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Background: The DNA-repair enzyme Artemis is essential for rearrangement of T- and B-cell receptors. Mutations in DCLRE1C , which encodes Artemis, cause Artemis-deficient severe combined immunodeficiency (ART-SCID), which is poorly responsive to allogeneic hematopoietic-cell transplantation., Methods: We carried out a phase 1-2 clinical study of the transfusion of autologous CD34+ cells, transfected with a lentiviral vector containing DCLRE1C , in 10 infants with newly diagnosed ART-SCID. We followed them for a median of 31.2 months., Results: Marrow harvest, busulfan conditioning, and lentiviral-transduced CD34+ cell infusion produced the expected grade 3 or 4 adverse events. All the procedures met prespecified criteria for feasibility at 42 days after infusion. Gene-marked T cells were detected at 6 to 16 weeks after infusion in all the patients. Five of 6 patients who were followed for at least 24 months had T-cell immune reconstitution at a median of 12 months. The diversity of T-cell receptor β chains normalized by 6 to 12 months. Four patients who were followed for at least 24 months had sufficient B-cell numbers, IgM concentration, or IgM isohemagglutinin titers to permit discontinuation of IgG infusions. Three of these 4 patients had normal immunization responses, and the fourth has started immunizations. Vector insertion sites showed no evidence of clonal expansion. One patient who presented with cytomegalovirus infection received a second infusion of gene-corrected cells to achieve T-cell immunity sufficient for viral clearance. Autoimmune hemolytic anemia developed in 4 patients 4 to 11 months after infusion; this condition resolved after reconstitution of T-cell immunity. All 10 patients were healthy at the time of this report., Conclusions: Infusion of lentiviral gene-corrected autologous CD34+ cells, preceded by pharmacologically targeted low-exposure busulfan, in infants with newly diagnosed ART-SCID resulted in genetically corrected and functional T and B cells. (Funded by the California Institute for Regenerative Medicine and the National Institute of Allergy and Infectious Diseases; ClinicalTrials.gov number, NCT03538899.)., (Copyright © 2022 Massachusetts Medical Society.)

Background: Hematopoietic stem cell transplantation (HSCT) is one of the treatments for hematologic malignancies. Numerous factors affect the HSCT outcome. The purpose of this study was to investigate the effect of post-HSCT administration of granulocyte colony-stimulating factor (post-G-CSF) on early neutrophil and platelet engraftment in allogeneic HSCT (allo-HSCT)., Material & Methods: The study was performed on 76 patients diagnosed with AML and ALL. All patients underwent allo-HSCT at Taleghani stem cell transplantation center, Tehran, Iran, from February 2016 to December 2018. Chemotherapy regimens based on patients' conditions were selected between myeloablative and reduced-intensity regimens., Results: Statistical analysis revealed that the number of administered G-CSF units after HSCT was a time-dependent variable. Statistical analysis before day +11 reported that patients who received G-CSF <14 units had three times better early neutrophil engraftment than those with G-CSF ≥14 (CI 95%, AHR = 3.03, p:0.002). CD3+ cells count <318.5 × 10 6 /kg was associated with fast platelet engraftment (CI 95%, AHR 2.28, p:0.01)., Conclusion: In this study, post-G-CSF stimulation was associated with early engraftment in a time- and dose-dependent manner. Administration of G-CSF beyond 14 units resulted in adverse effects on neutrophil early engraftment. It also appeared that with a reduction in CD3+ cell counts, the likelihood of GVHD decreases, and platelet engraftment occurs earlier. Further investigations in the future are required to determine the factors affecting the process of early engraftment., (© 2021 The Authors. Journal of Clinical Laboratory Analysis published by Wiley Periodicals LLC.)

Background/aims: To describe the clinicopathological and immunohistochemical characteristics of 10 patients representing a new entity of benign conjunctival myxoid stromal tumours., Methods: Retrospective review of clinical findings, histopathological and immunohistochemical studies identified 10 cases of low-grade conjunctival myxoid stromal tumours. Specimens were routinely processed and stained with H&E. Immunohistochemical stains for CD34, CD68, vimentin, S100, smooth muscle actin (SMA), myosin, desmin, actin, Bcl-2 and Ki-67 were performed. Specific stains for Alcian-blue periodic acid-Schiff (AB-PAS) and aldehyde fuchsin stains were also performed., Results: Ten patients with an average age of 45.6±11.1 years had a tender white or faint yellow to red mass on the bulbar conjunctiva. All the lesions were completely removed, and none of the patients relapsed. Histologically, all neoplasms consisted of spindle-shaped cells that showed signs of pseudonuclear inclusions, multinuclear cells and had no atypia. The stroma consisted of a large amount of mucus and was infiltrated with delicate to ropey collagens, a few mast cells and new vessels. Immunohistochemical stains were positive for CD34, vimentin and Bcl-2; partial positive for CD68; very low for Ki-67; and negative for S100, SMA, myosin, desmin and actin. AB-PAS suggested that the stroma was mucinous., Conclusions: These rare benign mesenchymal conjunctival tumours are mostly unilateral and occur in the bulbar conjunctiva. Complete resection is the radical treatment. These lesions are characterised by multiple spindle cells, a large amount of mucus, and sharing similar basic histopathological features with conjunctival myxoma and conjunctival stromal tumour. We suggest naming these lesions 'conjunctival myxoid stromal tumours'., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2019. No commercial re-use. See rights and permissions. Published by BMJ.)

Objective: High-doses of melphalan treatment with autologous stem cell transplantation in multiple myeloma (MM) remains a major treatment modality in suitable patients. A minimal dose of 2x10 6 /kg CD34+ cells is preferred to achieve engraftment. Some patients need multiple leukapheresis procedures to achieve the necessary number of CD34+ cells, but this can cause a high volume of stem cell product that cannot be given in a single day. Whether or not the number of infusion days affects engraftment has not been studied before. We aimed to evaluate the impact of reinfusion of stem cells on multiple days on engraftment results., Materials and Methods: Demographic features, CD34+ cell doses, neutrophil and platelet engraftment days, hospitalization days, and number of infusion days of 149 autologous transplantations of 143 MM patients were evaluated retrospectively., Results: The data of 143 MM patients who were transplanted were analyzed retrospectively. Median age was 55±8.5 (range: 26-70) years with a male/female ratio of 91/58. Hospitalization days for all patients were 24±6 (range: 14-50) days. Mean CD34+ cell number was (7.5±5.3)x10 6 /kg (range: 1.5-31x10 6 /kg). CD34+ cells were reinfused in 1 day in 80.5% (n=120) of the patients, 2 days in 18.2% of the patients (n=27), and 3 days in 1.3% of the patients (n=2). For 29 patients, reinfusion was applied in more than 1 day because of the high volume of stem cell product. We did not see any dimethyl sulfoxide toxicity, cardiac arrhythmia, or volume overload complications. Hypertensive attacks during infusion were easily controlled by furosemide treatment. In the group with multiple infusions, the infused CD34+ cell numbers had a mean of (4.8±2.8)x10 6 /kg, and in the single infusion group the mean was (8.1±5.5)x10 6 /kg. There were no statistical differences between the two groups regarding platelet and neutrophil engraftment days (p=0.850, r=0.820 and p=0.500, r=0.440). There was no statistical difference between the two groups for hospitalization days (p=0.060, r=0.050)., Conclusion: In cases with a high volume of stem cell product to acquire adequate stem cells, reinfusion can be safely applied across multiple days without any delay in engraftment.

Background: We investigated a correlation between electromechanical properties of the myocardium and response to CD34+ cell therapy in patients with chronic heart failure., Methods and Results: We enrolled 40 patients with ischemic cardiomyopathy (ICM) and 40 with nonischemic dilated cardiomyopathy (DCM). All patients were in New York Heart Association functional class III and had a left ventricular ejection fraction (LVEF) <40%. CD34+ cells were mobilized by granulocyte colony-stimulating factor and collected via apheresis. Electroanatomic mapping was performed to define areas of myocardial scar and hibernation, and CD34+ cells were injected transendocardially in the hibernating areas. Patient were followed for 6 months; responders were defined as patients with LVEF increase of >5%. At baseline, the groups did not differ in sex, LVEF, creatinine, N-terminal pro-B-type natriuretic peptide or electroanatomic parameters (scar area: 53 ± 18% in ICM vs 55 ± 23% in DCM [P = .83]; hibernating area: 23 ± 13% vs 22 ± 12% [P = .56]). At 6 months we found similar rates of responders in both groups (60% in ICM vs 65% in DCM [P = .95]). When compared with nonresponders, responders had less myocardial scar (47 ± 17% vs 58 ± 15% [P = .003])., Conclusions: In patients with chronic heart failure due to ICM and DCM we observed similar electroanatomic properties of the myocardium. In both groups, lower myocardial scar burden was associated with better clinical response to CD34+ cell therapy., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Rationale: Despite direct immediate intervention and therapy, ST-segment-elevation myocardial infarction (STEMI) victims remain at risk for infarct expansion, heart failure, reinfarction, repeat revascularization, and death., Objective: To evaluate the safety and bioactivity of autologous CD34+ cell (CLBS10) intracoronary infusion in patients with left ventricular dysfunction post STEMI., Methods and Results: Patients who underwent successful stenting for STEMI and had left ventricular dysfunction (ejection fraction≤48%) ≥4 days poststent were eligible for enrollment. Subjects (N=161) underwent mini bone marrow harvest and were randomized 1:1 to receive (1) autologous CD34+ cells (minimum 10 mol/L±20% cells; N=78) or (2) diluent alone (N=83), via intracoronary infusion. The primary safety end point was adverse events, serious adverse events, and major adverse cardiac event. The primary efficacy end point was change in resting myocardial perfusion over 6 months. No differences in myocardial perfusion or adverse events were observed between the control and treatment groups, although increased perfusion was observed within each group from baseline to 6 months (P<0.001). In secondary analyses, when adjusted for time of ischemia, a consistently favorable cell dose-dependent effect was observed in the change in left ventricular ejection fraction and infarct size, and the duration of time subjects was alive and out of hospital (P=0.05). At 1 year, 3.6% (N=3) and 0% deaths were observed in the control and treatment group, respectively., Conclusions: This PreSERVE-AMI (Phase 2, randomized, double-blind, placebo-controlled trial) represents the largest study of cell-based therapy for STEMI completed in the United States and provides evidence supporting safety and potential efficacy in patients with left ventricular dysfunction post STEMI who are at risk for death and major morbidity., Clinical Trial Registration: URL: http://www.clinicaltrials.gov. Unique identifier: NCT01495364., (© 2016 American Heart Association, Inc.)

The use of stem or progenitor cells from bone marrow, or peripheral or umbilical cord blood is becoming more common for treatment of diabetic foot problems. These cells promote neovascularization by angiogenic factors and they promote epithelium formation by stimulating cell replication and migration under certain pathological conditions. We investigated the role of CD34 + stem cells from human umbilical cord blood in wound healing using a rat model. Rats were randomly divided into a control group and two groups with diabetes induced by a single dose of 55 mg/kg intraperitoneal streptozocin. Scarred areas 5 mm in diameter were created on the feet of all rats. The diabetic rats constituted the diabetes control group and a diabetes + stem cell group with local injection into the wound site of 0.5 × 106 CD34 + stem cells from human umbilical cord blood. The newly formed skin in the foot wounds following CD34 + stem cell treatment showed significantly improvement by immunohistochemistry and TUNEL staining, and were closer to the wound healing of the control group than the untreated diabetic animals. The increase in FGF expression that accompanied the local injection of CD34 + stem cells indicates that FGF stimulation helped prevent apoptosis. Our findings suggest a promising new treatment approach to diabetic wound healing.

Background: In an open-label blinded study, we compared intracoronary and transendocardial CD34(+) cell transplantation in patients with nonischemic dilated cardiomyopathy., Methods and Results: Of the 40 patients with dilated cardiomyopathy, 20 were randomized to receive intracoronary injection and 20 received transendocardial CD34(+) cell delivery. In both groups, CD34(+) cells were mobilized by filgrastim, collected via apheresis, and labeled with technetium-99m radioisotope for single-photon emission computed tomographic imaging. In the intracoronary group, cells were injected intracoronarily in the artery supplying segments of greater perfusion defect on myocardial perfusion scintigraphy. In the transendocardial group, electroanatomic mapping was used to identify viable but dysfunctional myocardium, and transendocardial cell injections were performed. Nuclear single-photon emission computed tomographic imaging for quantification of myocardial retention was performed 18 hours thereafter. At baseline, groups did not differ in age, sex, left ventricular ejection fraction, or N-terminal pro-brain natriuretic peptide levels. The number of CD34(+) cells was also comparable (105 ± 31 × 10(6) in the transendocardial group versus 103 ± 27 × 10(6) in the intracoronary group, P=0.62). At 18 hours after procedure, myocardial retention was higher in the transendocardial group (19.2 ± 4.8%) than in the intracoronary group (4.4 ± 1.2%, P<0.01). At 6 months, left ventricular ejection fraction improved more in the transendocardial group (+8.1 ± 4.3%) than in the intracoronary group (+4.2 ± 2.3%, P=0.03). The same pattern was observed for the 6-minute walk test distance (+125 ± 33 m in the transendocardial group versus +86 ± 13 m in the intracoronary group, P=0.03) and N-terminal pro-brain natriuretic peptide (-628 ± 211 versus -315 ± 133 pg/mL, P=0.04)., Conclusions: In patients with dilated cardiomyopathy, transendocardial CD34(+) cell transplantation is associated with higher myocardial retention rates and greater improvement in ventricular function, N-terminal pro-brain natriuretic peptide, and exercise capacity compared with intracoronary route., Clinical Trial Registration Url: http://www.clinicaltrials.gov. Unique identifier: NCT01350310.

Rationale: Ischemic cardiovascular disease represents one of the largest epidemics currently facing the aging population. Current literature has illustrated the efficacy of autologous, stem cell therapies as novel strategies for treating these disorders. The CD34+ hematopoetic stem cell has shown significant promise in addressing myocardial ischemia by promoting angiogenesis that helps preserve the functionality of ischemic myocardium. Unfortunately, both viability and angiogenic quality of autologous CD34+ cells decline with advanced age and diminished cardiovascular health., Objective: To offset age- and health-related angiogenic declines in CD34+ cells, we explored whether the therapeutic efficacy of human CD34+ cells could be enhanced by augmenting their secretion of the known angiogenic factor, sonic hedgehog (Shh)., Methods and Results: When injected into the border zone of mice after acute myocardial infarction, Shh-modified CD34+ cells (CD34(Shh)) protected against ventricular dilation and cardiac functional declines associated with acute myocardial infarction. Treatment with CD34(Shh) also reduced infarct size and increased border zone capillary density compared with unmodified CD34 cells or cells transfected with the empty vector. CD34(Shh) primarily store and secrete Shh protein in exosomes and this storage process appears to be cell-type specific. In vitro analysis of exosomes derived from CD34(Shh) revealed that (1) exosomes transfer Shh protein to other cell types, and (2) exosomal transfer of functional Shh elicits induction of the canonical Shh signaling pathway in recipient cells., Conclusions: Exosome-mediated delivery of Shh to ischemic myocardium represents a major mechanism explaining the observed preservation of cardiac function in mice treated with CD34(Shh) cells.

Induction of antigen-specific T cell tolerance would aid treatment of diverse immunological disorders and help prevent allograft rejection and graft versus host disease. In this study, we establish a method of inducing antigen-specific T cell tolerance in situ in diabetic humanized mice and Rhesus monkeys receiving porcine islet xenografts. Antigen-specific T cell tolerance is induced by administration of an antibody ligating a particular epitope on ICAM-1 (intercellular adhesion molecule 1). Antibody-mediated ligation of ICAM-1 on dendritic cells (DCs) led to the arrest of DCs in a semimature stage in vitro and in vivo. Ablation of DCs from mice completely abrogated anti-ICAM-1-induced antigen-specific T cell tolerance. T cell responses to unrelated antigens remained unaffected. In situ induction of DC-mediated T cell tolerance using this method may represent a potent therapeutic tool for preventing graft rejection.

Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disorder with marginal therapeutic options. Degeneration of motor neurons in the primary motor cortex, brainstem and spinal cord lead to rapidly progressive paralysis and finally to death due to respiratory failure. As pharmacological therapies have failed to provide sufficient neuroprotective effects in ALS, transplantation of stem or progenitor cells is considered a promising treatment strategy. Cell transplantation approaches in ALS mainly aim to generate a neuroprotective environment for degenerating motor neurons by transplantation of non-neuronal cells, rather than to replace lost motor neurons. We present a 63-year-old male patient suffering from ALS who underwent intramedullary thoracic spinal cord implantation of human CD34(+) umbilical cord-derived haematopoietic progenitor cells with a three-year follow up after transplantation.

Neuronal damage induced by ongoing human immunodeficiency virus type 1 (HIV-1) infection was investigated in humanized NOD/scid-IL-2Rγ(c)(null) mice transplanted at birth with human CD34-positive hematopoietic stem cells. Mice infected at 5 months of age and followed for up to 15 weeks maintained significant plasma viral loads and showed reduced numbers of CD4(+) T-cells. Prospective serial proton magnetic resonance spectroscopy tests showed selective reductions in cortical N-acetyl aspartate in infected animals. Diffusion tensor imaging revealed structural changes in cortical gray matter. Postmortem immunofluorescence brain tissue examinations for neuronal and glial markers, captured by multispectral imaging microscopy and quantified by morphometric and fluorescence emission, showed regional reduction of neuronal soma and synaptic architectures. This was evidenced by loss of microtubule-associated protein 2, synaptophysin, and neurofilament antigens. This study is the first, to our knowledge, demonstrating lost neuronal integrity after HIV-1 infection in humanized mice. As such, the model permits studies of the relationships between ongoing viral replication and virus-associated neurodegeneration.

Background: The development of cell-based therapeutics for humans requires preclinical testing in animal models. The use of autologous animal products fails to address the efficacy of similar products derived from humans. We used a novel immunodeficient rat carotid injury model in order to determine whether human cells could improve vascular remodelling following acute injury., Methods: Human CD34+ cells were separated from peripheral buffy coats using automatic magnetic cell separation. Carotid arterial injury was performed in male Sprague-Dawley nude rats using a 2F Fogarty balloon catheter. Freshly harvested CD34+ cells or saline alone was administered locally for 20 minutes by endoluminal instillation. Structural and functional analysis of the arteries was performed 28 days later., Results: Morphometric analysis demonstrated that human CD34+ cell delivery was associated with a significant reduction in intimal formation 4 weeks following balloon injury as compared with saline (I/M ratio 0.79 +/- 0.18, and 1.71 +/- 0.18 for CD34, and saline-treated vessels, respectively P < 0.05). Vasoreactivity studies showed that maximal relaxation of vessel rings from human CD34+ treated animals was significantly enhanced compared with saline-treated counterparts (74.1 +/- 10.2, and 36.8 +/- 12.1% relaxation for CD34+ cells and saline, respectively, P < 0.05), Conclusion: Delivery of human CD34+ cells limits neointima formation and improves arterial reactivity after vascular injury. These studies advance the concept of cell delivery to effect vascular remodeling toward a potential human cellular product.

Development of topical microbicides that prevent sexual transmission of HIV is an active area of investigation. The purpose of this study was to test the ability of the potent membrane fusion inhibitor C34, an HIV gp41 antagonist, to block HIV infection of human Langerhans cells (LCs) in an epithelial environment that mimics a major route of HIV infection. We incubated freshly isolated epidermal explants containing LCs with various doses of C34 before, during, and after exposing explants to HIV. Although C34 only partially blocked HIV infection of LCs when pre-incubated with skin, it displayed full, dose-dependent inhibition when present during and after viral exposure. The poor protection from HIV infectivity in pre-incubated samples is consistent with mechanism of C34 inhibition and starkly contrasts to the full protection provided by PSC-RANTES, an entry inhibitor that prevents HIV gp120 interaction with its co-receptor CCR5. Real-time PCR confirmed that C34 blocked HIV infection of LCs before reverse transcription and inhibited LC-mediated transfer of virus to T cells. We conclude that C34, if used topically at susceptible mucosal sites, and if continually present, has the potential to block sexual transmission of HIV.

CD34+ progenitor cells hold promise for therapeutic neovascularization in various settings. In this study, the role of human peripheral blood CD34+ cells in neovascularization and inflammatory cell recruitment was longitudinally studied in vivo. Human CD34+ cells were incorporated in Matrigel, implanted subcutaneously in nude mice, and explanted after 2, 4, 7, or 14 days. Cell-free Matrigels served as controls. Histochemical analyses demonstrated that neovascularization occurred almost exclusively in CD34+ implants. Cellular and capillary density were increased in cell-loaded Matrigels after 2 days and further increased at 14 days. Human CD34+ cells did not incorporate in neovessels, but formed vWF+/CD31+/VEGF+ cell clusters that were present up to day 14. However, CD34+ cells induced host neovascularization, as demonstrated by increased presence of murine CD31+ and vWF+ vasculature from day 7 to 14. Moreover, recruitment of murine monocytes/macrophages was significantly enhanced in CD34+ implants at all time points. Gene expression of chemotactic cytokines MCP-1 and IL-8 was detected on CD34+ cells in vitro and confirmed immunohistochemically in cell-loaded explants at all time points. Our data indicate that human CD34+ cells, implanted in a hypoxic environment, generate an angiogenic niche by secreting chemotactic and angiogenic factors, enabling rapid neovascularization, possibly via recruitment of monocytes/macrophages.

Purpose: Nucleated cell dose is an important and modifiable factor in hematopoietic stem cell transplantation (HSCT), however its association with outcomes in the context of reduced intensity conditioning regimen (RIC) HSCT for adults with acute myelocytic leukemia (AML) is not known., Patients and Methods: From 1998 to 2003, 253 patients with de novo AML, received transplants with RIC and peripheral blood from a genoidentical donor. Median age was 55 years (range, 18 to 72) and the median follow-up was 17 months (range, 2 to 67). One hundred forty one patients received transplants in first remission (CR1), 47 received transplants in second remission (CR2), and 65 patients received transplants in a more advanced phase. Fludarabin-based RIC was used in 91% of patients and low-dose (< 4 Gy) total-body radiation in 23% of patients. The median nucleated and CD34 cell dose infused were 9.1x 10(8)/kg and 5.8x 10(6)/kg, respectively., Results: Overall, 2-year leukemia-free survival (LFS) was 41% +/- 4% and it was 46% +/- 5% for patients receiving a higher cell dose (> 9.1x 10(8)/kg) and 37% +/- 5% for the remainders (P = .03). Higher cell doses exclusively benefited patients who received transplantations in CR2 or beyond, with LFS of 47 +/- 8 versus 20 +/- 8, with no detectable effect for patients who received transplants in CR1. In a multivariate analysis of the overall patient population, higher nucleated cell dose cells were associated with higher LFS (P = .04), higher incidence of chronic graft-versus-host disease (P = .01), and there was a trend towards a lower relapse incidence (P = .06). Interestingly, CD34+ cell dose was not associated with any outcomes., Conclusion: Nucleated cell dose is an important factor that can be modified to improve results of RIC for patients with AML transplanted later than in CR1.

Current stem cell protocols for ischemic heart disease are limited by the small numbers of cells that can be obtained by bone marrow aspirate. To increase myocardial delivery of bone marrow stem cells in patients with chronic ischemic heart disease (CIHD), we used granulocyte colony stimulating factor (G-CSF) for bone marrow mobilization of CD34+ cells, enabling intracoronary infusion of large numbers of CD34+ stem cells. Patients with CIHD (n = 5) demonstrated significantly reduced numbers of CD34+ cells mobilized by G-CSF in comparison to age-matched controls. Sustained reduction in anginal symptoms and improvement in quality of life scores was seen in all patients following infusion of cells. Moreover, mean collateral flow grade at 12-month follow-up angiography significantly improved, indicating sustained myocardial neovascularization. No proliferative retinopathy was induced and no in-stent restenosis seen. However, in two patients with documented increase in collateral flow, complications arose, one developing an acute coronary syndrome and the other a lentigo maligna. These results demonstrate the feasibility of G-CSF mobilization, leukapheresis and intracoronary transfer of CD34+ stem cells in patients with CIHD, but longer-term studies are required to ensure that this protocol is safe and effective.

Objective: The purpose of this study was to determine cell dosage parameters for successful engraftment of human cord blood hematopoietic stem cells (HSC) using an in vivo assay system, and to determine if there are differences with donor gestational age., Study Design: HSCs were transplanted into nonobese diabetic-severe combined immunodeficient (NOD/SCID) mice. Donor cell dosage and gestational age ranges were 1 to 40 x 10(6) CD34+ cells per mouse, and 23 to 40 weeks, respectively. Recipient bone marrow was assessed for engraftment capacity of the HSCs., Results: There was increasing engraftment levels with increasing dosages of transplanted HSCs. When controlled for donor HSC dosage, engraftment levels using donor cord blood from earlier gestational ages were not different from that seen using later gestational ages., Conclusion: Similar dose responses are seen using HSCs derived from the late second trimester until term in engraftment potential in the NOD/SCID mouse model. Results from this study may be applicable to human postnatal and in utero transplantation studies.

Thrombo-occlusive cerebrovascular disease resulting in stroke and permanent neuronal loss is an important cause of morbidity and mortality. Because of the unique properties of cerebral vasculature and the limited reparative capability of neuronal tissue, it has been difficult to devise effective neuroprotective therapies in cerebral ischemia. Our results demonstrate that systemic administration of human cord blood-derived CD34(+) cells to immunocompromised mice subjected to stroke 48 hours earlier induces neovascularization in the ischemic zone and provides a favorable environment for neuronal regeneration. Endogenous neurogenesis, suppressed by an antiangiogenic agent, is accelerated as a result of enhanced migration of neuronal progenitor cells to the damaged area, followed by their maturation and functional recovery. Our data suggest an essential role for CD34(+) cells in promoting directly or indirectly an environment conducive to neovascularization of ischemic brain so that neuronal regeneration can proceed.

Previously, we have shown that patients with diffuse large B-cell lymphoma (DLBCL) transplanted with contaminated bone marrow (BM) generally have a poor outcome. Whether this is also the case when peripheral blood progenitor cell (PBPC) grafts are used is not known. Forty-three patients with chemosensitive DLBCL or follicular lymphoma grade 3 (FLgr3) were treated with high-dose therapy (HDT) and autologous stem cell support. Nine patients received purged grafts. Quantitative real-time polymerase chain reaction (QRT-PCR) for either the BCL2/IgH translocation or allele specific oligonucleotide (ASO) QRT-PCR for the immunoglobulin heavy chain (IgH) complementarity-determining region 3 were used. Nine of 25 (36%) PBPC grafts contained tumour cells as tested by QRT-PCR, including two grafts purged by CD34(+) cell enrichment combined with B-cell depletion. The level of contamination of the PBPC/CD34(+) cells ranged from 0 to 8.28%. No relationship could be shown between the total number of tumour cells infused and relapse. Patients receiving PCR-positive or PCR-negative PBPC grafts had similar progression-free survival (PFS) (P = 0.49). However, a significant difference was seen in PFS and overall survival (OS) for the patients given >/=6.1 x 10(6) CD34(+) cells/kg compared with those given <6.1 x 10(6) CD34(+) cells/kg (P = 0.01 and P < 0.05 respectively).

Purpose: The purpose of this work was to study the correlation of pancreatic phase Computed tomography (CT) enhancement, intratumoral microvessel density (MVD) and pathologic grades in pancreatic carcinoma and to evaluate the relationship between CT enhancement degree and the malignancy degree of pancreatic carcinoma., Methods: 34 patients with pancreatic carcinoma underwent CT scanning before resection. The enhancement degrees and forms of tumor were observed in pancreatic phase. The operational sample was stained with HE and CD34 marked by immunohistochemistry. MVD and histopathological grades of pancreatic carcinoma were examined. CT enhancement of the tumor, MVD counting in hot spot areas of neoplastic parenchymal cells and pathological grades of pancreatic carcinoma were comparatively analyzed., Result: Highly differentiated pancreatic adenocarcinoma was identified in 16 patients, moderately differentiated tumor in seven and poorly differentiated tumor in 11. Isodensity CT enhancement was demonstrated in 13 cases, slight low density enhancement in nine, slight low density enhancement along with small cyst lesion in nine and slight low density enhancement along with large cyst lesion in three. The counting of MVD with CD34 marked by immunohistochemistry in hot spot areas of neoplastic parenchyma cells were small in ten cases, medium in 16 and large in eight. The pathological grades correlated with CT enhancement of tumor (r=0.7857, P<0.001). The pathological grade correlated with MVD counting of tumor (r=0.3613, P<0.05). The CT enhancement of tumor correlated with MVD (r=0.6768, P<0.001)., Conclusion: There was obvious and significant correlation between CT enhancement, pathological grades and MVD numbers in the hot spot areas of tumor. The extent of CT enhancement was inversely proportional to the malignancy degree of pancreatic carcinoma, and inversely proportional to MVD numbers in the hot spot areas of neoplastic parenchyma. The MVD in the hot spot areas of neoplastic parenchyma cells could also reflect the prognosis of the patients, and was directly proportional to malignancy degree of pancreatic carcinoma.

Currently, no information is available regarding the influence of the different CD34+ cell subsets infused on the haematopoietic recovery, following non-myeloablative allogeneic peripheral blood stem cell transplantation (allo-PBSCT). We have explored, in a group of 13 patients receiving non-myeloablative allo-PBSCT from human leucocyte antigen-identical sibling donors, the influence of the total dose of CD34+ haematopoietic progenitor cells (HPC) infused, compared with that of the different CD34+ HPC and CD34- leucocyte subsets in the leukapheresis samples, on both engraftment and clinical outcome. The overall numbers of total CD34+ HPC (P = 0.002) and myelomonocytic-committed CD34+ HPC infused (P = 0.0002) were strongly associated with neutrophil recovery (> 1 x 109 neutrophils/l), the latter being the only independent parameter influencing neutrophil recovery. Regarding long-term engraftment, only the number of immature CD34+ HPC infused/kg correlated with the duration of hospitalization in the first 2 years after discharge (r = -0.75, P = 0.005). Both the overall amount of CD34+ HPC and the number of myelomonocytic CD34+ HPC infused showed a significant influence on the risk of graft-versus-host disease (GVHD). Thus, the overall probability of GVHD was 100%vs 25% for patients receiving >/= 5 x 106 CD34+ HPC or >/= 3.5 x 106 of myelomonocytic-committed CD34+ HPC vs lower doses (P = 0.013). None of the other CD34+ and CD34- cell subsets analysed correlated with development of GVHD. In summary, our results suggest that in non-myeloablative allo-PBSCT, high numbers of CD34+ HPC, especially the myelomonocytic-committed CD34+ progenitors, lead to rapid neutrophil engraftment. However, they also strongly impair clinical outcome by increasing the incidence of GVHD.

Thirty-six patients with multiple myeloma (23 PR1, nine PR2, four stable disease) were entered into a pilot study evaluating the use of CD34+-selected peripheral blood progenitor cell transplantation (PBPCT) following high-dose melphalan alone or high-dose melphalan and total body irradiation. Peripheral blood progenitor cells (PBPCs) were mobilized with cyclophosphamide and granulocyte colony stimulating factor (G-CSF). CD34+ selection using the Cellpro Ceprate-SC system was performed in 22 cases with an adequate yield in 20. 10 patients failed to mobilize sufficient cells to permit selection and in four cases selection was not performed for other reasons. 16 patients therefore received unselected PBPC. Tumour cell contamination was evaluated by IgH gene fingerprinting (fpPCR). Harvested PBPC were fpPCR positive in 13/20 CD34+-selected cases and remained positive after selection in seven. Harvested PBPC were studied in 9/16 patients receiving unselected cells; fpPCR was positive in five and negative in four. There was no difference in event-free survival (EFS) between the CD34+-selected group and the unselected group (median 21 and 26 months, respectively, P=ns). The CD34+-selection process therefore reduced contamination but did not eliminate it completely, and in this small non-randomized study there was no apparent clinical benefit of CD34+ selection.

Purpose: To assess hematologic recovery and procedure-related mortality in patients who received high-dose therapy with stem-cell support, in whom the peripheral-blood stem-cell (PBSC) collection fails (CD34+ cells < 1 x 10(6)/kg). The predictive value of granulocyte-monocyte colony-forming cell (GM-CFC) measurements and the value of bone marrow obtained after PBSC collection failure was assessed., Patients and Methods: The study group comprised 324 consecutive patients mobilized with granulocyte colony-stimulating factor (G-CSF) and cyclophosphamide (273 patients), G-CSF with other chemotherapy (37 patients), and G-CSF alone (14 patients). Between one and four aphereses were performed., Results: In 51 of 324 patients, there was failure to obtain 1 x 10(6)/kg CD34+ cells. Twenty-three patients had greater than 1 x 10(5)/kg GM-CFC; 22 patients proceeded to high-dose therapy. Neutrophil recovery occurred within 21 days, but platelet independence was delayed (> 28 days) in eight patients. Of 28 patients with less than 1 x 10(5)/kg GM-CFC, six received high-dose therapy with PBSC alone and five had delayed engraftment. Twelve patients with less than 1 x 10(5)/kg GM-CFC received high-dose therapy supported by bone marrow collected after PBSC collection failure. Eleven patients were assessable for engraftment; four patients had slow (> 21 days) or delayed (> 28 days) neutrophil recovery and eight patients had delayed platelet recovery. In the group of patients who received less than 1 x 10(5)/kg GM-CFC, there were five procedure-related deaths., Conclusion: This study shows that delayed hematologic recovery is frequent if less than 1 x 10(6)/kg CD34+ cells are infused after high-dose therapy, particularly with GM-CFC less than 1 x 10(5)/kg. The procedure-related mortality in this latter group is high. In most patients whose PBSC collection contains less than 1 x 10(5)/kg GM-CFC, the use of bone marrow cells does not improve engraftment, which suggests that poor PBSC mobilization usually indicates poor marrow function.

Circulating progenitor cells (CPCs) mobilized from bone marrow will replace the use of bone marrow transplantation because hematopoietic reconstitution is more rapid using the former technique. We report on early and late recovery of hematopoiesis after CPC transplantation in patients with non-Hodgkin's lymphoma (NHL) and analyze the role of variables possibly influencing engraftment. From December 1992 through September 1995, 57 consecutive NHL patients were enrolled in this study. Patients could be divided into 2 groups: the first comprised 32 patients with untreated diffuse large-cell lymphoma and unfavorable prognostic factors; the second comprised 25 patients with resistant or relapsing NHL of low-and high-grade histology. All patients received high-dose chemotherapy (carmustine, cytarabine, etoposide, and melphalan; BEAM) followed by CPC transplantation. In all, 25 patients were treated with granulocyte colony-stimulating factor (G-CSF) after CPC administration. The time to short-and long-term hematologic engraftment and variables correlating with multilineage long-term reconstitution were examined. The time to bilineage (neutrophils and platelets) hematopoietic reconstitution did not differ in G-CSF-treated and-untreated patients. In contrast, the time taken to reach a neutrophil count of 0.5 x 10(9)/1 and a WBC of 1 x 10(9)/1 was significantly shorter in G-CSF-treated patients. Overall, 33 patients achieved long-term, complete trilineage engraftment after a median of 117 days from CPC transplantation. The leukocyte count was the first parameter to reach full engraftment and hemoglobin was the last. According to Kaplan-Meier analysis, 80% of the patients are projected to reconstitute fully at 12 months after transplantation. Univariate and multivariate analyses showed that sustained, long-term hematopoiesis was significantly related to a younger age, an early bilineage reconstitution, and the quantity of CD34+ cells infused.