Objective To investigate the expression regulation of thyrotrophin-releasing hormone (TRH) and TRH receptor (TRH-R), and their role in the development of rat testis.Methods Oligonucleotide primers were designed from the sequences of rat hypothalamus prepro TRH (ppTRH) and pituitary TRH-R cDNA for reverse transcription polymerase chain reaction (RT-PCR). Specific fragments of ppTRH and TRH-R cDNA were cloned and sequenced. Expression plasmids containing ppTRH and TRH-R genes were then constructed, and expression was found in E.coli DH5-α. ppTRH and TRH-R mRNA in the testis was quantitated in RNA samples prepared from rats at different developmental stages by real time quantitative RT-PCR.Results The quantitative analyses demonstrated that no ppTRH and TRH mRNA could be detected at the earliest stage (day 8). ppTRH and TRH mRNA signals were detected on day 15 and increased progressively on days 20, 35, 60 and 90. Conclusion Our results suggest that rat testis could specifically express TRH and TRH-R, and the transcriptions of ppTRH and TRH-R genes in the rat testis were development-dependent. The acquirement of expressed products for ppTRH and TRH-R can be used for further research on the physiological significance of TRH and TRH-R expression in rat testis.

Objective To present a special case with the karyotype and molecular marker of acute myeloid leukemia (AML)-M2 who was induced to complete remission by all-trans retinoic acid (ATRA) alone.Methods A recently hospitalized young female patient with acute leukemia was initially diagnosed as M3 subtype based on morphological French-American-British (FAB) classification. Karyotype analysis using standard G and R banding techniques and RT-PCR were applied to further define the diagnosis. After primarily cultured bone marrow cells from the iliac aspiration were tested for in vitro induced differentiation, the patient was treated with oral all-trans retinoic acid alone, 60?mg per day until complete remission was achieved. Peripheral blood and bone marrow changes were monitored over the whole treatment course.Results The characteristic chromosomal aberration for M3, the t(15;17) reciprocal translocation, was not found while a t(8;21) translocation was verified. Furthermore, an amplified product of the AML-1/ETO fusion gene instead of the PML/RARα fusion gene was detected by RT-PCR and the diagnosis was corrected from M3 to M2. Primary cultured bone marrow cells can be fully induced to terminal differentiation after 4 days exposure to ATRA. A hematological complete remission was achieved after 40 days treatment with ATRA as a single therapeutic agent, suggesting an alternative pathway mediating ATRA-induced myeloid differentiation. Conclusion A leukemia patient with a subtype other than M3, such as M2 in this case, may also be induced to complete remission by the mechanism of ATRA-induced terminal differentiation. This implies that there may be a pathway other than PML/RARα fusion gene product which mediates ATRA-induced myeloid maturation in leukemia cells.

Objective To define the prevalence, severity and risk factors for lead poisoning in new immigrant children from the mainland of China to Hong Kong, China.Methods New immigrant children from the mainland of China under 18 years of age were invited to join the study. Their growth parameters and venous blood lead levels (BLL) were measured within 7 days of arrival. Those with elevated BLL i.e. >10?μg/dl (0.48?μmol/L) were assessed for signs, symptoms and risk factors of lead poisoning. Education on the prevention of lead poisoning and follow up BLL measurements were offered until their BLL normalized.Results Four hundred and fifty-seven children were recruited. Among them, 18.1% and 2.6% had BLL >0.48 and 0.71?μmol/L, respectively. None had BLL >0.96?μmol/L. Possible risk factors included contaminated drinking water (19%), herb ingestion (17.5%), pica, playing in dumping grounds, residing near paint factories or highways, habitual inhalation of car exhaust and cooking with petroleum. Symptoms and signs included abdominal pain, headache, short stature, and learning difficulties but did not correlate with BLL. None required treatment except for counseling on the avoidance of risk factors. About 94.7% showed a reduction in BLL 2-3 months after arrival and before counseling. All had normalized BLL by 9 months except 1 who had an X-ray feature of lead line.Conclusions Lead poisoning is common in new immigrant children from the mainland of China. Environmental factors were most important while behavioral factors like regular herb ingestion might have contributed to the persistence of high BLL in the minority.

Objective　To investigate the expression of the granulocyte-macrophage colony-stimulating factor (GM-CSF) and GM-CSF/IL-3/IL-5 receptor common beta chain (βc receptor) in an adult patient with idiopathic pulmonary alveolar proteinosis (PAP), so as to demonstrate the possible association of the GM-CSF and βc receptor with the pathogenesis of human PAP.Methods　The GM-CSF levels were measured with a commercial ELISA kit (sensitivity 5?pg/ml) and the βc receptor expression on the cell surface was detected by flow cytometry analysis. Reverse transcription-polymerase chain reaction (RT-PCR) analysis was employed to detect the expression of the GM-CSF mRNA and the βc receptor mRNA in peripheral blood mononuclear cells and alveolar macrophages. The entire coding regions of the GM-CSF cDNA and the βc receptor cDNA were sequenced by the Sanger dideoxy-mediated chain termination method to detect possible mutations.Results　The patient with PAP failed to release the GM-CSF protein either from circulating mononuclear cells or from alveolar macrophages. The expression of the GM-CSF mRNA was normal after the stimulation of lipopolysaccharide, whereas a point mutation at position 382 of the GM-CSF cDNA from “T" to “C" was revealed by cDNA sequencing, which caused a change in amino acid 117 of the protein from isoleucine to threonine. The βc receptor expression on the cell surface was normal, and the βc receptor mRNA expression and the sequence of the entire coding region of the βc receptor were also normal.Conclusions　The decreased GM-CSF production is associated with the pathogenesis of human PAP. A point mutation of the GM-CSF cDNA may contribute to the decreased GM-CSF production in our adult PAP patient. The mutation of the βc receptor in some of paediatric patients with PAP may not be a common problem in adult patients.

Objective To investigate the effect of overexpression of bcl-2 on ethanol-induced apoptosis of primary hepatocellular carcinoma (HCC) cells.Methods The retrovirus expression vector pDOR-SB containing human bcl-2 cDNA was introduced into a human HCC cell line HCC-9204 by liposome-mediated transfection. pDOR-transfected and non-transfected HCC-9204 cells were used as controls. The expression of Bcl-2 protein by transfected HCC-9204 cells was detected by the immunohistochemical method. Then the cells were cloned with the limited dilution method continually until a monoclonal cell strain whose positive rate of Bcl-2 protein was 100% detected by flow cytometry was obtained. The killing rates of cells were detected by Methabenzthiazuron assay after the treatment of 6% ethanol for 6?h. The extent of apoptosis was analyzed by transferase-mediated dUTP nick end labeling (TUNEL) staining and flow cytometry.Results Most of the pDOR-SB-transfected cells demonstrated Bcl-2 positive signals, while no signal was found in the controls. The positive rate of Bcl-2 protein detected by flow cytometry in the obtained monoclonal cell strain, which was named HCC-bcl2, was 100% after the cells had been cloned 3 times continually. The killing rate, TUNEL index and the scale of sub-G1 apoptotic peak in HCC-bcl2 cells were all significantly lower than those in the control cells.Conclusion Overexpression of Bcl-2 protein suppresses ethanol-induced apoptosis of the HCC cell line HCC-9204.

Objective　To investigate the effect of trans-acting factor(s) on rat glutathione S-transferase P1 gene (rGSTP1) transcription regulation in tumor cells.Methods　The binding of trans-acting factor(s) to two enhancers of the rGSTP1 gene, glutathione S-transferase P enhancer Ⅰ (GPEI) and glutathione S-transferase P enhancer Ⅱ-1 (GPEⅡ-1), was identified by an electrophoretic mobility shift assay (EMSA). The molecular weight of trans-acting factor was measured in a UV cross-linking experiment. Results　Trans-acting factor interacting with the core sequence of GPEI (cGPEI) were found in human cervical adenocarcinoma cell line (HeLa) and rat hepatoma cell line (CBRH7919). These proteins were not expressed in normal rat liver. Although specific binding proteins that bound to GPEⅡ-1 were detected in all three cell types, a 64 kDa binding protein that exists in HeLa and CBRH7919 cells was absent in normal rat liver. Conclusion　cGPEI, GPEII specific binding proteins expressed in HeLa and CBRH7919 cells may play an important role in the high transcriptional level of the rGSTP1 gene in tumor cells.

Objective　Protein induced by vitamin Kabsence or antagonistⅡ（PIVKAⅡ），also called des－gamma－carboxy prothro mbin（DCP），is a sensitive marker for the diagnosis of hepatocellular carcinoma（HCC）inJapan and the United States since the sensitive kits were available（1998）．PIVKAⅡis not usedin clinicaldiagnosis in China so far．The aimofthis study was to assess the diagnostic value of PIVKAⅡin Chinesepatients with HCC．Methods　Seru mPIVKAⅡandα－fetoprotein（AFP）levels were determined in 60 patients with HCCand 30patients with cirrhosis not carrying HCC．Results　The mean serumconcentration of PIVKAⅡin HCCpatients（784．3±1364．1，x±s）was higherthan thatin cirrhosis patients（16．1±31．7）；this difference was highly significant（P＜0．0001）．When thecutofflevelof 40 mAU／ml was used as the level of discriminating HCCfromcirrhosis，51．7％of patients（31／60）with HCChad PIVKAⅡvalues above this level（sensitivity）．Only 4 patients with cirrhosis hadsuch high PIVKAⅡlevels．Thus，the specificity of this test was 86．7％（26／30）．Total accuracy was62．2％［（31＋26）／（60＋30）］．Seven of 19 small HCCs（36．84％）had PIVKAⅡvalues above the cutofflevel．Concentrations of AFPabove 20 ng／ml were observed in 34 of60 patients with HCC（56．7％）and in11 patients with cirrhosis（36．7％）．Eleven of 26 patients with HCC（46．2％）withoutincreased AFPhadconcentrations of PIVKAⅡgreater than 40 mAU／ml．No significant correlation was found between serumlevels of AFPand PIVKAⅡthat were measured in 60 HCCpatients（rs＝0．101，P＝0．247）．Combiningthe information fromPIVKAⅡand AFPshowed anincrease of approximately 21．6％over AFPand 26．7％over PIVKAⅡalone．For small HCCpatients，combining the information from PIVKAⅡand AFPshowedan increase of approximately 15．8％over AFPalone and 21．1％over PIVKAⅡalone．Conclusion　PIVKAⅡis a useful early diagnostic marker for HCCand may be more sensitive whencombined with AFPin Chinese patients．

Objective　To investigate the distribution and clonality of T cell receptor (TCR) Vβ repertoire in patients with acute monoblastic leukemia (AML-M5).Methods　Expression of the TCR Vβ repertoire was analyzed using reverse transcription-polymerase chain reaction (RT-PCR), which amplified the complementarity determining region 3 of 24 TCR Vβ genes in peripheral blood from 9 cases with acute myclogenous leukemia subtype 5 or acute monoblastic leukemia (AML-M5). PCR products were further studied by genescan analysis to identify T cell clonality. Results　Expression of 1-10 Vβ subfamilies was found in samples from 9 patients. Genescan analysis showed that some Vβ subfamily products from 8 of 9 cases contained an oligoclonal peak. Oligoclonal T cells of the Vβ2 subfamily could be found in 6 patients with AML-M5. Conclusions　T cell clonality expansion was found in AML-M5 cases and were tendentious in the Vβ2 subfamily, suggesting a the specific immune response for leukemia cell (M5) associated antigen and may display antileukemia activity.

Objective　To assess the effect of tetrandrine (Tet) pulmonary targeting microspheres on hypoxic pulmonary hypertension and evaluate its selective action on pulmonary circulation.Methods　Twenty rats were exposed to hypoxic conditions for 3 weeks. Ten rats were used as normoxic controls. We administered Tet pulmonary targeting microspheres to 10 hypoxic rats and Tet aqueous solution to 10 hypoxic rats and the 10 control rats. Mean pulmonary arterial pressure (mPAP) was measured by a right cardiac catheterization, and mean systemic blood pressure (mSBP) was measured by left femoral catheterization. Results　Rats exposed to hypoxia developed pulmonary hypertension. The decrease in mPAP in rats treated with Tet pulmonary targeting microspheres was significantly greater than that in rats receiving Tet aqueous solution (P<0.05), and the effects were longer with Tet pulmonary targeting microspheres. Moreover, Tet pulmonary targeting microspheres, unlike Tet aqueous solution, did not decrease mSBP.Conclusion　Tet pulmonary targeting microspheres were more effective than Tet aqueous solution in treating hypoxic pulmonary hypertension and acted selectively on the pulmonary circulation.

Objectives To study the difference between gene expressions of high (H0-8910PM) and low (HO-8910) metastatic human ovarian carcinoma cell lines and screen novel associated genes by cDNA microarray. Methods cDNA retro-transcribed from equal quantities of mRNA derived from high and low metastatic tumor cells or normal ovarian tissues were labeled with Cy5 and Cy3 fluorescein as probes. The mixed probe was hybridized with two pieces of BioDoor 4096 double dot human whole gene chip and scanned with a ScanArray 3000 laser scanner. The acquired image was analyzed by ImaGene 3.0 software. Results A total of 355 genes with expression levels more than 3 times larger were found by comparing the HO-8910 cell with normal ovarian epithelial cells. A total of 323 genes with expression levels more than 3 times larger in HO-8910PM cells compared to normal ovarian epithelium cells were also detected. A total of 165 genes whose expression levels were more than two times those of HO-8910PM cells compared to their mother cell line (HO-8910) were detected. Twenty-one genes with expression levels >3 times were found from comparison of these two tumor cell lines.Conclusions cDNA microarray techniques are effective in screening differential gene expression between two human ovarian cancer cell lines (H0-8910PM; HO-8910) and normal ovarian epithelial cells. These genes may be related to the genesis and development of ovarian carcinoma. Analysis of the human ovarian cancer gene expression profile with cDNA microarray may help in gene diagnosis, treatment and prevention.

Objective　To confirm the genetic relation between Giardia lamblia (G. lamblia) isolates from different geographic regions of China and other countries.Methods　Genomic DNA were extracted from the trophozoites or cysts of Giardia lamblia. The triose phosphate isomerase (tim) gene was amplified using polymerase chain reaction (PCR) technique. PCR products were digested with endonuclease and sequenced. The data of sequencing were analyzed with the DNAstar software and compared with that of the isolates acquired from GenBank.Results　Of nine isolates of Giardia lamblia from China (C1, C2, CH2 and CH3), Cambodia (CAM), Australia (A1 and A2) and America (BP and CDC), respectively, 3 (A1, A2 and CAM) fit into Group 1 (WB), 2 (CH2 and CH3)) into Group 2, and 4 (C1, C2, BP and CDC) into Group 3 (GS). The results confirmed the genetic relatedness of G. lamblia isolates from all over the world.Conclusion　Genotyping isolates of G. Lamblia provides important information for establishing the phylogenetic relationship or for the epidemiological evaluation of the spreading of this organism.

Objective　To evaluate the efficacy and the mechanism of application of selective head cooling on neuronal morphological damage during postischemic reperfusion in a rabbit model.Methods　168 New Zealand rabbits were randomized into three groups. Group Ⅰ ［n=24, (38±0.5)℃, non-ischemic control］; Group Ⅱ ［n=72, (38±0.5)℃, normothermic reperfusion］; Group Ⅲ ［n=72, (28±0.5)℃, selective head cooling, initiated at the beginning of reperfusion). Animals in three subgroups (n=24, each) of Group Ⅱ and Group Ⅲ had reperfused lasting for 30, 180 and 360 min respectively. Using computerized image analysis technique on morphological changes of nucleus, the degree of neuronal damage in 12 regions were differentiated into type A (normal), type B (mild damaged), type C (severely damaged) and type D (necrotic). Fourteen biochemical parameters in brain tissues were measured.Results　As compared with Group Ⅰ, the counts of type A neuron decreased progressively, and those of type B, C and D increased significantly in Group Ⅱ during reperfusion (P<0.01). In Group Ⅱ, vasoactive intestinal peptide, b-endorphine, prostacyclin, T3 and Na+, K+-ATPase were correlated with the changes of type A; b-endorphine and thromboxane with type B; glucose and vasopressin with type C; Na+, K+-ATPase, glutamic acid, T3 and vasoactive intestinal peptide with type D (P<0.05). As compared with Group Ⅱ, the counts of type A increased, and those of type C and D significantly decreased in Group Ⅲ (P<0.01). In Group Ⅲ, Ca2+, Mg2+-ATPase were correlated with the changes of type A, C and D (P<0.01). Conclusion　Selective head cooling for sex hours during postischemic reperfusion does improve neuronal morphological outcomes in terms of morphological changes.

Objective To evaluate the immediate and follow-up results of percutaneous transluminal septal myocardial ablation (PTSMA) in patients with hypertrophic obstructive cardiomyopathy (HOCM). Methods Fifteen symptomatic, drug-refractory patients with HOCM underwent PTSMA procedures with application of a myocardial contrast echocardiography (MCE) intra-procedure. Before and after the procedure, clinical evaluations were obtained in all patients, who were followed up for a mean period of 8.6±3.8 (6-20) months.Results Immediate left ventricular outflow tract gradient (LVOTG) reduction was achieved (77.93±22?mm?Hg vs 14.8±15?mm?Hg, P<0.0001) after the procedure with a mean decrease of 5.75±2.87?mm?Hg of left ventricular end diastolic pressure (P<0.001). Follow up results revealed that ventricular remodelling occurred mainly 1-3 months after the procedure, but without evidence of ventricular dilation and contract dysfunction. Heart function (NYHA) was greatly improved (3.4±0.5 vs 1.1±0.4, P<0.001) and exercise endurance increased. A renewed increase of LVOTG was found in 2 patients during follow-up. Conclusions LVOTG was greatly decreased in HOCM patients undergoing a PTSMA procedure, and their symptoms were greatly improved without cardiac complications during follow-up. Sub-selection and re-opening of target vessels were the causes of renewed increase of LVOTG, and this can be avoided with the accumulation of experience. This is a promising method for the treatment of symptomatic patients with HOCM.

Objective To investigate the relationship between P16 gene (the tumor suppressor gene) and the bcl-2 gene (the apoptosis inhibitor gene) and the incidence and development of malignant eyelid tumors. Methods The streptavidin-biotin-peroxidase complex immunohistochemistry method was used to study the expression of P16 gene and the bcl-2 gene in 96 cases of malignant eyelid tumors. Results Among the 96 cases, there were 40 basal cell carcinomas (BCCs), 33 squamous carcinomas and 23 sebaceous carcinoma, with P16 protein positive (nuclear staining) rates 70%, 54.6% and 56.5%, respectively. The P16 positive rate was negatively correlated with the degree of tumor histological differentiation, and the rate difference between the high differentiated carcinomas was significant (P<0.05). Positive Bcl-2 protein expression was detected in the cytoplasm. All 40 BCC cases were Bcl-2 positive, and nearly all of the tumor cells showed positive cytoplasmic expression, while in the 33 squamous cell carcinoma cases only one showed positive focal reaction, and the staining in the other 32 cases was relatively faint. None of the 23 sebaceous carcinomas expressed Bcl-2. Conclusions The expression of the P16 protein was related to the occurrence and degree of differentiation of malignant eyelid tumors. The overexpression of the Bcl-2 protein suggests that suppression of apoptosis might play a role in the tumorigenesis of BCC.

Objectives To compare the difference in pulmonary function between Caucasians and Chinese and assess the best reference values of pulmonary function for Chinese adults. Methods Values for pulmonary function were predicted by tests on 4773 Chinese healthy subjects (male: 2560, female: 2213, aged 15-78 years) in six parts (north, northeast, northwest, east, southwest and south) of China. Prediction equations of the European Community for Steel and Coal (ECSC), other equations for overseas Chinese or for Caucasians were also selected. The regression coefficients of forced vital capacity (FVC), forced expiratory volume in one second (FEV1), FEV1/FVC, total lung capacity (TLC) and residual volume (RV) were summarized. Results ECSC predictions were closer to the Chinese ones than other selected equations. Comparison with ECSC predictions showed that on average the values for FVC, FEV1 were 5.3% smaller in Chinese males and 3.3% smaller in Chinese females, with the maximal differences in south China and the minimal differences in North China. RV and TLC in Chinese were lower than in Caucasians (males 4.8%, 5.5%, respectively; females 8.7% and 6.0%, respectively). Conversion factors were given for adjusting ECSC equations to fit Chinese. Conclusions For predicting values of pulmonary function in Chinese, we suggest to use the equations reported here. Alternatively, ECSC regression equations may be used with appropriate conversion factors.

Objective To evaluate the role of outer membrane protein (Omp) F-deficiency and active efflux in the accumulation of hydrophilic fluoroquinolones ciprofloxacin (CPLX) and lomefloxacin (LMLX) in resistant E. coli strains. Methods Fluoroquinolone accumulation in bacteria and the effect of active efflux were measured by a fluorescence method. The outer membrane proteins of the bacteria were analysed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). E. coli strains in this study included control strains JF701 and JF703 that are OmpC- or OmpF-deficient mutants of E. coli K-12, respectively, and the fluoroquinolone susceptible strain the fluoroquinolone susceptible strain of Escherichia coli (Ecs) and its in vitroselected resistant strains R2 and R256, and the clinical resistant isolates R5 and R6. Results The steady-state accumulation concentration of each drug in Ecs appeared to be the same as in JF701, while in the OmpF- deficient strain JF703, it was 1/5 CPLX or 1/2 LMLX lower than that in JF701, but JF703 was still susceptible to fluoroquinolones. On the other hand, compared with susceptible strains, a 2- to 10-fold decrease in the accumulation of each drug was found in the resistant strains except R2, in which the accumulation was slightly higher than in JF703. After the addition of 2,4-dinitrophenol (DNP), accumulation of each drug increased, especially in resistant strains, indicating that the function of the active efflux (pump) system in these bacteria had been enhanced dramatically. Furthermore, both OmpF and OmpC in Ecs, OmpF-deficiency in R2 and R256 and OmpC-deficiency in R5 and R6 were observed.Conclusion The decreased accumulation of hydrophilic fluoroquinolones in E. coli involved OmpF-deficiency and active efflux (pump), and the latter may be an important factor.

Objective To evaluate radial artery (RA) and internal mammary artery (IMA) grafts in coronary artery bypass and the use of color Doppler ultrasound in the peri-operative evaluation of IMA and radial-ulnar collateral circulation.Methods From June 1998 to June 2000, sixty cases of coronary bypass revascularization with RA and IMA were performed. Preoperatively, the radial-ulnar collateral circulation was evaluated with the modified Allen's test, color Doppler ultrasound and noninvasive oxygen saturation measurement. The IMA lumen and blood flow were measured at the first intercostal space with color Doppler ultrasound preoperatively and postoperatively.Results One patient (1.7%) died of serious cardiac arrhythmia on the fourth postoperative day. There were no arterial graft harvest related complications. Before harvesting, the ulnar artery blood flow was 30.78±9.71?ml/min, and it increased to 43.36±13.98?ml/min (40.87% increase, P<0.01) after the operation. Compared with the baseline, there was no obvious change of IMA blood flow postoperatively (P>0.05), but the systolic/diastolic flow ratio markedly decreased from 8.57±3.98?ml/min to 3.41±4.87?ml/min (P<0.01).Conclusions Arterial grafts can be safely used for coronary bypass revascularization with good results. The ulnar artery blood flow can increase compensatively after RA harvesting. The diastolic blood flow of grafted IMA markedly increased postoperatively. Color Doppler ultrasound was very helpful both in evaluating the radial-ulnar collateral circulation before RA harvesting and in assessing the patency of the grafted IMA after coronary artery bypass grafting (CABG).

Objective　To document morphological changes in hepatic microcirculation in liver tissue with hepatitis B and the pathogenesis of hepatic microcirculatory disturbances.Methods　Liver tissue samples were obtained from patients with hepatitis B by liver biopsy. These samples were examined with a light microscope and transmission electron microscope. Results　Hepatic microcirculatory disturbances existed in patients with hepatitis B, including those with normal liver function, manifested by red blood cell aggregation in sinusoids seen under light microscope and sinusoidal capillarization seen under electron microscope. Weibel-Palade bodies in sinusoidal endothelial cells were seen in 26 out of 53 cases. Intimate contacts were found between lymphocyte/Kupffer cells and sinusoidal endothelial cells. Conclusions　Hepatic microcirculatory disturbances exist in patients with hepatitis B .The appearance of Weibel-Palade bodies in sinusoidal endothelial cells may be a key step in the development of hepatic microcirculatory disturbances.

Objectives　To evaluate the relationship between microdeletion or mutation on the Y chromosome and Chinese patients with idiopathic azoospermia and severe oligozoospermia and to establish a molecular detection method.Methods　Microdeletion or mutation detection at the AZFa (sY84 and USP9Y), AZFb, AZFc/DAZ and SRY regions of the Y chromosome. Seventy-three azoospermia and 28 severe oligozoospermia patients were evaluated using PCR and PCR-SSCP techniques.Results　Twelve of 101 patients (12%) with the AZFc/DAZ microdeletion were found, including 8 with azoospermia (11%) and 4 with severe oligozoospermia (14.3%), and 1 patient had a AZFb and AZFc/DAZ double deletion. No deletions in the AZFa or SRY regions were found. No deletions in AZFa, AZFb, AZFc/DAZ or SRY regions were found in 60 normal men who had produced one or more children.Conclusions　Microdeletion on the Y chromosome, especially at its AZFc/DAZ regions, may be a major cause of azoospermia and severe oligozoospermia leading to male infertility in China. It is recommended that patients have genetic counseling and microdeletion detection on the Y chromosome before intracytoplasmic sperm injection.

Objective To evaluate the down stream involvement of the bile duct in hepatolithiasis.Methods Mechanical damage to bile duct epithelia and long standing cholangitis as result of hepatolithiasis play an important role in the carcinogenesis of bile duct epithelia and stricture of the intra- and extra-hepatic bile duct. Macromorphological and microscopic changes in bile duct mucosa of 100 consecutive patients with hepatolithiasis were investigated using intra- or post-operative cholangioscopy. Biopsy specimens of lesions obtained during cholangioscopy were studied with immunohistochemical staining and flow cytometry to determine proliferative activity and DNA content. Five cases of well-proven cholangiocarcinoma were simultaneously studied as controls.Results Of the 100 patients, those with chronic cholangitis accounted for 86% (86/100), proliferative lesions 11% (11/100), adenomatous polyps 1% (1/100), and adenocarcinoma 2% (2/100). The obvious mucosal lesion associated with hepatolithiasis was located down-stream of the bile duct, predominantly in the hilar region, e.g. orifices of the right/left hepatic duct and common hepatic duct (73% mucosa lesions in the hilar region). The intensity of cancer embryonic antigen stain and the proliferative cell nuclear antigen index increased with the development of bile duct lesions. Aneuploid DNA presented mainly in the high degree malignant adenocarcinomas (>80% of cases).Conclusions The obvious mucosal lesions associated with hepatolithiasis were located down-stream of the bile duct, predominantly in the hilar region (73% of mucosal lesions). The proliferative activity of examined bile duct mucosa lesions increased with the development of pathological deterioration, which may contribute to the development of hilar bile duct stricture and hilar cholangiocarcinoma.Department of Surgery (Lu SC, Yan LN, Rao LQ, Xia T and Gou JL), Department of Pathology (Zhang SY), Biotherapy Center of Oncological Department (Lei S), West China Hospital, Sichuan University, Chengdu 610041, China

Objective　To evaluate the changes in near infrared spectroscopy (NIRS) absorptive properties induced by cerebral ischemia.Methods　A dual wavelength (760?nm and 850?nm) NIRS system measuring total hemoglobin concentration changes was constructed. Twenty Sprague-Dawley rats were used to set up middle cerebral artery occlusion (MCAO) model that had a stable ischemia focus on the cortex. We used NIRS to localize the ischemia focus that was confirmed with magnetic resonance imaging (MRI) and triphenyltetrazonlium chloride brain staining. The cortical ischemia area and the geometric configuration of the NIRS topograms were compared with those from MRI and the anatomical samples for the same rat.Results　The reconstructed NIRS topograms showed that there was an optical density decreased area on the left cortex of the rats with MCAO model. The mean ischemia area as shown in NIRS images was 19.50?mm2 (19.50±0.35?mm2, n=20). Anatomical samples showed that the mean ischemic area located in the NIRS measurement area was 18.46?mm2　 (18.46±0.38?mm2, n=20). For MRI, the mean ischemia area located in the NIRS measurement area was 20.71?mm2 　(20.71±0.27?mm2, n=20). There were no significant differences among NIRS, MRI and anatomical samples (F(2,57)=2.47, P>0.05) for defining the ischemia area. The results showed that there was a significant correlation among NIRS, MRI (r=0.782, P<0.05) and anatomical sample (r=0.851, P<0.05) for the same cortical ischemia area. Meanwhile, the location and geometric configuration of the ischemia focus shown by NIRS were identified by MRI and anatomical samples, though the NIRS images had worse spatial resolution.Conclusions　NIRS showed a good agreement with MRI and anatomical samples in the ischemic area and location determination of the infarction focus for the rat MCAO model. The study suggests that NIRS can non-invasively trace cortical hemodynamic changes induced by ischemia in real time.

Objective To determine whether interleukin-1α and 1β gene polymorphism is associated with rheumatoid arthritis disease activity and bone mineral metabolism, and whether there is any relationship between IL-1β and rheumatoid arthritis (RA) motif gene. Methods IL-1 gene polymorphisms were analyzed in 65 RA patients who met American College of Radiology (ACR) criteria and 60 controls. From genomic DNA, 2 polymorphisms in each gene for IL1α-889 and IL-1β+3953 were typed by PCR-RFLP and HLA-DRB1 allele typing was also undertaken by PCR-SSOP. Some clinical and laboratory parameters were collected. The allelic frequencies and carriage rates were compared between RA patients and controls and between patients with active and quiescent disease. Comparison was also made between IL-1 polymorphism and parameters of bone mineral metabolism and between patients with the HLA-DRB1 RA motif plus IL-1β2 and patients without the two alleles. Fisher test and the analysis of variance was used to analyze the data.Results There was no significant difference in the frequency and carriage rate of IL-1α polymorphisms between RA patients and the controls. The β2/2 genotype of IL-1β was more common in female RA patients compared with controls (P=0.001). A lower carriage rate of IL-1β2 occurred in male RA patients (P=0.001). A higher carriage rate of IL-1α2 is associated with a higher ESR (P=0.008), HAQ score (P=0.03), and vit-D3 (P<0.001), but conversely a lower SJC (p=0.002), a lower RF (P=0.002) and a lower BMD at the lumbar spine (P=0.001). A higher frequency of IL-1α1 is associated with a lower CRP value (P=0.009). An increased IL-1β2 carriage is associated with active rheumatoid disease as indicated by a higher CRP (P<0.001), ESR (P<0.001) and pain score (P=0.001) and a higher BMD at the lumbar spine (P=0.007), lower vit-D3 and. Udpd/Crea level The presence of the HLA DRB1 RA motif and IL-1β allele 2 at same time did not contribute to disease activity.Conclution Polymorphisms of the IL-β gene may affect the RA occurrence. Carriage of IL-1β2 polymorphisms is associated with more active disease in RA and the presence of both the IL-1α2 and the IL-1β1 allele in RA influences bone resorption.

Objective To explore prospective diagnostic criteria for preleukemia.Methods A case control study was done comparing the discrepancies on clinical and laboratory features between patients with preleukemia and those with chronic aplastic anemia (CAA) or atypical paroxysmal nocturnal hemoglubinuria (a-PNH).Results There were eight variables of significance: (1) lymphocytoid micromegakaryocytes in the bone marrow; (2) immature granulocytes in the peripheral blood; (3) ≥2.0% myeloblasts in the bone marrow; (4) positive periodic acid schiff (PAS) stained nucleated erythrocytes; (5) myeloid differentiation index ≥1.8; (6) typical colonal karyotypic abnormalities; (7) negative sister chromatid differentiation; (8) cluster/colony ratio of granulocyte-macrophage colony-forming units (CFU-GM)>4.0. The following criteria were assigned: A: to meet variable one and at least two of the other seven variables and B: to meet at least four of the eight variables. All of the patients with preleukemia met either A or B and none of the patients with CAA or a-PNH did. Conclusions Preleukemia is different from CAA or a-PNH. It has its own clinical and laboratory features, which may be useful for its prospective diagnosis.

Objective　To investigate the effect of D-AP5 (D-2-amino-5-phosphonopentanoate, a specific NMDA-antagonist) on the increase of intracellular free Ca2+ concentration (［Ca2+］i) induced by glutamate in isolated cochlear inner hair cells (IHCs), and to detect the autoreceptors of the IHC membrane. Methods　When a laser scanning confocal microscope (LSCM) was used, the exogenous glutamate (Glu)-induced changes in ［Ca2+］i of isolated IHCs and OHCs of guinea pig cochlea were observed with fluo-3, a fluorescent probe for ［Ca2+］i. After D-AP5 or CNQX (6--cyano--7--nitroguinoxaline--2, 3--dione, a specific AMPA- antagonist) was administrated, the exogenous glutamate (Glu)-induced changes in ［Ca2+］i of isolated IHCs were recorded. Results　In the presence of a low concentration Glu (3.85?μmol/L), there was an increase of ［Ca2+］i in IHCs, whereas there was no change in OHCs. When 50?μmol/L of D-AP5 was administrated in advance, Glu did not induce a corresponding increase in ［Ca2+］i in IHCs, and 50?μmol/L of CNQX did not completely block the increase of ［Ca2+］i in IHCs. Conclusions　These results suggest that the autoreceptors existing in the IHC membrane are mainly of NMDA type, while there are relatively few AMPA receptors. Exogenous Glu is capable of increasing ［Ca2+］i in IHCs by acting on the NMDA autoreceptor of IHCs in a positive feedback manner.

Objective　To test the immunosuppressive effect of cyclosporine (Cs) in a polymer placed in the anterior chamber of corneal allograft recipients. Methods　Wistar inbred rats with vascularized corneas were recipients of corneal allografts from Sprague-Dawley donor rats. Rats underwent penetrating keratoplasty and were divided randomly into four groups: untreated control animals (UCA); Cs-polymer anterior chamber recipients (CPA); co-polymer subconjunctival recipients (CPS); and Cs-olive oil drop recipients (COO). Grafts were examined by slit lamp every 3 days and clinical conditions were scored. Cs concentration in the aqueous humor was assayed at 1, 2, and 4 weeks. At 1, 2 and 4 weeks after transplantation, the operated eyes were collected for histopathological evaluation of the grafts.Results　The median survival time of the allografts was 8.2±1.48 days for the UCA group, 11.4±2.50 days for the CPS group, and 17.0±2.00 days for the CPA group. There was a statistically significant difference (P<0.05) between survival time of the allografts in the animals of the CPA group compared to the other groups of graft recipients. Significantly higher concentrations of Cs were found in the eyes given an anterior chamber implant of Cs-polymer, compared to other treatment groups or untreated rats. A transient inflammatory response in the anterior chamber was observed in the CPA group. Conclusions　Cs-polymer placed in the anterior chamber significantly prolongs corneal allograft survival time in a high risk corneal graft rejection model. This intraocular delivery system may be a valuable adjunct for the suppression of immune graft rejection.

Objective　To identify the prevalence, incidence and trend of urinary incontinence in the elderly over a two-year period.Methods　We randomly selected 4187 elderly people using the State Electoral Data Base in South Australia. Questionnaire surveys were conducted in 1992 with a response rate of 53.4%. The following two surveys were conducted 12 and 24 months later. A total of 2087 elderly people completed all three surveys.Results　Urge incontinence had a higher prevalence rate, with 36.6% to 41.6% of the elderly having at least an occasional problem (with 7.5% to 9.6% having problems often), compared to 23.4% to 28.8% having at least an occasional problem with stress incontinence (with 3.3% to 5.0% having problems often). Combined incontinence and urge incontinence were more common than stress incontinence alone. The incidence rates of urge incontinence and stress incontinence were 19.8% (with 3.1% often) and 14.5% (with 1.9% often), respectively. The trends toward incontinence during the two-year period indicated increasing trends in the elderly.Conclusion　Urinary incontinence is an important health problem in the elderly and our data show that it has an increasing trend for future.

Objective　To evaluate the roles of Na+/H+ exchanger-1 （NHE-1）in the proliferation and apoptosis of pulmonary artery smooth muscle cells in rats. Methods　Twenty Wistar rats were randomized into control group and 3-week hypoxic group. Intracellular pH (pHi) of the smooth muscle was determined with fluorescence measurement of the pH-sensitive dye BCECF-AM， and the expression of NHE-1 mRNA was detected by reverse transcription polymerase chain reaction (RT-PCR). Primary culture of pulmonary artery smooth muscle cells in vitro was performed. In situ cell death detection kit (TUNEL) was used for studying the effect of specific NHE-1 inhibitor-dimethyl amiloride (DMA) on the apoptosis of muscle cells which had intracellular acidification. Results　pHi value and NHE-1 mRNA expression of pulmonary artery smooth muscle cells were significantly higher in the hypoxic group than in the control group (P<0.01, P<0.001). DMA elevated the apoptotic ratio remarkably. The effect was enhanced when DMA concentration increased and the time prolonged.Conclusions　With the function of adjusting pHi, NHE-1 may play an important role in the proliferation and apoptosis of pulmonary artery smooth muscle cells.

Objective　To evaluate the safety and effectiveness of direct gastroscopy for detecting gastric cancer.Methods　Clinical screening by direct gastroscopy was performed for gastric cancer (GC) from September 1985 to July 1998. 3048 elderly people were screened. Their age ranged from 60 to 93 years, and 2034 of the 3084 were followed up.Results　Ninety-two patients with gastric cancer were discovered by gastroscopy, representing 3.02% of the screened population. The rate of early gastric cancer (EGC) was 63.04% (58/92) of all gastric cancers detected. The rate was up to 79.59% (39/49) on follow-up, and was 74.14% (43/51) in asymptomatic patients with gastric cancer. The excision rate was 88.89% for patients with gastric cancer, and 100% for patients with early gastric cancer. The 5-year survival rate was 91.89% for patients with gastric cancer, and 96.30% for patients with early gastric cancer. Conclusion　Clinical screening and follow-up by direct gastroscopy in persons over 60 years of age are a safe and effective method for raising the 5-year survival and detection rate of gastric cancer, especially early gastric cancer.

Objective　To study the efficacy and safety of cutting balloons in coronary interventions.Methods　Twenty-two patients with 23 narrowed coronary arteries and 25 lesions enrolled the study, 17 of whom had 18 diseased coronary arteries and 18 lesions were in-stent restenosis. The average time from previous stenting was 7.60±3.53 months. The lesions were dilated with cutting balloons. Results　All of the lesions were dilated successfully by 5.16±2.30 inflations of a cutting balloon. The mean total duration of balloon inflation was 233.96±94.83 seconds at pressures up to 9.40±1.96 bars. The severity of vascular stenosis was lessened substantially (89.64±8.65% vs 17.60±17.15%， P=0.000) without severe complications. Three restenotic lesions were further dilated with conventional balloons, another one was stented again because of a dissection distal to the previous stent. Three primary lesions were stented for dissection or residual stenosis. Angina pectoris reoccurred in two patients in a mean follow-up period of 7.42±6.87 (range 0.5-20) months. Conclusion　Cutting balloon dilation is an effective and safe choice in interventions for coronary disease especially for in-stent restenosis.

Objective　To evaluate postoperative renal function in children with congenital moderate or severe hydronephrosis. Methods　99m Tc-labeled diethylenetriaminepenta-acetic acid scintigraphy was performed in 50 children with unilateral moderate or severe hydronephrosis to determine postoperative renal function. We also analyzed the factors influencing renal function recovery. Results　Average postoperative renal function in 50 cases was 40.62%±10.09%. Among them, 32% of patients had nearly normal renal function and differentiated renal function reached up to 45%. Average preoperative and postoperative renal function in 25 cases was 23.89%±11.65% and 39.33%±8.59% respectively and the increase of renal function was about 15.44%±11.18% (P=0.0003). Renal parenchyma thickness was negatively correlated with postoperative renal function (r=-0.62, P=0.0009). The follow-up period was positively correlated with postoperative renal function (r=0.58, P=0.0026). The patients'age had no correlation with renal function recovery (r=-0.05, P=0.80). Recovery of renal function in hydronephrosis with extrarenal pelvis was greater than that in hydronephrosis with intrarenal pelvis (P=0.016). Conclusions　Postoperative renal function in children with moderate or severe hydronephrosis can recover to normal. Recovery of renal function was more obvious in hydronephrosis with thinner renal parenchyma, longer follow-up period and extrarenal pelvis.

Malignant mixed Müllerian tumors (MMMTs) are highly aggressive neoplasms composed of sarcomatous and carcinomatous elements commonly encountered in the female genital tract and rarely occur in extragenital sites. Most of extragenital MMMTs develop from the peritoneum. To our knowledge, only 25 cases have been described in the literature. Among them, only 2 cases have arised in the major omentum.1 Here we report a case of a 67-year-old female with primary MMMT of the major omentum.

Nasopharyngeal carcinoma (NPC) is a high morbidity tumor of otorhinolaryngology. The usual clinical manifestations are well known by the specialists. But the bizarre and confusing nature of NPC has misled clinicians and contributed to delayed diagnosis. Recently our department treated a case with unusual nasopharyngeal carcinoma.

Purpose　To review evidence-based management of nephropathy in patients with type 2 diabetes.Data sources　A literature search (MEDLINE 1966 to 2000) was performed using the key word “diabetic nephropathy". Relevant book chapters were also reviewed.Study selection　Well-controlled, prospective landmark studies and expert review articles on diabetic nephropathy were selected.Data extraction　Data and conclusions from the selected articles that provide solid evidence to the optimal management of diabetic nephropathy were extracted and interpreted in light of our clinical research experience with many thousands of Hong Kong Chinese patients.Results　Hypertension, long diabetes duration, poor glycaemic control and central obesity are the most important risk factors. Microalbuminuria is a practical marker to predict overt nephropathy in type 2 diabetic patients. Risk factor modification, renal function monitoring and combined therapies are the current integrated approaches to manage patients with diabetic kidney disease. Optimal glycaemic control is the mainstay of treatment but effective antihypertensive therapy is also key to delaying the progression of diabetic nephropathy. Angiotensin-converting enzyme inhibitors and angiotensin Ⅱ receptor antagonists have important renoprotective actions independent of their blood pressure lowering actions. Conclusions　Diabetic nephropathy is the leading cause of end-stage renal disease worldwide. Monitoring renal function and screening for microalbuminuria will allow the identification of patients with nephropathy at a very early stage for intervention. Tight glycaemic control and aggressive antihypertensive treatment as well as the use of renin-angiotensin system inhibitors should substantially delay the progression of nephropathy.

Objective　To evaluate the safety and efficacy of the Amplatzer septal occluder for transcatheter closure in patients with secundum atrial septal defect (ASD Ⅱ).Methods　Patients with clinically confirmed ASD Ⅱ were recommended for transcatheter closure of ASD Ⅱ. Results　30 ASD Ⅱ patients (20 females) underwent transcatheter closure at a median age of 18.4 years (5-55 years). Both the stretched diameters of ASDs and the sizes of the devices were from 18 to 34?mm (25±7?mm). The successful placement rate was 100%. The rest shunt documented by color Doppler, was immediately after implantation in 40% of patients, in 9.9% after 24 hours, and in 3.3% trace at 3 months. No serious complications were observed. There was improvement in symptoms and in cardiac size. Septal motion abnormalities normalized in all patients after 3 months follow-up.Conclusion　The Amplatzer septal occluder is a safe and effective device for transcatheter closure of ASD Ⅱ. Long-term follow-up is still required before widespread clinical use can be recommended.