Alpha2-adrenergic regulation of NO production alters postoperative intestinal smooth muscle dysfunction in rodents

Alpha2-adrenergic receptor activation plays an important role in the development of postoperative ileus. Alpha2-adrenergic receptors also regulate nitric oxide (NO) production by the mononuclear phagocyte system. We have previously shown that intestinal manipulation leads to a significant increase in NO production by infiltrating monocytes within the intestinal muscularis. The purpose of this study was to investigate whether alpha2-adrenergic blockade with yohimbine would alter postsurgical intestinal smooth muscle dysfunction and NO production by infiltrating monocytes and macrophages within the intestinal muscularis. Rats underwent small bowel intestinal manipulation with or without yohimbine. In vivo gastrointestinal transit and in vitro jejunal circular muscle contractility was measured 24 h postoperatively. RT-PCR was used to detect inducible NO synthase (iNOS) expression. NO levels in tissue culture supernatants were measured. Immunohistochemistry was used to localize alpha2-adrenergic receptor expression in the intestinal muscularis. Yohimbine significantly decreased manipulation-induced delay in gastrointestinal transit and reversed the postoperative decrease in intestinal muscle contractility. Intestinal manipulation resulted in significant iNOS mRNA induction in the intestinal muscularis, which was markedly attenuated after yohimbine treatment. Yohimbine also significantly decreased the postoperative increase in NO released into intestinal muscularis tissue culture supernatant. Immunohistochemistry identified alpha2-adrenergic receptors on monocytes recruited postoperatively into the intestinal muscularis. This study demonstrates that alpha2-adrenergic receptor stimulation of the inflamed postoperative intestinal muscularis plays a significant role in aggravating postoperative ileus through an enhanced induction of iNOS mRNA and increased release of NO from manipulated intestinal muscularis.     

MHCPred: bringing a quantitative dimension to the online prediction of MHC binding

The accurate prediction of T cell epitopes is one of the key aspirations of immunoinformatics. We have developed a partial least squares-based, robust multivariate statistical method for the quantitative prediction of peptide binding to major histocompatibility complexes (MHCs), the principal checkpoint on the antigen presentation pathway. As a service to the immunobiology community, we have made a Perl implementation of the method available as a World Wide Web server.     

Coordination of anthocyanin decline and photosynthetic maturation in juvenile leaves of three deciduous tree species

Juvenile leaves in high-light environments commonly appear red as a result of anthocyanin pigments, which play a photoprotective role during light-sensitive ontogenetic stages. The loss of anthocyanin during leaf development presumably corresponds to a decreased need for photoprotection, as photosynthetic maturation allows leaves to utilize higher light intensities. However, the relationship between photosynthetic development and anthocyanin decline has yet to be quantitatively described. In this study, anthocyanin concentration was measured against photopigment content, lamina thickness, anatomical development, and photosynthetic CO(2) exchange in developing leaves of three deciduous tree species. In all species, anthocyanin disappearance corresponded with development of c. 50% mature photopigment concentrations, c. 80% lamina thickness, and differentiation of the mesophyll into palisade and spongy layers. Photosynthetic gas exchange correlated positively with leaf thickness and chlorophyll content, and negatively with anthocyanin concentration. Species with more rapid photosynthetic maturation lost anthocyanin earliest in development. Chlorophyll a/b ratios increased with leaf age, and were lower than those of acyanic species, consistent with a shading effect of anthocyanin. These results suggest that anthocyanin reassimilation is linked closely with chloroplast and whole-leaf developmental processes, supporting the idea that anthocyanins protect tissues until light processing and carbon fixation have matured to balance energy capture with utilization.     

Enhanced activity of NLRP3 inflammasome in peripheral blood cells of patients with active rheumatoid arthritis

IntroductionInterleukin-1β (IL-1β) is a major inflammatory cytokine, produced predominantly by innate immune cells through NLRP3-inflammasome activation. Both intrinsic and extrinsic danger signals may activate NLRP3. Genetic variations in NLRP3-inflammasome components have been reported to influence rheumatoid arthritis (RA) susceptibility and severity. We sought to assess the activity of NLRP3-inflammasome in patients with active RA compared to healthy individuals.MethodIntracellular protein expression of NLRP3, ASC, pro- and active caspase-1, pro- and active IL-1β was assessed by immunoblotting both at baseline and upon inflammasome activation. NLRP3 function (IL-1β secretion) was assessed upon priming of TLR2 (Pam(3)CysSK(4), TLR3 (poly(I:C)) or TLR4 (LPS) and ATP sequential treatment. We used caspase inhibitors (casp-1, 3/7 and 8) to assess their contribution to IL-1β maturation. All experiments were performed in whole blood cells.ResultsActive RA patients (n = 11) expressed higher basal intracellular levels of NLRP3 (p < 0.008), ASC (p < 0.003), active caspase-1 (p < 0.02) and pro-IL-1β (p < 0.001). Upon priming with TLR4 (LPS) and ATP, RA-derived cell extracts (n = 7) displayed increased expression of NLRP3 (p < 0.01) and active caspase-1 (p < 0.001). Secreted IL-1β in culture supernatants from whole blood cells activated with TLR4 (LPS) or TLR3 agonist (poly(I:C)) plus ATP was higher in RA patients (n = 20) versus controls (n = 18) (p < 0.02 for both). Caspase-1 inhibition significantly reduced IL-1β secretion induced by all stimuli, whereas caspase-8 inhibition affected only TLR4 and TLR3 cell priming.ConclusionPatients with active RA have increased expression of NLRP3 and NLRP3-mediated IL-1β secretion in whole blood cells upon stimulation via TLR3 and TLR4 but not TLR2. In these patients, IL-1β secretion seems to be predominately driven by caspase-1 and caspase-8. Targeting NLRP3 or downstream caspases may be of benefit in suppressing IL-1β production in RA.     

A global, myosin light chain kinase-dependent increase in myosin II contractility accompanies the metaphase-anaphase transition in sea urchin eggs

Myosin II is the force-generating motor for cytokinesis, and although it is accepted that myosin contractility is greatest at the cell equator, the temporal and spatial cues that direct equatorial contractility are not known. Dividing sea urchin eggs were placed under compression to study myosin II-based contractile dynamics, and cells manipulated in this manner underwent an abrupt, global increase in cortical contractility concomitant with the metaphase-anaphase transition, followed by a brief relaxation and the onset of furrowing. Prefurrow cortical contractility both preceded and was independent of astral microtubule elongation, suggesting that the initial activation of myosin II preceded cleavage plane specification. The initial rise in contractility required myosin light chain kinase but not Rho-kinase, but both signaling pathways were required for successful cytokinesis. Last, mobilization of intracellular calcium during metaphase induced a contractile response, suggesting that calcium transients may be partially responsible for the timing of this initial contractile event. Together, these findings suggest that myosin II-based contractility is initiated at the metaphase-anaphase transition by Ca2+-dependent myosin light chain kinase (MLCK) activity and is maintained through cytokinesis by both MLCK- and Rho-dependent signaling. Moreover, the signals that initiate myosin II contractility respond to specific cell cycle transitions independently of the microtubule-dependent cleavage stimulus.     

MHCPred: A server for quantitative prediction of peptide-MHC binding

Accurate T-cell epitope prediction is a principal objective of computational vaccinology. As a service to the immunology and vaccinology communities at large, we have implemented, as a server on the World Wide Web, a partial least squares-based multivariate statistical approach to the quantitative prediction of peptide binding to major histocom- patibility complexes (MHC), the key checkpoint on the antigen presentation pathway within adaptive cellular immunity. MHCPred implements robust statistical models for both Class I alleles (HLA-A*0101, HLA-A*0201, HLA-A*0202, HLA-A*0203, HLA-A*0206, HLA-A*0301, HLA-A*1101, HLA-A*3301, HLA-A*6801, HLA-A*6802 and HLA-B*3501) and Class II alleles (HLA-DRB*0401, HLA-DRB*0401 and HLA-DRB*0701). MHCPred is available from the URL: http://www.jenner.ac.uk/MHCPred.