Biomedical subjects

C N Abboud

Publications and source records attributed to C N Abboud.

At least 19 recordsLinked to original sources

PubMed2004

Phase I study for poor-prognosis lymphoma: augmentation of the "BEAM" regimen with escalating dose melphalan using amifostine cytoprotection and autologous hematopoietic stem cell transplantation--a preliminary report.

We and others have previously shown that the use of amifostine (Ethyol; MedImmune Inc, Gaithersburg, MD) can ameliorate certain regimen-related toxicities of high-dose melphalan (HD-MEL) in the autologous hematopoietic stem cell transplant setting. Our recent experience indicated that the maximum tolerated dose of HD-MEL plus autologous hematopoietic stem cell transplant could be increased from approximately 200 mg/m2 to at least 280 mg/m2 with amifostine. Although a dose-limiting toxicity was not clearly identified, atrial fibrillation was noted in several patients. Phase II trials using this regimen have been reported in lymphoma and myeloma. Nonetheless, it is unlikely that single agent therapy, regardless of dose, will be highly curative in advanced hematologic malignancy. Thus, we used amifostine to permit dose escalation of HD-MEL within the BEAM (BCNU/etoposide/arabinosylcytosine/HD-MEL) combination chemotherapy regimen before autologous hematopoietic stem cell transplant in selected patients with lymphoma. Patient entry at the starting dose (ie, HD-MEL 140 mg/m2) has been completed without the development of severe regimen-related toxicities. This trial is ongoing.

Adult↗

PubMed2003

Effects of the farnesyl transferase inhibitor R115777 on normal and leukemic hematopoiesis.

Patients with acute myelogenous leukemia or myelodysplastic syndrome may respond to farnesyl transferase inhibitors (FTIs) with partial or complete response rates noted in about 30% of such patients. FTIs prevent the attachment of a lipid farnesyl moiety to dependent proteins prior to their insertion into the plasma membrane and thereby prevent activity of these prenylation-dependent proteins, but their mechanism of tumor suppression remains unknown. Many patients receiving FTIs do experience myelosuppression. In this work, the in vitro effects of the FTI, R115777 on normal and leukemic hematopoiesis have been examined as have its effects on apoptosis induction and cell cycle profile in both leukemic blasts and normal CD34+ cells. R115777 was inhibitory to normal CD34+ cell proliferation and to leukemic blast cells, but did not affect long-term culture initiating cell frequency nor NOD-SCID reconstituting capacity. No induction of apoptosis or cell cycle changes were noted in AML blasts. These data suggest that myelosuppression with R115777 occurs largely at the intermediate to late progenitor stage of hematopoiesis and that cyclic use might avoid long-term marrow suppression.

Alkyl and Aryl Transferases↗

PubMed2001

Response of human CD34+ cells to CXC, CC, and CX3C chemokines: implications for cell migration and activation.

Ultrastructural studies of marrow and examination of the in vivo processes of stem cell homing and mobilization show that multipotential hematopoietic progenitors are able to traverse endothelial cells. The regulation of this process by various classes of chemokines was studied in this report, using an in vitro model of transendothelial migration. Human umbilical vein endothelial cells (HUVECs) or bone marrow-derived endothelial cells (BMECs) were grown to confluence on 3-microm microporous membrane inserts and placed in 24-well culture plates. CD34(+) cells isolated from normal volunteer donor marrow by immunoadsorption or magnetic bead selection techniques were added to the inserts and various individual chemokines were added to the lower chamber of the culture plates in serum-free conditions. After 24 h, the percentage of transmigrated cells was determined. A mean of 8.5% of unfractionated marrow CD34(+) populations migrated, and all chemokines tested, with the exception of macrophage inflammatory protein-1alpha (MIP-1alpha), had some positive effect on this migration. The greatest effects were seen with stroma-derived factor-1alpha (SDF-1alpha) and stroma-derived factor-1beta (SDF-1beta), with lesser effects noted for other chemokines and cytokines. When the CD34(+) population was subselected for expression of CD38, a greater fraction of the CD38(-) cells migrated as compared to the CD38(+) fraction. CD34(+) cells isolated from mobilized peripheral blood and cord blood also migrated in response to chemokines. Chemokines of the CC, CXC, and CX(3)C classes as well as other hematopoietic cytokines may modulate the process of stem cell transmigration of endothelial cells. Further understanding of this process may help elucidate the mechanism of stem cell mobilization and homing.

Antibodies↗

PubMed2000

MIP-1alpha and TGF-beta production in CD34+ progenitor-stromal cell coculture systems: effects of progenitor isolation method and cell-cell contact.

ABSTRACT Macrophage inflammatory protein-1alpha (MIP-1alpha) is a C-C chemokine which has antiproliferative effects on early hematopoietic progenitors and stimulatory effects on later progenitors. It also possesses chemotactic and activating properties for monocytes, macrophages, and T-cells. CD34+ progenitors isolated utilizing an avidin-biotin immunoadsorption column produced significant amounts of MIP-1alpha from 24 h onward when cultured in medium with 10% fetal calf serum (>200 pg/ml). Such production persisted through 96 h of culture and was greater when such progenitors were cocultured with a preformed marrow stromal layer (4000 pg/ml at 24 h). The production of MIP-1alpha declined over time of coculture with stromal layers, and stromal layers themselves produced minimal MIP-1alpha as detected by ELISA: <100 pg/ml. In contrast, CD34+ cells isolated by flow cytometry or by magnetic bead adsorption produced minimal MIP-1alpha (0-30 pg/ml). MIP-1alpha production also increased when cells isolated by these two methods were cocultured with stromal layers. The difference in MIP-1alpha production could not be accounted for by differences in purity of the CD34+ population between isolation methods nor on the basis of monocytic or lymphocytic contamination as assessed by the presence of CD14 or CD3 positive cells. CD34+ cells isolated by immune adsorption had increased expression of endothelial and mesenchymal associated antigens, however, suggesting that this subpopulation might account for the MIP-1alpha production observed. Freshly isolated CD34+ cells expressed MIP-1alpha message as assessed by RT-PCR and by in situ hybridization. Coculture of CD34+ cells isolated by any means with stromal cells increased transforming growth factor-beta (TGF-beta) production, in this case by the stromal layer itself. Both MIP-1alpha and TGF-beta have been found to influence cell cycle status and proliferation status of early hematopoietic progenitors, and both have potential effects on accessory cell function. These studies indicate that progenitor-stromal cell interactions may influence local cytokine output, thus potentially influencing progenitor cycling status and accessory cell activation. The method of isolation of CD34+ progenitors may influence secretion of certain cytokines and chemokines.

Animals↗

PubMed2000

Allogeneic bone marrow transplantation for children with acute leukemia: cytoreduction with fractionated total body irradiation, high-dose etoposide and cyclophosphamide.

Marrow-ablative chemo-radiotherapy followed by hematopoietic stem cell rescue from an allogeneic source improves outcomes for children with high-risk acute leukemia. The first effective pre-transplant preparative regimens consisted of high-dose cyclophosphamide (CY) and total body irradiation (TBI). Subsequent attempts have been made to improve leukemia-free survival, by adding other chemotherapy agents to these agents. In previous clinical studies of total body irradiation, etoposide, cyclophosphamide (TBI-VP-16-Cy) in adult allogeneic bone marrow transplantation, there has been a high incidence of severe regimen-related toxicity. In this study, we investigated the safety and efficacy of this combination in 41 children who received TBI (12-14 Gy), VP-16 (30 mg/kg), and CY (60 mg/kg x 2) and then either matched sibling or alternative donor transplants for acute leukemia. There was only one case of fatal regimen-related toxicity. The estimated 3-year event-free survival for patients with early or intermediate stage disease was 68% (53-88%). The estimated event-free survival of patients with advanced disease was 17% (5-59%). TBI-VP16-CY is safe in pediatric transplantation, and it has good efficacy for transplant recipients with less advanced disease. Bone Marrow Transplantation (2000) 25, 489-494.

Acute Disease↗

PubMed2000

Mobilization of peripheral-blood stem cells by concurrent administration of daniplestim and granulocyte colony-stimulating factor in patients with breast cancer or lymphoma.

PURPOSE: To evaluate the safety and hematopoietic activity of daniplestim administered concurrently with granulocyte colony-stimulating factor (G-CSF) for peripheral-blood stem-cell (PBSC) mobilization. PATIENTS AND METHODS: In the initial dose-escalation phase, 25 patients with adenocarcinoma of the breast (AB; 13 patients) or lymphoma (12 patients) were given daniplestim at doses ranging from 0.1 to 3.75 microgram/kg/d plus G-CSF 10 microgram/kg/d. In the randomized phase, 52 patients with AB (27 patients) or lymphoma (25 patients) were randomized within disease categories to the daniplestim dose chosen in the dose-escalation phase plus G-CSF 10 microgram/kg/d (D+G) or placebo plus G-CSF 10 microgram/kg/d (P+G) for up to 7 days. RESULTS: A daniplestim dose of 2. 5 microg/kg/d was chosen for further study because it was hematopoietically active and had an acceptable side-effect profile. In the randomized phase, in patients with AB, D+G was associated with a higher probability (P =.0696) of collecting >/= 2.5 x 10(6) CD34(+) cells/kg and significantly higher circulating CD34(+) cell counts (P =.0498) on days 6 through 9 after the initiation of dosing. The target level was more likely to be reached with additional leukaphereses in the patients given D+G. Patients given P+G did not benefit from additional leukaphereses beyond the first procedure. The type of mobilization did show a trend toward a shorter duration of neutropenia in the D+G group. The adverse events with D+G consisted largely of mild to moderate flu-like symptoms, including headache and fever, and occurred more frequently than with P+G. CONCLUSION: Daniplestim administered at 2.5 microgram/kg/d is tolerable and active when combined with G-CSF, and the combination may prove more effective than G-CSF alone in promoting the collection of adequate numbers of CD34(+) cells for PBSC infusion in patients with AB.

Adenocarcinoma↗

PubMed1999

Human immunodeficiency virus and the hematopoietic repertoire: implications for gene therapy.

The human immunodeficiency virus (HIV) infects primarily the hematopoietic and immune systems. At the onset of infection, an initial activation of the immune system occurs, with a subsequent suppression thereafter due to direct viral infection of cells, inhibitory effects of HIV proteins, an altered microenvironment with cytokine imbalance, and increased apoptosis of both infected and non-infected cells. The CD34+ hematopoietic stem cell, however, remains free of infection. Novel methods in gene therapy utilize viral vectors that can introduce genes with good efficiency into the non-dividing stem cell. Therefore, HIV-resistance genes can be introduced into stem cells using these vectors. This confers resistance to infection to their respective progeny, and concurrently allows for repopulation of the immuno-hematopoietic repertoire. Applications of this technology to the patient infected with HIV are discussed in the context of myeloablative therapy and stem cell rescue.

Antigens, CD34↗

PubMed1999

Treatment of chronic myelogenous leukemia with autologous bone marrow transplantation followed by roquinimex.

Unmanipulated autologous bone marrow transplant (ABMT) offers patients with chronic myelogenous leukemia (CML) a long-term survival of 10%, at best. Immunotherapy has a role in the myeloid leukemias, and there is increasing evidence that of all hematopoietic neoplasms, CML may be the most susceptible to immune regulation. Roquinimex is known to enhance T cell, NK cell and macrophage activity. A phase II study was initiated in March 1992 to evaluate the role of roquinimex in Ph chromosome-positive CML post ABMT. Patients were conditioned with busulfan/ cyclophosphamide followed by reinfusion of unmanipulated Ph-positive bone marrow stem cells (>1 x 108 NBC/kg). When engraftment of neutrophils (ANC) reached 100/microl, patients received oral roquinimex twice weekly, escalating to a maximal dose of 0.2 mg/kg in 2 weeks. Seventeen patients have entered the study; 11 in first chronic phase (CP1); two in second chronic phase (CP2) and four in accelerated phase (AP). All required significant myelosuppressive therapy prior to ABMT to maintain stable blood counts and most had also received prior interferon therapy. All patients survived the transplant. Subsequent toxicity consisted mainly of musculoskeletal aches and peripheral edema. Additionally, specific skin changes were observed including graft-versus-host-like disease and eccrine sweat gland necrosis. Eight out of 17 patients are alive 28-60 months post ABMT. Of the nine patients who died, two were in CP2 and three in AP. All patients in CP1 went into a complete hematological remission post ABMT and seven of the 11 patients had at least a major cytogenetic response (greater than 65% Ph-negative metaphases) at 1 year or beyond and four of the 11 patients had a complete cytogenetic response at 2 years or beyond. Cytogenetic response post transplant often developed over time and did not simply represent post ABMT engraftment with Ph-negative cells. The clinical and cytogenetic data in these patients are encouraging and suggest that roquinimex may have significant activity when given post ABMT to patients with Ph-positive CML.

Adult↗

PubMed1999

Analysis of factors that correlate with mucositis in recipients of autologous and allogeneic stem-cell transplants.

PURPOSE: To identify predictors of oral mucositis and gastrointestinal toxicity after high-dose therapy. PATIENTS AND METHODS: Mucositis and gastrointestinal toxicity were prospectively evaluated in 202 recipients of high-dose therapy and autologous or allogeneic stem-cell rescue. Of 10 outcome variables, three were selected as end points: the peak value for the University of Nebraska Oral Assessment Score (MUCPEAK), the duration of parenteral nutritional support, and the peak daily output of diarrhea. Potential covariates included patient age, sex, diagnosis, treatment protocol, transplantation type, stem-cell source, and rate of neutrophil recovery. The three selected end points were also examined for correlation with blood infections and transplant-related mortality. RESULTS: A diagnosis of leukemia, use of total body irradiation, allogeneic transplantation, and delayed neutrophil recovery were associated with increased oral mucositis and longer parenteral nutritional support. No factors were associated with diarrhea. Also, moderate to severe oral mucositis (MUCPEAK > or = 18 on a scale of 8 to 24) was correlated with blood infections and transplant-related mortality: 60% of patients with MUCPEAK > or = 18 had positive blood cultures versus 30% of patients with MUCPEAK less than 18 (P =.001); 24% of patients with MUCPEAK > or = 8 died during the transplantation procedure versus 4% of patients with MUCPEAK less than 18 (P =.001). CONCLUSION: Gastrointestinal toxicity is a major cause of transplant-related morbidity and mortality, emphasizing the need for corrective strategies. The peak oral mucositis score and the duration of parenteral nutritional support are useful indices of gastrointestinal toxicity because these end points are correlated with clinically significant events, including blood infections and treatment-related mortality.

Adolescent↗

PubMed1998

A RANTES-antibody fusion protein retains antigen specificity and chemokine function.

The successful eradication of cancer cells in the setting of minimal residual disease may require targeting of metastatic tumor deposits that evade the immune system. We combined the targeting flexibility and specificity of mAbs with the immune effector function of the chemokine RANTES to target established tumor deposits. We describe the construction of an Ab fusion molecule with variable domains directed against the tumor-associated Ag HER2/neu, linked to sequences encoding the chemokine RANTES (RANTES.her2.IgG3). RANTES is a potent chemoattractant of T cells, NK cells, monocytes, and dendritic cells, and expression of RANTES has been shown to enhance immune responses against tumors in murine models. RANTES.her2.IgG3 fusion protein bound specifically to HER2/neu Ag expressed on EL4 cells and on SKBR3 breast cancer cells as assayed by flow cytometry. RANTES.her2.IgG3 could elicit actin polymerization of THP-1 cells and transendothelial migration of primary T lymphocytes. RANTES.her2.IgG3 prebound to SKBR3 cells also facilitated migration of T cells. RANTES.her2.IgG3 bound specifically to the CCR5 chemokine receptor, as demonstrated by flow cytometry, and inhibited HIV-1 infection via the CCR5 coreceptor. RANTES.her2.IgG3, alone or in combination with other chemokine or cytokine fusion Abs, may be a suitable reagent for recruitment and activation of an expanded repertoire of effector cells to tumor deposits.

Amino Acid Sequence↗

PubMed1998

Autotransplantation for relapsed or refractory Hodgkin's disease: long-term follow-up and analysis of prognostic factors.

Seventy consecutive patients with refractory or relapsed Hodgkin's disease who received high-dose chemotherapy followed by autologous stem cell rescue were analyzed to identify clinically relevant predictors of long-term event-free survival. High-dose therapy consisted primarily of carmustine (BCNU), etoposide, cytarabine and cyclophosphamide (BEAC). The 5-year Kaplan-Meier event-free survival (EFS) for the entire cohort was 32% (95% confidence interval; 18-45%) with a median follow-up of 3.6 years (range 7 months-7.6 years). The most significant predictor of improved survival was the presence of minimal disease (defined as all areas < or =2 cm) at the time of transplant: the 5 years EFS was 46 vs 10% for patients with bulky disease (P = 0.0002). Other independent predictors identified by step-wise regression analysis included the presence of non-refractory disease and the administration of post-transplant involved-field radiotherapy (XRT). Treatment-related mortality occurred in 13 of 70 patients: nine patients (13%) died within the first 100 days, mainly from cardiopulmonary toxicity. However, only one of 24 patients (4%) transplanted during the last 4.5 years died from early treatment-related complications. While high-dose therapy followed by autotransplantation led to long-term EFS of 50% for patients with favorable prognostic factors, a substantial proportion of patients relapsed, indicating that new therapeutic strategies are needed.

Adult↗

PubMed1998

Effect of stromal cell coculture on progenitor cell expansion and myeloid effector function in vitro.

Stimulation of CD34(+)-enriched marrow or light density marrow with various growth factor combinations can generate granulocyte progenitors and mature neutrophils in vitro. In this work, we have examined the influence of irradiated marrow stromal layers on growth factor-induced myeloid and early multipotential progenitor expansion from enriched marrow CD34+ progenitors. We have also explored whether the addition of early-acting growth factors known to enhance myelopoiesis in long-term culture, such as fibroblast growth factor (b-FGF), insulin growth factor (IGF-1), c-kit ligand or stem cell factor (SCF), and flk-2flt-3 ligand (FL), can lengthen survival of CD34+ progenitors in these cultures. Stromal cell coculture resulted in greater numbers of total cells and CFU-GM at day 7 and day 14, but with the addition of multiple growth factors, these effects of stromal cell coculture were diminished. At day 14, generally < 1% of the expanded cells over stromal coculture conditions were CD34+, with up to 90% demonstrating CD15 positivity. Culture of CD34+ cells in the presence of early-acting growth factors did not cause significant expansion of CD34+ cells over a 14-day life span, even in the presence of marrow stromal cells. These data suggest that although stromal cell coculture for a period up to 14 days can enhance expansion of total cell numbers and CFU-GMs, stromal cell presence does not lead to expansion of CD34+ cells in these cultures and may diminish the number of clonogenic cells present when growth factors with differentiating capacity are present. Mature neutrophils harvested from such cultures are capable of chemotaxis, actin polymerization, and migration, suggesting a replete functional status.

Actins↗

PubMed1997

TGF-beta down-regulates stromal IL-7 secretion and inhibits proliferation of human B cell precursors.

Development of lymphoid progenitors in vivo requires interaction with a bone marrow stromal microenvironment containing multiple cytokines involved in the development of nonlymphoid hemopoietic lineages. We tested the effect of one such cytokine, TGF-beta, on the proliferation of early human clonogenic lymphoid progenitors using a stroma-dependent in vitro culture system. TGF-beta caused a dose-dependent inhibition of lymphoid progenitor colonies that was reversible at low TGF-beta doses by addition of exogenous IL-7 to the cultures. IL-7 was unable to reverse the inhibitory effect of higher TGF-beta concentrations or inhibition caused by IL-1alpha, IL-4, or TNF-alpha. Stromal IL-7 mRNA expression and protein secretion were markedly down-regulated by TGF-beta, suggesting that inhibition of stromal IL-7 secretion partially accounts for the inhibitory effect of TGF-beta on lymphopoiesis in this culture system. It is likely that higher TGF-beta concentrations do not inhibit lymphopoiesis by down-regulating IL-7 receptor expression, since this cytokine did not reduce IL-7R alpha or gamma c mRNA levels in normal B cell precursors. Since direct stromal contact is required for in vitro lymphopoiesis, the potential regulation of the IL-7 pathway by cell adhesion was examined. Adhesion of human B cell precursors to stroma did not alter stromal IL-7 expression or expression of IL-7R alpha or gamma c-chains by B cell precursors. These results indicate that TGF-beta is a significant negative regulator of stroma-dependent proliferation of early human lymphoid progenitors and acts in part by down-regulating stromal IL-7 secretion.

B-Lymphocytes↗

PubMed1997

Expression of interleukin-7 receptor by lineage-negative human bone marrow progenitors with enhanced lymphoid proliferative potential and B-lineage differentiation capacity.

Relatively little is known about the relationship of lymphoid-associated gene expression to the proliferation and differentiation potential of early human bone marrow lymphoid progenitors. Surface expression of interleukin-7 (IL-7) receptor-alpha (IL-7R alpha), a component of the high-affinity receptor for the lymphoid precursor growth factor IL-7, defined a CD34+ progenitor subset lacking the CD19+ pro-B phenotype but demonstrating markedly enhanced lymphoid clonogenic capacity and the ability to differentiate into pro-B cells in short-term culture. These progenitors expressed mRNA for the lymphoid-associated genes Ig beta, RAG-1, and PAX-5, and were uniformly TdT-positive (TdT+). In contrast, IL-7R alpha-/CD19-/ CD34+ progenitors had a 50-fold reduced lymphoid clonogenic capacity and did not differentiate into pro-B cells in short-term culture. Expression of TdT and the lymphoid-associated genes Ig beta and RAG-1, but not PAX-5, was detected in this fraction, although at lower levels than in the IL-7R alpha+ progenitors. In contrast to IL-7R alpha, loss of the stem cell factor receptor c-kit was associated with enhanced lymphoid clonogenic potential and increased B-lineage differentiation potential. These results indicate that IL-7R alpha expression defines entry into a developmental stage characterized by upregulation of multiple lymphoid-associated genes and enhanced fitness for B-lymphoid differentiation. The onset of IL-7R alpha and PAX-5 expression immediately before acquisition of CD19 is consistent with evidence suggesting upregulation of CD19 through pathways involving PAX-5 and IL-7.

Adult↗

PubMed1997

Autotransplantation for relapsed or refractory non-Hodgkin's lymphoma (NHL): long-term follow-up and analysis of prognostic factors.

One hundred and thirty-six patients autografted for relapsed or refractory non-Hodgkin's lymphoma (NHL) were evaluated to assess long-term event-free survival and to identify important prognostic factors. High-dose therapy consisted primarily of carmustine (BCNU), etoposide, cytarabine, and cyclophosphamide (BEAC) followed by unpurged autologous stem cell rescue. The 5-year Kaplan-Meier event-free survival (EFS) for the entire cohort was 34% (95% confidence interval: 24-44%) with a median follow-up of approximately 3 years (range 0-7.5 years). For patients entering with minimal disease (defined as all areas < or = 2 cm), the 5-year EFS was 40 vs 26% for those entering with bulky disease (P = 0.0004). In the multivariate analysis, minimal disease on entry and administration of involved-field XRT post-transplant were significantly associated with improved EFS; the latter association was observed mainly in the cohort of patients with bulky disease. The overall 100-day treatment-related mortality rate was 4.4% (3% for the last 71 patients). New strategies are needed to reduce the high rate of relapse (50-60%) following auto-transplantation for relapsed or refractory NHL.

Adolescent↗

PubMed1997

Successful treatment of a generalized human papillomavirus infection with granulocyte-macrophage colony-stimulating factor and interferon gamma immunotherapy in a patient with a primary immunodeficiency and cyclic neutropenia.

BACKGROUND: We describe a patient with generalized verrucosis secondary to human papillomavirus (HPV) type 2 infection and a primary immunodeficiency and cyclic neutropenia. Treatment, which was well tolerated, included granulocyte-macrophage colony-stimulating factor and interferon gamma (IFN-gamma). In vitro assays to assess responses of T lymphocytes to mitogens (ie, proliferation assay and IFN-gamma enzyme-linked immunosorbent assay) were performed. In situ hybridization and polymerase chain reaction were used to detect HPV DNA in skin biopsy specimens. OBSERVATIONS: The T lymphocytes of the patient showed a significant (P < .05, unpaired Student t test) defect in IFN-gamma production (the basis for initiating IFN-gamma therapy). The response to immunotherapy was confirmed by using molecular methods. Six months after the completion of immunotherapy, HPV DNA was undetectable in skin samples from clinically regressed warts (according to the results of in situ hybridization and polymerase chain reaction). CONCLUSIONS: Our patient had treatment-resistant generalized verrucosis for 13 years. Treatment with granulocyte-macrophage colony-stimulating factor and IFN-gamma may have reconstituted an immune response against HPV, resulting in the dramatic regression of her widespread warts.

Adolescent↗

PubMed1996

Stem cell factor and stromal cell co-culture prevent apoptosis in a subculture of the megakaryoblastic cell line, UT-7.

The megakaryoblastic cell line, UT-7, is dependent for its growth upon interleukin-3 (IL-3), erythropoietin, or granulocyte-macrophage colony stimulating factor (GM-CSF). A subculture of this line can be maintained in recombinant human c-kit ligand [stem cell factor (SCF)] at 100 ng/ml without requirement for other growth factors. Removal of this subculture from SCF results in rapid loss of viability and decreased proliferation. Cells grown in SCF also can be maintained in GM-CSF but not vice versa. In this work, we have characterized the SCF dependence of this UT-7 subculture. Stem cell factor removal results in apoptosis and a decline in viability which can be restored partially by re-addition of SCF, GM-CSF, or co-culture with adherent marrow stromal cells. Apoptosis in the factor-starved UT-7 population has been documented by light microscopy, electron microscopy and DNA analysis, showing the typical 180 base pair laddering characteristic of apoptosis. To quantitate the degree of apoptosis in the cell populations, and to assess whether apoptosis decreased with re-exposure of starved cells to growth factors or stroma, we utilized flow cytometry. This confirmed that exposure of previously factor-starved cells to stroma decreased the percentage of cells undergoing apoptosis. Co-culture with an SCF-deficient murine stromal cell line was also able to prevent apoptosis, suggesting contribution of other stromal cell factors. Experiments performed using trans-well inserts which do not allow cell passage, showed greatest viability of cells in contact with stroma, but viability was also improved in cells cultured in the presence of, but not in contact with, stromal cells compared to those cultured above plastic, suggesting a role for soluble stroma-produced substances. These data demonstrate that SCF alone can prevent apoptosis in cells dependent upon its presence for proliferation. Also, marrow stromal cells can serve as a partial substitute for growth factor in the prevention of apoptosis in these cells, probably due to constitutive presentation of SCF and other hematopoietic growth factors in both soluble and surface-bound forms.

Antibodies↗

PubMed1996

Flutamide-induced cyanosis refractory to methylene blue therapy.

Flutamide, an anti-androgen used in prostate cancer therapy, is also a derivative of aniline. Mild, usually asymptomatic, methaemoglobinaemia has been reported. We report a patient receiving flutamide therapy who developed cyanosis, dyspnoea and anaemia, initially attributed to marked methaemoglobinaemia by the CO-Oximeter method. An unsuccessful trial of methylene blue therapy led to the finding of marked sulphaemoglobinaemia. Sulphaemoglobinaemia has not previously been reported with flutamide use. Recognition of this association is important, given the refractoriness of sulphaemoglobinaemia to methylene blue therapy.

Aged↗