Long-term Effects of Regular Whole Blood Donation on Peripheral Blood CD34+ Cells Population with using Leukoreduction Filters.

Background & Objective: Trapped cell population in leukoreduction filters (LRFs) contains such a significant number of CD34+ hematopoietic stem cells that can be recovered to be used in research studies. Methods: Samples (n=20) were obtained from 10 first-time donors and 10 regular blood donors with more than 30 times blood donation. After separating leukocytes from LRFs by backflushing, total leukocyte number and differential count were determined in both groups using an automated haemocytometer. Then cell viability and CD34+ cell quantification were assessed using 7- amino-actinomycin D and fluorescent-labeled monoclonal antibodies using flow cytometry, respectively. Results: Total leukocyte count was 665±164.92×106 in the first-time blood donors and 883±233.89×106 in the regular donors, which were not significantly different (P=0.08). While the number of CD34+ cells was significantly reduced in the regular donors compared to the first-time donors (0.58±0.20×106/µL vs. 0.36±0.22×106/µL; P=0.034). There was no significant difference in terms of absolute neutrophil count (10.58±3.66×06 vs. 13.17±6.45×106/µL; P=0.349), lymphocytes (7.75±3.11×106 vs. 10.38±3.77×106 /µL; P=0.917), and monocytes (2.31±0.88×106 vs. 2.59±1.09×106/µL; P=0.591) between the first-time and regular donor groups, respectively. Based on the correlation coefficients, the participants' age had no significant effect on these variables. Conclusion: The results of this study depicted that regular blood donation reduces the number of CD34+ cells in the peripheral blood (PB) of regular donors while it has no significant effect on the ratio of myeloid to lymphoid cells of the two groups.

Optimizing the recovery of peripheral blood mononuclear cells trapped in leukoreduction filters - A comparison study

Abstract Introduction The isolation of captured peripheral blood mononuclear cells (PBMNCs) from leukoreduction filters (LRFs) can be of great importance in terms of bringing the lost cells back into use. Objective The aim of this study was to evaluate various methods based on their potential to recover the peripheral blood cells from LRFs with a focus on mononuclear cells (MNCs). Method For cell isolation from LRFs, three distinct methods (back-flushing, direct and vacuum pump) were compared through the calculation of the yield of isolated MNCs. The viability of extracted cells was determined by the flow cytometry technique. Moreover, the recovered MNCs were characterized regarding the presence of blood stem cell purification. The cell culture, microscopic observation, and immunophenotyping were employed to characterize the blood stem cells (hematopoietic, mesenchymal and progenitor endothelial stem cells). Results The yield of isolation obtained in the back-flushing, direct and vacuum pump methods were 17.7 ± 1.28, 17.3 ± 0.96 and 21.2 ± 0.90 percent, respectively. Although the highest potential for total blood cell recovery belonged to the vacuum pump method, the lowest cell viability (85.73 ± 4.84%) was observed in this method. However, the isolation process of the back-flushing and direct methods had less effect on cell viability. The characterization of the isolated MNCs displayed that the dominant positive phenotype was for CD34/CD45, indicating hematopoietic stem cells. In addition, the endothelial stem/progenitor cells were significantly detected as CD31/CD133 positive cells. Conclusion According to our results and considering the safety and efficiency potential of each of the applied methods, the back-flushing in comparison with the other methods can be considered a suitable procedure for MNC isolation from LRFs.

Optimizing the recovery of peripheral blood mononuclear cells trapped in leukoreduction filters - A comparison study.

INTRODUCTION: The isolation of captured peripheral blood mononuclear cells (PBMNCs) from leukoreduction filters (LRFs) can be of great importance in terms of bringing the lost cells back into use. OBJECTIVE: The aim of this study was to evaluate various methods based on their potential to recover the peripheral blood cells from LRFs with a focus on mononuclear cells (MNCs). METHOD: For cell isolation from LRFs, three distinct methods (back-flushing, direct and vacuum pump) were compared through the calculation of the yield of isolated MNCs. The viability of extracted cells was determined by the flow cytometry technique. Moreover, the recovered MNCs were characterized regarding the presence of blood stem cell purification. The cell culture, microscopic observation, and immunophenotyping were employed to characterize the blood stem cells (hematopoietic, mesenchymal and progenitor endothelial stem cells). RESULTS: The yield of isolation obtained in the back-flushing, direct and vacuum pump methods were 17.7 ±â€¯1.28, 17.3 ±â€¯0.96 and 21.2 ±â€¯0.90 percent, respectively. Although the highest potential for total blood cell recovery belonged to the vacuum pump method, the lowest cell viability (85.73 ±â€¯4.84%) was observed in this method. However, the isolation process of the back-flushing and direct methods had less effect on cell viability. The characterization of the isolated MNCs displayed that the dominant positive phenotype was for CD34/CD45, indicating hematopoietic stem cells. In addition, the endothelial stem/progenitor cells were significantly detected as CD31/CD133 positive cells. CONCLUSION: According to our results and considering the safety and efficiency potential of each of the applied methods, the back-flushing in comparison with the other methods can be considered a suitable procedure for MNC isolation from LRFs.

Gene expression analysis of noncoding PCA3 gene in patients with chronic myeloid leukemia.

INTRODUCTION AND OBJECTIVES: Nowadays, noncoding RNAs are of special significance to scientists. Among RNAs, long noncoding RNAs (lncRNAs) have an important role in gene expression regulation. Recent studies show aberrant lncRNAs expression in different types of cancer including blood malignancies. As such, lncRNAs could be a possible way for diagnosis and treatment of certain cancers. In the current study, the level of PCA3 gene expression in patients with chronic myeloid leukemia (CML) was compared with normal individuals to find out whether the level of PCA3 in CML patients is increased according to our hypothesis. MATERIALS AND METHODS: The buffy coat was collected from peripheral blood of thirty untreated CML patients (BCR-ABL positive) and twenty normal volunteers. RNA was extracted from white blood cells and cDNA was synthesized. The level of PCA3 gene expression was measured using quantitative reverse transcription-polymerase chain reaction. RESULTS: The results showed that PCA3 has expression in both normal and leukemic white blood cells. The data also revealed higher expression of PCA3 in leukemic patients, compared to the normal counterpart (P < 0.05). CONCLUSION: The unusual increase in PCA3 gene expression in CML patients suggests the need for more research on mechanisms of molecular pathways related to PCA3 which could help achieve better treatment.