We compared the utility of different adiposity measures in predicting indices of insulin sensitivity and secretion. At their pre-randomization visit for a 1 year, randomized, double masked, placebo controlled trial of metformin extended release, 47 (to date, of a planned 135) obese adolescents had a 7 point, 180 minute, 75-gm OGTT. Inclusion criteria included a BMI greater than the 95 th centile and weight of less than 137 kg. Subjects had a mean age of 14.6 (13-18) years. 21 (45%) were male and 55% were non-Hispanic white. Insulin sensitivity was estimated using the Composite Insulin Sensitivity Index [CISI, 1/((mg/dL)(µU/mL))]. Insulin secretion was calculated using the Corrected Insulin Release at glucose peak [CIRgp, ((µU/L)/(mg/dL) 2 )]. Total adiposity was measured using BMI and dual energy x-ray absorptiometry (DXA-whole body (WB)). Trunk adiposity was measured by DXA-TK and a CT slice aligned with the L4-L5 intervertebral disk. This CT slice was also used to partition intraperitoneal (CT-IP) and subcutaneous (CT-SQ) fat. While the mean body weights were similar (98.9 vs 100 kg), girls, as expected, had greater adiposity than boys (DX-WB 42[138}6% vs 36Ϯ6% fat). Moreover, non-Hispanic whites, while having similar adiposity, had greater insulin sensitivity (CISI 3.88Ϯ3.00 vs 3.38Ϯ4.55). CISI was highly correlated with fasting insulin concentrations (r = Ϫ0.95, p ≤ 0.0001). Intraperitoneal fat (CT-IP), while constituting only 14% of abdomen fat, had the strongest correlation with CISI of all the measures of adiposity (Table ).

were confirmed to be adipocytes by light microscopy. Gene expression levels were determined by quantitative real-time PCR. Results: Although adipocyte size measurements between IR and IS groups did not differ significantly, the IR group had a higher small to large cell ratio (1.66Ϯ 1.03 vs. 0.94 Ϯ 0.50, P ≤0.05). Adiponectin, PPAR␥1, PPAR␥2, and GLUT4 all showed 2-3 fold lower levels of expression in the IR group (n=5) compared to the IS group (n=5). In contrast, there were no significant differences in Adipsin and SREBP-1c expression in both groups. Conclusions: Our observation of a bimodal distribution of small and large cells in both IR and IS groups, and a larger ratio of small adipocytes in IR compared to IS individuals is a novel finding. Gene expression analysis suggests that there is probably a decrease in differentiation since GLUT4 and Adiponectin, known markers of terminal adipocyte differentiation, were significantly decreased in the IR group. However since SREBP-1c and Adipsin were not differentially expressed in both groups more work will need to be done to further characterize the adipocyte differences in the IR and IS groups and to determine the significance of adipocyte size and insulin resistance status.

Gonadotropinomas, which produce α, LHβ, and/or FSHβ subunits, represent 35% of pituitary tumors. They pose a diagnostic challenge, presenting predominantly in men with headaches, decreased libido and impotence. Little is known of their pathogenesis, and identification of tumor-specific genes may help elucidate the molecular mechanisms that are involved. We used the Affymetrix GeneChip Human Genome U133 Plus 2.0 Array to compare gene expression profiles in five gonadotropinomas with six pituitaries obtained from autopsies. Analysis with the Affymetrix GeneChip Operating System showed that 1549 of 39,000 genes examined were differentially expressed. We then used RT-PCR to analyze further several genes, which have been implicated in proliferation, that had unique expression patterns in our array. Bone morphogenetic proteins (BMPs) are growth factors involved in many aspects of tissue development and morphogenesis. Recently, members of the BMP family have been implicated in the production of FSH by gonadotropes and have been detected in gonadotropinomas. In contrast to prior literature, our microarray analysis showed that BMP-5 was selectively overexpressed 4.8-fold in gonadotropinomas compared with normal pituitary, which was confirmed by semi-quantitative RT-PCR. Thus, BMP-5 may have a previously undescribed role in FSH production and/or in gonadotrope proliferation. The promyelocytic leukemia zinc finger (PLZF) protein belongs to the POZ/zinc-finger family of transcription factors and acts as a negative regulator for GATA-2. GATA-2 and GATA-3 are found in pituitary cell lines and can regulate α-subunit gene expression, and GATA-2 has been detected in pituitary tumors. In our study, PLZF was underexpressed 13.9-fold in gonadotropinomas compared with normals, whereas GATA-2 and GATA-3 were overexpressed 6.7-fold and 26.5-fold, respectively. These data suggest that loss of the transcriptional repressor PLZF, together with activation of GATA family members, may be involved in the development of gonadotropinomas. Thus, various growth and transcription factors are differentially expressed in gonadotropinomas. The evaluation of candidate genes that emerge from these experiments provides a rational approach to investigate mechanisms underlying pituitary tumorigenesis.

Gonadotropinomas that make α, LHβ and/or FSHβ glycoprotein hormone subunits are the most prevalent type of pituitary macroadenoma. These tumors present a diagnostic challenge, as they are rarely associated with a characteristic clinical hypersecretory syndrome and thus difficult to recognize until large enough to cause mass effects. Mechanisms underlying the pathogenesis of gonadotropinomas have not been identified. Common oncogene mutations involved in other human cancers, such as Ras and Rb, are not involved in pituitary tumor formation. The aims of the current study were to elucidate molecular events responsible for the development of gonadotropinomas. We compared gene expression profiles between six normal human pituitaries and five glycoprotein-secreting tumors by cDNA microarray analysis. Pairwise comparisons of normal pituitaries and gonadotropinomas demonstrated consistent, greater than two-fold increased expression in 671 genes and decreased expression in 878 genes. Gene Ontology Database analysis was used to categorize differentially expressed genes into classes that may affect tumorigenic or cell proliferative properties. Relative to tumor versus normal, the following data were obtained: transcription factors (184 up/165 down), cell cycle regulation (104 up/62 down), apoptosis regulation (68 up/50 down), cancer genes (72 up/55 down), and signal transduction (158 up/240 down). Candidate genes were then selected based on the magnitude and consistency of change. RT-PCR was performed to confirm array-based results of differential expression in candidate genes. Deleted in Bladder Cancer Chromosome Region 1-Like (DBCCR1-L) is an attractive candidate as it was completely absent in all normal pituitary samples, but expressed in all tumor samples. DBCCR1-L is a novel gene. Its homologue, DBCCR1, is involved in G1 cell cycle transition and colon, breast, esophageal, ovarian, and bladder cancer. Investigation is underway to evaluate the consequences of DBCCR1-L knockdown and overexpression in pituitary cell lines and its effects on cell cycle progression. Our results demonstrate the differential expression of many genes, including DBCCR1-L, which may be involved in pituitary tumorigenesis. Future studies on candidate genes identified in this analysis may lead to the identification of tumor markers to detect, prognostically evaluate, and eventually medically treat gonadotropinomas. Sponsored by VA Merit to MEW and NIH and UCHSC Cancer Center Fellowships to DSM and MML