Abstract:Objective: To explore gene diagnosis of autosomal dominant polycystic kidney disease (ADPKDⅠ) and to look for the typical mutation to improve the gene diagnosis. Methods: Southern blot and PCR was used to observe the mutation condition of 3′end single copy region of ADPKDⅠ gene; Amplifing and analysing the microsatellite SM7 by PCR. Results: (1) After the probe AH4 was hybridized with 16 patients′ genomic DNA by Southern blot, the common 15 kb fragments were found in every one; (2) For 27 patients, 5.72 kb genomic DNA, which is between the probe AH4 and JH14, was amplified by PCR, and no 5. 5 kb genomic DNA deletion were found in this region; SM7 was amplified in 109 health persons, its PIC was 0.76, and was closely linked with ADPKD Ⅰ gene in 3 patients′ family. Conclusion:(1) No large genomic DNA segment deletion can be found frequently in ADPKD Ⅰ gene 3′end single copy region; (2) The PIC of SM7 is high, it can be used to make rapid gene diagnosis in about 70%？80%ADPKDⅠfamily.

Abstract:Objective：To study the functions of the first extracellular domain of β chemokine receptor 5(CCR5) NH2-terminal and to prepare its specific antibody F(ab′)2. Methods: Some β chemokine superfamily members were analyzed by computer and the least homologous domain of the extracellular loops were located. The first extracellular domain 114 nucleotides fragment was defined, sequenced and amplified by PCR，the expression vector pGEX-IN/NR5 of a recombinant GST fusion protein was constructed. After confirming the correctness of the inserted sequence，the transformation and expression of this fusion protein were performed in E. coli. The expression products of the fusion protein were purified and 2 New Zealand rabbits were immunized. An anti-CCR5 NH2-terminal antibody F (ab′)2 was prepared by protein A affinity chromatography，pepsin digestion and Sepharose-12 column chromatography. Results： Reduced, unreduced SDS-PAGE and FAX analysis demonstrated that this F (ab′)2 had high specificity to combine with CCR5. Conclusion：In this paper，we introduce a simple and quick method to get a specific antibody F (ab′)2 of certain functional domain. By that，not only can we get an important experiment material for studying gene expression，but also a good idea and technique to study other high similar superfamily members.

Abstract:Objective: To observe the effect of endothelin A receptor antagonist BQ610 on acute myocardial ischemic arrhythmia in cats and to study the role of endothelin in the development of acute ischemic arrhythmia. Methods: Acute myocardial ischemic arrhythmia was induced by the occlusion of the initial segment of left anterior descending part of the coronary artery. Arrhythmia during 30 min of the ischemia was observed and the effects of BQ610 on the arrhythmia was evaluated. Results: BQ610 dose-dependently antagonized the acute ischemic arrhythmia. Conclusion: Mediated by endothelin A receptor, endogenous endothelin may play an important role in the development of acute ischemic arrhythmia in cats.

Abstract:Objective：To study the expression of P16 gene and its correlation with clinical cases. Methods：The levels of P16 gene products were examined immunochemically in resected specimens of human laryngeal squamous cell carcinoma. Results： Carcinomas tested： 47.62%（20/42） showed positive staining in the lesions. 40.00%(4/10) in supraglottic cancer group, 57.89%(11/19) in glottic cancer group, 38.46%(5/13) in subglottic cancer group, 75.00%(12/16) in cell differentiation gradeⅠgroup, 30.77%(8/26) in cell differentiation grade Ⅱ？Ⅲ group, 20.00%(3/15) in lymph node metastasis group, 62.96%(17/27) in those without metastasis, 40.00%(8/20) in clinical stageⅠgroup, 54.54%(12/22) in stageⅡ？Ⅳgroup. Conclusion：The expression of P16 protein is related with malignant degree and lymph metastasis in human laryngeal carcinoma.

Abstract:Objective：To study the ultrastructural morphological changes at millisecond time course level in skeletal muscle during excitation-contraction coupling after electrical stimulation. Methods: Two-way infrared detectors with a computer to control the signal of electric stimulation and quick freezing fixation were employed in this test. The ultrastructure of toad skeletal muscle was studied by transmission electronic microscopy after its skeletal muscle was stimulated in 0.8, 5.6, 8.4 ms. The skeletal muscular ultrastructure at known millisecond time course were observed following quick-freezing after a single electrical stimulation. Results:The space between sarcolemma and muscle myofibril was strikingly narrowed from 0.8 ms to 8.4 ms after electric stimulation, and 2 holes were observed in the front of sarcoplasmic reticulum membrane at the same time. Conclusion: During excitation-contraction coupling, the sarcolemma has positional shaft and space changes with myofibril when depolarization occurs, besides, there are small holes on the sarcolemma when skeletal muscles contract.

Abstract:Objective: To study the protective effect of grafting NT-4 genetically modified cells on injury of spine motoneurons in rats. Methods: NT-4 genetically modified cells were obtained by infecting L-6TG cell in vitro with a retroviral vector pN2A containing the rat NT-4 cDNA gene. The rats with left sciatic nerve apocoped were taken as model for treatment by implanting NT-4 genetically modified cells. The right sciatic nerve was taken as control. The Nissl staining and ChE staining in motoneurons of spine were also studied. Results: There were significant differences between groups of NT-4 and control in the Nissl staining and ChE staining. Conclusion: Grafting NT-4 cells can prevent most motoneurons from death.

Abstract:Objective: To establish apoptosis model of cultured lung cancer cells. Methods: A549 human lung adenocarcinoma cells were treated in cultured fluid containing 20 μmol/L doxorubicin for 4 h. At different time points of post-incubation the detached cells were harvested for analysis by counting, trypan blue exclusion, acridine orange dying, electromicroscopy, DNA agarose gel electrophoresis and flow cytometry. Results: The morphology of the treated cells was characterized by positive trypan blue exclusion, membrane budding, cytoplasmic condensation, chromatin condensation, fragmentation, margination and apoptotic body formation. Agarose gel electrophoresis analysis revealed DNA cleavage (called DNA ladder). An apoptotic peak appeared in the analysis of flow cytometry. All those results suggested that apoptosis of A549 cells was successfully induced by doxorubicin. Most of the apoptotic cells appeared in the detached cell group after 10-12 h of post-incubation. Conclusion: Apoptosis of A549 lung cancer cells can be induced by doxorubicin.

Abstract:Objective: To provide a permanent wound coverage by using skin allografts in mouse model with fullthickness burn of 20%TBSA. Methods: The recipient mouse was treated with 9 Gy total body irradiation from 60Co source, followed by infusion of a mixed donor and recipient T-cell depleted bone marrow where a state of specific unresponsiveness to the skin allograft was induced without the need for chronic immunosuppression. Five experimental groups were divided: group Ⅰ, 20%TBSA burn of C57B/6 mice with skin allograft from a BALB/c mouse at 48 h without irradiation; group Ⅱ, similar to group Ⅰ but with irradiation; group Ⅲ, similar to group Ⅱ but with the infusion of mixed bone marrow; group Ⅳ, similar to group Ⅲ but with skin allograft from a C3H/He mouse; group Ⅴ, controlled C57BL/6 mice with homograft. Results: There were no statistical difference irranimal mortality in all groups except group Ⅱ, and no evidence of infectious morbidity and graft vs host disease were found. Mean skin allograft survival (mice died with intact graft were excluding) was as follows: group Ⅰ, (8.4±1.5) d; group Ⅱ, (17.5±3.7) d; group Ⅳ, (32.5±8.1) d, and group Ⅴ, 90 d; gorup Ⅲ, (78.2±5.6) d (P<0.05 vs group Ⅰ, Ⅱ and Ⅳ). Conclusion: C57BL/c mice exhibited specific transplantation unresponsiveness to the BALB/c mouse skin, but rejected the third party MHC-disparate C3H/He mouse skin. This study suggests the potential use of induced specific unresponsiveness to skin allografts for wound coverage in thermal injury.

Abstract:Objective: To study the mechanism of Traditional Chinese Medicine Tuojiagao in cleaning necrotic and denatured tissue in deep burn wound. Methods: Two groups of patients with deep burn wound were studied: (1) Traditional Chinese Medicine Tuojiagao group, (2) Ag-SD frost group. Tissue and exudate of the wound were collected. Macrophages were marked with anti-CD68 antibody in tissue slides. The bacteria of wound exudate were counted. Results: The sections from wounds after using Tuojiagao have more CD68+ cells (macrophages) than Ag-SD group, and no difference of bacteria count was found between the 2 groups. Conclusion: Tuojiagao could clean necrotic tissue through recruiting macrophages to scavenge and break down it.

Abstract:Objective: To study the effects of hypertension and aging on structure and function of coronary vasculature. Methods: Spontaneously hypertensive rats(SHR) and normotensive Wistar-Kyoto rats(WKY) strains at 4,16,26,40, and 55 weeks of age were studied. Maximal coronary flow(MCF) was measured using an isolated perfused rat heart model. Outer dimeter(OD), lumen dimeter(LD), cross-sectional area(CSA), and CSA/LD were analysed by computer. Results: LD demonstrated a significant hypertension related decline and was not effected by aging. CSA and CSA/LD in arterioles<100 μm demonstrated a significant hypertension-related increase and was not effected by aging. There was a significant hypertension- and age-related increase in CSA and CSA/LD in arterioles>100 μm. In addition, there was a significant hypertension- and age-related decline in MCF. The tight linear relation between CSA/LD in arterioles>100 μm and MCF. Conclusion: Both hypertension and aging are accompanied by structural and functional alterations of the cornoary vasculature. Structural alterations in arterioles>100 μm may be the main reason for MCF decrease.

Abstract:Objective: To compare the therapeutic efficacy between intravenous PGE1 and inhaled NO in the treatment of acute pulmonary hypertension. Methods: In 7 dogs, pulmonary hypertension was induced by infusion of the thromboxane analogue U46619 to observe the effect of intravenous administration of small dose PGE1 10 ng/(min*kg), large dose PGE1 50 ng/(min*kg),inhaled NO 4.0×10-5and inhaled NO plus small dose PGE1 on the hemodynamics of acute pulmonary hypertension. We measured the changes of hemodynamic variables after each drug given. Results: Intravenous administration of small dose PGE1 resulted in a decrease in MPAP by (17±8)%, MSAP by (19±10)%, and an increase of CO from (1.21±0.25) L/min to (1.83±0.27) L/min(P＜0.01). Intravenous administration of large dose PGE1 resulted in a decrease in MPAP by (29±7)%, MSAP by (41±11)%, and CO from (1.21±0.25) L/min to (1.04±0.38) L/min (P<0.05). Inhaled NO resulted in a decrease in MPAP by (23±6)%, but no significant changes of MSAP and CO were observed. Inhaled NO plus small dose PGE1 resulted in a decrease in PAPm by (31±7)%, SAPm by (14±9)%, and an increase of CO from (1.34±0.17) L/min to (2.01±0.34) L/min (P<0.01). Conclusion: Inhaled NO plus small dose PGE1 can not only decrease pulmonary artery pressure significantly, but also dilate the system artery and improve cardiac function, which will be useful to blood infusion of other tissues.

Abstract:Objective： To investigate the morphological changes of the cultured SAN cells in ischemia model and the protective effects of captopril on the SAN cells with ischemic injury. Methods: One hundred and fifty SD rats of 4 d old were used to develop primary cell culture of SAN cells. Then, living cells which included the cells in normal growing condition and in ischemic model were observed. In addition, living cells of SAN were fixed and stained with HE, and observed by light microscopy and transmission electromicroscopy(TEM). Results: The vitality, the morphologic characteristic and architecture of cultured SAN cells were slightly injured at 60 min,obviously injured at 90 min and seriously injured at 120 min following ischemia, whereas in the group treated with captopril the injuries were found to be significantly relieved. Conclusion：The ischemic injuries of cultured SAN cells in ischemia model were intensively developed with prolonged ischemia and the obvious protective effects of captopril on cultured SAN cells in ischemic model were observed.

Abstract:Objective: To study the effects of protein kinase C (PKC) inhibitors on the injury and proliferation of rat aortic vascular cells induced by fibrin fibrinogen degradation products (FFDP). Methods: The injury of rat aortic vascular endothelial cells (RAEC) was determined by the lactic dehydrogenase (LDH) release and the proliferation of rat aortic vascular smooth muscle cells (RASMC) by crystal violet staining. Results: FFDP obviously increased the LDH release of RAEC and the proliferation of RASMC. Quercetin and Ro 31-8220 inhibit the effects of FFDP on rat aortic vascular cells in a concentration-related manner. Conclusion: PKC inhibitors exert protective effect on the injury and proliferation of rat aortic vascular cells induced by FFDP.

Abstract:Objective: To study the red cell immune function and natural killer (NK) cell activity in the development of endometriosis. Methods: The red cell immune functions of 44 patients with endometriosis were assayed by means of erythrocyte C3b receptor rosette test and erythrocyte Ic rosette test. The NK cell activities in peripheral blood was assayed by means of MTT assay and compared with that of controls. Results: The RC3bR and NK cell activity of patients with endometriosis were lower than those of controls (P<0.05), and decreased as the stage of endometriosis increased. The RIcR of patients with stageⅢ,Ⅳendometriosis was higher than that of control (P<0.05). Positive linear relativity was found between RC3bR and NK cell activity in patients with endometriosis (r=0.665 9, P<0.01). Conclusion: The defects of red cell immune function and NK cell function may be related to the pathogenesis of the endometriosis.

Abstract:Objective: To study the role of potassium channel activator in treatment of asthma. Methods: Airway insufflation pressure measurement(AIP), bronchial alveolar lavage fluid(BALF) cells count and pathological analysis were employed to detect airway resistance variation and airway inflammatory cells infiltration following inhalation of ovalbumin. Results: BRL 55834(8 μg/kg) inhibited not only ovalbumin induced AIP increase but also inflammatory cells infiltration in sensitized guinea-pigs, it did not lower blood pressure. By contrast to BRL 55834, BRL 38227 (200 μg/kg) and aminophylline(25 mg/kg) inhibited ovalbumin induced inflammatory cells infiltration, but BRL 38227 lowered blood pressure markedly. Verapamil (0.5 mg/kg) had no effects on AIP increasing and inflammatory cells infiltration. Conclusions: BRL 55834, a selective potassium channel activator, not only decreases airway resistance, but also inhibits inflammatory cells infiltration, and this is beneficial for treatment of asthma.

Abstract:Objective: To evaluate the effects of a new antifungal agent, S1123, on controlling fungal infection on burn wound in vitro and in clinic. Methods: Compare their antifungal effects with 3 different antifungal drugs, naftifine, ketoconole and S1123 by testing their minimal inhibitory concentration (MIC). S1123 and a control drug were used to treat burn wound. The crusts were taken to culture before and after treatment for calculating the negative rate. One thousand nine hundred and twenty-seven patients suffered superficial mycoses on naval vessels were treated with S1123 cream. Results: S1123 was active in vitro against a broad spectrum of fungal and yeast pathogens. MIC of Aspergillus and Candida, 2 main pathogenic genura, was 0.156，0.625 μg/ml. In the clinical administration, invasive fungi of 79.71%burn wound disappeared in 3 d. Compared with the controls, the difference being signifficant statistically (P<0.01). The effective rate on superficial mycoses was 95.6%. Conclusion: S1123 is an new effective external antifungal drug.

Abstract:Objective:To put forward the concept of medical demand intensity(MDI) and to establish its measurement on the base of case mixed index(CMI) and all kinds of measurement of medical illness severity, it will help examine to which extent the health resources are needed by disease. Methods: All the cases of in-patients in 1996 in 3 hospitals were collected to caculate medical demand intensity index(MDII). Results: The current classification of hospital categories and ranks doesn′t suit the MDI degree and the efficiency of health resources. Conclusion: The resource allocation of health service should be based on the MDI， which gives the scientific basis for the reasonable proportion and structure of resource allocation in different type of hospitals， and also make it possible for efficient examination in hospitals.

Abstract:Objective: To study the expression of blood AFP mRNA in patients with recurrent or distant metastasis of human hepatocellular cacinoma (HCC). Methods: We examined 40 blood samples from patients with recurrent or distant metastasis HCC by nested RT-PCR to find out AFP mRNA. Results: AFP mRNA was detected in 7 blood samples from 19 HCC with recurrent HCC (36.8%) and in all HCC patients with distant metastasis(100%). Conclusions: AFP mRNA may be used as a marker of HCC recurrent and metastasis.