Age-related alterations in IL-1beta, TNF-alpha, and IL-6 concentrations in parotid acinar cells from BALB/c and non-obese diabetic mice.

IL-1beta, TNF-alpha, and IL-6 have been implicated in the destruction of parotid gland acinar cells (but not duct cells) in autoimmune sialoadenitis. Here we report the temporal alterations of these cytokines in parotid acinar cells that may lead to this specificity in cell death in the non-obese diabetic (NOD) mouse model for Sj?gren's syndrome. Immunohistochemistry on paraffin sections of parotid gland from 5- and 10-week-old BALB/c and NOD mice confirmed the presence of many peri-acinar lymphoid nodules but few T-cells and macrophages between acinar cells. RT-PCR on enzymatically dispersed mouse parotid acinar cells (MPACs) showed no bands for CD3varepsilon, CD20, or F4/80 regardless of mouse strain or age. By ELISA, MPACs from 10-week-old NODs showed a small but highly significant (p<0.003) increase in IL-1beta and a large significant decrease (p<0.008) in IL-6 compared to 5-week-old NODs. Norepinephrine-stimulated amylase release from MPACs was not different regardless of mouse strain or age. These data show that alterations in acinar cell production of IL-1beta and IL-6 in aging NODs precede periductal lymphoid aggregates and acinar cell secretory dysfunction. (J Histochem Cytochem 48:1033-1041,2000)     

Evaluation of the Leptospira interrogans Outer Membrane Protein OmpL37 as a Vaccine Candidate

The identification of potential vaccine candidates against leptospirosis remains a challenge. However, one such candidate is OmpL37, a potentially surface-exposed antigen that has the highest elastin-binding ability described to date, suggesting that it plays an important role in host colonization. In order to evaluate OmpL37’s ability to induce a protective immune response, prime-boost, DNA and subunit vaccine strategies were tested in the hamster model of lethal leptospirosis. The humoral immune response was evaluated using an indirect ELISA test, and the cytokine profile in whole blood was determined by quantitative real-time PCR. Unlike the DNA vaccine, the administration of recombinant OmpL37 induced a strong IgG antibody response. When individually administrated, both formulations stimulated a TNF-α mediated inflammatory response. However, none of the OmpL37 formulations or vaccination strategies induced protective immunity. Further studies are required towards the identification of new vaccine targets against leptospirosis.     

Predicting the Activity Coefficients of Free-Solvent for Concentrated Globular Protein Solutions Using Independently Determined Physical Parameters

The activity coefficient is largely considered an empirical parameter that was traditionally introduced to correct the non-ideality observed in thermodynamic systems such as osmotic pressure. Here, the activity coefficient of free-solvent is related to physically realistic parameters and a mathematical expression is developed to directly predict the activity coefficients of free-solvent, for aqueous protein solutions up to near-saturation concentrations. The model is based on the free-solvent model, which has previously been shown to provide excellent prediction of the osmotic pressure of concentrated and crowded globular proteins in aqueous solutions up to near-saturation concentrations. Thus, this model uses only the independently determined, physically realizable quantities: mole fraction, solvent accessible surface area, and ion binding, in its prediction. Predictions are presented for the activity coefficients of free-solvent for near-saturated protein solutions containing either bovine serum albumin or hemoglobin. As a verification step, the predictability of the model for the activity coefficient of sucrose solutions was evaluated. The predicted activity coefficients of free-solvent are compared to the calculated activity coefficients of free-solvent based on osmotic pressure data. It is observed that the predicted activity coefficients are increasingly dependent on the solute-solvent parameters as the protein concentration increases to near-saturation concentrations.     

Intracellular signals for developmental hemoglobin switching

We have detected trans-acting factors that regulate developmental hemoglobin switching by fusing erythroid cells of different developmental programs. Adult erythroid cells of one anuran species, Xenopus laevis, were fused with tadpole erythroid cells of another frog, Rana catesbeiana. In a second set of experiments, dimethyl sulfoxide-induced murine erythroleukemia cells, which express only adult mouse globins, were fused with Rana tadpole erythroid cells, which express only embryonic and fetal-like globins. Adult Rana globin gene expression was detected in both sets of transient heterokaryons at 6 hr after fusion. Dot blots and Northern blots of total RNA from the heterokaryons contained material that reacted with an adult Rana alpha-globin probe; newly synthesized adult Rana hemoglobin tetramers were detected with native polyacrylamide gel electrophoresis. These results show that developmental stage-specific transacting factors for globin genes can function across vertebrate classes (mammalia to amphibia) and suggest that the mechanisms that regulate developmental hemoglobin switching are highly conserved.     

Environmental design and its evaluation for intensively housed animals

It is argued that applied ethology has emerged as an empirical science with a limited base of theory. Two models are presented and used to examine problems of evaluation of husbandry design. The first model, of the process of adaptation to environments, predicts that long-term strain, learned helplessness and stereotypic behaviours are evidence of a failure to adapt appropriately and may thus acquire value as criteria for the suitability of long-term husbandry environments for animals. The model of the man-animal interface is used to examine systematically the components of environmental design for the process of evaluation. The features of an evaluation included small- and large-scale testing, the daily, weekly, seasonal and “settling in” schedules, the choice of stock and staff, and the physical features of an environment, including water, feed, floors, walls, shelter and the behavioural options available. Other discussions of the evaluative process include the role of hierarchy, staff, sensory input, postures, movements and their location, disruptions and stereotypics.     

Joint reaction forces at the first MTP joint in a normal elderly population

The calculation of net ankle, knee, and hip joint reaction forces is an often applied procedure in the analysis of gait. Except for very few studies, joint reaction forces have not been measured in other joints such as the fingers, wrist, elbow, shoulder and toes. In this study the joint reaction forces between the metatarsal head and the proximal phalanx and the metatarsal head and the sesamoids are calculated for the push off phase during gait. The results of ten normal elderly subjects show that the maximum resultant loads of the two articulations lie close to the longitudinal axis of the metatarsal. The knowledge of the magnitude and direction of the joint reaction forces of a normal elderly population will be essential for the design of an optimal fixation of an artificial anatomical first MTP joint.     

Inhibition of serotonin-amplified human platelet aggregation by ketanserin, ritanserin, and the ergoline 5HT2 receptor antagonists-LY53857, sergolexole, and LY237733

Human platelets are known to possess 5HT2 receptors which, when activated, amplify the aggregation response produced by other aggregating agents. Several 5HT2 receptor antagonists, including ketanserin and ritanserin, are known to antagonize serotonin-mediated aggregation of human platelets. In the present study, we document the ability of three ergoline 5HT2 receptor antagonists, LY53857, sergolexole, and LY237733, to antagonize the serotonergic component of the human platelet aggregation response. Potencies of the ergoline esters (LY53857 and sergolexole) and the ergoline amide (LY237733) to inhibit serotonin-amplified platelet aggregation responses were similar to the potencies of ketanserin and ritanserin under the conditions of our study. Furthermore, all five 5HT2 receptor antagonists were capable of fully inhibiting the serotonergic component of the platelet aggregation response. In contrast to these potent ergoline esters and amides, 1-isopropyl dihydrolysergic acid (up to 10(-5)M), a putative metabolite of the ergoline esters, was ineffective under these in vitro conditions. These data are consistent with the high potency of these ergolines as antagonists of 5HT2 receptors and further support the involvement of 5HT2 receptors on human platelets in the amplifying response to serotonin.     

Noise and the classical musician.

OBJECTIVES--To test the hypothesis that noise exposure may cause hearing loss in classical musicians. DESIGN--Comparison of hearing levels between two risk groups identified during the study by measuring sound levels. SETTING--Symphony orchestra and occupational health department in the west Midlands. MAIN OUTCOME MEASURES--Hearing level as measured by clinical pure tone audiometry. RESULTS--Trumpet and piccolo players received a noise dose of 160% and 124%, respectively, over mean levels during part of the study. Comparison of the hearing levels of 18 woodwind and brass musicians with 18 string musicians matched for age and sex did not show a significant difference in hearing, the mean difference in the hearing levels at the high (2, 4, and 8 KHz) audiometric frequencies being 1.02 dB (95% confidence interval -2.39 to 4.43). CONCLUSIONS--This study showed that there is a potential for occupational hearing loss in classical orchestral musicians.     

The human loricrin gene.

Loricrin is the major protein component of the cornified cell envelope of terminally differentiated mammalian epidermal (stratum corneum) cells. Using a specific human cDNA clone, we have isolated and characterized the human loricrin gene. We show that it has a very simple structure of a single intron of 1188 base pairs (bp) in the 5'-untranslated region; there are no introns in coding sequences. By use of rodent-human somatic cell hybrids, followed by in situ hybridization with a biotin-labeled genomic DNA clone, the single-copy gene maps to chromosome location 1q21. Polymerase chain reaction analyses of genomic DNAs from different individuals show that human loricrin consists of two allelic size variants, due to sequence variations in its second glycine loop domain, and these variants segregate in the human population by normal Mendelian mechanisms. Furthermore, there are multiple sequence variants within these two size class alleles due to various deletions of 12 bp (4 amino acids) in the major loop of this glycine loop domain. By use of a specific loricrin antibody, we show by immunogold electron microscopy that loricrin initially appears in the granular layer of human epidermis and forms composite keratohyalin granules with profilaggrin, but localizes to the cell periphery (cell envelope) of fully differentiated stratum corneum cells.