OBJECTIVETo evaluate the effects of processing methods of Hirudo.

METHODBoth water and alcohol extracts of Hirudo were studied according to Chinese Pharmacopeia (Edition 2005). The content of hypoxanthine in Hirudo was measured by high performance liquid chromatography (HPLC) and Hirudin was determined by thrombin.

RESULTThe contents of water, water soluble extraction, ethanol soluble extraction and hirudin in crude Hirudo are higher than those in processed Hirudo. But the contents of hypoxanthine in processed Hirudo is higher than in crude Hirudo.

CONCLUSIONHirudo fried by French chalk can decrease the active components with high intensive drug property, accordingly the toxicity of Hirudo was decreased. As a result, the effects of Hirudo as invigorate the circulation of blood and stimulate the menstrual flow were abated.

Animals ; Blood Circulation ; drug effects ; Ethanol ; chemistry ; Hirudins ; analysis ; Hirudo medicinalis ; chemistry ; Hypoxanthine ; analysis ; Menstrual Cycle ; drug effects ; Solubility ; Water ; chemistry

Adult ; Carcinoma, Renal Cell ; secondary ; Female ; Humans ; Kidney Neoplasms ; pathology ; Magnetic Resonance Imaging ; Vaginal Neoplasms ; secondary

【Objective】 To establish an enzyme-linked immunosorbent assay (ELISA) method for the determination of residual human coagulation factor Ⅺ in human prothrombin complex and validate the method. 【Methods】 Human factor Ⅺ was reacted with the capture antibody coated on the microtiter plate. After appropriate washing steps, biotinylated primary antibody was bound to the captured protein. Excess primary antibody was washed away and bound antibody was reacted with horseradish peroxidase conjugated streptavidin. TMB substrate was used for color development at 450 nm. The dilution reliability, accuracy, specificity, repeatability, intermediate precision, linearity, range and durability were verified. 【Results】 The verification results showed that the accuracy and specificity of this method met the experimental requirements, with an average recovery rate of 109.2% and RSD of 6.93%. The repeatability RSD was 6.78%, and the intermediate precision RSD was 6.75%, indicating good precision. The linear regression correlation coefficient of standard curve was 0.999 9, showing good accuracy and precision within the linear range. The durability was verified by the incubation time and the validity period of reagent kit opening. The results showed that the RSD of the incubation time change was 6.62%, indicating that the incubation time of this detection method was controlled between 28 to 32 minutes, and there was no significant impact on the results. The RSD of the detection results before and after the reagent kit was opened and stored under conditions for 7 days was 3.84%, indicating that the preservation of the reagent kit according to the conditions for 7 days after opening has no effect on the FⅪ detection results. Both indicated that the method had good durability. The dilution reliability results showed that there was a "hook" effect in the detection of FⅪ residue in human prothrombin complex, which could be solved by diluting 100 to 200 times. 【Conclusion】 This method can be used for the determination of FⅪ residues of human prothrombin complex in laboratory.

AlM: To study clinical reference value of retinal microvascular changes in patients with cerebral microbleeds ( CMBs) and discuss its clinical significance.
METHODS:From January 2012 to December 2013, 125 hospitalized patients were collected, including 81 cases were male, 44 cases were female, mean age 76. 3 ± 11. 2 years old. For all patients, functions of liver and kidney, blood - lipoids, blood sugar and blood biochemical examination were tested, and fundus photography and cerebral MR was done. According to the fundus camera eyes, retinal arteriolar equivalent ( RAE) , retinal venular equivalent ( RVE) , retinal vein diameter ratio ( AVR) and arteriovenous crossing sign ( AVN ) were identified, CMBs were classified with cerebral MRl. All the data were processed by SPSS statistical software.
RESULTS: The central retinal arteriolar equivalent (CRAE), central retinal venular equivalent (CRVE) and AVR values in the eyes were found no statistical difference (P<0. 05). Of CMBs classification, the grade 0 in 75 cases, 1 in 27 cases, 2 in 9 cases and 3 in 14 cases were included. The RVE, AVR and AVN and the different grades of CMBs had statistically significant correlation ( P<0. 01). The higher CMBs classification, the more obvious retinal microvascular changes were found. ln respectively to eliminate risk factors such as age, sex, blood glucose and blood pressure, AVR and AVN were still influencing factors for CMBs classification.
COCLUSlON: The results show that retinal microvascular changes, especially small retinal vein arteriovenous cross width, and arteriovenous crossing phenomenon, in which CMBs will happen more likely. After sex, age, hypertension and hyperglycemia in patients with traditional cardiovascular risk factors being ruled out, the retinal microvascular changes are still relatively factors of CMB's occurrence.

OBJECTIVETo investigate luciferase gene expression and pathological changes of submandibular glands (SMG) of rats after adenovirus-mediated gene transfer.

METHODSAdenovirus-mediated luciferase gene (AdCMVLuc, 10(8) pfu in 50 microl) was injected in to SMG of forty wistar rats. The SMGs were harvested for gene expression measurement and pathological study after 3 days, 1,2,4 and 8 weeks.

RESULTSPeak expression was observed in three days following administration of the vector however, gene expression in submandibular glands decreased rapidly. the pathological changes induced by retrograde injection of AdCMVLuc included: after 3 days to one week, compression of acini, dilation of terminal ducts; after two weeks, slight atrophy of a part of acini, increase of iteracinar distance and focal lymphocyte infiltration in lobules and interlobular ducts; after 4 weeks, recovery evidence was found in acini; after 8 weeks, normal acini and ducts were found. The ultrastructural changes included: 3 days, much more rough endoplasmic reticulum was found both in acini and duct epithelial cell; a lot of mucus drops were found in acini; after 1 week, microvillus decreased in duct epithelial cells, mitochondria increased significantly in acini; intercellular space was enlarged.

CONCLUSIONSAdenovirus-mediated gene transfer can produce biological proteins and induce marked inflammatory changes in rat SMG. The ultrastructural changes suggest high protein synthesis activity in the acinar cells after gene transfer.

Adenoviridae ; enzymology ; genetics ; Animals ; Gene Expression ; Gene Transfer Techniques ; Luciferases ; genetics ; Male ; Random Allocation ; Rats ; Rats, Wistar ; Submandibular Gland ; pathology ; virology

Objective To study the proliferation of in vitro cultured mouse chondrocytes exposed to different doses of fluoride.Methods The third generation of primary cultured chondrocytes were exposed to the concentrations of 0,5,10,20,40 mg/L fluoride for 10 days to observe the morphological changes under light microscope and electron microscope to counter the numbers of ehondrocytes and proliferating rote with the growth curve and MTT.Results After exposed to fluoride for 10 days,the proliferation was present in the chondrocytes of the 5,10,20 mg/L groups,and shrinked chromatine and apoptosed ehondrocytes were seen in 40 mg/L group.The absorbance was not significantly different between all groups(F=2.313,P＞0.05);after exposed to fluoride for 48 and 72 hours,there was a significant difference of proliferating ability among 0 mg/L(the contr01)group[(23.5±4.6)%,(29.9±1.7)%],5 mg/L group[(34.6±4.7)%,(45.3±5.9)%],10 mg/L group[(39.9±4.8)%.(56.8±5.5)%],20 mg/L group[(31.8±4.1)%,(38.3±6.5)%]and 40 mg/L group[(28.3±4.3)%,(33.4±4.8)%](F=11.401,25.671,P＜0.05).There wss a significant difference compared with the control group(P＜0.05)with that of 5 and 10 mg/L groups higher than that of 40 mg/L groups(P＜0.05).Conclusions Lower doses of fluoride improve the proliferation of in vitro mouse chondrocyte in a short exposing time,higher doses result in the opposite.

OBJECTIVETo investigate the efficacy of anti-idiotype antibody 3F6 and its single-chain variable fragment (3F6 ScFv) to induce humoral and cellular immune responses against small-cell-lung cancer (SCLC).

METHODS3F6 and 3F6 ScFv (Ab2) were used to immunize BALB/c mice. The reaction of antibodies (Ab3) with specific antigen on NCI-H128 cells was tested by ELISA and Western blot, and the antibody binding inhibition assays were performed by competitive Western blot. Cellular immunity against SCLC induced by Ab2 was detected by a delayed-type hypersensitivity response and mouse lymphocyte proliferation assay.

RESULTSThe sera immunized with Ab2 showed significant reaction (P < 0.001, as compared to control sera) with SCLC-specific antigen on NCI-H128 cells and specifically competed the binding of 2F7 (Ab1) to the specific antigen. DTH responses challenged with NCI-H128 cells were significantly (P < 0.001) stronger in mice immunized with Ab2 as compared to mice immunized with normal mouse IgG. T cell proliferation was significantly higher in Ab2-immunized mice (P < 0.05) than in control mice.

CONCLUSIONThe two kinds of anti-idiotypic antibodies successfully mimic the SCLC-specific antigen on NCI-H128 cell and induce strong humoral and cellular immune responses to SCLC-specific antigen in syngeneic mice. They may become novel vaccines against human small-cell-lung cancer and worthy of further investigation.

Animals ; Antibodies, Anti-Idiotypic ; immunology ; Antibodies, Neoplasm ; immunology ; Antigens, Neoplasm ; immunology ; Cancer Vaccines ; immunology ; Carcinoma, Small Cell ; immunology ; Cytotoxicity, Immunologic ; drug effects ; Immunoglobulin Fab Fragments ; immunology ; Immunoglobulin Fragments ; immunology ; Immunoglobulin Variable Region ; immunology ; Lung Neoplasms ; immunology ; Mice ; Mice, Inbred BALB C

Adolescent ; Adult ; Aged ; Antithyroid Agents ; administration & dosage ; adverse effects ; therapeutic use ; Biomarkers ; blood ; Female ; Hepatitis B ; complications ; pathology ; Hepatitis, Viral, Human ; complications ; pathology ; Humans ; Hyperthyroidism ; complications ; drug therapy ; Liver Function Tests ; Male ; Methimazole ; administration & dosage ; adverse effects ; therapeutic use ; Middle Aged ; Propylthiouracil ; administration & dosage ; adverse effects ; therapeutic use ; Retrospective Studies ; Severity of Illness Index ; Thyroid Function Tests ; Young Adult

OBJECTIVETo discuss the treatment of the windblow hand deformity.

METHODSWe treat 18 patients with operations step by step. First phase: to improve the thumb function; Second phase: shift the flexor digitorum superficialis of the middle and little finger to improve the ulnar drift of the digits; Third phase : to solve the flection deformity of the palm and digits.

RESULTSWe follow the patients for 6 - 72 months, only lost 3.11 of 15 patients have the thumb function recovered. 9 patients had their ulnar drift of the digits corrected, 2 patients partly corrected, 4 have relapse. 5 among the 15 patients were offered second operation, to have the ulnar arthrosis bursa contracted, using abduction splint, after rehabilitation 4 patients have the deformity corrected, 1 has partly corrected.

CONCLUSIONSOperation step by step is better for the complex windblow hand deformity, solve one major problem each time,and systemic rehabilitation therapy is needed for satisfied curative effect.

Child ; Child, Preschool ; Female ; Hand Deformities, Congenital ; surgery ; Humans ; Infant ; Male ; Reconstructive Surgical Procedures ; methods

BACKGROUNDTo construct recombinant retroviral vector expressing HBcAg in eukaryotic cells.

METHODSThe HBV core gene fragment was amplified by using PCR from pADR which contains complement nucleotide sequence of HBV subtype adr and cloned into retroviral expression plasmid pLXSN, then transfected into packing cell (PT67) with lipofec AMINE. After 2-3 weeks selection with G418, large colonies were isolated and transferred to individual plates. Virus-containing medium was collected and used to infect NIH3T3, EL4 and mouse bone marrow derived dendritic cells(DC). DNA was extracted from infected cells and tested by PCR. Indirect immunofluorescence and FACS were used to detect the expression of HBcAg. Cell mediated immunity of immunized C57BL/6 mice with transduced DC was analyzed.

RESULTSThe insertion of HBV core gene fragment in the recombinant retroviral plasmid was confirmed by PCR as well as enzyme digestion with EcoR1 and BamH2. The viral titer reached 3 x 10(5) CFU/ml. The result of PCR showed that the HBV core gene had been integrated into the genome of infected NIH3T3 cells. Indirect immunofluorescence and FACS analysis showed that HBcAg had been expressed in these cells. HBcAg specific CTL responses could be generated in mice immunized with retrovirus transduced DC.

CONCLUSIONSHBV core gene had been integrated into eukaryotic cells with retroviral vector and target gene had been expressed efficiently. These results may have some impact on gene therapy of chronic hepatitis B.

3T3 Cells ; Animals ; Cloning, Molecular ; Dendritic Cells ; metabolism ; Eukaryotic Cells ; metabolism ; Genetic Vectors ; Hepatitis B Core Antigens ; biosynthesis ; genetics ; Hepatitis B virus ; genetics ; Humans ; Mice ; Recombination, Genetic ; genetics ; Retroviridae ; genetics ; Transfection