OBJECTIVE: To explore whether the regulation of matrix metalloproteinase 9 (MMP-9)/ tissue inhibitors of MMPs (TIMPs) gene expression through histone acetylation is a possible mechanism by which electroacupuncture (EA) protects blood-brain barrier (BBB) integrity in a middle cerebral artery occlusion (MCAO) rat model.
METHODS: Male Sprague-Dawley rats were divided into four groups: the sham group, the MCAO group, the MCAO + EA (MEA) group, and the MCAO + EA + HAT inhibitor (HATi) group. The MCAO model was generated by blocking the middle cerebral artery. EA was applied to Baihui (GV20). Samples were collected 1 or 3 d after reperfusion. Neurological function scores and Evans blue extravasation were employed to evaluate the poststroke injury. The effect of EA on MMP-9/TIMPs gene expression was assessed by real-time fluorescence quantitative polymerase chain reaction (RT-qPCR) and chromatin immunoprecipitation (ChIP).
RESULTS: Our results showed that EA treatment prominently improved neurological function and ameliorated BBB disruption. The RT-qPCR assay showed that EA reduced the expression of MMP-9 and promoted TIMP-2 mRNA expression, but HATi reversed these effects of EA. In addition, ChIP results revealed that EA decreased the enrichment of H3K9ace/H3K27ace at MMP-9 promoters and notably stimulated the recruitment of H3K9ace/H3K27ace at TIMP-2 promoter.
CONCLUSIONS: EA treatment at Baihui (GV20) regulates the transcription of MMP-9 and TIMP-2 through histone acetylation modification in the acute stage of stroke, which preserves the structural integrity of the BBB in MCAO rats. These findings suggested that the histone acetylation-mediated transcriptional activity of target genes may be a crucial mechanism of EA treatment in stroke.

OBJECTIVE: To explore whether circulating matrix metalloproteinase-2 (MMP-2), MMP-9, MMP-9/neutrophil gelatinase-associated lipocalin, MMP-9/tissue inhibitor of metalloproteinase-1 (TIMP-1), MMP-14, TIMP-2 and TIMP-3 were associated with the severity and progression of diabetic retinopathy (DR) in patients with type 1 diabetes (T1D).
METHODS: Baseline and prospective analyses were conducted over a period of 10.5 person-years. In 2009, recruitment and biochemical analyses (MMPs, TIMPs, glycated haemoglobin (HbA1c), serum creatinine, macroalbuminuria) were performed. Fundus photography, performed at baseline and at follow-up in accordance with the regional screening programme, was compared after being categorised according to the International Clinical Diabetic Retinopathy Disease Severity Scale. \'DR progression at least one leve\' was calculated. High MMP-2 was defined as ≥178 ng/mL (≥75th percentile) and high TIMP-2 as ≥205 ng/mL (≥75th percentile). DR was dichotomised as \'at least moderate DR\' or \'no/mild DR\'.
RESULTS: The study included 267 participants, 57% of whom were men. At baseline, the prevalence of high MMP-2 (p=0.001) and high TIMP-2 (p=0.008) increased with the severity of DR. \'At least moderate DR\' (adjusted OR (AOR) 2.4, p=0.008) and macroalbuminuria (AOR 3.6, p=0.025) were independently associated with high MMP-2. \'At least moderate DR\' (AOR 2.3, p=0.009) and macroalbuminuria (3.4, p=0.031) were independently associated with high TIMP-2. DR progression occurred in 101 (46%) patients (p<0.001). HbA1c≥53 mmol/mol was associated with the progression of DR (crude OR 3.8, p=0.001). No other MMPs or TIMPs were linked to the severity or the progression of DR.
CONCLUSIONS: High levels of MMP-2 and TIMP-2 indicated more severe DR or diabetic nephropathy. Only HbA1c was associated with the progression of DR in 267 patients with T1D.

Cerebral amyloid angiopathy (CAA) is a highly prevalent and progressive pathology, involving amyloid-β (Aβ) deposition in the cerebral blood vessel walls. CAA is associated with an increased risk for intracerebral hemorrhages (ICH). Insight into the molecular mechanisms associated with CAA pathology is urgently needed, to develop additional diagnostic tools to allow for reliable and early diagnosis of CAA and to obtain novel leads for the development of targeted therapies. Tissue inhibitor of matrix metalloproteinases 4 (TIMP4) is associated with cardiovascular functioning and disease and has been linked to vascular dementia. Using immunohistochemistry, we studied occipital brain tissue samples of 57 patients with CAA (39 without ICH and 18 with ICH) and 42 controls, and semi-quantitatively assessed expression levels of TIMP4. Patients with CAA had increased vascular expression of TIMP4 compared to controls (p < 0.001), and in these patients, TIMP4 expression correlated with CAA severity (τb = 0.38; p = 0.001). Moreover, TIMP4 expression was higher in CAA-ICH compared to CAA-non-ICH cases (p = 0.024). In a prospective cross-sectional study of 38 patients with CAA and 37 age- and sex-matched controls, we measured TIMP4 levels in cerebrospinal fluid (CSF) and serum using ELISA. Mean CSF levels of TIMP4 were decreased in patients with CAA compared to controls (3.36 ± 0.20 vs. 3.96 ± 0.22 ng/ml, p = 0.033), whereas median serum levels were increased in patients with CAA (4.51 ng/ml [IQR 3.75-5.29] vs 3.60 ng/ml [IQR 3.11-4.85], p-9.013). Moreover, mean CSF TIMP4 levels were lower in CAA patients who had experienced a symptomatic hemorrhage compared to CAA patients who did not (2.13 ± 0.24 vs. 3.57 ± 0.24 ng/ml, p = 0.007). CSF TIMP4 levels were associated with CSF levels of Aβ40 (spearman r (rs) = 0.321, p = 0.009). In summary, we show that TIMP4 is highly associated with CAA and CAA-related ICH, which is reflected by higher levels in the cerebral vasculature and lower levels in CSF. With these findings we provide novel insights into the pathophysiology of CAA, and more specifically in CAA-associated ICH.

Echinops plants have received great attention for the treatment of many diseases due to pharmacological properties such as their antidiabetic, antioxidant, and anti-inflammatory characteristics. The major purpose of the present study was to investigate the cardioprotective benefits of Echinops cephalotes (Ech) against myocardial ischemia-reperfusion (MI/R) injury. Male Wistar rats were randomly allocated to three groups: sham, MI, and MI + Ech. The left coronary artery (LAD) was blocked for 30 minutes to induce MI. In the treatment group, rats were given 150 mg/kg/day of Ech extract for 28 days. Aqueous extracts were made from Echinops plants. To study heart function, fibrosis, cardiac damage indicators, and oxidative stress factors, echocardiography, Masson\'s trichrome staining, and biochemical tests were used. The expression of matrix metalloproteinase 2 and 9 (MMP2 and MMP-9) and tissue inhibitor of metalloproteinase (TIMP) was determined using Western blotting. Tissue damage was assessed using hematoxylin and eosin staining. MI group exhibited significantly reduced ejection fraction (EF) and fractional shortening (FS), enhanced levels of lactate dehydrogenase (LDH), creatine kinase MB (CK-MB), cardiac Troponin I (cTnI), and malondialdehyde (MDA), as well as a decrease in the Glutathione (GSH) tissue content, reduced activity of superoxide dismutase (SOD), increasing fibrosis, upregulations of MMP-2 and MMP-9, and reduction of TIMP compared to the sham group. The findings suggest that Ech in particular, could be a promising therapeutic agent to reduce the damage in MI by targeting oxidative stress and modulating the activities of matrix metalloproteinases and their tissue inhibitors.

OBJECTIVE: This study aims to evaluate the effects of intermittent compressive force (ICF) on the expression of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) by human periodontal ligament cells (hPDLCs).
METHODS: hPDLCs were subjected to ICF with a magnitude of 1.5 g/cm2 and loaded for 24 h. mRNA and protein expression of several MMPs and TIMPs were assessed using RT-PCR and ELISA analyses. An inhibitor of TGF-β (SB431542) was used to assess a possible role of TGF-β in the expression of MMPs and TIMPs under ICF.
RESULTS: mRNA and protein analyses showed that ICF significantly induced expression of TIMP1 and TIMP3, but decreased expression of MMP1. Incubation with the TGF-β inhibitor and applied to ICF showed a downregulation of TIMP3, but expression of MMP1 was not affected.
CONCLUSIONS: ICF is likely to affect ECM homeostasis by hPDLCs by regulating the expression of MMP1 and TIMPs. Moreover, TGF-β1 regulated expression of TIMP3. These findings suggest ICF may decrease the degradation of ECM and may thus be essential for maintaining PDL homeostasis.

Matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) play critical roles in regulating processes associated with malignant behavior. These endopeptidases selectively degrade components of the extracellular matrix (ECM), growth factors, and their receptors, contributing to cancer cell invasiveness and migratory characteristics by disrupting the basal membrane. However, the expression profile and role of various matrix metalloproteinases remain unclear, and only a few studies have focused on differences between diagnoses of brain tumors. Using quantitative real-time PCR analysis, we identified the expression pattern of ECM modulators (n = 10) in biopsies from glioblastoma (GBM; n = 20), astrocytoma (AST; n = 9), and meningioma (MNG; n = 19) patients. We found eight deregulated genes in the glioblastoma group compared to the benign meningioma group, with only MMP9 (FC = 2.55; p = 0.09) and TIMP4 (7.28; p < 0.0001) upregulated in an aggressive form. The most substantial positive change in fold regulation for all tumors was detected in matrix metalloproteinase 2 (MNG = 30.9, AST = 4.28, and GBM = 4.12). Notably, we observed an influence of TIMP1, demonstrating a positive correlation with MMP8, MMP9, and MMP10 in tumor samples. Subsequently, we examined the protein levels of the investigated MMPs (n = 7) and TIMPs (n = 3) via immunodetection. We confirmed elevated levels of MMPs and TIMPs in GBM patients compared to meningiomas and astrocytomas. Even when correlating glioblastomas versus astrocytomas, we showed a significantly increased level of MMP1, MMP3, MMP13, and TIMP1. The identified metalloproteases may play a key role in the process of gliomagenesis and may represent potential targets for personalized therapy. However, as we have not confirmed the relationship between mRNA expression and protein levels in individual samples, it is therefore natural that the regulation of metalloproteases will be subject to several factors.

The extracellular matrix (ECM) is a complex non-cellular three-dimensional macromolecular network present within all tissues and organs, forming the foundation on which cells sit, and composed of proteins (such as collagen), glycosaminoglycans, proteoglycans, minerals, and water. The ECM provides a fundamental framework for the cellular constituents of tissue and biochemical support to surrounding cells. The ECM is a highly dynamic structure that is constantly being remodeled. Matrix metalloproteinases (MMPs) are among the most important proteolytic enzymes of the ECM and are capable of degrading all ECM molecules. MMPs play a relevant role in physiological as well as pathological processes; MMPs participate in embryogenesis, morphogenesis, wound healing, and tissue remodeling, and therefore, their impaired activity may result in several problems. MMP activity is also associated with chronic inflammation, tissue breakdown, fibrosis, and cancer invasion and metastasis. The periodontium is a unique anatomical site, composed of a variety of connective tissues, created by the ECM. During periodontitis, a chronic inflammation affecting the periodontium, increased presence and activity of MMPs is observed, resulting in irreversible losses of periodontal tissues. MMP expression and activity may be controlled in various ways, one of which is the inhibition of their activity by an endogenous group of tissue inhibitors of metalloproteinases (TIMPs), as well as reversion-inducing cysteine-rich protein with Kazal motifs (RECK).

There has been limited research on assessing metalloproteinases (MMPs) 1, 2, and 7, as well as their tissue inhibitors (TIMPs) 1, 2, 3, and 4 in the context of polytrauma. These proteins play crucial roles in various physiological and pathological processes and could be a reliable tool in polytrauma care. We aimed to determine their clinical relevance. We assessed 24 blunt polytrauma survivors and 12 fatalities (mean age, 44.2 years, mean ISS, 45) who were directly admitted to our Level I trauma center and spent at least one night in the intensive care unit. We measured serum levels of the selected proteins on admission (day 0) and days 1, 3, 5, 7, and 10. The serum levels of the seven proteins varied considerably among individuals, resulting in similar median trend curves for TIMP1 and TIMP4 and for MMP1, MMP2, TIMP2, and TIMP3. We also found a significant interrelationship between the MMP2, TIMP2, and TIMP3 levels at the same measurement points. Furthermore, we calculated significant cross-correlations between MMP7 and MMP1, TIMP1 and MMP7, TIMP3 and MMP1, TIMP3 and MMP2, and TIMP4 and TIMP3 and an almost significant correlation between MMP7 and TIMP1 for a two-day-lag. The autocorrelation coefficient reached statistical significance for MMP1 and TIMP3. Finally, lower TIMP1 serum levels were associated with in-hospital mortality upon admission. The causal effects and interrelationships between selected proteins might provide new insights into the interactions of MMPs and TIMPs. Identifying the underlying causes might help develop personalized therapies for patients with multiple injuries. Administering recombinant TIMP1 or increasing endogenous production could improve outcomes for those with multiple injuries. However, before justifying further investigations into basic research and clinical relevance, our findings must be validated in a multicenter study using independent cohorts to account for clinical and biological variability.

Targeting metalloproteinases (MPs) has been the center of attention for developing therapeutics due to their contribution to a wide range of diseases, including cancer, cardiovascular, neurodegenerative disease, and preterm labor. Protein-based MP inhibitors offer higher stability and selectivity, which is critical for developing efficient therapeutics with low off-target effects. Tissue inhibitors of metalloproteinases (TIMPs), natural inhibitors of MPs, and antibodies provide excellent protein scaffolds for engineering selective or multispecific MP inhibitors. Advances in protein engineering and design techniques, such as rational design and directed evolution using yeast display to develop potent MP inhibitors, are discussed, including but not limited to loop grafting, swapping, and counterselective selection.

BACKGROUND: The prevalence of NAFLD is rapidly increasing. NAFLD can progress to NASH, fibrosis, cirrhosis, and HCC, which will soon become the main causes of liver transplantation. To date, no effective drug for NASH has been approved by the Food and Drug Administration. This is partly due to the lack of reliable human in vitro models. Here, we present a novel human liver spheroid model that can be used to study the mechanisms underlying liver fibrosis formation and degradation.
RESULTS: Such spheroids, which contain hepatocytes, stellate cells, KC, and LSECs, spontaneously develop fibrosis that is exacerbated by treatment with free fatty acids. Conditioned medium from activated LSECs caused similar activation of fibrosis in spheroids containing primary human hepatocyte and NPCs, indicating the action of soluble mediators from the LSECs. Spheroids containing LSECs treated with free fatty acids produced tissue inhibitor of metalloproteinases inhibitor 1, a matrix metalloproteinases inhibitor important for fibrosis progression. Tissue inhibitor of metalloproteinases inhibitor 1 knockdown using siRNA led to a reduction in collagen and procollagen accumulation, which could be partially rescued using a potent matrix metalloproteinases inhibitor. Interestingly, tissue inhibitor of metalloproteinases inhibitor 1 was found to be expressed at higher levels, specifically in a subtype of endothelial cells in the pericentral region of human fibrotic livers, than in control livers.
CONCLUSIONS: Potential anti-NASH drugs and compounds were evaluated for their efficacy in reducing collagen accumulation, and we found differences in specificity between spheroids with and without LSECs. This new human NASH model may reveal novel mechanisms for the regulation of liver fibrosis and provide a more appropriate model for screening drugs against NASH.