Tissue Inhibitor Of Metalloproteinase-1 mRNA Research Articles

Impact of tissue inhibitor of metalloproteinases-3 genetic variants on clinicopathological characteristics of urothelial cell carcinoma.

To investigate the distribution of single nucleotide polymorphism (SNP) of tissue inhibitor of metalloproteinases-3 (TIMP-3) in patients with/without urothelial cell carcinoma (UCC), three loci of TIMP-3 SNPs (rs9862 C/T, rs9619311 T/C, rs11547635 C/T) were genotyped via TaqMan allelic discrimination for 424 UCC patients and 848 non-UCC participants. Furthermore, the TIMP-3 mRNA expression and its correlation with clinical characters of urothelial bladder carcinoma was analyzed using The Cancer Genome Atlas database (TCGA). The distribution of all 3 studied SNPs of TIMP-3 was insignificantly different between the UCC and non-UCC groups. However, significantly lower tumor T status was found in TIMP-3 SNP rs9862 CT + TT variant than the wild type (OR: 0.515, 95% CI: 0.289-0.917, P = 0.023). Moreover, the muscle invasive tumor type was significantly correlated to the TIMP-3 SNP rs9619311 TC + CC variant in the non-smoker subgroup (OR: 2.149, 95% CI: 1.143-4.039, P = 0.016). With the TIMP-3 expression data provided in TCGA, significantly higher TIMP-3 mRNA expression was observed in UCC with high tumor stage (P < 0.0001), high tumor T status (P < 0.0001) and high lymph node status (P = 0.0005). In conclusions, TIMP-3 SNP rs9862 variant is associated with lower tumor T status of UCC while TIMP-3 SNP rs9619311 variant is correlated to muscle invasive UCC development in non-smoker.

Anti-aging Effect of Mixed Extract from Medicinal Herbs

Purpose: This study was conducted to confirm the applicability of a mixed herbal extract (MHE) as an anti-aging cosmetic ingredient by investigating its skin anti-aging activities in vitro and in vivo.Methods: In this study, we prepared MHE using an ultrasonic extraction containing Forsythiae fructus, Tribuli fructus, Solomon’s seal, Siberian ginseng, Ponciri fructus and Ginseng. We investigated the anti-aging effect of the MHE for skin in dermal fibroblasts. The anti-aging activity was determined by the type I collagen synthesis levels. Matrix metalloproteinase-1 (MMP1) and tissue inhibitor of metalloproteinases 1 (TIMP1) mRNA levels were measured by qRT-PCR. MMP1 protein levels were evaluated by blotting analysis. Clinical tests of skin moisture, elasticity, texture, and wrinkles were performed using cosmetics containing 1% MHE.Results: The MHE induced the upregulation of pro-collagen type I synthesis and TIMP1 mRNA expression. The MHE led to the downregulation of MMP1 mRNA levels and protein levels. Furthermore, after skin application of cosmetics containing 1% MHE, skin hydration, elasticity, texture, and crow’s feet were improved 4 weeks after the treatment.Conclusion: MHE has an anti-aging effect by promoting collagen synthesis and suppressing MMP1 gene expression in vitro, and it has a skin improvement effect in vivo. Therefore, the MHE was shown to have value as a functional cosmetic ingredient.

MiR-106b-5p Promotes Malignant Behaviors of Cervical Squamous Cell Carcinoma Cells by Targeting TIMP2.

The objective of this study was to investigate modulatory mechanism of miR-106b-5p and tissue inhibitor of metalloproteinases 2 (TIMP2) on cervical squamous cell carcinoma cells. Differentially expressed genes in CSCC were analyzed via bioinformatics analysis. The targeting impact of miR-106b-5p on TIMP2 was validated through dual-luciferase assay and RNA immunoprecipitation assay. MiR-106b-5p level and TIMP2 mRNA level were assessed via qRT-PCR. TIMP2 protein level was measured via western blot. Malignant behaviors of CSCC cells were evaluated by functional experiments. The EMT and apoptosis-related proteins were determined via western blot. MiR-106b-5p was noticeably elevated in CSCC cells. Its downstream target was TIMP2. MiR-106b-5p and TIMP2 levels were inversely correlated. MiR-106b-5p overexpression fostered malignant phenotypes of CSCC cells, and vice versus. TIMP2 overexpression weakened the promotive impact of forced expression of miR-106b-5p on CSCC cell growth. EMT was facilitated by forced expression of miR-106b-5p. MiR-106b-5p regulates the progression of CSCC cells via targeting TIMP2, which may provide novel value for development of therapeutic targets for CSCC.

Phenotypic and Functional Transformation in Smooth Muscle Cells Derived from a Superficial Thrombophlebitis-affected Vein Wall

Superficial thrombophlebitis (ST) is a frequent pathology, but its exact incidence remains to be determined. This study tested the hypothesis whether relationships exist among smooth muscle cells (SMCs) derived from ST, varicose great saphenous veins (VGSVs), and normal great saphenous veins (GSVs). Forty-one samples of ST, VGSVs, and GSVs were collected. SMCs were isolated and cultured. Proliferation, migration, adhesion, and senescence in SMCs from the three vein walls were compared by various methods. Bax, Bcl-2, caspase-3, matrix metalloproteinase-2 (MMP-2), MMP-9, tissue inhibitor of metalloproteinase-1 (TIMP-1), and TIMP-2 messenger RNA (mRNA) and protein expressions were detected by fluorescence quantitative PCR and Western blot. An obvious decrease in cytoskeletal filaments was observed in thrombophlebitic vascular smooth muscle cells (TVSMCs). The quantity of proliferation, migration, adhesion, and senescence in TVSMCs was significantly higher than in varicose vascular smooth muscle cells and normal vascular smooth muscle cells (NVSMCs) (all P < 0.05). Bax and caspase-3 mRNA and protein expression were decreased, while Bcl-2 mRNA and protein expression were increased in the TVSMCs compared with the varicose vascular smooth muscle cells and the NVSMCs (all P < 0.05). MMP-2, MMP-9, TIMP-1, and TIMP-2 mRNA and protein expression were significantly increased in the TVSMCs compared with the VVGSVs and the NVSMCs (all P < 0.05). SMCs derived from ST are more dedifferentiated and demonstrate increased cell proliferation, migration, adhesion, and senescence, as well as obviously decreased cytoskeletal filaments. These results suggest that the phenotypic and functional differences could be related to the presence of atrophic and hypertrophic vein segments during the disease course among SMCs derived from ST, VGSVs, and GSVs.

The pathomechanism of human myxomatous valvular degeneration at the mechanical and cellular level.

The purpose of this study was to explore the pathomechanism of human myxomatous valve degeneration by investigating changes in the phenotype of valvular cells, the metabolism of the extracellular matrix and their mechanical properties. Mitral valve specimens were harvested from patients who had undergone valve replacement, and divided into two groups: patients with a myxomatous mitral valve and a control group. Histological investigation showed that the morphology of the extracellular matrix was looser and less coordinated in myxomatous valves than in controls. α-SMA (α-smooth muscle actin) and Vimentin were positive and DNA (deoxyribonucleic acid) assay of leaflets and expression of SMemb (embryonic smooth muscle myosin heavy chain), MMP-13 (matrix Metalloproteinases-13), MMP-1 mRNA (messenger Ribonucleic Acid) of the myxomatous valves were increased while the hydroxyproline content, expression of TIMP-1 (tissue inhibitor of metalloproteinase-1) mRNA and mechanical properties were decreased compared with controls. Compared to the quiescent interstitial cells in non-myxomatous valves, interstitial cells in myxomatous valves exhibit myofibroblast activation and express excessive levels of matrix metalloproteinases. The balance between MMP/TIMP was disrupted. We conclude that overactivation of VICs (Valvular interstitial cells) and the imbalance of MMP/TIMP could be important features of the pathomechanism of myxomatous mitral valve degeneration.

Matrix metalloproteinase 7 (MMP-7) and tissue inhibitor of metalloproteinases-2 (TIMP-2) downregulation complements PALG1 oncogene overexpression in pleomorphic adenoma pathogenesis

Pleomorphic adenoma (PA) is the most common neoplasm of salivary glands and consists of epithelial and mesenchymal components. Although a benign lesion, it harbors a potential for recurrence and malignant transformation. Also, due to its histological diversity and unpredictive behavior PA can represent both diagnostic and therapeutic challenge. Matrix metalloproteinases (MMPs) are well known modifiers of extracellular matrix (ECM) PA component and in conjunction with their endogenous tissue inhibitors (TIMPs) may influence PA tumor biology. PLAG1 oncogene also has an important role in PA; however, neither the exact mechanisms of its influence nor its interactions with other genes are completely elucidated. The aims of this study were to assess the expression of PLAG1, MMP-2, MMP-7, MMP-9, TIMP-1 and TIMP-2 genes in PAs, and find a possible association of gene expression levels with clinical/epidemiological parameters of PA patients. Relative mRNA levels were assessed using Quantitative real-time PCR analyses in 15 PAs of the parotid gland and 5 normal salivary glands (NSGs). A statistically significant overexpression of PLAG1 was observed in PA compared to NSG samples (P=0.010); PA had 5.48 times higher mRNA levels than NSG. Out of the three analyzed MMP genes, significantly lower levels of MMP-7 were found in PA patients (P=0.026). TIMP2 was also downregulated in PA samples, compared to NSGs (P=0.040). MMP-7 and TIMP-2 mRNA levels were decreased 2.95 and 2.85 times respectively, in PA samples. No association was found between gene expression and clinical/epidemiological PA parameters. Our results suggest that PLAG1 overexpression with concomitant MMP7 and TIMP2 downregulation may contribute to PA development.

Leonurus sibiricus root extracts decrease airway remodeling markers expression in fibroblasts.

Bronchial asthma is believed to be provoked by the interaction between airway inflammation and remodeling. Airway remodeling is a complex and poorly understood process, and controlling it appears key for halting the progression of asthma and other obstructive lung diseases. Plants synthesize a number of valuable compounds as constitutive products and as secondary metabolites, many of which have curative properties. The aim of this study was to evaluate the anti-remodeling properties of extracts from transformed and transgenic Leonurus sibiricus roots with transformed L. sibiricus roots extract with transcriptional factor AtPAP1 overexpression (AtPAP1). Two fibroblast cell lines, Wistar Institute-38 (WI-38) and human fetal lung fibroblast (HFL1), were incubated with extracts from transformed L. sibiricus roots (TR) and roots with transcriptional factor AtPAP1 over-expression (AtPAP1 TR). Additionally, remodeling conditions were induced in the cultures with rhinovirus 16 (HRV16). The expressions of metalloproteinase 9 (MMP)-9, tissue inhibitor of metalloproteinases 1 (TIMP-1), arginase I and transforming growth factor (TGF)-β were determined by quantitative polymerase chain reaction (qPCR) and immunoblotting methods. AtPAP1 TR decreased arginase I and MMP-9 expression with no effect on TIMP-1 or TGF-β mRNA expression. This extract also inhibited HRV16-induced expression of arginase I, MMP-9 and TGF-β in both cell lines (P<0·05) Our study shows for the first time to our knowledge, that transformed AtPAP1 TR extract from L. sibiricus root may affect the remodeling process. Its effect can be attributed an increased amount of phenolic acids such as:chlorogenic acid, caffeic acid or ferulic acid anddemonstrates the value of biotechnology in medicinal research.

Biology of Tissue Inhibitor of Metalloproteinase 3 (TIMP3), and Its Therapeutic Implications in Cardiovascular Pathology.

Tissue inhibitor of metalloproteinase 3 (TIMP3) is unique among the four TIMPs due to its extracellular matrix (ECM)-binding property and broad range of inhibitory substrates that includes matrix metalloproteinases (MMPs), a disintegrin and metalloproteinases (ADAMs), and ADAM with thrombospondin motifs (ADAMTSs). In addition to its metalloproteinase-inhibitory function, TIMP3 can interact with proteins in the extracellular space resulting in its multifarious functions. TIMP3 mRNA has a long 3’ untranslated region (UTR) which is a target for numerous microRNAs. TIMP3 levels are reduced in various cardiovascular diseases, and studies have shown that TIMP3 replenishment ameliorates the disease, suggesting a therapeutic potential for TIMP3 in cardiovascular diseases. While significant efforts have been made in identifying the effector targets of TIMP3, the regulatory mechanism for the expression of this multi-functional TIMP has been less explored. Here, we provide an overview of TIMP3 gene structure, transcriptional and post-transcriptional regulators (transcription factors and microRNAs), protein structure and partners, its role in cardiovascular pathology and its application as therapy, while also drawing reference from TIMP3 function in other diseases.

Neuroprotective effect of neuregulin-1β on spinal cord ischemia reperfusion injury

Objective: This study was designed to see if neuregulin-1β (NRG-1β) plays a protective role in spinal cord ischemia and reperfusion injury (SCII). Design: Animal research. Setting: China. Participants: NA. Interventions: Forty-eight SD rats were randomly divided into control group (n = 16), SCII model group (n = 16) and NRG-1β-treated group (n = 16). In control group, the abdominal aorta was isolated but not clipped. The rats in NRG-1β-treated group were treated with 10μg/kg NRG-1β during developing SCII model. Outcome Measures: Neurological scores were evaluated. At 3, 6, 12 and 24 h after the reperfusion, rats were killed. Pathological changes of spinal cord were assessed with HE staining, and immunohistochemical staining of matrix metalloproteinases-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-1). MMP-9 and TIMP-1 mRNA levels were assessed using real-time PCR. Results: NRG-1β reduced the damage of SCII in the rats. The expression of MMP-9 protein and mRNA in NRG-1β treatment group was significantly lower than the model group (P < 0.05) at 6 h, 12 h and 24 h after the perfusion. The expression of TIMP-1 protein and mRNA in the treatment group was significantly higher than the model group at 12 h and 24 h after the perfusion. Conclusion: NRG-1β reduced the reperfusion damage in rat model of SCII, in which process MMP-9 and TIMP-1 were probably involved.

Kupffer-derived matrix metalloproteinase-9 contributes to liver fibrosis resolution.

Kupffer cells (KCs) contribute to liver fibrosis resolution by production of a large spectrum of matrix metalloproteinases (MMPs). MMP9 is a major MMP expressed by KCs. However, its role in liver fibrosis resolution remains unclear. In this study, rodent liver fibrosis was induced by intraperitoneal thioacetamide (TAA) and the resolution process was initiated by TAA withdrawal. The role of KC-derived MMP9 in fibrolysis was investigated by adoptive transfer of KCs with or without MMP9 following their depletion. The levels of serum alanine aminotransferase (ALT) and hepatic cytokines were measured during fibrosis regression. The mRNA levels of MMPs and tissue inhibitor of metalloproteinases (TIMPs) were analyzed as well. It was found that removing KCs delayed fibrosis resolution. Adoptive transfer of KCs from WT animals promoted liver fibrosis resolution, compared with transfer of KCs from MMP9-/- mice. Depletion of KCs also resulted in prolonged liver wound healing, which was reversed partially by transferred KCs from either WT or MMP9-/- mice. Likewise, the absence of KCs led to reduction in MMPs mRNA levels and elevation in TIMPs mRNA levels. The expression patterns of MMPs or TIMPs were restored by adoptive transfer of the wild-type but not MMP9-/- KCs. In addition, liver fibrosis resolution was accelerated in MMP9-/- mice by adoptive transferred KCs from WT animals, compared to the KCs from MMP9-/- mice. Overall, KC-derived MMP9 plays a critical role in fibrosis resolution, which might serve as the foundation for developing anti-fibrosis therapy.

Expression and role of microRNA-1271 in the pathogenesis of osteosarcoma.

The aim of the current study was to investigate the expression and role of microRNA (miR)-1271 in the pathogenesis of osteosarcoma, and the associated underlying mechanisms. Tissue samples from 45 patients with osteosarcoma were collected, while the 143B, MG-63 and U-2 OS osteosarcoma cell lines were also cultured. The expression levels of miR-1271 in the tissues and cells were detected with reverse transcription-quantitative polymerase chain reaction, and 143B osteosarcoma cells were subjected to miR-1271 manipulation. In addition, the cell proliferation, cell cycle progression, and migration and invasion abilities were assessed by Cell Counting Kit-8 assay, flow cytometry and Transwell chamber assay, respectively. Tissue inhibitor of metalloproteinases 2 (TIMP2) expression level was also detected with western blot analysis. Dual-luciferase reporter assay was performed to investigate the interaction between miR-1271 and TIMP2. The results revealed that miR-1271 expression was significantly elevated in the osteosarcoma tissue and was closely correlated with the clinical TNM staging. The expression levels of miR-1271 were also upregulated in the osteosarcoma cells, with the highest expression observed in 143B cells. Inhibition of miR-1271 significantly inhibited the cell proliferation, G1/S phase transition, and the migration and invasion abilities of 143B cells, while it also resulted in upregulated TIMP2 expression in these cells. Furthermore, overexpression of TIMP2 significantly inhibited the cell proliferation, G1/S phase transition, and migration and invasion abilities of 143B cells. Dual-luciferase reporter assay demonstrated that miR-1271 targeted on the 3′-untranslated region of TIMP2 mRNA. In conclusion, the expression levels of miR-1271 were significantly elevated in osteosarcoma tissues and cells. miR-1271 downregulated the expression of TIMP2 to promote the proliferation and enhance the migration and invasion abilities of 143B osteosarcoma cells, functioning as an oncogene.

Modified Xiaochaihu Decoction () Promotes Collagen Degradation and Inhibits Pancreatic Fibrosis in Chronic Pancreatitis Rats.

To investigate the effect of Modified Xiaochaihu Decoction (MXD, ) on collagen degradation in rats with chronic pancreatitis (CP). Rats were injected dibutyltin dichloride (DBTC, 7 mg/kg of body weight) into the right caudal vein to induce CP model. Thirty heallhy male Wistar rats were randomly divided into three groups by a random number table: the control, the model and the treatment groups. Rats of treatment group were administered MXD (10 g/kg of body weight) orally once daily starting from the day post-model establishment. Pancreatic tissues were harvested after 28-day feeding and fibrosis was evaluated by picro-sirius red staining. The contents of collagen type I and III were detected using enzymelinked immunosorbent assay (ELISA), the expression of matrix metalloproteinase 13 (MMP13) and tissue inhibitor of metalloproteinase 1 (TIMP1) was analyzed by Western blot and real-time polymerase chain reaction (PCR). The fibrosis scoring of pancreatic tissues, the concentrations of collagen type I and III, the expression levels of MMP13 and TIMP1 proteins and mRNA in the model group were all increased compared with the control group (P<0.05). After treatment with MXD, the fibrosis scoring of pancreatic tissues, the concentrations of collagen type I and III, the expression levels of MMP13 proteins and mRNA in the teatment group were all decreased compared with the model group (P<0.05), but there were no significant differences in the expression levels of TIMP1 proteins and mRNA (P>0.05). MXD could promote collagen degradation and reverse pancreatic fibrosis in CP rats via a mechanism involve up-regulation of MMP13 expression.

The urine biomarker panel [IGFBP7]x[TIMP-2] (NephroCheck® parameter) does not correlate with IGFBP7 and TIMP-2 gene expression in urinary sediment.

BackgroundAcute kidney injury (AKI) is frequently observed in serious infections, following nephrotoxic medication, surgery and trauma. Here we tested whether the detection of two recently identified biomarkers for AKI, Tissue Inhibitor of Metalloproteinase-2 (TIMP-2) and Insulin-Like Growth Factor Binding Protein 7 (IGFBP7), depends on the expression of these proteins in cells of the urinary sediment.MethodWe collected urine samples of 33 kidney transplant recipients and 14 non-transplanted patients who all had AKI (stages 1–3 according to KDIGO), and measured [IGFBP7]x[TIMP-2] using the NephroCheck® Astute1 40 ™ meter. Concomitantly, we analyzed IGFBP7 and TIMP-2 mRNA expression by quantitative polymerase chain reaction (qPCR) from urinary sediment of the same patients, and correlated the results with [IGFBP7]x[TIMP-2] (protein), by linear regression analysis. We also determined the association between [IGFBP7]x[TIMP-2] and estimated glomerular filtration rate (eGFR), and between IGFBP7 and TIMP-2 mRNA expression and markers of inflammation. Light microscopy and confocal immunofluorescence served to illustrate changes in the urinary sediment over the time course of renal function improvement.ResultsOf the 47 analyzed AKI patients, 14 presented with ascending urinary tract infection. Serum creatinine (sCr), blood urea nitrogen (BUN) and eGFR in all patients were 3.9±2.28 mg/dL, 47.59±23.1 mg/dL and 22.88±16.0 mL/min/1.73m2, respectively, on average ±standard deviation. [IGFBP7]x[TIMP-2] was 2.33±9.95 (ng/ml)2/1000, and did not associate with IGFBP7 and TIMP-2 gene expression (r = -0.0220, p = 0.4216; respectively r = 0.0972, p = 0.1909). [IGFBP7]x[TIMP-2] did not associate with eGFR; IGFBP7 and TIMP-2 mRNA expression. Improvement of renal function went along with disappearance of casts, decrease in aquaporin1 positive renal epithelial cells and leukocytes from the urinary sediment.ConclusionThe gene expression pattern of IGFBP7 and TIMP-2 from urinary sediment, which contains desquamated renal tubular epithelial cells, did not correlate with [IGFBP7]x[TIMP-2] protein, indicating that IGFBP7 and TIMP-2 measured in the NephroCheck® test originated predominantly from intact but stressed cells of the kidney itself.

Protective effect of glucosamine cyclohexyl ester on osteoarthritis in rat via targeting expressions of matrix metalloproteinase and tissue inhibitor of metalloproteinases-1

Purpose: To investigate the therapeutic effect of glucosamine cyclohexyl ester on osteoarthritis (OA) in a rat model.Methods: Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot assays were used to analyze the effect of glucosamine cyclohexyl ester on changes in mRNA and protein expressions of matrix metalloproteinase and tissue inhibitor of metalloproteinases-1 in isolated rat chondrocytes, and in a rat model of OA. The rat model of OA was prepared by injecting monoiodoacetate to Sprague-Dawley rats via intra-articular route.Results: Treatment of the chondrocytes with glucosamine cyclohexyl ester for 48 h prevented interleukin-1 β (IL-1β)-mediated increases in mRNA and protein expressions in matrix metalloproteinases-1, -3 and -13, and also blocked IL-1β-induced decreases in mRNA and protein expressions of tissue inhibitor of metalloproteinase-1. Glucosamine cyclohexyl ester treatment also blocked the onset of morphological changes such as irregular surface, adhesion of tissues andpresence of osteophytes in the femoral condyle surface of the OA rats. Mankin score for control, OA and glucosamine cyclohexyl ester treatment groups were 0.98 ± 0.15, 8.35 ± 0.88 and 2.39 ± 0. 67 (p = 0.002), respectively. Treatment of OA rats with glucosamine cyclohexyl ester also inhibited increases in the activities of matrix metalloproteinases-1, -3 and -13, and decreases of tissue inhibitor of metalloproteinase-1 mRNA and protein expressions. Treatment of chondrocytes and OA rats with IL-1β caused no significant changes in the levels of H3K27 and H4K8.Conclusion: These results show that glucosamine cyclohexyl ester prevents OA by targeting the expressions of matrix metalloproteinases-1, -3 and -13 and tissue inhibitor of metalloproteinases-1.Keywords: Metalloproteinases, Interleukin, Mankin score, Osteoarthritis, Cartilage

Reduced collagen accumulation and augmented MMP-2 activity in left ventricle of old rats submitted to high-intensity resistance training.

Progressive fibrosis is a hallmark of the aging heart. Age-related fibrosis is modulated by endurance exercise training; however, little is known concerning the influence of resistance training (RT). Therefore we investigated the chronic effects of high-intensity RT on age-associated alterations of left ventricle (LV) structure, collagen content, matrix metalloproteinase-2 (MMP-2), and extracellular matrix-related gene expression, including transforming growth factor-β (TGF-β). Young adult (3 mo) and aged (21 mo) male Wistar rats were submitted to a RT protocol (ladder climbing with 65, 85, 95, and 100% load), three times a week for 12 wk. Forty-eight hours posttraining, arterial systolic and diastolic pressure, LV end-diastolic pressure (LVEDP) and dP/dt were recorded. LV morphology, collagen deposition, and gene expression of type I (COL-I) and type III (COL-III) collagen, MMP-2, tissue inhibitor of metalloproteinases-1 (TIMP-1), and TGF-β1 were analyzed by quantitative reverse transcriptase-PCR. MMP-2 content was assessed by zymography. Increased collagen deposition was observed in LV from aged rats. These parameters were modulated by RT and were associated with increased MMP-2 activity and decreased COL-I, TGF-β1, and TIMP-1 mRNA content. Despite the effect of RT on collagen accumulation, there was no improvement on LVEDP and maximal negative LV dP/dt of aged rats. Cardiomyocyte diameter was preserved in all experimental conditions. In conclusion, RT attenuated age-associated collagen accumulation, concomitant to the increase in MMP-2 activity and decreased expression of COL-I, TGF-β1, and TIMP-1 in LV, illustrating a cardioprotective effect of RT on ventricular structure and function.NEW & NOTEWORTHY We demonstrated the beneficial resistance-training effect against age-related left ventricle collagen accumulation in the left ventricle, which was associated with decreased type I collagen (COL-I), transforming growth factor-β1 (TGF-β1), and tissue inhibitor of metalloproteinases-1 (TIMP-1) gene expression and matrix metalloproteinase-2 (MMP-2) activity. Our findings suggest for the first time the potential effects of resistance training in modulating collagen accumulation and possibly fibrosis in the aging heart.

Abstract 168: Loss of function mutation in TIMP2 gene accelerates tumorigenesis and mortality in murine model of lung cancer through EGFR signaling

Abstract The aim of this study is to determine the effect of the tissue inhibitor of metalloproteinase-2 (TIMP-2) on lung tumorigenesis and investigate the underlying molecular mechanisms using an orthotopic mouse model with a loss of function mutation in the Timp2 gene (T2M). T2M and wildtype (WT) control mice were given 1x106 Lewis lung carcinoma cells transfected with luciferase (LL/2-Luc-M38, Caliper) in 50μL PBS via intratracheal installation. IVIS imaging revealed a higher tumor burden in T2M mice compared to WT littermates, suggesting that loss of function of the Timp2 gene enhances tumor growth (p&amp;lt;0.05). We also conducted a Kaplan-Meier analysis to determine the effect of this mutation on mortality following cancer development. We found that LL/2 tumor-induced mortality was significantly higher in T2M mice compared to WT controls (p = 0.01). Histologic analysis and H&amp;E staining of lung tissue sections revealed a significant increase in the number of tumor nodules of T2M mice compared to WT controls (p &amp;lt;0.01). CD31 staining showed a significant increase in microvessel density (MVD) in T2M mice compared to WT controls (p&amp;lt;0.01). Given that VEGF is a primary driver of tumor neovascularization, we determined mRNA levels of VEGF expression in healthy and tumor bearing mice by qRTPCR analysis. Interestingly, basal VEGF expression levels were increased in lungs of both non-tumor bearing and tumor bearing T2M mice compared to WT controls (p&amp;lt;0.01). VEGF is the direct downstream target of HIF-2α, a transcription factor implicated in tumor hypoxia and expression correlates with decreased overall survival in non-small cell lung cancer (NSCLC) patients. Similarly, an examination of TIMP-2 and HIF-2α mRNA expression in a small cohort of patient samples revealed decreased TIMP-2 and increased HIF2α in NSCLC tumors compared to surrounding non-tumor lung tissue. Hypoxia induces EGFR signaling through HIF-2α. Thus, we determined the effect of mutated T2M on the EGFR signaling pathway. We found increased levels of EGFR phosphorylation as well as downstream ERK and Akt activation in tumor bearing and non-tumor bearing T2M lungs compared to WT controls. In conclusion, these findings offer new avenues for TIMP-2 research in regulating hypoxic mediators within the tumor microenvironment through EGFR signaling, suggesting TIMP-2 as a novel bio-therapeutic for lung cancer therapy. * Corresponding author This work is supported by center for cancer research, intramural research program, NCI/NIH research grants ZIA BC011204 &amp; ZIA SC 009179 to WGSS Citation Format: Sarvesh Kumar, Sandra Jensen, David Peeney, Ananda Chowdhury, Beiyang Wei, William G. Stetler-Stevenson. Loss of function mutation in TIMP2 gene accelerates tumorigenesis and mortality in murine model of lung cancer through EGFR signaling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 168. doi:10.1158/1538-7445.AM2017-168

Protective effect of Shenmai injection on knee articular cartilage of osteoarthritic rabbits and IL-1β-stimulated human chondrocytes.

Shenmai injection (SMI) has been widely used as a therapy to treat a number of diseases. However, its anti-osteoarthritic properties have not yet been fully investigated. In the present study, the protective effect of SMI on knee articular cartilage of anterior cruciate ligament transected rabbits and interleukin-1β (IL-1β)-stimulated human chondrocytes was investigated. For the in vivo study, knee osteoarthritis (KOA) was induced in female New Zealand white rabbits by anterior cruciate ligament transection (ACLT) in the knee of right hind limb. Rabbits either underwent sham surgery or ACLT surgery. Out of the rabbits receiving ACLT surgery, half of the rabbits received one 0.3 ml Shenmai intra-articular injection in the knee per week for four weeks, following ACLT surgery. The other rabbits received the same volume of normal saline solution. The cartilage was subsequently collected for histological evaluation. For the in vitro study, cultured human chondrocytes were treated with 10 ng/ml IL-1β in the presence or absence of 5 and 2% (v/v) SMI for 24 h. Nitric oxide (NO) and prostaglandin E2 (PGE2) levels in cell culture supernatant were assessed using a Griess reaction and ELISA respectively. The mRNA expression of cyclooxgenase-2 (COX-2), inducible nitric oxide synthase (iNOS), matrix metalloproteinase (MMP)-1, MMP-13 and tissue inhibitors of metalloproteinase-1 (TIMP-1) in chondrocytes were detected by reverse transcription-quantitative polymerase chain reaction. The results of the current study revealed that treatment with SMI ameliorated cartilage degradation in the ACLT rabbit model, and decreased levels of NO and PGE2. Furthermore, treatment with SMI decreased levels of COX-2, iNOS, MMP-1 and MMP-13 mRNA expression and increased TIMP-1 mRNA expression in IL-1β-stimulated human chondrocytes. These results indicate that SMI suppresses inflammation and ameliorated cartilage degradation, making it a potential and promising therapeutic option to treat KOA.

Reduced Scleral TIMP-2 Expression Is Associated With Myopia Development: TIMP-2 Supplementation Stabilizes Scleral Biomarkers of Myopia and Limits Myopia Development.

The purpose of this study was to determine the endogenous regulation pattern of tissue inhibitor of metalloproteinase-2 (TIMP-2) in the tree shrew sclera during myopia development and investigate the capacity of exogenous TIMP-2 to inhibit matrix metalloproteinase-2 (MMP-2) in vitro and both scleral collagen degradation and myopia development in vivo. TIMP-2 expression in the sclera during myopia development was assessed using polymerase chain reaction. In vitro TIMP-2 inhibition of MMP-2 was investigated using a gelatinase activity plate assay and zymography. Tree shrews were injected with a collagen precursor before undergoing monocular form deprivation and concurrent daily subconjunctival injections of either TIMP-2 or vehicle to the form-deprived eye. In vivo ocular biometry changes were monitored, and scleral tissue was collected after 12 days and assayed for collagen degradation. The development of myopia was associated with a mean reduction in TIMP-2 mRNA expression after 5 days of form deprivation (P < 0.01). Both activation and activity of MMP-2 were inhibited by TIMP-2 with an IC50 of 10 to 20 and 2 nM, respectively. In vivo exogenous addition of TIMP-2 significantly reduced myopia development (P < 0.01), due to reduced vitreous chamber elongation (P < 0.01). In vivo TIMP-2 treatment also significantly inhibited posterior scleral collagen degradation relative to vehicle-treated eyes (P < 0.01), with levels similar to those in control eyes. Myopia development in mammals is associated with reduced expression of TIMP-2, which contributes to increased degradative activity in the sclera. It follows that replenishment of this TIMP-2 significantly reduced the rate of both scleral collagen degradation and myopia development.

Correlation between the mRNA expression of tissue inhibitor of metalloproteinase-1 and apparent diffusion coefficient on diffusion-weighted imaging in rats' liver fibrosis

To explore the correlation between the apparent diffusion coefficient (ADC) and mRNA expression of tissue inhibitor of metalloproteinase-1 (TIMP-1) in different stages of liver fibrosis in rats. Methods: A model of liver fibrosis in rats was established by intraperitoneal injection of high-fat diet combined with porcine serum. After drug administration for 4 weeks, 48 rats served as a model group and 12 rats served as a control group, then they underwent diffusion weighted imaging (DWI) scanning. The value of ADC was calculated at b value=800 s/mm2. The rats were sacrificed and carried out pathologic examination after DWI scanning immediately. The mRNA expression of TIMP-1 was detected by real time-polymerase chain reaction (RT-PCR). The rats of hepatic fibrosis were also divided into a S0 group (n=4), a S1 group (n=11), a S2 group (n=12), a S3 group (n=10), and a S4 group (n=9) according to their pathological stage. The value of ADC and the expression of TIMP-1 mRNA among the different stage groups of liver fibrosis were compared, and the correlation between ADC and the TIMP-1 mRNA were analyzed. Results: The ADC value and the TIMP-1 mRNA expression were significantly different between the control group and the liver fibrosis group (F=46.54 and 53.87, P<0.05). There were significant differences in the value of ADC between every two groups (all P<0.05), except the control group vs the S1 group, the S1 group vs the S2 group, and the S2 group vs the S3 group (all P>0.05). For the comparison of TIMP-1 mRNA, there was no significant difference between the S1 group and the S2 group, the S3 group and the S4 group (both P>0.05). There were significant differences among the rest of the groups (all P<0.05). Rank correlation analysis showed that there was a negative correlation between the ADC value and the TIMP-1 mRNA expression (r=-0.76, P<0.01). Conclusion: When the value of ADC decreases in the progress of rats' liver fibrosis, the mRNA expression of TIMP-1 increases gradually, and there is a negative correlation between them.

Plasma levels of the tissue inhibitor matrix metalloproteinase-3 as a potential biomarker in oral cancer progression.

Oral cancer is the most common malignancy with poor prognosis and is the fourth most common cancer in men in Taiwan. The tissue inhibitor of metalloproteinase-3 (TIMP3) acts as a tumor suppressor gene by inhibiting the growth, angiogenesis, migration, and invasion of cancer cells. However, few studies have examined the association of plasma TIMP3 levels with oral squamous cell carcinoma (OSCC), and the role of plasma TIMP3 levels in OSCC progression is still unclear. We measured the plasma TIMP3 levels of 450 OSCC patients and 64 healthy controls by using a commercial enzyme-linked immunosorbent assay. We also analyzed TIMP3 mRNA levels of 328 OSCC patients and 32 normal tissues from The Cancer Genome Atlas (TCGA) dataset. Our results revealed that plasma TIMP3 levels were significantly lower in patients with OSCC than in healthy controls (p < 0.001). Moreover, plasma TIMP3 levels in patients with OSCC were significantly associated with the tumor stage and tumor status but not with the lymph node status, metastasis, and cell differentiation. To verify our findings, we also examined TCGA bioinformatics database and discovered similar results for the association with the pathological stage of OSCC. In conclusion, our results suggest that plasma TIMP3 is a potential biomarker for predicting the tumor stage and T status in patients with OSCC.