The Emergent Power of Human Cellular vs Mouse Models in Translational Hair Research

Different animal models have been used for hair research and regeneration studies based on the similarities between animal and human skins. Primary knowledge on hair follicle (HF) biology has arisen from research using mouse models baring spontaneous or genetically engineered mutations. These studies have been crucial for the discovery of genes underlying human hair cycle control and hair loss disorders. Yet, researchers have become increasingly aware that there are distinct architectural and cellular features between the mouse and human HFs, which might limit the translation of findings in the mouse models. Thus, it is enticing to reason that the spotlight on mouse models and the unwillingness to adapt to the human archetype have been hampering the emergence of the long-awaited human hair loss cure. Here, we provide an overview of the major limitations of the mainstream mouse models for human hair loss research, and we underpin a future course of action using human cell bioengineered models and the emergent artificial intelligence.This work was supported by FEDER (Fundo Europeu de Desenvolvimento Regional) funds through COMPETE 2020 (POCI, Programa Operacional Competividade e Internacionalização) and Portugal 2020 in the framework of the project 70201-SI I&DT EMPRESAS EM COPROMOÇÃO. E.L. was supported by CEECIND/00654/2020 grant from FCT—Fundação para a Ciência e a Tecnologia, I.P

Foxm1 modulates cell non-autonomous response in zebrafish skeletal muscle homeostasis

Foxm1 is a master regulator of the cell cycle, contributing to cell proliferation. Recent data have shown that this transcription factor also modulates gene networks associated with other cellular mechanisms, suggesting non-proliferative functions that remain largely unexplored. In this study, we used CRISPR/Cas9 to disrupt foxm1 in the zebrafish terminally differentiated fast-twitching muscle cells. foxm1 genomic disruption increased myofiber death and clearance. Interestingly, this contributed to non-autonomous satellite cell activation and proliferation. Moreover, we observed that Cas9 expression alone was strongly deleterious to muscle cells. Our report shows that foxm1 modulates a muscle non-autonomous response to myofiber death and highlights underreported toxicity to high expression of Cas9 in vivo.This study was supported by the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation program (ERC-2015-StG-680156-ZPR). FEDER (Fundo Europeu de Desenvolvimento Regional) funds through the COMPETE 2020—Operational Programme for Competitiveness and Internationalization (POCI), Portugal 2020 and by Portuguese funds through FCT (Fundação para a Ciência e a Tecnologia) in the framework of the project POCI-01-0145-FEDER-031120 (PTDC/BIA-CEL/31120/2017); and POCI-01-0145-FEDER-007274 i3S framework project co-funded by COMPETE 2020/PORTUGAL 2020 through FEDER. E.L. and J.B. are supported by FCT (J.B.: Grant CEECIND/03482/2018; E.L.: Grants CEECIND/00654/2020 and IF/00916/2014). F.J.F. (PD/BD/105745/2014) is a PhD fellow from FCT

Candidate alzheimer’s disease biomarker mir-483-5p lowers tau phosphorylation by direct erk1/2 repression

MicroRNAs have been demonstrated as key regulators of gene expression in the etiology of a range of diseases including Alzheimer’s disease (AD). Recently, we identified miR-483-5p as the most upregulated miRNA amongst a panel of miRNAs in blood plasma specific to prodromal, early-stage Alzheimer’s disease patients. Here, we investigated the functional role of miR-483-5p in AD pathology. Using TargetScan and miRTarBase, we identified the microtubule-associated protein MAPT, often referred to as TAU, and the extracellular signal-regulated kinases 1 and 2 (ERK1 and ERK2), known to phosphorylate TAU, as predicted direct targets of miR-483-5p. Employing several functional assays, we found that miR-483-5p regulates ERK1 and ERK2 at both mRNA and protein levels, resulting in lower levels of phosphorylated forms of both kinases. Moreover, miR-483-5p-mediated repression of ERK1/2 resulted in reduced phosphorylation of TAU protein at epitopes associated with TAU neurofibrillary pathology in AD. These results indicate that upregulation of miR-483-5p can decrease phosphorylation of TAU via ERK pathway, representing a compensatory neuroprotective mechanism in AD pathology. This miR-483-5p/ERK1/TAU axis thus represents a novel target for intervention in AD.This work has been supported by the Polish National Science Centre grant OPUS 2018/29 /B/NZ7/02757, by the EU Horizon 2020 FETOPEN grant, agreement no 737390 (ArrestAD), and by the funding from the Polish Ministry of Science and Higher Education within 2016-2020 funds for the implementation of international projects (agreement no 3548/H2020/COFUND/2016/2). A.F. research internship in Portugal was supported by Erasmus fellowship

FOXM1 repression increases mitotic death upon antimitotic chemotherapy through BMF upregulation

Inhibition of spindle microtubule (MT) dynamics has been effectively used in cancer treatment. Although the mechanisms by which MT poisons elicit mitotic arrest are fairly understood, efforts are still needed towards elucidating how cancer cells respond to antimitotic drugs owing to cytotoxicity and resistance side effects. Here, we identified the critical G2/M transcription factor Forkhead box M1 (FOXM1) as a molecular determinant of cell response to antimitotics. We found FOXM1 repression to increase death in mitosis (DiM) due to upregulation of the BCL-2 modifying factor (BMF) gene involved in anoikis, an apoptotic process induced upon cell detachment from the extracellular matrix. FOXM1 binds to a BMF intronic cis-regulatory element that interacts with both the BMF and the neighbor gene BUB1B promoter regions, to oppositely regulate their expression. This mechanism ensures that cells treated with antimitotics repress BMF and avoid DiM when FOXM1 levels are high. In addition, we show that this mechanism is partly disrupted in anoikis/antimitotics-resistant tumor cells, with resistance correlating with lower BMF expression but in a FOXM1-independent manner. These findings provide a stratification biomarker for antimitotic chemotherapy response.This work was supported by: FEDER (Fundo Europeu de Desenvolvimento Regional) funds through the COMPETE 2020 - Operational Programme for Competitiveness and Internationalization (POCI), Portugal 2020 and by Portuguese funds through FCT (Fundação para a Ciência e a Tecnologia) in the framework of the project POCI-01-0145-FEDER-031120 (PTDC/BIA-CEL/31120/ 2017); and POCI-01-0145-FEDER-007274 i3S framework project co-funded by COMPETE 2020/ PORTUGAL 2020 through FEDER. S.V. and F.F. were supported by FCT fellowships SFRH/BD/125017/2016 and PD/BD/105745/2014. E.L. was supported by an FCT Investigator Grant (IF/00916/2014). U.B-D. and G.L. were supported by the Azrieli Faculty Fellowship (to U.D.-D.) and the DoD CDMRP Career Development Award (CA191138 to U.B.-D.). J.B. was supported by an FCT Investigator Grant (CEECIND/03482/2018) and the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation programme (ERC-2015-StG-680156-ZPR)

Reprocessamento de dispositivos médicos de uso único: resultados clínicos e financeiros

ABSTRACT - Introduction: Excellent results regarding clinical efficacy and cost reduction are achieved by using certified reprocessed single-use medical devices (SUMDs). This explains why this practice is common in most hospitals across the USA and Germany. However, this is not a common practice at a national level and there are no reports regarding its use in Portugal. We present our experience using these methods at Centro Hospitalar de São João (CHSJ) in Porto. Objective: The aim of this study was to compare the clinical results and the financial impact of the use of 2 SUMDs, original and reprocessed, namely the linear suture machine GIA CovidienTM and Harmonic ACE® scissors. Method: A group of 733 patients operated in 2014 was evaluated. Of these patients, 316 were operated on with reprocessed SUMDs and 417 with original SUMDs. Variables referring to the clinical and financial results were analyzed through clinical and management information provided by Unidade de Gestão Autónoma de Cirurgia. A comparison between groups was performed using the χ2 test and the Mann-Whitney test. Results: Indicators related to clinical efficacy show that the use of these SUMDs, professionally reprocessed, did not represent any added risk in comparison to the use of original devices. Regarding costs, there is a very significant difference between the use of a new medical device and that of a reprocessed one. In the case of the Harmonic ACE® scissors and of the linear suture machine GIA CovidienTM, savings were up to 50% per device. Conclusion: This study, the first in Portugal, confirms the economic advantages of reprocessing these 2 devices. The financial benefit was obtained with maintenance of the same clinical results as the ones achieved using original devices. These results are in line with the published literature, proving the validity of using SUMDs after professionally certified reprocessing.RESUMO - Introdução: Os excelentes resultados da eficácia clínica e da redução de custos, obtidos com o reprocessamento certificado dos Dispositivos Médicos de Uso Único (DMUU) justificam que seja prática corrente na maior parte dos hospitais, nos Estados Unidos da América e na Alemanha. No entanto, a nível Nacional ainda não é prática comum, não existindo bibliografia sobre a experiência nacional. Apresentamos os resultados obtidos com esta prática no Centro Hospitalar de São João (CHSJ), Porto. Objetivo: Comparar os resultados clínicos e o impacto financeiro da reutilização de dispositivos médicos de uso único, reprocessados profissionalmente. Foram utilizados dois dispositivos de “uso único,” a máquina de sutura linear GIA CovidienTM e a tesoura Harmonic ACE®. Método: Analisamos um grupo de 733 doentes operados em 2014 que inclui todos os doentes operados no referido período e em que foram utilizados estes dispositivos. Destes doentes 316 foram operados com DMUU reprocessados, e 417 com DMUU originais. As variáveis referentes aos resultados clínicos e financeiros foram analisadas através dos respetivos registos clínicos e da informação de gestão fornecida pela Unidade de Gestão Autónoma de Cirurgia. A comparação entre grupos foi realizada pela aplicação dos testes de Qui-quadrado e Mann-Whitney. Resultados: Os indicadores relativos à eficácia clínica, comprovaram que o uso destes DMUU reprocessados profissionalmente utilizados nas intervenções cirúrgicas não representou qualquer risco acrescido em comparação com os dispositivos originais. Em termos financeiros, há uma diferença muito significativa entre a aquisição de um dispositivo médico novo ou reprocessado. A diferença no caso da tesoura Harmonic ACE® e na máquina de sutura linear GIA CovidienTM acarretou poupanças cerca de 50%. Conclusão: Este primeiro estudo, realizado em Portugal, confirma as vantagens económicas do reprocessamento destes dois dispositivos. O benefício financeiro foi obtido com manutenção da mesma qualidade clínica que se obteve com os dispositivos originais. Os resultados obtidos estão em conformidade com os publicados na literatura, o que confirma que a utilização de alguns dispositivos médicos de uso único após reprocessamento profissional é um método eficiente.info:eu-repo/semantics/publishedVersio

A tumor suppressor role of the Bub3 spindle checkpoint protein after apoptosis inhibition.

Most solid tumors contain aneuploid cells, indicating that the mitotic checkpoint is permissive to the proliferation of chromosomally aberrant cells. However, mutated or altered expression of mitotic checkpoint genes accounts for a minor proportion of human tumors. We describe a Drosophila melanogaster tumorigenesis model derived from knocking down spindle assembly checkpoint (SAC) genes and preventing apoptosis in wing imaginal discs. Bub3-deficient tumors that were also deficient in apoptosis displayed neoplastic growth, chromosomal aneuploidy, and high proliferative potential after transplantation into adult flies. Inducing aneuploidy by knocking down CENP-E and preventing apoptosis does not induce tumorigenesis, indicating that aneuploidy is not sufficient for hyperplasia. In this system, the aneuploidy caused by a deficient SAC is not driving tumorigenesis because preventing Bub3 from binding to the kinetochore does not cause hyperproliferation. Our data suggest that Bub3 has a nonkinetochore-dependent function that is consistent with its role as a tumor suppressor

A Conserved Arginine-Rich Motif within the Hypervariable N-Domain of Drosophila Centromeric Histone H3 (CenH3CID) Mediates BubR1 Recruitment

Centromere identity is determined epigenetically by deposition of CenH3, a centromere-specific histone H3 variant that dictates kinetochore assembly. The molecular basis of the contribution of CenH3 to centromere/kinetochore functions is, however, incompletely understood, as its interactions with the rest of centromere/kinetochore components remain largely uncharacterised at the molecular/structural level.Here, we report on the contribution of Drosophila CenH3(CID) to recruitment of BubR1, a conserved kinetochore protein that is a core component of the spindle attachment checkpoint (SAC). This interaction is mediated by the N-terminal domain of CenH3(CID) (NCenH3(CID)), as tethering NCenH3(CID) to an ectopic reporter construct results in BubR1 recruitment and BubR1-dependent silencing of the reporter gene. Here, we also show that this interaction depends on a short arginine (R)-rich motif and that, most remarkably, it appears to be evolutionarily conserved, as tethering constructs carrying the highly divergent NCenH3 of budding yeast and human also induce silencing of the reporter. Interestingly, though NCenH3 shows an exceedingly low degree of conservation, the presence of R-rich motives is a common feature of NCenH3 from distant species. Finally, our results also indicate that two other conserved sequence motives within NCenH3(CID) might also be involved in interactions with kinetochore components.These results unveil an unexpected contribution of the hypervariable N-domain of CenH3 to recruitment of kinetochore components, identifying simple R-rich motives within it as evolutionary conserved structural determinants involved in BubR1 recruitment

MAPK-Activated Protein Kinase 2 Is Required for Mouse Meiotic Spindle Assembly and Kinetochore-Microtubule Attachment

MAPK-activated protein kinase 2 (MK2), a direct substrate of p38 MAPK, plays key roles in multiple physiological functions in mitosis. Here, we show for the first time the unique distribution pattern of MK2 in meiosis. Phospho-MK2 was localized on bipolar spindle minus ends and along the interstitial axes of homologous chromosomes extending over centromere regions and arm regions at metaphase of first meiosis (MI stage) in mouse oocytes. At metaphase of second meiosis (MII stage), p-MK2 was localized on the bipolar spindle minus ends and at the inner centromere region of sister chromatids as dots. Knockdown or inhibition of MK2 resulted in spindle defects. Spindles were surrounded by irregular nondisjunction chromosomes, which were arranged in an amphitelic or syntelic/monotelic manner, or chromosomes detached from the spindles. Kinetochore–microtubule attachments were impaired in MK2-deficient oocytes because spindle microtubules became unstable in response to cold treatment. In addition, homologous chromosome segregation and meiosis progression were inhibited in these oocytes. Our data suggest that MK2 may be essential for functional meiotic bipolar spindle formation, chromosome segregation and proper kinetochore–microtubule attachments

Dual Role of Topoisomerase II in Centromere Resolution and Aurora B Activity

Chromosome segregation requires sister chromatid resolution. Condensins are essential for this process since they organize an axial structure where topoisomerase II can work. How sister chromatid separation is coordinated with chromosome condensation and decatenation activity remains unknown. We combined four-dimensional (4D) microscopy, RNA interference (RNAi), and biochemical analyses to show that topoisomerase II plays an essential role in this process. Either depletion of topoisomerase II or exposure to specific anti-topoisomerase II inhibitors causes centromere nondisjunction, associated with syntelic chromosome attachments. However, cells degrade cohesins and timely exit mitosis after satisfying the spindle assembly checkpoint. Moreover, in topoisomerase II–depleted cells, Aurora B and INCENP fail to transfer to the central spindle in late mitosis and remain tightly associated with centromeres of nondisjoined sister chromatids. Also, in topoisomerase II–depleted cells, Aurora B shows significantly reduced kinase activity both in S2 and HeLa cells. Codepletion of BubR1 in S2 cells restores Aurora B kinase activity, and consequently, most syntelic attachments are released. Taken together, our results support that topoisomerase II ensures proper sister chromatid separation through a direct role in centromere resolution and prevents incorrect microtubule–kinetochore attachments by allowing proper activation of Aurora B kinase