Heather L. Burrows

POSTTREATMENT EXPOSURE TO CAMPTOTHECIN ENHANCES THE LETHAL EFFECTS OF X-RAYS ON RADIORESISTANT HUMAN MALIGNANT MELANOMA CELLS

Little is known about the molecular mechanisms responsible for the survival recovery process(es) (known as potentially lethal damage repair), which occurs in mammalian cells following ionizing radiation. Previously, we presented data indicating a role for the DNA unwinding enzyme, topoisomerase I, in DNA repair. We now demonstrate that camptothecin, a specific inhibitor of topoisomerase I, causes dramatic radiosensitization of an extremely resistant human melanoma (U1-Mel) cell line. Camptothecin radiosensitized U1-Mel cells when it was administered either during or immediately following x-irradiation. U1-Mel cells were optimally radiosensitized with 4 microM camptothecin for a period of 4-6 hrs after x-irradiation. Enhanced cell killing by camptothecin was proportional to the initial extent of damage created by x-irradiation; the higher the dose of ionizing radiation, the greater the radiosensitization. The apparent synergy observed with camptothecin and x-rays was irreversible; camptothecin-treated U1-Mel cells were not able to carry out PLDR in a 48 hr period after the drug was removed. We hypothesize that the administration of camptothecin causes lesion modification through a topoisomerase I-mediated mechanism. These data support a role for topoisomerase I in DNA repair and indicate that camptothecin, or more effective derivatives, may have clinical use.

Genealogy of the Anterior Pituitary Gland: Tracing a Family Tree.

The anterior lobe of the pituitary gland is derived from the oral ectoderm early in gestation. A variety of techniques have been used to understand how early precursor cells differentiate to form the five major cell types that populate the adult anterior lobe. Current evidence suggests that corticotropes arise from a lineage distinct from that of the other four cell types. The cells of the other lineage branch - thyrotropes, gonadotropes, somatotropes and lactotropes - appear to be related because of their dependence on common transcription factors and the frequent occurrence of cells that produce multiple hormones. While thyrotropes arise through two routes, the lineage related to somatotropes and lactotropes appears to be the most important for hormone production. Each cell type can populate the organ and function in the absence of the other cell types, except for lactotropes, which have a strong dependence on somatotropes. Our current knowledge of anterior pituitary cell lineage relationships may contribute to a better understanding of the origin of pituitary adenomas and tumors.

Cre-mediated recombination in the pituitary gland.

Summary: Organ‐specific expression of a cre recombinase transgene allows for the analysis of gene function in a particular tissue or cell type. Using a 4.6 kb promoter from the mouse glycoprotein hormone α‐subunit (αGSU or Cga) gene, we have generated and characterized a line of transgenic mice that express cre recombinase in the anterior and intermediate lobes of the pituitary gland. Utilizing a cre‐responsive reporter transgene, αGSU‐cre transgene expression was detected in the pituitary primordium and in all five cell types of the adult anterior pituitary. αGSU‐cre transgene activity was also detected in the cardiac and skeletal muscle. Little or no activity was evident in the gonads, adrenal glands, brain, ventromedial hypothalamus, or kidneys. The αGSU‐cre transgenic mice characterized here will be a valuable tool for examining gene function in the pituitary gland. genesis 28:167–174, 2000.

Mapping of the mouse corticotropin-releasing hormone receptor 2 gene (Crhr2) to Chromosome 6

Species: Mouse Locus name: growth-retarded Locus symbol: grt Map position: The grt is localized on mouse Chromosome (Chr) 5, 59 cM distal from the centromere: centromere-D5Mit254-18.0 +_ 3.1 cM-grt, D5Mit240, D5Mit314, D5Mit338-15.3 +_ 2.9 cMD5Mit29-telomere Method of mapping: (C57BL/6 x DW/J-grt)F 1 • DW/J-grt backcross mice. Haplotype analysis is shown in Fig. 1A. Database deposit information: MGD accession number, MGDJNUM-40511 Molecular reagents: Microsatellite primers for PCR used in this study were all purchased from Research Genetics, Inc. (Huntsville, Ala., USA). Allele detection: Genotype of the grt/grt or grt/+ was determined by weighing mice at 6 weeks of age. Linkage analysis was performed in 150 backcross progeny by the grt genotype with DW/J-grt allele for several microsatellite loci representative of each chromosome. Since some of microsatellite loci localized on Chr 5 showed a cross-linkage to the grt allele, other microsatellite loci on Chr 5 were further examined. Finally, three microsatellite alleles were found to show no recombination with the grt allele (Fig. 1). Thus, the grt locus was mapped on 59 cM distal from the centromere of Chr 5, where human Chr 22ql 1.2-q13.1 shows synteny. Discussion: Several mouse mutants exhibiting dwarfism have been reported. The responsible genes for them have already been mapped and cloned [1-6]; dw (Chr 16), df (Chr 11), lit (Chr 6), and hyt (Chr 12). Mutant mice exhibiting dwarfism occurred recently in our DW/J-+/dw colony. Their phenotype was not as the dw phenotype in physiological and histological studies [7,8]; dwarfism in this mouse was thought to be caused by an abnormality in the thyroid function, whereas the primary cause in dw mice was functional deficiency in the transcription factor, Pit-l , specifically localized in the pituitary. Thus, we termed this putative locus growth-retarded (grt). In this report, the grt locus was shown to be localized on Chr 5, verifying that the grt gene is different from the dw or other murine mutant genes causing dwarfism reported so far.

Localization of the corticotropin-releasing hormone receptor gene on mouse Chromosome 11

rDepartment of Human Genetics, The University of Michigan Medical School, Ann Arbor, Michigan 48109-0618, USA 2Graduate Program in Cellular and Molecular Biology, The University of Michigan Medical School, Ann Arbor, Michigan 48109-0618, USA 3Department of Biological Chemistry, The University of Michigan Medical School, Ann Arbor, Michigan 48109-0618, USA 4Mental Health Research Institute, The University of Michigan Medical School, Ann Arbor, Michigan 48109-0618, USA

20 – Pituitary Gland Development

Pituitary gland is a small endocrine gland located at the base of the brain under the optic chiasm and the arachnoid membrane, in a bony indentation called the sella turcica. Its name comes from the Greek hypophysis, which means undergrowth. The role of the pituitary gland is the regulated synthesis and secretion of polypeptide hormones that are essential for the development and function of many other organs in the body. Releasing hormones and inhibiting factors reach the anterior pituitary via hypothalamic neurons that terminate in the capillary beds of the median eminence, just dorsal to the pituitary gland. These capillary beds are connected to the hypophyseal portal vessels that nourish the anterior pituitary. In response to these stimulatory stimulatory factors, pituitary hormones are released into hypophyseal portal blood vessels and carried through the blood stream to their target organs. An intriguing feature of pituitary development is the initial organization of the cell types into discrete patches and the loss of this spatial organization as the organ expands. The rodent pituitary is composed of three lobes. The posterior pituitary (neurohypophysis or pars nervosa) is derived from neural ectoderm and contains the nerve terminals that secrete oxytocin and vasopressin. The intermediate lobe (pars intermedia) and anterior lobe are both derived from oral ectoderm. The bulk of the anterior lobe (pars distaIis) is at the same level as the intermediate lobe, but a portion of it known as the pars tuberalis extends dorsally along the pituitary stalk.

Motivational Advising Workshop: Utilizing Motivational Interviewing Theory to Facilitate and Engage Intrinsic Motivation to Change Learners’ Behavior

Introduction Motivational interviewing (MI) is a counseling method that utilizes a patients own motivation to effect personal change. MI has been applied routinely and successfully to managing medical conditions (e.g., substance abuse). Employing MI techniques to engage medical learners (termed motivational advising [MA]) may help them overcome professional and/or personal challenges limiting their career development. Methods Medical educators from four academic medical centers developed a module focused on teaching fellow educators MI theory and techniques for MA using didactic and interactive components. Participants participated in facilitated role-plays to practice MA delivery techniques and observed videos of a traditional advisor-advisee interaction as well as an MA-focused engagement. A postworkshop survey was used to evaluate the workshop. Results In a survey of 48 educators attending the workshop at two medical conferences, over 80% of respondents demonstrated an interest in learning more about MA. Additionally, over 60% indicated that they would seek opportunities to practice and/or implement MA with their advisees. Knowledge of the technical components of MA also increased significantly in pre- and posttest analysis. Discussion This module introducing the concept of MA was well received by medical educators and was viewed as a valuable tool in advising medical learners. The provided components enable replication of this workshop in other settings with or without an expert in MI techniques. Although the workshop has been conducted with physicians involved in medical education, it would be applicable to other health professionals who advise trainees such as nursing, dentistry, pharmacy, or veterinary medicine.