The anthracycline antibiotics, daunorubicin, doxorubicin, and epirubicin, which are widely used for treatment of malignancies, have been evaluated for their effect on angiogenesis in relation to the inhibition of collagenase type IV reported previously. In the chick chorioallantoic membrane (CAM) system of angiogenesis, anthracyclines inhibited vascular density at doses of 5-20 micrograms/disc as well as collagenous protein biosynthesis, which is a reliable index of angiogenesis. Similarly, all three anthracyclines inhibited tube formation in the in vitro system of angiogenesis using human umbilical vein endothelial cells (HUVECs) plated on Matrigel. The inhibition was dose-dependent and caused 50% inhibition at concentrations of 2.5-15 micrograms/mL. At concentrations of anthracyclines which prevented tube formation and angiogenesis, there were no cytotoxic effects, as evidenced by methylene blue uptake, and the growth of these endothelial cells was not inhibited. The experimental antitumor agent titanocene dichloride inhibited collagenase type IV from Walker 256 carcinosarcoma with IC50 approximately 0.2 mM. Titanocene also prevented angiogenesis in the CAM and tube formation by HUVECs on Matrigel at concentrations that were without effect on growth or cytotoxicity of endothelial cells or Walker 256 cells in culture. The antiangiogenic effect of the aforementioned antitumor agents at therapeutically attainable concentrations may explain, at least in part, their antitumor properties because angiogenesis is an essential process for tumor growth and metastasis. The antiangiogenic effect is, however, unrelated to metalloproteinase inhibition because higher concentrations are required for that effect than for inhibition of angiogenesis.

We report a dynamic light scattering study on protein suspensions of bovine lens homogenates at conditions (pH and ionic strength) similar to the physiological ones. Light scattering data were collected at two temperatures, 20 and 37 degrees C, over a wide range of concentrations from the very dilute limit up to the dense regime approaching the physiological lens concentration. A comparison with experimental data from intact bovine lenses was advanced, revealing differences between dispersions and lenses at similar concentrations. In the dilute regime, two scattering entities were detected and identified with the long-time self-diffusion modes of alpha-crystallins and their aggregates, which naturally exist in lens nucleus. Upon increasing protein concentration, significant changes in time correlation function were observed starting at approximately 75 mg ml(-1), where a new mode originating from collective diffusive motions becomes visible. Self-diffusion coefficients are temperature insensitive, whereas the collective diffusion coefficient depends strongly on temperature revealing a reduction of the net repulsive interparticle forces with decreasing temperature. While there are no rigorous theoretical approaches on particle diffusion properties for multicomponent, nonideal hard sphere polydispersed systems, as the suspensions studied here, a discussion of the volume fraction dependence of the long-time self-diffusion coefficient in the context of existing theoretical approaches was undertaken. This study is purported to provide some insight into the complex light scattering pattern of intact lenses and the interactions between the constituent proteins that are responsible for lens transparency. This would lead to understand basic mechanisms of specific protein interactions that lead to lens opacification (cataract) under pathological conditions.

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Two populations of proteochondroitins were isolated from 4 M guanidine hydrochloride extracts of squid skin by a combination of ion exchange, gel chromatography and density gradient centrifugation. The proteoglycans, M , 4.8 x lo5 and 2.8 x lo5, contained four and two chondroitin chains respectively and unusual oligosaccharides with uronic acid and sulphate groups, and had different amino acid and neutral sugar composition. The chondroitin chains isolated after alkaline borohydride treatment contained varying amounts of glucose, galactose, mannose, fucose and xylose, most likely as branches. Both proteoglycans were antigenic to the rabbit and showed considerable cross-reactivity as assessed by competition experiments using the ELISA technique. The proteoglycans reacted neither with exogenous hyaluronic acid nor with each other to form aggregates.The skin of squid contains two types of proteoglycans. One, which contains only chondroitin and is extracted with water and high-speed homogenization [l], and the other which contains only over-sulphated chondroitin sulphate and is completely extracted with 4 M guanidine hydrochloride (GdnHCl) together with about 85% of the tissue chondroitin proteoglycans [2].The two types of proteoglycans have been separated from 4 M GdnHCl extract by ion-exchange chromatography, and those containing chondroitin to two partly overlapping populations by gel chromatography [2]. Subsequent experiments, however, employing SDSjPAGE showed that both chondroitin proteoglycan populations contained a variety of proteins/ glycoproteins which hinder further studies. It was therefore necessary to proceed to further purification of these proteoglycans in order to study in detail their macromolecular structure and properties, and to the elucidate possible relationship between the two populations.In the present study we report the purification of chondroitin proteoglycans, their chemical composition, macromolecular structure and immunological properties. MATERIALS AND METHODS MaterialsThe skin from freshly caught squids (Zllex illecebrosus coidentii) was powdered in solid C 0 2 , freeze-dried and stored at -20 "C. GdnHCl grade I, benzamidine hydrochloride, 6-aminohexanoic acid, phenylmethanesulphonyl fluoride, Nethylmaleimide, chondroitinase AC, and Fluorinert FC-40 were from Sigma. Urea as a 6 M solution was passed through a mixed ion-exchange resin, Zerolit DM-F (BDH), at 4°CCorrespondence to C. P. Tsiganos, Laboratory of Biochemistry, Abbreviation. GdnHC1, guanidine hydrochloride.Enzyme. Chondroitinase AC (EC 4.2.2.5).

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