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Biomedical subjects

F McLaughlin

Publications and source records attributed to F McLaughlin.

13 recordsLinked to original sources
PubMed2002

Glucocorticoid-induced osteopenia in the mouse as assessed by histomorphometry, microcomputed tomography, and biochemical markers.

Glucocorticoids are potent anti-inflammatory molecules used in the treatment of asthma, rheumatoid arthritis, inflammatory bowel disease, and other inflammatory and dermatological diseases, as well as in posttransplantation immunotherapy. Although glucocorticoids have been prescribed for many years, their potential side effects, when administered orally, can prevent their long-term use. The most serious side effect observed in the clinic is glucocorticoid-induced osteoporosis (GIOP). To develop a small animal model to characterize glucocorticoid-induced bone loss, we carried out a series of experiments using BALB/c mice given daily intraperitoneal doses of the synthetic glucocorticoid, dexamethasone. Following dexamethasone treatment, the mice became osteopenic, with highly significant decreases in bone formation rate and mineral apposition rate, as assessed by standard histomorphometry. Moreover, 3 week treatment with dexamethasone resulted in a decrease in trabecular thickness and trabecular number with an increase in surface-to-volume ratio of trabeculae in the distal femur, as measured using microcomputed tomography (micro-CT). The serum bone formation marker, osteocalcin, was dose-dependently decreased in all mice treated with dexamethasone and showed a parallel extent of regulation to the bone formation rate changes. In addition, serum levels of leptin, recently identified as playing a role in the regulation of bone mass, increased following dexamethasone treatment. BALB/c mice therefore represent a useful model system in which the detrimental effects of glucocorticoids on bone can be studied.

Animals↗
PubMed2001

Combined genomic and antisense analysis reveals that the transcription factor Erg is implicated in endothelial cell differentiation.

It has recently been shown that the transcription factor Erg, an Ets family member, drives constitutive expression of the intercellular adhesion molecule 2 (ICAM-2) in human umbilical vein endothelial cells (HUVECs) and that its expression is down-regulated by the pleiotropic cytokine tumor necrosis factor alpha (TNF-alpha). To identify other Erg target genes and to define its function in the endothelium, a combined approach of antisense oligonucleotides (GeneBloc) and differential gene expression was used. Treatment of HUVECs with Erg-specific GeneBloc for 24, 48, and 72 hours suppressed Erg mRNA and protein levels at all time points. Total RNA extracted from HUVECs treated with Erg-specific or control GeneBloc was analyzed for differences in gene expression using high-density, sequence-verified cDNA arrays containing 482 relevant genes. Inhibition of Erg expression resulted in decreased expression of ICAM-2, as predicted. Four more genes decreased in Erg-deficient HUVECs were the extracellular matrix proteins SPARC and thrombospondin, the adhesive glycoprotein von Willebrand factor, and the small GTPase RhoA. Each of these molecules has been directly or indirectly linked to angiogenesis because of its role in vascular remodeling, adhesion, or shape change. Therefore, the role of Erg in vascular remodeling was tested in an in vitro model, and the results showed that HUVECs treated with Erg GeneBloc had a decreased ability to form tubulelike structures when grown on Matrigel. These results suggest that Erg may be a mediator of the TNF-alpha effects on angiogenesis in vivo.

Antigens, CD↗
PubMed1999

Chromatin structure and transcriptional regulation of the stem cell leukaemia (SCL) gene in mast cells.

The stem cell leukaemia (SCL) gene is a member of the basic helix-loop-helix family of transcription factors and is essential for the development of all haematopoietic lineages. SCL is expressed in pluripotent haematopoietic stem cells and also following commitment to the erythroid, mast and megakaryocytic lineages. The mechanisms responsible for this pattern of expression are poorly understood, but are likely to illuminate the molecular basis for stem cell development and lineage commitment. Here we present the first description of the regulation of the SCL gene in mast cells. In this study we systematically analysed the chromatin structure of a 45 kb region of the murine SCL locus in mast cells. The pattern of DNase 1 and restriction endonuclease hypersensitive sites in mast cells was distinct from, but overlapped with, the pattern previously described in erythroid and primitive myeloid cells. Each potential regulatory element was tested using transient reporter assays to assess their functional significance in mast cells. These studies identified two potent enhancers, one of which was downstream of the SCL gene. Further characterisation of this 3' enhancer demonstrated that it required the presence of two distinct DNase 1 hypersensitive sites for full activity, and that it was capable of stimulating transcription from both promoter 1a and 1b. Since the 3' enhancer is active in both erythroid and mast cells, it will now be important to see whether it is independently activated in these lineages, or whether it is also active in haematopoietic stem cells.

Animals↗
PubMed1999

Characterisation of the tumour necrosis factor (TNF)-(alpha) response elements in the human ICAM-2 promoter.

ICAM-2 is a cell surface adhesion molecule constitutively expressed on the endothelium, involved in leukocyte recruitment into tissues. We recently showed that pro-inflammatory cytokines tumour necrosis factor (TNF)-(alpha) and interleukin (IL)-1(beta) down-regulate ICAM-2 expression at the transcriptional level. Here we investigate the elements in the ICAM-2 promoter required for the TNF-(alpha)-mediated down-regulation. Site directed mutagenesis of the ICAM-2 promoter implicated three consensus sites for Ets transcription factors in basal activity; two of these sites were also involved in the TNF-(alpha)-induced down-regulation. Electrophoretic mobility shift assays (EMSA) performed in human umbilical vein endothelial cells (HUVEC) showed that all three Ets binding sites (EBS) bind nuclear proteins. TNF-(alpha) treatment (10 ng/ml for 24 hours) decreased binding to the double -135/-127EBS, but not to the -44EBS. The Ets family member Erg was found to be constitutively expressed in HUVEC, and TNF-(alpha) down-regulated Erg protein levels. Furthermore, an Erg cDNA transactivated the ICAM-2 promoter when transiently transfected into both HeLa cells and HUVEC. Protein expression of ICAM-2 and Erg was found to be similarly regulated by TNF-(alpha) in an ex vivo artery model. These data suggest that constitutive endothelial genes ICAM-2 and Erg are on the same pathway of cytokine-dependent regulation of gene expression.

Antigens, CD↗
PubMed1998

Tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta down-regulate intercellular adhesion molecule (ICAM)-2 expression on the endothelium.

Leukocyte recruitment is a crucial step in inflammation. Inflammatory stimuli upregulate the expression of some endothelial adhesion molecules, such as E-selectin or ICAM-1, but not of others such as ICAM-2. ICAM-2, a constitutively expressed endothelial ligand for beta2 integrins LFA-1 and Mac-1, is involved in leukocyte adhesion to resting endothelium and in transmigration in vitro, however its role in inflammation is unclear. We have studied the effect of TNF-alpha and IL-1beta on ICAM-2 expression on human umbilical vein endothelial cells (HUVECs). Prolonged treatment (24 h) of HUVECs with TNF-alpha (10 ng/ml) or IL-1beta (34 ng/ml) reduced ICAM-2 surface expression to 50% of control, while interferon (IFN)-gamma had no effect. The loss in ICAM-2 surface expression correlated with a reduction of ICAM-2 mRNA to approximately 40% of control after 24 h of cytokine treatment. The activity of an ICAM-2 promoter reporter plasmid transfected into HUVECs was down-regulated by TNF-alpha and IL-1beta to similar values. Thus inflammatory cytokines inhibit ICAM-2 transcription, despite the absence of known cytokine-responsive elements in the promoter. Immunocytochemistry on HUVEC monolayers showed that ICAM-2 expression, mainly at the cell junctions in resting cells, was markedly decreased by cytokine treatment. This data suggest that ICAM-2 expression on the endothelium may be regulated during inflammation.

Antibodies, Monoclonal↗
PubMed1997

Transcription of the SCL gene in erythroid and CD34 positive primitive myeloid cells is controlled by a complex network of lineage-restricted chromatin-dependent and chromatin-independent regulatory elements.

The SCL gene (also known as TAL-1) encodes a basic helix-loop-helix transcription factor that is essential for the development of all haematopoietic lineages, and ectopic expression of which results in T cell leukaemia. SCL is expressed in normal pluripotent haematopoietic stem cells and its expression is maintained during differentiation along erythroid, mast and megakaryocytic lineages, but is extinguished following commitment to other cell types. The mechanisms responsible for this pattern of expression are poorly understood, but are likely to illuminate the molecular basis for stem cell development and lineage commitment. We have identified multiple lineage-restricted DNase I hypersensitive sites in a 45 kb region spanning the murine SCL locus. Committed erythroid cells and CD34 positive primitive myeloid cells exhibited both shared and unique DNase I hypersensitive sites whereas none were found in T cells. The function of each hypersensitive site was studied using both transient and stable reporter assays in erythroid, primitive myeloid and T cells. Multiple positive and negative regulatory elements were characterised and found to display lineage-specificity, promoter-specificity and/or chromatin-dependence. These results represent the first description of key components of a complex network of regulatory elements controlling SCL expression during haematopoiesis.

Animals↗
PubMed1997

Distinct mechanisms direct SCL/tal-1 expression in erythroid cells and CD34 positive primitive myeloid cells.

The SCL/tal-1 gene (hereafter designated SCL) encodes a basic helix-loop-helix transcription factor which is pivotal for the normal development of all hematopoietic lineages and which is expressed in committed erythroid, mast, and megakaryocytic cells as well as in hematopoietic stem cells. The molecular basis for expression of SCL in stem cells and its subsequent modulation during lineage commitment is of fundamental importance for understanding how early "decisions" are made during hematopoiesis. We now compare the activity of SCL promoters 1a and 1b in erythroid cells and in CD34 positive primitive myeloid cells. SCL mRNA expression in CD34 positive myeloid cells did not require GATA-1. Promoter 1a activity was weak or absent in CD34 positive myeloid cells and appeared to correlate with the presence or absence of low levels of GATA-1. However, promoter 1b, which was silent in committed erythroid cells, was strongly active in transient assays using CD34 positive myeloid cells, and functioned in a GATA-independent manner. Interestingly, RNase protection assays demonstrated that endogenous promoter 1b was active in both erythroid and CD34 positive myeloid cells. These results demonstrate that fundamentally different mechanisms regulate the SCL promoter region in committed erythroid cells and in CD34 positive myeloid cells. Moreover these observations suggest that in erythroid, but not in CD34 positive myeloid cells, promoter 1b required integration in chromatin and/or additional sequences for its activity. Stable transfection experiments showed that both core promoters were silent following integration in erythroid or CD34 positive myeloid cells. Our data therefore indicate that additional regulatory elements were necessary for both SCL promoters to overcome chromatin-mediated repression.

Animals↗
PubMed1995

Lineage-restricted regulation of the murine SCL/TAL-1 promoter.

The SCL/TAL-1 gene encodes a basic helix-loop-helix transcription factor that is expressed in multipotent hematopoietic progenitors before lineage commitment. Its expression is maintained during differentiation along erythroid, mast, and megakaryocytic lineages, but is repressed after commitment to nonexpressing lineages. To begin to address the molecular mechanisms underlying this complex pattern of expression, we have studied the regulation of the murine SCL promoter in erythroid and T-cell lines. Analysis of the methylation and chromatin structure of the SCL promoter region showed that SCL mRNA expression correlated with DNase hypersensitive sites and methylation status of the promoter. Transient reporter assays showed that promoter 1a was active in erythroid cells but not in T cells. Sequences between -187 and +26 were sufficient for lineage-restricted activity of promoter 1a. A joint promoter construct containing both promoter 1a and promoter 1b also exhibited lineage-restricted activity. Conserved GATA (-37), MAZ (+242), and ETS (+264) motifs were all shown to contribute to SCL promoter activity in erythroid cells, but several other motifs were not required for full promoter activity. The pattern of complexes binding to the +242 MAZ and +264 ETS sites were the same in erythroid and T cells. However, GATA-1 bound the -37 GATA site in erythroid cells, whereas in T cells GATA-3 was only able to bind weakly, if at all. Moreover, GATA-1 but not GATA-2 or GATA-3 was able to transactivate SCL promoter 1a in a T-cell environment. These results suggest that inactivity of SCL promoter 1a in T cells reflected the absence of GATA-1 rather than the presence of trans-dominant negative regulators.

Amino Acid Sequence↗
PubMed1994

Transcription factors and the regulation of haemopoiesis: lessons from GATA and SCL proteins.

One of the central issues of developmental biology concerns the molecular mechanisms whereby a multipotent cell gives rise to distinct differentiated progeny. Differences between specialised cell types reflect variations in their patterns of gene expression. The regulation of transcription initiation is an important control point for gene expression and it is, therefore, not surprising that transcription factors play a pivotal role in mammalian development and differentiation. Haemopoiesis offers a uniquely tractable system for the study of lineage commitment and differentiation. The importance of transcription factors in the normal regulation of haemopoiesis is underlined by the frequency with which transcription factors are targeted by leukaemogenic mutations. Studies of the function and regulation of haemopoietic transcription factors, especially those expressed in lineage-restricted patterns, should greatly increase our understanding of the molecular control of haemopoiesis. In this review we have focused on insights provided by recent studies of the GATA and SCL proteins.

Animals↗
PubMed1991

Preventing low birth weight: marketing and volunteer outreach.

Low birth weight (LBW) contributes to infant mortality and continues to be a public health concern in the United States. A community-based prenatal program was established to prevent low birth weight through early and continuous prenatal care. Two key dimensions of the program were marketing and the use of volunteer neighborhood outreach workers. This paper explores the role of the program's public health nurses in relation to these dimensions and makes recommendations regarding marketing and use of outreach workers.

Arizona↗
PubMed1983

Developmental patterns of serum luteinizing hormone, gonadal and adrenal steroids in the sooty mangabey (Cercocebus atys).

Serum levels of luteinizing hormone (LH), testosterone, dehydroepiandrosterone sulfate (DHAS), androstenedione and cortisol were determined in multiple samples from 86 sooty mangabeys of varying ages (0-17 years). Testosterone, androstenedione, DHAS and cortisol were measured by radioimmunoassay; LH was determined by in vitro bioassay. Serum LH concentrations were elevated in neonates (less than 6 months) and in animals older than 72 months of age. The higher LH levels were associated with increased circulating concentrations of testosterone in males but not females. The pubertal rise in serum testosterone at approximately 55-60 months of age in males was coincident with rapid body growth. No pubertal growth spurt was observed in females. Serum levels of androstenedione and DHAS were highest during early postnatal life (less than 6 months) with androstenedione exceeding 600 ng/dl in males and 250 micrograms/dl in females, but declined rapidly in both sexes to a baseline of 150 ng/dl by 19 months of age. Serum androstenedione did not fluctuate significantly in adult animals. The pattern of age-related changes in serum DHAS paralleled those of serum androstenedione, whereas serum cortisol values did not change significantly with age. Developmental changes in serum LH, testosterone and body weight suggest that the sooty mangabey matures substantially later than the rhesus monkey. The pattern of serum gonadal and adrenal steroids during sexual maturation is similar to that seen in the baboon with no evidence of an adrenarche.

Aging↗