1999年 · 第112卷第05期

OBJECTIVE To evaluate the effect of lamivudine on the loss of serum hepatitis B virus (HBV) DNA, HBeAg/antiHBe seroconversion and ALT levels in chronic hepatitis B patients and its safety profile and tolerance compared with placebo. METHODS Four hundred and twenty-nine patients with chronic HBV infection as defined by positive HBsAg, HBeAg and HBV DNA were enrolled and randomized into lamivudine and placebo groups. Three hundred and twenty-two patients received lamivudine 100 mg daily and 107 patients received placebo treatment for 12 weeks. Then, all patients were offered a further 9-month open label lamivudine treatment. The efficacy and safety were evaluated with clinical, biochemical, hematological and virological parameters. RESULTS During the 12-week treatment period, 92.2% of lamivudine treated patients became HBV DNA negative (below 1.6 pg/ml) compared with only 14.1% of those receiving placebo (P<0.01). At the end of 12 week, the sustained negative rate for HBV DNA in the lamivudine treated group was 78.5% compared with the placebo group (11.1%; P<0.01). There was a trend to a high proportion of patients treated with lamivudine to lose HBeAg (8.1%) and develop antiHBe (10.2%) than treated with placebo (5.3% and 6.4% respectively), but this difference was not statistically significant. Patients with elevated ALT levels at baseline became normal in 60. 3% of the lamivudine treated group compared with the placebo group where only 27.5% were normal (P<0.01). Lamivudine was well tolerated in a dose of (100 mg daily) and the overall incidence of adverse events was similar to that of the placebo. CONCLUSIONS Lamivudine (100 mg daily) is very effective in the inhibition of HBV replication, indicated by the rapid loss of serum HBV DNA, and often accompanied by a decrease of serum ALT levels. Lamivudine is well tolerated without severe adverse events during treatment.

OBJECTIVE To develop a universal quantitative immunoenzyme assay (EIA) for detecting amplified products of nucleic acid and its application in hepatitis C virus (HCV). METHODS The appropriate cycle number of amplification was selected to stop polymerase chain reaction (PCR) before the "plateau stage". At the same time, primers HCV (3) of the second PCR were modified with biotin so that the amplified products were labeled. The products were diluted and subsequently added to the streptavidin-coated wells, and the biotinylated products were captured, followed by denaturation of NaOH, and non-biotinylated strands were removed. Hybridization was performed by adding the specific probe labeled with fluorescein. Finally antifluorescein horse radish peroxidase (HRP) conjugates were added, after washing, 3, 3', 5, 5',-tetramethylbenzidine (TMB) was added to the wells and then measured on a microplate reader. RESULTS EIA detection of amplified products of HCV showed that this assay was rapid, sensitive, specific and accurate. Correlation between the initial number of viral template and the EIA of amplified products was good. We also prospectively investigated the response to interferon in five patients with HCV coinfection. Results showed that this assay could be used as a guidance to the clinical therapy in directing the use of antiviral drugs. CONCLUSIONS This assay could be widely used as a universal technique for the quantitative detection of amplified products of all nucleic acid (such as virus, bacterium) and other human genes (such as HLA B27), it has vast vistas.

OBJECTIVE To investigate the vertical transmission rate of Chlamydia trachomatis (CT) in Chongqing, China. METHODS Specimens taken from 278 women and from their 79 infants were examined by cell culture, polymerase chain reaction (PCR) and DNA sequence analysis. Chlamydia trachomatis was isolated in McCoy cell culture. CT DNA was extracted with a modified Nal method. After cloning, recombinant plasmids were used for sequence analysis with the dideoxy chain termination method. RESULTS 10.8% (30/278) of the cervical cultures of pregnant women were positive for Chlamydia trachomatis, while the positive rate tested by PCR was 14.0% (39/278). The vertical transmission rate of Chlamydia trachomatis was 55.0% (11/20). The incidences of conjunctivitis and pneumonia in infants with Chlamydia trachomatis positive mothers were 27.3% and 18.2%, respectively. DNA sequences of Chlamydia trachomatis isolated from the cervix of a mother and the nasopharynx of her baby were identical. CONCLUSIONS Chlamydia trachomatis infection is quite common in Chongqing, China. Our report is the first report of CT vertical transmission proved by DNA sequence analysis.

OBJECTIVE To evaluate the relative contribution of every part of vascular wall to lumen area loss after angioplasty. METHODS Angioplasty was performed on rabbits iliac atherosclerosis model. Histomorphometric RESULTS of histological cross-sectional areas from target vessels of animals killed immediately after angioplasty (acute group, n = 20) were compared with those of the same areas from animals killed 4 weeks after the procedure (chronic group, n = 23), when restenosis occurred in this model. If taking 30% diameter decrease as restenosis criteria, the chronic group was further divided into two subgroups (nonrestenosis subgroup [NRS, n = 9] and restenosis subgroup [RS, n = 14]) and their histomorphometric RESULTS were also compared. RESULTS The lumen area decreased by 0.48 mm2 from acute group to chronic group (acute group 0.86±0.23 mm2 vs chronic group 0.38±0.20 mm2, P<0.001). Over the same period intimal area increased by 0.12 mm2 and the area circumscribed by inner elastic lamina (IEL area) decreased by 0.36 mm2. Intimal hyperplasia could only explain 25% of the later lumen area loss. We further found a more significant IEL area decrease in RS subgroup than in NRS subgroup (0.68±0.35 mm2 vs 0.94±0.14 mm2, P<0.05), despite the same intimal area (0.39±0.16 mm2 vs 0.41±0.31 mm2, P = NS). The different lumen area between RS subgroup and NRS subgroup (0.27±0.10 mm2 vs 0.55±0.10 mm2, P<0.001) was predominantly due to the greater IEL shrinking in RS subgroup. CONCLUSIONS Our data indicate that later lumen area loss after angioplasty results from both intimal hyperplasia and arterial shrinking (remodeling), but the latter plays a much greater role.

OBJECTIVE To determine the function of, in vivo, renin and its role in the pathogenesis of hypertension. METHODS A renin-2 gene restriction map was constructed by endonuclease digestion and Southern blotting hybridization. Transgenic rats were produced via microinjection method. RESULTS The 24 kb fragments containing mouse full-length ren-2 and its flanking sequence were cleaved by single enzymes (EcoR I, Kpn I and BamH I) and combined enzymes (EcoR I/Kpn I, Kpn I/BamH I and BamH I/EcoR I), respectively. The digests were electrophoresed in 0.8% agarose plates and transferred onto NC membranes. Radioactive 735 bp and 1400 bp probes obtained from half and full-length renin-1 cDNA were used in southern blotting hybridization. According to the electrophoresis and hybridization patterns, a ren-2 restriction map was constructed. 1603 fertilized rat ova after injection with purified 24 kb renin-2 gene were implanted into the oviducts of 81 pseudopregnant recipients in about 20 ova per female rat. 306 progenies were obtained from 50 foster mothers (average of pregnancies was 56.6%). 248 survived pups were identified by PCR analysis and Southern hybridization, and eight positive rats were found to be the transgenic rats (founder, F). All of them carried long fragments (24 kb) of renin-2 gene with normal blood pressure. Preliminary breeding and screening were carried out in the founder. Total survival pups (17.8%) and overall efficiencies (1%) were harvested as the same as those reported in the literatures. A systemic observation and the problems occurred during production of transgenic rats were also described besides the technique procedure used in this study. CONCLUSIONS Mapping of full-length murine ren-2 can be used in investigation of the structure and function of the gene. The results denoted that the ren-2 transgenic rats were successfully established in this study and the technique used in the production of transgenic rats was proved to be valid in leading to wide-spread application of transgenic technique to many other related researches.

OBJECTIVE To investigate the change characteristics of neuropeptide Y (NPY) release during acute myocardial ischemia period. METHODS The animal test was carried out in in situ perfused guinea pig hearts with intact sympathetic innervation. Electrical stimulation-evoked exocytotic release of NPY during ischemia and reperfusion was tested by radioimmunoassay (RIA). The plasma NPY concentrations were measured in patients with acute myocardial infarction (AMI) and angina pectoris (AP) in different times. RESULTS Electric stimulation of the left ganglion in guinea pig heart evoked an exocytic release of neuropeptide Y. Stimulation after 20 minutes of global ischemia (S2), compared with control period stimulation (S1) produced the inhibition of NPY to a certain extent (S2/S1: 0.72, P<0.05), whereas the inhibition of NPY release disappeared after 5 minutes reperfusion (with S2/S1 of 1.01, P>0.05). Ischemia alone, without the electric stimulation, did not apparently induce NPY release. The clinical test found that the plasma NPY level was increased significantly during the acute ischemia attack period of coronary heart disease (CHD). The plasma NPY level reached peak (136.3±66.5 pg/ml) in patients during the first day after AMI. It began to decrease from the third day and came to normal level in the end of the first week. The plasma NPY level was 159.3±98.5 pg/ml in AP patients during angina attack. After two weeks treatment, the plasma NPY level was decreased to 118.9±54.3 pg/ml (P<0.05). CONCLUSIONS The NPY release of global ischemia have some relation with sympathetic nerve activity. At the early stage of ischemia, NPY release is inhibited to some degree and the inhibition factors will fade away on reperfusion. NPY interferes with the pathogenesis and the pathophysiolgy.

OBJECTIVE To determine whether extraadrenal tissues synthesize aldosterone in addition to vascular tissue and brain. METHODS Ex vivo kidney perfusion was performed in normal Wistar rats, ACEI pretreated and adrenalectomized rats prior to the perfusion experiment. After equilibration for 30 minutes, 120 ml of perfusate was collected and subjected to reverse-phase HPLC and then aldosterone was measured by RIA. By RT-PCR and Southern blot the expression of aldosterone synthase gene-CYP11B2 mRNA was studied in both kidney tissue and cultured renal tubular epithelial cell, lung and liver tissues. In situ hybridization was used to identify the cell types of liver and lung expressing CYP11B2 mRNA. RESULTS Production of aldosterone in the kidney perfusate was not changed in adrenalectomized rats although it was decreased in the group pretreated with ACEI perindopril. By RT-PCR and Southern blot the expression of CYP11B2 mRNA was demonstrated in both kidney tissue and cultured renal tubular epithelial cell. We have also identified CYP11B2 mRNA expression in liver and lung of rats. In situ hybridization showed that CYP11B2 mRNA was localized in the endoplasm of liver fat-storing cell (Ito cells) and type II alveolar cells of lung. CONCLUSIONS These studies prove that kidney, liver and lung are able to produce aldosterone.

OBJECTIVE To develop a new technique based on long distance polymerase chain reaction (LD-PCR) to replace Southern blotting method to detect Factor VIII (FVIII) gene inversion leading to severe Hemophilia A (HA) and carrier. METHODS Four primers P, Q, A&B were designed and synthesized. P&Q is specific for 5' and 3' flanking regions of F8A1 respectively. A&B is specific for 5' and 3' flanking regions of F8A2/F8A3 respectively. LD-PCR with 3 primers and 3 temprature was set up, optimized and used to detect the inversion. RESULTS The LD-PCR with primers P, Q, A&B, P, Q&B and P, Q&A can be used to detect the gene inversion and discriminate carrier from wild type. A blind analysis of 53 DNA samples from HA families was carried out by the LD-PCR and Southern blotting respectively. Two sets of the RESULTS were completely identical. They were 23 cases of inversion, 27 cases of wild type and 3 cases of carriers. The sensitivity and specificity of LD-PCR are both 100%. Three inversion hemizygotes and 4 female carriers were identified from 5 HA families by the LD-PCR technology. CONCLUSIONS The LD-PCR with primer P, Q&B or P, Q, A&B can be used to detect the gene inversion and the carrier of inversion. Compared with Southern blotting, this technique is simple, rapid, inexpensive, more sensitive, accurate and non-isotopic.

OBJECTIVE To investigate the effects of ischemic preconditioning (PC) and ATP sensitive K+ channels (KATP(+) opener nicorandil on reperfusion arrhythmias and electrophysiology. METHODS Langendorff-perfused rat hearts were subjected to ischemic PC with three cycles of 2 minutes of global ischemia or infusion of KATP+ opener nicorandil with subsequent 5 minutes global ischemia and reperfusion. The incidence of reperfusion arrhythmias, ventricular fibrillation threshold (VFT), effective refractory period (ERP) and monophasic action potential duration (MAPD) of the left and right ventricles were compared to those from control rat hearts. RESULTS The results indicated that PC reduced the incidence of total arrhythmias and ventricular fibrillation during reperfusion (P<0.05, vs controls). PC markedly delayed the onset of arrhythmia after reperfusion (P<0.01, vs controls). PC significantly enhanced the VFT values during reperfusion and shortened the ERP and the MAPD during ischemia. VFT was restored more rapidly than that in controls. KATP+ opener nicorandil neither reduced the incidence of total arrhythmias and VF nor delayed arrhythmia onset. Nicorandil shortened ERP and MAPD90 without enhancing the VFT values, and VFT returned to normal as slowly as that in controls. CONCLUSIONS We conclude that PC protects the globally ischemic rat hearts from reperfusion arrhythmias. The antiarrhythmic effect of PC is likely to be related to a significant increase of VFT. KATP+ opener nicorandil has no potential antiarrhythmic action and KATP+ channels may not play a major role in the antiarrhythmic effects of ischemic PC in isolated rat hearts.

OBJECTIVE To study the clinical results of hepatic radioembolization with Yttrium-90 (90Y) glass microspheres in the treatment of primary liver cancer. METHODS Seventeen patients with liver cancer were treated with glass microspheres from August 1996 to May 1998. Hepatic radioembolization with 90Y and lipiodol-ultrafluid was used. Percutaneous port-catheter system (PCS) implantations via femoral artery were performed in 12 patients. RESULTS In the 17 patients, their mean ratio of absorbed doses between tumor and normal liver was 2.4:1. CT showed a significant reduction in tumor size in 11 of the 17 patients. Average survival was 19.5 months. The indwelling catheters of all the 12 patients were patent and no catheter tip locations were found. CONCLUSIONS 90Y glass microsphere is one of the best radioisotopes. Not only good responses to the therapy of 90Y glass microspheres can be achieved in patients with metastatic liver cancer, but also in those with primary liver cancer, specially the localized or hypervascular mass. The patients with massive arterioportal shunt should not be limited to this form of radiation therapy. The percutaneous PCS implantation via the femoral artery is a new passageway for the treatment of primary liver cancer with 90Y glass microspheres and other interventional therapy.

Purpose To summarize the major achievements of Chinese work on the epidemiology and prevention of hemorrhagic fever with renal syndrome (HFRS) in recent years, and to give a general review on the present situation of HFRS in China. Data sources Reviews and papers published in Chinese journals, relevant to the objectives, written or collected by the author; and the research work of the author. Study selection An outline was drafted according to the purpose given above, and the relevant materials and data were grouped together into different items outlined. Data extraction All the materials and data extracted were published or specially assessed. Results HFRS was first recognized in the northeastern China in 1931. It has been found prevalent also in many other parts of China since 1955, and presently, 28 out of 31 provinces (autonomous regions, or municipalities) have been proved to be its endemic areas. The total number of cases is 1,256,431 from 1950 to 1997, with 44,304 death (3.53%). 50-100 thousands of cases could be registered annually since 1981 when the presence of the Rattus-type HFRS was first identified serologically in China, with the highest peak in 1986 (115,985 cases). Three types of endemicity have been differentiated: the Apodemus-type, the Rattus-type and the mixed type of the two, by their peculiar seasonal distributions of cases and by methods of serotyping (HI, Mabs serotyping kits). The epidemiologic features of the Rattus-type HFRS are quite different from that of the Apodemus-type. 67 species of vertebrates were found to harbor hantavirus antigen or antibodies, but the chief or primary reservoir hosts are Apodemus agrarius and Rattus norvegicus only. Besides the commonly recognized enzootic mode of transmission (via contacts with the reservoir rodents or their excreta), mite transmission (including certain species of gamasid mites, and chigger mites) has been identified as the potential vectors and reservoir hosts with HFRS. Vertical transmission had been found in pregnant patients with HFRS, and in all three chief host rodents (Apodemus agrarius, Rattus norvegicus, the laboratory rats). The relatively high inapparent infection rates in the population of endemic areas of the Rattus-type HFRS after big outbreaks (8%-20%) are suggested to play a significant role in the gradual decline of the incidence of HFRS in that areas. Three kinds of inactivated vaccines against HFRS (the golden hamster kidney cell vaccine, the Mongolian gerbil kidney cell vaccine and the purified suckling mouse brain vaccine) have been successively developed and proved highly effective in the prevention of HFRS. Conclusion Chinese workers on the epidemiology and prevention of HFRS in China have accomplished great achievements since 1981. As the relatively high annual incidence of HFRS sustained in recent years, control measures have to be reinforced. Many problems of HFRS and hantavirus infections are still not well understood or unclear, such as the poly-host nature, and the multi-modes of transmission, the natural history of hantaviruses and their genetic variation occurred in nature etc. The task for effective control of HFRS and the research on its epidemiology in China should be held on a high priority.