Expression of FoxC, FoxF, FoxL1, and FoxQ1 genes in the dogfish scyliorhinus canicula defines ancient and derived roles for fox genes in vertebrate development

In the human genome, members of the FoxC, FoxF, FoxL1, and FoxQ1 gene families are found in two paralagous clusters. Here we characterize all four gene families in the dogfish Seyliorhinus canicula, a member of the cartilaginous fish lineage that diverged before the radiation of osteichthyan vertebrates. We identify two FoxC genes, two FoxF genes, and single FoxQ1 and FoxL1 genes, demonstrating cluster duplication preceded the radiation of gnathostomes. The expression of all six genes was analyzed by in situ hybridization. The results show conserved expression of FoxL1, FoxF, and FoxC genes in different compartments of the mesoderm and of FoxQ1 in pharyngeal endoderm and its derivatives, confirming these as ancient sites of Fox gene expression, and also illustrate multiple cases of lineage-specific expression domains. Comparison to invertebrate chordates shows that the majority of conserved vertebrate expression domains mark tissues that are part of the primitive chordate body plan.

The amphioxus FoxQ1 gene is expressed in the developing endostyle

The FoxQ1 genes form a distinct group within the Fox (also known as forkhead) gene family. We have isolated a gene from the amphioxus Branchiostoma floridae that encodes a forkhead domain with high identity to FoxQ1 genes in other chordates. Molecular phylogenetic analysis places AmphiFoxQ1 in a robust grouping with vertebrate FoxQ1 genes and with Ciona intestinalis Ci-FoxQ1. This group is separate from that containing AmphiFoxQ2, which instead groups with other invertebrate Fox genes. The expression of AmphiFoxQ1 was analysed by whole mount in situ hybridisation. The results show that AmphiFoxQ1 expression is confined to the developing endoderm, and specifically marks the endostyle and associated peripharyngeal bands of amphioxus larvae. Ci-FoxQ1 is also expressed in the endostyle, highlighting this as a conserved site of FoxQ1 gene expression in basal chordates. (C) 2004 Elsevier B.V. All rights reserved.

Tissue distribution of quiescin Q6/sulfhydryl oxidase (QSOX) in developing mouse

Quiescin Q6/sulfhydryl oxidases (QSOX) are revisited thiol oxidases considered to be involved in the oxidative protein folding, cell cycle control and extracellular matrix remodeling. They contain thioredoxin domains and introduce disulfide bonds into proteins and peptides, with the concomitant hydrogen peroxide formation, likely altering the redox environment. Since it is known that several developmental processes are regulated by the redox state, here we assessed if QSOX could have a role during mouse fetal development. For this purpose, an anti-recombinant mouse QSOX antibody was produced and characterized. In E-13.5, E-16.5 fetal tissues, QSOX immunostaining was confined to mesoderm- and ectoderm-derived tissues, while in P1 neonatal tissues it was slightly extended to some endoderm-derived tissues. QSOX expression, particularly by epithelial tissues, seemed to be developmentally-regulated, increasing with tissue maturation. QSOX was observed in loose connective tissues in all stages analyzed, intra and possibly extracellularly, in agreement with its putative role in oxidative folding and extracellular matrix remodeling. In conclusion, QSOX is expressed in several tissues during mouse development, but preferentially in those derived from mesoderm and ectoderm, suggesting it could be of relevance during developmental processes.

Notch signaling controls the balance of ciliated and secretory cell fates in developing airways

Although there is accumulated evidence of a role for Notch in the developing lung, it is still unclear how disruption of Notch signaling affects lung progenitor cell fate and differentiation events in the airway epithelium. To address this issue, we inactivated Notch signaling conditionally in the endoderm using a Shh-Cre deleter mouse line and mice carrying floxed alleles of the Pofut1 gene, which encodes an O-fucosyltransferase essential for Notch-ligand binding. We also took the same conditional approach to inactivate expression of Rbpjk, which encodes the transcriptional effector of canonical Notch signaling. Strikingly, these mutants showed an almost identical lung phenotype characterized by an absence of secretory Clara cells without evidence of cell death, and showed airways populated essentially by ciliated cells, with an increase in neuroendocrine cells. This phenotype could be further replicated in cultured wild-type lungs by disrupting Notch signaling with a gamma-secretase inhibitor. Our data suggest that Notch acts when commitment to a ciliated or non-ciliated cell fate occurs in proximal progenitors, silencing the ciliated program in the cells that will continue to expand and differentiate into secretory cells. This mechanism may be crucial to define the balance of differentiated cell profiles in different generations of the developing airways. It might also be relevant to mediate the metaplastic changes in the respiratory epithelium that occur in pathological conditions, such as asthma and chronic obstructive pulmonary disease.

XRCC1 haploinsufficiency in mice has little effect on aging, but adversely modifies exposure-dependent susceptibility

Oxidative DNA damage plays a role in disease development and the aging process. A prominent participant in orchestrating the repair of oxidative DNA damage, particularly single-strand breaks, is the scaffold protein XRCC1. A series of chronological and biological aging parameters in XRCC1 heterozygous (HZ) mice were examined. HZ and wild-type (WT) C57BL/6 mice exhibit a similar median lifespan of similar to 26 months and a nearly identical maximal life expectancy of similar to 37 months. However, a number of HZ animals (7 of 92) showed a propensity for abdominal organ rupture, which may stem from developmental abnormalities given the prominent role of XRCC1 in endoderm and mesoderm formation. For other end-points evaluated-weight, fat composition, blood chemistries, condition of major organs, tissues and relevant cell types, behavior, brain volume and function, and chromosome and telomere integrity-HZ mice exhibited by-and-large a normal phenotype. Treatment of animals with the alkylating agent azoxymethane resulted in both liver toxicity and an increased incidence of precancerous lesions in the colon of HZ mice. Our study indicates that XRCC1 haploinsufficiency in mammals has little effect on chronological longevity and many key biological markers of aging in the absence of environmental challenges, but may adversely affect normal animal development or increase disease susceptibility to a relevant genotoxic exposure.

Aspectos morfológicos do saco vitelino em roedores da subordem Hystricomorpha: paca (Agouti paca) e cutia (Dasyprocta aguti)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Structural and ultrastructural characteristics of the yolk syncytial layer in Prochilodus lineatus (Valenciennes, 1836) (Teleostei; Prochilodontidae)

The yolk syncytial layer (YSL) has been regarded as one of the main obstacles for a successful cryopreservation of fish embryos. The purpose of this study was to identify and characterize the YSL in Prochilodus lineatus, a fish species found in southeastern Brazil and considered a very important fishery resource. Embryos were obtained through artificial breeding by hormonal induction. After fertilization, the eggs were incubated in vertical incubators with a controlled temperature (28 degrees C). Embryos were collected in several periods of development up to hatching and then fixed with 2% glutaralclehyde and 4% paraformaldehyde in 0.1 M sodium phosphate buffer (pH 7.3). Morphological analyses were carried out under either light, transmission or scanning electron microscopy. The formation of the YSL in P. lineatus embryos starts at the end of the cleavage stage (morula), mainly at the margin of the blastoderm, and develops along the embryo finally covering the entire yolk mass (late gastrula) and producing a distinct intermediate zone between the yolk and the endodermal cells. The YSL was characterized by the presence of microvilli on the contact region with the yolk endoderm. A cytoplasmic mass, full of mitochondria, vacuoles, ribosomes, endomembrane nets and euchromatic nuclei, indicated a high metabolic activity. This layer is shown as an interface between the yolk and the embryo cells that, besides sustaining and separating the yolk, acts as a structure that makes it available for the embryo. The structural analyses identified no possible barriers to cryoprotectant penetration.

Alterações anatômicas foliares em plantas de Brachiaria subquadripara submetidas à aplicação de herbicidas

O presente trabalho teve por objetivo avaliar a eficiência do controle dos herbicidas diquat, glyphosate e imazapyr e as alterações anatômicas do limbo foliar provocadas por eles em plantas de Brachiaria subquadripara. As plantas foram cultivadas em caixas-d'água sob condições de campo e, quando atingiram estádio de pleno desenvolvimento vegetativo, foram pulverizadas com soluções de diquat a 400 g ha-1, glyphosate a 4.320 g ha-1 em associação com o adjuvante Aterbane a 0,5% v v-1 e imazapyr a 750 g ha-1. Além disso, utilizou-se uma testemunha sem aplicação de herbicida. Foram realizadas avaliações de controle e das seguintes características anatômicas quantitativas das regiões da nervura central e da internervural do limbo foliar, além da região do colmo: porcentagem da epiderme adaxial e abaxial, porcentagem da endoderme, porcentagem do feixe vascular, porcentagem de lacunas do aerênquima, porcentagem de parênquima, espessura foliar, porcentagem da epiderme do colmo, porcentagem do câmbio vascular e diâmetro do colmo. Os herbicidas diquat e imazapyr promoveram o maior número de alterações nos caracteres anatômicos quantitativos das regiões da nervura central e internervural do limbo foliar e da região do colmo das plantas. Contudo, com exceção do diquat, os herbicidas foram eficientes no controle das plantas de B. subquadripara, porém todos eles permitiram a rebrota delas.

Alterações anatômicas foliares em Eichhornia crassipes submetidas à aplicação de herbicidas

O presente trabalho teve por objetivo avaliar a eficiência do controle dos herbicidas 2,4-D, diquat e glyphosate e as alterações anatômicas do limbo foliar provocadas pelos produtos em plantas de Eichhornia crassipes, coletadas nos reservatórios do complexo CESP. As plantas foram cultivadas em caixas-d'água sob condições de campo e, quando atingiram estádio de pleno desenvolvimento vegetativo, foram pulverizadas com soluções de diquat a 400 g i.a.ha-1, 2,4-D a 1.340 g e.a. ha-1 e glyphosate a 4.320 g e.a. ha-1 em associação com o adjuvante Silwet L-77 a 0,01% v v-1. Foi utilizada uma testemunha sem aplicação de herbicida. A seguir, foram realizadas avaliações de controle e das seguintes características anatômicas quantitativas das regiões da nervura central e da internervural do limbo foliar: porcentagem da epiderme adaxial e abaxial, porcentagem da endoderme, porcentagem do feixe vascular, porcentagem de lacunas do aerênquima, porcentagem de parênquima e espessura foliar (µm). Com base nos resultados, os principais caracteres anatômicos quantitativos das regiões da nervura central e internervural do limbo foliar que sofreram alterações após a aplicação dos herbicidas foram a porcentagem da epiderme adaxial, a porcentagem da endoderme e a espessura foliar. Os herbicidas diquat e 2,4-D foram os que mais promoveram alterações nos caracteres anatômicos quantitativos das regiões da nervura central e internervural do limbo foliar das plantas de E. crassipes. Apenas o glyphosate apresentou controle de 100% das plantas aos 22 dias após a aplicação, quando comparado com o observado na testemunha sem herbicida.

Cryogenic preservation of embryos of Prochilodus lineatus (Valenciennes, 1836) (Characiforme; Prochilodontidae)

While the freezing techniques of mammal embryos have been providing promising results, the cryopreservation of teleostean eggs and embryos have remained unsuccessful up to now. Therefore, this work aimed to develop a procedure of cryogenic preservation of embryos of Prochilodus lineatus and to observe, at both structural and ultrastructural levels, the morphological alterations that took place after the application of freezing/thawing techniques. The embryos at the morula stage could not tolerate exposure to the cryoprotectants ethylene glycol monomethyl ether, propylene glycol monomethyl ether, methanol, dimethyl sulphoxide and propylene glycol, presenting 100% of mortality. Embryos at the 4- to 6-somites stage tolerated exposure to propylene glycol and dimethyl sulphoxide, and the results revealed no significant differences (alpha = 0.05) regarding survival from both treatments. None of the freezing, thawing and hydration protocols was effective on preserving embryo viability. The ultrastructural analyses of frozen and thawed embryos showed that cells from ectoderm, somites, notochord and endoderm were structurally intact, with well preserved nuclei and mitochondria. The yolk globules were able to tolerate the freezing process, but the yolk syncytial layer was unorganized, displaying an electron-dense and compacted appearance, collapsed reticules, nuclei with modified chromatin and ruptures on the plasmatic membrane at the contact zone with endoderm. It might be concluded that the procedures tested for freezing were unable to avoid the formation of intracellular ice crystals, leading to drastic morphological modifications and making P. lineatus embryos unviable.

Ultrastructure features of the midgut of the female adult Amblyomma cajennense ticks Fabricius, 1787 (Acari: Ixodidae) in several feeding stages and subjected to three infestations

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Extração e caracterização do amido de diferentes genótipos de bananeira

A banana, uma das principais herbáceas cultivadas em grande quantidade no mundo e importante fonte de energia para as pessoas, tem obtido interesse como matéria-prima para a produção de farinha e fécula para uso alimentício e outros fins industriais, devido ao elevado conteúdo de amido nos frutos verdes. O trabalho teve como objetivo caracterizar os frutos verdes de sete genótipos de bananeira quanto à composição química e avaliar o conteúdo de amilose, propriedades de pasta e térmicas dos amidos destes genótipos. Os resultados obtidos mostraram diferenças significativas na composição química dos frutos, com maior teor de amido na cultivar Figo Cinza. As análises dos amidos mostraram valores de amilose de 26,68 a 33,95% e propriedades de pasta com elevado pico de viscosidade, baixa resistência à temperatura e agitação (quebra de viscosidade) e tendência à retrogradação para todos os amidos. Nas propriedades térmicas, os amidos mostraram um endoderma, temperatura de gelatinização na faixa de 60 a 73ºC com DH variando de 11,6 a 16,99 J g-1 na gelatinização e 4,11 a 7,97 J g-1 na retrogradação.

Caracterização das membranas fetais em búfalas no terço inicial da gestação

Este estudo visou à caracterização das membranas fetais em búfalas (Bubalus bubalis, Linnaeus 1758) no terço inicial da gestação. As membranas fetais foram analisadas macroscópica e microscopicamente (luz e microscopia eletrônica de transmissão). O córion possui uma camada simples de células circulares, com núcleos de forma esférica, denominadas trofobláticas; há outro tipo celular, as células trofoblásticas gigantes, com dois ou mais núcleos. Ambas possuem uma grande quantidade de vesículas no citoplasma e retículo endoplasmático à microscopia de transmissão. O alantóide possui vasos preenchidos com eritrócitos, e contêm células alongadas, que formam um epitélio estratificado simples. O âmnion é uma membrana transparente, ou esbranquiçada; constituído por epitélio estratificado simples. A diferença principal entre o alantóide e o âmnion é que o último é avascular. O saco vitelínico é uma membrana opaca que desaparece durante a gestação; é a única membrana que não está em contato com as outras e apresenta três tipos diferentes de células que dão forma a três camadas distintas (endoderma, mesotélio, mesênquima).

Morfoanatomia do sistema subterrâneo de Smallanthus sonchifolius (Poepp. & Endl.) H. Robinson (Asteraceae)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Placentation in the capybara (Hydrochaerus hydrochaeris), agouti (Dasyprocta aguti) and paca (Agouti paca)

Placentae of three hystricimorph rodents-capybara, agouti and paca-were examined by conventional histology, immunohistochemistry for cytokeratin and vimentin, and TUNEL staining. The placentae were divided into lobules of labyrinthine syncytium separated by interlobular and marginal trophoblast. The subplacenta comprised cytotrophoblasts, supported on lamellae of allantoic mesoderm, and syncytiotrophoblast. The central excavation was still apparent in the definitive placenta of capybara. In agouti and paca, the decidua of the junctional zone formed a mesoplacenta comprising a capsule and a pedicle. Towards term the pedicle formed a tenuous attachment between placenta and uterine wall comprising a few maternal vessels surrounded by degraded tissue. In paca placenta, it was shown by TUNEL staining that breakdown of this tissue occurred by apoptosis. The visceral yolk sac was highly villous and, in agouti, the yolk sac villi were extremely long. Lateral to its attachment to the placenta, the fetal surface was covered with non-vascular yolk sac endoderm. A layer of spongiotrophoblast cells was interposed between the endoderm and the marginal trophoblast. (C) 2002 Elsevier B.V. Ltd. All rights reserved.