Generation of T cells from cytokine-mobilized peripheral blood and adult bone marrow CD34+ cells

Abstract The present study compared the T-cell progenitor content of CD34+ lineage (Lin)- cells isolated from normal adult bone marrow (ABM) and mobilized peripheral blood (MPB). Both cell populations were found to differentiate into T cells when injected into human fetal thymi implanted into severe combined immunodeficient mice. Cytokine-MPB cells were less efficient than ABM cells in engrafting in the fetal human thymus, although both gave rise to thymocytes with identical phenotypes based on the analysis of CD1a, CD3, CD4, and CD8 expression. Thymocytes derived from adult CD34+ Lin- cells were capable of fully differentiating into mature CD3+ T cells expressing either the T-cell receptor (TCR) gamma delta or the TCR alpha beta (the later associated with CD4 or CD8), showing that the T-cell progenies of adult CD34+ cells were polyclonal and functional. Our data indicate that human MPB CD34+ cells are qualitatively identical to their BM counterparts, and demonstrate the existence of T-lymphoid progenitor cell activity in MPB.

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Generation of T cells from cytokine-mobilized peripheral blood and adult bone marrow CD34+ cells

Blood ◽

10.1182/blood.v84.1.104.bloodjournal841104 ◽

1994 ◽

Vol 84 (1) ◽

pp. 104-110

Author(s):

AH Galy ◽

S Webb ◽

D Cen ◽

LJ Murray ◽

J Condino ◽

...

Keyword(s):

Bone Marrow ◽

T Cells ◽

T Cell ◽

Peripheral Blood ◽

Cell Activity ◽

Immunodeficient Mice ◽

Adult Bone Marrow ◽

Cd34 Cells ◽

Adult Bone ◽

Mobilized Peripheral Blood

The present study compared the T-cell progenitor content of CD34+ lineage (Lin)- cells isolated from normal adult bone marrow (ABM) and mobilized peripheral blood (MPB). Both cell populations were found to differentiate into T cells when injected into human fetal thymi implanted into severe combined immunodeficient mice. Cytokine-MPB cells were less efficient than ABM cells in engrafting in the fetal human thymus, although both gave rise to thymocytes with identical phenotypes based on the analysis of CD1a, CD3, CD4, and CD8 expression. Thymocytes derived from adult CD34+ Lin- cells were capable of fully differentiating into mature CD3+ T cells expressing either the T-cell receptor (TCR) gamma delta or the TCR alpha beta (the later associated with CD4 or CD8), showing that the T-cell progenies of adult CD34+ cells were polyclonal and functional. Our data indicate that human MPB CD34+ cells are qualitatively identical to their BM counterparts, and demonstrate the existence of T-lymphoid progenitor cell activity in MPB.

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Human fetal liver MSCs are more effective than adult bone marrow MSCs for their immunosuppressive, immunomodulatory and Foxp3+ T regs induction capacity

10.21203/rs.3.rs-40561/v2 ◽

2020 ◽

Author(s):

Yi Yu ◽

Alejandra Vargas Valderrama ◽

Zhongchao Han ◽

Georges Uzan ◽

Sina Naserian ◽

...

Keyword(s):

Bone Marrow ◽

T Cells ◽

T Cell ◽

Immune Responses ◽

Molecular Mechanisms ◽

Fetal Liver ◽

Cell Contact ◽

Adult Bone Marrow ◽

Wide Range ◽

Adult Bone

Abstract Background: Mesenchymal stem cells (MSCs) display active capacities of suppressing or modulating harmful immune responses through diverse molecular mechanisms. These cells are under extensive translational efforts as cell therapies for immune-mediated diseases and transplantations. A wide range of preclinical studies and limited number of clinical trials using MSCs have not only shown promising safety and efficacy profiles but have also revealed changes in regulatory T cell (T reg) frequency and function. However, the mechanisms underlying this important observation are not well understood. Cell-to-cell contact, production of soluble factors, reprogramming of antigen presenting cells to tolerogenic phenotypes have emerged as possible mechanisms by which MSCs produce an immunomodulatory environment for T reg expansion. We and others demonstrated that adult bone-marrow (BM)-MSCs suppress adaptive immune responses directly by inhibiting the proliferation of CD4+ (“helper”) and CD8+ (“cytotoxic”) T cells but also indirectly through induction of Tregs. In parallel we demonstrated that fetal liver (FL)-MSCs displays much longer-lasting immunomodulatory properties compared to BM-MSCs, by inhibiting directly the proliferation and activation of CD4+ and CD8+ T cells. Therefore, we investigated if FL-MSCs exert their strong immunosuppressive effect also indirectly through induction of T regs.Methods: MSCs were obtained from FL and adult BM and characterized according to their surface antigen expression, their multilineage differentiation and their proliferation potential. Using different in-vitro combinations, we performed co-cultures of FL or BM-MSCs and murine CD3+CD25-T cells to investigate immunosuppressive effects of MSCs on T cells and to quantify their capacity to induce functional T regs. Results: We demonstrated that although both types of MSC exhibit similar phenotypic profile and differentiation capacity, FL-MSCs have significantly higher proliferative capacity and ability to suppress both CD4+ and CD8+ murine T cell proliferation and to modulate them towards less active phenotypes than adult BM-MSCs. Moreover, their substantial suppressive effect was associated with an outstanding increase of functional CD4+CD25+Foxp3+ T regs compared to BM-MSCs.Conclusions: These results highlight the immunosuppressive activity of FL-MSCs on T cells and show for the first time that one of the main immunoregulatory mechanisms of FL-MSCs passes through active and functional T reg induction.

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Delineation of T-progenitor cell activity within the CD34+ compartment of adult bone marrow

Blood ◽

10.1182/blood.v85.10.2770.bloodjournal85102770 ◽

1995 ◽

Vol 85 (10) ◽

pp. 2770-2778 ◽

Cited By ~ 35

Author(s):

AH Galy ◽

D Cen ◽

M Travis ◽

S Chen ◽

BP Chen

Keyword(s):

Bone Marrow ◽

T Cell ◽

Progenitor Cells ◽

Progenitor Cell ◽

Cell Activity ◽

Cell Potential ◽

Hematopoietic Stem ◽

Adult Bone Marrow ◽

Hla Dr ◽

Adult Bone

T-cell production is largely dependent on the presence of a thymus gland where CD34+ precursors mature into T lymphocytes. Prethymic stages of T-cell development are less defined. Therefore, this study aims to delineate T-progenitor cell potential within the CD34+ Lineage-- (Lin-) cell compartment of adult bone marrow (ABM). Fractionation of CD34+ Lin-ABM cells with CD45RA, Thy-1, CD38, and HLA-DR failed to absolutely segregate T-cell reconstituting ability, indicating broad distribution of T-progenitor cell potential. Titration experiments showed that low numbers of CD34+ Lin- CD45RA+ (RA+) cells had greater thymus repopulating ability than CD34+ Lin- CD45RA- cells (RA-). The great majority (> 95%) of RA+ cells expressed CD38, HLA-DR and 70% to 90% of RA+ cells lacked Thy-1 surface expression. RA+ cells contained colony-forming unit granulocyte-macrophage (CFU-GM) progenitor cells but were depleted of erythroid potential, did not provide hematopoietic reconstitution of human bone fragments implanted into SCID mice, and did not efficiently maintain CD34+ cells with secondary clonogenic potential in bone marrow cultures. Thus, RA+ cells are oligopotent (nonprimitive) CD34+ progenitors with T-cell reconstituting ability. In contrast, these same assays indicated that CD34+ Lin- CD45RA- cells (RA- cells) comprised hematopoietic stem cells (HSC) with primitive multilineage (T, B, myeloid, and erythroid) hematopoietic potential. It was confirmed that HSC-containing populations, such as CD34+ Lin- CD45RA- Thy-1+ cells had thymus repopulating ability. Culture of RA-cells on murine bone marrow stromal cells in the presence of interleukin (IL)-3, IL-6, and leukemia inhibitory factor (LIF) generated CD34+ CD45RA+ progeny engrafting in a secondary severe combined immunodeficiency (SCID)-hu thymus assay. Altogether, our results underscore the fact that T-cell reconstituting potential can be dissociated from HSC activity. Furthermore, we speculate that HSC might develop into the T lineage indirectly, via differentiation into an intermediate oligopotent CD34+ CD45RA+ stage. Finally, T-progenitor cells can be cultured in vitro.

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Flow Cytometric Detection of the Expression of CXCR4 on CD34+ Cells and the Distribution of Lymphocyte Subsets in Allogeneic Hematological Grafts.

Blood ◽

10.1182/blood.v106.11.5195.5195 ◽

2005 ◽

Vol 106 (11) ◽

pp. 5195-5195

Author(s):

Lulu Lu ◽

Yongping Song ◽

Baogen Ma ◽

Xiongpeng Zhu ◽

Xudong Wei ◽

...

Keyword(s):

Bone Marrow ◽

Flow Cytometry ◽

T Cells ◽

Cord Blood ◽

Peripheral Blood ◽

Lymphocyte Subsets ◽

Color Flow ◽

Hematopoietic Stem ◽

Cd34 Cells ◽

Mobilized Peripheral Blood

Abstract Background and objectives: Normal human bone marrow (BM), cord blood (CB) and mobilized peripheral blood (MPB) are the most commonly used sources for allogeneic hematopoietic stem cell transplantation (HSCT). The aim of this study was to detect the expression of CXCR4 on CD34+ cells and to assess the distribution of lymphocyte subsets in each type allograft. Methods: CD34+ cells were separated from BM (n=30), CB (n=30) and MPB (n=30) by the CD34 MultiSort Kit immunomagnetic bead system. The expression of CXCR4 on CD34+cells was assayed by double color flow cytometry. The lymphocyte subsets in each type of allograft were detected by three-color flow cytometry. The groups of monoclonal antibodies were used as the following: CXCR4-PE/CD34−Pecy5, CD8−FITC/CD4−R-PE/CD3−TC, CD45RA-FITC/CD45RO-PE/CD4−Pecy5, CD45RA-FITC/CD45RO-PE/CD8−Pecy5, and CD3−FITC/CD16+56-PE. Isotype-specific antibodies were used as controls. Results: The expression of CXCR4 of cord blood and mobilized peripheral blood CD34+ cells was lower than that of bone marrow cells (BM 40.21%±6.72%, CB 20.93%±3.96%, MPB 20.93%±3.96%, P <0.05). The difference between cord blood and mobilized peripheral blood was not significant (P>0.05). The CD3+CD8low and CD3+CD4−CD8low subsets were higher in BM than that of CB and MPB (BM 8.61%±1.40%, CB 3.31%±0.88%, MPB 5.11%±0.76%,P<0.01). The relative frequencies of the naïve CD45RA+ CD45RO− phenotype among CD4+ and CD8high T cells were highest in CB, and it was higher in MPB than in BM grafts (BM 28.09%±4.52%, 41.86 %±3.31%; CB83.83%±12.24%, 86.69%±6.12%; MPB 43.58%±4.54%, 57.64%±4.77%, P<0.01). Naïve T cells (CD45RA+ CD45RO−) were mobilized preferentially compared to memory T cells (CD45RA− CD45RO+)(P <0.01); The relative frequencies of NKT (CD3+CD16+56+) among lymphocytes were lower in CB than that in BM and MPB (CB 0.77±0.19, BM4.15±1.10, MPB 4.13±0.84, P<0.01). Conclusion: BM, CB and MPB allografts differ widely in cellular makeup of CD34+ cells and lymphocyte subsets, which are associated with the distinct characteristics after allogeneic HSCT from different allogeneic hematological sources.

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Human Fetal Liver MSCs are More Effective Than Adult Bone Marrow MSCs for Their Immunosuppressive, Immunomodulatory and Foxp3+ T regs Induction Capacity

10.21203/rs.3.rs-40561/v1 ◽

2020 ◽

Author(s):

Yi Yu ◽

Alejandra Vargas Valderrama ◽

Zhongchao Han ◽

Georges Uzan ◽

Sina Naserian ◽

...

Keyword(s):

Bone Marrow ◽

T Cells ◽

T Cell ◽

Immune Responses ◽

Molecular Mechanisms ◽

Fetal Liver ◽

Cell Contact ◽

Adult Bone Marrow ◽

Wide Range ◽

Adult Bone

Abstract Background: Mesenchymal stem cells (MSCs) display active capacities of suppressing or modulating harmful immune responses through diverse molecular mechanisms. These cells are under extensive translational efforts as cell therapies for immune-mediated diseases and transplantations. A wide range of preclinical studies and limited number of clinical trials using MSCs have not only shown promising safety and efficacy profiles but have also revealed changes in regulatory T cell (T reg) frequency and function. However, the mechanisms underlying this important observation are not well understood. Cell-to-cell contact, production of soluble factors, reprogramming of antigen presenting cells to tolerogenic phenotypes have emerged as possible mechanisms by which MSCs produce an immunomodulatory environment for T reg expansion. We and others demonstrated that adult bone-marrow (BM)-MSCs suppress adaptive immune responses directly by inhibiting the proliferation of CD4+ (“helper”) and CD8+ (“cytotoxic”) T cells but also indirectly through induction of Tregs. In parallel we demonstrated that fetal liver (FL)-MSCs displays much longer-lasting immunomodulatory properties compared to BM-MSCs, by inhibiting directly the proliferation and activation of CD4+ and CD8+ T cells. Therefore, we investigated if FL-MSCs exert their strong immunosuppressive effect also indirectly through induction of T regs.Methods: MSCs were obtained from FL and adult BM and characterized according to their surface antigen expression, their multilineage differentiation and their proliferation potential. Using different in-vitro combinations, we performed co-cultures of FL or BM-MSCs and murine CD3+CD25-T cells to investigate immunosuppressive effects of MSCs on T cells and to quantify their capacity to induce functional T regs. Results: We demonstrated that although both types of MSC exhibit similar phenotype profile and differentiation capacity, FL-MSCs have significantly higher proliferative capacity and ability to suppress both CD4+ and CD8+ murine T cell proliferation and to modulate them towards less active phenotypes than adult BM-MSCs. Moreover, their substantial suppressive effect was associated with an outstanding increase of functional CD4+CD25+Foxp3+ T regs compared to BM-MSCs.Conclusions: These results highlight the immunosuppressive activity of FL-MSCs on T cells and show for the first time that one of the main immunoregulatory mechanisms of FL-MSCs passes through active and functional T reg induction.

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T Cell-Depleted Related Haploidentical Peripheral Blood Stem Cell Transplantation in a Patient with Fanconi Anemia. Cagliari Experience.

Blood ◽

10.1182/blood.v104.11.5175.5175 ◽

2004 ◽

Vol 104 (11) ◽

pp. 5175-5175

Author(s):

Maria Adele Sanna ◽

Maria Grazia Orofino ◽

Fausto Cossu ◽

Maria Carmen Addari ◽

Antonio Piroddi ◽

...

Keyword(s):

Bone Marrow ◽

T Cells ◽

Stem Cell ◽

T Cell ◽

Stem Cell Transplantation ◽

Cell Transplantation ◽

Peripheral Blood ◽

Bone Marrow Failure ◽

Unrelated Donor ◽

Cd34 Cells

Abstract Stem cell transplantation is presently the best treatment for Fanconi Anaemia (FA) patients developing bone marrow failure. 70% of success is reported in patients with a HLA identical sibling whereas the outcome for HSCT in those transplanted from unrelated donors is in the range of 29–43%, graft rejection, GVHD and regimen related toxicity beeing the main causes of failure. This results limited the ability to perform marrow transplantation other than HLA identical siblings for this disease. Recently a fludarabine based cytoreductive regimen has been successfully used in T cell depleted haploidentical/mismatched transplant of FA patients. We report a case of a 7 year old boy with bone marrow failure since 1999. Androgens treatment was uneffective, no HLA identical family donor was available and the search for a suitable marrow or cord blood unrelated donor was unsuccessful. After 4 years he underwent T-cell depleted haploidentical PBSCT from his father. Conditioning regimen was: fludarabine 30 mg/mq from day −6 to day −3, cytoxan 300 mg/mq from day −6 to day −3, rabbit ATG (3.75 mg/kg) from day −5 to day −3. GvHD prophylaxis consisted of cyclosporine 1 mg/kg from day −1. The donor received G-CSF 8 ug/kg/dose twice daily for 6 days and underwent leukapheresis on day 5 and 6. Donor stem cells were depleted of T cells by positive selection of CD34+ cells using the Clinimacs device according to the suggested procedures (Milteny Biotec). On day 0, 15.3x106 x kg CD34+ cells were infused with 1.5 x 105 CD3 + cells. The clinical postransplant course was uneventful. Neutrophil engraftment ( >0.5 x 109 ) occurred on day 14, platelet count >100x109 on day 15. He was discharged on day 39 without signs of GVHD. Molecular analysis of DNA-VNTRs at 1, 3, 6, 9, 12 months showed >95% donor chimerism on peripheral blood. At 14 months after transplantation the patient is well, normal blood cell count (WBC 5.4 x 109/l, Hb 13.6 gr /dl, platelets 293x 109 /l). Count of T-cells are reported in the normal reference range ( CD3+ :1865 ug/l, CD8+ :1026ug/l, CD19+ :732ug/l, CD56+: 452). Karnofsky score is 100%. Conclusion: the case reported shows that the fludarabine based regimen and the infusion of a high number of T-cell depleted CD34+ was successful in absence of peri-transplant complications and can be proposed for the cure of FA patients at high risk of clonal disease and without HLA-matched sibling donor.

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A multifactorial analysis of umbilical cord blood, adult bone marrow and mobilized peripheral blood progenitors using the improved ML-IC assay

Experimental Hematology ◽

10.1016/j.exphem.2004.10.016 ◽

2005 ◽

Vol 33 (2) ◽

pp. 165-172 ◽

Cited By ~ 27

Author(s):

Koen Theunissen ◽

Catherine M. Verfaillie

Keyword(s):

Bone Marrow ◽

Cord Blood ◽

Umbilical Cord ◽

Umbilical Cord Blood ◽

Peripheral Blood ◽

Multifactorial Analysis ◽

Adult Bone Marrow ◽

Adult Bone ◽

Mobilized Peripheral Blood

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Human fetal liver MSCs are more effective than adult bone marrow MSCs for their immunosuppressive, immunomodulatory, and Foxp3+ T reg induction capacity

Stem Cell Research & Therapy ◽

10.1186/s13287-021-02176-1 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Yi Yu ◽

Alejandra Vargas Valderrama ◽

Zhongchao Han ◽

Georges Uzan ◽

Sina Naserian ◽

...

Keyword(s):

Bone Marrow ◽

T Cells ◽

Immune Responses ◽

Molecular Mechanisms ◽

Fetal Liver ◽

Cytotoxic T Cells ◽

Cell Contact ◽

Adult Bone Marrow ◽

Adult Bone

Abstract Background Mesenchymal stem cells (MSCs) exhibit active abilities to suppress or modulate deleterious immune responses by various molecular mechanisms. These cells are the subject of major translational efforts as cellular therapies for immune-related diseases and transplantations. Plenty of preclinical studies and clinical trials employing MSCs have shown promising safety and efficacy outcomes and also shed light on the modifications in the frequency and function of regulatory T cells (T regs). Nevertheless, the mechanisms underlying these observations are not well known. Direct cell contact, soluble factor production, and turning antigen-presenting cells into tolerogenic phenotypes, have been proposed to be among possible mechanisms by which MSCs produce an immunomodulatory environment for T reg expansion and activity. We and others demonstrated that adult bone marrow (BM)-MSCs suppress adaptive immune responses directly by inhibiting the proliferation of CD4+ helper and CD8+ cytotoxic T cells but also indirectly through the induction of T regs. In parallel, we demonstrated that fetal liver (FL)-MSCs demonstrates much longer-lasting immunomodulatory properties compared to BM-MSCs, by inhibiting directly the proliferation and activation of CD4+ and CD8+ T cells. Therefore, we investigated if FL-MSCs exert their strong immunosuppressive effect also indirectly through induction of T regs. Methods MSCs were obtained from FL and adult BM and characterized according to their surface antigen expression, their multilineage differentiation, and their proliferation potential. Using different in vitro combinations, we performed co-cultures of FL- or BM-MSCs and murine CD3+CD25−T cells to investigate immunosuppressive effects of MSCs on T cells and to quantify their capacity to induce functional T regs. Results We demonstrated that although both types of MSC display similar cell surface phenotypic profile and differentiation capacity, FL-MSCs have significantly higher proliferative capacity and ability to suppress both CD4+ and CD8+ murine T cell proliferation and to modulate them towards less active phenotypes than adult BM-MSCs. Moreover, their substantial suppressive effect was associated with an outstanding increase of functional CD4+CD25+Foxp3+ T regs compared to BM-MSCs. Conclusions These results highlight the immunosuppressive activity of FL-MSCs on T cells and show for the first time that one of the main immunoregulatory mechanisms of FL-MSCs passes through active and functional T reg induction.

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A comparison of murine T-cell–depleted adult bone marrow and full-term fetal blood cells in hematopoietic engraftment and immune reconstitution

Blood ◽

10.1182/blood.v99.1.364 ◽

2002 ◽

Vol 99 (1) ◽

pp. 364-371 ◽

Cited By ~ 16

Author(s):

Benny J. Chen ◽

Xiuyu Cui ◽

Gregory D. Sempowski ◽

Maria E. Gooding ◽

Congxiao Liu ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

T Cells ◽

T Cell ◽

Cord Blood ◽

Blood Cells ◽

Immune Reconstitution ◽

Full Term ◽

Fetal Blood ◽

Adult Bone

Umbilical cord blood has been increasingly used as a source of hematopoietic stem cells. A major area of concern for the use of cord blood transplantation is the delay in myeloid and lymphoid recovery. To directly compare myeloid and lymphoid recovery using an animal model of bone marrow and cord blood as sources of stem cells, hematopoietic engraftment and immune recovery were studied following infusion of T-cell–depleted adult bone marrow or full-term fetal blood cells, as a model of cord blood in a murine allogeneic transplantation model (C57BL/6 [H-2b] → BALB/c [H-2d]). Allogeneic full-term fetal blood has poorer radioprotective capacity but greater long-term engraftment potential on a cell-to-cell basis compared with T-cell–depleted bone marrow. Allogeneic full-term fetal blood recipients had decreased absolute numbers of T, B, and dendritic cells compared with bone marrow recipients. Splenic T cells in allogeneic full-term fetal blood recipients proliferated poorly, were unable to generate cytotoxic effectors against third-party alloantigens in vitro, and failed to generate alloantigen-specific cytotoxic antibodies in vivo. In addition, reconstituting T cells in fetal blood recipients had decreased mouse T-cell receptorδ single-joint excision circles compared with bone marrow recipients. At a per-cell level, B cells from fetal blood recipients did not proliferate as well as those found in bone marrow recipients. These results suggest that full-term fetal blood can engraft allogeneic hosts across the major histocompatibility barrier with slower hematopoietic engraftment and impaired immune reconstitution.

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