Identification of engrailed promoter elements essential for interactions with a stripe enhancer in Drosophila embryos.

BACKGROUND: The structures and functions of promoter sequences of most genes have been analysed using in vitro transcription and/or cultured cell systems, neither possessing tissue-specific enhancers. Promoter-enhancer interactions in vivo, in particular, during ontogeny, are still poorly understood. RESULTS: We have established a new method for the assessment of promoter activity in cells that participate in fly body formation, using the UAS/GAL4 system. A functional analysis was then conducted on the promoter sequence of the engrailed gene in Drosophila embryos. A 38-bp-long sequence, terminating with an initiator or RNA start site and a downstream promoter element, was found to be capable of receiving activation signals from the engrailed stripe enhancer. Transcriptional efficiency was improved significantly by the presence of upstream promoting elements, most functionally replaceable with synthetic GAGA factor binding sites. CONCLUSIONS: We identified the in vivo minimum promoter of engrailed and demonstrated that the GAGA factor binding sites serve primarily as quantitative elements which augment transcriptional efficiency. Evidence was also obtained that indicated that not only enhancer but also promoter sequences were involved in the determination of the tissue-specificity of gene expression.

Cellular and molecular dynamics in exercise-induced urticarial vasculitis lesions.

BACKGROUND: Based on the histologic findings of fully developed lesions, leukocytoclastic vasculitis has been regarded as the histologic criterion for differentiating urticarial vasculitis from urticaria. Nevertheless, because the early lesions have not been examined histologically, the key biological events leading to the development of leukocytoclastic vasculitis remain unknown. To address this issue, urticarial vasculitis lesions induced by physical exercise were sequentially examined histologically and immunohistochemically in a patient over the course of 24 hours. Serum levels of various cytokines also were determined in parallel. OBSERVATIONS: At 3 hours after exercise challenge, the number of identifiable mast cells decreased and the first cell type that appeared around the vessels was the eosinophil. The serum tumor necrosis factor alpha level was strikingly increased as compared with that before challenge. Intense expression of E-selectin was also induced at 3 hours. The deposition of eosinophil peroxidase was observed at 3 hours and reached maximum deposition at 10 hours. The eosinophil peroxidase deposits preceded the prominent influx of neutrophils and the subsequent deposits of neutrophil elastase. CONCLUSION: The extracellular deposition of eosinophil granule proteins, in addition to the deposition of immune complexes and a variety of cytokines from the infiltrating cells, appears to be one of the key biological events that determines whether urticarial lesions resolve without causing vasculitis or develop into vasculitis.

Successful treatment of Schamberg's disease with pentoxifylline.

Three patients with Schamberg's disease were treated with pentoxifylline, 300 mg daily for 8 weeks. A significant response was observed within 2 to 3 weeks. One patient had recurrence after discontinuation of this treatment; but promptly responded to resumption of therapy. No adverse effects were noted in any patients. We investigated the expression pattern of adhesion molecules in relation to perivascular accumulation of lymphocytes in specimens obtained before and after treatment with pentoxifylline in these patients. Our results suggest that pentoxifylline exerts its therapeutic effects on Schamberg's disease at the level of T-cell adherence to endothelial cells and keratinocytes.

Time-course analyses of exercise-induced lesions in a patient with urticarial vasculitis.

Using immunoperoxidase techniques, we examined the histological changes of exercise-induced lesions of urticarial vasculitis over a 24 h period. At 3 h, endothelial swelling and an eosinophilic infiltrate was seen. Neutrophils were increased in number by 10 h. Typical leucocytoclastic vasculitis with red blood cell extravasation developed by 24 h. Maximal deposition of eosinophil peroxidase was at 10 h compared to significant neutrophil elastase deposits at 24 h. This demonstrates the importance of the eosinophil in the development of the lesions of urticarial vasculitis.

Induction of a mirror-image duplication of anterior wing structures by localized hedgehog expression in the anterior compartment of Drosophila melanogaster wing imaginal discs.

The segment polarity gene hedgehog (hh) encodes a secretory protein involved in cell-cell communication in Drosophila melanogaster. The hh gene is expressed in the posterior compartment and is essential for the establishment and maintenance of the anterior/posterior-compartment boundary of each embryonic parasegment [Ingham, P.W., Nature 366 (1993) 560-562]. To clarify possible hh functions in adult appendage formation, we isolated a fly line (h9D) associated with a wing malformation from among fly lines with an hh transgene whose expression is under the control of trapped enhancers. In h9D flies, the ectopic expression of hh occurred in the anterior edge of wing pouch in the wing disc. This abnormal hh expression resulted in not only a mirror-image duplication and ectopic outgrowth in the anterior wing compartment, but also the ectopic expression of patched and decapentaplegic, strongly suggesting that the hh product serves as a morphogen or an inducer essential for wing development, including the proximal/distal axis formation.

Structure and expression of hedgehog, a Drosophila segment-polarity gene required for cell-cell communication.

The complete nucleotide sequence of the coding region of hedgehog (hh), a segment-polarity gene in Drosophila melanogaster, was determined. The gene was found to include three exons which would encode a 421- (or 471-) amino acid (aa) polypeptide with a long hydrophobic stretch. The hh mRNA was about 2.3 kb long and expressed throughout development. The hh expression in an embryo occurred in stripes, while that in imaginal discs occurred in the posterior compartment. As a whole, the spatial expression pattern of hh mRNA was very similar to that of engrailed (en), a homeobox gene required for the formation of the anterior-posterior compartment boundary. Unlike en, no hh expression was observed in the central nervous system.

Histological changes in recurrent soft tissue sarcomas: analysis of 56 patients.

Histological changes in 56 patients with recurrent soft tissue sarcomas (STS) were analysed. The original tumors included 23 malignant fibrous histiocytomas, five liposarcomas, four leiomyosarcomas, four synovial sarcomas, four neurogenic sarcomas, four dermatofibrosarcomas and 12 other assorted tumor types. The histological features of the recurrent tumors which underwent changes included increased cellularity, an increased number of mitotic figures, extension of sclerotic areas and the appearance of a storiform pattern; these were found in 30%, 43%, 41% and 29% of cases, respectively. No case showed a histological change from one subtype to another. A follow-up study revealed increased cellularity and mitotic counts in the recurrent tumors to be signs of an unfavorable prognosis.