Quantification of nucleated red blood cells in allogeneic marrow graft and impact of processing on recovery

BACKGROUND: In allogeneic bone marrow transplantation (BMT), a higher nucleated and CD34+ cell dose has been reported to improve various outcomes. Other cell types, such as lymphocyte subsets, also influenced BMT results. While nucleated red blood cells (NRBCs) represent a subset of bone marrow (BM) cell subpopulation, the question of their quantification in BM grafts and the impact of BM processing on their recovery has not been addressed. STUDY DESIGN AND METHODS: In a prospective study on 77 BM products, NRBCs were enumerated by flow cytometry and the recovery analyzed after manipulation. Because NRBCs could compromise white blood cell count, the impact of NRBC count on CD34+ cell percentage and total nucleated cell (TNC) dose were also determined. RESULTS: The mean percentage of NRBCs in BM grafts was 21.6 percent (range, 7.8%-40.9%). Mean NRBC recoveries after BM concentration or RBC depletion were 98.4 and 28.7 percent, respectively, close to those obtained for TNC cells (88.6 and 31.3%, respectively). When corrected with NRBC count, the mean percentages of corrected CD34+ cell and TNC dose were significantly modified when compared with uncorrected values, whatever the type of BM manipulation. CONCLUSION: Our data show that NRBC quantification might be of importance to improve quality control of BM products and to evaluate the influence of NRBCs cell dose on outcomes after BMT.     

High in-hospital mortality of intensive care patients with nucleated red blood cells in blood.

The detection of nucleated red blood cells (NRBCs) in blood of patients suffering from a variety of severe diseases is known to be highly associated with increased mortality. Blood analyzers to routinely measure NRBC concentrations are now available. However, the diagnostic and prognostic significance of this parameter for intensive care patients has not been evaluated. Using a Sysmex XE-2100 analyzer, NRBC concentrations were determined in blood samples from 421 patients treated in intensive care units (general and accident surgery, cardiothoracic surgery, and internal medicine) of a university hospital. NRBCs were found at least once in 19.2% of all patients. The mortality of NRBC-positive patients (n=81) was 42.0% (n=34); this was significantly higher (p<0.001) than the mortality of NRBC-negative patients (5.9%, n=340). The NRBC concentration was 115+/-4x10(6)/l (median 40x10(6)/l; range 20-2930x10(6)/l) at initial detection of NRBCs in the blood. Mortality increased with increasing NRBC concentration and increasing frequency of occurrence. With regard to in-hospital mortality, NRBCs in blood showed sensitivity and specificity of 63.0% and 87.2%, respectively. The detection of NRBCs is highly predictive of death, the odds ratio after adjustment for other laboratory prognostic indicators being 1.01 (p<0.01) for each increase in the NRBC concentration of +1x10(6)/l. NRBCs were detected for the first time, on average, 13 days (median 8 days) before death. The routine analysis of NRBCs in blood is of high prognostic power with regard to in-hospital mortality of critically ill patients. Therefore, this parameter may serve as an early indicator for patients at increased mortality risk.     

Cesarean section due to fetal distress increases the number of stem cells in umbilical cord blood

BACKGROUND: Umbilical cord blood (UCB) can be used as hematopoietic stem cell source for transplantation. The success of a transplantation is highly correlated with the number of total nucleated cells (TNCs) and CD34+ cells in the UCB. Certain obstetric factors increase the yield of stem cells in the UCB. It is necessary to evaluate optimal conditions in labor to decrease the rate of sample rejection due to low cell count. No data exist regarding the difference between primary and secondary Cesarean sections in terms of efficacy of stem cell harvesting. STUDY DESIGN AND METHODS: Seventy-nine consecutive UCB units from women who had a Cesarean section between 1997 and 2003 were included. The number of TNCs, CD34+ cells, colony-forming units (CFUs), white blood cells (WBCs), nucleated red blood cells (NRBCs), and the total collection volume were compared between cases with primary and secondary Cesarean section. RESULTS: UCB obtained after a Cesarean section due to fetal distress has significantly higher numbers of TNCs, CD34+ cells, NRBCs, and WBCs compared to elective Cesarean section. Of the cases with secondary Cesarean section due to fetal distress, 67 percent resulted in UCB units with sufficient TNC numbers (> or =80 x 10(7) TNCs) compared to 42 percent of the cases with primary Cesarean section. CONCLUSION: Fetal distress increases the number of hematopoietic stem cells mobilized into UCB. Particular effort should be made to collect UCB from newborns who experienced fetal distress.     

Increased numbers of total nucleated and CD34+ cells in blood group O cord blood: an analysis of neonatal innate factors in the Korean population

BACKGROUND: We analyzed neonatal factors that could affect hematopoietic variables of cord blood (CB) donated from Korean neonates. STUDY DESIGN AND METHODS: The numbers of total nucleated cells (TNCs), CD34+ cells, and CD34+ cells/TNCs of CB in neonates were compared according to sex, gestational age, birth weight, birth weight centile for gestational age, and ABO blood group. RESULTS: With 11,098 CB units analyzed, blood group O CB showed an increased number of TNCs, CD34+ cells, and CD34+ cells/TNCs compared with other blood groups. Although TNC counts were lower in males, no difference in the number of CD34+ cells was demonstrated because the number of CD34+ cells/TNCs was higher in males. An increase in the gestational age resulted in an increase in the number of TNCs and decreases in the number of CD34+ cells and CD34+ cells/TNCs. The numbers of TNCs, CD34+ cells, and CD34+ cells/TNCs increased according to increased birth weight centile as well as birth weight. CONCLUSION: CB with blood group O has unique hematologic variables in this large‐scale analysis of Korean neonates, although the impact on the storage policies of CB banks or the clinical outcome of transplantation remains to be determined.     

Increased Nucleated RBCs in Cord Blood: Not an Exclusion Criterion but a Quality Indicator for Hematopoietic Progenitor Cell Transplantation

Increased nucleated red blood cell (NRBC) counts have been reported to be associated with adverse fetal outcomes, and cord blood units (CBUs) with increased NRBC counts require a 2^nd questionnaire to determine their suitability for transplantation. However, a recent study demonstrated a positive correlation of NRBCs with CD34⁺ cells and total nucleated cells (TNCs). We evaluated the association between the NRBC count and hematopoietic progenitor cell (HPC) content (TNC and CD34⁺ cell counts) in Korean full-term newborn CBUs. In addition, we assessed whether an increased NRBC count is associated with newborn health problems that impair CBU safety. Among the 32,876 units processed from May 2006 to December 2018, a total of 23,385 CBUs with a TNC count ≥ 7 × 10⁸ and reliable perinatal information were analyzed to assess the association of the NRBC count with CBU parameters, and the newborns associated with 457 CBUs that required the 2^nd questionnaire due to an increased NRBC (≥ 15 NRBCs/100 WBCs) were assessed at one year for health problems that threatened CBU safety. The majority of the CBUs that required the 2^nd questionnaire due to an increased NRBC count (96.9%) were determined to be suitable for transplantation. Those with an increased NRBC count showed significantly higher CD34⁺ cell and TNC counts and a higher rate of transplantation (P < 0.001, < 0.001 and 0.025, respectively). NRBCs showed a significant positive correlation with TNCs and CD34⁺ cells and a significant negative correlation with birth weight (all P < 0.001; adjusted r = 0.185, 0.369 and - 0.029, respectively). In the multiple linear regression analysis, NRBCs showed independent and positive correlations with TNCs and CD34⁺ cells after adjustments for birth weight and gestational age (all P < 0.001; β = 0.182, adjusted R² = 0.053 and β = 0.367, adjusted R² = 0.418). An increased NRBC count in full-term normal delivery is a surrogate marker of HPCs in CBUs rather than an exclusion criterion for CBU safety. Moreover, providing the NRBC count together with the NRBC-corrected TNC count will be useful for clinicians to select CBUs for transplantation.     

Daily monitoring of nucleated red blood cells in the blood of surgical intensive care patients

BACKGROUND: In intensive care patients the appearance of nucleated red blood cells (NRBC) in blood is associated with a variety of severe diseases. Generally, when NRBCs are detected in the patients' blood the prognosis is poor. MATERIAL AND METHODS: In the present study the detection of NRBCs was used for a day-to-day monitoring of 284 surgical intensive care patients. RESULTS: NRBCs were found at least once in 32.0% of all patients. The mortality of NRBC-positive patients was 44.0% (40/91); this was significantly higher (P<0.001) than the mortality of NRBC-negative patients (4.2%, 8/193). With regard to intensive care mortality, NRBCs in blood showed sensitivity and specificity of 83.3% and 78.9%, respectively. The area under curve (C-statistic) was 0.86. Mortality increased with increasing NRBC concentration. All patients with more than 2000 NRBCs/microl died. Moreover, mortality increased with increasing frequency of occurrence. When after first detection of NRBCs in blood, during the further course of intensive care treatment the NRBCs have disappeared from the circulation, the mortality again decreased to values of NRBC-negative patients. The detection of NRBCs is associated with an increased mortality rate, the odds ratio after adjustment for other laboratory prognostic indicators being 1.8 (P<0.001) for each increase in the NRBC category (1-40/microl; 41-80/microl; 81-240/microl, >240/microl). NRBCs were detected for the first time, on average, 9 days (median 5 days) before death. CONCLUSIONS: The routine analysis of NRBC in blood is of potential prognostic power with regard to mortality of critically ill patients. Therefore, this parameter could possibly serve as a daily indicator of patients at high mortality risk.     

Predictive value of circulating immature cell counts in peripheral blood for timing of peripheral blood progenitor cell collection after G-CSF plus chemotherapy-induced mobilization

BACKGROUND: Enumeration of CD34+ cells in peripheral blood (PB) before apheresis predicts the number of CD34+ cells collected, although flow cytometric techniques used are complex and expensive. In an attempt to determine the optimal timing for peripheral blood progenitor cell (PBPC) collection, the usefulness of circulating immature cell (CIC) counts in PB was evaluated. STUDY DESIGN AND METHODS: CIC counts in PB and CD34+ cell counts in the apheresis product from 249 collections were assessed, and the relationship between these two parameters was evaluated by with the Pearson rank correlation analysis, the Fisher exact test, and the U-test. RESULTS: CIC counts were correlated significantly with the number of CD34+ cells per kg of patient's body weight in the apheresis product (Pearson rank correlation analysis: r = 0.635, p < 0.0001). When a level of 1 x 10(9) CICs per L was selected as a cutoff value, the sensitivity and specificity for collecting more than 1 x 10(6) CD34+ cells per kg of body weight were 75.7 and 85.5 percent, respectively. CONCLUSION: The present study strongly suggests that the number of CICs in PB may estimate the number of CD34+ cells collected. The data indicate that CIC counts above 1 x 10(9) per L can be used as a good predictor for PBPC collections containing more than 1 x 10(6) CD34+ cells per kg of body weight in a single apheresis procedure.     

Preterm birth and the availability of cord blood for HPC transplantation

BACKGROUND: Cord blood from deliveries at term can be used for HPC transplantation. The objective of this study was to determine the amounts of cord blood nucleated cells (NCs) and HPCs that were collectable from preterm deliveries. STUDY DESIGN AND METHODS: Cord blood collected from preterm deliveries between 22 and 36 weeks of gestation was compared with regard to volume, NC count (/mL), CD34+ cell count (/mL), and the NC and CD34+ cell counts per cord blood sample and at different gestational ages. RESULTS: A correlation was found between gestational age and NC count (r = 0.52, p<0.001), and an inverse relation was found between gestational age and CD34+ cell count (r = - 0.68, p<0.001). The CD34+ cell count per cord blood sample was independent of gestational age (r = - 0.13, p = NS), and no significant difference between early (22-32 week) and late (33-36 week) preterm deliveries was found (p = 0.870). Comparison with published data from cord blood transplantations revealed that up to one-third of preterm samples contained at least as many NCs (or CD34+ cells) as the median cell dose transplanted (calculated for the median recipient weight) in the respective study. Furthermore, 77 percent of all preterm samples contained at least 1 x 10(7) NCs (and 42% at least 1 x 10(5) CD34+ cells) per kg for transplantation in a recipient of 20-kg body weight, which corresponds to the lower threshold of cells per kg in the graft recommended by Eurocord. CONCLUSION: Preterm delivery should not be a reason to exclude cord blood collection if allogeneic cord blood transplantation in a sibling is planned.     

Ten-year quality control of a semiautomated procedure of cord blood unit volume reduction

BACKGROUND: Volume reduction of cord blood units decreases the cost of cryogenic storage. This study reports the analysis of a 10-year quality control program of a semiautomated cord blood volume reduction procedure.
STUDY DESIGN AND METHODS: Cord blood was collected in a plastic bag containing 29 mL citrate-phosphate-dextrose, centrifuged at 2124 ×  g for 12 minutes, and processed with a semiautomated device. The procedure was aimed at removing most red blood cells and plasma and concentrating hematopoietic progenitors in the buffy coat (BC), thus reducing the unit volume and saving cryogenic space. Finally, the BC was cryopreserved with an equal volume of 20 percent dimethyl sulfoxide. Total nucleated cells (TNCs) were counted before and after processing in the 4311 units banked from 1998 through 2007, whereas CD34+ cells and colony-forming units–granulocyte-macrophage (CFU-GM) were counted in 420 random units from 2001 through 2007.
RESULTS: Mean postvolume reduction annual recoveries of TNCs, CD34+ cells, and CFU-GM ranged from 82.8 ± 12.3 (standard deviation) to 91.4 ± 6.4 percent, from 87.8 ± 14.1 to 95.2 ± 23.8 percent, and from 101.5 ± 51.4 to 117.8 ± 59.5 percent, respectively. Very strong correlations were found (r > 0.87) between postprocessing versus preprocessing TNCs, CD34+ cells, and CFU-GM; a moderate correlation between initial TNC count and unit's volume (r = 0.51); and no correlation between TNC percentage of recovery in the BC and initial unit's volume. The latter data indicate that most TNCs concentrate in the BC.
CONCLUSIONS: The semiautomated procedure of cord blood unit volume reduction used in this study provides high and stable cellular recoveries during several years of routine cord blood banking.     

Poor prognosis indicated by nucleated red blood cells in peripheral blood is not associated with organ failure of the liver or kidney.

BACKGROUND: The appearance of nucleated red blood cells (NRBCs) in peripheral blood is associated with a variety of severe diseases. When NRBCs are detected in blood, this is generally associated with increased mortality. METHODS: In a prospective study, NRBCs and other laboratory parameters were measured daily in the peripheral blood of surgical intensive care patients. The appearance of NRBCs was analyzed in relation to laboratory indicators of organ injury. RESULTS: A total of 284 surgical intensive care patients were included in this study. The mortality of NRBC-positive patients was 44.0% (40/91). This was significantly higher (p<0.001) than the mortality of NRBC-negative patients (4.2%, 8/193). Mortality increased with the NRBC concentration and the length of the NRBC-positive period. Multiple logistic regression analysis of several other clinical and laboratory risk indicators revealed a significant association between NRBCs and increased mortality, with an odds ratio of 1.95 (95% CI 1.35-2.82; p<0.001) for each increment in NRBC category (0, 1-40, 41-80, 81-240 and >240 NRBC/microL). After the initial detection of NRBCs in blood, there were no significant increases in creatinine concentrations or alanine aminotransferase activity. However, the appearance of NRBCs coincided with increasing C-reactive protein and thrombocyte concentrations. CONCLUSIONS: The detection of NRBCs in blood of surgical intensive care patients is of prognostic power with regard to patient mortality. This prognostic significance of NRBCs was independent of some clinical and other laboratory risk parameters. The appearance of NRBCs in blood was not associated with kidney failure or lesion of the liver.     

The predictive value of white cell or CD34+ cell count in the peripheral blood for timing apheresis and maximizing yield

BACKGROUND: The collection of peripheral blood stem and progenitor cells (PBPCs) for transplantation can be time-consuming and expensive. Thus, the utility of counting CD34+ cells and white cells (WBCs) in the peripheral blood was evaluated as a predictor of CD34+ cell yield in the apheresis component. STUDY DESIGN AND METHODS: The WBC and CD34+ cell counts in the peripheral blood and the apheresis components from 216 collections were assessed. Sixty-three patients underwent mobilization with chemotherapy plus filgrastim, and 17 patients and 14 allogeneic PBPC donors did so with filgrastim alone. The relationship between the number of WBC and CD34+ cells in the peripheral blood and in the apheresis component was analyzed by using rank correlation and linear regression analysis. RESULTS: The correlation coefficient for CD34+ cells per liter of peripheral blood with CD34+ cell yield (x 10(6)/kg) was 0.87 (n = 216 collections). This correlation existed for many patient and collection variables. However, patients with acute myeloid leukemia had fewer CD34+ cells in the apheresis component at any level of peripheral blood CD34+ cell count. Components collected from patients with CD34+ cell counts below 10 x 10(6) per L in the peripheral blood contained a median of 0.75 x 10(6) CD34+ cells per kg. When the WBC count in the blood was below 5.0 x 10(9) per L, the median number of CD34+ cells in the peripheral blood was 5.6 x 10(6) per L (range, 1.0-15.5 x 10(6)/L). A very poor correlation was found between the WBC count in the blood and the CD34+ cell yield (p = 0.12, n = 158 collections). CONCLUSION: The number of CD34+ cells, but not WBCs, in the peripheral blood can be used as a predictor for timing of apheresis and estimating PBPC yield. This is a robust relationship not affected by a variety of patient and collection factors except the diagnosis of acute myeloid leukemia. Patients who undergo mobilization with chemotherapy and filgrastim also should undergo monitoring of peripheral blood CD34+ cell counts, beginning when the WBC count in the blood exceeds 1.0 to 5.0 x 10(9) per L.     

Technical and safety aspects of blood and marrow transplantation using G-CSF mobilized family donors

An allogeneic transplantation programme using immunoselected blood progenitor and bone marrow CD34+ cells has been established. Thirteen healthy HLA-matched, MLC negative sibling donors received two doses of 5 micrograms kg-1 G-CSF (s.c. daily) for 5 days. On days 4 and 5, large-volume mononuclear cell aphereses were performed (COBE Spectra) and on day 5 one unit of autologous blood was obtained. Mononuclear cells were pooled and cryopreserved after CD34+ cell-immunoselection on day 5. Bone marrow (BM) of the same donors was procured under routine conditions 10-45 days later (median: 27 days). The final graft consisted of blood CD34+ cells with either complete BM (n = 5) or immunoselected BM CD34+ cells (n = 8). The present paper describes the progenitor cell mobilization and apheresis protocol and analyzes the cell loss by BM and peripheral blood progenitor cell (PBPC) donation. Considerably larger amounts of mononuclear cells (CD45+), T-lymphocytes (CD3+) and platelets were lost by the apheresis as compared to bone marrow without apparent immediate clinical consequences for the donors. Owing to cross-cellular contamination of the apheresis concentrate, blood platelet count (PC) significantly decreased (mean PC after the second apheresis 116 x 10 microL-1); furthermore on average 3.04 x 10(10) CD3+ cells were removed by two apheresis sessions. This loss did not lead to long-term total lymphocyte count changes (2370 microL-1 versus 1889 microL-1) as observed during the long-term follow-up of 7/13 donors (mean 290 days). Subjectively, the PBPC collections were better accepted than BM donations in all but one family donor.     

TRANSPLANTATION AND CELLULAR ENGINEERING: Association of stress‐related perinatal factors and cord blood unit hematopoietic progenitors is dependent on delivery mode

BACKGROUND: Perinatal characteristics, variably utilized in cord blood (CB) selection for banking, affect CB hematopoietic progenitor cells (HPCs). The association between perinatal stress factors and CB unit HPCs was evaluated. STUDY DESIGN AND METHODS: Umbilical arterial (UA) pH, absolute and relative birth weight (BW) and placental weight (PW), and PW/BW ratio of 167 healthy, full‐term infants were compared with CB unit prefreeze total nucleated cells (TNCs), total CD34+ (TCD34+) cells, and total colony‐forming unit (CFU‐TOT) number. Cesarean section (C‐section, n = 104) and vaginal delivery subgroups were also analyzed. RESULTS: UA pH (median, 7.28; range, 7.04‐7.40) correlated with CB unit CFU‐TOT number (n = 166; r = ?0.32, p < 0.0001), TCD34+ cells (r = ?0.31, p < 0.0001), and TNCs (r = ?0.29, p = 0.0002). Similarly, BW, PW, and PW/BW ratio correlated with HPCs. In multiple linear regression analysis, CFU‐TOT number was predicted by collected CB TNCs and UA pH in vaginal deliveries (R² = 0.53), in contrast with TNCs, PW, and BW in C‐sections (R² = 0.37). TCD34+ cells were predicted by adding UA pH (vaginal deliveries, R² = 0.75) or PW (C‐sections, R² = 0.36) to collected CB TNCs. CONCLUSIONS: Stress‐related perinatal factors, particularly UA pH, are associated with CB unit HPCs and may improve unit selection. Multiple linear regression models may prove useful for predicting HPCs. Mode of delivery affects model choice; UA pH has a strong effect on HPCs in vaginal deliveries.     

Evaluation of an algorithm based on peripheral blood hematopoietic progenitor cell and CD34+ cell concentrations to optimize peripheral blood progenitor cell collection by apheresis

BACKGROUND: Quantification of peripheral blood (PB) CD34+ cells is commonly used to plan peripheral blood progenitor cell (PBPC) collection but is time-consuming. Sysmex has developed a hematology analyzer that can quickly identify a population of immature hematopoietic cells (HPCs) according to cell size, cell density, and differential lysis resistance, which may indicate the presence of PBPCs in PB. This prospective study has evaluated the potential of such method to predict the PBPC mobilization. STUDY DESIGN AND METHODS: A total of 141 patients underwent PBPC mobilization. PB HPCs and PB CD34+ cells were simultaneously quantified with a hematology analyzer (SE2100, Sysmex) and flow cytometry, respectively. The number of blood volumes processed was then based on PB CD34+ cell concentration. RESULTS: The optimal PB HPC level able to predict a minimal level of 10 x 10(6) PB CD34+ cells per L was 5 x 10(6) per L with positive and negative predictive values of 0.93 and 0.36 percent, respectively. For this cutoff point, sensitivity and specificity were 0.81 and 0.65, respectively. The median number of blood volumes processed according to the PB CD34+ cell count allowed us to perform only one apheresis procedure for a majority of patients. CONCLUSION: PB HPC quantification is very useful to quickly determine the initiation of PBPC apheresis especially for patients with higher concentrations. For patients exhibiting a lower HPC count (<5 x 10(6)/L), other parameters such as a CD34 test may be needed. Such a policy associated with a length of apheresis adapted to the richness in the PB CD34+ cells allows for optimizing the organization of centers with an improvement in patient comfort and economical savings.     

引物延伸预扩增后STR分型在鉴定母血中胎儿有核红细胞的应用

目的 探讨一种鉴定母血中胎儿有核红细胞（NRBC）的方法，为无创性产前基因诊断创造必要条件。方法 联苯胺染色识别孕妇外周血中NRBC，经显微操作获取，并以引物延伸预扩增（PEP）对单个NRBC进行全基凶组扩增后，用短串联重复序列（STR）分析其基因型，与父母基因型比对，确定该细胞的来源。结果 28例轻型β地中海贫血孕妇外周血样本中每例发现NRBC4～13个／5ml，经鉴定每例有胎儿NRBC2～8个／5ml，约43．6％的NRBC来源于胎儿。结论 PEP后STR基因型分析能有效鉴定孕妇外周血中NRBC的来源，使应用单个胎儿NRBC进行产前基因诊断成为可能。     

Evaluation of Quality Parameters for Cord Blood Donations

SUMMARY: BACKGROUND: Umbilical cord blood (CB) is widely used for hematopoietic stem cell transplantation and holds promise for the development of innovative medicinal products. In order to find out whether the conditions for collection and storage before processing might have an impact on the quality of CB preparations, viability and the clonogenic potential were assessed. METHODS: CB was collected under field conditions. Flow cytometry was used to determine leukocytes, CD34/CD45+ cells, viability, and nucleated red blood cells (NRBC). Clonogenic activity was determined using isolated mononuclear cells (MNC). RESULTS: Neither plasma citrate concentrations nor storage temperature (within 24 h) affected cell viability or colony formation. After storage for 49-80 h, leukocyte viability declined by about 16% compared to CB stored up to 24 h. In contrast, the clonogenic activity and CD34/CD45+ cell content were not affected. A higher gestational age was associated with a lower yield of clonogenic activity compared to midterm deliveries. NRBC varied widely (median 7.3%; range 0.63-17.3%) without relation to gestational age or colony formation. There was a close correlation between the percentage of viable CD34/CD45+ cells and colony formation (r = 0.77 for CFU-GM; r = 0.75 for CFU-C). CONCLUSIONS: The content of viable CD34/CD45+ cells represents the clonogenic activity of CB preparations. Therefore, determination of viable CD34/CD45+ cells should be generally performed as a routine quality control assay.     

Association of cord blood platelet characteristics and hematopoietic progenitor cells

BACKGROUND: Total nucleated cell (TNC) dose is associated with neutrophil and platelet (PLT) engraftment after cord blood (CB) transplantation and thus is used for selection of CB for banking. The goal of this study was to evaluate the internal relationships of CB PLT characteristics, TNC, and the hematopoietic progenitor cell (HPC) content of CB units. STUDY DESIGN AND METHODS: HPC and TNC counts of 167 CB units processed with an automated cell separation system were compared with CB PLT count and mean PLT volume (MPV). Megakaryocyte progenitors (CFU-MK) were cultured from a subset of units (n = 24). RESULTS: PLT concentration correlated with MPV (r = -0.39), which was also associated with both TNC and total CD34+ cells before and after processing (r = 0.37 and 0.35 and r = 0.41 and 0.42, respectively). In addition, MPV was associated with HPC counts in the CB unit. The p value was less than 0.001 for all associations. PLT count was inversely associated with markers of hematopoietic potential. Median removal of PLTs during processing was 62 percent (range, 40%-84%). All 24 CB units of the subset exhibited CFU-MK growth. In multivariate linear regression analysis, MPV improved prediction of the HPC content of the CB unit compared to prediction with CB volume and nucleated cell concentration only. CONCLUSION: Mean PLT volume correlated with current markers of CB hematopoietic potential and is potentially useful for evaluating CB collections for banking. The question of the clinical significance of PLT characteristics in CB transplantation remains unanswered.     

Influence of mode of birth and collection on WBC yields of umbilical cord blood units

BACKGROUND: The aim of this study was to determine the influence of mode of birth and umbilical cord blood (CB) collection before (in utero) or after delivery of the placenta (ex utero) on total number of WBCs and CD34+ cells in CB units. STUDY DESIGN AND METHODS: Consecutively donated, banked CB units were assessed for net volume, WBC concentration, total number of WBCs, proportion of CD34+ cells, and total number of CD34+ cells. These parameters were then correlated with the mode of birth and the mode of CB collection relative to the delivery of the placenta. RESULTS: A significantly higher CB volume was seen following cesarean section (n = 61) than following vaginal delivery (n = 157; median volume, 76 vs. 63 mL, respectively; p < 0.0001). In contrast, CB from vaginal delivery had a significantly higher WBC concentration compared with CB from cesarean section (medians, 17.1 x 10(9) and 13.6 x 10(9) WBCs/L, respectively; p < 0.0001). The mode of birth did not influence the proportion of CD34+ cells. A correlation was demonstrated between the total number of CD34+ cells and the total number of WBCs. As a consequence of the opposing effects on volume and WBC counts by cesarean section and vaginal delivery, there were no significant differences in the total number of WBCs or CD34+ cells for the CB units with mode of delivery in this study. No significant differences were found in CB with mode of CB collection (in utero [n = 58] or ex utero [n = 99]) following vaginal delivery. CONCLUSIONS: The mode of birth influences the CB WBC concentration and volume collected and should be taken into consideration for establishing any acceptance limits for CB units to be banked. There were no differences in CB with in utero or ex utero collections.     

The excessive numbers of total nucleated cells does not affect the performance of the CliniMACS

This prospective study was carried out in healthy donors and patients. The performance of the CliniMACS was evaluated with the comparison of the numbers of total nucleated cell (TNC) within and over the capacity of the normal scale column. In addition, large vs. normal scale column and manual vs. automated washing systems were also compared. A total of 44 selections were done. Eighteen normal scale selections were done with initial TNC numbers over 6 x 10(10) and 14 selections were performed below this number. None of the cases had CD34+ cell numbers over the capacity. Flow cytometry was used to check each separation performance for purity and recovery of CD34+ cells along with T- and B-cell depletion level parameters. All healthy donors had significantly better mean purity and recovery of CD34+ cells, and T- and B-cell depletion status than that of patients with values 95 vs. 85%, P: 0.006; 77 vs. 58%, P: 0.004; 4.55 log vs. 4.06 log, P: 0.004; 3.19 log vs. 2.63 log, P: 0.01, respectively. However, the performance of the system was not dependent on using the normal or large-scale column; automated or manual washing systems; and initial TNC numbers above (>6 x 10(10), range: 7.05-21.84 x 10(10), mean: 12.32 x 10(10)) or within (<6 x 10(10), range: 0.86-5.89 x 10(10), mean: 4.15 x 10(10)) the column capacity. In conclusion, the performance of the CliniMACS is more efficient in healthy donors than in patients. However, the performance of the system did not change as long as the numbers of CD34+ cells (range: 0.34-5.87 x 10(8)) were not exceeding the column capacity despite that more than 6 x 10(10) TNCs were used.     

人脐血间充质干细胞促进脐血CD34+细胞在NOD/SCID小鼠体内植入的实验研究

目的探讨人脐血间充质干细胞(MSC)联合人脐血CD34+细胞移植,能否促进CD34+细胞在NOD/SCID小鼠体内植入及加速其造血恢复.方法NOD/SCID小鼠于60Co 2.5 Gy照射后24h内由尾静脉输注胎儿脐血CD34+细胞1×105/只(低细胞量移植组)或1×106/只(高细胞量移植组),联合移植组同时输注脐血MSC 1×106/只.动态观察移植后小鼠外周血白细胞、血红蛋白和血小板恢复情况,于移植后第8周用流式细胞术检测存活小鼠骨髓中人CD45+、CD45+CD3+、CD45+CD19+和CD45+CD33+细胞的含量.结果①低细胞量移植时,联合移植组的植入率明显高于单纯移植组,分别为26.02%和16.52%(P＜0.05);高细胞量移植时,联合移植组和单纯移植组的植入率相近,分别为43.71%和39.23%(P＞0.05).②高细胞量联合移植组和单纯移植组的存活率分别为80%和70%;低细胞量联合移植组和单纯移植组的存活率分别为70%和50%.③无论高细胞量还是低细胞量组,联合移植小鼠白细胞、血红蛋白和血小板的恢复明显早于单纯移植组.④移植8周后,小鼠骨髓中人CD45+CD19+、CD45+CD33+细胞含量在低细胞量移植时,联合移植组高于单纯移植组;但在高细胞量移植时,两组之间差异无统计学意义.CD45+CD41a+细胞的含量无论在低细胞量和高细胞量移植时,联合移植组均高于单纯移植组.各组小鼠骨髓中CD45+CD3+细胞的含量均较少,且各组之间差异无统计学意义.结论①低细胞量移植时,人脐血MSC联合移植可提高人脐血CD34+细胞在小鼠体内的植入率.②人脐血MSC与人脐血CD34+细胞联合移植,加速NOD/SCID小鼠各系造血恢复,提高移植小鼠存活率.③MSC联合移植可促进人脐血CD34+细胞在NOD/SCID小鼠体内向粒系、B淋巴系和巨核系定向分化.