Chloride Intracellular Channel 1 Expression Research Articles

Multi-omics analysis reveals CLIC1 as a therapeutic vulnerability of gliomas.

Introduction: Despite advances in comprehending cancer biology, malignant gliomas remain incurable. The present work conducted a multi-omics analysis for investigating the significance of chloride intracellular channel 1 (CLIC1) in gliomas. Methods: Multi-omics data of glioma covering transcriptomics, genomics, DNA methylation and single-cell transcriptomics from multiple public cohorts were enrolled for analyzing CLIC1. In vitro experiments were conducted to measure apoptosis and cell mobility in U251 and U373 glioma cells following transfection of CLIC1 siRNAs. Results: Elevated CLIC1 expression was proven to stably and independently estimate worse survival outcomes. CLIC1 expression was higher in more advanced stage, wild-type IDH and unmethylated MGMT samples. Tumorigenic and anticancer immunity pathways were remarkably enriched in CLIC1-up-regulated tumors. Additionally, CLIC1 was positively linked with cancer-immunity cycle, stromal activation, DNA damage repair and cell cycle. Suppressing CLIC1 resulted in apoptosis and attenuated cell motility of glioma cells. More frequent genomic alterations were found in CLIC1-up-regulated tumors. CLIC1 expression presented a remarkably negative connection to DNA methylation. High CLIC1 expression samples were more sensitive to camptothecin, cisplatin, doxorubicin, erlotinib, paclitaxel, rapamycin, clofarabine, tanespimycin, methotrexate, everolimus, TAK-733, trametinib and AZD8330. Tumors with upregulated CLIC1 presented abundant immune cell infiltration, higher expression of immune-checkpoints and -modulators and similar transcriptome profiling, indicative of well response to immune-checkpoint blockade (ICB). Nevertheless, due to elevated TIDE score, tumors with CLIC1 upregulation appeared to be resistant to ICB. Single-cell analysis unveiled that CLIC1 was expressed ubiquitously in tumor cells and tumor microenvironment. Conclusions: Overall, CLIC1 was a promising treatment vulnerability in glioma.

Chloride Intracellular Channel Protein 1 Expression and Angiogenic Profile of Liver Metastasis of Digestive Origin

Chloride intracellular channel 1 (CLIC1) is involved in cell migration and metastasis. The histological growth patterns of liver metastasis are as follows: desmoplastic (d-HGP), replacement (r-HGP), pushing (p-HGP), and mixed. The aim of this study was to evaluate the relation between HGP, angiogenesis, and CLIC1 expression. Materials and Methods: A total of 40 cases of primary tumors and their LM: d-HGP (12 cases), r-HGP (13 cases), and p-HGP (15 cases), were evaluated through simple and double immunostaining. CLIC1 assessment was conducted as follows: scores of 0 (less than 10% of positive cells), 1 (10-30%), 2 (30-50%), or 3 (more than 50%) were assigned. Heterogeneous CLIC1 expression was found. CLIC1 in primary tumors correlated with grade G for all cases of LM with a p-HGP (p = 0.004). The CLIC1 score for LMs with an r-HGP correlated with grade G of the corresponding primary tumor (p = 0.027). CLIC1 and CD34+/Ki67+ vessels (p = 0.006) correlated in primary tumors. CLIC1 in primary tumors correlated with CD34+/Ki67+ vessels of LMs with a d HGP (p = 0.024). Conclusions: The CLIC1 score may have prognostic value, mainly for LMs with a p-HGP and r-HGP, and therapeutic value for LMs with a d-HGP.

Comprehensive analysis of clinical prognosis and CLIC1 immune invasion in lung adenocarcinoma.

Background:Chloride intracellular channel 1 (CLIC1) plays an important role in the process of cell epithelial transport, and is also involved in tumor invasion and metastasis. Due to its aberrant expression in cancer, the mechanism of action of CLIC1 in cancer has been carefully studied. In this study, we tried to investigate the relationship between CLIC1 and lung adenocarcinoma (LUAD).Methods:The RNA-sequencing data and clinical information of CLIC1 in lung adenocarcinoma were collected from the the cancer genome altas (TCGA) database and analyzed with R software. Paired t test and Mann–Whitney U test were used to detect differences between LUAD tissue and adjacent normal tissue, and the pROC software package performed reactive oxygen species (ROC) curves to detect cutoff values for CLIC1. The expression of CLIC1 in normal human tissues was extracted from the human protein altas (HPA) database, and analyzed clinical proteomic tumor analysis consortium by using UALCAN programme. The relationship between CLIC1 and LUAD was explored by enrichment analysis using gene oncology and Kyoto encyclopedia of genes and genomes. The tumor immunity estimation resource (TIMER) and integrated repository portal for tumor-immune system interactions (TISIDB) databases were used to analyze the correlation between CLIC1 and LUAD immune cell infiltration. Survival analysis of CLIC1 in LUAD was assessed by the PrognoScan database.Results:Compared with normal tissues, both mRNA (messenger Ribose Nucleic Acid) and protein of CLIC1 were overexpressed in LUAD, which was associated with shorter overall survial (OS). In addition, CLIC1 expression was in connection with some clinical-pathological characteristics like tumor node metatasis stages and lymph node metastases. What’s more, CLIC1 may play a role in the immune infiltration of LUAD.Conclusion:In summary, CLIC1 is up-regulated in LUAD and is associated with tumor metastasis, tumor staging, and OS. It may be regarded as a novel marker for prognostic judgement in LUAD.

Chloride Intracellular Channel 1 is a Potential Biomarker for Breast Cancer.

PurposeMultiple reports have demonstrated that highly expressed chloride intracellular channel 1 (CLIC1) exists in a range of malignant tumors and is involved in proliferation, invasion, and migration of cancer cells. There are few studies on CLIC1 and breast cancer (BC). The purpose of this research was to evaluate the expression level of CLIC1 in BC and its impact on prognosis of BC patients.Patients and MethodsDifferences in CLIC1 expression levels in 25 pairs of BC and corresponding paracancerous specimens were tested by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot (WB). Immunohistochemistry (IHC) was performed to discuss the relevance between CLIC1 expression in BC tissue chips and clinicopathological parameters of BC patients. The effect of CLIC1 expression on patient prognosis was evaluated by Kaplan–Meier survival curve and Cox regression analysis. Receiver operating characteristic (ROC) curve assessed the diagnostic performance of CLIC1 for BC.ResultsThe experimental results of qRT-PCR and WB demonstrated that CLIC1 was highly expressed in BC tissues. IHC results showed that overexpression of CLIC1 was strictly correlated with tumor size, TNM classification, pathological grade, lymph node metastasis and Ki67. Patients with lower CLIC1 expression had longer overall survival (OS) and progression-free survival (PFS). Cox regression analysis and ROC curve confirmed that CLIC1 could independently influence the prognosis of BC patients and might have diagnostic efficiency.ConclusionOverexpressed CLIC1 is closely related to the progression of BC and the poor prognosis of the patients, suggesting that it may act as a potential biological diagnostic index for BC.

CLIC1 Drives Angiogenesis in Hepatocellular Carcinoma by Modulating VEGFA.

Background: Chloride intracellular channel 1 (CLIC1) is upregulated in hepatocellular carcinoma (HCC). The present study aimed to investigate the role of CLIC1 in HCC angiogenesis. Materials and Methods: Immunohistochemistry (IHC) was used to test the expression of CLIC1 and CD34 in 67 pairs of HCC and paracarcinoma tissues. The prognosis data of the patients were used to analyze the clinical relevance of CLIC1. We built a coculture system of HCC cells and endothelial cells to explore the migration of endothelial cells. Conditioned media (CMs) from HCC cells was then collected to assess endothelial cell migration. Experiments were then conducted to confirm the relationship between CLIC1 and angiogenesis in a subcutaneous tumor model. Results: CLIC1 expression was higher in HCC tumor tissues than in paracarcinoma tissues. Patients with increased CLIC1 expression showed a higher microvascular density (MVD; P = .013). Kaplan-Meier curves indicated that patients with lower expression of CLIC1 had better overall survival (P < .001) and recurrence-free survival (P = .046). Vascular endothelial growth factor A (VEGFA) in CMs from CLIC1-knockdown cells was lower than in the control group, while VEGFA in CMs from CLIC1 overexpression cells was higher than in the control group. CMs from CLIC1 overexpression cell lines promote the in vitro migration of EA.hy926 cells. Meanwhile, adding Bevacizumab to CMs from CLIC1 overexpression cells significantly inhibited this migration. The growth of xenograft tumors derived from CLIC1-knockdown Huh7 cells was restrained compared with the control group (P < .001). IHC staining showed MVD was higher in tumors with CLIC1 overexpression. Conclusion: CLIC1 is a promising biomarker for predicting the prognosis of HCC patients, and expression of CLIC1 correlates with angiogenesis in HCC through regulating VEGFA

Chloride Intracellular Channel 1 Expression Is Associated With Poor Prognosis of Lung Adenocarcinoma

Chloride intracellular channel 1 (CLIC1) is a member of the chloride channel protein family. The aim of this study was to clarify the role of CLIC1 in lung adenocarcinoma. The expression levels of CLIC1 in 74 patients with completely resected lung adenocarcinoma were analyzed by immunohistochemistry. Overall survival was assessed in relation to the expression level of CLIC1. Moreover, in the lung cancer cell lines A549 and PC9, CLIC1 expression was inhibited by small interfering RNA. The function of CLIC1 was analyzed in these cell lines. High expression of CLIC1 was associated with short overall survival compared to low expression (p=0.0327). Multivariate analysis revealed that CLIC1 expression was an independent prognostic factor. Knockdown of CLIC1 inhibited cell proliferation and migration through suppression of the p38 MAPK signaling pathway in A549 and PC9 cells. CLIC1 may be a useful prognostic factor in lung adenocarcinoma.

Acetylation-stabilized chloride intracellular channel 1 exerts a tumor-promoting effect on cervical cancer cells by activating NF-κB.

Cervical cancer remains a major cause of cancer-related death in women, especially in developing countries. Previously, we found that the acetylation levels of chloride intracellular channel 1 (CLIC1) at lysine 131 were increased in cervical cancer tissues using a label-free proteomics approach. The aim of this study was to further determine the role of CLIC1 expression and its acetylation in cervical cancer. CLIC1 expression and its implications for the prognosis of cervical cancer were analyzed using primary patient samples and cells, and the Gene Expression Profiling Interactive Analysis (GEPIA) database (gepia.cancer-pku.cn). The effect of CLIC1 on cervical cancer cells was evaluated using Cell Counting Kit (CCK)-8, flow cytometry, scratch wound healing, transwell, Western blotting and co-immunoprecipitation (Co-IP) assays. In vivo tumor growth was assessed using mouse xenograft models. We found that CLIC1 expression was increased in cervical cancer tissues and cells and that patients with a high CLIC1 expression tended to havea shorter overall survival time. Knockdown of CLIC1 significantly reduced in vitro cervical cancer cell proliferation, migration and invasion, and in vivo tumorigenesis. At the molecular level, we found that nuclear factor kappa B (NF-κB) activity was positively regulated by CLIC1. Pyrrolidine dithiocarbamate (PDTC), an inhibitor of NF-κB, attenuated the tumor-promoting effect of CLIC1. Moreover, we found that CLIC1 acetylation at K131 was upregulated in cervical cancer cells, which stabilized CLIC1 by inhibiting its ubiquitynation. Substitution of K131 inhibited CLIC1 ubiquitynation and promoted in vitro cervical cancer cell proliferation, migration and invasion, and in vivo tumor growth. In addition, we found that acetyltransferase HAT1 was responsible for CLIC1 acetylation at K131. Our data indicate that CLIC1 acts as a tumor promoter in cervical cancer, suggesting a potential treatment strategy for cervical cancer by regulating CLIC1 expression and/or acetylation.

CLIC1 knockout inhibits invasion and migration of gastric cancer by upregulating AMOT-p130 expression.

To explore the regulatory relationship between Chloride intracellular channel 1 (CLIC1) and Angiomotin (AMOT)-p130, and reveal the role of AMOT-p130 in gastric cancer (GC). Immunohistochemistry was performed to analyze the expression of CLIC1 and AMOT-p130 in GC tissues and adjacent tissues. The expression of AMOT-p130 upon CLIC1 silencing was analyzed using RT-PCR, western blot, and immunofluorescence in GC cells. Transwell and wound-healing assays were performed to detect migration and invasion in GC cells. The changes in EMT-related proteins were detected using western blot. Our study found that high CLIC1expression was significantly associated with low AMOT-p130 expression in GC tissues. Silencing CLIC1 expression in MGC-803 cells (MGC-803 CLIC1 KO) and AGS cells (AGS CLIC1 KO) decreased the invasive and migratory abilities of tumor cells, which were induced by the upregulation of AMOT-p130. Subsequently, we demonstrated that AMOT-p130 inhibits the invasive and migratory abilities of GC cells by inhibiting epithelial-mesenchymal transition. Our study suggests that AMOT-p130 could inhibit epithelial-mesenchymal transition in GC cells. CLIC1 may participate in the metastatic progression of GC by downregulating the expression of AMOT-p130.

Establish cisplatin-resistant gastric cancer cell line MGC-803/cisplatin and reveal its mechanism

Objective To establish a cisplatin-resistant gastric cancer cell line MGC-803/cisplatin (DDP) and to explore its drug resistance mechanism. Methods The cisplatin was used to induce the drug resistance of MGC-803 cells. The half maximal inhibitory concentration (IC50) of the drug-resistant strains was detected by the cell counting kit-8 (CCK-8) method. The expression of chloride intracellular channel 1 (CLIC1) was detected by Western blotting analyses. The wild-type CLIC1 plasmid and the short hairpin RNA (shRNA) plasmid targeting CLIC1 were constructed. After Lipo3000 was transfected into gastric cancer cells, the IC50 of gastric cancer cells to cisplatin was detected, and then the intracellular Cl-concentration was detected by MQAE fluorescent probe. Results The cisplatin-resistant cell line MGC-803/DDP was successfully induced with an IC50 of (7.02±0.13) mg/L, while the IC50 of MGC803 was (1.29±0.09) mg/L, (t=36.090, P<0.05). The CLIC1 protein of MGC803/DDP was up-regulated by (2.27±0.15) times relative to MGC803 (t=7.841, P<0.05). After silencing the CLIC1 gene of MGC803/DDP, the IC50 of the CON, CN, and KD groups were (6.96±0.09), (6.93±0.15) and (3.02±0.20) mg/L, respectively, which was significantly decreased in the KD group (F=0.209, P<0.05). After overexpression of the CLIC1 gene in MGC803 cells, the IC50 of the CON, CN, and OE groups were (1.35±0.07), (1.25±0.07) and (4.77±0.12) mg/L, respectively, and the OE group was significantly elevated (F=0.508, P<0.05). The concentration of chloride ion was detected by MQAE. The relative fluorescence intensity of MGC-803, MGC-803/OE, MGC-803/DDP and MGC-803/DDP-KD were (1.02±0.03), (0.61±0.02), (0.67±0.01) and (1.39±0.02), respectively. The concentration of Cl- in MGC-803/OE cells was higher than that in MGC-803 (t=10.800, P<0.05), while the concentration of Cl- in MGC-803/DDP was higher than that in MGC-803 and MGC-803/DDP-KD, and the concentration of Cl- in KD group was the lowest (F=229, P<0.05). Conclusion The CLIC1 gene can induce the resistance of gastric cancer cell line MGC-803 to cisplatin in vitro. The mechanism may be related to the up-regulation of CLIC1 expression, which leads to an increase of intracellular chloride ion concentration. Key words: Gastric cancer; Chloride intracellular channel 1; Cisplatin; Drug resistance

MicroRNA‑124 negatively regulates chloride intracellular channel1 to suppress the migration and invasion of liver cancer cells.

The dysregulation of microRNAs (miRNAs) is associated with the development and progression of a variety of cancers, including liver cancer. Aberrant expression of miRNA (miR)-124 has been demonstrated in liver cancer, but its functional mechanism in liver cancer is still largely unknown. Metastasis of liver cancer is one of the most common causes of mortality. The present study showed that miR-124 inhibited the proliferation, migration and invasion of liver cancer cells. Furthermore, chloride intracellular channel 1 (CLIC1) was identified as a novel target of miR-124 in liver cancer cells. Overexpression of miR-124 reduced CLIC1 expression at both the protein and mRNA levels in liver cancer cells. Downregulation of CLIC1 decreased the migration and invasion of liver cancer cells without affecting cell proliferation. Taken together, these results showed that CLIC1 is a critical target for miR-124-mediated inhibitory effects on cell migration and invasion. Thus, miR-124 or suppression of CLIC1 may have diagnostic value and therapeutic potential for the treatment of human liver cancer.

Intracellular Chloride Ion Channel Protein-1 Expression in Clear Cell Renal Cell Carcinoma.

Chloride intracellular channel 1 (CLIC1) represents a promising target for personalized therapy. Our aim was to assess CLIC1 expression in clear cell renal cell carcinoma (cc RCC) and identify its possible prognostic role. Fifty cases of cc RCC were evaluated and selected for immunohistochemistry. CLIC1 expression was correlated with tumor grade, invasion and heterogeneity. A total of 87.5% of the cases were CLIC1 positive, with either a homogeneous (31.42%) or a heterogeneous (68.57%) pattern. Low, mild and strong CLIC1 expressing tumors were defined based on nuclear (N), cytoplasmic (C), membrane (M) or combinations of them (NC, NM, CM, NCM) in terms of CLIC1 distribution. A significant correlation was found between tumor grade and percent of positive tumor cells (p=0.017). For G3 tumors, CLIC1 cytoplasmic expression was strongly correlated with high expression status (p=0.025) and tumor heterogeneity (p=0.004). CLIC1 expression was also correlated with metastasis (p=0.046). We defined four cc RCC groups depending on G, CLIC1 expression and pattern: i) G3/NM/low CLIC1+, ii) G2/CM/mild CLIC1+ iii) G1 or G2/NM or CM /high CLIC1+, and iv) G2/M /high CLIC1.

CLIC1 Cooperates with Integrins to Promote Thrombus Formation and Angiogenesis

Chloride intracellular channel 1 (CLIC1) is a member of a family of six highly homologous membrane proteins (CLIC1-6), which have been shown to be co-regulated with integrins suggesting their involvement in cell adhesion, migration and proliferation. CLIC1 is a metamorphic protein that functions as an oxidoreductase in the cytoplasm as well as an ion channel in the cell membrane. CLIC1 is upregulated in angiogenic endothelial and metastatic tumor cells. In addition, studies in CLIC1 knockout mice have shown that CLIC1 promotes platelet function. Here, we hypothesize that CLIC1 supports cell adhesive functions in platelets as well as endothelial cells and in doing so mediates critical vascular biological processes such as thrombus formation, vascular repair and angiogenesis.To assess the role of CLIC1 in endothelial cells, we monitored human umbilical venous endothelial cells (HUVEC) after transfection with CLIC1 Smartpool® or non-targeted siRNA by phase contrast microscopy. This experiment revealed a significant anti-proliferative effect in CLIC1 knockdown cells in conventional 2-dimensional culture that coincided with reduced cell spreading. The deficit in spreading was reiterated after embedding the CLIC1-depleted HUVEC in a 3-dimensional matrix of fibrin clot and this loss in anchorage led to a dramatic increase in cell death.Immunocytochemistry and fluorescence microscopy of HUVEC 10 minutes after initial cell attachment showed strong CLIC1 expression in nascent lamellipodia, where it colocalized with F-actin. Recruitment of CLIC1 to the cell membrane was associated with lamellipodia formation and was lost at later stages of adhesion when HUVEC began to generate stress fibers. CLIC1 cell membrane expression was inhibited after knocking down integrin β3 or treating HUVEC with the small molecule CLIC1 inhibitor IAA94. Notably, inhibiting CLIC1 or integrin β3 caused a significant reduction of F-actin assembly in lamellipodia suggesting that both molecules cooperate during the initial phase of cell spreading.Paralleling our experiments with HUVEC, we observed that CLIC1 co-localizes with F-actin in the leading edges as well as in the cortical ring of newly attached platelets. Moreover, using flow cytometry, we detected robust CLIC1 cell surface expression in platelets after treatment with ADP, which was enhanced after addition of the β3 integrin ligand fibrinogen and reduced when we blocked integrins with peptides representing the integrin recognition motif RGD. These data suggest a close coordination of CLIC1 membrane recruitment with the adhesive machinery in activated platelets. Moreover, inhibition of CLIC1 relocation to the platelet membrane with IAA94 hindered activation of integrin αIIbβ3, which resulted in impaired platelet aggregation.Lastly, we tested whether CLIC1 inhibition has antithrombotic effects in vivo. To this end, we assessed vasoocclusion in a murine dorsal skinfold chamber model where blood flow was monitored by intravital fluorescence microscopy. Intraperitoneal injections of mice with the CLIC1 inhibitor IAA94 (20mg/kg) 19 hours and 1 hour before the photochemical induction of thrombus formation caused a significant delay in vaso-occlusion in response to CLIC1 inhibition compared to vehicle-treated controls indicating that CLIC1 inhibition has a significant anti-thrombotic effects in vivo. Together, our data show that CLIC1 cooperates with integrins during cell adhesion and as such mediates functions related to platelet homeostasis, thrombus formation, and angiogenesis. DisclosuresNo relevant conflicts of interest to declare.

Chloride intracellular channel 1 as a switch among tumor behaviors in human esophageal squamous cell carcinoma.

Background: Recent studies have reported important roles for chloride intracellular channel 1 (CLIC1) in various cancers; however, its involvement in esophageal squamous cell carcinoma (ESCC) remains unclear. The aim of the present study was to investigate the role of CLIC1 in human ESCC. Methods: CLIC1 expression in human ESCC cell lines was analyzed by Western blotting. Knockdown experiments were conducted with CLIC1 siRNA, and their effects on cell proliferation, the cell cycle, apoptosis, migration, and invasion were analyzed. The gene expression profiles of cells were analyzed using a microarray analysis. An immunohistochemical analysis was performed on 61 primary tumor samples obtained from ESCC patients who underwent esophagectomy. Results: ESCC cells strongly expressed CLIC1. The depletion of CLIC1 using siRNA inhibited cell proliferation, induced apoptosis, and promoted cell migration and invasion. The results of the microarray analysis revealed that the depletion of CLIC1 regulated apoptosis via the TLR2/JNK pathway. Immunohistochemistry showed that CLIC1 was present in the cytoplasm of carcinoma cells, and that the very strong or very weak expression of CLIC1 was an independent poor prognostic factor. Conclusions: The present results suggest that the very strong expression of CLIC1 enhances tumor survival, while its very weak expression promotes cellular movement. The present study provides an insight into the role of CLIC1 as a switch among tumor behaviors in ESCC.

Expression and prognostic value of CLIC1 in epithelial ovarian cancer.

The clinical significance of the chloride intracellular channel 1 (CLIC1) protein in ovarian cancer is yet to be determined. The present study aimed to investigate the association between CLIC1 expression, and clinicopathological features and prognosis of patients with epithelial ovarian cancer. In this retrospective study, CLIC1 level was determined by reverse transcription-quantitative polymerase chain reaction, western blotting and immunohistochemical staining. The association between CLIC1 expression and clinicopathological characteristics were evaluated. Progression-free survival and overall survival were assessed by univariate, and multivariate analyses. mRNA and protein levels of CLIC1 were significantly higher in cancerous tissues than in healthy ovarian tissues (P<0.001). CLIC1 signals in epithelial ovarian cancer tissues were significantly higher than that in healthy tissues (P<0.001). CLIC1 expression was significantly higher in higher-grade tumors than in low-grade tumors (P<0.001). Moreover, overexpression of CLIC1 was associated with cisplatin resistance (P<0.001). CLIC1 expression was an independent factor that predicted shorter progression-free survival (P=0.006) and overall survival (P=0.002) for patients with epithelial ovarian cancer. These findings indicate that CLIC1 is overexpressed and is associated with poor prognosis in patients with epithelial ovarian cancer.

Decreased expression of hsa‑miR‑372 predicts poor prognosis in patients with gallbladder cancer by affecting chloride intracellular channel 1.

It has been reported that hsa‑microRNA (miRNA/miR)‑372 functions as a tumor suppressor or oncogene in various digestive system tumors, however, its roles in gallbladder cancer (GBC) are yet to be established. The present study aimed to determine the expression and clinical relevance of hsa‑miR‑372 in GBC. The expression of hsa‑miR‑372 in 80 pairs of human GBC tissues and adjacent normal gallbladder tissues was measured by reverse transcription‑quantitative polymerase chain reaction. Subsequently, the associations between hsa‑miR‑372 expression levels and the clinicopathological characteristics of patients with GBC were determined using χ2 test. Furthermore, Kaplan‑Meier method and Cox regression analysis were performed to evaluate the association between hsa‑miR‑372 expression and the prognosis of patients with GBC. Furthermore, a dual‑luciferase reporter assay and western blot analysis were performed to predict and verify the target gene of hsa‑miR‑372. The results demonstrated that markedly lower hsa‑miR‑372 expression was observed in GBC tissues, which was associated with poor prognosis in patients with GBC. Downregulated expression of hsa‑miR‑372 was negatively associated with tumor histological grade, tumor‑node‑metastasis stage, lymph node metastasis and distant metastasis, however, no association was observed between reduced hsa‑miR‑372 expression and patient gender, age, tumor size and gallbladder stones. Multivariate Cox regression analysis revealed that hsa‑miR‑372 expression, histological grade and lymph node metastasis were independent prognostic factors for overall survival in patients with GBC. Chloride intracellular channel 1 (CLIC1) was previously reported to be an effective biomarker for predicting the prognosis of GBC. Notably, the results of the present study indicated that CLIC1 may be a direct target gene of hsa‑miR‑372. In conclusion, the current study provides the first statistically convincing evidence that downregulation of hsa‑miR‑372 may occur in GBC tissues, which may be associated with aggressive and progressive tumor behavior by affecting CLIC1 expression.

Chloride intracellular channel 1 regulates migration and invasion in gastric cancer by triggering the ROS-mediated p38 MAPK signaling pathway.

Mol Med Rep 12:[Related article:] 8041–8047, 2015; DOI: 10.3892/mmr.2015.4459 Following the publication of this article, an interested reader drew to our attention anomalies associated with the presentation of Fig. 4A. In the original presentation of the Figure, different representations of the same photomicrograph were inadvertently used for the panels labelled 'N', 'H-R', 'H-R + NAC' and 'H-R + SB203580'. This error arose as a consequence of an incorrect ordering of our original image numbers. A corrected version of Fig. 4A is provided below, featuring the appropriate data for each of the five panels listed above. This Figure was intended to show how, compared with the normoxic control group, the invasiveness of the SGC-7901 gastric cancer cells was markedly increased under hypoxia-reoxygenation (H-R) conditions, although pretreatment with N-acetyl cysteine (NAC; 30 mmol/l) or indanyloxyacetic acid 94 (IAA-94; 40 μmol/l), inhibited the invasiveness resulting from H-R exposure. Similarly, treatment with the inhibitor of p38 mitogen-activated protein kinase, SB203580 (10 μmol/l), also inhibited the cell invasiveness induced under the H-R conditions. We sincerely apologize for this mistake, and thank the reader of our article who drew this matter to our attention. Furthermore, we regret any inconvenience this mistake has caused.

CLIC1 overexpression is associated with poor prognosis in pancreatic ductal adenocarcinomas.

Clinical significance of chloride intracellular channel 1 (CLIC1) in pancreatic ductal adenocarcinomas (PDAC) remains largely unknown. This study was performed to assess the expression of CLIC1 in benign and malignant pancreatic lesions, and to assess its clinicopathological significance. Tissue samples from resected PDAC (n = 70) and their matched normal pairs were evaluated for CLIC1 expression by immunohistochemical staining. Their expression was correlated with different clinicopathological parameters. CLIC1 expression was significantly higher (67.1%) in PDAC than in adjacent control tissues (25.7%, P < 0.001). High CLIC1 levels were associated with the histological grade (P < 0.001) and tumor size (P < 0.001); but not with sex, age, tumor-node-metastasis (TNM) stage, tumor location, or lymph node metastasis (P < 0.05). Univariate Kaplan-Meier analysis showed that a positive CLIC1 expression was associated with a decreased overall survival (P < 0.01). Multivariate cox regression analysis showed that CLIC1 expression and lymph node metastasis were independent risk factors for disease-free survival. The expression of CLIC1 might be closely related to the carcinogenesis, clinical biological behaviors, and prognosis of pancreatic ductal adenocarcinomas.

Chloride intracellular channel 1 regulates migration and invasion in gastric cancer by triggering the ROS-mediated p38 MAPK signaling pathway.

Chloride intracellular channel 1 (CLIC1) has been demonstrated to be overexpressed in gastric cancer, and elevated CLIC1 expression levels are markedly associated with the processes of tumor cell migration and invasion. However, the regulatory mechanism and signaling pathway underlying these processes have remained to be elucidated. The present study examined the impact of N-acetyl cysteine (NAC), indanyloxyacetic acid (IAA)-94 and SB203580, inhibitors of reactive oxygen species (ROS), as well as CLIC1 and p38 mitogen-activated protein kinase (MAPK) on the migration and invasion of SGC-7901 gastric cancer cells in a hypoxia-reoxygenation (H-R) microenvironment. The results demonstrated that intracellular ROS and CLIC1 levels were increased under H-R conditions, and that functional inhibition of CLIC1 significantly decreased the H-R-elevated ROS generation and p-p38 MAPK levels in SGC-7901 cells, as well as inhibited the migration and invasion of SGC-7901 cells. In addition, the expression levels of MMP-2 and MMP-9 were inhibited by NAC, IAA-94 and SB203580. These results indicated that CLIC1 regulates gastric cancer-cell migration and invasion via the ROS-mediated p38 MAPK signaling pathway.

Chloride intracellular channel 1 participates in migration and invasion of hepatocellular carcinoma by targeting maspin.

Our previous proteomic research found that chloride intracellular channel 1 (CLIC1) was upregulated in hepatocellular carcinoma (HCC) tissues with portal vein tumor thrombus. The present study aimed to determine the role of CLIC1 in HCC invasion. Immunohistochemistry was used to explore protein expression of CLIC1 in 15 cirrhotic tissues and 69 pairs of HCC and paracarcinoma tissues. Small interfering RNA (siRNA) and plasmids were transfected into HepG2 and SMMC7721 cells, and the in vitro function of CLIC1 in these cells were assessed with cell counting kit-8 assays, cell apoptosis assays, scratch assays, and transwell assays. Microarray analysis was also performed to further explore the candidate genes related to CLIC1. Our results confirmed that upregulated CLIC1 expression was significantly correlated with vascular invasion (P = 0.034) in HCC tissues. Knockdown of CLIC1 decreased cell viability and the invasive potency of HepG2 cells, whereas CLIC1 overexpression resulted in an opposite effect in SMMC7721 cells. Microarray analysis identified 618 genes that were differentially expressed (fold change ≥ 2, P < 0.05) between HepG2 cells transfected with CLIC1 siRNA and the negative control. Further studies indicate that knockdown of CLIC1 increased maspin expression and reduced vascular endothelial growth factor (VEGF), matrixmetalloproteinase-2 (MMP2), MMP9, MMP11, and MMP12 expression. In contrast, overexpression of CLIC1 decreased maspin expression and increased VEGF, MMP2, MMP12, and MMP13 expression. CLIC1 protein expression is significantly correlated with vascular invasion, and the present study suggests a previously unknown mechanism of CLIC1-mediated control of HCC invasiveness by targeting maspin.

CLIC1 overexpression is associated with poor prognosis in gallbladder cancer.

The chloride intracellular channel 1 (CLIC1) gene family is a recently identified class of Cl channel proteins. Although CLIC1 involvement is well established in some cancers such as gastric cancer and colon cancer, its expression pattern in gallbladder cancer (GBC) remains unknown. The aim of our study was to investigate the expression of CLIC1 in relation to progression and prognosis of GBC. Eight fresh gallbladder cancers paired with adjacent non-tumour tissues were quantified using real-time PCR and Western blot. Tissue samples from resected gallbladder cancer (n = 75) and cholelithiasis (n = 75) were evaluated for CLIC1 expression by immunohistochemical staining. Their expression was correlated with different clinicopathological parameters. CLIC1 expression was significantly higher (62.7%) in gallbladder cancer than in cholelithiasis (21.3%, p < 0.001). CLIC1 levels were associated with the histological grade, TNM stage and perineural invasion (p < 0.05), but not with patient age, sex, lymph node metastasis or gallbladder stones (p > 0.05). Univariate Kaplan-Meier analysis showed that a positive CLIC1 expression was associated with a decreased overall survival (p < 0.001). Multivariate Cox regression analysis showed that CLIC1 expression and histological grade were independent risk factors for overall survival. Therefore, the expression of CLIC1 is closely related to the progression of GBC and may be used as an effective marker for predicting the prognosis of this disease.