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The development of the gastrointestinal tract in newborn calves is essential for sufficient nutrient uptake. An intensive milk feeding during the neonatal period may impair the rumen development in calves. The aim of this study was to investigate effects of milk replacer (MR) feeding in unlimited amounts for the first 5 wk of age on the gastrointestinal growth and development in preruminant calves at wk 9 of age. Twenty-eight newborn Holstein and Holstein × Charolais crossbred calves (19 male and 9 female) were fed MR ad libitum (ADLIB) or in restricted amounts (6 L per day; RES) until wk 5 of age. Thereafter, the MR intake of ADLIB was gradually reduced at wk 6 and 7, and all calves received 6 L of MR per day until wk 9 of age. In wk 9, calves were slaughtered and carcass and organ weight as well as rumen papilla size in the atrium, ventral sac, and ventral blind sac, and villus size of the mucosa in the small intestine (duodenum; proximal, mid, and distal jejunum; and ileum) were determined. The expression of mRNA associated with the local insulin-like growth factor (IGF) system was measured in the rumen epithelium. Ad libitum MR feeding increased MR intake and growth in ADLIB without influencing concentrate intake compared with RES. Carcass weight in wk 9 was greater in ADLIB than in RES. The density of the rumen papillae in the atrium and ventral blind sac was greater in RES than in ADLIB calves, but surface area of the epithelium was not different between groups in the investigated regions of the rumen. The mRNA abundance of IGF1 in the atrium tended to be greater and the IGFR1 mRNA abundance in the ventral sac tended to be lower in the ADLIB than in the RES feeding group. The rumen pH and volatile fatty acid concentrations were not affected by MR feeding intensity. In mid-jejunum, villus circumference was greater in ADLIB than in RES calves. In the distal jejunum, villus surface area and the villus height/crypt depth ratio were greater and the villus circumference and height tended to be greater, whereas crypt depth was smaller in ADLIB than in RES calves. The findings from this study indicate that ad libitum MR feeding for 5 wk of age followed by its gradual reduction promotes growth performance without any negative influence on gastrointestinal growth and development in dairy calves at 9 wk of age.

Scientific proof for flavonoids as a health tool in calf nutrition is inconsistent. We investigated the effects of the most abundant flavonoid, quercetin, and of a green tea extract (GTE) containing various catechins on the metabolic and antioxidative traits in dairy calves to clarify their potential health-promoting effects. Male newborn German Holstein calves (n=7 per group) received either no flavonoid (control group), 10mg of quercetin equivalents as quercetin aglycone or as rutin/kg of body weight (BW) per day, or 10mg/kg of BW per day of a GTE from d 2 to 26 of life. The supplements were provided with the morning and evening feeding. The calves were fed colostrum and milk replacer, and BW, feed intake, and health status were evaluated daily. Blood samples were collected from a jugular vein on d 1, 5, 12, 19, and 26 before the morning feeding to investigate the metabolic and antioxidative status of the calves. The growth performance and health status remained unchanged, but the GTE-fed calves had fewer loose feces than the controls. The plasma concentrations of quercetin changed over time and were higher in the rutin-fed group than in the control group, whereas the catechins were below the detection limit. The plasma Trolox equivalent antioxidative capacity and ferric reducing ability of plasma were measured as markers for plasma antioxidative capacity. The concentrations of Trolox equivalent antioxidative capacity increased, whereas ferric reducing ability of plasma decreased after the first day of life in all the groups. The oxidative stress markers in the plasma were measured as thiobarbituric acid reactive substances and F2-isoprostanes, but these did not indicate treatment or time effects. The plasma concentrations of total protein, albumin, urea, lactate, glucose, and nonesterified fatty acids and of insulin and cortisol varied over time, but no group differences were caused by the flavonoid supplementation. In summary, orally administered quercetin and catechins at the dosages used in the present study resulted in weak effects on health and no effects on the metabolic and antioxidative status of newborn dairy calves.

Polyphenols, such as flavonoids, are secondary plant metabolites with potentially health-promoting properties. In newborn calves flavonoids may improve health status, but little is known about the systemically availability of flavonoids in calves to exert biological effects. The aim of this study was to investigate the oral bioavailability of the flavonol quercetin, applied either as quercetin aglycone (QA) or as its glucorhamnoside rutin (RU), in newborn dairy calves. Twenty-one male newborn German Holstein calves were fed equal amounts of colostrum and milk replacer according to body weight. On d 2 and 29 of life, 9 mg of quercetin equivalents/kg of body weight, either fed as QA or as RU, or no quercetin (control group) were fed together with the morning meal. Blood samples were taken before and 0.5, 1, 1.5, 2, 2.5, 3, 4, 5, 6, 12, 24, and 48 h after feed intake. Quercetin and quercetin metabolites with an intact flavonol structure (isorhamnetin, tamarixetin, and kaempferol) were analyzed in blood plasma after treatment with glucuronidase or sulfatase by HPLC with fluorescence detection. Maximum individual plasma concentration was depicted from the concentration-time-curve on d 2 and 29, respectively. Additional blood samples were taken to measure basal plasma concentrations of total protein, albumin, urea, and lactate as well as pre- and postprandial plasma concentrations of glucose, nonesterified fatty acids, insulin, and cortisol. Plasma concentrations of quercetin and its metabolites were significantly higher on d 2 than on d 29 of life, and administration of QA resulted in higher plasma concentrations of quercetin and its metabolites than RU. The relative bioavailability of total flavonols (sum of quercetin and its metabolites isorhamnetin, tamarixetin, and kaempferol) from RU was 72.5% on d 2 and 49.6% on d 29 when compared with QA (100%). Calves fed QA reached maximum plasma concentrations of total flavonols much earlier than did RU-fed calves. Plasma metabolites and hormones were barely affected by QA and RU feeding in this experiment. Taken together, orally administrated QA resulted in a greater bioavailability of quercetin than RU on d 2 and 29, respectively, and quercetin bioavailability of quercetin and its metabolites differed markedly between calves aged 2 and 29 d.

A dog model of stable mixed hematopoietic chimerism was established in which leukocyte-antigen-identical littermates receive nonmyeloablative total body irradiation before hematopoietic cell transplantation and postgrafting immunosuppression with mycophenolate mofetil and cyclosporine. Unmodified donor lymphocyte infusion (DLI) into stable mixed chimeras failed to increase donor chimerism, while DLI from donors sensitized to recipient minor-histocompatibility antigens promptly converted all recipients to complete donor chimerism. This established a model for studying approaches to enhance the graft-versus-host (GVH)-effect, a potential surrogate for graft-versus-leukemia activity. We asked if interleukin-2 (IL-2) given after unmodified DLI could result in reliable conversion to complete donor chimerism. IL-2, 4 x 10(5) IU/kg/day, was administered to six mixed chimeric dogs for 14 days. Four dogs received unmodified DLI with IL-2. At 20-40 weeks after DLI, all dogs remained mixed chimeras. For the two recipients of IL-2 only, mixed chimerism also remained unchanged. These results show that IL-2 given with DLI after nonmyeloablative transplantation in dogs is not effective in reliably converting mixed to complete donor chimerism.

Summary. We hypothesized that patients undergoing major ABO-incompatible non-myeloablative haematopoietic stem cell transplantation (nm-HSCT) might experience prolonged haemolysis after transplant due to the delayed disappearance of host plasma cells producing anti-donor isohaemagglutinins (HAs). To address this question, we analysed data from 107 consecutive patients transplanted with allogeneic peripheral blood stem cells from human leucocyte antigen-matched (related, n = 84; unrelated, n = 23) donors after non-myeloablative conditioning (200 cGy total body irradiation ± fludarabine). In total, 23 out of the 107 patients received major or major/minor ABO-incompatible transplants. Red blood cell (RBC) transfusion requirements during the first 120 d post transplant were higher in major ABO-mismatched than in ABO-matched recipients (0·12 vs 0·03 median units RBC concentrate/d, P = 0·04). Two patients developed transient pure red cell aplasia, which had resolved spontaneously by 9 months after transplant. Major ABO incompatibility did not influence rates of engraftment. Patients with sustained engraftment experienced gradual declines of anti-donor HAs, and the estimated median time to reaching IgM and IgG titres of

Severe hemolytic anemia in Basenji dogs secondary to pyruvate kinase (PK) deficiency can be corrected by marrow allografts from healthy littermates after a conventional high-dose myeloablative conditioning regimen. The nonmyeloablative conditioning regimen used here, which consisted of a sublethal dose of 200 cGy total body irradiation before and immunosuppression with mycophenolate mofetil and cyclosporine after a dog leukocyte antigen (DLA)-identical littermate allograft, has been found to be effective in establishing stable mixed donor/host hematopoietic chimerism in normal dogs. We explored the feasibility of nonmyeloablative marrow allografts for the treatment of canine PK deficiency and studied the effect of stable allogeneic mixed hematopoietic chimerism on the natural course of the disease. Five affected dogs received transplants, of which 3 dogs had advanced liver cirrhosis and myelofibrosis. Both complications were presumed to be due to iron overload. All 5 dogs showed initial engraftment. Two rejected their grafts after 6 weeks but survived with completeautologous marrow recovery and return of the disease. One died from liver failure on day 27 with 60% donor engraftment. Two dogs have shown sustained mixed donor/host chimerism for more than a year with 85% and 12% donor hematopoietic cells, respectively. Overall clinical response correlated with the degree of donor chimerism. The dog with the low degree of chimerism achieved partial resolution of hemolysis, but the disease symptoms persisted as manifested by increasing iron overload resulting in progression of marrow and liver fibrosis. The dog with the high degree of donor chimerism achieved almost complete resolution of hemolysis with a decrease of marrow iron content and resolution of marrow fibrosis. These observations suggest that mixed hematopoietic chimerism can be relatively safely established in dogs with PK deficiency even in the presence of advanced liver cirrhosis. However, although effective in correcting or delaying the development of myelofibrosis, a low degree of mixed chimerism was not sufficient to prevent continued hemolysis of red blood cells of host origin. Complete donor chimerism appears necessary to achieve a long-term cure. Biol Blood Marrow Transplant 2001;7(1):14-24.

Stable mixed chimerism can be established in dogs given a sublethal dose of 200 cGy total body irradiation (TBI) before and immunosuppression with mycophenolate mofetil (MMF) and cyclosporine (CSP) for 28 and 35 days, respectively, after dog leukocyte antigen-identical marrow transplantation. Most likely, the role of pretransplant TBI was to provide host immunosuppression, since stable mixed chimerism was also achieved in MMF/CSP-treated dogs when 450 cGy irradiation, targeted to cervical, thoracic, and upper abdominal lymph nodes, was substituted for TBI. When TBI was reduced from 200 to 100 cGy, all grafts were rejected within 3 to 12 weeks. Here, we asked whether stable engraftment after 100 cGy TBI could be accomplished by first reducing the intensity of host immune responsiveness with help of the fusion peptide CTLA4Ig, which blocks T-cell costimulation through the B7-CD28 signal pathway. Accordingly, recipient T cells were activated with intravenous (IV) injections of 106 donor peripheral blood mononuclear cells (PBMC)/kg per day on days −7 to −1 before 100 cGy TBI, with concurrent administration of CTLA4Ig 4 mg/kg/d IV. All 7 dogs so treated showed initial mixed chimerism. Two rejected their allografts after 8 and 20 weeks, respectively, and survived with autologous marrow recovery; 1 mixed chimera was unevaluable because of death at 3 weeks from intussusception; and 4 showed persisting mixed chimerism, including unirradiated marrow and lymph node spaces, for now more than 46 to 70 weeks after transplant. Data support the hypothesis that stable marrow allografts can be established by combining nonmyeloablative pretransplant host immunosuppression with posttransplant host and donor cell immunosuppression using MMF/CSP.

Stable mixed donor/host hematopoietic chimerism can be accomplished in dog leukocyte antigen (DLA)-identical littermate dogs given sublethal (200 cGy) total-body irradiation (TBI) before and immunosuppression with mycophenolate mofetil (MMF) and cyclosporine (CSP) after transplant (Blood 89:3048, 1997). Studies were based on the hypothesis that drugs that prevent graft-versus-host disease (GVHD) after transplant also suppress host-versus-graft (HVG) reactions and thereby enhance engraftment. Here, we asked whether pretransplant TBI provided marrow space for the graft to home or caused host immunosuppression. To address the questions, recipients were given pretransplant irradiation to cervical, thoracic, and abdominal lymph nodes (except pelvis), DLA-identical littermate marrow grafts, and MMF/CSP posttransplant. Six dogs that received 450 cGy irradiation showed initial engraftment. Two rejected their grafts after 8 and 18 weeks, 1 died with GVHD and engraftment, and 3 are alive as mixed chimeras after 57 to 97 weeks. Four dogs given 200 cGy irradiation also showed initial engraftment, but rejected their grafts after 10 to 18 weeks. Mixed chimerism was present in nonirradiated marrow and lymph node spaces and involved granulocytes, T cells, and monocytes. While other explanations are possible, results seem consistent with the hypothesis that pretransplant radiation provides host immunosuppression, and grafts can create their own marrow space. These data set the stage for the development of novel transplant regimens that substitute immunosuppressive for cytotoxic agents.

The construction of an apparatus for electrochemical investigations using an IBM personal microcomputer is presented. This apparatus can be used in a wide range of electrochemical methods, stationary and dynamic. A special construction solution allows for application in electrochemical impedance spectroscopy measurements, particularly using the fast Fourier transform method. The apparatus contains a potentiostat connected with signal generators for the polarization of the working electrode and two identical analogue-to-digital conversion channels. For data transmission between this apparatus and the microcomputer, two parallel transmission ports and a special communication interface are used. Also, the detailed description of the construction of the analogue-to-digital conversion channels, including a special module for direct voltage compensation in the input analogue signal, is given, together with the final technical specification of the apparatus.

BACKGROUND: Data on red blood cell (RBC) alloantibody appearance after hematopoietic progenitor cell transplantation (HPCT) from an ABO-matched donor are limited. CASE REPORT: A 32-year-old Caucasian man with chronic myeloid leukemia, never transfused, conditioned with BuCy120, received granulocyte–colony-stimulating factor–mobilized peripheral blood progenitor cells from his HLA-identical sister. Graft-versus-host disease (GVHD) prophylaxis consisted of cyclosporine A and methotrexate. The patient engrafted showing complete donor chimerism and developed Grade 3 acute GVHD and subsequently extensive chronic GVHD. He received 2 units of RBCs on Days 13 and 14 without clinical signs of hemolysis. RESULTS: Patient was 0 Rh–K–JK(a–) and donor was 0 Rh+K–JK(a–). Pretransplant alloantibody screening was negative in the patient. In the donor, anti-E was detected before donation, together with anti-Jka and anti-Dib 4, 9, and 12 months after donation, likely triggered by RBC transfusion 2 months before cell harvest. In the patient, anti-Jka was detected on Day +25 and later anti-E and anti-Dib as well. In both, a Dia/Dia genotype was identified. Both parents were Di(a+b+). CONCLUSIONS: Three kinds of non-ABO alloantibodies were detected, including anti-Dib described for the first time after HPCT. The rapid development of antibodies in the recipient despite intensive immunosuppression suggests their production by the donor cells primed by the RBC transfusion before HPC harvest. Their production by the residual host cells cannot be unequivocally excluded, however.

Background Stable mixed-donor-host-hematopoietic chimerism can serve as a platform for adoptive immunotherapy. Infusions of donor lymphocytes (DLI) sensitized against hematopoietic cells converted mixed hematopoietic into full-donor chimerism in dog-leukocyte antigen (DLA)-identical littermates. Whether sensitization against tissue of solid organs leads to organ-specific immunity that can be transferred by DLI was unknown and was investigated in these experiments using the kidney as target. Methods. DLA-identical recipients with established stable mixed-donor-host-hematopoietic chimerism were used. In five pairs, hematopoietic stem-cell transplant (HSCT) donors were sensitized by kidney transplantation from the respective chimeras. In a second group, five HSCT donors received vaccinations that were generated from kidney cells of the respective mixed chimeras. Twenty-eight days after sensitization, DLI were administered to the mixed-hematopoietic chimeras. Results. All HSCT donors rejected their kidney grafts from their mixed-chimeric recipients within 22 to 45 days. DLI caused no sustained graft-versus-kidney effects in the mixed-chimeric recipients. However, DLI donors sensitized by kidney transplantation converted 4 of 5 mixed chimeras into virtually complete (>95%) donor-type chimeras, compared with 1 of 5 mixed chimeras given DLI by vaccination from sensitized donors. Conclusion. Although DLA-identical kidney grafts from mixed-hematopoietic chimeras were readily rejected by their HSCT donors, subsequent transfusions of sensitized-donor lymphocytes into mixed chimeras converted mixed to all-donor chimerism but failed to induce graft-versus-kidney effects. Vaccination strategies in lieu of kidney grafts failed to convert mixed chimerism.

The development of nonmyeloablative allogeneic hematopoietic stem cell transplantation (HSCT) from the preclinical studies to the clinic has permitted the treatment of a larger number of patients who previously had not been candidates for the standard approach with myeloablation. This includes older patients and those patients who had contraindications to intensive cytotoxic regimens. Regimen-related toxicities (RRT) after myeloablation result in prolonged periods of hospitalization and the development of significant morbidity and potential mortality including veno-occlusive disease and idiopathic interstitial pneumonitis. Severe RRT can generally be avoided after nonmyeloablative conditioning. A graft-versus-leukemia reaction is critical to the eradication of many hematological malignancies after transplantation and has been shown to be effective in those patients who have been infused with lymphocytes from the donor (DLI) after relapse.

We hypothesized that patients undergoing major ABO-incompatible non-myeloablative haematopoietic stem cell transplantation (nm-HSCT) might experience prolonged haemolysis after transplant due to the delayed disappearance of host plasma cells producing anti-donor isohaemagglutinins (HAs). To address this question, we analysed data from 107 consecutive patients transplanted with allogeneic peripheral blood stem cells from human leucocyte antigen-matched (related, n = 84; unrelated, n = 23) donors after non-myeloablative conditioning (200 cGy total body irradiation +/- fludarabine). In total, 23 out of the 107 patients received major or major/minor ABO-incompatible transplants. Red blood cell (RBC) transfusion requirements during the first 120 d post transplant were higher in major ABO-mismatched than in ABO-matched recipients (0.12 vs 0.03 median units RBC concentrate/d, P = 0.04). Two patients developed transient pure red cell aplasia, which had resolved spontaneously by 9 months after transplant. Major ABO incompatibility did not influence rates of engraftment. Patients with sustained engraftment experienced gradual declines of anti-donor HAs, and the estimated median time to reaching IgM and IgG titres of1:1 was at least 133 d in evaluable patients, approximately twice longer than reported after myeloablative conditioning. There was a strong correlation between degrees of donor chimaerism in erythroid burst-forming units, granulocyte macrophage colony-forming units and granulocytes, indicating that donor erythroid engraftment, defined by early erythroid progenitors, was as prompt as myeloid engraftment. In conclusion, our data suggest that major ABO-incompatibility is not a barrier to successful non-myeloablative HSCT.

A retrospective analysis of granulocyte colony-stimulating factor (G-CSF)–mobilized peripheral blood mononuclear cell (G-PBMC) products harvested from healthy donors indicates significant variability in both the absolute number and relative proportion of CD34, CD3, and CD14 cells obtained. This report examined whether variations in the cellular composition of G-PBMC products correlated with clinical outcomes after myeloablative allogeneic transplantation. The numbers of CD34, CD3, and CD14 cells infused into 181 human leukocyte antigen (HLA)–identical sibling recipients were analyzed with respect to tempo of engraftment, acute graft-versus-host-disease (GVHD), clinical extensive chronic GVHD, overall survival, and disease relapse. Neither acute GVHD, overall survival, nor disease relapse was statistically significantly associated with CD34, CD3, or CD14 cell doses or the CD14 to CD3 ratio. CD3 and CD14 cell doses and CD14 to CD3 ratios did not correlate with the tempo of neutrophil and platelet engraftment. However, increasing CD34 cell numbers were significantly associated with accelerated neutrophil (P = .03) and platelet (P = .01) engraftment. Higher doses of CD34 cells (> 8.0 × 106/kg) were also associated with a significantly increased hazard of clinical extensive chronic GVHD (HR = 2.3, 95% confidence interval [CI] 1.4-3.7,P = .001), but neither CD3 nor CD14 doses were statistically significantly associated with chronic GVHD. It was concluded that CD34 cell dose in G-PBMC grafts appears to affect both the engraftment kinetics and the development of clinical extensive chronic GVHD in HLA-identical sibling recipients but without a demonstrable impact on survival, relapse, and acute GVHD. Given the morbidity associated with extensive chronic GVHD, efforts to further accelerate engraftment in HLA-matched sibling transplants by increasing CD34 cell number in G-PBMC products may be counterproductive.

Genetically modified donor T cells with an inducible “suicide” gene have the potential to improve the safety and availability of allogeneic hematopoietic stem cell transplantation by enhancing engraftment and permitting control of graft-versus-host disease (GVHD). However, several clinical studies of gene-modified T cells have shown limited to no in vivo function of the ex vivo expanded T cells. Using the well-established dog model of allogeneic marrow transplantation, the question was asked if retrovirally transduced, donor derived, ex vivo expanded cytotoxic T lymphocytes (CTLs) that are recipient specific could enhance engraftment of dog leukocyte antigen (DLA)–haploidentical marrow following a single dose of 9.2 Gy total body irradiation and no postgrafting immunosuppression. In this setting, only 4 of 11 control recipients of DLA-haploidentical marrow without added CTLs engrafted. CTLs did not enhance engraftment of CD34+ selected peripheral blood stem cells. However, recipient-specific CTLs enhanced engraftment of DLA-haploidentical marrow in 9 of 11 evaluable recipients (P = .049). All dogs that engrafted developed multiorgan GVHD. To facilitate in vivo tracking, 8 dogs received CTLs transduced with a retroviral vector encoding green fluorescent protein (GFP) and neomycin phosphotransferase (neo). Recipients that engrafted had sharp increases in the numbers of circulating GFP+ CTLs on days +5 to +6 after transplantation. GFP+ CTLs isolated from blood were capable of recipient-specific lysis. At necropsy, up to 7.1% of CD3+ cells in tissues were GFP+ and polymerase chain reaction in situ hybridization for neoshowed infiltration of transduced CTLs in GVHD-affected organs. These results show that ex vivo expanded, transduced T cells maintained in vivo function and enhanced marrow engraftment.

Clinical observations show that older patients do not tolerate high-dose chemoradiotherapy as well as younger patients. It is unclear whether this is due to age-related differences in their responses to hematopoietic injury or to differential toxicities to other organs. In the present study, 6 young (0.5 years) and 6 elderly (8 years) dogs were challenged with 7 repeated nonlethal doses of 50 or 100 cGy total body irradiation (TBI) each (total 550 cGy), and 21 days of recombinant canine granulocyte–colony stimulating factor (rcG-CSF) after the last TBI dose. Recoveries of absolute neutrophil, platelet, and lymphocyte counts after each TBI dose, responses to rcG-CSF treatment, and telomere lengths in neutrophils were compared before and after the study. No differences were found in recoveries of neutrophils, platelets, or in responses to rcG-CSF among young and old dogs. In contrast, recoveries were suggestively worse in younger dogs. After rcG-CSF, platelet recoveries were poor in both groups compared with previous platelet recoveries (P

Stable mixed donor/host hematopoietic chimerism was uniformly achieved in dogs given 200 cGy total body irradiation (TBI) before and immunosuppression with mycophenolate mofetil (MMF) for 28 days and cyclosporine (CSP) for 35 days after transplantation of marrow from dog leukocyte antigen-identical littermates. When the TBI dose was lowered to 100 cGy, donor marrow engraftment in 6 dogs was only transient, lasting 3 to 12 weeks. In this study, we asked whether stable engraftment in this model could be achieved: (1) by substituting recombinant canine granulocyte-colony-stimulating factor-mobilized peripheral blood mononuclear cells (G-PBMCs) for marrow and (2) by extending CSP administration from 35 to 100 days. Eighteen dogs were given G-PBMC grafts and MMF for 28 days. Eight of the 18 dogs received CSP for 35 days and 10 for 100 days. We found that substituting G-PBMCs for marrow did not increase the incidence of stable allogeneic engraftment (P = .11). However, increasing the duration of posttransplantation immunosuppression with CSP from 35 to 100 days favorably influenced stable donor engraftment (P = .06). Biol Blood Marrow Transplant 2001;7(9):513-6.

Stable mixed donor/host hematopoietic chimerism can be uniformly established in dogs conditioned with 200 cGy TBI before dog leukocyte antigen (DLA)-identical marrow transplantation and immunosuppressed with a short course of mycophenolate mofetil (MMF) and cyclosporine (CSP) after the transplantation. A further decrease in the TBI dose to 100 cGy or the elimination of MMF in this model results in graft rejection. Here we asked whetherthe addition of G-CSF-mobilized peripheral blood mononuclear cells (G-PBMC) to marrow grafts would enhance donor engraftment in dogs conditioned with 100 cGy TBI and given postgrafting immunosuppression with CSP alone. Using this model, 7 of 9 dogs given only marrow cells rejected their grafts within 8 to 17 weeks after transplantation. In contrast, the addition of unmodified G-PBMC to marrow grafts resulted in stable mixed donor/host chimerism in 5 of 8 dogs studied (P = .06). However, addition of the CD3-depleted fraction of G-PBMC, which contained both CD34 cells and CD14 cells, resulted in engraftment in only 1 of 7 recipients. We conclude that adding G-PBMC to marrow grafts replaced the requirement of MMF and 100 cGy of TBI, and that CD3 cells were required to facilitate engraftment of marrow cells in DLA-identical recipients, whereas the additional CD34 cells present in G-PBMC were not sufficient for this effect. Biol Blood Marrow Transplant 2001;7(11):613-9.