An essential role for calmodulin in regulating human T cell aggregation

After activation of T cells with either CD3 antibodies or phorbol esters, we have found that T cell-cell aggregation, integrin-dependent actin reorganisation and cell spreading are strongly suppressed by any of three structurally different calmodulin antagonists, without any effect on the amount of CD11/CD18 integrin binding to the actin cytoskeleton. However, only T cell receptor-induced, and not phorbol ester-induced, aggregation and cell spreading are prevented by inhibitors of phosphatidylinositide (PI) 3-kinase. These results suggest that PI 3-kinase lies upstream of calmodulin in the signalling pathway leading to T cell aggregation, cell spreading and actin reorganisation and that cell spreading and actin reorganisation are essential for T cell adhesion.     

Mice Lacking beta2-Integrin Function Remain Glucose Tolerant in Spite of Insulin Resistance,Neutrophil Infiltration and Inflammation

Beta2-integrins are important in leukocyte trafficking and function, and are regulated through the binding of cytoplasmic proteins, such as kindlin-3, to their intracellular domain. Here, we investigate the involvement of beta2-integrins in the regulation of metabolic disease using mice where the kindlin-3 binding site in the beta2-integrin cytoplasmic tail has been mutated (TTT/AAA-beta2-integrin knock-in (KI) mice), leading to expressed but dysfunctional beta2-integrins and significant neutrophilia in vivo. Beta2-integrin KI mice fed on a high fat diet showed normal weight gain, and normal accumulation of macrophages and lymphocytes in white adipose tissue (WAT) and liver, but increased neutrophil numbers especially in WAT. In addition, beta2-integrin KI mice fed on a high fat diet showed significantly increased peripheral insulin resistance in response to high-fat feeding. However, this was associated with improved glucose disposal following glucose load. Interestingly, beta2-integrin KI neutrophils produced more elastase in vitro, in response to stimulation. Beta2-integrin KI mice displayed variability of tissue inflammatory status, with liver and WAT exhibiting little or no difference in inflammation compared to high fat fed controls, whereas skeletal muscle demonstrated a raised inflammatory profile in association with higher elastase levels and diminished signalling through the IRS1-PKB pathway. In conclusion, although expression of dysfunctional beta2-integrins increased neutrophil production and infiltration into tissue, skeletal muscle was the most affected tissue exhibiting evidence of higher neutrophil activity and insulin resistance. Thus, beta2-integrins modulate glucose homeostasis during high fat feeding predominantly through actions on skeletal muscle to affect metabolic phenotype in vivo.     

Cubicle Refusal and Rearing Accommodation as Possible Mastitis Risk Factors in Cubicle-Housed Dairy Heifers

Fifty-nine of the 65 dairy farms with cubicle sheds in the Norwegian county of Oppland in 1990 were included in a study of rearing accommodation, cubicle refusal and mastitis incidence. The farmers recorded the favoured resting location of the individual cows and heifers throughout the final week of pregnancy as well as during calving. The observations were matched with individual records of mastitis cases during the first 30 days after calving. Mastitis incidence in the heifers was analysed by logistic regression using rearing accommodation and cubicle refusal as independent variables, controlling for herd as a clustering factor. Cubicle refusal was found in 29% of the heifers, but in only 3% of older cows. The results of the analysis indicated a tendency for cubicle refusal to be associated with an increased mastitis incidence among the heifers (OR = 2.2, c.i._95%OR = 0.9–5.4, P = 0.08). Cubicle refusal accounted for 21% (0–32%) of the mastitis cases in the study population (PAF = 0.21).     

Cubicle Refusal in Norwegian Dairy Herds

In order to survey the behaviour of choosing the alley area instead of a cubicle as a lying place (cubicle refusal), a questionnaire was sent to the 273 dairy farms in Norway known to keep cows in cubicle housing systems. Sixty-six percent of the farmers contacted were included in the study. The median herd size was 18 cows (range 7–118). More than 85% of the herds had sheds providing one or more cubicles per cow. The mean herd occurrence of cubicle refusal was 6%, but showed great variation (range 0–55%). Regression analysis showed a significant association between rearing heifers in slatted floor pens and an increased cubicle refusal occurrence (p = 0.02, R² = 0.05), while herd size, use of litter, or cubicle-to-animal ratio were not found to be associated with cubicle refusal. The practice of rearing heifers in slatted floor pens accounted for about one half of the observed cubicle refusal (etiologic fraction = 0.51).     

ABE production from corn: a recent economic evaluation

This article details an economic assessment of butanol production from corn using the newly developed hyper-butanol-producing strain of Clostridium beijerinckii BA101. Butanol is produced in batch reactors and recovered by distillation. For a plant with 153,000 metric tons of acetone, butanol, and ethanol (ABE) production capacity, the production equipment cost and total working capital cost is US$33.47×10⁶ and US$110.46×10⁶, respectively. Based on a corn price (C _p) of US$79.23 ton⁻¹ (US$2.01 bushel⁻¹), an ABE yield of 0.42 (g ABE/g glucose) butanol price is projected to be US$0.34 kg⁻¹. An improved yield of 0.50 will reduce this price to US$0.29 kg⁻¹. Assumptions, such as by-product credit for gases and complete conversion of corn steep liquor (CSL) to fermentation by-products, have been taken into consideration. An increased price of corn to US$197.10 ton⁻¹ would result in a butanol price of US$0.47 kg⁻¹. A grass-rooted plant would result in a butanol price of US$0.73 kg⁻¹ (C _p US$79.23 ton⁻¹). In a worst case scenario, the price of butanol would increase to US$1.07 kg⁻¹ (C _p 197.10 ton⁻¹ for a grass-rooted plant and assuming no credit for gases). This is based on the assumption that corn price would not increase to more than US$197.10 ton⁻¹. Journal of Industrial Microbiology & Biotechnology (2001) 27, 292–297. Received 12 September 2000/ Accepted in revised form 12 January 2001     

Systematic implications of male caudal morphology in ascaridoid nematode parasites

In view of earlier results, suggesting a considerable stability in the number and distribution patterns of caudal papillae and papillae-like structures in the male of some ascaridoid species, these features are analysed more generally within the superfamily Ascaridoidea. They include the single pair of phasmids on the tail, four pairs of distal papillae some of which are considered in some cases to occur anterior to the level of the cloaca, two pairs of paracloacal papillae, a median papilla or plaque, and numerous pairs of proximal papillae some of which may be situated posterior to the level of the cloaca. The exceptions to this general pattern and the systematic and phylogenetic significance of caudal morphology are discussed.     

Pathogenicity of the gill artery worm Philometra obturans (Nematoda) in northern pike (Esox lucius) in southwest Finland

The pathogenicity of female Philometra obturans (Prenant, 1886) was investigated. Fish samples were taken in May and June 2001 from Lake Littoistenj?rvi, in southwest Finland. The intensity of infection varied between 1 and 4 worms, these being found in the afferent artery of the 2 outermost gill arches. The prevalence was 33.9% (n = 59). Host condition and tissue reactions were investigated through dissection and analysis of histopathology. The parasites obstructed the gill arteries, and the arterial wall was stretched around intact worms. Hypertrophy of the arterial wall was observed, especially when only disintegrating remains of the parasites were present. Infected pike were in a poorer condition after catch, transportation, and holding in aquaria and thus less tolerant to stress than uninfected pike. This was correlated with the number of infected gill arteries (P < 0.05). The results show that infection with this nematode has a detrimental effect on the host when combined with environmental or physiological stress and may result in host mortality.     

Specific integrin alpha and beta chain phosphorylations regulate LFA-1 activation through affinity-dependent and -independent mechanisms

Integrins are adhesion receptors that are crucial to the functions of multicellular organisms. Integrin-mediated adhesion is a complex process that involves both affinity regulation and cytoskeletal coupling, but the molecular mechanisms behind this process have remained incompletely understood. In this study, we report that the phosphorylation of each cytoplasmic domain of the leukocyte function-associated antigen-1 integrin mediates different modes of integrin activation. alpha Chain phosphorylation on Ser1140 is needed for conformational changes in the integrin after chemokine- or integrin ligand-induced activation or after activation induced by active Rap1 (Rap1V12). In contrast, the beta chain Thr758 phosphorylation mediates selective binding to 14-3-3 proteins in response to inside-out activation through the T cell receptor, resulting in cytoskeletal rearrangements. Thus, site-specific phosphorylation of the integrin cytoplasmic domains is important for the dynamic regulation of these complex receptors in cells.     

Neural diffusivity and pre-emptive epileptic seizure intervention

The propagation of epileptic seizure activity in the brain is a widespread pathophysiology that, in principle, should yield to intervention techniques guided by mathematical models of neuronal ensemble dynamics. During a seizure, neural activity will deviate from its current dynamical regime to one in which there are significant signal fluctuations. In silico treatments of neural activity are an important tool for the understanding of how the healthy brain can maintain stability, as well as of how pathology can lead to seizures. The hope is that, contained within the mathematical foundations of such treatments, there lie potential strategies for mitigating instabilities, e.g. via external stimulation. Here, we demonstrate that the dynamic causal modelling neuronal state equation generalises to a Fokker-Planck formalism if one extends the framework to model the ways in which activity propagates along the structural connections of neural systems. Using the Jacobian of this generalised state equation, we show that an initially unstable system can be rendered stable via a reduction in diffusivity–i.e., by lowering the rate at which neuronal fluctuations disperse to neighbouring regions. We show, for neural systems prone to epileptic seizures, that such a reduction in diffusivity can be achieved via external stimulation. Specifically, we show that this stimulation should be applied in such a way as to temporarily mirror the activity profile of a pathological region in its functionally connected areas. This counter-intuitive method is intended to be used pre-emptively–i.e., in order to mitigate the effects of the seizure, or ideally even prevent it from occurring in the first place. We offer proof of principle using simulations based on functional neuroimaging data collected from patients with idiopathic generalised epilepsy, in which we successfully suppress pathological activity in a distinct sub-network prior to seizure onset. Our hope is that this technique can form the basis for future real-time monitoring and intervention devices that are capable of treating epilepsy in a non-invasive manner.     

In vitro glucocorticoid sensitivity is associated with clinical glucocorticoid therapy outcome in rheumatoid arthritis

Introduction

Genetic and disease-related factors give rise to a wide spectrum of glucocorticoid (GC) sensitivity in rheumatoid arthritis (RA). In clinical practice, GC treatment is not adapted to these differences in GC sensitivity. In vitro assessment of GC sensitivity before the start of therapy could allow more individualized GC therapy. The aim of the study was to investigate the association between in vitro and in vivo GC sensitivity in RA.

Methods

Thirty-eight early and 37 established RA patients were prospectively studied. In vitro GC sensitivity was assessed with dexamethasone-induced effects on interleukin-2 (IL-2) and glucocorticoid-induced leucine zipper (GILZ) messenger RNA expression in peripheral blood mononuclear cells (PBMCs). A whole-cell dexamethasone-binding assay was used to measure number and affinity (1/K_D) of glucocorticoid receptors (GRs).In vivo GC sensitivity was determined by measuring the disease activity score (DAS) and health assessment questionnaire disability index (HAQ-DI) score before and after 2 weeks of standardized GC treatment.

Results

GR number was positively correlated with improvement in DAS. IL-2-EC₅₀ and GILZ-EC₅₀ values both had weak near-significant correlations with clinical improvement in DAS in intramuscularly treated patients only. HAQ responders had lower GILZ-EC₅₀ values and higher GR number and K_D.

Conclusions

Baseline cellular in vitro glucocorticoid sensitivity is modestly associated with in vivo improvement in DAS and HAQ-DI score after GC bridging therapy in RA. Further studies are needed to evaluate whether in vitro GC sensitivity may support the development of tailor-made GC therapy in RA.