MMP-9 mRNA Research Articles

Semaphorin 4C Plays a Key Role in Colorectal Cancer Cells Development

Objective: Purpose of this work was to discuss effects and mechanisms of Sema 4C in colon cancer development. Results: Sema4C were significantly upregulated in cancer tissues (P <0.001). Following transfection, the expression levels of Sema4C mRNA were significantly downregulated in the si-Sema4C groups compared with those in the si-negative control groups of both HT-29 and SW620 cells (both P <0.001). The apoptotic rate was significantly increased, while the invasive and wound healing rates were significantly suppressed in the si-Sema4C groups of HT-29 and SW620 cells (both P < 0.001). The results of the RT-qPCR and western blotting analyses revealed that PI3K, AKT, MMP-2 and MMP-9 mRNA and protein expression levels, respectively, were significantly downregulated, while caspase-3 mRNA and protein expression levels were significantly upregulated in the si-Sema4C groups of HT-29 and SW620 cells. Conclusion: The findings of the present study demonstrated that the knockdown of Sema4C expression suppressed CRC cell biological activities by regulating the PI3K/AKT signaling pathway. Therefore, Sema4C may act as an oncogene in CRC.

Transforming Growth Factor β1/Vascular Endothelial Growth Factor Signaling Pathway Regulates Chondrocytes Dedifferentiation

This study investigated the interrelation between TGF-β and VEGF during CH dedifferentiation. A high dose of TGF-β1 was used to induce CHs dedifferentiation, and the effect of the TGF-β receptor inhibitor containing ALK1 and ALK5 on VEGF expression was explored. A VEGF inhibitor was used to investigate whether it prevented TGF-β1-induced CHs dedifferentiation. Low-dose TGF-β1 contributed to the steady-state of CHs, but a high dose significantly decreased collagen II and SOX-9 expression and increased collagen X, Runx2, and MMP-13 mRNA expression, accompanied by a marked upregulation of VEGF. ALK5 inhibition exacerbated the side effect caused by high-dose TGF-β1, while ALK1 inhibition had the opposite effect. Suppression of ALK1 and ALK5 contributed to the prevention of the VEGF level under high-dose TGF-β1 conditions. The suppression of VEGF protected the content of chondrogenic genes and suppressed the expression of dedifferentiated genes. In conclusion, high-dose TGF-β1 upregulated VEGF expression by ALK1 and ALK5 activation, resulting in CHs dedifferentiation. Suppression of ALK1 is an effective way to interrupt the TGF-β1/VEGF signaling pathway induced CH dedifferentiation.

Altered choline level in atherosclerotic lesions: Upregulation of choline transporter-like protein 1 in human coronary unstable plaque.

Inflammatory activity and hypoxia in atherosclerotic plaques are associated with plaque instability and thrombotic complications. Recent studies show that vascular cell metabolism affects atherogenesis and thrombogenicity. This study aimed to identify the metabolites in macrophage-rich unstable plaques that modulate atherogenesis and serve as potential markers of plaque instability. Atherosclerotic plaques were induced by balloon injury in the iliofemoral arteries of rabbits fed on a conventional or 0.5% cholesterol diet. At 3 months post-balloon injury, the arteries and cardiac tissues were subjected to histological, quantitative real-time polymerase chain reaction, and metabolomic analyses. The identified metabolite-related proteins were immunohistochemically analyzed in stable and unstable plaques from human coronary arteries. The factors modulating the identified metabolites were examined in macrophages derived from human peripheral blood mononuclear cells. Metabolomic analysis revealed that choline and guanine levels in macrophage-rich arteries were upregulated compared with those in non-injured arteries and cardiac tissues. Vascular choline levels, but not guanine levels, were positively correlated with the areas immunopositive for macrophages and tumor necrosis factor (TNF)-α and matrix metalloproteinase (MMP) 9 mRNA levels in injured arteries. In human coronary arteries, choline transporter-like protein (CTL) 1 was mainly localized to macrophages within plaques. The area that was immunopositive for CTL1 in unstable plaques was significantly higher than that in stable plaques. Intracellular choline levels were upregulated upon stimulation with TNF-α but were downregulated under hypoxia in cultured macrophages. Administration of choline upregulated the expression of TNF-α and CTL1 mRNA in cultured macrophages. The transfection of CTL1 small interfering RNA decreased CTL1, TNF-α, and MMP9 mRNA levels in cultured macrophages. These results suggest that choline metabolism is altered in macrophage-rich atherosclerotic lesions and unstable plaques. Thus, CTL1 may be potential markers of plaque instability.

Cornelian Cherry (Cornus mas L.) Iridoid and Anthocyanin-Rich Extract Reduces Various Oxidation, Inflammation, and Adhesion Markers in a Cholesterol-Rich Diet Rabbit Model.

Atherogenesis leads to the development of atherosclerosis, a progressive chronic disease characterized by subendothelial lipoprotein retention and endothelial impairment in the arterial wall. It develops mainly as a result of inflammation and also many other complex processes, which arise from, among others, oxidation and adhesion. Cornelian cherry (Cornus mas L.) fruits are abundant in iridoids and anthocyanins-compounds with potent antioxidant and anti-inflammatory activity. This study aimed to determine the effect of two different doses (10 mg and 50 mg per kg of body weight, respectively) of iridoid and anthocyanin-rich resin-purified Cornelian cherry extract on the markers that are important in the progress of inflammation, cell proliferation and adhesion, immune system cell infiltration, and atherosclerotic lesion development in a cholesterol-rich diet rabbit model. We used biobank blood and liver samples that were collected during the previous original experiment. We assessed the mRNA expression of MMP-1, MMP-9, IL-6, NOX, and VCAM-1 in the aorta, and the serum levels of VCAM-1, ICAM-1, CRP, PON-1, MCP-1, and PCT. The application of the Cornelian cherry extract at a dose of 50 mg/kg bw resulted in a significant reduction in MMP-1, IL-6, and NOX mRNA expression in the aorta and a decrease in VCAM-1, ICAM-1, PON-1, and PCT serum levels. The administration of a 10 mg/kg bw dose caused a significant decrease in serum ICAM-1, PON-1, and MCP-1. The results indicate the potential usefulness of the Cornelian cherry extract in the prevention or treatment of atherogenesis-related cardiovascular diseases, such as atherosclerosis or metabolic syndrome.

Analyses of Transcriptomics upon IL-1β-Stimulated Mouse Chondrocytes and the Protective Effect of Catalpol through the NOD2/NF-κB/MAPK Signaling Pathway

The overall objective of this study was to investigate the mechanism of inflammation on chondrocyte injury and the protective effect of catalpol on chondrocytes in an inflammatory environment. Chondrocytes were isolated and cultured from the knee joints of three-day-old newborn mice. Alcian Blue staining and the immunocytochemistry staining of type II collagen were used to identify the purity of chondrocytes. Primary chondrocytes were stimulated by IL-1β (10 ng/mL) and subjected to transcriptome analysis. Differentially expressed genes (DEGs) were further analyzed based on Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. In this experimental study, we performed the viability assay to determine the effects of different concentrations of catalpol on the cell viability of chondrocytes. Chondrocytes were seeded in six-well plates and exposed to 10 μM catalpol 2 h prior to treatment with IL-1β (10 ng/mL). Quantitative real-time (qPCR) and Western blotting were performed to evaluate the RNA and protein expression, respectively. Based on the results of transcriptomics analysis, we found the NOD2 signaling pathway, the NF-kappa B signaling pathway, and the MAPK signaling pathway showed significant changes in chondrocyte damage caused by inflammation. Catalpol (10 μM and 100 μM) could significantly reduce NO, IL-6, IL-1β, and TNF-α in supernatant of chondrocytes. Catalpol significantly inhibited the mRNA expression of IL-1, IL-6, and IL-12 in chondrocytes induced by IL-1β. Catalpol markedly inhibited MMP3, MMP13 mRNA, and protein levels. Catalpol could significantly reduce TNF-α mRNA levels in inflammatory chondrocytes. Inflammation causes significant increases in mRNA levels and protein levels of NOD2, mRNA levels, and protein levels were markedly suppressed by catalpol. In addition, catalpol could significantly increase IKBα protein levels and significantly lower intranuclear P65 levels. Catalpol significantly lowered the phosphorylation protein levels of ERK, p38, and JNK. Our transcriptomic analysis demonstrated that the activation of NOD2 and its downstream pathways, NF-κB and MAPK, is an important cause of the inflammatory injury to chondrocytes induced by IL-1β. Catalpol inhibited the activation of the NOD2 signaling pathway, which reduced the phosphorylation of ERK, p38, and JNK, inhibited the degradation of IκBα, inhibited p65 translocation into the nucleus, reduced the release of inflammatory cytokines, and attenuated the inflammatory damage to chondrocytes.

Inhibitory effect of non-alcoholic steatohepatitis on colon cancer liver metastasis.

The incidence of non-alcoholic steatohepatitis (NASH) is dramatically increasing, but the effect of NASH on colon cancer liver metastasis (CLM) is controversial. The aim of this study was to investigate the impact and mechanism of action of NASH on CLM using a western diet (WD)-fed mouse model. Six-week-old male C57BL/6J mice were used. They were divided into the WD group and control group with normal diet. MC38 colon cancer cells were injected into the spleen at 2, 6, 8 and 16 weeks, and mice were killed at 2 weeks after injection to evaluate hepatic steatosis, fibrosis, metastasis and mRNA/protein expression in the liver. Only mice fed a WD for 16 weeks showed hepatic fibrosis. These mice showed significantly higher alanine aminotransferase and total cholesterol levels compared with the control group (p<0.05). The WD group showed significantly lower tumor number and smaller tumor diameter (p<0.05). In the WD group, expression of SAA1, IL6, STAT3 and MMP9 mRNA in the liver was significantly lower than in the control group (p<0.05). Serum amyloid A1 protein expression was also lower in the WD group. The WD-fed NASH mouse model showed an inhibitory effect on CLM. Suppressed interleukin-6/signal transducer and activator of transcription 3 signaling and serum amyloid A/matrix metalloproteinase 9 expression may affect this phenomenon.

Cinnamaldehyde modulates host immunoinflammatory responses in rat ligature-induced periodontitis and peripheral blood mononuclear cell models.

Cinnamaldehyde is a natural product with anti-inflammatory and immune-modulatory properties, known to regulate host responses to bacterial stimuli. This study aimed to investigate the effects of cinnamaldehyde on ligature-induced periodontitis in rats, and its impact on the modulation of human peripheral blood mononuclear cells (PBMC). Male Wistar rats were assigned into three groups:i) control: no ligature+vehicle; ii) ligature: ligature+vehicle; and iii) ligature+cinnamaldehyde (50mg/kg); all treatments by daily oral gavage. After 14days of induced periodontitis, the hemimandibles were collected for bone loss evaluation. The gingival levels of IL-1β, MMP-9 and iNOS mRNA were evaluated. Nitric oxide (NO) was measured in both rat saliva and plasma. PBMC were stimulated with Aggregatibacter actinomycetemcomitans (Aa) in the presence or absence of cinnamaldehyde (5, 20 e 40µM), and cytokine production was quantified in cell supernatant. Proliferating lymphocytes were taken for flow cytometer reading, while culture supernatants were used for IFN-γ and IL-10 assessment. The ligature group had both increased alveolar bone loss and gingival expression of IL-1β, MMP-9 and iNOS compared to the control group. All parameters were attenuated by cinnamaldehyde treatment. Lower salivary but not plasma NO was detected in the cinnamaldehyde compared to the ligature group. Aa-stimulated PBMCs treated with cinnamaldehyde produced less IL-1β; the compound also attenuated lymphocyte proliferation in a dose-dependent manner, as well as cell IL-10 production. Cinnamaldehyde treatment reduced periodontal bone loss, and downregulated key inflammatory mediators and human PBMC responses, pointing to novel potential therapeutic effects of this compound.

Downregulated ETV4 inhibits the proliferation, migration, and invasion of trophoblast cells in preeclampsia.

Preeclampsia is a pregnancy complication that can lead to severe adverse maternal and fetal outcomes, but the mechanisms underlying the development of preeclampsia are not fully understood. This study shows that ETV4 plays an essential role in the proliferation, invasion, and migration of trophoblast cells by regulating MMP-2 and MMP-9 and is involved in the pathogenesis of preeclampsia. Preeclampsia (PE) is a pregnancy complication that can lead to severe adverse maternal and fetal outcomes. However, the mechanisms underlying the development of PE are not fully understood. ETS Variant Transcription Factor 4 (ETV4) plays an important role in cell proliferation, migration, and invasion. In this study, we aimed to explore the potential function of ETV4 in placental trophoblast cells. We analyzed the expression and location of ETV4 in PE and uncomplicated placental tissues using RT-qPCR, Western blotting, immunohistochemistry, and immunofluorescence staining. The results showed that both the mRNA and protein levels of ETV4 were markedly decreased in PE placental tissues compared with placental tissues from women with uncomplicated pregnancies (P < 0.05). Then, the effects of ETV4 on HTR-8/SVneo and Bewo cell proliferation, migration, and invasion were evaluated by MTT, 5-ethynyl-2-deoxyuridine (EdU), wound healing, and Transwell assays, respectively. The results showed that ETV4 knockdown inhibited both HTR-8/SVneo and Bewo cell proliferation, migration, and invasion (P < 0.05). Conversely, overexpression of ETV4 promoted both HTR-8/SVneo and Bewo cell proliferation, migration, and invasion (P < 0.05). We then measured the expression of MMP-2 and MMP-9 in HTR8/SVneo cells. We found that ETV4 knockdown decreased the mRNA and protein expression of MMP-2 and MMP-9, while ETV4 overexpression increased MMP-2 and MMP-9 mRNA and protein expression (P < 0.05). In conclusion, ETV4 plays an essential role in the proliferation, invasion, and migration of trophoblast cells by regulating MMP-2 and MMP-9. Our findings provide novel insight into the mechanisms underlying the occurrence of PE.

Vitamin A- and D-Deficient Diets Disrupt Intestinal Antimicrobial Peptide Defense Involving Wnt and STAT5 Signaling Pathways in Mice.

Vitamin A and D deficiencies are associated with immune modulatory effects and intestinal barrier impairment. However, the underlying mechanisms remain unclear. C57BL/6J mice were fed either a diet lacking in vitamin A (VAd), vitamin D (VDd) or a control diet (CD) for 12 weeks. Gut barrier function, antimicrobial peptide (AMP) defense and regulatory pathways were assessed. VAd mice compared to CD mice showed a reduced villus length in the ileum (p &lt; 0.01) and decreased crypt depth in the colon (p &lt; 0.05). In both VAd- and VDd-fed mice, ileal α-defensin 5 (p &lt; 0.05/p &lt; 0.0001 for VAd/VDd) and lysozyme protein levels (p &lt; 0.001/p &lt; 0.0001) were decreased. Moreover, mRNA expression of lysozyme (p &lt; 0.05/p &lt; 0.05) and total cryptdins (p &lt; 0.001/p &lt; 0.01) were reduced compared to controls. Furthermore, matrix metalloproteinase-7 (Mmp7) mRNA (p &lt; 0.0001/p &lt; 0.001) as well as components of the Wnt signaling pathway were decreased. VAd- and VDd-fed mice, compared to control mice, exhibited increased expression of pro-inflammatory markers and β-defensins in the colon. Organoid cell culture confirmed that vitamins A and D regulate AMP expression, likely through the Jak/STAT5 signaling pathway. In conclusion, our data show that vitamin A and D regulate intestinal antimicrobial peptide defense through Wnt and STAT5 signaling pathways.

Development on potential skin anti-aging agents of Cosmos caudatus Kunth via inhibition of collagenase, MMP-1 and MMP-3 activities

BackgroundSkin aging is associated with degradation of collagen by matrix metalloproteinases (MMPs), which leads to loss of skin elasticity and formation of wrinkles. Cosmos caudatus Kunth (CC) has been traditionally claimed as an anti-aging agent in Malaysia. Despite its well-known antioxidant activity, the anti-aging properties of CC was not validated. PurposeThis study aimed to investigate the anti-aging potential of CC extracts and fractions, particularly their inhibition of collagenase, MMP-1 and MMP-3 activities in human dermal fibroblasts CCD-966SK, followed by isolation, identification and analysis of their bioactive constituents. Study design and methodsDPPH assay was firstly used to evaluate the antioxidant activity throughout the bioactivity-guided fractionation. Cell viability was determined using MTS assay. Collagenase activity was examined, while MMP-1 and MMP-3 expression were measured using qRT-PCR and western blotting. Then, chemical identification of pure compounds isolated from CC fractions was done by using ESIMS, 1H and 13C NMR spectroscopies. HPLC analyses were carried out for bioactive fractions to quantify the major components. ResultsThroughout the antioxidant activity-guided fractionation, fractions CC-E2 and CC-E3 with antioxidant activity and no toxicity towards CCD-966SK cells were obtained from CC 75% ethanol partitioned layer (CC-E). Both fractions inhibited collagenase activity, MMP-1 and MMP-3 mRNA and protein expression, as well as NF-κB activation induced by TNF-α in CCD-966SK cells. 14 compounds, which mainly consists of flavonoids and their glycosides, were isolated. Quercitrin (14.79% w/w) and quercetin (11.20% w/w) were major compounds in CC-E2 and CC-E3, respectively, as quantified by HPLC. Interestingly, both fractions also inhibited the MMP-3 protein expression synergistically, compared with treatment alone. ConclusionThe quantified CC fractions rich in flavonoid glycosides exhibited skin anti-aging effects via the inhibition of collagenase, MMP-1 and MMP-3 activities, probably through NF-κB pathway. This is the first study reported on MMP-1 and MMP-3 inhibitory activity of CC with its chemical profile, which revealed its potential to be developed as anti-aging products in the future.

Resveratrol suppresses lipopolysaccharide-mediated activation of osteoclast precursor RAW 264.7 cells by increasing miR-181a-5p expression

Resveratrol (Res) has anti-inflammation and antiosteoporosis functions. We evaluated the effect of Res on osteoclast differentiation by releasing inflammatory cytokines from osteoclast precursor RAW 264.7 cells stimulated by lipopolysaccharide (LPS). In the study, LPS (1ng/L) was used to induce the Raw 264.7 inflammatory injury model in vitro. A total of 25ng/mL M-CSF + 30ng/mL RANKL or plus 1μg/L LPS was used to induce osteoclastogenesis in the experiments. We utilized the Cell Counting Kit-8 assay to measure the relative cell survival of RAW 264.7 cells. Then, enzyme-linked immunosorbent assays were utilized to measure the abundance of inflammatory markers, such as interleukin-1 beta (IL-1β), tumor necrosis factor-alpha (TNF-α), and IL-6. Subsequently, Western blot analysis was applied to assess the abundance of phosphorylated transforming growth factor beta-activated kinase 1 (P-TAK1) protein, TNF receptor-associated factor 6 (TRAF6), nuclear factor-κB inhibitor protein (IκB), phosphorylated IκB-α(P-IκB-α), and nuclear factorκB65 (NF-κB65). mRNA expression levels of miR-181a-5p, TRAF6, specific gene calcitonin receptor (CTR), activated T nuclear factor 1 (NFATC1), cathepsin K (CTSK), and matrix metalloproteinase (MMP)-9 were determined via a real-time polymerase chain reaction. Osteoclast bone resorption function was determined. Finally, tartrate-resistant acid phosphatase (TRAP) staining was performed.The results found that Compared with the model group, the degrees of expressions of supernatant inflammatory factors TNF-α, IL-1β, and IL-6 were substantially attenuated in the Res treatment group (p< 0.05). Furthermore, the extent of miR-181a-5p expression in the RAW 264.7 cells significantly increased, whereas P-IκB-α, P-TAK1, NF-κB65, and TRAF6 expressions significantly decreased in the Res treatment group as opposed to the model group (p< 0.05). The CTR, NFATC1, MMP-9, CTSK, and TRAP mRNA expression levels were substantially reduced during osteoclast differentiation and bone resorption in the Res treatment group.The results suggest that Res can reduce the RAW 264.7 cell differentiation into osteoclasts and relieve LPS-stimulated osteoporosis, and the underlying mechanism may be associated with the Res-inhibited activity of the TRAF6/TAK1 pathway through the increased miR-181a-5p expression.

Wogonin Restrains the Malignant Progression of Lung Cancer Through Modulating MMP1 and PI3K/AKT Signaling Pathway.

Wogonin, a natural flavonoid compound, represses cancer cell growth and induces cancer cell apoptosis in diverse malignancies. However, the function of Wogonin in lung cancer cells and its regulatory mechanism deserve to be identified. A549 and H460 cells were treated with Wogonin, and the cell growth, apoptosis, migration and invasion were measured by CCK-8 and EdU, flow cytometry and Transwell assays. The targeted genes of Wogonin and lung cancer were identified from the TCMSP and Genecards databases, respectively. The STRING database and Cytoscape software were used to establish a PPI network and screen hub genes. GO and KEGG analysis was conducted to explore the functions and signal pathways related to the hub genes. MMP1 expression in lung cancer was analyzed using the UALCAN databases, and GSEA was performed utilizing LinkedOmics. Gelatin zymography assay was used to detect MMP1 activity. MMP1 mRNA expression was detected by qRT-PCR. Besides, MMP1, p-AKT and c-Myc protein were detected by Western Blot assay. Wogonin could suppress the proliferation, migration and invasion of A549 and H460 cells and induce apoptosis. GO and KEGG enrichment analysis revealed the hub genes were mostly enriched in re-entry into the mitotic cell cycle and apoptosis. The expression of MMP1 was markedly upregulated in lung squamous cell carcinoma, lung adenocarcinoma tissues, and lung cancer cell lines. Wogonin could significantly inhibit MMP1 expression and activity, and overexpression of MMP1 significantly reversed the effect of Wogonin on the malignant phenotypes of A549 and H460 cells. Wogonin inhibited the expression of p-AKT and c-Myc protein by regulating MMP1. Wogonin can repress lung cancer cells' growth and metastatic potential and promote cell apoptosis via repressing MMP1 expression and modulating PI3K/AKT signaling pathway.

PS-BPB01-3: ANGIOTENSIN (1–7)-MEDIATED PREVENTIVE EFFECT OF ANGIOTENSIN II TYPE 1 RECEPTOR ANTAGONISTS ON ABDOMINAL AORTIC ANEURYSM PROGRESSION IN OPG-KO MICE

Objective: Hypertension is one of the major risk factors for onset of abdominal aortic aneurysm (AAA), which is a local enlargement of the abdominal aorta. Therefore, anti-hypertensive drugs, including angiotensin (Ang) II type 1 receptor blocker (ARB), have been expected to limit AAA growth. However, the efficacy of ARB against AAA in clinical settings is controversial. In the present study, we attempted to elucidate a mechanism by which ARB inhibits AAA progression. Design and method: To examine the effect of ARB against AAA progression, olmesartan (20 mg/kg/day) was administered orally to male wild-type mice and osteoprotegerin (Opg)-knockout (KO) mice, from two weeks before the AAA induction by using a CaCl2 application on the abdominal aortae to the sacrifice date. Since Opg could bind to tumor necrosis factor-related apoptosis-inducing ligand (Trail) for inhibiting its activity, deficiency of the Opg gene has been reported to induce overactivation of the Trail induced-c-Jun N-terminal kinase (Jnk)/matrix metalloproteinase 9 (Mmp9) pathway in CaCl2-induced mouse AAA model, resulting in serious AAA progression. Here, we tested whether olmesartan could prevent AAA progression associated with Trail/Jnk/Mmp9 pathway. Results: We found that olmesartan prevented excessive aortic enlargement and serious degenerative change of the aortic tissue in Opg-KO mice at six weeks after AAA induction, but not affected AAA formation in wild-type mice. In Opg-KO mice, olmesartan reduced co-localized expression of phosphorylated Jnk and Mmp9 in the tunica media. Then, we tested whether Ang (1–7), the blood level of which is increased by ARB administration, is associated with the preventive effect of olmesartan, because Ang (1–7) had been reported to reduce the Jnk phosphorylation. Consistent with previous studies, olmesartan increased serum level of Ang (1–7) in our AAA model. In vascular smooth muscle cells, Ang (1–7) inhibited the phosphorylation of Jnk and the increase of downstream Mmp9 mRNA expression induced by Trail. Furthermore, administration of an Ang (1–7) antagonist, A779, to Opg-KO mice diminished the preventive effect of ARB against the AAA progression. Notably, olmesartan slightly lowered blood pressure in Opg-KO mice, while A779 did not affect the blood pressure-lowering effect of olmesartan. Conclusions: Our results indicate that ARB prevents the AAA progression in Opg-KO mice via the inhibition of Trail-induced Jnk-Mmp9 pathway. Furthermore, it is suggested that the increase of Ang (1–7) may play a key role in the preventive effect of ARB on the AAA progression.

PATTERN OF MMP2 AND MMP9 EXPRESSION DEPENDS ON BREAST CANCER PATIENTS' AGE.

Despite the large number of studies devoted to the study of the features of tumor microenvironment in breast cancer (BCa), presently there is no consensus on the features of MMP-2 and MMP-9 expression in the tumor tissue of BCa patients depending on the age. The aim of the study was to investigate the relationship between MMP-2 and -9 expression at the protein and mRNA levels in BCa tissues and the clinical and pathological features of BCapatientsin different age groups. The expression level of MMP-2 and -9in the BCa tissue of patients of two age groups (< 45 years and > 45 years) was studied using the bioinformatics method (UALCAN database), immunohistochemical method, and real-time PCR. It was established that a characteristic feature of BCa in young patients is the low level of MMP2 mRNA against the background of increased expression of this gelatinase at the protein level, as well as decreased expression of MMP9 at both the mRNA and protein levels. When analyzing the correlation of the gelatinase expression indices in BCa tissue of young patients, depending on the clinical and pathological features, a significantly lower level of MMP-2 expression was recorded in BCa cases of stage II compared to the indices of stage I cases. High expression of MMP-2 and -9 was recorded in BCa tissue in node-positive cases and the basal molecular BCa subtype. The identified relationship between the expression of the studied gelatinases and such indices of BCa malignancy as its stage, positive regional lymph node status, and the molecular BCa subtype in young patients indicates the need for further research of the features of the tumor microenvironment to predict the cancer aggressiveness.

Sidaxue, a traditional Guizhou Miao herbal medicine formula, reduces necroptosis and synovial vascular proliferation in rats with collagen-induced arthritis by inhibiting the RIPK1/RIPK3/MLKL pathway

To explore the inhibitory effect of Sidaxue (SX), a traditional Guizhou Miao herbal medicine formula, on necrotic apoptosis and synovial angiogenesis in rats with collagen-induced arthritis (CIA) and the role of the RIPK1/RIPK3/MLKL pathway in mediating this effect. Forty-two SD rats were randomized into 6 groups (n=7), including a normal control group, a CIA model group, 3 SX treatment groups at low (10 g/kg), moderate (20 g/kg) and high (40 g/kg) doses, and a GTW treatment group. CIA rat models were established by subcutaneous injections of bovine type II collagen, and the treatments were administered daily by gavage for 21 days. The rats were observed for swelling of the hind limb joints, which was rated using the arthritis index (AI) score on a weekly basis. Serum levels of TNF-α, IL-1β and IL-17 in the rats were detected using ELISA, and the pathological changes in the synovium were observed with HE staining. Real-time PCR was performed to detect the mRNA expression levels of VEGF, MMP-9, Ang-1, RIPK1, RIPK3, and caspase-8 in the synovial tissues, and the protein expressions of VEGF, MMP9, Ang-1, Stat-3, RIPK1, RIPK3, MLKLl, p-MLKL and caspase-8 were detected using Western blotting. Compared with those in CIA model group, the rats receiving treatment with GTW and SX showed milder swelling of the hind limb joints with significantly lower AI scores (P < 0.05). In CIA model group, a large number of inflammatory cells were observed in the synovium with obvious damages of the tissue structure. In the drug treatment groups, inflammatory cell infiltration, synovial angiogenesis and synovial hyperplasia were alleviated, and the therapeutic effects were obviously enhanced as SX dose increased. Compared with those in the model group, the rats treated with GTW and high-dose SX showed significantly decreased serum levels of IL-1β, IL-17 and TNF-α (P < 0.05), lower mRNA and protein expressions of RIPK1, RIPK3, VEGF, Ang-1, and MMP9 (P < 0.05), higher expressions of caspase-8 (P < 0.01), and obviously lowered expression of Stat-3 protein and phosphorylation level of MLKL (P < 0.05). SX can improve synovial angiogenesis in CIA rats possibly by inhibiting the activation of RIP1/RIP3/MLKL signaling pathway and down-regulating the expression of the vascular growth factors VEGF, Ang-1, MMP9, and Stat-3.

Effect of Secretome-Hypoxia Mesenchymal Stem Cells on Regulating SOD and MMP-1 mRNA Expressions in Skin Hyperpigmentation Rats

BACKGROUND: Ultraviolet B (UVB) radiation is the main factor causing hyperpigmentation. MSC secretome contains bioactive soluble molecules such as cytokines and growth factors that can accelerate skin regeneration. However, the molecular role of the secretome in hyperpigmentation is still unclear. AIM: This study aimed to determine the effect of secretome hypoxia mesenchymal stem cells (S-HMSC) gel on the expression of superoxide dismutase (SOD) and matrix metalloproteinases (MMP-1) genes in skin tissue of hyperpigmented rats induced by UVB light exposure. MATERIALS AND METHODS: Experimental research with post-test only control group. The control, base gel, T1 and T2 groups were UVB irradiated 6 times in 14 days at 302 nm with an minimal erythema dose of 390 mJ/cm2, respectively, while sham group did not receive UVB exposure. T1 was given 100 uL of S-HMSC gel and T2 was given 200 uL of S-HMSC gel every day for 14 days, while base gel received base gel. On day 15, skin tissue was isolated and analyzed for SOD and MMP-1 expression using qRT-PCR. RESULTS: The relative expression of the SOD gene in the treatment group (P1 = 0.47 ± 0.20, P2 = 1.22 ± 0.47) increased with increasing dose compared to the control group (UVB = 0.05 ± 0.01, Base gel = 0.05 ± 0.02). The relative expression of the MMP-1 gene in the treatment group (T1 = 5.82 ± 1.16, T2 = 2.86 ± 1.57) decreased with increasing dose compared to the control group (Control = 10.10 ± 2.31, and Base gel = 9.55 ± 1.29). CONCLUSION: Administration of S-HMSC gel can increase SOD gene expression and decrease MMP-1 gene expression in skin tissue of hyperpigmented rats model induced by UVB light.

Amentoflavone-Enriched Selaginella rossii Protects against Ultraviolet- and Oxidative Stress-Induced Aging in Skin Cells.

Selaginellaceae plants are used in cosmetics to limit skin aging. This study is the first to investigate the anti-aging effects of Selaginella rossii (SR) on ultraviolet B (UVB)- and oxidative stress-induced skin cells. The 95% ethanol extract of Selaginella rossii (SR95E) contained much higher amounts of amentoflavone (AMF), an active compound, than other Selaginellaceae plants and was more effective in inhibiting matrix metalloproteinase (MMP)-1 expression in CCD-986sk fibroblasts. SR95E significantly decreased UVB-induced MMP-1, MMP-2, MMP-3 and MMP-9 expression and enhanced procollagen type I C-peptide content and mRNA expression of collagen type I alpha (COL1A)1 and COL1A2 in CCD-986sk fibroblasts. In HaCaT keratinocytes, SR95E treatment also dose-dependently decreased UVB-induced MMP-1 concentration and MMP-1, MMP-2, MMP-3 and MMP-9 mRNA expression. Moreover, SR95E treatment markedly inhibited UVB-induced c-Jun N-terminal kinase and p38 mitogen-activated protein kinase signaling and nuclear factor kappa-B signaling in HaCaT cells. Furthermore, SR95E and AMF markedly regulated the 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH)-induced expression of cellular senescence-related markers, including p16, p21 and LMNB1, in HaCaT cells. Overall, this study indicates that SR may have potential as a functional material on preventing UVB- and AAPH-induced skin aging and wrinkles.

Gene Expression Linked to Reepithelialization of Human Skin Wounds.

Our understanding of the regulatory processes of reepithelialization during wound healing is incomplete. In an attempt to map the genes involved in epidermal regeneration and differentiation, we measured gene expression in formalin-fixed, paraffin-embedded standardized epidermal wounds induced by the suction-blister technique with associated nonwounded skin using NanoString technology. The transcripts of 139 selected genes involved in clotting, immune response to tissue injury, signaling pathways, cell adhesion and proliferation, extracellular matrix remodeling, zinc transport and keratinocyte differentiation were evaluated. We identified 22 upregulated differentially expressed genes (DEGs) in descending order of fold change (MMP1, MMP3, IL6, CXCL8, SERPINE1, IL1B, PTGS2, HBEGF, CXCL5, CXCL2, TIMP1, CYR61, CXCL1, MMP12, MMP9, HGF, CTGF, ITGB3, MT2A, FGF7, COL4A1 and PLAUR). The expression of the most upregulated gene, MMP1, correlated strongly with MMP3 followed by IL6 and IL1B. rhIL-1β, but not rhIL-6, exposure of cultured normal human epidermal keratinocytes and normal human dermal fibroblasts increased both MMP1 mRNA and MMP-1 protein levels, as well as TIMP1 mRNA levels. The increased TIMP1 in wounds was validated by immunohistochemistry. The six downregulated DEGs (COL7A1, MMP28, SLC39A2, FLG1, KRT10 and FLG2) were associated with epidermal maturation. KLK8 showed the strongest correlation with MKI67 mRNA levels and is a potential biomarker for keratinocyte proliferation. The observed gene expression changes correlate well with the current knowledge of physiological reepithelialization. Thus, the gene expression panel described in this paper could be used in patients with impaired healing to identify possible therapeutic targets.

Treponema pallidum protein Tp0136 promoting MMPs/TIMPs imbalance via PI3K, MAPK and NF-κB signalling pathways in HDVSMCs

The invasive capability of Treponema. pallidum is central to its infection process. Matrix metalloproteinases (MMPs), which are specifically inhibited by the tissue inhibitors of metalloproteinases (TIMPs), play a pivotal role in promoting pathogenic invasion by destroying tissue barriers within the body. This study aimed to explore the effect of T. pallidum protein Tp0136 on the balance of MMPs/TIMPs in human dermal vascular smooth muscle cells (HDVSMCs) and the related underlying mechanisms. A number of in vitro studies were conducted to access the impact of recombinant Tp0136 protein on the balance of MMPs/TIMPs in HDVSMCs. The involvement of the PI3K, MAPK, and NF-κB signaling pathways in this process was also investigated. Tp0136 induced the mRNA and protein expressions of MMP1 in HDVSMCs in a concentration-dependent way. In addition, MMP1/TIMP1 and MMP1/TIMP2 ratios were also increased. Furthermore, the study demonstrated that treatment of HDVSMCs with Tp0136 activated the PI3K, MAPK, and NF-κB signaling pathways. Inhibition of PI3K, JNK, P38, and NF-κB, suppressed MMP1 expression and reduced the induction of MMP1/TIMP1 and MMP1/TIMP2 ratios by Tp0136. These findings demonstrate that Tp0136 enhanced the expression of MMP1 involving the PI3K, MAPK, and NF-κB signaling pathways in HDVSMCs, and thus generated the unbalance of MMPs/TIMP, which could contribute to the early spread of T. pallidum and pathogenesis of syphilis.

Hypoxia inducible factor-1α mediates the mechanism of the Hedgehog pathway in tendinopathy repair by Asperosaponin VI.

Our previous study found that asperosaponin VI (ASA VI) has a positive effect on the repair of tendinopathy. However, its molecular biological mechanism is unclear. To investigate the role of hypoxia inducible factor-1α (HIF-1α) in mediating the hedgehog (Hh) pathway in tendinopathy repair by ASA VI. A total of 36 2-month-old female SD rats were classified into the normal group (NG, n=10) and tendinopathy model group (n=26). The tendinopathy model group was further divided into the model group (MG), ASA VI group (AG), and triamcinolone acetonide+lidocaine group (TG). Compared with those in the MG group, IL-1 mRNA was significantly downregulated and IL-4 and IL-10 were increased in the AG group (P<0.01). The mRNA expression levels of MMP3, TIMP3, VEGF-A, KDR, and VWF mRNA decreased (P<0.01). Immunofluorescence staining revealed that CD31/endomucin levels were significantly attenuated. Scx, Mkx, EYA1, EYA2, COL1, COL3, and TNC mRNA levels showed significant differences (P<0.01). Immunofluorescence staining suggested the upregulation of Scx and the downregulation of Sox9. Shh, Ptch1, Smo, Gli1, Cyc-D1, Cyc-E1, and c-Myc mRNA levels were downregulated (P<0.01). The protein expression levels of Gli 1, Shh, and Ptch1 decreased significantly (P<0.01). The immunofluorescence staining levels of Shh, Ptch, and Gli 1 significantly decreased. ASA VI inhibits local vascular hyperproliferation and downregulates the HIF-1α/Hh pathway to promote the tendinous differentiation of tendon stem/progenitor cells and the repair of tendinopathy. The effect of ASA VI on HIF-1α levels may be an effective target in the treatment of tendinopathy.