Nuclear magnetic resonance spectroscopy to determine the micellar cholesterol in human bile.

The cholesterol of gallstones comes from the vesicular rather than the micellar phase of bile. Progress in this field has been limited because conventional analytical methods disturb the distribution of cholesterol between the two phases. The resonance of the cholesterol C6 proton occurs at a chemical shift of 5.4 ppm, to be shown by 2D NMR to be specific for biliary cholesterol, and arises only from the micellar mode. Thus integration of the C6 proton resonance peak area provides a direct non-invasive determination of the cholesterol distribution in human bile.     

Adaptation in Skate Photoreceptors

Receptor potentials were recorded extracellularly from the all-rod retina of the skate after the application of sodium aspartate. This agent suppresses the responses of proximal elements, but leaves relatively unaffected the electrical activity of the photoreceptors (a-wave) and pigment epithelium (c-wave). Since the latter develops too slowly to interfere with the receptor response, it was possible to isolate receptor potentials and to compare their behavior in light and dark adaptation with earlier observations on the S-potential, b-wave, and ganglion cell discharge. The results show that the photoreceptors display the full complement of adaptational changes exhibited by cells proximal to the receptors. Thus, it appears that visual adaptation in the skate is governed primarily by the photoreceptors themselves. Of particular interest was the recovery of sensitivity in the presence of background fields that initially saturate the receptor potential. Analysis of this recovery phase indicates that a gain-control mechanism operates within the receptors, at a distal stage of the visual process.     

Production and Purification of Human Fibroblast Collagenase (MMP-1) Expressed in the Methylotrophic YeastPichia pastoris

The cDNA that encodes the proenzyme form of human fibroblast collagenase (proMMP-1) was expressed in the methylotrophic yeastPichia pastoris.The proMMP-1 encoding DNA was fused to theSaccharomyces cerevisiaepre-pro α-mating factor secretion signal in theP. pastorispPIC9 expression plasmid, transformed into strain GS115 (His⁻), and His⁺Mut^s(slow methanol utilization) transformants were selected. Full-length proenzyme and processed forms of the protein could be detected in yeast culture supernatants following shake flask and 10-liter fermentations. The protein was purified to greater than 95% homogeneity. The recombinant proMMP-1 was comparable to the native fibroblast material based on (i) migration of the full-length molecule as a 52-kDa protein on reducing SDS–PAGE, (ii) correct N-terminal amino acid sequence, (iii) activation of the full-length molecule by 4-amino-phenylmercuric acetate to yield processed protein species, (iv) degradation of gelatin as monitored by zymogram gels, and (v) enzymatic activity. These data suggest that theP. pastorisexpression system offers a convenient and efficient means to produce and purify MMP-1.     

Adolescent idiopathic scoliosis as developmental instability

Adolescent idiopathic scoliosis is the most common spinal deformity affecting children, with a prevalence from mass screening programmes of 1–3%. Despite centuries of study, it remains a problem with no generally accepted theory of aetiology, and disagreement on its natural history and management. Because the deformity consists ultimately of gross left-right asymmetry, a study was undertaken to test the hypothesis that it might be a manifestation of developmental instability. Palmar dermatoglyphics in 112 normal subjects, 62 with non-scoliosis trunk asymmetry and 85 with defined adolescent idiopathic scoliosis were examined and both the absolute right-left difference and the ratio of this to the total were considered. There was increased fluctuating asymmetry of atd difference in those with any asymmetry, scoliotic or not, and increased directional asymmetry of ab and cd ridge counts only in those with pure scoliosis. This suggests that, at adolescence, developmental instability may result in a loss of symmetry in growth, and that in the presence of an increased developmental left-right gradient, this may be of sufficient severity to be classified as deformity and come to the attention of orthopaedic surgeons. This interpretation changes the focus of many previous observations on scoliosis and raises the prospect that developmental stability in humans has relevance to problems hitherto restricted to clinical practice.     

Isolation of multiple substances in amniotic fluid that regulate amnion prostaglandin E2 production: The effects of gestational age and labor

The present study was undertaken to evaluate the effects of whole amniotic fluid (AF) and fractions of AF on amnion cell prostaglandin E2 (PGE2) production. Amnion cells were grown to confluence and then incubated in the presence of AF, or fractions thereof, obtained at 17-19 weeks gestation (MID), at term prior to the onset of labor (NIL), and at term after spontaneous onset of labor (LABOR). All whole AFs were stimulatory to amnion cell PGE2 production (p less than 0.001) but the stimulation by NIL and LABOR AFs was significantly greater (p less than 0.001) than stimulation by the MID AF. Fractionation of the AFs from the three groups (n = 9-10 per group) revealed multiple discernable peaks of stimulatory activity in each group. The majority of peaks had retention times that were similar among the three groups, and peak stimulatory activities were greater in NIL and LABOR samples than in MID samples.