No abstract

The maturation state of dendritic cells (DC) is regarded as a control point for the induction of peripheral tolerance or autoimmunity. Experimental autoimmune orchitis (EAO) serves as a model to investigate inflammatory-based testicular impairment, which ranks as a significant cause of male infertility. This work aimed to determine whether DC enrichment occurs organotypically in testicular draining lymph nodes (TLN) compared with LN draining the site of immunization (ILN) and thus contributes to the pathogenesis of autoimmune orchitis. In this regard, we quantified and characterized the DC from TLN and ILN in rats with EAO. Flow cytometric analysis showed a significant increase in the percentage of DC (OX62+) only in TLN from EAO rats compared with normal (N) and adjuvant control (C) groups. The number of DC from ILN and TLN expressing CD80, CD86 and major histocompatibility complex (MHC) II was comparable among N, C and experimental (E) groups at 30 and 50 days after the first immunization. However, TLN DC from EAO rats (50 days) showed an increase in mean fluorescence intensity for MHC II compared with N, C and E groups (30 days). The mRNA expression level of IL-10 and IL-12p35 was significantly upregulated in enriched DC fraction from TLN in EAO rats with no significant changes observed in ILN DC. The expression of IL-23p19 mRNA remained unchanged. Functional data, using proliferation assays showed that EAO-DC from TLN, but not from ILN, significantly enhanced the proliferation of naïve T cells compared with C-DC. In summary, our data suggest that the DC in TLN from orchitis rats are mature, present antigens to T cells and stimulate an autoimmune response against testicular antigens, thus causing immunological infertility.

the common toad Rhinella arenarum is widely distributed in Argentina, where it is utilised as an autochthonous model in ecotoxicological research and environmental toxicology. However, the lack of a reference genome makes molecular assays and gene expression studies difficult to carry out on this non-model species. to address this issue, we performed a genome-wide transcriptome analysis on R. arenarum larvae through massive RnA sequencing, followed by de novo assembly, annotation, and gene prediction. We obtained 57,407 well-annotated transcripts representing 99.4% of transcriptome completeness (available at http://rhinella.uncoma.edu.ar). We also defined a set of 52,800 highconfidence lncRNA transcripts and demonstrated the reliability of the transcriptome data to perform phylogenetic analysis. our comprehensive transcriptome analysis of R. arenarum represents a valuable resource to perform functional genomic studies and to identify potential molecular biomarkers in ecotoxicological research. Amphibians are poikilothermic vertebrates with morphological and ecological adaptations that allow them to occupy diverse terrestrial environments associated with humid ecosystems 1,2. They are the only terrestrial vertebrates that preserve free-living larvae and produce large oocytes with a transparent vitelline membrane that allows for the direct observation of the different stages of embryonic development. These characteristics have been exploited in various research areas such as toxicology, physiology, ecology, and evolution 3-7. The South American common toad Rhinella arenarum [ex. Bufo arenarum (Hensel, 1867)] is amply distributed in Argentina and breeds in shallow-water areas such as ponds and ditches 5,8,9. Amphibian research models can be easily and inexpensively established. However, only six anuran genomes are available to date: Pyxicephalus adspersus 10 , Nanorana parkeri 11 , Rana catesbeiana 6 , Rhinella marina (Bioproject: PRJEB24695, ID: 445546), Xenopus laevis 12 , and Xenopus tropicalis 13. Furthermore, several conserved morphological characteristics shared by anurans make both taxonomic classification and phylogenetic analysis difficult to perform 14. This stresses the need for combining novel genomic information with morphological and karyological data, as well as mitochondrial DNA sequencing, in order to improve accuracy in phylogenetic studies. Next Generation Sequencing (NGS) provides a cost-effective and rapid method to sequence and analyse complete genomes. However, amphibians have a very high DNA content and a large proportion of repetitive and non-coding sequences 15 ; thus, whole-genome assembly is still expensive and bioinformatically challenging. In contrast, high-throughput RNA-sequencing (RNA-Seq) is an affordable NGS technique that provides a convenient platform for transcript profiling and transcriptome sequencing in non-model amphibian species like R. arenarum 16,17. Here, we report for the first time the de novo assembly of R. arenarum transcriptome using massive RNA-Seq, followed by gen...

Arsenic (As), a natural element of ecological relevance, is found in natural water sources throughout Argentina in concentrations between 0.01 mg/L and 15 mg/L. The autochthonous toad Rhinella arenarum was selected to study the acute toxicity of As and the biochemical responses elicited by the exposure to As in water during its embryonic development. The median lethal concentration (LC50) value averaged 24.3 mg/L As and remained constant along the embryonic development. However, As toxicity drastically decreased when embryos were exposed from heartbeat-stage on day 4 of development, suggesting the onset of detoxification mechanisms. Given the environmental concentrations of As in Argentina, there is a probability of exceeding lethal levels at 1% of sites. Arsenic at sublethal concentrations caused a significant decrease in the total antioxidant potential but generated an increase in endogenous glutathione (GSH) content and glutathione S-transferase (GST) activity. This protective response might prevent a deeper decline in the antioxidant system and further oxidative damage. Alternatively, it might be linked to As conjugation with GSH for its excretion. The authors conclude that toad embryos are more sensitive to As during early developmental stages and that relatively high concentrations of this toxic element are required to elicit mortality, but oxidative stress may be an adverse effect at sublethal concentrations.