Immunoelectron microscopic studies reveal differences in distribution of sialo-oligosaccharide receptors for Mycoplasma pneumoniae on the epithelium of human and hamster bronchi.

Long-chain sialo-oligosaccharides with poly-N-acetyllactosamine backbones (Ii antigen type) are major host cell receptors for the human pathogen Mycoplasma pneumoniae. Previous immunofluorescence studies of the human bronchial epithelium, using sequence-specific monoclonal antibodies to the branched I-type and linear i-type backbones, have indicated that sialylated and nonsialylated long-chain sequences of both types are richly expressed on the ciliated cells, where they are polarized at the apical aspects. These sequences are lacking in the goblet cells. In the present study, the display of these oligosaccharides has been investigated by electron microscopy (immunogold labelling) in the human bronchial epithelium and in that of the hamster, an animal model commonly used for M. pneumoniae infection. In the human bronchial epithelium, the long-chain branched sequences have been detected along the entire length of the cilia and on microvilli, whereas the linear sequences are confined to the microvilli and the basal aspects of the cilia. On the ciliated epithelial cells of the hamster, by contrast, the branched and linear sequences (sialo- and asialo-) have been detected exclusively on microvilli. A further striking difference is that in the hamster these structures are expressed in abundance on the goblet cells and in the intracellular globules. We suggest that the latter finding may partly explain the relatively large doses of M. pneumoniae required to establish experimental infection in the hamster, as the receptor-bearing secreted mucus may have a protective role in binding to the microorganisms, leading to their clearance by bronchociliary action.     

New reference allelic ladders to improve allelic designation in a multiplex STR system

This paper reports the composition of a new reference allelic ladder mixture for use with a multiplex DNA profiling system consisting of six short tandem repeat loci. The loci included in this mixture are HUMTH01, D21S11, D18S51, D8S1179, HUMVWAF31/A, HUMFIBRA/FGA and an amelogenin sex test. Sequence analysis of individual ladder alleles was carried out and allelic designations made in accordance with the recommendations of the International Society of Forensic Haemogenetics (1992; 1994). A series of rare alleles which increase the range of alleles previously reported were identified. By including some of the rare alleles into the ladder marker system, we have significantly improved the ability to identify new alleles in unknown samples. Received: 12 August 1997 / Received in revised form: 7 November 1997     

Post-capillary venules in the “milky spots” of the greater omentum are the major site of plasma protein and leukocyte extravasation in rodent models of peritonitis

Intraperitoneal injection of inflammatory agents in the mouse and rat causes plasma protein and leukocyte extravasation into the peritoneal cavity. Following an intraperitoneal injection of zymosan A, the milky spots of the omentum were the only abdominal sites detected where intravenously administered Monastral Blue labeled interendothelial cell gaps responsible for plasma extravasation. In addition, when colored microspheres were intraventricularly administered to quantify blood flow, the omentum was the only abdominal organ which showed an increase in blood flow during zymosan A peritonitis. A combination of light and electron microscopy, plus measurement of myeloperoxidase activity (a marker of neutrophil accumulation) demonstrated that the omental milky spots are the major route through which leukocytes migrate into the peritoneal cavity. Identical structures in the pleura likewise are the sites of protein leakage into the pleural cavity. In contrast, selective sites of protein and cellular extravasation could not be detected in the synovial lining of the inflamed knee joint.     

Antibiograms of resistant Gram-negative bacteria from Scottish CF patients.

BACKGROUND: Over a 19-month pilot phase, 93 multiply resistant Gram-negative isolates from Scottish cystic fibrosis patients were sent to a referral laboratory for further investigation. METHODS: In common with the referring diagnostic laboratories, disc diffusion testing was carried out. Antibiotic susceptibility testing was also established by MIC methodology. NCCLS methods were used throughout. Twenty antibiotics were tested. RESULTS: Comparing disc diffusion results against MIC results, there were 167 (14%) major errors. By MIC, Pseudomonas aeruginosa (n = 59), Stenotrophomonas maltophilia (n = 16), Burkholderia cepacia (n = 10) and Alcaligenes xylosoxidans (n = 7) were susceptible to 18%, 11%, 4% and 35% of the antibiotics tested, respectively. Colistin and tobramycin were the most active agents against P. aeruginosa with 60% and 49%, respectively, testing susceptible. Minocycline and gentamicin were most active against S. maltophilia with 58% and 18%, respectively, testing susceptible. B. cepacia were most susceptible to co-trimoxazole (10%) and ciprofloxacin (10%). Five and six of the seven A. xylosoxidans isolates were susceptible to piperacillin and imipenem, respectively. CONCLUSIONS: Improved methods for susceptibility testing of such clinical isolates need to be employed in routine diagnostic laboratories. Levels of resistance in referred isolates were very high and similar to those described in the USA.     

Aminoguanidine selectively inhibits inducible nitric oxide synthase.

1. Endotoxin induces nitric oxide synthase in vascular tissue, including rat main pulmonary artery. Currently available agents that cause inhibition of nitric oxide synthase are relatively non-selective between the constitutive and inducible forms of the enzyme. 2. Aminoguanidine caused a dose-dependent increase in phenylephrine-induced tension in intact and endothelium-denuded pulmonary artery rings from endotoxin-treated rats, but had no effect on sham-treated controls. 3. Contraction caused by aminoguanidine in endothelium-denuded vessels from endotoxin-treated rats was unaffected by indomethacin (10 microM), and by cimetidine and mepyramine (both 10 microM), excluding an effect of aminoguanidine mediated by arachidonic acid metabolites or histamine. 4. Contraction caused by aminoguanidine in endothelium-denuded vessels from endotoxin-treated rats was abolished by L-arginine (2 mM) and L-NG-monomethyl arginine (300 microM), but unaffected by D-arginine and D-NG-monomethyl arginine, suggesting that its action is mediated by the L-arginine/nitric oxide pathway.(ABSTRACT TRUNCATED AT 250 WORDS)     

Keratoacanthoma: the value of photographic follow-up in conservative management

The management of keratoacanthoma is controversial. Two cases are presented to confirm the value of conservative treatment. The presence of hairs within keratoacanthomas as a diagnostic feature is discussed. Correspondence to: R.W. Griffiths     

Mechanical characteristics of skinned and intact muscle fibres from the giant barnacle,Balanus nubilus

Intact muscle fibres fromBalanus nubilus develop tensions of up to 600 kN sd m⁻² during electrical stimulation. The rise of tension occurs with a half-time (177 ms at 12° C) about fivefold longer than that of tetanised frog muscle at the same temperature. The response of myofibrillar bundles to a rapid stretch resembles that of frog muscle but has a y_o value (i.e. the size of an instantaneous release necessary to just discharge tension) which is ca. 2.5 times smaller, and phase 2 of the tension transient (the “quick phase”) occurs at a rate comparable to that of frog muscle. In contrast, the ATPase activity (0.018 mmoles · kg wet weight⁻¹ · s⁻¹) of this preparation and its maximum shortening velocity (0.15–0.16 muscle lengths · s⁻¹) are both at least fivefold slower than frog muscle. These findings can be accounted for by a cross-bridge cycle in barnacle muscle in which events prior and subsequent to the tension generating step(s) occur at a rate at least fivefold slower than comparable steps in frog muscle, but the step(s) associated with tension development occur at similar rates in the two preparations. Since the rate of mechanical relaxation in barnacle muscle is modified in the presence of intracellular calcium buffers and by depolarisation-induced elevation of the free calcium during the relaxation phase, it is proposed that the time course of relaxation is not determined exclusively by the kinetics of the cross-bridge cycle, but is also dependent on the free calcium concentration during relaxation.     

Analysis of the operation of the SCD Response intermittent compression system

The work assessed the performance of the Kendall SCD Response intermittent pneumatic compression system for deep vein thrombosis prophylaxis, which claimed to set its cycle according to the blood flow characteristics of individual patient limbs. A series of tests measured the system response in various situations, including application to the limbs of healthy volunteers, and to false limbs. Practical experimentation and theoretical analysis were used to investigate influences on the system functioning other than blood flow. The system tested did not seem to perform as claimed, being unable to distinguish between real and fake limbs. The intervals between compressions were set to times unrealistic for venous refill, with temperature changes in the cuff the greatest influence on performance. Combining the functions of compression and the measurement of the effects of compression in the same air bladder makes temperature artefacts unavoidable and can cause significant errors in the inter-compression interval.     

Effect of prostaglandin E2 on agonist-stimulated cAMP accumulation in the distal convoluted tubule isolated from the rabbit kidney

The effects of calcitonin, vasoactive intestinal peptide (VIP), parathyroid hormone (PTH) and isoprenaline on intracellular cAMP accumulation were determined in the distal tubule (DCT) microdissected from collagenase-treated rabbit kidney. In DCTb (the initial bright portion) calcitonin (10 ng/ml) elicited a highly reproducible response 203.7±19.1 fmol cAMP mm^–1 4 min^–1 (SE, N=13) whereas VIP-induced cAMP accumulation was less and more variable from one experiment to another (1 M, 97.2±17.8 fmol mm^–1 4 min^–1, SE, N=12). When used in combination, these two agonists were non-additive, indicating stimulation of a single pool of cAMP in DCTb. In DCTg, (granular) which consists of at least two cell types, PTH (100 nM) elicited a marked, reproducible accumulation of cAMP (154.3±27.0 fmol mm^–1 4 min^–1; SE, N=5). Isoprenaline (1 M) and VIP (1 M) induced much smaller increases in cAMP levels 20.9±2.7 and 29.4±4.1 fmol mm^–1 4 min^–1 (SE, N=5) respectively, and, when used in combination, were non-additive, demonstrating that VIP and isoprenaline are active on the same cell type. In DCTb, prostaglandin E₂ (PGE₂) inhibited both calcitoninand VIP-stimulated cAMP accumulation (calcitonin 57.8±2.7% inhibition, SE, N=16; VIP, 80.6±2.1% inhibition, SE, N=5). The EC₅₀ values for calcitonin were 1.21±0.33 ng/ml and 1.83±0.25 ng/ ml (SD, N=3) in the absence and presence of PGE₂ (300 nM) respectively with an IC₅₀ for PGE₂ of 26.3±6.3 nM (SE, N=4). In contrast, no effects of PGE₂ were seen in DCTg vis à vis PTH, isoprenaline or VIP. The percentage inhibition of calcitonin-stimulated cAMP accumulation by PGE₂ was of the same order in the presence of isobutylmethylxanthine (an inhibitor of all types of phosphodiesterase), Ro 20-1724 (inhibitor of low-K _m cAMP-specific phosphodiesterase) or in the absence of inhibitor. Preincubation of DCTb with pertussis toxin for up to 8 h in different experimental conditions did not relieve the inhibition by PGE₂. Protein kinase C activation by phorbol ester did not attenuate calcitonin responses. These data demonstrate that the inhibition by PGE₂ of cAMP production is restricted to the initial portion (DCTb) of the distal convoluted tubule and is effective on both calcitonin and VIP responses. When tested in the presence of Ro 20-1724, ionomycin, A₁-adenosine, ₂-adrenergic and muscarinic agonists were without effect on calcitoninand PTH-stimulated cAMP accumulation in DCTb and DCTg respectively.