Site of Blood Vessel Damage and Relevance of CD18 in a Murine Model of Immune Complex-Mediated Vasculitis

How neutrophils (polymorphonuclear neutrophils, PMNs) damage vessels in leukocytoclastic vasculitis (LcV) mediated by immune complexes (ICs) is unclear. If degradative enzymes and oxygen radicals are released from PMNs while adhering to the inner side of the vessel wall, they could be washed away by the blood stream or neutralized by serum protease inhibitors. We investigated if in LcV PMNs could damage vessels from the tissue side after transmigration. We used CD18-deficient (CD18−/−) mice because the absence of CD18 excludes transmigration of PMNs. When eliciting the Arthus reaction in ears of CD18−/− mice, deposition of ICs was not sufficient to recruit PMNs or to induce IC-mediated LcV. Injection of PMNs intradermally in CD18−/− mice allowed us to investigate if bypassing diapedesis and placing PMNs exclusively on the abluminal side leads to vascular destruction. We found that injected PMNs gathered around perivascular ICs, but did not cause vessel damage. Only intravenous injection of wild-type PMNs could re-establish the Arthus reaction in CD18−/− mice. Thus, PMNs cause vessel damage during diapedesis from the luminal side, but not from the perivascular space. We suggest that in order to shield the cytotoxic products from the blood stream, ICs induce particularly tight interactions between them, PMNs and endothelial cells

TFIIH mutations can impact on translational fidelity of the ribosome

TFIIH is a complex essential for transcription of protein-coding genes by RNA polymerase II, DNA repair of UV-lesions and transcription of rRNA by RNA polymerase I. Mutations in TFIIH cause the cancer prone DNA-repair disorder xeroderma pigmentosum (XP) and the developmental and premature aging disorders trichothiodystrophy (TTD) and Cockayne syndrome (CS). 50% of TTD cases are caused by TFIIH mutations. Using TFIIH mutant patient cells from TTD and XP subjects we can show that the stress-sensitivity of the proteome is reduced in TTD, but not in XP. Using three different methods to investigate the accuracy of protein synthesis by the ribosome, we demonstrate that translational fidelity of the ribosomes of TTD, but not XP cells, is decreased. The process of ribosomal synthesis and maturation is affected in TTD cells and can lead to instable ribosomes. Isolated ribosomes from TTD patients show an elevated error rate when challenged with oxidized mRNA, explaining the oxidative hypersensitivity of TTD cells. Treatment of TTD cells with N-acetyl cysteine normalized the increased translational error-rate and restored translational fidelity. Here we describe a pathomechanism that might be relevant for our understanding of impaired development and aging-associated neurodegeneration

Loss of Proteostasis Is a Pathomechanism in Cockayne Syndrome

Retarded growth and neurodegeneration are hallmarks of the premature aging disease Cockayne syndrome (CS). Cockayne syndrome proteins take part in the key step of ribosomal biogenesis, transcription of RNA polymerase I. Here, we identify a mechanism originating from a disturbed RNA polymerase I transcription that impacts translational fidelity of the ribosomes and consequently produces misfolded proteins. In cells from CS patients, the misfolded proteins are oxidized by the elevated reactive oxygen species (ROS) and provoke an unfolded protein response that represses RNA polymerase I transcription. This pathomechanism can be disrupted by the addition of pharmacological chaperones, suggesting a treatment strategy for CS. Additionally, this loss of proteostasis was not observed in mouse models of CS. Cockayne syndrome is a devastating childhood progeria. Here, Alupei et al. show that cells from CS patients have reduced translation accuracy and elevated ROS, leading to generation of unstable proteins and activation of ER stress. Reducing ER stress by chemical chaperones in these cells rescues RNA polymerase I activity and protein synthesis

Role of β2-integrins for homing and neovascularization capacity of endothelial progenitor cells

The mechanisms of homing of endothelial progenitor cells (EPCs) to sites of ischemia are unclear. Here, we demonstrate that ex vivo–expanded EPCs as well as murine hematopoietic Sca-1+/Lin− progenitor cells express β2-integrins, which mediate the adhesion of EPCs to endothelial cell monolayers and their chemokine-induced transendothelial migration in vitro. In a murine model of hind limb ischemia, Sca-1+/Lin− hematopoietic progenitor cells from β2-integrin–deficient mice are less capable of homing to sites of ischemia and of improving neovascularization. Preactivation of the β2-integrins expressed on EPCs by activating antibodies augments the EPC-induced neovascularization in vivo. These results provide evidence for a novel function of β2-integrins in postnatal vasculogenesis

Adaptive Immune Response to Model Antigens Is Impaired in Murine Leukocyte-Adhesion Deficiency-1 Revealing Elevated Activation Thresholds In Vivo

Absence of β2 integrins (CD11/CD18) leads to leukocyte-adhesion deficiency-1 (LAD1), a rare primary immunodeficiency syndrome. Although extensive in vitro work has established an essential function of β2 integrins in adhesive and signaling properties for cells of the innate and adaptive immune system, their respective participation in an altered adaptive immunity in LAD1 patients are complex and only partly understood in vivo. Therefore, we investigated adaptive immune responses towards different T-dependent antigens in a murine LAD1 model of β2 integrin-deficiency (CD18−/−). CD18−/− mice generated only weak IgG responses after immunization with tetanus toxoid (TT). In contrast, robust hapten- and protein-specific immune responses were observed after immunization with highly haptenated antigens such as (4-hydroxy-3-nitrophenyl)21 acetyl chicken γ globulin (NP21-CG), even though regularly structured germinal centers with specificity for the defined antigens/haptens in CD18−/− mice remained absent. However, a decrease in the hapten/protein ratio lowered the efficacy of immune responses in CD18−/− mice, whereas a mere reduction of the antigen dose was less crucial. Importantly, haptenation of TT with NP (NP-TT) efficiently restored a robust IgG response also to TT. Our findings may stimulate further studies on a modification of vaccination strategies using highly haptenated antigens in individuals suffering from LAD1

The proangiogenic capacity of polymorphonuclear neutrophils delineated by microarray technique and by measurement of neovascularization in wounded skin of CD18-deficient mice

Growing evidence supports the concept that polymorphonuclear neutrophils (PMN) are critically involved in inflammation-mediated angiogenesis which is important for wound healing and repair. We employed an oligonucleotide microarray technique to gain further insight into the molecular mechanisms underlying the proangiogenic potential of human PMN. In addition to 18 known angiogenesis-relevant genes, we detected the expression of 10 novel genes, namely midkine, erb-B2, ets-1, transforming growth factor receptor-beta(2) and -beta(3), thrombospondin, tissue inhibitor of metalloproteinase 2, ephrin A2, ephrin B2 and restin in human PMN freshly isolated from the circulation. Gene expression was confi rmed by the RT-PCR technique. In vivo evidence for the role of PMN in neovascularization was provided by studying neovascularization in a skin model of wound healing using CD18-deficient mice which lack PMN infi ltration to sites of lesion. In CD18-deficient animals, neo- vascularization was found to be signifi cantly compromised when compared with wild- type control animals which showed profound neovascularization within the granulation tissue during the wound healing process. Thus, PMN infiltration seems to facilitate inflammation mediated angiogenesis which may be a consequence of the broad spectrum of proangiogenic factors expressed by these cells. Copyright (c) 2006 S. Karger AG, Basel