Biomedical subjects

A M Robert

Publications and source records attributed to A M Robert.

At least 19 recordsLinked to original sources

PubMed2006

Effect of high glucose concentration on corneal collagen biosynthesis.

The effect of high glucose concentration (3 g/l) on bovine corneal total protein and collagen biosynthesis was studied, using 3H-proline incorporation in explant cultures with protein and collagen determinations. The high glucose concentration increased the incorporation of 3H-proline in total corneal proteins as well as in collagens. The specific radioactivity of stromal collagens was strongly increased in these conditions. Mannitol was used to control the osmotic effect of the high glucose concentration, both at 1 and 3 g/l concentrations. Mannitol did not increase the incorporation of 3H-proline in total proteins or collagens, but on the contrary decreased it. The high glucose concentration decreased the excretion of neosynthesized proteins and collagens in the culture medium, but did not affect the total protein or collagen content of the corneas. The strong increase in the specific radioactivity of corneal collagens in the presence of 3 g/l glucose suggests an increased turnover of collagens in diabetic corneas. The increased biosynthesis of collagens together with their decreased elimination in the extracellular compartment can create the conditions for the formation and accumulation of advanced glycation endproducts by the Maillard reaction. This can induce and stimulate the liquefaction of the vitreous body leading to sight-threatening disorders such as diabetic retinopathy, retinal detachment, glaucoma, cataract formation and age-related macular degeneration.

Animals↗

PubMed2005

MMP-type endopeptidase activity in the cornea. Its evolution during organ culture storage at the Eye Bank. Effect of hyaluronan.

About 46% of total corneas obtained from donors in the French Eye Bank cannot be grafted for several reasons as loss of endothelium or other. Corneal cells express proteolytic enzymes, essentially matrix metallo-proteinase MMP-2 and MMP-9. In presence of hyaluronan and some other GAG-s their activity increases as could be shown on keratocyte cultures. Hyaluronan concentration increases during in vitro preservation and can represent a serious hazard for corneal conservation. The control of MMP-release and activation might well be one of the factors involved in graft deterioration. We could show however that only a slight fraction (< or =12%) of total, relatively high endopeptidase activity of the cornea is released in the media during storing. It appears therefore that most of the proteolytic activity determined in corneal extracts remains confined to the stroma and might not represent an important risk for preservation, at least for the endothelium.

Cornea↗

PubMed2005

Effect of procyanidolic oligomers of Vitis vinifera on the biosynthesis and excretion of corneal glycosaminoglycans.

Glycosaminoglycans (GAG) as hyaluronan and proteoglycans (PG) as dermatan sulfate (DSPG) and keratan sulfate (KSPG) play important role in the structure and physiological functions of cornea. Several corneal pathologies are associated with qualitative and quantitative alterations of GAG and/or PG biosynthesis. For these reasons we investigated the effect of procyanidolic oligomers (PCO) on the biosynthesis and excretion of GAG-s in bovine corneal explant cultures. This was accomplished by the determination of their chemical components, hexoses, hexosamines and uronic acids as well as the incorporation of (35)S-sulfate in the presence and without PCO in the culture medium. In presence of 1 mg/ml PCO total hexoses, hexosamines and uronic acids increased at 5 and 24 h of incubation, hexoses less than the two other components. Stimulation of biosynthesis concerned essentially the uronic acid containing GAG-s, DS and hyaluronan. (35)S-sulfate incorporation decreased in presence of PCO, showing a decreased sulfation of GAG-s. The most striking effect was however the strongly increased excretion of neosynthesized GAG-s in the culture medium. This effect could be beneficial in decreasing corneal hydration in inflammatory conditions, together with the previously demonstrated protection of corneal macromolecules against proteolytic degradation.

Animals↗

PubMed2005

[Effect of procyanidolic oligomers on corneal collagen fibrillogenesis].

PURPOSE: We showed in a previous study that procyanidolic oligomers (PCOs) from grape seeds effectively protect corneal stroma against degradation by bacterial collagenase. Here we report the study of the effect of PCOs on protein and collagen composition of cornea and on the biosynthesis of corneal collagens. METHODS: Bovine corneas were used in explant cultures. We quantitatively determined total proteins and collagen as well as the incorporation of 3H-proline in separated collagen types. Collagens type I, V, and VI were separated and quantitated. In order to understand some of the results obtained, we studied the interaction of PCO with collagen type I separately. RESULTS: In the absence of PCOs, collagen typing and 3H-proline incorporation yielded the expected results for a normal cornea, with the usual proportion of the three major types of collagens. In presence of PCOs at 1 mg/ml and after 24 h incubation, total proteins and collagens decreased, as did papain-extractable collagens. Proteins in the final residue solubilized in 1 M KOH-80% v/v aqueous ethanol increased. The proportion of the three principal collagens was also modified: type I became preponderant, and the proportions of the two others (type V and VI) decreased. The study of the interaction of collagen type I with PCOs showed that 30% of total PCOs do not interact with collagen, 20% interact reversibly, and 50% of PCOs are strongly and irreversibly fixed. This strongly fixed fraction could not be separated from collagen by either column chromatography or collagenase or KOH in aqueous ethanol. CONCLUSIONS: Bovine corneas in explant cultures in presence of PCOs undergo a modification of their protein and collagen content, with a concurrent modification of the proportion of collagens types I, V, and VI. Collagen type I predominates, and the two other types decrease. Study of the collagen type I-PCO interaction showed that roughly 50% of PCOs become irreversibly fixed to collagen, resisting collagenase or other methods of separation. This strong PCO-collagen type I interaction explains the decrease in solubility.

Animals↗

PubMed2005

Effect of a preparation containing a fucose-rich polysaccharide on periorbital wrinkles of human voluntaries.

BACKGROUND: Skin aging is accompanied by wrinkle formation. At some sites, as the periorbital skin, this is a relatively early phenomenon, variable from one woman to the other and even between the right and left eye. PURPOSE: We tested the effect of a preparation with fucose-rich oligo- and polysaccharide (FROP-3) on periorbital wrinkles (crow's feet) on a team of voluntaries. METHOD: The efficiency of a preparation containing as active principle a fucose-rich polysaccharide in a base-cream for the treatment of temporal periorbital wrinkles on 20 voluntary women was tested as follows: the women applied the cream twice a day for 4 weeks. Negative replicas were taken on the periorbital skin before and after 4 weeks of application of the cream. The results were evaluated by semi-automated morphometry on the plastic replicas and expressed as the 'wrinkle-factor' obtained by multiplying total wrinkle length in millimeters by the average wrinkle width. RESULTS: After 4 weeks of treatment, there was a significant improvement of the periorbital wrinkles for the majority of the voluntary team who tested the cream. On the 29 periocular wrinkles examined 65% showed an improvement at the end of the treatment. In two cases, the improvement was 100% on one eye (disappearance of the crow's feet) and of 75% and 79% on the other eye. On six eyes, there was no significant change after the end of the treatment, and on four eyes a worsening was observed. The age-dependent modifications showed that improvement was independent of age and was the result of individual skin reactions to the cream. The occurrence and the severity of crow's feet is highly individual, and differs from one side of the face to the other for the same person. These strong individual variations explain the highly variable results, not only from one woman to the other but also comparing one side of the face to the other for the same person. CONCLUSION: Under the effect of 4 weeks of treatment with the FROP-3-containing cream, most of the voluntaries had their periorbital wrinkles attenuated and some women showed a total regression of crow's feet on one of their eyes.

Adult↗

PubMed2004

Effects of L-fucose and fucose-rich oligo- and polysaccharides (FROP-s) on collagen biosynthesis by human skin fibroblasts. Modulation of the effect of retinol, ascorbate and alpha-tocopherol.

The action of L-fucose and a fucose-rich oligosaccharide (FROP-3) on skin explant cultures and fibroblast cell cultures, alone or together with three vitamins (A, C, E, often used in topical preparations) was studied, using as criteria collagen accumulation and collagen and non-collagen protein biosynthesis. In order to confirm the results observed, several variants of the chemical procedure were used. Collagen accumulation in cell cultures and (3)H-proline incorporation in proteins and collagen gave comparable results. L-Fucose produced a slight but significant inhibition of collagen accumulation and of incorporation of radioactive proline in cell associated collagen and also modified the ratio of excreted (soluble) to cell- or tissue-bound collagen and non-collagen proteins. The oligosaccharide (FROP-3) did not produce a comparable inhibition of collagen biosynthesis. Both L-fucose and FROP-3 modulated the action of the two above-mentioned vitamins in most experimental conditions used. The combined action of the three vitamins (all-trans retinol, ascorbate, alpha-tocopherol) with L-fucose and even more so with FROP-3 can be considered as favourable for the modulation of the biosynthetic activity of fibroblasts.

Ascorbic Acid↗

PubMed2004

Pharmacology of skin aging. Stimulation of glycosaminoglycan biosynthesis by L-fucose and fucose-rich polysaccharides, effect of in vitro aging of fibroblasts.

The effect of L-fucose and fucose-rich polysaccharides (FROP-s [Biomed. Pharmacother., 2003; 57: 187-94]) was investigated, using human skin fibroblast cultures at several passages. The cells were incubated with radioactive glucosamine for 24 h, followed by the determination of individual glycosaminoglycans (GAG-s) by selective hydrolysis using specific enzymes. The effect of L-fucose and of FROP-3 [Biomed. Pharmacother., in 2003; 57: 187-94], both at 1 and 10 microg/ml, added to the culture medium, was investigated. L-Fucose stimulated the incorporation of the tracer in heparan sulfates by fibroblasts at the ninth passage by 20%. FROP-3 stimulated incorporation in keratan sulfates by 45% by fibroblasts of the fifth passage. This effect was identical at both concentrations tested. For fibroblasts at the ninth passage FROP-3 stimulated incorporation in dermatan sulfate. This effect was dose dependent of the order of +67% at 1 microg/ml and +128% at 10 microg/ml. Incorporation in hyaluronan was also stimulated by about +27%. These stimulations of GAG-biosynthesis might play a role in the increase of total skin thickness of hairless rats treated with L-fucose, as well as in several other favorable results recorded for FROP-3 such as the increased hydration (resistance to pressure) and elasticity of human skin (Robert C, Robert AM, Robert L).

Cells, Cultured↗

PubMed2004

Effect of L-fucose and fucose-rich polysaccharides on elastin biosynthesis, in vivo and in vitro.

With increasing age elastic fibres in human skin are progressively lysed and skin elasticity is also decreasing. Still there is an age-dependent increase of elastic fibre surface density, mostly due to an alteration of the fibres. The present experiments were undertaken to explore if L-fucose and fucose-rich polysaccharides (FROP-s) could influence elastin biosynthesis. We show here, that topical application of a fucose-containing preparation to the skin of hairless rats increased after 4 weeks the elastic fibre surface density by about 40%, shown by quantitative morphology. Using human skin fibroblasts in explant cultures, the addition of L-fucose or of FROP-3 increased the biosynthesis of immunoprecipitable tropoelastin by about 40%. No increase was found however of desmosine-isodesmosine in skin explant cultures after 72 h of incubation. The effect of L-fucose and FROP-3 on the biosynthesis of collagen and non-collagen proteins excreted by the skin explant cultures was also investigated. L-fucose, but not FROP-3, decreased collagen biosynthesis but both increased non-collagen protein biosynthesis. These results show that L-fucose and FROP-3 stimulate tropoelastin biosynthesis in vitro, and elastic fibre formation in vivo. This stimulation concerns also several non-collagen proteins secreted by skin explant cultures. Elastic fibre formation necessitates the simultaneous synthesis of several microfibrillar glycoproteins as well as of tropoelastin. The increased elastic fibre density in the in vivo experiments suggests that this is indeed achieved by L-fucose and FROP-3, further demonstrating their efficiency in the control of age-dependent modifications of connective tissues in general and of skin in particular.

Administration, Cutaneous↗

PubMed2003

Aging, from basic research to pathological applications.

This introductory chapter tentatively draws a parallel between some of the highlights of basic aging research and their incidence for the understanding of age-associated pathologies. Some of the most important mechanisms described for cell- and tissue-aging, such as limited cell-proliferation, "parasitic" reactions, such as non-enzymatic glycation, ROS-production, uncontrolled proteolysis involving matricryptic sites, receptor loss and uncoupling appear to be the most promising links between those two approaches to fundamental and clinical gerontology. Some of the "parasitic", epigenetic, post-translational reactions lead to vicious circles which over the years might well produce cell- and tissue-damage as seen in most age-associated diseases. There is little doubt that the better understanding of these relationships will also prove helpful for pharmacological research as related to the age-associated pathologies.

Aged↗

PubMed2003

Effect of a fucose-rich polysaccharide preparation on the age-dependent evolution of the skin surface micro-relief.

It was demonstrated previously that the evaluation of the microdepressionary skin surface relief by semi-automated computerised morphometry enables the determination of several of its geometrical parameters, such as, the average number of polygons in a given microscopic field, and/or the average surface of the polygons. Using this procedure, it could be shown in a study on about 100 persons, males and females, aged from 6 months to 89 years that these parameters undergo age-related alterations. These alterations are faster at younger ages, slow down at middle ages and accelerate after 50-60 years again. Taken at a sun-protected site, the interior face of the forearm, close to the elbow, this test reflects the effect of chronological ageing of the skin surface micro-relief. In the present work we tested the effect of a preparation containing a fucose-rich polysaccharide as active principle on the microdepressionary skin surface micro-relief evaluated by a morphometric method, comparing results before treatment and after 4 weeks of treatment. We could demonstrate on 20 female volunteers, aged from 39 to 71 years, that after 4 weeks of treatment with the abovementioned preparation there was a significant improvement of the skin surface relief, as shown by the displacement of geometrical characteristics of 17 treated skins out of 20, towards a "younger" pattern, as for instance the increase of the average number of polygons in a given microscopic field. This improvement was less than 15% of the untreated value for two persons, and more than 15% for 15 women out of 20. The average improvement was 37.16%, and for the 15 persons showing more than 15% improvement, it was nearly 50%, corresponding to a decrease of apparent age by 10-15 years. These results suggest that a treatment with the fucose-rich polysaccharide preparation can slow down human skin ageing and even can reverse age-dependent skin alterations.

Adult↗

PubMed2002

Regulation of elastase-type endopeptidase activity, MMP-2 and MMP-9 expression and activation in human dermal fibroblasts by fucose and a fucose-rich polysaccharide.

Tissue loss during ageing and age-dependent pathologies are the result of a disturbed regulation of proteolytic activities. Elastase-type endopeptidases, especially MMP-2 and -9, play an important role in this respect. Dermal fibroblast cultures and skin explant cultures were used in order to measure the efficiency of fucose and fucose-rich polysaccharides to downregulate the elastase-type endopeptidase activity. Fucose and fucose-rich polysaccharides were shown to downregulate this elastase-type activity, the basic activity and also the hyaluronan or kappa-elastin-stimulated activity. In skin explant cultures, we could demonstrate that fucose and fucose-rich polysaccharides produced an inhibition of the activation of the pro-form to the active form of MMP-9. Here, we show that mono-, di-, oligo- and polysaccharides acting on the elastin-laminin receptor and/or on the fucose-mannose receptor are efficient inhibitors of such enzymes by downregulating elastase-type endopeptidase activity, both at the level of their biosynthesis and at the level of the activation of the pro-enzymes. Fucose and fucose-rich polysaccharide preparations were shown to be efficient modulators of MMP-2 and MMP-9, activity with potential therapeutic applications in age-related pathologies accompanied by tissue loss.

Child, Preschool↗

PubMed2002

[Protection of cornea against proteolytic damage. Experimental study of procyanidolic oligomers (PCO) on bovine cornea].

AIM OF THE STUDY: The purpose of this work was the development of an antiprotease treatment for the cornea and later for other ocular tissues. Indeed, many proteases participate in various ocular pathologies and no really effective treatments are currently available. METHODS: We worked on bovine corneas in organ cultures, incubated in the presence and absence of bacterial collagenase. We used procyanidolic oligomers from grape seeds (PCO) as the antiproteolytic agent, added to the cultures in two different concentrations. RESULTS: The corneas incubated in the presence of collagenase were quickly attacked and their degradation was practically complete after 24 hours. With the low concentration used (0.066 mg/ml), proteolysis was only slightly inhibited. With the higher concentration of PCO (1 mg/ml), the collagenolysis was completely prevented and the corneas were completely preserved. CONCLUSION: These results show that the protective effect of the PCO against proteolytic attack, demonstrated previously on other tissues, proved to be effective also for the cornea in vitro. An adequate method of administration in clinical practice must now be developed in order to obtain an effective clinical antiproteolytic treatment.

Animals↗

PubMed2001

Chemotactic penetration of keratocytes in ePTFE polymer in vitro.

Expanded polytetrafluoroethylene (ePTFE) is used as a support for artificial corneas. Implanted in corneas, most of the time this polymer is colonized by corneal host cells. The absence of colonization often coincides with extrusion of the polymer. Therefore, we decided to introduce keratocytes into ePTFE in vitro before implantation. Because keratocytes do not spontaneously enter ePTFE, we used several chemoattractants, separately and in a mixture, to stimulate the penetration of cultured keratocytes into the polymer. The influence of the passage number on cell penetration was also studied. No significant differences were observed up to the seventh passage, although seventh-passage cells penetrated somewhat more slowly than younger cells. Satisfactory results were obtained with four of the tested chemotactic factors: IL-6, type alpha transforming growth factor (TGF-alpha), platelet derived growth factor isoform BB (PDGF-BB), and fibroblast growth factor-2 (FGF-2). Under our experimental conditions, two to more than six million keratocytes entered the polymer discs with a volume of 706.5 mm(3) in the presence of these four chemoattractants. TGF-alpha was the most efficient and was selected for further in vitro and in vivo studies.

Biocompatible Materials↗

PubMed2001

Corneal collagens.

Cornea is a highly differentiated tissue rich in extracellular matrix (ECM) specifically distributed in space in order to insure its dual role--transparency and protection of inner eye-tissues. Corneal ECM is especially rich in collagens. Since the characterisation of a number of distinct collagen types it appeared that most of them are present in the cornea. Their synthesis follows a specific program of sequential expression of the different collagen types to be synthesised during the development and maturation of the cornea. The precise regulation of the diameter and orientation of fibers, and of the interfibrillar spaces is partially at least attributed to interactions between glycosaminoglycans and collagens. The 'program' of vectorial collagen synthesis and GAG-collagen interactions changes also with age and in several pathological conditions as corneal dystrophies and wound healing. The Maillard reaction, especially in diabetes, is one of these important factors involved in age-dependent modifications of corneal structure and function. Far from being inert, corneal collagens were shown to have relatively short half-lives. The biosynthesis of corneal collagens was studied also during wound healing. The refibrillation of wounded corneas does not follow the original 'program' of ECM-synthesis as shown by the comparative study of wound healing using biochemical and morphometric methods. This review recapitulates briefly previous and recent studies on corneal collagens in order to present to clinicians and scientists an overview of the state of the art of this important field at the intersection of eye research and matrix biology.

Aging↗

PubMed2001

Effect of an alpha-blocker (Nicergoline) and of a beta-blocker (Acebutolol) on the in vitro biosynthesis of vascular extracellular matrix.

The effect of an alpha-blocking agent and of a beta-blocking agent on the biosynthesis of extracellular matrix macromolecules of the arterial wall was investigated. Rabbit aorta explants were cultured up to 48 hours with radioactive proline, lysine or glucosamine. In presence of these drugs, at concentration shown to be effective for the inhibition of platelet-endothelial cell interactions (10(-7) M), the incorporation of 14C proline in total macromolecular proline was higher than in macromolecular hydroxyproline suggesting a relatively higher rate of biosynthesis of non-collagenous proteins as compared to collagens. The alpha-blocking increased the incorporation of 14C proline in collagenous and non-collagenous proteins after 18 hours of incubation. beta-blocking also increased the incorporation of proline in macromolecular proline and hydroxyproline as compared to control cultures. Both increased the incorporation of 3H glucosamine in newly synthesised glycosaminoglycans. beta-blocking increased mainly the neosynthesis of heparan sulphate, alpha-blocking that of hyaluronan. The incorporation of 14C-lysine in crosslinked, insoluble elastin was not modified. These experiments confirm that alpha and beta-blocking agents can influence not only the tonus of aortic smooth muscle cells but also the relative rates of biosynthesis of extracellular matrix macromolecules. This effect should be taken in consideration for the evaluation of the long range effect of alpha and beta-blocking drugs on the vascular wall.

Acebutolol↗

PubMed2001

Effect of procyanidolic oligomers on the permeability of the blood-brain barrier.

Blood-brain barrier (BBB) is the site of regulatory mechanisms which control the exchange of substances between the brain and the blood through the wall of 'true' brain capillaries with tight junctions between endothelial cells. In some pathological situations the permeability of the BBB is increased because of a partial proteolytic degradation of some constituents of the capillary basement lamina. In such cases it is important to restore normal permeability. The effect of procyanidolic oligomers (PCO) on the BBB was investigated in vivo with quantitative morphologic procedures. We also investigated the action of this drug on collagen and basement lamina constituents (Matrigel) in vitro. Collagenase injected in lateral brain ventricles was shown to increase BBB permeability. Per os administration of PCO to rats greatly increased the resistance of brain capillaries to bacterial collagenase, as shown by the inhibition of the diffusion of fluorescein-isothiocyanate-marked dextran particles from the blood-stream into the brain tissues. Calf skin collagen pretreated in vitro with PCO became more resistant to the hydrolytic action of collagenase. Similar, even more intense protective effect was seen when basal lamina constituents containing type IV collagen was incubated with PCO before exposure to pronase. These in vitro effects may partly explain the in vivo protective effect of PCO against the alteration of brain capillaries by i.v. injected bacterial collagenase.

Animals↗

PubMed1999

Condom practices of urban teens using Norplant contraceptive implants, oral contraceptives, and condoms for contraception.

OBJECTIVES: The availability of long-acting hormonal birth control methods has created new contraceptive options for adolescents. The purpose of this study was to determine whether teens initiating these methods use condoms less frequently than teens using oral contraceptive pills or condoms alone and may therefore be at an increased risk of acquiring sexually transmitted infections. STUDY DESIGN: To investigate ongoing condom behavior in teens using levonorgestrel (Norplant) contraceptive implants, oral contraceptives, and condoms alone, we examined data from a 2-year prospective cohort study of 399 urban teens. The study consisted of 3 clinic-based cohorts of adolescent female contraceptive users: Norplant contraceptive implants (n = 200), oral contraceptives (n = 100), and condoms alone (n = 99). Data were collected at an admission interview and at 1- and 2-year follow-up from method continuers. RESULTS: Norplant contraceptive implant users were less likely than oral contraceptive or condom users to report condom use at last sex or consistent condom use at 1- and 2-year follow-up. The implant group showed a significant decrease in condom use from admission to 2 years after method initiation. The proportion of implant users self-reporting new sexually transmitted infections at 2-year follow-up, however, was not significantly greater than that of oral contraceptive or condom users. CONCLUSIONS: Our findings indicate that teen users of Norplant contraceptive implants are less likely to use condoms than teens who choose oral contraceptives but, probably because of differences in sexual behavior, are no more likely to self-report sexually transmitted infections. Our findings also indicate that teens who choose oral contraceptives and condoms do not use them consistently enough to avoid pregnancies or sexually transmitted infections.

Adolescent↗

PubMed1998

Elastin-elastase-atherosclerosis revisited.

This review proposes reinvestigation of a topic studied in the author's laboratory over the last decades concerning the age-dependent modifications of the vascular extracellular matrix (ECM) as related to atherogenesis and its recognized risk-factors: blood lipids, lipoproteins. Most salient previous results are confronted with recent publications in this field. Age-dependent modifications of the vascular wall discussed in this review include upregulation of elastolytic enzymes, demonstrated for the first time in the vascular wall in this laboratory, matrix biosynthesis and receptor function. The progressive deposition of lipids in elastic tissues as well as the addition of lipoproteins or lipids to cell and organ cultures were shown to modify matrix biosynthesis and upregulate elastase expression. Lipid-elastin interactions exhibit a great deal of specificity as shown by the nature and amount of lipids accumulating in elastin in vivo and in vitro. Recent epidemiological studies (the EVA study) enables the confrontation of blood lipid parameters with matrix related components (serum elastase and inhibitors, elastin peptides, fibronectin) in the same blood samples. The elastin laminin receptor present on vascular cells was shown to trigger NO dependent vasodilation, and downregulation of cholesterol synthesis. Both of these functions decrease or disappear with age except the upregulation of elastase release which is preserved and increased. Recent experiments extended these findings to T-lymphocytes present also in the atherosclerotic plaque. Finally several recent publications are analyzed which give more precision on the cellular mechanisms underlying the above-described modifications.

Adult↗