Rationale, study design, and analysis plan of the Alveolar Recruitment for ARDS Trial (ART): Study protocol for a randomized controlled trial

Background: Acute respiratory distress syndrome (ARDS) is associated with high in-hospital mortality. Alveolar recruitment followed by ventilation at optimal titrated PEEP may reduce ventilator-induced lung injury and improve oxygenation in patients with ARDS, but the effects on mortality and other clinical outcomes remain unknown. This article reports the rationale, study design, and analysis plan of the Alveolar Recruitment for ARDS Trial (ART). Methods/Design: ART is a pragmatic, multicenter, randomized (concealed), controlled trial, which aims to determine if maximum stepwise alveolar recruitment associated with PEEP titration is able to increase 28-day survival in patients with ARDS compared to conventional treatment (ARDSNet strategy). We will enroll adult patients with ARDS of less than 72 h duration. The intervention group will receive an alveolar recruitment maneuver, with stepwise increases of PEEP achieving 45 cmH(2)O and peak pressure of 60 cmH2O, followed by ventilation with optimal PEEP titrated according to the static compliance of the respiratory system. In the control group, mechanical ventilation will follow a conventional protocol (ARDSNet). In both groups, we will use controlled volume mode with low tidal volumes (4 to 6 mL/kg of predicted body weight) and targeting plateau pressure <= 30 cmH2O. The primary outcome is 28-day survival, and the secondary outcomes are: length of ICU stay; length of hospital stay; pneumothorax requiring chest tube during first 7 days; barotrauma during first 7 days; mechanical ventilation-free days from days 1 to 28; ICU, in-hospital, and 6-month survival. ART is an event-guided trial planned to last until 520 events (deaths within 28 days) are observed. These events allow detection of a hazard ratio of 0.75, with 90% power and two-tailed type I error of 5%. All analysis will follow the intention-to-treat principle. Discussion: If the ART strategy with maximum recruitment and PEEP titration improves 28-day survival, this will represent a notable advance to the care of ARDS patients. Conversely, if the ART strategy is similar or inferior to the current evidence-based strategy (ARDSNet), this should also change current practice as many institutions routinely employ recruitment maneuvers and set PEEP levels according to some titration method.Hospital do Coracao (HCor) as part of the Program 'Hospitais de Excelencia a Servico do SUS (PROADI-SUS)'Brazilian Ministry of Healt

Molecular characterization of the circadian clock elements during the development and senecence of Apis mellifera

O ciclo circadiano é um sistema adaptativo e vantajoso que permite a antecipação dos organismos e sincronização de suas atividades fisiológicas frente às variações ambientais que ocorrem ao longo do dia. Seu funcionamento molecular acontece pela geração dos ritmos circadianos, os quais surgem a partir da expressão cíclica dos genes do relógio em um feedback autoregulatório. Em insetos, os ritmos circadianos apresentam função importante em coordenar o timing do desenvolvimento e o comportamento. Nos últimos anos, estudos desvendaram que o relógio molecular de insetos sociais apresenta um funcionamento mais similar ao relógio de mamíferos do que com outros insetos como Drosophila melanogaster. Em especial, abelhas sociais têm sido ótimos modelos para investigar como os ritmos circadianos são modulados de acordo com as interações sociais entre os indivíduos, a plasticidade comportamental e a divisão social do trabalho entre as operárias. Operárias jovens (nutrizes) cuidam da cria no interior da colônia e geralmente apresentam uma atividade arrítimica ao longo de 24 horas, enquanto as abelhas mais velhas (forrageiras) são rítmicas e desenvolvem atividades complexas no ambiente externo. Nesse trabalho, nós caracterizamos os perfis de expressão dos genes do relógio period (per), cryptochrome mammalian-like (cry-m), clock (clk), cycle (cyc), timeout 2 (tim2), par domain protein 1 (pdp1), vrille (vri) e clockwork orange (cwo) durante todo o desenvolvimento de abelhas operárias de Apis mellifera. Verificamos que os genes do relógio são expressos antes mesmo da formação do sistema nervoso central no embrião e que seus transcritos podem ser herdados maternalmente. No desenvolvimento de larvas e pupas, revelamos que estes genes são diferencialmente expressos entre as fases investigadas e, com exceção dos genes cwo e tim2, todos respondem ao tratamento com o Hormônio Juvenil (HJ) na fase de pupa de olho branco. A resposta positiva dos genes clk, cyc e pdp1 frente ao tratamento hormonal pode estar relacionada com o envolvimento destes nas vias que respondem à sinalização do HJ, interagindo com os genes Kruppel (Kr-h1) e Methoprene-tolerant (MET). No desenvolvimento adulto, vimos que os genes per e cry-m são potenciais marcadores da plasticidade comportamental e divisão social do trabalho. Em um experimento usando single-cohort colony, estes genes apresentam níveis transcricionais que não oscilam em cabeças de operárias jovens (3 e 7 dias) ao longo de 24 horas, comparado aos níveis de expressão que oscilam de forma robusta em abelhas mais velhas (15 e 25 dias). Ainda, reconstruímos redes de interação proteína-proteína e miRNA-mRNA onde foram identificadas potenciais moléculas que atuam modulando os genes do relógio em nível póstranscricional e traducional. Dentre elas, validamos as interações entre os miRNA-34 e seus sítios de ligação que estão presentes nas 3`UTRS dos genes cyc e cwo, através do ensaio por luciferase, revelando que este miRNA é um regulador negativo da expressão desses genes. Pela primeira vez, realizamos uma análise ampla dos genes do relógio em um inseto social, além de identificar novas moléculas que podem atuar modulando os ritmos circadianos. Nosso trabalho demonstra a importâcia das abelhas sociais como modelos ideais para desvendar os mecanismos moleculares que regem os ritmos circadianos não só em abelhas, como também em outros organismos, inclusive mamíferos.The circadian clock is an advantageous adaptive system that enables organisms to anticipate and syncronize their biological activities during the daily environmental changes. The circadian clock acts through the ontogeny of circadian rhythms, which are generated by the cyclic expression of the clock genes in an autregulatory feedback loop. In insects, the circadian rhythms have important roles in the coordination of the developmental timing and behavior, interacting with the endocrine system. In the last years, researchers revealed that the molecular clock of social insects is more similar to mammals than to insects. In particular, the social honeybee is an excellent model to investigate how the circadian rhythms are modulated accordingly to the social context, behavioral plasticity, and taskrelated activities. While young bees (nurses) work arrhythmically around the clock inside the colony in brood-care activities, old bees (foragers) need to be strongly rhythmic to develop complex tasks. In this work, we characterized the expression patterns of the clock genes period (per), cryptochrome mammalian-like (cry-m), clock (clk), cycle (cyc), timeout 2 (tim2), par domain protein 1 (pdp1), vrille (vri) e clockwork orange (cwo) in the entire development of Apis mellifera. Our results revealed that the clock genes are expressed before the formation of the central nervous system in embryos and that their transcripts might be inherited maternally. The clock genes are diferentially modulated during the larval and pupal development and, except for tim2 and cwo, all of them respond to the treatment with Juvenile Hormone (JH) in white-eyed pupae. The positive response to JH by clk, cyc and pdp1 might be related to the involvement of these genes on the pathways of the JH signaling, interacting with Kruppel (Kr-h1) and Methoprene-tolerant (MET) genes. In the adult development, the clock genes per and cry-m are potential molecular markers of the behavioral plasticity and division of labor in a single-cohort colony, once they did not exhibit transcriptional oscillations in heads of young bees (3 and 7 days-old) during 24h, compared to the robust transcriptional oscillation in old bees (15 and 25 days-old). Additionally, we reconstructed protein-protein and miRNA-mRNA interaction networks and identified putative molecules involved in the post-transcriptional and translational regulation of the clock genes. Among those molecules, we validated interactions between the miR-34 and its binding sites in the 3`UTR of cyc and cwo by luciferase assay, showing that this miRNA is a negative regulator of both clock genes. We showed for the first time a broad analysis of the circadian clock elements in a social insect, and also identified news molecules with potential to act as modulators of the circadian rhythms. This work expands the knowledge about the biological roles of the circadian clock in honeybees. Our work also contributes to highlight the importance of honeybees as an ideal model to uncover the molecular mechanisms that govern the circadian rhythms, not only in bees, but in other organisms, including mammals

Molecular characterization of the circadian clock elements during the development and senecence of Apis mellifera

O ciclo circadiano é um sistema adaptativo e vantajoso que permite a antecipação dos organismos e sincronização de suas atividades fisiológicas frente às variações ambientais que ocorrem ao longo do dia. Seu funcionamento molecular acontece pela geração dos ritmos circadianos, os quais surgem a partir da expressão cíclica dos genes do relógio em um feedback autoregulatório. Em insetos, os ritmos circadianos apresentam função importante em coordenar o timing do desenvolvimento e o comportamento. Nos últimos anos, estudos desvendaram que o relógio molecular de insetos sociais apresenta um funcionamento mais similar ao relógio de mamíferos do que com outros insetos como Drosophila melanogaster. Em especial, abelhas sociais têm sido ótimos modelos para investigar como os ritmos circadianos são modulados de acordo com as interações sociais entre os indivíduos, a plasticidade comportamental e a divisão social do trabalho entre as operárias. Operárias jovens (nutrizes) cuidam da cria no interior da colônia e geralmente apresentam uma atividade arrítimica ao longo de 24 horas, enquanto as abelhas mais velhas (forrageiras) são rítmicas e desenvolvem atividades complexas no ambiente externo. Nesse trabalho, nós caracterizamos os perfis de expressão dos genes do relógio period (per), cryptochrome mammalian-like (cry-m), clock (clk), cycle (cyc), timeout 2 (tim2), par domain protein 1 (pdp1), vrille (vri) e clockwork orange (cwo) durante todo o desenvolvimento de abelhas operárias de Apis mellifera. Verificamos que os genes do relógio são expressos antes mesmo da formação do sistema nervoso central no embrião e que seus transcritos podem ser herdados maternalmente. No desenvolvimento de larvas e pupas, revelamos que estes genes são diferencialmente expressos entre as fases investigadas e, com exceção dos genes cwo e tim2, todos respondem ao tratamento com o Hormônio Juvenil (HJ) na fase de pupa de olho branco. A resposta positiva dos genes clk, cyc e pdp1 frente ao tratamento hormonal pode estar relacionada com o envolvimento destes nas vias que respondem à sinalização do HJ, interagindo com os genes Kruppel (Kr-h1) e Methoprene-tolerant (MET). No desenvolvimento adulto, vimos que os genes per e cry-m são potenciais marcadores da plasticidade comportamental e divisão social do trabalho. Em um experimento usando single-cohort colony, estes genes apresentam níveis transcricionais que não oscilam em cabeças de operárias jovens (3 e 7 dias) ao longo de 24 horas, comparado aos níveis de expressão que oscilam de forma robusta em abelhas mais velhas (15 e 25 dias). Ainda, reconstruímos redes de interação proteína-proteína e miRNA-mRNA onde foram identificadas potenciais moléculas que atuam modulando os genes do relógio em nível póstranscricional e traducional. Dentre elas, validamos as interações entre os miRNA-34 e seus sítios de ligação que estão presentes nas 3`UTRS dos genes cyc e cwo, através do ensaio por luciferase, revelando que este miRNA é um regulador negativo da expressão desses genes. Pela primeira vez, realizamos uma análise ampla dos genes do relógio em um inseto social, além de identificar novas moléculas que podem atuar modulando os ritmos circadianos. Nosso trabalho demonstra a importâcia das abelhas sociais como modelos ideais para desvendar os mecanismos moleculares que regem os ritmos circadianos não só em abelhas, como também em outros organismos, inclusive mamíferos.The circadian clock is an advantageous adaptive system that enables organisms to anticipate and syncronize their biological activities during the daily environmental changes. The circadian clock acts through the ontogeny of circadian rhythms, which are generated by the cyclic expression of the clock genes in an autregulatory feedback loop. In insects, the circadian rhythms have important roles in the coordination of the developmental timing and behavior, interacting with the endocrine system. In the last years, researchers revealed that the molecular clock of social insects is more similar to mammals than to insects. In particular, the social honeybee is an excellent model to investigate how the circadian rhythms are modulated accordingly to the social context, behavioral plasticity, and taskrelated activities. While young bees (nurses) work arrhythmically around the clock inside the colony in brood-care activities, old bees (foragers) need to be strongly rhythmic to develop complex tasks. In this work, we characterized the expression patterns of the clock genes period (per), cryptochrome mammalian-like (cry-m), clock (clk), cycle (cyc), timeout 2 (tim2), par domain protein 1 (pdp1), vrille (vri) e clockwork orange (cwo) in the entire development of Apis mellifera. Our results revealed that the clock genes are expressed before the formation of the central nervous system in embryos and that their transcripts might be inherited maternally. The clock genes are diferentially modulated during the larval and pupal development and, except for tim2 and cwo, all of them respond to the treatment with Juvenile Hormone (JH) in white-eyed pupae. The positive response to JH by clk, cyc and pdp1 might be related to the involvement of these genes on the pathways of the JH signaling, interacting with Kruppel (Kr-h1) and Methoprene-tolerant (MET) genes. In the adult development, the clock genes per and cry-m are potential molecular markers of the behavioral plasticity and division of labor in a single-cohort colony, once they did not exhibit transcriptional oscillations in heads of young bees (3 and 7 days-old) during 24h, compared to the robust transcriptional oscillation in old bees (15 and 25 days-old). Additionally, we reconstructed protein-protein and miRNA-mRNA interaction networks and identified putative molecules involved in the post-transcriptional and translational regulation of the clock genes. Among those molecules, we validated interactions between the miR-34 and its binding sites in the 3`UTR of cyc and cwo by luciferase assay, showing that this miRNA is a negative regulator of both clock genes. We showed for the first time a broad analysis of the circadian clock elements in a social insect, and also identified news molecules with potential to act as modulators of the circadian rhythms. This work expands the knowledge about the biological roles of the circadian clock in honeybees. Our work also contributes to highlight the importance of honeybees as an ideal model to uncover the molecular mechanisms that govern the circadian rhythms, not only in bees, but in other organisms, including mammals

Expressão de microRNAs em formas larvais e adultas de Schistosoma mansoni.

Programa de Pós-Graduação em Biotecnologia. Núcleo de Pesquisas em Ciências Biológicas, Pró-Reitoria de Pesquisa e Pós Graduação, Universidade Federal de Ouro Preto.A esquistossomose é uma doença debilitante causada por platelmintos do gênero Schistosoma. Devido ao seu ciclo de vida complexo, posição evolutiva e dimorfismo sexual, o parasita Schistosoma mansoni é um modelo interessante para investigar mecanismos de regulação da expressão gênica. MicroRNAs (miRNAs) são pequenas moléculas de RNA endógeno que regulam a expressão gênica em nível pós-transcricional pela interação específica a RNA mensageiros alvos. O conhecimento atual de miRNAs em S. mansoni é limitado e baseado em predições computacionais e sequenciamento de nova geração a partir de bibliotecas de vermes adultos. No presente estudo, nós caracterizamos os perfis de expressão de nove microRNAs maduros nas formas larvais e adultas do parasita usando qRT-PCR. Além disso, foram identificados os genes alvos desses miRNAs utilizando abordagens computacionais. Nossos resultados mostraram padrões de expressão diferenciais para os miRNAs: sma-miR-2162-3p, sma-miR-250, sma-miR-92a, sma-miR-new_2-5p, sma-miR-new_4-3p, sma-miR-new_4-5p, sma-miR-new_12-5p, sma-miR-new_13-5p e sma-miR-new-16-3p. A análise computacional revelou alvos de miRNAs envolvidos em processos celulares importantes como a via de sinalização TGF-β, metabolismo de glicose e lipídeos, proteína Tetraspanina (TSP), proteína Catepsina B1 (CB1) e proteína VAL-6 (“Venom-allergen-like 6 protein”). Além disso, grande parte dos genes preditos estão envolvidos com o processo de fosforilação oxidativa, sugerindo que pelo menos em parte a expressão dos genes que codificam para subunidades do complexo NADH desidrogenase, citocromo c oxidase e ATP sintase são regulados por miRNAs. Observamos também, seis alvos associados ao sistema ubiquitina-proteassoma, sugerindo que esses miRNAs possam regular esse importante processo biológico no parasita. Tomados em conjunto, podemos concluir que os miRNAs analisados são diferencialmente expressos e atuam fortemente como reguladores da expressão gênica em nível pós-transcricional.Schistosomiasis is a debilitating disease that is caused by Platyhelminths of the genus Schistosoma. Due to its complex life cycle, evolutional position and sexual dimorphism, schistosomes can serve as an interesting model to investigate mechanisms of gene regulation. MicroRNAs (miRNAs) are short endogenous RNA molecules that regulate gene expression at the post-transcriptional level by targeting mRNA transcripts. Current knowledge of Schistosoma mansoni miRNAs is limited and is based on computational predictions and next-generation sequencing through adult worms libraries. In this study, we validated the expression profiles of thirteen mature miRNAs in different life cycle stages of the parasite using qRT-PCR and identified the miRNA target genes using a computational approach. Our results showed differential expression patterns of the miRNAs: sma-miR-2162-3p, sma-miR-250, sma-miR-92a, sma-miR-new_2-5p, sma-miR-new_4-3p, sma-miR-new_4-5p, sma-miR-new_12-5p, sma-miR-new_13-5p e sma-miR-new-16-3p. The computational analysis revealed miRNA target genes that are related to important biological processes, such as TGF-β signalling, glucose and lipid metabolism, tetraspanin protein (TSP), cathepsin B 1 protein (CB1) and Venom allergen-like 6 protein (VAL-6). Furthermore, most of the target genes that were found are linked to oxidative phosphorylation, suggesting that at least in part the expression of NADH dehydrogenase, cytochrome c oxidase e ATP synthase genes are regulated by microRNAs. We also observed six miRNA target genes that are involved in the proteasome-ubiquitin protein degradation pathway, suggesting that miRNAs can regulate this important biological process in the parasite. Together, our results lead us to conclude that the microRNAs analysed are differentially expressed and act in post-transcriptional regulation of genes

In silico analysis and developmental expression of ubiquitin-conjugating enzymes in Schistosoma mansoni.

Ubiquitin-conjugating enzymes (Ub-E2) perform the second step of ubiquitination and, consequently, are essential for regulating proteolysis and for modulating protein function, interactions and trafficking. Previously, our group demonstrated the crucial role of ubiquitination and the Ubproteasome pathway during the Schistosoma mansoni life cycle. In the present investigation, we used a homology-based genome-wide bioinformatics approach to identify and molecularly characterise the Ub-E2 enzymes in S. mansoni. The putative functions were further investigated through molecular phylogenetic and expression profile analyses using cercariae, adult worms, eggs and mechanically transformed schistosomula (MTS) cultured in vitro for 3.5 h or 1 or 3 days. We identified, via in silico analysis, 17 Ub-E2 enzymes with conserved structural characteristics: the beta-sheet and the helix-2 form a central core bordered by helix-1 at one side and helix-3 and helix-4 at the other. The observed quantitative differences in the steady-state transcript levels between the cercariae and adult worms may contribute to the differential protein ubiquitination observed during the parasite?s life cycle. This study is the first to identify and characterise the E2 ubiquitin conjugation family in S. mansoni and provides fundamental information regarding their molecular phylogenetics and developmental expression during intra-mammalian stages

Characterization of export receptor exportins (XPOs) in the parasite Schistosoma mansoni.

Several proteins and different species of RNA that are produced in the nucleus are exported through the nuclear pore complexes, which require a family of conserved nuclear export receptors called exportins (XPOs). It has been reported that the XPOs (XPO1, XPO5, and XPOT) are directly involved in the transport processes of noncoding RNAs from the nucleus to the cytoplasm and/or from cytoplasm to the nucleus. All three genes are present in fungi, plants, and deuterostome metazoans. However, protostome metazoan species lack one of the three genes across evolution. In this report, we have demonstrated that all three XPO proteins are present in the parasite protostome Schistosoma mansoni . As this parasite has a complex life cycle presenting several stages in different hosts and environments, implying a differential gene regulation, we proposed a genomic analysis of XPOs to validate their annotation. The results showed the conservation of exportin family members and gene duplication events in S. mansoni .We performed quantitative RT-PCR, which revealed an upregulation of SmXPO1 in 24 h schistosomula (sixfold when compared with cercariae), and similar transcription levels were observed for SmXPO5 and SmXPOT in all the analyzed stages. These three XPO proteins have been identified for the first time in the protostome clade, which suggests a higher complexity in RNA transport in the parasite S. mansoni. Taken together, these results suggest that RNA transport by exportins might control cellular processes during cercariae, schistosomula, and adult worm development

ACETATO DE TRIANCINOLONA COMO PRÉ-INDUTOR DO PARTO DE RECEPTORAS DE EMBRIÃO NELORE PRODUZIDOS IN VITRO INDUCTION OF PARTURITION USING TRIAMCINOLONE PRETREATMENT IN PREGNANT RECIPIENTS OF IN VITRO PRODUCED EMBRYO

Este estudo teve como objetivo avaliar o efeito de diferentes doses e momentos da gestação para a administração do acetato de triancinolona (TRI) na pré-indução de partos. Utilizaram-se como modelo experimental receptoras de embriôes Nelore produzidos in vitro (PIV). Cento e setenta e cinco receptoras gestantes foram distribuídas em quatro grupos experimentais de acordo com o tempo gestacional (280 ou 285 dias) e a dose de TRI (1mg/60 Kg ou 1mg/100 Kg de Peso Vivo). Avaliaram-se o momento da gestação, as taxas de sincronização e de assistência aos partos, a viabilidade e o peso dos bezerros e ainda a taxa de retenção de placenta. A dose de TRI não alterou os parâmetros avaliados (p>0,05). Houve maior sincronização dos partos (p<0,05) quando o TRI foi administrado aos 280 dias em relação aos 285 dias de gestação. Os partos de bezerros mais pesados (?42 kg) necessitaram maior assistência (p?0,05). Trata-se de produtos que apresentaram menor viabilidade (p?0,05) quanto aos partos de bezerros mais leves. Conclui-se que, independente da dose de TRI, a pré-indução aos 280 dias de gestação resulta em maior sincronização e melhor previsibilidade dos nascimentos, quando comparada à pré-indução aos 285 dias, sem comprometer a viabilidade dos bezerros.<br /><br />PALAVRAS-CHAVES: Corticoide de longa ação, indução de parto, retenção de placenta. The aim of the present study was to evaluate the effect of two doses of acetonide triamcinolone (TRI) at two distinct moments of gestation in the pre induction of parturition. Embryo recipients pregnant of the in vitro produced embryo were utilized as experimental model. A total of 175 pregnant embryo recipients were assigned into one of the four groups according to gestational period (280 or 285 days) and utilized dose of the TRI (1mg/60Kg or 1mg/100kg of body weight). The synchronization and assistance rates of the calving, the viability and body weight of calves at calving as well as retained placenta rate were evaluated. The TRI dose did not effect any variables responses (p>0.05). Least spread of the calving moment was observed in the recipients treated at the 280 days of gestation (p?0.05). The cow calving calf with ?42kg needed more assistance to calving (p?0.05) and these calves had lower viability than calves with <42Kg of the body weight (p?0.05). The data suggested that, independent of dose of the TRI, the pre induction with TRI at 280 days of gestation promote a good synchronization in the calving with a high predictable calving time without compromise the calf viability when compared with the pre induction at 285 days of gestation.<br /><br />KEY WORDS: Induction of parturition, long action corticoid, retention placenta.<br /&gt

Exploring integument transcriptomes, cuticle ultrastructure, and cuticular hydrocarbons profiles in eusocial and solitary bee species displaying heterochronic adult cuticle maturation.

Differences in the timing of exoskeleton melanization and sclerotization are evident when comparing eusocial and solitary bees. This cuticular maturation heterochrony may be associated with life style, considering that eusocial bees remain protected inside the nest for many days after emergence, while the solitary bees immediately start outside activities. To address this issue, we characterized gene expression using large-scale RNA sequencing (RNA-seq), and quantified cuticular hydrocarbon (CHC) through gas chromatography-mass spectrometry in comparative studies of the integument (cuticle plus its underlying epidermis) of two eusocial and a solitary bee species. In addition, we used transmission electron microscopy (TEM) for studying the developing cuticle of these and other three bee species also differing in life style. We found 13,200, 55,209 and 30,161 transcript types in the integument of the eusocial Apis mellifera and Frieseomelitta varia, and the solitary Centris analis, respectively. In general, structural cuticle proteins and chitin-related genes were upregulated in pharate-adults and newly-emerged bees whereas transcripts for odorant binding proteins, cytochrome P450 and antioxidant proteins were overrepresented in foragers. Consistent with our hypothesis, a distance correlation analysis based on the differentially expressed genes suggested delayed cuticle maturation in A. mellifera in comparison to the solitary bee. However, this was not confirmed in the comparison with F. varia. The expression profiles of 27 of 119 genes displaying functional attributes related to cuticle formation/differentiation were positively correlated between A. mellifera and F. varia, and negatively or non-correlated with C. analis, suggesting roles in cuticular maturation heterochrony. However, we also found transcript profiles positively correlated between each one of the eusocial species and C. analis. Gene co-expression networks greatly differed between the bee species, but we identified common gene interactions exclusively between the eusocial species. Except for F. varia, the TEM analysis is consistent with cuticle development timing adapted to the social or solitary life style. In support to our hypothesis, the absolute quantities of n-alkanes and unsaturated CHCs were significantly higher in foragers than in the earlier developmental phases of the eusocial bees, but did not discriminate newly-emerged from foragers in C. analis. By highlighting differences in integument gene expression, cuticle ultrastructure, and CHC profiles between eusocial and solitary bees, our data provided insights into the process of heterochronic cuticle maturation associated to the way of life