MMP-9 gene deletion mitigates microvascular loss in a model of ischemic acute kidney injury

Microvascular rarefaction following an episode of acute kidney injury (AKI) is associated with renal hypoxia and progression toward chronic kidney disease. The mechanisms contributing to microvascular rarefaction are not well-understood, although disruption in local angioregulatory substances is thought to contribute. Matrix metalloproteinase (MMP)-9 is an endopeptidase important in modifying the extracellular matrix (ECM) and remodeling the vasculature. We examined the role of MMP-9 gene deletion on microvascular rarefaction in a rodent model of ischemic AKI. MMP-9-null mice and background control (FVB/NJ) mice were subjected to bilateral renal artery clamping for 20 min followed by reperfusion for 14, 28, or 56 days. Serum creatinine level in MMP-9-null mice 24 h after injury [1.4 (SD 0.8) mg/dl] was not significantly different from FVB/NJ mice [1.5 (SD 0.6) mg/dl]. Four weeks after ischemic injury, FVB/NJ mice demonstrated a 30-40% loss of microvascular density compared with sham-operated (SO) mice. In contrast, microvascular density was not significantly different in the MMP-9-null mice at this time following injury compared with SO mice. FVB/NJ mice had a 50% decrease in tissue vascular endothelial growth factor (VEGF) 2 wk after ischemic insult compared with SO mice. A significant difference in VEGF was not observed in MMP-9-null mice compared with SO mice. There was no significant difference in the liberation of angioinhibitory fragments from the ECM between MMP-9-null mice and FVB/NJ mice following ischemic injury. In conclusion, MMP-9 deletion stabilizes microvascular density following ischemic AKI in part by preserving tissue VEGF levels.     

Targeted deletion of MMP-9 attenuates myocardial contractile dysfunction in heart failure

Chronic volume overload (VO) on the left ventricle (LV) augments redox stress and activates matrix metalloproteinase (MMP) which causes the endocardial endothelial-myocyte (EM) disconnection leading to myocardial contractile dysfunction. VO-induced MMP-9 activation impairs cardiac functions, in part by endothelial endocardial apoptosis, but the role of MMP-9 on EM functions remains obscure. We conjecture that chronic VO activates MMP-9 and causes EM uncoupling. Arteriovenous fistula (AVF) was created in genetically identical wild type (WT) mice (FVB/NJ) and MMP-9 knockout mice (MMP-9KO, FVB.Cg-MMP9(tm1Tvu)/J). Sham-operated mice were used as controls. Before experimentation the phenotype analysis of MMP-9KO mice was carried out. In-gel-gelatin zymography for MMP-9 activation was performed on LV homogenates. The EM functions were determined on LV rings using tissue myobath. We report a decrease in MMP-9 activity in left ventricular myocardial extracts in MMP-9 deficient mice after AVF. The responses to drugs affecting cardiac functions (acetylcholine (Ach), nitroprusside and bradykinin) were attenuated in AVF mice suggesting the impairment of EM coupling. Interestingly, the EM functions were restored in the MMP-9 deficient mice after AVF. We suggest a direct cause-and-effect relationship between MMP-9 activation and EM uncoupling in LV myocardium after chronic VO and the possible involvement of MMP-9 in myocardial contractile performance.     

Mice lacking the gene encoding for MMP-9 and resistance artery reactivity

OBJECTIVES: To define the link between the deletion of gene encoding for metalloproteinase 9 and resistance artery reactivity, we studied in vitro smooth muscle and endothelial cell function in response to pressure, shear stress, and pharmacological agents. BACKGROUND: Matrix metalloproteinases play a crucial role in the regulation of extracellular matrix turnover and structural artery wall remodeling. METHODS: Resistance arteries were isolated from mice lacking gene encoding for MMP-9 (KO) and their control (WT). Hemodynamic, pharmacology approaches, and Western blot analysis were used in this study. RESULTS: The measurement of blood pressure in vivo was similar in KO and WT mice. Pressure-induced myogenic tone, contractions to angiotensin-II and phenylephrine were similar in both groups. The inhibition of MMP2/9 ((2R)-2-[(4-biphenylylsulfonyl) amino]-3-phenylpropionic acid) significantly decreased myogenic tone in WT and had no effect in KO mice. Relaxation endothelium-dependent (flow-induced- dilation 41.3+/-0.6 vs. 21+/-1.6 at 10 microl/min in KO and WT mice, respectively, P<0.05) and eNOS expression were increased in KO compared to WT mice. The inhibition of eNOS with L-NAME significantly decreased endothelium response to shear stress, which was more pronounced in KO mice resistance arteries (-26.83+/-2.5 vs. -15.84+/-2.3 at 10 microl/min in KO and WT, respectively, P<0.05). However, the relaxation to exogenous nitric oxide-donor was similar in both groups. CONCLUSION: Our study provides evidence of a selective effect of MMP-9 on endothelium function. Thus, MMP-9 gene deletion specifically increased resistance artery dilation endothelium-dependent and eNOS expression. Based on our results, MMP-9 could be a potential therapeutic target in cardiovascular disease associated with resistance arteries dysfunction.     

Matrix metalloproteinase inhibitors attenuate endotoxemia induced cardiac dysfunction: A potential role for MMP-9

Enhanced cardiac generation of peroxynitrite contributes to septic cardiomyopathy. Since matrix metalloproteinases (MMPs) are activated in vitro by peroxynitrite, we hypothezised that MMPs may contribute to cardiac mechanical dysfunction in sepsis. Rats were injected (i.p.) with either lipopolysaccharide (LPS, 4mg/kg) or vehicle. MMP inhibitors, either Ro 31-9790 (20 mg/kg), doxycycline (4mg/kg), or vehicle were administered i.p. 30 min after LPS. At 6 h, when the symptoms of endotoxemia peak, hearts were excised and perfused as working hearts with Krebs-Henseleit buffer at 37°C. Cardiac work (cardiac output x peak systolic pressure product) was measured. Perfusate and ventricle samples were analyzed by gelatin zymography to quantify MMP activity.Cardiac function was significantly depressed in LPS-treated rats compared to control rats (control: 55 ± 4, LPS: 26 ± 6 mmHg*mL*min^–1). LPS also caused a loss of 72 kDa MMP-2 activity in the ventricles and the perfusate. Although MMP-9 activity was not detected in the ventricles, LPS resulted in an increase in perfusate 92 kDa MMP-9 activity. The MMP inhibitors significantly improved cardiac function of LPS-treated rats (Ro31-9790: 38 ± 3, doxycycline: 51 ± 3 mmHg*mL*min^–1), had no effect on the loss of MMP-2 activity, and significantly reduced the MMP-9 activity in the perfusate. These results demonstrate, for the first time, that LPS induced cardiac dysfunction is associated with a loss in ventricular MMP-2 activity and the release of MMP-9 from the heart. MMP inhibitors can significantly preserve cardiac mechanical function during septic shock.     

MMP-9 gene variants increase the risk for non-atopic asthma in children

Background

Atopic and non-atopic wheezing may be caused by different etiologies: while eosinophils are more important in atopic asthmatic wheezers, neutrophils are predominantly found in BAL samples of young children with wheezing. Both neutrophils as well as eosinophils may secrete matrix metalloproteinase 9 (MMP-9). Considering that MMP-9 plays an important role in airway wall thickening and airway inflammation, it may influence the development of obstructive airway phenotypes in children. In the present study we investigated whether genetic variations in MMP-9 influence the development of different forms of childhood asthma.

Methods

Genotyping of four HapMap derived tagging SNPs in the MMP-9 gene was performed using MALDI-TOF MS in three cross sectional study populations of German children (age 9-11; N = 4,264) phenotyped for asthma and atopic diseases according to ISAAC standard procedures. Effects of single SNPs and haplotypes were studied using SAS 9.1.3 and Haploview.

Results

SNP rs2664538 significantly increased the risk for non-atopic wheezing (OR 2.12, 95%CI 1.40-3.21, p < 0.001) and non-atopic asthma (OR 1.66, 95%CI 1.12-2.46, p = 0.011). Furthermore, the minor allele of rs3918241 may be associated with decreased expiratory flow measurements in non-atopic children. No significant effects on the development of atopy or total serum IgE levels were observed.

Conclusions

Our results have shown that homozygocity for MMP-9 variants increase the risk to develop non-atopic forms of asthma and wheezing, which may be explained by a functional role of MMP-9 in airway remodeling. These results suggest that different wheezing disorders in childhood are affected differently by genetic alterations.     

Mitochondrial dysfunction in hearing loss

Mitochondrial pathology plays an important role in both inherited and acquired hearing loss. Inherited mitochondrial DNA mutations have been implicated in both syndromic and non-syndromic hearing loss, as well as in predisposition to aminoglycoside ototoxicity. Acquired mitochondrial dysfunction in the absence of mitochondrial DNA mutations has also been proposed as playing an important role in noise-induced and toxin-induced hearing loss. Presbycusis, the hearing loss associated with aging, may be caused by mitochondrial dysfunction resulting from the accumulation of acquired mitochondrial DNA mutations and other factors. The pathophysiological mechanisms and clinical implications of these findings are discussed.     

Selective disruption of MMP-2 gene exacerbates myocardial inflammation and dysfunction in mice with cytokine-induced cardiomyopathy

Tumor necrosis factor-alpha (TNF-alpha) plays a pathophysiological role in the development and progression of heart failure. Matrix metalloproteinase (MMP)-2 is involved in extracellular matrix remodeling. Recent evidence suggests a protective role for this protease against tissue inflammation. Although MMP-2 is upregulated in the failing heart, little is known about its pathophysiological role. We thus hypothesized that ablation of the MMP-2 gene could affect cardiac remodeling and failure in TNF-alpha-induced cardiomyopathy. We crossed transgenic mice with cardiac-specific overexpression of TNF-alpha (TG) with MMP-2 knockout (KO) mice. Four groups of male and female mice were studied: wild-type (WT) with wild MMP-2 (WT/MMP(+/+)), WT with MMP-2 KO (WT/MMP(-/-)), TNF-alpha TG with wild MMP-2 (TG/MMP(+/+)), and TG with MMP-2 KO (TG/MMP(-/-)). The upregulation of MMP-2 zymographic activity in TG/MMP(+/+) mice was completely abolished in TG/MMP(-/-) mice, and other MMPs and tissue inhibitors of metalloproteinase were comparable between groups. Survival was shorter for male TG/MMP(-/-) than TG/MMP(+/+) mice. Female TG/MMP(-/-) mice were more severely affected than TG/MMP(+/+) mice with diminished cardiac function. Myocardial TNF-alpha and other proinflammatory cytokines were increased in TG/MMP(+/+) mice, and this increase was similarly observed in TG/MMP(-/-) mice. The extent of myocardial infiltrating cells including macrophages was greater in TG/MMP(-/-) than in TG/MMP(+/+) mice. Selective ablation of the MMP-2 gene reduces survival and exacerbates cardiac failure in association with the increased level of myocardial inflammation. MMP-2 may play a cardioprotective role in the pathogenesis of cytokine-induced cardiomyopathy.     

Functional polymorphism located in MMP-9 gene promoter is strongly associated with obesity

The adipose tissue expansion is accompanied by remodeling of extracellular matrix performed by matrix metalloproteinases (MMPs). Higher plasma and tissue MMP-9 levels are found in obese; therefore, we evaluated if the functional C(-1562)T polymorphism (rs3918242) located in promoter region of the MMP-9 gene is associated with obesity in women. We studied 112 lean and 114 obese women. Plasma MMP-9 and tissue inhibitor of MMP-9 (TIMP)-1 were measured using enzyme-linked immunosorbent assay. We found different genotype frequencies between lean and obese women (p=0.008), prevailing T-allele in obese (2.3-fold). However, although obese women present higher levels of plasma MMP-9, lack of modulation by the polymorphism was found (all p>0.05). Our findings suggest that C(-1562)T polymorphism may contribute to pathogenetic mechanisms involved in the development of obesity in women.     

Association of MMP-2 and MMP-9 with clinical outcome of neurocysticercosis

Matrix metalloproteinases (MMPs) are the major endopeptidases involved in proteolysis of blood brain barrier (BBB) during central nervous system (CNS) infections. The present study detected serum levels and activities of MMP-2 and MMP-9 in patients with neurocysticercosis (NCC) and their association with symptomatic disease. In total, 68 individuals with NCC (36 symptomatic patients with active seizures and 32 asymptomatic individuals) and 37 healthy controls were enrolled for the study. Serum MMP-2 and MMP-9 levels and their activities were measured by ELISA and gel zymography respectively. Mean serum MMP-2 levels (ng/ml) were higher both in asymptomatic and symptomatic NCC cases compared to healthy controls. However, significantly higher levels of serum MMP-9 (ng/ml) were detected only in symptomatic NCC patients compared to asymptomatic NCC cases and healthy controls. Levels of both MMPs positively correlated with symptomatic NCC. Serum MMP-2 activities were significantly higher in symptomatic and asymptomatic NCC compared to healthy controls whereas serum MMP-9 activity was significantly associated with symptomatic NCC compared to healthy controls and asymptomatic NCC. In conclusion, the elevated level of MMP-9 in serum appears to play an important role in the development of symptoms i.e. active seizures in patients with NCC. However, further studies are needed to elucidate its precise role in disease pathogenesis.     

基质金属蛋白酶-9 与脑出血的关系

基质金属蛋白酶(MMPs)是一族锌离子依赖性内肽酶,具有降解细胞外基质的作用,而后者是构成血管基底膜的主要成分。MMPs参与了伤口愈合、动脉硬化发生、肿瘤细胞扩散等过程。MMP-9是MMPs中的重要成员,与脑血管病关系密切,在脑出血的发生、发展过程中起到了重要的作用。因此,监测MMP-9表达水平的变化可能对脑出血的发生、发展及预后产生重要影响;而降低MMP-9表达水平,则可能成为治疗脑出血的新途径。     

MMP-2、MMP-9及VEGF在子宫内膜异位症中的表达及临床意义

目的:研究基质金属蛋白酶2(MMP-2)、基质金属蛋白酶9(MMP-9)、血管内皮生长因子(VEGF)表达在子宫内膜异位症发生发展中的作用.方法:应用免疫组化二步法检测EMs患者异位内膜36例、在位内膜36例及正常内膜中的MMP-2、MMP-9、VEGF的表达情况.结果:MMP-2、MMP-9、VEGF在异位内膜组中阳性表达率分别为94.4%、91.7%和91.7%,显著高于在位内膜组、正常内膜组(P＜0.05);而在位内膜组和正常内膜组差异无统计学意义(P＞0.05).结论:检测MMP-2、MMP-9、VEGF的表达可用于判断子宫内膜异位症的侵袭转移和评估预后.     

UVA-mediated downregulation of MMP-2 and MMP-9 in human epidermal keratinocytes

While human dermal fibroblasts increase the expression and secretion of distinct matrix metalloproteinases (MMPs) in response to ultraviolet (UV) irradiation, much less is known about regulation of MMPs with regard to normal human epidermal keratinocytes (NHEK). In this in vitro study, the effect of ultraviolet A (UVA) irradiation on gelatinase expression and secretion by NHEK was investigated. Irradiation of NHEK with non-toxic doses of UVA resulted in a dose-dependent downregulation of MMP-2 (gelatinase A) and MMP-9 (gelatinase B). A single dose of 30JUVA/cm(2) lowered MMP-2 activity to 26% and MMP-9 activity to 33% compared with mock-irradiated cells at 24h after irradiation. Downregulation of MMP-2 and MMP-9 steady-state mRNA levels was observed at 4h after UVA irradiation. The inhibitory effect of UVA on gelatinases was mediated by UVA-generated singlet oxygen (1O(2)). These findings suggest an inverse response to UVA irradiation in NHEK than in fibroblasts.     

UV-B induced production of MMP-2 and MMP-9 in human corneal cells

The purpose of this study was to determine the production of metalloproteinases (MMP) 2 and 9 following UV-B irradiation in human corneal epithelial cells and fibroblasts. Epithelial cells and fibroblasts were separated from human donor corneas and exposed to UV-B lamp irradiation for 20, 40, 80 and 120 s. Media samples were collected at 8, 24, 48 and 72 h and gelatinase A and B production was assayed by the ELISA test. Statistical significance of production was assessed by the paired t-test. Increased production of MMP-2 was found in human corneal fibroblasts in response to UV-B irradiation. A statistically significant production of MMP-2 was not observed in human corneal epithelial cells following UV-B exposure. We did not detect any increase in MMP-9 after irradiation in either epithelial cells or fibroblasts. MMP-2 is produced by the corneal fibroblasts in the acute phase after UV-B irradiation. MMP-9 is not released in vitro following UV-B irradiation damage and therefore does not directly participate in the pathophysiology of acute photokeratitis.