Poster abstracts.

V International Symposium on Animal Biology of Reproduction, October 8-11, 2014, Campinas, SP,
Brazil.

Protein profile of rete testis fluid from Morada Nova rams

S.D. Sousa1, M.F. Van Tilburg1, R.V. Oliveira1, T.A. Silva1, R.B.F. Melo1, A.A. Araújo2, A.A.A.N. Moura1
1
Laboratory of Animal Physiology, Department of Animal Science, Federal University of Ceará; 2State University of Ceará,
Fortaleza, CE, Brazil.

Seminal plasma is a complex secretion composed by fluids from accessory sex glands, epididymis and testicles, and
its molecular composition is capable of modulating sperm function. Proteins are the most abundant organic
compounds in seminal plasma and play a crucial role in processes related to fertilizing capacity of sperm, and can be
considered as potential molecular markers of fertility. The rete testis fluid consists of the secretions of the
seminiferous tubules and epithelial cells of the rete testis and serves partly as a vehicle for movement of sperm in the
epididymis. Few studies have been conducted on the protein profile of this fluid and its functional significance
remains largely speculative. The aim of the study was to analyze the protein profile of rete testis fluid. The testes
were obtained from five slaughtered Morada Nova rams. The testes were dissected and the head of the epididymis
was separated to gain access to the efferent ducts. The fluid from the efferent ducts was obtained by testis massage
and then collected using a pipette. Thereafter, the fluid was subjected to two centrifugations: 1500 x g for twenty
minutes at 4°C and 5000 x g for thirty minutes at 4°C, respectively. The supernatant was collected and precipitated
with acetone for two hours at -20°C. It was then centrifuged at 5000 x g for forty minutes at 5°C, and then,
supernatant was discarded and the pellet was dried overnight in a freezer at 4°C. Posteriorly, the pellet was
resuspended in sample buffer. Protein concentration of the rete testis fluid was determined by the Bradford assay.
Four hundred micrograms of each sample were subjected to isoelectric focusing in strips of 13 cm (pH 4-7). The
second dimension was conducted on SDS-PAGE 15%. The obtained gels were scanned with an ImageScanner II
(GE Lifesciences, USA) and analyzed using the PDQuest® version 8.0.1 (Bio-Rad Laboratories, USA). In the gels
237 ± 46.4 spots (mean ± SD) were detected, where 66.2% of the proteins were found above 40 kDa, and 55.2% of
the proteins in the pH range between 5.0 and 5.9. The fourteen more intense spots with a molecular mass between 66
and 70kDa and pH range between 5.4 and 6.6 corresponded to 25% of total proteins. Based on approximated
molecular protein weights, those proteins could correspond to isoforms of albumin (68 kDa), which was also
identified by Fritz et al. (1987) in the rete testis fluid in levels ranging from 11% to 17% of total protein. Our results
show that the rete testis fluid from Morada Nova rams has a greater amount of high molecular weight proteins and
albumin is probably the major protein in this fluid.

Financial support: UFC, Funcap and CNPq

E-mail: arlindo.moura@gmail.com

178 Anim. Reprod., v.12, n.1, p.178, Jan./Mar. 2015