scholarly journals Human peripheral blood mononuclear cells enriched in endothelial progenitor cells via quality and quantity controlled culture accelerate vascularization and wound healing in a porcine wound model

2018 ◽  
Vol 27 (7) ◽  
pp. 1068-1079 ◽  
Author(s):  
Makiko Kado ◽  
Rica Tanaka ◽  
Kayo Arita ◽  
Kayoko Okada ◽  
Rie Ito-Hirano ◽  
...  
Keyword(s):  
Wound Healing ◽  
Progenitor Cells ◽  
Endothelial Progenitor Cells ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Skin Defect ◽  
Endothelial Progenitor ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells

The transplantation of endothelial progenitor cells (EPCs) is used to promote wound angiogenesis. In patients with chronic wounds and accompanying morbidities, EPCs are often compromised in number and function. To overcome these limitations, we previously developed a quality and quantity controlled (QQ) culture system to enrich peripheral blood mononuclear cells (PBMNCs) in EPCs. To evaluate the wound healing efficacy of mononuclear cells (MNCs) harvested after QQ culture (QQMNCs), preclinical studies were performed on large animals. MNCs harvested from the blood of healthy human subjects were cultured in the presence of angiogenic cytokines and growth factors in a serum-free medium for 7 days. A total of 5 × 106 QQMNCs per full-thickness skin defect or control saline was injected into wounds induced in cyclosporine-immunosuppressed pigs. EPC colony-forming assays revealed a significantly higher number of definitive (partially differentiated) EPC colony-forming units in QQMNCs. Flow cytometry evaluation of QQMNC surface markers showed enrichment of CD34+ and CD133+ stem cell populations, significant reduction in CCR2+ cell percentages, and a greater than 10-fold increase in the percentage of anti-inflammatory M2-type macrophages (CD206+ cells) compared with PBMNCs. Wounds treated with QQMNCs had a significantly higher closure rate. Wounds were harvested, frozen, and sectioned at day 21 postoperatively. Hematoxylin and eosin staining revealed that the epithelization of QQMNC-treated wounds was more advanced than in controls. Treated wounds developed granulation tissue with more mature collagen and larger capillary networks. CD31 and human mitochondrial co-staining confirmed the presence of differentiated human cells within newly formed vessels. Real-time polymerase chain reaction (PCR) showed upregulation of interleukin 6 (IL-6), IL-10, and IL-4 in the wound bed, suggesting paracrine activity of the transplanted QQMNCs. Our data demonstrate for the first time that QQ culture of MNCs obtained from a small amount of peripheral blood yields vasculogenic and therapeutic cells effective in wound healing.

scholarly journals EFFECT OF FLAVONOIDS ON OXIDATIVE STRESS, APOPTOSIS, AND CELL MARKERS OF PERIPHERAL BLOOD-DERIVED ENDOTHELIAL PROGENITOR CELLS: AN IN VITRO STUDY

2021 ◽  
pp. 39-42
Author(s):  
WAHYU WIDOWATI ◽  
RIMONTA F. GUNANEGARA ◽  
TERESA LILIANA WARGASETIA ◽  
HANNA SARI WIDYA KUSUMA ◽  
SEILA ARUMWARDANA ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Progenitor Cells ◽  
Endothelial Progenitor Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
In Vitro Study ◽  
Endothelial Progenitor ◽  
Peripheral Blood Mononuclear ◽  
Cell Markers

Objective: Circulating EPCs (endothelial progenitor cells) play a role in neovascularization and vascular repair. Oxidative stress impairs endothelial progenitor. Flavonoid is a phytochemical compound for antioxidant activity. Flavonoid effects toward oxidative stress, apoptosis, and expression of the cell markers on EPCs are not fully understood. This study was aimed to elucidate the effects of quercetin, kaempferol, and myricetin toward oxidative stress, apoptosis, and cell markers of peripheral blood-derived-EPCs. Methods: EPCs (endothelial progenitor cells) were isolated from peripheral blood mononuclear cells (PBMNCs) using cultivation under EPCs spesific media. Oxidative stress in EPCs was induced by H2O2 and then treated by quercetin, kaempferol, and myricetin. Cytotoxicity was measured by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay, while intracellular reactive oxygen species (ROS), apoptosis and characterization of cells, which expressed CD133 and KDR, was measured using flow cytometry. Results: Quercetin, kaempferol, and myricetin at concentration 12.50 µmol/l were not toxic on EPCs as the cells viability were 96.11±4.03%, 95.42±7.75%, and 94.22±9.49%, respectively. Flavonoids decreased intracellular ROS level in EPCs (quercetin: 14.38±1.47%, kaempferol: 20.21±6.25%, and myricetin: 13.88±4.02%) compared to EPCs treated with H2O2 (30.70%±1.04). Percetage of EPCs apoptosis was not significantly different among each treatment. Immunophenotyping showed the increasing of CD133 and KDR expression in EPCs treated with flavonoids. Conclusion: Quercetin, kaempferol, and myricetin were safe for EPCs, decreased ROS levels, and increased CD133 and KDR expression. However, the flavonoids did not significantly affect EPCs apoptosis.

scholarly journals Secretome of Peripheral Blood Mononuclear Cells Enhances Wound Healing

PLoS ONE ◽  
2013 ◽  
Vol 8 (3) ◽  
pp. e60103 ◽  
Author(s):  
Michael Mildner ◽  
Stefan Hacker ◽  
Thomas Haider ◽  
Maria Gschwandtner ◽  
Gregor Werba ◽  
...  
Keyword(s):  
Wound Healing ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells

scholarly journals Thawed and Washed Apheresis Products Keep an Excellent CD34+ Cells Viability for 24 Hours Using Either Voluven or Normal Saline Plus Albumin

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 5738-5738
Author(s):  
Miguel Blanquer Blanquer ◽  
Carmen Alguero ◽  
Pilar Menchon ◽  
Assumpta Ferrer ◽  
Pilar Martínez Avilés ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Peripheral Blood Mononuclear Cells ◽  
Normal Saline ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Peripheral Blood Mononuclear ◽  
Time Points ◽  
Cd34 Cells ◽  
Blood Mononuclear Cells ◽  
The Mean

Abstract Cryopreservation of products rich in progenitor cells is mandatory for the feasibility of autologous hematopietic progenitor cells transplants. The most used product nowadays is the apheresis of mobilized peripheral blood mononuclear cells. However, cryopreservation implies the use of cryoprotectant molecules, such as DMSO, that can be toxic for the patients. Moreover, the thawing of the product goes with some unavoidable cell death and liberation of cytoplasmic content such as cytokines to the medium. And so, adverse reactions during the product infusion are not infrequent. Our group has demonstrated that washing the thawed apheresis products both with Voluven or with normal saline plus 5% albumin (NSA) is able to almost completely avoid infusion reactions without losing CD34+ cells. Here we wanted to know whether it also had the benefit of extending CD34+ viability for 24 hours after washing. METHODS: We thawed 3 spare peripheral blood mononuclear cells apheresis products that had been cryopreserved with 9%DMSO. Ten mL of each product were separated and the remaining volume was splitted in 2 bags to be washed either with Voluven or NSA. Sepax 2 smartwash automatic program was used to wash the cells. The washed cells were stored at 4ºC and a sample was taken and immediately analyzed at 0h, 1h, 2h, 4h and 24h. At the same time points 10 mL of each bag were separated and kept 30 min. at room temperature (RT) before being analyzed. A blood count was performed on all the samples. Flow cytometry was used to measure CD45+ and CD34+ cells viability by 7AAD staining. RESULTS: The mean CD45+ cells viability was 69% after thawing the cells ,81% immediately after washing them with Voluven, and 81%, 80%, 79% and 73% 1h, 2h, 4h and 24h after the wash. When using NSA the mean CD45+ viability was 79%, 92%, 93%, 92%, 93% and 87% at the same time points. When the samples were kept for 30 additional minutes at RT, the mean CD45+ viability was 78%, 81%, 84%, 82%, 84% and 81% with Voluven, and 80%, 92%, 94%, 91%, 95% and 86% with NSA. Regarding CD34+ cells, when washed with Voluven the mean viability was 46%, 87%, 87%, 92%, 83% and 69%, while it was 61%, 91%, 91%, 92%, 89% and 84% when NSA was used. The mean CD34+ cells viability results after 30' at RT for each time point were 79%, 90%, 82%, 91%, 82% and 81% with Voluven and 85%, 90%, 90%, 94%, 93% and 86% with NSA. Moreover the mean viable CD34+ cells recovery at 24h was 90.94% for Voluven and 87,88% for NSA. CONCLUSIONS: We have obtained an excellent stability of the CD34+ cells viability for 24h after washing the mobilized mononuclear cells apheresis products both with Voluven and with NSA when the products are stored at 4ºC. Moreover, the viability is not affected when the cells are kept for an additional 30' at RT. The viable CD34+ cells loss at 24h was scarce. Disclosures Blanquer Blanquer: Pfizer: Research Funding.

CD34+ progenitor cells mobilized by natalizumab are not a relevant reservoir for JC virus

2010 ◽  
Vol 17 (2) ◽  
pp. 151-156 ◽  
Author(s):  
Clemens Warnke ◽  
Vsevolod Smolianov ◽  
Thomas Dehmel ◽  
Marcel Andrée ◽  
Hartmut Hengel ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Jc Virus ◽  
Viral Dna ◽  
Peripheral Blood Mononuclear ◽  
Cd34 Cells ◽  
Blood Mononuclear Cells ◽  
Natalizumab Treatment

Background: Progressive multifocal leukoencephalopathy (PML) is associated with natalizumab treatment in patients with multiple sclerosis (MS). It has been hypothesized that natalizumab mobilizes JC virus (JCV)-infected haematopoietic progenitor cells mediating viraemia and subsequently this disease. Objective: The objective of this study was to investigate peripheral haematopoietic progenitor cells for evidence of JCV DNA in MS patients treated with natalizumab. Methods: We assessed JCV and cytomegalovirus (CMV) DNA in magnetically separated CD34+ haematopoietic progenitor cells, peripheral blood mononuclear cells and plasma of 67 natalizumab-treated patients with MS and six PML patients. Results: Viral DNA was not detectable in CD34+ haematopoietic progenitor or peripheral blood mononuclear cells from any sample. Two plasma samples from patients with MS while undergoing natalizumab treatment were JCV-positive. In one case clinically manifest PML developed 8 months thereafter. Conclusions: Our findings do not support the hypothesis that natalizumab mobilizes JC virus-infected CD34+ cells from the bone marrow mediating JC viraemia. Notably, JC viraemia was detected in one patient with MS prior to developing clinical PML. This warrants further study.

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