Biomedical subjects
R A Preti
Publications and source records attributed to R A Preti.
Effect of interface/offset (I/O) adjustment on collection efficiency using the Fenwal CS3000 Plus Blood Cell Separator for peripheral blood progenitor cell collection.
Peripheral blood progenitor cells (PBPC) reside within the mononuclear cell (MNC) component of the blood and can be collected using a number of apheresis devices, including the Fenwal CS3000 Plus Blood Cell Separator. Increased MNC collection efficiency, therefore, may reduce the number of apheresis required to achieve collection goals. In this study, patients were divided into groups by absolute MNC count to determine the effect of interface detector offset (I/O) adjustment on MNC collection efficiency. Apheresis products from 104 procedures collected using a standard I/O setting of 100 were compared with 121 collections for which the I/O setting was adjusted according to the preapheresis MNC count. Adjustment of the I/O setting in this manner had no statistically significant impact on the per kilogram dose of MNC collected. The data did show that MNC collection efficiency was reduced as both the MNC count and I/O setting increased, as the collection efficiency was greatest for patients with the lowest peripheral MNC counts and was inversely correlated with the preapheresis MNC count. Although contamination of the product with platelets was drastically reduced at higher I/O settings, there was a concomitant rise in RBC contamination. We conclude that a standard setting of 100 is preferable to adjustment of the I/O setting as a function of the preapheresis MNC count.
CD34+CD33- cells influence days to engraftment and transfusion requirements in autologous blood stem-cell recipients.
PURPOSE: To evaluate the reliability of CD34/CD33 subset enumeration as a predictor of hematopoietic repopulating potential in autologous blood stem-cell transplantation and to determine which patient and treatment-related factors affect the timing, quantity, and type of blood stem cells mobilized. PATIENTS AND METHODS: We analyzed blood stem-cell collections from 410 consecutive cancer patients who received mobilization therapy and evaluated factors, including CD34+ subset quantities, that might influence engraftment kinetics and transfusion requirements in autologous blood stem-cell recipients. RESULTS: The majority of patients (97%) mobilized CD34+33- cells, which were usually collected in the greatest quantity on the first day of apheresis. Patients who received only growth factor mobilized the highest percentage of CD34+33- cells. Extensive prior chemotherapy limited the collection of CD34+33- cells. In addition to patient diagnosis (P < .006) and total CD34+ cell dose (P = .0001), CD34+33- cell dose (P < .005) and percentage of CD34+33- cells (P < .005) were identified as independent factors significantly predictive of engraftment kinetics. CD34+33- cell dose (R2 < or = .177; P < .0001) was a strong and the only significant predictor of RBC and platelet transfusion requirements. Furthermore, independent of the total CD34+ cell dose, as the CD34+33- cell dose increased, days to neutrophil recovery, days to platelet recovery, and transfusion requirements decreased. CONCLUSION: These findings show that CD34+33- cells are readily collected in most cancer patients and significantly influence engraftment kinetics and transfusion requirements in autologous blood stem-cell recipients. CD34+33- cell quantity of the blood stem-cell graft appears to be a more reliable predictor of hematopoietic recovery rates than total CD34+ cell quantity in this setting.
Single and double autotransplants for relapsing/refractory Hodgkin's disease: results of two consecutive trials.
To evaluate a strategy of one cycle of dose-intensive chemotherapy for patients with Hodgkin's disease in sensitive relapse and two cycles for those with refractory disease, 122 patients received dose-intensive chemotherapy followed by autotransplant in two consecutive studies. Patients with refractory disease were offered a second transplant with different conditioning in the absence of progression or excessive toxicity. CR was present after treatment in 46% while 16% died in the peritransplant period. Of 41 patients with primary refractory disease and 42 with refractory relapse, 24 and 21 respectively received a second cycle. Of these 45 refractory patients, 12 were in CR and 11 in PR after the first cycle and 10 of these 11 in PR achieved a durable CR with the second transplant. The CR rate is 37% in patients with refractory relapse and 19% in those with primary refractory disease. At a median follow-up of 4 years, median survival is 45 months. Progression-free survival of the refractory patients who could receive a second cycle was similar to that of patients with sensitive disease. A sequential transplant strategy is feasible. A subgroup of patients with refractory disease can achieve long-term survival after sequential BMT.
An intralaboratory quality control program for quantitation of CD34+ cells by flow cytometry.
This study was undertaken to develop a quality control protocol to monitor instrument, operator, and CD34 assay performance. A dual level control system was established by cryopreserving aliquots of cells from peripheral blood progenitor cell (PBPC) collections exhibiting different percentages of CD34+ cells. Twenty-five samples from each control specimen were assayed to establish a control range (mean +/- 2 SD). Levey-Jennings graphs were prepared for each control specimen to plot multiple measurements of CD34%. No significant differences were observed between fresh or cryopreserved PBPC aliquots in terms of light scatter properties or CD34 antigen density within the gated cell population. Cryopreserved PBPC samples are ideal for serving as a positive methodology control for daily CD34 analysis. Furthermore, such a system can help identify problems with assay reagents, sample preparation technique, or incorrect data analysis.
Optimization of CD34+ cell selection using immunomagnetic beads: implications for use in cryopreserved peripheral blood stem cell collections.
Isolation of CD34+ cells from bone marrow, umbilical cord blood, and mobilized peripheral blood stem cell (PBSC) collections has many potential clinical benefits. The aim of this study was to evaluate the use of the ISOLEX 300 system to select hematopoietic precursors and determine the effectiveness at depleting contaminating tumor cells from cryopreserved/thawed PBSC. Median recovery of CD34+ cells and CFU-GM colonies was 71% and 51.5%, respectively, using a protocol optimized for our laboratory. A mean 2.9 log10 decrease in contaminating breast carcinoma cells was seen after the selection process. Selected CD34+ cells underwent a second round of cryopreservation/thawing while retaining 85.6% viability and 72.3% recovery of CFU-GM colonies.
New York State guidelines for cord blood banking. Department of Health.
In response to the profound interest in the transplantation of umbilical cord blood, the New York State (NYS) Department of Health embarked on a regulatory strategy designed to provide guidance to the practitioner while allowing for the flexibility required by the inherent developmental nature of the technology. The result was the development of the Guidelines for Collection, Processing, and Storage of Cord Blood Stem Cells, adherence to which is optional. It is anticipated that through the use of such guidelines as a supplement to the existing hematopoietic progenitor cell (HPC) regulations for tissue banks, practitioners may provide to potential patients the safest and most efficacious cord blood products currently available, and information gained through the use of these products will be of optimal value toward the goal of continuous improvement in the delivery of this new technology.
The effect of UVB irradiation on proliferative activity and cell growth potential of cord blood.
To determine the degree of graft versus host disease (GVHD) prophylaxis that might be necessary if cord blood (CB) transplantation is more widely applied, we compared human cord blood mononuclear cell (CBMC) and adult peripheral blood mononuclear cell (PBMC) proliferative responses and stimulatory capabilities; to examine the utility of UVB irradiation for GVHD prophylaxis, we compared proliferative responses, antigen-presenting cell (APC) stimulatory functions, and cytokine production by untreated and UVB-irradiated CBMCs. The two cell types, CBMC and PBMC, proliferated equally both in response to phytohemagglutinin (PHA) and alloantigen in mixed lymphocyte culture (MLC). Cord blood stimulatory function in MLC was significantly (P < 0.05) reduced to 60% of PBMC stimulatory capability. Ultraviolet-B irradiation at a dose of 100 J/m2 of CBMCs significantly (P < 0.01) inhibited PHA stimulation by 79.4%, reduced responder activity in MLC by 75.8%, and inhibited stimulatory activity in MLC by 55.6% as compared with the activity shown by untreated CBMCs. The same dose of UVB preserved 59.9% of CFU-GM and 65.9% of BFU-E colony growth as compared with untreated CBMCs. Production of lymphokines (IL-2, GM-CSF, LIF, and gamma-IFN) by PHA-stimulated CBMCs was decreased, but monokine (IL-1 beta and IL-6) production was unchanged. We conclude that UVB irradiation at a dose of 100 J/m2 inhibits CB lymphocyte activation and preserves the cellular growth potential of CB hematopoietic progenitor cells.
The combined use of soybean agglutinin and immunomagnetic beads for T lymphocyte subset depletion of bone marrow allografts: a laboratory analysis.
The prevention of graft-versus-host disease by T-lymphocyte depletion of allografts prior to bone marrow transplantation has resulted in an increase in graft failure/rejection and relapse of disease. Evidence for the selective roles of specific T-lymphocyte subsets in each of the engraftment, graft-versus-host disease, and disease relapse processes has been presented, but sufficient clinical verification to support any hypothesis in this regard is lacking. In this paper we describe a convenient and highly flexible clinical laboratory method for depletion of specific T-lymphocytes in controllable quantities by the use of select monoclonal antibodies. Almost 3 log10 removal of CD2+ and CD8+ cells without significant loss of hematopoietic progenitors (CFU-GM, BFU-E, and CD34+ cells) can be reproducibly achieved. This method, employing soybean agglutination and immunomagnetic beads, is potentially adaptable to depletion of any cell subset or any tumor cell for which unique cell-surface antigen characteristics have been defined. In addition, our protocol is equally suited to the positive selection of stem cells and hematopoietic progenitors bearing the CD34 surface antigen.
A two-phase approach to B lymphocyte purging of autologous bone marrow grafts for patients with malignant lymphoma contaminated bone marrow.
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Selection and purification of CD34+ cells using monoclonal antibody and ferrofluids.
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Peripheral blood mononuclear mobilization with sargramostim (GM-CSF).
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Computerization of the bone marrow processing laboratory: current progress and future directions.
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Umbilical cord blood infusion in a patient for correction of Wiskott-Aldrich syndrome.
A 2 3/4 year old male with thrombocytopenia secondary to Wiskott-Aldrich Syndrome (WAS) and a history of two intracranial hemorrhages as well as hemolytic anemia and neutropenia received a placental blood infusion from an HLA-identical female sibling born by caesarian section at 35 weeks gestation. The patient was prepared with Thiotepa and Cytoxan and received a nucleated cell dose of 3.0 x 10(7)/kg. Cyclosporin A and Methylprednisolone was given for graft versus host disease (GVHD) prophylaxis. An ANC of 0.5 x 10(9)/L and 1.0 x 10(9)/L were achieved on post-transplant days 18 and 28, respectively. Platelet recovery was rapid with a platelet count > or = 100 x 10(9)/L on day +39. On posttransplant day +11, the patient developed an erythematous rash consistent with grade I acute GVHD that resolved without therapy. He was discharged day on +60 and has remained free of infections with a normal platelet count off all immunosuppression therapy 30+ months post-transplantation. Chimerism studies performed on peripheral blood mononuclear cells by fluorescent in situ hybridization indicated that the percentage of donor cells ranged between 55 and 80%. The phenotype and function of peripheral blood lymphocytes are completely normal and the patient has responded in vivo with production of antibodies to both diphtheria and tetanus immunizations. This study demonstrates the feasibility of collecting placental blood after a multiple birth delivery and the ability of umbilical cord blood to provide complete hematopoietic and immunologic reconstitution in a patient with WAS.
Clinical and laboratory comparison study of refrigerated and cryopreserved bone marrow for transplantation.
Refrigerated storage for short-term preservation of bone marrow is an alternative to cryopreservation where chemotherapeutic regimens include drugs with short in vivo half-lives. We performed a clinical and laboratory comparison of bone marrow stored at 4 degrees C for up to 9 days to bone marrow cryopreserved at -90 degrees C for autotransplantation. After adjusting for the confounding effects of disease type or sex, no clinically meaningful variation in post-transplant course between refrigerated storage and cryopreserved was found. Therefore, the data presented in this study suggest that the clinical recovery indices following transplantation between the two storage groups are essentially equivalent. One potential advantage to refrigerated storage, however, is that it may provide an opportunity for extended exposure to growth factors and/or purging agents in vitro prior to transplantation. To prepare for an in vitro analysis of this hypothesis, we concentrated the stem cell population and compared the nucleated cell recovery, viability and colony forming potential following refrigerated storage of whole bone marrow and buffy coat to cryopreserved bone marrow stored for the same interval. While the nucleated cell recovery for cryopreserved marrow was significantly greater than for refrigerated storage, the viability and colony forming potential of the refrigerated storage was superior or equivalent, independent of prior processing.
ISHAGE bone marrow processing survey: report on an international information gathering process. International Society for Hematotherapy Graft Engineering.
Significant amounts of information are currently available within the database generated by the responses to the Bone Marrow Processing Survey. As additional Surveys are returned, the data will continue to be entered into a Lotus spreadsheet, until a more sophisticated database with a programmed interface becomes available. The readership is encouraged to enter or edit the databank by returning a completed or amended Survey to the Society. Copies of the form are available from the Society or can be found in the first issue of the Journal of Hematotherapy.
Hemopoietic stem cell processing: comparison of progenitor cell recovery using the Cobe 2991 cell washer and the Haemonetics V50 apheresis system.
Using 24 bone marrow (BM) harvests intended for cryopreservation and transplantation, we compared the use of the Cobe 2991 cell washer (2991) and the Haemonetics V50 apheresis system (HV50) for automated BM processing. Our in vitro data indicate that while the mononuclear cell (MNC) concentration of the HV50 product was significantly greater than that of the 2991, the overall MNC recovery of the two products was equivalent. In addition, although the concentration of CFU-GM and BFU-E in the products was equivalent, recovery of these progenitors in the 2991 product was significantly greater than those of the HV50 product. There was no significant difference in either the final product concentration or the overall recovery of cells bearing the primitive myeloid antigens, CD33 or CD34, between the two methods. The HV50 product volume, the red cell and the granulocyte mass were significantly lower than those of the 2991. We conclude that the advantages gained through the use of each machine should be evaluated within the context of the specific intention for the graft. Future advances in the identification and understanding of the primitive stem cell will aid in attempts to evaluate the methods used to isolate these cells.
Hematopoiesis on nylon mesh templates. I. Long-term culture of rat bone marrow cells.
Rodent hematopoietic cells have been perpetuated in long-term bone marrow culture (LTBMC), provided that they were plated onto a pre-established layer of bone marrow stromal cells (fibroblasts, reticular cells, adipocytes, macrophages, etc.). This monolayer-type system supports the self-renewal of murine pluripotent stem cells and produces substantial numbers of progenitors and mature cells of the myeloid lineage. In an effort to increase the growth potential of cells in LTBMC, an adherent matrix of stromal cells was established on a pretreated nylon screen template. Subsequent seeding of hematopoietic cells onto this matrix occurred both on the surface of the adherent layer and in the interstices formed by the developing stromal cells. A three dimensional growth pattern of hematopoietic colonies and clusters was observed. Mature cells and late stage precursors of the myeloid and erythroid series were observed in the non-adherent layer for the duration of the experiment (39 weeks). Cells similar in appearance to small lymphocytes were also seen in both the non-adherent and adherent layers. Cells associated with the adherent layer of this LTBMC system displayed the ability to reconstitute hematopoiesis in the irradiated host. It is suggested that because of its three-dimensional nature the nylon mesh LTBMC system possesses a greater hematologic potential per unit area than monolayer-type LTBMC.