Fgfs direct embryogenesis of several organs, including the lung, limb, and anterior pituitary. Here we report male-to-female sex reversal in mice lacking Fibroblast growth factor 9 (Fgf9), demonstrating a novel role for FGF signaling in testicular embryogenesis. Fgf9(-/-) mice also exhibit lung hypoplasia and die at birth. Reproductive system phenotypes range from testicular hypoplasia to complete sex reversal, with most Fgf9(-/-) XY reproductive systems appearing grossly female at birth. Fgf9 appears to act downstream of Sry to stimulate mesenchymal proliferation, mesonephric cell migration, and Sertoli cell differentiation in the embryonic testis. While Sry is found only in some mammals, Fgfs are highly conserved. Thus, Fgfs may function in sex determination and reproductive system development in many species.

Objective Systemic corticosteroids are known to induce osteoporosis and increase the risk of fractures in adults and children. Inhaled corticosteroids have been shown to increase the risk of osteoporosis and fractures in adults at risk. However, long-term prospective studies in children to assess risks of multiple short courses of oral corticosteroids and chronic inhaled corticosteroids have not been done. Thus, we assessed the effects of multiple short courses of oral corticosteroids and long-term inhaled corticosteroids on bone mineral accretion over a period of years. Patients and Methods This was a cohort followup study for a median of 7 years of children with mild to moderate asthma initially randomized into the Childhood Asthma Management Program (CAMP) trial. Serial dual-energy x-ray absorptiometry (DEXA) scans of the lumbar spine for bone mineral density (BMD) were performed in all patients. Annual bone mineral accretion was calculated in 531 boys and 346 girls with asthma aged 5–12 years at baseline (84% of the initial cohort). Results Oral corticosteroid bursts produced a dose-dependent reduction in bone mineral accretion (0.052, 0.049, and 0.046 gm/cm2/year, p=0.0002) and an increase in risk of osteopenia (10%, 14% and 21%, p=0.02) for 0, 1–4, and 5+ courses, respectively, in males but not females. Cumulative inhaled corticosteroid use was associated with a small decrease in bone mineral accretion in males (p=0.05) but not females, but no increased risk of osteopenia. Conclusion Multiple oral corticosteroid bursts over a period of years can produce a dose-dependent reduction in bone mineral accretion and increased risk of osteopenia in children with asthma. Inhaled corticosteroid use has the potential for reducing bone mineral accretion in male children progressing through puberty but this risk is likely to be outweighed by the ability to reduce the amount of oral corticosteroids used in these children.

The rat intestinal fatty acid binding protein (I-FABP) gene has been used as a model to study temporal and spatial differentiation of the gut epithelium while its protein product has been used as a model for examining the atomic details of noncovalent fatty acid-protein interactions. We have isolated the mouse I-FABP gene (Fabpi) and determined its nucleotide sequence. Comparisons of the orthologous mouse, rat, and human I-FABP genes revealed three conserved domains in their otherwise divergent 5' nontranscribed sequences. RNA blot hybridization and multilabel immunocytochemical methods were used to compare the developmental stage-specific patterns of activation of the rat and mouse genes. In addition, Fabpi expression in enterocytes was examined as a function of their differentiation along the crypto-to-villus and duodenal-to-colonic axes of the small intestine. Based on the similar temporal and geographic patterns of mouse and rat I-FABP expression described here and the results of our earlier studies of transgenic mice containing rat Fabpi/human growth hormone fusion genes, we propose that one of the conserved domains, spanning nucleotides -500 to -419 in mouse Fabpi, and/or a 14-bp element, are necessary for establishing and maintaining its region-specific expression along the duodenal-to-colonic axis of the perpetually renewing gut epithelium. Finally, predictions of the structure of mouse I-FABP using the refined 2.0 A model of rat I-FABP, suggest that a proline found at position 69 of the mouse, but not rat, protein may affect its ligand binding properties.

The region of Saccharomyces cerevisiae chromosome III centromere-distal to the PGK gene is the site of frequent chromosome polymorphisms. We have sequenced this region from fragments of chromosome III isolated from three different yeast strains, GRF88, CN31C and CF4-16B. The sequence analysis demonstrates that these polymorphisms are associated with the presence of Ty and delta elements and defines a region of the chromosome which is a hot-spot for transposition events (the RAHS). The three strains can be arranged into a logical evolutionary series in which successive transposition and recombination events insert Ty elements and fuse them with consequent deletions of chromosome and of transposon sequences. The influence of such events on yeast genome evolution is discussed.

An 8-week-old infant presented to the emergency department with lethargy, tachycardia, and a blood glucose concentration of 1.8 mmol/L. After admission, hypoglycemia recurred on 3 additional occasions. Initial urinalysis results were negative for ketones, and the results of additional laboratory tests did not support the diagnosis of cortisol or growth hormone deficiency, oral hypoglycemic ingestion, or an inborn error of metabolism. Difficulty restoring and maintaining glucose concentrations along with a transient response to glucagon during 1 hypoglycemic episode suggested hyperinsulinism. In 1 hypoglycemic episode, elevated insulin and low C-peptide concentrations suggested exogenous insulin administration, but 2 subsequent blood samples obtained during hypoglycemia contained appropriately decreased concentrations of insulin. The insulin immunoassay initially used in this case (Roche ElecSys/cobas [Roche Diagnostics, Indianapolis, IN]) was insensitive to insulin analogs. Two additional immunoassays, 1 with intermediate (Immulite [Siemens, Deerfield, IL]) and 1 with broad (radioimmunoassay [Millipore, Inc, Billerica, MA]) reactivity to insulin analogs were used to characterize insulin in each of the critical blood samples. Samples obtained during hypoglycemia displayed a graded reactivity similar to that observed in type 1 diabetic patients prescribed insulin analogs, whereas a sample obtained from the patient and a control subject during euglycemia showed equal reactivity among the 3 assays. These data suggested administration of insulin analog to the child, and further characterization of insulin by using tandem mass spectrometry confirmed the presence of Humalog. The child was subsequently placed in foster care with no further recurrence of hypoglycemia.

We have tested the clones used in the European Yeast Chromosome III Sequencing Programme for possible artefacts that might have been introduced during cloning or passage through Escherichia coli. Southern analysis was performed to compare the BamHI, EcoRI, HindIII and PstI restriction pattern for each clone with that of the corresponding locus on chromosome III in the parental yeast strain. In addition, further enzymes were used to compare the restriction maps of most clones with the map predicted by the nucleotide sequence (Oliver et al., 1992). Only four of 506 6-bp restriction sites predicted by the sequence were not observed experimentally. No significant cloning artefacts appear to disrupt the published sequence of chromosome III. The restriction patterns of six yeast strains have also been compared. In addition to two previously identified sites of Ty integration on chromosome III (Warmington et al., 1986; Stucka et al., 1989; Newlon et al., 1991), a new polymorphic site involving Ty retrotransposition (the Far Right-Arm transposition Hot-Spot, FRAHS) has been identified close to CRY1. On the basis of simple restriction polymorphisms, the strains S288C, AB972 and W303-1b are closely related, while XJ24-24a and J178 are more distant relatives of S288C. A polyploid distillery yeast is heterozygous for many polymorphisms, particularly on the right arm of the chromosome.

An in vivo system has been developed for examining These villi can be rapidly identified in fixed whole-mount preparations of intestine using the a-L-fucose-specific Ulex europaeus agglutinin type I (UEA-I) lectin. They appear striped because UEA-I recognizes a cell-surface carbohydrate polymorphism between the inbred strains used to generate the chimeric animals. The strength of this system derives from the fact that two gut epitheL;al populations can be compared and contrasted that occupy virtually identical positions along the crypt-to-villus and duodenal-to-colonic axes within the same animal and differ only by the presence or absence of a single gene product. The band of blastocyst-derived epithelium in these striped, polydonal villi can be used as an internal control to assess the biological effect of the transfected gene product produced in the adjacent stripe of ES-derived cells. The system can be used for either gain-of-function or loss-of-function experiments.

Signaling through fibroblast growth factor receptors (FGFRs) is critical for the development and patterning of the vertebrate skeleton. Gain-of-function alleles of fgfr2 and fgfr3 have been linked to several dominant skeletal disorders in humans, while null mutations in fgfr3 result in the overgrowth of long bones in a mouse model system. Interestingly, the expression pattern of fgfr3 in growth plate chondrocytes overlaps that of the parathyroid hormone (PTH)-related peptide (PTHrP) receptor, a signaling molecule that also regulates endochondral ossification. The coincident expression of these two receptors suggests that their signaling pathways may also interact. To gain insight into the regulatory mechanism(s) that govern the expression of the fgfr3 gene in chondrocytes, we have identified a cellspecific transcriptional regulatory element (CSRh) by measuring the activity of various promoter fragments in FGFR3-expressing (CFK2) and nonexpressing (RCJ) chondrocyte-like cell lines. Furthermore, we demonstrate that activation of PTH/PTHrP receptors, either by stimulation with PTH or through the introduction of activating mutations, represses CSRh-mediated transcriptional activity. Finally, the transcriptional repression of the CSRh element was mimicked by treatment with forskolin, 8-bromo-cAMP, and 3-isobutyl-1-methylxanthine or by overexpression of the catalytic subunit of protein kinase A. Together, these data suggest that protein kinase A activity is a critical factor that regulates fgfr3 gene expression in the proliferative or prehypertrophic compartment of the epiphyseal growth plate. Furthermore, these results provide a possible link between PTHrP signaling and fgfr3 gene expression during the process of endochondral ossification.Elaboration of the vertebrate skeleton occurs via two overlapping, yet distinct developmental pathways. Intramembranous ossification, the primary pathway for flat bone development, relies upon the direct differentiation of condensed mesenchyme into osteoblasts. These osteoblasts then secrete various matrix components until they are encapsulated by calcified bone, thus linking the rate of intramembranous bone growth to the rate of osteoblast differentiation. Alternatively, endochondral ossification, the primary pathway for formation of the axial and appendicular skeleton, differs from intramembranous ossification in that condensed mesenchymal cells (chondrocytes) elaborate a complex cartilaginous template as they progress through a series of developmental stages at the epiphyseal growth plate. The epiphyseal growth plate is organized into distinct cellular compartments: the resting zone, which serves as a renewable source of chondrocytes; the proliferative zone, where rapid cell division results in stacked columns of chondrocytes; and the hypertrophic zone, where the cells terminally differentiate, hypertrophy, and secrete a specialized matrix. After the encapsulated hypertrophic chondrocytes mature, the associated extracellular matrix is rapidly invaded by blood vessels and bone-for...