Dibutyl phthalate affects the recovery, size, and viability of pig testicular tissue ectopically grafted in immunocompromised mice

Abstract

Phthalic acid esters, or phthalates, are used as plasticizers and are present in polyvinyl chloride products, solvents, lubricating oils, and personal care products. Phthalates are not covalently bound to these products and easily leach into the environment, resulting in ubiquitous exposure of humans and animals. Phthalates have been shown to have anti‐androgenic effects in rodents, which makes it an important disruptor of testicular maturation and function. Some studies have been performed in larger mammals, including primates; however, the results are still inconclusive. The studies that are necessary to elucidate the effects of phthalates on the testes of non‐rodent species are logistically difficult and ethically prohibitive. One alternative to assess the effects of these compounds on the testes of non‐rodent species is using mice as bioincubators to grow testicular tissue of large animals and then expose these mice to those compounds. In this technique, small pieces of immature testes transplanted from large animals into adult castrated immunocompromised mice, survive and fully mature. In order to investigate the effects of this compound on the testes of other species, we grafted piglet testicular pieces (~1 mg, 8 pieces / mice) under the skin of adult castrated NCr nude mice. After two weeks of grafting, mice (n=10) were treated daily by oral gavage for 26 weeks (the approximate time in which pig testicular tissue reaches full maturity in mice) with 10 mg/kg of Dibutyl phthalate (DBP) in corn oil as vehicle. As control, we exposed an equal number of mice to the same volume of corn oil. The experimental dose of DBP used is equivalent to dose exposures that may be encountered by humans (for example, children exposed to medical supplies in the intensive care unit). At sacrifice, neither the weight of mice nor their liver were affected by treatment (P>0.05), indicating that effects of treatment on the grafted tissue were a result of direct DBP exposure and not due to indirect systemic toxicity induced by this compound. The rate of graft recovery was affected by treatment; 26.78 vs 58.33 % of the transplanted pieces were recovered from the treated and control mice, respectively (P<0.05). The graft weight was also significantly reduced by DBP; 17.16 ± 4.33 vs 43.62 ± 5.36 mg (P<0.05). Moreover, the rate of graft viability (defined as the presence of viable seminiferous tubules in the graft) was significantly reduced in treated mice as well (30.00 vs 70.00 % in treated and control groups, respectively, P<0.05). Some seminiferous tubules in grafts of both groups of mice reached full maturity (full spermatogenesis; presence of elongated spermatids in the seminiferous epithelium), the number of these tubules was not affected by DBP; 6.41 vs 8.89 % (P>0.05). In summary, exposure to DBP at a dose that mimics possible environmental exposures, reduced the recovery, size, and viability of the piglet testicular tissue ectopically grafted into the mice, but not its maturity. Future studies will be aimed to determine the effects of DBP on the function of the grafts (e.g., steroidogenesis). These results suggests that the testes of non‐rodent animals are also susceptible to the detrimental effects observed in rodents after expose to DBP.Support or Funding InformationMidwestern University intramural funds