Plasminogen Activator Production Research Articles

Fibrin and Fibrinolytic Enzyme Cascade in Thrombosis: Unravelling the Role.

Blood clot formation in blood vessels (thrombosis) is a major cause of life-threatening cardiovascular diseases. These clots are formed by αA-, βB-, and ϒ-peptide chains of fibrinogen joined together by isopeptide bonds with the help of blood coagulation factor XIIIa. These clot structures are altered by various factors such as thrombin, platelets, transglutaminase, DNA, histones, and red blood cells. Various factors are used to dissolve the blood clot, such as anticoagulant agents, antiplatelets drugs, fibrinolytic enzymes, and surgical operations. Fibrinolytic enzymes are produced by microorganisms (bacteria, fungi, etc.): streptokinase of Streptococcus hemolyticus, nattokinase of Bacillus subtilis YF 38, bafibrinase of Bacillus sp. AS-S20-I, longolytin of Arthrobotrys longa, versiase of Aspergillus versicolor ZLH-1, etc. They act as a thrombolytic agent by either enhancing the production of plasminogen activators (tissue or urokinase types), which convert inactive plasminogen to active plasmin, or acting as plasmin-like proteins themselves, forming fibrin degradation products which cause normal blood flow again in blood vessels. Fibrinolytic enzymes may be classified in two groups, as serine proteases and metalloproteases, based on their catalytic properties, consisting of a catalytic triad responsible for their fibrinolytic activity having different physiochemical properties (such as molecular weight, pH, and temperature). The analysis of fibrinolysis helps to detect hyperfibrinolysis (menorrhagia, renal failure, etc.) and hypofibrinolysis (diabetes, obesity, etc.) with the help of various fibrinolytic assays such as a fibrin plate assay, fibrin microplate assay, the viscoelastic method, etc. These fibrinolytic activities serve as a key aspect in the recognition of numerous cardiovascular diseases and can be easily produced on a large scale with a short generation time by microbes and are less expensive.

Gelatinized PLCL Electrospun Membrane for the Prevention of Postoperative Abdominal Adhesion Through Fibrinolysis Activation

AbstractAbdominal adhesion is a common complication after surgery, which causes pain to patient and increases the risks of reoperation. Electrospinning polyester membrane serving as a physical barrier is used for preventing abdominal adhesion; however, the efficacy is limited due to the lacking of bioactivity necessary for antiadhesion. Here, hydrophobic poly(l‐lactide‐co‐caprolactone) (PLCL) electrospinning membranes are hydrophilized with photo‐crosslinked methacrylic anhydride gelatin (GelMA) hydrogel for postoperative abdominal adhesion prevention. The composite membranes show fibrous network architecture, good cytocompatibility, and hydrophilicity than commercial poly(d,l‐lactide acid) (PDLLA) film. Importantly, the composite membrane provides superior antiadhesion effect over PLCL membrane alone or commercial PDLLA film, against abdominal wall–cecum adhesion models in Sprague Dawley (SD) rats. The underlying mechanism is that PLCL/GelMA membrane successfully promotes the secretion of matrix metalloproteinases 9 (MMP‐9) that can reduce the collagen deposition in extracellular matrix (ECM). Furthermore, the composite membrane significantly upregulates the expression of tissue‐type plasminogen activator (t‐PA) and downregulates the production of plasminogen activator inhibitor‐1 (PAI‐1), leading to the activation of fibrinolytic system, which inhibits the formation of adhesive tissues. Collectively, gelatinized PLCL electrospun membrane is a promising bioactive barrier for preventing postsurgery abdominal adhesions.

Production and market comparison of urokinase and streptokinase as effective and cheap fibrinolytic agents for treatment of cardiovascular diseases

Failure of hemostasis and the formation of blood clots in the arteries are the main reasons that provoke the onset of cardiovascular diseases (CVDs) such as myocardial infarction and ischemic stroke. Cardiovascular diseases have become the primary cause of deaths and disabilities across the globe. Therefore, this problem needs to be addressed with urgency. The disintegration of blood clots requires fibrinolytic agents, which are involved in thrombolysis. Streptokinase and urokinase are fibrinolytic enzymes; the former is primarily produced from microbial sources and the latter is isolated from urine, respectively. Streptokinase and urokinase have been in use for a long time to treat cardiovascular diseases. This review explains in detail the comparison of employing streptokinase and urokinase for the said purpose in a cost-effective manner. The recombinant production of both the agents has been discussed in detail. Furthermore, the efficacy of both the agents has been compared based upon their side effects and retention time in the body. A thorough study has been made to compare the influence of using both the agents on the health of cardiovascular patients in the last decade.

Improved cultivation of Chinese hamster ovary cells in bioreactor with reciprocal mixing

We have constructed a new bioreactor with reciprocal mixing that is better suited for the cultivation of delicate animal cells. In-silico simulation (computational fluid dynamics) suggested both maximum and average shear stresses in the bioreactor with reciprocal mixing to be remarkably lower than in a conventional bioreactor with rotary mixing. Although we could not find any difference in growth speed and cell density between the bioreactors with reciprocal and rotary mixing, we did find cell viability in the reciprocal-mixing bioreactor to be retained longer than in the rotary-paddle bioreactor. This implied that cell culture in a bioreactor with reciprocal mixing could be prolonged for the production of target proteins. Leakage of lactate dehydrogenase activity into the culture medium was suppressed much more in the reciprocal-mixing bioreactor than in the rotary-paddle one. Production of human tissue plasminogen activator in the former system was also observed to be much higher than in the latter. Therefore, a bioreactor with reciprocal mixing was concluded to be better suited for the cultivation of animal cells and efficient production of proteins, such as antibody drugs and various growth factors.

Resilience of the Internal Mammary Artery to Atherogenesis: Shifting From Risk to Resistance to Address Unmet Needs.

Fueled by the global surge in aging, atherosclerotic cardiovascular disease reached pandemic dimensions putting affected individuals at enhanced risk of myocardial infarction, stroke, and premature death. Atherosclerosis is a systemic disease driven by a wide spectrum of factors, including cholesterol, pressure, and disturbed flow. Although all arterial beds encounter a similar atherogenic milieu, the development of atheromatous lesions occurs discontinuously across the vascular system. Indeed, the internal mammary artery possesses unique biological properties that confer protection to intimal growth and atherosclerotic plaque formation, thus making it a conduit of choice for coronary artery bypass grafting. Its endothelium abundantly expresses nitric oxide synthase and shows accentuated nitric oxide release, while its vascular smooth muscle cells exhibit reduced tissue factor expression, high tPA (tissue-type plasminogen activator) production and blunted migration and proliferation, which may collectively mitigate intimal thickening and ultimately the evolution of atheromatous plaques. We aim here to provide insights into the anatomy, physiology, cellular, and molecular aspects of the internal mammary artery thereby elucidating its remarkable resistance to atherogenesis. We propose a change in perspective from risk to resilience to decipher mechanisms of atheroresistance and eventually identification of novel therapeutic targets presently not addressed by currently available remedies.

Effects of Feeding OmniGen-AF<sup>&reg</sup> during Superovulation on <i>in Vitro</i> Development of Embryos Recovered from Donor Beef Cows

Embryo quality is crucial when selecting embryos for transfer. Variation in quality may be attributed to poor oocytes, semen, stress, inflammation, and potential immune system dysregulation. OmniGen-AF&reg (OG) feeding supports immune system function and animal health. Our laboratory recently reported lower percent degenerate embryos recovered and increased plasma progesterone in beef cattle donors fed OG during superovulation. In vitro development of embryos recovered from donor cows fed OG prior to collection is presented here. Embryos were recovered from 24 beef cows assigned to four treatment groups: 0 g OG/hd/d and 200 mg Folltropin&reg-V (FSH) (0/200); 0 g OG/hd/d and 400 mg FSH (0/400), 56 g OG/hd/d, 200 mg FSH (56/200) and 56 g OG/hd/d and 400 mg FSH (56/400). Good to excellent quality early blastocysts were cultured for 8 d. and development through hatching, embryonic volume and plasminogen activator (PA) production were quantified. The complete protocol was repeated 90 - 120 d later as Replicate 2. Optimal development was observed by embryos recovered from 0/200 cows where percent blastocysts hatching was greater (P compared to 56/200 and 0/400 cows and embryonic volume was greatest (P was similar (P > 0.10) to 56/400 cows and embryos recovered from 56/400 cows in Replicate 1 produced more (P in vitro embryo development similar to that observed for 0/200 cows.

CLINICAL AND NEUROLOGICAL PECULIARITIES AND THE LEVEL OF AUTOANTIBODIES TO NEUROSPECIFIC ANTIGENS IN PATIENTS WITH CHRONIC HEMOBLASTOSIS

The purpose of this study was to determine the level of autoantibodies to neurospecific antigens and their relationship with clinical manifestations of the nervous system lesions in chronic hematological malignancies. Materials and methods. The study included 86 patients with chronic hemoblastosis divided into 3 groups: group 1 consisted of patients with chronic lymphoid leukaemia (n = 30), group 2 involved patients with chronic myeloid leukaemia (n = 30), group 3 included patients with multiple myeloma (n = 26). 21 inpatients of the neurological hospital without immunological and hematological disorders were selected as a control group. We investigated the anamnestic data on the presence of a clinical diagnosis of the nervous system lesions before and after the establishing a cancerous hematological diagnosis, carried out a complete neurological examination of the patients with chronic hematoblastosis to determine main neurological syndromes. The level of peripheral blood autoantibodies to neurospecific antigens was evaluated by the enzyme immunoassay. The level of circulating immune complexes was determined by the method of selective precipitation of antigen-antibody complexes. The findings of the study were statistically processes by using Microsoft Excel 2019 and IBM SPSS Statistics 26.0 software. The data obtained were processed by descriptive statistics methods. The study has demonstrated that encephalopathy (70%) and radiculopathy (40%) develop statistically significantly more often in the patients with chronic lymphoid leukaemia. The patients with chronic myeloid leukaemia are found to have encephalopathy (43%) and polyneuropathy (20%). The development of multiple myeloma is statistically significantly accompanied by the occurrence of radiculopathy (50%) and polyneuropathy (53%). Conclusion. The nervous system lesions are found considerably frequent in the patients with chronic hematological malignancies. The development of secondary encephalopathy in the patients with chronic lymphoid leukaemia may develop as a consequence of paraneoplastic syndrome due to the excessive production of cytokines and plasminogen activators. The predominance of radiculopathy in chronic myeloid leukaemia can be reliably explained by the high severity of the proliferative syndrome with a tendency to the production of localized chlorine, which can compress the corresponding roots of the peripheral nerves. However, peripheral lesions of nervous system are more typical for patients with multiple myeloma that may be due to hyperparaproteinemia combined with the chemotherapy of the underlying disease. Along with this, chronic lymphoid leukaemia and chronic myeloid leukaemia cause an increase in the circulating immune complexes level. Immunological changes indicate significant damage to nerve fibres and considerably rapid destruction of neurons in all forms of chronic hemoblastosis.

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.

Biosíntesis del agente fibrinolítico uroquinasa por Enterococcus gallinarum aislado de sardine

Fibrinogen degradation by thrombin leads to the formation of fibrin clots in the blood which results in cardiovascular diseases due to which certain fibrinolytic agents are used to treat cardiovascular complications. Urokinase is being favored as a fibrinolytic agent due to its high fibrin specificity and fewer side effects. Therefore, the main objective of this research was the isolation of potent urokinase producing bacteria from soil, seawater, and fermented food samples. Thirty-five isolates that were capable of producing urokinase were isolated using sixty samples and subjected to secondary screening for urokinase estimation. Bacterial isolate QGA-20 that showed maximum enzyme activity 52.6±0.03 FU/ml/min was identified by 18s RNA sequencing as Enterococcus gallinarum which has never been reported before for urokinase production. Different physical and cultural parameters such as fermentation medium, incubation time, temperature, pH, carbon source, nitrogen source, inoculum size, and trace elements were optimized for submerged fermentation to synthesize urokinase. Maximum production of urokinase 81.3±0.02 FU/ml/min was obtained with M-5 fermentation media at 96 hrs of incubation, 37oC temperature, 7 pH, sucrose as carbon, and soya flour as a nitrogen source, 3% inoculum size and CaCl2 as a trace element. In in vitro studies, urokinase enzyme was also applied for the disintegration of clot successfully. This study revealed a potent urokinase producer, reported first time, with appreciable clot lysis ability.

Pathophysiology of atherosclerosis: Association of risk factors and treatment strategies using plant-based bioactive compounds.

Under physiological conditions, endothelial cells act as protective barrier which prevents direct contact of blood with circulating factors via production of tissue plasminogen activator. Risk factors of metabolic disorders are responsible to induce endothelial dysfunction and may consequently lead to prognosis of atherosclerosis. This article summarizes the process of atherosclerosis which involves number of sequences including formation and interaction of AGE-RAGE, activation of polyol pathway, protein kinase C, and hexosamine-mediated pathway. All these mechanisms can lead to the development of oxidative stress which may further aggravate condition. Different pharmacological interventions are being used to treat atherosclerosis, however, these might be associated with mild to severe side effects. Therefore, plant-based bioactive compounds having potential to combat and prevent atherosclerosis in diabetic patients are attaining recent focus. By understanding process of development and mechanisms involved in atherosclerotic plaque formation, these bioactive compounds can be better option for future therapeutic interventions for atherosclerosis treatment. PRACTICAL APPLICATIONS: Atherosclerosis is one of major underlying disorders of cardiovascular diseases which occur through multiple mechanisms and is associated with metabolic disorders. Conventional therapeutic interventions are not only used to treat atherosclerosis, but are also commonly associated with mild to severe side effects. Therefore, nowadays, bioactive compounds having potential to combat and prevent atherosclerosis in diabetic patients are preferred. By understanding mechanisms involved in atherosclerotic plaque formation, bioactive compounds can be better understood for treatment of atherosclerosis. In this manuscript, we have focused on treatment strategies of atherosclerosis using bioactive compounds notably alkaloids and flavonoids having diverse pharmacological and therapeutic potentials with special focus on the mechanism of action of these bioactive compounds suitable for treatment of atherosclerosis. This manuscript will provide the scientific insights of bioactive compounds to researchers who are working in the area of drug discovery and development to control pathogenesis and development of atherosclerosis and its associated cardiometabolic disorders.

β-Sitosterol regulated microRNAs in endothelial cells against an oxidized low-density lipoprotein.

β-Sitosterol is a natural compound widely found in many vegetable oils, nuts, and plant medicines; it lowers the cholesterol levels, enhances the production of plasminogen activators, and exhibits anticancer and antiatherogenic effects. However, the direct endothelial protection of β-sitosterol against an oxidized low-density lipoprotein (ox-LDL) is not well understood. In the present study, β-sitosterol significantly inhibited cell apoptosis (P < 0.01), increased cell migration (P < 0.01), improved energy metabolism (P < 0.05) and improved morphology after ox-LDL (50 μg ml-1) exposure following β-sitosterol (2 μg mL-1) treatment in human aortic endothelial cells (HAECs ). A total of 691 differentially expressed (DE) mRNAs were identified (579 were upregulated and 112 were downregulated, fold change ≥2.0, P < 0.05) after 24 h of β-sitosterol administration in transcriptome sequencing (β-sitosterol vs. ox-LDL), which suggested that β-sitosterol reversed 62.32% change in mRNAs induced by ox-LDL. DE mRNAs are enriched mainly in focal adhesion, ribosomes, eukaryotic translation elongation, etc. Considering that one of the enrichment is 3'-UTR-mediated translational regulation, we explored DE microRNA (miRNA). The miRNA-seq data proposed 87 up-regulated and 58 down-regulated miRNAs (fold change ≥2.0, P < 0.05) in miRNA-seq (β-sitosterol vs. ox-LDL), suggesting that β-sitosterol reversed 76.67% change in miRNAs induced by ox-LDL. The DE miRNA-DE mRNA coexpression network focused on ribosomes, cell cycle, oxidative phosphorylation, PI3K-Akt signaling pathway, TNF signaling pathway, ErbB signaling pathway, and mTOR signaling pathway. Consequently, miRNAs might be the targets of β-sitosterol and play vital roles in transcriptional regulation in endothelial protective and antiatherogenic effects against ox-LDL.

Stimulation of Phagocytic Activity in Cultured Human Corneal Fibroblasts by Plasminogen.

Plasminogen has been detected in the corneal stroma after tissue injury and interacts with corneal fibroblasts during wound healing. We examined the effect of plasminogen on phagocytic activity of corneal fibroblasts. Cultured human corneal fibroblasts were exposed to plasminogen and then incubated with fluorescent microparticles before measurement of phagocytic activity by confocal microscopy or flow cytometry. The binding of corneal fibroblasts to immobilized plasminogen was quantitated with a real-time biomolecular interaction assay. The production of urokinase-type plasminogen activator (uPA), matrix metalloproteinases (MMPs), and IL-1β by corneal fibroblasts was measured by fibrin zymography, by immunoblot analysis or gelatin zymography, or with an enzyme-linked immunosorbent assay, respectively. Plasminogen increased phagocytic activity of corneal fibroblasts in a concentration- and time-dependent manner, with the maximal effect apparent at 30 μg/mL and 24 hours. Corneal fibroblasts bound to immobilized plasminogen in a manner dependent on time and cell number, and the stimulatory effect of plasminogen on phagocytic activity was blocked in the presence of epsilon-aminocaproic acid, an inhibitor of plasminogen binding to cell surface receptors. Plasminogen-induced phagocytic activity was not associated with changes in the production of uPA, MMPs, or IL-1β by corneal fibroblasts. Plasminogen induced phagocytic activity in corneal fibroblasts in a manner dependent on its binding to the cell surface. This effect was not associated with increased production of proteases or IL-1β. Thus, plasminogen may promote the clearance of foreign particles or damaged tissue components by corneal fibroblasts early after tissue injury.

Fiber-Reinforced Silk Composite for Enhanced Urokinase Production Using High-Density Perfusion Culture and Bioactive Molecule Supplementation.

Urokinase plasminogen activator (uPA) has been extensively used as a thrombolytic drug in cases of myocardial infarction, thromboembolism, and ischemic brain stroke. Media optimization and high-density perfusion culture are the decisive factors that facilitate enhanced urokinase production in a conditioned medium. In this study, we have aimed for a high-density perfusion culture of HT1080, a human fibrosarcoma cell line, by formulating optimal media for enhanced urokinase productivity. Four scaffold variants were fabricated from silk fibroin and microfibers of Bombyx mori (BM) and Antheraea assamensis (AA) and physico-chemically characterized. Field emission scanning electron microscopy studies revealed a heterogeneous distribution of pores with interconnected networks supporting cell infiltration, attachment, and long-term viability. AA-based fiber-reinforced scaffolds (ASAF) demonstrated superior mechanical strength, integral stability, and increased cell proliferation as compared to pure silk scaffolds. Media formulation was accomplished by limiting serum concentration (2% FBS) and supplementing with 20 μg/mL arginine and 20 ng/mL TGF-β1 to retain the stationary phase of cells and augment the urokinase production. A perfusion bioreactor culture of HT1080-laden scaffolds in the presence of formulated media was performed for improving the production of urokinase, with a maximum activity of 432 U/L. Also, gene expression analysis revealed that the individual silk scaffolds have different effects on regulating the expression of plasminogen activator urokinase and plasminogen activator urokinase receptor. In brief, our results suggest that a perfusion bioreactor culture of HT1080-laden ASAF scaffolds in formulated media promotes an increased urokinase production, such that it can be further used as a novel 3D matrix platform for industrial production of the lifesaving uPA drug.

The Role of Abnormal Hemostasis and Fibrinolysis in Morbidity and Mortality of Acute Promyelocytic Leukemia.

Despite the improved therapeutic advances in the management of acute promyelocytic leukemia (APL), a significant early mortality during induction, also referred to as early death (ED), remains an obstacle for further improvement in outcome. Hemorrhagic complications are the most common cause of morbidity and mortality. Perturbed hemostatic dysfunction is present as the result of abnormalities in both the coagulation and the fibrinolytic systems. The activation of coagulation is distinct from the classical disseminated intravascular coagulation. Multiple abnormalities in the fibrinolytic system have recently been identified. The most significant change is increased production of tissue plasminogen activator (tPA) and its receptor annexin A2 by the APL promyelocytes. Among the hemorrhagic complications, intracranial hemorrhage predominates. The pathogenesis of this catastrophic event is elucidated by new evidence of adverse effect of tissue plasminogen activator (tPA) on the brain, including both the plasmin-dependent and plasmin-independent pathways. In order to address the hemorrhagic complications, a thorough understanding of the hemostatic dysfunction is essential. In this article, our current concept of the abnormal hemostasis in APL is reviewed. The failure to reduce the early death rate, despite the introduction of effective therapy, will also be discussed.

Production of human tissue-type plasminogen activator (htPA) using in vitro cultured transgenic pig mammary gland cells

Tissue plasminogen activator (tPA) is a protein involved in the breakdown of blood clots. We have previously produced a human tPA (htPA)-overexpressing transgenic pig using a mammary gland-specific promoter. In this study, we have established a transgenic pig mammary gland cell line that produces recombinant htPA. The mammary gland cells grew well and retained their character over long periods of culture. There was no difference in the extent of apoptosis in transgenic cells compared to wild-type mammary gland cells. In addition, the transgenic mammary gland cells expressed and secreted htPA into the conditioned media at a concentration similar to that in milk. This transgenic cell line represents a simple and ethical method for recombinant htPA production.

In silico and in vivo analyses of the mutated human tissue plasminogen activator (mtPA) and the antithetical effects of P19 silencing suppressor on its expression in two Nicotiana species

Human tissue-type plasminogen activator is one of the most important therapeutic proteins involved in the breakdown of blood clots following the stroke. A mutation was found at position 1541 bp (G514E) and the mutated form was cloned into the binary vector pTRAc-ERH. In silico analysis showed that this mutation might have no significant effect on the active site of the tissue plasminogen activator enzyme. Accordingly, zymography assay confirmed the serine protease activity of the mutated form and its derivatives. The expression of the mutated form was verified with/without co-agroinjection of the P19 gene silencing suppressor in both Nicotiana tabacum and N. benthamiana. The ELISA results showed that the concentration of the mutated form in the absence of P19 was 0.65% and 0.74% of total soluble protein versus 0.141% and 1.36% in the presence of P19 in N. benthamiana and N. tabacum, respectively. In N. tabacum, co-agroinjection of P19 had the synergistic effect and increased the mutated tissue plasminogen activator production two-fold higher. However, in N. benthamiana, the presence of P19 had the adverse effect of five-fold reduction in the concentration. Moreover, results showed that the activity of the mutated form and its derivatives was more than that of the purified commercial tissue plasminogen activator.

Estramustine Phosphate Inhibits TGF-β-Induced Mouse Macrophage Migration and Urokinase-Type Plasminogen Activator Production.

Transforming growth factor-beta (TGF-β) has been demonstrated as a key regulator of immune responses including monocyte/macrophage functions. TGF-β regulates macrophage cell migration and polarization, as well as it is shown to modulate macrophage urokinase-type plasminogen activator (uPA) production, which also contributes to macrophage chemotaxis and migration toward damaged or inflamed tissues. Microtubule (MT) cytoskeleton dynamic plays a key role during the cell motility, and any interference on the MT network profoundly affects cell migration. In this study, by using estramustine phosphate (EP), which modifies MT stability, we analysed whether tubulin cytoskeleton contributes to TGF-β-induced macrophage cell migration and uPA expression. We found out that, in the murine macrophage cell line RAW 264.7, EP at noncytotoxic concentrations inhibited cell migration and uPA expression induced by TGF-β. Moreover, EP greatly reduced the capacity of TGF-β to trigger the phosphorylation and activation of its downstream Smad3 effector. Furthermore, Smad3 activation seems to be critical for the increased cell motility. Thus, our data suggest that EP, by interfering with MT dynamics, inhibits TGF-β-induced RAW 264.7 cell migration paralleled with reduction of uPA induction, in part by disabling Smad3 activation by TGF-β.

Evaluating the Role of the Na+‐H+ Exchanger Isoform 1 (NHE1) in Lysophosphatidic Acid and Urokinase‐type Plasminogen Activator Stimulation of Ovarian Cancer Cells

Lysophosphatidic Acid (LPA) is a bioactive lipid which stimulates an increase in cell proliferation and migration in a range of normal and cancer lines in part through the activation of both the RhoA/Rock and Ras/ErK signaling pathways. Plasma levels of LPA are elevated in ovarian cancer patients even in the early stages of the disease where it is believed to participate in pathogenesis. In patients with more advanced stages of ovarian cancer, the higher the levels of LPA found in the blood and ascites is a negative prognostic indicator. In ovarian cancer, LPA enhances cell proliferation and promotes cell survival by preventing apoptosis and anoikis. LPA also decreases the sensitivity of the ovarian cancer cells to cisplatin and other chemotherapy drugs. While LPA signaling can directly enhance ovarian cancer progression, it can also act by stimulating the production and release of urokinase‐type plasminogen activator (uPA). uPA is a serine protease that binds to a plasma membrane receptor stimulating activation of both the RhoA/Rock and Ras/ErK signaling pathways. Studies in non‐small cell lung cancer cells have shown that LPA and uPA activate the Na+‐H+ Exchanger Isoform 1 (NHE1) leading to an increase in cell proliferation and migration. NHE1 is an 815 amino acid transmembrane protein that exchanges 1 extracellular Na+ for 1 intracellular H+ leading to the pH inversion common to cancer cells in a developing tumor. Activation of NHE1 is one of the initial steps in the development of the transformed phenotype of cancer cells. NHE1 can be phosphorylated and activated by Rock and kinases in the Ras/Erk signaling pathway. Recently, increases in NHE1 expression was demonstrated to play an important role in cancer progression in ovarian cancer patients. In this project, we specifically explore the relationship between LPA and uPA signaling to NHE1 in the regulation of cell proliferation, stress fiber formation, and cell migration in three ovarian cancer cell lines, CAOV‐3, SKOV‐3, and OVCAR‐3.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Lectin BS-I inhibits cell migration and invasion via AKT/GSK-3β/β-catenin pathway in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is most common malignant cancer worldwide; however, the mortality rate of HCC remains high due to the invasion and metastasis of HCC. Thus, exploring novel treatments to prevent the invasion of HCC is needed for improving clinical outcome of this fatal disease. In this study, we identified lectin from Bandeiraea simplicifolia seeds (BS‐I) binds to metastasis‐associated HCC cell surface glycans by a lectin microarray and inhibits HCC cell migration and invasion through downregulating the matrix metalloproteinase 2 (MMP2), matrix metalloproteinase 9 (MMP9) and urokinase‐type plasminogen activator (uPA) production. These effects of BS‐I were mediated by inhibiting the activation of AKT/GSK‐3β/β‐catenin pathway and depended on specificity of lectin BS‐I binding to GalNAc. GSK3β inhibitors rescued BS‐I‐mediated inhibition of migration and invasion of HCC cell. Further, we identified that lectin BS‐I interacts with sGrp78, affects membrane localization of sGrp78 and attenuates the binding of sGrp78 and p85 to inhibit the activation of AKT/GSK‐3β/β‐catenin pathway. Overexpression of Grp78 or P85 rescues BS‐I‐mediated inhibition of migration and invasion of HCC cell. These findings demonstrated for the first time that BS‐I can act as a novel potential drug to prevent the invasion of HCC.

Endothelial cells promote triple-negative breast cancer cell metastasis via PAI-1 and CCL5 signaling.

Endothelial cells (ECs) in the tumor microenvironment have been reported to play a more active role in solid tumor growth and metastatic dissemination than simply providing the physical structure to form conduits for blood flow; however, the involvement of ECs in the process of triple-negative breast cancer (TNBC) metastasis has not been addressed. Here, we demonstrate that ECs-when mixed with TNBC cells-could increase TNBC cell metastatic potency. After treatment with TGF-β to induce endothelial-mesenchymal transition (EMT), TNBC cells could produce plasminogen activator inhibitor-1 (PAI-1) and stimulate the expression and secretion of the chemokine, CCL5, from ECs, which then acts in a paracrine fashion on TNBC cells to enhance their migration, invasion, and metastasis. CCL5, in turn, accelerates TNBC cell secretion of PAI-1 and promotes TNBC cell metastasis, thus forming a positive feedback loop. Moreover, this enhanced metastatic ability is reversible and dependent on CCL5 signaling via the chemokine receptor, CCR5. Of importance, key features of this pathway are manifested in patients with TNBC and in The Cancer Genome Atlas database. Taken together, our results suggest that ECs enhance EMT-induced TNBC cell metastasis via PAI-1 and CCL5 signaling and illustrate the potential of developing new PAI-1- and CCL5-targeting therapy for patients with TNBC.-Zhang, W., Xu, J., Fang, H., Tang, L., Chen, W., Sun, Q., Zhang, Q., Yang, F., Sun, Z., Cao, L., Wang, Y., Guan, X. Endothelial cells promote triple-negative breast cancer cell metastasis via PAI-1 and CCL5 signaling.