The P-selectin gene is highly polymorphic: reduced frequency of the Pro715 allele carriers in patients with myocardial infarction

P-selectin is an adhesion molecule, expressed at the surface of activated cells, that mediates the interaction of activated endothelial cells or platelets with leukocytes. P-selectin expression is increased in atherosclerotic plaques, and high plasma levels of this molecule have been observed in patients with unstable angina. We investigated the P-selectin gene as a possible candidate for myocardial infarction (MI). The P-selectin gene is situated on chromosome 1q21-q24, spans >50 kb and contains 17 exons. The sequences of the 5'-flanking region and exons of 40 alleles from patients with MI were screened for polymorphisms using polymerase chain reaction/single-strand conformation polymorphism (PCR-SSCP) and sequencing. Thirteen polymorphisms were identified: five in the 5'-flanking and eight in the exonic sequences. Four polymorphisms (Ser290Asn, Asn562Asp, Leu599Val and Thr715Pro) predicted a change in the amino acid sequence of the P- selectin protein. All P-selectin polymorphisms as well as a common E- selectin polymorphism, Ser128Arg which has been reported as being associated with an increased risk of premature coronary heart disease (CHD), and is in tight linkage disequilibrium with several P-selectin polymorphisms, were investigated in 647 patients with MI and 758 control subjects from four regions of France and Northern Ireland (the ECTIM study). The entire set of P-selectin polymorphisms provided a heterozygosity of 91%. The polymorphisms were tightly associated with one another and displayed patterns of linkage disequilibrium suggesting the existence of highly conserved ancestral haplotypes. The five polymorphisms in the 5'-flanking region of the gene were unrelated to MI or any relevant phenotype measured in the ECTIM study. We inferred that the four missense variants identified in the coding region predicted eight common forms of the P-selectin protein. The Pro715 allele which characterizes one of these forms was less frequent in France than in Northern Ireland ( P < 0.002) and in cases than in controls ( P < 0.002; P < 0.02 after correction for the number of tests). We conclude that the P-selectin gene is highly polymorphic and hypothesize that the Pro715 variant may be protective for MI. Whether this variant affects the properties of the P-selectin protein in a way which is compatible with this hypothesis needs to be checked experimentally.      

Glycoproteins present in human follicular fluid that inhibit the zona- binding capacity of spermatozoa

Previous studies have suggested that human follicular fluid contains factors that reduce the zona-binding capacity of spermatozoa. The present study provides further evidence of the existence of such factors. Using the hemizona binding assay (HZA), we have shown that the inhibitory effect of human follicular fluid on the zona-binding capacity of spermatozoa is concentration-dependent, an inhibitory effect being detected when the concentration of human follicular fluid was > or = 10%. A 1% concentration of human follicular fluid did not possess this inhibitory activity. Heating human follicular fluid at 56 degrees C for 30 min did not affect its inhibitory properties; treatment with proteinase-K abolished such inhibition. Human follicular fluid was fractionated sequentially by concanavalin-A affinity chromatography, Mono Q ion-exchange chromatography and Superose-12 gel filtration. The zona binding inhibitory activity resided in the fraction which bound to the lectin and Mono Q column and contained molecules with native molecular weights of 32 and 192 kDa. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis analysis suggested that the 192 kDa glycoprotein was a tetramer, while the 32 kDa glycoprotein remained as a single molecular species under denaturing conditions. We conclude that two glycoproteins were responsible for the zona binding inhibitory activity of human follicular fluid. The physiological role of these factors remains unclear.      

The use of laparoscopic ovarian electrocautery in preventing cancellation of in-vitro fertilization treatment cycles due to risk of ovarian hyperstimulation syndrome in women with polycystic ovaries

Fifty women with polycystic ovaries took part in a prospective randomized study. All women required treatment by in-vitro fertilization (IVF) for reasons other than anovulation. They had all previously undergone ovarian stimulation with gonadotrophin therapy which had failed to result in pregnancy or had been abandoned due to high risk of developing ovarian hyperstimulation syndrome (OHSS). Twenty-five women were treated by long-term pituitary desensitization followed by gonadotrophin therapy, oocyte retrieval and embryo transfer (group 1). Twenty-five women underwent laparoscopic ovarian electrocautery after pituitary desensitization followed by gonadotrophin therapy, oocyte retrieval and embryo transfer (group 2). A significantly higher number of women in group 1 had to have the treatment cycle abandoned due to impending or actual OHSS, determined by endocrine and clinical findings. In addition, the development of moderate or severe OHSS in completed cycles was higher in group 1. The pregnancy rate and miscarriage rates in the two treatment groups were similar. The authors propose that laparoscopic ovarian electrocautery is a potentially useful treatment for women who have previously had an IVF treatment cycle cancelled due to risk of OHSS or who have suffered OHSS in a previous treatment cycle.      

Epidemiology and molecular characterization of Streptococcus pyogenes recovered from scarlet fever patients in central Taiwan from 1996 to 1999

One hundred seventy-nine Streptococcus pyogenes isolates recovered from scarlet fever patients from 1996 to 1999 in central Taiwan were characterized by emm, Vir, and pulsed-field gel electrophoresis (PFGE) typing methods. The protocols for Vir and PFGE typing were standardized. A database of the DNA fingerprints for the isolates was established. Nine emm or emm-like genes, 19 Vir patterns, and 26 SmaI PFGE patterns were detected among the isolates. Among the three typing methods, PFGE was the most discriminatory. However, it could not completely replace Vir typing because some isolates with identical PFGE patterns could be further differentiated into several Vir patterns. The prevalent emm types were emm4 (n = 81 isolates [45%]), emm12 (n = 64 [36%]), emm1 (n = 14 [8%]), and emm22 (n = 13 [7%]). Some emm type isolates could be further differentiated into several emm-Vir-PFGE genotypes; however, only one genotype in each emm group was usually predominant. DNA from nine isolates was resistant to SmaI digestion. Further PFGE analysis with SgrAI showed that the SmaI digestion-resistant strains could be derived from indigenous strains by horizontal transfer of exogenous genetic material. The emergence of the new strains could have resulted in an increase in scarlet fever cases in central Taiwan since 2000. The emm sequences, Vir, and PFGE pattern database will serve as a basis for information for the long-term evolutionary study of local S. pyogenes strains.     

Human papillomavirus typing with a polymerase chain reaction-based genotyping array compared with type-specific PCR

BACKGROUND: Type-specific persistence of human papillomavirus (HPV) infection can cause invasive cervical cancer. OBJECTIVES: To evaluate the efficacy of HPV detection and typing with a general polymerase chain reaction (PCR)-based genotyping array and to compare it with a type-specific PCR assay. STUDY DESIGN: Four hundred and thirty-three cervical samples were tested with a modified MY11/GP6+ PCR-based reverse-blot assay (EasyChip HPV Blot; King Car, Taiwan [hereafter HPV Blot]) and with 20 genotypes of L1-type-specific PCR (HPV-6, -11, -16, -18, -31, -33, -35, -39, -45, -51, -52, -53, -56, -58, -59, -62, -66, -68, -70, and -71 [CP8061]). RESULTS: The concordance of the two tests in determining HPV positivity was 96.8% (419/433), with a Cohen's kappa=0.93 (95% CI: 0.90-0.97) and McNemar's test of P=1.0, which indicates excellent agreement. The overall concordance of the two tests in the identification of type-specific HPV was 91.0% (394/433). Sensitivity (90-100%), specificity (99.2-100%), and accuracy (98.6-100%) rates of HPV Blot against the gold standard were satisfactory for HPV-16, -18, -58, -33, -52, -39, -45, -31, -51, -70 while HPV-71 (63.6%) had suboptimal sensitivity. Though the kappa values between the two tests for many individual genotypes could not be reliably calculated because of low positivity, the kappa values for HPV-16, -52, and -58 were excellent (0.93, 0.96, and 0.95, respectively). CONCLUSION: The modified MY11/GP6+ PCR-based HPV Blot assay is accurate and sensitive for detection and genotyping of HPV in cervical swab samples.     

Epidemiologic relationship between fluoroquinolone-resistant Salmonella enterica Serovar Choleraesuis strains isolated from humans and pigs in Taiwan (1997 to 2002)

The emergence of ciprofloxacin-resistant Salmonella enterica serovar Choleraesuis in recent years has become an important public health issue in Taiwan. The resistant strains that cause human infections are considered to be from pigs. In this study, we characterized 157 swine and 42 human Salmonella serovar Choleraesuis isolates by pulsed-field gel electrophoresis (PFGE) and drug susceptibility testing to investigate the epidemiologic relationship among the isolates. By PFGE analyses, two major clusters (clusters GA and GB) were identified. Isolates in cluster GA were of both human and swine origins, while those in cluster GB were from pigs only. Among the various genotypes identified, genotype gt-1a was the most prevalent, which was found in 71% (30 of 42) and 48% (76 of 157) of human and swine isolates, respectively. The susceptibility tests for the 106 gt-1a isolates identified 44 susceptibility profiles and showed that 73% of human isolates and 34% of swine isolates were resistant to three fluoroquinolones (ciprofloxacin, enrofloxacin, and norfloxacin). Our findings indicate that a clonal group of Salmonella serovar Choleraesuis may have been circulating in human and swine populations in Taiwan for years and that the fluoroquinolone-resistant Salmonella serovar Choleraesuis strains most likely evolved from a gt-1a clone that emerged in 2000 and that then caused widespread infections in humans and pigs. Nevertheless, it is still debatable whether those Salmonella infections in humans are caused by isolates derived from pigs, on the basis of the higher fluoroquinolone and other antimicrobial resistance percentages in human isolates than in pig isolates.     

Radioimmunoscintigraphy and radioimmunotherapy of renal cell carcinoma xenografts

From a panel of monoclonal antibodies (MAb) reactive with human renal cell carcinoma generated by this laboratory, three (designated A6H, C5H, and D5D) were selected for in vivo studies with a nude mouse xenograft model. These studies included 131I- and 111In-labeled MAb radioimmunoscintigraphy and 131I-labeled MAb radioimmunotherapy. In the imaging studies, these radiolabeled MAb allowed visualization of subcutaneous xenografts larger than 40 mg and subrenal capsule xenografts smaller than 20 mg. Comparisons of tumor to non-tumor tissue radiolabel distribution yielded unusually high ratios and depended on the MAb-xenograft combination. The 111In-radiolabeled A6H showed increased accumulation in the liver compared with 131I-A6H, but this still did not necessitate background subtraction for good visualization of small, subrenal capsule renal cell carcinoma xenografts. Radioimmunotherapy studies with 131I-A6H in BALB/c nu/nu mice bearing established renal cell carcinoma xenografts showed a prolonged (greater than 90 days) regression in tumor burden and possible "cures," whereas three sets of control mice showed progressive and rapid increases in tumor size. These studies indicated that MAb, which show good tissue biodistribution and high imaging sensitivity, could also be capable of delivering effective radiotherapy to the tumor when "human equivalent" radiolabeled-MAb doses are used.