A novel embryogenetic mechanism for Currarino''s triad: inadequate dorsoventral separation of the caudal eminence from hindgut endoderm

We studied the effect of sulfasalazine on anaphylaxis. Sulfasalazine dose-dependently inhibited systemic anaphylaxis induced by compound 48/80 in rats. Sulfasalazine also inhibited local anaphylaxis activated by anti-dinitrophenyl (anti-DNP) IgE. Moreover, sulfasalazine dose-dependently inhibited histamine release in the peritoneal mast cells activated by compound 48/80 or anti-DNP IgE. All of these effects were comparable to those of the disodium cromoglycate (reference drug) tested. When sulfasalazine was added, the level of cAMP in rat peritoneal mast cells transiently and significantly increased about 6-fold compared with that of basal cells. Our studies provide evidence that sulfasalazine may be beneficial in the treatment of anaphylaxis.     

Combinational effects of vitamin D3 and retinoic acid (all trans and 9 cis) on proliferation, differentiation, and programmed cell death in two small cell lung carcinoma cell lines

The effects of a combination of vitamin D3 [1,25(OH)2D3] and retinoic acid (RA) on proliferation, differentiation, and apoptosis of the human small cell lung carcinoma (SCLC) cell lines NCI-H82 and NCI-H209 were evaluated. Cell proliferation was inhibited by 1,25(OH)2D3 and RA alone. The combination of 1,25(OH)2D3 and the cis form of retinoic acid resulted in an additive decrease in cell proliferation and the induction of apoptosis in various concentrations. Moreover, 3H-thymidine incorporation was inhibited and the number of viable cells was decreased. The characteristics of the apoptotic cells were examined and confirmed by morphologic analysis, light and electron microscopy, and fluorescence detection. It was concluded that 1,25(OH)2D3 and RA exert additive effects on the inhibition of proliferation and the induction of apoptosis in both the NCI-H82 and the NCI-H209 SCLC cell lines. This finding has important implications for the use of retinoids and 1,25(OH)2D3 in cancer prevention and in the therapy of small cell lung carcinoma.     

Heterogenous response of B lymphocytes to transforming growth factor-beta in B-cell chronic lymphocytic leukaemia: correlation with the expression of TGF-beta receptors

We investigated the potential role of transforming growth factor-beta (TGF-beta) on spontaneous and cytokine-induced proliferation of B-cell chronic lymphocytic leukaemia (B-CLL) cells in vitro. Purified B lymphocytes from 21 B-CLL patients were cultured for 5 d in the presence of medium alone, IL-2 and/or IL-10, in the presence or absence of TGF-beta, and proliferation was measured by 3H-thymidine incorporation. TGF-beta inhibited B-cell proliferation in the majority of patients (15/21) but no inhibition was detected in 6/21 patients whatever the type of stimulant used. Addition of neutralizing antibodies to TGF-beta increased spontaneous and cytokine-induced proliferation; this effect was dose dependent and specific because addition of an irrelevant chicken IgG had no effect on B-CLL proliferation. In resistant patients, neutralizing antibodies to TGF-beta did not increase the proliferation. The expression of TGF-beta receptors on B-CLL cells was significantly lower than the one observed on normal CD5+ B lymphocytes for which the sensitivity to TGF-beta inhibition was more marked than in CLL. In addition, we found a strong correlation between the response of leukaemic B cells to TGF-beta inhibitory action and the expression of TGF-beta receptors on these cells. In summary, TGF-beta appears to function in CLL as a negative regulator of B lymphocytes but loss of responsiveness to this factor accompanied by a decrease of TGF-beta receptor expression, might provide a selective advantage to B-CLL lymphocytes.     

Influence of antirheumatic drugs on nitric oxide and interleukin 8 production in human articular chondrocytes

OBJECTIVE: To determine whether antirheumatic drugs can inhibit chondrocyte nitric oxide and interleukin 8 (IL-8) production. METHODS: IL-1beta stimulated human chondrocytes were incubated with indomethacin, methotrexate, sulfasalazine, dexamethasone, and methylprednisolone. Nitric oxide was detected as nitrite; IL-8 was detected by a radioimmunoassay method. RESULTS: Nitric oxide production was partly (< or = 50%) inhibited and IL-8 almost completely suppressed (> 80%) by dexamethasone and methylprednisolone. Sulfasalazine in pharmacological concentration and indomethacin slightly increased IL-8. No effect of methotrexate on nitric oxide or IL-8 production was found. CONCLUSION: Dexamethasone and methylprednisolone are inhibitors of human chondrocyte nitric oxide production, although to a lesser extent than for IL-8 production. Indomethacin, sulfasalazine, and methotrexate had no major influence on these mediators.     

A radiographic quality control system for the dental office

The sensitivity of 18 permanent hemopoietic cell lines to Cyclosporin A (CsA) was tested in a 3H-thymidine incorporation rate assay. Two human T cell lines (Molt4 and CEM) were significantly inhibited by a CsA concentration of 0.5 microgram/ml. Not affected at all or only inhibited by 10 to 20 times higher CsA concentrations were: three human B cell lines (4413a, Daudi, Raji), a monkey B cell line (B95-8), a mouse plasmocytoma line (X63-Ag8/653), a human non-B T cell line (Reh), four human myeloid lines (HL-60, ML-1, ML-2, ML-3), a human myelomonocytic line (Karpas 230), four human monoblastic lines (U 937, SU-DHL-1, THP-1, Karpas 241) and a human erythroid line (K 562). It therefore seems that among permanently growing hemopoietic cells a cell type specificity for T cells also exists.     

The Mayo Clinic-Canadian Cooperative trial of sulfasalazine in active multiple sclerosis

OBJECTIVE: To determine whether sulfasalazine is better than placebo in slowing disability progression in MS. METHODS: In this randomized, double-blind, placebo-controlled phase III trial, 199 patients with active relapsing-remitting (n = 151) or progressive (n = 48) MS were evaluated at 3-month intervals for a minimum of 3 years (94% completed 3 years of follow-up; mean follow-up, 3.7 years). MRI studies were performed at 6-month intervals on a subset of 89 patients. RESULTS: Sulfasalazine failed to slow or prevent disability progression as measured by the primary outcome (confirmed worsening of the Expanded Disability Status Scale [EDSS] score by at least 1.0 point on two consecutive 3-month visits). Sulfasalazine influenced favorably a number of secondary outcomes during the first 18 months of the trial (e.g., annualized relapse rate, proportion of relapse-free patients; progressive subgroup only: rate of EDSS progression at 1 and 2 years, median time to EDSS progression) but these positive findings were not sustained into the second half of the trial. CONCLUSIONS: Sulfasalazine does not prevent EDSS score progression in the subset of MS patients studied by this protocol. Treatments may improve relapse-related outcomes in MS, at least temporarily, without providing sustained slowing of EDSS progression. Phase III MS trials should be of sufficient length to determine a meaningful impact on disease course.     

Sphingosine 1-phosphate mobilizes sequestered calcium, activates calcium entry, and stimulates deoxyribonucleic acid synthesis in thyroid FRTL-5 cells

Sphingosine 1-phosphate (SPP) potently mobilizes sequestered calcium and is a mitogen in several cell types. In the present investigation, we have evaluated the effect of SPP on intracellular free calcium concentration ([Ca2+]i) and synthesis of DNA in thyroid FRTL-5 cells. SPP rapidly and transiently mobilized sequestered calcium and stimulated entry of extracellular calcium. The entry of calcium, but not the mobilization, was in part inhibited by pretreatment with pertussis toxin (Ptx), and by activation of protein kinase C. SPP did not stimulate the production of inositol 1,4,5-trisphosphate. SPP stimulated the incorporation of 3H-thymidine in a time- and dose-dependent manner. The effect was not inhibited by Ptx. Furthermore, SPP stimulated the activation of the proto-oncogene c-fos. SPP rapidly tyrosine-phosphorylated an approximately 66 kDa protein. This phosphorylation persisted for at least 1 h. Pretreatment of the cells with genistein abolished the SPP-evoked tyrosine phosphorylation, and attenuated the SPP-evoked increase in [Ca2+]i. Furthermore, the SPP-evoked activation of Na+-H+ exchange was inhibited by genistein. The phosphorylation was not attenuated by pretreatment of the cells with Ptx. SPP per se did not affect cellular cAMP levels but attenuated the TSH-evoked increase in cAMP. As the effect of SPP might be due to activation of phospholipase D, we tested whether phosphatidic acid (PA) mobilized calcium or stimulated the incorporation of 3H-thymidine. PA mobilized sequestered calcium but did not stimulate calcium entry. PA very modestly enhanced the incorporation of 3H-thymidine. Our results suggest, that SPP stimulates DNA synthesis and activates entry of calcium in FRTL-5 cells. The effect on calcium entry appears to be dependent, at least in part, on one or several tyrosine kinases.     

The repertoire of rheumatoid factor-producing B cells in normal subjects and patients with rheumatoid arthritis

OBJECTIVE: To compare the B cell repertoire of normal individuals and patients with rheumatoid arthritis (RA) and, specifically, to identify precursor B cells with the potential to secrete rheumatoid factor (RF) and to understand the T helper cell requirements for the production of this autoantibody. METHODS: Frequencies of precursors of IgM-, IgG-, and RF-producing B cells were measured in a limiting-dilution system. Two distinct sources of T cell help were compared. T cell help was provided by anti-CD3-activated CD4+ human T cell clones, or T cell-B cell interaction was facilitated by the bacterial super-antigen staphylococcal enterotoxin D (SED). RESULTS: A subset of 2-14% of peripheral blood B cells secreted IgM and IgG in SED-driven cultures. The SED-responsive B cell subpopulation was present at 10 times higher frequency in normal donors compared with RA patients. However, the repertoires were very similar, particularly for RF+ precursors, which represented approximately one-third of all SED-responsive B cells. In normal individuals, most of these RF+ precursor B cells did not respond to anti-CD3-activated T helper cells, with only a very small fraction of B cells activated by anti-CD3-driven helper cells maturing into RF-secreting B cells (from 1 of 182 to 1 of 889 IgM-producing B cells). This subset was expanded approximately 50-fold in RA patients. CONCLUSION: Normal subjects and RA patients share a pool of B cells which secrete RF when activated in the presence of SED and T helper cells. These B cells are frequent and obviously anergic in normal individuals. The B cell subset with the potential to produce RF when help is provided in noncognate T-B interaction (anti-CD3-driven T cells) is considerably expanded in RA patients, probably reflecting an increased responsiveness of such B cells to helper signals.     

Angiopeptin inhibits thymidine incorporation by explants of porcine coronary arteries

Angiopeptin, a stable octapeptide analog of somatostatin, inhibits proliferation in a variety of cancer cell lines. We studied the effect of angiopeptin on 3H-thymidine uptake into ring segments from the porcine coronary tree. The incorporation of 3H-thymidine into segments of porcine left anterior descending (LAD) coronary artery was time dependent and reached a plateau after 48 h. The addition of angiopeptin (48.1 and 96.2 nM) to the culture medium significantly inhibited 3H-thymidine incorporation into the segments by 36.7 +/- 10.1% and 48.3 +/- 2.3% of the control, respectively. Forskolin (100 microM), inhibited 3H-thymidine incorporation (52.7 +/- 10.1%) to the same degree as did angiopeptin (96.2 nM). Incubation of the segments with 125I-labeled angiopeptin, for 2 h at 37 degrees C, showed angiopeptin uptake to be time dependent and exhibited a first-order kinetics, reaching equilibrium after 30 min. Autoradiographic studies showed a uniform distribution of angiopeptin within the endothelium, media, and adventitia. Most of the labeling was associated with the nuclei of the cells. Angiopeptin, after 30-min incubation, did not significantly modify the basal levels of cyclic adenosine monophosphate (cAMP). In contrast, forskolin (100 microM) elicited a 50-fold increase of the basal levels of cAMP. These results indicate that in addition to its endocrine effects, angiopeptin reduces the rate of proliferation by acting directly on the vessel wall.     

Negative regulation of the neu promoter by the SV40 large T antigen

We investigated the effect of transforming growth factor-beta 1 (TGF beta) on the proliferation and differentiation of cultured acute promyelocytic leukemia (APL) cells with the chromosomal t(15;17) translocation obtained from four patients to determine the role of TGF beta on growth and differentiation of APL cells. DNA synthesis, determined by 3H-thymidine uptake, was inhibited in the presence and absence of granulocyte colony-stimulating factor (G-CSF) in a dose-dependent manner by TGF beta in APL cells obtained from three of the four cases. TGF beta and G-CSF did not significantly affect the differentiation of APL cells, but all-trans retinoic acid (RA) induced morphological and functional differentiation in all APL cells tested. G-CSF markedly enhanced RA-induced granulocytic differentiation in APL cells obtained from all four cases. In cells in which TGF beta inhibited DNA synthesis, it also inhibited RA-induced granulocytic differentiation of APL cells and, to a greater degree, granulocytic differentiation induced by RA plus G-CSF. These results suggest that TGF beta is a negative regulator of the proliferation and differentiation of APL cells. The significance of TGF beta as an endogenous regulator in differentiation therapy with RA of APL patients is discussed.     

Heparan sulfate proteoglycan on endothelium efficiently induces integrin-mediated T cell adhesion by immobilizing chemokines in patients with rheumatoid synovitis

OBJECTIVE: To clarify the role of heparan sulfate proteoglycan (HSPG) and chemokines in integrin-mediated T cell adhesion to endothelial cells in the synovium of patients with rheumatoid arthritis (RA). METHODS: Endothelial cells were purified from RA synovium. Expression of heparan sulfate, chemokines, and adhesion molecules on the endothelium was assessed by immunohistochemical analysis or flow cytometry. The effects of chemokines and heparan sulfate on T cell adhesion to RA endothelium were estimated with relevant antibodies and signaling inhibitors. Production of chemokines from synovial T cells was detected by Northern blotting and enzyme-linked immunosorbent assay. RESULTS: The endothelium in RA synovium highly expressed HSPG. The soluble form of chemokines, macrophage inflammatory protein 1beta (MIP-1beta), induced T cell adhesion to the endothelial cells. When MIP-lalpha and MIP-1beta were immobilized on RA endothelial cells, a more efficient integrin-mediated adhesion of T cells was induced compared with their soluble form. The induced T cell adhesion was reduced by pretreatment with either heparitinase, anti-MIP-lalpha antibody, or anti-MIP-lbeta antibody, indicating that these chemokines were bound to heparan sulfate on the cells. T cell adhesion was also inhibited by pertussis toxin, wortmannin, and cytochalasin B. MIP-lalpha and MIP-1beta were found on vessels in RA synovium in vivo, which were spontaneously produced from T cells purified from RA synovium. CONCLUSION: Endothelial cells in RA synovium characteristically express HSPG, which is involved in T cell integrin triggering by "posting" chemokines, which are produced by synovial T cells, and by "relaying" them to their receptors on T cells, which activate G protein-dependent phosphoinositide 3-kinase and actin-dependent integrin triggering.     

Ribosomal structure: RNA-protein and protein-protein interactions

Vitamin A and its derivatives have been postulated to play an important role in renal tubulogenesis and compensatory hypertrophy. This study examined the effects of two carboxylic derivatives of vitamin A on Lewis lung carcinoma-porcine kidney-1 (LLC-PK1) renal tubular epithelial cell mito- and motogenesis and cell size. It was found that all-trans and 13-cis retinoic acids exerted modest, dose-dependent effects to stimulate incorporation of 3H-thymidine into acid-precipitable material of LLC-PK1 cells. The effects of all-trans retinoic acid to promote 3H-thymidine uptake in LLC-PK1 cells modestly enhanced that seen with acidic fibroblastic growth factor. Similar findings of these two retinoic acid derivatives to promote 3H-thymidine uptake and to enhance 3H-thymidine uptake stimulated by another growth factor (platelet-derived growth factor BB) were also observed in cultured bovine aortic smooth muscle cells. Both retinoic acids promoted healing of denuded areas made within confluent monolayers of serum-starved LLC-PK1 cells. All-trans retinoic acid also stimulated recovery of mechanically denuded areas within bovine aortic smooth muscle monolayers. Neither all-trans nor 13-cis retinoic acids s affected cell size as assessed by forward light scatter with flow cytometry, suggesting lack of effect to induce hypertrophy. These results demonstrate that two carboxylic acid derivatives of vitamin A are capable of stimulation of basal and growth factor-induced incorporation of 3H-thymidine uptake into acid-precipitable material and healing of denuded areas in disparate cell types. These findings are compatible with a role for vitamin A and its analogues in the tissue repair process.     

Cell cycle regulation of human pancreatic cancer by tamoxifen

BACKGROUND: Clinical trials have suggested a survival advantage for selected patients with metastatic pancreatic cancer treated with tamoxifen. We sought to identify the molecular mechanism by which tamoxifen inhibits human pancreatic cancer cell (HPCC) growth. METHODS: HPCCs were grown in tamoxifen and growth inhibition was determined by 3H-thymidine uptake and by the MTT assay; changes in cell viability were determined by cell counts. Cell cycle alterations were evaluated by FACS, and the induction of apoptosis was evaluated using the TUNEL assay. Total cellular RNA was isolated after tamoxifen treatment, and Northern blot analysis was performed for p21waf1. RESULTS: Tamoxifen inhibited HPCC growth as measured by inhibition of 3H-thymidine incorporation and by the MTT assay. However, there was no decrease in the total number of viable cells after 6 days of treatment with 10 microM of tamoxifen and no evident apoptosis, confirming the absence of a cytotoxic effect. Cell cycle analysis revealed cellular arrest in the G0/G1 phase, which correlated with p21waf1 mRNA upregulation in response to tamoxifen treatment. CONCLUSIONS: Tamoxifen inhibits HPCC growth by inducing G0/G1 arrest with an associated increase in p21waf1 mRNA expression. Tamoxifen is an effective inhibitor of HPCC growth in vitro and warrants further in vivo study.     

Effect of lipoproteins on cultured human mesangial cells

It was recently reported that low-density lipoprotein (LDL) promotes mesangial cell proliferation, and oxidized LDL is cytotoxic for mesangial cells. However, there have been few studies about the effects of other lipoproteins on mesangial cells. Accordingly, we investigated the effect of various lipoproteins on cultured human mesangial cells using 3H-thymidine (3H-TdR) incorporation and cell counting assays. We also investigated the levels of several cytokines in mesangial cell culture supernatants after stimulation by the lipoproteins. Addition of very-low-density lipoprotein (VLDL) at concentrations up to 100 micrograms/mL, intermediate-density lipoprotein (IDL) at up to 50 micrograms/mL, and LDL at up to 50 micrograms/mL induced the proliferation of cultured human mesangial cells, whereas cell growth was inhibited at higher concentrations. Oxidized LDL caused a concentration-dependent decrease of 3H-TdR incorporation. High-density lipoprotein (HDL) had no proliferative effective effect at any concentration. Exposure to VLDL, IDL, LDL, or a high concentration of HDL enhanced the secretion of interleukin-6, platelet-derived growth factor, and transforming growth factor-beta by mesangial cells, whereas tumor necrosis factor-alpha secretion was stimulated by oxidized LDL. These finding indicate that triglyceride (TG)-rich lipoproteins (VLDL and IDL) promote mesangial cell proliferation as well as LDL, whereas oxidized LDL has the reverse effect. These effects of lipoproteins may be related to modulation of various cytokines. Accordingly, TG-rich lipoproteins, LDL, and oxidized LDL may be involved in mesangial cell proliferation and injury in patients with mesangial proliferative glomerulonephritis.     

The contribution of "time-of-flight" MRI-angiography in the study of neurovascular interactions (hemifacial spasm and trigeminal neuralgia)

Rat tracheal epithelial cells were cultured and the effects of LPS and TNF alpha on cell morphology, rate of proliferation and NO synthase activity were studied. NO synthase activity was determined by measuring the accumulation of 3H-L-citrulline during incubation of confluent monolayer with 3H-L-arginine. In untreated cells no significant 3H-L-citrulline formation was detected, and bradykinin and the calcium ionophore A 23187 failed to stimulate 3H-L-citrulline formation excluding a constitutively expressed, calcium-dependent NO synthase activity. After culturing the cells for 18 h in the presence of LPS (10 micrograms/ml) and TNF alpha (500 U/ml) a marked formation of 3H-L-citrulline could be detected, which was largely inhibited by N(G)-monomethyl-L-arginine (L-NMMA) indicating the induction of NO synthase activity which could be prevented by dexamethasone. Exposure of confluent monolayer to LPS and TNF alpha for up to 4 days resulted in a reduction in cell density by 20% within 1 to 2 days and in additional marked changes in cell morphology. The normal honeycomb-like structure of the culture was lost and a considerable number of cells developed "dendritic' outgrowths. These morphological changes as well as the reduction in cell density was attenuated by dexamethasone, but not by the NO synthase inhibitor L-NMMA. The rate of cell proliferation was determined in non-confluent cultures 24 h after passage by determination of the incorporation of tritium into DNA during 24 h of incubation with 3H-thymidine. 3H-thymidine incorporation was reduced by about 40-45% when LPS or TNF alpha was present during exposure to 3H-thymidine, and by about 65%, when LPS and TNF alpha were present in combination. Neither L-NMMA nor dexamethasone significantly affected the 3H-thymidine incorporation nor the inhibitory effects of LPS and TNF alpha. In conclusion, airway epithelial cells are markedly affected by LPS and TNF alpha and the various responses (changes in the cell morphology, inhibition of cell proliferation and induction of NO synthase activity) appear to be caused by different (dexamethasone-sensitive and -insensitive), cellular mechanisms. An enhanced formation of endogenous NO may not be responsible for the observed morphological changes or the inhibition of cell proliferation.     

Arterial heparan sulfate proteoglycans inhibit vascular smooth muscle cell proliferation and phenotype change in vitro and neointimal formation in vivo

PURPOSE: The aim of this study was to determine whether heparan sulfate proteoglycans (HSPGs) from the normal arterial wall inhibit neointimal formation after injury in vivo and smooth muscle cell (SMC) phenotype change and proliferation in vitro. METHODS: Arterial HSPGs were extracted from rabbit aortae and separated by anion-exchange chromatography. The effect of HSPGs, applied in a periadventitial gel, on neointimal formation was assessed 14 days after balloon catheter injury of rabbit carotid arteries. Their effect on SMC phenotype and proliferation was measured by point-counting morphometry of the cytoplasmic volume fraction of myofilaments (Vvmyo) and 3H-thymidine incorporation in SMCs in culture. RESULTS: Arterial HSPGs (680 microg) reduced neointimal formation by 35% at 14 days after injury (P=.029), whereas 2000 microg of the low-molecular-weight heparin Enoxaparin was ineffective. HSPGs at 34 microg/mL maintained subconfluent primary cultured SMCs with the same high Vvmyo (52.1%+/-13.8%) after 5 days in culture as did cells freshly isolated from the arterial wall (52.1%+/-15.1%). In contrast, 100 microg/mL Enoxaparin was ineffective in preventing phenotypic change over this time period (Vvmyo 38.9%+/-14.6%, controls 35.9%+/-12.8%). HSPGs also inhibited 3H-thymidine incorporation into primary cultured SMCs with an ID50 value of 0.4 microg/mL compared with a value of 14 microg/mL for Enoxaparin (P< .01). CONCLUSION: When used periadventitially in the rabbit arterial injury model, natural arterial HSPGs are effective inhibitors of neointimal formation. In vitro, the HSPGs maintain SMCs in a quiescent state by inhibiting phenotypic change and DNA synthesis. This study suggests that HSPGs may be a natural agent for the treatment of clinical restenosis.     

Epstein-Barr virus (EBV) can immortalize B-cll cells activated by cytokines

B-type of chronic lymphocytic leukemia (B-CLL) cells are inert to the potent transforming action of Epstein-Barr virus (EBV). The mitogenic action of Staphylococcus aureus Cowan I (SAC), MP6-thioredoxin, and interleukin 2 (IL-2), agents previously shown to induce proliferation in normal as well as in B-CLL cells, lifted this block, and EBV-positive cell lines could be established. It was not possible to establish cell lines of leukemic origin from cultures that were incubated with EBV alone or cytokine mix alone. CLL-cells infected with EBV only, expressed the viral nuclear antigen complex (EBNA), but not the viral latent membrane protein (LMP). They were not activated as measured by cell size and 3H-thymidine incorporation. In contrast, cells incubated with EBV and cytokine mix expressed both EBNA and LMP in parallel with enlargement and increased 3H-thymidine incorporation. These results emphasize that LMP expression is a prerequisite for growth transformation and immortalization and that cytokine activation signals are required for its expression in B-CLLs. Cells incubated with SAC/MP6-thioredoxin/IL-2 did not express any of the viral antigens, but were activated with regard to the mentioned parameters. Nine cell lines were established from six patients. From each of the three patients, we obtained 'twin'-pair lines: one corresponding to the malignant cell and the other to a normal B-lymphoblastoid cell. Thus, malignant and normal B-cell counterparts, from the very same donor, are at hand for comparative studies. The cell lines have been carried out for more than 12 months in culture. We conclude that B-CLL that are refractory to EBV-transformation can be rendered susceptible through in vitro cytokine activation.     

Relative biological effectiveness (RBE) of 252Cf fission neutrons for the induction of chromosome damage in human spermatozoa

Severe congenital insulin resistance in the syndrome of leprechaunism is caused by mutations in the insulin receptor gene. We report a patient with leprechaunism who was homozygous for a mutation resulting in the absence of cell surface insulin receptors. To determine whether the receptor for Insulin-like growth factor-I (IGF-I) is involved in the phenotype of leprechaunism, we studied the effect of insulin and of IGF-I on cells from this patient. The patient had a homozygous C-->T substitution at base pair 8212 in exon 12 of the insulin receptor gene, creating a premature stop codon. This nonsense mutation is in the extracellular portion of the receptor and truncates the insulin receptor proximal to its transmembrane anchor, resulting in the absence of cell surface insulin receptors. This finding indicates that complete absence of the insulin receptor is compatible with life. Secondly, DNA synthesis was studied in skin derived fibroblasts in response to increasing concentrations of either insulin or Insulin-like growth factor-I (IGF-I), and was assessed by 3H-thymidine incorporation. In this patient's cells, both of these hormones increased 3H-thymidine incorporation, and the effect was blocked by alpha-IR3, a monoclonal antibody that blocks activation of the IGF-I receptor. These findings confirmed the absence of the insulin receptor and indicated that insulin acts here through activation of the IGF-I receptor. These data support the contention that the phenotypic and metabolic abnormalities of leprechaunism result from the combination of lack of insulin receptor action and over-activation by insulin of the type 1 IGF receptor.     

Stimulation of leukaemic cells from adult T-cell leukaemia patients with bacterial superantigens

Bacterial superantigens stimulate T cells in a manner that is restricted to the Vbeta of the T-cell receptor. We examined the ability of adult T-cell leukaemia (ATL) cells to respond to these superantigens. Mononuclear cells from 10 patients were cultured with staphylococcal enterotoxin A (SEA), staphylococcal enterotoxin B (SEB) or toxic shock syndrome toxin-1 (TSST-1), and their response was determined by MTT assay and 3H-thymidine incorporation assay. Cells from six patients showed a specific response to a single superantigen. In two cases the cells responded to TSST-1 and bore Vbeta2, the known target of TSST-1. In three cases the cells responded to SEA with one bearing Vbeta9, a target of SEA, and one bearing Vbeta16. In one case the cells responded to SEB. Most of the cells which proliferated in response to superantigens were determined genetically to be leukaemic. The response to TSST-1 was inhibited by anti-Vbeta2 antibody. The responding cells showed a strongly enhancement expression of interleukin-2 receptor. These findings indicate that leukaemic cells from a proportion of ATL patients have an ability to respond to T-cell receptor-dependent superantigens. This suggests that bacterial infection in such patients may contribute to the expansion of ATL cells.