Expression Of Urokinase-type Plasminogen Activator Research Articles

MicroRNA-181b attenuates lipopolysaccharide-induced inflammatory responses in pulpitis via the PLAU/AKT/NF-κB axis

ObjectiveThis study aimed to investigate the role and underlying mechanisms of microRNA (miRNA)-181b in the inflammatory response in pulpitis. MethodsQuantitative reverse-transcription polymerase chain reaction (qRT-PCR), fluorescence in situ hybridization (FISH), and immunofluorescence techniques were used to determine the miRNA-181b and urokinase-type plasminogen activator (PLAU) expression levels in inflamed human dental pulp tissues (HDPTs) and lipopolysaccharide (LPS)-stimulated human dental pulp cells (hDPCs). The targets of miRNA-181b were identified and confirmed using a bioinformatics analysis, RNA sequencing, and dual-luciferase gene reporter assays. The effect of miRNA-181b or PLAU on proinflammatory cytokine expression in hDPCs was examined using qRT-PCR and western blotting. RNA sequencing was conducted to examine the signaling pathways implicated in miRNA-181b-mediated pulpitis. Western blotting and qRT-PCR were used to determine the miRNA-181b /PLAU/AKT/NF-κB signaling axis in pulpitis. A rat pulpitis model was created to observe the histopathological changes in the dental pulp tissue after the topical application of miRNA-181b agomir. ResultsA significant decrease in miRNA-181b and an increase in PLAU were observed in HDPTs compared to the healthy controls, and these two factors showed a negative correlation. MiRNA-181b directly targeted PLAU. The miRNA-181b inhibitor resulted in a significant upregulation of IL-1β, IL-6 and TNF-α, whereas the knockdown of PLAU reversed this proinflammatory effect. Conversely, PLAU overexpression prevented the anti-inflammatory effects of the miRNA-181b mimics. Mechanistically, miRNA-181b inhibited the AKT/NF-κB pathway by targeting PLAU. In vivo application of the miRNA-181b agomir to inflamed pulp tissue alleviated inflammation. ConclusionMiRNA-181b targets PLAU, negatively regulating pro-inflammatory cytokine expression via the AKT/NF-κB signaling pathway.

Reduction of invasion and cell stemness and induction of apoptotic cell death by Cinnamomum cassia extracts on human osteosarcoma cells.

Cinnamomum cassia possesses antioxidative activity and induces the apoptotic properties of various cancer types. However, its effect on osteosarcoma invasion and cancer stemness remains ambiguous. Here, we examined the molecular evidence of the anti-invasive effects of ethanoic C. cassia extracts (CCE). Invasion and migration were obviously suppressed after the expression of urokinase-type plasminogen activator and matrix metalloprotein 2 in human osteosarcoma 143B cells were downregulated. CCE reversed epithelial-to-mesenchymal transition (EMT) induced by transforming growth factor β1 and downregulated mesenchymal markers, such as snail-1 and RhoA. CCE suppressed self-renewal property and the expression of stemness genes (aldehyde dehydrogenase, Nanog, and CD44) in the 143B cells. CCE suppressed cell viability, reduced the colony formation of osteosarcoma cancer cells, and induced apoptotic cell death in the 143B cells, as indicated by caspase-9 activation. The xenograft tumor model of immunodeficient BALB/c nude mice showed that CCE administered in vivo through oral gavage inhibited the growth of implanted 143B cells. These findings indicated that CCE inhibited the invasion, migration, and cancer stemness of the 143B cells. CCE reduced proliferation of 143B cell possibly because of the activation of caspase-9 and the consequent apoptosis, suggesting that CCE is a potential anticancer supplement for osteosarcoma.

A curcumin analog CA-5f inhibits urokinase-type plasminogen activator and invasive phenotype of triple-negative breast cancer cells.

Triple-negative breast cancer (TNBC) is one of the most aggressive types of breast cancer with poor outcomes. Patients with TNBC cannot benefit from targeted therapies such as Tamoxifen and Herceptin. The aim of the present study was to seek a preventive or therapeutic agent with a potential inhibitory effect on aggressive progression of TNBC. Anticancer effect of a natural compound curcumin have been demonstrated, however, development of more effective curcumin analogs with better bioavailability is needed. We investigated if a curcumin analog CA-5f could inhibit the invasive phenotype of TNBC cell lines in the present study. Treatment with CA-5f inhibited the viability of MDA‑MB‑231 and Hs578T TNBC cells, possible by inducing apoptosis. The invasive phenotypes of these cells were inhibited by CA-5f in a concentration-dependent manner. Protein expression of urokinase-type plasminogen activator (uPA), a serine protease known to degrade the extracellular matrix and lead to invasion, was markedly decreased by CA-5f in Hs578T cells. However, mRNA level of uPA was not altered by CA-5f, implicating that the effect of CA-5f was not through transcriptional regulation. Of note, CA-5f upregulated plasminogen activator inhibitor type (PAI)-1, which is known to inhibit uPA by interacting with urokinase-type plasminogen receptor, in TNBC cells. Taken together, these results demonstrated that CA-5f significantly inhibited the invasive phenotype of TNBC cells, possibly by decreasing the protein level of uPA through upregulating PAI-1. Our results may provide useful information on developing CA-5f as a potential therapeutic agent against malignant progression of TNBC.

Synthetic Fibrin-Derived Bβ15-42 Peptide Delays Thrombus Resolution in a Mouse Model.

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Gossypol Reduces Metastasis and Epithelial-Mesenchymal Transition by Targeting Protease in Human Cervical Cancer.

Metastasis is the most prevalent cause of cancer-associated deaths amongst patients with cervical cancer. Epithelial-mesenchymal transition (EMT) is essential for carcinogenesis, and it confers metastatic properties to cancer cells. Gossypol is a natural polyphenolic compound with anti-inflammation, anti-oxidant, and anticancer activities. In this study, we investigated the antimetastatic and antitumour effects of gossypol on human cervical cancer cells (HeLa and SiHa cells). Gossypol exerted a strong inhibition effect on the migration and invasion of human cervical cancer cells. It reduced the focal adhesion kinase (FAK) pathway-mediated expression of matrix metalloproteinase-2 and urokinase-type plasminogen activator, subsequently inhibiting the invasion of SiHa cells. In addition, gossypol reversed EMT induced by transforming growth factor beta 1 (TGF-[Formula: see text]1) and up-regulated epithelial markers, such as E-cadherin but significantly suppressed Ras homolog family member (Rho)A, RhoB, and p-Samd3. The tail vein injection model showed that gossypol treatment via oral gavage reduced lung metastasis. Gossypol also decreased tumour growth in vivo in the nude mouse xenograft model. All these findings suggest that gossypol suppressed the invasion and migration of human cervical cancer cells by targeting the FAK signaling pathway and reversing TGF-[Formula: see text]1-induced EMT. Hence, gossypol warrants further attention for basic mechanistic studies and drug development.

Anthocyanins: Promising Natural Compounds for Prevention and Treatment of Oral Squamous Cell Carcinoma

Background: Oral squamous cell carcinoma (OSCC) is the most common oral cancer. In recent decades, numerous studies have focused on the effects of new herbal and natural bioactive substances or pharmaceutical products on different cancer types. Some investigations have shown that anthocyanins can inhibit carcinogenesis in various ways, and some others focused on the effects of anthocyanins on squamous cell carcinoma (SCC). Methods: An electronic search of the literature was carried out in PubMed, Web of Science, and Scopus databases using the keywords “anthocyanins” and “oral cancer” or “oral squamous cell carcinoma”. Articles relevant to our topic (except for review articles) were included in this study. Results: The reviewed articles showed the inhibitory and suppressive effects of anthocyanins on OSCC, which were exerted by inhibiting NF-κB, p-Akt/PI3k, and JAK/STAT3 pathways, the induction of caspase-1 and caspase-3 overexpression, pyroptosis, mitochondrial-mediated apoptosis, and reduction of expression and production of matrix metalloproteinases and urokinase-type plasminogen activator. Conclusions: Based on the results, the anti-proliferative and apoptotic effects of anthocyanins on OSCC were dominant. Their antimetastatic effect was also reported. Therefore, anthocyanins seem to be promising natural compounds for the prevention and treatment of OSCC. In vivo studies and clinical trials are recommended to further confirm these results.

Ephrin-B1 Is a Novel Biomarker of Bladder Cancer Aggressiveness. Studies in Murine Models and in Human Samples.

Bladder cancer (BC) is the ninth most common cancer worldwide, but molecular changes are still under study. During tumor progression, Epithelial cadherin (E-cadherin) expression is altered and β-catenin may be translocated to the nucleus, where it acts as co-transcription factor of tumor invasion associated genes. This investigation further characterizes E-cadherin and β-catenin associated changes in BC, by combining bioinformatics, an experimental murine cell model (MB49/MB49-I) and human BC samples. In in silico studies, a DisGeNET (gene-disease associations database) analysis identified CDH1 (E-cadherin gene) as one with highest score among 130 BC related-genes. COSMIC mutation analysis revealed CDH1 low mutations rates. Compared to MB49 control BC cells, MB49-I invasive cells showed decreased E-cadherin expression, E- to P-cadherin switch, higher β-catenin nuclear signal and lower cytoplasmic p-Ser33-β-catenin signal, higher Ephrin-B1 ligand and EphB2 receptor expression, higher Phospho-Stat3 and Urokinase-type Plasminogen Activator (UPA), and UPA receptor expression. MB49-I cells transfected with Ephrin-B1 siRNA showed lower migratory and invasive capacity than control cells (scramble siRNA). By immunohistochemistry, orthotopic MB49-I tumors had lower E-cadherin, increased nuclear β-catenin, lower pSer33-β-catenin cytoplasmic signal, and higher Ephrin-B1 expression than MB49 tumors. Similar changes were found in human BC tumors, and 83% of infiltrating tumors depicted a high Ephrin-B1 stain. An association between higher Ephrin-B1 expression and higher stage and tumor grade was found. No association was found between abnormal E-cadherin signal, Ephrin-B1 expression or clinical-pathological parameter. This study thoroughly analyzed E-cadherin and associated changes in BC, and reports Ephrin-B1 as a new marker of tumor aggressiveness.

Matrix protease production, epithelial-to-mesenchymal transition marker expression and invasion of glioblastoma cells in response to osmotic or hydrostatic pressure

Both hydrostatic and osmotic pressures are altered in the tumour microenvironment. Glioblastoma (GBM) is a brain tumour with high invasiveness and poor prognosis. We hypothesized that physical and osmotic forces regulate glioblastoma (GBM) invasiveness. The osmotic pressure of GBM cell culture medium was adjusted using sodium chloride or water. Alternatively, cells were subjected to increased hydrostatic force. The proteolytic profile and epithelial–mesenchymal transition (EMT) were investigated using zymography and real-time qPCR. The EMT markers assessed were Snail-1, Snail-2, N-cadherin, Twist and vimentin. Invasion was investigated in vitro using extracellular matrix-coated Transwell inserts. In response to osmotic and mechanical pressure, GBM cell lines U87 and U251 and patient-derived neural oncospheres upregulated the expression of urokinase-type plasminogen activator (uPA) and/or matrix metalloproteinases (MMPs) as well as some of the EMT markers tested. The adherent cell lines invaded more when placed in media of increased osmolality. Therefore, GBM respond to osmotic or mechanical pressure by increasing matrix degrading enzyme production, and adopting a phenotype reminiscent of EMT. Better understanding the molecular and cellular mechanisms by which increased pressure promotes GBM invasiveness may help to develop innovative therapeutic approaches.

Fibrinolytic Activity of Circulating Microvesicles Is Associated with Progression of Breast Cancer.

The fibrinolytic system plays an important role in breast cancer, favoring progression through extracellular-matrix degradation, angiogenesis, apoptosis and cellular proliferation. The expression of urokinase-type plasminogen activator (uPA) in breast cancer tissue is widely recognized as an unfavorable prognostic factor. However, fibrinolytic activity associated with uPA cannot be reliably measured in the blood because of the rapid inhibition of uPA by plasminogen activator inhibitor-1 (PAI-1). By contrast, circulating microvesicles (Mvs) in peripheral blood protect bound enzymes from inhibition. Mvs are extracellular vesicles, released from various types of cells, and their size fluctuates between 100 and 1,000 nm. Mvs carry DNA, RNA, miRNA, and proteins, thereby serving as a source of horizontal communication between cells. We investigated whether fibrinolytic activity on circulating Mvs reflects breast cancer progression. The study population consisted of 13 patients with breast cancer and 13 healthy women. The cancer patients included 4 patients in remission, 3 patients with locally advanced cancer, and 6 with metastatic disease. Mvs were isolated from peripheral blood, quantified by a protein concentration method, and their fibrinolytic potential was measured by their capacity to generate plasmin. Although the quantity of Mvs found in patients with cancer and healthy individuals was similar, plasmin generated on Mvs was twice the amount in patients with metastasis than in healthy women (P < 0.05), underlying the value of this distinctive parameter. The data suggest that in breast cancer patients, higher fibrinolytic activity of circulating Mvs could be related to progression and metastasis of breast cancer.

Tazarotene-Induced Gene 1 (TIG1) Interacts with Serine Protease Inhibitor Kazal-Type 2 (SPINK2) to Inhibit Cellular Invasion of Testicular Carcinoma Cells.

Tazarotene-induced gene 1 (TIG1) encodes a protein that is a retinoid-regulated tumor suppressor. TIG1 is expressed in most normal tissues, and downregulation of TIG1 expression in multiple cancers is caused by promoter hypermethylation. Kazal-type serine protease inhibitor-2 (SPINK2) is a serine protease inhibitor, and the SPINK protein family has been shown to inhibit the expression of urokinase-type plasminogen activator (uPA). In addition, increased levels of uPA and the uPA receptor were observed in testicular cancer tissues. This study demonstrated that TIG1 interacts with SPINK2 in NT2/D1 testicular carcinoma cells. TIG1 and SPINK2 were highly expressed in normal testis tissues, while low expression levels of TIG1 and SPINK2 were found in testicular cancer tissues. TIG1 inhibited cell invasion, migration, and epithelial–mesenchymal transition (EMT) of NT2/D1 cells. SPINK2 enhanced TIG1-regulated uPA activity and EMT suppression, while silencing SPINK2 alleviated TIG1-mediated EMT regulation, cell migration, and invasion. Therefore, the results suggest that the interaction between TIG1 and SPINK2 plays an important role in the inhibition of testicular cancer cell EMT, and suppression is mediated through downregulation of the uPA/uPAR signaling pathway.

Inhibition by Epigallocatechin Gallate of IL-1-Induced Urokinase-Type Plasminogen Activator Expression and Collagen Degradation by Corneal Fibroblasts.

The proinflammatory cytokine interleukin (IL)-1 is implicated in corneal ulceration and promotes collagen degradation by corneal fibroblasts cultured in a three-dimensional (3D) collagen gel. Epigallocatechin-3-gallate (EGCG), the principal polyphenol in extracts of green tea, has various beneficial health effects, some of which appear to be mediated through direct or indirect inhibition of protease activity. We therefore examined the effect of EGCG on IL-1β-induced collagen degradation by corneal fibroblasts embedded in a collagen gel. Human corneal fibroblasts were cultured in a type I collagen gel. Collagen degradation was assessed by measurement of hydroxyproline in acid hydrolysates of culture supernatants. The expression of urokinase-type plasminogen activator (uPA) was examined by real-time and RT-PCR analysis and by fibrin zymography, and that of the collagenase matrix metalloproteinase 1 (MMP1) was detected by immunoblot analysis. EGCG inhibited IL-1β-induced, plasminogen-dependent collagen degradation by corneal fibroblasts in a concentration-dependent manner. It also attenuated the IL-1β-induced expression of uPA at both mRNA and protein levels. EGCG inhibited the IL-1β-induced conversion of exogenous plasminogen to plasmin as well as the plasminogen-dependent activation of pro-MMP1 in the 3D cultures without a substantial effect on pro-MMP1 abundance. EGCG inhibits IL-1β-induced collagen degradation by corneal fibroblasts, with this effect likely being mediated by suppression of the upregulation of uPA, the uPA-mediated conversion of plasminogen to plasmin, and the plasmin-mediated activation of pro-MMP1. EGCG thus warrants further investigation as a potential treatment for corneal ulcer.

β-catenin nuclear translocation represses thyroid cancer stem cells differentiating into cells with sodium-iodine symporter functional expression

Objective: To confirm whether β-catenin nuclear translocation in thyroid cancer stem cells can differentiate into thyroid cancer cells without functional membrane expression of sodium-iodine transporter (NIS) and be resistant to iodide 131 treatment. Methods: Thyroid cancer stem cells were firstly isolated as a side population (SP) from human thyroid cancer cell line FTC133. The SP cells from FTC133 were transfected with β-catenin, and then differentiated. The cells were further collected for Western blot, Transwell and MTT assay to investigate the epithelial-mesenchymal transition (EMT) characteristics, tumor growth, invasion, and iodine uptake potency in vitro. Functional NIS expression and iodide uptake in differentiated cells were detected with immunofluorescent staining and iodide uptake assay, respectively. Subcutaneous severe combined immunodeficient (SCID) mice tumor model was induced with differentiated cancer cells to explore the in vivo effect of radioiodine treatment. Further immunohistochemical staining was performed to reveal the changes of functional proteins involved in tumor radioiodine treatment. Results: Side population was isolated from FTC133 accounting for about 0.03%, with high expression of stem cell markers and decreased expression of differentiated cell markers. Western blot showed prominent EMT phenotype in the differentiated cells from β-catenin transfected stem cell model, with absence of epithelial expression of E-cadherin and cytokeratin 18, as well as abnormal expression of vimentin,fibronectin and urokinase-type plasminogen activator. Moreover,compared with cells differentiated from untransfected or empty plasmid transfected stem cells, in vitro proliferation markedly increased 85.4% and 81.0%, respectively (both P<0.01); while in vitro invasion augmented 78.8% and 84.4%, respectively (both P<0.01). Immunofluorescent staining identified that, after transfected with β-catenin, differentiated cells underwent β-catenin nuclear translocation and NIS localization transferred from membrane to plasma, compared with cells from untransfected or empty plasmid transfected stem cells. Cell iodide uptake in vitro decreased about 52.8% and 45.2%, respectively (both P<0.01). Furthermore, in vivo experiment further demonstrated that, cells differentiated from β-catenin transfected stem cells were found with much higher tumor proliferation,tumor growth rate and larger tumor mass after radioiodine 131 treatment (both P<0.05). Conclusion: Induction of β-catenin nuclear translocation in stem cells may generate differentiated thyroid cancer cells that are not sensitive to radioiodine treatment.

Peptide P7 inhibits the bFGF-stimulated proliferation and invasion of SKOV3 cells.

Peptide P7 specifically binds with basic fibroblast growth factor (bFGF) to inhibit the proliferation and invasion of numerous types of cancer cell. However, this effect has remained to be demonstrated in ovarian cancer-derived cell lines. In the present study, the protein P7 was used treat bFGF-stimulated SKOV3 epithelial ovarian cancer cells to explore the therapeutic potential of P7. An MTT and a scratch wound assay were used to respectively evaluate the proliferation and migration of bFGF-stimulated SKOV3 cells treated with P7. Reverse transcription-quantitative polymerase chain reaction analysis was used to detect the gene expression of urokinase-type plasminogen activator (uPA), as well as matrix metallopeptidase (MMP)-2 and −9, which have a role in cell migration/invasion. The morphology and proliferation of SKOV3 cells were not significantly affected by different concentrations of P7. However, P7 had an obvious inhibitory effect on the proliferation and migration of bFGF-stimulated SKOV3 cells. Treatment with P7 significantly lowered the gene expression of uPA, MMP-2 and MMP-9 compared with that in the control group. In conclusion, the present results suggested that P7, which, at least in part, acts through inhibition of bFGF, may have a potential therapeutic application in epithelial ovarian cancer.

Influence of miR-34a on preeclampsia through the Notch signaling pathway.

The aim of this study was to investigate the influence of micro-ribonucleic acid-34a (miR-34a) on preeclampsia through the Notch signaling pathway. The expressions of miR-34a, Notch-1, Notch-2, and Notch-3 in the placenta of 39 preeclampsia patients and 42 normal patients were detected by immunohistochemistry and Reverse Transcription-Polymerase Chain Reaction (RT-PCR). The correlations between miR-34a expression with the expressions of Notch-1, Notch-2 and Notch-3 were analyzed, respectively. Besides, placental trophoblasts were isolated from preeclampsia patients and cultured in vitro. The expressions of miR-34a, Notch-1, Notch-2 and Notch-3 in placental trophoblasts were analyzed. Furthermore, the influences of miR-34a on the protein expressions of Notch-1, Notch-2, Notch-3, and hairy and enhancer of split-1 (Hes-1) in the Notch signaling pathway were analyzed by Luciferase reporter gene assay and Western blotting. The role of Notch in trophoblast invasion was investigated through the Notch inhibitors. In addition, its influence on the expression of urokinase-type plasminogen activator (uPA) was studied by miR-34a overexpression. The expressions of miR-34a and Notch-1 were correlated with preeclampsia in the placentas of preeclampsia patients and normal patients to a certain degree. The expression of miR-34a in preeclamptic placenta was significantly higher than that of the normal placenta (p<0.05). However, Notch-1 expression was markedly lower in preeclamptic placenta (p<0.05). No significant differences were found in the expressions of Notch-2 and Notch-3 between the two types of placentas (p>0.05). MiR-34a had a remarkable negative correlation with Notch-1 expression in the Notch family (p<0.001, r=-0.5775). RT-PCR results revealed that the mRNA expression of miR-34a in placental trophoblasts of patients with preeclampsia was notably higher than that of normal people (p<0.01). However, Western blotting demonstrated that the protein expressions of Notch-1, Notch-2 and Notch-3 exhibited the opposite results. Additionally, the protein expression of Notch-1, Notch-2, Notch-3 and Hes-1 in trophoblasts transfected with pre-miR-34a was significantly decreased. The treatment with Notch inhibitors markedly reduced the trophoblast invasion. Furthermore, miR-34a overexpression or intracellular domain of Notch (ICN) overexpression regulated uPA expression. MiR-34a regulates uPA system through the Notch signal transduction, thereby regulating the invasion of placental trophoblasts in patients with preeclampsia.

Expression of urokinase-type plasminogen activator and its receptor in squamous cell carcinoma of the oral tongue.

Urokinase-type plasminogen activator (uPA) and its receptor (uPAR) act in the proteolysis of basement membrane and extracellular matrix structures, facilitating tumor invasion. The purpose of this study was to evaluate the relationship between these proteins and clinicopathological parameters in squamous cell carcinoma of the oral tongue (SCCOT). Sixty cases of SCCOT were submitted to immunohistochemistry and analyzed semiquantitatively at the invasion front and in the tumor core. The results were associated with lymph node metastasis, clinical stage, locoregional recurrence, clinical outcome and histological grade of malignancy. A higher expression of uPA was observed in cases of tumors of high-grade versus low-grade malignancy (p = 0.010). Moreover, the cases with the worst pattern of invasion presented an overexpression of uPA (p = 0.011). The presence of locoregional recurrence was associated with uPAR (p = 0.039), and the expression of both biomarkers was much higher at the invasion front than in the tumor core (p < 0.001). The results suggest uPA and uPAR are involved in the progression and aggressiveness of SCCOT, mainly at the tumor-host interface.

Involvement of RSK1 activation in malformin-enhanced cellular fibrinolytic activity

Pharmacological interventions to enhance fibrinolysis are effective for treating thrombotic disorders. Utilizing the in vitro U937 cell line-based fibrin degradation assay, we had previously found a cyclic pentapeptide malformin A1 (MA1) as a novel activating compound for cellular fibrinolytic activity. The mechanism by which MA1 enhances cellular fibrinolytic activity remains unknown. In the present study, we show that RSK1 is a crucial mediator of MA1-induced cellular fibrinolysis. Treatment with rhodamine-conjugated MA1 showed that MA1 localizes mainly in the cytoplasm of U937 cells. Screening with an antibody macroarray revealed that MA1 induces the phosphorylation of RSK1 at Ser380 in U937 cells. SL0101, an inhibitor of RSK, inhibited MA1-induced fibrinolytic activity, and CRISPR/Cas9-mediated knockout of RSK1 but not RSK2 suppressed MA1-enhanced fibrinolysis in U937 cells. Synthetic active MA1 derivatives also induced the phosphorylation of RSK1. Furthermore, MA1 treatment stimulated phosphorylation of ERK1/2 and MEK1/2. PD98059, an inhibitor of MEK1/2, inhibited MA1-induced phosphorylation of RSK1 and ERK1/2, indicating that MA1 induces the activation of the MEK-ERK-RSK pathway. Moreover, MA1 upregulated the expression of urokinase-type plasminogen activator (uPA) and increased uPA secretion. These inductions were abrogated in RSK1 knockout cells. These results indicate that RSK1 is a key regulator of MA1-induced extracellular fibrinolytic activity.

Downregulation of SPINK13 Promotes Metastasis by Regulating uPA in Ovarian Cancer Cells

Background/Aims: Ovarian cancer (OC) is the fifth leading cause of cancer-related death in women, and it is difficult to diagnose at an early stage. The purpose of this study was to explore the prognostic biological markers of OC. Methods: Univariate Cox regression analysis was used to identify genes related to OC prognosis from the Cancer Genome Atlas(TCGA) database. Immunohistochemistry was used to analyse the level of SPINK13 in OC and normal tissues. Cell proliferation, apoptosis and invasion were performed using MTT assay, flow cytometric analysis and Transwell assay, respectively. Results: We identified the Kazal-type serine protease inhibitor-13 (SPINK13) gene related to OC prognosis from the Cancer Genome Atlas (TCGA) database by univariate Cox regression analysis. Overexpression of SPINK13 was associated with higher overall survival rate in OC patients. Immunohistochemistry showed that the level of SPINK13 protein was significantly lower in OC tissues than in normal tissues (P < 0.05).In vitro experiments showed that the overexpression of SPINK13 inhibited cellular proliferation and promoted apoptosis. Moreover, SPINK13 inhibited cell migration and epithelial to mesenchymal transition (EMT). SPINK13 was found to inhibit the expression of urokinase-type plasminogen activator (uPA), while recombinant uPA protein could reverse the inhibitory effect of SPINK13 on OC metastasis. Conclusion: These results indicate that SPINK13 functions as a tumour suppressor. The role of SPINK13 in cellular proliferation, apoptosis and migration is uPA dependent, and SPINK13 may be used as a potential biomarker for diagnosis and targeted therapy in OC.

Inflammation-driven colon neoplasmatogenesis in uPA-deficient mice is associated with an increased expression of Runx transcriptional regulators

Deregulation of the bone morphogenetic protein (BMP) pathway has been documented in colorectal cancer (CRC). Previously, we investigated possible associations between urokinase-type plasminogen activator (uPA) deficiency and expression of extracellular constituents of BMP signaling in a newly developed mouse model of inflammation-driven intestinal neoplasmatogenesis, in which chronic colitis and CRC are induced using dextran sodium sulfate (DSS). In this report, we explored the contribution of intracellular components of Smad-mediated BMP signal transduction using the same model. Interestingly, upon DSS treatment, we noticed an overexpression of Runx1/2/3 transcription factors in both wild-type and uPA-deficient mice. Moreover, Runx1 and Runx2 expression levels exhibited an even higher increase in DSS-treated/uPA-deficient mice as compared to DSS-treated/wild-type animals. In all experimental conditions, in situ investigation of Runx-expressing cell types, revealed detection of all three Runx in the immune cells, yet in the DSS-treated/uPA-deficient mice Runx1 and Runx2 were also identified in the preneoplastic epithelium of advanced high-grade dysplasia and carcinoma in-situ colonic lesions. Finally, the uPA-deficient pro-tumorigenic colitic microenvironment exhibited increased levels of the Runx-induced target genes Snai2, Bim and Claudin1, known to have a role in tumor development and progression. These findings suggest that the absence of uPA correlates with increased levels of Runx transcriptional regulators in a way that promotes inflammation-associated carcinogenesis.

Anticancer Effects of Colchicine on Hypopharyngeal Cancer

Colchicine is an alkaloid widely used for the treatment of inflammatory diseases, such as gout. It suppresses cell division by inhibiting mitosis. We investigated the anticancer effects of colchicine on human hypopharyngeal cancer cells and the mechanisms underlying its anticancer effects. XTT cell proliferation assay showed that colchicine inhibited the growth and proliferation of human hypopharyngeal cancer cells (FaDu and SNU1041) in a dose- and time-dependent manner. Colchicine also inhibited the migration, invasion, and adhesion of hypopharyngeal cancer cells in a dose-dependent manner. The levels of mRNA expression and activity of matrix metalloproteinase-9 (MMP9) and urokinase-type plasminogen activator (uPA) decreased after treatment with colchicine. Further investigation revealed that colchicine inhibited the phosphorylation of the FAK/SRC complex and paxillin. Tumor volume ratios in colchicine-treated mice (0.1 mg/kg, every 2 days for 14 days) increased less than in control mice. To our knowledge, this is the first report showing that colchicine can suppress cell invasion, migration, and adhesion through reduced expression of MMP9, the uPA system, and the FAK/SRC complex. Colchicine has the potential to prevent disease progression in hypopharyngeal cancer and may have application as an adjunctive treatment.

PATZ1 knockdown enhances malignant phenotype in thyroid epithelial follicular cells and thyroid cancer cells.

This study was designed to examine the involvement of PATZ1 in carcinogenesis and dedifferentiation of thyroid cancer. Immunohistochemistry on clinical specimens indicated nuclear PATZ1 expression in all normal thyroid glands and adenomatous goiter, while nuclear PATZ1 expression decreased along with the dedifferentiation of thyroid cancer. Knockdown of nuclear PATZ1 by siRNA in an immortalized normal follicular epithelial cell line (Nthy-ori 3-1) altered cellular morphology and significantly increased cell proliferation, migration, and invasion. In addition, the expression of urokinase-type plasminogen activator (uPA), matrix metalloproteinase (MMP) 2, MMP9, and MMP11 was increased by PATZ1 knockdown in Nthy-ori 3-1 cells. When PATZ1 was silenced in differentiated thyroid cancer (DTC) cell lines (TPC-1 and FTC-133), proliferation, cellular motility, and expression of uPA and MMPs were significantly increased. Forced expression of exogenous PATZ1 decreased proliferation, cellular motility, and the expression of uPA and MMPs in ATC cell lines (ACT-1 and FRO). In thyroid cancer cell lines, PATZ1 functioned as a tumor suppressor regardless of p53 status. Moreover, the ratio of nuclear PATZ1 positive tumors was significantly decreased in ATC irrespective of p53 status. Our study demonstrates that PATZ1 knockdown enhances malignant phenotype both in thyroid follicular epithelial cells and thyroid cancer cells, suggesting that PATZ1 functions as a tumor suppressor in thyroid follicular epithelial cells and is involved in the dedifferentiation of thyroid cancer.