Expression of matrix metalloproteinases 2, 7 and 9, and their tissue inhibitors 1 and 2, in developing rabbit tracheae

BACKGROUND: Structural changes in the developing conducting airway impact the rigidity of the airway, altering the airway's ability to sustain its shape during ventilation. The developmental changes in airway compliance oppose the changes in compliance of the developing lung; thus the expression profiles of matrix modeling proteins likely are also opposite in these developing organs. OBJECTIVES: To determine the profiles of matrix metalloproteinases (MMPs) -2, -7, and -9 and tissue inhibitors (TIMPs) -1 and -2 in the developing trachea and test the hypothesis these profiles would contrast those previously reported for the lung. METHODS: Rabbits tracheae were harvested at 21 days of gestation, 3 and 17 days postgestation and at adulthood. Tissue homogenates were analyzed by substrate zymography for the activity of MMPs, and reverse zymography for TIMPs. Immunostainings on neonatal lamb tracheal rings were used to localize MMP-2 and 9. RESULTS: Analysis revealed an age-dependent decrease in total MMP-2 quantity and the ratio of active to latent forms. TIMP-2 shows a time-dependent increase throughout airway development. Total MMP-9 and TIMP-1 quantities were unchanged across these ages, although MMP-9 protein was found predominantly in its latent form during development and predominantly in its active form during adulthood. Respiratory epithelial cells reacted positive for both MMP-2 and 9 and trachealis muscle fibers were positive for MMP-2. No MMP-7 expression was identified in the rabbit airway. CONCLUSIONS: The opposing developmental patterns in MMP-2 expression between the airway and lung lead to speculation regarding the role of MMP-2 activity on changes in organ compliance.     

基质金属蛋白酶在小鼠病毒性心肌炎中的表达及肾素-血管紧张素系统阻断后的变化

目的探讨基质金属蛋白酶-3（MMP-3）及金属蛋白酶组织抑制因子-1（TIMP-1）在病毒性心肌炎（VM）小鼠中的作用及肾素一血管紧张素系统抑制剂（RASI）的治疗效果。方法100只Balb／c小鼠随机分为对照组（A组）、模型组（B组）、假干预组（C组）、福辛普利组（D组）和缬沙坦组（E组）5组,每组20只。B～E组小鼠腹腔接种0．1ml的柯萨奇病毒B3（CVB3）悬液建立VM模型,A组腹腔注射不含CVB3的0．1mlHep-2细胞冻溶液作对照组。C组、D组及E组于注射CVB324h后分别予生理盐水、福辛普利、缬沙坦灌胃,第14天处死小鼠,心脏切片HE染色计算病理积分,氯氨T法测定心肌胶原含量,RT—PCR法检测心肌MMP．3、TIMP-1mRNA的表达。结果与A组比较,B组、C组心肌MMP-3表达明显上调,TIMP-1表达明显下调,心肌胶原含量显著增高（P均〈0．05）;与B组比较,D组、E组心肌MMP-3表达明显下调,TIMP-1表达明显上调,心肌胶原含量显著减少（P均〈0．05）。结论MMP-3／TIMP-1参与VM小鼠的心肌病理过程,可能是导致心肌胶原重构的重要因素之一。福辛普利及缬沙坦能明显下调MMP-3的表达、上调TIMP-1的表达,逆转心肌胶原重构。     

Dysregulated expression of matrix metalloproteinases and their inhibitors may participate in the pathogenesis of pre-eclampsia and fetal growth restriction

Background

Trophoblast invasion into the maternal endometrium serves an important function in human pregnancy. Dysregulation of the finely controlled process of trophoblast invasion can result in a wide spectrum of pregnancy abnormalities.

Aims

We aimed to elucidate the relationship between the expression of matrix metalloproteinases and pregnancy complication.

Study design

The study group consisted of placental bed biopsy tissues obtained from normal vaginal deliveries (N = 15), normal cesarean deliveries (N = 15), pre-eclampsia (N = 24) and fetal growth restriction (FGR) (N = 10). We evaluated the expressions of MMP-2, -8, -9, -11, -19, -15 (MT2-MMP), -16 (MT3-MMP), and -24 (MT5-MMP), as well as TIMP-1 and -3, by applying Western blot and immunohistochemistry methods.

Subjects

Human placental tissues were used for this study.

Outcome measures

The expressions of MMP-2, -8, -9, -11, -19, -15 (MT2-MMP), -16 (MT3-MMP), and -24 (MT5-MMP), as well as TIMP-1 and -3 in human placenta tissues.

Results

Compared with those in normal pregnancies, the expression of MMP-2, -8, -9 and -11 was downregulated in villous tissues of pre-eclampsia and FGR cases (p < 0.05). TIMP-1 and -3 were increased in pre-eclampsia and FGR (p < 0.05). No significant difference was found between normal vaginal deliveries and cesarean deliveries.

Conclusions

We speculate that the change in invasion-associated proteinase expression will affect placental development and may thus contribute to the development of complicated pregnancies.     

川崎病患儿血清中hs-CRP与基质金属蛋白酶及抑制剂联合检测的临床意义

目的：检测基质金属蛋白酶(MMP)-2、MMP-9、基质金属蛋白酶抑制剂(TIMP)-1及hs-CRP在川崎病患儿血清中的表达,并探讨其与冠状动脉损伤的关系。方法：选取151例川崎病患儿作为观察组（无冠状动脉损伤40例,有冠状动脉损伤111例）,60例健康儿童作为对照组,采用酶联免疫吸附实验(ELISA)检测血清中MMP-2、MMP-9和TIMP-1含量,应用终点散射比浊法检测血清hs-CRP的含量。结果：MMP-2,MMP-9及hs-CRP含量在冠脉损伤组及无冠脉损伤组与对照组比较差异均有显著性,在冠脉损伤组量最高（P<0.05）;川崎病患儿中MMP-2、MMP-9及TIMP-1之间的检测量呈正相关（P<0.05）。结论：MMP-2、MMP-9、TIMP-1及hs-CRP在川崎病的发生发展中可能起重要作用,MMP-2、MMP-9及hs-CRP的联合检测可能对判断病变程度有重要帮助。［中国当代儿科杂志,2009,11（12）：989-991］     

病毒性心肌炎心肌基质金属蛋白酶活性的动态变化及其与心功能和心肌胶原的关系

目的 探讨病毒性心肌炎(viral myocarditis,VM)心肌基质金属蛋白酶(matrixmetalloproteinases,MMPs)活性的动态变化及其与心功能、心肌胶原的关系。方法 50只6周龄雄性DBA/2小鼠腹腔接种0.14 ml的CVB3建立VM模型,15只同周龄同品系小鼠腹腔接种0.14 ml的Eagle's液作为对照组。VM组于接种后3、7、10、21、30 d,对照组于30 d均采用酶谱法测定心肌基质金属蛋白酶2(MMP-2)和9(MMP-9)的活性;超声心动图测定心脏收缩功能:主动脉血流峰值流速(Vp)、主动脉血流速度积分(Vi);心脏石蜡切片HE染色计算病理积分以及氯氨T法测定心肌的胶原含量。结果 VM小鼠心肌MMP-2、MMP-9的活性表现为一过性增高,以10 d时为最高,并且10 d时Vp、Vi亦明显低于对照组(P<0.05);心肌胶原含量在21 d、30 d时显著增高(P<0.05);MMP-2、MMP-9与病理积分均呈正相关(r分别为0.801,0.821,均P<0.05)、与Vp、Vi呈负相关(r分别为-0.649,-0.683和-0.711,-0.755,均P<0.05)。结论 VM小鼠心肌MMPs过度激活并伴有继发的胶原含量升高,且MMPs活性与心肌病理损害、心功能下降密切相关。MMPs参与了心肌炎的病理改变,可能是导致间质改变、心功能异常的因素之一,提示心肌MMPs是VM心脏间质病变的关联因素,可用来作为判断VM小鼠心肌间质损害程度及估计心脏功能的     

Elastolytic matrix metalloproteinases and coronary outcome in children with Kawasaki disease

Kawasaki disease (KD) is a multisystem vasculitis that leads to coronary artery damage and aneurysm formation. Elastolytic matrix metalloproteinases (MMPs) have been implicated in the pathogenesis of arterial aneurysms. To determine the relationship between circulating levels of elastolytic MMPs and development of coronary artery aneurysms in children with KD, we partnered studies done in affected children with an animal model of disease. In affected children, circulating protein levels and enzymatic activity of MMP-2 and MMP-9 did not have a statistically significant relationship with coronary artery outcome after adjusting for demographic characteristics, and clinical and laboratory features. Although matrix-degrading proteolytic activity was specific for affected mice and localized to inflamed coronary artery segments, the enzymatic activity in the systemic circulation of affected and control mice were not different. Similar to affected children, peripheral blood levels of MMP-9 enzymatic activity did not correlate with coronary artery disease in the animal model. Therefore, circulating levels of MMPs known to act locally may not be useful biomarkers of disease. This is especially relevant to enzymatic activity that is tightly regulated at multiple levels including the local tissue environment.     

Matrix metalloproteinase-9 and tissue inhibitors of metalloproteinases 1 in influenza-associated encephalopathy

Matrix metalloproteinase-9 (MMP-9) and tissue inhibitors of metalloproteinases 1 (TIMP-1) play important roles in the function of the blood-brain barrier. Serum MMP-9 and TIMP-1 concentrations were determined in influenza virus infection with or without neurologic complications. Our results suggest that an imbalance between MMP-9 and TIMP-1 damages the blood-brain barrier and promotes febrile seizure or encephalopathy in influenza virus infection.     

Relevance of tenascin-C and matrix metalloproteinases in vascular abnormalities in murine hypoplastic lungs

BACKGROUND: Tenascin-C (TN-C), an extracellular matrix glycoprotein, is crucial to cell-migration, proliferation, apoptosis and remodeling of tissues, with a potential role in pathobiology of pulmonary hypertension. Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) are crucial to the integrity of the extracellular matrix. TN-C and MMPs are counter-regulatory molecules, which influence the vascular integrity through modulations of elastin. We have a murine model of pulmonary hypoplasia with coexistent diaphragmatic hernia, vascular abnormalities and excessive arterial smooth muscle cell (SMC) proliferation. OBJECTIVES: Our objective was to investigate modulations of TN-C and MMPs in hypoplastic lungs and their possible contribution to the observed pulmonary vascular abnormalities. METHODS: We addressed our objectives by pursing immunoblotting and immunohistochemistry and zymography/reverse zymography to assess the alterations in activities of MMPs and their inhibitors. RESULTS: We observed significant down-regulation of MMP-9 activity in hypoplastic lungs at the later fetal developmental stages, whereas MMP-2 activity assessed by gelatin zymography remained unaltered. Reverse zymography revealed up-regulation of activities of TIMP-1, -2, -3 and -4 in hypoplastic lungs during later fetal development, with pronounced increases in TIMP-3 and -4 activities. Furthermore, immunoblot analyses and immunohistochemistry revealed that TN-C protein was down-regulated in developing hypoplastic lungs, compared to normal lungs. CONCLUSIONS: (1)TN-C is known to inhibit vascular SMC proliferation. But, decrease in TN-C in hypoplastic lungs may support the observed arterial SMC proliferation. (2) Our studies showed that in hypoplastic lungs the SMC apoptosis is not affected, thus suggesting that SMC proliferation and apoptosis may be two separate processes in pulmonary hypoplasia with coexistent diaphragmatic hernia. Together, our data showed an imbalance in the extracellular matrix proteins, which may contribute to the pulmonary vascular abnormalities.     

缺氧缺血性脑损伤新生大鼠脑组织基质金属蛋白酶的动态变化及对细胞凋亡的影响

目的探讨缺氧缺血性脑损伤(HIBD)新生大鼠脑组织基质金属蛋白酶(MMPs)的动态变化及其与细胞凋亡的关系。方法将72只7日龄W istar大鼠随机分为假手术组和HIBD组,后者根据处死时间分为6 h、24 h、48 h、5天和14天组,每组12只。采用免疫组化方法测定MMP-2、MMP-9和金属蛋白酶组织抑制因子-1(TIMP-1)的表达,用流式细胞术测定脑细胞凋亡率。结果 HIBD组缺氧缺血侧大脑组织MMP-2于损伤6 h增高,5天再次升高,且峰值高于6 h组,与假手术组相比,除24 h组,其余差异均有统计学意义(P<0.05);MMP-9正常情况下有一定表达,HIBD 24 h表达达高峰,后逐渐下降,与假手术组相比,除14天组,差异均有统计学意义(P<0.05);TIMP-1正常情况下与MMP-9保持动态平衡,HIBD时两者比例失衡,后期表达增加两者平衡恢复,与假手术组相比,除6 h组和14天组,差异均有统计学意义(P<0.05)。HIBD组缺氧缺血侧脑细胞凋亡率24 h轻度升高,5天达高峰,与假手术组相比,除6 h组,差异均有统计学意义(P<0.05)。结论新生大鼠HIBD后MMP-2、MMP-9和TIMP-1的表达有不同程度变化,可能参与了HIBD的发病与修复。     

新生鼠窒息后心肌基质金属蛋白酶活性与心肌损害的相关性

目的探讨新生鼠窒息后心肌组织心肌基质金属蛋白酶（MMPs）表达与心肌损害的相关性。方法60只7—10日龄Wistar鼠,随机分成对照组（D组）、窒息组（A、B、C3组,分别为窒息复氧后1、7、14d）,每组15只。窒息组模拟新生鼠常压窒息模型,测定cTnI水平;心肌组织MMPs-3、9活性;心肌病理积分及胶原含量。结果cTnI（ng／m1）A组（0．3680±0．40）明显高于D组（0．0783±0．06）P〈0．05,B组下降（0．1889-i-0．15）P仍〈0．05,C组降至正常（0．1338±0．07）P〉0．05;心肌组织MMPs-3活性A组（0．1847±0．04）,B组最高（O．2780±0．05）,C组（0．2014±0．05）,与D组（0．1213±0．03）比较P均〈0．05;MMPs-9活性A、B、C组逐渐增高（0．1502±0．01、0．1715±0．01、0．1868±0．03、P均〈0．05）;心肌胶原含量A、B、C组逐渐增高（35．59±1．09、38．94±0．67、40．69±0．75ng／ml分别为P〉0．05、P〈0．05、P〈0．05）;MMPs-3、9表达与心肌病理积分呈正相关（r分别为0．669、0．667,P均〈0．05）,与胶原含量呈正相关（r分别为0．482、0．679,P均〈0．05）。结论新生鼠窒息后心肌MMPs-3、9过度激活,继发胶原含量增高,MMPs-3、9活性与心肌病理损害密切相关。提示MMPs参与了窒息后心肌损害,可能是心肌间质重塑的因素之一。     

The pathophysiology of coronary artery aneurysms in Kawasaki disease: role of matrix metalloproteinases

Kawasaki disease is an acute inflammatory syndrome that takes the form of systemic vasculitis, and predominantly affects children. Important complications of this disease are coronary artery dilation and aneurysm formation. Recent studies indicate that Kawasaki disease patients have elevated expression, activity, or protein levels of matrix metalloproteinases (MMPs), and suggest that imbalances in MMPs or MMP/tissue inhibitor of MMP (TIMP) play important pathophysiological roles in the development of coronary artery lesions in this disease. However, it remains unclear whether MMP activities at the site of coronary artery lesions are indeed increased. Further studies on the effects of MMP inhibition on coronary outcome are needed to define the roles of MMPs and TIMPs in the formation of coronary artery lesions in Kawasaki disease; findings of such studies may support the use of MMP inhibitors for the prevention of coronary artery complications in patients with this disease.     

Remodeling of rat neonatal pulmonary artery: role of matrix metalloproteinases.

We studied whether rapid thinning of large pulmonary arteries of neonatal rats is associated with breakdown of collagen. Pulmonary artery extracts from fetal to 21 days of age were assayed for collagen content and matrix metalloproteinases. Within 3 days postpartum, no changes in collagen content, collagenolytic activity, or levels of stromelysin-l or gelatinase A were observed. After day 3, collagen content and total proteolytic activity increased with little change in matrix metalloproteinase expression. Thus, collagen was not degraded, and the late increases in collagen and total proteolytic activity were probably growth related. Unlike adult rats in which collagen is broken down after reversal of hypoxic pulmonary artery remodeling, collagen is not broken down in neonatal pulmonary arteries during adaptation to extrauterine life.     

The pathophysiology of coronary artery aneurysms in Kawasaki disease: role of matrix metalloproteinases.

Kawasaki disease is an acute inflammatory syndrome that takes the form of systemic vasculitis, and predominantly affects children. Important complications of this disease are coronary artery dilation and aneurysm formation. Recent studies indicate that Kawasaki disease patients have elevated expression, activity, or protein levels of matrix metalloproteinases (MMPs), and suggest that imbalances in MMPs or MMP/tissue inhibitor of MMP (TIMP) play important pathophysiological roles in the development of coronary artery lesions in this disease. However, it remains unclear whether MMP activities at the site of coronary artery lesions are indeed increased. Further studies on the effects of MMP inhibition on coronary outcome are needed to define the roles of MMPs and TIMPs in the formation of coronary artery lesions in Kawasaki disease; findings of such studies may support the use of MMP inhibitors for the prevention of coronary artery complications in patients with this disease.     

The role of matrix metalloproteinases -9 and -2 in development of neonatal chronic lung disease

AIM: Matrix metalloproteinases (MMPs) -9 and -2 degrade type-IV collagen, a major constituent of lung basement membrane, and may have a role in the pathogenesis of neonatal chronic lung disease (CLD). We determined factors influencing MMP levels in neonatal bronchoalveolar lavage (BAL) fluid to establish whether an imbalance between MMP and its inhibitor could be implicated in CLD. METHODS: We measured MMP-9 and -2 and tissue inhibitor of metalloproteinase-1 (TIMP-1) levels in 316 BAL fluid samples from 121 babies of gestational ages 23 to 42 wk over the first 14 d of life to determine effects of gestation and postnatal age. Median MMP-9, -2, TIMP-1 and MMP-9/TIMP-1 ratio in BAL were further studied in a subgroup of 85 babies <33 wk gestation to determine their ability to predict CLD and to establish effects of antenatal corticosteroid therapy (ANCS). RESULTS: MMP-9, -2 and TIMP levels did not vary with postnatal age over the first week. Median MMP-9 levels and MMP-9/TIMP-1 ratio increased with decreasing gestation in preterm babies. The MMP-9/TIMP-1 ratio was higher in babies who developed CLD, implying a proteinase/antiproteinase imbalance, but this association disappeared when controlled for gestational age. ANCS had no effect on BAL fluid MMP or TIMP levels. CONCLUSION: MMPs may have a role in the development of lung injury and fibrosis, but estimating their levels in the first week of life does not help with prediction of CLD.     

雷公藤甲素对实验性肺动脉高压肺脏基质金属蛋白酶MMP2和MMP9的影响

目的：雷公藤甲素能缓解实验性大鼠肺动脉高压,该实验观察雷公藤甲素对肺动脉高压血管重构模型的影响并探讨与基质金属蛋白酶的关系。方法：60只SD大鼠随机分为6组：①正常对照组;②模型组;③连续治疗组;④连续治疗组的安慰剂对照组;⑤延迟治疗组;⑥延迟治疗组的安慰剂对照组。术后5周通过心导管术测得血液动力学指标,并用弹力纤维染色观察肺小动脉的病理学变化。用免疫组织化学染色显示MMP2 和MMP9在肺血管上的阳性分布和半定量,荧光定量PCR（FS-PCR）行肺组织MMP2和MMP9的定量。结果:平均肺动脉压(m PAP） 为38.10±1.20 vs 16.70±1.16 mmHg和心室重量变化［RV/(LV+S)%］为（62.45±5.28）% vs （22.76±3.01）%及血管阻塞积分(VOS） 1.736±0.080 vs 0.000±0.000,②组较①组明显增高(P<0.01);MMP2和MMP9的表达量在肺血管壁(MMP2为 1.48±0.35 vs 0.00±0.00; MMP9 1.95±0.22 vs 0.00±0.00)和肺组织(MMP2 0.85±0.67 vs 5.50±0.67. MMP9 0.85±0.67 vs 4.95±0.69)均显示②组较①组明显增高（P<0.05）;雷公藤甲素连续治疗组和延迟治疗组各指标均较模型组和各自的安慰剂组减低,且连续治疗效果更好（mPAP 20.80±1.03 vs 26.20±1.03 mmHg,P<0.05）。结论: 雷公藤甲素能延缓大鼠左肺切除加野百合碱注射诱导的肺动脉高压和右心室肥厚,并能抑制新生内膜形成,可能与抑制MMPs的活性增高有关。［中国当代儿科杂志,2007,9（5）：479-483］