Matrix Metalloproteinase-9 Expression Research Articles

Protective Mechanism of Ethyl Gallate against Intestinal Ischemia-Reperfusion Injury in Mice by in Vivo and in Vitro Studies Based on Transcriptomics.

Intestinal ischemia-reperfusion injury (IIRI) is a common clinical disease that can be life-threatening in severe cases. This study aimed to investigate the effects of ethyl gallate (EG) on IIRI and its underlying mechanisms. A mouse model was established to mimic human IIRI by clamping the superior mesenteric artery. Transcriptomics techniques were used in conjunction with experiments to explore the potential mechanisms of EG action. Intestinal histomorphological damage, including intestinal villi damage and mucosal hemorrhage, was significantly reversed by EG. EG also alleviated the oxidative stress, inflammation, and intestinal epithelial apoptosis caused by IIRI. 2592 up-regulated genes and 2754 down-regulated genes were identified after EG treatment, and these differential genes were enriched in signaling pathways, including fat digestion and absorption, and extracellular matrix (ECM) receptor interactions. In IIRI mouse intestinal tissue, expression of the differential protein matrix metalloproteinase 9 (MMP9), as well as its co-protein NF-κB-p65, was significantly increased, while EG inhibited the expression of MMP9 and NF-κB-p65. In Caco-2 cells in an established oxygen-glucose deprivation/reperfusion model (OGD/R), EG significantly reversed the decrease in intestinal barrier trans-epithelial electrical resistance (TEER). However, in the presence of MMP9 inhibitors, EG did not reverse the decreasing trend in TEER. This study illustrates the protective effect and mechanism of action of EG on IIRI and, combined with in vivo and in vitro experiments, it reveals that MMP9 may be the main target of EG action. This study provides new scientific information on the therapeutic effects of EG on IIRI.

TNF-α‒Mediated Keratinocyte Expression and Release of Matrix Metalloproteinase 9: Putative Mechanism of Pathogenesis in Stevens‒Johnson Syndrome/Toxic Epidermal Necrolysis

Stevens‒Johnson syndrome and toxic epidermal necrolysis (SJS/TEN) are severe cutaneous adverse drug reactions characterized by widespread keratinocyte cell death and epidermal detachment. At present, there is little understanding of how the detachment occurs or how it is abrogated by the TNF-α inhibitor etanercept, an effective SJS/TEN treatment. RNA sequencing was used to identify upregulated transcripts in formalin-fixed paraffin-embedded SJS/TEN skin biopsies. Epidermal matrix metalloproteinase 9 (MMP9) expression was assessed by immunohistochemistry in skin biopsies and cultured human skin explants exposed to serum from patients with cutaneous adverse drug reactions. TNF-α‒induced MMP9 expression and activity and its abrogation by etanercept were determined using the HaCaT immortalized keratinocyte cell line. Epidermal MMP9 expression was significantly higher in SJS/TEN skin (70.6%) than in healthy control skin (0%) (P= 0.0098) and nonbullous skin reactions (10.7%) (P= 0.0002). SJS/TEN serum induced significant MMP9 expression and collagenase activity in healthy skin explants, which was reduced by etanercept. Etanercept was also able to negate the TNF-α‒induced MMP9 expression in the HaCaT cell line. Data suggest that elevated epidermal MMP9 expression and collagenase activity are a putative pathogenic mechanism in SJS/TEN, which is limited by etanercept. Modulation of MMP9 expression and activity represents, to our knowledge, a previously unreported therapeutic target for the treatment of SJS/TEN.

β-hydroxybutyrate improves cognitive impairment caused by chronic cerebral hypoperfusion via amelioration of neuroinflammation and blood-brain barrier damage

BackgroundVascular cognitive impairment (VCI) is the second most common type of dementia after Alzheimer's disease (AD) in elderly people. Chronic cerebral hypoperfusion (CCH) is the early pathophysiological basis of VCI. β-Hydroxybutyrate (BHB) is one of the important components of ketone bodies, an intermediate product of endogenous energy metabolism, which can mitigate neuroinflammation in stroke and neurodegenerative diseases. The present study aimed to investigate whether BHB can improve cognitive impairment caused by CCH and the underlying mechanism. MethodsThe CCH model was established by permanent bilateral common carotid artery occlusion (2VO). CCH rats were intraperitoneally injected with BHB (1.5 mmol/kg/d) every day for 8 consecutive weeks from 2 weeks before surgery. The hippocampal blood flow of rats was measured by using a laser Doppler velocimetry. Used the Morris water maze test (MWM) to assess spatial learning and memory of rats, and harvested brain tissues for molecular, biochemical, and pathological tests. ResultsWe found that BHB intervention for 8 weeks could effectively restore hippocampal blood flow and improve spatial learning and memory in CCH rats. BHB can protect the blood-brain barrier (BBB), as manifested by reducing the ultrastructural damage and leakage of the BBB, restoring the expression of tight junction-related proteins and reducing the expression of Matrix Metalloproteinases-9 (MMP-9). Additionally, after BHB intervention, microglia activation was reduced, oligodendrocyte motility was active, and the expression levels of pro-inflammatory factors such as tumor necrosis factor (TNF-α), interleukin-1β (IL-1β), nuclear factor-κB (NF-κB) and advanced glycation end-products (RAGE) were lower, which also indicated that BHB had a beneficial effect in mitigating neuroinflammation. ConclusionBHB can improve the cognitive impairment caused by CCH. The potential mechanisms of BHB may be through reducing neuroinflammation and protecting BBB.

Warm acupuncture stimulation improves cartilage damage and motor function by regulating JAK2/STAT3 signaling pathway in rabbits with knee osteoarthritis

To observe the effect of warm acupuncture on the expression of Janus protein tyrosine kinase 2/signal transducer and activator of transcription 3 (JAK2/STAT3) signaling pathway and inflammatory factors of articular cartilage in rabbits with knee osteoarthritis (KOA), so as to explore its underlying mechanisms in improving KOA. New Zea-land rabbits were randomly divided into blank, model, warm acupuncture and medication groups (12 rabbits in each group). The KOA model was prepared by using the right hind limb tubular plaster extension fixation method. The rabbits in the warm acupuncture group received acupuncture of "Neixiyan"(EX-LE4),"Waixiyan"(ST35),"Heding"(EX-LE2) and "Zusanli"(ST36), followed by attaching an ignited moxa-stick segment to the acupuncture-handle. The treatment was conducted for 15 min, once a week for 4 weeks. The rabbits in the medication group received gavage of diclofenac sodium solution(0.35 mg/kg), once daily for 4 weeks. The dysfunction severity state of the rabbit's knee-joint was evaluated using Lequesne scale (0-3 points), and the histopathological changes of cartilage were observed under microscope after H.E. staining and the state of distribution of chondrocytes in different layers and the extracellular matrix was assessed using Mankin score (0-6 points). The contents of serum interleukin (IL)-6, tumor necrosis factor-α (TNF-α) and matrix metalloproteinase-9 (MMP-9) were measured by using ELISA, and the expression levels of p-JAK2/JAK2, p-STAT3/STAT3 and MMP-9 in knee cartilage tissue were detected using Western blot. Compared with the blank group, the Lequesne score, Mankin score, and the contents of serum IL-6, TNF-α and MMP-9, and the ratios of p-JAK2/JAK2 and p-STAT3/STAT3, and the expression level of MMP-9 protein in knee cartilage tissue were significantly increased in the model group (P<0.01). In comparison with the model group, the Lequesne score, Mankin score, contents of serum IL-6, TNF-α and MMP-9, and the ratios of p-JAK2/JAK2 and p-STAT3/STAT3, and the expression of MMP-9 protein in knee cartilage tissue were notably decreased in both the warm acupuncture and medication groups (P<0.01,P<0.05). The levels of Lequesne score, Mankin score, contents of serum IL-6, TNF-α and MMP-9, and the ratios of p-JAK2/JAK2 and p-STAT3/STAT3 in knee cartilage tissue were significantly lower in the warm acupuncture group than in the medication group (P<0.01, P<0.05). No significant difference was found between the warm acupuncture and medication groups in the expression of MMP-9 protein (P>0.05). Outcomes of H.E. showed injury of the perichondrium of knee joint, obvious reduction of the cartilage matrix staining, cystic changes, clustered and disordered arrangement and severe pyknosis and necrosis of the surface cells with reduction of number of cells and increase of vacuoles in the model group, which was milder in both warm acupuncture and medication groups. Warm acupuncture can improve motor function and reduce cartilage injury in KOA rabbits, which may be related to its functions in inhibiting the secretion of pro-inflammatory factors and regulating JAK2/STAT3 signaling and downregulating MMP-9 expression in the cartilage tissue.

Retinal protection by fungal product theissenolactone B in a sodium iodate-induced AMD model through targeting retinal pigment epithelial matrix metalloproteinase-9 and microglia activity

Age-related macular degeneration (AMD) is the leading cause of low vision and blindness for which there is currently no cure. Increased matrix metalloproteinase-9 (MMP-9) was found in AMD and potently contributes to its pathogenesis. Resident microglia also promote the processes of chronic neuroinflammation, accelerating the progression of AMD. The present study investigates the effects and mechanisms of the natural compound theissenolactone B (LB53), isolated from Theissenia cinerea, on the effects of RPE dysregulation and microglia hyperactivation and its retinal protective ability in a sodium iodate (NaIO3)-induced retinal degeneration model of AMD. The fungal component LB53 significantly reduces MMP-9 gelatinolysis in TNF-α-stimulated human RPE cells (ARPE-19). Similarly, LB53 abolishes MMP-9 protein and mRNA expression in ARPE-19 cells. Moreover, LB53 efficiently suppresses nitric oxide (NO) production, iNOS expression, and intracellular ROS levels in LPS-stimulated TLR 4-activated microglial BV-2 cells. According to signaling studies, LB53 specifically targets canonical NF-κB signaling in both ARPE-19 and BV-2 microglia. In an RPE-BV-2 interaction assay, LB53 ameliorates LPS-activated BV-2 conditioned medium-induced MMP-9 activation and expression in the RPE. In NaIO3-induced AMD mouse model, LB53 restores photoreceptor and bipolar cell dysfunction as assessed by electroretinography (ERG). Additionally, LB53 prevents retinal thinning, primarily the photoreceptor, and reduces retinal blood flow from NaIO3 damage evaluated by optic coherence tomography (OCT) and laser speckle flowgraphy (LSFG), respectively. Our results demonstrate that LB53 exerts neuroprotection in a mouse model of AMD, which can be attributed to its anti-retinal inflammatory effects by impeding RPE-mediated MMP-9 activation and anti-microglia.

Fusobacterium nucleatum induces colon anastomosis leak by activating epithelial cells to express MMP9.

Despite advances in anastomotic techniques and perioperative care, the incidence of anastomotic leak (AL) has not substantially decreased over time. Although it is known that AL etiology is multifactorial and the mechanisms involved remain unclear, there is accumulating evidence pointing at AL related to gut microbiota. We firstly performed a clinical study to analyze the gut microbiota between colorectal cancer patients who developed AL and those who did not (nAL) using 16S-rRNA sequencing and quantitative real-time PCR to identify AL risk bacterial taxa. Then we built a rat anastomosis model and performed a bacteria transplantation to ensure the cause-effect relationship. The anastomotic healing score was used to evaluate the healing of anastomosis. In addition, we assessed the adhesion ability of bacteria by staining with fluorescein isothiocyanate and attachment assay. The expression of matrix metalloproteinase 9 (MMP9) was detected by western blot, and the activity was detected by gelatin zymography. We found that the abundance and positive rate of Fusobacterium nucleatum (Fn) were higher in the AL patients. Exposure of the rat's colon anastomosis to Fn contributes to the loss of submucosa collagen I and III, leading to AL's pathogenesis. Fn can attach to the gut epithelial cells and stimulate intestinal MMP9 expression in vitro and in vivo. We further confirmed that these effects of Fn depended on the E-cadherin/β-catenin signaling pathway. This work demonstrates that Fn attaches and then stimulates the expression of epithelial cells MMP9 by the E-cadherin/β-catenin signaling pathway. These effects contribute to collagen break down in the intestinal tissue, finally leading to AL.

AMP-activated protein kinase inhibition in fibro-adipogenic progenitors impairs muscle regeneration and increases fibrosis.

Following muscle injury, fibro-adipogenic progenitors (FAPs) are rapidly activated and undergo apoptosis at the resolution stage, which is required for proper muscle regeneration. When excessive FAPs remain, it contributes to fibrotic and fatty infiltration, impairing muscle recovery. Mechanisms controlling FAP apoptosis remain poorly defined. We hypothesized that AMP-activated protein kinase (AMPK) in FAPs mediates their apoptosis during the muscle regeneration. To test, AMPKα1fl/fl PDGFRαCre mice were used to knock out AMPKα1 in FAPs. Following AMPKα1 knockout, the mice were injected with phosphate-buffered saline or glycerol to induce muscle injury. Tibialis anterior muscle and FAPs were collected at 3, 7 and 14days post-injury (dpi) for further analysis. We found that AMPKα1 deletion in FAPs enhanced p65 translocation to the nuclei by 110% (n=3; P<0.01). AMPKα1 knockout group had a higher gene expression of MMP-9 (matrix metalloproteinase-9) by 470% (n=3; P<0.05) and protein level by 39% (n=3; P<0.05). Loss of AMPKα1 up-regulated the active TGF-β1 (transforming growth factor-β1) levels by 21% (n=3; P<0.05). TGF-β promoted apoptotic resistance, because AMPKα1-deficient group had 36% lower cleaved Caspase 3 (cCAS3) content (n=3; P<0.05). Fibrotic differentiation of FAPs was promoted, with increased collagen protein level by 54% (n=3; P<0.05). Moreover, obesity decreased phosphorylation of AMPK by 54% (n=3; P<0.05), which decreased cCAS3 in FAPs by 44% (n=3; P<0.05) and elevated collagen accumulation (52%; n=3; P<0.05) during muscle regeneration. These data suggest that AMPK is a key mediator of FAPs apoptosis, and its inhibition due to obesity results in fibrosis of regenerated muscle.

Bumetanide Rescues Aquaporin-4 Depolarization via Suppressing β-Dystroglycan Cleavage and Provides Neuroprotection in Rat Retinal Ischemia-Reperfusion Injury

Aquaporin-4 (AQP4) regulates retinal water homeostasis and participates in retinal oedema pathophysiology. β-dystroglycan (β-DG) is responsible for AQP4 polarization and can be cleaved by matrix metalloproteinase-9 (MMP9). Retinal oedema induced by ischemia–reperfusion (I/R) injury is an early complication. Bumetanide (BU) has potential efficacy against cytotoxic oedema. Our study investigated the effects of β-DG cleavage on AQP4 and the roles of BU in a rat retinal I/R injury model. The model was induced by applying 110 mm Hg intraocular pressure to the anterior eye chamber. BU and U0126 (a selective ERK inhibitor) were intraperitoneally administered 15 and 30 min, respectively, before I/R induction. Rhodamine isothiocyanate extravasation detection, quantitative real-time PCR, transmission electron microscopy, hematoxylin-eosin staining, immunofluorescence staining, western blotting, and TUNEL staining were performed. AQP4 lost its polarization in the retinal perivascular domain as a result of β-DG cleavage. BU rescued AQP4 depolarization, suppressed AQP4 protein expression, attenuated retinal cytotoxic oedema, and downregulated β-DG and AQP4 mRNA expression. BU suppressed glial responses and mitochondria-mediated apoptotic protein expression, including that of Caspase-3 and Cyto C, raised the Bcl-2/Bax ratio, and lowered the number of apoptotic cells in the retina. Both BU and U0126 downregulated p-ERK and MMP9 expression. Thus, BU treatment suppressed β-DG cleavage, recovered AQP4 polarization partially via inhibiting ERK/MMP9 signaling pathway, and possess potential neuroprotective efficacy in the rat retinal ischemia–reperfusion injury model.

MMP-7 expression is associated with a higher rate of tumor spread through air spaces in resected lung adenocarcinomas

ObjectivesThe spread through air spaces (STAS) of adenocarcinoma (ADC) is a unique pattern for local invasion, which comprises the spread of tumor cells within air spaces beyond the tumor edge without a direct connection with the primary tumor. Matrix metalloproteinase-7 (MMP-7), a secreted proteolytic enzyme that degrades various extracellular matrix components and other substrates, regulates several pathophysiological processes as well as the occurrence and development of cancers in humans. Here, we retrospectively analyzed a cohort of Japanese patients with treatment-naive, surgically-resected lung ADC to assess whether MMP-7 is associated with STAS development and if it could be used as a predictor of STAS. Materials and methodsWe performed histological evaluation using hematoxylin and eosin staining and immunohistochemical analysis using microarrays. Thereafter, we scored the examined tissues for immune markers to identify significant tumor STAS predictors. ResultsWe identified that high MMP-7 expression is an independent predictor of a high STAS incidence. Multivariate analysis revealed that MMP-7 expression was correlated with tumor behavior and poor prognosis. Furthermore, STAS remained significantly associated with a higher risk of ADC recurrence. ConclusionThe development of tumor STAS could be promoted by the functioning of MMP-7. This study could be a crucial basis for future investigations on the detection of tumor STAS.

Sodium houttuyfonate against cardiac fibrosis attenuates isoproterenol-induced heart failure by binding to MMP2 and p38

BackgroundHeart failure (HF), caused by stress cardiomyopathy, is a major cause of mortality. Cardiac fibrosis is an essential structural remodeling associated with HF; therefore, preventing cardiac fibrosis is crucial to decelerating the progression of HF. Sodium houttuyfonate (SH), an extract of Houttuynia cordata, has a potent therapeutic effect on hypoxic cardiomyocytes in a myocardial infarction model. PurposeTo investigate the preventative and therapeutic effects of SH during isoproterenol (ISO)-induced HF and explore the pharmacological mechanism of SH in alleviating HF. MethodsWe analyzed the overlapping target genes between SH and cardiac fibrosis or HF using a network pharmacology analytical method. We verified the suppressive effect of SH on ISO-induced proliferation and activation of cardiac fibroblasts by immunohistochemical staining and histological analysis in an isoproterenol-induced HF mouse model. Additionally, we investigated the effect of SH by evaluating fibrosis and cardiac remodeling markers. To further decipher the pharmacological mechanism of SH against cardiac fibrosis and HF, we performed a molecular docking analysis between SH and hub common target genes. ResultsThere were 20 overlapping target genes between SH and cardiac fibrosis and 32 overlapping target genes between SH and HF. The 16 common target genes of SH against cardiac fibrosis and HF included MMP2 (matrix metalloproteinase 2), and p38. SH significantly inhibited the ISO- or TGF-β-induced expression of Col1α (collagen 1), α-SMA (smooth muscle actin), MMP2, TIMP2 (tissue inhibitor of metalloproteinase 2), TGF-β (transforming growth factor), and Smad2 phosphorylation. Moreover, both ISO- and TGF-β-induced p38 phosphorylation was inhibited. Molecular docking analysis showed that SH forms a stable complex with MMP2 and p38. ConclusionsIn addition to protecting cardiomyocytes, SH directly inhibits cardiac fibroblast activation and proliferation by binding to MMP2 and p38, subsequently delaying cardiac fibrosis and HF progression. Our prevention- and intervention-based approaches in this study showed that SH inhibited the development of stress cardiomyopathy-mediated cardiac fibrosis and HF when SH was administered before or after the initiation of cardiac stress.

C/EBP-β contributes to pig endometrial LE receptivity by targeting cell remodeling genes during implantation.

Transforming the endometrial luminal epithelium (LE) into a receptive state is a requisite event for successful embryo implantation. This study suggests the role of a transcription factor in regulating endometrial LE receptivity. The endometrial luminal epithelium (LE) undergoes extensive remodeling during implantation to establish receptivity of the uterus in response to the conceptus signals, such as interleukin 1β (IL1B). But the mechanisms remain to be fully understood. This study investigated the role of CCAAT/enhancer-binding protein β (C/EBP-β) in regulating pig endometrial LE receptivity. Our results showed that C/EBP-β was expressed and activated only in the endometrial LE in an implantation-dependent manner. In addition, C/EBP-β was highly activated at the pre-attachment stage compared to the attachment stage, and its activation was correlated with the expression of IL1B-dependent extracellular signal-regulated kinases1/2-p90 ribosomal S6 kinase signaling axis. Subsequent chromatin immunoprecipitation (ChIP)-sequencing analysis revealed that the binding of C/EBP-β within the promoter was positively associated with the transcription of genes related to cell remodeling. One such gene is matrix metalloproteinase 8 (MMP8), which is responsible for extracellular matrix degradation. The expression of MMP8 was abundant at the pre-attachment stage but dramatically declined at the attachment stage in the endometrial LE. Consistent with C/EBP-β, the expression and activation of MMP8 were limited to the endometrial LE in an implantation-dependent manner. Using ChIP-qPCR and electrophoresis mobility shift assay approaches, we demonstrated that C/EBP-β regulated the expression of the MMP8 gene during implantation. Furthermore, we detected that MMP8 and one of its substrates, type II collagen, showed a mutually exclusive expression pattern in pig endometrial LE during implantation. Our findings indicate that C/EBP-β plays a role in pig endometrial LE receptivity by regulating cell remodeling-related genes, such as MMP8, in response to conceptus signals during implantation.

Upregulation of KLHL17 promotes the proliferation and migration of non-small cell lung cancer by activating the Ras/MAPK signaling pathway

Analysis of the Gene Expression Profiling Interactive Analysis (GEPIA) database revealed that Kelch-like 17 (KLHL17) is overexpressed in non-small cell lung cancer (NSCLC) including adenocarcinoma (ADC) and squamous cell carcinoma (SCC). We therefore explored the role of KLHL17 in the development and progression of NSCLC. Immunohistochemistry and western blotting showed that KLHL17 expression was significantly higher in the tumor tissues from 173 patients with NSCLC, compared with the corresponding non-neoplastic tissue. In addition, upregulated KLHL17 expression was positively correlated with tumor size, lymph node metastasis and tumor node metastasis (TNM) stage, and affected the overall survival (OS) of patients with NSCLC. Consistent with clinical samples, in vitro studies demonstrated that KLHL17 expression was higher in various cell lines of NSCLC (A549, H1299, H460 and SK cells) as compared to normal human bronchial epithelial cells (HBE cells). Overexpression of KLHL17 in the cell lines of NSCLC with KLHL17-Flag plasmid promoted the proliferation and migration of tumor cells, which was associated with elevated activation of Rat sarcoma/Mitogen-activated protein kinases (Ras/MAPK) signaling and increased expression of cyclin D1, cyclin D-dependent kinases 4 (CDK4), matrix metalloproteinase 2 (MMP2) and Ras homolog gene family member A (RhoA). In contrast, knockdown of KLHL17 in the cell lines of NSCLC using KLHL17 small interfering RNA suppressed the proliferation and migration of tumor cells, in association with reduced activation of Ras/MAPK signaling and decreased expression of cyclin D1, CDK4, MMP2 and RhoA. Moreover, treatment of tumor cells with Ras inhibitor salirasib prevented KLHL17-induced Ras/MAPK activity as well as tumor proliferation and migration. These results suggest that upregulated KLHL17 in NSCLC promotes the proliferation and migration of tumor by activating Ras/MAPK signaling pathway. Therefore, KLHL17 may be a novel therapeutic target for the treatment of NSCLC.Non-small cell lung cancer (NSCLC) is the most common type of lung cancer and is associated with limited treatment options. This study elucidates the role of KLHL17 in the development and progression of NSCLC using clinical samples and NSCLC cell lines. The results show that upregulated KLHL17 in NSCLC promotes the proliferation and migration of tumor by activating Ras/MAPK signaling pathway, and suggest that KLHL17 may be a novel therapeutic target for the treatment of NSCLC.

Expression and clinical significance of miR-17-5p in tumor tissues of patients with colorectal cancer

At present, there is a lack of novel biomarkers for the early diagnosis and targeted therapy of colorectal cancer (CRC). The current study investigated the expression of miR-17-5p in colorectal tumors and adjacent tissues, and examined the effects of miR-17-5p on the cellular growth of the colon cancer cell line HCT-116. A total of 30 paired tissue specimens were obtained from patients with CRC who were admitted to the Department of General Surgery V of the First Affiliated Hospital, Gannan Medical College between December 2019 and January 2021. The clinical information for the corresponding patients was collated. Real-time fluorescence quantitative polymerase chain reaction (RT-qPCR) was performed to detect the expression of miR-17-5p in tumor tissues and the paracancerous tissues. The relationship between miR-17-5p and clinicopathology was analyzed. The CRC cell line HCT-116 was transfected with miR-17-5p mimics and inhibitors using liposomes. The effects of miR-17-5p on cell proliferation, invasion and migration, and apoptosis were determined using colon formation assays, transwell and scratch assays, and flow cytometry, respectively. The effects of miR-17-5p on CD44 (homing cell adhesion molecule), MMP2 (matrix metalloproteinase 2), BCL2 (B-cell lymphoma-2), BAX (BCL2-associated X), and E-cadherin protein and gene expression were investigated using Western blot and RT-qPCR, respectively. MiR-17-5p expression was higher in tumor tissues compared to the normal adjacent tissues. While miR-17-5p expression levels were unrelated to age nor gender, they were related to the degree of differentiation, stage, lymph node metastasis, and tumor size in patients with CRC. Upregulation of miR-17-5p promoted CRC cell proliferation, metastasis, and invasion, while inhibiting apoptosis. However, downregulation of miR-17-5p had the opposite effect. Furthermore, miR-17-5p upregulation increased E-cadherin, CD44, MMP2, and BCL2 expression while decreasing BAX expression, whereas miR-17-5p downregulation had the opposite effect. MiR-17-5p is highly expressed in tumor tissues and correlates with increased proliferation, invasion, and migration, as well as reduced apoptosis in HCT-116 cells. Indeed, miR-17-5p may be a potential indicator for the early diagnosis of CRC and may guide clinical targeted therapy.

Diagnostic and prognostic values of MMP-9 expression in ovarian cancer: A study based on bioinformatics analysis and meta-analysis.

This study aims to explore the expression of matrix metalloproteinase-9 (MMP-9) associated with both diagnostic and prognostic value in ovarian cancer by meta-analysis and bioinformatics analyses. We investigated the prognostic value of MMP-9 expression in ovarian cancer based on The Cancer Genome Atlas. Five databases were used to collect records about MMP-9 expression related to diagnostic and prognostic values in ovarian cancer from inception to June 2022. Using Stata 15.0 software, hazard ratio (HR) and odds ratio (OR) were calculated as the effect index of prognosis. We chose the pooled sensitivity, specificity, and area under the curve (AUC) to judge the diagnostic utility of MMP-9 for ovarian cancer. A total of 23 studies on prognosis, and five studies on diagnosis were entered into the meta-analysis. These suggest that high MMP-9 expression was detrimental to the overall survival of patients with ovarian cancer (HR = 1.34; 95% confidence interval (CI) 1.08∼1.66; P<0.01). High MMP-9 expression increased the risk of tumor stage (OR = 3.66; 95% CI 1.89∼7.07), but was not related to the tumor grade of ovarian cancer (P>0.05). The pooled analysis of serum MMP-9 diagnosing for ovarian cancer gave the pooled sensitivity, specificity, and AUC the values of 0.72 (95% CI 0.61∼0.81), 0.81 (95% CI 0.77∼0.85), and 0.84 (95% CI 0.81∼0.87), respectively. High MMP-9 expression can increase the tumor stage, and a correlation exists between high MMP-9 expression and poor prognosis in patients with ovarian cancer. Also, serum MMP-9 has a good diagnostic value for ovarian cancer.

Guggulsterone inhibits migration and invasion through proteasomal and lysosomal degradation in human glioblastoma cells

Glioblastoma multiforme (GBM) is a deadly brain malignancy, and current therapies offer limited survival benefit. The phytosterol guggulsterone (GS) has been shown to exhibit antitumor efficacy. This study aimed to investigate the effects of GS on migration and invasion and its underlying mechanisms in human GBM cell lines. After GS treatment, the survival rate of GBM cells was reduced, and the migration and invasion abilities of GBM cells were significantly decreased. There was also concomitant decreased expression of focal adhesion complex, matrix metalloproteinase-2 (MMP2), MMP9 and cathepsin B. Furthermore, GS induced ERK phosphorylation and autophagy, with increased p62 and LC3B-II expression. Notably, treatment of in GBM cells with the proteasome inhibitor MG132 or the lysosome inhibitor NH4Cl reversed the GS-mediated inhibition of migration and invasion. In an orthotopic xenograft mouse model, immunohistochemical staining of brain tumor tissues demonstrated that MMP2 and cathepsin B expression was reduced in GS-treated mice. GS treatment inhibited GBM cell migration and invasion via proteasomal and lysosomal degradation, suggesting its therapeutic potential in clinical use in the future.

Therapeutic Hypothermia Combined with Hydrogen Sulfide Treatment Attenuated Early Blood–Brain Barrier Disruption and Brain Edema Induced by Cardiac Arrest and Resuscitation in Rat Model

Brain injury remains a major problem in patients suffering cardiac arrest (CA). Disruption of the blood–brain barrier (BBB) is an important factor leading to brain injury. Therapeutic hypothermia is widely accepted to limit neurological impairment. However, the efficacy is incomplete. Hydrogen sulfide (H2S), a signaling gas molecule, has protective effects after cerebral ischemia reperfusion injury. This study showed that combination of hypothermia and H2S after resuscitation was more beneficial for attenuated BBB disruption and brain edema than that of hypothermia or H2S treatment alone. CA was induced by ventricular fibrillation for 4 min. Hypothermia was performed by applying alcohol and ice bags to the body surface under anesthesia. We used sodium hydrosulphide (NaHS) as the H2S donor. We found that global brain ischemia induced by CA and cardiopulmonary resuscitation (CPR) resulted in brain edema and BBB disruption; Hypothermia or H2S treatment diminished brain edema, decreased the permeability and preserved the structure of BBB during the early period of CA and resuscitation, and more importantly, improved the neurologic function, increased the 7-day survival rate after resuscitation; the combination of hypothermia and H2S treatment was more beneficial than that of hypothermia or H2S treatment alone. The beneficial effects were associated with the inhibition of matrix metalloproteinase-9 expression, attenuated the degradation of the tight junction protein occludin, and subsequently protected the structure of BBB. These findings suggest that combined use of therapeutic hypothermia and hydrogen sulfide treatment during resuscitation of CA patients could be a potential strategy to improve clinical outcomes and survival rate.

EF-24 inhibits TPA-induced cellular migration and MMP-9 expression through the p38 signaling pathway in cervical cancer cells.

Diphenyl difluoroketone (EF-24), a synthetic curcumin analog, has enhanced bioavailability over curcumin. EF-24 acts more powerful bioactivity for anti-inflammatory and anti-cancer activity. However, the effects and mechanism of EF-24 on cervical cancer has not been fully investigated. Herein, this study evaluated the effects of EF-24 on TPA-induced cellular migration of cervical cancer. The results showed that EF-24 substantially reduced the cellular migration and cellular invasion of the HeLa and SiHa cells. Moreover, gelatin zymography, western blotting analyses and real-time PCR revealed that EF-24 suppressed Matrix metalloproteinase-9 (MMP-9) activity, protein expression and mRNA levels. Mechanistically, EF-24 inhibited the phosphorylation of the p38 signaling pathway. In conclusion, EF-24 inhibited TPA-induced cellular migration and cellular invasion of cervical cancer cell lines through modulating MMP-9 expression via downregulating signaling p38 pathway and EF-24 may have potential to serve as a chemopreventive agent of cervical cancer.

Serum amyloid A1 recruits neutrophils to the invasive front of T1 colorectal cancers.

The tumor microenvironment plays an essential role in the development and progression of colorectal cancer (CRC). We recently reported that crosstalk between CRC cells and tumor-associated macrophages (TAMs) via serum amyloid A1 (SAA1) promotes invasion by T1 CRCs. In the present study, we aimed to clarify the role of neutrophils in early CRCs. Immunohistochemical analysis of CD66b, chemokine CXC motif ligand 8 (CXCL8 or interleukin-8, IL-8) and matrix metalloproteinase-9 (MMP-9) was performed using primary T1 CRCs (n=49). The HL-60 human promyelocytic leukemia cell line and THP-1 human monocytic leukemia cell line were used to obtain neutrophil-like and macrophage-like cells, respectively. Boyden chamber assays were used to analyze cell migration and invasion, and quantitative RT-PCR was used to analyze gene expression. Immunohistochemical analysis revealed accumulation of neutrophils at the SAA1-positive invasive front of T1 CRCs. Experiments using HL-60 cells suggested that treatment with SAA1 induced neutrophil migration and expression of CXCL8 and MMP-9 in neutrophils and that neutrophils promote CRC cell migration and invasion. Immunohistochemistry confirmed accumulation of CXCL8- or MMP-9-positive neutrophils at the SAA1-positive invasive front of T1 CRCs. Moreover, co-culture experiments using CRC, THP-1 and HL-60 cells suggested that CRC cells activated by macrophages upregulate CXCL8 and MMP-9 in neutrophils. Our results suggest that interplay between macrophages and CRC cells leads to recruitment of neutrophils to the invasive front of T1 CRCs and that SAA1 secreted by CRC cells activate neutrophils to promote invasion.

Astragalus polysaccharide protects against blood-brain barrier damage in MCAO rats by inhibiting P2X7R channel

To investigate the protective effect of astragalus polysaccharide (APS) against blood-brain barrier in a rat model of middle cerebral artery occlusion (MCAO) and the role of P2X7R channel in the protective mechanism. In rat microglial cell models of oxygen and glucose deprivation (OGD) or ATP treatment, the formation of blood-brain barrier in vitro was assessed using the leak test, and the effect of APS on the permeability of the blood-brain barrier was determined using LC-MS. In 12 SD rats, MCAO model was established followed by treatment with intraperitoneal injection of normal saline (n= 6) or APS (45 mg/kg, n=6) for 3 consecutive days, with another 6 rats without MCAO receiving saline injections as the control group. The permeability of the blood-brain barrier of the rats was evaluated by determining Evans blue (EB) extravasation, and ATP content in the brain tissue was detected using ELISA; the expression levels of matrix metalloproteinase-9 (MMP-9) and P2X7R in the brain tissue were detected with Western blot. In the in vitro cell model of OGD or ATP treatment, APS treatment obviously promoted the repair of blood-brain barrier integrity. In the rat models, the EB content in the brain tissue and the blood-brain barrier permeability increased significantly in MCAO+saline group and MCAO+APS group as compared with those in the control group (P < 0.01). Compared with saline treatment, APS treatment significantly decreased EB content in the brain tissue and improved the blood-brain barrier permeability in the MCAO rats (P < 0.05). MCAO caused a significant reduction of ATP content and obviously increased the expression levels of MMP-9 and P2X7R in the brain tissue of the rats (P < 0.01), and these changes were significantly alleviated after APS treatment (P < 0.01 or 0.05). APS can protect the brain tissue of MCAO rats by stabilizing the internal environment, down-regulating the expression of MMP-9 and improving the permeability of blood-brain barrier under cerebral ischemia and hypoxia, and its mechanism may involve the inhibition of P2X7R channel.

The diagnostic and prognostic potential of the EGFR/MUC4/MMP9 axis in glioma patients

Glioblastoma is the most aggressive form of brain cancer, presenting poor prognosis despite current advances in treatment. There is therefore an urgent need for novel biomarkers and therapeutic targets. Interactions between mucin 4 (MUC4) and the epidermal growth factor receptor (EGFR) are involved in carcinogenesis, and may lead to matrix metalloproteinase-9 (MMP9) overexpression, exacerbating cancer cell invasiveness. In this study, the role of MUC4, MMP9, and EGFR in the progression and clinical outcome of glioma patients was investigated. Immunohistochemistry (IHC) and immunofluorescence (IF) in fixed tissue samples of glioma patients were used to evaluate the expression and localization of EGFR, MMP9, and MUC4. Kaplan–Meier survival analysis was also performed to test the prognostic utility of the proteins for glioma patients. The protein levels were assessed with enzyme-linked immunosorbent assay (ELISA) in serum of glioma patients, to further investigate their potential as non-invasive serum biomarkers. We demonstrated that MUC4 and MMP9 are both significantly upregulated during glioma progression. Moreover, MUC4 is co-expressed with MMP9 and EGFR in the proliferative microvasculature of glioblastoma, suggesting a potential role for MUC4 in microvascular proliferation and angiogenesis. The combined high expression of MUC4/MMP9, and MUC4/MMP9/EGFR was associated with poor overall survival (OS). Finally, MMP9 mean protein level was significantly higher in the serum of glioblastoma compared with grade III glioma patients, whereas MUC4 mean protein level was minimally elevated in higher glioma grades (III and IV) compared with control. Our results suggest that MUC4, along with MMP9, might account for glioblastoma progression, representing potential therapeutic targets, and suggesting the ‘MUC4/MMP9/EGFR axis’ may play a vital role in glioblastoma diagnostics.