Purification of a basic somatomedin, from human plasma Cohn fraction IV-1, with physicochemical and radioimmunoassay similarity to somatomedin-C and insulin-like growth factor

1979 ◽  
Vol 57 (11) ◽  
pp. 1289-1298 ◽  
Author(s):  
R. Marvin Bala ◽  
B. Bhaumick
Keyword(s):  
Growth Factor ◽  
Human Plasma ◽  
Protein Separation ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Extraction Procedure ◽  
Molecular Size ◽  
Protein Band ◽  
Insulin Like Growth Factor ◽  
Igf I

A basic somatomedin (SM) was purified from human plasma Cohn fraction IV-1 using an initial acid–ethanol–acetone extraction procedure followed by alternating molecular size or charge protein separation techniques. The final recovery of SM bioactivity was approximately 2% of that present in the starting Cohn fraction. The purified SM has an approximate molecular weight of 7500, pI 8.6, 4000 SM bioactivity units per milligram (as measured by a hypophysectomized rat bioassay) and a parallel approximately equipotent radioimmunoassay dose–response curve to SM-C and insulin-like growth factor-I (IGF-I). Sodium dodecyl sulfate – polyacrylamide gel electrophoresis of this purified SM revealed a single protein band. The preliminary determination of the amino acid sequence of the N terminus suggested that this SM preparation was over 75% pure and the first five N-terminal amino acids were identical with those of IGF-I.

scholarly journals Insulin-like growth factor-I and -II and their binding proteins in human ejaculates

2003 ◽  
Vol 22 (3) ◽  
pp. 221-227 ◽  
Author(s):  
Anna Nikolic-Judith ◽  
Olgica Nedic ◽  
Nevena Mihailovic ◽  
Ivona Baricevic
Keyword(s):  
Growth Factor ◽  
Binding Proteins ◽  
Polyacrylamide Gel Electrophoresis ◽  
Molecular Size ◽  
Insulin Like Growth Factor ◽  
Igf I ◽  
Ionic Bonds ◽  
Igf Binding ◽  
Highly Correlated ◽  
Igf Binding Protein

The insulin-like growth factor (IGF)/IGF-binding protein (IGFBP) system was investigated in ejaculates from thirty men aged 23-46 years. Mean ejaculate volume was 3.3 mL (range 1-6 mL). Viscosity was increased moderately in six samples and greatly in four more. The number of motile sperm (0-165 million/mL; mean 49; median 39) was highly correlated with total number (P < 0.001). Concentrations of IGF-I and -II determined by displacement radioimmunoassay were positively correlated (P = 0.001). IGF-I ranged from 2.7 to 29.7 nmol/L (mean 9.6; median 6.5) and increased with viscosity. IGF-II ranged from 5.7 to 39.0 nmol/L (mean 16.0; median 13.5). IGFBP-2 and putative IGFBP-4 were detected by autoradiography of ligand blots after polyacrylamide gel electrophoresis. These binding proteins were stable in ejaculates stored at -18 ?C with protease inhibitors and remained associated with higher molecular weight complexes during molecular size chromatography with 1 mol/L acetic acid. They eluted at least partially in the expected fractions with 1 mol/L sodium chloride but further attempts at purification were unsuccessful due to progressive degradation. It is suggested that ionic bonds between IGFBP-2 and other components, such as proteoglycans, provide protection from proteolysis in a similar way as shown for IGFBP-1 and alpha2-macroglobulin.

Receptors for IGF-I, but not for IGF-II, on proximal colon epithelial cell apical membranes

1989 ◽  
Vol 257 (1) ◽  
pp. E27-E34 ◽  
Author(s):  
D. J. Pillion ◽  
J. F. Haskell ◽  
J. A. Atchison ◽  
V. Ganapathy ◽  
F. H. Leibach
Keyword(s):  
Growth Factor ◽  
Epithelial Cell ◽  
Polyacrylamide Gel Electrophoresis ◽  
Proximal Colon ◽  
Cross Linking ◽  
Insulin Like Growth Factor ◽  
Igf I ◽  
Apical Membranes ◽  
Colon Epithelial Cell ◽  
Binding Subunit

Rabbit proximal colon epithelial cell apical membranes, which are known to contain receptors for insulin, were isolated by a Ca2+-precipitation technique. Binding assays with 125I-insulin-like growth factor I (IGF-I) revealed the presence of specific high-affinity binding sites, with 50% inhibition of binding observed at a concentration of 13.7 ng/ml IGF-I. In contrast, 50% inhibition of 125I-IGF-I binding was observed at an insulin concentration of 1.37 micrograms/ml, suggesting that 125I-IGF-I was not binding to insulin receptors present in this tissue. Cross-linking studies revealed an 125I-IGF-I binding subunit of relative molecular weight (Mr) of 130,000 under reducing conditions on docecyl sulfate-polyacrylamide gel electrophoresis that was similar to the IGF-I binding subunit in human placental membranes (Mr 140,000). Binding and cross-linking studies with 125I-insulin-like growth factor II (IGF-II), however, failed to reveal a specific receptor for this peptide in colon epithelial cell membranes. These results establish the coexistence of receptors for IGF-I and insulin, but not IGF-II, on rabbit proximal colon epithelial cell apical membranes and demonstrate that colon epithelial cells are capable of selective synthesis of various peptide hormone receptors.

Evaluation of a radioimmunoassay for somatomedin-C/insulin-like growth factor I (Sm-C/IGF-I) in human plasma

1990 ◽  
Vol 188 (3) ◽  
pp. 253-260 ◽  
Author(s):  
Daniel Giannella-Neto ◽  
Ana Mercedes Cavaleiro ◽  
Rosa Sadoyama ◽  
Bernardo Leo Wajchenberg ◽  
E.Martin Spencer
Keyword(s):  
Growth Factor ◽  
Human Plasma ◽  
Insulin Like Growth Factor ◽  
Somatomedin C ◽  
Factor I ◽  
Igf I ◽  
Growth Factor I

scholarly journals Differential effect of growth hormone and insulin-like growth factor-I, insulin-like growth factor-II, and insulin on Ig production and growth in human plasma cells

Blood ◽  
1994 ◽  
Vol 83 (6) ◽  
pp. 1569-1574 ◽  
Author(s):  
H Kimata ◽  
A Yoshida
Keyword(s):  
Growth Hormone ◽  
Growth Factor ◽  
Human Plasma ◽  
Plasma Cell ◽  
Plasma Cells ◽  
Thymidine Uptake ◽  
Insulin Like Growth Factor ◽  
Cell Responses ◽  
Factor I ◽  
Igf I

The effect of human growth hormone (GH) and insulin-like growth factor- I (IGF-I), IGF-II, and insulin on human plasma cell responses was studied. GH enhanced Ig production and thymidine uptake in the human plasma cell lines, IM-9 and AF-10. IGF-I, but not IGF-II or insulin, also enhanced Ig production and proliferation in them. However, enhancement by GH was not mediated by IGF-I, because enhancement was blocked by anti-GH antibody (Ab), but not by Ab to IGF-I or IGF-I receptor. Conversely, the enhancement by IGF-I was blocked by either Ab to IGF-I or IGF-I receptor, but not by anti-GH Ab. GH and IGF-I also enhanced production of IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, and IgM and thymidine uptake in PCA-1+ plasma cells generated in vitro. Again, enhancement by GH was specifically blocked by anti-GH Ab, whereas enhancement by IGF-I was specifically blocked by either Ab to IGF-I or IGF-I receptor. These results indicate that GH and IGF-I may play important roles in plasma cell responses.

Isolation and partial characterization of six somatomedin-like peptides from human plasma Cohn fraction IV

1986 ◽  
Vol 111 (2) ◽  
pp. 271-284 ◽  
Author(s):  
Werner F. Blum ◽  
Michael B. Ranke ◽  
Jürgen R. Bierich
Keyword(s):  
Growth Factor ◽  
Human Plasma ◽  
Cell Membranes ◽  
Reversed Phase ◽  
Insulin Like Growth Factor ◽  
C Peptide ◽  
Sds Page ◽  
Igf I ◽  
Basic Peptides ◽  
Acidic Peptides

Abstract. Six somatomedin-like peptides were purified from human plasma Cohn fraction IV by a six-step procedure which included ethanol precipitation, reversed-phase extraction, gel filtration, chromatofocusing and reversed-phase high pressure liquid chromatography (HPLC). Purification was monitored with a competitive protein binding assay using a crude preparations of somatomedin carrier protein. The peptides isolated were homogeneous by reversed-phase HPLC and sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). Their apparent isoelectric points determined by chromatofocusing were 9.2 (Sm I), (Sm II), 8.2 (Sm III), 6.7 (Sm IV), 6.3 (Sm V), and 6.15 (Sm VI). SDS-PAGE under reducing conditions revealed that they are composed of a single peptide chain with apparent molecular weights of 6800 for Sm I, II and IV and 6400 for Sm III, V, and VI. They were equally potent in the porcine costal cartilage in vitro bioassay. The basic peptides (Sm I–III) were significantly more active in radioimmunoassays for somatomedin C (SmC) and insulin-like growth factor I C-peptide (IGF-I (30–41)), while only the slightly acidic peptides were active in a radioimmunoassay for insulin-like growth factor II C-peptide (IGF-II (33–40)). When receptor binding was tested with human placental cell membranes and Sm III as tracer, the basic peptides were significantly more potent than Sm IV–VI. With rat liver cell membranes and Sm V as tracer the slightly acidic peptides were more potent. These findings suggest 1) that human plasma may contain other somatomedin-like peptides besides the major components IGF-I/SmC and IGF-II, and 2) that the basic peptides are structurally related to IGF-I/SmC and the slightly acidic peptides are related to IGF-II.

scholarly journals PENAMBAHAN PROTEIN INSULIN LIKE GROWTH FACTOR-I COMPLEX DALAM PENGENCER PEMBEKUAN SEMEN TERHADAP KUALITAS SPERMATOZOA KAMBING PADA WAKTU EKUILIBRASI

2011 ◽  
Vol 5 (2) ◽  
Author(s):  
Suherni Susilowati ◽  
Tatik Hernawati
Keyword(s):  
Growth Factor ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Polyacrylamide Gel ◽  
Insulin Like Growth Factor ◽  
Factor I ◽  
Plasma Séminal ◽  
Igf I ◽  
Native Polyacrylamide Gel Electrophoresis ◽  
Growth Factor I

Penelitian ini bertujuan mengisolasi protein Insulin Like Growth Factor-I (IGF-I) complex untuk meningkatkan kualitas semen beku kambing pada waktu ekuilibrasi setelah penambahan protein IGF-I complex. Penelitian ini terdiri atas 2 tahap yaitu isolasi protein IGF-I complex dari plasma seminal kambing dan aplikasi terhadap prosesing pembekuan semen. Pada tahap pertama dilakukan identifikasi IGF-I complex dengan menggunakan gel native polyacrylamide gel electrophoresis dan isolasi IGF-I complex. Pada tahap kedua dilakukan aplikasi penambahan protein IGF-I complex pada prosesing semen beku. Semen dikoleksi dengan menggunakan vagina buatan dan kemudian disentrifus selama 5 menit dengan kecepatan 1800 rpm. Kemudian semen dibagi menjadi tiga kelompok. Pada kelompok I, II, dan III ditambahkan protein IGF-I complex masing-masing 0, 12, dan 618 ng/ 3X10 sperma. Selanjutnya dilakukan ekuilibrasi selama 1 jam dan dilanjutkan dengan evaluasi motilitas, viabilitas, dan membran sperma. Hasil penelitian menunjukkan perbedaaan motilitas, viabilitas, dan membran sperma yang signifikan (P<0,05) di antara tiga kelompok perlakuan. Dapat disimpulkan bahwa penambahan protein IGF-I complex dapat meningkatkan kualitas semen beku kambing pada fase ekuilibrasi.

Chemistry, structure, and function of insulin-like growth factors and their receptors: a review

1984 ◽  
Vol 62 (11) ◽  
pp. 1237-1245 ◽  
Author(s):  
James F. Perdue
Keyword(s):  
Growth Factor ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Insulin Receptors ◽  
Cross Reactivity ◽  
Blood Levels ◽  
Protein Kinase Activity ◽  
Binding Studies ◽  
Normal Children ◽  
Igf I

Human serum contains two classes of somatomedins (Sin) with similar three-dimensional structures and mass (i.e., ~7500 kdaltons (kd)), but that can be distinguished from each other by their isoelectric point (pI) values. Those with alkaline pI's include the presumably identical molecules of insulin-like growth factor I (IGF-I), SmA, SmC, and basic Sm. In humans, the neutral–acidic class of Sm is represented by IGF-II. The sera of rats and other animals contain similar classes of Sm. In normal children and adults, plasma levels of both IGF-I and -II are under growth hormone control, although only elevated concentrations of the former can be positively correlated with adolescent skeletal growth. Elevated blood levels of IGF-II or an embryonic form of it, however, are correlated with fetal development of rats, sheep, and possibly in humans. As evidenced from competitive radioreceptor binding studies and affinity-labelling techniques, receptors that bind insulin and IGF-I are immunologically, structurally, and functionally related: (a) monoclonal and polyclonal antibodies to insulin receptors recognize determinants on receptors that bind IGF-I; (b) both receptors have 140-, 95-, and possibly 45-kd subunits that are interchain disulfide-bonded to form glycoprotein complexes of relative masses (Mr) 300 000 – 400 000; and (c) occupancy of the 140-kd subunits by hormone or growth factor stimulates the phosphorylation of tyrosine residues on the 95-kd subunit by receptor-associated protein kinases. Receptors that bind IGF-II are very different from insulin or IGF-I receptors: (a) there is no evidence of immunological cross-reactivity to the insulin receptors or of intrinsic protein kinase activity and (b) estimates of the molecular weight of the detergent-solubilized IGF-II receptor from its hydrodynamic properties or from its electrophoretic mobility during quantitative or sodium dodecyl sulfate–polyacrylamide gel electrophoresis established it to be a single-chained glycoprotein of MT 220 000 held together by intrachain disulfide bonds.

Distinct receptors for insulin-like growth factor I in rat renal glomeruli and tubules

1988 ◽  
Vol 255 (4) ◽  
pp. E504-E512 ◽  
Author(s):  
D. J. Pillion ◽  
J. F. Haskell ◽  
E. Meezan
Keyword(s):  
Growth Factor ◽  
Specific Binding ◽  
Sodium Dodecyl ◽  
Insulin Like Growth Factor ◽  
Receptor Subunit ◽  
Iron Oxide Particles ◽  
Factor I ◽  
Renal Glomeruli ◽  
Igf I ◽  
Growth Factor I

Purified preparations of renal glomeruli and tubules were obtained by a procedure involving perfusion of rat kidneys with magnetic iron oxide particles to selectively separate the iron-containing glomeruli from the nonmagnetic tubules. Detergent-soluble extracts of both renal glomerular and tubular membranes showed high-affinity, specific binding of 125I-labeled insulin-like growth factor I (125I-IGF-I), whereas degradation of this peptide hormone was minimal during a 90-min incubation at 22 degrees C in the presence of 2.5 mM EDTA and 5 mM N-ethylmaleimide. The affinity of these receptors for IGF-I appeared identical in the two types of renal tissue, since 50% inhibition of 125I-IGF-I binding to both glomerular and tubular tissue occurred in the presence of approximately 3 x 10(-9) M unlabeled IGF-I. In contrast, insulin was much less effective at blocking 125I-IGF-I binding to either tissue, with 1 x 10(-6) M insulin required to produce 50% inhibition of binding. Relative to 125I-IGF-I binding, 125I-insulin binding to glomerular and tubular tissue was significantly lower per milligram protein. 125I-IGF-I was specifically cross-linked to a glomerular receptor subunit that migrated as two discrete bands with relative molecular weight (Mr) of 140,000-150,000 on sodium dodecyl sulfate polyacrylamide gels in the presence of 40 mM dithiothreitol. In contrast, 125I-IGF-I was cross-linked to a tubular receptor subunit that migrated as two discrete bands but at a slightly different position, with Mr of 120,000-140,000.(ABSTRACT TRUNCATED AT 250 WORDS)

Comparison of receptors for insulin-like growth factor II from various rat tissues

1987 ◽  
Vol 115 (1) ◽  
pp. 35-NP ◽  
Author(s):  
J. E. Taylor ◽  
C. D. Scott ◽  
R. C. Baxter
Keyword(s):  
Growth Factor ◽  
Ph Dependence ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Subcutaneous Fat ◽  
Cross Reactivity ◽  
Rat Tissues ◽  
Insulin Like Growth Factor ◽  
Major Band ◽  
Factor Ii

ABSTRACT Receptors for insulin-like growth factor II (IGF-II) have been compared in solubilized microsomal membranes from rat lung, brain, kidney, heart, epididymal and subcutaneous fat, ovary, testis and adrenals. Highest binding/μg protein was seen with testicular membranes. Receptors from all tissues showed high affinity for human IGF-II (mean association constant = 65 litres/nmol) and a high degree of specificity (mean IGF-I cross-reactivity 0·3%; no cross-reactivity with insulin). Affinity labelling followed by sodium dodecyl sulphate polyacrylamide gel electrophoresis showed binding was only to a type-II IGF receptor, with a major band seen at a molecular weight of about 230 000 in lung, brain, kidney and testis, and 240 000 in heart, fat and adrenal gland. All tissues showed broad or bimodal pH dependence of binding, with optima seen at about pH 6 and pH 9. Mild stimulation of IGF-II binding by low calcium concentrations (1–2 mmol/l) was seen in all tissues, although higher concentrations were inhibitory in the brain. It was concluded that IGF-II receptors from different rat tissues, when studied under uniform conditions, show similar binding affinities but differences in size and regulation which might be missed if receptors are examined in separate studies. J. Endocr. (1987) 115, 35–41

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