Diabetes mellitus is a major health concern, affecting more than 5% of the population. Here we describe a potential novel therapeutic agent for this disease, FGF-21, which was discovered to be a potent regulator of glucose uptake in mouse 3T3-L1 and primary human adipocytes. FGF-21-transgenic mice were viable and resistant to diet-induced obesity. Therapeutic administration of FGF-21 reduced plasma glucose and triglycerides to near normal levels in both ob/ob and db/db mice. These effects persisted for at least 24 hours following the cessation of FGF-21 administration. Importantly, FGF-21 did not induce mitogenicity, hypoglycemia, or weight gain at any dose tested in diabetic or healthy animals or when overexpressed in transgenic mice. Thus, we conclude that FGF-21, which we have identified as a novel metabolic factor, exhibits the therapeutic characteristics necessary for an effective treatment of diabetes.

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A major pre-P-amyloid proteinsps (APP,,,) processing activity from Alzheimer's disease brain extracts was identified and found to be indistinguishable from the activity of cathepsin D.APP,, processing activity cleaved APP,,, into a series of fragments that reacted on immunoblots to a monoclonal antibody (C286.8a) against p-amyloid-( 1 -7)-peptide and cleaved N-dansyl-APP-(591-601)-amide at the Glu-Val and Met-Asp bonds. Fragments of 5.5 kDa and 10-12 kDa were formed from the cleavage of APP,,, by cathepsin D at the Glu593-Val594 bond, and had the same Nterminus as a minor form of P-amyloid released by cells. Abbreviations. APP,,,, pre-P-amyloid protein,,, isoform; [N5", L5'6]APP,,,, APP,,, with lysine and methionine at positions 595 and 596 replaced by asparagine and leucine, respectively; dansyl, 5-(dimethylamino)-napthalene-1-sulfonyl; N-dansyl-APP-(591-601)-amide, N-dansyl-ISEVKMDAEFR-amide; C286.8a, IgG,, mAb recognizing P-amyloid residues at positions 1-7; PhMeSO,F, phenylmethylsulfonyl fluoride; E-64, trans-epoxysuccinyl-L-leucylamido(4-guanidino)butane; P-2, post-I5000 g pellet.Enzymes. Cathepsin D (EC 3.4.23.5); type I1 site-specific deoxyribonucleases (EC 3.1.21.4).-~ processing enzyme, the activity of which is influenced by a mutation associated with the Swedish pedigree of familial Alzheimer's disease. MATERIALS AND METHODSHuman cathepsin D from Calbiochem was pure by SDS/ PAGE, 93% accurate by amino acid analysis, and contained only cathepsin D derived N-termini, the majority of which were from equimolar amounts of the mature light (GPI-PEVLKNY-), and heavy (GGVKVERQVF-) chains. Pepstatin A and heparin agarose were from Sigma. Mono-Q columns were from Pharmacia. Rabbit antiserum to human cathepsin D was from Dako Corp. IgG was purified from rabbit sera using Avid AL [23] and coupled to CnBr-activated Sepharose 4B [24]. The methods and instrumentation for peptide synthesis [25], amino acid analysis [26] and mass spectrocopy [26] were as cited previously. The protein content of crude extracts was determined by the Bradford assay [27].

The risk of cardiovascular disease and osteoporosis drastically increases at the onset of menopause. Phytoestrogens have been suggested to inhibit bone loss and protect the cardiovascular system, in part by improving lipid profiles. The purpose of the present study was to examine the effects of flaxseed, a rich source of the phytoestrogens called lignans, on lipid metabolism and biomarkers of bone turnover in postmenopausal women. Postmenopausal women who were not on hormone replacement therapy were assigned to one of two treatment groups in a double-blind randomized study. Women were asked to consume 40 g of either ground flaxseed or wheat-based comparative control regimen daily for 3 months. In addition, all subjects received 1,000 mg calcium and 400 IU vitamin D daily. Flaxseed supplementation lowered (P < 0.05) both serum total cholesterol and non-high-density lipoprotein cholesterol by 6%, whereas the comparative control regimen had no such effect. Flaxseed regimen reduced serum levels of both low-density- and high-density-lipoprotein cholesterol by 4.7% and triglyceride by 12.8%, albeit not statistically significant. Serum apolipoprotein A-1 and apolipoprotein B concentrations were significantly (P < 0.005) reduced by 6 and 7.5%, respectively, by the flaxseed regimen. Markers of bone formation and resorption were not affected by either of the treatments. The findings of this study indicate that flaxseed supplementation improves lipid profiles but has no effect on biomarkers of bone metabolism in postmenopausal women.

Soy isoflavones, independent of the protein source, prevent hypercholesterolemia and the formation of atherosclerotic lesions induced by ovarian hormone deficiency in hamsters. The antiatherogenic mechanisms of isoflavones need further investigation.

Certain precursor proteins (APP751 and APP770) of the amyloid beta-protein (AP) present in Alzheimer's disease contain a Kunitz-type serine protease inhibitor domain (APPI). In this study, the domain is obtained as a functional inhibitor through both recombinant (APPIr) and synthetic (APPIs) methodologies, and the solution structure of APPI is determined by 1H 2D NMR techniques. Complete sequence-specific resonance assignments (except for P13 and G37 NH) for both APPIr and APPIs are achieved using standard procedures. Ambiguities arising from degeneracies in the NMR resonances are resolved by varying sample conditions. Qualitative interpretation of short- and long-range NOEs reveals secondary structural features similar to those extensively documented by NMR for bovine pancreatic trypsin inhibitor (BPTI). A more rigorous interpretation of the NOESY spectra yields NOE-derived interresidue distance restraints which are used in conjunction with dynamic simulated annealing to generate a family of APPI structures. Within this family, the beta-sheet and helical regions are in good agreement with the crystal structure of BPTI, whereas portions of the protease-binding loops deviate from those in BPTI. These deviations are consistent with those recently described in the crystal structure of APPI (Hynes et al., 1990). Also supported in the NMR study is the hydrophobic patch in the protease-binding domain created by side chain-side chain NOE contacts between M17 and F34. In addition, the NMR spectra indicate that the rotation of the W21 ring in APPI is hindered, unlike Y21 in BPTI, showing a greater than 90% preference for one orientation in the hydrophobic groove.

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