The expression of polyspecific transporters in renal tumors and their role in chemotherapeutical treatment

Nierenzellkarzinome (RCC) sind normalerweise Chemotherapie-resistent. Diese Chemoresistenz kann durch Gabe von Zytostatika überwunden werden, für welches die Karzinomazellen einen Aufnahmetransporter exprimieren. In der vorliegenden Arbeit wurde sowohl die Expression unterschiedlicher löslichen Carrier Transporter (SLC) in verschiedenen RCC-Proben als auch ihre Fähigkeit, mit verschieden Chemotherapeutika zu interagieren, untersucht. Dafür wurden fünf Nierenkarzinoma-Linien, gesundes Gewebe und Nierentumorgewebe mittels RT-PCR und TaqMan real-time-PCR analysiert. In zwei der untersuchten Zelllinien, A498 und 786-O, konnte eine signifikant erhöhte Expression von SLC22A3 (hOCT3) detektiert werden. Die Aufnahme des organischen Kations [3H]MPP (4-Methylpyrididium-Iodid) in diese Zellen und in stabil transfizierte hOCT3-exprimierende CHO-Zellen (CHO-hOCT3) wurde durch die Agenzien Irinotekan, Vincristin und Mephalan inhibiert. Die durch Dixon-Plots bestimmten Ki-Werte für Irinotekan und Vincristin und Mephalan sind 1,72+-0,45 μM, 17,0+-4,81 μM und 366,0+-51,0 μM. Die zytotoxischen Aktivitäten dieser Drogen wurden durch [3H]-Thymidin-Inkorporation und MTT-Assays in CHO-hOCT3, A498 (hohe Expression von hOCT3) und ACHN (niedrige Expression von hOCT3) Zellen überprüft. Das Wachstum der CHO-hOCT3 Zellen wurde im Vergleich zu nicht-transfizierten CHO Zellen durch Irinotekan um 20% und durch Vincristin um 50% inhibiert. Mephalan löste eine 20-30%ige Wachstumsinhibierung in hOCT3 transfizierten im Vergleich zu nicht-transfizierten Zellen aus. Ähnliche Ergebnisse wurden für A498 und ACHN Zellen erhalten. Damit unterstützen die in dieser Arbeit erhaltenen Daten die Hypothese, wonach die Sensitivität von Tumorzellen für Chemotherapeutika von der Expression von Transporter-Proteinen abhängig ist, welche die spezifische Akkumulation der Drogen in den Zielzellen vermitteln

Relationship between stretch zone parameters and fracture toughness of heat-resistance steel

Predicting the product a customer would like to buy is an increasingly important field of study and there are several different recommender system models that are used to make recommendations for users. Deep learning has shown effective results in a variety of predictive tasks but there haven’t been much research concerning its usage in recommender systems. This thesis studies the effectiveness of using a long short term memory implementation (LSTM) of a recurrent neural network (RNN) as a recommender system by comparing it to one of the most common recommender system implementations, the matrix factorization method. A radio playlist dataset is used to train both the LSTM and the matrix factorization models with the intent of generating accurate predictions. We were unable to create a LSTM model with good performance and due to that we are unable to make any significant conclusions regarding whether or not LSTM networks outperform matrix factorization models

Pattern of mRNA expression of β-defensins in basal cell carcinoma

BACKGROUND: Although the human β-defensins hBDs today seem to have diverse functional activities in innate antimicrobial immunity, a few reports also indicated an altered expression of these antimicrobial peptides (AMPs) in tissues of cancers such as oral squamous cell carcinoma. The present work was aimed on the study of hBD gene expression in basal cell carcinoma (BCC) which is the most common cancer in humans. METHODS: Twenty-two non-ulcerated BCCs (12 nodular type, 10 superficial type) have been analysed for the presence of hBD (1–3) mRNA by quantitative real-time RT-PCR. As controls, non-lesional skin specimens of BCC patients as well as samples of healthy subjects were assessed by RT-PCR. RESULTS: hBD-1 levels in healthy controls and non-lesional skin of BCC patients were significantly (P < 0.05) higher than the levels observed in tumour tissue. Moreover, BCCs showed significantly (P < 0.05) increased mRNA expression of hBD-2 as compared to controls. There was no significant (P > 0.05) difference between lesional mRNA levels for hBD-3 and those levels observed in controls. The mRNA expression of hBDs (1–3) found in nodular and superficial BCCs did not significantly (P > 0.05) differ. CONCLUSION: The gene expression patterns of hBD-1 and hBD-2 are for the first time shown to be significantly altered in non-ulcerated BCCs as compared to intra-individual and inter-individual controls, respectively. The present findings may indicate that beside the antimicrobial activity of AMPs, hBDs may also play a role in the pathogenesis of BCC. However, functional and immunohistological studies investigating hBDs in patients with BCC are needed to confirm our data

Downregulation of organic cation transporters OCT1 (SLC22A1) and OCT3 (SLC22A3) in human hepatocellular carcinoma and their prognostic significance

<p>Abstract</p> <p>Background</p> <p>Organic cation transporters (OCT) are responsible for the uptake and intracellular inactivation of a broad spectrum of endogenous substrates and detoxification of xenobiotics and chemotherapeutics. The transporters became pharmaceutically interesting, because OCTs are determinants of the cytotoxicity of platin derivates and the transport activity has been shown to correlate with the sensitivity of tumors towards tyrosine kinase inhibitors. No data exist about the relevance of OCTs in hepatocellular carcinoma (HCC).</p> <p>Methods</p> <p>OCT1 (<it>SLC22A1</it>) and OCT3 (<it>SLC22A3</it>) mRNA expression was measured in primary human HCC and corresponding non neoplastic tumor surrounding tissue (TST) by real time PCR (n = 53). Protein expression was determined by western blot analysis and immunofluorescence. Data were correlated with the clinicopathological parameters of HCCs.</p> <p>Results</p> <p>Real time PCR showed a downregulation of <it>SLC22A1 </it>and <it>SLC22A3 </it>in HCC compared to TST (p ≤ 0.001). A low <it>SLC22A1 </it>expression was associated with a worse patient survival (p < 0.05). Downregulation was significantly associated with advanced HCC stages, indicated by a higher number of T3 tumors (p = 0.025) with a larger tumor diameter (p = 0.035), a worse differentiation (p = 0.001) and higher AFP-levels (p = 0.019). In accordance, <it>SLC22A1 </it>was less frequently downregulated in tumors with lower stages who underwent transarterial chemoembolization (p < 0.001) and liver transplantation (p = 0.001). Tumors with a low <it>SLC22A1 </it>expression (< median) showed a higher <it>SLC22A3 </it>expression compared to HCC with high <it>SLC22A1 </it>expression (p < 0.001). However, there was no significant difference in tumor characteristics according to the level of the <it>SLC22A3 </it>expression.</p> <p>In the western blot analysis we found a different protein expression pattern in tumor samples with a more diffuse staining in the immunofluorescence suggesting that especially OCT1 is not functional in advanced HCC.</p> <p>Conclusion</p> <p>The downregulation of OCT1 is associated with tumor progression and a worse patient survival.</p

Signaling through the primary cilium

© 2018 Wheway, Nazlamova and Hancock. The presence of single, non-motile "primary" cilia on the surface of epithelial cells has been well described since the 1960s. However, for decades these organelles were believed to be vestigial, with no remaining function, having lost their motility. It wasn't until 2003, with the discovery that proteins responsible for transport along the primary cilium are essential for hedgehog signaling in mice, that the fundamental importance of primary cilia in signal transduction was realized. Little more than a decade later, it is now clear that the vast majority of signaling pathways in vertebrates function through the primary cilium. This has led to the adoption of the term "the cells's antenna" as a description for the primary cilium. Primary cilia are particularly important during development, playing fundamental roles in embryonic patterning and organogenesis, with a suite of inherited developmental disorders known as the "ciliopathies" resulting from mutations in genes encoding cilia proteins. This review summarizes our current understanding of the role of these fascinating organelles in a wide range of signaling pathways

Xenopus Pkdcc1 and Pkdcc2 Are Two New Tyrosine Kinases Involved in the Regulation of JNK Dependent Wnt/PCP Signaling Pathway

Protein Kinase Domain Containing, Cytoplasmic (PKDCC) is a protein kinase which has been implicated in longitudinal bone growth through regulation of chondrocytes formation. Nevertheless, the mechanism by which this occurs remains unknown. Here, we identified two new members of the PKDCC family, Pkdcc1 and Pkdcc2 from Xenopus laevis. Interestingly, our knockdown experiments revealed that these two proteins are both involved on blastopore and neural tube closure during gastrula and neurula stages, respectively. In vertebrates, tissue polarity and cell movement observed during gastrulation and neural tube closure are controlled by Wnt/Planar Cell Polarity (PCP) molecular pathway. Our results showed that Pkdcc1 and Pkdcc2 promote the recruitment of Dvl to the plasma membrane. But surprisingly, they revealed different roles in the induction of a luciferase reporter under the control of Atf2 promoter. While Pkdcc1 induces Atf2 expression, Pkdcc2 does not, and furthermore inhibits its normal induction by Wnt11 and Wnt5a. Altogether our data show, for the first time, that members of the PKDCC family are involved in the regulation of JNK dependent Wnt/PCP signaling pathway.Fundacao Ciencia e Tecnologia - IP; IBB/CBME, LA [PTDC/BIA-BCM/69912/2006, Pest-OE/EQB/LA0023/2013]; FCTinfo:eu-repo/semantics/publishedVersio

Die Funktion von PTK7 in der Neuralleistenzellmigration in Xenopus laevis

PTK7 ist ein neuer Bestandteil des Signalwegs der planaren Zellpolarität (PCP), welcher die Polarität der Innenohrhaarzellen und den Neuralrohrschluss reguliert. Der Signalmechanismus jedoch ist weitgehend unbekannt. In Xenopus wird PTK7 im Neuralrohr sowie in den Neuralleistenzellen exprimiert, was eine Funktion in der Regulation morphogenetischer Bewegungen anzeigt. Für den Neuralrohrschluss ist dieses bereits demonstriert worden, doch die Rolle von PTK7 bei der Migration der Neuralleistenzellen ist unklar. Das Ziel dieser Doktorarbeit war es, zu analysieren, ob PTK7 die Neuralleistenzellmigration beeinflusst und wie dieses mit dem PCP-Signalweg in Verbindung steht, um Zellbewegungen zu regulieren. Die Untersuchung von Protein Ko-lokalisationen in ektodermalen Explantaten von Xenopus Embryonen hat gezeigt, dass dsh durch PTK7 zur Plasmamembran transloziert wird. Durch Ko-Immuno- präzipitationsexperimente konnte weiterhin gezeigt werden, dass PTK7 ein Bestandteil des fz7-dsh Komplexes ist, der für die fz7-abhängige dsh Membranrekrutierung und Phosphorylierung notwendig ist. Da unsere Daten zeigten, dass die PTK7-dsh Interaktion nicht direkt ist, wurde eine Tandem-Massenspektrometrie Analyse durchgeführt, um neue Bindungspartner von PTK7 zu identifizieren. Als Bindungspartner von PTK7 wurde receptor of activated PKC 1 (RACK1) identifiziert. RACK1 hat ein ähnliches Expressionsmuster wie PTK7 und in Immuno-präzipitationsexperimenten wurde eine Bindung von RACK1 und PTK7 gezeigt. Der Funktionsverlust von RACK1 führt ebenso wie der von PTK7 zu Defekten beim Neuralrohrschluss. In den gleichen Experimenten wurde außerdem gezeigt, dass RACK1 für die durch PTK7 vermittelte dsh Lokalisierung benötigt wird. Die Analyse der Protein-Protein-Interaktionen sowie Ko-Lokalisationsstudien zeigten, dass RACK1 mit PKCδ1 interagiert und der PKCδ1-RACK1 Komplex die PTK7-abhängige dsh Membranrekrutierung vermittelt. Der Funktionsverlust von PTK7 demonstriert, dass dieses Protein für die Neuralleistenzellmigration erforderli ch ist. Durch die Expression des kompletten PTK7 Proteins sowie von Deletionsmutanten unter der Kontrolle des neuralleistenzellspezifischen Slug-Promotors, konnte gezeigt werden, dass PTK7 speziell in migrierenden Neuralleistenzellen benötigt wird. Eine genetische Interaktion von PTK7 mit dsh zur Regulation von Neuralleistenzellmigration wurde gezeigt

Dev Dyn

Semaphorins are major regulators of morphogenesis and are involved in a variety of processes ranging from the guidance of cell migration to the development of cancer. Since semaphorins were first characterized as repulsive neuronal guidance cues, their expression has been best documented in the nervous system. However, broader studies are lacking. Here, we describe the expression of 13 members of the semaphorin family and two neuropilin receptors during early Xenopus laevis development. No particular expression pattern defines any of the semaphorin classes, but many are dynamically expressed in distinct areas undergoing morphogenetic cell movements like the developing mesoderm and the migrating neural crest. Furthermore, the complementary expression patterns of Sema3A/Nrp1 and Sema3F/Nrp2 are maintained across hundreds of millions of years, possibly indicating a conserved role in the guidance of migrating neural crest cells

Дослідження надпружної поведінки NiTi сплаву за циклічного навантаження

Методом диференціальної сканувальної калориметрії досліджено температуру прямих і зворотних фазових переходів нікельтитанового сплаву Ni55,8Ti44,2. Зіставлення температур фазових переходів підтверджує зворотний характер зміни кристалографічної структури досліджуваного матеріалу. Під час нагрівання зразка фазовий перехід відбувається в діапазоні температур між -60,5°С та -38,7°С, а температура переходу становить - 45,7°С. Таким чином, температури початку і завершення аустенітної фази складали відповідно As = -60,5°С і Af = - 38,7°С. Розроблено методику й досліджено вплив розмаху напруження на закономірності деформування одновісним розтягом і функціональні властивості нікельтитанового сплаву за температури 0 °С в середовищі талого льоду. Характеристики механічних властивостей і вплив циклічного навантаження на функціональні властивості сплаву досліджували за одновісного розтягу циліндричних зразків діаметром 4 мм і довжиною робочої ділянки 12,5 мм, які були вирізані з прутка 8 mm. Частота навантаження за синусоїдальної форми циклу складала 0,5 Hz. Коефіцієнт асиметрії циклу навантаження min max r s / s 0 (тут min max s , s – найменше і найбільше значення переміщення штоку). При температурі вище температури закінчення мартенситно – аустенітного перетворення СПФ, в умовах контрольованого переміщення затискачів, вплив циклічного навантаження на максимальне напруження загалом можна охарактеризувати ділянками зміцнення, знеміцнення, стабілізації і стрімкого падіння максимального напруження, яке спричинене появою та поширенням макротріщини. Для усіх значень початкового максимального напруження, упродовж перших десяти циклів навантаження, спостерігається стрімке зменшення розмаху деформації, потім – ділянка стабілізації розмаху деформації або менш інтенсивного її зменшення, після якої йде ділянка спаду, що завершується руйнуванням зразка. Зі збільшенням максимального напруження у першому циклі навантаження від 509 MPa до 605 MPa збільшується значення розмаху деформації.The phase transformation temperatures of pseudoelastic NiTi alloy were defined by differential scanning calorimetry. The effect of the stress range on the functional properties of the NiTi alloy under uniaxial tension in ice water at a temperature of 00С was studied. The cylindrical specimens with 4 mm in diameter and a gage length of 12.5 mm were tested under static and cyclic loading with frequency of 0.5 Hz. All cyclic tests were performed under the crosshead displacement controlled condition on the STM100 machine. At the temperature above Af the effect of the cyclic loading on the maximum stress in general could be characterized by several stages: strengthening, softening, stabilization and rapid decrease of the maximum stress, which is caused by the initiation and macrocrack growth. With the increase of the maximal stress in the first cycle from 509 MPa to 605 MPa, the strain range also increases