DYNAMICS OF SERUM PROTEIN DURING THE ESTROUS CYCLE OF GOATS BRED IN BRAZIL AND NATURALLY INFECTED BY CAPRINE ARTHRITIS ENCEPHALITIS VIRUS

The aim of the present study was to evaluate variations in proteinogram occurring during the estrus cycle of animals infected or not by caprine arthritis encephalitis. Forty blood samples were collected from female goats in different phases of the estrus cycle (estrus, proestrus, metaestrus and diestrus). Samples were classified as positive (n = 5) and negative (n = 5), according to the results of the survey of antibodies against caprine arthritis encephalitis as performed by immunodiffusion in agar gel. Samples were also used in the determination of albumin, total protein and alpha, beta and gammaglobulin by means of electrophoresis and biuret method, respectively. Electrophoresis showed that estrus, proestrus, metaestrus and diestrus positive animals presented total protein mean equal to 7.96 ± 0.69, 7.67 ± 1.14, 7.77 ± 0.36, 7.48 ± 0.83; mean albumin equal to 3.22 ± 0.41, 3.11 ± 0.68, 3.30 ± 0.67, 3.28 ± 0.57; mean alphaglobulin equal to 0.70 ± 0.09, 0.70 ± 0.12, 0.69 ± 0.12, 0.61 ± 0.11; mean betaglobulin 1 equal to 0.85


INTRODUCTION
Caprine arthritis encephalitis (CAEV) is caused by a virus of the family Retroviridae, subfamily Orthoretrovirinae, genus Lentivirus (BUCHEN-OSMOND, 2004).It is a single-stranded virus that replicates by means of the production of deoxyribonucleic acid (DNA), and depends on intermediate reverse transcriptase.This kind of virus may insert its genetic material in the genome of the host (PUGH, 2002).
The virus enters monocytes and macrophages, inducing persistent infection.Restricted replication enables the virus to remain latent in monocytes, undetected by the host immune system, leading to many debilitating, severe, progressive diseases.The virus quickly replicates in the beginning of the infection, activating the immune system, which limits, but does not eliminate the virus.Infected macrophages express viral proteins similar to the antigens of the main histocompatibility complex, which are recognized by T lymphocytes and stimulate the production of cytokines.These macrophages may be more susceptible to activation, inducing lymphocyte and macrophage proliferation.Lymphocyte proliferation determines the pathological changes characteristic of CAE infection.Transmission routes reported are the ingestion of milk or colostrum from an infected dam, and horizontal transmission after a long period of contact.The virus preferentially affects joints, mammary gland, lungs and brain, where the immune system of the host tends to manifest chronic inflammation (BOHLAND & D' ANGELINO, 1999;PUGH, 2002).
CAE is one of the diseases most responsible for the limitation to the development of productive goat breeding in São Paulo and in Brazil.In 2003, the Brazilian herd was constituted of 9,581,653 head (INSTITUTO BRASILEIRO DE G EOGRAFIA E ESTATISTICA, 2004).Prevalence rates of the disease show that the agent is disseminated throughout the country, as an enzootic disease that is present in most of the geo-economic regions and herds.CAE leads to important losses, due to the discard of premature kids, mortality and decrease in productivity (BOHLAND, 1998).
Some organic changes or even the intensity of the inflammatory response may be influenced by the phase of the estrus cycle in which the animal is (HAENLEIN, 2002).The hypothesis that this phenomenon is even greater influenced by the occurrence of chronic diseases such as CAE should be further studied.
The estrus cycle of female goat lasts 21 days, in average.Its phases, proestrus, estrus, metaestrus and diestrus, last, respectively, 2 to 3 days, 30 to 40 hours, 2 to 3 days and 13 to 15 days (RAPOSO, et al., 1999).These phases occur in a cyclic and sequential manner, except for anestrus, which has to be considered in this species, for goats are seasonal polyestrous animals.
Due to the biological meaning (KANEKO,1997) and multiple functions of proteins in the body (such as being part of the cell structure, tissues, organs, hormones, enzymatic catalysts, antibodies, and others), the evaluation of serum levels of total proteins and their fractions (albumin, alphaglobulin, betagalobulin 1, betaglobulin 2 and gammaglobulin), as detected by electrophoresis, and the influence caused by the estrus cycle and/or chronic, multisystemic diseases (BIRGEL JUNIOR et al., 2001;GARCIA et al., 2002), such as caprine arthritis encephalitis, is an important tool aiding clinical diagnosis.
The objective of the present study was to evaluate the dynamics of serum proteins during the estrus cycle of female goats affected and unaffected by caprine arthritis encephalitis.

MATERIAL AND METHOD
This study used 10 female Saanen and Toggenburg female goats bred in an intensive system and fed goat food, hay and mineral supplement.
These animals were separated into two groups according to their status as determined by immunodiffusion in agar gel antibodies against caprine arthritis encephalitis virus (CUTLIP et al., 1977).
For the evaluation of serum protein dynamics, blood samples were collected by a puncture in the external jugular vein, with 10 mL Vacutainer tubes.They were centrifuged for 15 minutes and serum was separated and fractionated in two aliquots stored at -20º C until they were to be analyzed.
Total proteins were quantified in an automatic biochemical analyzer using the biuret method (KANEKO, 1997).Electrophoresis fractioning of serum proteins was performed in cellulose acetate strips, according to the techniques described by FRIEDMAN (1961).
Data on the determination of total protein and its fractions by means of electrophoresis (albumin, alphaglobulin, betaglobulin 1, betaglobulin 2 and gammaglobulin) were submitted to variance analysis (ANOVA) and to the Student Newman-Keuls test to Dynamics of serum protein during the estrous cycle of goats bred in Brazil and naturally infected by caprine arthritis encephalitis virus.compare pair of means at a 5% significance level (α = 0.05).
Statistical analyses were performed by means of the Graphpad Instat software.

RESULTS
Values observed for the different protein fractions of adult goats positive or negative for caprine arthritis encephalitis were inside the normal range expected for the species and age (BIRGEL & ARAÚJO, 1968).
Results obtained in the present study are presented in Table 1 and no influence of the period of estrus cycle was detected.

DISCUSSION
Interference of retrovirosis in humoral immune responses have been studied in other species (BIRGEL JUNIOR et al., 2001;GARCIA et al., 2002), mainly in relation to enzootic bovine leucosis (EBL).These authors studied the dynamics of serum proteins not only as markers of acute inflammatory process, but also as an indirect way to evaluate humoral response.
BOHLAND & D'ANGELINO (1999) described the clinical and epidemiological aspects of caprine arthritis encephalitis and its importance to Brazilian goat breeding.They did not identify any information on the variations observed with the estrus cycle.

Table 1 -
Average values, standard deviation and contrast between averages obtained in the goats' serum with or without falling CAEV in cycle estral different phases.State of São Paulo, 2004.