Biomedical subjects

M P Jacob

Publications and source records attributed to M P Jacob.

At least 19 recordsLinked to original sources

PubMed2001

Differential expression of matrix metalloproteinases after stent implantation and balloon angioplasty in the hypercholesterolemic rabbit.

BACKGROUND: Intimal hyperplasia is the principal mechanism of in-stent restenosis. Matrix metalloproteinases (MMPs) play a key role in intimal growth after balloon angioplasty (BA). Little is known, however, about MMP expression after stent implantation (ST). We investigated whether MMP9 and MMP2 are differentially expressed after ST and BA. METHODS AND RESULTS: Hypercholesterolemic rabbits underwent ST and BA in the right and left iliac arteries, respectively. The expression of MMPs and their inhibitors (TIMPs) was studied at various time points in the injured arteries by use of zymography, reverse transcription-polymerase chain reaction, and immunohistochemistry. MMP2, but not MMP9, was constitutively expressed in uninjured arteries. MMP9 expression was rapidly induced after injury, whereas the increase in MMP2 expression was delayed. At all time points, pro-MMP9 activity and MMP9 mRNA levels were >/=2-fold (ANOVA, P=0.002) and >/=3-fold (P<0.0001) higher after ST than after BA, respectively. Active MMP9 was detected only after ST. Although the increases in MMP2 mRNA levels were of similar magnitudes after ST and BA, pro-MMP2 activity was slightly higher 7 and 30 days after ST, and MMP2 activity was >/=2-fold higher 7 to 60 days after ST (P=0.002). No difference in TIMP expression was observed between stented and balloon-injured arteries. Cellular distributions of MMPs and TIMP1 were similar after ST and BA. Early inflammatory cell recruitment and 30-day intimal growth were more severe after ST. CONCLUSIONS: Stent implantation results in more intense and sustained expression of MMP9 and activation of MMP2 than balloon angioplasty.

Angioplasty, Balloon↗

PubMed2001

Smooth muscle cell modulation and cytokine overproduction in varicose veins. An in situ study.

The exact aetiology and physiopathology of varicose disorders remain unclear. The aim of the present work was to study, in situ, the morphology and composition of cellular and matrix components in varicose veins compared with control veins and to identify factors that could contribute to varicose remodelling. A combined histological, immunohistochemical, and biochemical approach was used. Longitudinal sections of varicose (n=12) and control veins (n=9) were studied to assess the organization, structure, and phenotype of smooth muscle cells; the localization of microvascular endothelial cells; the distribution of connective tissue proteins; and the localization of cytokines. These cytokines were further quantified by ELISA. Considerable heterogeneity of the varicose vein wall was observed, with a succession of hypertrophic and atrophic segments, presenting severe disorganization of the medial layer and numerous areas of intimal thickening. In hypertrophic portions, medial smooth muscle cells showed marked alterations suggesting modulation from a contractile to a proliferative and synthetic phenotype; furthermore, the number of vasa vasorum was increased. In contrast, in atrophic portions, both cellular and matrix components were decreased. TGFbeta1 (p< or =0.005) and bFGF (p< or =0.001) were increased and VEGF was not significantly modified in varicose veins when the results were expressed per mg of DNA. These results show that phenotypic modulation of smooth muscle cells, altered extracellular matrix metabolism, and angiogenesis are the main mechanisms contributing to the morphological and functional modifications of varicose remodelling. The increased expression of bFGF and TGFbeta1 by varicose vein cells may play a pivotal role in the hypertrophy of the venous wall, but the exact mechanism leading to aneurysmal dilatations remains to be elucidated.

Adult↗

PubMed2001

Effects of interleukin-10 on monocyte/endothelial cell adhesion and MMP-9/TIMP-1 secretion.

OBJECTIVE: Monocyte adhesion to endothelial cells and subsequent secretion of matrix metalloproteinases (MMPs) by activated macrophages are key events in arteriosclerosis and restenosis. We tested the hypothesis that interleukin-10 (IL-10), a potent anti-inflammatory cytokine, inhibits monocyte-endothelial cell interactions. METHODS: The effect of IL-10 on monocyte/endothelial cell adhesion, as well as on the expression of MMP-9 and the tissue inhibitor of MMP-9, TIMP-1, were first tested in vitro in coculture systems. In addition, we used an ex vivo binding assay to study the inhibitory effect of IL-10 on monocyte adhesion to carotid arteries obtained from either normal, or L-nitro arginine-methyl ester (L-NAME)-treated rats. The effect of IL-10 on the expression of monocyte adhesion molecules (CD18 and CD62-L) was studied by flow cytometry. RESULTS: IL-10 (150 ng/ml) inhibits monocyte adhesion to endothelial cells (by 35%) and to carotid arteries (by 40 and 50%, in normal and L-NAME-treated rats, respectively), via direct modulation of the expression of CD18 and CD62-L. Moreover, IL-10 dose-dependently decreases MMP-9 activity and increases TIMP-1 levels in coculture systems, both at the transcriptional level. CONCLUSIONS: Our results suggest that IL-10 is an important modulator of monocyte-endothelial cell interactions.

Analysis of Variance↗

PubMed2001

Age-related alterations in the signal transduction pathways of the elastin-laminin receptor.

With aging we assist to alterations in the vascular structure and function. One important factor in these vascular wall changes is the degradation of the elastin fibre major protein: elastin. Elastin peptides derived from the degradation are present in human sera. Elastin peptides induce on fibroblasts, phagocytic cells, lymphocytes, smooth muscle cells and endothelial cells, a variety of biological effects mediated by the elastin-laminin receptor which has been demonstrated to be present on the membrane of these cells. The transduction pathway of the ELR receptor involves the activation of phospholipase C (PLC) by a pertussis toxin sensitive G-protein. PLC induces the production of inositol trisphosphate (IP3) leading to the increase of the intracellular free calcium on one hand, and of diacylglycerol (DAG) which stimulates the translocation to the membrane of PKC leading to the phosphorylation of members of the MAPK family, such as p42/p44 MAPK. A progressive age dependent uncoupling of the elastin-laminin receptor occurs impairing its transduction pathway and which results in alteration of the calcium signaling and loss in calcium homeostasis of the cells. These alterations in the signal transduction of the elastin-laminin receptor result in modified activities of parenchymal and phagocytic cells with aging, such as free radical production and elastase release. Thus, these age-related alterations in the elastin-laminin receptor signal transduction may be involved in the atherogenesis.

Aging↗

PubMed2001

Extracellular matrix remodeling in the vascular wall.

The extracellular matrix provides a structural framework essential for the functional properties of vessel walls. The three dimensional organization of the extracellular matrix molecules--elastin, collagens, proteoglycans and structural glycoproteins--synthesized during fetal development--is optimal for these functions. Early in life, the vessel wall is subjected to injury: lipid deposition, hypoxia, enzyme secretion and reactive oxygen species production during inflammatory processes, and the extracellular matrix molecules are hydrolyzed by proteases--matrix metalloproteinases, leukocyte elastase, etc. In uninjured arteries and veins, some proteases are constitutively expressed, but through the control of their activation and/or their inhibition by inhibitors, these proteases have a very low activity. During the occurrence of vascular pathologies--atherosclerosis, hypertension, varicosis, restenosis, etc.--the balance between proteases and their inhibitors is temporally destroyed through the induction of matrix metalloproteinase gene expression or the secretion of enzymes by inflammatory cells. Smooth muscle cells, the most numerous cells in vascular walls, have a high ability to respond to injury through their ability to synthesize extracellular matrix molecules and protease inhibitors. However, the three dimensional organization of the newly synthesized extracellular matrix is never functionally optimal. In some other pathologies--aneurysm--the injury overcomes the responsive capacity of smooth muscle cells and the quantity of extracellular matrix decreases. In conclusion, care should be taken to maintain the vascular extracellular matrix reserve and any therapeutic manipulation of the protease/inhibitor balance must be perfectly controlled, because an accumulation of abnormal extracellular matrix may have unforeseen adverse effects.

Aneurysm↗

PubMed2001

[Regulation of elastin synthesis].

Elastin is the main protein of elastic fibers and confers the property of elastic recoil to the tissues such as arteries, lung, elastic cartilage,... Elastin synthesis goes through several steps: gene transcription, alternative splicing of pre-mRNA, mRNA translation, hydroxylation of some proline residues of the newly synthesized protein-tropoelastin-, association of with a 67 kDa chaperone protein, secretion of tropoelastin molecules in the extracellular space, and their deposition on the microfibrillar scaffold which contains fibrillin 1, fibrillin 2, MAGP 1 and MAGP 2,.... After the synthesis of cross-links-lysinonorleucine, desmosine, isodesmosine-, elastin becomes insoluble and elastic. The elastogenic pathway is regulated at many levels. The most recently described regulatory mechanism of elastin synthesis is the control of elastin mRNA stability. Elastogenesis is well controlled during development and aging but remains responsive to external factors such as soluble compounds-cytokines, vitamins, hormones,...- and hemodynamic stress. In order to ensure its function, both quantity and quality of elastin should be and should remain optimal in elastic tissues.

Alternative Splicing↗

PubMed2001

[The elastin-laminin receptor].

Elastin is a major component of the extracellular matrix. Elastin peptides derived from its degradation are present in human sera. Elastin peptides induce on fibroblasts, phagocytic cells, lymphocytes, smooth muscle cells and endothelial cells, a variety of biological effects mediated by the elastin-laminin receptor which has been demonstrated to be present on the membrane of these cells. The transduction pathway of the ELR receptor involves the activation of phospholipase C (PLC) by a pertussis toxin sensitive G-protein. PLC induces the production of inositol trisphosphate (IP3) leading to the increase of the intracellular free calcium on one hand, and of diacylglycerol (DAG) which stimulates the translocation to the membrane of PKC leading to the phosphorylation of members of the MAPK family, such as p42/p44 MAPK. Considering the multiple biological effects of ELR the elucidation of the complexity of the signaling pathways will help to better modulate it, mainly in pathological situations such as atherosclerosis.

Animals↗

PubMed2000

Increased TIMP/MMP ratio in varicose veins: a possible explanation for extracellular matrix accumulation.

Primary varicose veins are functionally characterized by venous back-flow and blood stagnation in the upright position. Dilatation and tortuosity provide evidence for progressive venous wall remodelling, with disturbance of smooth muscle cell/extracellular matrix organization. Affected areas are not uniformly distributed, some areas being hypertrophic, whereas others are atrophic or unaffected. In 12 varicose veins and ten control veins, the proteolytic enzyme/inhibitor balance which may participate in the remodelling of the venous wall was investigated. For this purpose, the presence and enzymatic activity of matrix metalloproteinases (MMP-2, MMP-9), tissue inhibitors of MMPs (TIMP-1, TIMP-2), urokinase-type (uPA) and tissue-type (tPA) plasminogen activators (PAs), and plasminogen activator inhibitor-1 (PAI-1) were quantified by western blot and gelatin or plasminogen-casein zymography. In addition, MMP-2, TIMP-1, TIMP-2, and PAI-1 levels were measured by ELISA. A high TIMP-1 level and a low MMP-2 level/activity were found in varicose veins (p<0.005), resulting in a three-fold increase in the TIMP-1/MMP-2 ratio in varicose versus control veins. Levels of PAs (uPA and tPA) as well as PAI-1 were both lower in varicose veins (p<0.005), with minimal change in the PAI/PA ratio. These results demonstrate that varicose veins are characterized by a higher than normal TIMP/MMP ratio, which may facilitate extracellular matrix accumulation in the diseased venous wall.

Adult↗

PubMed2000

Keratinocytes influence the maturation and organization of the elastin network in a skin equivalent.

Elastic fibers form a complex network that contributes to the elasticity of connective tissues. Alterations in the elastic fiber network are involved in several disease affecting organs in which compliance of the connective tissue is essential: skin, main vasculature, lung, joints, muscle, and ligament. The aim of our work was to study the deposition, maturation, and organization of elastic fiber components in a dermal equivalent model consisting of collagen-GAG-chitosan seeded with fibroblasts. The influence of keratinocytes was studied in parallel, thus constituting a skin equivalent model. These models were examined by transmission electron microscopy (TEM) and by immunohistochemistry to determine the staining patterns of fibrillin-1 and elastin proteins representative of the microfibrillar framework and of the elastic fibers, respectively. After 2 mo of fibroblast culture in the dermal equivalent, elastin was undetectable, whereas fibrillin-1 staining was weak and microfibrils were infrequently observed by TEM. In the skin equivalent, fibrillin-1 and elastin were detected by immunostaining 15 d after epidermization and TEM revealed the typical structure and organization of the elastic network in the dermis, with elastin deposition on the microfibrillar scaffold. This in vitro skin equivalent model is to our knowledge the first in which elastic fibers have been detected, thus demonstrating the influence of keratinocytes on the maturation and organization of the elastic network.

Antibodies↗

PubMed2000

Molecular plasticity of vascular wall during N(G)-nitro-L-arginine methyl ester-induced hypertension: modulation of proinflammatory signals.

It has previously been reported that hypertension induced by the chronic blockade of NO production is characterized by a proinflammatory phenotype of the arterial wall associated with a periarterial accumulation of inflammatory cells. In the present study, the cellular and molecular mechanisms involved in the luminal and perivascular accumulation of inflammatory cells were evaluated in the aortas of N(G)-nitro-L-arginine methyl ester (L-NAME)-treated rats. Because the medial layer remains intact, putative markers of the resistance of the vascular wall to cell migration and to oxidative stress were also explored. For this purpose, monocyte adhesion, cytokine expression, superoxide anion production, and nuclear factor-kappa B (NF-kappa B) activation were assessed in the aortas of L-NAME-treated rats. Expressions of tissue inhibitor of metalloproteinases-1 (TIMP-1) and heme oxygenase-1 (HO-1) in the aortic wall were also studied as possible markers of such resistance. Chronic blockade of NO production increased ex vivo monocyte adhesion to the endothelium, increased the production of superoxide anions, and activated the NF-kappa B system. In concert with this modification of the redox state of the vascular wall in L-NAME-treated rats, the expression of proinflammatory cytokines interleukin-6, monocyte chemoattractant protein-1, and macrophage colony-stimulating factor was increased. In parallel, expressions of both TIMP-1 and HO-1 were increased. All these changes were prevented by treatment with an angiotensin-converting enzyme inhibitor (Zofenopril). Hypertension associated with a proinflammatory phenotype of the vascular wall induced by blockade of NO production could be due to an increase in oxidative stress, which, in turn, activates the NF-kappa B system and increases gene expression. In parallel, the arterial wall overexpresses factors such as TIMP-1 and HO-1, which could participate in the resistance to cell migration and oxidative stress.

Angiotensin-Converting Enzyme Inhibitors↗

PubMed1999

Influence of elastin gene polymorphism on the elastin content of the aorta: A study in 2 strains of rat.

The elastin content in the thoracic aorta of male Brown-Norway (BN) rats is 31.4+/-1.2% (dry weight), whereas that of male LOU rats is 37.2+/-1.0%. A similar difference in the elastin content of the thoracic aorta is also observed in female animals. Furthermore, in the thoracic aorta of young, growing rats as well as in cultured aortic smooth muscle cells, the steady-state level of elastin mRNA is significantly lower in the BN than in the LOU strain. These results suggested that 1 or more genes control the elastin mRNA level and the elastin content in the aortas of BN and LOU rats. A possible relationship between a polymorphism in the elastin gene and the elastin content of the aorta was tested. For this purpose, the aortic elastin content was measured in F(1) and F(2) generations bred from LOU and BN rats and was compared with that of the F(0) (parental) generation. A polymorphic marker located in intron 25 of the elastin gene has been used to genotype the F(2) rats. The degree of genetic determination of aortic elastin content was estimated to be 73% in the F(2) cohort, but the elastin locus accounts for only 3. 9% of the total variance in aortic elastin content. Other genes are thus responsible for the major part of the observed interstrain difference by regulating the transcription of the gene, the stability of elastin mRNA, and/or posttranslational events.

Alleles↗

PubMed1998

The secondary structure and architecture of human elastin.

The presented work constitutes the first structural characterization of both insoluble human elastin and its solubilized form, kappa-elastin. Structural data were reached following the use of Fourier transform infrared, near infrared Fourier transform Raman and circular dichroism optical spectroscopic methods and their quantitative analysis permitted us to estimate approximately 10% alpha-helices, approximately 35% beta-strands and approximately 55% undefined conformations in the global secondary structure of insoluble human elastin in the solid state. Following the use of the LINK method, the probable local distribution of the secondary-structure elements along the sequence was determined and compared to that obtained for bovine elastin, the historical standard of elastin. This comparison led us to propose a globular architecture for the human elastomer and permitted us to delineate some elements of its structure-elasticity relationship.

Amino Acid Sequence↗

PubMed1998

Action of tropoelastin and synthetic elastin sequences on vascular tone and on free Ca2+ level in human vascular endothelial cells.

The elastic properties of extensible tissues such as arteries and skin are mainly due to the presence of elastic fibers whose major component is the extracellular matrix protein elastin. Pathophysiological degradation of this protein leads to the generation of elastin peptides that have been identified in the circulation in the ng/mL to microg/mL range. Similar concentrations of an elastin peptide preparation (kappa-elastin) were previously demonstrated to induce, among other biological actions, a dose- and endothelium-dependent vasorelaxation mediated by the elastin/laminin receptor and by endothelial NO production. To determine the elastin sequence(s) responsible for vasomotor activity and to learn more about possible signaling pathways, we have compared the action of different concentrations (10(-13) to 10(-7) mol/L) of recombinant human tropoelastin, eight synthetic elastin peptides, and a control peptide (VPVGGA) on both rat aortic ring tension and [Ca2+]i of cultured human umbilical vein endothelial cells. No vasoactivity could be detected for VPVGGA and for the elastin-related sequences VGVGVA, PGVGVA, and GVGVA. Tropoelastin, VGV, PGV, and VGVAPG were found to induce an endothelium- and dose-dependent vasorelaxation and to increase endothelial [Ca2+]i, whereas PVGV and VGVA produced these effects only at low concentration (10(-11) mol/L). A likely candidate for mediating the elastin peptide-related effects is the elastin/laminin receptor, since the presence of lactose strongly inhibited the vasoactivity associated with these compounds. Our results show that although the flanking amino acids modulate its activity, VGV seems to be the core sequence recognized by the elastin receptor.

Animals↗

PubMed1998

Localization of elastin mRNA and TGF-beta1 in rat aorta and caudal artery as a function of age.

Several in vitro studies have previously demonstrated that the addition of TGF-beta to aortic smooth muscle cells or skin fibroblasts stimulates elastin synthesis. It is not clear however whether, in vivo, TGF-beta participates in the regulation of elastin synthesis, especially in physiological conditions. The aim of our study was to explore the localization of elastin mRNA and TGF-beta1 in the rat thoracic aorta (an elastic artery) and caudal artery (a muscular artery). Elastin mRNA was localized by in situ hybridization and quantified using Northern blot analysis. TGF-beta1 was detected using immunohistochemistry. The study was carried out as a function of age (rats of 3, 10, 20, and 30 months). We observed that TGF-beta1 immunoreactivity is present predominantly, but not exclusively, at the sites of elastin synthesis as determined by elastin mRNA detection: in smooth muscle cells in the aorta and in endothelial cells in the caudal artery. The ability of exogenously added TGF-beta1 (0.001-10 ng/ml) to modulate the steady-state levels of elastin mRNA in primary cultures of endothelial cells, smooth muscle cells, and fibroblasts isolated from the thoracic aorta was also studied. At the highest concentration used, elastin mRNA levels increased 5-fold in endothelial cells and 11-fold in smooth muscle cells. The demonstration that TGF-beta1 immunoreactivity is present at the sites of elastin synthesis in the thoracic aorta and in the caudal artery and the observation that TGF-beta1 induces an increase in elastin mRNA levels in cultured endothelial cells and smooth muscle cells suggest that TGF-beta1 may be implicated, at least in part, in the physiological regulation of elastin gene expression.

Aging↗

PubMed1998

Epitope specificity of monoclonal and polyclonal antibodies to human elastin.

Polyclonal (pAb) and monoclonal (mAb) anti-human aorta elastin antibodies were reacted with a series of overlapping hexapeptides along the human tropoelastin sequence covering exons 2-7 and 23-36 from the N-terminus to the C-terminus, advancing 1 amino acid residue each time. ELISA indicated reactive epitopes. mAb A2.1 recognized sequences containing Ala-Lys, mAb G8.1, A7.1 and pAb, hydrophobic sequences. None of them reacted with the hexapeptide VGVAPG, or with desmosine or isodesmosine. pAb L85 reacted with a His-containing sequence coded in exon 26A. pAb kappaE(L), kappaE(S) and L85 reacted with the Cys-containing sequence of exon 36. A synthetic 14-residue peptide containing the three proximal tyrosines coded in exon 13 did not react with any of the antisera tested. It appears therefore that the most frequently recognized epitopes are hydrophobic sequences. One polyclonal antibody detected several isoforms of tropoelastin in the medium of cultured vascular smooth muscle cells. Monoclonal and polyclonal antibodies stained elastic fibers on tissue sections, suggesting that the epitopes recognized are available on the native fibers for reaction with the antibodies.

Amino Acid Sequence↗

PubMed1998

Biological effects of elastin peptides.

Atherosclerosis is clearly one of the most life-threatening diseases and a major cause of morbidity and mortality in industrialized countries. Typical arterial lesions contain both cells originating from the blood (monocytes/macrophages) and locally-recruited smooth muscle cells. The structure of the artery is profoundly disrupted. Degradation of arterial elastin fibers results in loss of elasticity, and several elastin peptides are released that can interact with various cells via an increasingly well-characterized elastin receptor. Elastin receptor-mediated reactions that are of obvious physiologic importance include vasodilating effects and induction of mesenchymal cell adhesion to elastin fibers. Other effects are potentially harmful, such as increased elastase production, free radical release, induction of LDL oxidation, and stimulation of endogenous cholesterol production. These deleterious effects become predominant during aging as a result of chronic exposure of the elastin receptor to circulating elastin peptides. This review describes the results of recent investigations into the biological effects of elastin peptides.

Animals↗

PubMed1998

Interactions of elastin fibers with fibroblasts a time-lapse cinemicrographic study.

Adhesion of cells to the extracellular matrix is mediated by structural glycoproteins such as fibronectin and laminin, and also elastonectin, whose role is to ensure binding of elastin fibers to cells. Interactions between elastin fibers and human skin fibroblasts cultured in a Rose chamber were investigated by using cinemicrography to observe elastin fiber attachment, detachment, and displacement over a five-day period. Elastin fiber displacement over the cell layer resulted in aggregation, which was measured using morphometry. The total number of isolated elastin fibers or aggregates decreased between 1 h and 8 h and remained stable thereafter. During the same time interval, significant decreases occurred in the numbers of isolated fibers and small aggregates (perimeter < 0.268 mm; surface area < 894 microns 2), whereas larger aggregates were formed. After 15 hours of interaction, none of the aggregates had a perimeter greater than 0.536 mm, consistent with an increase in aggregate compacting. These data demonstrate that elastin-cell interactions do not occur at random. These interactions may play a pivotal role in morphogenesis and in maintaining the integrity of elastic tissues such as the arterial wall, lungs, and skin.

Cells, Cultured↗

PubMed1997

Effect of lithium on superoxide production and intracellular free calcium mobilization in elastin peptide (kappa-elastin) and FMLP stimulated human PMNS. Effect of age.

The effect of lithium pretreatment on superoxide anion production and intracellular free calcium levels was investigated in polymorphonuclear leukocytes (PMN) from middle-aged and old individuals after stimulation by elastin peptides or FMLP. K-elastin (KE) significantly stimulated the production of superoxide anion by PMNs from middle-aged subjects, while this stimulation decreased with age and was absent in PMNs of elderly arteriosclerotic patients. Li pretreatment slightly increased this stimulating effect of KE in PMNs from middle-aged subjects and elderly arteriosclerotic patients, while slightly decreased in healthy elderly subjects. Moreover, Li was able to increase superoxide anion production even in the absence of KE, but this effect decreased also in PMNs of healthy and arteriosclerotic elderly patients. FMLP significantly increased superoxide anion production in all age-groups, but this effect was further amplified by Li only in PMNs of middle-aged subjects. In aged individuals Li pretreatment slightly decreased the effect of FMLP and had no effect in arteriosclerotic patients. Ca-mobilization induced by KE was inhibited by Li pretreatement in each age group. This inhibition by Li was much weaker in FMLP-stimulated PMNs. Li pretreatment did however modify the shape of the Ca-transient curves in FMLP stimulated leukocytes suggesting a qualitative modification of ion channel regulation. No such shape change of Ca-transient curves was observed after KE stimulation of Li pretreated PMNs. It appears that the regulation of these two receptors is differently affected by Li treatment.

Adult↗