A 26-year-old male patient with pachydermoperiostosis is reported. He had severe anemia with myelofibrosis. Treatment with iron, prednisolone, oxymethorone and 1 a (OH)D3were not satisfactory. But steroid pulse therapy with parenteral iron improved his anemia and pancytopenia, but was not sufficient to relieve the bone marrowfibrosis or splenomegaly. The mechanismof anemia which was considered to be multifactorial including gastro-intestinal bleeding associated with peptic ulcer or erosion and bone marrow failure due to myelofibrosis, is discussed.
A potent growth-promoting polypeptide, the prostate-derived growth factor (PrDGF), has been purified to apparent homogeneity from acid extracts of rat prostatic tissue using ion-exchange, reverse-phase, and gel-permeation chromatography. PrDGF migrates as a single protein-staining band in NaDodSO4/PAGE in precise correspondence to extractable PrDGF activity in nonstained NaDodSO4 gels. PrDGF is acidand heat-stable but is sensitive to reduction or protease treatment. PrDGF is an acidic (pI 5.0) protein of =25 kDa in NaDodSO4/polyacrylamide gels and of =4-8 kDa in reduced NaDodSO4/polyacrylamide gels. PrDGF stimulates the linear incorporation of [methyl-3H] (20,21). We now report the purification and partial characterization ofa growth factor from prostate glands of rats that appears to differ from other previously characterized growth factors by chemical and physiological criteria (3-9, 11, 12, 19, 22-28). MATERIALS Frozen rat prostates were from Pel-Freez Biologicals and normal rat kidney (NRK) cells were from the American Type Culture Collection. Human plasma-derived serum (PDS) was prepared from human platelet-poor plasma (28). Bovine pancreatic trypsin and NaIO4 were from Sigma. Pronase was from Calbiochem-Behring. Na125I (2 mCi, 17 Ci/mg; 1 Ci = 37 GBq) and [methyl-3H]thymidine (79.4 Ci/mmol) were from New England Nuclear. Sephadex G-75 (200-400 mesh), DEAE-Sephadex (A-50), and sulfopropyl Sephadex (SPSephadex) (C-25) were from Pharmacia.The preparative TSK-DEAE-5-PW column and molecular mass standards were from Bio-Rad. The TSK G2000 SW column and ampholytes (pH range, 3.5-10) were from LKB, and the C18 HPLC column was from Vydac (Hesperia, CA).All solvents were HPLC grade and all chemicals were reagent grade. METHODS Protein Purification. Nine hundred grams of rat prostate was homogenized at 40C in 3000 ml of 1.0 M acetic acid, centrifuged at 13,500 x g for 30 min, dialyzed against 10 mM ammonium acetate (pH 5.5), and passed through an SPSephadex C-25 column equilibrated with the same buffer. The extracts were adjusted to pH 7.0 with ammonium hydroxide and passed through a DEAE-Sephadex A-50 column in the same buffer, and the active fractions were pooled, precipitated with 70% ammonium sulfate, dialyzed against 1.0 M acetic acid, and lyophilized. The lyophilized sample was dissolved in 50 ml of 1.0 M acetic acid and fractionated with a 5 cm x 90 cm Sephadex G-75 column (fine, 200-400 mesh) equilibrated with 1.0 M acetic acid. The active fractions were pooled and lyophilized, and 6.3 mg of the lyophilized sample ('t33% of total protein) was dissolved in 5 ml of 10 mM ammonium acetate (pH 7.0) and separated with a Bio-Gel TSK-DEAE-5-PW column using a linear ammonium acetate gradient (0.01-0.8 M). Three activity peaks were observed-I, II, and III-in order of elution, dialyzed against 1.0 M acetic acid, and lyophilized. Peak II samples from separate DEAE HPLC columns were dissolved in 0.1% trifluoroacetic acid, loaded onto a C18 column, and eluted with a linear gradient of 0-80% acetonitrile in 0.1% ...
Glycoprotein V (GPV) is a membrane-associated, 82 Kd platelet glycoprotein that is hydrolyzed during thrombin activation to yield 69 Kd fragment. We have developed a rapid and simple method for isolation of the protein from platelet extracts using a combination of gel permeation, anion-exchange, and lectin affinity chromatography. The partial amino acid sequence was determined by analysis of peptides generated by digestion of the S-carboxyamido-methylated protein with Achromobacter protease I or cyanogen bromide. The sequence shows a remarkable periodicity of leucine residues, which is homologous to the consensus sequence of a highly diversified protein super-family with a common repetitive module. Thrombin cleavage site was determined to be located at the C-terminal region of GPV by analysis of the products separated by sizing and reversed-phase high performance liquid chromatography. By lectin blot analysis, the existence of mucin-type carbohydrate chains was indicated, as well as the existence of asparagine-linked carbohydrate chains shown by the amino acid sequence analysis. From these data, we report a structural model of GPV that is analogous to glycoprotein Ib.
SummaryTime course change of the platelet cytoskeletal protein component in the Triton X-100 insoluble fraction after stimulation was analyzed in Hermansky-Pudlak syndrome and thrombasthenia.In Hermansky-Pudlak syndrome (HPS), a 31 kDa protein, myosin, actin, and a 100 kDa protein assembled as in the normal platelets at the shape change and release reaction phases after ADP or collagen stimulation, suggesting that, the deficient dense granule content do not lead to an abnormal platelet cytoskeletal protein assembly. In thrombasthenia (Type I), myosin increased at the shape change and release reaction phases as it does in normal platelets, but actin and the 100 kDa protein increased only at the initial activation phase, and then subsequently decreased to the level of the resting phase. The actin-binding protein (ABP) and the 31 kDa protein increased a little following stimulation. Similar cytoskeletal protein change after stimulation were found in normal platelets which were prevented from the aggregation process by chelating the external Ca2+ or by using synthetic decapeptide of fibrinogen γ-chain of carboxyl terminus. The decreased platelet cytoskeletal protein assembly in thrombasthenia or in platelets stimulated without aggregation, was derived from a loss of the platelet aggregation process due to the defect of GP Ilb-IIIa complex or an interaction failure between GP IIb-IIIa complex and fibrinogen. The interaction between platelets and either fibrinogen or fibrin can induce a more stable platelet cytoskeletal protein assembly, however, agonistic stimulation without these interactions cannot do it directly.
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