Biomedical subjects

Alan Flint

Publications and source records attributed to Alan Flint.

6 recordsLinked to original sources

PubMed2006

Muscle engraftment of myogenic progenitor cells following intraarterial transplantation.

Cell-based therapy continues to be a promising avenue for the treatment of Duchenne muscular dystrophy (DMD), an X-linked skeletal muscle-wasting disease. Recently, we demonstrated that freshly isolated myogenic progenitors contained within the adult skeletal muscle side population (SP) can engraft into dystrophic fibers of nonirradiated mdx(5cv) mice after intravenous transplantation. Engraftment rates, however, have not been therapeutically significant, achieving at most 1% of skeletal muscle myofibers expressing protein from donor-derived nuclei. To enhance the engraftment of transplanted myogenic progenitors, an intraarterial delivery method was adapted from a previously described procedure. Cultured, lentivirus-transduced skeletal muscle SP cells, derived from mdx(5cv) mice, were transplanted into the femoral artery of noninjured mdx(5cv) mice. Based on the expression of microdystrophin or green fluorescent protein (GFP) transgenes in host muscle, sections of the recipient muscles exhibited 5%-8% of skeletal muscle fibers expressing donor-derived transgenes. Further, donor muscle SP cells, which did not express any myogenic markers prior to transplant, expressed the satellite cell transcription factor, Pax7, and the muscle-specific intermediate filament, desmin, after extravasation into host muscle. The expression of these muscle-specific markers indicates that progenitors within the side population can differentiate along the myogenic lineage after intraarterial transplantation and extravasation into host muscle. Given that femoral artery catheterization is a common, safe clinical procedure and that the transplantation of cultured adult muscle progenitor cells has proven to be safe in mice, our data may represent a step toward the improvement of cell-based therapies for DMD and other myogenic disorders.

Animals↗

PubMed2005

A comparison of the psychometric properties of the hooked on nicotine checklist and the modified Fagerström tolerance questionnaire.

We compared the psychometric properties of the Hooked on Nicotine Checklist (HONC) and the Modified Fagerström Tolerance Questionnaire (MFTQ). Adolescent current smokers (n = 215) completed both instruments three times, at baseline and 6- and 12-month follow-up. Internal consistency of the HONC was high (alpha = 0.92), as was its stability over the follow-up interval (intraclass correlation (ICC) = 0.93 over 6 months and 0.91 over 1 year). Internal consistency of the MFTQ was acceptable (alpha = 0.83), and its stability over the follow-up interval was similar to that reported previously (ICC = 0.79 at 6 months and 0.76 at 1 year). The HONC predicted smoking at both follow-up points, while the MFTQ did so only at 6 months. The HONC compared favorably with the MFTQ in all respects. The most important advantage of the HONC is that it is measuring a clearly defined concept, diminished autonomy over tobacco, which begins when the sequelae of tobacco use present a barrier to quitting.

Adolescent↗

PubMed2004

Demystifying SP cell purification: viability, yield, and phenotype are defined by isolation parameters.

Side population (SP) cells isolated from bone marrow, skeletal muscle, and skin have been shown to engraft in dystrophic muscle. However, there have been questions on the phenotypical heterogeneity, tissue of origin, and relationships among SP cell populations extracted from different tissues. Studies on bone marrow SP cells have followed a consistent protocol for their isolation and results obtained are concordant. In contrast, protocols for the isolation of muscle SP cells vary greatly, and consequently reports on their phenotype, differentiation potential and origin have been inconsistent. To address this controversy, we demonstrate that isolation parameters, such as tissue dissociation, cell counting, Hoechst concentration, and stringency in the selection of SP cells, have an effect on the yield, viability, and homogeneity of SP cells derived from bone marrow, skeletal muscle, and skin. In this paper, we demonstrate that SP cells isolated from the bone marrow are distinct from SP cells extracted from skeletal muscle and skin tissues. This study offers an explanation for the controversy surrounding muscle SP cells, provides a detailed standardized protocol for their isolation, and highlights basic guidelines for reproducible and reliable isolation of SP cells from any tissue.

Animals↗

PubMed2004

Systemic delivery of human microdystrophin to regenerating mouse dystrophic muscle by muscle progenitor cells.

Cell-based therapy for Duchenne muscular dystrophy patients and mdx mice has proven to be a safe but ineffective form of treatment. Recently, a group of cells called muscle side population (SP) cells have been isolated based on their ability to efflux the DNA-binding dye Hoechst. To understand the potential of skeletal muscle SP cells to serve as precursors for muscle, SP cells from the two mice strains mdx(5cv) and C57BL/6N were isolated, transduced, and transplanted. Under coculture conditions with myogenic cells, some cells within the SP cell population can give rise to early Pax7-positive satellite cells and other later stage myogenic cells. Transduced SP cells were transplanted via the tail vein and were shown to successfully deliver enhanced GFP and human microdystrophin to the skeletal muscle of nonirradiated mdx(5cv) mice, thus demonstrating their ability to travel through the capillaries and enter into damaged muscle. These results demonstrate that i.v. delivery of genes via SP cells is possible and that these SP cells are capable of recapitulating the myogenic lineage. Because this approach shows definitive engraftment by using autologous transplantation of noninjured recipients, our data may have substantial implications for therapy of muscular dystrophy.

Animals↗

PubMed2003

Skeletal muscle engraftment potential of adult mouse skin side population cells.

Adult bone marrow and skeletal muscle have been shown to contain a subpopulation of cells, called side population (SP) cells, that can be isolated with the fluorescence-activated cell sorter. We used a similar method to identify SP cells in the skin of adult mice. These cells express surface markers similar to SP cells isolated from skeletal muscle, but differ from bone marrow SP cells and do not express hematopoietic markers. When transplanted into nonirradiated mdx mice, nuclei from donor skin SP cells are found within myofibers that express dystrophin. Thus, adult skin SP cells can engraft in dystrophic skeletal muscle even in the absence of total body irradiation.

Animals↗

PubMed2002

AC133-2, a novel isoform of human AC133 stem cell antigen.

Human AC133 antigen, also called CD133, was recently identified as a hematopoietic stem cell marker. However, the molecular structure and function of this protein has remained unclear. Here we cloned and identified a novel isoform of AC133, which we named AC133-2. In comparison to the reported AC133 cDNA, which is referred to herein as AC133-1, a small exon of 27 nucleotides is deleted in AC133-2 by alternative mRNA splicing. Similar to the previously characterized AC133 antigen, recombinant AC133-2 expressed in 293 cells was glycosylated and transported to plasma membrane. AC133-2 mRNA was found predominant in a variety of human fetal tissue, adult tissues, and several carcinomas. In contrast, AC133-1 mRNA was more prominent in fetal brain and adult skeletal muscle but was not detected in fetal liver and kidney, adult pancreas, kidney, and placenta, suggesting different roles for the two isoforms in fetal development and mature organ homeostasis. Here, we demonstrate that AC133-2 is the isoform expressed on hematopoietic stem cells derived from fetal liver, bone marrow, and peripheral blood. The results indicate that AC133-2, not AC133-1, has been the cell surface antigen recognized by anti-AC133 monoclonal antibodies that are used for isolation of hematopoietic stem cells. To further investigate its expression in other stem cell populations, we found that AC133-2 co-expressed with beta(1) integrin in the basal layer of human neonatal epidermis. AC133-2(+)/beta(1) integrin(+) cells proliferated and differentiated in culture, which coincided with a loss of AC133-2 and gain in a terminal differentiation marker involucrin. Taken together, these results suggest that AC133-2 is expressed in multiple stem cell niches and may provide a means to isolate specific stem cell subpopulations from human tissues.

AC133 Antigen↗