Quantitative assays are needed to characterize the multilineage engraftment potential of clinical hematopoietic grafts. After we observed a dose-response relationship between the number of human hematopoietic cells transplanted into nonobese diabetic-scid/scid (NOD/SCID) mice and the number of human CD45+ cells recovered in the chimeras’ marrows and spleens, we sought to develop a multiple linear regression model that allows quantitative comparisons of human cell engraftment in vivo.
Methods
We used this NOD/SCID xenotransplant model to compare the engraftment potential of cord blood vs. adult marrow or mobilized blood, after either of 2 commonly used clinical graft engineering procedures: CD34+ cell selection or T cell depletion.
Results
The engraftment per transplanted cell was >20 fold higher for cord blood cells, as compared to hematopoietic cells from adults. However, there was no difference in engraftment per CD34+ cell transplanted between marrow and mobilized blood. Levels of human cell engraftment from all sources could be increased by administration of human hematopoietic growth factors to human/mouse chimeras after transplantation. Correlation analysis of the number of human CD13+ myeloid cells and CD19+ B lymphoid cells in the chimeras’ marrows 8 weeks after transplantation provided evidence that almost all the human myeloid and B lymphoid cells were derived from the same primitive precursor cells.
Conclusions
These findings and assay may be useful in the development of clinical hematopoietic cell therapies.
AppelbaumFR. Allogeneic stem cells transplantation for the treatment of hematologic malignancies: current indications and challenges. J. Rheumatol1997;48:41–5.
2.
TerritoM.The use of autologous transplantation in the treatment of malignant disorders. J. Rheumatol1997;48:36–40.
3.
StorbR.Hematopoietic stem cell transplantation in nonmalignant diseases. J Rheumatol1997;48:30–5.
4.
FlakeAW, ZanjaniED. In utero hematopoietic stem cell transplantation. JAMA1997;278:932–7.
5.
CollinsN.A, GeeA, RowleyS, ShpallE.Are we ready for regulation? ISHAGE Regulatory Action Group. International Society for Hematotherapy and Graft Engineering. J. Hematother1995;4:1–2.
6.
SutherlandHJ, HoggeDE, PhillipsGL PML, EavesAC, EavesCJ. Quantitation, mobilization, and clinical use of long-term culture-initiating cells in blood cell autografts. J Hematother1995;4:3–10.
7.
PflumioF, IzacB, KatzA, ShultzLD, VainchenkerW, CoulombelL.Phenotype and function of human hematopoietic cells engrafting immune-deficient CB17-severe combined immunodeficiency mice and nonobese diabetic-severe combined immunodeficiency mice after transplantation of human cord blood mononuclear cells. Blood1996;88:3731–40.
8.
WangJCY, DoedensM, DickJE. Primitive human hematopoietic cells are enriched in cord blood compared with adult bone marrow or mobilized peripheral blood as measured by the quantitative in vivo SCID-repopulating cell assay. Blood1997;89:3919–24.
9.
ConneallyE, CashmanJ, PetzerA, EavesC.Expansion in vitro of transplantable human cord blood stem cells demonstrated using a quantitative assay of their lymphoid-myeloid repopulating activity in nonobese diabetic-scid/scid mice. Proc Natl Acad Sci1997;94:9836–41.
10.
LeungW, ChenAR, CivinCI. Use of purified stem and progenitor cells for transplantation. In: AtkinsonK, ed. Clinical Bone Marrow Transplantation. Cambridge: Cambridge University Press, 1998. In press.
11.
MartinPJ, HorowitzMM. T cell depletion of donor marrow. In: AtkinsonK, ed. Clinical Bone Marrow Transplantation. Cambridge: Cambridge University Press, 1994;653–7.
12.
PiantadosiS.Factorial designs. In: PiantadosiS, ed. Clinical trials: a methodologic approach. New York: John Wiley & Sons, 1997;385–400.
13.
GluckmanE, RochaV, Boyer-ChammardA, LocatelliF, ArceseW, PasquiniR, OrtegaJ, SouilletG, FerreiraE, LaporteJ, FernandezM, ChastangC.Outcome of cord blood transplantation from related and unrelated donors. N Engl J Med1997;337:373–81.
14.
MavroudisD, ReadE, Cottler-FoxM, CourielD, MolldremJ, CarterC, YuM, DunbarC, BarrettJ.CD34+ cell dose predicts survival, posttransplant morbidity, and rate of hematologic recovery after allogeneic marrow transplants for hematologic malignancies. Blood1996;88:3223–9.
15.
KorblingM, HuhYO, DurettA, MirezaN, MillerP, EngelH, AnderliniP, Van BesienK, AndreeffM, PrzepiorkaD, DeisserothAB, ChamplinRE. Allogeneic blood stem cell transplantation: peripheralization and yield of donor-derived primitive hematopoietic progenitor cells (CD34+ Thy-1 dim) and lymphoid subsets, and possible predictors of engraftments and graft-versus-host disease. Blood1995;86:2842–8.
16.
HulleySB, CummingsSR. Total sample size required when using the correlation coefficient. In: HulleySB, CummingsSR, eds. Designing Clinical Research. Baltimore: Williams & Wilkins, 1988;218.
17.
ShultzLD, SchweitzerPA, ChristiansonSW, GottB, SchweitzerIB, TennentBT, McKennaS, MobraatenL, RajanTV, GreinerDL, LeiterEH. Multiple defects in innate and adaptive immunologic function in NOD/LtSz-scid mice. J Immunol1995;154:180–91.
18.
CohenJ.Statistical power analysis for the behavioral sciences. Hillsdale, NJ. 1988.
19.
Passos-CoelhoJL, RossAA, MossTJ, DavisJM, HuelskampA, NogaST, DavidsonNE, KennedyMJ. Absence of breast cancer cells in single-day peripheral blood progenitor cell collection after priming with Cyclophosphamide and granulocyte-macrophage colony-stimulating factor. Blood1995;85:1138.
20.
CivinCI, Almeida-PoradaG, LeeMJ, OlweusJ, TerstappenLWMM, ZanjaniED. Sustained, retransplantable, multilineage engraftment of highly purified adult human bone marrow stem cells in vivo. Blood1996;88:4102–9.
21.
RamirezM, RottmanGA, ShultzLD, CivinCI. Mature human hematopoietic cells in donor bone marrow complicate interpretation of stem/progenitor cell assays in xenogeneic hematopoietic chimeras. Exp Hematol1998;26:332–44.
22.
BoggsDR. The total marrow mass of the mouse: a simplified method of measurement. Am J Hematol1984;16:277–86.
23.
TrischmannTM, SchepersKG, CivinCI. Measurement of CD34+ cells in bone marrow by flow cytometry. J Hematother1993;2:305–13.
24.
CarowCE, HarringtonMA, BroxmeyerHE. Measurement of CD34+ cells in bone marrow by flow cytometry. J. Hematother1993;2:305–13.
25.
HockingRR. The analysis and selection of variables in linear regression. Biometrics1976;32:1–50.
26.
DraperNR, SmithH.Fitting a straight line by least squares. In: DraperNR, SmithH, eds. Applies regression analysis. New York: John Wiley & Sons, 1981;1–69.
27.
EverittBS. Repeated measures analysis of variance and the analysis of covariance. In: EverittB, ed. Statistical methods for medical investigation. New York: Oxford University Press, 1989;57–67.
28.
WaldA.The fitting of straight lines if both variables are subject to error. Ann Math Statist1940;1:284–300.
29.
RamseyJB. Tests for specification errors in classical linear least squares regression analysis. JR Statist Soc Series1969;31:350–71.
30.
CookRD, WeisbergS.Diagnostics for heteroscedasticity in regression. Biometrika1983;70:1–10.
31.
WilcoxonF.Individual comparisons by ranking methods. Biometrics1945;1:80–3.
32.
SpearmanC.The proof and measurement of association between two things. Am J Psychol1904;15:72–101.
33.
HarrisonDE, CraigTJ, ZhongRK, ClintonMA. Primitive hematopoietic stem cells: direct assay of most productive populations by competitive repopulation with simple binomial, correlation and covariance calculations. Exp Hemato.1993;21:206–19.
34.
AtkinsonK.Hematopoietic reconstitution posttransplant. In: AtkinsonK, eds. Clinical Bone Marrow Transplantation. Cambridge: Cambridge University Press, 1994;31–41.
35.
WeisdorfDJ, VerfaillieCM, DaviesSM, FilipovichAH, WagnerJEJr, MillerJS, BurroughsJ, RamsayNK, KerseyJH, McGlavePB. Hematopoietic growth factors for graft failure after bone marrow transplantation: a randomized trial of granulocyte-marcrophage colony-stimulating factor (GM-CSF) versus sequential GM-CSF plus granulocyte-CSF. Blood1995;85:3452–6.
36.
SchueningF, BeanMA, DeegHJ, StorbR.Prevention of graft failure in patients with aplastic anemia. Bone Marrow Transplant1993;12:548–9.
37.
Henslee-DowneyPJ. Mismatched bone marrow transplantation. Curr Opin Oncol1995;7:115–11.
38.
NogaSJ, VogelsangGB, SeberA, DavisJM, SchepersK, HessAD, JonesRJ. CD34+ stem cell augmentation of allogeneic, elutriated marrow grafts improves engraftment but cyclosporing A is still required to reduce GVHD and morbidity. Transplant Proc. 1997;29:728–32.
39.
BrennerMK. Gene transfer to hematopoietic cells. N Engl J Med1996;335:337–9.
40.
BambachBJ, MoserHW, BlakemoreK, CorsonVL, GriffinCA, NogaSJ, PerlmanEJ, ZuckermanR, WengerDA, JonesRJ. Engraftment following in utero bone marrow transplantation for globoid cell leukodystrophy. Bone Marrow Transplant.1997;19:399–402.
41.
OrlicD, BodineDM. What defines a pluripotent hematopoietic stem cell (PHSC): will the real PHSC please stand up. Blood1994;84:3991–4.
42.
MorrisonSJ, ShahNM, AndersonDJ. Regulatory mechanisms in stem cell biology. Cell1997;88:287–98.
43.
MorrisonSJ, WandyczAM, AkashiK, GloversonA, WeissmanIL. The aging of hematopoietic stem cells. Nature Med1996;2:202–6.
44.
MorrisonSJ, ProwseKR, HoP, WeissmanIL. Telomerase activity in hematopoietic cells is associated with self-renewal potential. Immunity1996;5:207–16.
45.
MayaniH, LansdorpPM. Thy-1 expression is linked to functional properties of primitive hematopoietic progenitor cells from human umbilical cord blood. Blood1994;83:2410–7.
46.
VaziriH, DragowskaW, AllsoppRC, ThomasTE, HarleyCB, LansdorpPM. Evidence for a mitotic clock in human hematopoietic stem cells: loss of telomeric DNA with age. Proc Natl Acad Sci1994:91:9857–60.
47.
VormoorJ, LapidotT, PflumioF, RisdonG, PattersonB, BroxmeyerHE, DickJE. Immature human cord blood progenitors engraft and proliferate to high levels in severe combined immunodeficient mice. Blood1994;83:2489–97.
48.
HoganCJ, ShpallEJ, McNultyO, McNieceI, DickJE, ShultzLD, KellerG.Engraftment and development of human CD34+-enriched cells from umbilical cord blood in NOD/LtSz-scid/scid mice. Blood1997;90:85–96.
49.
NishiN, IshikawaR, InoueH, NishikawaM, KakedaM, YoneyaT, TsumuraH, OhashiH, YamaguchiY, MotokiK, SudoT, MoriKJ. Granulocyte-colony stimualting factor and stem cell factor are the crucial factors in long-term culture of human primitive hematopoietic cells supported by a murine stromal cell line. Exp Hematol1996;24:1312–21.
50.
Dawson-SaundersB, TrappRG. Association and prediction. In: Dawson-SaundersB, TrappRG, eds. Basic and clinical biostatistics. Norwalk, Connecticut: Appleton & Lange. 1994;164–5.
