Objective To explore the proliferation-promoting effect of bovine mammary gland epithelial cells (BMECs) co-cultured with umbilical cord mesenchymal stem cells (UC-MSCs) in serum-free culture mediuum.Methods Bovine UC-MSCs and BMECs were selected for co-culturing in direct or indirect contact.In the direct contact culture groups, UC-MSCs and BMECs were co-cultured at concentration ratios of 2∶1, 1∶1, 1∶2, 1∶3, 1∶4, 1∶5, and 1:10, respectively.In the indirect contact culture group, the supernatant of UC-MSCs was used as the conditioned medium to re-suspend BMECs.In the control groups, UC-MSCs and BMECs were cultured alone.The cell growth status in each group was observed at 0, 4, 8, 12, 24, 36, 48, 60, 72 h after culture, and cell proliferation was detected by cell counting kit-8 (CCK-8) assay.Results At 48 h, the optical density of the conditioned medium-BMECs group was significantly higher compared with the control groups (P<0.05).Meanwhile, the optical density in the direct contact group at a concentration ratio of 1∶2 reached the peak, which was extremely significantly higher compared with the control groups (P<0.01) and significantly higher compared with the other direct contact culture groups and the conditioned medium-BMECs group (P<0.05).Conclusions Co-culture of UC-MSCs and BMECs in serum-free culture medium is capable to promote the proliferation of BMECs, and the co-culture by cell-to-cell contact has a better effect.The optimal concentration ratio of UC-MSCs to BMECs is 1∶2, and the optimal culture time is 48 h.

BACKGROUND: In the process of limb allograft, apoptosis of target cell is one of the main mechanisms of dysfunction of allograft, which might lead to the failure of allotransplantation. It is assumed that immunosuppressant may relate with cell apoptosis. OBJECTIVE: To investigate effects of FK506 on Bcl-2 mRNA and Bax mRNA expressions and cell apoptosis in rat limb allograft. DESIGN, TIME AND SETTING: The randomized controlled animal trial was performed at the Animal Experimental Center of First Hospital of Harbin Medical University from June 2005 to November 2006. MATERIALS: Fifty-six clean-grade healthy male SD recipient rats and 56 Wistar donor rats were selected. FK506 was product of Fujisawa, Japan (No. 100143G). METHODS: Right hind limb was separated from the upper segment of thigh of SD rat (donor), and washed using heparin saline. The recipient rates underwent limb allotransplantation from allogenetic Wistar to establish injury model. The recipients were randomly divided into two groups (n=28): immunosuppressant group was injected with FK506 1 mg/kg per day, and the control group was not given any immunosuppressant. The right hind limb including skin, subcutaneous tissues, muscles and femoral arteriolar-venular tissue mass were harvested on postoperative days 1, 3, 5, and 7. MAIN OUTCOME MEASURES: Bcl-2 Mrna and Bax mRNA expression were detected using reverse transcription polymerase chain reaction (RT-PCR), and cell apoptosis was detected using in situ terminal deoxynucleotidyl transferase mediated deoxyuridine triphosphate nick end labeling technique. RESULTS: Fifty-six model rats were included in final analysis. On the 3, 5, and 7 days after FK506 injection, Bcl-2 mRNA expression in immunosuppressant group was significantly higher than the control group (t=7.18-21.20, P

To explore the experience in teaching of interns in orthopedic,we put emphasis on the knowledge teaching of medical ethics and medical dispute in addition to the teaching of expertise of orthopedic.

The interaction between the antibody and the corresponding target molecule determines the characteristics of immunoassay. In this study, a single chain variable fragment antibody (scFv4C7) derived from the hybridoma strain 4C7 were prepared via genetic engineering technique. The recognition properties of scFv4 C7 was determined and compared to those of the parent monoclonal antibody by indirect competitive enzyme-linked immunosorbent assay(ic-ELISA). Three dimensional structure of the scFv4C7 was presented by Swiss-Model, and sulfathiazole ( STZ) was docked to the scFv4C7 model to obtain the structure of the binding complex. The results from the ic-ELISA showed that the binding properties of scFv4C7 were comparable with the parent monoclonal antibody and STZ was almost completely buried in a deep binding pocket formed by the heavy chain and light chain of the antibody. The interaction between STZ and scFv4C7 was more closely related to the heavy chain and the complementarity-determining region ( CDR ) H3 loop played more important role than other CDR loops. The study preliminary provided the necessary structural information for the preparation of antibody with broader specificity and higher affinity.

Objective To evaluate the clinical application of May-Grunwald-Giemsa staining followed by fluorescence in situ hybridization (MGG-FISH) technique in the differentiation diagnosis of Ph-chromosome positive acute lymphoid leukemia (Ph + ALL) from chronic myeloid leukemia in lymphoid blast crisis(CML-LBC). Methods The bone marrow smears of 4 patients with Ph+ ALL, 4 patients with CML-LBC, 1 patient with CML in myelocytic blast crisis complicated with lymphoma and 1 patient with CML in mixed blast crisis were assayed with the MGG-FISH technique in which the spectrum green labeled BCR and spectrum orange labeled ABL dual color dual fusion probes were used. Based on the morphological classification, the percentages of BCR-ABL positive cells were subsequently determined respectively in the erythroid, myeloid and lymphoid hneages for the 10 specimens. Results According to the MGG-FISH analysis, the erythroid lineage was not involved in the 4 Ph+ ALL specimens without BCR/ABL positive cells. While the BCR/ABL positive percentage of myeloid cells was 11% (1/9), 8% (1/12), 0% (0/8) and 10% (1/10) respectively and that of lymphoid cells was 97% (76/78), 98% (87/89), 98% (97/99) and 97% (75/77) respectively. On the other hand, the BCR/ABL positive percentage was 100% (8/8), 91% (10/11), 82% (9/11), 88% (7/8) in the erythroid lineage, 89% (8/9), 96% (94/98), 100% (47/47), 98% (40/41)in the myeloid lineage and 96% (78/81), 93% (52/56), 96% (68/71), 95% (58/61) in the lymphoid lineage respectively for the 4 CML-LBC specimens. The BCR/ABL positive percentages of the other 2 specimens were all above 80% and through MGG-FISH analysis we also identified the source of the malignant clones and ascertained the diagnosis of the 2 patients. Conclusions The MGG-FISH technique has proved useful in providing rapid and precise differentiation between Ph + ALL and CML-LBC. The source of the malignant clones can also be analyzed by this technique.

BACKGROUND: Limb allograft is a sort of composite tissues allotrans plantation(CTA), some researches showed that the apoptosis of target cell is one of the main mechanism of the dysfunction of allograft.OBJECTIVE: To investigate the characteristic of cell apoptosis in acute rejection of limb allograft in rats based on limb allograft model.DESIGN: A randomized controlled trial using the experimental animals as the objects.SETTING: Experimental animal center Laboratory of a hospital of a medical university MATERIALS: The experiment was done in the Experimental Animal Central Laboratory of the First Affiliated Hospital of Harbin Medical University from November 2003 to May 2004. Totally 56 healthy and male SD rats and 28 Wister rats were involved with body mass of 200 to 250 g. The rats were provided by the Experimental Animal Center of the First Affiliated Hospital of Harbin Medical University. They were randomly divided into two groups:transplantation group with 28 Wistar rats and 28 SD rats and control group with 28 SD rats.INTERVENTIONS: The transplantation group of SD rats underwent limb allotransplantation from allogenetic Wistar. The control group of SD rats underwent limb replantation. The expression of acute rejected in limb allografts was observed. The limb grafts were harvested atday 1, 3, 5 or 7 after transplantation. Histopathological rejection grade of each tissue rejection was performed with hematoxylin-eosin staining. Apoptotic cells were detected by TUNEL (terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling) and apoptotic index(AI) was calculated.MAIN OUTCOME MEASURES: Primary results: ① istopathological grade of acute rejection in limb allografts ② The relationship between apoptosis and acute rejection in limb allograft in rats; Secondary results:General condition of rats in each group.RESULTS: The limb grafts showed edema and erythema and the skin became red at day(3.43 ±0.79) after transplantation. The average survival time was(7.42 ± 1.72) days. The acute rejection in skin was the strongest. On the day 3, 5 and 7 after operation, the histopathological rejection grades of skins in the transplantation group were(1.14±0.38) ,(2.28 ±0.48) and(2.86 ±0.38) grades respectively. They were significantly different from that of muscle and nerve( P ＜ 0.05 ) . The apoptotic cells in allografts were mainly infiltrating lymphocytes in subcutaneous tissues and then the muscle cells. All was positively correlated with acute rejection grade in limb allograft .CONCLUSION: Apoptosis was involved in acute rejection of limb allograft in rat. The apoptotic index can be used as a quantitative index to estimate the injury of grafts.

Objective To investigate the radiosensitizing effect of 17AAG-cypate micelles on human non-small cell lung cancer A549 cells and its possible mechanism.Methods (1) A single-hit multi-target model formula was used to analyze the radiosensitizing effects of 17AAG-M and 17AAG-cypate-M.(2) The effects of 17AAG-cypate-M on the viability of A549 cells under laser and X-ray irradiation were analyzed by MTT assay.(3) The effect of the drugs on the cell senescence was observed by β-galactosidase staining assay.(4) The effects of different treatment conditions on DNA damage repair were analyzed by γ-H2AX immunofluorescence staining assay.(5) The expression of p-Erk1/2 and p-Akt was measured by Western blot.The paired t test was used for analyzing the differences between groups.Results Compared with the X-ray irradiation group,the X-ray+17AAG-cypate-M group had a lower mean lethal dose and a sensitization enhancement ratio greater than 1,indicating that 17AAG-cypate-M had a radiosensitizing effect.Compared with the 17AAG-M group,the 17AAG-cypate-M group showed significantly lower cell viability (P<0.01),a significantly higher percentage of aging cells (P<0.01),and significantly further delayed DNA damage repair (P<0.01).And the 17AAG-cypate-M group had lower expression of p-Erk1/2 and p-Akt than the 17AAG-M group.Conclusions Compared with 17AAG-M,17AAG-cypate-M has a higher radiosensitizing effect on A549 cells.The mechanism might be inducing the cell senescence,delaying DNA damage repair,and inhibiting the expression of p-Erk1/2 and p-Akt.

Objective To investigate the clinical effect of compound Qianglizhenxian pill on epilepsy. Methods 94 patients with epilepsy were treated with compound Qianglizhenxian pill, and every course lasted three months. After treatment, checking patients' electroencephalogram again, evaluating the clinical effect according to degrees and numbers of attack. Results After 12 months of treatment, 90.43% of patients got an obvious improvement, while 6.38% for better, 3.19% for validity and no invalid. Conclusions There is a great improvement for patients with epilepsy after taking the medicine. The effective control rate is 92.62% with no toxication on blood system, liver and kidney.

Objective To investigate the effect of CT total tumor perfusion parameters in the identification of benign and malignant lung lesions. Methods 80 patients with pulmonary space-occupying lesions who were treated in our hospital from December 2015 to May 2017 were enrolled. After pathological histological diagnosis, 52 cases of malignant lesions and 28 cases of benign lesions were confirmed. And they all underwent CT whole tumor perfusion scan and perfusion imaging analysis. And the blood flow, blood volume, average transit time and surface permeability were obtained.Some cases of histopathological specimens were treated by monoclonal CD34 staining method, and the microvascularrich areas in the specimens were selected for blood vessel counting. The morphology of blood vessels was observed and microvessel density was obtained. Results The levels of BF, BV, MTT and PS in benign lesions were significantly lower than those in patients with malignant lesions. The best diagnostic threshold, sensitivity and specificity of BF in lung occupying lesions was> 49.12 mL/100 mL, 80.00％ and 52.40％. The best diagnostic threshold, sensitivity and specificity of BV was 6.68 m L/100 mL, 70.00％ and 81.00％. The best diagnostic threshold, sensitivity and specificity of MTT was3.82 m L/100 mL, 97.50 ％ and 52.40％. The best diagnostic threshold, sensitivity and specificity of PS was 7.25 mL/100 mL, 92.50％ and 90.50％. The malignant lesion microvessel density count in the malignant lesions was significantly higher than that of benign lesions, and the difference was statistically significant (P<0.05). Conclusion CT total tumor perfusion parameters have a great auxiliary effect on the diagnosis of benign and malignant lung occupying lesions. Among them, PS and BV have the strongest sensitivity and are worthy of clinical promotion.

Objective To study the therapeutic effects of Ag85A plasmid DNA vaccines in a mouse model of multi-drug resistant-(MDR-) Mycobacterium tuberculosis infection. Methods BALB/c mice were infected with Mycobacterium tuberculosis clinical strain HB361 with isoniazid and rifampin resist-ance by intratail-vein injection and were subsequently divided into 6 groups. At the third day after infection, the mice were treated with saline (group A), vector pVAX1 (greup B), rifampin (group C), vaccae (group D), Ag85A plasmid DNA vaccines (group E),rifampin and Ag85A plasmid DNA vaccines (group F) for 60 d. The lungs and spleens from the mice were taken and their pathological changes, weight and number of myeobacterial colony were examined at the third week after the end of treatment. Results At third week af-ter the end of treatment, the gross pathological observation and histopathological examination in lung showed that the lung lesions were limited, the profile of the alveoli was relatively clear, and normal structure could be seen in 2/3 areas of the lung sections in group D, E and F. The extent of lung lesion was 50% in group D,20% in group E and F. The pathological changes in group A, B, and C were more severer than those in group D, E and F. Compared with group A, the colony-forming units (CFU) in the lungs from mice in group D,E and F decreased 52%, 68%, 78%, respectively. The CFU in the spleens from mice in group D,E and F decreased 48%, 65%, 79%, respectively. Conclusion Ag85A plasmid DNA vaccines alone or Ag85A plasmid DNA vaccines along with chemotherapy have significant therapeutic effects on the mouse model of MDR-Mycobacterium tuberculosis infection.