Statins induce regulatory T cell recruitment via a CCL1 dependent pathway

The statins, a group of inhibitors of the 3-hydroxy-3-methylglutaryl coenzyme A reductase, are reported to influence a variety of immune system activities through 3-hydroxy-3-methylglutaryl coenzyme A reductase-dependent and -independent mechanisms. How statin treatment regulates immune system function in vivo nonetheless remains to be fully defined. We analyzed the immunomodulatory effects of lovastatin in a Candida albicans-induced delayed-type hypersensitivity reaction in mice. In this model, lovastatin administration reduced the acute inflammatory response elicited by C. albicans challenge. This anti-inflammatory activity of lovastatin was associated with a shift from a Th1 to a Th2 immune response, as well as an increase in the percentage of regulatory T cells at the inflammation site and in the regional draining lymph node. The lovastatin-induced increase in regulatory T cells in the inflamed skin was dependent on expression of CCL1, a chemokine that is locally up-regulated by statin administration. The anti-inflammatory effect of lovastatin was abrogated in CCL1-deficient mice. These results suggest that local regulation of chemokine expression may be an important process in statin-induced modulation of the immune system.

Visualization of alveolar recruitment in a porcine model of unilateral lung lavage using 3He-MRI

BACKGROUND: In the acute respiratory distress syndrome potentially recruitable lung volume is currently discussed. (3)He-magnetic resonance imaging ((3)He-MRI) offers the possibility to visualize alveolar recruitment directly. METHODS: With the approval of the state animal care committee, unilateral lung damage was induced in seven anesthetized pigs by saline lavage of the right lungs. The left lung served as an intraindividual control (healthy lung). Unilateral lung damage was confirmed by conventional proton MRI and spiral-CT scanning. The total aerated lung volume was determined both at a positive end-expiratory pressure (PEEP) of 0 and 10 mbar from three-dimensionally reconstructed (3)He images, both for healthy and damaged lungs. The fractional increase of aerated volume in damaged and healthy lungs, followed by a PEEP increase from 0 to 10 mbar, was compared. RESULTS: Aerated gas space was visualized with a high spatial resolution in the three-dimensionally reconstructed (3)He-MR images, and aeration defects in the lavaged lung matched the regional distribution of atelectasis in proton MRI. After recruitment and PEEP increase, the aerated volume increased significantly both in healthy lungs from 415 ml [270-445] (median [min-max]) to 481 ml [347-523] and in lavaged lungs from 264 ml [71-424] to 424 ml [129-520]. The fractional increase in lavaged lungs was significantly larger than that in healthy lungs (healthy: 17% [11-38] vs. lavage: 42% [14-90] (P=0.031). CONCLUSION: The (3)He-MRI signal might offer an experimental approach to discriminate atelectatic vs. poor aerated lung areas in a lung damage animal model. Our results confirm the presence of potential recruitable lung volume by either alveolar collapse or alveolar flooding, in accordance with previous reports by computed tomography.

Divergent adaptation of hepatitis C virus genotypes 1 and 3 to human leukocyte antigen-restricted immune pressure

Many hepatitis C virus (HCV) infections worldwide are with the genotype 1 and 3 strains of the virus. Cellular immune responses are known to be important in the containment of HCV genotype 1 infection, and many genotype 1 T cell targets (epitopes) that are presented by host human leukocyte antigens (HLAs) have been identified. In contrast, there is almost no information known about the equivalent responses to genotype 3. Immune escape mechanisms used by HCV include the evolution of viral polymorphisms (adaptations) that abrogate this host-viral interaction. Evidence of HCV adaptation to HLA-restricted immune pressure on HCV can be observed at the population level as viral polymorphisms associated with specific HLA types. To evaluate the escape patterns of HCV genotypes 1 and 3, we assessed the associations between viral polymorphisms and specific HLA types from 187 individuals with genotype 1a and 136 individuals with genotype 3a infection. We identified 51 HLA-associated viral polymorphisms (32 for genotype 1a and 19 for genotype 3a). Of these putative viral adaptation sites, six fell within previously published epitopes. Only two HLA-associated viral polymorphisms were common to both genotypes. In the remaining sites with HLA-associated polymorphisms, there was either complete conservation or no significant HLA association with viral polymorphism in the alternative genotype. This study also highlights the diverse mechanisms by which viral evasion of immune responses may be achieved and the role of genotype variation in these processes. CONCLUSION: There is little overlap in HLA-associated polymorphisms in the nonstructural proteins of HCV for the two genotypes, implying differences in the cellular immune pressures acting on these viruses and different escape profiles. These findings have implications for future therapeutic strategies to combat HCV infection, including vaccine design.

Inhibition of Notch signaling induces extensive intussusceptive neo-angiogenesis by recruitment of mononuclear cells

Notch is an intercellular signaling pathway related mainly to sprouting neo-angiogenesis. The objective of our study was to evaluate the angiogenic mechanisms involved in the vascular augmentation (sprouting/intussusception) after Notch inhibition within perfused vascular beds using the chick area vasculosa and MxCreNotch1(lox/lox) mice. In vivo monitoring combined with morphological investigations demonstrated that inhibition of Notch signaling within perfused vascular beds remarkably induced intussusceptive angiogenesis (IA) with resultant dense immature capillary plexuses. The latter were characterized by 40 % increase in vascular density, pericyte detachment, enhanced vessel permeability, as well as recruitment and extravasation of mononuclear cells into the incipient transluminal pillars (quintessence of IA). Combination of Notch inhibition with injection of bone marrow-derived mononuclear cells dramatically enhanced IA with 80 % increase in vascular density and pillar number augmentation by 420 %. Additionally, there was down-regulation of ephrinB2 mRNA levels consequent to Notch inhibition. Inhibition of ephrinB2 or EphB4 signaling induced some pericyte detachment and resulted in up-regulation of VEGFRs but with neither an angiogenic response nor recruitment of mononuclear cells. Notably, Tie-2 receptor was down-regulated, and the chemotactic factors SDF-1/CXCR4 were up-regulated only due to the Notch inhibition. Disruption of Notch signaling at the fronts of developing vessels generally results in massive sprouting. On the contrary, in the already existing vascular beds, down-regulation of Notch signaling triggered rapid augmentation of the vasculature predominantly by IA. Notch inhibition disturbed vessel stability and led to pericyte detachment followed by extravasation of mononuclear cells. The mononuclear cells contributed to formation of transluminal pillars with sustained IA resulting in a dense vascular plexus without concomitant vascular remodeling and maturation.

Increased vascularization during early healing after biologic augmentation in repair of chronic rotator cuff tears using autologous leukocyte- and platelet-rich fibrin (L-PRF): a prospective randomized controlled pilot trial

HYPOTHESIS We hypothesized that arthroscopic rotator cuff repairs using leukocyte- and platelet-rich fibrin (L-PRF) in a standardized, modified protocol is technically feasible and results in a higher vascularization response and watertight healing rate during early healing. METHODS Twenty patients with chronic rotator cuff tears were randomly assigned to 2 treatment groups. In the test group (N = 10), L-PRF was added in between the tendon and the bone during arthroscopic rotator cuff repair. The second group served as control (N = 10). They received the same arthroscopic treatment without the use of L-PRF. We used a double-row tension band technique. Clinical examinations including subjective shoulder value, visual analog scale, Constant, and Simple Shoulder Test scores and measurement of the vascularization with power Doppler ultrasonography were made at 6 and 12 weeks. RESULTS There have been no postoperative complications. At 6 and 12 weeks, there was no significant difference in the clinical scores between the test and the control groups. The mean vascularization index of the surgical tendon-to-bone insertions was always significantly higher in the L-PRF group than in the contralateral healthy shoulders at 6 and 12 weeks (P = .0001). Whereas the L-PRF group showed a higher vascularization compared with the control group at 6 weeks (P = .001), there was no difference after 12 weeks of follow-up (P = .889). Watertight healing was obtained in 89% of the repaired cuffs. DISCUSSION/CONCLUSIONS Arthroscopic rotator cuff repair with the application of L-PRF is technically feasible and yields higher early vascularization. Increased vascularization may potentially predispose to an increased and earlier cellular response and an increased healing rate.

DAPK2 positively regulates motility of neutrophils and eosinophils in response to intermediary chemoattractants

The tight regulation of granulocyte chemotaxis is crucial for initiation and resolution of inflammation. Here, we show that DAPK2, a Ca(2+)/CaM-sensitive serine/threonine kinase known to modulate cell death in various cell types, is a novel regulator of migration in granulocytes. We demonstrate that human neutrophils and eosinophils express DAPK2 but unlike other leukocytes, no DAPK1 or DAPK3 protein. When DAPK activities were blocked by inhibitors, we found that neither granulocyte lifespan nor phagocytosis was affected. However, such pharmacological inactivation of DAPK activity abolished motility of granulocytes in response to intermediary but not end-target chemoattractants ex vivo. The defect in chemotaxis in DAPK2-inactive granulocytes is likely a result of reduced polarization of the cells, mediated by a lack of MLC phosphorylation, resulting in radial F-actin and pseudopod formation. As neutrophils treated with DAPKi also showed reduced recruitment to the site of inflammation in a mouse peritonitis model, DAPK2 may be a novel target for anti-inflammatory therapies.

Recruitment of EB1, a master regulator of microtubule dynamics, to the surface of the Theileria annulata schizont

The apicomplexan parasite Theileria annulata transforms infected host cells, inducing uncontrolled proliferation and clonal expansion of the parasitized cell population. Shortly after sporozoite entry into the target cell, the surrounding host cell membrane is dissolved and an array of host cell microtubules (MTs) surrounds the parasite, which develops into the transforming schizont. The latter does not egress to invade and transform other cells. Instead, it remains tethered to host cell MTs and, during mitosis and cytokinesis, engages the cell's astral and central spindle MTs to secure its distribution between the two daughter cells. The molecular mechanism by which the schizont recruits and stabilizes host cell MTs is not known. MT minus ends are mostly anchored in the MT organizing center, while the plus ends explore the cellular space, switching constantly between phases of growth and shrinkage (called dynamic instability). Assuming the plus ends of growing MTs provide the first point of contact with the parasite, we focused on the complex protein machinery associated with these structures. We now report how the schizont recruits end-binding protein 1 (EB1), a central component of the MT plus end protein interaction network and key regulator of host cell MT dynamics. Using a range of in vitro experiments, we demonstrate that T. annulata p104, a polymorphic antigen expressed on the schizont surface, functions as a genuine EB1-binding protein and can recruit EB1 in the absence of any other parasite proteins. Binding strictly depends on a consensus SxIP motif located in a highly disordered C-terminal region of p104. We further show that parasite interaction with host cell EB1 is cell cycle regulated. This is the first description of a pathogen-encoded protein to interact with EB1 via a bona-fide SxIP motif. Our findings provide important new insight into the mode of interaction between Theileria and the host cell cytoskeleton.

Slit2 prevents neutrophil recruitment and renal ischemia-reperfusion injury.

Neutrophils recruited to the postischemic kidney contribute to the pathogenesis of ischemia-reperfusion injury (IRI), which is the most common cause of renal failure among hospitalized patients. The Slit family of secreted proteins inhibits chemotaxis of leukocytes by preventing activation of Rho-family GTPases, suggesting that members of this family might modulate the recruitment of neutrophils and the resulting IRI. Here, in static and microfluidic shear assays, Slit2 inhibited multiple steps required for the infiltration of neutrophils into tissue. Specifically, Slit2 blocked the capture and firm adhesion of human neutrophils to inflamed vascular endothelial barriers as well as their subsequent transmigration. To examine whether these observations were relevant to renal IRI, we administered Slit2 to mice before bilateral clamping of the renal pedicles. Assessed at 18 hours after reperfusion, Slit2 significantly inhibited renal tubular necrosis, neutrophil and macrophage infiltration, and rise in plasma creatinine. In vitro, Slit2 did not impair the protective functions of neutrophils, including phagocytosis and superoxide production, and did not inhibit neutrophils from killing the extracellular pathogen Staphylococcus aureus. In vivo, administration of Slit2 did not attenuate neutrophil recruitment or bacterial clearance in mice with ascending Escherichia coli urinary tract infections and did not increase the bacterial load in the livers of mice infected with the intracellular pathogen Listeria monocytogenes. Collectively, these results suggest that Slit2 may hold promise as a strategy to combat renal IRI without compromising the protective innate immune response.