Resolving the role of podoplanin in the motility of papillary thyroid carcinoma-derived cells using RNA sequencing.

The intracellular level of podoplanin (PDPN), a transmembrane protein of still unclear function, is frequently altered in metastatic tumors. High expression of PDPN is frequently observed in papillary thyroid cancer (PTC) specimens. Similarly, PTC-derived cell lines (BCPAP and TPC1, harboring the BRAF V600E mutation and RET/PTC1 fusion, respectively), also present enhanced PDPN yield. We previously reported that depletion of PDPN impairs migration of TPC1 cells, but augments metastasis of BCPAP cells. Interestingly, this phenomenon stays in contrast to the migratory pattern observed for wild-type cells, where TPC1 exhibited higher motility than BCPAP cells. Here, we aimed to elucidate the potential role of PDPN in regulation of molecular mechanisms leading to the diverse metastatic features of the studied PTC-derived cells. We consider that this phenomenon may be caused by alternative regulation of signaling pathways due to the presence of the mutated BRAF allele or RET/PTC1 fusion. The high-throughput RNA sequencing (RNA-seq) technique was used to uncover the genes and signaling pathways affected in wild-type and PDPN-depleted TPC1 and BCPAP cells. We found that changes in the expression of various factors of signaling pathways, like RHOA and RAC1 GTPases and their regulators, are linked with both high PDPN levels and presence of the BRAF V600E mutation. We imply that the suppressed motility of wild-type BCPAP cells results from overactivation of RHOA through natively high PDPN expression. This process is accompanied by inhibition of the PI3K kinase and consequently RAC1, due to overactivation of RAS-mediated signaling and the PTEN regulator.

Simvastatin Coadministration Modulates the Electrostatically Driven Incorporation of Doxorubicin into Model Lipid and Cell Membranes.

Understanding the interactions between drugs and lipid membranes is a prerequisite for finding the optimal way to deliver drugs into cells. Coadministration of statins and anticancer agents has been reported to have a positive effect on anticancer therapy. In this study, we elucidate the mechanism by which simvastatin (SIM) improves the efficiency of biological membrane penetration by the chemotherapeutic agent doxorubicin (DOX) in neutral and slightly acidic solutions. The incorporation of DOX, SIM, or a combination of them (DOX:SIM) into selected single-component lipid membranes, zwitterionic unsaturated 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), neutral cholesterol, and negatively charged 1,2-dimyristoyl-sn-glycero-3-phospho-l-serine (DMPS) was assessed using the Langmuir method. The penetration of neutral lipid monolayers by the codelivery of SIM and DOX was clearly facilitated at pH 5.5, which resembles the pH conditions of the environment of cancer cells. This effect was ascribed to partial neutralization of the DOX positive charge as the result of intermolecular interactions between DOX and SIM. On the other hand, the penetration of the negatively charged DMPS monolayer was most efficient in the case of the positively charged DOX. The efficiency of the drug delivery to the cell membranes was evaluated under in vitro conditions using a panel of cancer-derived cell lines (A172, T98G, and HeLa). MTS and trypan blue exclusion assays were performed, followed by confocal microscopy and spheroid culture tests. Cells were exposed to either free drugs or drugs encapsulated in lipid carriers termed cubosomes. We demonstrated that the viability of cancer cells exposed to DOX was significantly impaired in the presence of SIM, and this phenomenon was greatly magnified when DOX and SIM were coencapsulated in cubosomes. Overall, our results confirmed the utility of the DOX:SIM combination delivery, which enhances the interactions between neutral components of cell membranes and positively charged chemotherapeutic agents.

Molecular Factors and Mechanisms Driving Multidrug Resistance in Uropathogenic Escherichia coli-An Update.

The rapid emergence of multidrug-resistant (MDR) bacteria indisputably constitutes a major global health problem. Pathogenic Escherichia coli are listed among the most critical group of bacteria that require fast development of new antibiotics and innovative treatment strategies. Among harmful extraintestinal Enterobacteriaceae strains, uropathogenic E. coli (UPEC) pose a significant health threat. UPEC are considered the major causative factor of urinary tract infection (UTI), the second-most commonly diagnosed infectious disease in humans worldwide. UTI treatment places a substantial financial burden on healthcare systems. Most importantly, the misuse of antibiotics during treatment has caused selection of strains with the ability to acquire MDR via miscellaneous mechanisms resulting in gaining resistance against many commonly prescribed antibiotics like ampicillin, gentamicin, cotrimoxazole and quinolones. Mobile genetic elements (MGEs) such as transposons, integrons and conjugative plasmids are the major drivers in spreading resistance genes in UPEC. The co-occurrence of various bacterial evasion strategies involving MGEs and the SOS stress response system requires further research and can potentially lead to the discovery of new, much-awaited therapeutic targets. Here, we analyzed and summarized recent discoveries regarding the role, mechanisms, and perspectives of MDR in the pathogenicity of UPEC.

Diagnostic and Clinical Value of Specific Autoantibodies against Kelch-like 12 Peptide and Nuclear Envelope Proteins in Patients with Primary Biliary Cholangitis.

Primary biliary cholangitis (PBC) is a chronic autoimmune liver disease characterized by the presence of antimitochondrial and antinuclear antibodies in patients' serum. Here, we analyzed the reactivity of autoantibodies against a novel autoantigen, kelch-like 12 (KLHL12) protein, in a cohort of 138 PBC and 90 non-PBC patients. Additionally, we compared the reactivity of KLHL12 with antinuclear envelope antibodies: anti-gp210, anti-p62, and anti-LBR. Commercially available kits and an 'in-house' ELISA were used in the studies. Antinuclear envelope antibodies were detected in 65% of PBC patients and the presence of these antibodies was observed more frequently in patients diagnosed with later stages (III/IV) of PBC, according to Ludwig's classification (p < 0.05) and were found to correlate with a higher concentration of bilirubin. Overall, anti-KLHL12 antibodies were found more frequently in PBC patients than in non-PBC controls (p < 0.001). Anti-KLHL12 antibodies were detected in 36% of the tested PBC cohort, including PBC patients negative for antimitochondrial antibodies. Presence of anti-KLHL12 was also associated with a higher concentration of bilirubin and correlated with fibrosis (p < 0.05). Anti-KLHL12 antibodies were detected in 30% of PBC individuals positive for antinuclear envelope antibodies, while anti-KLHL12 and antinuclear envelope antibodies were found in 17% of all PBC cases. Concluding, our data confirm that antibodies against the KLHL12 protein are highly specific for PBC and when used in combination with other markers, may significantly increase the diagnosis of PBC.

Novel Inhibitor-Based Therapies for Thyroid Cancer-An Update.

Thyroid cancers (TCs) are the most common tumors of the endocrine system and a constant rise in the number of TC cases has been observed for the past few decades. TCs are one of the most frequent tumors in younger adults, especially in women, therefore early diagnosis and effective therapy are especially important. Ultrasonography examination followed by fine needle biopsy have become the gold standard for diagnosis of TCs, as these strategies allow for early-stage detection and aid accurate qualification for further procedures, including surgical treatment. Despite all the advancements in detection and treatment of TCs, constant mortality levels are still observed. Therefore, a novel generation line of targeted treatment strategies is being developed, including personalized therapies with kinase inhibitors. Recent molecular studies on TCs demonstrate that kinase inhibitor-based therapies might be considered as the most promising. In the past decade, new kinase inhibitors with different mechanisms of action have been reported and approved for clinical trials. This review presents an up-to-date picture of new approaches and challenges of inhibitor-based therapies in treatment of TCs, focusing on the latest findings reported over the past two years.

Au@Pt Core-Shell Nanoparticle Bioconjugates for the Therapy of HER2+ Breast Cancer and Hepatocellular Carcinoma. Model Studies on the Applicability of 193mPt and 195mPt Radionuclides in Auger Electron Therapy.

193mPt and 195mPt radionuclides are therapeutically attractive Auger electron emitters with notably high Auger electron yield per decay. The present paper summarizes the first step of research on the applications of core-shell (Au@Pt) nanoparticles for electron Auger therapy of HER2+ (human epidermal growth factor receptor 2) breast cancer and hepatocellular carcinoma. Gold nanoparticles (30 nm) were synthesized covered with a platinum shell at high efficiency (>80%) and were further evaluated for in vitro studies such as binding affinity, internalization and cytotoxicity. To find the mechanism(s) responsible for platinum cytotoxicity in HepG2 cells, the platinum concentration in isolated cell nuclei and cytoplasm was determined using ICP-MS (inductively coupled plasma mass spectrometry). Lack of platinum in cell nuclei suggests that the cytotoxic effect is associated with the generation of reactive oxygen species (ROS) and reactive nitrogen species (RNS). Studies carried out on the SKOV-3 cell line with the use of a synthesized targeting bioconjugate (Au@Pt-PEG-trastuzumab) revealed a high affinity of this preparation to HER2+ cells, its internalization, its placement in the perinuclear area and partial intranuclear location. The specific binding for HER2 negative cells, MDA-MB-231, was negligible and Au@Pt-PEG-trastuzumab did not enter these cells. The results obtained are promising and warrant future investigation of Auger electron therapy using 193mPt and 195mPt based radiopharmaceuticals.

Analysis of Autoantibodies against Promyelocytic Leukemia Nuclear Body Components and Biochemical Parameters in Sera of Patients with Primary Biliary Cholangitis.

Primary biliary cholangitis (PBC) is a chronic autoimmune liver disease characterized by immune-mediated destruction of intrahepatic bile ducts and the presence of specific antibodies. The aim of the study was to examine the diagnostic significance of antibodies against promyelocytic leukemia nuclear body (PML NB) components such as Sp100, Sp140, and PML in a cohort of PBC patients and compare the results with biochemical and histological parameters. Serum samples were collected from 93 PBC patients. Anti-Sp100 and anti-PML antibodies were assessed using commercially available kits, anti-Sp140 using developed "in-house" ELISA test. Anti-Sp140, anti-Sp100, and anti-PML antibodies were present in 25 (27%), 37 (40%), and 29 (31%) PBC patients, respectively. Anti-PML NB positive patients also showed increased concentration of bilirubin and alkaline phosphatase (p < 0.05). In the group with the presence of at least two types of these antibodies, more frequent deaths or transplantations were observed. A correlation between the presence of antibodies and histological grade (OR = 2.55 p = 0.039) was established. Patients with bilirubin > 1.1 mg/dL at the time of diagnosis had a significantly shorter time of survival than patients with bilirubin ≤ 1.1 mg/dL (HR 5.7; 95% C.I., 2.7, 12.3; p < 0.001). Our data confirm very high specificity of anti-PML NB antibodies, which can expand the laboratory diagnostic capabilities of PBC. We found an association between positive reactivity of autoantibodies directed against components of PML nuclear bodies and higher concentrations of bilirubin and alkaline phosphatase, but the main prognostic marker of survival remains serum bilirubin.

The Role of miRNA-7 in the Biology of Cancer and Modulation of Drug Resistance.

MicroRNAs (miRNAs, miRs) are small non-coding RNA (ncRNA) molecules capable of regulating post-transcriptional gene expression. Imbalances in the miRNA network have been associated with the development of many pathological conditions and diseases, including cancer. Recently, miRNAs have also been linked to the phenomenon of multidrug resistance (MDR). MiR-7 is one of the extensively studied miRNAs and its role in cancer progression and MDR modulation has been highlighted. MiR-7 is engaged in multiple cellular pathways and acts as a tumor suppressor in the majority of human neoplasia. Its depletion limits the effectiveness of anti-cancer therapies, while its restoration sensitizes cells to the administered drugs. Therefore, miR-7 might be considered as a potential adjuvant agent, which can increase the efficiency of standard chemotherapeutics.

Lipid membranes exposed to dispersions of phytantriol and monoolein cubosomes: Langmuir monolayer and HeLa cell membrane studies.

The interactions of liquid-crystalline nanoparticles based on lipid-like surfactants, glyceryl monooleate, monoolein (GMO) and 1,2,3-trihydroxy-3,7,11,15-tetramethylhexadecane, phytantriol (PT) with selected model lipid membranes prepared by Langmuir technique were compared. Monolayers of DPPC, DMPS and their mixture DPPC:DMPS 87:13 mol% were used as simple models of one leaflet of a cell membrane. The incorporation of cubosomes into the lipid layers spread at the air-water interface was followed by surface-pressure measurements and Brewster angle microscopy. The cubosome - membrane interactions lead to the fluidization of the model membranes but this effect depended on the composition of the model membrane and on the type of cubosomes. The interactions of PT cubosomes with lipid layers, especially DMPS-based monolayer were stronger compared with those of GMO-based nanoparticles. The kinetics of incorporation of cubosomal material into the lipid layer was influenced by the extent of hydration of the polar headgroups of the lipid: faster in the case of smaller, less hydrated polar groups of DMPS than for strongly hydrated uncharged choline of DPPC. The membrane disrupting effect of cubosomes increased at longer times of the lipid membrane exposure to the cubosome solution and at larger carrier concentrations. Langmuir monolayer observations correspond well to results of studies of HeLa cells exposed to cubosomes. The larger toxicity of PT cubosomes was confirmed by MTS. Their ability to disrupt lipid membranes was imaged by confocal microscopy. On the other hand, PT cubosomes easily penetrated cellular membranes and released cargo into various cellular compartments more effectively than GMO-based nanocarriers. Therefore, at low concentrations, they may be further investigated as a promising drug delivery tool.

Trastuzumab Modified Barium Ferrite Magnetic Nanoparticles Labeled with Radium-223: A New Potential Radiobioconjugate for Alpha Radioimmunotherapy.

Barium ferrite nanoparticles (BaFeNPs) were investigated as vehicles for 223Ra radionuclide in targeted α-therapy. BaFe nanoparticles were labeled using a hydrothermal Ba2+ cations replacement by 223Ra with yield reaching 61.3 ± 1.8%. Radiolabeled nanoparticles were functionalized with 3-phosphonopropionic acid (CEPA) linker followed by covalent conjugation to trastuzumab (Herceptin®). Thermogravimetric analysis and radiometric method with the use of [131I]-labeled trastuzumab revealed that on average 19-21 molecules of trastuzumab are attached to the surface of one BaFe-CEPA nanoparticle. The hydrodynamic diameter of BaFe-CEPA-trastuzumab conjugate is 99.9 ± 3.0 nm in water and increases to 218.3 ± 3.7 nm in PBS buffer, and the zeta potential varies from +27.2 ± 0.7 mV in water to -8.8 ± 0.7 in PBS buffer. The [223Ra]BaFe-CEPA-trastuzumab radiobioconjugate almost quantitatively retained 223Ra (>98%) and about 96% of 211Bi and 94% of 211Pb over 30 days. The obtained radiobioconjugate exhibited high affinity, cell internalization and cytotoxicity towards the human ovarian adenocarcinoma SKOV-3 cells overexpressing HER2 receptor. Confocal studies indicated that [223Ra]BaFe-CEPA-trastuzumab was located in peri-nuclear space. High cytotoxicity of the [223Ra]BaFe-CEPA-trastuzumab bioconjugate was confirmed by radiotoxicity studies on SKOV-3 cell monolayers and 3D-spheroids. In addition, the magnetic properties of the radiobioconjugate should allow for its use in guide drug delivery driven by magnetic field gradient.

Combinatory Treatment with miR-7-5p and Drug-Loaded Cubosomes Effectively Impairs Cancer Cells.

BACKGROUND: Multidrug resistance (MDR) is an emerging problem in the treatment of cancer. Therefore, there is a necessity for novel strategies that would sensitize tumor cells to the administered chemotherapeutics. One of the innovative approaches in fighting drug-resistant tumors is the treatment of cancer with microRNA (miRNA), or the use of cubosomes (lipid nanoparticles) loaded with drugs. Here, we present a study on a novel approach, which combines both tools. METHODS: Cubosomes loaded with miR-7-5p and chemotherapeutics were developed. The effects of drug- and miRNA-loaded vehicles on glioma- (A172, T98G), papillary thyroid- (TPC-1) and cervical carcinoma-derived (HeLa) cells were analyzed using molecular biology techniques, including quantitative real-time PCR, MTS-based cell proliferation test, flow cytometry and spheroids formation assay. RESULTS: The obtained data indicate that miR-7-5p increases the sensitivity of the tested cells to the drug, and that nanoparticles loaded with both miRNA and the drug produce a greater anti-tumor effect in comparison to the free drug treatment. It was found that an increased level of apoptosis in the drug/miRNA co-treated cells is accompanied by an alternation in the expression of the genes encoding for key MDR proteins of the ABC family. CONCLUSIONS: Overall, co-administration of miR-7-5p with a chemotherapeutic can be considered a promising strategy, leading to reduced MDR and the induction of apoptosis in cancer cells.

Thyroid Peroxidase Revisited - What's New?

Thyroid peroxidase (TPO) is an enzyme that participates in thyroid hormone biosynthesis. TPO is also a major autoantigen in autoimmune thyroid diseases (AITD). In this review, we summarize the latest developments in the field of TPO research. We present the current understanding of immunodominant serologic determinants, frequency of TPO-specific autoantibodies in the population, as well as genetic and environmental factors contributing to their development. Moreover, we report recent progress in the clinical utilities of TPO autoantibody testing, including thyroid dysfunctions and extra-thyroidal disorders.

Replication fidelity in E. coli: Differential leading and lagging strand effects for dnaE antimutator alleles.

DNA Pol III holoenzyme (HE) is the major DNA replicase of Escherichia coli. It is a highly accurate enzyme responsible for simultaneously replicating the leading- and lagging DNA strands. Interestingly, the fidelity of replication for the two DNA strands is unequal, with a higher accuracy for lagging-strand replication. We have previously proposed this higher lagging-strand fidelity results from the more dissociative character of the lagging-strand polymerase. In support of this hypothesis, an E. coli mutant carrying a catalytic DNA polymerase subunit (DnaE915) characterized by decreased processivity yielded an antimutator phenotype (higher fidelity). The present work was undertaken to gain deeper insight into the factors that influence the fidelity of chromosomal DNA replication in E. coli. We used three different dnaE alleles (dnaE915, dnaE911, and dnaE941) that had previously been isolated as antimutators. We confirmed that each of the three dnaE alleles produced significant antimutator effects, but in addition showed that these antimutator effects proved largest for the normally less accurate leading strand. Additionally, in the presence of error-prone DNA polymerases, each of the three dnaE antimutator strains turned into mutators. The combined observations are fully supportive of our model in which the dissociative character of the DNA polymerase is an important determinant of in vivo replication fidelity. In this model, increased dissociation from terminal mismatches (i.e., potential mutations) leads to removal of the mismatches (antimutator effect), but in the presence of error-prone (or translesion) DNA polymerases the abandoned terminal mismatches become targets for error-prone extension (mutator effect). We also propose that these dnaE alleles are promising tools for studying polymerase exchanges at the replication fork.

Selected CD molecules and the phagocytosis of microvesicles released from erythrocytes ex vivo.

BACKGROUND AND OBJECTIVES: The accumulation of microvesicles in erythrocyte concentrates during storage or irradiation may be responsible for clinical symptoms such as inflammation, coagulation and immunization. Our aim was to determine whether any of the cluster of differentiation (CD) molecules responsible for important functions are present on microvesicles, and if their expression level is dependent on the storage period of erythrocyte concentrates. MATERIAL AND METHODS: Erythrocyte microvesicles were isolated from 'fresh' (2nd day) and 'old' (42nd day) stored erythrocyte concentrates. Qualitative cytometric analysis of 0·5 µm, erythrocyte-derived, PS-exposing vesicles was performed using the annexin V-FITC, anti-CD235a-PE antibody and calibrated beads. The microvesicles were also visualized under a confocal microscope. The expression of the molecules CD235a, CD44, CD47, CD55, CD59 and of phosphatidylserine (PS) was compared using flow cytometry. Measurements of microvesicle phagocytosis by human monocytes were carried out using a flow cytometer and a confocal microscope. RESULTS: The analysis of the microvesicles with calibration beads allowed us to identify these structures with a diameter of about 0·5 µm in the 'fresh' and 'old' samples. At day 2, the microvesicles had elevated expression levels of CD47, reduced expression levels of PS, CD55 and CD59. The phagocytosis index was higher for the microvesicles isolated from the 42-day-old erythrocyte concentrates. CONCLUSION: This research may bring us closer to understanding the factors responsible for erythrocyte ageing and to evaluate the quality of stored red blood concentrates intended for transfusion.

Trastuzumab-Modified Gold Nanoparticles Labeled with 211At as a Prospective Tool for Local Treatment of HER2-Positive Breast Cancer.

Highly localized radiotherapy with radionuclides is a commonly used treatment modality for patients with unresectable solid tumors. Herein, we propose a novel α-nanobrachytherapy approach for selective therapy of human epidermal growth factor receptor 2 (HER2)-positive breast cancer. This uses local intratumoral injection of 5-nm-diameter gold nanoparticles (AuNPs) labeled with an α-emitter (211At), modified with polyethylene glycol (PEG) chains and attached to HER2-specific monoclonal antibody (trastuzumab). The size, shape, morphology, and zeta potential of the 5 nm synthesized AuNPs were characterized by TEM (Transmission Electron Microscopy) and DLS (Dynamic Light Scattering) techniques. The gold nanoparticle surface was modified by PEG and subsequently used for antibody immobilization. Utilizing the high affinity of gold for heavy halogens, the bioconjugate was labelled with 211At obtained by α irradiation of the bismuth target. The labeling yield of 211At was greater than 99%. 211At bioconjugates were stable in human serum. Additionally, in vitro biological studies indicated that 211At-AuNP-PEG-trastuzumab exhibited higher affinity and cytotoxicity towards the HER2-overexpressing human ovarian SKOV-3 cell line than unmodified nanoparticles. Confocal and dark field microscopy studies revealed that 211At-AuNP-PEG-trastuzumab was effectively internalized and deposited near the nucleus. These findings show promising potential for the 211At-AuNP-PEG-trastuzumab radiobioconjugate as a perspective therapeutic agent in the treatment of unresectable solid cancers expressing HER2 receptors.

MIX2: A Novel Natural Multi-Component Modulator of Multidrug-Resistance and Hallmarks of Cancer Cells.

Multidrug resistance is one of the key barriers suppressing the effectiveness of drug therapies of malignant tumors. Here, we report a study on the effect of a mix of natural extracts (MIX2) prepared from fresh fruits of Prunus spinosa, Crataegus monogyna, Sorbus aucuparia, and Euonymus europaeus on the classic hallmarks of cancer cells and the expression of multidrug resistance proteins. In the studies, HeLa and T98G cell lines, and classic methods of molecular biology, including RT-qPCR, Western blot, flow cytometry, and confocal imaging, were used. Additionally, migration, adhesion, and proliferation assays were performed. The obtained results indicate that the MIX2 cocktail presents strong anti-cancer properties. MIX2 is not toxic, but at the same time significantly alters the migration, proliferation, and adhesion of tumor cells. Furthermore, it was found that cells exposed to the mixture presented a significantly reduced expression level of genes associated with MDR, including ABCB1, which encodes for glycoprotein P. In vitro data showed that MIX2 effectively sensitizes tumor cells to doxorubicin. We postulate that modulation of the multidrug resistance phenotype of tumors with the use of MIX2 may be considered as a safe and applicable tool in sustaining drug delivery therapies of malignancies.

Podoplanin (PDPN) affects the invasiveness of thyroid carcinoma cells by inducing ezrin, radixin and moesin (E/R/M) phosphorylation in association with matrix metalloproteinases.

BACKGROUND: Podoplanin (PDPN) is a mucin-type transmembrane glycoprotein specific to the lymphatic system. PDPN expression has been found in various human tumors and is considered to be a marker of cancer. We had previously shown that PDPN expression contributes to carcinogenesis in the TPC1 papillary thyroid cancer-derived cell line by enhancing cell migration and invasiveness. The aim of this study was to determine the effect of PDPN down-regulation in another thyroid cancer-derived cell line: BcPAP. METHODS: In order to determine the effects of PDPN on malignant features of BcPAP cells (harboring the BRAFV600E mutated allele) and TPC1 cells (carrying the RET/PTC1 rearrangement), we silenced PDPN in these cells using small interfering RNA (siRNA). The efficacy of PDPN silencing was confirmed by qRT-PCR and Western blotting. Then, we tested the motility and invasiveness of these cells (using scratch test and Transwell assay), their growth capacities F(cell cycle analysis, viability, clonogenic activity) and apoptosis assays), adhesion-independent colony-formation capacities, as well as the effect of PDPN silencing on MMPs expression and activity (zymography). RESULTS: We found that PDPN-induced cell phenotype depended on the genetic background of thyroid tumor cells. PDPN down-regulation in BcPAP cells was negatively correlated with the migration and invasion, in contrast to TPC1 cells in which PDPN depletion resulted in enhanced migration and invasiveness. Moreover, our results suggest that in BcPAP cells, PDPN may be involved in the epithelial-mesenchymal transition (EMT) through regulating the expression of the ezrin, radixin and moesin (E/R/M) proteins, MMPs 9 and MMP2, remodeling of actin cytoskeleton and cellular protrusions. We also demonstrated that PDPN expression is associated with the MAPK signaling pathway. The inhibition of the MAPK pathway resulted in a decreased PDPN expression, increased E/R/M phosphorylation and reduced cell migration. Additionally, PDPN depleted BcPAP cells treated with inhibitors of MEK1/2 kinases (U0126) or of the BRAF V600E protein (PLX4720) had reduced motility, similar to that previously observed in TPC1 cells after PDPN knock-down. CONCLUSIONS: Altogether, our data suggest that PDPN may play an important role in the control of invasion and migration of papillary thyroid carcinoma cells in association with the E/R/M, MMPs and MAPK kinases.

Abnormal spontaneous activity in primary myopathic disorders.

INTRODUCTION: Reproducible non-insertional spontaneous activity (SA), with the exception of endplate activity, is an unequivocal sign of abnormality and is one of the most useful findings obtained on electromyography. METHODS: In this retrospective study we analyzed occurrence and distribution of abnormal SA in 151 patients with genetically confirmed myopathies. RESULTS: Complex repetitive discharges (CRDs) occurred more frequently than fibrillation potentials (fibs) and positive sharp waves (PSWs) in centronuclear myopathy (CNM) and limb-girdle muscular dystrophy type 2A (LGMD-2A), whereas fibs/PSWs were observed more often in desminopathy and facioscapulohumeral dystrophy (FSHD). Abnormal SA was commonly found in CNM (66.7%) and desminopathy (61.5%), occasionally in Duchenne (DMD) and Becker muscular dystrophy (BMD) (45.2% and 27.6%, respectively), but rarely in FSHD (14.9%) and LGMD-2A (12.0%). CONCLUSIONS: Abnormal SA probably occurs more frequently in disorders associated with structural changes in muscle fibers. Screening for SA may be a valuable tool for diagnosis of non-myotonic myopathies. Muscle Nerve 56: 427-432, 2017.

The role of prospero homeobox 1 (PROX1) expression in follicular thyroid carcinoma cells.

The prospero homeobox 1 (Prox1) transcription factor is a key player during embryogenesis and lymphangiogenesis. Altered Prox1 expression has been found in a variety of human cancers, including papillary thyroid carcinoma (PTC). Interestingly, Prox1 may exert tumor suppressive or tumor promoting effect, depending on the tissue context. In this study, we have analyzed Prox1 expression in normal and malignant human thyroid carcinoma cell lines. Moreover, we determined the effect of Prox1 silencing and overexpression on the cellular processes associated with the metastatic potential of tumor cells: proliferation, migration, invasion, apoptosis and anchorage-independent growth, in the follicular thyroid carcinoma (FTC) FTC-133 cell line. We found that Prox1 expression was significantly higher in FTC-derived cells than in PTC-derived cells and normal thyroid, and it was associated with the PI3K/Akt signaling pathway. In the FTC-133 cells, it was associated with cell invasive potential, motility and wound closure capacities, but not with proliferation or apoptosis. Modifying Prox1 expression also induced substantial changes in the cytoskeleton structure and cell morphology. In conclusion, we have shown that Prox1 plays an important role in the development of FTC and that its suppression prevents, whereas its overexpression promotes, the malignant behavior of thyroid follicular cancer cells.

Diagnostic value of blink reflex in multisystem atrophy, progressive supranuclear palsy and Parkinson disease.

UNLABELLED: Abnormal blink reflex (BR) is a result of reticular brainstem pathways dysfunction and seems to be one of the features of brain degenerative disorders. The aim of the study was to estimate the diagnostic value of blink reflex in neurodegenerative diseases such as: multisystem atrophy (MSA), progressive supranuclear palsy (PSP) and Parkinson disease (PD). Material consisted of 99 patients with clinically probable MSA (51), PSP (28) and PD (20). MSA patients were divided into two subgroups, with dominant cerebellar (MSA-C) and parkinsonian signs (MSA-P). The mean age of patients was 64.9 years (47-79 years); males - 55.3%. Blink reflex was obtained in a typical way. RESULTS: The significant differences in mean values of blink reflex latencies between PD and other subgroups (MSA-P, MSA-C, PSP) were found, but all of them were in normal range. In individual patients with PD and PSP (50% and 18%, respectively) delayed R2 latencies were recorded. CONCLUSIONS: The most frequently abnormal blink reflexes, comparing the MSA, PSP and PD groups, were present in PD patients. We postulate that this may be explained by pathological influence of nigrostriatal pathway on the circuit linking the basal ganglia, cerebellum and brainstem.