Identification of differentially expressed genes in the testis of Sprague-Dawley rats treated with di(n-butyl) phthalate.
The aim of this study was to identify the di(n-butyl) phthalate (DBP)-induced differentially expressed genes (DEGs) using a novel annealing control primer system in the testes of Sprague-Dawley male rats. Animals (4 weeks of age) were administered orally either corn oil only (vehicle control) or DBP (250, 500, or 750 mg/kg/day) for 30 days. Total RNA was isolated from the rat testes and GeneFishing PCR was used to determine the differential gene expression levels. Using this technique, a total of 59 DEG mRNA fragments were observed in the testes treated with DBP 750 mg/kg/day compared to vehicle control. Of these 59 genes, 31 genes were significantly altered after exposing rats to high dose DBP (750 mg/kg/day), and their sequences cloned. Based on the Basic Local Alignment Search Tool (BLAST), 4 expressed sequence tags (EST), 27 cloned genes (Insl3, pgrp, H1SHR, etc.) and 3 genes (LDHA, lactate dehydrogenase A; Spag4, sperm associated antigen 4 and PBR, peripheral-type benzodiazepine receptor) were found to be involved in spermatogenesis and steroidogenesis. In addition, the expression patterns of the steroidogenesis-related genes such as scavenger receptor class B-1 (SR-B1), steroidogenic acute regulated protein (StAR), P450 side chain cleavage (P450scc), CYP17, and CYP19 were further analyzed by RT-PCR. Significant increases in the mRNA levels of steroidogenesis-related genes (PBR, SR-B1, StAR, P450scc, and CYP17) were observed in the high dose DBP-treated rats. However, DBP significantly decreased the CYP19 mRNA levels compared with controls. DBP (750 mg/kg/day) significantly increased the TR-alpha1 and PPARgamma expression in testes, whereas the AR and ERbeta protein levels were significantly reduced in the same group. These data indicate that the steroidogenesis- or spermatogenesis-related genes identified in this study may provide insights into the molecular mechanisms underlying environmental pollutants-mediated male infertility.
Ryu
Ju Young
JY
Laboratory of Molecular Toxicology, College of Pharmacy, Pusan National University, San 30, Jangjun-Dong, Gumjung-Ku, Busan, South Korea.
Lee
Byung Mu
BM
Kacew
Sam
S
Kim
Hyung Sik
HS
D000284:Administration, Oral; D000367:Age Factors; D000818:Animals; D015153:Blotting, Western; D003993:Dibutyl Phthalate / Q000008:administration & dosage / Q000633:toxicity*; D004305:Dose-Response Relationship, Drug; D047629:Estrogen Receptor beta / Q000235:genetics / Q000378:metabolism; D020869:Gene Expression Profiling / Q000379:methods*; D005786:Gene Expression Regulation / Q000187:drug effects; D018776:Genes, erbA / Q000187:drug effects / Q000235:genetics; D008297:Male; D009929:Organ Size / Q000187:drug effects; D047493:PPAR alpha / Q000235:genetics / Q000378:metabolism; D010968:Plasticizers / Q000008:administration & dosage / Q000633:toxicity; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D011944:Receptors, Androgen / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013091:Spermatogenesis / Q000187:drug effects / Q000235:genetics; D013737:Testis / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology
D047629:Estrogen Receptor beta; D047493:PPAR alpha; D010968:Plasticizers; D012333:RNA, Messenger; D011944:Receptors, Androgen; D003993:Dibutyl Phthalate