Professor JIANG Yu-ren,a famous pediatrist of TCM in present age,has been engaged in clinic,teaching and scientifi c research of TCM pediatrics for 60 years.His achievement is remarkable.Especially,he has rich clinical experience in acute febricity of paediatrics,such as critical encephalitis B and serious measles complicated by pneumonia.He puts forward the rule of treating measles with favourable prognosis by method of dispersing-promoting,cooling-relieving and nourishing ying,treating measles with reverse prognosis by method of clearing heat,cooling blood and reviving yang and treating measles complicated by pneumonia should use the method of promoting pathogenic factors,removing toxic substance and relieving exhaustion.

Directed evolution was conducted to improve the thermostability of lipase from Rhizopus chinensis CCTCC M201021. Mutations were introduced by two rounds of error-prone PCR and mutant lipase was selected by fast-blue RR top agar screening. Two positive variants were selected in the first-round and four in the second-round screening process. Ep2-4 was proved as the most thermostable lipase and its DNA sequencing revealed three amino acid substitutions: A129S, P168L and V329A. Compared with the parent, its half-life at 60 degrees C was 5.4- times longer and T50 was 7.8 degrees higher. Purified lipase of Ep2-4 was characterized and the result shows that its thermostability improved without compromising enzyme activity. According to the mimicked protein structure, mutation A129S formed a hydrogen bond with Gln133 and improved the thermostability by increasing the hydrophilicity and polarity of protein; mutation P168L by forming a hydrophobic bond with the nearby Leu164.
Cloning, Molecular ; Directed Molecular Evolution ; methods ; Enzyme Stability ; genetics ; Hot Temperature ; Industrial Microbiology ; Lipase ; chemistry ; genetics ; Mutation ; Pichia ; genetics ; metabolism ; Polymerase Chain Reaction ; methods ; Protein Engineering ; methods ; Rhizopus ; enzymology

Objective To make sure whether or not Bcrp1 is the marker of cervical cancer stem cells or not by studying the invasive ability and formation of tumors of Bcrp1 + phenotype HeLa cells.Methods The tumor cell migration and invasion assay were used by boyden chamber to identify the invasive ability of Bcrp1 + phenotype HeLa cells.The formation of tumors in vivo experiments were completed,in which the two groups of cells with different concentrations were inoculated in non obese diabetes-severe combined immunodeficiency disease ( NOD/SCID ) mice ( 1 × 104,1 × 105,1 × 106/ml ) and the differences of time,rate and volume in the formation of tumors between two groups were observed.Results ( 1 ) In the invasion assay,the amount of cells that invaded through the artificial basement membrane in Bcrp1 + group were 99 ± 14,which was significantly greater than those in Bcrp1- group ( 57 ± 13,P ＜ 0.05 ) ; the length of the Bcrp1 + group was ( 366 ± 52 ) μm,which was significantly greater than the Bcrp1 - group ( 301 ± 54) μm ( P ＜ 0.05 ).( 2 ) Following transplantation of 1 × 104 cells,only the Bcrp1 + cells formed tumors in NOD/SCID mice.When 1 × 105 or 1 × 106 cells were transplanted,the tumor incidence and the tumor mass were greater in the Bcrp1 + groups than those in the Bcrp1 - groups ( P ＜ 0.05 ).Conclusion Bcrp1 + HeLa cell have the greater capacity of invasive and the tumorigenicity,which may contain cancer stem cells.

Objective To compare the clinical effect of needle knife therapy and traction for treatment of cervical spondylosis .Methods 156 patients with nerve root type of cervical spondylosis patients definitely diag -nosed,randomly divided into two groups ,the treatment group of 78 cases,with small needle knife therapy;the control group of 78 cases,the cervical traction therapy ,combined with traditional Chinese medicine .The clinical efficacy was compared between the two groups .Results The treatment group 70 cases were cured(89.6%),the total efficiency of 96.9%;control group 44 cases were cured(56.7%),the total efficiency of 83.3%.The cure rate and total effective rate was significantly better than the control group (χ2 =22.02,11.57,all P<0.05).Conclusion The needle knife therapy to treat cervical vertebra disease is better than that of cervical traction ,has good clinical curative effect .

Patients with primary liver cancer are always associated with hypersplenism .The traditional treatment of hypersplenism is splenectomy .With the development of interventional imaging ,partial splenic emboli-zation has been used extensively in clinical practice .Hepatocellular carcinoma with hypersplenism patients by par-tial splenic embolization can obviously improve the blood , reduce the complications of portal hypertension .PSE combined with other treatment can prolong the survival of patients with hepatocellular carcinoma .In this paper ,the clinical application of PSE is reviewed .

Objective To set up the quality standard of Shenbai concentrated pills. Methods Radix Astragali, Radix Paeoniae Rubra and Rhizoma Imperatea in Shenbai concentrated pills were identified with the method of TLC. The concentration of salvianolic acid B was determined with the method of HPLC. Results For TLC, the chromatogram spots of Radix Astragali, Radix Paeoniae Rubra and Rhizoma Imperatea were well separated and without interference in their negative controls. Quantitative analysis of HPLC showed that the average recovery of salvianolic acid B was 99.74% (RSD=2.08%) with the RSD of precision, reproducibility and stability was 1.74%, 1.19% and 1.70%, respectively. Conclusions The methods set up by this study are easy, accurate and stable, and can be used as the quality control standard of Shenbai concentrated pills.

White spot syndrome virus(WSSV), Taura syndrome virus(TSV)and Infectious hypodermal and haematopoietic necrosis virus(IHHNV)are three shrimp viruses responsible for major pandemics affecting the shrimp farming industry. Shrimps samples were collected from 12 farms in Zhejiang province, China, in 2008 and analyzed by PCR to determine the prevalence of these viruses. From the 12 sampling locations, 8 farms were positive for WSSV, 8 for IHHNV and 6 for both WSSV and IHHNV. An average percentage of 57.4% of shrimp individuals were infected with WSSV, while 49.2% were infected with IHHNV. A high prevalence of co-infection with WSSV and IHHNV among samples was detected from the following samples: Bingjiang(93.3%), liuao(66.7%), Jianshan(46.7%)and Xianxiang(46.7%). No samples exhibited evidence of infection with TSV in collected samples. This study provides comprehensive information of the prevalence of three shrimp viruses in Zhejiang and may be helpful for disease prevention control in this region.

ObjectiveTo study tear film functions of diabetes mellitus with pterygium before and after surgery.MethodsNinety patients with single side pterygium were divided into 3 groups with 30 cases each:simple pterygium (treatment group A ),pterygium combination with type 1 diabetes mellitus (treatment group B) and pterygium combination with type 2 diabetes mellitus(treatment group C).Ninety patients without pterygium were divided into 3 groups with 30 cases each:normal (control group A),type 1 diabetes mellitus (control group B) and type 2 diabetes mellitus (control group C ).Tear film functions were compared in each group before surgery,while tear film functions in treatment group B and C were compared between before and after surgery.ResultsBefore surgery,there was no significant difference among each group in lacrimal secretion test.Every group's tear film break-up time(BUT) was abnormal except control group A.Cornea fluorescein staining (CFS) score in treatment group B compared with control group B,and treatment group C compared with control group C were significant differences [ (9.08 ± 0.48 ) scores vs.(3.43 ± 0.51 )scores and (10.31 ±0.94) scores vs. (7.91 ±0.51 ) scores,P＜0.05].There was significant difference in mucus fern test(MFT) between treatment group B and control group B (P＜0.05).Tear clearance rate (TCR) were all abnormal except control group A.Before surgery compared with after surgery between treatment group B and treatment group C,there was no significant difference in lacrimal secretion test (P＞0.05 ),but there were significant differences in BUT,CFS score and MFT,the TCR were normal.Conclusion Diabetes mellitus with pterygium will aggravate damage in tear film functions,and these damaged functions could be recovered through pterygium surgery with conjunctival limbal autograft.

Objective To make sure whether Bcrp1 is the marker of cervical cancer stem-like cells or not by studying the characterization of Bcrp1+ HeLa cells.Methods Immunofluorescence stained flow cytometry and electron microacope were used to sort and observe uhrastructures of Bctp1+ and Bcrp1- HeLa cells.Flow cytometry wag used to identify the cycle and the rate of apoptosis with annexin V in two group cells.The expression of proliferating cell nuclear antigen (PCNA)and caspase-3 were tested using western blot methed.Results (1)There were 7.1% Bcrp1+ cells and 92.9% Bcrol- cells in HeLa cells.Bcrp1+ HeLa cells were large in size of nuclear and nucleoli are clear.and there were rich of cytomicrosome and rough endoplasmic reticulum.After sorted and cultured for 24,48,72 hours,the adhesion in Bcrp1+ cells were 72.8%,81.1%,80.4%,respectively.While,they were 3.3%,18.7%,12.6%at each time for Bcrpl- cells(all P<0. 05 ). (2) There are more S phase cells in Bcrp1+ cells than that in Bcrp1- cells (54. 1% vs 21.1%, P <0. 05) ,while the percentage of G0/G1 and G2/M in Bcrp1 - cells were highter than those in Bcrp1 + cells (53.0% vs 44. 4% ,25.9% vs 1.5% ; all P <0. 05 ). The rate of apoptosis in Bcrp1+ cells was lower than that in Bcrp1 - cells (0. 2% vs 5.3%, P < 0. 05 ). ( 3 ) The expression of PCNA in Bcrp1 + cells was higher than that in Bcrp1- cells (3140 vs 2255, P< 0. 05 ), while the expression of caspase-3 of Bcrpl + cells was lower than that in Bcrp1 - cells ( 1970 vs 3551, P < 0. 05 ). Conclusion There are more vigor and ability of proliferation and lower rate of apeptosis in Bcrp1 + HeLa cells than those in Bcrp1 - cells ,which may be some characters of cervical cancer stem cells.

Objective To investigate the approach and clinical efficacy of second repair surgery according to the nose and mouth contour design of unilateral cleft lip.Methods Based on the characteristics of anatomy,mechanics and deformity of the cleft lip,we designed a surgical incision of nasolabial contour lines with reference to the horizontal and central vertical lines and the contour.After careful investigation of classic procedures and clinical practice,unilateral cleft lip was repaired by overall stage Ⅱ surgical procedures.Results Application of this surgical method with satisfactory clinical efficacy had been achieved.Conclusions The second stage repair personalized surgical methods of unilateral cleft lip designed according to the nose and mouth contour can achieve maximal correction of malformation and beautiful appearance.