Immunophenotypic and functional recovery following stem-cell transplantation.
Explore the source record for details and available documents.
Antigens, Surface↗
Biomedical subjects
L S Lamb
Publications and source records attributed to L S Lamb.
In vitro interactions between gamma deltaT cells, DC, and CD4+ T cells; implications for the immunotherapy of leukemia.
BACKGROUND: Gamma deltaT cells contribute to immune defense against infectious organisms and some malignancies, but the process of activation and proliferation of these cells is not well understood. It is known that the immune response of gamma deltaT cells is not MHC-dependent, but is likely based on direct recognition of surface peptides and non-peptide ligands. This study examined whether DCs and CD4(+) T cells can participate in the activation of gamma deltaT cells. METHOD: Peripheral blood gamma deltaT cells were co-cultured with CD34-derived autologous DCs and CD4(+) T cells using contact-dependent cultures and transwell systems. Proliferation, immunophenotyping, and cytotoxicity assays determined the extent of gamma deltaT cell proliferation and cytotoxicity. RESULTS: Human gamma deltaT cells expanded 221.3 +/- 76-fold in cultures with DCs, and 165.7 +/- 76.6-fold with CD4(+) T-cells alone. Proliferation was enhanced (1949.8 +/- 261.3-fold) when gamma deltaT cells were cultured with both DC and CD4(+) T cells. Proliferation was contact-dependent, and resulted in the expansion of V delta1+ or V delta2+ cells cytotoxic against several leukemic cell-lines, but not against allogeneic PHA-induced lymphoid blasts. Ligation of the T-cell receptor with anti-pan-delta Ab significantly up-regulated cytotoxicity against K562, KBM-5 and KG1a, and normal BM, but not against Molt-4, allogeneic EBV-transfected B cells and allogeneic PHA-blasts. Minimal cytotoxic activity was shown against allogeneic marrow colony-forming units granulocyte-macrophage and erythrocyte colony-forming units. CONCLUSION: DCs can participate in the activation of gamma deltaT cells against specific autologous targets, and cytotoxicity can be enhanced by further stimulation via the T-cell receptor.
CD11c Antigen↗
Human gammadelta(+) T lymphocytes have in vitro graft vs leukemia activity in the absence of an allogeneic response.
Refractory acute lymphoblastic leukemia (ALL) is often incurable, and relapse rates following allogeneic bone marrow transplantation (BMT) remain high. We have reported that patients who develop increased numbers of gammadelta(+) T cells soon after BMT are significantly less likely to relapse. We now show in seven donor/recipient pairs that donor-derived Vdelta1(+)CD4(-)CD8(-)gammadelta(+) T cells are activated and proliferate in response to recipient primary ALL blasts. In addition, these cells have been shown to bind and lyse the recipient ALL blasts. Separately, gammadelta(+) T cells proliferate poorly or not at all in mixed lymphocyte culture against HLA-mismatched unrelated stimulator cells. These observations suggest that allogeneic gammadelta(+) T cells could be an effective immunotherapeutic strategy against refractory disease without the risk of graft-versus-host disease.
Cytotoxicity, Immunologic↗
Expression of CD134 (0X-40) on T cells during the first 100 days following allogeneic bone marrow transplantation as a marker for lymphocyte activation and therapy-resistant graft-versus-host disease.
CD134 (OX-40) is an activation-associated antigen which functions as a costimulatory receptor for CD4+ T cells. In order to determine the expression of CD134 during immune recovery following allogeneic bone marrow transplantation (BMT), we measured its expression on T cells and T cell subsets during the first 100 days following BMT in 26 patients. CD4+CD134+ T could be seen approximately 14 days following BMT cells in patients who did not develop GvHD which required therapy (n = 20). The percentage of CD4+CD134+ cells continued to increase up to the fourth week following BMT to a maximum of 40-50% of CD4+ T cells (normal = 1-8%). Two patients who developed Grade I-II GvHD and who responded to treatment with pulsed high-dose methylprednisolone (MPD) showed a decline of approximately 40% in CD4+CD134+ T cells was seen within 48 hours of treatment. Four patients who developed GvHD that was not responsive to MPD and who later developed high IV GvHD showed increasing CD4+CD134+ T cells up to 85% of the CD4+ T cells. Additionally, rapid increases in CD134+ T cells following antibody-based T cell therapy were associated with GvHD recurrence. In no cases was the percentage of CD134+ CD4+ T cells predictive of clinical GvHD. In this exploratory study, we have shown that CD134, although not predictive of the initial onset of GvHD, may be a useful tool for monitoring the response to early GvHD therapy and identification of patients at risk for reemergence of GvHD who may benefit from anti-T cell therapy. Cytometry (Comm. Clin. Cytometry) 38: 238-243, 1999.
Biomarkers↗
Cryopreserved normal macrophages as a control for assays of macrophage function.
Explore the source record for details and available documents.
Animals↗
Phenotypic and functional reconstitution of peripheral blood lymphocytes following T cell-depleted bone marrow transplantation from partially mismatched related donors.
Myeloablative chemotherapy followed by transplantation of a T cell-depleted bone marrow graft from a partially mismatched related donor provides a potentially curative option for patients with leukemia and other disorders of hematopoiesis, although the patient is faced with a period of sustained immunodeficiency as well as pharmacologic immunosuppression as a result of prophylaxis against graft-versus-host disease. Thirty patients who received one to three antigen T cell-depleted mismatched grafts were evaluated for immune reconstitution. The percentage and numbers of cells expressing lymphocyte subset antigens were determined by flow cytometry at 14, 28, 60, 100, 180, 270 and 365 days post-BMT and at 6 month intervals thereafter. Lymphocyte reconstitution was characterized by the early appearance of natural killer cells and a low percentage of both T and B cells. During the first year after BMT, the number of NK cells remained constant while T and B cells gradually returned to normal numbers and proportions. Response to the lymphocyte mitogen phytohemagglutinin returned to normal over the course of 2 years, while the response to concanavalin A was slightly depressed and the response to pokeweed mitogen became supranormal at about 1.5 years and continued to increase. These data suggest the need for long-term immunophenotypic monitoring as well as prolonged infection surveillance and prophylaxis.
B-Lymphocytes↗
Donor leukocyte infusion for treatment of graft rejection post partially mismatched related donor bone marrow transplant.
Graft rejection following bone marrow transplantation is more common in patients who receive their grafts from alternative donors and whose marrow is T cell depleted. Rejection in these patients is mediated by persistent host cells that interfere with successful establishment of donor-derived hematopoietic recovery. We describe a patient with chronic myelogenous leukemia in accelerated phase who rejected a T cell-depleted bone marrow graft, 2 months following partially mismatched related donor bone marrow transplant. Unmanipulated peripheral blood donor leukocyte infusion, without additional chemotherapy or immunosuppressive therapy resulted in complete hematopoietic recovery. Cytogenetics and RFLP demonstrated hematopoietic donor chimerism. The patient did not develop graft-versus-host disease.
Adult↗
De novo chronic graft-versus-host disease presenting as hemolytic anemia following partially mismatched related donor bone marrow transplant .
Chronic graft-versus-host disease (cGVHD) is a disease of immune dysregulation that resembles an autoimmune disease. It usually involves the skin, mucosal and serosal surfaces and, less commonly, the hematopoietic system. We report hemolytic anemia (HA) as the primary manifestation of de novo cGVHD in recipients of partially mismatched related donor transplants. Five of 40 eligible patients developed HA at a median of 168 days post-transplant. Recipients were mismatched for one to three major HLA antigens. Conditioning therapy consisted of total body irradiation, etoposide, Ara-C, cycle-phosphamide and steroids. GVHD prophylaxis included partial T cell depletion, using anti alpha/beta CD3 antibody (T10B9) and complement, in addition to post-transplant immunosuppression. At presentation, all patients were receiving cyclosporine with or without low-dose steroids. Along with a mean Hb of 7.1 g%, patients had an increased reticulocyte count, a mild raised lactic dehydrogenase and a positive Coombs' test (in 2/5 patients). Four patients had also demonstrated a decrease in platelet count. Treatment was initiated with high-dose steroids and intravenous gamma globulin and response was observed within 1 week. Awareness of this presentation of cGVHD and early therapeutic intervention can result in successful reversal of presumed immune-mediated red cell and platelet destruction.
Adrenal Cortex Hormones↗
Flow cytometric cell sorting combined with molecular chimerism analysis to detect minimal recurrent leukemia: good news and bad news.
Allogeneic BMT offers the possibility of cure for a variety of hematopoietic malignancies, but disease relapse remains a major cause of treatment failure. This report describes two cases in which flow cytometric cell sorting (FACS) and molecular chimerism analysis were combined to increase the sensitivity of minimal residual disease (MRD) detection. In the first case this approach was used to demonstrate that a suspicious phenotype was not recurrent leukemia, thus preventing the use of potentially toxic therapy. In the second case the recurrence of a leukemia which was undetectable by conventional analysis was confirmed. The potential benefits of combining these MRD detection methods are discussed.
Adolescent↗
Increased frequency of TCR gamma delta + T cells in disease-free survivors following T cell-depleted, partially mismatched, related donor bone marrow transplantation for leukemia.
Recent interest has focused on the function of gamma delta + T cells in immune responses. However, their role in allogeneic bone marrow transplantation (BMT) remains undefined. We report on a group of 43 leukemia patients who survived for at least 100 days following transplantation using partially HLA-mismatched grafts from related donors that were T cell depleted with the anti-TCR alpha beta monoclonal antibody T10B9.1A-31 and complement. Ten patients (23.2%) were found to have an increased (> or = 10%) proportion of gamma delta + T cells in the peripheral blood at 60-270 days after BMT. All of these patients remain alive, and 9 (90% of patients with > or = 10% gamma delta + cells) are free of disease at 2.5 years compared with a disease-free survival probability of 31% among patients with a normal proportion and concentration of gamma delta + T cells. No other factor was found to be independently associated with improved survival in these patients. These data suggest a possible association between an increase in the percentage and number of gamma delta + T cells and improved disease-free survival following transplantation from a partially mismatched related donor.
Bone Marrow Transplantation↗
Acute rejection of marrow grafts in patients transplanted from a partially mismatched related donor: clinical and immunologic characteristics.
Bone marrow transplantation (BMT) from a partially mismatched related donor (PMRD) provides a treatment option for patients lacking a matched sibling donor. T lymphocyte depletion of the graft reduces the risk of severe graft-versus-host disease, but may increase the risk of graft failure. We evaluated the pattern of acute graft rejection in eight patients receiving PMRD BMT with respect to the conditioning therapy, diagnosis, age and sex of donor and recipient, degree of HLA mismatch, and peripheral blood immunophenotype at the time of graft failure. All grafts were partially depleted of T lymphocytes. Marrow grafts infused into patients who experienced acute rejection did not differ significantly in nucleated cell dose, degree of T lymphocyte depletion, T cell dose, or CFU-GM/kg infused, from those received by 31 patients who showed durable engraftment. In three of four patients who rejected their grafts, and had sufficient peripheral blood cells for immunophenotyping, a CD3+CD8+ T lymphocyte phenotype was predominant at the time of acute rejection. In one patient rejection was associated with a predominant population of CD3+CD4+ T lymphocytes. Rejection was significantly associated with chronic myelogeneous leukemia and in patients mismatched by more than two antigens.
Adult↗
In vitro expansion and characterization of dendritic cells derived from human bone marrow CD34+ cells.
Dendritic cells (DC), as professional antigen-presenting cells, play a major role in stimulating naive T cell responses in vivo and in vitro, and may exacerbate or modulate T lymphocyte-mediated reactions, such as interactions between a hematopoietic graft and the recipient, eg GVHD and graft-versus-leukemia. Here, we describe a two-stage cell culture system for expansion of functionally active human DC from CD34+ marrow precursors. Optimal outgrowth was achieved by initially culturing CD34+ cells for 5 days in medium containing GM-CSF, MGF and TNF-alpha. Substitution of CD40L and IL-4 for TNF-alpha during a subsequent 5-day subculture increased DC content, such that by 10 days the cultures contained approximately 40% DC as determined by immunophenotype and morphology. An increase in DC purity to 84% at 10 days was achieved by immunomagnetic separation for CD1a+ cells from 5-day cultures and subculturing these cells in medium with IL-4 and CD40L. Reversing the sequence of growth factors during culture and subculture decreased the yield and purity of DC. Expression of CD80 and CD86 was enhanced by adding CD40L and IL-4, and the DC showed stimulatory activity in MLC. In conclusion, we have described a simple two-stage culture system to generate functional DC from CD34+ marrow precursors.
Antigens, CD34↗
Pneumococcal clearance function of the intact autotransplanted spleen.
OBJECTIVE: To determine if the venous drainage of the spleen into the portal circulation is essential for its ability to protect against encapsulated bacterial challenge. DESIGN AND INTERVENTIONS: Three groups of dogs were randomly assigned to undergo either sham laparotomy, splenectomy, or splenectomy with autotransplantation of the intact spleen into the pelvis and formation of vascular anastomoses to the iliac vessels. Two weeks post-operatively, the dogs received a sublethal intravenous injection of Streptococcus pneumoniae type 25. OUTCOME MEASURES: Bacterial clearances and inflammatory damage to the liver. Bacterial clearance of the autotransplanted spleen should be no different from that of a sham-operation spleen and significantly different from that of a splenectomized animal. In addition, immunologic function of the autotransplanted spleen should not differ from that of a sham-operation spleen in the degree of inflammatory damage to the liver. RESULTS: No differences in bacterial clearance function were found between the animals that had undergone sham laparotomy or splenic autotransplantation. However, bacterial clearance in the splenectomized animals was significantly impaired. Histologic examination of the liver 2 weeks after the bacterial challenge revealed high-grade inflammatory damage to the livers of splenectomized dogs, intermediate liver damage in dogs that underwent autotransplantation, and essentially no damage in dogs that underwent sham laparotomy. Autotransplanted spleens were essentially nonreactive, lacking actively proliferating germinal centers, whereas splenic tissue from sham-operation animals showed reactivity. CONCLUSION: Although bacterial clearance function is unchanged in autotransplanted spleens, this method still does not fully protect the liver from inflammatory damage.
Analysis of Variance↗
Morphologic and functional characteristics of alveolar macrophages following cryopreservation.
Cryopreservation of nucleated blood and bone marrow mononuclear cells has been used both to preserve such cells for clinical, diagnostic, or research use and to eliminate them as passengers in frozen tissue destined for transplantation. Although techniques for macrophage and monocyte cryopreservation have been described, little work has been done on the functional state of these cells following frozen storage. Using rabbit alveolar macrophages (AM), we have shown that AM can be frozen, stored, and recovered without morphologic changes. Furthermore, freshly isolated and cryopreserved AM do not differ in their adherence characteristics or the organization of their actin cytoskeleton. Cryopreserved normal AM also retained inducible functions such as superoxide anion (O2-) release and production of tumor necrosis factor (TNF) and interleukin-1 (IL-1). In order to further define the effects of cryopreservation on macrophage biology, we have also frozen macrophages which have been primed in vitro, and have demonstrated that the primed state remains at prefreezing levels after thawing and subsequent analysis. These data help define the effects of cryopreservation on cell function and establish parameters for the clinical and experimental evaluation of cryopreserved mononuclear phagocytes.
Actins↗
Responsibilities in point-of-care testing. An institutional perspective.
The development of "user-friendly" laboratory analyzers, combined with the need for rapid assessment of critical care patients, has led to the performance of in vitro diagnostic testing at the point of care. This strategy has been well received by most physicians who desire rapid turnaround times for laboratory tests, especially in the critical care areas. Since the primary care-giver in most critical care units is the registered nurse, much point-of-care testing has been delegated to nursing personnel. This has resulted in questions of authority, responsibility, regulation, and conflict resolution. These areas are discussed along with alternative strategies, such as vacuum transport, stat and/or satellite laboratories, unit-based phelebotomists, or "super techs," designed to achieve the goals of bedside or near-bedside testing in the critical care setting.
Centralized Hospital Services↗
Characterization of acute bone marrow graft rejection in T cell-depleted, partially mismatched related donor bone marrow transplantation.
The purpose of this study was to characterize the phenotype and clonality of the T cell population in patients who experience acute rejection (AR) following bone marrow transplantation (BMT) from a partially mismatched related donor (PMRD). Phenotypic analysis was performed using flow cytometry, assignment of donor/host lineage by cytogenetics or HLA-specific flow cytometry, and analysis of the T cell receptor (TCR) by reverse-transcriptase polymerase chain reaction (RT-PCR). We have previously reported the initial appearance in the blood of AR patients of host CD8+brightCD3low T cells that progressively express increasing amounts of CD3+ cells. We now report that this cell population can differentiate into either a cytotoxic T cell phenotype (CD3+CD8+HLA-DR+CD57-) usually associated with AR of grafts from matched unrelated donors or a suppressor T cell phenotype (CD3+CD8+CD57+HLA-DR-) usually associated with AR of grafts from matched sibling donors. Analysis of the TCR V beta subsets from two patients revealed sorted host CD3+CD8+ cells (purity 90-95%) from the first patient to express V beta 18 almost exclusively. In a second patient with late rejection (55 days post-BMT), the CD3+CD8+ cells were predominantly restricted to V beta 1, 5.1, 7, 9, and 18. Although CD3+CD8+ T cells are known to be associated with AR, cytotoxic and suppressor lineages in AR from the same type of BMT and clonal distribution of T cells in AR have not been reported. Preliminary results suggest that V beta expression in AR of PMRD grafts is restricted and host T cell phenotype may vary. Further studies will investigate whether specific mismatches correlate with specific V beta usage and/or host T cell phenotype.
Base Sequence↗
Current nursing practice of point-of-care laboratory diagnostic testing in critical care units.
BACKGROUND: The development of user-friendly laboratory analyzers, combined with the need for rapid assessment of critically ill patients, has led to the performance of in vitro diagnostic testing at the point of care by personnel without formal laboratory training. OBJECTIVES: To determine the range of laboratory testing performed by critical care nurses and their attitudes toward this role. METHODS: A survey of critical care nursing consultants was conducted, using a modified Likert scale, to assess objective measures of point-of-care testing practice in critical care units and to determine nurses' attitudes toward the practice of point-of-care testing. Statistical analysis was performed to determine significant trends in responses. RESULTS: Of the units responding to the survey, 35% used critical care nurses exclusively to perform point-of-care testing, 32.5% used laboratory technicians and critical care nurses, and 25% used other personnel. Of critical care nurses performing laboratory testing, 95.5% performed blood glucose analysis; 18.7%, arterial blood gas analysis; 4.5%, electrolyte analysis; 4.5%, hematology profiles; and 22.7%, other testing. Most agreed that stat tests were not reported promptly, thereby necessitating bedside testing. Respondents indicated that they would prefer that laboratory personnel operate in vitro diagnostic equipment and that requirements for critical care nurses to perform laboratory testing detracted from other patient care duties. CONCLUSIONS: Most nurses who perform point-of-care testing responded that it was necessary and helpful in patient management. However, they would prefer, because of their other patient care responsibilities, that laboratory personnel take this responsibility.
Attitude of Health Personnel↗
Effect of dietary copper on colonic tumor production and aortic integrity in the rat.
Gastrointestinal malignancy has been associated with aortic aneurysmal disease in humans, while metabolic derangement of copper has been incriminated as a possible promotor of aneurysmal development of the aorta. An animal model utilizing the carcinogen 1,2-dimethylhydrazine (DMH) was selected to evaluate levels of dietary copper on both colonic tumor production and morphologic changes in the rat aorta. Six groups, each including 10 Sprague-Dawley rats, received 16 weekly doses (20 mg/kg) of DMH beginning at 4 weeks of age. Groups were maintained on either normal (25 ppm), low (0.6 ppm), or high (100 ppm) copper chow during the entire experimental period. After 25 weeks, all animals were sacrificed to assess colonic tumor production and to perform scanning (SEM) and transmission electron microscopic (TEM) studies of the rat aorta. Results showed a significant increase in colonic tumor production (3.14 +/- 0.39 tumors per centimeter colon) in rats treated with low-copper chow and DMH when compared with rats on normal chow and DMH (0.74 +/- 0.07 tumors per centimeter colon) and animals maintained on high-copper diets and DMH (0.76 +/- 0.08 tumors per centimeter colon). In addition, morphologic study showed disruption of the intima and media in rats maintained on low-copper diet alone, and also on low-copper diet plus DMH. The results of this study showed that DMH and low dietary copper significantly increase (P less than 0.001) the yield of colonic tumors and produce loss of aortic integrity when studied morphologically. Copper levels may be important in the association of neoplasia and aneurysm formation in the clinical setting.
1,2-Dimethylhydrazine↗