissue injury.

RhoGDI2 induced malignant phenotypes of pancreatic cancer cells via regulating Snail expression.

Rho GDP dissociation inhibitor 2 (RhoGDI2) has been shown to contribute to the aggressive phenotypes of human cancers, such as tumor metastasis and chemoresistance. This study aimed to assess the effects of RhoGDI2 on tumor progression and chemoresistance in pancreatic cancer cells. The expression of RhoGDI2 in pancreatic cancer cells was detected by Western blot analysis. Gain-of-function and loss-of-function approaches were done to examine the malignant phenotypes of the RhoGDI2-expressing or RhoGDI2-depleting cells. The correlation between RhoGDI2 and Snail was also analyzed. Differential expression of RhoGDI2 protein in pancreatic cancer cell lines was identified. Gain-of-function and loss-of-function experiments showed that RhoGDI2 induced the malignant phenotypes of pancreatic cancer cells, including proliferation, migration, invasion, and gemcitabine (GEM) chemoresistance. The upregulation of RhoGDI2 stimulated the expression of Snail, resulting in the altered expression of epithelial marker E-cadherin and mesenchymal marker Vimentin, which were characteristics of the tumorigenic activity of epithelial-mesenchymal transition. The expression of RhoGDI2 and Snail was upregulated in clinical tumor samples, and higher expression of RhoGDI2 or Snail was significantly associated with poor patient survival in pancreatic ductal adenocarcinoma (PDAC). The findings indicated that RhoGDI2 promoted GEM resistance and tumor progression in pancreatic cancer and that RhoGDI2 might be a potential therapeutic target in patients with PDAC.

Knockdown of RhoGDI2 represses human gastric cancer cell proliferation, invasion and drug resistance via the Rac1/Pak1/LIMK1 pathway.

Gastric cancer (GC) is the fifth most common primary malignancy in humans. Rho GDP dissociation inhibitor 2 (RhoGDI2) is overexpressed in multiple cancer types, but the role of RhoGDI2 in GC has not been elucidated. This study aims to determine the level of RhoGDI2 in GC and to confirm the effect of its inhibition or overexpression on GC cell migration, invasion and chemosensitivity. RhoGDI2 level is significantly enhanced in human GC tissue samples in comparison with normal gastric epithelium and corresponding para-cancerous samples. The expression of RhoGDI2 is correlated with clinicopathological parameters and prognosis. Transfection in combination with miRNA targeting of RhoGDI2 in GC cell lines remarkably downregulates GC cell migration and invasion and reduces the mRNA levels of Rac1, Pak1 and LIMK1. The inhibition of RhoGDI2 downregulates GC cell migration and invasion by attenuating the EMT cascade via the Rac1/Pak1/LIMK1 pathway. Knockdown of RhoGDI2 is a potential therapeutic strategy for GC.

Overexpression of RhoGDI2 correlates with the progression and prognosis of pancreatic carcinoma.

Rho GDP dissociation inhibitor 2 (RhoGDI2) has been found to be a regulator of tumor metastasis. However, the expression of RhoGDI2 and its clinicopathological significance as well as the pathway of RhoGDI2 in tumor metastasis have yet to be investigated. To investigate the role of RhoGDI2 in the progression and prognosis of pancreatic carcinoma (PC), the expression of RhoGDI2 in human PC tissues was examined and compared with the clinicopathological characteristics and prognosis. Moreover, the relationship between RhoGDI2 and E-cadherin was examined. The results indicated that RhoGDI2 was overexpressed in PC tissues and associated with clinicopathological characteristics, including clinical stage and lymph node metastasis. Patients with a RhoGDI2‑negative expression had a significantly longer survival time than those with a RhoGDI2‑positive expression. Additionally, the expression of RhoGDI2 was negatively correlated with the expression of E-cadherin in PC tissues. Taken together, the findings suggest that RhoGDI2 is important in the progression and prognosis of PC, and may be used as a potential prognostic biomarker and a therapeutic target for PC.

RhoGDI2 is associated with HGF-mediated tumor invasion through VEGF in stomach cancer.

RhoGDP dissociation inhibitor 2 (RhoGDI2) has been identified as a regulator of tumor metastasis; however, its role in cancer remains controversial. The aims of this study were to analyze RhoGDI2 in gastric cancer growth and metastasis, and to determine its possible signaling pathway. The level of expression of RhoGDI2 was further confirmed by real time RT-RCR and Western blot analysis. Transfection of cells with RhoGDI2 shRNA resulted in no effects of cell proliferation, as determined with MTT assays. In an in vitro invasion assay, significantly fewer cells transfected with RhoGDI2 shRNA, compared with control cells, were able to invade across a Matrigel membrane barrier. The role of RhoGDI2 in the level of HGF-induced up-regulation of vascular endothelial growth factor (VEGF) was measured by knockdown of RhoGDI2 with RhoGDI2 shRNA and a chromatic immuno-precipitation assay. The levels of RhoGDI2 and VEGF were up-regulated in cells treated with HGF in a dose-dependent manner. HGF-induced up-regulation of VEGF was repressed by RhoGDI2 knockdown. HGF-induced upregulation of phosphorylated ERK and P38 levels was inhibited in RhoGDI2 knockdown cells. HGF enhanced the binding activity of RhoGDI2 to the VEGF promoter in control cells, but not in RhoGDI2-shRNA cells. Findings of this study also showed a statistically significant difference in the mean RhoGDI2 level before and after surgery (p < 0.01) and the mean level of RhoGDI2 before surgery showed a statistically significant difference depending on lymphatic, neural invasion and stage (p < 0.05). In conclusion, RhoGDI2 might play an important role in up-regulation of VEGF induced by HGF and contributes to HGF-mediated tumor invasion and metastasis, which may serve as a promising target for gastric cancer therapy.

Depletion of RhoGDI2 expression inhibits the ability of invasion and migration in pancreatic carcinoma.

Rho GDP dissociation inhibitor 2 (RhoGDI2) has been identified as a regulator of tumor metastasis, although its role in tumor progression remains controversial. In this study, we examined the expression of RhoGDI2 in PC tissues and cell lines. To investigate the function of RhoGDI2 in PC cells, RhoGDI2 expression was depleted in PANC-1 and Patu8988 cells by small interfering RNA (siRNA). RhoGDI2 was found to be overexpressed in pancreatic carcinoma (PC) tissues and PC cell lines. Additionally, the results showed that depletion of RhoGDI2 significantly inhibited cell motility and invasion in vitro, but did not affect cell proliferation. The clinical study together with the experimental data confirmed that RhoGDI2 modulated the expression of matrix metalloproteinase 2 (MMP2). Taken together, findings of the present study indicated that RhoGDI2 is involved in pancreatic tumor malignancy and metastasis. Thus, RhoGDI2 is a potential target for the gene therapy of PC.

VEGF‐C mediates RhoGDI2‐induced gastric cancer cell metastasis and cisplatin resistance

Rho GDP dissociation inhibitor 2 (RhoGDI2) expression is correlated with tumor growth, metastasis and chemoresistance in gastric cancer. However, the mechanisms by which RhoGDI2 promotes tumor cell survival and metastasis remain unclear. In this study, we clearly demonstrate that RhoGDI2 upregulates VEGF-C expression and RhoGDI2 expression is positively correlated with VEGF-C expression in human gastric tumor tissues as well as parental gastric cancer cell lines. VEGF-C depletion suppressed RhoGDI2-induced gastric cancer metastasis and sensitized RhoGDI2-overexpressing cells to cisplatin-induced apoptosis in vitro and in vivo. Secreted VEGF-C enhanced gastric cancer cell invasion and conferred cisplatin resistance to RhoGDI2-overexpressing cells. We also show that RhoGDI2 positively regulates Rac1 activity in gastric cancer cells. Inhibition of Rac1 expression suppressed RhoGDI2-induced VEGF-C expression, and this inhibition was associated with decreased invasiveness and increased sensitivity to cisplatin in RhoGDI2-overexpressing cells. Our results indicate that RhoGDI2 might be a potential therapeutic target for simultaneously reducing metastasis risk and enhancing chemotherapy efficacy in gastric cancer.

642 VEGF-C Mediates RhoGDI2-induced Gastric Cancer Cell Metastasis and Cisplatin Resistance

Rho GDP dissociation inhibitor 2 (RhoGDI2) expression is correlated with tumor growth, metastasis and chemoresistance in gastric cancer. However, the mechanisms by which RhoGDI2 promotes tumor cell survival and metastasis remain unclear. In this study, we clearly demonstrate that RhoGDI2 upregulates VEGF-C expression and RhoGDI2 expression is positively correlated with VEGF-C expression in human gastric tumor tissues as well as parental gastric cancer cell lines. VEGF-C depletion suppressed RhoGDI2-induced gastric cancer metastasis and sensitized RhoGDI2-overexpressing cells to cisplatin-induced apoptosis in vitro and in vivo. Secreted VEGF-C enhanced gastric cancer cell invasion and conferred cisplatin resistance to RhoGDI2-overexpressing cells. We also show that RhoGDI2 positively regulates Rac1 activity in gastric cancer cells. Inhibition of Rac1 expression suppressed RhoGDI2-induced VEGF-C expression, and this inhibition was associated with decreased invasiveness and increased sensitivity to cisplatin in RhoGDI2-overexpressing cells. Our results indicate that RhoGDI2 might be a potential therapeutic target for simultaneously reducing metastasis risk and enhancing chemotherapy efficacy in gastric cancer.

Changes of neuronic apoptosis, Rho GDP dissociation inhibitor 2 and Bcl-2 in brain tissue of neonatal rats with hypoxic-ischemic brain damage

Objective To investigate the effect of Rho GDP dissociation inhibitor 2 （RhoGDI2） and Bcl-2 in pathogenesis of hypoxic-ischemic brain damage （HIBD）. Methods Neonatal 7-day-old Sprague Dawley rats were randomly divided into sham-operation control group, HIBD 6 h group and HIBD 48 h group （ n = 10 per group）. The apoptosis rate of brain cell was measured by flow cytometer and the expression of RhoGDI2 mRNA and Bcl-2 mRNA were detected by Real-time RT-PCR. Results （ 1 ） The ligated cerebral hemisphere of neonatal rats showed obvious edema at 48 h after hypoxia-ischemia. （ 2 ） Apoptotic cell appeared at 6 h in HIBD group,the apoptosis rate was （ 1.40 ±0. 12） %. The apoptosis rate obviously increased to （ 15.86 ±0.98 ） % at 48 h after HIBD, which showed a significant increase compared to sham-operation control group （ P 〈 0.01 ）. （ 3 ） The expressions of both RhoGDI2 mRNA and Bcl-2 mRNA were 4. 12 ±0. 74 and 2. 55±0. 65 respectively in sham-operation control group. In HIBD group, the expressions of both RhoGDI2 mRNA and Bcl-2 mRNA began to decrease at 6 h after HIBD （3. 19±0. 77,1.96±0. 36） and decreased furthermore at 48 h after HIBD （ 1.04±0. 18,1.06 ±0. 17 ）. The differences of expression levels among three groups were statistically significant （P 〈 0. 01 ）. （4） The expression of RhoGDI2 mRNA positively correlated with the expression of Bcl-2 mRNA （ r = 0. 831, P 〈 0. 05 ）. Conclusion With the emerging of apoptosis after HIBD, the expressions of both RhoGDI2 mRNA and Bcl-2 mRNA are decreased. The imbalance of expression of RhoGDI2 is involved in pathogenesis of HIBD by regulating Bcl-2 expression. Key words: Hypoxic-ischemic brain damage; Rho GDP dissociation inhibitor 2 ; Bcl-2 ; Apoptosis ; Rat, newborn

PLCγ is required for RhoGDI2-mediated cisplatin resistance in gastric cancer

Rho GDP dissociation inhibitor 2 (RhoGDI2) is a regulator of the Rho family GTPases. Recent work from our laboratory suggests that RhoGDI2 expression potentially enhances resistance to cisplatin as well as promotes tumor growth and malignant progression in gastric cancer. In this study, we demonstrate that phospholipase C-gamma (PLCγ) is required for RhoGDI2-mediated cisplatin resistance and cancer cell invasion in gastric cancer. The levels of phosphorylated PLCγ are markedly enhanced in RhoGDI2-overexpressing SNU-484 cells and, by contrast, repressed in RhoGDI2-depleted MKN-28 cells. Depletion of PLCγ expression or inhibition of its activity not only significantly increases cisplatin-induced apoptosis but also suppresses the invasive ability of RhoGDI2-overexpressing SNU-484 cells. Taken together, our results suggest that PLCγ plays a key role in RhoGDI2-mediated cisplatin resistance and cell invasion in gastric cancer cells.

RhoGDI2 antagonizes ovarian carcinoma growth, invasion and metastasis

Previous studies described functional roles for Rho GDP dissociation inhibitor 2 (RhoGDI2) in bladder, gastric and breast cancers. However, only limited expression and no functional analyses have been done for RhoGDI2 in ovarian cancer. We determined RhoGDI2 protein expression and function in ovarian cancer. First, western blot analysis was performed to determine the expression levels of RhoGDI2 in ovarian cells lines. RhoGDI2 but not RhoGDI1 protein expression levels varied widely in ovarian carcinoma cell lines, with elevated levels seen in Ras-transformed ovarian epithelial cells. Next, immunohistochemistry was performed to detect RhoGDI2 expression in patient samples of ovarian cysts and ovarian cancer with known histological subtype, stage, grade and outcome. RhoGDI2 protein was significantly overexpressed in high-grade compared to low-grade ovarian cancers, correlated with histological subtype, and did not correlate with stage of ovarian cancer nor between carcinomas and benign cysts. Unexpectedly, stable suppression of RhoGDI2 protein expression in HeyA8 ovarian cancer cells increased anchorage-independent growth and Matrigel invasion in vitro and in tail-vein lung colony metastatic growth in vivo. Finally, we found that RhoGDI2 stably-associated preferentially with Rac1 and suppression of RhoGDI2 expression resulted in decreased Rac1 activity and Rac-associated JNK and p38 mitogen-activated protein kinase signaling. RhoGDI2 antagonizes the invasive and metastatic phenotype of HeyA8 ovarian cancer cells. In summary, our results suggest significant cell context differences in RhoGDI2 function in cancer cell growth.

Abstract 1582: Rho GDP dissociation inhibitor 2 (RhoGDI2) plays a functionally important role on EGFR signaling pathway in adenocarcinoma cells of lung cancer

Abstract It has been known that Rho GDP dissociation inhibitor 2 (RhoGDI2) is a regulator of Rho family GTPase. We identified a RhoGDI2 protein through the protein expression profile and then investigated its functional role on EGFR downstream signal in NSCLC (non-small cell lung cancer). Three adenocarcinoma cell lines were selected according to the EGFR and K-RAS mutation status; PC9 (E476-A750 del in exon 19, wild type K-RAS), H1975 (T790M in exon 20 and L858R in exon 21, wild type K-RAS), A549 (wild type EGFR, K-RAS mutation in codon 12). Among 342 protein spots detected on the 2D gel, 134 spots were initially selected as differentially expressed proteins using proteome analysis. It was noteworthy that the ubiquitination-related proteins were highly expressed in EGFR TKIs-sensitive PC9 cells. On the other hand, the EGFR TKIs-resistant H1975 cells harboring both T790M and L858R mutations expressed the signal transduction-related proteins such as Zinc finger protein, G protein-coupled receptor and Ras oncogene family protein. We observed that p-EGFR (Y1047), p-AKT, and p-ERK were highly activated in PC9 cells in contrast to no or weak activation of these signaling proteins in A549 and H1975 cells. In PC9 cell, the expression of RhoGDI2 was significantly reduced after gefitinib treatment. On the contrary, its expression was enhanced by gefitinib in H1975 cells. Accordingly, RhoGDI2 of the final six protein spots was selected for further study. In six NSCLC cell lines, RhoGDI2 mRNA and protein expressions were evaluated using RT-PCR and Western blot analysis, respectively. The highest expression level was observed in PC-9 cells. When RhoGDI2 siRNA was transfected in PC9 cells, it was shown that RhoGDI2 protein expression was markedly reduced. Additionally, in this transfected PC-9 cells, no changes in the expression of p-ERK and truncated caspase-3 were detected, but total EGFR and p-AKT expression were increased. When gefitinib was added in the PC9 cells transfected with RhoGDI siRNA, the expression level of total EGFR was less decreased compared to that of PC-9 cells transfected with GAPDH (positive control). The RhoGDI2 siRNA transfected PC-9 cells treated with gefitinib showed more increased expression level of actin compared to that of positive control. We observed that the metastatic potential of the RhoGDI2 siRNA transfected PC-9 cells with lack of its expression was significantly increased in transwell migration assay. Taken together, it is suggested that RhoGDI2 protein is an important down-stream molecule of EGFR signaling pathway in PC9 cells harboring EGFR sensitive mutation, and plays a functional role in cell migration and metastasis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1582.

Up-regulation of RhoGDI2 in Human Breast Cancer and Its Prognostic Implications

Recent research has identified many genes and proteins that play specific roles in the process of systemic metastasis in various types of cancer. Rho GDP dissociation inhibitor 2 (RhoGDI2) has been shown to inhibit metastasis in human bladder cancer, but its role in breast cancer is controversial. We examined the regulation and clinical significance of RhoGDI2 in Korean breast cancer patients by using proteomic approaches. By using a proteomic approach, we observed an increased expression of RhoGDI2 in human breast cancer tissues when compared to that of the normal breast tissues, and we validated its up-regulation in an independent cohort of 8 breast cancer patients. The clinical implication of a RhoGDI2 expression was investigated in 57 breast cancer patients by performing immunohistochemistry. RhoGDI2 did not show a significant association with the tumor size, lymph node metastasis, the histologic grade or the hormone receptor status. However, the patients with RhoGDI2-expressing tumors had significantly shorter disease-free survival (p=0.043; hazard ratio, 3.87) and distant metastasis-free survival (p=0.039; hazard ratio, 5.15). Our results demonstrated a potential role of RhoGDI2 as a poor prognostic marker as well as a potential therapeutic target. The pro-metastatic nature of RhoGDI2 shown in our study may indicate its organ-specific role in cancer metastasis.

RhoGDI2 Expression Is Associated with Tumor Growth and Malignant Progression of Gastric Cancer

Rho GDP dissociation inhibitor 2 (RhoGDI2) has been identified as a regulator of Rho family GTPase. However, there is currently no direct evidence suggesting whether RhoGDI2 activates or inhibits Rho family GTPase in vivo (and which type), and the role of RhoGDI2 in tumor remains controversial. Here, we assessed the effects of RhoGDI2 expression on gastric tumor growth and metastasis progression. Proteomic analysis was done to investigate the tumor-specific protein expression in gastric cancer and RhoGDI2 was selected for further study. Immunohistochemistry was used to detect RhoGDI2 expression in clinical samples of primary gastric tumor tissues which have different pathologic stages. Gain-of-function and loss-of-function approaches were done to examine the malignant phenotypes of the RhoGDI2-expressing or RhoGDI2-depleting cells. RhoGDI2 expression was correlated positively with tumor progression and metastasis potential in human gastric tumor tissues, as well as cell lines. The forced expression of RhoGDI2 caused a significant increase in gastric cancer cell invasion in vitro, and tumor growth, angiogenesis, and metastasis in vivo, whereas RhoGDI2 depletion evidenced opposite effects. Our findings indicate that RhoGDI2 is involved in gastric tumor growth and metastasis, and that RhoGDI2 may be a useful marker for tumor progression of human gastric cancer.