Biocompatible adipose extracellular matrix and reduced graphene oxide nanocomposite for tissue engineering applications.

Despite the immense need for effective treatment of spinal cord injury (SCI), no successful repair strategy has yet been clinically implemented. Multifunctional biomaterials, based on porcine adipose tissue-derived extracellular matrix (adECM) and reduced graphene oxide (rGO), were recently shown to stimulate in vitro neural stem cell growth and differentiation. Nevertheless, their functional performance in clinically more relevant in vivo conditions remains largely unknown. Before clinical application of these adECM-rGO nanocomposites can be considered, a rigorous assessment of the cytotoxicity and biocompatibility of these biomaterials is required. For instance, xenogeneic adECM scaffolds could still harbour potential immunogenicity following decellularization. In addition, the toxicity of rGO has been studied before, yet often in experimental settings that do not bear relevance to regenerative medicine. Therefore, the present study aimed to assess both the in vitro as well as in vivo safety of adECM and adECM-rGO scaffolds. First, pulmonary, renal and hepato-cytotoxicity as well as macrophage polarization studies showed that scaffolds were benign invitro. Then, a laminectomy was performed at the 10th thoracic vertebra, and scaffolds were implanted directly contacting the spinal cord. For a total duration of 6 weeks, animal welfare was not negatively affected. Histological analysis demonstrated the degradation of adECM scaffolds and subsequent tissue remodeling. Graphene-based scaffolds showed a very limited fibrous encapsulation, while rGO sheets were engulfed by foreign body giant cells. Furthermore, all scaffolds were infiltrated by macrophages, which were largely polarized towards a pro-regenerative phenotype. Lastly, organ-specific histopathology and biochemical analysis of blood did not reveal any adverse effects. In summary, both adECM and adECM-rGO implants were biocompatible upon laminectomy while establishing a pro-regenerative microenvironment, which justifies further research on their therapeutic potential for treatment of SCI.

Widespread occurrence of chitinase-encoding genes suggests the Endozoicomonadaceae family as a key player in chitin processing in the marine benthos.

Chitin is the most abundant natural polymer in the oceans, where it is primarily recycled by chitin-degrading microorganisms. Endozoicomonadaceae (Oceanospirillales) bacteria are prominent symbionts of sessile marine animals, particularly corals, and presumably contribute to nutrient cycling in their hosts. To reveal the chitinolytic potential of this iconic, animal-dwelling bacterial family, we examined 42 publicly available genomes of cultured and uncultured Endozoicomonadaceae strains for the presence of chitinase-encoding genes. Thirty-two of 42 Endozoicomonadaceae genomes harbored endo-chitinase- (EC 3.2.1.14), 25 had exo-chitinase- (EC 3.2.1.52) and 23 polysaccharide deacetylase-encoding genes. Chitinases were present in cultured and uncultured Endozoicomonadaceae lineages associated with diverse marine animals, including the three formally described genera Endozoicomonas, Paraendozoicomonas and Kistimonas, the new genus Candidatus Gorgonimonas, and other, yet unclassified, groups of the family. Most endo-chitinases belonged to the glycoside hydrolase family GH18 but five GH19 endo-chitinases were also present. Many endo-chitinases harbored an active site and a signal peptide domain, indicating the enzymes are likely functional and exported to the extracellular environment where endo-chitinases usually act. Phylogenetic analysis revealed clade-specific diversification of endo-chitinases across the family. The presence of multiple, distinct endo-chitinases on the genomes of several Endozoicomonadaceae species hints at functional variation to secure effective chitin processing in diverse micro-niches and changing environmental conditions. We demonstrate that endo-chitinases and other genes involved in chitin degradation are widespread in the Endozoicomonadaceae family and posit that these symbionts play important roles in chitin turnover in filter- and suspension-feeding animals and in benthic, marine ecosystems at large.

Interfacing reduced graphene oxide with an adipose-derived extracellular matrix as a regulating milieu for neural tissue engineering.

Enthralling evidence of the potential of graphene-based materials for neural tissue engineering is motivating the development of scaffolds using various structures related to graphene such as graphene oxide (GO) or its reduced form. Here, we investigated a strategy based on reduced graphene oxide (rGO) combined with a decellularized extracellular matrix from adipose tissue (adECM), which is still unexplored for neural repair and regeneration. Scaffolds containing up to 50 wt% rGO relative to adECM were prepared by thermally induced phase separation assisted by carbodiimide (EDC) crosslinking. Using partially reduced GO enables fine-tuning of the structural interaction between rGO and adECM. As the concentration of rGO increased, non-covalent bonding gradually prevailed over EDC-induced covalent conjugation with the adECM. Edge-to-edge aggregation of rGO favours adECM to act as a biomolecular physical crosslinker to rGO, leading to the softening of the scaffolds. The unique biochemistry of adECM allows neural stem cells to adhere and grow. Importantly, high rGO concentrations directly control cell fate by inducing the differentiation of both NE-4C cells and embryonic neural progenitor cells into neurons. Furthermore, primary astrocyte fate is also modulated as increasing rGO boosts the expression of reactivity markers while unaltering the expression of scar-forming ones.

Decreasing sperm quality in mice subjected to chronic cannabidiol exposure: New insights of cannabidiol-mediated male reproductive toxicity.

Cannabidiol (CBD) is a natural cannabinoid present in the Cannabis sativa plant, widely prescribed as an anticonvulsant drug, especially for pediatric use. However, its effects on male reproduction are still little investigated. Therefore, the present study assessed the effects of CBD on the spermatogenesis and sperm quality. For this, twenty-one-day-old Swiss mice received CBD for 34 consecutive days by gavage at doses of either 15 or 30 mg/kg. Chronic exposure to CBD decreased the frequency of stages VII-VIII and XII of spermatogenesis and an increase in the frequency of stage IX were noted. Furthermore, the seminiferous epithelium height reduced at stage IX and increased at stage XII in both CBD-treated groups. There was a significant rise of sperm DNA damage, while no genotoxic effects were observed in leukocytes. The activities of superoxide dismutase and catalase decreased, while malondialdehyde levels increased in the sperm of mice treated with a higher dose of CBD. Mice exposed to 30 mg/kg of CBD showed a reduction in the mobile spermatozoa percentage and in curvilinear velocity, while straight line and average path velocity decreased in both treated groups. The number of acrosome-intact spermatozoa declined in the CBD 30 group, and the number of abnormal acrosomes raised in both CBD groups. On the other hand, the weight of reproductive organs, sperm count, and hormone levels were not affected by CBD treatment. These findings show that dysregulation of the endocannabinoid system by CBD can reduce sperm quality. The mechanisms responsible may be associated with disorders during spermatogenesis, especially during the final stages of nuclear remodelling and assembly of acrosome. However, changes in mitochondrial function, as well as the reduction on the antioxidant enzyme activities during epididymal transit, at least partly, may also be involved.

Characterization of the mechanical and biological properties of a new alumina scaffold.

PURPOSE: In this work, an alumina scaffold was produced through a new method to be used in a near future as a bone substitute. METHODS: In vitro and in vivo studies were performed in order to characterize the mechanical and biological properties of the scaffold. RESULTS: The results obtained showed that this scaffold has high mechanical resistance and a porous surface that allows human osteoblast cells to adhere and proliferate. The in vivo studies revealed no systemic reaction. CONCLUSIONS: The alumina scaffold produced herein has the mechanical and biological properties that are compatible with its application in bone therapy.

Influence of land use changes on water chemistry in streams in the State of São Paulo, southeast Brazil.

Streamwater is affected by several processes in the watershed including anthropogenic activities that result in changes in water quality as well as in the functioning of these stream ecosystems. Therefore, this work aims to evaluate the concentration of major ions (Ca(2+), Mg(2+), Na(+), K(+), NH4(+), NO3(-), NO2(-), Cl(-), SO4(2-), PO4(3-), HCO3(-)) in streams in the state of São Paulo (southeast Brazil). The sampling sites are located at undisturbed (ombrophilus dense forest, semideciduous forest and savanna - cerrado) and disturbed areas (pasture, urbanization and sugar cane crops). Streamwater chemistry varied according to land use change and, in general, was higher in disturbed sites. Streams located in undisturbed sites at Ribeira de Iguape/Alto Paranapanema watershed (streams 1, 2 and 3) seem to be regulated by soil characteristics, as the disturbed streams located at the same watershed covered by pasture (stream 7) showed high concentration for the most of the variables. Exception to streams located at Pontal do Paranapanema watershed where both disturbed (stream 8) and undisturbed streams (stream 4 and 5) presented similar patterns for almost all variables measured.

Challenges in clinical and laboratory diagnosis of androgen insensitivity syndrome: a case report.

INTRODUCTION: Androgen is a generic term usually applied to describe a group of sex steroid hormones. Androgens are responsible for male sex differentiation during embryogenesis at the sixth or seventh week of gestation, triggering the development of the testes and penis in male fetuses, and are directed by the testicular determining factor: the gene SRY (sex determining region on Y chromosome) located on the short arm of chromosome Y. The differentiation of male external genitalia (penis, scrotum and penile urethra) occurs between the 9th and 13th weeks of pregnancy and requires adequate concentration of testosterone and the conversion of this to another more potent androgen, dihydrotestosterone, through the action of 5α-reductase in target tissues. CASE PRESENTATION: This report describes the case of a teenage girl presenting with a male karyotype, and aims to determine the extension of the mutation that affected the AR gene. A Caucasian girl aged 15 was referred to our laboratory for genetic testing due to primary amenorrhea. Physical examination, karyotype testing and molecular analysis of the androgen receptor were critical in making the correct diagnosis of complete androgen insensitivity syndrome. CONCLUSIONS: Sex determination and differentiation depend on a cascade of events that begins with the establishment of chromosomal sex at fertilization and ends with sexual maturation at puberty, subsequently leading to fertility. Mutations affecting the AR gene may cause either complete or partial androgen insensitivity syndrome. The case reported here is consistent with complete androgen insensitivity syndrome, misdiagnosed at birth, and consequently our patient was raised both socially and educationally as a female. It is critical that health care providers understand the importance of properly diagnosing a newborn manifesting ambiguous genitalia. Furthermore, a child with a pseudohermaphrodite phenotype should always undergo adequate endocrine and genetic testing to reach a conclusive diagnosis before gender is assigned and surgical interventions are carried out. Our results show that extreme care must be taken in selecting the genetic tools that are utilized for the diagnosis for androgen insensitivity syndrome.

[Development and reproduction of Sipha flava (Forbes) (Hemiptera: Aphididae) at different temperatures]. / Desenvolvimento e reprodução de Sipha flava (Forbes) (Hemiptera: Aphididae) em diferentes temperaturas.

The aphid Sipha flava (Forbes) is a pest on elephant grass, but little is known about its biology. The objective of this work was to evaluate the temperature effects on the development, survival and reproduction of S. flava fed on Pennisetum purpureum. Twelve-hour-old nymphs were individualized on sections of elephant grass blades and maintained at 12, 16, 20, 24, 28 and 32 degrees C+/-1 degrees C, UR 70+/-10% and 12 h photophase. A total of 150 nymphs were used per treatment divided in 30 replicates, using a totally random design. The lower threshold temperatures (LTT) for first, second, third and fourth instars were 0.83, 1.05, 3.01 and 4.98 degrees C, respectively, indicating a change in thermal requirements as the development progress. The LTT for the whole nymphal stage was 2.08 degrees C, pointing to the tolerance of this species to low temperatures. A significant reduction in survival was observed at high temperatures (28 and 32 degrees C). Although the reproductive periods were longer and insects lived longer at 12 degrees C as compared with those at higher temperatures, the total fecundity was substantially reduced. The overall life cycle duration was almost twice as long at 12 degrees C than at 24 degrees C. The greatest daily production of nymphs and greatest number of nymphs produced overall occurred at 24 degrees C. The temperatures of 20 degrees C and 24 degrees C were more suitable to S. flava development and reproduction.

Desenvolvimento e reprodução de Sipha flava (Forbes) (Hemiptera: Aphididae) em diferentes temperaturas / Development and reproduction of Sipha flava (Forbes) (Hemiptera: Aphididae) at different temperatures

O afídeo Sipha flava (Forbes) ocasiona injúrias em capim-elefante e sua biologia é pouco conhecida. O objetivo deste trabalho foi avaliar o efeito da temperatura no desenvolvimento, sobrevivência e reprodução de S. flava alimentada com Pennisetum purpureum. Ninfas com até 12h de idade foram individualizadas sobre seções foliares de capim-elefante e mantidas em condições controladas (12, 16, 20, 24, 28 e 32ºC ± 1ºC, UR 70 ± 10 por cento e fotofase de 12h). O total de 150 ninfas foi usado por tratamento, divididas em 30 repetições, em delineamento inteiramente casualizado. A temperatura base (Tb) foi de 0,83, 1,05, 3,01 e 4,98 para ninfas de primeiro, segundo, terceiro e quarto ínstares, respectivamente, indicando exigências térmicas distintas para os diferentes estádios de desenvolvimento. A Tb foi de 2,08 para a fase ninfal, verificando-se a tolerância a baixas temperaturas. Em temperaturas elevadas (28 e 32ºC), constatou-se redução significativa na sobrevivência. Embora tenha sido constatado maior período reprodutivo e longevidade a 12ºC, quando comparado às temperaturas mais altas, a fecundidade total foi substancialmente reduzida. A duração do ciclo de vida foi cerca de duas vezes maior a 12ºC em relação a 24ºC. A maior produção diária e total de ninfas foi a 24ºC. As temperaturas de 20ºC e 24ºC foram as mais favoráveis para o desenvolvimento e reprodução de S. flava.

The aphid Sipha flava (Forbes) is a pest on elephant grass, but little is known about its biology. The objective of this work was to evaluate the temperature effects on the development, survival and reproduction of S. flava fed on Pennisetum purpureum. Twelve-hour-old nymphs were individualized on sections of elephant grass blades and maintained at 12, 16, 20, 24, 28 and 32ºC ± 1ºC, UR 70 ± 10 percent and 12h photophase. A total of 150 nymphs were used per treatment divided in 30 replicates, using a totally random design. The lower threshold temperatures (LTT) for first, second, third and fourth instars were 0.83, 1.05, 3.01 and 4.98ºC, respectively, indicating a change in thermal requirements as the development progress. The LTT for the whole nymphal stage was 2.08ºC, pointing to the tolerance of this species to low temperatures. A significant reduction in survival was observed at high temperatures (28 and 32ºC). Although the reproductive periods were longer and insects lived longer at 12ºC as compared with those at higher temperatures, the total fecundity was substantially reduced. The overall life cycle duration was almost twice as long at 12ºC than at 24ºC. The greatest daily production of nymphs and greatest number of nymphs produced overall occurred at 24ºC. The temperatures of 20ºC and 24ºC were more suitable to S. flava development and reproduction.