Antibodies generated by a novel DNA vaccination identify the MHC class III encoded BAT2 polypeptide

Recombinant vaccines containing Ii sequences were employed to elicit an antibody response. Gene gun immunisation of mice with the recombinant Ii-antigen-encoding vectors induced antigen-specific antibodies. Antibody levels were substantially elevated when the DNA construct was extended by a sequence encoding the protease inhibitory domain of the invariant chain isoform Ii41. Employing this approach, we raised antibodies specific for a novel MHC class III encoded protein. The antibodies identify the 216 kDa BAT2 polypeptide. Immunostaining of embryonic tissue sections showed specific expression, especially in central nervous tissue, and suggests BAT2 as a novel differentiation marker.     

Potency requirements of rabies vaccines administered intradermally using the Thai Red Cross regimen: investigation of the immunogenicity of serially diluted purified chick embryo cell rabies vaccine

To determine the minimum vaccine potency per intradermal dose required to elicit an adequate immune response using the Thai Red Cross (TRC) regimen (2-2-2-0-1-1), healthy volunteers received 0.1 mL volumes of PCECV containing decreasing amounts of antigen. Subjects also received HRIG to evaluate potential interference with antibody production. Results indicated that when each 0.1 mL intradermal dose of PCECV contained antigen corresponding to 0.32 IU per intramuscular dose, every subject had titers above 0.5 IU/mL by day 14. These results confirm that the current World Health Organization (WHO) recommendations for vaccine potency (2.5 IU per intramuscular dose) are sufficient for use in the Thai Red Cross intradermal regimen.     

Distinct mixtures of muscarinic receptor subtypes mediate inhibition of noradrenaline release in different mouse peripheral tissues, as studied with receptor knockout mice

The muscarinic heteroreceptors modulating noradrenaline release in atria, urinary bladder and vas deferens were previously studied in mice in which the M(2) or the M(4) muscarinic receptor genes had been disrupted. These experiments showed that these tissues possessed both M(2) and non-M(2) heteroreceptors. The analysis was now extended to mice in which either the M(3), both the M(2) and the M(3), or both the M(2) and the M(4) genes had been disrupted (M(3)-knockout, M(2/3)-knockout and M(2/4)-knockout). Tissues were preincubated with (3)H-noradrenaline and then stimulated electrically (20 pulses per 50 Hz). In wild-type atria, carbachol (0.01-100 microM) decreased the electrically evoked tritium overflow by maximally 60-78%. The maximum inhibition of carbachol was reduced to 57% in M(3)-knockout and to 23% in M(2/4)-knockout atria. Strikingly, the effect of carbachol was abolished in M(2/3)-knockout atria.In wild-type bladder, carbachol (0.01-100 microM) reduced the evoked tritium overflow by maximally 57-71%. This effect remained unchanged in the M(3)-knockout, but was abolished in the M(2/4)-knockout bladder. In wild-type vas deferens, carbachol (0.01-100 microM) reduced the evoked tritium overflow by maximally 34-48%. The maximum inhibition of carbachol was reduced to 40% in the M(3)-knockout and to 18% in the M(2/4)-knockout vas deferens. We conclude that the postganglionic sympathetic axons of mouse atria possess M(2) and M(3), those of the urinary bladder M(2) and M(4), and those of the vas deferens M(2), M(3) and M(4) release-inhibiting muscarinic receptors.     

Total synthesis of two novel subpicomolar sarco/endoplasmatic reticulum Ca2+-ATPase inhibitors designed by an analysis of the binding site of thapsigargin

Analysis of molecular interaction fields based on the published crystal structure of thapsigargin bound to the sarco/endoplasmatic reticulum Ca(2+)-ATPase and analysis of the volume and shape of the ligand binding site and of the SERCA-thapsigargin interactions have enabled design of two new compounds inhibiting SERCA in the subpicomolar range. The two inhibitors were synthesized using (S)-carvone as starting material and found to be 3 and 10 times more potent than thapsigargin.     

Decreased bone turnover and deterioration of bone structure in two cases of pycnodysostosis

Pycnodysostosis is an uncommon human genetic disorder characterized by osteosclerosis of the skeleton, short stature, and bone fragility. The disease results from mutations in the cathepsin K gene, a lysosomal cysteine protease highly expressed in osteoclasts and crucial for the degradation of organic matrix from mineralized bone. Recently, interest has focused on a pharmaceutical inhibition of cathepsin K to prevent bone loss. However, little is known about the cellular activity or material quality of bone in pycnodysostosis. In the present study, transiliac bone biopsies from two affected individuals, aged 5 and 21 yr, were investigated using light microscopy, quantitative backscattered electron imaging, and small angle x-ray scattering. Results were compared with published age-matched reference data. The mutations in the cathepsin K gene of both patients were identified, including one novel defect. Both individuals had severe osteosclerosis, and their biopsies displayed multinucleated osteoclasts apposed to areas of demineralized matrix as well as bone-lining cells adjacent to this undigested collagen left over by osteoclasts. The homogeneity of the mineralized matrix was markedly disturbed due to large inclusions of mineralized cartilage residues. Histomorphometric evaluation showed a quantitative decrease in static parameters of bone formation. In contrast and despite deficient cathepsin K activity, osteoclastic parameters were close to normal range. At the nanostructural level, there was a marked increase in the mean thickness of the mineral particles, reflecting decreased bone remodeling. Examination of the trabecular structure revealed that the lamellae were highly disordered, which was also apparent from a poor alignment of mineral crystals oriented along the longitudinal axis of collagen fibrils. Taken together, these results strongly suggest that functional cathepsin K is important for balanced bone turnover, and enzyme deficiency results in a profound deterioration of bone quality with respect to trabecular architecture and lamellar arrangement, which is presumably the reason for bone fragility in pycnodysostosis.     

Amniotic cavity cultures, blood cultures, and surface swabs in preterm infants--useful tools for the management of early-onset sepsis?

AIMS: To evaluate the potential benefit of amniotic fluid and amniotic/placental membrane cultures for the management of early-onset sepsis in preterm infants. METHODS: The results of amniotic cavity cultures obtained during cesarean section and of peripheral blood cultures and surface swabs obtained from the preterm infant at the time of admission were analyzed with respect to the diagnosis of clinical sepsis in 221 preterm infants <34 weeks of gestation. RESULTS: 136 (61.5%) patients had negative amniotic cavity culture results or growth of contaminants, 56 (25.3%) had growth of Ureaplasma urealyticum, and 29 (13.1%) of other pathogens. The corresponding numbers for surface swabs were 82.8%, 11.6%, and 5.6%. A positive blood culture was found in only two neonates. Fifty-four patients (24.4%) had clinical early-onset sepsis. Patients with amniotic cavity culture results that were positive for other pathogens were significantly more likely to experience clinical sepsis than patients with negative culture results (51.7% vs 15.1%, OR 6.1, p<0.0001). Regarding surface swabs, this correlation did not reach statistical significance. CONCLUSION: The strong association between positive amniotic cavity culture results and clinical early-onset sepsis supports the existence of a causal relation and provides evidence for the potential value of amniotic and/or placental membrane sampling in the management of early-onset sepsis in preterm infants. Surface swabs add no additional information and hence should not be performed routinely.