LINC01116 Promotes Migration and Invasion of Oral Squamous Cell Carcinoma by Acting as a Competed Endogenous RNA in Regulation of MMP1 Expression.

Oral squamous cell carcinoma (OSCC) has increasingly become a worldwide health concern, and its survival rate has not been much improved partially due to a deficiency of precise molecular markers. Dysregulation of LINC01116, a long noncoding RNA sequence, has been observed in several types of cancer. However, the role played by LINC01116 in OSCC has not yet been fully elaborated. This study explored how LINC01116 was involved in the regulation of OSCC progression by analyzing expressions of LINC01116 in OSCC patients. The findings demonstrated upregulation of LINC01116 in OSCC tissues as opposed to regular oral mucosa, and overexpression of LINC01116 was correlated with advanced tumor status. LINC01116 knockdown using shRNA markedly reduced the OSCC cell invasion and migration in vitro. Moreover, the expression of LINC01116 was negatively correlated with that of microRNA-9-5p (miR-9). Luciferase reporter and loss-of-function assays demonstrated that LINC01116 functioned as a competing endogenous RNA (ceRNA) that could effectively sponge miR-9, thus regulating the derepression of matrix metalloproteinase 1 (MMP1). Furthermore, we confirmed that LINC01116 knockdown did not affect the expression of MMP1 messenger RNA (mRNA). Collectively, it is demonstrated in this study that overexpression of LINC01116 can promote the OSCC progression. The LINC01116-miR-9-MMP1 axis provides a novel insight into the OSCC pathogenesis and offers potential therapeutic targets against OSCC.

A Practical Approach for the In Vitro Safety and Efficacy Assessment of an Anti-Ageing Cosmetic Cream Enriched with Functional Compounds.

(1) Background: Cosmeceuticals are topical products applied to human skin to prevent skin ageing and maintain a healthy skin appearance. Their effectiveness is closely linked to the compounds present in a final formulation. In this article, we propose a panel of in vitro tests to support the efficacy assessment of an anti-ageing cream enriched with functional compounds. (2) Methods: biocompatibility and the irritant effect were evaluated on reconstructed human epidermis (RHE) and corneal epithelium (HCE) 3D models. After a preliminary MTT assay, normal human dermal fibroblasts (NHDF) and keratinocytes (HaCaT) were used to evaluate the extracellular matrix (ECM) protein synthesis, and interleukin-6 (IL-6) and metalloproteinase-1 (MMP-1) production. (3) Results: data collected showed good biocompatibility and demonstrated the absence of the irritant effect in both 3D models. Therefore, we demonstrated a statistical increase in collagen and elastin productions in NHDF cells. In HaCaT cells, we highlighted an anti-inflammatory effect through a reduction in IL-6 levels in inflammatory stimulated conditions. Moreover, the reduction of MMP-1 production after UV-B radiation was demonstrated, showing significant photo-protection. (4) Conclusion: a multiple in vitro assays approach is proposed for the valid and practical assessment of the anti-ageing protection, anti-inflammatory and biocompatible claims that can be assigned to a cosmetic product containing functional compounds.

EMMPRIN expression is associated with metastatic progression in osteosarcoma.

BACKGROUND: Extracellular matrix metalloproteinase inducer (EMMPRIN), a cell-surface glycoprotein, is overexpressed in several cancer types. EMMPRIN induces a metastatic phenotype by triggering the production of matrix metalloproteinase proteins (MMPs) such as MMP1 and MMP2, and vascular endothelial growth factor (VEGF) in cancer cells and the surrounding stromal cells. The purpose of this study was to investigate the expression and role of EMMPRIN in osteosarcoma. METHODS: The level of EMMPRIN expression was evaluated using reverse transcriptase polymerase chain reaction (RT-PCR) in 6 tumor-derived osteosarcoma cell lines and compared with that in normal osteoblasts. To study the prognostic significance of EMMPRIN expression, immunohistochemistry was carried out in prechemotherapy biopsies of 54 patients. siRNA knockdown of EMMPRIN in SaOS-2 cells was conducted to explore the role of EMMPRIN. To study the role of EMMPRIN in tumor-stromal interaction in MMP production and invasion, co-culture of SaOS-2 cells with osteoblasts and fibroblasts was performed. Osteosarcoma 143B cells were injected into the tail vein of BALB/c mice and lung metastasis was analyzed. RESULTS: EMMRIN mRNA expression was significantly higher in 5 of 6 (83%) tumor-derived cells than in MG63 cells. 90% of specimens (50/54) stained positive for EMMPRIN by immunohistochemistry, and higher expression of EMMPRIN was associated with shorter metastasis-free survival (p = 0.023). Co-culture of SaOS-2 with osteoblasts resulted in increased production of pro-MMP2 and VEGF expression, which was inhibited by EMMPRIN-targeting siRNA. siRNA knockdown of EMMPRIN resulted in decreased invasion. EMMPRIN shRNA-transfected 143B cells showed decreased lung metastasis in vivo. CONCLUSIONS: Our data suggest that EMMPRIN acts as a mediator of osteosarcoma metastasis by regulating MMP and VEGF production in cancer cells as well as stromal cells. EMMPRIN could serve as a therapeutic target in osteosarcoma.

Transcriptome analysis of human dermal fibroblasts following red light phototherapy.

Fibrosis occurs when collagen deposition and fibroblast proliferation replace healthy tissue. Red light (RL) may improve skin fibrosis via photobiomodulation, the process by which photosensitive chromophores in cells absorb visible or near-infrared light and undergo photophysical reactions. Our previous research demonstrated that high fluence RL reduces fibroblast proliferation, collagen deposition, and migration. Despite the identification of several cellular mechanisms underpinning RL phototherapy, little is known about the transcriptional changes that lead to anti-fibrotic cellular responses. Herein, RNA sequencing was performed on human dermal fibroblasts treated with RL phototherapy. Pathway enrichment and transcription factor analysis revealed regulation of extracellular matrices, proliferation, and cellular responses to oxygen-containing compounds following RL phototherapy. Specifically, RL phototherapy increased the expression of MMP1, which codes for matrix metalloproteinase-1 (MMP-1) and is responsible for remodeling extracellular collagen. Differential regulation of MMP1 was confirmed with RT-qPCR and ELISA. Additionally, RL upregulated PRSS35, which has not been previously associated with skin activity, but has known anti-fibrotic functions. Our results suggest that RL may benefit patients by altering fibrotic gene expression.

miR-202-3p negatively regulates MMP-1 to inhibit the proliferation, migration and invasion of lung adenocarcinoma cells.

Lung adenocarcinoma (LUAD) is one of the common cancers. Studies show that MMP-1 is involved in tumor progression, yet relevant regulatory mechanism in LUAD remains to be further elucidated. Here, we demonstrated from bioinformatics analysis for GEO data that MMP-1 was differentially up-regulated in LUAD. miR-202-3p, identified as the upstream regulator of MMP-1 by both bioinformatics and dual-luciferase assays, was differentially down-regulated in LUAD and presented a negative correlation with MMP-1. Following cell biological experiments proved that knocking down the expression of MMP-1 inhibited the proliferation, migration and invasion of LUAD cells, while overexpressed miR-202-3p posed a similar suppressive effect on cancer progression. Additionally, rescue assay further identified that overexpression of MMP-1 attenuated the suppressive effect of up-regulated miR-202-3p on malignant progression of LUAD cells. In all, this research suggests a mechanism by which MMP-1 under the regulation of miR-202-3p modulates the proliferation, migration and invasion of LUAD cells, which may contribute to the development of new therapeutic strategies.

Epigallocatechin-3-Gallate Suppresses the Expression of TNF-α-Induced MMP-1 via MAPK/ERK Signaling Pathways in Human Dermal Fibroblasts.

Deeper wrinkles and loss of elasticity are one of the skin-aging symptoms. Collagen breakdown by matrix metalloproteinase-1 (MMP-1), which is induced by reactive oxygen species (ROS) and pro-inflammatory cytokines, has been known to be responsible for these skin-aging symptoms. Therefore, much attention has been paid to chemicals to suppress the MMP-1 activity. Epigallocatechin-3-gallate (EGCG), catechin rich in green tea, has been reported to show antioxidant and protect skin from various stimuli such as UV and chemicals. In this study, we evaluated the inhibitory effect of EGCG on MMP-1 gene expression and secretion in tumor necrosis factor-α (TNF-α)-treated human dermal fibroblast cells (Hs68 cells). Pre-treatment with EGCG (10 and 20 µM) suppressed TNF-α-induced MMP-1 expression and secretion. EGCG also reduced the phosphorylation of extracellular signal regulated kinase (ERK) significantly but not that of p38 activation and c-Jun N-terminal kinase (JNK). Besides, EGCG (10 and 20 µM) showed the inhibitory effect on mitogen-activated protein extracellular kinase (MEK) and Src phosphorylation which is reported to be upstream signal proteins of ERK signal pathway. Based on these results, EGCG might have potential activity to slow down the skin-aging through inhibition of collagen breakdown, which remains to be elucidated.

Lysophosphatidic Acid Augments the Gene Expression and Production of Matrix Metalloproteinases-1 and -3 in Human Synovial Fibroblasts in Vitro.

Rheumatoid arthritis (RA) is an inflammatory disease with joint dysfunction following cartilage degradation. The level of lysophosphatidic acid (LPA) has been reported to be augmented in human synovial fluid from patients with RA. However, it remains to be elucidated whether LPA participates in cartilage destruction. In the present study, we have demonstrated that the production of promatrix metalloproteinases (proMMPs)-1 and -3 was augmented along with an increase of extracellular signal-regulated kinase (ERK)1/2 phosphorylation through LPA receptor 1 (LPAR1) in human synovial fibroblasts. These results suggest that LPA transcriptionally increases MMP production by the activation of an LPAR1/ERK1/2 signal pathway in human synovial fibroblasts. Thus, LPA is likely to be a pathological candidate for cartilage degradation in RA.

Spatial expression of metallothionein, matrix metalloproteinase-1 and Ki-67 in human epidermal wounds treated with zinc and determined by quantitative immunohistochemistry: A randomised double-blind trial.

Reepithelialisation is fundamental to wound healing, but our current understanding largely relies on cellular and animal studies. The aim of the present randomised double-blind three-arm controlled trial was to correlate genuine epidermal wound healing with key proteins and topical zinc treatment in humans. Sixty wounds were produced using deroofed suction blisters in 30 healthy volunteers and randomised to topical zinc sulphate (n = 20), placebo (n = 20), or control (n = 20) treatment for 4 days. All wounds with perilesional skin were processed for automatic immunostaining of paraffin tissue sections with monoclonal antibodies against Ki-67, metallothionein (MT) and matrix metalloproteinase (MMP)-1. Protein expression was quantified by automated digital image analysis. Epidermal Ki-67 and MT labelling indices were increased in keratinocytes in the neoepidermis (∼1.1 mm) and at the wound edge (0.5 mm) compared to normal skin. Increased MMP-1 immunostaining was restricted to the neoepidermis. MT was robustly upregulated in the upper dermis of the wounds. Zinc treatment enhanced MMP-1 expression beneath the neoepidermis via paracrine mechanisms and MT under the neoepidermis and in the nonepithelialised wound bed via direct actions of zinc as indicated by the induction of MT2A mRNA but not MMP-1 mRNA in cultured normal human dermal fibroblasts by zinc sulphate. The present human study demonstrates that quantitative immunohistochemistry can identify proteins involved in reepithelialisation and actions of external compounds. Increased dermal MT expression may contribute to the anti-inflammatory activities of zinc and increased MMP-1 levels to promote keratinocyte migration.

Filifactor alocis and Tumor Necrosis Factor-Alpha Stimulate Synthesis of Visfatin by Human Macrophages.

There is little known about the effect of the periodontopathogen Filifactor alocis on macrophages as key cells of the innate immune defense in the periodontium. Therefore, the aim of the present study was to investigate the effect of F. alocis and additionally of the pro-inflammatory cytokine tumor necrosis factor-alpha (TNFα) on visfatin and other pro-inflammatory and proteolytic molecules associated with periodontitis in human macrophages. The presence of macrophage markers CD14, CD86, CD68, and CD163 was examined in gingival biopsies from healthy individuals and periodontitis patients. Human macrophages were incubated with F. alocis and TNFα for up to 2 d. The effects of both stimulants on macrophages were determined by real-time PCR, ELISA, immunocytochemistry, and immunofluorescence. F. alocis was able to significantly stimulate the synthesis of visfatin by human macrophages using TLR2 and MAPK pathways. In addition to visfatin, F. alocis was also able to increase the synthesis of cyclooxygenase 2, TNFα, and matrix metalloproteinase 1. Like F. alocis, TNFα was also able to stimulate the production of these proinflammatory and proteolytic molecules. Our results highlight the pathogenetic role of F. alocis in periodontal diseases and also underline the involvement of visfatin in the aetiopathogenesis of periodontitis.

Prevention of intra-abdominal adhesions by a hyaluronic acid gel; an experimental study in rats.

BACKGROUND: In 80% to 90% of the patients intra-abdominal adhesions occur after abdominal surgery, which can cause small-bowel obstruction, chronic abdominal pain, female infertility and difficulty during reoperation. A novel crosslinked hyaluronic acid gel is evaluated regarding its anti-adhesive capacities in an ischemic button model in rats. METHOD: 51 adult, male Wistar rats from a registered breeder, received eight ischemic buttons each and were treated with hyaluronic acid gel (HA, HyaRegen©), hyaluronic acid carboxymethylcellulose (HA-CMC, Seprafilm©) or no anti-adhesive barrier. After 14 days, the animals were sacrificed and adhesions were scored macroscopically. The number of buttons and organs involved in adhesions were recorded. Per animal, one button with adhesions and one without adhesions was explanted for qPCR analysis. Mann-Whitney U, Fisher's exact and Wilcoxon signed rank test were used for data analysis. A p-value of 0.05 was considered significant. RESULTS: Macroscopic evaluation of adhesion formation did not differ between the groups. The number of organs involved in adhesions in the HA gel group was significantly lower compared to HA-CMC (p = .041) and the control group (p = .012). A significantly, 1.36-fold higher clec10a (p = 0.25), 1.80-fold higher cd163 (p = 0.003) and 5.14-fold higher mmp1 expression (p = 0.028) was found in ischemic buttons with adhesions compared to buttons without adhesions. CONCLUSION: HA gel application reduces the number of organs involved in adhesions in an ischemic button model, but no overall reduction in adhesion formation was encountered. Macrophage subtype 2 polarization and high mmp1 expression are associated with adhesion formation. Further investigation is needed in the exact pathophysiologic process of adhesion formation and the role of macrophage polarization.

The expression, purification, and substrate analysis of matrix metalloproteinases in Drosophila melanogaster.

Matrix metalloproteinases (MMPs) are evolutionarily conserved extracellular matrix proteinases. Genetic analysis of the Drosophila MMPs, Mmp1 and Mmp2, in vivo reveal that they play vital roles in tissue remodeling. Although the catalytic domain (CD) undertakes most MMP functions, few studies have sought to demonstrate the biochemical properties of the CDs of fly MMPs. Here, we identified the overexpression, purification, and refolding of the CDs of Drosophila Mmp1 and Mmp2 for biochemical studies. Zymography assays and substrate degradation analysis showed that both Mmp1-CD and Mmp2-CD were able to digest casein, gelatin, fibronectin, collagen (types I, IV, and V), while Mmp2-CD showed much higher degradation activity compared with Mmp1-CD. Moreover, human collagen III could be degraded by Mmp1-CD but not Mmp2-CD, and rat collagen I and laminin could be degraded by Mmp2-CD but not Mmp1-CD, suggesting that Drosophila Mmp1 and Mmp2 might have overlapping yet distinct substrate specificity. Using synthetic fluorescent substrates, we further demonstrated that the enzymatic activity of Mmp1-CD and Mmp2-CD could be inhibited by human tissue inhibitors of metalloproteinases (TIMPs). These results reveal the context of the cooperative yet distinct roles of Mmp1 and Mmp2 in tissue remodeling.

Matrix metalloproteinases MMP-1, MMP-2, and MMP-13 are overexpressed in primary nodular melanoma.

BACKGROUND: The spread and invasion of malignant melanoma cells involve degradation and reorganization of the extracellular matrix by the activation of several matrix metalloproteinases (MMPs). This study analyzed the expression of MMP-1, MMP-2, and MMP-13 proteins in primary nodular melanoma (NM) and dysplastic nevi (DN) as a significant risk factor for melanoma development. The secondary goal was to analyze the correlation of MMPs protein expression in NM with tumor invasion, BRAF V600 mutation status, and overall survival. METHODS: Immunohistochemistry for MMP-1, MMP-2, and MMP-13 was performed on nodular melanoma (n = 52) and dysplastic nevi (n = 28) on tissue microarray (TMA). BRAF V600 mutation analysis on NM samples was performed by the Sanger sequencing method. RESULTS: A high level of MMPs expression in NM samples (>30%) compared with DN (<8%) was statistically significant (P < 0.001). BRAF V600 mutations were detected in 15 of 39 (38.5%) NM samples. This study revealed an interesting finding that MMP-1 and MMP-13 protein expression in the BRAF V600 mutated melanomas were significantly lower than in the BRAF V600 wild type (P < 0.05). CONCLUSION: Cox analysis revealed that Clark categories, Breslow thickness, and MMP-1 high protein expression are predictive factors for shorter overall survival (P < 0.05).

Leonurine Hydrochloride Suppresses Inflammatory Responses and Ameliorates Cartilage Degradation in Osteoarthritis via NF-κB Signaling Pathway.

Leonurine hydrochloride (LH) has been reported to exhibit a number of biological properties such as suppression of inflammation. This study aimed to examine whether the progression of osteoarthritis (OA) could be delayed by the administration of LH in an OA model. Rat chondrocytes were treated with LH under the condition of TNF-α-induced inflammation. After that, real-time PCR and Western blotting were conducted to evaluate relative gene/protein expression levels. For the in vivo study, rats were randomly allocated to a control group (anterior cruciate ligament transection (ACLT) surgery, treatment with saline) and LH group (ACLT surgery, treatment with LH). Articular cartilage degeneration was assessed by histological evaluation. It was found that LH significantly suppressed the expression of MMP-1, MMP-3, MMP-13, IL-6, and ADAMTS-5 in cells via the NF-κB signaling pathway. In addition, it is revealed that intra-articular injection of LH significantly ameliorated cartilage degeneration in a rat OA model. Taken together, these results indicate that LH attenuates progression of OA by inhibition of inflammation via the NF-κB signaling pathway and represents a potential preventive therapy for OA.

Prostaglandin E2 Induces Skin Aging via E-Prostanoid 1 in Normal Human Dermal Fibroblasts.

Collagen type I production decreases with aging, leading to wrinkles and impaired skin function. Prostaglandin E2 (PGE2), a lipid-derived signaling molecule produced from arachidonic acid by cyclo-oxygenase, inhibits collagen production, and induces matrix metallopeptidase 1 (MMP1) expression by fibroblasts in vitro. PGE2-induced collagen expression inhibition and MMP1 promotion are aging mechanisms. This study investigated the role of E-prostanoid 1 (EP1) in PGE2 signaling in normal human dermal fibroblasts (NHDFs). When EP1 expression was inhibited by EP1 small interfering RNA (siRNA), there were no significant changes in messenger RNA (mRNA) levels of collagen, type I, alpha 1 (COL1A1)/MMP1 between siRNA-transfected NHDFs and siRNA-transfected NHDFs with PGE2. This result showed that EP1 is a PGE2 receptor. Extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation after PGE2 treatment significantly increased by ~2.5 times. In addition, PGE2 treatment increased the intracellular Ca2+ concentration in NHDFs. These results indicated that PGE2 is directly associated with EP1 pathway-regulated ERK1/2 and inositol trisphosphate (IP3) signaling in NHDFs.

Long-term studies on the integration of acellular porcine dermis as an implant shell and the effect on capsular fibrosis around silicone implants in a rat model.

Acellular dermal matrices have recently increasingly been used in alloplastic breast reconstruction with silicone breast implants. Among these matrices, acellular porcine dermis (APD) is frequently applied, but long-term data on tissue integration and capsular fibrosis formation are still missing. Silicone prostheses with (group A) and without (group B) APD as an implant-covering shell were implanted in male Lewis rats. At 3, 12, and 52 weeks after implantation, the constructs were explanted. Molecular biological and immunohistochemical analyses were performed afterwards. On comparing the collagenous layer and the newly formed myofibroblast-rich layer around the implants of both groups, it became apparent that in group A, these layers were thinner, followed by a lower expression of TGFß1 after 12 and 52 weeks. Further, in this group, at the endpoint of 52 weeks, a lower amount of CD68-positive cells in the collagenous and myofibroblast-rich layers were observed and the expression of TNFα was reduced, while the number of Ki67-positive cells was significantly higher with time. Furthermore, MMP1 expression in group A was lower than that in group B, and the calculated ratio of MMP1:TIMP1 expression was higher. The long-term results clearly show a reduction in inflammatory and fibrotic tissue reaction when APD is used to cover silicone prostheses. These experimental data will be of considerable importance for implant-based breast surgery, as they indicate a potential benefit in the reduction of capsular fibrosis formation of an interposition of APD between the recipient and the silicone implant.

Vitexin alleviates interleukin-1ß-induced inflammatory responses in chondrocytes from osteoarthritis patients: Involvement of HIF-1α pathway.

It has been reported that vitexin has anti-inflammatory effects in osteoarthritis (OA) rats. However, the effects of vitexin on interleukins-1ß (IL-1ß)-stimulated OA patient-derived chondrocytes have not been reported. The purpose of this study was to investigate the anti-inflammatory effects of vitexin on IL-1ß-stimulated human osteoarthritis chondrocytes and to reveal the involvement of hypoxia-inducible factor 1α (HIF-1α) pathway. Enzyme-linked immunosorbent assay, quantitative real-time PCR and Western blotting assays were employed. ELISA results demonstrated that the proinflammatory cytokine levels of interleukins-6 (IL-6) and tumour necrosis factor α (TNF-α) in the serum and synovial fluid and HIF-1α level in the synovial fluid were significantly elevated in OA patients compared to normal healthy subjects. Moreover, the Western blotting results indicated that the protein expression of HIF-1α was significantly higher in the cartilage tissues of OA patients. OA patient-derived chondrocytes were stimulated by IL-1ß and treated with different concentration of vitexin for 24 hours. Vitexin showed no cytotoxicity and increased the survival of chondrocytes under IL-1ß stimulation. Vitexin suppressed IL-1ß-induced production of NO and prostaglandin E2 (PGE2 ) in chondrocytes culture. The treatment of vitexin significantly inhibited IL-1ß-induced expressions of proinflammatory cytokine levels of IL-6, TNF-α, matrix metalloproteinase (MMP)-1, MMP-3 and MMP-13. Furthermore, Western blotting results demonstrated that HIF-1α is involved in vitexin's protective effects on IL-1ß-stimulated injuries in OA patient-derived chondrocytes. Our study demonstrates that vitexin alleviates IL-1ß-induced inflammatory responses in chondrocytes from osteoarthritis patients, which may be attributed partly to the inhibition of HIF-1α pathway.

Sudachitin Inhibits Matrix Metalloproteinase-1 and -3 Production in Tumor Necrosis Factor-α-Stimulated Human Periodontal Ligament Cells.

Sudachitin, a polymethoxylated flavonoid found in the skin of Citrus sudachi, is a biologically active substance. The aim of this study was to examine whether sudachitin could be used to inhibit the expression of matrix metalloproteinase (MMP)-1 and MMP-3, which are involved in the destruction of periodontal tissues in periodontal lesions, in tumor necrosis factor (TNF)-α-stimulated human periodontal ligament cells (HPDLC). Sudachitin suppressed TNF-α-induced MMP-1 and MMP-3 production in HPDLC. On the other hand, it enhanced tissue inhibitor of metalloproteinase (TIMP)-1 expression. The level of Akt phosphorylation in the TNF-α-stimulated HPDLC was decreased by sudachitin treatment. Moreover, an Akt inhibitor reduced MMP-1 and MMP-3 production and increased TIMP-1 production. These findings indicate that sudachitin reduces MMP-1 and MMP-3 production in TNF-α-stimulated HPDLC by inhibiting the Akt pathway.

Silencing Y-box binding protein-1 inhibits triple-negative breast cancer cell invasiveness via regulation of MMP1 and beta-catenin expression.

Y-box binding protein-1 (YB-1), an important transcription and translation regulator protein, is known to increase cancer cell invasiveness and spreading. Here, we report its role in breast cancer, particularly in mediating cell invasion in triple-negative breast cancer (TNBC). YB-1 stable knockdown (shYB-1) significantly reduced the invasive potential of MDA-MB-231 TNBC cells in 2D and 3D (spheroid) cultures. Whole proteome mass spectrometry analysis showed an enrichment of cell adhesion and cell to matrix interaction proteins, notably, matrix metalloproteinase-1 (MMP1) and beta-catenin (CTNNB1), which are known to play critical roles in cancer metastasis. shYB-1 cells exhibited substantial downregulation of MMP1 and CTNNB1 mRNA and protein expression, with reduced MMP1 enzyme activity. YB-1 was also observed to bind to the promoter of MMP1 and overexpression of MMP1 plasmid in shYB-1 cells increased cell invasion. Finally, analysis of tumour samples from the Gene Expression-Based Outcome for Breast Cancer Online (GOBO) database revealed that high gene expressions of YBX1, MMP1 and CTNNB1 predict for a significantly lower 10-year distant metastasis free survival. Altogether, this study shows that YB-1 mediates breast cancer invasion and metastasis via regulation of MMP1 and beta-catenin.

Fibrogenic Gene Expression in Hepatic Stellate Cells Induced by HCV and HIV Replication in a Three Cell Co-Culture Model System.

Retrospective studies indicate that co-infection of hepatitis C virus (HCV) and human immunodeficiency virus (HIV) accelerates hepatic fibrosis progression. We have developed a co-culture system (MLH) comprising primary macrophages, hepatic stellate cells (HSC, LX-2), and hepatocytes (Huh-7), permissive for active replication of HCV and HIV, and assessed the effect of these viral infections on the phenotypic changes and fibrogenic gene expression in LX-2 cells. We detected distinct morphological changes in LX-2 cells within 24 hr post-infection with HCV, HIV or HCV/HIV in MLH co-cultures, with migration enhancement phenotypes. Human fibrosis microarrays conducted using LX-2 cell RNA derived from MLH co-culture conditions, with or without HCV and HIV infection, revealed novel insights regarding the roles of these viral infections on fibrogenic gene expression in LX-2 cells. We found that HIV mono-infection in MLH co-culture had no impact on fibrogenic gene expression in LX-2 cells. HCV infection of MLH co-culture resulted in upregulation (>1.9x) of five fibrogenic genes including CCL2, IL1A, IL1B, IL13RA2 and MMP1. These genes were upregulated by HCV/HIV co-infection but in a greater magnitude. Conclusion: Our results indicate that HIV-infected macrophages accelerate hepatic fibrosis during HCV/HIV co-infection by amplifying the expression of HCV-dependent fibrogenic genes in HSC.

Molecular expression patterns in the synovium and their association with advanced symptomatic knee osteoarthritis.

OBJECTIVE: Osteoarthritis (OA) is a major source of knee pain. Mechanisms of OA knee pain are incompletely understood but include synovial pathology. We aimed to identify molecular expression patterns in the synovium associated with symptomatic knee OA. DESIGN: Snap frozen synovia were from people undergoing total knee replacement (TKR) for advanced OA, or from post-mortem (PM) cases who had not sought help for knee pain. Associations with OA symptoms were determined using discovery and validation samples, each comprising TKR and post mortem (PM) cases matched for chondropathy (Symptomatic or Asymptomatic Chondropathy). Associations with OA were determined by comparing age matched TKR and PM control cases. Real-time quantitative PCR for 96 genes involved in inflammation and nerve sensitisation used TaqMan® Array Cards in discovery and validation samples, and protein expression for replicated genes was quantified using Luminex bead assay. RESULTS: Eight genes were differentially expressed between asymptomatic and symptomatic chondropathy cases and replicated between discovery and validation samples (P<0.05 or >3-fold change). Of these, matrix metalloprotease (MMP)-1 was also increased whereas interleukin-1 receptor 1 (IL1R1) and vascular endothelial growth factor (VEGF) were decreased at the protein level in the synovium of symptomatic compared to asymptomatic chondropathy cases. MMP1 protein expression was also increased in OA compared to PM controls. CONCLUSION: Associations of symptomatic OA may suggest roles of MMP1 expression and IL1R1 and VEGF pathways in OA pain. Better understanding of which inflammation-associated molecules mediate OA pain should inform refinement of existing therapies and development of new treatments.