Kinetics of peripheral blood CD34-positive cells and the optimum timing for harvesting peripheral blood stem cells during BEP chemotherapy in patients with testicular germ cell tumor

PURPOSE: Recently, high-dose chemotherapy with peripheral blood stem cell (PBSC) rescue has been developed for poor risk testicular germ cell cancer. In this study, we investigated the optimum timing for harvesting PBSCs with the use of bleomycin + etoposide + cisplatin (BEP) chemotherapy, which is a well known first-line regimen for the testicular cancer. MATERIAL AND METHOD: Peripheral blood CD34-positive cell ratios were measured during a total of 10 courses of BEP chemotherapy in 6 patients with metastatic germ cell cancer between 1996 and 1998. We performed 4 apheresis in 3 patients during this period. Recombinant human granulocyte-colony stimulating factor (rhG-CSF) was administrated from the day on which the neutrophil count decreased less than 1,000/microliter. RESULTS: The peripheral blood CD34-positive cell ratios became maximum (3.0-24.6%; average 10.0%) on the day 18 to 21 (median day 19) of BEP chemotherapy with rhG-CSF administration. The maximum ratios of peripheral blood CD34 positive cells were achieved when the number of leukocyte were 6,880-23,600/microliter and exceeded 6,000/microliter after the 18th day of BEP chemotherapy. The average number of collected CD34 positive cells was 9.5 x 10(6)/kg at a single apheresis, and 12.6 x 10(6)/kg per patient. CONCLUSION: Efficient hematopoietic progenitor cells were mobilized by BEP chemotherapy with rhG-CSF administration of first-line setting. Our results suggest that the optimum timing of PBSCs harvest is the day when the numbers of leukocyte exceed 6,000/microliter after the 18th day of BEP chemotherapy and the following day.     

Hematologic reconstitution following high-dose and supralethal chemoradiotherapy using stored, noncryopreserved autologous hematopoietic stem cells

Although cryopreservation is the standard for autotransplantation, it has logistic and financial disadvantages in undeveloped countries such as Colombia. In 47 patients, peripheral blood was refrigerated at 4 degrees C up to 144 h before autotransplantation. For mobilization, 27 men and 20 women of median age 37 years affected with hematologic malignancies received G-CSF. The 17 patients in Group 1 showed pre-refrigeration CFU-GM of 2.62 x 10(5)/kg (range 0.36 to 16.6 x 10(5)/kg) and at re-infusion, 1.36 x 10(5)/kg (range 0 to 6.32 x 10(5)/kg) of 83% viability (range, 78% to 96%). These patients showed >0.5 x 10(9)/L granulocytes on day +11 (range, 9 to 15) and >20 x 10(9)/L platelets on day +16 (range, 11 to 44). The 25 patients in Group 2 showed CD34 of 3.9 x 10(6)/kg (range, 0.16 to 9 x 10(6)/kg) and mononuclear cell count (MNC) of 8.7 x 10(8)/kg, reaching >0.5 x 10(9)/L granulocytes at day +13 (range, 10 to 17) and >20 x 10(9)/L on day +15 (range, 14 to 20). Among the 5 patients in Group 3, the average of MNC of 12.7 x 10(8)/kg was reached and >0.5 x 10(9)/L granulocytes on day 11 (range, 10 to 16) and >20 x 10(9)/L on day 14 (range, 10 to 18). No differences were observed between the groups. Refrigeration of stem cells appears to be a simple, effective, and inexpensive method that should be considered for autotransplants within a few days of harvesting when resources are limited for long-term storage.     

Peripheral blood stem cell harvest for patients with germ cell tumors

From January 1996 to December 1999, fifteen patients with germ cell tumors underwent peripheral blood stem cell harvest during 15 courses of bleomycin, etoposide, cisplatin (BEP), 4 courses of etoposide, ifosfamide, cisplatin (VIP) and 3 courses of high-dose etoposide mobilization at Nagoya University Hospital. We performed 29 aphereses during BEP, eight during VIP, and six during high-dose etoposide. Although we were able to harvest 4.4 x 10(6)/kg of median CD34 positive cells per apheresis during BEP, the number of stem cells (more than 4 x 10(6)/kg of CD34 positive cells), which are needed for tandem high-dose chemotherapy, could not be obtained during four courses of BEP. For three patients in whom white blood cell counts at nadir were 2,000/microL or more, however, the required number of CD34 positive cells were harvested. VIP provided only 1.7 x 10(6)/kg of median CD34 positive cells per apheresis, while, 7.3 x 10(6)/kg of CD34 positive cells were harvested during high-dose etoposide mobilization. The dose of G-CSF was a significant factor for the number of CD34 positive cells harvested during BEP (p = 0.02); however, there might be some relationship between the harvest and the number of the peripheral white blood cells on the day of apheresis (p = 0.08), the day to start G-CSF (p = 0.13), or the day to initiate apheresis (p = 0.27). Based on our experience, it is recommended that 5 micrograms/kg of G-CSF should be started from the 14th or 15th day of BEP until the last apheresis and that aphereses should be performed between the 19th and 21st day, especially at the days when the peripheral white blood cell count increases beyond 10,000/microL.     

Second mobilization of peripheral blood progenitor cells in patients with poor first mobilization

The aim of this study was to analyse the efficacy of 2 second mobilization (MB) protocols in 2 groups of patients who failed to obtain enough peripheral blood progenitor cells (PBPC) in the first MB. In 1 group (8 patients), 10 microg/kg of G-CSF was administered, and in the other group (8 patients), a double dosage (10 microg/kg twice a day) was administered. Both groups of patients received Cyclophosphamide (1.5 g/kg) 10 days before the apheresis. No difference was found among both groups of patients in diagnosis, previous chemotherapy, and time elapsed after the first MB. Administration of higher doses of G-CSF decreased the number of apheresis needed in the second MB to complete 2 x 10(6)/kg of CD34+ cells. It also increased the number of patients who achieved sufficient CD34+, namely, 75% versus 50%.     

Two-step ultra high-dose chemotherapy with peripheral blood stem cell autotransplantation for refractory testicular cancer: a case report]

A 35-year-old male had advanced nonseminomatous germ cell tumor (stage IIIC, embryonal cell carcinoma) which proved refractory to conventional PVB combined chemotherapy. He was then treated with an ultra high-dose chemotherapy consisting of carboplatin (1.5 g/m2) and etoposide (1.3 g/m2), followed by the transplantation of peripheral blood stem cells (PBSCT) with a total of 1.9 x 10(5)/kg granulocyte colony-forming cells (CFU-GM). Because he developed lung metastasis, escalated doses of carboplatin (2.0 g/m2), and etoposide (1.8 g/m2) combined with cyclophosphamide (7.0 g/m2) were given with peripheral blood stem cell transplant of 3.2 x 10(5)/kg CFU-GM. He has remained free of any recurrence without maintenance therapy.     

Cotreatment with darbepoetin and granulocyte colony-stimulating factor is efficient to recruit proangiogenic cell populations in patients with acute myocardial infarction

To determine whether newer combination cytokine treatment with granulocyte colony-stimulating factor (G-CSF) and darbepoetin can improve efficacy of stem cell therapy, we evaluated safety and peripheral blood stem/progenitor cell (PBSC) mobilizing effects of combination cytokine in comparison with G-CSF alone in patients with acute myocardial infarction (AMI). We randomized 60 patients with AMI into two groups under 2:1 ratio; combination treatment with darbepoetin and G-CSF (n = 41: Combicytokine group) and the G-CSF alone (n = 19: G-CSF group). After coronary angioplasty, G-CSF was treated for 3 days with dose of 10 μg/kg/day in both groups. Only in the combicytokine group, additional single intravenous injection of 4.5 μg/kg of darbepoetin was administrated immediate after coronary angioplasty. Combination cytokine treatment was well tolerated as was G-CSF alone. PBSCs were obtained by apheresis for intracoronary infusion after completion of cytokine treatment and were analyzed by flow cytometry. The purity of proangiogenic cells was higher in combination cytokine group than the G-CSF group. Specifically, proportion of CD34(+)/KDR(+) endothelial progenitor cells, CD3(+)/CD31(+) angiogenic T cells and Tie2(+)/CXCR4(+) cells in apheresis products were higher in the combicytokine group. These meant that the combicytokine treatment recruited PBSCs in higher purity and fewer unwanted inflammatory cells than G-CSF alone in apheresis products. Combination treatment with darbepoetin and G-CSF is safe and more efficient to mobilize and recruit proangiogenic cells than G-CSF alone in patients with AMI. (Trial registration: www.ClinicalTrials. gov identifier: NCT00501917).     

Mobilization and Collection of Peripheral Blood Stem Cells: Guidelines for Blood Volume to Process, Based on CD34-Positive Blood Cell Count in Adults and Children

We report 189 mobilizations and 489 collections of peripheral blood stem cells (PBSC) performed in 139 autologous transplantation patients and in 28 donors for allogeneic transplantations whose ages ranged from 2-68 years. We observed a correlation (P < .001; Pearson's coefficient 0.64) between CD34-positive cells and granulocyte-macrophage colony-forming units examined to estimate PBSC. In a subset of 287 collections (97 adults and 49 children) we obtained peripheral blood (PB) CD34-positive cell counts at 2 to 4 hours before leukapheresis. We noted a correlation between PB CD34-positive cell counts before leukapheresis and the number of CD34-positive cells per kilogram of body weight collected in the whole apheresis of the day (P < .001; Pearson's coefficient 0.82). An even better correlation was obtained between PB CD34-positive cells preapheresis and the yield of each individual blood volume (BV) processed (P < .001; Pearson's coefficient 0.87). Healthy donors and patients in each age group behaved similarly. In addition, the collection yield was greater among children than adults. These findings allowed us to develop a simple predictive model to estimate the BV to process for a target dose of CD34-positive cells per kilogram, based on the level of PBSC before apheresis in children and adults.     

Addition of cladribine to induction/consolidation regimen does not impair peripheral blood stem cell mobilization and bone marrow harvest for autotransplantation in acute myeloid leukemia patients

BACKGROUND: The previous study by the Polish Adult Leukemia Group has demonstrated that addition of cladribine to standard DNR+AraC induction potentiates the antileukemic activity. The goal of this study was to compare the efficacy of bone marrow or peripheral blood hematopoietic cell collection in patients who obtained remission after daunorubicine plus cytarabine induction with cladribine (DAC-7) or without addition of cladribine (DA-7) in preparation for autotransplantation. PATIENTS AND METHODS: Sixty-six patients aged 41 years (range, 17-58 years) were included in this study: 33 cases in the DAC-7 and 33 in the DA-7 arm. Hematopoietic cells were collected from the bone marrow (ABMT, n = 29) or from the peripheral blood (ABCT, n = 37) using cytopheresis after administration of AraC (2 x 2 g/m2) on days 1, 3, 5 and subsequent G-CSF (10 microg/kg) from day 7 as mobilization therapy. RESULTS: The numbers of harvested CD34+ cells were similar in the DAC-7 and DA-7 pretreated patients both after harvesting from peripheral blood (2.55 x 10(6)/kg vs 2.5 x 10(6)/kg) and from bone marrow (1.62 x 10(6)/kg vs 1.55 x 10(6)/kg), respectively. The proportion of patients with sufficient material for autologous bone marrow transplantation was higher in the DAC-7 compared with the DA-7 arm. All patients engrafted; hematopoietic recovery was similar in both subgroups. CONCLUSION: Addition of cladribine to a standard DA induction does not impair the harvesting of hematopoietic cells and their engraftment after autotransplantation.     

Feasibility and long-term results of autologous PBSC transplantation in recurrent undifferentiated nasopharyngeal carcinoma

BACKGROUND: Recurrent undifferentiated nasopharyngeal carcinoma (UNPC) is a chemosensitive illness. Here we report long-term results of high-dose chemotherapy (HDC) as late intensification, with autologous peripheral blood stem cell (PBSC) support. METHODS: Six patients (5 men, 1 woman; median age 41years; median ECOG PS = 0) with recurrent UNPC (local, 2; local + nodal, 2; bone metastasis, 2) have been enrolled. All patients had been previously treated with neoadjuvant chemotherapy and radiotherapy; 3 of 4 local relapses had received a re-irradiation. Every patient received three courses of cisplatin + epirubicin and 1 cycle of epirubicin followed by PBSC collection. A median of 7.2 x 10(6)/kg (range, 4.5-18) CD34+ cells were reinfused. HDC was according ICE scheme: ifosfamide, 2.5 g/m(2)/d, + carboplatin, 300 mg/m(2)/d, + VP-16, 300 mg/m(2)/d days 1 through 4. RESULTS: After conventional chemotherapy, we had 1 CR (16%), 3 PR (50%), and 2 NC (34%). After HDC, we had 4 CR (66%),1 PR (17%), and 1 MR (17%). Toxicity was manageable. After a median follow-up of 30 months (range, 14-50), two patients are alive without disease (34%), one is alive with bone disease (16%), and three (50%) died of disease at 16, 18, and 24 months. CONCLUSIONS: HDC has an acceptable toxicity, can convert PR in CR, and seems effective, with long-lasting CRs.     

Optimizing umbilical cord blood collection: impact of obstetric factors versus quality of cord blood units

INTRODUCTION: The main limitation factor for wide use of umbilical cord blood units (UCBs) as a source of hematopoietic progenitors for transplantation is cell dose. International standard guidelines recommend 2 x 10(7)/kg as the minimal nucleated cell dose for UCB transplantation for adults and 3.7 x 10(7)/kg for children. Therefore it is important to the optimize donor selection and the collection method so as to achieve high cell doses. In this study our main purpose was to determine whether obstetric factors influence UCBs collected. STUDY DESIGN: The study involved 304 UCBs collected from January to December 2004. The UCBs were collected after donor selection based on international criteria for cord blood banking. We analyzed UCB biological features such as collected volume, total nucleated cells (TNC), and CD34-positive cells, and obstetric factors. RESULTS: First, our study showed by multivariate analysis that infant weight was the main factor that influenced biologic features of UCB collected such as total volume (P = .000), TNC (P = .000), CD34 total count (P = .003), and CFU-GM (P = .004). Placental weight > 600 g produced a better volume (P = .007) and increased TNC (P = .056). Gestational age > 39 weeks enhanced CD34% (P = .016). Regarding route of delivery, we found that cesarean section produced higher volume and reduced WBC count compared to vaginal delivery, regarding cord length, it increased TNC (P = .037). And last, we noticed that female infants increased WBC (P = .013) and CD34(+) total count (P = .019) more than male ones. CONCLUSIONS: Our results confirm that volume and TNC are influenced by several obstetric factors, such as greater infant and placental weight, predicting a better collection.     

-80℃非程控冷冻保存人外周血干细胞的效果观察

目的 探讨-80℃非程控冷冻保存人外周血干细胞(PBSC)的效果.方法 41份PBSC标本,39份采自恶性肿瘤患者,2份采自正常供者.对于恶性肿瘤患者,干细胞采集前采用针对性化疗方案联合粒细胞集落刺激因子或粒-单细胞集落刺激因子进行动员.将采集的干细胞与采用120 g/L羟乙基淀粉、体积分数为10%二甲基亚砜及80 g/L人血清白蛋白组成的冷冻保护剂按1:1混合,然后将其直接放入-80℃低温冰箱内保存.测定保存前以及保存后不同时间的单个核细胞(MNC)、有核细胞(NC)、CD34+细胞、粒一单核细胞系集落形成单位(CFU-GM)和红系爆式集落形成单位(BFU-E)等的回收率以及细胞的台盼蓝拒染率.结果 经-80℃非程控直接冷冻保存1个月至10年,PBSC的台盼蓝拒染率、NC和MNC回收率的变化较小(P>0.05),而CD34+细胞、CFU-GM和BFU-E回收率在冻存5～10年后下降较明显(P＜0.05),但仍分别达89.6%、85.1%和83.7%.34例患者经相应处理后,回输经-80℃下保存13～53 d(平均19 d)的干细胞,患者于细胞回输后11～27 d(平均14.7 d)获得造血功能重建.结论 在采用终浓度为60 g/L 羟乙基淀粉、体积分数为5%二甲基亚砜及40 g/L人血清白蛋白组成的冷冻保护剂的情况下,-80℃非程控冷冻法适于PBSC的长期保存,效果较理想.     

The effect of G-CSF-stimulated donor marrow on engraftment and incidence of graft-versus-host disease in allogeneic bone marrow transplantation

Graft-versus-host disease (GVHD) and infection are major obstacles to successful allogeneic bone marrow transplantation (allo-BMT). In an attempt to improve the results of HLA-identical sibling BMT, we investigated the effect of accelerating hemopoietic reconstitution and reducing acute GVHD (aGVHD) in allo-BMT receiving G-CSF-stimulated donor marrow and the preliminary biological mechanism. The donors of 30 patients (study group) with leukemia were given G-CSF 3-4 microg/kg/d for 7 doses prior to marrow harvest. The results of subsequent engraftment in the recipients were compared with those of 18 patients without G-CSF (control group). Five donors themselves were studied to assess the effects of G-CSF on the hematopoietic progenitor cells and lymphocyte subsets in the bone marrow (BM). We observed that the stimulated BM yielded higher numbers of nucleated cells as well as CFU-GM and CD34+ cells (p<0.01), and that hemopoietic reconstitution was accelerated. The median number of days of granulocyte count exceeding 0.5x10(9)/L and platelet count exceeding 20x10(9)/L was 16 (range 10-23 d) and 18.5 (range 13-31 d), respectively (control group: median 22 d, range 13-29 d and median 23 d, range 17-34 d; p=0.001). The incidence of grade II-IV severe aGVHD was very low, with only 1 case (3.3%) with acute grade II aGVHD limited to the skin in the study group. Five of 18 patients in the control group manifested grade II-IV severe aGVHD (27.8%, p=0.02). The number of T-lymphocyte subsets in the harvested BM using G-CSF stimulation was changed. In the G-CSF-stimulated marrow group, CD4+ decreased and CD8+ increased significantly (p=0.02). The changes of progenitor cells and T-lymphocyte subsets in donors' BM from pre- and post-G-CSF stimulation showed that the percentage of CD4+ reduced (p=0.04) and that of CD8+ increased (p=0.06), while that of CD34+ also increased (p=0.002). The incidence of chronic GVHD and relapse had no significant difference between both groups. These results indicate that allo-BMT in BM G-CSF priming can accelerate engraftment and minimize the incidence of severe aGVHD. There is a trend in favor of improved transplantation-related mortality.     

Transplantation of human peripheral blood stem cells into fetal rhesus monkeys (Macaca mulatta)

BACKGROUND: Methods for assessing engraftment efficiency have been explored in a primate xenogeneic model of in utero hematopoietic stem cell transplantation. METHODS: Human peripheral blood stem cells (PBSC) were obtained by leukapheresis from a human male donor after 4 days of administration of recombinant human granulocyte-colony stimulating factor (5 microg/kg/ day). PBSC were enriched for the CD34+ population with and without T-cell depletion. The resulting mononuclear cells consisted of two cell populations, one that was stem cell enriched (0.83% CD3+ cells, 95% CD34+; group 1) and one that was stem cell enriched and T-cell depleted (<0.03% CD3+ cells, 98% CD34+; group 2). Four fetal monkeys (two per group) received either two or four i.p. injections (approximately 5x10(6) cells/injection) via ultrasound guidance every other day over a 7-day period (gestational days 50, 52, 54, and 56). One fetus in each group also received i.p. recombinant human stem cell factor (25 microg/kg) and recombinant human granulocyte-colony stimulating factor (10 microg/kg) posttransplant every 10 days from gestational day 60-150. RESULTS: Four healthy newborns were delivered at term, and specimens were analyzed by polymerase chain reaction for the human Y chromosome (birth, monthly to 6 months; blood, marrow, progenitor assays). Polymerase chain reaction results were positive for all four newborns in all specimens assessed, and flow cytometric analysis for human CD45 in marrow showed engraftment ranging from 0.1-1.7%. There was no evidence of graft-versus-host disease in any of the animals. CONCLUSION: These studies show that (1) multilineage engraftment of human PBSC can be achieved in the fetal rhesus recipient, (2) the rhesus fetus appears to tolerate relatively high numbers of human CD3+ cells, and (3) healthy chimeric rhesus infants can be delivered at term after multiple in utero procedures.     

供者应用粒细胞集落刺激因子促进异基因骨髓移植受者造血重建的临床研究

目的评价供者应用粒细胞集落刺激因子（G-CSF）以增加骨髓中CD34+细胞的效果和对异基因骨髓移植造血重建的影响。方法供者采髓前应用G-CSF250μg/d,连用7d,随后对15例白血病患者进行异基因骨髓移植,观察植入物单个核细胞、CD3     

Neutropenic enterocolitis in acute myeloid leukemia

In this report we focus on the importance of an accurate diagnosis of gastrointestinal complications during chemotherapy for acute myeloid leukemia. The leukemic infiltrtion of the digestive system may cause mucosal ulcers which can lead to bleeding or perforation. The immune system deficiency in this cohort of patients may result in necrotic enterocolitis (leukemic typhlitis), perianal inflammation, abscesses, and peritonitis. We describe a 37-year old male who presented in June 2004 with 2-month history of fever, weakness and sore throat, treated with antibiotic therapy. Physical examination demonstrated palor. The peripheral blood count at admittance was as follow: Hemoglobin 87 g/l, WBC 63 x 10(9)/l, and platelets 56 x 10(9)/l. The peripheral blood differential count showed: myeloblasts 4%, polymorphonuclear neutrophils (PMN) 20%, monocytes 60%, lymphocytes 16%. The diagnosis of acute myeloid leukemia (AML) was confirmed by bone marrow aspirate, which presented an almost total infiltration by monocytoid blasts, AML type M5 according to FAB classification. Immunophenotypic evaluation by flow cytometry showed that the blast cells reacted with antibodies to CD33, CD13, CD14, CD64, CD15, cytogenetics showed normal karyotype. Induction treatment consisting of cytarabine 2 x 200 mg intravenously in push on days 1-8, vepeside 200 mg i.v. on days 1-5, adriblastine 90 mgon days 1,3 and 5. On day 15 of chemotherapy the patient got fever 38.5 degrees C, abdominal pain and diarrhea (10 stools daily). Broad-spectrum antibiotic therapy with ceftriaxone and amikacin was promptly instituted but condition worsened, abdominal pain extended to all abdomen while the fever and diarrhea persisted. Ultrasonography on day 18 documented bowel wall thickness of colic tract, part of duodenum and jejunum. Owing to suspicion of neutropenic enterocolitis, antibiotic therapy intensified with teicoplanin, fluconazole, metronidazole and pipril. Patient was neutropenic and thrombocytopenic, although daily platelet transfusion from a single donor were given. We started with granulocyte colony stimulating factor (G-CSF) 5 g/kg, which was adiminstered for 7 days. After 7 days neutrophil value reached 1 x 10(9)/l, but fever persisted, abdominal distension and diarrhea progressively improved. The fever peristed and central venous catheter was removed on day 30. After removal of the catheter the patient was getting better: the fever disappeared. The blood count showed Hb 91 g/l, WBC 3,4 x 10(9)/l, platelet 114 x 10(9)/l and normal leukocyte differential count. We emphesize the importance of collaboration between the hematologist and the surgeon in monitoring gastrointestinal complications during and after chemotherapy for acute leukemias and value of abdominal ultrasonography evaluation.     

T-cell reconstitution after unmanipulated, CD8-depleted or CD34-selected nonmyeloablative peripheral blood stem-cell transplantation

BACKGROUND: We have previously shown that CD8 depletion or CD34 selection of peripheral blood stem cells (PBSC) reduced the incidence of acute graft-versus-host disease (GvHD) after nonmyeloablative stem-cell transplantation (NMSCT). In this study, we analyze the effect of CD8 depletion or CD34 selection of the graft on early T-cell reconstitution. METHODS: Nonmyeloablative conditioning regimen consisted in 2 Gy total-body irradiation (TBI) alone, 2 Gy TBI and fludarabine, or cyclophosphamide and fludarabine. Patients 1 to 18 received unmanipulated PBSC, patients 19 to 29 CD8-depleted PBSC, and patients 30 to 35 CD34-selected PBSC. RESULTS: T-cell counts, and particularly CD4+ and CD4CD45RA+ counts, remained low the first 6 months after nonmyeloablative stem-cell transplantation (NMSCT) in all patients. CD34 selection (P<0.0001) but not CD8 depletion of PBSC significantly decreased T-cell chimerism. Donor T-cell count was similar in unmanipulated compared with CD8-depleted PBSC recipients but was significantly lower in CD34-selected PBSC recipients (P=0.0012). T cells of recipient origin remained stable over time in unmanipulated and CD8-depleted PBSC patients but expanded in some CD34-selected PBSC recipients between day 28 and 100 after transplant. Moreover, whereas CD8 depletion only decreased CD8+ counts (P<0.047), CD34 selection reduced CD3+(P<0.001), CD8+(P<0.016), CD4+ (P<0.001), and CD4+CD45RA+ (P<0.001) cell counts. T-cell repertoire was restricted in all patients on day 100 after hematopoietic stem-cell transplantation but was even more limited after CD34 selection (P=0.002). CONCLUSIONS: Despite of the persistence of a significant number of T cells of recipient origin, T-cell counts were low the first 6 months after NMSCT. Moreover, contrary with CD8 depletion of the graft that only affects CD8+ lymphocyte counts, CD34 selection dramatically decreased both CD8 and CD4 counts.     

High-dose chemotherapy followed by peripheral blood stem cell transplantation in advanced extragonadal germ cell tumor--a case report]

We reported the experience of high-dose chemotherapy (HDC) combined with peripheral stem cell transplantation (PBSCT) in 29 years-old man with advanced retroperitoneal germ cell tumor accompanied with left supraclavicular lymph node metastases, who obtained complete remission after comprehensive treatment. The initial levels of serum AFP, hCG and beta-hCG were high at 30.2 ng/ml, 14,000 mIU/ml and 66 ng/ml, respectively. After 3 courses of chemotherapy (BEP regimen), while left supraclavicular lymph node swelling was disappeared, the retroperitoneal mass lesion persisted on CT scan. Not all of 3 markers fell to the normal range. After myelosuppressive chemotherapy (etoposide 500 mg/m2 Day 1-3), PBSCs were collected by two consecutive apheresises on Day 17 and 18. In total, 19.5 x 10(6)/kg CD 34 positive cells were obtained. The patient underwent PBSCT (all CD 34 positive cells were infused) on Day 0 following HDC (CBDCA 250 mg/m2/day, etoposide 300 mg/m2/day, IFM 1.5 g/m2/day, Day-7(-)-3, respectively). He became severely leukopenic and thrombopenic with nadir of 200/microliter on Day 6 and 2 x 10(4)/microliter on Day 2, respectively. By administration of platelet transfusion and G-CSF, the white blood cell counts and thrombocyte counts recovered to 6,400/microliter and 4.1 x 10(4)/microliter on Day 10, respectively. Microbiologically enterocolic and respiratory tract infections occurred with elevated body temperature (> 40 degrees C). Antibiotic and antimycotic treatments were continued until disappearance of all clinical and microbiological evidence. He was kept for 10 days in clean room. After HDC, all markers fell to the normal range, but the retroperitoneal residual mass still persisted. Resection of the residual mass and retroperitoneal lymph node dissection were performed with pathological examination revealing tissue necrosis without viable cell. The patient has survived with no sign of the disease for 9 months.     

动员自体骨髓干细胞对大鼠急性肾小管坏死的治疗

目的：观察粒细胞集落刺激因子联合干细胞因子动员骨髓干细胞的作用、骨髓干细胞是否具有向损伤肾组织归巢的能力及其在肾脏组织中的分布,初步探讨粒细胞集落刺激因子联合干细胞因子是否具有促进急性肾小管坏死修复的作用。方法：160只8～10周龄雄性SD大鼠随机分为4组：对照组,模型组、G-CSF＋SCF治疗组、G-CSF＋SCF对照组,检测：（1）外周血白细胞总数及单个核细胞中CD34＋细胞百分比的变化;（2）尿NAG酶检测;（3）肾脏组织病理学改变;（4）肾组织CD34＋细胞表达变化。结果：（1）G-CSF＋SCF治疗组和G-CSF＋SCF对照组外周血中白细胞数、CD34＋细胞百分比于第5天达高峰,与对照组、模型组相比,差异有统计学意义（P〈0.05）,以后逐渐下降;相应地,G-CSF＋SCF治疗组肾组织内CD34＋细胞较对照组、模型组也明显增多（P〈0.05）。（2）手术后第5、10、17天,G-CSF＋SCF治疗组尿NAG酶、肾脏病理学改变均明显好于模型组（P〈0.05）。第24天G-CSF＋SCF治疗组尿NAG酶、肾脏病理学改变基本恢复正常,而模型组仍异常。第31天各组间尿NAG酶、肾脏病理学改变其差异无统计学意义。结论：（1）粒细胞集落刺激因子和干细胞因子联合应用对缺血再灌注损伤诱发急性肾小管坏死大鼠的骨髓干细胞有显著的动员作用。（2）骨髓干细胞能在损伤的肾小管归巢和定居,并可能参与损伤肾组织的修复。（3）粒细胞集落刺激因子和干细胞因子联合应用能在一定程度上加速急性肾小管坏死后肾功能的修复。     

自体外周血干细胞移植治疗患者下肢动脉缺血性疾病的随访研究

目的对16例自体外周血干细胞（PBSC）移植治疗下肢动脉缺血性溃疡的患者进行长期的随访。方法自2004年3月至2007年2月，我科利用自体外周血干细胞技术治疗了16例患有严重下肢动脉缺血性溃疡的患者，对患者进行长期的包括肢体疼痛、活动情况、下肢动脉磁共振血管成像（MRA）等随访检查。使用G—CSF5-6天动员患者骨髓干细胞进入周围循环，第六天使用细胞分离器采集周围血干细胞，然后肌注到患肢缺血的肌肉内。结果移植后经过10～45个月的追踪随访，15例患者静息痛缓解，11例溃疡愈合，4例失访，间歇性跛行时间及距离延长。结论通过对白体外周血干细胞移植治疗患者下肢动脉缺血性溃疡的随访显示，应用自体外周血干细胞移植技术治疗下肢动脉缺血性疾病是可行且有效的，是治疗肢体缺血性疾病的一种新手段，值得进一步研究和探讨。     

Identification of hematopoietic cell populations from the dermal papillae of human hair follicles

Hematopoietic stem cell (HSC) activity has been identified from the hair follicles (HFs) in mice; however, it has not been identified in human HFs. We used immunohistochemistry and flow cytometry to identify cultured dermal papilla (DP) cells expressing CD45 to test for hematopoietic activity in colony-forming assays of granulocyte/macrophage hematopoietic progenitors (CFU-GM). Occasional CD45-positive cells were detected in cultured DP cells. After in vitro stimulation with IL-3, granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor (G-CSF) for 7 days, about 1% of the cells were CD45-positive by flow cytometry analysis, an fifty-fold expansion in cell numbers. We further examined whether mesenchymal stem/progenitor cells reside in human dermal papillae. Cultured DP papilla cells incubated with monoclonal antibodies to remove the CD45 positive cells were induced into multilineage differentiation with the formation of CFU-GM. Our findings preliminarily indicate that human dermal papilla contain at least a CD45-positive hematopoietic cell population and a mesenchymal stem/progenitor cell population.