This review focuses on the analysis and evaluation of the diverse senescence markers suggested to prime red blood cells (RBC) for clearance in humans. These tags develop in the course of biochemical ...and structural alterations accompanying RBC aging, as the decrease of activities of multiple enzymes, the gradual accumulation of oxidative damage, the loss of membrane in form of microvesicles, the redistribution of ions and alterations in cell volume, density, and deformability. The actual tags represent the penultimate galactosyl residues, revealed by desialylation of glycophorins, or the aggregates of the anion exchanger (band 3 protein) to which anti-galactose antibodies bind in the first and anti-band 3 naturally occurring antibodies (NAbs) in the second case. While anti-band 3 NAbs bind to the carbohydrate-free portion of band 3 aggregates in healthy humans, induced anti-lactoferrin antibodies bind to the carbohydrate-containing portion of band 3 and along with anti-band 3 NAbs may accelerated clearance of senescent RBC in patients with anti-neutrophil cytoplasmic antibodies (ANCA). Exoplasmically accessible phosphatidylserine (PS) and the alterations in the interplay between CD47 on RBC and its receptor on macrophages, signal regulatory protein alpha (SIRPalpha protein), were also reported to induce erythrocyte clearance. We discuss the relevance of each mechanism and analyze the strength of the data.
In addition to transporting oxygen and carbon dioxide to and from the tissues, a range of other functions are attributed to red blood cells (RBCs) of vertebrates. Diseases compromising RBC ...performance in any of these functions warrant in-depth study. Furthermore, the human RBC is a vital host cell for the malaria parasite. Much has been learned from classical biochemical approaches about RBC composition and membrane organization. Here, we use mass spectrometry (MS)–based proteomics to characterize the normal RBC protein profile. The aim of this study was to obtain the most complete and informative human RBC proteome possible by combining high-accuracy, high-sensitivity protein identification technology (quadrupole time of flight and Fourier transform MS) with selected biochemical procedures for sample preparation. A total of 340 membrane proteins and 252 soluble proteins were identified, validated, and categorized in terms of subcellular localization, protein family, and function. Splice isoforms of proteins were identified, and polypeptides that migrated with anomalously high or low apparent molecular weights could be grouped into either ubiquitinylated, partially degraded, or ester-linked complexes. Our data reveal unexpected complexity of the RBC proteome, provide a wealth of data on its composition, shed light on several open issues in RBC biology, and form a departure point for comprehensive understanding of RBC functions.
Abstract Immunoglobulins may have been developed in evolution to provide specificity for clearing body waste in the first animals with three germ layers. Tissue homeostasis in vertebrates comprises ...clearance of proteins released from lysed cells, elimination of altered plasma proteins, of senescent and apoptotic cells. Rather specific IgM and IgG naturally occurring antibodies (NAbs) to cytoplasmic and cytoskeletal proteins bind to proteins released from lysing cells and the IgG NAbs are slightly upregulated upon demand. Some of these NAbs along with complement have devastating effects when massive amounts of intracellular proteins are released during an infarct or an ischemia/reperfusion experiment. IgM NAbs to neoepitopes on plasma proteins/lipids help clear denatured proteins and are protective. IgG NAbs to an exposed protein, band 3 from red blood cells, bind to oligomerized band 3 and due to an affinity for C3 within their framework preferentially form C3b2–IgG complexes from nascent C3b. Thus, anti-band 3 NAbs gain potency by using avidity and generating a potent precursor of the amplifying C3 convertase. IgM NAbs to neoepitopes, which are generated by oxidized lipids forming Schiff bases with proteins, are protective and help clear this waste in atherosclerosis, but IgG antibodies (NAbs?) of the same specificity promote disease.
Antibodies with germline or close to germline configuration exist in vertebrates, and these so-called ‘naturally occurring auto-antibodies’ (NAb) are directed to self and altered self components. ...Such NAbs have been attracting increasing interest because several of them, including some in their recombinant forms, have therapeutic potential. Whereas a large number of IgM and IgG NAbs have tissue homeostatic roles, others modulate and regulate cellular and enzyme properties. This review describes some of these NAbs and emphasizes how these low-titer, low-affinity NAbs interact with self and altered self and show functional potency in homeostasis and regulation, in addition to in diseases such as infarction and systemic inflammatory response syndrome.
Naturally occurring anti-band 3 antibodies (anti-band 3 NAbs) are directed against the 55-kDa chymotryptic fragment of the anion transport protein (band 3) of red blood cells (RBCs). They bind to ...senescent and oxidatively stressed RBCs and induce their selective clearance. These IgG NAbs exist at low concentrations, and have a weak affinity that prevents them from actively recruiting second binding sites. Cellular senescence or oxidative damage induces a cascade of biochemical events that results in the detachment of band 3 from the cytoskeleton and in clustering of band 3 protein by bound hemichromes and Syk kinase. Clustered band 3 proteins allow bivalent binding of anti-band 3 NAbs. Bivalently bound anti-band 3 NAbs have the unique capacity to stimulate C3b deposition by preferentially generating C3b2-IgG complexes, which act as potent C3 convertase precursors of the alternative complement pathway. Antibody binding not only to clustered, but also to oligomerized band 3 protein further increases if the human plasma also contains induced anti-lactoferrin antibodies. These bind to the polylactosaminyl oligosaccharide, a carbohydrate that exists in lactoferrin and in the 38-kDa fragment of band 3 protein. Anti-lactoferrin antibodies are found primarily in plasma of patients with autoimmune diseases and who have anti-neutrophil cytoplasmic antibodies (ANCA).
In sepsis death follows an excessive inflammatory response involving cytokines and complement that is activated primarily via the amplifying C3/C5 convertase. Excessive stimulation of complement ...amplification requires IgG-containing or F(ab′)2-containing immune complexes (IC) that capture dimeric C3b on one of their heavy chains or heavy chain fragments. The ability of IgG-IC to capture dimeric C3b by the Fab portion is dependent on an affinity for C3 within the Fab portion, but outside the antigen-binding region. This property is rare among IgG NAbs. In contrast to this, the lack of the Fc portion renders the Fab regions of any F(ab′)2-IC accessible to nascent C3b, but dimeric C3b deposits only if F(ab′)2-IC form secondary IC with anti-hinge NAbs that rigidify the complex and thereby promote deposition of dimeric C3b. Both types of complexes, C3b2-IgG-IC and C3b2-F(ab′)2-IC/anti-hinge NAbs, are potent precursors of alternative C3 convertases and stimulate complement amplification along with properdin up to 750 times more effectively than C3b and properdin. F(ab′)2 fragments are not normally generated, but are formed from NAbs by enzymes from pathogens and neutrophils in sepsis. Unlike IgG-IC F(ab′)2-IC are not cleared by Fc-receptor dependent processes and circulate long enough to form secondary IC with anti-hinge NAbs that rigidify the complexes such that they capture dimeric C3b and gain the potency to stimulate complement amplification.
Membranoproliferative glomerulonephritis type II (MPGN II) is a rare disease characterized by the deposition of abnormal electron-dense material within the glomerular basement membrane of the kidney ...and often within Bruch's membrane in the eye. The diagnosis is made in most patients between the ages of 5 and 15 yr, and within 10 yr, approximately half progress to end-stage renal disease, occasionally with the late comorbidity of visual impairment. The pathophysiologic basis of MPGN II is associated with the uncontrolled systemic activation of the alternative pathway (AP) of the complement cascade. In most patients, loss of complement regulation is caused by C3 nephritic factor, an autoantibody directed against the C3 convertase of the AP, but in some patients, mutations in the factor H gene have been identified. For the latter patients, plasma replacement therapy prevents renal failure, but for the majority of patients, there is no proven effective treatment. The disease recurs in virtually all renal allografts, and a high percentage of these ultimately fail. The development of molecular diagnostic tools and new therapies directed at controlling the AP of the complement cascade either locally in the kidney or at the systemic level may lead to effective treatments for MPGN II.
We identified 11 human pedigrees with dominantly inherited hemolytic anemias in both the hereditary stomatocytosis and spherocytosis classes. Affected individuals in these families had an increase in ...membrane permeability to Na and K that is particularly marked at 0 °C. We found that disease in these pedigrees was associated with a series of single amino-acid substitutions in the intramembrane domain of the erythrocyte band 3 anion exchanger, AE1. Anion movements were reduced in the abnormal red cells. The 'leak' cation fluxes were inhibited by SITS, dipyridamole and NS1652, chemically diverse inhibitors of band 3. Expression of the mutated genes in Xenopus laevis oocytes induced abnormal Na and K fluxes in the oocytes, and the induced Cl transport was low. These data are consistent with the suggestion that the substitutions convert the protein from an anion exchanger into an unregulated cation channel.
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