Biomedical subjects

Anthony L Cook

Publications and source records attributed to Anthony L Cook.

8 recordsLinked to original sources

PubMed2005

Co-expression of SOX9 and SOX10 during melanocytic differentiation in vitro.

Investigations into pigment cell biology have relied on the ability to culture both murine and human melanocytes, numerous melanoma cell lines and more recently, murine and human melanoblasts. Melanoblast culture requires medium supplemented with a range of growth factors including Stem Cell Factor, Endothelin-3 and Fibroblast Growth Factor-2, withdrawal of which causes the cells to differentiate into melanocytes. Using the human melanoblast culture system, we have now examined the expression and/or DNA binding activity of several transcription factors implicated in melanocytic development and differentiation. Of these, the POU domain factor BRN2 and the SOX family member SOX10 are both highly expressed in unpigmented melanocyte precursors but are down-regulated upon differentiation. In contrast, the expression levels of the previously described MITF and PAX3 transcription factors remain relatively constant during the melanoblast-melanocyte transition. Moreover, BRN2 ablated melanoma cells lack expression of SOX10 and MITF but retain PAX3. A novel finding implicates a second SOX protein, SOX9, as a potential melanogenic transcriptional regulator, as its expression level is increased following the down-regulation of BRN2 and SOX10 in differentiated melanoblasts. Our results suggest that a complex network of transcription factor interactions requiring proper temporal coordination is necessary for acquisition and maintenance of the melanocytic phenotype.

Cell Differentiation↗

PubMed2004

Gene-expression profiling reveals distinct expression patterns for Classic versus Variant Merkel cell phenotypes and new classifier genes to distinguish Merkel cell from small-cell lung carcinoma.

Merkel cell carcinoma (MCC) is a rare aggressive skin tumor which shares histopathological and genetic features with small-cell lung carcinoma (SCLC), both are of neuroendocrine origin. Comparable to SCLC, MCC cell lines are classified into two different biochemical subgroups designated as 'Classic' and 'Variant'. With the aim to identify typical gene-expression signatures associated with these phenotypically different MCC cell lines subgroups and to search for differentially expressed genes between MCC and SCLC, we used cDNA arrays to profile 10 MCC cell lines and four SCLC cell lines. Using significance analysis of microarrays, we defined a set of 76 differentially expressed genes that allowed unequivocal identification of Classic and Variant MCC subgroups. We assume that the differential expression levels of some of these genes reflect, analogous to SCLC, the different biological and clinical properties of Classic and Variant MCC phenotypes. Therefore, they may serve as useful prognostic markers and potential targets for the development of new therapeutic interventions specific for each subgroup. Moreover, our analysis identified 17 powerful classifier genes capable of discriminating MCC from SCLC. Real-time quantitative RT-PCR analysis of these genes on 26 additional MCC and SCLC samples confirmed their diagnostic classification potential, opening opportunities for new investigations into these aggressive cancers.

Biomarkers, Tumor↗

PubMed2004

Use of florfenicol in non-human primates.

Studies were undertaken to determine if florfenicol, an antimicrobial agent structurally similar to chloramphenicol, could be used as an effective broad spectrum antibiotic for the treatment of bacterial infections in primates. Florfenicol was developed as an injectable antibiotic for use in cattle on an every other day dosing schedule. Its broad spectrum activity, long duration of action following i.m. administration, and its safety as compared with chloramphenicol made it an attractive antibiotic for use in non-human primates. Previous studies had shown that florfenicol is effective against common primate pathogens such as Salmonella, Klebsiella, Escherichia coli, Bordetella bronchiseptica, Streptococcus pneumoniae, Staphylococcus spp., and Yersinia pseudotuberculosis. We performed experiments on a total of 15 macaques. The animals were given florfenicol at 50 mg/kg i.m. and blood samples taken at various time points. Serum was evaluated for florfenicol absorption. Necropsies were also performed to determine if major organs were affected and to determine the effects of i.m. injection of florfenicol. We determined that florfenicol given every 48 hours in rhesus macaques results in blood levels that were acceptable for therapeutic use. The effect on muscle tissue of i.m. injection was similar to ketamine and normal saline. There were no gross lesions observed and no changes with tissues submitted for histology. Our work shows that with further studies, florfenicol may be useful when injectable antibiotic therapy is required in non-human primates.

Analysis of Variance↗

PubMed2004

Spontaneous uterine leiomyosarcoma in a rhesus macaque.

A 12-year-old female Rhesus macaque (Macaca mulatta) presented with a uterine enlargement during physical exam. Exploratory celiotomy revealed an intrauterine mass that extended into the cervix. A cystic lesion surrounded and enveloped the left ovary. Because of numerous adhesions and the size of the mass, the animal was euthanized. Microscopically, the neoplasia showed small to medium-sized spindle-shaped cells which formed bundles. The cells resembled smooth muscle, with elongated nuclei, indistinct nucleoli, and scant to moderate quantities of eosinophilic cytoplasm. Mitotic activity was low (0 to 2 mitotic figures/high-power field). Immunohistochemistry revealed that the cells stained for smooth muscle actin. In light of the biological activity (locally invasive) and immunohistochemical findings, a diagnosis of leiomyosarcoma of the myometrium was made. Grossly and histologically, uterine leiomyosarcoma can resemble uterine leiomyoma. Although uterine leiomyosarcoma represents a small percentage of uterine tumors, it should always be considered as a rule out for uterine leiomyoma (fibroids).

Animals↗

PubMed2004

Beta 2 microglobulin and resistance to murine respiratory mycoplasmosis.

Mycoplasma pulmonis (MP) infection causes murine respiratory mycoplasmosis (MRM) in mice. This study was designed to determine whether C57BL/6 beta 2 microglobulin knockout (KO) mice (B2m-/-) differed from C57BL/6 B2m(+/+) wild-type (WT) mice in the early events of acute infection and to confirm previous studies that suggested loss of beta 2 microglobulin (B2m) expression was correlated with increased disease susceptibility. Experimentally infected mice were necropsied at various time points after infection. Quantitative cultures were performed on lung, trachea, and nares at each time point. Tissues were collected for histopathology. Both genotypes had marked clearance of MP from all sites within the first 24 h post-infection (p.i.). There were no differences between KO and WT mice in the ability to clear MP from the lung, nares, or trachea during the first 48 h p.i. During chronic infection, there was a significant difference among days p.i. for number of MP isolated from the lungs, nares, and trachea (P<0.05). There were significant differences (P<0.05) between genotypes for number of MP isolated from the lung. Numbers of MP isolated from KO mice were greater than from WT mice at day 10 through day 28 p.i. There was no significant overall difference between genotypes for number of MP isolated from the nares or trachea. However, increased numbers of MP were isolated from the nares of KO mice at days 21 and 28 p.i. and from the trachea of KO mice at days 7 and 21 p.i. During chronic infection, KO mice had increased facial scratching and altered respiration as well as marked weight loss compared with those of WT mice. The KO and WT mice differed (KO more severe) with respect to the presence and extent of neutrophilic exudate in the lung. Differences approached significance for consolidation of lung parenchyma but not for lymphoid infiltration, tending to be more severe in KO mice. Loss of B2m does not appear to affect clearance of MP during the critical first 48 h of infection but does seem to affect chronic infection, which may be because of the loss of CD8(+) Tc1 cells, the CD1-B2m complex, or both.

Analysis of Variance↗

PubMed2003

Screening of human primary melanocytes of defined melanocortin-1 receptor genotype: pigmentation marker, ultrastructural and UV-survival studies.

Recent population studies have demonstrated an association with the red-hair and fair-skin phenotype with variant alleles of the melanocortin-1 receptor (MC1R) which result in amino acid substitutions within the coding region leading to an altered receptor activity. In particular, Arg151Cys, Arg160Trp and Asp294His were the most commonly associated variants seen in the south-east Queensland population with at least one of these alleles found in 93% of those with red hair. In order to study the individual effects of these variants on melanocyte biology and melanocytic pigmentation, we established a series of human melanocyte strains genotyped for the MC1R receptor which included wild-type consensus, variant heterozygotes, compound heterozygotes and homozygotes for Arg151Cys, Arg160Trp, Val60Leu and Val92Met alleles. These strains ranged from darkly pigmented to amelanotic, with all strains of consensus sequence having dark pigmentation. UV sensitivity was found not to be associated with either MC1R genotype or the level of pigmentation with a range of sensitivities seen across all genotypes. Ultrastructural analysis demonstrated that while consensus strains contained stage IV melanosomes in their terminal dendrites, Arg151Cys and Arg160Trp homozygote strains contained only stage II melanosomes. This was despite being able to show expression of tyrosinase and tyrosinase-related protein-1 markers, although at reduced levels and an ability to convert exogenous 3,4-dihydroxyphenyl-alanine (DOPA) to melanin in these strains.

Alleles↗

PubMed2003

Human melanoblasts in culture: expression of BRN2 and synergistic regulation by fibroblast growth factor-2, stem cell factor, and endothelin-3.

The BRN2 transcription factor (POU3F2, N-Oct-3) has been implicated in development of the melanocytic lineage and in melanoma. Using a low calcium medium supplemented with stem cell factor, fibroblast growth factor-2, endothelin-3 and cholera toxin, we have established and partially characterised human melanocyte precursor cells, which are unpigmented, contain immature melanosomes and lack L-dihydroxyphenylalanine reactivity. Melanoblast cultures expressed high levels of BRN2 compared to melanocytes, which decreased to a level similar to that of melanocytes when cultured in medium that contained phorbol ester but lacked endothelin-3, stem cell factor and fibroblast growth factor-2. This decrease in BRN2 accompanied a positive L-dihydroxyphenylalanine reaction and induction of melanosome maturation consistent with melanoblast differentiation seen during development. Culture of primary melanocytes in low calcium medium supplemented with stem cell factor, fibroblast growth factor-2 and endothelin-3 caused an increase in BRN2 protein levels with a concomitant change to a melanoblast-like morphology. Synergism between any two of these growth factors was required for BRN2 protein induction, whereas all three factors were required to alter melanocyte morphology and for maximal BRN2 protein expression. These finding implicate BRN2 as an early marker of melanoblasts that may contribute to the hierarchy of melanocytic gene control.

Antigens, Neoplasm↗

PubMed2002

Proneural and proneuroendocrine transcription factor expression in cutaneous mechanoreceptor (Merkel) cells and Merkel cell carcinoma.

Merkel cells form part of the peripheral neuroendocrine system of the skin and act as mechanoreceptors in touch response. Merkel cell carcinoma (MCC) is a rare, aggressive disease with similarities to small cell lung cancer (SCLC), which is also of neuroendocrine origin. We previously identified a novel DNA binding protein complex specific for MCC suspension cell lines, termed Merkel nuclear factor (MNF) by its binding to the POU-IV family DNA binding consensus sequence. Here we report that MNF contains the POU-IV family member Brn-3c and that Brn-3c is expressed in normal Merkel cells. Additionally, Brn-3c protein reactivity is restricted to a subset of MCC biopsies and is not seen in biopsies revealing adherent, variant cell lines lacking neuroendocrine markers. Recently, proper development of murine Merkel cells was shown to require the proneural basic helix-loop-helix transcription factor, atonal family member, MATH1. We demonstrate a correlation between Brn-3c and HATH1 reactivity in MCC biopsies and cell lines with retention of neuroendocrine phenotype. In SCLC, the related basic helix-loop-helix transcription factor HASH1 is responsible for neuroendocrine phenotype, but HASH1 transcripts were not detected in MCC cell lines. We propose that HATH1 and Brn-3c may form a transcriptional hierarchy responsible for determining neuroendocrine phenotype in Merkel cells and that lack of Brn-3c and/or HATH1 in MCC may indicate a more aggressive disease requiring closer patient follow-up.

Basic Helix-Loop-Helix Proteins↗