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G Ruiu

Publications and source records attributed to G Ruiu.

At least 19 recordsLinked to original sources
PubMed1999

Study of the glycosylation of apolipoprotein H.

Apolipoprotein H is a single chain polypeptide composed of 326 amino acids highly glycosylated. Its carbohydrate content is approximately 19% of the molecular weight. We show that it is rich in sialic acid linked alpha (2-6) to galactose or N-acetylgalactosamine. Sialic acid is not alpha (2-3) linked to galactose. Galactose is beta (1-4) linked to N-acetylglucosamine and beta (1-3) linked to N-acetylgalactosamine. Carbohydrate O-linked chains (mainly sialic acid) are alpha (2-6) linked to galactose or N-acetylgalactosamine. Galactose is also organised in O-linked chains and beta (1-4) linked to N-acetylglucosamine and beta (1-3) linked to acetylgalactosamine. Concanavalin A lectin was used to isolate two groups of apolipoprotein H molecules bearing biantennary and truncated hybrids and high mannose and hybrid oligosaccharides. Apolipoprotein H fails to bind lysine-Sepharose. Our results thus show that it presents truncated hybrid or hybrid-type carbohydrate chains which bear few unmasked mannose residues as a terminal sugar. Biochemical analysis of carbohydrate structures conducted on single isoforms separated through IEF revealed that no specific carbohydrate complex is bound to a single isoform.

Chromatography, Affinity↗
PubMed1999

The binding of apolipoprotein H (beta2-Glycoprotein I) to lipoproteins.

Beta2-glycoprotein I has a high affinity for triglyceride-rich particles, activates lipoprotein lipase, and is also defined as an apolipoprotein H. Previous studies have shown that apolipoprotein H is a regular structural component of the major classes of lipoproteins. In view of these findings, we analyzed the interactions of apolipoprotein H with lipoproteins in the fasting plasma of eight normal, seven hypertriglyceridemic, and seven hypercholesterolemic subjects. After rate-zonal, density gradient ultracentrifugation, apolipoprotein H was little distributed among the different density fractions, and most of it was recovered in the last fraction that contained the lipoprotein-free plasma. A small percentage (4-13%) of the apolipoprotein H associated with plasma lipoproteins was detected at the density ranging from 1.090 to 1.225 g/ml. This result means that apolipoprotein H is little associated with lipoproteins.

Case-Control Studies↗
PubMed1999

Apolipoprotein H is not affected by in vitro glycosylation.

Increased nonenzymatic glycosylation of all major classes of apolipoproteins has been demonstrated in diabetes. In this work we deal with the in vitro nonenzymatic glycosylation of apolipoprotein H, whose role in lipid metabolism is still poorly understood and whose levels increase in diabetes. Apolipoprotein H was isolated from human plasma and purified through a combination of affinity chromatography and continuous elution electrophoresis. The in vitro glycosylation was performed by incubating purified apolipoprotein H with high concentration of glucose. Our results indicate that the in vitro nonenzymatic glycosylation has no effect on the physical properties of apolipoprotein H, despite the fact that this apolipoprotein contains a high number of lysine residues. Since the in vitro concentration of glucose was far higher than the levels normally found in diabetic subjects, it is unlikely for apolipoprotein H to become glycosylated in diabetes.

Blotting, Western↗
PubMed1998

Pharmacokinetic effects of conversion to a new formulation of cyclosporin A in rheumatoid arthritis patients.

In this study we aimed at evaluating the modifications in the pharmacokinetic profile of cyclosporin A (CyA) after conversion from standard formulation (CyA-ST) to a new formulation (CyA-NF, Sandimmun Neoral) in patients with rheumatoid arthritis (RA). It was an open, crossover study that involved 15 RA patients who were on stabilized treatment with CyA-ST. The patient continued receiving CyA-ST (mean dose of 3.0 +/- 0.7 mg/kg per day) for 3 weeks and then converted 1:1 to CyA-NF for a further 3 weeks. CyA pharmacokinetics were established on day 1 (CyA-ST evaluation) and +21 (CyA-NF evaluation). The results showed that the bioavailability of CyA-NF was greater than that of CyA-ST (AUC tau, bss: 3335 +/- 1300 vs 2667 +/- 1155 ng.h/ml, P = 0.0073; AUC tau, bss ratio 1.26 +/- 0.40 vs 1.0 as reference, P < 0.05), with higher and earlier peak blood concentrations (Cmax: 677 +/- 256 vs 475 +/- 213 ng/ml, P = 0.0329; tmax: 1.5 +/- 0.7 vs 2.6 +/- 1.6 h, P = 0.0720). The pharmacokinetic profile of CyA-NF showed greater between-patient reproducibility (lower CV% for all of the considered parameters). In conclusion, when using CyA-NF instead of CyA-ST, greater and more constant exposure to CyA should be expected.

Adolescent↗
PubMed1997

Apolipoprotein H levels in diabetic subjects: correlation with cholesterol levels.

To assess the relationship between apolipoprotein H (apo H) plasma levels and lipid metabolism in diabetes mellitus, we have examined the correlation between apo H plasma concentration and the main plasma lipid levels in 127 non-insulin-dependent (NIDDM) and 118 insulin-dependent (IDDM) diabetes mellitus patients. The data are compared with those in 286 nondiabetics. Our data show a significant increase in plasma apo H in diabetic as opposed to nondiabetic subjects (NIDDM, 29.9 +/- 10.8 mg/dL; IDDM, 31.3 +/- 9.9; controls, 22.5 +/- 7.7; F = 53.3, P = .0001). The relation between plasma lipids and apo H was simultaneously evaluated in the three groups with inclusion of diabetes, sex, body mass index (BMI), and age as covariates in the model. This analysis showed a strong positive correlation (P = .0009) between apo H and total cholesterol, and a weaker positive correlation with triglycerides ([TGs] P = .016). The correlation between apo H and hemoglobin A1c (HbA1c) levels in diabetics (P = .03) highlights the importance of glycemic control for plasma levels of this apoprotein, which is highly glycated. Although the role of apo H in lipid metabolism is still uncertain, recent investigations on the possible relation between plasma apo H levels and increased plasma lipids and thrombotic risk could explain the increased atherosclerotic risk in diabetic patients.

Adult↗
PubMed1997

Characterization of the carbohydrate structures of apolipoprotein H through concanavalin A affinity chromatography.

Apolipoprotein H, also known as beta 2-Glycoprotein I, is a single chain highly glycosylated polypeptide of 326 amino acids. The carbohydrate content of apolipoprotein H is approximately 19% of the molecular weight. Some studies have described the main oligosaccharides forming the glycosylated chains but the carbohydrate inner structures of apolipoprotein H has not been investigated yet. This gap should be filled being glycosylation a very important process which is able to regulate the structure and the biological functions of proteins. Lectins are proteins which specifically bind carbohydrate structures. Affinity chromatography of glycoproteins on immobilized lectins, such as Concanavalin A (Con A), has been proved to be a useful method for oligosaccharide fractionation. N-Linked oligosaccharide structures were shown to interact with Con A according to their branching properties. In the present study, we analyzed the patterns of Con A elution of apolipoprotein H isolated from human plasma. Using Con A affinity chromatography we show that apolipoprotein H has a high degree of heterogeneity in its glycosylated structure. It allowed one to isolate two groups of apolipoprotein H molecules bearing biantennary and truncated hybrids and high mannose and hybrid oligosaccharides. Since Con A affinity chromatography allows fractionation of molecules differing in the extent of carbohydrate branching irrespective of the sialyl residues, we can conclude that mannose residues are masked with other sugars such as galactose-beta (1-4)N-acetylglucosamine, galactose-beta (1-3)N-acetyl-galactosamine and sialic acid linked alpha (2-6) to galactose or to N-acetylgalactosamine, or capped with sulfated residues. Thus, according to our results apolipoprotein H presents truncated hybryd or hybrid-type carbohydrate chains which bear few unmasked mannose residues as terminal sugar. Moreover, isoelectrofocusing of apolipoprotein H forms fractionated on Con A demostrates that weakly bound material presents a predominance of more acidic isoforms than that firmly bound to the lectin, indicating that weakly bound fractions contain molecules which are more negatively charged and that Con A is able to separate glycosylated forms which are not discriminated by isoelectrofocusing.

Apolipoproteins↗
PubMed1997

Characterization and representative structures of N-oligosaccharides bound to apolipoprotein H.

We studied the structure of N-linked carbohydrates bound to apolipoprotein H by a combination of two methods which make use of lectins. Digoxigenin-labelled lectins are used for the structural characterization of carbohydrate chains of glycoproteins. Concanavalin A lectin affinity chromatography was used to analyse apolipoprotein H according to the characteristics of its carbohydrate chain inner to sialic acid residues. Our results from digoxigenin-labelled lectins analysis showed that apolipoprotein H gave positive bands to SNA, DSA, GNA, PNA and AAA lectins. Apolipoprotein H gave a negative band when reacted with MAA lectin. When we applied apolipoprotein H onto the Concanavalin A lectin column no detectable amounts of protein were eluted with Concanavalin A buffer. After adding a buffer with low sugar concentration (10 mM glucoside) a large amount of apolipoprotein H was recovered. These molecules of apolipoprotein H weakly bound to the lectin. When a higher sugar concentration (500 mM mannoside) was added most of the sample applied was eluted. These molecules of apolipoprotein H firmly bound to the column having high affinity for the lectin. These results combined with those coming from the digoxigen-labeled lectins method enable us to understand the inner structure of carbohydrate chains with their outer branches. Molecules of apolipoprotein H which weakly bind to Concanavalin A could bear complex N-glycans organized in biantennary or truncated hybrid structures. Firmly bound apolipoprotein H referred to molecules rich in N-glycan hybrid structures. They have an outer branch belonging to the high mannose carbohydrate chains which explain the ability to bind to the column and an other main branch bearing the sequence galactose beta-(1-4)-N-acetylglucosamine beta-(1-2) mannose. Galactose could be the terminal sugar or, alternatively, be masked with sialic acid alpha-(2-6) terminally linked.

Amino Acid Sequence↗
PubMed1997

Qualitative analysis of the carbohydrate composition of apolipoprotein H.

The specific binding of digoxigenin-labeled lectins to carbohydrate moieties is used to characterize the carbohydrate chains bound to apolipoprotein H. Our results show that apolipoprotein H is rich in sialic acid linked alpha(2-6) to galactose or N-acetylgalactosamine. Sialic acid is not alpha(2-3)-linked to galactose. Galactose is beta(1-4)-linked to N-acetylglucosamine and beta(1-3)-linked to N-acetylgalactosamine. High-mannose N-glycan chains are barely detectable. After N-glycosidase F treatment the molecular weight is substantially reduced. The main band is 32,500 daltons. Carbohydrate O-linked chains, which are mainly represented by sialic acid, are alpha(2-6)-linked to galactose or N-acetylgalactosamine. Galactose is also organized in O-linked chains and it is beta(1-4)-linked to N-acetylglucosamine and beta(1-3)-linked to acetylgalactosamine. Biochemical analysis of carbohydrate structures reveals that no specific carbohydrate complex is bound to a single isoform.

Acetylgalactosamine↗
PubMed1997

Influence of APOH protein polymorphism on apoH levels in normal and diabetic subjects.

Apolipoprotein (apo)H (also known as beta 2 glycoprotein-I) is a glycoprotein synthesized by liver cells and it is present in the blood associated with plasma lipoproteins. APOH displays a genetically determined structural polymorphism: three alleles (APOH*1, APOH*2, APOH*3) at a single locus on chromosome 17 code for different isoforms, and population studies have shown that APOH*2 is the most frequent allele. This paper assesses the relation between APOH phenotypes and plasma apoH levels in a population composed of 278 healthy subjects (243 H2/2, 32 H3/2, 2 H3/3, 1 H2/1; allele frequencies APOH*1 0.002, APOH*2 0.934, APOH*3 0.064) and 245 diabetics (212 H2/2, 30 H3/2, 3 H3/3; allele frequencies APOH*2 0.927 and APOH*3 0.073). Determination of apoH levels by competitive ELISA gave a mean value of 26.3 +/- 9.8 mg/dl for all subjects, 22.6 +/- 7.7 in normals vs 30.6 +/- 10.3 in diabetics (p = 0.0001), and 23.0 +/- 7.9, 19.3 +/- 5.4 and 18.5 +/- 3.5 mg/dl for H2/2, H3/2 and H3/3 in normals and 31.1 +/- 10.1, 28.2 +/- 10.8 and 15.7 +/- 9.0 mg/dl in diabetics, respectively. ANCOVA of the adjusted data revealed a significant difference in apoH levels for the three phenotypes in both the normal subjects (p = 0.01) and the diabetics (p = 0.02). ANCOVA of the whole samples of subjects, controlling for diabetes as well as age, sex and total cholesterol, indicated a substantial effect of phenotype, independent of the other variables (p = 0.0007).

Adult↗
PubMed1997

[A new infusion method for a prostacyclin analogue].

BACKGROUND: Iloprost, a prostaglandin I2, is chemically stable and it has been successfully used by intravenous infusion in severe limb ischemia. Usually Iloprost is diluted in 0.9% sodium chloride solution and infused intravenously for six hours each day for 28 days in hospital. METHODS: In the present study after the first three days of infusion with a traditional pump in hospital, a home pump has been utilised for the infusion of Iloprost at home. This device allows the continue infusion of Iloprost at a flow rate of 2 ml/h for six days, then the pump is filled with a new solution. The home pump consists of a protective shell in polycarbonate (10 x 12 cm), 270 ml of volume, inside there is a balloon reservoir (3 membranes) which is filled with Iloprost. The structure of Iloprost does not change into the home pump as evidenced by HPLC studies and its continue infusion allows plasmatic high levels of its active isomers during the 28 days of therapy. In 30 patients, 25 men and 5 women (mean age 61 years) with Fontaine stage IIB (6), III (5) and IV (19) POAD Iloprost has been infused with the home pump. The follow-up period was 1 to 16 months. RESULTS: The results have shown 4 major amputations and 1 death, in 9 patients complete pain relief and ulcer healing, and in 6 patients only improvement in relief of rest pain and ulcers. CONCLUSIONS: All the patients appreciated this system of infusion because they had a normal life; in addition it is less expensive because the patients stay in hospital only 3 days.

Female↗
PubMed1996

Postprandial triglyceride-rich lipoprotein changes in elderly and young subjects.

To determine whether an increased risk for atherosclerosis in older humans is related to changes in postprandial lipoprotein metabolism, we compared the dynamic profiles (0-10 hours) of triglyceride (Tg)-rich lipoproteins and the Tg content in VLDL subfractions in elderly and young subjects after an oral fat load. The plasma Tg response curves displayed significant differences between the groups at all times. Postprandial triglyceridemia was quantified from the plasma response curves as an incremental area, and was significantly different in the two groups (young subjects 231.9 +/- 199.6 vs elderly subjects 511.0 +/- 305.6 mg/dL x 10 hr, p = 0.036). The more scattered VLDL-Tg values were significantly different compared to values at baseline and 6 hours after fat load. Tg baselines in the four VLDL subfractions (expressed as percentages) were higher in the larger particles (B Sf = 175-400) in the elderly subjects, and in the smaller, denser particles (D Sf = 20-100) in the young subjects. In both groups, postprandial hyperlipidemia increased the Tg content of the larger, less dense particles (Sf more than 400), and reduced that of the denser particles. These variations usually coincided with the plasma Tg and VLDL peaks: 63% to 70% above the Tg baseline between the 2nd and 4th hour in all the young subjects: 48% to 68% above the baseline between the 4th and the 6th hour in all the elderly subjects. Total cholesterol variations showed no significant differences between the two groups at any time. All subjects tested for the missense mutation at codon 188 of the human lipoprotein lipase (LPL) gene resulted noncarriers of LPL mutant alleles. Our data show that, after a fatty meal, healthy elderly subjects tend to present prolonged postprandial hypertriglyceridemia, suggesting an atherogenic behavior of their lipid metabolism.

Adult↗
PubMed1996

Apolipoprotein H: a two-step isolation method.

A new method for the purification of apolipoprotein H by affinity chromatography followed by continuous-elution electrophoresis is described. It is both simpler and less complicated than the chromatographic and electrophoretic methods usually used. In addition, apolipoprotein H is isolated in a pure, structurally uncleaved form. This is of importance, as impairment has been detected in commercial preparations. The separation and purification of apolipoprotein H is a necessary prelude to its quantitative determination and phenotyping, and hence the clarification of its physiopathological mechanisms in lipid metabolism.

Amino Acid Sequence↗
PubMed1995

Apolipoprotein E allele frequencies in an Italian population: relation to age and lipid profile.

Apo E phenotype and plasma Tg, Chol, LDL-Chol, HDL-Chol and Apo B levels were determined in a sample of 228 healthy Italian subjects (124 men and 104 women) aged 18-93. The allele frequencies were: epsilon 2 = 0.070; epsilon 3 = 0.829; epsilon 4 = 0.101 (among the lowest values in the literature). Division of the sample into four age groups indicated that epsilon 4 frequency decreased with age to 0 in persons aged over 75. Covariance analysis of the influence of each allele on plasma lipids showed that epsilon 4 was significantly associated with the highest Chol, LDL-Chol and Apo B levels. These data are evidence of the influence of epsilon 4 on Chol metabolism in an Italian population. They also show that its frequency decreases with age.

Adolescent↗
PubMed1995

Evaluation of new small barium alginate microcapsules.

Microencapsulation of islets of Langerhans has been proposed in order to prevent immune rejection and possible recurrence of autoimmune disease. This study introduces a fast simple one-step microencapsulation procedure which allows the production of small sized barium-alginate beads. The volume of the microcapsules produced was approximately that of the encapsulated islets. Consequently, the insulin kinetics and the oxygen diffusion were favoured, while the transplanted tissue volume was reduced. Electron microscopy and immunoisolating testing were performed to evaluate the molecular cut-off, the physical and chemical characteristics of these microcapsules. Immunohistochemical staining and perifusion experiments of microencapsulated pancreatic islets showed their viability after the encapsulation procedure as well as in vivo experiments. In fact, microencapsulated porcine islets were implanted intraperitoneally into streptozotocin-diabetic rats. The xenografts reversed the hyperglycemic state and functioned for a period ranging from 9 to 385 days. The low mannuronic acid concentration and the purity grade of the alginate, exerted a combined influence on the capsule biocompatibility as in vivo studies showed.

Alginates↗
PubMed1994

Influence of apolipoprotein H polymorphism on levels of triglycerides.

Human apolipoprotein H (apo H) displays a genetically determined structural polymorphism: three alleles (H*1, H*2 and H*3) on chromosome 17 code for the six phenotypes (three homozygotes and three heterozygotes). The effect of apolipoprotein polymorphism on individual variations in plasma lipoprotein levels has been underscored in recent years. Since apo H is involved in metabolism of triglycerides (Tg), its phenotype could affect Tg levels. This paper reports an investigation of apo H phenotypes in a sample of 217 subjects of the Italian population by means of isoelectrofocussing followed by immunoblotting. The levels of the main lipid parameters were evaluated in relation to phenotype and other influential factors. Analysis of covariance disclosed a significant association between Tg levels (log transformed) and phenotype (F = 8.27, P = 0.004). Comparison of Tg levels between bearers of the two most frequent phenotypes (H2/2 and H3/2) divided by sex and age classes revealed significantly higher levels in male H3/2 heterozygotes (P = 0.0053) and in H3/2 subjects aged less than 50 (P = 0.0095). Our data support the view that there is an association between hypertriglyceridaemia and apo H polymorphism, especially with the H*3 allele.

Apolipoproteins↗
PubMed1994

Plasma beta-thromboglobulin and platelet factor 4 are not increased in insulin-dependent diabetic patients with microalbuminuria.

To evaluate the possibility that platelet dysfunctions contribute to the cardiovascular risk of microalbuminuric insulin-dependent diabetic (IDD) patients, we have measured beta-thromboglobulin (BTG) and platelet factor 4 (PF4) in 74 IDD patients with different degrees of albuminuria (8 macro-, 36 micro- and 30 normoalbuminuric) and in 30 non-diabetic control subjects. BTG values (20.4 +/- 1.5 SEM in normo-, 22.2 +/- 1.2 in micro-, 101.1 +/- 2.9 in macroalbuminuric patients and 21.8 +/- 1.1 IU/ml in control subjects) were significantly higher (P < 0.001) in the macroalbuminuric patients, but similar among the other groups. These results suggest that platelet hyperactivation is not present in the microalbuminuric stage of diabetic nephropathy, only in overt nephropathy.

Adult↗