Biomedical subjects

Gay M Crooks

Publications and source records attributed to Gay M Crooks.

6 recordsLinked to original sources

PubMed2003

Dynamic tracking of human hematopoietic stem cell engraftment using in vivo bioluminescence imaging.

The standard approach to assess hematopoietic stem cell (HSC) engraftment in experimental bone marrow transplantation models relies on detection of donor hematopoietic cells in host bone marrow following death; this approach provides data from only a single time point after transplantation for each animal. In vivo bioluminescence imaging was therefore explored as a method to gain a dynamic, longitudinal profile of human HSC engraftment in a living xenogeneic model. Luciferase expression using a lentiviral vector allowed detection of distinctly different patterns of engraftment kinetics from human CD34+ and CD34+CD38- populations in the marrow NOD/SCID/beta 2mnull mice. Imaging showed an early peak (day 13) of engraftment from CD34+ cells followed by a rapid decline in signal. Engraftment from the more primitive CD34+CD38- population was relatively delayed but by day 36 increased to significantly higher levels than those from CD34+ cells (P <.05). Signal intensity from CD34+CD38-engrafted mice continued to increase during more than 100 days of analysis. Flow cytometry analysis of bone marrow from mice after death demonstrated that levels of 1% donor cell engraftment could be readily detected by bioluminescence imaging; higher engraftment levels corresponded to higher image signal intensity. In vivo bioluminescence imaging provides a novel method to track the dynamics of engraftment of human HSC and progenitors in vivo.

Animals↗

PubMed2003

Ikaros isoform x is selectively expressed in myeloid differentiation.

The Ikaros gene is alternately spliced to generate multiple DNA-binding and nonbinding isoforms that have been implicated as regulators of hematopoiesis, particularly in the lymphoid lineages. Although early reports of Ikaros mutant mice focused on lymphoid defects, these mice also show significant myeloid, erythroid, and stem cell defects. However, the specific Ikaros proteins expressed in these cells have not been determined. We recently described Ikaros-x (Ikx), a new Ikaros isoform that is the predominant Ikaros protein in normal human hematopoietic cells. In this study, we report that the Ikx protein is selectively expressed in human myeloid lineage cells, while Ik1 predominates in the lymphoid and erythroid lineages. Both Ik1 and Ikx proteins are expressed in early human hematopoietic cells (Lin(-)CD34(+)). Under culture conditions that promote specific lineage differentiation, Ikx is up-regulated during myeloid differentiation but down-regulated during lymphoid differentiation from human Lin(-)CD34(+) cells. We show that Ikx and other novel Ikaros splice variants identified in human studies are also expressed in murine bone marrow. In mice, as in humans, the Ikx protein is selectively expressed in the myeloid lineage. Our studies suggest that Ikaros proteins function in myeloid, as well as lymphoid, differentiation and that specific Ikaros isoforms may play a role in regulating lineage commitment decisions in mice and humans.

Animals↗

PubMed2003

Albumin-expressing hepatocyte-like cells develop in the livers of immune-deficient mice that received transplants of highly purified human hematopoietic stem cells.

Rodent bone marrow cells can contribute to liver. If these findings are applicable to humans, marrow stem cells could theoretically be harvested from a patient and used to repair his/her damaged liver. To explore this potential, CD34(+) or highly purified CD34(+)CD38(-)CD7(-) human hematopoietic stem cells from umbilical cord blood and bone marrow were transplanted into immunodeficient mice. One month after transplantation, carbon tetrachloride (CCl(4)) was administered into the mice to induce liver damage and hepatocyte proliferation. Mice were analyzed in comparison with CCl(4)-injured mice that did not receive transplants and noninjured controls that received transplants with the same stem cell populations, one month after liver damage. Human-specific albumin mRNA and protein were expressed in the mouse liver and human albumin was detected in the serum of mice that had received CCl(4) injury. Human alpha-fetoprotein was never expressed, but in some mice, human cytokeratin 19 was expressed, which may indicate bile duct development in addition to the albumin-secreting hepatocyte-like cells. Human albumin was not expressed in the starting stem cell populations in injured mice that did not receive transplants nor in noninjured mice that had received transplants of human stem cells. Human albumin expression was detected only in CCl(4)-treated mice that received transplants of human stem cells, and recovery was increased by administration of human hepatocyte growth factor 48 hours after the CCl(4)-mediated liver injury. Our studies provide evidence that human "hematopoietic" stem/progenitor cell populations have the capacity to respond to the injured liver microenvironment by inducing albumin expression.

Animals↗

PubMed2002

Human hematopoietic lineage commitment.

The ultimate goal of developmental immunology is to understand the normal processes that give rise to the immune system in order to diagnose and develop effective treatments for diseases that occur as a consequence of immune system defects. Central to achieving this goal is understanding the complex interplay between microenvironmental signals and transcription factors that direct human hematopoietic differentiation and lineage commitment. The ability to isolate highly purified populations of human hematopoietic cells at critical points in differentiation make it possible to answer very specific questions about the hematopoietic process and lineage restriction. This review describes the use of surface immunophenotypes to identify human hematopoietic cells at particular points in differentiation or with particular patterns of lineage restriction. Culture models are discussed in the context of the ability to detect, characterize and determine the lineage potential of human hematopoietic stem cells and progenitors. Variations in hematopoeises that correspond to ontogeny will be examined. Potential roles for the HOX and Ikaros proteins in human lineage commitment will be considered. Also included will be discussion of a number of factors that provide challenges to experimental design, to experimental interpretation, and to the development of a comprehensive model of human hematopoiesis.

Cell Differentiation↗

PubMed2002

Second hematopoietic stem cell transplantation in pediatric patients: overall survival and long-term follow-up.

Despite potent intensive conditioning regimens, hematopoietic stem cell transplantation (HSCT) may fail because of either relapse of the malignancy or the rejection of the graft. We report on 27 pediatric patients who received a second HSCT from an allogeneic donor for relapsed malignancy or graft failure. One-year, 5-year, and 10-year probabilities of survival for all patients were 53%, 36%, and 24%, respectively. Twenty patients received second HSCTs for relapsed malignancy, of whom 6 were alive and disease free at the time of this report. Seven patients received a second HSCT for graft failure, of whom 3 were alive and well as of this report. Twenty-five patients were tested for immune reconstitution following their second HSCT. Sixteen patients developed antigen-specific T-lymphocyte responses; the median time to development of antigen-specific responses was 13 months. There was no significant neurocognitive decline in patients tested 1 to 3 years following their second HSCT. Endocrine evaluations revealed deficiencies in growth hormone (7 patients), gonadal function (3 patients), and thyroid function (2 patients). Three patients developed significant abnormalities of tooth development, including absence of secondary teeth. These results show that a second HSCT offers curative therapy for selected pediatric patients whose first HSCT failed. Although toxicity is considerable following a second transplantation, the major causes of mortality continue to be relapse and infection.

Adolescent↗

PubMed2002

Expression from second-generation feline immunodeficiency virus vectors is impaired in human hematopoietic cells.

Vectors based on the feline immunodeficiency virus (FIV) have been developed as an alternative to those based on another lentivirus, human immunodeficiency virus-1 (HIV-1), because of theoretical safety advantages. We compared the efficiency of gene transfer and expression in human and feline hematopoietic progenitors using second-generation HIV-1 and FIV-based vectors. Vector pairs were tested using either human cytomegalovirus or murine phospho-glycerate kinase (PGK) internal promoters and were pseudotyped with the vesicular stomatitis virus G protein (VSV-G). Vector proviral copy numbers were similar in human and feline hematopoietic primary cells and cell lines transduced by HIV-1 or FIV vectors, demonstrating that both vectors are able to transfer genes efficiently to these cell types. HIV-1 vectors were well expressed in human primary hematopoietic cells and cell lines. However, transgene expression from FIV vectors was almost undetectable in human hematopoietic cells. In contrast, the FIV vector was expressed well in primary hematopoietic feline cells and human non-hematopoietic cells, demonstrating that low transgene expression from the FIV vector is a phenomenon specific to human hematopoietic cells. Northern blot analysis demonstrated decreased vector transcript levels in human CEM cells transduced with FIV relative to cells transduced with HIV-1, despite high vector copy numbers. No evidence of vector transcript instability was seen in studies of transduced CEM cells treated with actinomycin D. We conclude that FIV vectors can transfer genes into human hematopoietic cells as effectively as HIV-1 vectors, but that unknown elements in the current FIV backbone inhibit expression from FIV vectors in human hematopoietic cells.

Antigens, CD34↗