Kidney Antigens Research Articles

Evidence for expression of Ly-6.2 on non-bone marrow-derived cells in kidney, skin and connective tissues.

Mouse alloantigen Ly-6.2 is detectable in various non-lymphoid tissues such as kidney, but it is not clear whether or not this expression is due to bone-marrow derived passenger leukocytes. To determine whether non-marrow derived cells express Ly-6.2, we examined the expression of this antigen in kidney and on isolated connective tissue and epidermal cells. Studies in radiation chimeras demonstrated that the kidney did not become Ly-6.2 positive when negative animals were reconstituted with positive marrow. Thus, passenger leukocytes cannot account for the renal expression of Ly-6.2, indicating that most of this antigen is on non-marrow-derived (parenchymal) cells in kidney. Various isolated cell types--fibroblasts, osteocytes, chondrocytes and skin epidermal cells--were found to be Ly-6.2 positive. Indeed, absorption and cytotoxicity results suggested that the amount of Ly-6.2 on fibroblasts exceeded the amount of an H-2 antigen on these cells. Comparison of fibroblasts to lymphocytes indicated that fibroblasts had 13--60 times more Ly-6.2 than spleen cells and three times more than PHA blasts. The results indicate that the Ly-6.2 detected in non-lymphoid tissues is predominantly on the parenchymal or connective tissue elements of those tissues.

Expression of Ia-like antigens on the vasculature of human kidney

The distribution of Ia-like antigens in human kidney has been investigated by indirect immunofluorescence staining of frozen sections with monoclonal antibodies to framework determinants of Ia-like antigens. These antigens are localized by patchy and interrupted linear fluorescence along the endothelium of glomerular efferent arterioles and capillaries, intertubular cortical and medullary capillaries as well as the vasa recta. Ia-like antigens are undetectable, however, on the endothelia of arcuate and interlobular arteries and on tubular epithelial cells. The expression of Ia-like antigens on kidney endothelial cells may account for the role of Ia-like antigens in kidney allograft rejection.

Localization of HLA-ABC and DR antigens in human kidney.

Monoclonal antibodies to human monomorphic class I and class II major histocompatibility complex (MHC) determinants have been used with immunofluorescence and immunoperoxidase techniques, to localize these antigens in normal human kidneys. HLA-DR antigen was located in the glomeruli (probably on endothelium as well as the mesangium) and within the cells of cortical and medullary tubules. Dendritic cells in the renal interstitium stained brightly for the DR antigen and could be distinguished from the staining of capillary endothelium. The vascular endothelium of large vessels stained less densely for the HLA-DR antigen than for HLA-ABC antigens. The glomeruli stained intensely for the HLA-ABC antigens and diffuse staining of HLA-ABC antigens was also noted within renal tubular cells.

Sensitive radioimmunoassays for 7 S collagen and laminin: Application to serum and tissue studies of basement membranes

Radioimmunoassays were developed for the basement membrane components, 7 S collagen and fragments of the noncollagenous protein laminin, which allowed quantitative analysis of as little as 0.1–0.4 ng of these proteins. Similar materials could be detected by these assays in serum, in kidney digests, and in the medium of cell cultures obtained from mice and rats. Distinct changes in the amounts of antigen in serum and kidney were observed during aging and in mice inoculated with a basement membrane tumor.

Entactin, a novel basal lamina-associated sulfated glycoprotein.

A sulfated glycoprotein, entactin, of apparent molecular weight 158,000 has been isolated from an extracellular basement membrane-like matrix. This matrix is elaborated in cell culture by a mouse endodermal cell line. Antibodies prepared in rabbits against this sulfated glycoprotein react with mouse and rat basement membranes from a variety of tissues. These antibodies also react in a specific manner with a discrete component of mouse and rat kidney glomeruli. The electrophoretic mobility of this component is identical to that of entactin. The mouse kidney antigen, as shown by immunoelectron microscopic studies, is predominantly localized at the surface of epithelial cells of tubules and glomeruli adjacent to the basement membrane. Some antigen is also present in the basal lamina adjacent to the epithelial cells. Entactin is distinct from the basement membrane-associated protein GP-2, a protein similar to laminin. Entactin differs from GP-2 in electrophoretic mobility, cyanogen bromide peptide fragmentation pattern, immunological cross-reactivity, and incorporation of H235SO4. Entactin is insensitive to treatment with chrondroitinase ABC. It is suggested that this molecule plays a role in the interaction of the extracellular matrix and the cell surface.

1529 MINIMAL CHANGE NEPHROTIC SYNDROME (MCNS): AN AUTOIMMUNE DISEASE?

The presence of low-level tissue directed antibodies (AAb's) and immune complexes (IC) were determined in 12 MCNS and 5 mesangial proliferative glomerulonephritis (MesPGN) patients with biopsy proven disease. The AAb's were detected by immunofluor-escence (IF) against human cadaveric tissues and the IC detected by 3 different methods and isolated by polyethylene glycol precipitation/sedimentation. The isolated antibodies from the IC were also tested by IF for AAb activity against the same tissue antigen battery. The presence of such AAb activity was demonstrated in 8/12 MCNS and in all of the MesPGN sera. In the MCNS group 7/8 of the AAb positive sera also contained IC with corresponding AAb activity (2/7 directed against smooth muscle antigens and 5/7 directed against kidney tubular microsomes). The MesPGN group all reacted against kidney basement membrane antigens but no other tissue antigen. Of these 3/5 also demonstrated the presence of IC which contained AAb reactive with the same kidney antigen. The presence of such AAb's and their formation of IC indicates that these antibodies may have a functional role in the initial immunological insult to the kidney.

Development of a radioimmunoassay for a high molecular mass tubular antigen in urine — Its application for early detection of tubular damage

In order to isolate urinary kidney antigens, the gammaglobulin fraction of an antiserum against human kidney cortex plasma membranes was coupled to Sepharose 4B. By immun ospecific affinity chromatography an antigen fraction was obtained from the urine of a patient suffering from severe kidney disease. After gel filtration, the main fraction, eluted with the exclusion volume of a Sephadex G-200 column and enriched 16 000-fold, was labelled with 131I and used in a radioimmunoassay system. Soluble kidney antigens, presumably of proximal tubular origin, could be detected and quantified by the assay system in urine samples of patients with various diseases. The samples did not need to be treated, either concentrated or dialyzed, before application. The results of our experiments show a correlation between antigen excretion and kidney damage. Rejection episodes in patients with kidney transplants could be recognized early by enhanced antigen excretion. Potentially nephrotoxic drugs caused antigen excretion as well. In normal, healthy subjects output of the antigen was very low. The assay system might be of value for monitoring renal diseases.

Ultrastructural localization of blood group A antigen in normal human kidneys

Blood group A antigen has been shown by immunofluorescence to be present on vascular endothelia ( J. Bariety, R. Oriol, N. Hinglais, M. Zanetti, R. Bretton, A. M. Dalix and C. Mandet, 1980 , Kidney Int. 17, 820—826; E. J. Holborow, P. C. Brown, L. E. Glynn, M. D. Hawes, G. A. Greshan, T. E. O'brien, and R. R. A. Coombs, 1960 , Brit. J. Exp. Pathol. 41a, 430—437; A. E. Szulman, 1960 , J. Exp. Med. 111B, 785—789), distal convoluted tubules (Bariety et al.), and, in secretors, on deep collecting tubules (Bariety et al.: Szulman) of normal human kidneys. The present immunoelectron microscopic study was performed on 15 nephrectomy specimens, 13 of which were from blood group A patients including 12 secretors and I nonsecretor. Purified rabbit anti-A antibodies were used with peroxidase-labeled sheep anti-rabbit lgG F(ab')2 antibodies in a double-step procedure. The reaction product was shwon on plasma membranes and Golgi apparatus of all types of vascular endothelia, on convoluted distal tubules, and on collecting tubules of both secretor and nonsecretor patients. However, in the secretor patients, labelling was more intense and extensive as it was present on a thick apical glycocalyx, on numerous intracytoplasmic areas, and in the basement membrane. These results localize blood group A antigen at the ultra-structural level and suggest an in situ synthesis of blood group A substance in the kidney.

Expression of Ia in mouse kidney.

Expression of Ia antigens in mouse kidney was studied by absorption of diluted anti-Ia sera with crude suspensions of kidney issue. Specific absorption of anti-Ia activity was seen for all Ia specificities tested: Ia.1,2, Ia.3, Ia.4,5,12, Ia.7, Ia.8, Ia.9, Ia.15, and Ia.16. Certain polyspecific antisera (against the I-Ak products Ia.1,2,3,15) were more difficult to absorb than oligo-specific antisera against other Ia specificities (e.g., Ia.7 and Ia.9). This observation may indicate that polyspecific sera are less absorbable by limited numbers of antigenic sites because of steric hindrance, although differences in the extent of antigen expression in kidney have not been excluded. Ia absorption could be demonstrated either in microcytotoxicity or in 51Cr release assays. Both mechanically disrupted and enzyme-disrupted kidney tissue suspensions absorbed Ia specifically, although the former method was used routinely. As estimated from the efficiency of absorption, the amount of Ia in kidney was small, about 2 to 5% of that in spleen. One kidney was equivalent in absorptive capacity to about 3 X 10(6) splenocytes, and to greater than 3 X 10(6) buffy coat cells. Comparisons of the rates of absorption indicated that the amount of Ia in kidney was less than the amount of an H-2K or D alloantigen. Ia was expressed in kidney in an immunogenic form, since animals immunized repeatedly with I region-incompatible kidney tissue produced anti-Ia antibodies. Thus, Ia antigens are expressed in and are immunogenic in mouse kidney and can be studied by conventional serological techniques.

Effect of lymphocytes sensitized to kidney antigens, injected into female mice, on renal function of their progeny

Effect of lymphocytes sensitized to kidney antigens, injected into female mice, on renal function of their progeny

Developmental aspects A, B, H, Lewis, I, i and Pr antigens in human kidney

Summary Blood group antigens are widely distributed in human tissues. Numerous studies were carried out on adult kidneys, the aim of this work being to compare the distribution of ABH, I, i, Lewis and Pr antigens in adult kidneys with that of three month embryos. In actual fact, at this stage of development, the same structures as those observed in the adult can be seen. The study was carried out by means of the indirect immunofluorescence technique. In the adult, ABH antigens were evidenced on the cells of the vascular endothelium and in the distal, and collecting tubules of secretor subjects only. Lewis antigen was absent in the vascular structures but present in the cells of the distal and collecting tubules of all the subjects with the same Le gene. A high marker heterogeneity was noted inside a same tubule. I and i antigens were observed in the membrane and cytoplasm of some cells from the thick part of the loop of Henle and in some distal and collecting tubule cells. Pr antigen was present in the glomerulus and the distal and collecting tubules. In the three month fetus, ABH and Pr antigens were observed in the same structures. I antigen was absent, whereas i antigen was present in the cells of the distal and collecting tubules. However, the expression of the Lewis specificities was virtually inexistent. Therefore, at the age of three months, when the development of the organs is finished, the embryonic tissues possess the same markers as the adult with the exception of I and Lewis antigens. The results observed in the kidneys of the 14 week fetuses were similar to those observed in the adults (8, 10). Nevertheless, the staining of the apical membrane of the cells appeared stronger than in the adults. Lewis antigens were not shown on the endothelial cells of all the vessels. These antigens were only present in the membrane and cytoplasm of distal and collecting tubules [5]. Whilst the Lewis enzyme was clearly demonstrated in the glomerulus from Lewis positive kidneys, Lewis antigen was not found in the glomerulus [5, 9]. Although A, B, and H antigens were present on some kidney structures of the three month fetuses, Lewis antigens were only found in a small quantity. I and i antigens were only studied on the adult kidneys. In a former study, Feizi [2], using the method of inhibiting precipitation, did not evidence I and i antigens in the kidney. Moreover, in a recent paper, using indirect immunofluorescent techniques, Lenhart et al. [4] showed I and i antigens in different structures of adult kidney. As regards localization, our results come close to those observed by Lenhart et al. [4]. I and i antigen were also found mainly in the cytoplasm and on the surface of the epithelium of distal tubules ; no I and i antigens were found in the glomerulus. The fetuses of three months were completely lacking in I antigen. Whilst i antigen was expressed, its strength seems almost identical to that found in the adult. In contrast to Lenhart et al. [4], Pr antigens were found in the collecting and distal tubules. Nevertheless, some anti-Pr gave a very slight staining of the tubule, whereas the glomerulus were distinctly positive. Thus the main difference between the adult and fetus kidneys was the absence of I and Lewis antigens on the epithelium of the distal and collecting tubules.

THE DISTRIBUTION AND PROPERTIES OF BLOOD GROUP P ANTIGENS IN HUMAN ORGANS

1) The distribution of blood group P1 antigens in human organs was investigated. P1 antigen was found in blood group P1 kidney. Both group P1 and P2 kidneys contained much P antigens and lung, spleen and heart contained small amount of P antigens.2) The each fraction corresponding to globoside and CTH (ceramide trihexoside) isolated from kidney showed P and Pk activities respectively.3) Treatment of the globoside fraction from kidney with human liver's crude enzyme preparation (contained s-N-acetylgalactosaminidase) caused the loss of P specificity and the appearance of Pk specificity.As the above results, it was confirmed that blood group P antigens in kidney were identical with those in erythrocytes.

Reactivity of Paul‐Bunnell Type Heterophile Antibody in Sera from Infectious Mononucleosis Patients with the Surface of Lymphoid Cells Carrying Epstein‐Barr Virus Genomes

The possible presence of Paul-Bunnell (PB) antigen on Epstein-Barr virus (EBV)-transformed lymphocytes were investigated. Of 23 EBV genome-positive lymphoblastoid cell lines tested all but one absorbed PB type antibody from the serum of an infectious mononucleosis patient. The one EBV-negative B cell line tested did not absorb the heterophile antibody. PB antibody, purified by an immunoadsorbent procedure using beef cell antigen, reacted with the EBV producer P3HR-1 cell line in an indirect membrane immunofluorescence test and was shown to be IgM antibody. Titers of heterophile agglutinin and reactivity with the cell surface were reduced to the same degree by absorption with beef cell antigen but not with guinea pig kidney antigen. PB antibody was distinct from IgM antibody against the EBV-determined membrane antigen, since the latter was not absorbed by beef cell antigen. PB antibody was also reactive with other EBV-positive B cell lines (QIMR-WIL, NC-37, and Raji) which were free of surface IgM. No reaction occurred with the nonproducer P3HR-1 line, a null cell line, and two T cell lines. The results suggest the presence of PB antigen on most EBV-transformed B lymphocytes, and its appearance in each of the transformed lymphocytes of patients with acute infectious mononucleosis.

Studies on “Liver-Specific” Antigens

Sera of patients with chronic liver disease were tested for antibodies against the soluble liver cell membrane antigens LSP and LP-2 as well as particulate liver cell membrane proteins, and against KSP, the kidney equivalent of LSP In a technique for antibody-dependent, cellular cytotoxicity, 3 H-proline-labeled, antigen-coated marine mastocytoma target cells were incubated with the patient's serum in the presence of human, Fc-receptor-bearing, polymorphonuclear leukocytes, acting as effector cells. As compared to normal control sera, the serum of a significant number of patients with chronic hepatitis contained antibodies against one or more of the four antigens tested, including the kidney antigen, KSP. Lysis of antigen-coated target cells could be inhibited by the test antigen and by other antigens against which the patient's serum contained antibodies. However, cross-inhibition by KSP could not always be recorded, which suggested that liver antigens contain an organ-specific component. Substitution of test sera by anti-LSP, -KSP, or -LP-2 sera, resulted in Lysis of all test antigen-coated target cells but had no effect on uncoated, human serum albumin-, and bovine serum albumin-coated mastocytoma cells. These results indicate that, in most cases, sensitization in chronic liver disease is not an organ-specific phenomenon and that liver cell membrane components contain common antigenic determinants that are also shared by the kidney antigen, KSP. The pathogenetic role of LSP and other liver antigens in chronic hepatitis must, therefore, be evaluated with caution. However, differential immunity to specific antigens of the liver and other organs may be of diagnostic and prognostic significance.

Argininosuccinase from bovine brain: Isolation and comparison of catalytic, physical, and chemical properties with the enzymes from liver and kidney

Homogeneous argininosuccinase has been isolated from bovine brain: compared to liver and kidney argininosuccinases from the same species, the catalytic activity (1400 U/mg). molecular weight of the fully active form (202,000 by gel filtration), and the minimum molecular weight (50, 000 in sodium dodecyl sulfate and mercaptoethanol) were in agreement with published liver and kidney enzyme values from this laboratory. That the brain enzyme is composed of four identical, or closely similar, polypeptide chains is supported by peptide maps analyzed after tryptic or cyanogen bromide cleavage. One-fourth the number of peptide fragments were produced as compared to the total number of susceptible residues per mole. The number of peptides containing other specific residues, or methionyl residues, were consistently one-fourth of the total considered. As maps of peptide fragments prepared from the brain enzyme were also superimposable, or nearly so, on liver enzyme maps, the four polypeptide chains from both sources were closely similar to each other in amino acid sequence. Distribution of the 16 sulfhydryl groups, as based on titration with Ellman's reagent, was in accord with the liver enzyme: Four sulfhydryl groups reacted without affecting catalytic activity, a second group of 4 became accessible on cold dissociation of the tetramers to catalytically inactive dimers, and the final 8 became accessible in strong dissociating agents. On analysis, Km values and negative homotropic interactions with substrate were in accord with liver enzyme kinetics. Immunological studies indicated a ciose resemblance in antigenic properties. The brain enzyme, as antigen, was fully crossreactive in the formation of precipitin bands with rabbit antibody to either liver or kidney enzymes already known to be mutually cross-reactive. The antibody to the liver enzyme was an effective inhibitor of brain enzyme activity comparable to inhibition of the homologous liver and kidney antigen.

MACROPHAGE-LYMPHOCYTE INTERACTION IN GRAVES' DISEASE AND HASHIMOTO'S THYROIDITIS

The formation of macrophage-lymphocyte rosettes was studied in lymphocyte cultures from patients with Graves' disease; Hashimoto's thyroiditis, other thyroid diseases and control subjects; the cultures were incubated with normal human thyroid and other non-specific antigens. At the end of incubation, the cell pellets were smeared on slides, stained with Wright's stain and the number of rosettes determined under the microscope. The membrane immunofluorescence technique was employed to identify whether the surrounding lymphocytes were T- or B-lymphocytes. In Graves' disease and Hashimoto's thyroiditis, the mean percentages of rosette formation with crude thyroid antigen were 0.98 +/- 0.22% (mean +/- SEM), and 1.15 +/- 0.25%, respectively. These values were significantly higher than those of control lymphocytes (0.03 +/- 0.02%). Lymphocytes from other thyroid diseases also gave higher values than controls. Kidney antigen, used as a control antigen, gave negative results in Graves' disease and other thyroid diseases, but in Hashimoto's thyroiditis, the mean percentage was of borderline significance. In the direct immunofluorescent staining study using fluorescein-conjugated goat anti-human Ig determinants, including the Fab fraction of anti-human IgG, it appeared that both B- and T-lymphocytes were involved in the rosettes, although B-lymphocytes were more numerous. These results indicate that in patients with Graves' disease and Hashimoto's thyroiditis, a probable immune reaction with thyroid antigen can be demonstrated by macrophage-lymphocyte rosette formation.

Hemocoagulation disorders in kidney allotransplants, caused by preformed antirenal antibodies

Kidney allografts were transplanted through cervical blood vessels to dogs, normal or with induced glomerulonephritis (those producing antirenal antibodies). Fifteen minutes after the introduction of the allogenic kidney into the recipient's circulation, a sharp decline in the anti-kidney antibody titer was observed in the blood in flowing out of the transplanted kidney. It was accompanied by a lowering in the platelet count, and by a decrease in the content of fibrinogen and complement. Considerable amount of FDP appeared. A high degree of correlation was found between the immunological alterations and hemocoagulative indices. Examined by means of light microscopy, allograft tissue samples, withdrawn three hours after the transplantation, demonstrated the typical hyperacute rejection phenomen: fibrin deposition in the capillaries of the glomeruli, and fibrinoid impregnation of the walls of large and medium size interstice arteries. Immunoglobulins eluted from the transplants precipitated kidney antigen specifically, and if fluorescent-dye-labeled, caused luminescence in the capillar loops of glomeruli and the walls of larger vessels. These findings suggest an explanation for acute kidney allotransplant damages occuring soon after the operation in patients with glomerulonephritis even in the absence of preexistent lymphocytotoxic antibodies.

An Ly-like specificity with extensive nonlymphoid expression

The characteristics of a strong mouse alloantigen with renal, bone marrow, and lymphoid expression were studied. This antigen is probably identical to that currently designated Ly-6.2. It was defined by the high-titered (1:1000) cytotoxic activity of three different antisera against peripheral lymphocyte target cells from DBA/2, DBA/1, and a variety of other strains. In the F(2) and four backcross generations the genetic control of this specificity segregated as a single autosomal dominant gene. In lymphoid tissues the predominant expression was on T cells but 10-30% of B cells were lysed by these antisera. The specificity was expressed strongly in kidney, as shown by sequential absorption, in amounts equal to or greater than the amount in lymphoid tissues. Comparison to the rate of absorption of H-2 by kidney indicated that this antigen may be expressed in amounts comparable to an H-2 antigen in kidney. Immunization with kidney tissue resulted in a strong cytotoxic antibody response. The number of bone marrow cells expressing this antigen (40-50%) was well beyond what could be accounted for by T lymphocytes in bone marrow. In addition, a nonlymphoid tumor, the P815Y mastocytoma, was positive by cytotoxicity and by absorption. The extensive nonlymphoid expression and antigenic strength of Ly-6.2 raises the possibility that this serologically defined lymphocyte alloantigen will have histocompatibility effects when allografts of the appropriate tissues are examined.

Partial Purification of Human Kidney Antigens

A chromatographic method using immunoaffinity has been utilized in the search for kidney-specific antigens. After repeated absorptions by liver proteins coupled to a column of Sepharose CN-Br of a rabbit anti-human kidney serum, several antibodies remained. When these antibodies were coupled to a similar column and used for immunoaffinity chromatography, 8 kidney antigens, studied as peptides with molecular weights of 33,000, 80,000, 98,000 and 110,000-125,000 were obtained. They should not be found in similarly treated liver extracts.

Species Specificity in Thermostable and Ethanol Insoluble Tissue Antigens

In a previous experiment, rabbits and goats were immunized with boiled and ethanol precipitated (BE) bovine kidney antigen, and the specificity of the antibodies produced was compared (Andersen 1975). The caprine sera were species specific while the rabbit sera, however, cross-reacted with BE antigens from other ruminant species. Sera from 2 rabbit littermates differed somewhat in that 1 serum seemed to be mainly species specific giving only weak reactions against BE antigens from kidney and spleen from other ruminants, whilst the other serum was more organ specific and reacted equally with homologous and heterologous kidney antigens.