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Purpose: The purpose was to evaluate if there are differences between tears of contact lens (CL) wearers of different materials detectable by measuring the hydrodynamic diameter of tear components through photon correlation spectroscopy (PCS).Methods: Tears of 59 CL wearers and tears of 39 non-wearers were collected by glass capillary. Wearers were divided into groups depending on the CL material: (i) hydrogels of II FDA group (H-II, 15 subjects), (ii) hydrogels of IV FDA group (H-IV, 13 subjects), (iii) silicone hydrogels (SH, 31 subjects). PCS analyses were performed at 25 °C on samples diluted with deionized water with tear concentration (10 ± 2)% V/W to obtain, for each subject, the average hydrodynamic diameter (dH,avg) of tear components by analyzing intensity fluctuations in time of scattered light.Results: Means of dH,avg calculated on each group were found, on increasing order, to be 256 nm (std dev 18 nm) for non-wearers, 297 nm (std dev 45 nm) for H-II, 360 nm (std dev 76 nm) for SH, and 391 nm (std dev 85 nm) for H-IV with statistical differences between each group of wearers compared to non-wearers and between groups of wearers except between SH and H-IV.Conclusions: PCS reveals the differences between tears of CL wearers of different materials, not only between tears of wearers and non-wearers. [ABSTRACT FROM AUTHOR]

By exposing Igrov-1 human ovarian cancer cells to increasing concentrations of Ecteinascidin-743 (ET-743), either for a short or prolonged time, we obtained sublines resistant to ET-743 which overexpress Pgp. The most resistant clone (Igrov-1/25 ET) was evaluated for biological and pharmacological characterizations. The increased Pgp levels of Igrov-1/25 ET were not due to amplification of the mdr-1 gene but to increased mRNA levels. No increase in other multidrug resistance-related proteins such as MRP or LRP was observed in Igrov-1/25 ET. The IC50 values of ET-743 against Igrov-1/25 ET was approximately 50 times higher than the parental cell line. Resistance was not reversed while maintaining the cell line in drug-free medium for at least 24 months. Igrov-1/25 ET was cross-resistant to Doxorubicin and VP16 while it was equally sensitive to L-PAM, MNNG, CPT and only marginally less sensitive to Cis-DDP and Oxaliplatin compared to the parental cell line. Igrov-1/25 ET exposed to Doxorubicin retained this drug much less, mainly because of a more efficient drug efflux. The cyclosporine analogue SDZ PSC-833 reversed the resistance of Igrov-1/25 ET to ET-743, without any enhancement of the drug activity against the parental Igrov-1 cell line. Igrov-1/25 ET exhibits typical features of cell lines overexpressing the mdr-1 gene and can be a potentially useful tool in selecting ET-743 non-cross-resistant analogues as well as to investigate methods to counteract resistance to this drug. [ABSTRACT FROM AUTHOR]

Thiocoraline, a new anticancer agent derived from the marine actinomycete Micromonospora marina, was found to induce profound perturbations of the cell cycle. On both LoVo and SW620 human colon cancer cell lines, thiocoraline caused an arrest in G1 phase of the cell cycle and a decrease in the rate of S phase progression towards G2/M phases, as assessed by using bromodeoxyuridine/DNA biparametric flow cytometric analysis. Thiocoraline does not inhibit DNA-topoisomerase II enzymes in vitro, nor does it induce DNA breakage in cells exposed to effective drug concentrations. The cell cycle effects observed after exposure to thiocoraline appear related to the inhibition of DNA replication. By using a primer extension assay it was found that thiocoraline inhibited DNA elongation by DNA polymerase a at concentrations that inhibited cell cycle progression and clonogenicity. These studies indicate that the new anticancer drug thiocoraline probably acts by inhibiting DNA polymerasea activity. [ABSTRACT FROM AUTHOR]

Summary Isohomohalichondrin B (IHB), a novel marine compound with anti-tumoral activity, extracted from the Lissodendorix sponge, inhibits GTP binding to tubulin, preventing microtubule assembly. Cell cycle perturbations and apoptosis induced by IHB were investigated on selected human cancer cell lines by using flow cytometric and biochemical techniques. Monoparameter flow cytometric analysis showed that 1 (h IHB exposure caused a delayed progression through S-phase, a dramatic block in G)2(M phase of the cell cycle and the appearance of) tetraploid cell population in LoVo, LoVo/DX, MOLT-4 and K562 cells. At 24 (h after IHB exposure, the majority of cells blocked in G)2(M were in) prophase as assessed by morphological analysis and by the fact that they expressed high levels of cyclin A/cdc2 and cyclin B1/cdc2. At 48 h, all cells were tetraploid as assessed by biparameter cyclin A/DNA and cyclin B1/DNA content analysis. Apoptotic death was detected in both leukaemic MOLT-4 and K562 cells, which express wild-type and mutated p53 respectively, when the cells were blocked in mitotic prophase. In conclusion, IHB is a novel potent anti-tumour drug that causes delayed S-phase progression, mitotic block, tetraploidy and apoptosis in cancer cell lines. [ABSTRACT FROM AUTHOR]

Objective: To investigate methodological aspects in body fat (BF) measurements in 7-to-10-year-old children. Study design: Systematic review of the literature. Methods: The studies were chosen from the PubMed and Scielo databases according to a protocol that defined: inclusion criteria; a search and quality-assessment strategy; and information extraction. Results: 27 studies published from 2004 to 2014 were included. The literature describes skin-fold measurements and dual energy X-ray absorptiometry (DEXA) as being the reference methods most widely used in the assessment of the ability of methods to identify BF. The most commonly-used statistical analyses were the Pearson correlation coefficient, and sensitivity and specificity performance analyses. The comparison between the tested methods and the references showed that body mass index (BMI) and waist circumference (WC) are strongly correlated to BF as calculated by bioelectrical impedance or skinfolds, and that there is a moderate positive correlation with percent body fat as calculated by DEXA, air-displacement plethysmography (ADP) or isotope dilution. There was a moderate positive correlation between weight-to-height ratio (WtHR) and BF, as estimated by ADP and skinfolds. Performance studies suggest that BMI and WC are very specific but less sensitive methods. Conclusions: The results of this systematic review show favourable evidence for the use of anthropometric indicators -- above all BMI and WC-in the measurement of BF, when more accurate techniques such as DEXA and ADP are not feasible. They also demonstrate features that make them advantageous for epidemiological studies in a child population, since they are easy and safe to obtain and well tolerated by the children. [ABSTRACT FROM AUTHOR]

Summary: Background: Gap‐junctional intercellular communication is crucial for epidermal cellular homeostasis. Inability to establish melanocyte–keratinocyte contact and loss of the intercellular junction's integrity may contribute to melanoma development. Connexins, laminins and desmocollins have been implicated in the control of melanoma growth, where their reduced expression has been reported in metastatic lesions. Objectives: The aim of this study was to investigate connexin 31·1 (GJB5) expression and identify any association with BRAF mutational status, prognosis of patients with melanoma and mitogen‐activated protein kinase (MAPK) inhibitor (MAPKi) treatment. Methods: GJB5 expression was measured at RNA and protein level in melanoma clinical samples and established cell lines treated (or not) with BRAF and MEK inhibitors (MEKi), as well as in cell lines which developed MAPKi resistance. Findings were further validated and confirmed by analysis of independent datasets. Results: Our analysis reveals significant downregulation of GJB5 expression in metastatic melanoma lesions compared with primary ones and in BRAF‐mutated vs. BRAF‐wildtype (BRAFWT) melanomas. Likewise, GJB5 expression is significantly lower in BRAFV600E compared with BRAFWT cell lines and increases on MAPKi treatment. MAPKi‐resistant melanoma cells display a similar expression pattern compared with BRAFWT cells, with increased GJB5 expression associated with morphological changes. Enhancement of BRAFV600E expression in BRAFWT melanoma cells significantly upregulates miR‐335‐5p expression with consequent downregulation of GJB5, one of its targets. Furthermore, overexpression of miR‐335‐5p in two BRAFWT cell lines confirms specific GJB5 protein downregulation. Reverse transcriptase quantitative polymerase chain reaction analysis also revealed upregulation of miR‐335 in BRAFV600E melanoma cells, which is significantly downregulated in cells resistant to MEKi. Our data were further validated using the TCGA_SKCM dataset, where BRAF mutations associate with increased miR‐335 expression and inversely correlate with GJB5 expression. In clinical samples, GJB5 underexpression is also associated with patient overall worse survival, especially at early stages. Conclusions: We identified a significant association between metastases/BRAF mutation and low GJB5 expression in melanoma. Our results identify a novel mechanism of gap‐junctional protein regulation, suggesting a prognostic role for GJB5 in cutaneous melanoma. Whatis already known about this topic? GJB5 expression has never been studied in melanoma.Although there is very limited knowledge about connexins in melanoma, these types of gap‐junction proteins have recently been linked with late stages of tumorigenesis and metastasis and are considered as tumour suppressors. Whatdoes this study add? This study establishes a significant association between BRAFV600E, metastases and GJB5 downregulation in melanoma.GJB5 underexpression also correlates with worse patient overall survival. Therefore, the data support a prognostic role for GJB5 in cutaneous melanoma. Whatis the translational message? This study highlights the importance of monitoring the integrity of connexin and junctional proteins during melanomagenesis, providing novel therapeutic target options for melanoma treatment, as well as a novel prognostic biomarker to predict melanoma progression. Linked Comment: J.E. Fromme and P. Zigrino. Br J Dermatol 2022; 186:13–14. Plain language summary available online [ABSTRACT FROM AUTHOR]

RAS mutations occur frequently in human cancer and activated RAS signalling contributes to tumour development and progression. Apart from its oncogenic effects on cell growth, active RAS has tumour-suppressive functions via its ability to induce cellular senescence and apoptosis. RAS is known to induce p53-dependent cell cycle arrest, yet its effect on p53-dependent apoptosis remains unclear. We report here that apoptosis-stimulating protein of p53 (ASPP) 1 and 2, two activators of p53, preferentially bind active RAS via their N-terminal RAS-association domains (RAD). Additionally, ASPP2 colocalises with and contributes to RAS cellular membrane localisation and potentiates RAS signalling. In cancer cells, ASPP1 and ASPP2 cooperate with oncogenic RAS to enhance the transcription and apoptotic function of p53. Thus, loss of ASPP1 and ASPP2 in human cancer cells may contribute to the full transforming property of RAS oncogene. [ABSTRACT FROM AUTHOR]

Background:Novel prognostic biomarkers and therapeutic strategies are urgently required for malignant melanoma. Ecto-5-prime-nucleotidase (NT5E; CD73) overexpression has been reported in several human cancers. The mechanism(s) underlying deregulated expression and the clinical consequences of changes in expression are not known.Methods:We used RT-PCR, qPCR, methylation-specific PCR and pyrosequencing to analyse expression and regulation of NT5E in malignant melanoma cell lines and primary and metastatic melanomas.Results:NT5E is subject to epigenetic regulation in melanoma. NT5E mRNA is downregulated by methylation-dependent transcriptional silencing in the melanoma cell lines SKMel2, SKMel23, WM35, Mel501, Mel505 and C81-61 and expression is reactivated by azacytidine. In contrast, the CpG island is unmethylated and the gene expressed in cultured normal melanocytes. In clinical cases of melanoma, methylation in the NT5E CpG island occurs in both primary and metastatic melanomas and correlates with transcriptional downregulation of NT5E mRNA. Relapse with metastatic disease, particularly to the visceral sites and brain, is more common in primary melanomas lacking NT5E methylation. Primary melanomas with methylation in NT5E show limited metastatic potential or more commonly metastasise predominantly to nodal sites rather than viscera and brain (P=0.01).Conclusion:Deregulation of NT5E expression in melanoma occurs via epigenetic changes in the NT5E CpG island. Confirmation of our results in larger clinical series would support the candidacy of NT5E as a clinical biomarker in melanoma, which could be applied in both primary and relapsed disease. Inhibition of NT5E may have therapeutic potential in melanoma, particularly in patients with more aggressive disease metastatic to viscera or the brain. [ABSTRACT FROM AUTHOR]