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The presence of prolactin (PRL) mRNA in the mammary gland, placenta, and pituitary gland of lactating and pregnant rats was investigated by polymerase chain reaction (PCR). Polyadenylated RNA was prepared from total RNA samples by oligo(dT)-cellulose chromatography, and complementary cDNAs were synthesized. A standardized amount of cDNA from each sample was used as the template in a Taq PCR under high-stringency conditions. PCR amplified a signal with the predicted size of approximately 375 bp in mammary and pituitary glands of lactating and pregnant rats, and in placentae of pregnant rats. This band specifically hybridized with a probe overlapping the entire sequence of the mature rat (r) PRL mRNA in Southern blot analysis. When the rPRL-specific primers were used, PCR revealed no signal in the liver or in lactating mammary gland explants cultured in vitro for 48 h, while the same cDNA preparations gave strong signals for beta-actin. The viability of the mammary gland explants was also suggested by their ability to secrete immunoreactive casein in vitro. PRL mRNA was localized in the epithelium of alveoli and ducts of the lactating mammary gland by in situ hybridization. These data provide evidence that the PRL gene is expressed in the mammary gland of pregnant and lactating rats, and suggest that the mammary gland might contribute to PRL in milk by de novo synthesis. Thus, while the placenta is an exogenous source of PRL-like activities for the fetus in utero, the mammary gland might take over this function after birth.

Fatty acid binding proteins (FABPs) are a multigene family of small intracellular proteins that bind hydrophobic ligands. In this report we describe the cloning and expression pattern of a novel member of this gene family that is specifically expressed in the developing and adult nervous system and thus was designated brain (B)-FABP. B-FABP is closely related to heart (H)-FABP with 67% amino acid identity. B-FABP expression was first detected at mouse embryonic day 10 in neuroepithelial cells and its pattern correlates with early neuronal differentiation. Upon further development, B-FABP was confined to radial glial cells and immature astrocytes. B-FABP mRNA and protein were found in glial cells of the peripheral nervous system such as satellite cells of spinal and cranial ganglia and ensheathing cells of the olfactory nerve layer from as early as embryonic day 11 until adulthood. In the adult mouse brain, B-FABP was found in the glia limitans, in radial glial cells of the hippocampal dentate gyrus and Bergman glial cells. These findings suggest a function of B-FABP during neurogenesis or neuronal migration in the developing nervous system. The partially overlapping expression pattern with that of cellular retinoid binding proteins suggests that B-FABP is involved in the metabolism of a so far unknown hydrophobic ligand with potential morphogenic activity during CNS development.

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Mammary derived growth inhibitor (MDGI) is a member of the family of cytoplasmic fatty acid binding proteins (FABPs), which bind hydrophobic ligands such as fatty acids, retinoids, eicosanoids and prostaglandines. MDGI and an 11 amino acid MDGI-derived conserved Cterminal peptide (P108) inhibits growth of normal mammary epithelial cells in tissue and organ culture, but fails to inhibit proliferation of many breast cancer cell lines in vitro. Here, the e ects of peptide P108 on tumor growth of MCF-7, MDA-MB468 and MDA-MB231 human breast cancer cell lines in nude mice were tested. To deliver P108 into tumors, a novel peptide production system was applied for expression and secretion of small bioactive peptides in mammalian cells. Functional di erentiation was observed in MCF-7 and MDA-MB468 cells upon P108 expression. In addition, EGF-dependent colony formation in soft agar by MDA-MB468 cells was inhibited by secreted P108. Tumor growth in athymic nude mice was suppressed in all three cell lines tested. Furthermore, P108 expressed by MCF-7/P108 cells caused paracrine tumor growth inhibition of MDA-MB231 cells. These results indicate that breast cancer inhibition by P108 is independent of binding to hydrophobic ligands and is perhaps mediated by interference with EGF-dependent signaling pathways.

Antiangiogenesis therapy has become a potentially promising tool to inhibit tumor growth by targeting an essential yet untransformed tissue component. Identifying the factors involved and understanding the mechanisms required for tumor angiogenesis will facilitate efficient and specific targeting. In neurofibromas, tumor growth is facilitated by a genetically and cytologically diverse mixture of cell types, including Schwann cells, fibroblast, mast cells, and neurons where nf-/- Schwann cells are most likely the tumorigenic cell type. The matrix forming nf+/- cells may provide a permissive environment, facilitating tumor development, perhaps by providing landscaping factors such as the angiogenic molecules fibroblast growth factor-2, platelet-derived growth factor, endothelial growth factor, vascular endothelial growth factor, and midkine, which have been detected in neurofibromas. Systemic overexpression of specific factors such as midkine owing to loss of one nf allele might further lower the overall threshold for tumorigenesis and development of a tumor vasculature. Targeting these heparin-binding growth factors might inhibit not only angiogenesis but also proliferation of tumor cells because most of these factors also stimulate proliferation of neurofibroma-derived Schwann cells. We discuss the role of specific secreted molecules for angiogenesis in tumors of neurofibromatosis 1 and possible Approaches for their targeting. Furthermore, results are discussed that demonstrate the efficacy of antiangiogenesis targeting to inhibit growth of neurofibrosarcomas in experimental animal models.

Neurofibromatosis type 1 is an autosomal dominantly inherited disease predisposing to a multitude of tumors, most characteristically benign plexiform neurofibromas and diffuse cutaneous neurofibromas. We investigated the presence and distribution of somatic mitochondrial DNA (mtDNA) mutations in neurofibromas and in nontumor tissue of neurofibromatosis type 1 patients. MtDNA alterations in the entire mitochondrial genome were analyzed by temporal temperature gradient gel electrophoresis followed by DNA sequencing. Somatic mtDNA mutations in tumors were found in 7 of 19 individuals with cutaneous neurofibromas and in 9 of 18 patients with plexiform neurofibromas. A total of 34 somatic mtDNA mutations were found. All mutations were located in the displacement loop region of the mitochondrial genome. Several plexiform neurofibromas from individual patients had multiple homoplasmic mtDNA mutations. In cutaneous neurofibromas, the same mtDNA mutations were always present in tumors from different locations of the same individual. An increase in the proportion of the mutant mtDNA was always found in the neurofibromas when compared with nontumor tissues. The somatic mtDNA mutations were present in the Schwann cells of the analyzed multiple cutaneous neurofibromas of the same individual. The observed dominance of a single mtDNA mutation in multiple cutaneous neurofibromas of individual patients indicates a common tumor cell ancestry and suggests a replicative advantage rather than random segregation for cells carrying these mutated mitochondria.

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