[ effects of trifluralin on LH, FSH, and testosterone levels and testis histological changes in adult rats]

Trifluralin is a dinitroaniline herbicide and is used for controlling of annual grasses and broadleaf weeds in agriculture. It enters into the plants through developing roots and stops plant cells from dividing and elongation [meristemic inhibitor]. According to entrance of this herbicide into the body via fresh vegetables, in this research the effects of trifluralin on reproductive parameters of the male wistar rats including serum LH, FSH, testosterone, and changes in testicular tissue and body weight were investigated. In this experimental study, forty male wistar rats with weight of 250 +/- 5g were randomly divided in 5 groups, including control, sham [received normal saline as a solvent], and three experimental groups receiving 500, 1000 and 2000 mg/kg oral trifluralin, respectively. It was administered orally once a day for 16 days. After 16 days, body and testis weight were measured and blood samples were taken from heart and used for measurement of LH, FSH and testosterone levels. To evaluate histological changes, testes were removed and weighed and, after obtaining tissue section, stained by Hematoxilin-Eosine. Serum testosterone, FSH, and LH levels showed significant decrease in experimental groups [p<0.001, p<0.05]. There was significant decrease in the number of germinal and somatic cells in testis in experimental groups [p<0.05]. There was also a significant decrease in body and testis weight, and concentration of sperms in experimental groups [p<0.05]. Oral administration of trifluralin could decrease gonadotropins and testosterone levels and impair steroidogenesis and spermatogenesis by oxidative stress and production of free radicals

Rat liver perfusion and hepatocytes isolation

Perfused rat liver and isolated hepatocytes are useful to study specific metabolism of this organ. Traditionally, mechanical, chemical and enzymatic methods have been used to isolate hepatocytes, and it appears that a mixed method is the preferred choice. The rat liver perfused in situe in three subsequent steps, 10 minutes with Krebs-Ringer solution containing ethylene diamine tetra-acetic acid [EDTA, 2mM] without calcium and magnesium, and 1 minute with the same solution, however, without EDTA in open non-recirculatory pathway. This followed by 10 minutes with Krebs-Ringer solution, containing collagenase [200 IU/mL] in closed recirculatory system. The hepatocytes were isolated, collected and incubated [6-8 X 10[6] cells/mL] for 3-hours in Krebs-Ringer solution, under the atmosphere of O[2]:CO[2] [95:5]. The cell viability was assessed with tripan blue exclusion and lactate dehydrogenase [LDH] leakage. Assay of LDH activity, the marker of cytosolic enzyme, show that more than 92% and 88% of the enzyme had been retained in the cells, with less than 8% and 12% leakage from the damaged cells in the beginning and end of incubation, respectively. Microscopic evaluation showed that the majority of the cells had intact plasma membrane without vacuoles. When the cells were stained with dye, less than 5% of vital dye included these cells. The results showed that perfusion of the rat liver with the solution containing calcium chealator and collagenase can yield about 10 mL of hepatocytes with a viability of more than 90%