The growth potentiating effects of the insulin-like growth factor (IGF)-I and IGF-II are modulated by a family of six insulin-like growth factor binding proteins (IGFBPs). Despite the similarity in amino acid sequences of the IGFBPs, their effects on the growth of bone cells differ. Studies on the molecular mechanisms for IGFBP-4 actions revealed that coincubation of bone cells with IGFBP-4 and 125I-IGF-I or 125I-IGF-II decreased the binding of both of these ligands in a dose-dependent manner. In addition, IGFBP-4 decreased the binding of IGF-I tracer to purified type I IGF receptor. These data in conjunction with data showing that IGFBP-4 had no effect on cell proliferation induced by analogs of IGF-I or IGF-II, which exhibited > 100-fold reduced affinity for binding to IGFBP-4 suggest that IGFBP-4 may inhibit IGF action by preventing the binding of ligand to its membrane receptor. In contrast to IGFBP-4, IGFBP-5 treatment increased the binding of IGF tracer to bone cells but did not increase the binding of 125I-IGF-I to type I IGF receptor. Studies on the mechanism by which IGFBP-5 increased the binding of 125I-IGF tracer to bone cells suggest that IGFBP-5 could facilitate IGF binding by a mechanism in which IGFBP-5 has cell surface binding sites independent of IGF receptors. These data in conjunction with the findings that IGFBP-5 potentiated cell proliferation even in the presence of those same IGF analogs that exhibited > 200-fold reduced affinity for binding to IGFBP-5, suggest that IGFBP-5 may in part stimulate bone cell proliferation by an IGF-independent mechanism involving IGFBP-5-specific cell surface binding sites.

A series of substituted 2,3-dihydrothiazolo[2,3-a]isoindol-5(9bH)-ones and related compounds 1-73 were synthesized and evaluated for their ability to inhibit reverse transcriptase (RT) of the human immune deficiency virus 1 (HIV-1) and replication of HIV-1 in MT2 cells. The antiviral activity of these compounds depends on the stereoselective configuration of the substituent in position 9b. Structure-activity studies were done within these series of compounds to determine the optimum substituents for antiviral activity. The most potent inhibitors were found in the class of 2,3-dihydrothiazolo[2,3-a]isoindol-5(9bH)-ones bearing a phenyl ring system in position 9b optionally substituted with one or two methyl groups or a chlorine atom in position 8. The most active analogues (R)-(+)-1, (R)-(+)-6, (R)-(+)-13, (R)-(+)-26, and (R)-(+)-53 inhibit the HIV-1 RT with an IC50 between 16 and 300 nM and an IC50 between 10 and 392 nM in MT2 cells, respectively.

The signaling pathways of bone morphogenic protein 2 (BMP-2) and Sonic hedgehog (Shh) are related during embryogenesis. Both proteins have been implicated as important components during osteogenic differentiation; e.g., considering their in vitro effects in the pluripotent C3H10T/1/2 cell system. Also, BMP-2 has been frequently reported to stimulate adipogenesis as well as osteogenesis in these cells. We investigated the relative potencies of Shh and BMP-2 with regard to adipogenesis. We performed differentiation experiments by stimulating C3H10T1/2 cells with BMP-2, Shh, or a combination. We monitored adipocyte-like differentiation via gene expression analysis and cytologic staining. An adipocytic phenotype was observed in BMP-2-treated cells, as shown by upregulation of two adipocytic marker mRNAs, PPAR-gamma and aP2, and by staining of lipid-filled cell vesicles with Oil Red O. In contrast, no adipocyte-like differentiation could be detected either after treatment with Shh or after exposure to a combination of Shh and BMP-2. Our results demonstrate for the first time that Shh and BMP-2 have contrary effects on adipocyte-like differentiation. Whereas BMP-2 promotes the adipocytic lineage, Shh suppresses the expression of the BMP-2-induced fat-cell phenotype.

The Epstein-Barr virus (EBV) immediate early gene product BZLF1 was localized by indirect immunofluorescence to the cytoplasm of the basal epithelial layer at the lateral border and dorsum of tongue in human immunodeficiency virus-infected and -seronegative patients. Two biopsies of oral hairy leukoplakia revealed a sporadic cytoplasmic staining of the BHRF1 and BRLF1 gene products in the basal epithelial layer. The widespread presence ofBZLF1 in the basal epithelial layer indicated that this cell layer contained EBV DNA and was probably directly infected by EBV. Nuclear localization of the immediate early and early gene products BZLF1, BHRF1, BRLF1, and BMLF1 was limited to oral hairy leukoplakia in human immunodeficiency virus-seropositive patients and revealed a codistribution with the virus capsid antigen. Our results indicate that the epithelium of the tongue is a potential reservoir for EBV and that in heavily immunocompromised patients EBV may move from the cytoplasm to the nucleus with increasing differentiation and be coactivated there during the terminal differentiation ofepithelial cells at the lateral border and dorsum of tongue. The cellular and viral factors that determine whether EBV infection is latent or productive are mainly unknown and depend on the host cell (10). EpsteinThe concept of strict B-lymphocyte tropism has been central to discussion of the biology of EBV since the discovery of this herpesvirus. The observation that epithelial cells in nasopharyngeal carcinoma are latently infected with EBV and that lytic expression of EBV occurs in epithelial cells of the parotid gland (11) and other sites of the human body (12) clearly points to a complex biology of this herpesvirus.Virus binding to B cells occurs via a specific interaction between the major envelope glycoprotein gp340 and the C3d receptor molecule (CD21; refs. 13 and 14) and it seems that the specificity of gp340-CD21 binding plays a major role in determining the viral B lymphotropism (15).The presence of EBV in the upper two-thirds of the epithelium in oral hairy leukoplakia (OHL) was originally described by Greenspan et al. (16) and was later confirmed by ultrastructural studies (17), immunohistochemistry, and in situ hybridization (18)(19)(20).The means by which EBV gains access to epithelial cells in general and the questions of which epithelial cell layer in OHL could be infected by EBV and of how the virus reaches the replicating cells in OHL have been the subject of numerous studies (15,(21)(22)(23)(24). Different modes of infection have been suggested: Receptor molecules similar but not identical to CD21 have been reported from undifferentiated epithelial cells of the oropharynx (22,25). It is possible that basal epithelial cells in OHL could be infected through such receptors or via cell fusion (21) by EBV-infected B lymphocytes from peripheral blood. Production of EBV in OHL could be initiated later as a consequence of cellular differentiation (26). It has been shown that human epithelial cells of normal nasopha...

Thjazolo-iso-indolinone derivatives with high specificity toward the reverse transcriptase (RT) of human immunodeficiency virus type 1 (HIV-1) were identified. The most potent compound, BM +51.0836, inhibited HIV-1 RT at a 50%o inhibitory concentration of 90 nM in vitro. In cell culture assays, similar 50%o inhibitory concentrations were obtained with high specificity for HIV-1. These substances were equally active against a zidovudine-resistant isolate. No antiviral effect was observed with an HIV-2 isolate. HIV-1 isolates resistant to the thiazolo-iso-indolinones were generated in cell culture, and the nucleotide sequences of the respective RT genes were analyzed subsequently. Comparison Fig. lb], a substance with 10-fold lower 50% inhibitory concentrations (IC50s) than the lead compound BM 21.1298 and higher antiviral activity than 0-TIBO and nevirapine (BI-RG-587). MATERIALS AND METHODSCompounds. The thiazolo-iso-indolinones were synthesized by Boehringer Mannheim GmbH. A detailed description of the synthesis will be published elsewhere. Zidovudine (AZT; Wellcome), nevirapine (BI-RG-587; Boehringer Ingelheim), and 0-TIBO (Janssen) were also synthesized by the chemical department of Boehringer Mannheim GmbH. The inhibitors were dissolved in dimethyl sulfoxide, and the mixture was added to the RT assay mixture (the dimethyl sulfoxide concentration was less than 5%). For cell culture assays, stock solutions were prepared by dissolving the inhibitors in 50% dimethyl sulfoxide-50% culture medium. In the cell culture supernatant, the dimethyl sulfoxide concentration was less than 1%.Nonradioactive Bl-1 RT assay. Selective inhibitors of HIV-1 RT were identified by using a previously published high-efficiency screening system (4). The assay contains purified HIV-1 RT (11) expressed in Escherichia coli, an in vitro transcript of the HIV-1 long terminal repeat (21), and the primer-binding site as the template and as well as an 18-mer oligonucleotide as the primer. Inhibition of RT was determined by a nonradioactive RT assay (ELISA) with biotin-and digoxigenin-labeled nucleoside triphosphates (4). In brief, 1 ,ug