Genetic alterations in benign lesions:Chronic gastritis and gastric ulcer

AIM： To investigate the occurrence of chromosome 3, 7, 8, 9, and 17 aneuploidies, TPS3 gene deletion and p53 protein expression in chronic gastritis, atrophic gastritis and gastric ulcer, and their association with H pylori infection. METHODS： Gastric biopsies from normal mucosa （NM, n = 10）, chronic gastritis （CG, n = 38）, atrophic gastritis （CAG, n=13） and gastric ulcer （GU, n=21） were studied using fluorescence in situ hybridization （FISH） and immunohistochemical assay. A modified Giemsa staining technique and PCR were used to detect Hpylori. An association of the gastric pathologies and aneuploidies with Hpylori infection was assessed. RESULTS： Aneuploidies were increasingly found from CG （21%） to CAG （31%） and to GU （62%）, involving mainly monosomy and trisomy 7, trisomies 7 and 8, and trisomies 7, 8 and 17, respectively. A significant association was found between H pylori infection and aneuploidies in CAG （P=0.0143） and GU （P=0.0498）. No TP53 deletion was found in these gastric lesions, but p53-positive immunoreactivity was detected in 45% （5/11） and 12% （2/17） of CG and GU cases, respectively. However, there was no significant association between p53 expression and H pylori infection. CONCLUSION： The occurrence of aneuploidies in benign lesions evidences chromosomal instability in early stages of gastric carcinogenesis associated with Hpylori infection, which may confer proliferative advantage. The increase of p53 protein expression in CG and GU may be due to overproduction of the wild-type protein related to an inflammatory response in mucosa.

Maspin subcellular expression in wild-type and mutant TP53 gastric cancers

BACKGROUND Although the role of p53 in the evolution and prognosis of gastric cancer(GC)has been extensively examined,the exact mechanism of action is incompletely understood.In the last years,p53-target genes were supposed to be involved in the p53 pathway.One of them is the tumor-suppressor gene Maspin,which codifies the protein with the same name.Maspin activity depends on its subcellular localization.To our knowledge,the possible role of TP53 gene in Maspin subcellular localization,in GC cells,has not yet been studied in a large number of human samples.AIM To evaluate the possible role of wild-type and mutated p53 in Maspin subcellular localization.METHODS The present study included 266 consecutive patients with GC in which TP53 gene status,and mutations in exons 2 to 11,respectively,were analyzed and correlated with immunohistochemical expression of p53 and Maspin.RESULTS None of the 266 cases showed mutations in exon 9.The rate of TP53 mutations was 33.83%.The mutation rate was slightly higher in distally-located GCs,with a lower degree(≤5 buds/high power fields)of dyscohesivity(P<0.01).The wildtype cases had a longer survival,compared with mutant GCs,especially in patients without lymph node metastases,despite the high depth of tumor infiltration(P=0.01).The Dukes-MAC-like staging system was proved to have the most significant independent prognostic value(P<0.01).The statistical correlations proved that TP53 gene mutations in exon 7 might induce knockdown of Maspin,but wild-type p53 can partially restore nuclear Maspin expression and decrease the metastatic potential of gastric adenocarcinoma cells.CONCLUSION Downregulated Maspin might be induced by mutations in exon 7 of the TP53 gene but wild-type p53 can partially restore nuclear Maspin expression.These findings should be proved in experimental studies.

Construction of the Eukaryotic Expression Vector for Outer Membrane Protein Tp92 from Treponema pallidum and Its Preliminary Study on the Immune Responses in New Zealand Rabbits

To construct the recombinant plasmid of eukaryotic expression containing Tp92 gene from Treponema pallidum and study its immunogenicity in New Zealand white rabbits. Tp92 gene was amplified from the genomic DNA of T. pallidum by polymerase chain reaction (PCR) and subcloned into appropriate site of pcDNA3.1(+) vector. After identification by sequencing and restrictive enzyme digestion, the recombinant plasmid was transfected into HeLa cells using liposome, and the expressed protein was identified by immunocytochemistry and Western blotting. After verifying that the Tp92 antigen gene fragment could be expressed in HeLa cells, 100?μg of recombinant plasmids [pcDNA3.1(+)-Tp92], 100 μg of control plasmids [pcDNA3.1(+)] or 0.5 ml PBS buffer were administered in 3 groups of New Zealand white rabbits (6 rabbits/group), and the booster immunizations were employed at 2-week interval for 3 times. ELISA assay was used for the quantitative detection of the specific antibody in the sera of rabbits, and the proliferation response of spleen cells was detected by MTT assay. It was found that the target gene Tp92 segment about 2103 bp was obtained, and the DNA sequence of Tp92 gene constructed in pcDNA3.1 (+) vector was consistent with the published nucleotide sequence. The homologies of the nucleotide and putative amino acid sequences of Tp92 gene between T.pallidum subsp. pallidum Nichols and various pathogenic treponeme strains were 95.5%-100%. The analysis of immunocytochemistry and Western blotting showed that Tp92 gene segment constructed in pcDNA3.1(+) vector could express a fusion protein with a calculated molecular mass of 77 kDa in HeLa cells and the expressed protein could react with positive blood serum from syphilis patient. The specific antibody IgG titers were observed and the highest titer was 1∶1024 in rabbits after 3 times with pcDNA3.1(+)-Tp92. The proliferation response of spleen cells were significantly higher than that of rabbits injected with pcDNA3.1(+) ( P <0.05). The successful expression of the eukaryotic expression plasmid of Tp92 gene from T. pallidum was obtained in eukaryotic system and strong responses of humoral and cellular immunity was evoked by DNA vaccine of pcDNA3.1(+)-Tp92 in rabbits thus establishing a solid basis for the future studies in the biological activities and for the development of the syphilis DNA vaccine.

TP53 Mutations and Chemotherapy Response to Neoadjuvant Metotrexate, Cisplatin and Adryamicin Chemotherapy in Resected Osteosarcoma

Osteosarcoma is a rare and highly malignant tumor that usually affects adolescents and young adults. Despite current management protocols, up to half of patients succumb to the disease. Moreover, there is no well-characterized molecular marker for diagnosis and prognosis. TP53 alterations have been associated with a poor prognosis in many cancers. The aim of this retrospective work was to find out whether TP53 functional status predicts response to neoadjuvant chemotherapy and thus may help treatment decision for osteosarcoma patients. Seventeen biopsies of osteosarcoma patients receiving primary metotrexate, cisplatin and adryamicin chemotherapy followed by surgery were analyzed. TP53 exons 5-9 mutations were screened. Among 17 biopsies, 4 (23.5%) displayed TP53 mutations: 3 deletions and one single-nucleotide substitution. The presence of TP53 gene mutation does not correlate with resistance to chemotherapy according to histological Rosen grade and nevertheless is associated with patient’s age in a significant manner (p 0.05). The presence of non-mutated TP53 is not entirely specific for a good prognosis. We found no evidence that TP53 mutations predict chemoresistance in osteosarcoma patients more over the overall survival curve, followed for more than 12 years, showing no difference between patients with tumors harboring wild type or mutated TP53 gene (p 0.5). Further analysis to identify other genes that can influence chemotherapy response and clinical outcome in osteosarcoma is needed to improve patient treatment.

Currently Clinical Views on Genetics of Wilson＇s Disease

Objective： Tile objective of this study was to review the research on clinical genetics of Wilson＇s disease （WD）. Data Sources： We searched docunlents from PubMed and Wanfang databases both in English and Chinese up to 2014 using the keywords WD in combination with genetic,ATP7B gene, gene mntation, genotype, phenotype. Study Selection： Publications about the ATP7B gene and protein timction associated with clinical features were selected. Results： Wilson＇s disease, also named hepatolenticular degeneration, is an autosomal recessive genetic disorder characterized by abnormal copper metabolism caused by mutations to the copper-transporting gene A TP7B. Decreased biliary copper excretion and redtlced incorporation of copper into apoeeruloplasmin caused by defunctionalization of ATP7B protein lead to acct, mulation of copper in many tissues and organs, including liver; brain, and cornea, finally restllting in liver disease and extrapyramidal symptoms. It is the most common genetic neurological disorder in the onset of adolescents, second to muscular dystrophy in China. Early diagnosis and medical therapy are of great significance tbr improving the prognosis of WD patients. However, diagnosis of this disease is usually diffict.lt because of its complicated phenotypes. In the last l0 years, an increasing number of clinical studies have used molecular genetics techniques. Improved diagnosis and prediction ol＇the progression of this disease at the molecular level will aid in the development of more individualized and effective intervcntions, which is a key to transition from molecular genetic research to the clinical study. Conclusions： Clinical genetics studies are necessary to understand the mechanism underlying WD at the molecular level from the genotype to the phenotype. Clinical genetics research benefits newly emerging medical treatments including stem cell transplantation and gone therapy for WD patients.