Glutamate (mGluR-5) gene expression in brain regions of streptozotocin induced diabetic rats as a function of age: role in regulation of calcium release from the pancreatic islets in vitro

Metastatic renal cell carcinoma (RCC) is highly resistant to conventional systemic treatments, including chemotherapy, radiotherapy and hormonal therapies. Previous studies have shown over-expression of EGFR is associated with high grade tumors and a worse prognosis. Recent studies suggest anticancer therapies targeting the EGFR pathway have shown promising results in clinical trials of RCC patients. Therefore, characterization of the level and localization of EGFR expression in RCC is important for target-dependent therapy. In this study, we investigated the clinical significance of cellular localization of EGFR in human normal renal cortex and RCC. RCC and adjacent normal kidney tissues of 63 patients were obtained for characterization of EGFR expression. EGFR protein expression was assessed by immunohistochemistry on a scale from 0 to 300 (percentage of positive cells × staining intensity) and Western blotting. EGFR membranous staining was significantly stronger in RCC tumors than in normal tissues (P < 0.001). In contrast, EGFR cytoplasmic staining was significantly higher in normal than in tumor tissues (P < 0.001). The levels of membranous or cytoplasmic EGFR expression in RCC tissues were not correlated with sex, tumor grade, TNM stage or overall survival (P > 0.05). These results showed abundant expression of membranous EGFR in RCC, and abundant expression of cytoplasmic EGFR in normal tissues. EGFR expression in RCC was mostly located in the cell membrane, whereas the EGFR expression in normal renal tissues was chiefly seen in cytoplasm. Our results suggest different locations of EGFR expression may be associated with human renal tumorigenesis.     

Down-regulation of HAI-1 is associated with poor-differentiation status of colorectal cancer

Hepatocyte growth factor activator inhibitor type 1 (HAI-1) is a Kunitz-type serine protease inhibitor which is widely expressed in epithelial cells. The purpose of this study is to investigate the relationship between HAI-1 expression and differentiation status of colorectal epithelia. The expression of HAI-1 in clinical samples of both cancerous and normal colorectal tissues was evaluated by immunohistochemical staining. An in vitro epithelial differentiation model of Caco-2 cell was established, and the characteristics of differentiation of Caco-2 cells were observed by transmission electron microscopy. The expression of HAI-1 in Caco-2 cells at different differentiation stages was examined by western blot. Immunohistochemical staining of 52 human colorectal cancer tissues showed a definite correlation between HAI-1 expression and differentiation status: IHC score (mean ± SE) of HAI-1 was higher for well- or moderately-differentiated colorectal cancer tissues than for poorly-differentiated colorectal cancer tissues, with significant differences in HAI-1 positive rate (P < 0.01 and P < 0.05 for well-differentiated vs. poorly-differentiated and moderately-differentiated vs. poorly-differentiated, respectively). Immunohistochemical staining of normal colorectal tissues showed positive HAI-1 expression in well-differentiated epithelial cells whereas the under-differentiated crypt cells showed very weak HAI-1 staining signals. The result of western blot also showed the gradual increasing of HAI-1 expression during the process of Caco-2 differentiation in vitro. HAI-1 expression correlates with the differentiation status of colorectal epithelia and could serve as a differentiation marker.     

Association of the plasma riboflavin levels and riboflavin transporter (C20orf54) gene statuses in Kazak esophageal squamous cell carcinoma patients

To evaluate the association of the plasma riboflavin level in Kazak esophageal cancer patients and their riboflavin transporter (C20orf54) gene statuses. Plasma riboflavin levels were detected by high performance liquid chromatography in Kazak patients with esophageal squamous cell carcinoma (ESCC) and healthy controls. C20orf54 mRNA and protein expression were analyzed by real-time fluorogenic quantitative polymerase chain reaction and immunohistochemistry in samples from 61 ESCC patients consisting of both tumor and normal tissue, respectively. C20orf54 mRNA expression was decreased in ESCC (0.279 ± 0.102) than in normal counterpart tissue (0.479 ± 0.287; P = 0.049) significantly. Tumors exhibited low C20orf54 protein expression (42.6, 26.2, 18.0 and 13.1 % for no C20orf54 staining, weak staining, medium staining and strong staining, respectively), which was significantly lower than that in the normal mucous membrane (13.1, 26.2, 41.0 and 19.7 % for no C20orf54 staining, weak staining, medium staining and strong staining, respectively). Defective expression of C20orf54 in tumor cells was significantly associated with poor differentiation. However, other parameters such as depth of invasion and lymph node metastasis had no significant relationship with C20orf54 expression. The average blood concentration of riboflavin was 2.6468 ± 1.3474 ng/ml in ESCC patients lower than control group (4.2960 ± 3.2293 ng/ml, P = 0.015). A positive correlation of plasma riboflavin levels with defective expression of C20orf54 protein was found in ESCC patients (F = 8.626; P = 0.038). Defective expression of C20orf54 is associated with the development of Kazak esophageal squamous cell carcinoma and this may represent a mechanism underlying the decreased plasma riboflavin levels in ESCC.     

SFRP2 and Slug Contribute to Cellular Resistance to Apoptosis in Hypertrophic Scars

Hypertrophic scars (HS) are skin disorders which occur after wounding and thermal injury. Our previous studies have suggested that secreted frizzled-related protein 2 (SFRP2) is involved in HS formation and that the suppression of SFRP2 promotes apoptosis of hypertrophic scar fibroblasts (HSFBs). However, the mechanisms have not been clarified. Previous studies revealed that Slug expression inhibits cell apoptosis, in vitro and in vivo, and SFRP2 regulates the expression of Slug in cervical cancer cells. In the present study, we quantified differential expression levels of expression of SFRP2 and Slug in HS and normal skin tissues by immunohistochemistry, both of which have important anti-apoptosis roles. Furthermore, a short hairpin RNA approach was adopted to investigate the potential function of SFRP2 and Slug in HSFB apoptosis. Cell apoptosis was detected using fluorescence-activated cell sorting and Caspase-3 activity was assayed by spectrophotometry. This study demonstrates that SFRP2 expression, as well as Slug, is dramatically up-regulated in HS relative to normal skin tissues, and the Slug expression is positively correlated with SFRP2. Slug expression was down-regulated in SFRP2-deficient cells, and the down-regulation of Slug expression increased sensitivity to apoptosis which was induced through a caspase-3-dependent pathway. The infected cells with reduced levels of Slug were tested for the expression of apoptosis-related genes (Bcl-2, Bax and PUMA) which were previously identified as Slug targets. Bcl-2 expression was down-regulated in Slug-deficient cells. In conclusion, SFRP2 appears to interact with Slug to affect the apoptosis of hypertrophic scar fibroblasts.     

胸水中肺腺癌细胞与血清中PTN多效蛋白分子生物学的研究

通过检测PTN蛋白在肺腺癌患者术前血清标本及相对应的恶性胸水肺腺癌细胞2种不同标本中的表达及对比其表达的差异,探讨其诊断意义.利用Western-blot免疫印迹方法检测50例恶性胸水及相对应的术前血清,并对肺腺癌细胞进行石蜡包埋、免疫细胞化学检查.同时分别以10例正常献血者血清、20例胸水良性增生细胞作为对照.肺腺癌患者血清和恶性胸水细胞中PTN蛋白的表达分别高于对照组PTN蛋白的表达,恶性胸水中PTN蛋白的表达59.0％ (49/83)高于肺腺癌患者血清中PTN蛋白的表达32.5％ (27/83).差异均具有统计学意义(P＜0.05),恶性胸水肺腺癌细胞中的PTN蛋白表达和波形蛋白Vimentin呈正相关关系(P ＜0.01,r =0.728),而与钙粘连蛋白E-ca呈负相关.PTN蛋白在肺腺癌患者血清和恶性胸水细胞标本中高表达,恶性胸水肺腺癌细胞中PTN蛋白的表达高于血清中PTN蛋白的表达,肺腺癌细胞中PTN蛋白的表达与波形蛋白Vimentin表达相一致,肺腺癌细胞在转移过程中已发生了向间质细胞转化EMT的过程,同时增强了肺腺癌细胞的高侵袭性,而恶性胸水肺腺癌细胞PTN蛋白的高表达更促进了肺腺癌细胞的转移.提示对未发生胸水转移的肺腺癌患者进行血清中PTN蛋白的检测,对已发生胸水转移的肺腺癌患者同样要检测PTN蛋白,以期提高肺腺癌患者的诊断率.     

Reduced expression of NGEP is associated with high-grade prostate cancers: a tissue microarray analysis

New gene expressed in prostate (NGEP) is a newly diagnosed prostate-specific gene that is expressed only in normal prostate and prostate cancer cells. Discovery of tissue-specific markers may promote the development of novel targets for immunotherapy of prostate cancer. In the present study, the staining pattern and clinical significance of NGEP were evaluated in a series of prostate tissues composed of 123 prostate cancer, 19 high-grade prostatic intraepithelial neoplasia and 44 samples of benign prostate tissue included in tissue microarrays using immunohistochemistry. Our study demonstrated that NGEP localized mainly in the apical and lateral membranes and was also partially distributed in the cytoplasm of epithelial cells of normal prostate tissue. All of the examined prostate tissues expressed NGEP with a variety of intensities; the level of expression was significantly more in the benign prostate tissues compared to malignant prostate samples (P value <0.001). Among prostate adenocarcinoma samples, a significant and inverse correlation was observed between the intensity of NGEP expression and increased Gleason score (P = 0.007). Taken together, we found that NGEP protein is widely expressed in low-grade to high-grade prostate adenocarcinomas as well as benign prostate tissues, and the intensity of expression is inversely proportional to the level of malignancy. NGEP could be an attractive target for immune-based therapy of prostate cancer patients as an alternative to the conventional therapies particularly in indolent patients.     

Involvement of ZEB1 and E-cadherin in the invasion of lung squamous cell carcinoma

This study intended to investigate the expression of the ZEB1 and E-cadherin proteins in lung squamous cell carcinoma (LSCC) tissues and to examine the clinicopathological correlation between protein levels and LSCC. RT-PCR and Western blot were used to examine the expression of ZEB1 and E-cadherin mRNAs and proteins in LSCC tissues as well as in adjacent normal tissues, and then analyze the relationship between the clinicopathological characteristics and the expression changes of ZEB1 and E-cadherin mRNAs in LSCC. In addition, RNAi was used to knockdown the expression of the ZEB1 gene in Human HCC827 cells; subsequently, changes in the invasive ability of the resultant cells were studied. The positive rates of ZEB1 and E-cadherin mRNAs in LSCC tissues were 69.2 and 38.5 %, respectively. They differed significantly from the corresponding positive rates in the adjacent normal lung tissues (15.4 and 80.8 %, p < 0.05). There was a negative correlation between the protein levels of ZEB1 and E-cadherin in LSCC tissues (r = -0.714, p < 0.001); in addition, it was found that ZEB1 protein expression in LSCC tissues was significantly higher than that in the neighboring normal lung tissues (p < 0.05), and its expression was also significantly higher in patients with lymph node metastases and distant metastases compared to those patients without metastatic disease (p < 0.05). On the contrary, E-cadherin expression was significantly lower in LSCC tissues than that in the neighboring normal tissue (p < 0.05). It was lower in patients with lymph node metastasis and distant metastasis compared to patients without metastatic disease (p < 0.05). However, the expression of ZEB1 and E-cadherin was independent of gender, age, tumor size, or tumor differentiation level (p > 0.05). Transfection of ZEB1 siRNA into HCC827 cells significantly reduced the ZEB1 protein level (p < 0.01) and significantly elevated E-cadherin levels (p < 0.01). Moreover, significantly less ZEB1 siRNA-transfected cells migrated through Transwell chambers in the LSCC tissue than that in the control groups (untransfected or transfected with control siRNA, p < 0.01). The expression of the ZEB1 gene in LSCC tissues is downregulated with the expression of E-cadherin. On the other hand, the expression of siRNA against ZEB1 promotes E-cadherin expression and suppresses the invasive ability conferred by E-cadherin. In conclusion, our data suggested that overexpression of the ZEB1 gene is possibly associated with the occurrence, development, invasion of LSCC.     

Overexpression of integrin-linked kinase (ILK) is associated with tumor progression and an unfavorable prognosis in patients with colorectal cancer

Integrin-linked kinase (ILK), an intracellular serine-threonine kinase, has been reported to be overexpressed in multiple types of human malignancies, including colorectal cancer (CRC). The prognostic value of ILK in CRC, however, remains unknown. In the present study, expression of ILK in 25 paired primary CRC samples and adjacent noncancerous tissues were quantified using real-time PCR and Western blotting. ILK protein expression was analyzed in 102 archived, paraffin-embedded CRC samples using immunohistochemistry. The correlation between ILK expression and clinicopathological factors was evaluated by the χ² test. Patients’ overall survival was analyzed by Kaplan–Meier method. We found that both ILK mRNA and protein expression levels were significantly up-regulated in primary CRC samples compared with their corresponding normal tissues. Immunohistochemical analysis revealed relative high expression of ILK in 43 of 102 (42.2 %) primary CRC samples. Statistical analysis showed a significant correlation of ILK expression with tumor differentiation, lymph node metastasis, tumor invasion, and tumor-node-metastasis stage. Patients with tumors displaying high-level ILK expression showed significantly shorter overall survival (P = 0.028, log-rank test). More importantly, multivariate analysis indicated that high ILK protein expression was an independent prognostic factor for CRC patients (P = 0.026). Taken together, our data suggest that ILK overexpression is associated with tumor progression and a poor prognosis in CRC patients and may represent a novel potential prognostic marker for patients with CRC.     

Prognostic significance of survivin expression in renal cell cancer and its correlation with radioresistance

Survivin, an important inhibitor of apoptosis, has been found to play an important role in the initiation, progression, and chemoradioresistance of human malignancies. Previously, we have reported that upregulation of survivin in oral squamous cell carcinoma correlates with poor prognosis and chemoresistance. The aim of this study was to assess prognostic significance of survivin protein expression in RCC and analyze its correlation with radiosensitivity of RCC cells. RT-PCR and Western blot assays were performed to detect survivin mRNA and protein expression in normal human kidney epithelial cell line (HKEC) or RCC cell lines. The expression of survivin mRNA in RCC and corresponding nontumor kidney tissues was also detected by RT-PCR. Immunohistochemistry was performed to determine survivin protein expression in 75 cases of RCC tissue samples. Moreover, the association of survivin protein expression with clinicopathogical factors and prognosis of RCC patients was statistically analyzed. Small interfering RNA was used to knockdown the endogenous survivin expression in RCC cell line (ACHN) and evaluate the effects of survivin knockdown on proliferation, apoptosis, and radiosensitivity of RCC cell line. RCC cells showed sufficient expression of survivin mRNA and protein, but the expression of survivin gene was not detected in normal HKEC. Moreover, the expression level of survivin mRNA in RCC tissues was significantly higher than that in corresponding nontumor kidney tissues. The immunostaining of survivin protein was mainly located in cytoplasm of RCC tumor cells. Tumor pathological stage (P = 0.028), grade (P = 0.004), and lymph node metastasis (P = 0.017) of RCC patients were significantly correlated with survivin protein expression. In addition, patients with high survivin levels had a significantly shorter overall survival than those with low levels (P < 0.001), and the expression of survivin protein was an independent prognostic factor for RCC patients (P = 0.008). The expression of survivin gene could be reduced in RCC cell line and survivin knockdown could inhibit growth and enhance in vivo radiosensitivity of RCC cell line by inducing apoptosis enhancement. Taken together, the status of survivin protein expression may be an independent factor for predicting the prognosis of RCC patients and tumor-specific survivin knockdown combined with radiotherapy will be a potential strategy for RCC therapy.     

Expression and Biologic Significance of Adiponectin Receptors in Papillary Thyroid Carcinoma

Obesity is associated with a higher incidence of thyroid cancer. Adiponectin is one of the most abundant adipokines with a pleiotropic role in metabolism and in the development and progression of cancer. It has been shown that circulating adiponectin level is inversely associated with the risk of thyroid cancer. This study aimed to investigate the possible association between the expression of adiponectin receptors (AdipoR1 and AdipoR2) and clinicopathological variables in papillary thyroid cancer. We found that protein levels of AdipoR1 and AdipoR2 were increased in some thyroid cancer specimens compared with adjacent normal thyroid tissues. Thyroid cancer cells expressed AdipoR1 and AdipoR2, which were attenuated by histone deacetylase inhibitors valproic acid and trichostatin A. Adiponectin stimulated AMP-activated protein kinase phosphorylation in thyroid cancer cells. We further determined the expression of AdipoR1 and AdipoR2 by immunohistochemical staining in primary tumor samples and metastatic lymph nodes. AdipoR1 was expressed in 27 % of primary tumors and AdipoR2 in 47 %. Negative expression of both adiponectin receptors was significantly associated with extrathyroidal invasion, multicentricity, and higher TNM stage. There was a trend toward decreased disease-free survival in patients with negative tumor expression of AdipoR1 and AdipoR2 (log-rank P = 0.051). Collectively, overexpression of adiponectin receptors was observed in some tumor tissues of papillary thyroid cancer and was associated with a better prognosis.     

miR-145-5p attenuates hypertrophic scar via reducing Smad2/Smad3 expression

The study was designed to explore the underlying mechanism of micro ribonucleic acids (miR)-145-5p in the process of hypertrophic scar (HS). The difference in the relative content of miR-145-5p between HS and adjacent normal skin collected from 5 patients was detected via RT-PCR. Expressions of Smad2 and Smad3 with or without TGF-β1 was detected by western blotting. Fibroblasts apoptosis rate was examined by Annexin V/Propidium Iodide double staining. HS fibroblasts (HSFs) were isolated from HS tissues, cultured and then divided into control group, miR-145-5p inhibitor group (transfected with miR-145-5p inhibitor) and miR-145-5p mimic group (transfected with miR-145-5p plasmid) based on different treatment methods. Next, CCK-8 was employed to examine the function of miR-145-5p in HSF proliferation. Luciferase assay was conducted to confirm whether Smad2/3 were direct targets of miR-145-5p, and RT-PCR was done to measure the expression of miR-145-5p, Smad2/Smad3 and fibrosis-related genes of fibroblasts in three groups. Wound injury mice model was established to determine the function of miR-145-5p in regulating scar formation. miR-145-5p was found lowly expressed in HS tissues. Compared with Control group, miR-145-5p mimic decreased the levels of Smad2/3, arrested the activation and proliferation of HSFs and induced HSFs apoptosis. Overexpressing miR-145-5p achieved the contrary results. Smad2/3 was confirmed as the target of miR-145-5p. Moreover, miR-145-5p mimic decreased the recruitment of fibroblasts in vivo and decreased the expression of fibrosis-related genes after wound injury. In conclusion, miR-145-5p arrests the development of fibrogenesis and decreases HS formation by reducing the expression of Smad2/3. miR-145-5p may be an optional novel molecular target for treating HS.     

Methylation status and protein expression of RASSF1A in breast cancer patients

Recently genetics and epigenetics alterations have been found to be characteristic of malignancy and hence can be used as targets for detection of neoplasia. RAS association domain family protein 1A (RASSF1A) gene hypermethylation has been a subject of interest in recent researches on cancer breast patients. The aim of the present study was to evaluate whether RASSF1A methylation status and RASSF1A protein expression are associated with the major clinico-pathological parameters. One hundred and twenty breast cancer Egyptian patients and 100-control subjects diagnosed with benign lesions of the breast were enrolled in this study. We evaluated RASSF1A methylation status in tissue and serum samples using Methyl specific PCR together with RASSF1A protein expression in tissues by immunohistochemistry. Results were studied in relation to known prognostic clinicopathological features in breast cancer. Frequency of RASSF1A methylation in tissues and serum were 70 and 63.3 % respectively and RASSF1A protein expression showed frequency of 46.7 %. There was an association between RASSF1A methylation in tissues, serum and loss of protein expression in tissues with invasive carcinoma, advanced stage breast cancer, L.N. metastasis, ER/PR and HER2 negativity. RASSF1A methylation in serum showed high degree of concordance with methylation in tissues (Kappa = 0.851, P < 0.001). RASSF1A hypermethylation in tissues and serum and its protein expression may be a valid, reliable and sensitive tool for detection and follow up of breast cancer patients.     

Cholesteatoma Fibroblasts Promote Epithelial Cell Proliferation through Overexpression of Epiregulin

To investigate whether keratinocytes proliferate in response to epiregulin produced by subepithelial fibroblasts derived from middle ear cholesteatoma. Tissue samples were obtained from patients undergoing tympanoplasty. The quantitative polymerase chain reaction and immunohistochemistry were performed to examine epiregulin expression and localization in cholesteatoma tissues and retroauricular skin tissues. Fibroblasts were cultured from cholesteatoma tissues and from normal retroauricular skin. These fibroblasts were used as feeder cells for culture with a human keratinocyte cell line (PHK16-0b). To investigate the role of epiregulin in colony formation by PHK16-0b cells, epiregulin mRNA expression was knocked down in fibroblasts by using short interfering RNA and epiregulin protein was blocked with a neutralizing antibody. Epiregulin mRNA expression was significantly elevated in cholesteatoma tissues compared with that in normal retroauricular skin. Staining for epiregulin was more intense in the epithelial cells and subepithelial fibroblasts of cholesteatoma tissues than in retroauricular skin. When PHK16-0b cells were cultured with cholesteatoma fibroblasts, their colony-forming efficiency was 50% higher than when these cells were cultured with normal skin fibroblasts. Also, knockdown of epiregulin mRNA in cholesteatoma fibroblasts led to greater suppression of colony formation than knockdown in skin fibroblasts. Furthermore, the colony-forming efficiency of PHK16-0b cells was significantly reduced after treatment with an epiregulin neutralizing antibody in co-culture with cholesteatoma fibroblasts, but not in co-culture with skin fibroblasts. These results suggest that keratinocyte hyperproliferation in cholesteatoma is promoted through overexpression of epiregulin by subepithelial fibroblasts via epithelial–mesenchymal interactions, which may play a crucial role in the pathogenesis of middle ear cholesteatoma.     

Cross-talk between macrophages,smooth muscle cells,and endothelial cells in response to cigarette smoke: the effects on MMP2 and 9

The aim of the present study was to analyze the expression of sex-determining region Y-related high mobility group box 4 (SOX4) in non-small cell lung cancer (NSCLC) and its correlation with clinicopathologic characteristics, including the survival of NSCLC patients. To observe initially the expression status of SOX4 in lung squamous cell carcinoma and adenocarcinoma at gene expression omnibus. The expression of SOX4 mRNA and protein was examined in NSCLC tissues and normal lung tissues through real-time PCR and immunohistochemistry. Meanwhile, the relationship of SOX4 expression levels with clinical characteristics of 168 NSCLC patients was analyzed by immunohistochemistry. Univariate and multivariate analyses were performed to determine the association between SOX4 expression and prognosis of NSCLC patients. In our results, SOX4 expression was increased in NSCLC tissues compared with paired normal lung tissues in microarray data (GSE3268). SOX4 mRNA and protein expression were markedly higher in NSCLC tissues than in normal lung tissues (P = 0.001 and P = 0.001, respectively). Using immunohistochemistry, high levels of SOX4 protein were positively correlated with status of differentiated degree (high vs. middle, P = 0.004; high vs. low, P < 0.001), clinical stage (I–II vs. III–IV, P < 0.001), T classification (T1–T2 vs. T3–T4, P = 0.004), N classification (N0–N1 vs. N2–N3, P = 0.002), and M classification (M0 vs. M1, P = 0.011) in NSCLC. Moreover, the higher level of SOX4 expression was markedly correlated with poor overall survival in NSCLC patients (P < 0.001). Multivariate analysis suggested that increased SOX4 expression was a poor independent prognostic predictor for NSCLC patients (P = 0.002). In conclusion, SOX4 plays an important role on NSCLC progression and prognosis and may serve as a convictive prognostic biomarker for NSCLC patients.     

MicroRNA-192 regulates hypertrophic scar fibrosis by targeting SIP1

Hypertrophic scar (HS) is a fibro-proliferative disorder which is characterized by excessive deposition of collagen and accumulative activity of myofibroblasts. Increasing evidences have demonstrated miRNAs play a pivotal role in the pathogenesis of HS. MiR-192 is closely associated with renal fibrosis, but its effect on HS formation and skin fibrosis remains unknown. In the study, we presented that miR-192 was up-regulated in HS and HS derived fibroblasts (HSFs) compared to normal skin (NS) and NS derived fibroblasts (NSFs), accompanied by the reduction of smad interacting protein 1 (SIP1) expression and the increase of Col1, Col3 and α-SMA levels. Furthermore, we confirmed SIP1 was a direct target of miR-192 by using luciferase reporter assays. Meanwhile, the overexpression of miR-192 increased the levels of Col1, Col3 and α-SMA. The synthesis of collagen and more positive α-SMA staining were also observed in bleomycin-induced dermal fibrosis model of BALB/c mice treated with subcutaneous miR-192 mimics injection, whereas the inhibition of miR-192 decreased the expression of Col1, Col3 and α-SMA. Moreover, SIP1 siRNA could enhance the levels of Col1, Col3 and α-SMA, showing that the effect of knockdown SIP1 was similar to miR-192 mimics, and the phenomenon manifested miR-192 regulated HS fibrosis by targeting SIP1. Together, our results indicated that miR-192 was a critical factor of HS formation and facilitated skin fibrosis by targeting directly SIP1.     

Reduced expression of SOCS2 and SOCS6 in hepatocellular carcinoma correlates with aggressive tumor progression and poor prognosis

To investigate the clinical significance of suppressor of cytokine signaling (SOCS)-2 and SOCS6 in human hepatocellular carcinoma (HCC). The expression levels of SOCS2 and SOCS6 mRNA and protein in tumor, para-tumor and normal liver tissues were detected in 106 HCC patients by real-time quantitative RT-PCR (qRT-PCR) and Western blot. According to qRT-PCR and western blot analyses, we first found that both the expression levels of SOCS2 and SOCS6 mRNA and protein in HCC were significantly lower than those in para-tumor (both P < 0.001) and normal liver tissues (both P < 0.001). Then, the correlation analysis showed that both SOCS2 and SOCS6 protein downregulation were significantly correlated with advanced TNM stage (both P < 0.001) and high serum AFP (P = 0.008 and 0.01, respectively). Especially, the reduced expression of SOCS2 more frequently occurred in HCC patients with vascular invasion (P = 0.03), and that of SOCS6 was also associated with tumor recurrence (P = 0.01). Moreover, HCC patients with low expression of SOCS2 and SOCS6 had significantly shorter overall (P = 0.008 and 0.01, respectively) and disease-free survival (both P = 0.01). Furthermore, multivariate analysis showed that both SOCS2 and SOCS6 downregulation were independent prognostic factors of overall (P = 0.01 and 0.03, respectively) and disease-free survival (P = 0.01 and 0.03, respectively) in HCC. Our data demonstrate for the first time that SOCS2 and SOCS6 expression were remarkably reduced in HCC and may be served as potential prognostic markers for patients with this deadly disease.     

LMP gene promoter hypermethylation is a mechanism for its down regulation in Kazak’s esophageal squamous cell carcinomas

Abnormal hypermethylation of CpG islands not only associated with tumor suppressor genes can lead to repression of gene expression, but also contribute to escape of the tumor from immune surveillance and contribute significantly to tumorigenesis. In the present study, we studied the hypermethylation of low molecular-weight protein (LMP) gene and its regulation on protein expression in biopsies from resected tissues from Kazak’s esophageal squamous cell carcinoma (ESCC) patients and their neighboring normal tissues. LMP2 and LMP7 genes promoter region methylation sequences were maped in esophageal cancer cell line Eca109 by bisulfite-sequencing PCR and quantitative detection of methylated DNA from 30 pairs of Kazak’s ESCC and adjacent normal tissues by MassARRAY (Sequenom, San Diego, CA, USA) and LMP2 and LMP7 protein expression were analyzed with immunohistochemistry. In Eca109, we identified 6 CG sites methylated from all of 22 CpG sites of LMP7 gene. However, no methylation was found for LMP2. The analysis of the data resulted from the quantitative analysis of single CpG site methylation by Sequenom MassARRAY platform, has shown that the methylation level between two groups CpG sites (CpG_5, CpG_9, CpG_20, CpG_21 and CpG_20) from CpG_1, CpG_2, CpG_3, CpG_4, CpG_5, CpG_6, CpG_7, CpG_8, CpG_9, CpG_10.11, CpG_12.13.14, CpG_15.16.17.18, CpG_19, CpG_20, CpG_21 and CpG_22 significant differences between ESCC and neighboring normal tissues. The analysis of methylation level of whole target CpG fragment indicated that the methylation level of LMP7 was significant higher in ESCC (0.0517 ± 0.0357) than in neighboring normal tissues (0.0380 ± 0.0214, P < 0.05). there was a tendency of decreasing the LMP7 proteins expression as the increasing the methylation level of LMP7 gene promoter regions (F = 7.69, P = 0.041). The LMP7 gene promoter methylation and protein downregulation were correlated at high extent in Kazakh’s ESCC patients, and may explain the epigenetic regulation on gene expression.