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Biomedical subjects

F Levi-Schaffer

Publications and source records attributed to F Levi-Schaffer.

At least 73 records · Page 4Linked to original sources
PubMed1995

Modulations of histamine release from mast cells by interleukin-2 is affected by nedocromil sodium.

We have previously demonstrated that histamine release from immunologically activated mast cells (MC) is enhanced by their preincubation (1 h) with interleukin-2(IL-2), and that IL-2 induces slow-chronic histamine release by MC in long-term cultures (6 days). In the present study we assessed whether nedocromil sodium can interfere with IL-2 modulation of MC histamine release. IL-2 enhancing effects nedocromil sodium activity were studied in cocultures of rat peritoneal MC with 3T3 fibroblasts (MC/3T3). MC/3T3 were preincubated for 1 h with IL-2 (50 micrograms/ml) and activated with either rabbit anti-rat IgE or compound 48/80. In chronic experiments MC/3T3 were long-term (5-6 days) incubated with IL-2 (50 micrograms/ml). Nedocromil sodium was used at 10(-5) M. MC activation both when added during the preincubation period (no tachyphylaxis was present) and when added together with the MC activators (30-50% inhibition). Washing out IL-2 before addition of the anti-IgE antibodies did not affect its histamine-release enhancing activity. Removal of nedocromil sodium before addition of the stimulus completely abrogated its effect. Continuous presence of IL-2 in the culture medium enhanced spontaneous histamine release by 37% and this effect was completely abolished in the presence of nedocromil sodium. Furthermore, nedocromil sodium decreased MC basal histamine release by 23% in long-term cocultures. Since IL-2 is known to be elevated in some pathological conditions, our results show that nedocromil sodium inhibits MC activation in an in vitro system which may represent a close resemblance to the in vivo allergic response.

Animals↗
PubMed1995

Activated mast cells are fibrogenic for 3T3 fibroblasts.

The extent of mast cell direct involvement in fibrosis is not defined as yet. In the present study we assessed whether long-term co-culture (up to 7 d) of functionally active rat peritoneal mast cells with 3T3 mouse fibroblasts and mass cell activation can affect fibroblast proliferation and collagen production. Co-culture of subconfluent 3T3 fibroblasts with resting mast cells or with mast cells stimulated by alpha IgE (1:35) or repeatedly activated by low concentrations of compound 48/80 (0.25-0.75 microgram/ml) did not alter fibroblast proliferation. However, fibroblast proliferation was increased significantly (100-130%) when mast cells were repeatedly activated with higher concentrations of compound 48/80 (1-3 micrograms/ml). Repeated mast cell activation by compound 48/80 (0.25 microgram/ml) caused a twofold increase in collagen production and this was reduced by 63% by the mast cell stabilizer nedocromil sodium (10(-5) M). At the same time, co-culture of 3T3 fibroblasts with unstimulated or immunologically activated mast cells did not modulate their collagen production. In conclusion, we have demonstrated that mast cell activation, under certain conditions, can enhance significantly 3T3 fibroblast proliferation and collagen production, thus indicating a direct mast cell involvement in the fibrotic processes.

3T3 Cells↗
PubMed1995

Challenge of mast cells with increasing amounts of antigen induces desensitization.

BACKGROUND: In vivo rush desensitization is a procedure widely employed to quickly desensitize allergic patients by administering increasing doses of the offending antigen at short intervals. The mechanism(s) underlying this process and the possible role of mast cells in it have not been well delineated. OBJECTIVE: To define an ex vivo model for rush desensitization utilizing the mast cell-fibroblast coculture system. METHODS: Rat peritoneal mast cells were cultured on 3T3 fibroblasts and were preincubated with saturating or suboptimal concentrations of IgE anti-DNP antibodies. Mast cells were then repeatedly challenged at 30 min intervals, with increasing amounts (0.001-100 ng/mL) of the antigen DNP-HSA, in the presence of calcium ions. Parallel control cultures were stimulated only once by each antigen concentration. In another set of experiments, mast cells were repeatedly activated with increasing concentrations (0.1-1000 ng/mL) of compound 48/80. Supernatants and cell sonicates were assessed for histamine content and the percentage of histamine released was calculated. RESULTS: When saturating concentrations of IgE anti-DNP antibodies were used, mast cells challenged repeatedly with DNP-HSA did not release significant amounts of histamine up to an antigen concentration of 1 ng/mL. At this stage they were partially desensitized, releasing only 108.3 +/- 17.1 ng histamine/plate (7.9 +/- 0.8%). A marked desensitization was observed at optimal antigen concentration (100 ng/mL), where experimental mast cells released only 45.6 +/- 10.9 ng/plate, compared with 661.9 +/- 48.2 ng/plate in firstly challenged cultures. Desensitization was probably not due to mast cells histamine depletion, since the cells still contained large amounts of histamine (579.5 +/- 108.6 ng/plate) at the end of the procedure. A similar pattern of desensitization was observed when mast cells were preincubated with a suboptimal concentration of IgE antibodies. Activation of mast cells with increasing amounts of the IgE-independent secretagogue, compound 48/80, did not cause desensitization since at each concentration both repetitively challenged and control cultures released similar amounts of histamine. Furthermore, challenge of antigen-desensitized mast cells with compound 48/80 caused the release of 75.9 +/- 4.9% histamine comparable to the percentage of histamine released from controls (79.5 +/- 6.7). CONCLUSION: Repeated exposure of mast cells to gradually increasing amounts of antigen induces their refractoriness. This observation would suggest a role for mast cells in rush desensitization procedure in vivo. Our coculture system may serve as a useful model for studying this process.

3T3 Cells↗
PubMed1995

Ketotifen therapy in chronic graft-versus-host disease (cGVHD): effect on mast cells and fibroblasts.

Current treatment options for cGVHD are limited. Mast cells (MC) and fibroblasts have been shown to play a role in the murine model of cGVHD. Ketotifen is an anti-H-1 antihistamine with MC-stabilizing properties. We therefore treated eight patients with cGVHD with ketotifen (6 micrograms/day for 3 months). Three additional age- and sex-matched cGVHD patients served as controls. MC count and activation state in cGVHD skin biopsies and the in vitro effect of peripheral blood mononuclear cell (PBMC) supernatants on (i) histamine release by MC; (ii) 3T3-fibroblast proliferation; and (iii) prostaglandin E2 (PGE2) production, were evaluated. Ketotifen therapy resulted in clinical improvement in 4/8 patients, stabilization of the disease in 2/8, while in 2/8 patients the cGVHD progressed and they died of bacterial sepsis. Side effects were minimal. In the skin biopsies the number of MC was found to be 0.58 +/- 0.17 (n = 8) (field x 400) and the MC looked degranulated (toluidine blue staining). Following ketotifen therapy MC number was increased to 1.2 +/- 0.28 (n = 8) (P < 0.05). PBMC supernatants of cGVHD patients stimulated histamine release from cultured rat MC (n = 8) (2.7 +/- 0.5 micrograms/ml; normal values are 2.1 +/- 0.4 micrograms/ml, n = 5). Ketotifen therapy reduced the histamine release level to the normal range (2.0 +/- 0.5 micrograms/ml, P < 0.05) (n = 8). Ketotifen therapy had no significant effect on: (i) 3T3 fibroblast proliferation which was suppressed by cGVHD PBMC supernatants; (ii) the elevated PGE2 production which we observed when fibroblasts were incubated with the PBMC supernatants. These results indicate that ketotifen may play a therapeutic role in cGVHD.

3T3 Cells↗
PubMed1995

Cytokine dysregulation in chronic graft versus host disease.

Cytokines play a key role in the pathogenesis of chronic Graft versus Host Disease (cGVHD) and various studies have shown abberant production of cytokines by immune cells from GVHD patients. Based on these findings and others showing that high TNF levels precede the development of GVHD, we evaluated inflammatory cytokine levels following BMT and during the development of cGVHD. In this study, patients undergoing bone marrow transplantation (BMT) who consequently developed chronic GVHD were analyzed as to their cytokine production during cGVHD and this was correlated with their clinical manifestations. A positive correlation was found between the severity as well as the number of major clinical complications and high levels of inflammatory cytokines (IL-1 beta IL-6 and TNF alpha) compared to control patients or to normal donors. Patients undergoing BMT who did not develop GVHD, did not produce high levels of IL-1 beta IL-6 or TNF. High levels of cytokines may be used as a tool for assessing novel therapeutic modalities and response to GVHD treatment.

Adolescent↗
PubMed1994

Mast cells and fibroblasts: two interacting cells.

Mast cell (MC) fibroblast interactions may have a role in health and disease. We analyzed the relationships between these cells by utilizing our in vitro model in which mucosal (MMC) and connective tissue (CTMC) type MC were cocultured long-term with different fibroblasts. Mouse 3T3 fibroblasts were used to provide a normal microenvironment for MC, while fibroblasts derived from mouse with chronic graft-versus-host disease (cGVHD) provided a fibrotic one. We found that both 3T3 and cGVHD fibroblasts maintain CTMC viability, phenotype and functional activity. When MMC were cocultured with 3T3 or cGVHD fibroblasts, they changed their phenotype towards that of CTMC. On the other hand, MCs were found to affect fibroblast properties. Coculture of MMC on 3T3 monolayers was shown to increase Forsmann antigen production and collagen synthesis and stimulate fibroblast proliferation. Resting CTMC or CTMC activated by anaphylactic stimuli induced 3T3 and cGVHD fibroblasts to proliferate more. In addition, CTMC activation increased collagen production by 3T3 fibroblasts. In conclusion, fibroblasts were found to regulate MC survival and differentiation, whereas MCs were shown to affect the biochemical properties of fibroblasts, which can lead to fibrosis.

3T3 Cells↗
PubMed1994

Mast cells in experimental allergic encephalomyelitis: characterization, distribution in the CNS and in vitro activation by myelin basic protein and neuropeptides.

Mast cells (MC) have been implicated in the pathogenesis of experimental allergic encephalomyelitis (EAE). In order to further evaluate their role, several morphological and functional studies were performed. Semiquantitative counts of histological sections showed a significant reduction in MC numbers in EAE brains. In addition, a higher proportion of EAE MC (about 50-70%) appeared degranulated compared with about 20% degranulation in controls. Central nervous system (CNS) MC exhibited staining properties of connective tissue MC and about 98% of them, both in diseased and control rats, were located in the thalamus. They were not present in the spinal cord and did not relate to EAE lesions. In vitro incubation of peritoneal MC (of connective tissue phenotype) with either MBP, or with neuropeptides such as substance P or bradykinin resulted in release of beta-hexosaminidase and histamine. The latter responses were similar in both EAE and control rats. It is suggested that the decrease in number and in granular content of CNS MC in EAE may reflect prior in vivo activation. The fact that MC were activated by MBP and by neuropeptides in vitro suggests a possible mechanism of MC activation in EAE.

Animals↗
PubMed1993

Proliferation and functional responses of bone marrow-derived mast cells after activation.

To investigate the behavior of mast cells following activation we have studied IL-3-dependent mouse bone marrow-derived mast cells (BMMC). Proliferation of BMMC presensitized with IgE anti-DNP antibodies was assessed after immunological stimulation with DNP-HSA. Proliferative potential of activated BMMC, as detected by [3H]thymidine incorporation, was similar to that of control BMMC during the first 6 days after activation while on Day 9 activated cells incorporated more [3H]thymidine. Both activated and control BMMC numbers slightly decreased 3-4 days after challenge with antigen and by Day 5 started to increase, reaching a maximum at Day 9. Immunologic activation of BMMC presensitized with IgE antibodies led to a profound loss of responsiveness to rechallenge with the same antigen 2-24 hr and 3-5 days later (70-100% inhibition of beta-hexosaminidase release). After 12 days cells were still partially unresponsive (20% inhibition). On the other hand, when BMMC were incubated again with IgE prior to rechallenge, desensitization was less pronounced and was resolved by Day 12. Activation of BMMC with an optimal concentration of thrombin induced only a partial unresponsiveness upon rechallenge 2-4 hr after activation (30% inhibition of beta-hexosaminidase release), while activation with calcium ionophore did not induce any desensitization. In both control and experimental BMMC cell-associated histamine remained stable for 12 days after challenge. We conclude that BMMC undergo a period of unresponsiveness which is more pronounced after immunological activation in contrast to nonimmunological ones, similarly to what previously shown for rat peritoneal mast cells.

Animals↗
PubMed1993

Interaction between mast cells and glial cells: an in vitro study.

Brain mast cells (MC) are located in close proximity to glial cells and it has been suggested that they belong to the connective tissue phenotype. To determine whether the local microenvironment provided by glial cells can influence mouse bone marrow-derived MC (BMMC), the putative counterpart of mucosal MC, we co-cultured these two cell types. BMMC numbers, morphology, histochemical properties and histamine content as well as glial cell morphology and function were evaluated up to 21 days. Our data indicate that BMMC adhere, proliferate, survive and can be activated to release histamine on the glial cell monolayers without changing their phenotype. Co-cultured glial cells preserve their morphological appearance and function throughout the culture period. These data indicate that central nervous system (CNS) glial cells do not induce phenotypic changes in BMMC and do not interfere with their viability and function.

Animals↗
PubMed1993

Investigations on the role of inflammatory and anti-inflammatory agents on the treatment of murine B cell leukemia by recombinant human interleukin-2.

Human recombinant interleukin-2 (rIL-2) was used in an effort to find effective biological therapy against the murine B-cell leukemia (BCL1), a spontaneous, nonimmunogenic, highly lethal leukemia of Balb/c origin. High dose rIL-2 (10(5) Cetus units 3 times a day for 5 days) was proven to be curative for mice inoculated with up to 10(4) BCL1 cells. Considering the fact that mice which showed more clinical signs of rIL-2 related toxicity had higher leukemia-free survival, and in view of the fact that BCL1 is a totally nonimmunogenic tumor, we have investigated the possible role of nonspecific effects of inflammatory and anti-inflammatory agents in order to speculate on the possible therapeutic role of inflammation on the overall effect of rIL-2 in the treatment of BCL1. According to the results presented in this work, it appears that although mediators of inflammation may play some role against leukemia, their overall effect in comparison with high-dose rIL-2 is relatively insignificant.

Animals↗
PubMed1992

Serosal mast cells maintain their viability and promote the metabolism of cartilage proteoglycans when cocultured with chondrocytes.

OBJECTIVE: To determine the consequences of mast cell (MC)-chondrocyte interactions. METHODS: Cocultured cells were analyzed histochemically, morphologically, biochemically, and functionally. RESULTS: Cocultured MC adhered to the chondrocytes and remained viable. Chondrocytes cocultured with nonactivated MC produced more proteoglycans than did chondrocytes cultured alone, and these proteoglycans possessed an intact hyaluronic acid-binding region. In contrast, most of the proteoglycans produced by chondrocytes cocultured with activated MC were degraded. CONCLUSION: These studies indicate that a complex interaction occurs in which the nonactivated MC stimulates biosynthesis and the activated MC degrades cartilage proteoglycans.

Animals↗
PubMed1992

Effect of coculture of rodent mast cells with murine chronic graft-versus-host disease (cGVHD)-derived fibroblasts.

We investigated whether murine chronic graft-versus-host disease (cGVHD)-derived skin fibroblasts maintain the viability and functional activity of rat peritoneal connective tissue-mast cells (CTMCs) and whether they affect the change in phenotype of mouse bone marrow-derived mast cells (BMMCs). Skin fibroblasts were isolated before the development of fibrosis (day 5), during overt fibrosis (day 28), and after recovery from fibrosis (day 120). cGVHD fibroblasts of days 5 and 28 exhibited enhanced proliferation, a property that was maintained through several subcultures. CTMCs adhered to the same extent, did not divide, and maintained their viability in all the different cultures. When CTMCs were activated with compound 48/80, they released approximately 80% of their histamine content, indicating that coculture with cGVHD fibroblasts did not adversely affect CTMC function. The amount of histamine found in the medium of 8 days CTMC/cGVHD fibroblast coculture was similar to that found in control culture. These findings suggest that the degranulation of dermal MCs, characteristic of cGVHD, is not due to a direct activating effect of the cGVHD fibroblasts on the MCs. BMMCs seeded on cGVHD fibroblasts acquired the capacity for safranin staining and increased their histamine content, indicative of a change to CTMCs. Thus, cGVHD fibroblasts are able to provide a microenvironment adequate for maintaining viability and activity of CTMCs and for promoting maturation and change in phenotype of BMMCs.

Animals↗
PubMed1991

Mast cell activity in experimental allergic encephalomyelitis.

The number and functional reactivity of peritoneal mast cells (MCs) were evaluated in rats with experimental allergic encephalomyelitis (EAE). Cells were counted following staining with toluidine blue and activation was measured by B-hexosaminidase (B-hex) release. The number of detectable MCs and their capacity to release B-hex decreased significantly by 40 and 65%, respectively, as compared with normal controls just prior to the onset of clinical signs. These values returned to normal on clinical recovery. Preliminary data on MC counts performed on histological sections of rat brains with EAE suggested a similar pattern of response, i.e., an early decrease prior to disease onset with subsequent normalization on recovery. In an attempt to modify the course of EAE, rats were treated with the MC stabilizing agent nedocromil or with the MC activating agent, compound 48/80. Nedocromil induced a slight delay in the onset of EAE, but only when administered at the time of EAE induction. Compound 48/80 did not seem to affect the clinical course of the disease. Our results suggest that MCs are involved in the pathogenesis of EAE and may contribute to the induction of the disease rather than to the effector phase and its clinical expression.

Animals↗
PubMed1991

Desferal (desferrioxamine)--a novel activator of connective tissue-type mast cells.

The effect of Desferal (desferrioxamine), an iron-chelating agent with allergic side effects, was examined on human basophils and rodent mast cells (MCs) in vitro and in the human skin. Even at a high concentration (100 mg/ml), the drug neither induced histamine release (HR) from human basophils nor primed these cells to release higher amounts of histamine when they were activated with f-met-peptide or anti-IgE antibodies. In contrast, in all seven subjects studied, intradermal injection of Desferal (0.1 mg/ml to 100 mg/ml) elicited classic wheal-and-flare responses. Ingestion of 10 mg of cetirizine, an H1 antagonist, 3 hours before the intracutaneous administration of Desferal, significantly reduced the diameters of both wheal-and-flare reactions, indicating that the drug caused local HR. Desferal also induced HR from rat peritoneal MCs in vitro but had no effect on mouse bone marrow-derived MCs. These results suggest that Desferal has a direct, IgE-independent, stimulatory effect on connective tissue-type MCs. Thus, it may be used as a positive control in skin testing.

Animals↗