Expansion and Multipotencial Differentiation of Mesenchymal Stem Cells Isolated from Patients after High Dose Chemotherapy.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 4259-4259
Author(s):  
Karen L. Prata ◽  
Maristela D. Orellana ◽  
Aparecida M. Fontes ◽  
Karina R. Solano ◽  
Simone Kashima ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
High Dose Chemotherapy ◽  
High Dose ◽  
Cd34 Cells ◽  
Significant Difference ◽  
Cell Cycle Status ◽  
Supportive Function

Abstract Background. High dose chemotherapy (HDCT) followed by autologous PBSC rescue has been increasingly used for the treatment of several human diseases. However, little is known on the extent of this therapy on the marrow mesenchymal stem cells (MSCs). Aims. To evaluate the feasibility of expansion and multipotencial differentiation of MSCs isolated from patients after HDCT. Patients and Methods. Twelve lymphoma’s patients (LP) free of disease in bone marrow (BM) were enrolled in the study. They were submitted to BEAM’s protocol with autologous PBSC rescue 28 to 1836 days before the sample collection. Six normal bone marrow donors (ND) were used as controls. The LP and ND median age were 37.5 (range 22–49) and 31.5 years old (range 23–42), respectively. MSCs were isolated by plastic adherence and expanded ex vivo by cultivation in flasks with α-MEM with 15% fetal bovine serum. Media was changed every 3–4 days. At 90% confluence, the cells were re-plated and expanded. The isolation efficiency, colony-forming unit-fibroblast (CFU-F) frequency, growth kinetics, phenotypic characteristics, cell cycle status, multi-lineage differentiation capacity as well as hematopoiesis-supportive function were determined and compared with those of ND-MSCs. This study protocol and the consent form were approved by the institution ethics committees. Results. The results were analyzed by Mann-Whitney test and are expressed as median (range) to LP and ND, respectively. MSCs were successful isolated from all BM samples collected for this study. The cell population showed typical fibroblast-like morphology, appearing as an adherent, spindle shaped cell layer and growing to confluence after a few weeks of culture. The number of CFU-F found at 14 days of culture were 0.94 (0.00–3.75) and 1.25 (0.13–9.25) x10−5 nucleated cells (p = 0.4421). The doubling time between the 1st and 2nd passages was 80.66 (34.08–195.35) and 46.30 (36.36–270.59) hours (p = 0.1025). The cell clones proliferated extensively until 8.17 (1.81–28.27) and 18.11 (11.85–27.48) population doublings (p = 0.0668) in 71.50 (46–88) and 81 (57–103) cultivation days (p = 0.1505). Immunophenotypically, these cells were positive for the CD73, CD105, CD90, CD29, CD13, CD44, CD49e, CD54, HLA-class 1 and Stro-1 markers and negative for CD34, CD45, CD14, CD51/61, HLA-DR and KDR. Regarding the cell cycle status, 85.63 (63.19–92.17) and 82.41 (82.19–87.02) % were in GO-G1 phase (p = 1,000), while only 12.17 (3.33–36.81) and 10.67 (6.59–12.05) % were in S phase (p = 0,6828). All samples tested were capable of differentiating along adipogenic, osteogenic and chondrogenic lineages in vitro, demonstrated by morphology, cyto- and imunohistochemistry or RT-PCR reaction (PPARg and osteopontin genes expression). After co-culture with CD34+ cord blood cells for 1 and 4 weeks, no significant difference CD34+ expansion or colony-forming cells (BFU-E or CFU-GM) were observed between the CD34+ cells/LP-MSCs and CD34+ cells/ND-MSCs co-cultures with cytokines or not. Interpretation and Conclusions. Our results demonstrate that is possible to cultivate and expand MSCs with multipotential differentiation capabilities and hematopoiesis-supportive function from patients after HDCT. Despite there were no significant differences in the median values between LP and ND, the comparative study indicates a possible damage in MSCs by HDCT.

Study on Telomerase Activity of Human Bone Marrow Mesenchymal Stem Cells.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4255-4255
Author(s):  
Jingyuan Li ◽  
Xiaoyu Lai ◽  
Huang He
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Telomerase Activity ◽  
Negative Control ◽  
Human Bone ◽  
Human Bone Marrow ◽  
Significant Difference

Abstract Human mesenchymal stem cells(hMSCs) have multiple differentiate potential, and it can differentiate into adipocytes, osteogenic cells, chondrocyte and neural cells et al. It has been reported that telomerase activity in hMSCs is negative, but it is still controversial and telomerase activity in hMSCs-derived adipocytes has not been reported. We investigate the telomerase activity in hMSCs before and after their committed differentiation into adipocytes in vitro and cryopreservation. hMSCs were isolated from normal human bone marrow fellowed by cell culture in DMEM with low glucose containing 10% FBS. The FACS was performed to examine the expression of cell surface molecules and analyze cell cycle of primary hMSCs.Then some of hMSCs were induced to differentiate into adipocytes in vitro by being treated with adipocytic medium fellowed by being stained with oil red O, and the others were cryopreserved in liguid nitrogon for three months. TRAP assay(telomerase repeat amplification protocol assay)was employed to detect telomerase activity in those hMSCs. T293 cells and α-Interferon were analyzed with each test as an additional positive control and negative control respectively. Telomerase activity was expressed as a percentage of the relative telomerase activity (RTA) of the hMSCs relative to the RTA of T293 cells. The results indicated the cells were positive for SH2, SH3, CD90 and negative for CD34, CD45, AC133. It was showed that the majority of primary hMSCs(85%) was at cell cycle of G0/G1 phase and the minority of hMSCs was at S, G2 or M phase. 80% hMSCs was orange adipocytes after they were treated with adipocytic medium for 3–4weeks. Telomerase activity was negative in hMSCs both at the beginning of culture and at the later stages during cell expansion,telomerase activity in hMSCs-passage 1–3(n=10) and hMSCs-passage 4–7(n=9) made no significant difference(1.46±0.83% vs 1.46±0.67%, p=0.99). Cryopreservation did not affect the telomerase activity in hMSCs. Telomerase activity in fresh hMSCs(n=13) and frozen hMSCs(n=6) made no significant difference(1.41±0.44% vs 1.51±1.07%, p=0.64). Telomerase activity in hMSCs-derived adipocytes(n=3) was significantly higher than in hMSCs(n=19)( 11.8±2.52% vs 1.46. ±0.67%, p<0.00001). It is concluded that hMSCs are telomerase-negative, and the stage of culture or cryopreservation does not affect their telomerase activity. After being induced to differentiated into adipocytes, hMSCs telomerase activity is upregulated.

Rapid Hematopoietic Recovery After Coinfusion of Autologous-Blood Stem Cells and Culture-Expanded Marrow Mesenchymal Stem Cells in Advanced Breast Cancer Patients Receiving High-Dose Chemotherapy

2000 ◽  
Vol 18 (2) ◽  
pp. 307-307 ◽  
Author(s):  
Omer N. Koç ◽  
Stanton L. Gerson ◽  
Brenda W. Cooper ◽  
Stephanie M. Dyhouse ◽  
Stephen E. Haynesworth ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Cancer Patients ◽  
High Dose Chemotherapy ◽  
High Dose ◽  
Human Mscs ◽  
Breast Cancer Patients ◽  
Hematopoietic Recovery

PURPOSE: Multipotential mesenchymal stem cells (MSCs) are found in human bone marrow and are shown to secrete hematopoietic cytokines and support hematopoietic progenitors in vitro. We hypothesized that infusion of autologous MSCs after myeloablative therapy would facilitate engraftment by hematopoietic stem cells, and we investigated the feasibility, safety, and hematopoietic effects of culture-expanded MSCs in breast cancer patients receiving autologous peripheral-blood progenitor-cell (PBPC) infusion. PATIENTS AND METHODS: We developed an efficient method of isolating and culture-expanding a homogenous population of MSCs from a small marrow-aspirate sample obtained from 32 breast cancer patients. Twenty-eight patients were given high-dose chemotherapy and autologous PBPCs plus culture-expanded MSC infusion and daily granulocyte colony-stimulating factor. RESULTS: Human MSCs were successfully isolated from a mean ± SD of 23.4 ± 5.9 mL of bone marrow aspirate from all patients. Expansion cultures generated greater than 1 × 106 MSCs/kg for all patients over 20 to 50 days with a mean potential of 5.6 to 36.3 × 106 MSCs/kg after two to six passages, respectively. Twenty-eight patients were infused with 1 to 2.2 × 106 expanded autologous MSCs/kg intravenously over 15 minutes. There were no toxicities related to the infusion of MSCs. Clonogenic MSCs were detected in venous blood up to 1 hour after infusion in 13 of 21 patients (62%). Median time to achieve a neutrophil count greater than 500/μL and platelet count ≥ 20,000/μL untransfused was 8 days (range, 6 to 11 days) and 8.5 days (range, 4 to 19 days), respectively. CONCLUSION: This report is the first describing infusion of autologous MSCs with therapeutic intent. We found that autologous MSC infusion at the time of PBPC transplantation is feasible and safe. The observed rapid hematopoietic recovery suggests that MSC infusion after myeloablative therapy may have a positive impact on hematopoiesis and should be tested in randomized trials.

scholarly journals High-Dose 111In Induces G1 Cell Cycle Arrest and Cell Death in Rat Bone Marrow Mesenchymal Stem Cells

2012 ◽  
Vol 46 (2) ◽  
pp. 81-88 ◽  
Author(s):  
Bok-Nam Park ◽  
Wooyoung Shim ◽  
Young Hwan Ahn ◽  
Jae-Ho Lee ◽  
Young-Sil An ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Bone Marrow ◽  
Cell Death ◽  
Mesenchymal Stem Cells ◽  
High Dose ◽  
Cycle Arrest ◽  
Marrow Mesenchymal Stem Cells ◽  
G1 Cell Cycle Arrest ◽  
Rat Bone

scholarly journals Bone marrow mesenchymal stem cells promote remyelination in spinal cord by driving oligodendrocyte progenitor cell differentiation via TNFα/RelB-Hes1 pathway: a rat model study of 2,5-hexanedione-induced neurotoxicity

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Shuangyue Li ◽  
Huai Guan ◽  
Yan Zhang ◽  
Sheng Li ◽  
Kaixin Li ◽  
...  
Keyword(s):  
Spinal Cord ◽  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Statistical Significance ◽  
High Dose ◽  
Sprague Dawley ◽  
Oligodendrocyte Progenitor ◽  
Hes1 Expression ◽  
Marrow Mesenchymal Stem Cells

Abstract Background N-hexane, with its metabolite 2,5-hexanedine (HD), is an industrial hazardous material. Chronic hexane exposure causes segmental demyelination in the peripheral nerves, and high-dose intoxication may also affect central nervous system. Demyelinating conditions are difficult to treat and stem cell therapy using bone marrow mesenchymal stem cells (BMSCs) is a promising novel strategy. Our previous study found that BMSCs promoted motor function recovery in rats modeling hexane neurotoxicity. This work aimed to explore the underlying mechanisms and focused on the changes in spinal cord. Methods Sprague Dawley rats were intoxicated with HD (400 mg/kg/day, i.p, for 5 weeks). A bolus of BMSCs (5 × 107 cells/kg) was injected via tail vein. Demyelination and remyelination of the spinal cord before and after BMSC treatment were examined microscopically. Cultured oligodendrocyte progenitor cells (OPCs) were incubated with HD ± BMSC-derived conditional medium (BMSC-CM). OPC differentiation was studied by immunostaining and morphometric analysis. The expressional changes of Hes1, a transcription factor negatively regulating OPC-differentiation, were studied. The upstream Notch1 and TNFα/RelB pathways were studied, and some key signaling molecules were measured. The correlation between neurotrophin NGF and TNFα was also investigated. Statistical significance was evaluated using one-way ANOVA and performed using SPSS 13.0. Results  The demyelinating damage by HD and remyelination by BMSCs were evidenced by electron microscopy, LFB staining and NG2/MBP immunohistochemistry. In vitro cultured OPCs showed more differentiation after incubation with BMSC-CM. Hes1 expression was found to be significantly increased by HD and decreased by BMSC or BMSC-CM. The change of Hes1 was found, however, independent of Notch1 activation, but dependent on TNFα/RelB signaling. HD was found to increase TNFα, RelB and Hes1 expression, and BMSCs were found to have the opposite effect. Addition of recombinant TNFα to OPCs or RelB overexpression similarly caused upregulation of Hes1 expression. The secretion of NGF by BMSC and activation of NGF receptor was found important for suppression of TNFα production in OPCs. Conclusions  Our findings demonstrated that BMSCs promote remyelination in the spinal cord of HD-exposed rats via TNFα/RelB-Hes1 pathway, providing novel insights for evaluating and further exploring the therapeutical effect of BMSCs on demyelinating neurodegenerative disease.

scholarly journals Melatonin Inhibits the Ferroptotic Pathway in Rat Bone Marrow Mesenchymal Stem Cells by Activating the PI3K-AKT-mTOR Signaling Axis to Attenuate Steroid-induced Osteoporosis

2021 ◽  
Author(s):  
meng li ◽  
ning yang ◽  
li hao ◽  
wei zhou ◽  
lei li ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Mammalian Target Of Rapamycin ◽  
Mtor Signaling ◽  
High Dose ◽  
Treatment Pathways ◽  
Marrow Mesenchymal Stem Cells

Abstract ObjectivesSteroid-induced osteoporosis (SIOP) is a secondary osteoporosis, which is a systemic bone disease characterized by low bone mass, bone microstructure damage, increased bone fragility, and easy fracture. However, the specific mechanism remains unclear. Glucocorticoid-induced death of osteoblasts and bone marrow mesenchymal stem cells (BMSCs) is an important factor in SIOP. Ferroptosis is an iron-dependent programmed cell death that differs from apoptosis, cell necrosis, and autophagy, which can be induced by many factors. Herein, we aimed to explore whether glucocorticoids (GCs) cause ferroptosis in BMSCs and determine possible treatment pathways and mechanisms of action. Melatonin (MT), a hormone secreted by the pineal gland, displays strong antioxidant abilities to scavenge free radicals and alleviates ferroptosis in many tissues and organs. MethodsIn this study, we used high-dose dexamethasone (DEX) to observe whether glucocorticoids induced ferroptosis in BMSCs. We then assessed whether MT can inhibit the ferroptotic pathway, thereby providing early protection against GC-induced SIOP, and investigated the signaling pathways involved.ResultsIn vitro experiments showed that MT intervention significantly improved GC-induced ferroptosis in BMSCs and significantly improved SIOP in vivo. Pathway analysis showed that MT improves ferroptosis by activating the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) axis. MT upregulates expression of PI3K, which is an important regulator of ferroptosis resistance. PI3K activators mimic the anti-ferroptosis effect of MT, but after blocking the PI3K pathway, the effect of MT is weakened. Obviously, MT can protect against SIOP induced by GC. Notably, even after GC-induced ferroptosis begins, MT can confer protection against SIOP. ConclusionOur research confirms that GC-induced ferroptosis is closely related to SIOP. Melatonin can inhibit ferroptosis by activating the PI3K-AKT-mTOR signaling pathway, thereby reducing the occurrence of steroid-induced osteoporosis. Therefore, MT may provide a novel strategy for preventing and treating SIOP.

scholarly journals Phase I Trial of Intra-arterial Administration of Autologous Bone Marrow-Derived Mesenchymal Stem Cells in Patients with Multiple System Atrophy

2021 ◽  
Vol 2021 ◽  
pp. 1-10
Author(s):  
Seok Jong Chung ◽  
Tae Yong Lee ◽  
Yang Hyun Lee ◽  
KyoungWon Baik ◽  
Jin Ho Jung ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Multiple System Atrophy ◽  
Phase I ◽  
Dose Group ◽  
Low Dose ◽  
Autologous Bone Marrow ◽  
High Dose ◽  
Autologous Bone

Background. This study is aimed at investigating the safety and tolerability of the intra-arterial administration of autologous bone marrow-derived mesenchymal stem cells (BM-MSCs) in patients with multiple system atrophy- (MSA-) cerebellar type (MSA-C). Methods. This was a single-center, open-label phase I clinical trial in patients with MSA-C. A three-stage dose escalation scheme (low-dose, 3.0 × 10 5 cells/kg; medium-dose, 6.0 × 10 5 cells/kg; high-dose, 9.0 × 10 5 cells/kg) was applied to determine the maximum tolerated dose of intra-arterial administration of BM-MSCs based on the no-observed-adverse-effect level derived from the toxicity study. The occurrence of adverse events was evaluated 1 day before and 1, 14, and 28 days after BM-MSC therapy. Additionally, we assessed changes in the Unified MSA Rating Scale (UMSARS) score 3 months after BM-MSC treatment. Results. One serious adverse drug reaction (ADR) of leptomeningeal enhancement following the intra-arterial BM-MSC administration occurred in one patient in the low-dose group. The safety review of the Internal Monitoring Committee interpreted this as radiological evidence of the blood-brain barrier permeability for MSCs. No other ADRs were observed in the medium- or high-dose groups. In particular, no ischemic lesions on diffusion-weighted images were observed in any of the study participants. Additionally, the medium- and high-dose groups tended to show a slower increase in UMSARS scores than the low-dose group during the 3-month follow-up. Conclusion. The present study confirmed that a single intra-arterial administration of autologous BM-MSCs is a safe and promising neuroprotective strategy in patients with MSA-C.

scholarly journals PTPN21 Overexpression Promotes Osteogenic and Adipogenic Differentiation of Bone Marrow-Derived Mesenchymal Stem Cells but Inhibits the Immunosuppressive Function

2019 ◽  
Vol 2019 ◽  
pp. 1-19 ◽  
Author(s):  
Huafang Wang ◽  
Xiaohang Ye ◽  
Haowen Xiao ◽  
Ni Zhu ◽  
Cong Wei ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Bone Marrow ◽  
Endothelial Cells ◽  
Mesenchymal Stem Cells ◽  
Tumor Cells ◽  
Vascular Endothelial Cells ◽  
Adipogenic Differentiation ◽  
Target Cells ◽  
Vascular Endothelial

Protein tyrosine phosphatases (PTPs) act as key regulators in various cellular processes such as proliferation, differentiation, and migration. Our previous research demonstrated that non-receptor-typed PTP21 (PTPN21), a member of the PTP family, played a critical role in the proliferation, cell cycle, and chemosensitivity of acute lymphoblastic leukemia cells. However, the role of PTPN21 in the bone marrow microenvironment has not yet been elucidated. In the study, we explored the effects of PTPN21 on human bone marrow-derived mesenchymal stem cells (BM-MSCs) via lentiviral-mediated overexpression and knock-down of PTPN21 in vitro. Overexpressing PTPN21 in BM-MSCs inhibited the proliferation through arresting cell cycle at the G0 phase but rendered them a higher osteogenic and adipogenic differentiation potential. In addition, overexpressing PTPN21 in BM-MSCs increased their senescence levels through upregulation of P21 and P53 and dramatically changed the levels of crosstalk with their typical target cells including immunocytes, tumor cells, and vascular endothelial cells. BM-MSCs overexpressing PTPN21 had an impaired immunosuppressive function and an increased capacity of recruiting tumor cells and vascular endothelial cells in a chemotaxis transwell coculture system. Collectively, our data suggested that PTPN21 acted as a pleiotropic factor in modulating the function of human BM-MSCs.

Transplantation of Peripheral Blood Stem Cells Mobilized by Chemotherapy and Single Dose Pegylated G-CSF in Patients with Multiple Myeloma: Equivalence of 6 mg and 12 mg Pegfilgrastim.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2868-2868 ◽  
Author(s):  
Ingmar Bruns ◽  
Ulrich Steidl ◽  
Christof Scheid ◽  
Kai Hübel ◽  
Roland Fenk ◽  
...  
Keyword(s):  
Stem Cells ◽  
Multiple Myeloma ◽  
Stem Cell ◽  
Single Dose ◽  
Peripheral Blood ◽  
High Dose Chemotherapy ◽  
High Dose ◽  
Cytotoxic Therapy ◽  
Cd 34 ◽  
Cd34 Cells

Abstract To date the most effective treatment for patients (pts) with multiple myeloma consists of conventional induction chemotherapy followed by either single or tandem high-dose chemotherapy and autologous blood stem cell transplantation. Collection of sufficient numbers of hematopoietic stem cells is essential for high-dose chemotherapy. Current regimens for stem cell mobilization are based on daily subcutaneous injections of human recombinant G-CSF starting shortly after cytotoxic therapy. Here we examined the use of polyethyenglycole (PEG)-conjugated G-CSF (pegfilgrastim) at two different doses in patients with stage II or III multiple myeloma. Patients received induction therapy with 2–4 cycles ID or VAD. Following cytotoxic therapy with cyclophosphamide (4g/m2) we administered either a single dose of 6 mg pegfilgrastim (n=10 pts; median age: 55 years), 12 mg pegfilgrastim (n=12 pts; median age: 51 years) or daily doses of 8,5 μg/kg unconjugated G-CSF (filgrastim) (n=12 pts; median age: 51 years). The growth factor was given on day 4 (range 2–5 days) in the “6 mg pegfilgrastim group”, on day 5 (range 2–7 days) in the “12 mg pegfilgrastim group” and on day 4 (range 3–6 days) in the “filgrastim group” after cyclophosphamide. Numbers of CD34+ cells were determined during leukocyte recovery and harvested by large volume apheresis using a cobe spectra blood cell separator. Pegfilgratim was associated with an earlier leukocyte recovery both at the 6mg dose (median 12 days, range 8–16 days) and the 12mg dose (median 12 days, range 7–16 days) as compared to filgrastim (median 14 days, range 11–15 days, p=0.04). Similarily, the peripheral blood CD34+ cell peak occurred earlier in patients who received pegfilgrastim (median 12 days, range 11–18 days versus median 15 days, range 12–18). On the other hand the peripheral blood CD 34+ peak did not differ significantly between the three groups (median 129/μl with 6 mg pegfilgrastim, range 30–433, median 78/μl with 12 mg pegfilgrastim, range 20– 1055 and median 111/μl with filgrastim, range 28–760, p=0.95). With a median of 1.0x10E7 CD34+ cells per kg (range 5.8x10E6-1.9x10E7) in the “6 mg pegfilgrastim group”, 7.4x10E6 CD34+ cells per kg (median, range 4.9x10E6- 3.8x10E7) in the “12 mg pegfilgrastim group” and 10.8x10E6 CD34+ cells per kg (median, range 5.0x10E6-8.7x10E7) in the “filgrastim group” there were no significant differences in the total number of harvested CD34+ cells. Following high-dose therapy with melphalan (200 mg/m2) and autografting leukocyte and platelet reconstitution was similar within all groups. In summary, a single dose of pegfilgrastim after high dose cyclophosphamide is capable of mobilizing a sufficient number of CD 34+ cells for succesful autografting and sustained hematological reconstitution in patients with multiple myeloma. No difference could be observed between 6 mg and 12 mg of pegfilgrastim. Our data provide the basis for randomized studies evaluating the optimal dose and timing of pegfilgrastim as well as long-term outcome in larger cohorts of patients.

Impact of Disease and Mobilizing Agents on Initial and Remobilization Failure.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 5222-5222
Author(s):  
John F. DiPersio ◽  
Angela Smith ◽  
Dianne Sempek ◽  
Albert Baker ◽  
Steven Jiang ◽  
...  
Keyword(s):  
Failure Rate ◽  
Surrogate Marker ◽  
High Dose Chemotherapy ◽  
Disease State ◽  
High Dose ◽  
Failure Rates ◽  
Gm Csf ◽  
Scheffe Test ◽  
Cd34 Cells ◽  
Significant Difference

Abstract Background: High-dose chemotherapy with autologous stem cell transplantation (ASCT) is a widely used treatment strategy in lymphoma and myeloma; however, no standard approach for the mobilization of peripheral hematologic stem and progenitor cells (HSPCs) has been established. Levels of circulating CD34+ cells, a surrogate marker for mobilization efficiency, vary widely between pts, and may be influenced by disease state, prior therapy, and/or mobilization regimen. Methods: The Washington University (St. Louis, MO) transplantation database includes clinical parameters from 407 multiple myeloma (MM), 562 non-Hodgkin’s Lymphoma (NHL), and 164 Hodgkin’s disease (HD) pts who received an ASCT between 1995 and 2006. A retrospective analysis of this large (1133 pts) population was conducted to determine factors associated with mobilization efficiency. Mobilization failure was defined as collection of < 2 × 10^6 CD34+ cells/kg within 5 apheresis days. Statistical analysis included analysis of variance (ANOVA) with Scheffe Test to determine differences in mobilization between the various mobilization regimens (G-CSF, G-CSF/chemotherapy, G-/GM-CSF, G-CSF/AMD3100). Results: All pts were included in the analysis; 87% received G-CSF alone as the initial mobilization regimen. Mobilization failure rates are summarized in Table 1. NHL and HD pts had an approx. 4-fold higher failure rate than MM pts. The combination of G-CSF with chemotherapy increased the median CD34+ yield compared to G-CSF alone, although no obvious impact on the failure rate was noted in this relatively small group of pts. Remobilization was associated with high failure rates in NHL (79.2%), HD (77.1%), and MM (73.3%). Pooled collections were <2 × 10^6 CD34+/kg in 33.6%, 37.1%, and 36.7% of failed mobilizers, respectively. ANOVA analysis indicated a significant difference in outcome based on remobilization regimen. A post hoc comparison using the Scheffe Test determined that G-CSF mobilization failures remobilized with G-CSF plus AMD3100 collected significantly more CD34+ cells than G-CSF-failures remobilized with either G-CSF, G/GM-CSF or G-CSF/chemo (1-way ANOVA: F(3, 233) = 27.878, F0.5(3, 233).05 = 2.643, p < .0001). The compared groups did not significantly differ in initial mobilization efficiency with G-CSF (as determined by ANOVA and Scheffe Test). Conclusions: The mobilization failure rate is substantially higher in NHL and HD pts than MM pts. Pts who fail initial mobilization are highly likely to fail a 2nd mobilization, regardless of disease state. As the combination of chemotherapy to G-CSF may not be sufficient to reduce failure rates, alternative mobilization strategies are needed to improve HSPC collection, particularly in NHL/HD pts and failed mobilizers. First mobilization failure rates (< 2×10^6 CD34+/kg) Mobilization regimen N Failures Median yield (×10^6) 95% C.I (×10^6) * Incl. pts mobilized w. alternative regimens NHL G-CSF 471 26.5% 2.89 2.76–3.04 G-CSF/Chemo 35 22.9% 4.68 2.8–8.53 All* 564 28.7% HD G-CSF 130 26.2% 3.01 2.75–3.37 G-CSF/Chemo 12 16.7% 5.38 2.35–9.52 All* 165 24.8% MM G-CSF 386 6.5% 4.62 4.16–4.98 G-CSF/Chemo 17 5.9% 8.52 4.46–16.3 All* 409 6.6%

Hematopoiesis in the Elderly: Age-Associated Effects in Frequency, Function, and Gene Expression of Human Hematopoietic Stem Cells.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1505-1505
Author(s):  
Wendy W. Pang ◽  
Elizabeth A. Price ◽  
Irving L. Weissman ◽  
Stanley L. Schrier
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Stem Cells ◽  
Bone Marrow ◽  
Expression Profiles ◽  
Frequency Function ◽  
Hematopoietic Stem ◽  
Hematopoietic System ◽  
Cell Cycle Status

Abstract Abstract 1505 Poster Board I-528 Aging of the human hematopoietic system is associated with an increase in the development of anemia, myeloid malignancies, and decreased adaptive immune function. While the hematopoietic stem cell (HSC) population in mouse has been shown to change both quantitatively as well as functionally with age, age-associated alterations in the human HSC and progenitor cell populations have not been characterized. In order to elucidate the properties of an aged human hematopoietic system that may predispose to age-associated hematopoietic dysfunction, we evaluated and compared HSC and other hematopoietic progenitor populations prospectively isolated via fluorescence activated cell sorting (FACS) from 10 healthy young (20-35 years of age) and 8 healthy elderly (65+ years of age) human bone marrow samples. Bone marrow was obtained from hematologically normal young and old volunteers, under a protocol approved by the Stanford Institutional Review Board. We determined by flow cytometry the distribution frequencies and cell cycle status of HSC and progenitor populations. We also analyzed the in vitro function and generated gene expression profiles of the sorted HSC and progenitor populations. We found that bone marrow samples obtained from normal elderly adults contain ∼2-3 times the frequency of immunophenotypic HSC (Lin-CD34+CD38-CD90+) compared to bone marrow obtained from normal young adults (p < 0.02). Furthermore, upon evaluation of cell cycle status using RNA (Pyronin-Y) and DNA (Hoechst 33342) dyes, we observed that a greater percentage of HSC from young bone marrow are in the quiescent G0- phase of the cell cycle compared to elderly HSC, of which there is a greater percentage in G1-, S-, G2-, or M-phases of the cell cycle (2.5-fold difference; p < 0.03). In contrast to the increase in HSC frequency, we did not detect any significant differences in the frequency of the earliest immunophenotypic common myeloid progenitors (CMP; Lin-CD34+CD38+CD123+CD45RA-), granulocyte-macrophage progenitors (GMP; Lin-CD34+CD38+CD123+CD45RA+), and megakaryocytic-erythroid progenitors (MEP; Lin-CD34+CD38+CD123-CD45RA-) from young and elderly bone marrow. We next analyzed the ability of young and elderly HSC to differentiate into myeloid and lymphoid lineages in vitro. We found that elderly HSC exhibit diminished capacity to differentiate into lymphoid B-lineage cells in the AC6.21 culture environment. We did not, however, observe significant differences in the ability of young and elderly HSC to form myeloid and erythroid colonies in methylcellulose culture, indicating that myelo-erythroid differentiation capacity is preserved in elderly HSC. Correspondingly, gene expression profiling of young and elderly human HSC indicate that elderly HSC have up-regulation of genes that specify myelo-erythroid fate and function and down-regulation of genes associated with lymphopoiesis. Additionally, elderly HSC exhibit increased levels of transcripts associated with transcription, active cell-cycle, cell growth and proliferation, and cell death. These data suggest that hematopoietic aging is associated with intrinsic changes in the gene expression of human HSC that reflect the quantitative and functional alterations of HSC seen in elderly bone marrow. In aged individuals, HSC are more numerous and, as a population, are more myeloid biased than young HSC, which are more balanced in lymphoid and myeloid potential. We are currently investigating the causes of and mechanisms behind these highly specific age-associated changes in human HSC. Disclosures: Weissman: Amgen: Equity Ownership; Cellerant Inc.: ; Stem Cells Inc.: ; U.S. Patent Application 11/528,890 entitled “Methods for Diagnosing and Evaluating Treatment of Blood Disorders.”: Patents & Royalties.

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