Matrix Metalloproteinase-9 mRNA Expression Research Articles

Preventive and therapeutic effects of Yiqi Huayu Recipe on degeneration of articular cartilage in rats with osteoarthritis

To study the efficacy and possible mechanism of Yiqi Huayu Recipe (YQHYR), a compound traditional Chinese herbal medicine, in preventing and treating degeneration of the articular cartilage in rats with osteoarthritis (OA). A total of 90 one-month-old SD rats were randomly divided into normal control group, untreated group and YQHYR group, with 30 rats in each group. The osteoarthritis was induced by shoulder disarticulation plus upright posture in rats. The rats in YQHYR group were treated with YQHYR at 5-, 7- and 9-month old (4, 6 and 8 months after the surgery) for one month. Ten rats in each subgroup were sacrificed at 6-, 8- and 10-month old (5, 7 and 9 months after the surgery) respectively and the knee joint samples were harvested for detection. Safranin-O/fast green staining was performed to examine the morphology of articular cartilage. The expressions of type II collagen (Col2A1), aggrecan-1 (Agc1), matrix metalloproteinase-13 (MMP-13) and tissue inhibitor of metalloproteinase-1 (TIMP-1) mRNAs were evaluated by real-time fluorescent quantitation polymerase chain reaction. Histological analysis showed that the structure of articular cartilage was seriously destroyed in rats in the untreated group. On the contrary, the degenerative changes of the articular cartilage in the YQHYR group were dropped off. Real-time fluorescent quantitation polymerase chain reaction showed that expressions of Agc1, TIMP-1 and Col2A1 mRNAs were up-regulated in 6- and 10-month-old rats in the YQHYR group as compared with those in the untreated group (P<0.01, P<0.05), but there was no significant difference in expression of MMP-13 mRNA between the YQHYR group and the untreated group. The expression of Agc1 mRNA was up-regulated and the expression of MMP-13 mRNA was down-regulated in the 8-month-old rats in YQHYR group as compared with those in the untreated group (P<0.01). YQHYR can promote the synthesizing of aggrecan and type II collagen in chondrocytes and delay articular cartilage degradation in OA rats.

Effects of BRAFV600E mutation on the invasion capacity of human melanoma cells.

Objective To investigate the effect of BRAFV600E mutation on the invasion capacity of a human melanoma cell line, A375. Methods Plasmids containing short hairpin RNAs (shRNA) specific for BRAF gene were prepared in previous study, and used to transfect A375 cells. Those cells transfected with negative plasmid and untransfected cells served as the controls. Transwell chambers were used to examine the invasion ability of melanoma cells in vitro. RT-PCR and Western blotting were performed to detect the mRNA and protein expressions of matrix metalloproteinase 2 (MMP-2) and vascular endothelial growth factor (VEGF), respectively, before and after the transfection. The activity of MMP-2 was also studied with sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Results Compared with the negative control, the specific shRNA decreased the mRNA and protein expressions of MMP-2 by 35% and 85%, respectively, and those of VEGF by 45% and 14%, respectively. Additionally, the number of cells invading through Matrigel chambers reduced by 69% in those cells transfected with the positive plasmid. Conclusions The mutant BRAFV600E has the potential to enhance the invasion capacity of melanoma cells, whereas specific shRNA could suppress the increase in metastasis capacity likely by inhibiting the production of VEGF and MMP. Key words: Melanoma; RNA, small interfering; Matrix metalloproteinase 2; Vascular endothelial growth factors; Cell line, tumor; BRAF gene

Matrix metalloproteinase-1 expression induced by IL-1β requires acid sphingomyelinase

Matrix metalloproteinase-1 (MMP-1) is increased in inflammatory conditions leading to destruction of extracellular matrix. Many inflammatory stimuli activate sphingomyelinases (SMases), which generate ceramide. We aimed to define the relevance and type of SMase responsible for the regulation of MMP-1. Acid sphingomyelinase (ASM)-deficient human fibroblasts failed to phosphorylate extracellular signal-regulated kinase (ERK), or upregulate MMP-1 mRNA and protein expression upon stimulation with interleukin-1 beta (IL-1β), whereas phosphorylation of p38 mitogen-activated protein kinase and IL-8 production remained unaffected. Transfection of ASM restored MMP-1 production. Addition of exogenous SMase was sufficient to restore activation of ERK and increase MMP-1 mRNA. Inhibition of ASM with imipramine completely abrogated MMP-1 induction. The results suggest that IL-1β-induced expression of MMP-1 is dependent on ASM.

Correlation between MMPs and their inhibitors in breast cancer tumor tissue specimens and in cell lines with different metastatic potential

BackgroundThe metastatic disease rather than the primary tumor itself is responsible for death in most solid tumors, including breast cancer. The role of matrix metalloproteinases (MMPs), tissue inhibitors of MMPs (TIMPs) and Reversion-inducing cysteine-rich protein with Kazal motifs (RECK) in the metastatic process has previously been established. However, in all published studies only a limited number of MMPs/MMP inhibitors was analyzed in a limited number of cell lines. Here, we propose a more comprehensive approach by analyzing the expression levels of several MMPs (MMP-2, MMP-9 and MMP-14) and MMP inhibitors (TIMP-1, TIMP-2 and RECK) in different models (five human breast cancer cell lines, 72 primary breast tumors and 30 adjacent normal tissues).MethodsWe analyzed the expression levels of MMP-2, MMP-9 and MMP-14 and their inhibitors (TIMP-1, TIMP-2 and RECK) by quantitative RT-PCR (qRT-PCR) in five human breast cancer cell lines presenting increased invasiveness and metastatic potential, 72 primary breast tumors and 30 adjacent normal tissues. Moreover, the role of cell-extracellular matrix elements interactions in the regulation of expression and activity of MMPs and their inhibitors was analyzed by culturing these cell lines on plastic or on artificial ECM (Matrigel).ResultsThe results demonstrated that MMPs mRNA expression levels displayed a positive and statistically significant correlation with the transcriptional expression levels of their inhibitors both in the cell line models and in the tumor tissue samples. Furthermore, the expression of all MMP inhibitors was modulated by cell-Matrigel contact only in highly invasive and metastatic cell lines. The enzyme/inhibitor balance at the transcriptional level significantly favors the enzyme which is more evident in tumor than in adjacent non-tumor tissue samples.ConclusionOur results suggest that the expression of MMPs and their inhibitors, at least at the transcriptional level, might be regulated by common factors and signaling pathways. Therefore, the multi-factorial analysis of these molecules could provide new and independent prognostic information contributing to the determination of more adequate therapy strategies for each patient.

Time profiles of the expression of metalloproteinases, tissue inhibitors of metalloproteases, cytokines and collagens in hamsters infected with Opisthorchis viverrini with special reference to peribiliary fibrosis and liver injury

The liver fluke Opisthorchis viverrini is endemic in southeastern Asia, and causes cholangiocarcinoma and liver fibrosis. We investigated the time profile of the expression of matrix metalloproteinases (MMPs) and tissue inhibitors of MMPs (TIMPs) in relation to peribiliary fibrosis in O. viverrini-infected hamsters. Hepatic mRNA expression of MMPs, TIMPs, cytokines and collagens I and III was assessed by quantitative reverse transcription-PCR. Zymography and immunohistochemistry were also used to examine MMPs-2 and -9 expression. After infection, an increase of peribiliary fibrosis was time-dependent. Opisthorhis viverrini-induced gene expression in hamster liver, with increased mRNA expression levels of IL-1β, TNF-α, TGF-β, and collagens I and III, was observed at 21 days p.i. Expression of MMPs-2, -13 and -14 and TIMPs-1 and -3 genes, was significantly higher at 1 month, and maximal levels of most MMPs (MMPs-2, -9, -13 and -14) were observed at 2 months p.i. The cytoplasmic levels of MMP-2 and MMP-9 were similar to mRNA expression. Immunohistochemistry revealed that MMP-9 was expressed mainly in the cytoplasm of inflammatory cells at the invasive front of the fibrous area. In contrast, the highest levels of mRNA expression of TIMPs-2 and -3, and TGF-β were observed 10 months p.i. Concentration of TIMP-2 protein in the plasma correlated with its transcriptional level ( r = 0.320, P = 0.040). Peribiliary fibrosis correlated positively with liver hydroxyproline content ( r = 0.846, P < 0.001), plasma hydroxyproline concentration ( r = 0.770, P < 0.001), plasma TIMP-2 level ( r = 0.335, P = 0.046), and mRNA expression levels of MMP-7 ( r = 0.511, P = 0.006), TIMP-1 ( r = 0.320, P = 0.040), TIMP-2 ( r = 0.428, P = 0.026), and TIMP-3 ( r = 0.553, P = 0.003). This study suggests that expression of MMPs is associated with an inflammatory reaction in the early phase and TIMPs expression at the late phase may contribute to both fibrosis and liver injury. MMPs and TIMPs may serve as diagnostic markers for the severity of O. viverrini-induced liver injury.

Effects of angiopoietin-1 on attachment and metastasis ofhuman gastric cancer cell line BGC-823

To evaluate the effects of angiopoietin-1 (Ang-1) on adhesion of gastric cancer cell line BGC-823 and expression of integrin beta1, CD44V6, urokinase-type plasminogen activator (uPA) and matrix metalloproteinase-2 (MMP-2). BGC-823 cells were transfected transiently with adenovirus-Ang-1 (Ad-Ang-1). Cells transfected transiently with adenovirus-green fluorescent protein (Ad-GFP) and untransfected cells were used as a negative and blank control group, respectively. The cell adhesion rate between cell and extracellular matrix (ECM) was determined by cell adhesion assay. To investigate whether Ang-1 could reinforce gastric carcinoma metastasis, we performed migration and invasion assays in BGC-823 cells. The mRNA and protein expression of integrin beta1, CD44V6, uPA and MMP-2 were detected by reverse transcription polymerase chain reaction and Western blotting, respectively. The expression of integrin beta1 and CD44V6 was measured by immunohistochemistry. BGC-823 cells were transfected successfully. The adhesion rate increased significantly in the Ad-Ang-1 group (P < 0.05). The Ad-Ang-1-transfected group had a significant increase in migration and invasion compared with that of the mock-transfected and Ad-GFP groups. The mRNA and protein expression of integrin beta1, CD44V6, uPA and MMP-2 in the Ad-Ang-1 group was higher than that in the Ad-GFP and blank control groups (P < 0.05). Compared with mock-transfected and Ad-GFP groups, integrin beta1 and CD44V6 expression intensity greatly increased (P < 0.05). Transfection of Ang-1 into human gastric cancer cell line BGC-823 can significantly increase expression of integrin beta1 and CD44V6, by which cell adhesion and metastasis to the ECM are promoted.

Endometrial mRNA expression of matrix metalloproteinases, their tissue inhibitors and cell adhesion molecules in unexplained infertility and implantation failure patients

The aim of this study was to analyse whether some cases of unexplained infertility and implantation failure after IVF could be explained by different expression levels of the matrix metalloproteinases (MMP-2, 9), their tissue inhibitors (TIMP-2, 3) and intercellular (ICAM-1) and vascular (VCAM-1) adhesion molecules in endothelial cells. Total RNA was extracted from the endometrial tissues of 41 women (unexplained infertile, group 1, n = 15; fertile volunteers, group 2, n = 15 and patients with implantation failure after IVF, group 3, n = 11). MMP-2, MMP-9, TIMP-2, TIMP-3, ICAM-1 and VCAM-1 mRNA expression levels were measured quantitatively using real-time polymerase chain reaction. In the endometrium from women with unexplained infertility and implantation failure after IVF, MMP-2 and TIMP-3 expression were significantly decreased when compared with the fertile group ( P ≤ 0.05 and P. 0.001 respectively). In addition, a marked decrease was observed in the expression of VCAM-1 in women with unexplained infertility. These results suggest that the expression of gelatinase A (MMP-2), TIMP-3 and VCAM-1, at least at the transcriptional level, might be regulated by common factors and signalling pathways. The present study adds new and important data in this field and highlights the complex preparation of the endometrium for implantation at the molecular lev

Propylthiouracil Attenuates Monocrotaline-Induced Pulmonary Arterial Hypertension in Rats

Propylthiouracil (PTU) enhances nitric oxide production and inhibits smooth muscle cell proliferation, suggesting a possible role in the prevention of pulmonary arterial hypertension (PAH). The 30 male Sprague-Dawley rats were randomized to receive saline injection only (group 1), monocrotaline (MCT) (70 mg/kg) only (group 2) or MCT + 0.1% PTU in drinking water (group 3) given on day 5 after MCT administration. By day 35, western blot showed lower connexin43 (Cx43) and membranous protein kinase C-epsilon expressions in the right ventricle (RV) of group 2 animals than in the other groups (all P<0.05). Conversely, Cx43 expression in the lung was higher in group 2 than in other groups (all P<0.02). Additionally, mRNA expressions of matrix metalloproteinase-9, tissue necrotic factor-alpha, and caspase-3 were higher, whereas Bcl-2 and endothelial nitric oxide synthase were lower, in the lungs and RV of group 2 rats than in the other groups (all P<0.05). Moreover, the numbers of alveolar sacs and lung arterioles were also reduced in group 2 than in other groups (all P<0.05), and RV systolic pressure and RV weight were increased in group 2 compared with other groups (all P<0.001). PTU effectively attenuates complications associated with MCT-induced PAH.

Lentivirus-mediated shRNA interference targeting STAT3 inhibits human pancreatic cancer cell invasion

To investigate RNA interference targeting signal transducer and activator of transcription-3 (STAT3) on invasion of human pancreatic cancer cells. We constructed three plasmids of RNA interference targeting the STAT3 gene. After LV (lentivirus)-STAT3siRNA (STAT3 small interfering RNA) the vector was transfected into the human pancreatic cell line, SW1990 and cell proliferation was measured by the MTT assay. Flow cytometry was used to assess cell cycle. Vascular endothelial growth factor (VEGF) and matrix metalloproteinase-2 (MMP-2) mRNA and protein expression were examined by quantitative PCR and western blotting, respectively. The invasion ability of SW1990 cells was determined by cell invasion assay. We successfully constructed the LV-STAT3siRNA lentivirus vector and proved that it can suppress expression of STAT3 gene in SW1990 cells. RNA interference of STAT3 by the LV-STAT3siRNA construct significantly inhibited the growth of SW1990 cells, in addition to significantly decreasing both VEGF and MMP-2 mRNA and protein expression. Moreover, suppression of STAT3 by LV-STAT3siRNA decreased the invasion ability of SW1990 cells. The STAT3 signaling pathway may provide a novel therapeutic target for the treatment of pancreatic cancer since it inhibits the invasion ability of pancreatic cancer cells.

Suppression of matrix metalloproteinase-2 via RNA interference inhibits pancreatic carcinoma cell invasiveness and adhesion

To investigate the inhibitory effects of RNA interference (RNAi) on expression of matrix metalloproteinase-2 (MMP-2) gene and invasiveness and adhesion of human pancreatic cancer cell line, BxPC-3. RNAi was performed using the vector (pGPU6)-based small interference RNA (siRNA) plasmid gene silence system to specifically knock down MMP-2 expression in pancreatic cancer cell line, BxPC-3. Four groups of different specific target sequence in coding region of MMP-2 and one non-specific sequence were chosen to construct four experimental siRNA plasmids of pGPU6-1, pGPU6-2, pGPU6-3 and pGPU6-4, and one negative control siRNA plasmid of pGPU6 (-). MMP-2 expression was measured by reverse transcription polymerase chain reaction (RT-PCR) and Western blot. Cell proliferation and apoptosis were examined by methyl thiazolyl tetrazolium (MTT) and flow cytometry, respectively. The abilities of adhesion and invasion were detected by cell adhesion assay and cell invasion assay using Transwell chambers. The expression of MMP-2 was inhibited and the inhibitory effects of different sequence varied. pGPU6-1 group had the most efficient inhibitory effect, followed by pGPU6-2 and pGPU6-3 groups. Invasiveness and adhesion were more significantly reduced in pGPU6-1, pGPU6-2 and pGPU6-3 groups as compared with pGPU6 (-) and blank control groups. However, no difference concerning cell proliferation and apoptosis was observed after transfection between experiment groups and control groups. RNAi against MMP-2 successfully inhibited the mRNA and protein expression of MMP-2 in the pancreatic cancer cell line, BxPC-3, leading to a potent suppression of tumor cell adhesion and invasion without affecting cell proliferation and apoptosis. These findings suggest that the RNAi approach towards MMP-2 may be an effective therapeutic strategy for the clinical management of pancreatic tumor.

Double-stranded RNA poly(I:C) enhances matrix metalloproteinase mRNA expression in human nasal polyp epithelial cells

Conclusion: The significant up-regulation of matrix metalloproteinase (MMP)-9 mRNA, which is not modulated by tissue inhibitor of metalloproteinase (TIMP)-1, is an additional source of increased proteolytic activity in virus-infected upper airways that might contribute to the exacerbation of chronic rhinosinusitis with nasal polyps. Objectives: Chronic rhinosinusitis is often exacerbated by viral infection. We hypothesized that a disruption of the mechanisms that regulate the activity of MMPs during viral infection is one possible mechanism responsible for the exacerbation. In the present study we attempted to achieve a better understanding of MMP expression in nasal epithelial cells after viral infection. Materials and methods: Human nasal epithelial cells were isolated from nasal polyp specimens obtained during endoscopic endonasal surgery in chronic rhinosinusitis patients. We investigated the expression of MMP-2, MMP-9, and TIMP-1 mRNA in primary human nasal polyp epithelial cells after dsRNA stimulation. Results: Among the genes whose expression was evaluated, only expression of MMP-9 mRNA increased significantly after dsRNA stimulation.

Resveratrol inhibits MMP-9 expression by up-regulating PPAR α expression in an oxygen glucose deprivation-exposed neuron model

Resveratrol ( trans-3,4′,5-trihydroxystilbene, Res) is a natural polyphenol. A recent experiment confirmed that Res can selectively activate both peroxisome proliferators-activated receptors (PPAR) α and γ. In addition, Res can protect neurons by matrix metalloproteinase-9 (MMP-9) down-regulation. The relationship between Res, MMP-9 and PPAR α or γ was studied in an oxygen glucose deprivation-exposed neuron model. It showed that Res can inhibit mRNA and protein expression of MMP-9, while it up-regulates the expression of PPAR α and γ. The effect of Res on both PPAR α and MMP-9 can be offset partially by MK886. However, PPAR γ antagonist GW9662 had little effect on MMP-9 expression. These results suggest that Res can inhibit MMP-9 expression by up-regulating PPAR α.

Transactivation by Runt related factor-2 of matrix metalloproteinase-13 in astrocytes

We have previously shown functional expression by osteoblasts of signaling machineries required for neurotransmission in the brain. In this study, we have evaluated possible functional expression of different osseous genes in the brain. In embryonic and adult mouse brains, mRNA expression was invariably seen for the master regulator of osteoblastic differentiation Runt related factor-2 (Runx2), in addition to the partner protein core binding factor-β and their targets such as osteopontin (OPN) and matrix metalloproteinase-13 (MMP13), but not for collagen-I or osteocalcin. In pluripotent P19 progenitor cells, Runx2 mRNA expression was drastically increased along with mRNA expression of an astrocytic marker, but not with neuronal marker mRNA expression. Both mRNA and corresponding protein were detected for Runx2 in cultured rat neocortical astrocytes and astrocytic C6 glioma cells. In C6 glioma cells, transient overexpression of Runx2 significantly increased mRNA expression of MMP13, but not of OPN. Moreover, transient overexpression of Runx2 significantly increased luciferase activity in C6 glioma cells transfected with the reporter plasmid linked to a wild-type Runx2 binding element in the MMP13 promoter, but not in cells with a mutated element. These results suggest that Runx2 signal input may lead to transactivation of MMP13 gene without affecting OPN expression in astrocytes.

Edaravone, a Free Radical Scavenger, Inhibits MMP-9–Related Brain Hemorrhage in Rats Treated With Tissue Plasminogen Activator

Intracerebral hemorrhage, induced by recombinant tissue plasminogen activator (rtPA) in ischemic stroke, is attributable to the increased activity of matrix metalloproteinase-9 (MMP-9). Patients with acute infarct benefit from the neuroprotective drug edaravone, a free radical scavenger. We examined the mechanisms by which edaravone may help to suppress rtPA-induced brain hemorrhage. Male Wistar rats weighing 250 to 280 g were subjected to 3-hour transient middle cerebral artery occlusion (MCAO) and divided randomly into 3 groups. Immediately after reperfusion, 1 group was intravenously injected with 10 mg/kg rtPA, another with rtPA plus 3 mg/kg edaravone, and the 3rd group received no treatment. We assessed the hemorrhage volume and the activity of MMP-9 in the brain 24 hours postischemia. We also studied the activity of MMP-9, its mRNA expression, and nuclear factor-kappa B (NF-kappaB) activity in rtPA-stimulated human microvascular endothelial cells (HBECs). The degree of hemorrhage and the level of endothelial cell-derived MMP-9 were elevated in rats treated with rtPA alone and attenuated in rats treated with rtPA plus edaravone. In rtPA-stimulated HBECs, edaravone suppressed the activity and mRNA expression of MMP-9 in a dose-dependent manner. Edaravone also inhibited NF-kappaB activation. We demonstrate that edaravone inhibits rtPA-induced cerebral hemorrhage in the ischemic brain of rats via the inhibition of MMP-9 expression in vivo, which is substantiated by inhibition of MMP-9 expression and NF-kappaB activation in HBECs. Edaravone may render thrombolytic therapy safer for the administration of rtPA in patients with ischemic stroke.

Stat3 Is Involved in Angiotensin II-Induced Expression of MMP2 in Gastric Cancer Cells

The expression of matrix metalloproteinase-2 (MMP2) has been linked with tumor invasion, angiogenesis, and metastasis. It has been reported that angiotensin II (Ang II) can induce MMP2 expression in gastric cancer cells. However, the molecular basis for Ang II regulates MMP2 expression in gastric cancer cells remains unclear. The aim of our study is to explore whether angiotensin II could induce MMP2 expression mediated through the Stat signaling pathway and its potential mechanism. Electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP)-binding assays was employed to determine the DNA-STAT binding activity. MMP2 and VEGF expression was analyzed with real-time PCR and Western blots. To examine the role of Stat3 in angiotensin II-induced MMP2 expression, A JAK-specific inhibitor and AG490 were used. Angiotensin II activated STAT-DNA binding activity in dose-dependent manners in gastric cancer cells. AG490 markedly inhibited angiotensin II-induced Stat3 activation and the expression of MMP2 and VEGF in gastric cancer cells. These results indicate that Stat3 may at least in part mediate angiotensin II-induced MMP2 mRNA expression in human gastric cancer cells. The activation of the JAK/Stat3 signaling pathway plays an important role in the progression of gastric cancer and that blockade of JAK/Stat3 signals may provide a novel therapeutic strategy for gastric cancer.

Ceramide accelerates ultraviolet-induced MMP-1 expression through JAK1/STAT-1 pathway in cultured human dermal fibroblasts

Ultraviolet (UV) irradiation accelerates formation of ceramide through hydrolysis of sphingomyelin and de novo synthesis. Here, we investigated the effects of ceramide on UV-induced matrix metalloproteinase-1 (MMP-1) expression in human dermal fibroblasts. Our results showed that acidic-sphingomyelinase (aSMase) and MMP-1 mRNA expression were increased by UV irradiation. Treatment of D609 (aSMase inhibitor) decreased the level of basal and UV-induced MMP-1 expression. On the other hand, basal and UV-induced MMP-1 expression was increased through induction of intracellular ceramide by D-MAPP, a ceramidase inhibitor. Our results also showed that MMP-1 protein expression was dose-dependently increased by C(2)-ceramide or SMase treatment. The activation of ceramide pathway by C(2)-ceramide enhanced phosphorylation of signal transducer and activators of transcription-1 (STAT-1), whereas ceramide-induced MMP-1 expression was potently prevented by piceatannol; Janus kinase (JAK1) inhibtor; and WHI-P131, a specific inhibitor of JAK3; but not by AG490, JAK 2 inhbitor, in human dermal fibroblasts. We also found that UV induced the phosphorylation of STAT-1, and UV-induced MMP-1 expression was significantly decreased by JAK1 inhibitor, piceatannol. Overall, we demonstrate that induction of intracellular ceramide by UV may activate MMP-1 gene expression via JAK1/STAT-1 pathway. Therefore, we suggest that targeted modulation of the ceramide signaling pathway may offer a novel therapeutic approach for inhibiting MMP-1 expression, which is a causing gene of skin aging.

Temporal and quantitative analysis of expression of metalloproteinases (MMPs) and their endogenous inhibitors in atherosclerotic lesions.

Matrix metalloproteinases (MMPs) play an important role in the pathogenesis of vascular diseases, such as atherosclerosis, plaque rupture and aneurysms. Although several MMPs have been demonstrated in the lesions of atherosclerosis, their expression profiles during the initiation and progression of lesions have not been fully determined. We hypothesized that the expression of various MMPs, along with their endogenous inhibitors, may be differentially regulated dependent upon the lesion progression. Therefore, we made a temporal and quantitative analysis of the mRNA and protein expression of MMPs and tissue inhibitors of metalloproteinases expressed in the different stages of atherosclerotic lesions of rabbits and humans. We found that MMP-1, MMP-12 and MMP-13 expression was nearly absent in the normal arterial wall, but was remarkably increased with lesion progression. Furthermore, the expression of these MMPs in the lesions was closely associated with intimal macrophages and monocyte chemoattractant protein-1 expression, suggesting that the intimal macrophages are the major source of production of these MMPs. MMP-3 and MT1-MMP were also significantly upregulated in the early-stage lesions and fatty streaks compared to the normal aortas of rabbits. Our results indicate that MMP-1, -12, and -13 derived from intimal macrophages may play a pivotal role in both lesion initiation and progression, and therefore are potential therapeutic targets for the treatment of plaque rupture and aneurysm formation.

UV‐induced DNA damage initiates release of MMP‐1 in human skin

Destruction of collagen is a hallmark of photoaging. The major enzyme responsible for collagen 1 digestion, matrix metalloproteinase-1 (MMP-1), is induced by exposure to sunlight. To study the molecular trigger for this induction, human skin was ultraviolet-B (UVB)-irradiated and treated with liposome-encapsulated DNA repair enzymes. The photolyase-mediated DNA repair of epidermal UV damage was associated with a reduction of MMP-1 mRNA and protein expression in both the epidermal and dermal compartments of the skin. The role of the epidermal cells in MMP-1 induction in the fibroblasts was examined when human epidermal keratinocytes were irradiated with UVB and their media were transferred to unirradiated human dermal fibroblasts. Transfer of media from irradiated keratinocytes to unirradiated fibroblasts enhanced MMP-1 mRNA and protein. Thus, UV damage to keratinocytes of the epidermis may participate in the destruction of collagen in the dermis by release of soluble mediators that signal fibroblasts to release MMP-1. The MMP-1 induction was reduced when the keratinocytes were treated with DNA repair enzymes T4 endonuclease V or UV endonuclease prior to transfer of the media to fibroblasts. This implies that UVB, which deposits most of its energy on the chromatin of the epidermal keratinocytes and to a lesser extent in the upper dermis, has a significant role in photoaging. DNA damage in the keratinocytes initiates one of the signals for MMP-1 release, and enhancing DNA repair can reduce MMP-1 expression in human skin cells and tissue.

Expression profiles of matrix metalloproteinase 9 in teleost fish provide evidence for its active role in initiation and resolution of inflammation

Matrix metalloproteinase 9 (MMP-9) belongs to a family of zinc-dependent endopeptidases. As a consequence of its ability to cleave structural extracellular matrix molecules, mammalian MMP-9 is associated with vital inflammatory processes such as leucocyte migration and tissue remodelling and regeneration. Interestingly, MMP-9 genes have been identified in fish, but functional data are still limited and focus on the involvement of MMP-9 in embryonic development, reproduction and post-mortem tenderization. Here, we describe the involvement of MMP-9 in the innate immunity of carp. In carp, MMP-9 was most notably expressed in classical fish immune organs and in peritoneal and peripheral blood leucocytes, indicating a role of MMP-9 in immune responses. In our well-characterized zymosan-induced peritonitis model for carp, we analysed expression of the MMP-9 gene and the gelatinolytic levels of both pro- and activated forms of MMP-9. The biphasic profile of MMP-9 mRNA expression indicated involvement during the initial phase of inflammation and during the later phase of tissue remodelling. Also, in vitro stimulation of carp phagocytes with lipopolysaccharide or concanavalin A increased MMP-9 gene expression, with a peak at 24 hr. The increase of MMP-9 mRNA correlated with the peak of MMP-9 gelatinolytic level in culture supernatants. These results provide evidence for an evolutionarily conserved and relevant role of MMP-9 in the innate immune response.

Effects of cytomegalovirus infection on the mRNA expression of collagens and matrix metalloproteinases in gingival fibroblasts

The purpose of this in vitro investigation was to study the effects of human cytomegalovirus infection on the mRNA expression for collagens I and III and for matrix metalloproteinases 1 and 2 in gingival fibroblasts. Gingival fibroblasts were experimentally infected with the Towne strain of human cytomegalovirus and the kinetics of expression of mRNA for collagens I and III and for matrix metalloproteinases 1 and 2 was studied at different time-points. Total RNA was isolated at the indicated time, and the reverse transcription-polymerase chain reaction was used to analyze the level of mRNA expression. In addition, gingival specimens were obtained from 14 periodontitis and from three non-periodontitis subjects and mRNA analysis for collagens and metalloproteinases was carried out. Nested polymerase chain reaction was used to determine the presence or absence of human cytomegalovirus in subgingival samples from each subject. The infection of gingival fibroblasts with human cytomegalovirus during a 0-72-h period resulted in progressive reduction in the expression of mRNA for collagens I and III (p < 0.05). A higher concentration of human cytomegalovirus resulted in varying degrees of mRNA reduction, suggesting a virally mediated effect. Biopsies from human cytomegalovirus-positive individuals with periodontitis had a higher expression of mRNA for collagens I and III than biopsies from human cytomegalovirus-negative individuals. An up-regulation in the mRNA expression of matrix metalloproteinases 1 and 2 over time was observed (p < 0.05). Analysis of mRNA expression in gingival biopsies demonstrated higher expression of matrix metalloproteinase-1 in human cytomegalovirus-positive periodontitis specimens compared with human cytomegalovirus-negative periodontitis specimens. Altered expression of mRNA for collagens and metalloproteinases in human cytomegalovirus-infected gingival fibroblasts should be considered as possible modifying mechanisms in periodontitis-infected sites.