Use of AffiProbe HPV test kit for detection of human papillomavirus DNA in genital scrapes.

The presence of human papillomavirus (HPV) DNA in cervical and vaginal scrapes was analyzed by the AffiProbe HPV test kit (Orion Corp., Orion Pharmaceutica, Helsinki, Finland), which is a 1-day solution hybridization test for HPV type 6/11, 16, or 18. The AffiProbe test was compared with a commercially available dot blot test (ViraPap and ViraType tests; Life Technologies Inc., Gaithersburg, Md.). The study group consisted of 178 patients seen in a gynecological outpatient clinic. Altogether, 64 specimens (36 cervical and 28 vaginal scrapes) from 49 patients were positive by the AffiProbe test. Concurrently collected cervical scrapes from 174 patients were available for the reference test, which yielded 27 positive results for HPV type 6/11 or 16/18 and 25 positive results for HPV type 31/33/35. Agreement as to the presence of HPV type 6/11, 16, or 18 by the two tests was reached in 85% of the specimens. Eleven cervical specimens were positive by the AffiProbe test only, and nine cervical specimens were positive by the ViraType test only. Independent evidence obtained by the polymerase chain reaction, repeat examination, or the concurrent presence of HPV DNA in vaginal or vulval epithelium supported the AffiProbe and the ViraType test results for 6 of the 11 and 6 of the 9 specimens with discrepant results, respectively. Thus, the DNA tests had similar sensitivities for HPV type 6/11, 16, and 18 DNAs, but the results were obtained within 1 day by the AffiProbe test, whereas results for the ViraPap and ViraType analyses required from 4 days to 2 weeks.     

A role for the basal ganglia in nicotinic modulation of the blink reflex

Summary In humans and rats we found that nicotine transiently modifies the blink reflex. For blinks elicited by stimulation of the supraorbital branch of the trigeminal nerve, nicotine decreased the magnitude of the orbicularis oculi electromyogram (OOemg) and increased the latency of only the long-latency (R2) component. For blinks elicited by electrical stimulation of the cornea, nicotine decreased the magnitude and increased the latency of the single component of OOemg response. Since nicotine modified only one component of the supraorbitally elicited blink reflex, nicotine must act primarily on the central nervous system rather than at the muscle. The effects of nicotine could be caused by direct action on lower brainstem interneurons or indirectly by modulating descending systems impinging on blink interneurons. Since precollicular decerebration eliminated nicotine's effects on the blink reflex, nicotine must act through descending systems. Three lines of evidence suggest that nicotine affects the blink reflex through the basal ganglia by causing dopamine release in the striatum. First, stimulation of the substantia nigra mimicked the effects of nicotine on the blink reflex. Second, haloperidol, a dopamine (D₂) receptor antagonist, blocked the effect of nicotine on the blink reflex. Third, apomorphine, a D₂ receptor agonist, mimicked the effects of nicotine on the blink reflex.     

Bacterial persistence within erythrocytes: a unique pathogenic strategy of Bartonella spp

The genus Bartonella comprises human-specific and zoonotic pathogens responsible for a wide range of clinical manifestations, including Carrion's disease, trench fever, cat scratch disease, bacillary angiomatosis and peliosis, endocarditis and bacteremia. These arthropod-borne pathogens typically parasitise erythrocytes in their mammalian reservoir host(s), resulting in a long-lasting haemotropic infection. We have studied the process of Bartonella erythrocyte parasitism by tracking green fluorescent protein-expressing bacteria in the blood of experimentally infected animals. Following intravenous infection, bacteria colonise a yet enigmatic primary niche, from where they are seeded into the blood stream in regular intervals of approximately five days. Bacteria invade mature erythrocytes, replicate temporarily and persist in this unique intracellular niche for the remaining life span of the infected erythrocytes. A triggered antibody response typically results in an abrogation of bacteremia within 3 months of infection, likely by blocking new waves of bacterial invasion into erythrocytes. The recent establishment of genetic tools for Bartonella spp. permitted us to identify several putative pathogenicity determinants. Application of differential fluorescence induction technology resulted in the isolation of bacterial genes differentially expressed during infection in vitro and in vivo, including an unknown family of autotransporter proteins as well as a novel type IV secretion system homologous to the conjugation system of E. coli plasmid R388. Mutational analysis of a previously described type IV secretion system displaying homology to the virB locus of Agrobacterium tumefaciens provided the first example of an essential pathogenicity locus in Bartonella. Though required for establishing haemotropic infection, it remains to be demonstrated if this type IV secretion system is necessary for colonisation of the primary niche or for the subsequent colonisation of erythrocytes.     

Function-blocking antibodies to human vascular adhesion protein-1: a potential anti-inflammatory therapy

Human vascular adhesion protein-1 (VAP-1) is a homodimeric 170-kDa sialoglycoprotein that is expressed on the surface of endothelial cells and functions as a semicarbazide-sensitive amine oxidase and as an adhesion molecule. Blockade of VAP-1 has been shown to reduce leukocyte adhesion and transmigration in in vivo and in vitro models, suggesting that VAP-1 is a potential target for anti-inflammatory therapy. In this study we have constructed mouse-human chimeric antibodies by genetic engineering in order to circumvent the potential problems involved in using murine antibodies in man. Our chimeric anti-VAP-1 antibodies, which were designed to lack Fc-dependent effector functions, bound specifically to cell surface-expressed recombinant human VAP-1 and recognized VAP-1 in different cell types in tonsil. Furthermore, the chimeric antibodies prevented leukocyte adhesion and transmigration in vitro and in vivo. Hence, these chimeric antibodies have the potential to be used as a new anti-inflammatory therapy.     

Rewarding effects of the optical isomers of 3,4-methylenedioxy-methylamphetamine ('Ecstasy') and 3,4-methylenedioxy-ethylamphetamine ('Eve') measured by conditioned place preference in rats

3,4-methylenedioxy-methylamphetamine (MDMA) (‘Ecstasy’) and its analogue 3,4-methylenedioxy-methylamphetamine (MDE) (‘Eve’) are well known illicit street drugs mainly abused by young people. In spite of the actual research going on, the classification of their abuse potential remains unclear. Since secondary reinforcers are the main factors responsible for craving and relapse, the aim of our study was to assess the potency of MDMA and MDE in a second order reinforcement paradigm, i.e. conditioned place preference (CPP). For the general assessment of our study conditions, we compared MDMA with amphetamine. Unexpectedly, no significant CPP for MDMA was found in contrast to amphetamine. Detailed analysis of current literature led us to the working hypothesis that social environment is crucial for the development of CPP. In a subsequent experiment we tested the influence of housing conditions on CPP using MDMA and demonstrated that isolated animals show significant CPP compared to group-housed ones. In order to better understand the rewarding mechanisms of Ecstasy-derivatives, we tested both the racemic drugs and the pure isomers in the CPP paradigm. Both MDMA's optical isomers and racemic MDMA showed significant CPP without notable differences, while MDE and its isomers completely failed to show any significant CPP. In conclusion, the mechanism by which MDMA induces addiction is much more complicated than assumed so far and more pronounced in isolated animals. The fact that both optical isomers of MDMA led to CPP implies that at least two pathways by which MDMA induces craving behaviour exist.     

Role of gamma delta T cells in inflammatory bowel disease.

gammadelta T cells have previously been shown to play a protective role in various animal models of chronic inflammation (e.g., experimental autoimmune encephalomyelitis, collagen-induced arthritis, and non-obese diabetes). This immunoregulatory potential is exerted by synthesizing various anti-inflammatory cytokines and growth factors (e.g., transforming growth factor-beta). As the normal balance between inflammatory and regulatory cytokines is perturbed in inflammatory bowel disease (IBD) a protective effect of gammadelta T cells seems likely. This notion is supported by our finding of increased mortality of rats with 2,4,6-trinitrobenzene sulfonic acid-induced colitis following gammadelta T cell depletion. In contrast, no effect was observed after depletion of gammadelta T cells in a Crohn's disease animal model with terminal ileitis (TNF(DeltaARE) mice). Therefore, future studies must further define where in the intestinal immune system gammadelta T cells exert their protective function and how this can be used in the treatment of IBD.     

Impaired sodium excretion, decreased glomerular filtration rate and elevated blood pressure in endothelin receptor type B deficient rats

The renal endothelin (ET) system, particularly the ET type B receptor, has been implicated in the regulation of sodium excretion and glomerular filtration rate (GFR). We analyzed kidney morphology and function in a rat strain characterized by complete absence of a functional ETB receptor. Due to Hirschsprung's disease limiting lifetime in these rats, studies were performed in 23-day-old rats. Kidney size and morphology (glomerular and interstitial matrix content, glomerular size and cell density and intrarenal vascular morphology) were normal in ETB-deficient rats. There were also no evidence of altered kidney cell cycle regulation in these rats. GFR was significantly lower, by 72% (P<0.001), in homozygous ETB-deficient rats than in wild-type rats. Fractional sodium excretion was likewise markedly reduced by 84% in homozygous ETB-deficient rats (P<0.001 versus wild-type rats). Treatment with the specific epithelial sodium channel blocker amiloride led to a much higher increase in fractional sodium excretion in ETB-deficient rats (934.2+/-73% in ETB-deficient rats versus 297+/-20% in wild-type rats, expressed as percentage of corresponding placebo treated control; P<0.001). Mean arterial blood pressure was elevated by 7.9 mmHg in homozygous ETB-deficient rats (P<0.05 versus wild-type rats). Our study demonstrates that ETB-deficiency causes early onset kidney dysfunction characterized by a markedly reduced sodium excretion, decreased GFR, and slightly elevated blood pressure. The complete absence of the ETB receptor causes in the kidney--in contrast to the colon--a functional rather than a developmental, neural crest cell dependent disease, since kidney morphology was normal in ETB-deficient rats. The much higher increase in the fractional sodium excretion in ETB-deficient rats after pharmacological blockade of the epithelial sodium channel indicates that the decreased fractional sodium excretion in ETB-deficient rats is most probably due to a lack of the inhibitory property of the ETB receptor on the epithelial sodium channel activity.     

Ontogeny, differentiation and growth of the endocrine pancreas

The pancreas develops from the primitive foregut endoderm, which differentiates into ductal, acinar and endocrine cells. This complex process is probably replicated in the adult pancreas when endocrine cell renewal is required, as may be the case in diabetes mellitus. This review describes what is known about the morphogenesis of the endocrine pancreas during ontogeny and the mechanisms regulating its differentiation and growth. Received: 23 December 1999 / Accepted: 15 February 2000     

Comparative typing of Campylobacter jejuni by heat-stable serotyping and PCR-based restriction fragment length polymorphism analysis

Campylobacter jejuni has become the most common bacterial cause of human gastroenteritis worldwide. Rapid, discriminatory typing methods are required to identify potential clusters of infections. The major disadvantage of the well-evaluated and widely used Penner heat-stable serotyping method is the high level of nontypeability. The correlation of the types determined by the Penner heat-stable serotyping method and PCR-based restriction fragment length polymorphism (RFLP) analysis of the lipooligosaccharide (LOS) biosynthesis genes of C. jejuni was studied with 149 C. jejuni strains. Of these strains, 79 were patient strains belonging to 25 Penner serotypes, 60 were nontypeable patient strains, and 10 were reference strains. A 9.6-kb DNA fragment of the LOS gene cluster was amplified and digested with the restriction enzymes HhaI and DdeI. Altogether, 39 different RFLP types (including 30 HhaI profiles and 32 DdeI profiles) were identified. Type Hh1Dd1 was the most common type, with 36% of the strains and strains of 12 serotypes being of this type. A high level of discrimination was obtained, and a correlation between the Penner serotypes and the PCR-RFLP types could be seen. Also, variation in the LOS biosynthesis genes within a single Penner serotype was found. Although the PCR-RFLP method may not be sufficient to compensate for Penner serotyping, it can give valuable information about nontypeable strains and further characterize strains of common serotypes.