Study of differential expression of miRNAs in lung tissue of mice submitted to experimental infection by Paracoccidioides brasiliensis.

MicroRNAs (miRNAs) are small single stranded RNA sequences involved in post-transcriptional regulation of different biological and physiological processes. Paracoccidioidomycosis (PCM) is an infection caused by Paracoccidioides brasiliensis, and it is a major cause of mortality due to systemic mycoses in Brazil. To date, there have been few reports on the role of miRNAs in the immune response against fungi, especially PCM. The objective of this study was to evaluate the differential expression of miRNAs related to the inflammatory response associated with pulmonary infection by P. brasiliensis. For this purpose, lungs from BALB/c mice, intravenously infected with P. brasiliensis (2.7×107 yeast cells/ml, n = 12) and noninfected BALB/c mice (n = 8), were collected at the 28 and 56 day after infection. The lung parenchyma presented a great number of yeast cells, granulomas, and edema at 28 days and a framework of resolution of the inflammatory process after 56 days. The mRNAs gata-3, ror-γt, foxp3, and IL-6 were positively regulated at the moment at the 56 day, while the TGF-ß1 mRNA was positively regulated at both moments. The miRNAs 126a-5p, 340-5p, 30b-5p, 19b-3p, 221-3p, 20a-5p, 130a-3p, and 301a-3p, 466k presented the greatest increase in expression levels 28 days after infection, and the miRNAs let-7f-5p, let-7a-5p, 5p-26b, let-7e-5p and 369-3p, 466k presented a greater increase in levels of expression 56 days after infection. This study shows a set of differentially expressed miRNAs possibly involved in the immune response in mice during pulmonary infection by P. brasiliensis.

The production and characterization of anti-bothropic and anti-crotalic IgY antibodies in laying hens: a long term experiment.

Ophidian accidents represent a great public health problem in developing countries. Recent studies have shown that antibodies produced in laying hens could be an alternative method for producing antivenin in mammals. In this study we analyzed the production of IgY antibodies in laying hens inoculated with snake venom from the Bothrops and Crotalus genera over a 360-day period. IgY antibodies present in the serum and egg yolks were analyzed according to avidity, antigen recognition pattern and efficiency in neutralizing the venom. The levels of anti-bothropic and anti-crotalic IgY antibodies increased significantly after the third immunization, and remained at these levels until the end of the experiment. Significantly high avidity levels were observed for anti-bothropic IgY antibodies on the 142nd day and for anti-crotalic antibodies on the 232nd day after the first immunization. Anti-bothropic IgY antibodies recognized antigens with molecular masses ranging from 25 kDa to 50 kDa, whereas anti-crotalic IgY antibodies mainly recognized antigens with molecular masses of 14 kDa and 30 kDa. An increase in the antigens recognized by the antivenins was observed during the experimental period. Samples of bothropic IgY antivenin antibodies presented an efficiency of 290 µl/3 DL50, a potency of 0.307 mg/ml and a specific activity of 0.230. Samples of anti-crotalic IgY antibodies presented an efficiency of 246 µl/4 DL50, a potency of 0.829 mg/ml and a specific activity of 0.271. These results show that the administration of successive doses of the venoms for more than 6 months results in an antivenin with higher avidity that is able to recognize a greater number of antigens present in the venoms. These characteristics indicate a more efficient and potent antivenin than what has been described in other studies.