Acupuncture Points ; Adult ; Drugs, Chinese Herbal ; administration & dosage ; Ear ; anatomy & histology ; Female ; Humans ; Melanosis ; drug therapy ; Middle Aged ; Young Adult

Traditional recombination technology of bacteria chromosome and its limitation were introduced. The definition of Red recombination technology is put forward: a method of homologous recombination between foreign linear DNA and the target gene in chromosomes mediated by ? phage Red system. The linear DNA referred here is general PCR product or oligonucleotide, which has a 36~50bp homologous sequence with the target gene in chromosome at both flanking. Red recombination technology leaves out the in vitro DNA restriction enzyme digestion and link process, which makes the knockout and alternation of target gene in bacteria chromosome relatively easier, and becomes an effective method to exploring genes and constructing new strains gradually. The gene inactivation and alternation method aiming at bacteria chromosome applied to Red recombination system was summarized by the structure element, action mechanism, and strategy of recombination, advantage and developing prospect. The Red system includes three genes: bet (aka?), exo and gam (aka ?). Exo is a 5′→3′ exonuclease, which degrades the 5′ ends of linear DNA molecules. Bet is a single-stranded DNA binding protein that binds to the single stranded 3′ ends generated by Exo and promotes annealing to complementary DNA. Gam binds to the host RecBCD complex and inhibits its exonuclease activity. Red recombination system may be constructed in such plasmids as pKD20 and pKD46 or in chromosome of bacteria. Most bacteria are not readily transformable with linear DNA because of the presence of intracellular exonucleases that degrade linear DNA. But when bacteria cells are transformed with pKD20 or pKD46 plasmid, or integrated with a detective ? prophage, Red recombination enzymes may be expressed in host cells, which make linear DNA with 36~50bp extensions that are homologous to both flanking of target genes transform E.coli readily and knock-out or alternate target gene. The Red recombination method is not only useful in chromosomal gene inactivation in E.coli, but also in other bacteria or virus, such as Salmonella, Shigella flexneri and virus HaSNPV. With the proceeding research, Red system will be applied for more and more purposes, and contribute a lot for gene improvement and gene function investigation in the coming Postgenome Era.

Objective To determine effects of T-2 toxin and selenium on expression of aggrecanase in human chondrocyte.Methods Human chondrocytes were treated with T-2 toxin(0,1,10,20 μg/L),and/or sodium selenite(final concentration of selenium 0,0.1 mg/L) for 5 days.Aggrecan expression was determined by Western blotting,aggrecanase-1 and aggrecanase-2 mRNA levels were measured by RT-PCR.ResultsSelenium and T-2 toxin had effects on both aggrecan protein levels and its aggrecanases(include aggrecanase-1 and aggrecanase-2 ) mRNA levels(F =0.294,27.71 for aggrecan,F =107.45,362.25 for aggrecanase-l,F =34.68,22.26 for aggrecanase-2,respectively,all P ＜ 0.05),and there was interaction between selenium and T-2 toxin on aggrecan,aggrecanase-1 and aggrecanase-2 expression(F =79.99,230.76,388.33,all P ＜ 0.05).Furthermore,selenium presented significant antagonism to T-2 toxin on aggrecan,aggrecanase-1 and aggrecanase-2 expression.Aggrecan expression levels(0.278 ± 0.015,0.235 ± 0.029,0.195 ± 0.028,0.399 ± 0.028,0.299 ± 0.020,0.263 ±0.019) induced by both 1,10,20 μg/L T-2 toxin and 0,0.1 mg/L selenium were significantly decreased than the levels(0.472 ± 0.0358,0.197 ± 0.018,all P ＜ 0.05) in control group(0 mg/L toxin).Selenium partially blocked the effects induced by 1,10,and 20 μg/L T-2 toxin(all P＜ 0.05).One,10,20 μg/L T-2 toxin and 0,0.1 mg/L selenium increased both aggrecanase-1 mRNA levels(0.535 ± 0.033,1.071 ± 0.043,1.454 ± 0.058,1.057 ±0.048,0.555 ± 0.036,0.902 ± 0.045) and aggrecanase-2 mRNA levels(0.596 ± 0.038,0.656 ± 0.033,0.949 ±0.049,0.600 ± 0.040,0.453 ± 0.031,0.164 ± 0.011),when compared with control(0.481 ± 0.023,0.346 ±0.020 for aggrecanase-1 and 0.387 ± 0.020,1.021 ± 0.046 for aggrecanase-2,respectively,all P ＜ 0.05).Selenium partially blocked 10,20 μg/L T-2 toxins induced upregulation of aggrecanase-1 (all P ＜ 0.05) and aggrecanase-2 (all P ＜ 0.05 ).Conclusions These data suggest a possible molecular mechanism that T-2 toxin could induce cartilage matrix degradation through the upregulation of aggrecanases expression and enzyme activities.Trace element selenium has some protective effect on cartilage proteoglycan degradation induced by T-2 toxins.

Objective To explore the plague epidemical trend of nearly a 10 years data in Qinghai province to provide basis for making the prevention and control measures. Method The regional distribution and time distribution of animal and human plague, monitoring and plague foci of survey data in Qinghai from 2001 to 2010 were analyzed with Excel software 2003. Results In Qinghai province, a total of 167 strains of Yersinia pestis were isolated from infected animals and insects in 10 years. Yersinia pestis was mainly distributed in Wulan,Delinha, Geermu, and Tianjun, along the Qinghai-Xizang railway. Human plague was occurred every year from 2001 to 2010 except 2002, 2007, 2008, and 2010. In the 10 years, there were 37 plague cases and 16 of these cases died, the mortality was 43.24％. The plague cases were mainly distributed in Nangqian, Qumalai, Chenduo,Zhiduo, Xinghai, Tongde, Tianjun, Wulan and Qilian. And these cases were found mostly in the period from May to October, especially in the period from August to October. Major clinical type of the plague cases was lung-type (62.16％,23/37). Conclusions The plague epidemic situation in Qinghai province is still severe, animal plague occurred year after year, and human plague outbreaks occasionally. Monitoring and early warning in the key areas should be strengthened, and the comprehensive measures of plague prevention and control should be carried out to reduce the incidence and prevalence of plague.

Objective Occult hepatitis B infection of voluntary blood donors has been plagued in the serum screening.Determined the OBI through the highly sensitive detection methods Nest-PCR among the blood donors,and then learned occult HBV infection and analysed the genotypes of this area.Methods 10 080 serums of donors were determined respectively by the imported Abbott HBsAg kit and Beijing Wantai anti-HBc and anti-HBs reagents,obtained the gene and detected DNA sequences by the high sensitive Nest-PCR method.Results Among 10 080 cases of unpaid blood donors,108 cases were detected HBsAg positively by Abbott sensitivity kit ( positive rate of 1.07％ ),767 cases were anti-HBc single - positive ( positive rate of 7.67％ ).25 patients screened blood donors who tested negative for serum HBsAg and positive for HBV DNA in the 10080 cases.Occult HBV infection incidence rate was 0.25％.12 cases were HBV genotype C (48％),13 cases were genotype B (52％),and no other genotypes.Genotype B has no statistically significant difference to genotype C ( P ＞ 0.05 ).Sequence analysis showed that 5 patients in the HBsAg epitope "a" (aa124 - aa147 ) have mutation (20％).Conclusion The high proportion of occult hepatitis B infected among voluntary blood donors in our country.Also genotype and mutation was differences in different regions.

Objective To assess if the modulating effect of platelet-derived growth factor(PDGF)BB on p21~(WAF1) was mediated by upregulating transforming growth factor(TGF)-β_1 expression in vascular smooth muscle cells(VSMC).Methods TGF-β_1 mRNA and protein expressions were measured by reverse transeription-PCR and ELISA,the protein expressions of p21~(WAF1) and the downstream TGF-βsignalling including TGF-β type I receptor(ALK-5 in VSMC),Smurf2,pSmad2/3,Smad4,Smad7 were detected by Western blot.Results PDGF-BB significantly upregulated the expressions of TGF-β_1 at mRNA(0.79-fold)and protein(1.98-fold)levels in VSMC,significantly inhibited the expression of p21WAF1(-67±12)%,and enhanced the expressions of ALK-5,pSmad2/3,Smad4,Smurf2 protein by 1.21-fold.0.95-fold,0.69.fold and 2.55-fold respectively,inhibited Smad7 expression(-65±9)%,these alterations were partially restored by anti-TGF-β_1 neutralizing antibody.Conclusions These findings suggested that PDGF-BB inhibited p21~(WAF1) expression in VSMC partially through upregulating TGF-β_1 expression via PDGFBB and TGF-β signalling pathways.

OBJECTIVETo study effects of alternative transcripts of ING1 transfection on human cancer cell lines.

METHODSp47/ING1A and p33/ING1B expression vehicles were constructed and introduced into a human breast cancer cell line MCF-7 and a human lung cancer cell line PAa, both expressing wild-type p53 protein. Growth characteristics of the transfectants and potentially related genes were analyzed.

RESULTSThe levels of p47/ING1A and p33/ING1B protein elevated respectively in tumor cells of MCF-7 and PAa after transfected with p47/ING1A and p33/ING1B, and the latter was much higher than that of the former. Ectopic overexpression of p33/ING1B effectively blocked tumor cell growth and arrested cells in the G(0) approximately G(1) phase of the cell cycle (P < 0.01), while p47/ING1A gave no effect on cell growth or cell cycle. Tumor cells overexpressing p33/ING1B contained more p21(WAF1) protein than that of the control cells, with undisturbed p53 protein level.

CONCLUSIONSExpression of two different transcripts of ING1 may have different effects on tumor cell growth. p33/ING1B may cooperate with p53 in stimulating expression of p21(WAF1) gene, thus to arrest cell cycle and to inhibit tumor cell growth. p33/ING1B may be considered to be a candidate as a partner of p53 in gene therapy.

Adenocarcinoma ; genetics ; metabolism ; pathology ; Alternative Splicing ; Breast Neoplasms ; genetics ; metabolism ; pathology ; Cell Cycle ; Cell Cycle Proteins ; Cell Division ; Cell Line, Tumor ; Cyclin-Dependent Kinase Inhibitor p21 ; Cyclins ; biosynthesis ; DNA-Binding Proteins ; Genes, Tumor Suppressor ; Humans ; Inhibitor of Growth Protein 1 ; Intracellular Signaling Peptides and Proteins ; Lung Neoplasms ; genetics ; metabolism ; pathology ; Nuclear Proteins ; Protein Biosynthesis ; Proteins ; genetics ; Transfection ; Tumor Suppressor Protein p53 ; biosynthesis ; Tumor Suppressor Proteins

OBJECTIVETo summary the experience of the surgical comprehensive treatment of severe acute pancreatitis (SAP).

METHODSFrom July 1999 to December 2009, a total of 506 patients suffered SAP were admitted with a mean APACHE II score 12.8 ± 4.6. There were 270 male and 236 female, aged from 16 to 89 years, mean age 43 years. SAP patients were treated by the SAP treatment team which consisted of pancreatic specialized and multidisciplinary doctors. Two hundreds and thirty-four cases (46.2%) received non-operative treatment and 272 cases (53.8%) received surgical intervention.

RESULTSIn 506 cases, 445 patients were cured and 52 patients died (31 died in early stage, 21 died in later stage), 9 cases discharged automatically. The overall incidence of complication, overall mortality and overall curative rate were 29.4% (149/506), 10.3% (52/506) and 87.9% (445/506), respectively. The incidences of complication in non-operative group and in surgical intervention group were 27.8% (65/234) and 30.9% (84/272), respectively (P > 0.05). The mortality in non-operative group and in surgical intervention group were 9.4% (22/234) and 11.0% (30/272), respectively (P > 0.05). The curative rates in non-operative group and in surgical intervention group were 90.6% (212/234) and 85.7% (233/272), respectively (P > 0.05).

CONCLUSIONSPatients should be treated in ICU in the early phase of the disease when APACHE II score > 10. Pancreatic specialized and multidisciplinary team treatment, appropriate choice of timing, indication and procedure of surgical intervention and details of drainage are vital to the prognosis of SAP.

APACHE ; Acute Disease ; Adolescent ; Adult ; Aged ; Aged, 80 and over ; Female ; Humans ; Male ; Middle Aged ; Pancreatitis ; mortality ; surgery ; Prognosis ; Retrospective Studies ; Survival Rate ; Young Adult

OBJECTIVETo evaluate the reliability of the reverse facial artery-submental artery island myocutaneous flap for repairing the oral and maxillofacial defects.

METHODSEighteen oral and maxillofacial defects were repaired with reverse facial artery-submental artery island myocutaneous flaps following resection of malignant tumors. The ages of the patients ranged from 28 to 90 years; 11 were male and 7 were female. Primary sites of the lesions were the tongue squamous cell carcinoma (SCC) (7 cases), buccal mucosa SCC (4 cases), palate SCC (3 cases), oropharyngeal SCC (2 case), and facial skin basal cell carcinoma (2 cases). The sizes of skin paddle varied from a minimum of 4.0 cm x 12.0 cm to a maximum of 5.0 cm x 15.0 cm. The followed-up period was 9 to 18 months (11.8 months on average).

RESULTSThe postoperative outcome for the flaps was 17 cases surviving, one complete necrosis, and one temporary palsy of the marginal mandibular branch of the facial nerve. The form and function of recipient sites were well recovered. The donor site left a well-hidden scar. One case of cervical lymph node metastasis was observed.

CONCLUSIONSReverse facial artery-submental artery island myocutaneous flap is reliable for reconstructing medium-sized oral and maxillofacial defects.

Adult ; Aged ; Aged, 80 and over ; Facial Neoplasms ; surgery ; Female ; Follow-Up Studies ; Humans ; Male ; Middle Aged ; Mouth Neoplasms ; surgery ; Reconstructive Surgical Procedures ; methods ; Surgical Flaps ; Treatment Outcome

BACKGROUNDScavenger receptor that binds phosphatidylserine and oxidized lipoprotein/CXC chemokine ligand 16 (SR-PSOX/CXCL16) promotes foam cell formation through the tumor necrosis factor (TNF)-alpha mediated mechanism. Because chemokine CXCL16 could be expressed in atherosclerotic lesions and induce smooth muscle cell (SMC) proliferation, we presume that the monocyte SR-PSOX/CXCL16 detection in the patients' peripheral blood will be important for early diagnosis and prognosis of atherosclerosis (AS).

METHODSEnrolled in this study were 40 patients with acute coronary syndrome (ACS), including 20 patients with acute myocardial infarction (AMI) and 20 patients with unstable angina pectoris (UAP), and 20 normal controls. Monocytes in the peripheral blood were isolated, and the changes of expression of CXCL16/SR-PSOX mRNA were compared using reverse transcription-polymerase chain reaction (RT-PCR), with beta-actin as internal control. We compared the expression of CXCL16/SR-PSOX in the ACS subgroups, using Western-blot to analyze protein expression levels. Tissue sections were made from biopsy specimens taken from patients with infective endocarditis, liver cirrhosis, and lung cancer as well as normal controls. And the expression of CXCL16/SR-PSOX was analyzed with a confocal microscope.

RESULTSThe expression of CXCL16/SR-PSOX mRNA and protein in the monocytes of peripheral blood was significantly higher in ACS patients than in normal controls (P < 0.05); however, there was no significant difference in CXCL16/SR-PSOX expression between UAP group and AMI group (P > 0.05). Immunofluorescence showed that there were low expression of SR-PSOX in normal vascular endothelial cells and enhanced expression in every layer of the infected vessels, while spreading from endothelial cells to surrounding tissues as infection worsens. Confocal microscopy showed that the expression of SR-PSOX was enhanced in the infiltrated lymphocytes in liver cirrhosis, and that the expression level was proportionate to the degree of inflammation in the portal hepatis and folia.

CONCLUSIONSThe expression of CXCL16/SR-PSOX in the monocytes of peripheral blood was significantly higher in ACS patients than in the controls. CXCL16/SR-PSOX-mediated inflammation may contribute to the pathogenesis of ACS, and CXCL16 may play an important role in the pathogenesis and development of AS in humans.

Acute Coronary Syndrome ; immunology ; Blotting, Western ; Chemokine CXCL16 ; Chemokines, CXC ; blood ; genetics ; Coronary Angiography ; Fluorescent Antibody Technique ; Humans ; RNA, Messenger ; blood ; Receptors, Scavenger ; blood ; genetics