Codon 249 mutation in exon 7 of p53 gene in plasma DNA:maybe a new early diagnostic marker of hepatocellular carcinoma in Qidong risk area,China

AIM: One of the characteristics of hepatocellular carcinoma (HCC) in Qidong area is the selective mutation resulting in a serine substitution at codon 249 of the p53 gene (1,20),and it has been identified as a 'hotspot' mutation in heptocellular carcinomas occurring in populations exposed to aflatoxin and with high prevalence of hepatitis B virus carriers (2, 3, 9, 10, 16, 24). We evaluated in this paper whether this 'hotspot' mutation could be detected in cellfree DNA circulating in plasma of patients with hepatocellular carcinoma and cirrhosis in Qidong, China, and tried to illustrate the significance of the detection of this molecular biomarker.METHODS: We collected blood samples from 25hepatocellular carcinoma patients, 20 cirrhotic patients and 30 healthy controls in Qidong area. DNA was extracted and purified from 200 μl of plasma from each sample. The 249ser p53 mutation was detected by restriction digestion analysis and direct sequencing of exon-7 PCR products.RESULTS: We found in exon 7 of p53 gene G→T transversion at the third base of codon 249 resulting 249Arg→249ser mutation in 10/25 (40%) hepatocellular carcinoma cases,4/20 (20%) cirrhotics, and 2/30 (7 %) healthy controls.The adjusted odds ratio for having the mutation was 22.1(95 % CI, 3.2～91.7) for HCC cases compared to controls.CONCLUSION: These data show that the 249ser p53mutation in plasma is strongly associated with hepatocellular carcinoma in Qidong patients. We found this mutation was also detected, although it was at a much lower frequency,in plasma DNA of Qidong cirrhotics and healthy controls;We consider that these findings, together with the usual method of HCC diagnosis, will give more information in early diagnosis of HCC, and 249ser p53 mutation should be developed to a new early diagnostic marker for HCC.

Establishment of transgenic mice carrying gene encoding human zinc finger protein 191

AIM:Human zinc finger protein 191 (ZNF191) was cloned and characterized as a Krueppel-like transcription factor, which might be relevant to many diseases such as liver cancer,neuropsychiatric and cardiovascular diseases. Although progress has been made recently, the biological function of ZNF191 remains largely unidentified. The aim of this study was to establish a ZNF191 transgenic mouse model,which would promote the functional study of ZNF191.METHODS:Transgene fragments were microinjected into fertilized eggs of mice.The manipulated embryos were transferred into the oviducts of pseudo-pregnant female mice.The offsprings were identified by PCR and Southern blot analysis.ZNF191 gene expression was analyzed by RT-PCR.Transgenic founder mice were used to establish transgenic mouse lineages.The first generation (F1) and the second generation (F2) mice were identified by PCR analysis.Ten-week transgenic mice were used for pathological examination.RESULTS: Four mice were identified as carrying copies of ZNF191 gene.The results of RT-PCR showed that ZNF191 gene was expressed in the liver,testis and brain in one of the transgenic mouse lineages.Genetic analysis of transgenic mice demonstrated that ZNF191 gene was integrated into the chromosome at a single site and could be transmitted stably. Pathological analysis showed that the expression of ZNF 191 did not cause obvious pathological changes in multiple tissues of transgenic mice.CONCLUSION:ZNF191 transgenic mouse model would facilitate the investigation of biological functions of ZNF191 in vivo.

Multiple MR Imaging Techniques in the Diagnosis and Assessment of Resectability in Pancreatic Carcinoma

Objective: To study the value of multiple MR imaging techniques in the diagnosis of pancreatic carcinoma and the assessment of resectbility of the lesion. Methods: MR imaging was performed in 18 pa-tients with surgically and/or pathologically proven pancreatic carcinoma. GRE T1WI, TSE T2WI, GRE T1WI with fat suppression, delayed enhancement GRE T1WI, MRCP and 3D DCE MRA were used in MR scanning. Tumor involvement of the celiac trunk and its main branches, superior mesenteric artery,the portal, splenic and superior mesenteric veins were prospectively graded on a 0-4 scale based on cir-cumferential contiguity of tumor to vessel. Results: On GRE T1WI and TSE T2WI all the lesions showed slightly hypointense and hyperintense, respectively; On GRE T1WI with fat suppression, all the tumors obviously appeared hypointense; On delayed enhancement GRE T1WI, the lesions displayed irregularly circular enhancement in 14 patients and well-distributed enhancement in 4 patients. MRCP showed exten-sive bile and main pancreatic duct dilatation with typical "double-duct" sign in 8 patients. On 3D DCE MRA, we thought it was unresectable with more than half circumferential involvement of tumor to vessel,so that the portal, splenic and superior mesenteric veins were involved with 56% (10/18), 39% (7/18)and 67% (12/18), respectively. The celiac trunk and its main branches and superior mesenteric arteries were involved with 22% (4/18) and 17% (3/18), respectively. The pancreatic lesions in 2 cases could be completely resected in the evaluation of MR imaging, which was fitted to the findings of operation by pan-creatoduodenectomy. The pancreatic lesions in other 2 cases were partly, resected because there was tumor extension to superior mesenteric vein and/or artery. The tumors in the remaining 14 patients were too large and involved peripancreatic vessels or there were stomach or liver metastases, so these patients were only treated by choledochojejunostomy and gastrojejunstomy. Conclusion: The "all-in-one" MR approach including fast scanning sequences, fat suppression, MRCP and 3D DCE MRA provides the surgeon with diagnosis and assessment of resectability of tlm lesion prior to surgery of pancreatic carcinoma.