Progenitor Hematopoietic Cells Implantation Improves Functional Capacity of End Stage Coronary Artery Disease Patients with Advanced Heart Failure.

Background. Proangiogenic Hematopoietic Cells (PHC) which comprise diverse mixture of cell types are able to secrete proangiogenic factors and interesting candidate for cell therapy. The aim of this study was to seek for benefit in implantation of PHC on functional improvement in end stage coronary artery disease patients with advanced heart failure. Methods. Patients with symptomatic heart failure despite guideline directed medical therapy and LVEF less than 35% were included. Peripheral blood mononuclear cells were isolated, cultivated for 5 days, and then harvested. Flow cytometry and cell surface markers were used to characterize PHC. The PHC were delivered retrogradely via sinus coronarius. Echocardiography, myocardial perfusion, and clinical and functional data were analyzed up to 1-year observation. Results. Of 30 patients (56.4 ± 7.40 yo) preimplant NT proBNP level is 5124.5 ± 4682.50 pmol/L. Harvested cells characterized with CD133, CD34, CD45, and KDR showed 0.87 ± 0.41, 0.63 ± 0.66, 99.00 ± 2.60, and 3.22 ± 3.79%, respectively. LVEF was improved (22 ± 5.68 versus 26.8 ± 7.93, p < 0.001) during short and long term observation. Myocardial perfusion significantly improved 6 months after treatment. NYHA Class and six-minute walk test are improved during short term and long term follow-up. Conclusion. Expanded peripheral blood PHC implantation using retrograde delivery approach improved LV systolic function, myocardial perfusion, and functional capacity.

Structural basis for Fc gammaRIIa recognition of human IgG and formation of inflammatory signaling complexes.

The interaction of Abs with their specific FcRs is of primary importance in host immune effector systems involved in infection and inflammation, and are the target for immune evasion by pathogens. FcγRIIa is a unique and the most widespread activating FcR in humans that through avid binding of immune complexes potently triggers inflammation. Polymorphisms of FcγRIIa (high responder/low responder [HR/LR]) are linked to susceptibility to infections, autoimmune diseases, and the efficacy of therapeutic Abs. In this article, we define the three-dimensional structure of the complex between the HR (arginine, R134) allele of FcγRIIa (FcγRIIa-HR) and the Fc region of a humanized IgG1 Ab, hu3S193. The structure suggests how the HR/LR polymorphism may influence FcγRIIa interactions with different IgG subclasses and glycoforms. In addition, mutagenesis defined the basis of the epitopes detected by FcR blocking mAbs specific for FcγRIIa (IV.3), FcγRIIb (X63-21), and a pan FcγRII Ab (8.7). The epitopes detected by these Abs are distinct, but all overlap with residues defined by crystallography to contact IgG. Finally, crystal structures of LR (histidine, H134) allele of FcγRIIa and FcγRIIa-HR reveal two distinct receptor dimers that may represent quaternary states on the cell surface. A model is presented whereby a dimer of FcγRIIa-HR binds Ag-Ab complexes in an arrangement that possibly occurs on the cell membrane as part of a larger signaling assembly.

Inhibition of destructive autoimmune arthritis in FcgammaRIIa transgenic mice by small chemical entities.

The interaction of immune complexes with the human Fc receptor, FcgammaRIIa, initiates the release of inflammatory mediators and is implicated in the pathogenesis of human autoimmune diseases, including rheumatoid arthritis and systemic lupus erythematosus, so this FcR is a potential target for therapy. We have used the three-dimensional structure of an FcgammaRIIa dimer to design small molecule inhibitors, modeled on a distinct groove and pocket created by receptor dimerization, adjacent to the ligand-binding sites. These small chemical entities (SCEs) blocked immune complex-induced platelet activation and aggregation and tumor necrosis factor secretion from macrophages in a human cell line and transgenic mouse macrophages. The SCE appeared specific for FcgammaRIIa, as they inhibited only immune complex-induced responses and had no effect on responses to stimuli unrelated to FcR, for example platelet stimulation with arachidonic acid. In vivo testing of the SCE in FcgammaRIIa transgenic mice showed that they inhibited the development and stopped the progression of collagen-induced arthritis (CIA). The SCEs were more potent than methotrexate and anti-CD3 in sustained suppression of CIA. Thus, in vitro and in vivo activity of these SCE FcgammaRIIa receptor antagonists demonstrated their potential as anti-inflammatory agents for autoimmune diseases involving immune complexes.

Palmitoylation at Cys595 is essential for PECAM-1 localisation into membrane microdomains and for efficient PECAM-1-mediated cytoprotection.

The Ig-ITIM superfamily member, PECAM-1 acts as a negative regulator of ITAM-signalling pathways in platelets involving GPVI/FcR gamma chain and Fc?RIIa. This negative feedback loop involves regulation of collagen and GPVI-dependent aggregation events, platelet-thrombus-growth on immobilised collagen under flow and Fc?RIIa-mediated platelet responses. In this study, we show that PECAM-1 is selectively palmitoylated involving a thioester linkage with an unpaired cysteine residue at amino acid position 595 in its cytoplasmic domain. As palmitoylation is known to target proteins to membrane microdomains, we investigated the microdomain localisation for PECAM-1 in platelets and nucleated cells. In unstimulated platelets, approximately 20% of PECAM-1 is localised to Triton-insoluble microdomain fractions and it does not increase with platelet activation by collagen, collagen-related peptide, thrombin- or human-aggregated IgG. PECAM-1 is in close physical proximity with GPVI in platelet microdomains. Removal of platelet cytoskeleton prior to sucrose-density-gradient separation showed that PECAM-1 was associated with both the Triton-soluble and membrane skeleton in microdomain-associated fractions. Disruption of microdomains by membrane-cholesterol depletion resulted in loss of PECAM-1 localisation to membrane microdomains. Mutational analysis of juxtamembrane cysteine residue to alanine (C595A) of human PECAM-1 resulted in loss of palmitoylation and a sixfold decrease in association with membrane microdomains. Functionally, the palmitoylated cysteine 595 residue is required, in part, for efficient PECAM-1-mediated cytoprotection. These results show that cysteine 595 is required for constitutive association of PECAM-1 with membrane microdomains and PECAM-1-mediated cytoprotection, where it may act as a crucial regulator of signaling and apoptosis events.

Development of spontaneous multisystem autoimmune disease and hypersensitivity to antibody-induced inflammation in Fcgamma receptor IIa-transgenic mice.

OBJECTIVE: The major human Fc receptor, FcgammaRIIa, is the most widespread activating FcR. Our aim was to determine the role of FcgammaRIIa in a transgenic mouse model of immune complex-mediated autoimmunity and to characterize the development of spontaneous autoimmune disease. METHODS: Arthritis was induced in normal and FcgammaRIIa-transgenic mice by immunization with type II collagen (CII) or by transfer of arthritogenic anti-CII antibodies. Also, mice that spontaneously developed autoimmune disease were assessed by clinical scoring of affected limbs, histology and serology, and measurement of autoantibody titers and cytokine production. RESULTS: FcgammaRIIa-transgenic mice developed collagen-induced arthritis (CIA) more rapidly than did archetypal CIA-sensitive DBA/1 (H-2q) mice, while nontransgenic C57BL/6 (H-2b) mice did not develop CIA when similarly immunized. Passive transfer of a single dose of anti-CII antibody induced a more rapid, severe arthritis in FcgammaRIIa-transgenic mice than in nontransgenic animals. In addition, most immune complex-induced production of tumor necrosis factor alpha by activated macrophages occurred via FcgammaRIIa, not the endogenous mouse FcR. A spontaneous, multisystem autoimmune disease developed in aging (>20 weeks) transgenic mice (n = 25), with a 32% incidence of arthritis, and by 45 weeks, all mice had developed glomerulonephritis and pneumonitis, and most had antihistone antibodies. Elevated IgG2a levels were seen in mice with CIA and in those with spontaneous disease. CONCLUSION: The presence of enhanced passive and induced autoimmunity, as well as the emergence of spontaneous autoimmune disease at 20-45 weeks of age, suggest that FcgammaRIIa is a very important factor in the pathogenesis of autoimmune inflammation and a possible target for therapeutic intervention.

The role of FcgammaRIIa as an inflammatory mediator in rheumatoid arthritis and systemic lupus erythematosus.

Despite their essential role in host protection, immunoglobulins are also involved in autoimmune processes where antibodies recognize the host's own tissue, triggering inflammatory responses that result in extensive tissue damage. A complex interaction of genetic predisposition, together with environment factors, is thought to trigger immune dysfunction. Although recent studies have dissected the essential role of Fc receptors in autoimmune antibody mediated processes, the uniquely human FcgammaRIIa has not been studied in detail. This Fc receptor is of particular interest, as it is the most abundantly expressed Fc receptor in humans and is implicated in immune complex disease. Investigation of its role has been hampered to date due to lack of suitable animal models. This review examines the evidence for the direct role of this receptor in diseases such as systemic lupus erythematosus and rheumatoid arthritis.