The Possible Effect of ß-Blocker Use on the Circulating MMP-2/TIMP-2 System in Patients with Chronic Kidney Disease on Conservative Treatment.

Background: The purpose of the study was to determine whether the use of ß-adrenoceptor antagonists (ß-blockers) can affect metalloproteinase 2 (MMP-2) and its tissue inhibitor (TIMP-2) in patients with chronic kidney disease (CKD) on conservative treatment. Methods: The circulating MMP-2/TIMP-2 system, proinflammatory cytokines (tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6), and the marker of oxidative stress-Cu/Zn superoxide dismutase (Cu/Zn SOD)-were measured in 23 CKD patients treated with ß-blockers [ß-blockers (+)] and in 27 CKD patients not receiving the above medication [ß-blockers (-)]. Results: The levels of MMP-2, TIMP-2, and IL-6 were significantly lower in the ß-blockers (+) than in the ß-blockers (-) group, whereas Cu/Zn SOD concentrations were not affected by ß-blocker use. There was a strong, independent association between MMP-2 and TIMP-2 in both analyzed patient groups. In the ß-blockers (+) group, MMP-2 levels were indirectly related to the signs of inflammation, whereas in the ß-blockers (-) group, the alterations in the MMP-2/TIMP-2 system were associated with the oxidative stress marker and CKD etiology. Conclusions: This study is the first to suggest that the use of ß-blockers was associated with the reduction in IL-6 and the MMP-2/TIMP-2 system in CKD, providing a pharmacological rationale for the use of ß-blockers to reduce inflammation and abnormal vascular remodeling in CKD.

TIMP-3 Alleviates White Matter Injury After Subarachnoid Hemorrhage in Mice by Promoting Oligodendrocyte Precursor Cell Maturation.

Subarachnoid hemorrhage (SAH) is associated with high mortality and disability rates, and secondary white matter injury is an important cause of poor prognosis. However, whether brain capillary pericytes can directly affect the differentiation and maturation of oligodendrocyte precursor cells (OPCs) and subsequently affect white matter injury repair has still been revealed. This study was designed to investigate the effect of tissue inhibitor of metalloproteinase-3 (TIMP-3) for OPC differentiation and maturation. PDGFRßret/ret and wild-type C57B6J male mice were used to construct a mouse model of SAH via endovascular perforation in this study. Mice were also treated with vehicle, TIMP-3 RNAi or TIMP-3 RNAi + TIMP-3 after SAH. The effect of TIMP-3 on the differentiation and maturation of OPCs was determined using behavioral score, ELISA, transmission electron microscopy, immunofluorescence staining and cell culture. We found that TIMP-3 was secreted mainly by pericytes and that SAH and TIMP-3 RNAi caused a significant decrease in the TIMP-3 content, reaching a nadir at 24 h, followed by gradual recovery. In vitro, the myelin basic protein content of oligodendrocytes after oxyhemoglobin treatment was increased by TIMP-3 overexpression. The data indicates TIMP-3 could promote the differentiation and maturation of OPCs and subsequently improve neurological outcomes after SAH. Therefore, TIMP-3 could be beneficial for repair after white matter injury and could be a potential therapeutic target in SAH.

GnT-V-mediated aberrant N-glycosylation of TIMP-1 promotes diabetic retinopathy progression.

BACKGROUND: Vascular endothelial growth factor (VEGF) signaling pathway plays an important role in the progression of diabetic retinopathy (DR). The glycosylation modification process of many key functional proteins in DR patients is abnormal. However, the potential involvement of abnormal N-glycoproteins in DR progression remains unclear. METHODS: Glycoproteomic profiling of the vitreous humor was performed. The level of protein and N-glycoprotein was confirmed by Western blot and Lectin blot, respectively. The cell viability and migration efficiency were detected by CCK-8 and Transwell assay. Flow cytometry was conducted to analyze the level of cell apoptosis and reactive oxygen specie. Malondialdehyde, superoxide dismutase activity and VEGF content were detected by Enzyme linked immunosorbent assays. The interaction of metalloproteinase 1 (TIMP-1) with N-acetylglucosamine transferase V (GnT-V) was detected by GST pull-down. Hematoxylin and eosin staining and choroidal and retinal flat mount stained with fluorescein isothiocyanate-Dextran assay were used for functional research in vivo. RESULTS: We found that N-glycosylation was up-regulated in DR rats and high glucose (HG)-induced human retinal pigment epithelium cell line ARPE-19. HG-induced inhibited the viability of ARPE-19 cells and promoted cell apoptosis and oxidative stress (OS), but these effects were reversed with kifunensine treatment, GnT-V knockdown and TIMP-1 mutation. Additionally, GnT-V binds to TIMP-1 to promote N-glycosylation of TIMP-1. Over-expression of GnT-V inhibited the viability of ARPE-19 cells and promoted cell apoptosis, OS and VEGF release, which these effects were reversed with TIMP-1 mutation. Interestingly, over-expression of GnT-V promoted retinal microvascular endothelial cells (RMECs) angiogenesis but was revered with TIMP-1 mutation, which was terminally boosted by VEGF-A treatment. Finally, knockdown of GnT-V relieved DR progression. CONCLUSION: The findings indicate that GnT-V can promote RMECs angiogenesis and ARPE-19 cells injury through activation VEGF signaling pathway by increasing TIMP-1 N-glycosylation level, which provides a new theoretical basis for the prevention of DR.

The association between activation of the ERK1/2-NF-κB signaling pathway by TIMP2 expression and chronic renal allograft dysfunction in the CRAD rat model.

PURPOSE: The tissue inhibitor of metalloproteinase 2 (TIMP2), a natural inhibitor of matrix metalloproteinase (MMP), regulates inflammation, fibrosis, and cell proliferation. Chronic renal allograft dysfunction (CRAD) is a primary factor affecting the long-term survival of renal allografts. We assessed whether up-regulation of TIMP2 expression may affect the ERK1/2-NF-κB signaling pathway and CRAD development. METHODS: Lewis rats received orthotopic F344 kidney allografts to establish the classical CRAD model. The treatment group was injected with a lentivirus encoding a TIMP2-targeting small hairpin (sh)RNA (LTS) at 5 × 108 TU/ml monthly after kidney transplantation. A second CRAD group was injected with a lentivirus TIMP2-control vector (LTC). After 12 weeks, blood, urine, and kidney tissue were harvested to evaluate renal function and pathological examinations. Hematoxylin and eosin staining, Masson staining, and Periodic acid-Schiff staining were performed for renal histopathological evaluation according to the Banff criteria. TIMP2, phospho (p)-ERK1/2, p-p65 (NF-κB) expression levels were measured via immunohistochemical and Western blot analyses. RESULTS: Compared to the F344 and Lewis control groups, the expression of TIMP2, p-ERK1/2, and p-p65 were significantly higher in the CRAD and CRAD+LTC renal tissues (p < 0.05). There were also increased levels of serum creatinine, nitrogen, and 24 h urinary protein in these two groups (p < 0.05). Typical histopathological changes of CRAD were observed in the CRAD and CRAD+LTC groups. Administration of LTS effectively decreased the expression of TIMP2, p-ERK1/2, and p-P65, and reduced interstitial fibrosis and macrophage infiltration in the treatment group (p < 0.05). Additionally, MCP1 and ICAM-1, which are downstream cytokines of the NF-κB pathway, were also inhibited in the renal rat kidney from the LTS group (p < 0.05). Furthermore, renal function was well preserved in the LTS group compared to the CRAD group and CRAD+LTC group. CONCLUSION: A decrease of TIMP2 can alleviate the progression of inflammation in CRAD via inhibition of the ERK1/2-NF-κB signaling pathway.

Detrimental Effects of ApoE ε4 on Blood-Brain Barrier Integrity and Their Potential Implications on the Pathogenesis of Alzheimer's Disease.

Alzheimer's disease (AD) is a progressive neurodegenerative disease representing the most common type of dementia in older adults. The major risk factors include increased age, genetic predisposition and socioeconomic factors. Among the genetic factors, the apolipoprotein E (ApoE) ε4 allele poses the greatest risk. Growing evidence suggests that cerebrovascular dysfunctions, including blood-brain barrier (BBB) leakage, are also linked to AD pathology. Within the scope of this paper, we, therefore, look upon the relationship between ApoE, BBB integrity and AD. In doing so, both brain-derived and peripheral ApoE will be considered. Despite the considerable evidence for the involvement of brain-derived ApoE ε4 in AD, information about the effect of peripheral ApoE ε4 on the central nervous system is scarce. However, a recent study demonstrated that peripheral ApoE ε4 might be sufficient to impair brain functions and aggravate amyloid-beta pathogenesis independent from brain-based ApoE ε4 expression. Building upon recent literature, we provide an insight into the latest research that has enhanced the understanding of how ApoE ε4, secreted either in the brain or the periphery, influences BBB integrity and consequently affects AD pathogenesis. Subsequently, we propose a pathway model based on current literature and discuss future research perspectives.

Targeting multiple disease hallmarks using a synergistic disease-modifying drug combination ameliorates osteoarthritis via inhibition of senescence and inflammation.

AIMS: Osteoarthritis (OA), is a debilitating disease characterized by progressive cartilage degradation, synovial inflammation, and chondrocyte senescence. Various treatment agents independently targeting these hallmarks have been investigated. However, due to the complex multifaceted nature of OA, no disease-modifying osteoarthritis drugs are clinically available. In an attempt to overcome this, we developed a combinatorial approach and demonstrated the efficacy of TsC [Tissue inhibitor of metalloproteinase-3 (TIMP3) + sulfated carboxymethylcellulose (sCMC)] and piperlongumine (PL) combination for the amelioration of OA in a goat ex vivo OA model. MAIN METHODS: The efficacy of the drug combination was evaluated using the goat ex vivo OA explant model and results were validated in clinically relevant human OA cartilage explants. The chondroprotective effects were evaluated in terms of reduced inflammation and cartilage matrix loss, reduction in chondrosenescence, and reduced oxidative stress. KEY FINDINGS: A combination of TsC and PL (TsC-PL) significantly reduced inflammation, cartilage matrix loss, chondrosenescence, and oxidative stress in the goat ex vivo OA model and showed chondroprotective effects. Further, similar chondroprotective effects were observed in human OA cartilage. Additionally, the coefficient of drug interaction analysis indicated that the combination of TsC and PL had a synergistic effect in reducing matrix degrading proteases and inflammation (goat ex vivo OA model) and Reactive oxygen species (ROS) production (human OA cartilage). SIGNIFICANCE: Combinatorial treatment with TsC and PL demonstrated potential disease-modifying effects for the treatment of osteoarthritis via inhibition of inflammation and senescence and supports the usage of treatment strategies targeting multiple pathological factors of OA simultaneously.

Effect of eucalyptol on matrix metalloproteinase-9 and its tissue inhibitor in hypertensive rats.

Effects of eucalyptol, a key component of eucalyptus globules, on matrix metalloproteinase-9 (MMP-9) and its tissue inhibitor (TIMP-1), compared with lisinopril, were investigated in a model of hypertension induced by chronic intraperitoneal (IP) injection of low dose nicotine in rats. The hypertensive rats were randomly allocated to 4 groups (n=8): Positive control (PC, untreated), eucalyptol-treated group (1.0 mg/kg, IP), lisinopril-treated group (10 mg/kg, IP), and eucalyptol+lisinopril-treated group. Systolic blood pressure and plasma levels of MMP-9 and TIMP-1 were measured. All treatments decreased the elevated blood pressure and plasma levels of MMP-9 and TIMP-1 most significantly with the combination group which showed non-significant differences from the normal control group. Lisinopril reduced plasma levels of MMP-9 and TIMP-1 more significantly than eucalyptol. In conclusion, eucalyptol significantly decreased the increased plasma levels of MMP-9 and TIMP-1 in nicotine-induced hypertension in rats. Moreover, its combination with lisinopril exerted more significant effects compared to each drug alone. This makes this combination particularly useful in hypertension and related cardiovascular disorders where suppression of MMP-9 and TIMP-1 activities decreases the related complications and improves the overall morbidity and mortality. To our knowledge, the current data are novel, and may open the way for development of a co-delivery system of both drugs which could be beneficial in treatment of hypertension in chronic smokers.

Engineering minimal tissue inhibitors of metalloproteinase targeting MMPs via gene shuffling and yeast surface display.

Overexpression of specific matrix metalloproteinases (MMPs) has a key role in development of several diseases, such as cancer, neurological disorders, and cardiovascular diseases due to their critical role in degradation and remodeling of the extracellular matrix (ECM). Tissue inhibitors of metalloproteinases (TIMPs), a family of four in humans, are endogenous inhibitors of MMPs. TIMPs have a high level of sequence and structure homology, with a broad range of binding and inhibition to the family of MMPs. It is important to identify the key motifs of TIMPs responsible for inhibition of MMPs to develop efficient therapeutics targeting specific MMPs. We used DNA shuffling between the human TIMP family to generate a minimal TIMP hybrid library in yeast to identify the dominant minimal MMP inhibitory regions. The minimal TIMP variants screened toward MMP-3 and MMP-9 using fluorescent-activated cell sorting (FACS). Interestingly, several minimal TIMP variants selected after screening toward MMP-3cd or MMP-9cd, with lengths as short as 20 amino acids, maintained or improved binding to MMP-3 and MMP-9. The TIMP-MMP binding dissociation constant (KD ), in the nM range, and MMP inhibition constants (Ki ), in the pM range, of these minimal TIMP variants were similar to the N-terminal domain of TIMP-1 on the yeast surface and in solution indicating the potency of these minimal variants as MMP inhibitors. We further used molecular modeling simulation, and molecular docking of the minimal TIMP variants in complex with MMP-3cd to understand the binding and inhibition mechanism of these variants.

Loss of TIMP3, but not TIMP4, exacerbates thoracic and abdominal aortic aneurysm.

AIMS: Aorta exhibits regional heterogeneity (structural and functional), while different etiologies for thoracic and abdominal aortic aneurysm (TAA, AAA) are recognized. Tissue inhibitor of metalloproteinases (TIMPs) regulate vascular remodeling through different mechanisms. Region-dependent functions have been reported for TIMP3 and TIMP4 in vascular pathologies. We investigated the region-specific function of these TIMPs in development of TAA versus AAA. METHODS & RESULTS: TAA or AAA was induced in male and female mice lacking TIMP3 (Timp3-/-), TIMP4 (Timp4-/-) or in wildtype (WT) mice by peri-adventitial elastase application. Loss of TIMP3 exacerbated TAA and AAA severity in males and females, with a greater increase in proteinase activity, smooth muscle cell phenotypic switching post-AAA and -TAA, while increased inflammation was detected in the media post-AAA, but in the adventitia post-TAA. Timp3-/- mice showed impaired intimal barrier integrity post-AAA, but a greater adventitial vasa-vasorum branching post-TAA, which could explain the site of inflammation in AAA versus TAA. Severity of TAA and AAA in Timp4-/- mice was similar to WT mice. In vitro, Timp3 knockdown more severely compromised the permeability of human aortic EC monolayer compared to Timp4 knockdown or the control group. In aneurysmal aorta specimens from patients, TIMP3 expression decreased in the media in AAA, and in adventitial in TAA specimens, consistent with the impact of its loss in AAA versus TAA in mice. CONCLUSION: TIMP3 loss exacerbates inflammation, adverse remodeling and aortic dilation, but triggers different patterns of remodeling in AAA versus TAA, and through different mechanisms.

Metalloproteinase-2 in failed back surgery syndrome caused by epidural fibrosis: can it play a role in persistent pain?

Purpose: Failed Back Surgery Syndrome (FBSS) occurs in 10-40% of patients treated surgically due to disk herniation (DH). There are several factors that can cause a predisposition to FBSS, but the exact pathomechanism has not been elucidated. The aim of this study was to investigate Metalloproteinase-2 (MMP-2) and Tissue Inhibitor of Metalloproteinase-2 (TIMP-2) activities in a homogeneous group of FBSS patients with epidural fibrosis in comparison to its activity in patients with surgically treated DH. Methods: DH, FBSS, and control (CG) groups consisted of 30 subjects. The patients were assessed clinically by the Numerical Rating Scale (NRS), McGill Pain Questionnaire (SF -MPQ), Oswestry Disability Index (ODI), and Beck Depression Inventory (BDI). Serum concentrations of MMP-2 and TIMP-2 were measured by using the immunoenzymatic method. Results: There was a significantly higher MMP-2 expression (medians: 4797.49 vs. 2656.65; p < 0.0001) and TIMP-2 concentration (medians: 166.40 vs. 109.60; p < 0.0001) in the DH compared to the CG. Significantly higher MMP-2 expression (4219.95 vs. 2656.65; p < 0.0001) and TIMP-2 concentration (medians: 150.17 vs. 109.60; p = 0.0003) were also found in the FBSS compared to the CG. The activity of MMP-2, measured as MMP-2/TIMP-2, did not significantly change between the DH, FBSS, and CG. MMP2 expression (p < 0.0001) and TIMP-2 concentration (p < 0.0001) were significantly higher in the DH than FBSS. Conclusion: Results indicate the presence of a contribution of MMP-2 and TIMP-2 in DH and FBSS. Unchanged activity of MMP-2 can indicate an insufficiency in the MMP-2 repair system in both diseases. Lower MMP-2 expression and TIMP-2 concentration in the FBSS group can reflect the chronicity of the process.

Upregulated TIMP1 facilitates and coordinates myometrial contraction by decreasing collagens and cell adhesive capacity during human labor.

Myometrial contraction is one of the key events involved in parturition. Increasing evidence suggests the importance of the extracellular matrix (ECM) in this process, in addition to the functional role of myometrial smooth muscle cells, and our previous study identified an upregulated tissue inhibitor of metalloproteinase 1 (TIMP1) in human laboring myometrium compared to nonlabor samples. This study aimed to further explore the potential role of TIMP1 in myometrial contraction. First, we confirmed increased myometrial TIMP1 levels in labor and during labor with cervical dilation using transcriptomic and proteomic analyses, followed by real-time PCR, western blotting, and immunohistochemistry. Then, a cell contraction assay was performed to verify the decreased contractility after TIMP1 knockdown in vitro. To further understand the underlying mechanism, we used RNA-sequencing analysis to reveal the upregulated genes after TIMP1 knockdown; these genes were enriched in collagen fibril organization, cell adhesion, and ECM organization. Subsequently, a human matrix metalloproteinase (MMP) array and collagen staining were performed to determine the TIMPs, MMPs and collagens in laboring and nonlabor myometrium. A real-time cell adhesion assay was used to detect cell adhesive capacity. The results showed upregulated MMP8 and MMP9, downregulated collagens, and attenuated cell adhesive capacity in laboring myometrium, while lower MMP levels and higher collagen levels and cell adhesive capacity were observed in nonlabor. Moreover, TIMP1 knockdown led to restoration of cell adhesive capacity. Together, these results indicate that upregulated TIMP1 during labor facilitates and coordinates myometrial contraction by decreasing collagen and cell adhesive capacity, which may provide effective strategies for the regulation of myometrial contraction.

The predictive value of [TIMP-2]*[IGFBP7] in adverse outcomes for acute kidney injury: a systematic review and meta-analysis.

MATERIALS AND METHODS: Relevant articles published up to 17 June 2023 were retrieved from five databases (Cochrane Library/Embase/PubMed/SinoMed/Web of Science). The pre-established inclusion and exclusion criteria determined the selection of publications. Pooled sensitivity (SEN), specificity (SPE), diagnostic odds ratio, likelihood ratio, and summary receiver operating characteristic curve were employed to assess the predictive value. The presence or potential sources of heterogeneity were investigated via subgroup and SEN analyses. RESULTS: Ten published and eligible studies (1559 cases) were included in the evaluation for the capability of [TIMP-2]*[IGFBP7] to predict the poor prognosis of AKI through the random effect model. Pooled SEN, SPE, diagnostic odds ratio, and positive and negative likelihood ratios were 0.82 (95% CI: 0.77-0.86, I2 = 53.4%), 0.64 (95% CI: 0.61-0.67, I2 = 88.3%), 14.06 (95% CI: 7.31-27.05, I2 = 55.0%), 2.859 (95% CI: 2.15-3.77, I2 = 80.7%), and 0.28 (95% CI: 0.20-0.40, I2 = 35.0%), respectively. The estimated area under the curve was 0.8864 (standard error: 0.0306), and the Q* was 0.7970 (standard error: 0.0299). The endpoints and cutoff values were the main causes of heterogeneity. CONCLUSIONS: [TIMP-2]*[IGFBP7] is possible in predicting poor prognosis of AKI, but it is better to be applied along with other indicators or clinical risk factors.

Urinary cell cycle biomarkers for the prediction of renal non-recovery in patients with septic acute kidney injury: a prospective study.

BACKGROUND: Poor prognosis has been associated with the absence of renal recovery after acute kidney injury (AKI). This study aimed to investigate whether urinary biomarkers at 0 and 24 h could be used independently or in conjunction with a clinical model to predict renal non-recovery in septic AKI. METHODS: A prospective observational study was conducted to measure the urinary levels of insulin-like growth factor-binding protein 7 (IGFBP7) and tissue inhibitor of metalloproteinase-2 (TIMP-2) at the time of AKI diagnosis (0 h) and 24 h later. Renal non-recovery within 7 days was defined as the outcome. The predictive value of urinary biomarkers for renal non-recovery in septic AKI was assessed using the area under the curve (AUC). RESULTS: A total of 198 individuals with septic AKI were included in the final analysis. Among them, 38.9% (n = 77) did not experience renal recovery within 7 days. The combination of urinary IGFBP7 and TIMP-2 at the initial time point demonstrated prognostic value for non-recovery of renal function, with an AUC of 0.782. When [TIMP-2]*[IGFBP7] was measured at 0 h, the clinical prognostic model, incorporating AKI stage 2-3 and the non-renal sequential organ failure assessment score, showed an improved AUC of 0.822 (with a sensitivity of 88.3% and specificity of 59.5%). CONCLUSIONS: The combination of urinary [TIMP-2]*[IGFBP7] at 0 h exhibited moderate predictive ability for renal non-recovery in cases of septic AKI. However, there is potential to enhance the prognostic capabilities of the [TIMP-2]*[IGFBP7]-clinical prediction model.

Genome-wide interaction analysis of folate for colorectal cancer risk.

BACKGROUND: Epidemiological and experimental evidence suggests that higher folate intake is associated with decreased colorectal cancer (CRC) risk; however, the mechanisms underlying this relationship are not fully understood. Genetic variation that may have a direct or indirect impact on folate metabolism can provide insights into folate's role in CRC. OBJECTIVES: Our aim was to perform a genome-wide interaction analysis to identify genetic variants that may modify the association of folate on CRC risk. METHODS: We applied traditional case-control logistic regression, joint 3-degree of freedom, and a 2-step weighted hypothesis approach to test the interactions of common variants (allele frequency >1%) across the genome and dietary folate, folic acid supplement use, and total folate in relation to risk of CRC in 30,550 cases and 42,336 controls from 51 studies from 3 genetic consortia (CCFR, CORECT, GECCO). RESULTS: Inverse associations of dietary, total folate, and folic acid supplement with CRC were found (odds ratio [OR]: 0.93; 95% confidence interval [CI]: 0.90, 0.96; and 0.91; 95% CI: 0.89, 0.94 per quartile higher intake, and 0.82 (95% CI: 0.78, 0.88) for users compared with nonusers, respectively). Interactions (P-interaction < 5×10-8) of folic acid supplement and variants in the 3p25.2 locus (in the region of Synapsin II [SYN2]/tissue inhibitor of metalloproteinase 4 [TIMP4]) were found using traditional interaction analysis, with variant rs150924902 (located upstream to SYN2) showing the strongest interaction. In stratified analyses by rs150924902 genotypes, folate supplementation was associated with decreased CRC risk among those carrying the TT genotype (OR: 0.82; 95% CI: 0.79, 0.86) but increased CRC risk among those carrying the TA genotype (OR: 1.63; 95% CI: 1.29, 2.05), suggesting a qualitative interaction (P-interaction = 1.4×10-8). No interactions were observed for dietary and total folate. CONCLUSIONS: Variation in 3p25.2 locus may modify the association of folate supplement with CRC risk. Experimental studies and studies incorporating other relevant omics data are warranted to validate this finding.

Gene Expression of Clusterin, Tissue Inhibitor of Metalloproteinase-1, and Their Receptors in Retinal Pigment Epithelial Cells and Müller Glial Cells Is Modulated by Inflammatory Stresses.

Balanced activities of matrix metalloproteinases (MMPs) and their inhibitors are essential for photoreceptor (PR) cell survival. PR rod cell survival in rodent models of inherited retinitis pigmentosa (RP) is prolonged by recombinant tissue inhibitor of metalloproteinase (TIMP)-1 or clusterin (CLU) proteins. Retinal pigment epithelial cells (RPE) and Müller glia (MG) cells support PR cells. In human RPE and MG cell lines, we measured their mRNA levels of the two genes with quantitative real-time PCR (qRT-PCR) with interleukin (IL)-1ß treatment, a key pathological component in retinal degeneration. Endogenous CLU gene expression was significantly downregulated by IL-1ß in both cell types, whereas TIMP-1 expression was upregulated in MG cells, suggesting the transcriptional control of CLU is potentially more sensitive to inflammatory conditions. The expression levels of CLU endocytic receptors revealed that the low-density lipoprotein receptor-related protein 2 (LRP2) was upregulated only in MG cells by the treatment with no detectable change in RPE cells. Like LRP2, IL-1ß upregulated TIMP-1 receptor LRP1 expression in MG cells; however, it was decreased in the expression of RPE cells. These data suggest that the gene expression of CLU and TIMP-1 and their receptors may be dynamically modulated in inflammatory conditions.

TIMP3 induces gene expression partly through PI3K and their association with vascularization and heart rate.

Background: Tissue inhibitor of metalloproteinase 3 (TIMP3) was recently demonstrated capable to regulate some gene expression in a myocardial infarction model. Here we aim to explore the gene expression profile in TIMP3-treated cardiomyocytes and related potential cardiovascular functions. Methods: Total RNA extracted from cultured neonatal rat ventricular myocytes (NRVMs) were used for RNA sequencing analysis and real-time PCR. KEGG pathway enrichment assay and Ingenuity Pathway Analysis (IPA) were performed to study the signaling pathways and downstream effects. Western blot was used to detect phosphorylation of protein kinase B (Akt). A Cell Counting Kit-8 assay was employed to evaluate the proliferation of human umbilical vein endothelial cells (HUVECs). Contraction rate of NRVMs was measured with microscopy. Results: RNA sequencing data showed that expression of 2,526 genes were significantly modulated by recombinant TIMP3 (rTIMP3, 100 ng/ml) in NRVMs. Some differentially expressed genes (DEGs) were validated with real-time PCR. Several KEGG pathways including the phosphoinositide-3-kinase (PI3K)-Akt pathway were significantly regulated by rTIMP3. Phosphorylation of Akt was increased by rTIMP3 and a PI3K inhibitor LY294002 suppressed rTIMP3-induced up-regulation of some genes. Some DEGs were predicted by IPA to increase vascularization, and some to decrease heart rate. RTIMP3 could reduce the contraction rate of NRVMs and its conditioned media increased the proliferation of HUVECs. Conclusion: TIMP3 can regulate expression of multiple genes partly through PI3K. Some DEGs were associated with activation of vascularization and some with heart rate reduction. This study suggests that TIMP3 can potentially modulate cardiovascular functions via DEGs.

The usefulness of MMP-9, TIMP-1 and MMP-9/TIMP-1 ratio for diagnosis and assessment of COPD severity.

BACKGROUND: Inflammation, oxidative stress and an imbalance between proteases and protease inhibitors are recognized pathophysiological features of chronic obstructive pulmonary disease (COPD). The aim of this study was to evaluate serum levels of matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-1) in patients with COPD and to assess their relationship with lung function, symptom severity scores and recent acute exacerbations. METHODS: In this observational cohort study, serum levels of MMP-9 and TIMP-1 and the MMP-9/TIMP-1 ratio in the peripheral blood of COPD patients with stable disease and healthy controls were determined, and their association with lung function (postbronchodilator spirometry, body plethysmography, single breath diffusion capacity for carbon monoxide), symptom severity scores (mMRC and CAT) and exacerbation history were assessed. RESULTS: COPD patients (n = 98) had significantly higher levels of serum MMP-9 and TIMP-1 and a higher MMP-9/TIMP-1 ratio than healthy controls (n = 47) (p ≤ 0.001). The areas under the receiver operating characteristic curve for MMP-9, TIMP-1 and the MMP-9/TIMP-1 ratio for COPD diagnosis were 0.974, 0.961 and 0.910, respectively (all p < 0.05). MMP-9 and the MMP-9/TIMP-1 ratio were both negatively correlated with FVC, FEV1, FEV1/FVC, VC, and IC (all p < 0.05). For MMP-9, a positive correlation was found with RV/TLC% (p = 0.005), and a positive correlation was found for the MMP-9/TIMP-1 ratio with RV% and RV/TLC% (p = 0.013 and 0.002, respectively). Patients with COPD GOLD 3 and 4 presented greater MMP-9 levels and a greater MMP-9/TIMP-1 ratio compared to GOLD 1 and 2 patients (p ≤ 0.001). No correlation between diffusion capacity for carbon monoxide and number of acute exacerbations in the previous year was found. CONCLUSIONS: COPD patients have elevated serum levels of MMP-9 and TIMP-1 and MMP-9/TIMP-1 ratio. COPD patients have an imbalance between MMP-9 and TIMP-1 in favor of a pro-proteolytic environment, which overall indicates the importance of the MMP-9/TIMP-1 ratio as a potential biomarker for COPD diagnosis and severity.

Chronic Kidney Disease: Combined Effects of Gene Polymorphisms of Tissue Inhibitors of Metalloproteinase 3, Total Urinary Arsenic, and Blood Lead Concentration.

The tissue inhibitor of metalloproteinase 3 (TIMP3) is known to be an anti-fibrotic factor. Arsenic, lead, and cadmium exposure and selenium intake may affect TIMP3 expression. The downregulation of TIMP3 expression is related to kidney fibrosis. Genotypes of TIMP3 are related to hypertension and cardiovascular diseases. Therefore, this study explored whether TIMP3 polymorphism is associated with hypertension-related chronic kidney disease (CKD). In addition, the combined effects of TIMP3 polymorphism and total urinary arsenic, blood lead and cadmium, and plasma selenium concentrations on CKD, were investigated. This was a case-control study, with 213 CKD patients and 423 age- and sex-matched controls recruited. Polymerase chain reaction-restriction fragment length polymorphism was used to determine TIMP3 gene polymorphisms. The concentrations of urinary arsenic species, plasma selenium, and blood lead and cadmium were measured. The odds ratio (OR) of CKD in the TIMP3rs9609643 GA/AA genotype was higher than that of the GG genotype at high levels of total urinary arsenic and blood lead; the OR and 95% confidence interval (CI) were 0.57 (0.31-1.05) and 0.52 (0.30-0.93), respectively, after multivariate adjustment. High blood lead levels tended to interact with the TIMP3rs9609643 GG genotype to increase the OR of CKD, and gave the highest OR (95% CI) for CKD of 5.97 (2.60-13.67). Our study supports a possible role for the TIMP3rs9609643 risk genotype combined with high total urinary arsenic or with high blood lead concentration to increase the OR of CKD.

Myeloma Microenvironmental TIMP1 Induces the Invasive Phenotype in Fibroblasts to Modulate Disease Progression.

Tissue inhibitors of metalloproteinases (TIMPs) are endogenous matrix metalloproteinase inhibitors. TIMP1 is produced by cancer cells and has pleiotropic activities. However, its role and source in multiple myeloma (MM) are unclear. Here, we evaluated TIMP1 protein and mRNA levels in bone marrow (BM) plasma cells and assessed the effects of TIMP1 expression on fibroblast invasive capacity using three-dimensional spheroid cell invasion assays. TIMP1 mRNA and protein levels were elevated when patients progressed from monoclonal gammopathy of undetermined significance or smouldering myeloma to MM. Furthermore, TIMP1 levels decreased at complete response and TIMP1 protein levels increased with higher international staging. TIMP1 mRNA levels were markedly higher in extramedullary plasmacytoma and MM with t(4;14). Overall survival and post-progression survival were significantly lower in MM patients with high TIMP1 protein. Recombinant TIMP1 did not directly affect MM cells but enhanced the invasive capacity of fibroblasts; this effect was suppressed by treatment with anti-TIMP1 antibodies. Fibroblasts supported myeloma cell invasion and expansion in extracellular matrix. Overall, these results suggested that MM-derived TIMP1 induces the invasive phenotype in fibroblasts and is involved in disease progression. Further studies are required to elucidate the specific roles of TIMP1 in MM and facilitate the development of novel therapies targeting the TIMP1 pathway.