Clusterin (CLU) is expressed in a wide variety of human tissues and fluids. Overexpression of cytoplasmic clusterin (sCLU) has been implicated in cancer development and progression. The aim of the present study is to evaluate the association of sCLU overexpression with clinicopathological features of human gastric carcinomas (GC).We constructed a gastric cancer tissue microarray containing 173 primary gastric carcinomas and 70 paired non-neoplastic mucosa specimens. The expression of sCLU was studied by immunohistochemistry. The correlations between sCLU expression and clinicopathological features, p53 abnormality, as well as Ki67 activation were analyzed. Overexpressions of sCLU was detected in 28.5% (n=165) of primary GCs by immunohistochemical staining, but not in non-neoplastic mucosa. Clinical association study found that overexpression of sCLU was significantly correlated with lymph-node metastasis (p < 0.001), tumor invasion (p < 0.001) and TNM stage (p < 0.001). In Kaplan-Meier survival analysis, overexpression of sCLU was significantly correlated with unfavorable survival in advanced GCs (p < 0.03). Furthermore, the association of sCLU with abnormal expression of p53 was ascertained. These results suggested that overexpression of sCLU was involved in the progression of GC and it's oncogenic function might be associated with p53 abnormality. Overexpression of sCLU seems to be related with patient's shorter survival in late stage GC.

We used methylation-sensitive restriction fingerprinting (MSRF) to identify novel CpG (5'-CG-3' palindrome; p, phosphate group)-rich sequences that are methylated differentially between the hepatocellular carcinoma (HCC) genomes and adjacent nontumorous liver tissues. A 199-base-pair sequence methylated in HCC tumor tissue was isolated and showed high homology to the 5' CpG island of the zinc fingers and homeoboxes protein 2 (ZHX2) gene. By using bisulfite sequencing, we confirmed that hypermethylation of the 5' CpG island of ZHX2 occurred in some HCC and HepG2 cell lines but not in 6 normal liver tissue samples. By using methylation-specific polymerase chain reaction, we detected methylation of the 5' CpG island of ZHX2 in 46.9% of the HCCs. Reverse transcription-polymerase chain reaction demonstrated that ZHX2 messenger RNA (mRNA) was expressed in all 6 normal liver tissue samples but in only 13.3% of the methylated HCCs. Treatment of HepG2 with 5-aza-deoxycytidine could demethylate the promoter and increase ZHX2 mRNA expression. These results suggest that hypermethylation-mediated silencing of ZHX2 is an epigenetic event involved in HCC.

We used methylation-sensitive restriction fingerprinting (MSRF) to identify novel CpG (5'-CG-3' palindrome; p, phosphate group)-rich sequences that are methylated differentially between the hepatocellular carcinoma (HCC) genomes and adjacent nontumorous liver tissues. A 199-base-pair sequence methylated in HCC tumor tissue was isolated and showed high homology to the 5' CpG island of the zinc fingers and homeoboxes protein 2 (ZHX2) gene. By using bisulfite sequencing, we confirmed that hypermethylation of the 5' CpG island of ZHX2 occurred in some HCC and HepG2 cell lines but not in 6 normal liver tissue samples. By using methylation-specific polymerase chain reaction, we detected methylation of the 5' CpG island of ZHX2 in 46.9% of the HCCs. Reverse transcription-polymerase chain reaction demonstrated that ZHX2 messenger RNA (mRNA) was expressed in all 6 normal liver tissue samples but in only 13.3% of the methylated HCCs. Treatment of HepG2 with 5-aza-deoxycytidine could demethylate the promoter and increase ZHX2 mRNA expression. These results suggest that hypermethylation-mediated silencing of ZHX2 is an epigenetic event involved in HCC.

Objective: Hepatocellular carcinoma (HCC) is one of the most common malignant tumors with a high rate of mortality. Our previous study shows the expression of calponin 2 (CNN2) is up-regulated in hepatocellular carcinoma tissues, especially in metastatic ones. To better understand the role of CNN2 in HCC, RNA interference (RNAi) was used to explore its role in tumor growth and metastasis.Methods: Lentivirus-mediated CNN2-shRNA was transfected into SK-hep-1 cells, and the efficacy of CNN2 expression, cell migration, invasion, proliferation and cell cycles were evaluated by quantitative real-time polymerase chain reaction (qRT-PCR), Western blot (WB), Transwell assay, methyl thiazol tetrazolium assay and flow cytometry, respectively. SK-hep-1 cells transfected with Lentivirus-CNN2 shRNA were xenografted in Balb/C nude mice to explore the effect of CNN2-shRNA in tumor growth. Xenograft tumor tissues were examined for their histopathology, cell apoptosis, the expression of total protein and their corresponding phosphorylated protein of MEK1/2, ERK1/2, AKT, by hematoxylin and eosin stain (H & E staining), TUNEL assay, immunohistochemical technique, respectively.Results: Our research shows it is evident that CNN2 shRNA can effectively down-regulate the expressions of CNN2 mRNA and protein, inhibit cell proliferations, arrest cell cycles at the S phase and reduce cell migration and invasion. SK-hep-1 cells with CNN2 down-regulation have markedly attenuated tumor growth in nude mice. Xenograft tumor tissues have displayed typical tumor characteristics and no apoptosis is detected in shRNA group or in control group. No metastatic tumor was found in any group of nude mice. With CNN2 protein down-regulation, the protein of pMEK1/2 and pERK1/2 are effectively down-regulated, except pAKT, AKT, MEK1/2 and ERK1/2.Conclusions: CNN2 plays an important role in tumor growth and metastasis, possibly through MEK1/2-ERK1/2 signaling pathway. Our study illustrate that CNN2 might be a potential target in HCC molecular target therapy.

Rationale: Glycogen storage disease (GSD) type VI is a rare disease caused by the inherited deficiency of liver phosphorylase. Patient concerns: The proband, a 61-month-old Chinese boy, manifested intermittent hematochezia, growth retardation, hepatomegaly, damage of liver function, mild hypoglycemia, and hyperlactatemia. The other patient was a 107-month-old Chinese girl with growth retardation, hepatomegaly, mild hypoglycemia, and hyperlactatemia. In order to further confirm the diagnosis, we conducted a liver biopsy and detected blood samples for their gene using IDT exon chip capture and high-throughput sequencing. Diagnoses: According to the clinical symptoms, physical examination, laboratory examinations, liver biopsy, and the genetic test finding, the 2 patients were diagnosed GSD VI. Interventions: They were treated mainly with uncooked cornstarch. Outcomes: There were 2 mutations of PYGL gene in this pedigree. c.2467C>T (p. Q823X) and c.2178-2A>C occurred both in the proband and his second sister. Lessons: As a novel mutation, c.2178-2A>C enriches the mutation spectrum of PYGL gene. The different degrees of elevated lactate is an unusual phenotype in GSD VI patients. It is not clear if this is caused by the new mutation of c. 2178-2A > C. Long-term complications remains to be observed.

Senescence marker protein 30 (SMP30) has been identified as a tumor-related molecule of hepatocellular carcinoma (HCC). Its clinical significance and underlying mechanisms in HCC tissues, however, remain largely unexplored. We have demonstrated a preferentially expressed SMP30 in normal liver using a tissue microarray. By employing real-time quantitative PCR, two tissue microarrays and Oncomine database analysis, we have also shown that the SMP30 in HCC tissues has significantly reduced when compared with that in paired adjacent non-tumor tissues (P = 0.0037). The reduced expression of SMP30 is very noticeably related to larger tumor size (P = 0.012), enhanced TNM (P = 0.009) and worse survival (P < 0.0001) in HCC patients. The analyses using Cox regression have indicated that the decreased SMP30 expression is an independent risk to the reduced overall survival rate of HCC patients (P = 0.001), and the down-regulation of SMP30 in HCC might be mediated by DNA methylation. Moreover, genes co-expressed with SMP30 may affect the prognosis through apoptotic process, biological adhesion and blood coagulation by PANTHER analyses. Our studies have indicated that the SMP30 may serve as a candidate of HCC clinical prognostic marker and a potential therapeutic target.

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Anti-SMP30 antibody levels can be used as a biomarker for diagnosing HCC; marked results have been observed for patients with alpha-fetoprotein (AFP) negativity, in particular.