Lymphocyte predominance Hodgkin disease is characterized by recurrent genomic imbalances

Single-cell polymerase chain reaction (PCR) has been used as a tool to demonstrate clonality and B-cell origin of Reed-Sternberg (RS) cells in Hodgkin disease (HD). An analogous approach was used to investigate genomic imbalances in a (cyto)genetically poorly characterized subentity: lymphocyte predominance Hodgkin disease (LPHD). Nineteen cases of LPHD were selected for a comparative genomic hybridization (CGH) study. CGH was performed with degenerate oligonucleotide primed-PCR (DOP-PCR)-amplified DNA from 4-5 microdissected CD20+ malignant cells. All analyzed cases revealed a high number of genomic imbalances (average 10.8 per case), involving all chromosomes but the excluded 19, 22, and Y, indicating a high complexity of LPHD. The majority of detected aberrations were recurrent. Gain of 1, 2q, 3, 4q, 5q, 6, 8q, 11q, 12q, and X, and loss of chromosome 17 were identified in 36.8% to 68.4% of the analyzed cases. Some of them have also been found in non-Hodgkin lymphoma (NHL), and possibly represent secondary changes associated with disease progression. Gain of 2q, 4q, 5q, 6, 11q, however, are much more rarely observed in NHL and could be more specifically associated with LPHD. Particularly interesting is a frequent overrepresentation of chromosome arm 6q, a region usually deleted in NHL. Rearrangement of the BCL6 gene (3q27) demonstrated by cytogenetics and fluorescence in situ hybridization in 2 cases in this study suggests its contribution in pathogenesis of LPHD. In conclusion, the data show a consistent occurrence of genomic alterations in LPHD and highlight genomic regions that might be relevant for development and/or progression of this lymphoma entity.     

Unconstrained lateral diffusion of concanavalin A receptors on bulbous lymphocytes.

The lateral diffusion coefficient, D, of concanavalin A receptors and receptor complexes on the surface of lymphocytes and RDM4 lymphomas is enhanced by several orders of magnitude to D greater than 5 X 10(-9)cm2/sec by induction of swelling of the cells to bulbous form. Treatments with concanavalin A or 7-nitrobenz-2-oxa-1,3-diazole-phallacidin induce blebs and the bulbous form. The resulting separation of the plasma membrane from most of the F-actin cytoskeleton is accompanied by release of constraints on lateral diffusion of the cell surface receptors, which allows the diffusivity of these glycoproteins to increase nearly to the limit allowed by membrane viscosity.     

Concanavalin-A-induced transmembrane linkage of concanavalin A surface receptors to intracellular myosin-containing filaments.

With normal rat kidney cells in monolayer culture, we have studied the distribution on the cell surface of receptors for concanavalin A, and the distribution of the smooth muscle myosin-like protein inside the same cell, using specific fluorescence microscopic methods. The concanavalin A receptors were initially uniformly dispersed over the cell surface, but 20 min after the addition of concanavalin A at 37 degrees, the receptors showed a variety of nonuniform surface distributions, including extended parallel linear arrays. These arrays of receptors were found to be superimposed on the linear arrays of the intracellular myosin-containing filaments, indicating that a transmembrane linkage of the receptors and the filaments had occurred. This linkage required a lateral redistribution of concanavalin A receptors, since it did not occur with succinylated concanavalin A, but was subsequently induced if the cells that had been reacted with succinylated concanavalin A were then treated with antibodies to concanavalin A. The redistributions of concanavalin A receptors on the surfaces of these normal rat kidney cells, however, were much less extensive than the patching that was induced on the surfaces of the same cells infected with, and transformed by, Rous sarcoma virus.     

I and i antigens of human peripheral blood lymphocytes cocap with receptors for concanavalin A.

Surface immunofluorescence experiments using a human anti-i and two anti-I antisera have been performed on human peripheral blood lymphocytes. These are known to contain cold-reactive monoclonal IgM antibodies against the carbohydrate sequence: (formula: see text). A high proportion of B- and T-type lymphocytes express these I and i determinants. In the presence of anti-human immunoglobulin, the cold-reactive membrane-associated complexes of I-anti-I and i-anti-i become stabilized, and redistribution (with patching and capping) can be elicited at 37 degrees C. Dual fluorescence experiments have shown striking concordant staining of I or i (fluorescein) caps and patches with concanavalin A (rhodamine) reactive sites on normal and leukemic cells, suggesting that a proportion of I and i active structures of lymphocyte membranes are structurally associated or physiologically coupled with glycoproteins carrying oligosaccharides with branched mannosyl cores.     

Increased concanavalin A induced suppression in treated and untreated coeliac disease.

The generation of suppression by concanavalin A in peripheral blood mononuclear cells in treated and untreated coeliac subjects using an in vitro assay was found to be significantly increased when compared with controls. The response of peripheral blood mononuclear cells to the plant mitogen concanavalin A (con A) was also significantly depressed in both groups of coeliac patients. It is proposed that the depressed cell mediated immunity found in this and other studies in coeliac patients is because of increased suppression. The possible connection between these findings and the increased incidence of malignancy also found in coeliac disease is discussed.     

Combination chemotherapy of Hodgkin disease in children.

MOPP chemotherapy program was employed in 20 children with advanced Hodgkin disease (stages III and IV). Complete remissions were achieved in 13 patients (65%)within an average duration rate of 17.1 months varying from 2 to 40 months. If thses only one patient developed recurrency and died 10 months after the onset of treatment. The remaining 12 children are alive and well; an average survival rate since the onset of MOPP chemotherapy being 20 months. The patients with partial remission died from further progression of the disease within the period from 2 to 15 months. The study showed that histological type and mode of previous treatment had an influence on the results of chemotherapy used. The complications of MOPP chemotherapy were insignificant and easily controlled.     

Lymphocyte subpopulations in adult coeliac disease.

Rosetting techniques were used to estimate T and B cell subpopulations in the peripheral blood in patients with treated and untreated adult coeliac disease and in control subjects. In patients with untreated coeliac disease, T cell numbers were significantly lower than in controls or treated patients, although there was no difference in total lymphocyte counts. There was no significant difference in B cell numbers between treated and untreated patients, and the subpopulation which increased to replace the T cells in untreated patients comprised cells not identified by B or T cell markers. Total lymphocyte counts and lymphocyte subpopulations were affected by splenic atrophy. It is suggested that these effects might be caused by the loss of lymphocytes from the gastrointestinal tract in untreated coeliac disease.     

Lymphocyte thyroid hormone receptors in obesity

Triiodothyronine (T3)-receptor characteristics of isolated circulating human mononuclear cells have been studied in a group of obese patients who claimed to be unable to lose weight on conventional 4.2 MJ (1000 kcal) diets. The cells of the obese patients exhibited a lower receptor capacity than those of a control group of non-obese subjects but the difference was not significant. There was a significant fall (P less than 0.01) in receptor capacity in the obese patients after 12 weeks in a 1.34 MJ (320 kcal) per d formula diet and this provides a further mechanism whereby a fall in metabolic rate takes place in response to severe dietary restriction. Some patients who also received T3 (60 micrograms/d) in addition to the formula diet showed a further fall in the receptor numbers. These findings may partly account for the previously reported resistance to thyroid hormones in obesity.     

Lateral transport on cell membranes: mobility of concanavalin A receptors on myoblasts.

We report measurements of the lateral mobility of fluorescent labeled concanavalin A receptor complexes on the plasma membrane of cultured myoblasts of rat. Transport rates were measured by observing the recovery of fluorescence in a small region of the cell surface initially photobleached irreversibly by an intense, focused laser light pulse. Under different conditions we measured effective diffusion coefficients of the receptor complexes in the range 8 x 10(-12) less than D less than 3 x 10(-11) cm2/sec which is two orders of magnitude lower than we found for a fluorescent lipid probe, D approximately (8 +/- 3) x 10(-9) cm2/sec. This large difference and the presence of apparently immobile concanavalin A receptors suggests that factors beyond the fluoidity of the phospholipid bilayer membrane matrix control the rate of lateral transport of the complexes. Effective mobilities of the complexes decrease with increases in the valence, dose, and occupation time of the lectin on the membrane. These properties imply an aggregation of the lectin-receptor complexes. Mobilities are not influenced by azide, colchicine or preincubation at low temperature. Cytochalasin B and low temperatures, during the time of measurement, decrease the lateral transport rate.     

Isolation and partial characterization of concanavalin A receptors on cloned cytotoxic T lymphocytes.

A small set of concanavalin A (Con A)-binding glycoproteins was isolated from the surface membrane of cloned cytotoxic T lymphocytes (CTL) and partly identified using monoclonal antibodies. The binding of Con A by these glycoproteins on the CTL surface results in the secretion of gamma-interferon and in blocking the effector functions of the cells-namely, antigen-specific and lectin-dependent cytotoxicity. The Con A is evidently bound tightly to some surface structures ("Con A-receptors") that are required for the activation and cytotoxic activity of CTL. To isolate and identify these receptors, antibodies to Con A were used. After Con A was allowed to bind to radiolabeled cloned CTL (labeled with 125I or [35S]methionine or 3H-labeled amino acids), the cells were washed thoroughly, lysed in detergents and anti-Con A antibodies were added to bind to the Con A-receptor complexes. The resulting aggregates were adsorbed with protein A-bearing Staphylococci and the receptors were then specifically released from the pelleted bacteria by alpha-methyl-D-mannoside and analyzed by polyacrylamide gel electrophoresis under reducing conditions. Eight to nine labeled components were seen by autoradiography and with the aid of monoclonal antibodies to known T-cell surface molecules, four were identified as T200, lymphocyte function-associated antigen (LFA)-1, alpha- and beta-chains, and (on some clones) Lyt-2. Other components with Mr congruent to 160,000, 120,000, 46,000, 42,000, and 23,000 have not been identified. The procedures described here may have general application in the studies of the functional properties of other cell surface molecules.     

Lymphocyte subpopulations, lymphoblast transformation activity, and concanavalin A-induced suppressor activity in patients with ulcerative colitis and Crohn's disease

The following immunologic in vitro tests were applied on peripheral blood mononuclear cells (PBMC) from patients with chronic inflammatory bowel disease (IBD): concanavalin A (Con A)-induced suppressor test, Con A-activated lymphoblast transformation test, and spontaneous lymphoblast transformation test. Concomitant phenotypic characterization of subsets of PBMC was performed with monoclonal antibodies. Patients with ulcerative colitis and a control group with rheumatoid arthritis showed significantly reduced activity in the Con A-activated lymphoblast transformation test compared with healthy controls and patients with Crohn's disease. The distribution of PBMC subsets and the results of the other in vitro tests were similar for patients with IBD and healthy controls. Thus the decrease in Con A-activated lymphoblast activity was not due to an increased suppressor function as measured either by functional Con A-induced suppressor test or indirectly by T8 phenotype.     

Monovalent derivatives of concanavalin A.

Monovalent dimers of concanavalin A (Con A) have been prepared by a combination of succinylation and photoaffinity labeling. Partial derivatization of native Con A using the photoaffinity label, p-azidophenyl-alpha-D-mannopyranoside, followed by affinity chromatography yielded a fraction that consisted of dimers with a single saccharide-binding site at pH 5. These monovalent dimers formed divalent tetramers at pH 7. In order to achieve a monovalent dimer at this pH, the divalent tetramers were succinylated by previously developed methods. Ultracentrifugation, equilibrium dialysis, and chromatographic experiments indicated that the resultant preparations consisted mainly of monovalent dimers which showed subunit exchange to yield about 15% divalent dimers after 12 hr at physiological pH. Freshly prepared material failed to agglutinate sheep erythrocytes at concentrations 500-fold higher than native tetravalent Con A. In addition, they showed saturating dose-response curves of mitogenic stimulation of mouse splenic lymphocytes. These curves resembled those of divalent succinyl-Con A but not those of the native molecule. Further development of methods for preparing stable monovalent derivatives of Con A should allow a refined analysis of the effects of lectin valence at the cell surface.