Mitochondrial DNA(mtDNA),a double-stranded circular molecular,encodes several genes essential for mitochondrial and cellular functions despite its small amount of genetic information.Damage of mtDNA can lead to changes of cellular structures and functions and this fact is drawing more and more attention.This review summarized the recent research about mtDNA damage and its influence on cells.

AIM:To investigate the effect of RNA interference (RNAi)-mediated insulin-like growth factor 1 receptor ( IGF1R) gene silencing on the growth , migration, and invasion of hepatocellular carcinoma cells .METHODS:The most effective siRNA targeting IGF1R gene was designed and screened .After lentiviral expression vector pLVX-shR-NA2-IGF1R carrying the most effective siRNA sequence was constructed , it was transfected into 293T cells and packed into pLVX-shRNA2-IGF1R lentivirus.Huh7 and Hep3B cells were infected with the pLVX-shRNA2-IGF1R lentivirus to screen the positive clone Huh7 cells and Hep3B cells with the lentivirus .These Huh7 cells and Hep3B cells were cultured to ana-lyze the mRNA level of IGF1R, cell proliferation, cell cycle, cell apoptosis, cell migration/invasion, and the protein levels of IGF1R, Ki-67, p-AKT, p-ERK1, Gli1,β-catenin, cyclin D1, p21 and BCL-XL.RESULTS:The mRNA expression of IGF1R in Huh7 cells and Hep3B cells with pLVX-shRNA2-IGF1R lentivirus was significantly reduced .The proliferation of these cells was remarkably inhibited , and the number in G 1 phase was increased significantly .The percentages of apop-totic cells were increased markedly , and the number of cell migration/invasion was decreased markedly .The protein levels of IGF1R, Ki-67, p-AKT, p-ERK1, Gli1,β-catenin, cyclin D1, p21 and BCL-XL were decreased significantly compared with the blank control group and negative control group .CONCLUSION:The RNAi-mediated IGF1R gene silencing sig-nificantly suppresses the growth and the malignant biological characteristics of Huh 7 cells and Hep3B cells, which may be involved in the reduced protein levels of the above genes induced by down -regulation of IGF1R expression.

Cold perfusion of liver can significantly alleviate the ischemia-reperfusion injury caused by hepatic blood flow occlusion. We have modified the technique of cold perfusion of liver and applied it to total pancreatectomy for patients with pancreatic head carcinoma complicated with metastasis to the body and tail of pancreas and with portal invasion. After skeletonization of the hepatoduodenal ligament, the amputation of the portal vein and blockage of the superior mesentoric vein were performed before portal perfusion. Meanwhile, pancreatic head resection, duodenectomy, subtotal gastrectomy and partial resection of the superior mesenteric vein and portal vein were carried out. Superior mesenteric vein and portal vein bypass grafting was achieved with artificial vessels. The digestive tract was reconstructed after it was freed of the spleen and resection of the body and tail of pancreas to the left side of superior mesenteric vein, greater omentum and intestine from the end of the colon to splenic flexure of colon. The patient was followed up for 3 months, and the general condition was good, although diarrhea frequently occurred. No tumor metastasis occurred.

Ex-vivo liver resection is developed based on liver transplantation and technique of cold preservation of organs.It overcomes the shortcomings of time limit of warm ischemia and high technique demand of hepatectomy of tumors located at critical sites.A 58-year-old woman with hepatocellular carcinoma located close to the middle hepatic vein combined with invasion of right hepatic vein was admitted to Southwest Hospital.Because of the critical tumor site,conventional liver resection Wag assessed as impossible.Ex-vivo liver resection was performed,and a vessel patch from an organ wag harvested to repair the defect of the right hepatic vein,and then the liver remnant was subsequently autotransplanted.After operation,the patient recovered smoothly without venous outflow complication.Bile leakage wag observed on postoperative day 23,and the maximnm volume of intraperitoneal drainage wag 200 ml per 24 hours.Endoscopic nasobiliary drainage Was performed and the volume of intraperitoneal drainage gradually decreased to none.Liver function of the patient was back to normal and with no tumor recurrence at the end of 6 months of follow up.Ex-vivoliver resection is beneficial to patients with centrally located hepatocellular carcinoma with the involvement of hepatic vein and inferior vena cava.

Periampullary carcinoma is a rare malignant tumor of digestive system,and its accurate diagnosis is still difficult.From January 2007 to July 2012,12 patients with periampullary carcinoma had been admitted to the Southwest Hospital of Third Military Medical University,and the imaging data were retrospectively analyzed.The results of ultrasonography revealed that all tumors were hypoechoic.The tumor displayed hyperenhancement in 3 patients,isoenhancement in 1 patient,hypoenhancement in 8 patients during the arterial phase on contrastenhanced ultrasonography (CEUS),while the tumor displayed hypoenhancement in all patients during the venous phase.Magnetic resonance imaging (MRI) plain scanning showed duodenal papilla enlargement in 1 patient,ampullary tissue mass signal in 2 patients,tissue mass signal at distal common bile duct in 2 patients,the rest 7 patients did not show tissue mass signal.Lower biliary obstruction was the common manifestation of the 12 patients on magnetic resonance cholangiopancreatography (MRCP),intrahepatic and extrahepatic bile vine-like expansion in 4 patients,double duct sign in 2 patients,the bottom of common bile duct with filling defect in 2 patients and it revealed beak-like narrow in 1 patient.CEUS,MRI and MRCP could both play an important role as conventional methods in diagnosing periampullary carcinoma.

The aim of the present studies was to investigate the cardioprotection of late IP at 72 h and determine the involvement of iNOS, nNOS and COX-2 in this protection. Conscious rabbits were preconditioned with three cycles of 5-minute coronary occlusion/5-minute reperfusion. The myocardial infarct area in the rabbits preconditioned 72 h earlier was significantly smaller than that in control rabbits. The activity of lactate dehydrogenase (LDH) and the level of 6-Keto-PGF1alpha in the rabbits preconditioned 72 h earlier were lower than those in control rabbits. The left ventricular systolic pressure (LVSP) and maximal velocity of contraction and relaxation (+/- dP/dtmax) were improved in rabbits preconditioned 72 h earlier. The nNOS-selective inhibitors N-propyl-L-arginine and selective cyclooxygenase-2 (COX-2) inhibitor celecoxib completely blocked the protection of late IP at 72 h, whereas the iNOS selective inhibitor S-methybisothiourea had no effect. In conclusion, the cardioprotection observed in the final stage of late IP (72 hours) is mediated by nNOS or COX-2, but not by iNOS.
Animals ; Cyclooxygenase 2 ; metabolism ; physiology ; Ischemic Preconditioning, Myocardial ; Male ; Myocardial Infarction ; enzymology ; pathology ; prevention & control ; Myocardial Reperfusion Injury ; enzymology ; metabolism ; prevention & control ; Nitric Oxide Synthase Type I ; metabolism ; physiology ; Rabbits ; Random Allocation ; Time Factors

Objective To evaluate the clinical application of Vater ampulla-duodenal conjunction resection in the treatment of periampullary carcinoma. Methods From January 2005 to July 2006, 15 patients underwent this modus operandi, including carcinoma of duodenal papilla (6 cases), Vater ampulla (5 cases) and lower part of common bile duct (4 cases). The descending part of duodenum, Vater ampulla, head of pancreas and common bile duct were excised en bloc followed by reconstruction of GI conduit. Result One patient died of stress ulcer 2 months postoperatively, the 14 patients recovered uneventfully without any major complications, and 3-16 months follow-up found no tumor recurrence. Conclusion Vater ampulla-duodenal conjunction resection as a new surgical procedure provides enough tumor margin clearance while causing less trauma than standard pancreatoduodenectomy in selected cases of periampullary carcinoma.

Lipases are important biocatalysts that are widely used in food processing and bio-diesel production. However, organic solvents could inactivate some lipases during applications. Therefore, the efficient cloning and expression of the organic solvent-tolerant lipase is important to its application. In this work, we first found out an organic solvent-tolerant lipase from Staphylococcus saprophyticus M36 and amplified the 741 bp Lipase gene lip3 (GenBank Accession No. FJ979867), by PCR, which encoded a 31.6 kD polypeptide of 247 amino acid residues. But the lipase shared 83% identity with tentative lip3 gene of Staphylococcus saprophyticus (GenBank Accession No. AP008934). We connected the gene with expression vector pET-DsbA, transformed it into Escherichia coli BL21 (DE3), and obtained the recombinant pET-DsbA-lip3. With the induction by 0.4 mmol/L of isopropyl beta-D-thiogalactopyranoside at pH 8.0, OD600 1.0, 25 degrees C for 12 h, the lipase activity reached up to 25.8 U/mL. The lipase expressed was stable in the presence of methanol, n-hexane, and isooctane, n-heptane.
Amino Acid Sequence ; Base Sequence ; Cloning, Molecular ; Enzyme Stability ; Escherichia coli ; genetics ; metabolism ; Lipase ; biosynthesis ; genetics ; Molecular Sequence Data ; Organic Chemicals ; chemistry ; Recombinant Proteins ; biosynthesis ; genetics ; Solvents ; chemistry ; Staphylococcus saprophyticus ; enzymology

With the genomic DNA of strain EJS-3 as the template, we amplified the gene of fibrinolytic enzyme from Paenibacillus polymyxa (PPFE-I) by PCR. We purified the PCR product and ligated it into pMD19-T. After DNA sequencing, we cloned the PPFE-I gene into expression vector pET-DsbA and transformed it into Escherichia coli BL21(DE3). Upon induction of IPTG, we found that the activity of recombinant fibrinolytic enzyme fused with DsbA expressed in Escherichia coli was 228 IU/mL. SDS-PAGE analysis showed that the recombinant enzyme was soluble and accounted for about 18.4% of total cell protein. Western blotting demonstrated that the recombinant protein was DsbA-PPFE-I. We purified the recombinant enzyme by Ni affinity chromatography, thrombin digestion and sephadex G-100 gel-filtration, and identified the molecular weight of purified product to be 66.3 kDa with MALDI-TOF mass spectrometry. The purified enzyme exhibited distinct fibrinolytic activity on fibrin plate.
Antifibrinolytic Agents ; pharmacology ; Cloning, Molecular ; Escherichia coli ; genetics ; metabolism ; Fibrinolytic Agents ; metabolism ; Genetic Vectors ; genetics ; Paenibacillus ; chemistry ; enzymology ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; pharmacology

To carry out phylogenetic analysis for drug-resistance genes from clinical isolates of Helicobacter pylori (Hp) among patients with gastric diseases from Tibet, China.

METHODSHp strains were isolated and cultured from saliva and gastric mucosal tissues derived from patients with gastric diseases. Nine strains (including 5 isolated from oral tissues, 1 isolated from gastric tissues, and 3 representative strains of SS international standard strains used for animal models) were tested for common antibiotic resistance. Together with an ACTT 11637 international standard strain, these were subjected to re-sequencing to obtain drug-resistance genes. Such genes from various sources were compared with the resistance genes of Hp strains recorded by the NCBI website. Combined with results of drug-resistance experiments, correlation between molecular evolution and drug-resistance was analyzed.

RESULTSTesting of gastric mucosal tissues and salivary samples from 217 patients has found 89 Hp strains, which yielded a total infection rate of 41.01%. The resistance rates of 9 representative Hp strains for clarithromycin, amoxicillin, metronidazole, levofloxacin and tetracycline were 77.8%, 77.8%, 44.4%, 77.8%, and 77.8%, respectively. Compared with the reference strain, the similarity between clarithromycin-resistance genes was 99%, and that between amoxicillin- and metronidazole-resistance genes was 96%-97%. A2143G mutation was also found in clarithromycin-resistant genes of three Hp strains.

CONCLUSIONThe sensitivity of Hp to metronidazole is much higher in patients from Tibet region, and the sensitivity of Hp to clarithromycin, amoxicillin, levofloxacin and tetracycline is poor. Resistance mutations are consistent with drug resistance.