Reversal of basic fibroblast growth factor-mediated autocrine cell transformation by aromatic anionic compounds.

NIH-3T3 cells transfected with basic fibroblast growth factor (bFGF) fused to a signal peptide sequence (spbFGF cells) are transformed in vitro and tumorigenic in vivo. Treatment of spbFGF cells with low and nontoxic concentrations (0.5-2.5 micrograms/ml) of negatively charged, nonsulfated aromatic compounds (e.g., aurin tricarboxylic acid, 4-hydroxyphenoxyacetic acid) resulted in restoration of their normal proliferative rate, morphological appearance, and adhesion properties. Binding and cross-linking experiments using 125I-labeled bFGF revealed that these alterations were associated with an up-regulation of high affinity receptors bFGF receptors was induced by these compounds in spbFGF cells that were seeded on fibronectin to enforce a firm cell attachment and flattening. Thus, induction of spbFGF cell adhesion and spreading may not be related to restoration of normal bFGF-receptor interactions. Although the negatively charged aromatic compounds mimic many of the effects of heparin in other systems (e.g., release of heparin- and heparan sulfate-bound proteins, inhibition of heparanase), heparin, heparan sulfate, and dextran sulfate were not effective at the low concentrations of the anionic compounds used in the present study. Likewise, suramin, a sulfated aromatic molecule, was effective at toxic concentrations, 400-600-fold higher than the nonsulfated aromatic compounds. The development of defined, nontoxic anionic compounds may provide a new strategy to interfere with the autonomous and anchorage independent mode of cell growth involved in autocrine cell transformation and cancer.     

The relationship between lymphocyte transformation and immune responses. II. Correlations between transformation and humoral and cellular immune responses

Rabbits were immunized against purified proteins, tissue extracts or sheep erythrocytes, with or without Freund's adjuvant. Skin reactions of the delayed type were found only in animals given antigen with adjuvant, although all rabbits developed serum antibodies and most of them Arthus type reactions. Lymphocytes of all animals, however, showed similar transformation activity when cultured with the immunizing antigen.

Thus the blast transformation phenomenon correlates well with both cellular and humoral immune responses and not with the delayed type hypersensitivity only.

     

Chronic uveitis in Gaucher's disease

Thirty patients with Gaucher's disease have been examined in the authors' clinic; 29 of them suffered from the adult form and one had the juvenile type of the disease. The authors found intraocular abnormalities in four patients: three suffered from uveitis and in one, multiple vitreous opacities were observed. Two of the three patients had bilateral uveitis, while in one the uveitis was confined to the right eye only. In two of the three patients the manifestations of Gaucher's disease were present before the onset of uveitis. In one of the three, however, bilateral uveitis with spontaneous hyphema in the left eye was the presenting symptom. The diagnosis of Gaucher's disease in this case was arrived at during the work-up investigations for the etiology of the ocular manifestations. Initially the ocular symptoms in these three patients manifested as acute anterior uveitis, becoming chronic and involving also the posterior segments at a later stage.     

The role of cytokines in angiogenesis

Using the corneal model of neovascularization developed in their laboratory, the authors investigated the angiogenic potential of interleukin-1 (IL-1α and β), interleukin-6 (IL-6), interleukin-8 (IL-8), tumor necrosis factor (TNFα), transforming growth factor β (TGFβ), granulocyte macrophage-colony stimulating factor (GM-CSF), and interferon (IFN(γ) and IFN(γ)). Various concentrations of the tested cytokine were sequestered into Elvax-40 and implanted at 2.5 mm from the limbus within the corneal stroma of the rabbit eye. Three distinct groups of cytokines could be observed according to their angiogenic potential in this system. IL-1α was by far the most potent stimulator of neovascularization, inducing this process at concentrations as low as one nanogram per implant. TGFβ, TNFα and GM-CSF induced significant neovascularization only at concentrations of 500 nanograms per implant. IL-2, IL-6, IL-8, IFNα and IFN(γ) did not induce any significant neovascularization at the concentration tested (0.5 μg to 10 μg per implant).     

The role of cyclosporin eye drops in ocular inflammatory diseases

Fifty-nine patients were treated with cyclosporin A (CsA) eye drops for various diseases. Most patients with severe vernal keratoconjunctivitis responded well to the treatment. Treatment had little or no effect on anterior uveitis (acute and chronic) or on the fate of high-risk corneal grafts. The response of patients suffering from peripheral corneal melting, ocular pemphigoid or Stevens-Johnson syndrome was variable. Among these, some patients responded well to the treatment with CsA drops while others had no detectable clinical benefit. The large differences in the observed treatment responses may be interpreted to indicate that a different pathological mechanism may take place within a group of patients with similar diagnoses and clinical manifestations.     

Influence of interferons corneal angiogenesis induced by basic fibroblast growth factor and lipopolysaccharide

A controlled and reproducible angiogenic stimulus was induced in the rabbit cornea by Elvax-40 implants sequestering either 500 ng of lipopolysaccharide (LPS) or 500 ng of basic fibroblast growth factor (bFGF). The effect of IFNα and IFN(β) on angiogenesis was studied by inserting implants sequestering 500 ng of these cytokines (approximately 10(4) Units/implant) adjacent to the LPS or bFGF implants. Interferon-α or γ did not inhibit the bFGF-induced angiogenesis, and in most of these experiments an enhancing effect was observed. This enhancement was not statistically significant. The LPS-induced angiogenesis, however, was slightly inhibited in the presence of either interferon-α or γ but these differences were not statistically significant with p=0.1 only. Both cytokines were non angiogenic and there was no detectable difference between them regarding their effect on the angiogenic process induced by either bFGF or LPS.     

Low-dose Cyclosporin a in uveitis a long-term follow-up

Seventy patients suffering from bilateral endogenous chronic uveitis or ocular Behcet's disease have been treated with 5 mg/kg/day Cyclosporin A. All patients were followed at regular intervals for up to eight years. In this group of patients, we observed that CsA is an effective drug for the treatment of intraocular inflammation. Patients have needed continuous treatment with CsA for an average period of 31 months. Exacerbations of the inflammatory processes have been observed in 95% of the patients on initial attempts to lower the CsA dosage. These exacerbations were controlled either by local treatment or a combination of low-dose systemic corticosteroids. Elevation of the serum creatinine was initially observed in all patients when under the higher doses of CsA. However, only 15 patients (21.4%) had levels which were higher than the upper normal limit. Elevation of the bilirubin level of 50% or more above baseline was observed in 45 patients (64.3%) but only 11 patients (15.7%) showed levels which were above the upper normal limit. During this period of follow-up, cure was achieved in 25 patients and no exacerbations were observed up to five years after discontinuation of treatment.     

MAD2 associates with the cyclosome/anaphase-promoting complex and inhibits its activity

Cell cycle progression is monitored by checkpoint mechanisms that ensure faithful duplication and accurate segregation of the genome. Defects in spindle assembly or spindle-kinetochore attachment activate the mitotic checkpoint. Once activated, this checkpoint arrests cells prior to the metaphase-anaphase transition with unsegregated chromosomes, stable cyclin B, and elevated M phase promoting factor activity. However, the mechanisms underlying this process remain obscure. Here we report that upon activation of the mitotic checkpoint, MAD2, an essential component of the mitotic checkpoint, associates with the cyclin B-ubiquitin ligase, known as the cyclosome or anaphase-promoting complex. Moreover, purified MAD2 causes a metaphase arrest in cycling Xenopus laevis egg extracts and prevents cyclin B proteolysis by blocking its ubiquitination, indicating that MAD2 functions as an inhibitor of the cyclosome. Thus, MAD2 links the mitotic checkpoint pathway to the cyclin B destruction machinery which is critical in controlling the metaphase-anaphase transition.