Search PubMed⌕ Search

Biomedical subjects

M D Tilson

Publications and source records attributed to M D Tilson.

At least 55 records · Page 3Linked to original sources
PubMed1994

Matrix metalloproteinases in abdominal aortic aneurysm: characterization, purification, and their possible sources.

One of the most consistent observations in abdominal aortic aneurysm (AAA) disease is the disorganization and disruption of elastin and other matrix components of the aortic wall. The enzymatic basis for the biochemical features of AAA has been investigated beginning with the demonstration on substrate gel enzymography of a typical "profile" of proteinase activities in AAA tissue extracts which degrade gelatin, casein and elastin. A recombinant TIMP-1 affinity column was developed and three of the elastolytic/caseinolytic activities with approximate molecular weights of approximately 80 kDa, approximately 50 kDa and approximately 32 kDa were partially purified from these extracts. Affinity for rTIMP-1 suggests that these enzymes are members of the matrix metalloproteinase (MMP) family. High molecular weight forms of two MMPs, collagenase (MMP-1) and stromelysin-1 (MMP-3), were also isolated from the AAA tissue on this column; active forms of MMP-1 could be demonstrated by immunoblotting techniques in this preparation under reducing conditions. Infiltrating inflammatory cells are known sources of these proteolytic activities; analysis of these cell populations in the aneurysmal aortic wall using fluorescence-activated cell counting revealed a fifty-fold increase in macrophages (a well-known source of matrix-degrading enzymes) as well as a significant increase in lymphocytes.

Aortic Aneurysm, Abdominal↗
PubMed1994

Cytokines that activate proteolysis are increased in abdominal aortic aneurysms.

BACKGROUND: Abdominal aortic aneurysm (AAA) disease is characterized by an increase in proteolysis and loss of matrix components. The cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta), products of activated macrophages and T cells, are known to increase the production of matrix-degrading enzymes in some pathological states. METHODS AND RESULTS: Seven AAA and five control aortic tissue extracts were assayed for TNF-alpha and IL-1 beta with ELISA. TNF-alpha was elevated significantly in AAA extracts compared with controls (86 +/- 34 pg/mg of total protein versus 1 +/- 1 pg/mg of total protein; P < .001). IL-1 beta concentration also was significantly increased in the AAA specimens (48 +/- 14 pg/mg of total protein versus 12 +/- 5 pg/mg of total protein; P < .05). Immunoblotting demonstrated secreted forms of TNF-alpha in the AAA extracts, and possible membrane-bound forms were observed when the tissues were detergent-extracted. Known forms of IL-1 beta also were observed on immunoblots of AAA tissue extracts. CONCLUSIONS: The presence of TNF-alpha and IL-1 beta in AAA tissue underscores the importance of the infiltrating inflammatory cells present in the media and adventitia of aneurysmal aortic wall and further implicates an inflammatory process in the pathogenesis of AAA.

Aorta, Abdominal↗
PubMed1994

Possible key role for plasmin in the pathogenesis of abdominal aortic aneurysms.

BACKGROUND: Activation of proteolysis is characteristic of abdominal aortic aneurysm (AAA) disease, and by substrate gel enzymography with casein the most conspicuous proteinase of AAA wall has a molecular weight of approximately 80 kd. This activity has been resolved into separate metalloproteinase and serine proteinase (SP) components, by binding out the metalloproteinase by affinity to tissue inhibitor of metalloproteinases. Because plasmin plays a key role in activating members of the metalloproteinase family, the following experiments were done to test the hypothesis that the unknown SP is plasmin. METHODS: Immunoblots, immunoprecipitations, and immunohistochemistry were performed by conventional methods with a specific polyclonal antibody to plasminogen. RESULTS: Immunoblot analysis of extracts from AAA specimens (n = 7) revealed dense bands corresponding to known molecular forms of plasmin. Only trace amounts were detected in control aortic extracts (n = 4). When samples were equalized for total protein, the mean amount of immunoreactive material in the 80 kd band (in densitometric units) was 216 +/- 44 (SEM) for AAAs versus 27 +/- 15 (SEM) for controls (p < 0.01). The residual 80 kd SP activity on casein substrate gel enzymography was quenched by immunoprecipitation with the specific antibody. Immunohistochemistry revealed strong reactivity of the AAA wall. CONCLUSIONS: Because plasmin plays a key role in the cascade for activation of the matrix metalloproteinase (including collagenase and the metalloelastases), the present results suggest that plasmin may be important in the sequence of events leading to the destruction of aortic matrix in AAA.

Aortic Aneurysm, Abdominal↗
PubMed1994

Analysis of elastin cross-linking and the connective tissue matrix of abdominal aortic aneurysms.

BACKGROUND: Studies of the connective tissue matrix of abdominal aortic aneurysms (AAAs) have yielded conflicting results, and the glycosaminoglycan content has not been previously reported. The present work was done to evaluate the matrix components of AAAs, including the cross-link content of the residual elastin. METHODS: Aortic specimens from AAAs and controls were sequentially extracted with salt, Brij, and urea; and the residual pellets were the subject of further studies. Elastin was purified by hot alkali treatment; other matrix components were determined by conventional methods. RESULTS: Elastin content of the purified material was reduced in AAA. The cross-link content, desmosine+isodesmosine, was also reduced in AAA as a ratio to insoluble matrix dry weight. However, the cross-link content as a ratio to valine in the purified elastin was normal. The amino acid profiles of representative AAA and controls elastin preparations were similar to that of reference elastin. The amino acid content of the insoluble matrix of AAA revealed a significant reduction of protein (controls = 820 +/- 40 micrograms/mg versus AAA = 700 +/- 20 micrograms/mg, p < 0.05); the collagen content was unaltered. The content of glycosaminoglycan in AAA was noted to be significantly reduced (controls = 33.5 +/- 3.4 micrograms/mg versus AAA = 17.1 +/- 2.0 micrograms/mg, p < 0.05). CONCLUSIONS: The data do not support the hypothesis of a primary cross-link deficiency in elastin of AAA; but the reduced contents of protein and glycosaminoglycans in AAA suggests basic biochemical alterations in the diseased aorta that warrant further investigation.

Amino Acids↗
PubMed1993

Demonstration of interstitial collagenase in abdominal aortic aneurysm disease.

This study was performed to evaluate the presence of interstitial collagenase, now known as matrix metalloproteinase-1 (MMP-1), in specimens of abdominal aortic aneurysms (AAA). Eight AAA and four control infrarenal aortas were evaluated. After homogenization and extraction of soluble proteins, immunoblots of the extracts equalized for protein content were performed with a specific antibody to MMP-1. Under native conditions, immunoreactive material was distributed between M(r) 27 kDa to > 106 kDa. When the extracts were reduced and denatured, immunoreactive bands were detected in AAA at the expected M(r)'s of the secreted isoforms (57 and 52 kDa), whereas control aortic extracts had low levels of detectable immunoreactive material. Only extracts from AAA demonstrated significant immunoreactivity to the lower M(r) isoforms (22, 25, and 27 kDa), which correspond to reported cleavage products of MMP-1. Preliminary immunofluorescent studies of AAA localized MMP-1 to cells present in the adventitia of AAA. These findings will help to resolve disagreement in the recent literature regarding the presence of collagenolytic activity in AAA disease.

Aortic Aneurysm, Abdominal↗
PubMed1993

Analysis of the connective tissue matrix and proteolytic activity of primary varicose veins.

PURPOSE: Valvular incompetence and venous wall abnormalities have been suggested as primary etiologic factors responsible for the development of varicose veins. This study was conducted to evaluate the connective tissue constituents of greater saphenous varicosities. Proteolytic activity, a factor that can lead to matrix degradation and cause weakening and dilation of the venous wall, was also assessed. METHODS: The collagen and elastin contents of 16 nonthrombophlebitic greater saphenous varicose veins (VV) and seven normal greater saphenous veins (NV) were quantified. In addition, four duplex scanning-confirmed competent segments of greater saphenous veins (i.e., potential varicose veins [PV]) affected by varicosis at alternate sites were analyzed. Proteolytic activity was determined by zymography and radiolabeled substrate assay. RESULTS: The content of collagen was significantly increased in the VV and PV compared with NV (VV = 189 +/- 7 mg/gm, PV = 189 +/- 9 mg/gm vs NV = 144 +/- 10 mg/gm, p < 0.05). Conversely, the elastin content in the VV and PV was significantly reduced (VV = 53 +/- 3 mg/gm, PV = 50 +/- 4 mg/gm vs NV = 74 +/- 4 mg/gm, p < 0.05). The collagen to elastin ratio demonstrated an alteration in VV and PV compared with NV (VV = 3.7 +/- 0.3, PV = 3.9 +/- 0.4 vs NV = 2.0 +/- 0.2, p < 0.05). Casein and gelatin zymography did not demonstrate significant qualitative differences in the enzymatic activities among the three groups. Quantitative analysis of the elastase activity in the venous tissues was similarly not appreciably altered (VV = 5.1 +/- 0.2 U/gm, PV = 5.3 +/- 0.2 U/gm vs NV = 5.7 +/- 0.3 U/gm). CONCLUSION: A significant increase in the collagen content and a significant reduction in the elastin content of VV were demonstrated. The net increase in the collagen/elastin ratio is indicative of an imbalance in the connective tissue matrix. The biochemical profile of PV was similar to VV and significantly different from NV. These preliminary data support the presence of connective tissue abnormalities before valvular insufficiency. In addition, the absence of an increase in the proteolytic activity excludes enzymatic matrix degradation as an essential component in the formation of venous varicosities.

Collagen↗
PubMed1993

Expression and sequence of the gene for tissue inhibitor of metalloproteinases in patients with abdominal aortic aneurysms.

PURPOSE: The tissue inhibitors of metalloproteinases (TIMPs) are major inhibitors of several enzymes that are destructive to connective tissue, and TIMP-1 has been reported to be deficient in the wall of abdominal aortic aneurysms. This deficiency could represent failure of expression resulting from either local tissue conditions or mutation in the primary structure of the gene (or one of its regulatory elements). METHODS: Southern blotting techniques were used to examine the possibility of global deletions or inserts in the gene (14 patients); Northern blot techniques were performed to examine the expression of the mRNAs in cultured fibroblasts under basal conditions (six patients); and sequence analysis of the cDNA derived from fibroblast mRNAs was done after amplification by polymerase chain reaction (six patients). RESULTS: The Southern blots revealed a normal distribution of the known alleles of the gene without unique restriction-length polymorphisms. Fibroblast expression of TIMP mRNA was normal under basal conditions. Sequence analysis of the cDNAs revealed an identical-point polymorphism in two of the six patients (a single base pair substitution of C-->T at the third position in codon 101), but the amino acid was conserved. CONCLUSION: The studies reported here do not support the hypothesis that deficiency of TIMP-1 in specimens of aorta of patients with abdominal aortic aneurysms results from a primary genetic defect.

Aortic Aneurysm, Abdominal↗
PubMed1992

Characterization of an elastase from aneurysmal aorta which degrades intact aortic elastin.

Accumulating evidence suggests that abdominal aortic aneurysms (AAA) are due to a pathologic process which results in the destruction of aortic elastin and other matrix components. In this study, protein extractions were performed on both aneurysmal and normal aorta. Extracts were applied to frozen section of normal aorta either alone or in combination with 10 mM ethylenediaminetetraacetic acid, recombinant tissue inhibitor of metalloproteases, 10 mM zinc, and 5 mM phenylmethylsulfonyl fluoride, under conditions where calcium was removed from the buffer. After incubation, the sections were stained for elastin and evaluated by computerized morphometry. Aneurysm extracts, only in the presence of calcium, showed significant elastolytic activity characterized by destruction of intact elastic lamellae that was inhibited by ethylenediaminetetraacetic acid, the recombinant metalloprotease inhibitor, and zinc. Phenylmethylsulfonyl fluoride showed no inhibitory activity. Healthy aortic extract showed no elastolytic activity. This inhibitory profile is consistent with a metalloenzyme. We conclude that aneurysmal aorta contains elastolytic activity that is secondary to a metalloenzyme which is not present in normal aorta. This activity may play a role in the destruction of the elastin matrix that is seen in AAA's.

Aorta, Abdominal↗
PubMed1991

The role of inflammation in nonspecific abdominal aortic aneurysm disease.

The predominant pathologic feature of abdominal aortic aneurysm is elastin destruction, and elastin destruction may be mediated by inflammation. In this investigation serial sections of abdominal aortic aneurysm specimens were selectively stained to study the relationship between inflammation and elastin degradation. In addition, soluble aortic extracts were examined for the presence of immunoglobulins. An inflammatory infiltrate was present in 8 of 10 of the abdominal aortic aneurysm specimens examined. The infiltrate was mononuclear, commonly located at the junction of the media and adventitia; it did not codistribute with loss of elastin. The presence of an inflammatory component in abdominal aortic aneurysm was associated with a large amount of immunoglobulin in soluble extracts from aneurysmal tissue compared to atherosclerotic and normal control extracts. This study further characterizes the microscopic pathology of abdominal aortic aneurysm and describes the presence of immunoglobulin in soluble tissue extracts. In addition, the possible role of inflammation in abdominal aortic aneurysm as it relates to protease expression is detailed.

Aorta, Abdominal↗
PubMed1991

Decreased tissue inhibitor of metalloproteinases (TIMP) in abdominal aortic aneurysm tissue: a preliminary report.

Important concepts underlying the pathophysiology of abdominal aortic aneurysm (AAA) disease include a genetic predisposition, male predominance, and increased proteolysis. Proteolytic activity is carefully controlled by an abundance of protease inhibitors and the increased proteolysis may reflect a decrease in inhibitory activity. The recent assignment for the major tissue inhibitor of metalloproteinases (TIMP) to the X chromosome provides a potential link between the male predominance and increased proteolysis noted in AAA. The purpose of this investigation was to measure the amount of TIMP in normal and diseased aorta. A polyclonal antibody to recombinant human TIMP was produced in rabbit and used to establish an immunoassay. Immunoreactive TIMP in normal and diseased aorta was then measured. There was more TIMP in the matrix-associated fraction than in the soluble fraction. There was significantly less immunoreactive TIMP in aortic extracts from AAA than from control as measured both by Western blot and radioimmunoassay. These results suggest that a diminished amount of TIMP in the aortic matrix in AAA patients may contribute to the increased proteolysis observed in AAA.

Aorta, Abdominal↗
PubMed1991

Suggested standards for reporting on arterial aneurysms. Subcommittee on Reporting Standards for Arterial Aneurysms, Ad Hoc Committee on Reporting Standards, Society for Vascular Surgery and North American Chapter, International Society for Cardiovascular Surgery.

The literature on arterial aneurysms is subject to potential misinterpretation because of inconsistencies in reporting standards. The joint councils of the Society for Vascular Surgery and the North American Chapter of the International Society for Cardiovascular Surgery appointed an ad hoc committee to address this issue. This communication, prepared in response to the need for standardized reporting, defines and classifies arterial aneurysms and recommends standards for describing the causes, manifestations, treatment, and outcome criteria that are important when publishing data on aneurysmal disease.

Aneurysm↗
PubMed1990

Tissue inhibitor of metalloproteases (TIMP) is matrix associated in aortic tissue: report of a radioimmunoassay.

Tissue inhibitor of metalloproteinases (TIMP) is the major inhibitor of collagenase, gelatinase, proteoglycanase, stromelysin, and metalloelastases. An imbalance between proteases and inhibitors has been implicated in numerous disease processes including tumor invasion, rheumatoid arthritis, emphysema, and aortic aneurysm disease. The purpose of this investigation was to develop a polyclonal antibody to recombinant TIMP and establish an immunoassay to measure immunoreactive protein in normal and diseased tissues. A polyclonal antibody was produced in rabbit against recombinant human TIMP which was characterized and used to establish a radioimmunoassay. The assay was used to measure immunoreactive protein in fibroblast conditioned medium, human serum, and aortic extracts. There was more immunoreactive TIMP in matrix associated urea extracts than soluble salt extracts from human aorta, suggesting that TIMP is matrix associated. The sensitivity of the assay enables the specific measurement of this inhibitor in serum, fibroblast culture medium, and tissue extracts.

Aorta, Abdominal↗
PubMed1990

Hydrocortisone rapidly induces aortic rupture in a genetically susceptible mouse.

The Blotchy mouse has an X-linked trait that leads to aortic aneurysms and subsequent fatal rupture in nearly all affected male mice. Heterozygous female mice occasionally develop aneurysms, but they rarely rupture. Ten heterozygous female mice received 0.45 mg/mL of hydrocortisone acetate in drinking water. Within 2 weeks, 9 of 10 mice were dead (6 with proved aortic rupture, 3 with presumed rupture). The 10th mouse was documented to have an aortic aneurysm. A dose-response curve was generated. Hydrocortisone's effect was shown to be dose-dependent. In another experiment, normal female mice received 0.10 mg/mL of hydrocortisone acetate for 14 days. Two mice developed aneurysms, and the others developed aortic ectasia. These experiments establish the role of hydrocortisone in the induction of aortic rupture in a mouse with genetic susceptibility and the induction of aneurysms and ectasia in normal mice.

Animals↗