Subtypes of serum group specific component (Gc) in normal conditions and in pathology

In the framework of the ecogenetic research programme, the data are presented on the genetic polymorphism of the vitamin D-binding protein (Gc) in various USSR populations. Blood serum samples were studied, taken from the Russians of the town Yegorievsk, Moscow Region (p = 321) and 113 Russian patients with tuberculosis using the method of isoelectrofocusing. The information was obtained of the Gc frequencies in two population units of Buryats of Aginsky and Ost-Ordynsky Autonomous Districts of Chita and Irkutsk Regions, including the Olkhon island (on the lake Baikal), in totality, 593 individuals and 13 local groups. The position of the studied Russian and Buryat groups within the gene frequency co-ordinate space is well in line with the estimated area of their localization, with regard to the world distribution. Among the Buryat populations studied, there is distinct heterogeneity for which the factor Gc1F plays a leading role within the Gc system/responsible for 92% of all possible genetic variability. Gc factor frequencies in Buryats range within the following limits: 1F.-0.3864-0.6023, 1S-0.1895-0.4535, 2-0.1364-0.2581. For the Russians of Yegorievsk and the patients with tuberculosis of Moscow and Moscow Region following allele frequencies are established: 1-F0.1169, 1S-0.5476, 2-0.1364 and 1F-0.1106, 1S-0.5531, 2-0.3363, respectively, which indicates that no association exists between Gc variants and tuberculosis. The correlation of the Gc allele frequency distribution with the ratio of insulin-independent diabetes (type 2) world-wide indicates that expression of high frequency of diseases is accompanied with comparatively rare characteristic combination of frequencies of three Gc alleles.     

Gc (vitamin D binding protein) subtypes in rheumatoid arthritis

Summary Two hundred and six patients with rheumatoid arthritis were examined for their association with the subtypes of Gc (vitamin D binding protein). In patient groups there is 11% excess of individuals with Gc^*2 allele compared to the control group, giving a relative risk of 1.55. This risk increases with the humoral status of the individual. A possible physiological basis between the association of vitamin D binding protein and rheumatoid arthritis is discussed.     

Direct activation in vitro of mouse peritoneal macrophages by pyran copolymer (NSC 46015)

Normal resting macrophages were transformed to cytostatic effector cells in the presence of pyran copolymer (NSC 46015) in the culture medium. Macrophage “activation” to inhibit MBL-2 leukemia cell growth was sharply dose-dependent and required >24 hr after exposure to pyran. The observed growth inhibition appeared to result from a modification of the macrophages themselves, since neither allogeneic macrophages nor drug alone interfered with MBL-2 cell growth. The primary mechanism of cytostasis did not involve phagocytosis or soluble macrophage product(s). Similar activation was observed for poly(I)·poly(C) and to a lesser extent for dextran sulfate. It is suggested that the antitumor activity of these polyanions is due to their function as direct macrophage stimulants.     

Homology between the human vitamin D-binding protein (group specific component), alpha-fetoprotein and serum albumin

41 Amino acid long N-terminal sequences of the three major human vitamin D-binding proteins (group-specific components Gc1F, Gc1S and Gc2) were characterized: they were identical. By computer analyses, the alignment of this N-terminal sequence with several sequences of human serum pre-proalbumin and human pre-alpha-fetoprotein was established.     

Distribution of the group specific (Gc) serum component in the populations of the Markham Valley, New Guinea

The distribution of the Gc phenotypes was determined by immunoelectrophoresis amongst 486 inhabitants of nine villages of the Markham River Valley of New Guinea. The overall gene frequencies were Gc¹, 0.538; Gc², 0.351; Gc^Aborigine, 0.112. Gc^Aborigine occurred in all the villages, its frequency ranging from 0.041 to 0.187. The Gc² gene frequency also varied widely ranging from 0.167 to 0.491. No correlation could be found between altitude and the Gc distribution and there was an overlap in the gene frequencies between the Austronesian and non-Austronesian-speaking villages.     

Impaired macrophage activation in vitamin D3 deficiency: differential in vitro effects of 1,25-dihydroxyvitamin D3 on mouse peritoneal macrophage functions

1,25-Dihydroxyvitamin D3 (1,25(OH)2D3) is known to interact in vitro with mononuclear phagocytes. The purpose of this study was to determine the role of the steroid in macrophage activation in vivo. Peritoneal macrophages from normal and vitamin D3-deficient mice were obtained after i.p. injection of activating or eliciting agents. Cells obtained from vitamin D3-deficient mice exhibited defected capabilities to perform anti-tumor activities (cytostasis and cytolysis) and to form oxygen reduction products (H2O2 and O2-). On the other hand, the level of the lysosomal enzyme acid phosphatase was unaffected by vitamin D3 deficiency. In vitro, incubation of macrophages with 1,25(OH)2D3 enhanced their anti-tumor activities, but did not affect the cells' capacity to produce H2O2 and O2-, or acid phosphatase. Our results suggest that 1,25(OH)2D3 is essential for macrophage activation in vivo. However, in vitro, the hormone is only partially capable of affecting the macrophage functions, probably because of the maturation state of the cells.     

The isolation, characterization and amino terminal sequence of the vitamin D-binding protein (group specific component) from mouse plasma

1. In order to establish a homologous system in which to study the interaction of mouse vitamin D-binding protein (MVDBP) with mouse T-cell lymphocytes, we purified MVDBP from mouse plasma. 2. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) showed that purified MVDBP had an apparent relative molecular weight of 49,000. 3. Previous work in our laboratory has shown that purified rat vitamin D-binding protein (RVDBP) has an apparent relative molecular weight of 52,000. 4. The amino terminal amino acid sequence of MVDBP is shown below and compared with that of RVDBP. MVDBP: LeuGluArgGlyArgAspTyrGluLysAspLysValCysAsnGluLeuAlaMetLeuGlyLysGlu RVDBP: LeuGluArgGlyArgAspTyrGluLysAspLysValCysGlnGluLeuSerThrLeuGlyLysAsp AspPhe AspPhe While 21 out of 24 residues (87.5%) of the amino terminus of MVDBP are the same as those in RVDBP, residues 14, 17, 18 and 22 (underlined) are different. 5. The sedimentation coefficient of the protein, determined by sucrose density gradient ultracentrifugation, is 3.8 for MVDBP and 4.1 for the rat VDBP. 6. The MVDBP purified in this study exhibits only one isoform on isoelectric focusing; the isoelectric point was 4.87 as determined on pH 4.0-6.5 isoelectric focusing gels (IEF). 7. The binding of vitamin D3, 25-hydroxyvitamin D3 and three other analogs was investigated with a charcoal dextran assay.(ABSTRACT TRUNCATED AT 250 WORDS)     

Role of the maleyl-albumin receptor in activation of murine peritoneal macrophages in vitro

It has been previously demonstrated that maley-lated-BSA (maleyl-albumin) induces functional activation in murine peritoneal macrophages. Furthermore, maleyl-albumin has been shown to interact with two distinct sites on human monocytes; one site is the scavenger receptor, a 260-kDa oligomeric protein which recognizes modified forms of low density lipoprotein (LDL), and the second is a lower affinity site which has yet to be structurally characterized. In the present study, we wished to quantitatively assess the number and character of maleyl-albumin-binding sites on murine peritoneal macrophages and to determine which site or sites are involved in signaling the macrophage to undergo extensive functional development. Binding studies. demonstrate at least two distinct receptors for maleyl-albumin on murine peritoneal macrophages. Scatchard analyses of the binding isotherms reveal two sites characterized by dissociation constants (Kd) of 17.6 nM and 4.9 microM and maximal binding of 1.2 x 10(5) and 1 x 10(6) sites/cell, respectively. The contribution of the scavenger receptor, determined by binding analyses of malondialdehyde-LDL, is described by two sites with Kd of 39.4 pM and 9.6 nM, and maximal binding of 2.7 x 10(3) and 1.9 x 10(4) sites/cell, respectively. Maleyl-albumin blocks binding of malondialdehyde-LDL, whereas modified LDL fails to inhibit binding of maleyl-albumin. Maleyl-albumin, at concentrations producing lower affinity binding, stimulates tumor cytolysis, expression of mRNA encoding TNF, and suppression of INF-gamma-induced expression of Ia Ag. Malondialdehyde-LDL fails to elicit these responses. We conclude that macrophage activation produced by maleyl-albumin is mediated by interaction with the low affinity, high capacity binding site for maleyl-albumin rather than the scavenger receptor.     

Identification of novel adhesion proteins in mouse peritoneal macrophages.

When mouse peritoneal macrophages were made to adhere firmly on glass surface and then removed by sequential treatment with hypotonic triethanolamine and Nonidet P-40, a set of proteins were found to be left behind at the sites of adherent cells. Such glass-adherent proteins were detected as round or ellipsoidal patches of autofluorescence under a confocal laser microscope, and visualized ultrastructurally as aggregates of narrow threads of unique loop structures which were composed of linearly aligned particles of 22 +/- 2 nm in diameter. Lithium dodecylsulfate-polyacrylamide gel electrophoresis of the glass-adherent proteins showed two major bands, 12 kDa and 14 kDa, which always co-existed in any different sample. The polyclonal antibody raised against these two proteins specifically stained the glass-adherent proteins in situ. The adhesion of macrophages to glass was significantly blocked with Fab fragments of the antibody. The in situ cross-linking experiment suggested that these two proteins might be closely associated with each other to form complexes. Hence, these proteins can be reasonably considered to be responsible for non-specific adhesion of macrophages to glass.     

The vitamin D binding of the common and rare variants of the group-specific component (Gc)

Summary The vitamin D₃ binding properties of the common and rare Gc variants were examined. Vitamin D₃ labeled with ¹⁴C was added to serum. Gc phenotypes were demonstrated autoradiographically following separation by immunofixation electrophoresis on agarose. This qualitative analysis did not reveal differences in vitamin D₃ binding by the group-specific components of the common types Gc1-1, Gc 2-1, and Gc2-2. The double-band variants Gc Darmstadt, Gc Y/Ab, Gc Toulouse, Gc Norway, and Gc Caucasian were examined; the phenotypes Gc Ab-Ab, Gc Ab-1, Gc Ab-2, Gc T-1, Gc T-2, Gc Norw-2, and Gc 1-Cau showed normal D₃ binding. The double bands of Gc Darmstadt in the phenotype D-2 appeared somewhat weak. The singleband mutants Gc Wien, Gc Chippewa, Gc Opava, and Gc Z were analyzed; the phenotypes Gc W-1, Gc W-2, Gc Chip-1, Gc Chip-2, Gc 1-Op, Gc Op-2, Gc 1-Z, and Gc 2-Z showed normal D₃ binding. A mutant in the Gc system with clearly defective vitamin D₃ binding properties remains to be delineated.     

Population genetics of the group specific component (Gc) and phosphoglucomutase (PGM1) studied by isoelectric focusing

For the determination of the group-specific component (Gc) and phosphoglucomutase (PGM1) phenotypes, isoelectric focusing was performed on two samples, one of Jat Sikh of northwest India, the other of northeast English. The subtype frequencies of these two systems do not differentiate the two populations sampled. Synthesis of the existing data shows distinct PGM1 and Gc subtype frequencies in various ethnic and racial groups. The anthropological implication of these subtype frequencies is discussed.     

Role of a serum factor in enhancement of in vitro interactions between Plasmodium berghei sporozoites and hamster peritoneal macrophages

Interactions between Plasmodium berghei sporozoites and hamster peritoneal macrophages were studied. Hamster serum was shown to enhance the percentage of sporozoites tha attached to macrophages, thus confirming previous studies by other workers using mouse macrophages and mouse serum. The enhancement factor within hamster serum was concentrated by a fractionation procedure consisting of ammonium sulfate precipitation followed by Concanavalin A-Sepharose affinity chromatography. This serum fraction has been shown previously to contain component(s) that bind to P. berghei sporozoites.     

Polymorphic post-albumin of cattle and horse plasma identified as vitamin D binding protein (Gc protein)

Cattle and horse plasma samples of known post-albumin types were radiolabelled with ¹⁴C-vitamin D₃. These samples were then analysed by polyacrylamide gel electrophoresis, followed by autoradiography. The patterns observed were identical to those of post-albumin variants. The polymorphic post-albumin protein of cattle and horse was thus identified as the vitamin D binding protein and homologous to the Gc protein of human plasma.     

The distribution of the group specific component (Gc) in four endogamous groups of Punjab, North India

The paper reports the distribution of group specific component (Gc) types among four endogamous groups of Punjab: Jat Sikh, Khatri, Ramgarhia, and Ramdasia. A total of 418 individuals were tested for this polymorphism. The frequency of Gc1 alleles ranges between 0.7056 and 0.7636. These frequencies are compared with those obtained in other Indian populations.     

Evolutionary conservation of the linkage between the structural loci for serum albumin and vitamin D binding protein (Gc) in cattle

Evidence is presented for close genetic linkage between the structural loci for serum albumin and the vitamin D binding protein (Gc) in Belgian Blue and White cattle. Five recombinants were observed in a total of 342 informative offspring. The recombination frequency between the two loci was estimated as 1.5% +/- 0.9. The observed distribution of the haplotypes deviated from the expected one in the population, probably due to selection and significant linkage disequilibrium.     

Genetic polymorphism of the vitamin D binding protein (Gc) in the musk shrew (Suncus murinus)

In the musk shrew ( Suncus murinus ), the electrophoretic bands in the post-albumin region were identified as vitamin D binding protein (Gc) by the [³HI vitamin D₃ binding method. Three Gc phenotypes were distinguished from each other: a single faster band (Gc-A), a single slower band (Gc-B) and the double bands (Gc-AB). Results of mating experiments indicated that the Gc-A and Gc-B are controlled by two codominant alleles, Cc ^a and Gc ^b at an autosomal locus ( Cc ), respectively. It was noticed that, in the Gc-AB phenotypes, the Gc-B band was constantly more intense than the Gc-A band in the protein staining. The same tendency was also observed btween the homozygous Gc-A and Gc-B bands, and further, radioactivity of the Gc-B bound with [³H] vitamin D₃ was about twofold higher than that of the Gc-A. These results suggest that the Gc^b yields its protein product twofold more than the Gc ^a. No cross-reaction between the shrew proteins and a rabbit anti-human Gc protein was observed.     

Evolutionary conservation of the linkage between the structural loci for serum albumin and vitamin D binding protein (Gc) in cattle

Summary. Evidence is presented for close genetic linkage between the structural loci for serum albumin and the vitamin D binding protein (Gc) in Belgian Blue and White cattle. Five recombinants were observed in a total of 342 informative offspring. The recombination frequency between the two loci was estimated as 1.5%± 0.9. The observed distribution of the haplotypes deviated from the expected one in the population, probably due to selection and significant linkage disequilibrium.