The renoprotective effects
of endothelial
progenitor cells (EPC)
have been clearly identified
and... more The renoprotective effects of endothelial progenitor cells (EPC) have been clearly identified and many results have shown their successful use in neovascularization of ischemic tissues associated with blood vessel diseases. However, the availability of a sustainable source of EPC for large therapeutic applications remains critical. The aim of this work is to ensure the production of a specific EPC profile from mesenchymal stem cells (MSC). Non-adherent EPC were significantly amplified after 6 days of angiogenic growth factor activation. Three day-EPC showed an over expression of the heamatopoietic markers CD34 and CD45 with a commitment to endothelial lineage with the expression of CD31 and CD146 cell markers. MSC lost their mesenchymal profile by down-regulating the CD90 and CD105 expression. Cytokeratin 8/18 and von Willebrand Factor VIII were progressively expressed on EPC as well as the monocytic CD14. In six-day EPC, all of the CD31+ cells expressed CD14 and CD146. CD54, CD11a, and CD49d adhesion molecules were also strongly expressed in both 3-day and 6-day EPC. These findings suggest that MSC-derived EPC might be a unique population of CD34+CD45+C D14+CD31+CD146+ cells sharing both stemness and progenitor characteristics of endothelial cells and monocytes, and that this culture model improved EPC production and expansion for a potential future use in vascular diseases.
A recently described long-term culture system for early human progenitor cells was established wi... more A recently described long-term culture system for early human progenitor cells was established with the murine preadipocyte stromal line FBMD-1 grown in 96-well plates; cobblestone areas formed by inoculated hematopoietic cells are determined in a limiting dilution setting after five weeks' culture. To compare the capacity of cobblestone-area-forming cell (CAFC) formation by bone marrow and leukapheresis products in this system, mononuclear cells (MNC) of both origins were cultured. As related to CD34 + cell content, CAFC yields after five weeks' culture were in the same range in bone marrow and leukapheresis stemming from patients with efficient mobilization of hematopoietic cells. In purified CD34 + cell fractions, the CAFC yield per inoculated cell number was considerably higher than in MNC; however, if the CAFC number was related to the inoculated CD34 + cell number in MNC and after purification, the yield was four to eight times decreased in purified fractions. Addition of the mature cells brought the CAFC yield back up to the numbers obtained in the unseparated MNC fraction. By contrast, slightly more advanced progenitors per CAFC were found in cultures of purified hematopoietic cells from both origins than in whole MNC. The results suggest that mature human accessory cells give noticeable support to recruitment of early progenitors on this feeder but lead to lower yield of GM progenitors.
Since the success of the first umbilical cord blood (UCB) transplantation in a child with Fanconi... more Since the success of the first umbilical cord blood (UCB) transplantation in a child with Fanconi anaemia in 1989, great interests have emerged for this source of stem cells. UCB provide an unlimited source of ethnically diverse stem cells and an alternative for bone marrow (BM) and peripheral blood (PB) heamatopoietic stem cell transplantation. Thus, UCB and manipulated stem cells are now collected and banked according to international accreditation standards for listing on registries allowing rapid search and accessibility world wide. This work aims to identify problems limiting the creation of a Moroccan cord blood bank and to highlight opportunities and issues of a new legislation promoting additional applications of cell therapy.
Journal of Hematotherapy & Stem Cell Research, 2000
Human early hematopoietic progenitors from bone marrow (BM) and leukapheresis products (LP) are h... more Human early hematopoietic progenitors from bone marrow (BM) and leukapheresis products (LP) are highly proliferative in presence of accessory cells in standard culture on the murine FBMD-1 cell feeder with weekly addition of human interleukin-3 (HuIL-3) and granulocyte-colony stimulating factor (HuG-CSF). If however purified CD34+ cells are cultured under otherwise identical conditions, cobblestone areas (CAFC) formed by the same number of target cells are diminished by more than 1 log, as we showed previously. This suggests that mature cells are involved in growth of early progenitors. To determine whether this bystander effect is mediated by soluble growth factors, or by direct cell-to-cell contact with early progenitors, we stimulated mature plastic adherent cells separately and tested the resulting conditioned supernatant (ACS) on CAFC and colony-forming unit-granulocyte-macrophage (CFU-GM) production. In ACS-complemented standard cultures of purified CD34+ cells, the yield of CAFC was up to 1 log higher if compared to parallel cultures without ACS. Likewise, the CFU-GM production was enhanced in presence of ACS, especially in the adherent fraction of the culture. When CD34+ cell cultures were performed with ACS but without added interleukin-3 (IL-3) and granulocyte colony-stimulating factor (G-CSF), CAFC production was in the same range as if these growth factors were added alone. Addition of anti-G-CSF antibody (Ab) to ACS decreased CAFC recruitment significantly, whereas anti-IL-3 Ab had no significant effect. These findings suggest that ACS complemented with IL-3 and G-CSF replaces the accessory cells largely; this is not only due to presence of G-CSF, because ACS in combination with recombinant growth factors mounts CAFC yield higher than saturating amounts of growth factors alone do. There must be further synergizing soluble factors in the supernatant.
ABSTRACT A recently described long-term culture system for early human progenitor cells was estab... more ABSTRACT A recently described long-term culture system for early human progenitor cells was established with the murine preadipocyte stromal line FBMD-1 grown in 96-well plates; cobblestone areas formed by inoculated hematopoietic cells are determined in a limiting dilution setting after five weeks' culture. To compare the capacity of cobblestone-area-forming cell (CAFC) formation by bone marrow and leukapheresis products in this system, mononuclear cells (MNC) of both origins were cultured. As related to CD34+ cell content, CAFC yields after five weeks' culture were in the same range in bone marrow and leukapheresis stemming from patients with efficient mobilization of hematopoietic cells. In purified CD34+ cell fractions, the CAFC yield per inoculated cell number was considerably higher than in MNC; however, if the CAFC number was related to the inoculated CD34+ cell number in MNC and after purification, the yield was four to eight times decreased in purified fractions. Addition of the mature cells brought the CAFC yield back up to the numbers obtained in the unseparated MNC fraction. By contrast, slightly more advanced progenitors per CAFC were found in cultures of purified hematopoietic cells from both origins than in whole MNC. The results suggest that mature human accessory cells give noticeable support to recruitment of early progenitors on this feeder but lead to lower yield of GM progenitors.
ABSTRACT Highly purified human CD34+ cells produced up to 500× less CFU-GM in short term culture ... more ABSTRACT Highly purified human CD34+ cells produced up to 500× less CFU-GM in short term culture (STC), and 10× less week-5 CAFC in murine FBMD-1 feeder long term culture (LTC) than MNC as calculated per input CD34+ cells, according to previous results (1). Cocultured CD34− cells, separated from CD34+ cells by a dialysis membrane, re-established efficient colony growth in STC. We now tested efficiency of conditioned media from normal blood adherent cells (ACS) after stimulation with 7ng/ml GM-CSF for 48h. Addition of ACS from 9 healthy donors enhanced growth of CFU-GM in STC of purified mobilized blood CD34+ cells at individually variable degree (2,5–168.8% additional colony yield). 82% additional colonies resulted from pooled supernatants. When this pooled ACS was added weekly at 1% v/v to CD34+ cells growing in LTC, the yield of weeks-5 CAFC raised from 67,1 to 1234,6 CAFC per 105 CD34+ cells (determined by limiting dilution method and extrapolated); neutralizing antibodies against G-CSF and SCF did not change this enhancement. Fractions bound to anion-exchange columns enhanced early CAFC (week-2), while unbound fractions late (week-5) CAFC. We conclude that the supportive effect of mature adherent cells must be due to secreted factors, which can be separated by their different net electric charge. (1) Mature accessory cells influence long term growth of human hematopoietic progenitors on a murine stromal cell feeder layer. L. Mazini, E. Wunder, H. Sovalat et al.; Stem Cells 16, 404-412 (1998)
Developing countries, including Egypt and North African countries, need to improve their quality ... more Developing countries, including Egypt and North African countries, need to improve their quality of research by enhancing international cooperation and exchanges of scientific information, as well as competing for obtaining international funds to support research activities. Research must comply with laws and other requirements for research that involves human subjects. The purpose of this article is to overview the status of health research ethics in Egypt and North African countries, with reference to other Middle Eastern countries. The EU and North African Migrants: Health and Health Systems project (EUNAM) has supported the revision of the status of health research ethics in Egypt and North African countries, by holding meetings and discussions to collect information about research ethics committees in Egypt, and revising the structure and guidelines of the committees, as well as reviewing the literature concerning ethics activities in the concerned countries. This overview has revealed that noticeable efforts have been made to regulate research ethics in certain countries in the Middle East. This can be seen in the new regulations, which contain the majority of protections mentioned in the international guidelines related to research ethics. For most of the internationally registered research ethics committees in North African countries, the composition and functionality reflect the international guidelines. There is growing awareness of research ethics in these countries, which extends to teaching efforts to undergraduate and postgraduate medical students.
EFFECT OF ACTIVATED ADHERENT CELL
CONDITIONED MEDIUM ON HUMAN CD341 CELL
RECRUITMENT IN SHORT- AN... more EFFECT OF ACTIVATED ADHERENT CELL CONDITIONED MEDIUM ON HUMAN CD341 CELL RECRUITMENT IN SHORT- AND LONG TERM CULTURES
Highly purified human CD341 cells produced up to 5003 less CFU-GM in short term culture (STC), and 103 less week-5 CAFC in murine FBMD-1 feeder long term culture (LTC) than MNC as calculated per input CD341 cells, according to previous results (1). Cocultured CD342 cells, separated from CD341 cells by a dialysis membrane, re-established efficient colony growth in STC. We now tested efficiency of conditioned media from normal blood adherent cells (ACS) after stimulation with 7ng/ml GM-CSF for 48h. Addition of ACS from 9 healthy donors enhanced growth of CFU-GM in STC of purified mobilized blood CD341 cells at individually variable degree (2,5–168.8% additional colony yield). 82% additional colonies resulted from pooled supernatants. When this pooled ACS was added weekly at 1% v/v to CD341 cells growing in LTC, the yield of weeks-5 CAFC raised from 67,1 to 1234,6 CAFC per 105 CD341 cells (determined by limiting dilution method and extrapolated); neutralizing antibodies against G-CSF and SCF did not change this enhancement. Fractions bound to anion-exchange columns enhanced early CAFC (week-2), while unbound fractions late (week-5) CAFC. We conclude that the supportive effect of mature adherent cells must be due to secreted factors, which can be separated by their different net electric charge.
Umbilical cord blood (UCB) is commonly used as the main heamatopoietic stem cell (HSC) source for... more Umbilical cord blood (UCB) is commonly used as the main heamatopoietic stem cell (HSC) source for allogeneic transplantation. Immediately available when compared to bone marrow (BM), UCB is now widely processed and stored as rigorously qualified units. These units were firstly estimated before processing using specific criteria for the donor’s choice, and by parameters such as Total Nucleated Cells (TNC), Mononuclear Cells (MNC), CD34+ cells and Colony Forming Units (CFU) for final processing. Appropriate assessment of factors related to maternal, obstetrical and neonatal variables showed these variables as the mainly contributors to a better UCB quality management. This preliminary study aimed to identify the effects of parameters such as maternal age, weight, gravid status, delivery mode, neonatal weight and gender on the MNC count in more 18 years old aged consent, and 33 full-term gravid donors. MNC count did not depend on gravid status and maternal age and weight as well as delivery mode and gender. However, neonatal weight and MNC counts appeared slightly associated independently to maternal age. This work might be helpful in developing and controlling procedures prior to UCB manipulation and would be improved for more UCB eligibility after bank creation.
The renoprotective effects
of endothelial
progenitor cells (EPC)
have been clearly identified
and... more The renoprotective effects of endothelial progenitor cells (EPC) have been clearly identified and many results have shown their successful use in neovascularization of ischemic tissues associated with blood vessel diseases. However, the availability of a sustainable source of EPC for large therapeutic applications remains critical. The aim of this work is to ensure the production of a specific EPC profile from mesenchymal stem cells (MSC). Non-adherent EPC were significantly amplified after 6 days of angiogenic growth factor activation. Three day-EPC showed an over expression of the heamatopoietic markers CD34 and CD45 with a commitment to endothelial lineage with the expression of CD31 and CD146 cell markers. MSC lost their mesenchymal profile by down-regulating the CD90 and CD105 expression. Cytokeratin 8/18 and von Willebrand Factor VIII were progressively expressed on EPC as well as the monocytic CD14. In six-day EPC, all of the CD31+ cells expressed CD14 and CD146. CD54, CD11a, and CD49d adhesion molecules were also strongly expressed in both 3-day and 6-day EPC. These findings suggest that MSC-derived EPC might be a unique population of CD34+CD45+C D14+CD31+CD146+ cells sharing both stemness and progenitor characteristics of endothelial cells and monocytes, and that this culture model improved EPC production and expansion for a potential future use in vascular diseases.
A recently described long-term culture system for early human progenitor cells was established wi... more A recently described long-term culture system for early human progenitor cells was established with the murine preadipocyte stromal line FBMD-1 grown in 96-well plates; cobblestone areas formed by inoculated hematopoietic cells are determined in a limiting dilution setting after five weeks' culture. To compare the capacity of cobblestone-area-forming cell (CAFC) formation by bone marrow and leukapheresis products in this system, mononuclear cells (MNC) of both origins were cultured. As related to CD34 + cell content, CAFC yields after five weeks' culture were in the same range in bone marrow and leukapheresis stemming from patients with efficient mobilization of hematopoietic cells. In purified CD34 + cell fractions, the CAFC yield per inoculated cell number was considerably higher than in MNC; however, if the CAFC number was related to the inoculated CD34 + cell number in MNC and after purification, the yield was four to eight times decreased in purified fractions. Addition of the mature cells brought the CAFC yield back up to the numbers obtained in the unseparated MNC fraction. By contrast, slightly more advanced progenitors per CAFC were found in cultures of purified hematopoietic cells from both origins than in whole MNC. The results suggest that mature human accessory cells give noticeable support to recruitment of early progenitors on this feeder but lead to lower yield of GM progenitors.
Since the success of the first umbilical cord blood (UCB) transplantation in a child with Fanconi... more Since the success of the first umbilical cord blood (UCB) transplantation in a child with Fanconi anaemia in 1989, great interests have emerged for this source of stem cells. UCB provide an unlimited source of ethnically diverse stem cells and an alternative for bone marrow (BM) and peripheral blood (PB) heamatopoietic stem cell transplantation. Thus, UCB and manipulated stem cells are now collected and banked according to international accreditation standards for listing on registries allowing rapid search and accessibility world wide. This work aims to identify problems limiting the creation of a Moroccan cord blood bank and to highlight opportunities and issues of a new legislation promoting additional applications of cell therapy.
Journal of Hematotherapy & Stem Cell Research, 2000
Human early hematopoietic progenitors from bone marrow (BM) and leukapheresis products (LP) are h... more Human early hematopoietic progenitors from bone marrow (BM) and leukapheresis products (LP) are highly proliferative in presence of accessory cells in standard culture on the murine FBMD-1 cell feeder with weekly addition of human interleukin-3 (HuIL-3) and granulocyte-colony stimulating factor (HuG-CSF). If however purified CD34+ cells are cultured under otherwise identical conditions, cobblestone areas (CAFC) formed by the same number of target cells are diminished by more than 1 log, as we showed previously. This suggests that mature cells are involved in growth of early progenitors. To determine whether this bystander effect is mediated by soluble growth factors, or by direct cell-to-cell contact with early progenitors, we stimulated mature plastic adherent cells separately and tested the resulting conditioned supernatant (ACS) on CAFC and colony-forming unit-granulocyte-macrophage (CFU-GM) production. In ACS-complemented standard cultures of purified CD34+ cells, the yield of CAFC was up to 1 log higher if compared to parallel cultures without ACS. Likewise, the CFU-GM production was enhanced in presence of ACS, especially in the adherent fraction of the culture. When CD34+ cell cultures were performed with ACS but without added interleukin-3 (IL-3) and granulocyte colony-stimulating factor (G-CSF), CAFC production was in the same range as if these growth factors were added alone. Addition of anti-G-CSF antibody (Ab) to ACS decreased CAFC recruitment significantly, whereas anti-IL-3 Ab had no significant effect. These findings suggest that ACS complemented with IL-3 and G-CSF replaces the accessory cells largely; this is not only due to presence of G-CSF, because ACS in combination with recombinant growth factors mounts CAFC yield higher than saturating amounts of growth factors alone do. There must be further synergizing soluble factors in the supernatant.
ABSTRACT A recently described long-term culture system for early human progenitor cells was estab... more ABSTRACT A recently described long-term culture system for early human progenitor cells was established with the murine preadipocyte stromal line FBMD-1 grown in 96-well plates; cobblestone areas formed by inoculated hematopoietic cells are determined in a limiting dilution setting after five weeks' culture. To compare the capacity of cobblestone-area-forming cell (CAFC) formation by bone marrow and leukapheresis products in this system, mononuclear cells (MNC) of both origins were cultured. As related to CD34+ cell content, CAFC yields after five weeks' culture were in the same range in bone marrow and leukapheresis stemming from patients with efficient mobilization of hematopoietic cells. In purified CD34+ cell fractions, the CAFC yield per inoculated cell number was considerably higher than in MNC; however, if the CAFC number was related to the inoculated CD34+ cell number in MNC and after purification, the yield was four to eight times decreased in purified fractions. Addition of the mature cells brought the CAFC yield back up to the numbers obtained in the unseparated MNC fraction. By contrast, slightly more advanced progenitors per CAFC were found in cultures of purified hematopoietic cells from both origins than in whole MNC. The results suggest that mature human accessory cells give noticeable support to recruitment of early progenitors on this feeder but lead to lower yield of GM progenitors.
ABSTRACT Highly purified human CD34+ cells produced up to 500× less CFU-GM in short term culture ... more ABSTRACT Highly purified human CD34+ cells produced up to 500× less CFU-GM in short term culture (STC), and 10× less week-5 CAFC in murine FBMD-1 feeder long term culture (LTC) than MNC as calculated per input CD34+ cells, according to previous results (1). Cocultured CD34− cells, separated from CD34+ cells by a dialysis membrane, re-established efficient colony growth in STC. We now tested efficiency of conditioned media from normal blood adherent cells (ACS) after stimulation with 7ng/ml GM-CSF for 48h. Addition of ACS from 9 healthy donors enhanced growth of CFU-GM in STC of purified mobilized blood CD34+ cells at individually variable degree (2,5–168.8% additional colony yield). 82% additional colonies resulted from pooled supernatants. When this pooled ACS was added weekly at 1% v/v to CD34+ cells growing in LTC, the yield of weeks-5 CAFC raised from 67,1 to 1234,6 CAFC per 105 CD34+ cells (determined by limiting dilution method and extrapolated); neutralizing antibodies against G-CSF and SCF did not change this enhancement. Fractions bound to anion-exchange columns enhanced early CAFC (week-2), while unbound fractions late (week-5) CAFC. We conclude that the supportive effect of mature adherent cells must be due to secreted factors, which can be separated by their different net electric charge. (1) Mature accessory cells influence long term growth of human hematopoietic progenitors on a murine stromal cell feeder layer. L. Mazini, E. Wunder, H. Sovalat et al.; Stem Cells 16, 404-412 (1998)
Developing countries, including Egypt and North African countries, need to improve their quality ... more Developing countries, including Egypt and North African countries, need to improve their quality of research by enhancing international cooperation and exchanges of scientific information, as well as competing for obtaining international funds to support research activities. Research must comply with laws and other requirements for research that involves human subjects. The purpose of this article is to overview the status of health research ethics in Egypt and North African countries, with reference to other Middle Eastern countries. The EU and North African Migrants: Health and Health Systems project (EUNAM) has supported the revision of the status of health research ethics in Egypt and North African countries, by holding meetings and discussions to collect information about research ethics committees in Egypt, and revising the structure and guidelines of the committees, as well as reviewing the literature concerning ethics activities in the concerned countries. This overview has revealed that noticeable efforts have been made to regulate research ethics in certain countries in the Middle East. This can be seen in the new regulations, which contain the majority of protections mentioned in the international guidelines related to research ethics. For most of the internationally registered research ethics committees in North African countries, the composition and functionality reflect the international guidelines. There is growing awareness of research ethics in these countries, which extends to teaching efforts to undergraduate and postgraduate medical students.
EFFECT OF ACTIVATED ADHERENT CELL
CONDITIONED MEDIUM ON HUMAN CD341 CELL
RECRUITMENT IN SHORT- AN... more EFFECT OF ACTIVATED ADHERENT CELL CONDITIONED MEDIUM ON HUMAN CD341 CELL RECRUITMENT IN SHORT- AND LONG TERM CULTURES
Highly purified human CD341 cells produced up to 5003 less CFU-GM in short term culture (STC), and 103 less week-5 CAFC in murine FBMD-1 feeder long term culture (LTC) than MNC as calculated per input CD341 cells, according to previous results (1). Cocultured CD342 cells, separated from CD341 cells by a dialysis membrane, re-established efficient colony growth in STC. We now tested efficiency of conditioned media from normal blood adherent cells (ACS) after stimulation with 7ng/ml GM-CSF for 48h. Addition of ACS from 9 healthy donors enhanced growth of CFU-GM in STC of purified mobilized blood CD341 cells at individually variable degree (2,5–168.8% additional colony yield). 82% additional colonies resulted from pooled supernatants. When this pooled ACS was added weekly at 1% v/v to CD341 cells growing in LTC, the yield of weeks-5 CAFC raised from 67,1 to 1234,6 CAFC per 105 CD341 cells (determined by limiting dilution method and extrapolated); neutralizing antibodies against G-CSF and SCF did not change this enhancement. Fractions bound to anion-exchange columns enhanced early CAFC (week-2), while unbound fractions late (week-5) CAFC. We conclude that the supportive effect of mature adherent cells must be due to secreted factors, which can be separated by their different net electric charge.
Umbilical cord blood (UCB) is commonly used as the main heamatopoietic stem cell (HSC) source for... more Umbilical cord blood (UCB) is commonly used as the main heamatopoietic stem cell (HSC) source for allogeneic transplantation. Immediately available when compared to bone marrow (BM), UCB is now widely processed and stored as rigorously qualified units. These units were firstly estimated before processing using specific criteria for the donor’s choice, and by parameters such as Total Nucleated Cells (TNC), Mononuclear Cells (MNC), CD34+ cells and Colony Forming Units (CFU) for final processing. Appropriate assessment of factors related to maternal, obstetrical and neonatal variables showed these variables as the mainly contributors to a better UCB quality management. This preliminary study aimed to identify the effects of parameters such as maternal age, weight, gravid status, delivery mode, neonatal weight and gender on the MNC count in more 18 years old aged consent, and 33 full-term gravid donors. MNC count did not depend on gravid status and maternal age and weight as well as delivery mode and gender. However, neonatal weight and MNC counts appeared slightly associated independently to maternal age. This work might be helpful in developing and controlling procedures prior to UCB manipulation and would be improved for more UCB eligibility after bank creation.
Uploads
of endothelial
progenitor cells (EPC)
have been clearly identified
and many results
have shown their successful
use in neovascularization
of ischemic
tissues associated with
blood vessel diseases.
However, the availability
of a sustainable source of
EPC for large therapeutic
applications remains critical.
The aim of this
work is to ensure the
production of a specific
EPC profile from mesenchymal
stem cells (MSC).
Non-adherent EPC were
significantly amplified after 6 days of angiogenic
growth factor activation. Three day-EPC showed
an over expression of the heamatopoietic markers
CD34 and CD45 with a commitment to endothelial
lineage with the expression of CD31 and CD146
cell markers. MSC lost their mesenchymal profile by
down-regulating the CD90 and CD105 expression.
Cytokeratin 8/18 and von
Willebrand Factor VIII
were progressively expressed
on EPC as well
as the monocytic CD14.
In six-day EPC, all of the
CD31+ cells expressed
CD14 and CD146. CD54,
CD11a, and CD49d adhesion
molecules were also
strongly expressed in both
3-day and 6-day EPC.
These findings suggest
that MSC-derived EPC
might be a unique population
of CD34+CD45+C
D14+CD31+CD146+
cells sharing both stemness
and progenitor characteristics
of endothelial cells
and monocytes, and that this culture model improved
EPC production and expansion for a potential future
use in vascular diseases.
CONDITIONED MEDIUM ON HUMAN CD341 CELL
RECRUITMENT IN SHORT- AND LONG
TERM CULTURES
Highly purified human CD341 cells produced up to 5003 less
CFU-GM in short term culture (STC), and 103 less week-5 CAFC
in murine FBMD-1 feeder long term culture (LTC) than MNC as
calculated per input CD341 cells, according to previous results
(1). Cocultured CD342 cells, separated from CD341 cells by a
dialysis membrane, re-established efficient colony growth in STC.
We now tested efficiency of conditioned media from normal blood
adherent cells (ACS) after stimulation with 7ng/ml GM-CSF for
48h. Addition of ACS from 9 healthy donors enhanced growth of
CFU-GM in STC of purified mobilized blood CD341 cells at individually
variable degree (2,5–168.8% additional colony yield).
82% additional colonies resulted from pooled supernatants. When
this pooled ACS was added weekly at 1% v/v to CD341 cells
growing in LTC, the yield of weeks-5 CAFC raised from 67,1 to
1234,6 CAFC per 105 CD341 cells (determined by limiting dilution
method and extrapolated); neutralizing antibodies against
G-CSF and SCF did not change this enhancement. Fractions bound
to anion-exchange columns enhanced early CAFC (week-2), while
unbound fractions late (week-5) CAFC. We conclude that the supportive
effect of mature adherent cells must be due to secreted factors,
which can be separated by their different net electric charge.
Immediately available when compared to bone marrow (BM), UCB is now widely processed and stored as rigorously
qualified units. These units were firstly estimated before processing using specific criteria for the donor’s choice, and
by parameters such as Total Nucleated Cells (TNC), Mononuclear Cells (MNC), CD34+ cells and Colony Forming Units
(CFU) for final processing. Appropriate assessment of factors related to maternal, obstetrical and neonatal variables showed
these variables as the mainly contributors to a better UCB quality management. This preliminary study aimed to identify the
effects of parameters such as maternal age, weight, gravid status, delivery mode, neonatal weight and gender on the MNC
count in more 18 years old aged consent, and 33 full-term gravid donors. MNC count did not depend on gravid status and
maternal age and weight as well as delivery mode and gender. However, neonatal weight and MNC counts appeared slightly
associated independently to maternal age. This work might be helpful in developing and controlling procedures prior to
UCB manipulation and would be improved for more UCB eligibility after bank creation.
of endothelial
progenitor cells (EPC)
have been clearly identified
and many results
have shown their successful
use in neovascularization
of ischemic
tissues associated with
blood vessel diseases.
However, the availability
of a sustainable source of
EPC for large therapeutic
applications remains critical.
The aim of this
work is to ensure the
production of a specific
EPC profile from mesenchymal
stem cells (MSC).
Non-adherent EPC were
significantly amplified after 6 days of angiogenic
growth factor activation. Three day-EPC showed
an over expression of the heamatopoietic markers
CD34 and CD45 with a commitment to endothelial
lineage with the expression of CD31 and CD146
cell markers. MSC lost their mesenchymal profile by
down-regulating the CD90 and CD105 expression.
Cytokeratin 8/18 and von
Willebrand Factor VIII
were progressively expressed
on EPC as well
as the monocytic CD14.
In six-day EPC, all of the
CD31+ cells expressed
CD14 and CD146. CD54,
CD11a, and CD49d adhesion
molecules were also
strongly expressed in both
3-day and 6-day EPC.
These findings suggest
that MSC-derived EPC
might be a unique population
of CD34+CD45+C
D14+CD31+CD146+
cells sharing both stemness
and progenitor characteristics
of endothelial cells
and monocytes, and that this culture model improved
EPC production and expansion for a potential future
use in vascular diseases.
CONDITIONED MEDIUM ON HUMAN CD341 CELL
RECRUITMENT IN SHORT- AND LONG
TERM CULTURES
Highly purified human CD341 cells produced up to 5003 less
CFU-GM in short term culture (STC), and 103 less week-5 CAFC
in murine FBMD-1 feeder long term culture (LTC) than MNC as
calculated per input CD341 cells, according to previous results
(1). Cocultured CD342 cells, separated from CD341 cells by a
dialysis membrane, re-established efficient colony growth in STC.
We now tested efficiency of conditioned media from normal blood
adherent cells (ACS) after stimulation with 7ng/ml GM-CSF for
48h. Addition of ACS from 9 healthy donors enhanced growth of
CFU-GM in STC of purified mobilized blood CD341 cells at individually
variable degree (2,5–168.8% additional colony yield).
82% additional colonies resulted from pooled supernatants. When
this pooled ACS was added weekly at 1% v/v to CD341 cells
growing in LTC, the yield of weeks-5 CAFC raised from 67,1 to
1234,6 CAFC per 105 CD341 cells (determined by limiting dilution
method and extrapolated); neutralizing antibodies against
G-CSF and SCF did not change this enhancement. Fractions bound
to anion-exchange columns enhanced early CAFC (week-2), while
unbound fractions late (week-5) CAFC. We conclude that the supportive
effect of mature adherent cells must be due to secreted factors,
which can be separated by their different net electric charge.
Immediately available when compared to bone marrow (BM), UCB is now widely processed and stored as rigorously
qualified units. These units were firstly estimated before processing using specific criteria for the donor’s choice, and
by parameters such as Total Nucleated Cells (TNC), Mononuclear Cells (MNC), CD34+ cells and Colony Forming Units
(CFU) for final processing. Appropriate assessment of factors related to maternal, obstetrical and neonatal variables showed
these variables as the mainly contributors to a better UCB quality management. This preliminary study aimed to identify the
effects of parameters such as maternal age, weight, gravid status, delivery mode, neonatal weight and gender on the MNC
count in more 18 years old aged consent, and 33 full-term gravid donors. MNC count did not depend on gravid status and
maternal age and weight as well as delivery mode and gender. However, neonatal weight and MNC counts appeared slightly
associated independently to maternal age. This work might be helpful in developing and controlling procedures prior to
UCB manipulation and would be improved for more UCB eligibility after bank creation.