Zymographic analysis of latent and activated forms of matrix metalloproteinase-2 and -9 in synovial fluid: correlation to polymorphonuclear leukocyte infiltration and in response to infection

Makowski, Gregory S.; Ramsby, Melinda L.

doi:10.1016/s0009-8981(03)00015-9

Cited by 45 publications

(53 citation statements)

References 21 publications

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“…Although HMW MMP-9 enzymatic activity has been recognized in OA SF in earlier studies (30), the 125-kd MMP activity (24,25) has not previously been characterized. Since the apparent size of the HMW synovial fluid MMP activity noted in these and other studies was similar to the size reported for the NGAL-MMP-9 complex purified from human neutrophils (31) and also shown to be present in breast cancer patients (15), we investigated the possibility that this HMW (ϳ125 kd) MMP activity in OA SF might represent an NGAL-MMP-9 complex.…”

Section: Resultsmentioning

confidence: 99%

“…Furthermore, results of gelatin zymography revealed that SF samples obtained from patients with OA were rich in 72-kd, 92-kd, and 125-kd MMP activity (Figure 1). MMP activity at 92 kd and 72 kd has previously been determined to represent MMP-9 and MMP-2, respectively (9,24,31). All of the SF samples also contained the partially proteolyzed forms of MMP-9 (Ͻ92 kd), indicating the presence of active MMP-9 in the OA SF samples.…”

Section: Resultsmentioning

confidence: 99%

“…Previous studies have documented the presence of HMW MMP activity in the SF of arthritis patients (9,24,25), but these HMW gelatinases were not characterized. The NGAL-MMP-9 complex detected in OA SF may be from joint-infiltrating cells, or possibly from cytokine-stimulated chondrocytes and synovial cells.…”

Section: Discussionmentioning

confidence: 99%

“…26), but this high molecular weight (HMW) gelatinase activity has not been characterized. Since NGAL has been shown to bind with MMP-9 (15), the purpose of this study was to examine the expression of NGAL in OA joints, using a combination of molecular, immunologic, and biochemical approaches to determine whether NGAL forms a complex with MMP-9, giving rise to the HMW gelatinase activity in OA SF (24).…”

mentioning

confidence: 99%

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Neutrophil gelatinase–associated lipocalin is expressed in osteoarthritis and forms a complex with matrix metalloproteinase 9

Gupta¹,

Shukla²,

Cowland³

et al. 2007

Arthritis & Rheumatism

121

119

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Objective. Expression of matrix metalloproteinase 9 (MMP-9) is up-regulated in osteoarthritis (OA) and usually presents as multiple bands when synovial fluid (SF) from OA patients is analyzed by zymography. Among these bands is an ϳ125-130-kd band for high molecular weight (HMW) gelatinase, which has not been characterized. This study was undertaken to characterize the HMW MMP activity in OA SF.Methods. MMP activity in OA SF was determined by gelatin zymography. Recombinant MMPs were used to identify MMP activity on the zymogram. Western immunoblotting, immunoprecipitation, and immunodepletion analyses were performed using antibodies specific for human MMP-9 and human neutrophil gelatinase-associated lipocalin (NGAL). Human cartilage matrix degradation was determined by dimethylmethylene blue assay.Results. Zymographic analysis showed that the HMW gelatinase in OA SF comigrated with a purified NGAL-MMP-9 complex. Results of Western immunoblotting showed that the HMW gelatinase was also recognized by antibodies specific for human NGAL or human MMP-9. These same antibodies also immunoprecipitated the HMW gelatinase activity from OA SF. The NGAL-MMP-9 complex was reconstituted in vitro in gelatinase buffer. In the presence of NGAL, MMP-9 activity was stabilized; in the absence of NGAL, rapid loss of MMP-9 activity occurred. MMP-9-mediated release of cartilage matrix proteoglycans was significantly higher in the presence of NGAL (P < 0.05).Conclusion. Our findings demonstrate that the HMW gelatinase activity in OA SF represents a complex of NGAL and MMP-9. The ability of NGAL to protect MMP-9 activity is relevant to cartilage matrix degradation in OA and may represent an important mechanism by which NGAL may contribute to the loss of cartilage matrix proteins in OA.The developed world's aging population has experienced a dramatic increase in the incidence of joint dysfunction and osteoarthritis (OA), leading to a compromised quality of life. Aging, biomechanical stress, and oxidative stress, in addition to genetic factors and trauma, all appear to be associated with degenerative and progressive cartilage and bone changes in OA, particularly in the weight-bearing joints. Although OA is not considered an inflammatory disease by traditional standards, elevated levels of proinflammatory cytokines, such as interleukin-1␤ (IL-1␤) and tumor necrosis factor ␣, have been observed in OA synovial fluid (SF), supporting the notion that there is an inflammatory component associated with the pathogenesis of OA (for review, see refs. 1-3).The only cell type present in articular cartilage is the chondrocyte, which is embedded within an extracellular matrix (ECM) primarily composed of type II collagen and aggrecan. Chondrocytes arise from a mesenchymal progenitor cell and are responsible for the biosynthesis of ECM proteins and their transport into the space occupied by the ECM. Chondrocytes play an important role in cartilage homeostasis by maintaining the proper balance between anabolic pathways, which Supported in part by the NIH...

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

See 2 more Smart Citations

Neutrophil gelatinase–associated lipocalin is expressed in osteoarthritis and forms a complex with matrix metalloproteinase 9

Gupta¹,

Shukla²,

Cowland³

et al. 2007

Arthritis & Rheumatism

121

119

View full text Add to dashboard Cite

show abstract

“…Dried gels were scanned with Epson Stylus TX130 (Seiko Epson Corporation, Nagano, Japan) and band volumes were determined using software Image J, version 1.45 (National Institutes of Health, Bethesda, MD, USA). Human plasma prepared from clinically healthy volunteers was used as a positive control as reference of the latent forms of MMP-2 and MMP-9, and a 125-kDa NGAL-proMMP-9 complex (Kolkenbrock et al, 1996;Makowski and Ramsby, 2003).…”

Section: Gelatin Zymographymentioning

confidence: 99%

Fingerprinting of gelatinase subtypes for different topographic regions on non-retaining placenta of Holstein cows

Piamya

Tiantong

Chen

et al. 2015

Animal

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The contribution of matrix metalloproteinases (MMP) to timely discharge of the placenta from bovine uterus at parturition is yet inconclusive, partly because of the presence of multiple MMP forms in situ. In the current study, the expression of different gelatinase subtypes on non-retaining placentas of Holstein cows was fingerprinted by using gelatin zymography. Different topographic regions on the placenta were measured separately, including the placentome-like structure and the fetal and maternal sides of interplacentomal placenta, all sampled from the central and peripheral areas of the placenta, respectively. The spontaneously ruptured umbilical cords were cross-sectioned as fetus end, middle and placenta end also for separate measurement. Body fluids including blood samples from the parturient cows, their neonatal calves and umbilical cord, as well as fetal fluids and the first colostrum were measured concomitantly. Results showed multiple forms of gelatinases subtypes in the placenta tissues and body fluids, including neutrophil gelatinase-associated lipocalin (NGAL)-MMP-9 complex, both the latent and active forms of MMP-2 and MMP-9; of them, the latent forms were much more abundantly and frequently expressed than the active forms. NGAL-MMP-9 complex was more prevalently present in the body fluids than in the placenta tissues. No distinguishable pattern of the expression of any gelatinase subtype was observed among the placentome-like structure, interplacentomal placenta and umbilical cord, or between fetal and maternal sides. Nonetheless, for interplacentomal placenta, proMMP-9 expression was higher in the central than in the peripheral area. In addition, proMMP-2 expression was higher in the rupture end (fetus end) than the placenta end of the umbilical cord. In conclusion, the current validated gelatin zymography detected a gradient proMMP-9 expression on the non-retaining placenta of cows in reverse to the proximity to the umbilical insertion point, and a gradient proMMP-2 expression on a section of the umbilical cord in reverse to the proximity to the rupture site, suggesting roles played by gelatinases in normal discharge of the placenta at term.

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Left ventricular diastolic function in type 2 diabetes mellitus is associated with myocardial triglyceride content but not with impaired myocardial perfusion reserve

Korosoglou

Humpert

Ahrens

et al. 2011

Magnetic Resonance Imaging

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Purpose: To study myocardial perfusion reserve and myocellular metabolic alterations indicated by triglyceride content as possible causes of diastolic dysfunction in patients with type 2 diabetes mellitus, preserved systolic function, and without clinically evident coronary artery disease. Materials and Methods:Patients with type 2 diabetes mellitus (n ¼ 42) underwent cardiac magnetic resonance (CMR) for quantification of 1) myocardial contractility by strain-encoded MR (SENC); 2) myocardial triglyceride content by proton magnetic resonance spectroscopy ( 1 H-MRS); and 3) myocardial perfusion reserve during pharmacologic hyperemia. Age-matched healthy volunteers (n ¼ 16) also underwent CMR to acquire normal values for myocardial strain and perfusion reserve.Results: Stress CMR procedures were successfully performed in all subjects, and no regional inducible perfusion defects were observed in type 2 diabetes mellitus patients. Diastolic strain rate and myocardial perfusion reserve were significantly impaired in patients with type 2 diabetes mellitus compared to control subjects (P < 0.001 for both). Interestingly, impaired diastolic function in type 2 diabetes mellitus was not associated with impaired myocardial perfusion reserve (r ¼ 0.12, P ¼ NS). Conversely a significant association was observed between diastolic dysfunction and myocardial triglyceride content (r ¼ À0.71, P < 0.001), which proved to be independent of age, gender, diabetes duration, blood pressure, and fasting blood glucose. Conclusion:Myocardial steatosis may represent an early marker of diabetic heart disease, triggering subclinical myocardial dysfunction irrespective of myocardial perfusion reserve.

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Zymographic analysis of latent and activated forms of matrix metalloproteinase-2 and -9 in synovial fluid: correlation to polymorphonuclear leukocyte infiltration and in response to infection

Cited by 45 publications

References 21 publications

Neutrophil gelatinase–associated lipocalin is expressed in osteoarthritis and forms a complex with matrix metalloproteinase 9

Neutrophil gelatinase–associated lipocalin is expressed in osteoarthritis and forms a complex with matrix metalloproteinase 9

Fingerprinting of gelatinase subtypes for different topographic regions on non-retaining placenta of Holstein cows

Left ventricular diastolic function in type 2 diabetes mellitus is associated with myocardial triglyceride content but not with impaired myocardial perfusion reserve

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