Collagen type II and a thermo-responsive polymer of N-isopropylacrylamide induce arthritis independent of Toll-like receptors: a strong influence by major histocompatibility complex class II and Ncf1 genes

We established and characterized an arthritis mouse model using collagen type II (CII) and a thermo-responsive polymer, poly(N-isopropylacrylamide) (PNiPAAm). The new PNiPAAm adjuvant is TLR-independent, as all immunized TLR including MyD88-deficient mice developed an anti-CII response. Unlike other adjuvants, PNiPPAm did not skew the cytokine response (IL-1β, IFN-γ, IL-4, and IL-17), as there was no immune deviation towards any one type of immune spectrum after immunization with CII/PNiPPAm. Hence, using PNiPAAm, we studied the actual immune response to the self-protein, CII. We observed arthritis and autoimmunity development in several murine strains having different major histocompatibility complex (MHC) haplotypes after CII/PNiPAAm immunization but with a clear MHC association pattern. Interestingly, C57Bl/6 mice did not develop CII-induced arthritis, with PNiPAAm demonstrating absolute requirement for a classical adjuvant. Presence of a gene (Ncf1) mutation in the NADPH oxidation complex has a profound influence in arthritis and using PNiPAAm we could show that the high CIA severity in Ncf1 mutated mice is independent of any classical adjuvant. Macrophages, neutrophils, eosinophils, and osteoclasts but not mast cells dominated the inflamed joints. Furthermore, arthritis induction in the adjuvant-free, eosinophil-dependent Vβ12 DBA/1 mice could be shown to develop arthritis independent of eosinophils using CII/PNiPAAm. Thus, biocompatible and biodegradable PNiPAAm offers unique opportunities to study actual autoimmunity independent of TLR and a particular cytokine phenotype profile.     

A small molecule deubiquitinase inhibitor increases localization of inducible nitric oxide synthase to the macrophage phagosome and enhances bacterial killing

Macrophages are key mediators of antimicrobial defense and innate immunity. Innate intracellular defense mechanisms can be rapidly regulated at the posttranslational level by the coordinated addition and removal of ubiquitin by ubiquitin ligases and deubiquitinases (DUBs). While ubiquitin ligases have been extensively studied, the contribution of DUBs to macrophage innate immune function is incompletely defined. We therefore employed a small molecule DUB inhibitor, WP1130, to probe the role of DUBs in the macrophage response to bacterial infection. Treatment of activated bone marrow-derived macrophages (BMM) with WP1130 significantly augmented killing of the intracellular bacterial pathogen Listeria monocytogenes. WP1130 also induced killing of phagosome-restricted bacteria, implicating a bactericidal mechanism associated with the phagosome, such as the inducible nitric oxide synthase (iNOS). WP1130 had a minimal antimicrobial effect in macrophages lacking iNOS, indicating that iNOS is an effector mechanism for WP1130-mediated bacterial killing. Although overall iNOS levels were not notably different, we found that WP1130 significantly increased colocalization of iNOS with the Listeria-containing phagosome during infection. Taken together, our data indicate that the deubiquitinase inhibitor WP1130 increases bacterial killing in macrophages by enhancing iNOS localization to the phagosome and suggest a potential role for ubiquitin regulation in iNOS trafficking.     

Destabilization of YopE by the ubiquitin-proteasome pathway fine-tunes Yop delivery into host cells and facilitates systemic spread of Yersinia enterocolitica in host lymphoid tissue

Pathogenic Yersinia species inject a panel of Yop virulence proteins by type III protein secretion into host cells to modulate cellular defense responses. This enables the survival and dissemination of the bacteria in the host lymphoid tissue. We have previously shown that YopE of the Y. enterocolitica serogroup O8 is degraded in the host cell through the ubiquitin-proteasome pathway. YopE normally manipulates rearrangements of the actin cytoskeleton and triggers phagocytosis resistance. To shed light into the physiological role of YopE inactivation, we mutagenized the lysine polyubiquitin acceptor sites of YopE in the Y. enterocolitica serogroup O8 virulence plasmid. The resulting mutant strain escaped polyubiquitination and degradation of YopE and displayed increased intracellular YopE levels, which was accompanied by a pronounced cytotoxic effect on infected cells. Despite its intensified activity on cultured cells, the Yersinia mutant with stabilized YopE showed reduced dissemination into liver and spleen following enteral infection of mice. Furthermore, the accumulation of degradation-resistant YopE was accompanied by the diminished delivery of YopP and YopH into cultured, Yersinia-infected cells. A role of YopE in the regulation of Yop translocation has already been described. Our results imply that the inactivation of YopE by the proteasome could be a tool to ensure intermediate intracellular YopE levels, which may effectuate optimized Yop injection into host cells. In this regard, Y. enterocolitica O8 appears to exploit the host ubiquitin proteasome system to destabilize YopE and to fine-tune the activities of the Yop virulence arsenal on the infected host organism.     

Unique characteristics of motor adaptation during walking in young children

Children show precocious ability in the learning of languages; is this the case with motor learning? We used split-belt walking to probe motor adaptation (a form of motor learning) in children. Data from 27 children (ages 8-36 mo) were compared with those from 10 adults. Children walked with the treadmill belts at the same speed (tied belt), followed by walking with the belts moving at different speeds (split belt) for 8-10 min, followed again by tied-belt walking (postsplit). Initial asymmetries in temporal coordination (i.e., double support time) induced by split-belt walking were slowly reduced, with most children showing an aftereffect (i.e., asymmetry in the opposite direction to the initial) in the early postsplit period, indicative of learning. In contrast, asymmetries in spatial coordination (i.e., center of oscillation) persisted during split-belt walking and no aftereffect was seen. Step length, a measure of both spatial and temporal coordination, showed intermediate effects. The time course of learning in double support and step length was slower in children than in adults. Moreover, there was a significant negative correlation between the size of the initial asymmetry during early split-belt walking (called error) and the aftereffect for step length. Hence, children may have more difficulty learning when the errors are large. The findings further suggest that the mechanisms controlling temporal and spatial adaptation are different and mature at different times.     

Implementation and workflow for PET monitoring of therapeutic ion irradiation: a comparison of in-beam, in-room, and off-line techniques

An independent assessment of the dose delivery in ion therapy can be performed using positron emission tomography (PET). For that a distribution of positron emitters which appear as the result of interaction between ions of the therapeutic beam and the irradiated tissue is measured during or after the irradiation. Three concepts for PET monitoring implemented in various therapy facilities are considered in this paper. The in-beam PET concept relies on the PET measurement performed simultaneously to the irradiation by means of a PET scanner which is completely integrated into the irradiation site. The in-room PET concept allows measurement immediately after irradiation by a standalone PET scanner which is installed very close to the irradiation site. In the off-line PET scenario the measurement is performed by means of a standalone PET/CT scanner 10-30 min after the irradiation. These three concepts were evaluated according to image quality criteria, integration costs, and their influence onto the workflow of radiotherapy. In-beam PET showed the best performance. However, the integration costs were estimated as very high for this modality. Moreover, the performance of in-beam PET depends heavily on type and duty cycle of the accelerator. The in-room PET is proposed for planned therapy facilities as a good compromise between the quality of measured data and integration efforts. For facilities which are close to the nuclear medicine departments off-line PET can be suggested under several circumstances.     

Intraoperative cerebral high-intensity transient signals and postoperative cognitive function: a systematic review

Background

Much attention in the literature has focused on the relationship between perioperative microemboli during cardiac and vascular surgery and postoperative cognitive decline. Transcranial Doppler ultrasonography (TCD) has been used to measure high-intensity transient signals (HITS), which represent microemboli during cardiac, vascular, and orthopedic surgery. The purpose of this study was to systematically examine the literature with respect to HITS and postoperative cognitive function.

Methods

Systematic PubMed searches identified articles related to the use of TCD and cognitive function in the surgical setting.

Results

The literature remains largely undecided on the role of HITS and cognitive impairment after surgery, with most studies being underpowered to show a relationship. Although the cognitive effects of HITS may be difficult to detect, subclinical microemboli present potential harm, which may be modifiable.

Conclusions

TCD represents a tool for intraoperative cerebral monitoring to reduce the number of HITS during surgery.     

Effect of fiber distribution and realignment on the nonlinear and inhomogeneous mechanical properties of human supraspinatus tendon under longitudinal tensile loading

Tendon exhibits nonlinear stress–strain behavior that may be partly due to movement of collagen fibers through the extracellular matrix. While a few techniques have been developed to evaluate the fiber architecture of other soft tissues, the organizational behavior of tendon under load has not been determined. The supraspinatus tendon (SST) of the rotator cuff is of particular interest for investigation due to its complex mechanical environment and corresponding inhomogeneity. In addition, SST injury occurs frequently with limited success in treatment strategies, illustrating the need for a better understanding of SST properties. Therefore, the objective of this study was to quantitatively evaluate the inhomogeneous tensile mechanical properties, fiber organization, and fiber realignment under load of human SST utilizing a novel polarized light technique. Fiber distributions were found to become more aligned under load, particularly during the low stiffness toe‐region, suggesting that fiber realignment may be partly responsible for observed nonlinear behavior. Fiber alignment was found to correlate significantly with mechanical parameters, providing evidence for strong structure–function relationships in tendon. Human SST exhibits complex, inhomogeneous mechanical properties and fiber distributions, perhaps due to its complex loading environment. Surprisingly, histological grade of degeneration did not correlate with mechanical properties. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27:1596–1602, 2009