Cohick W S, Armstrong J D, Whitacre M D, Lucy M C, Harvey R W, Campbell R M
Department of Medicine, University of North Carolina School of Medicine, Chapel Hill 27599, USA.
Endocrinology. 1996 May;137(5):1670-7. doi: 10.1210/endo.137.5.8612500.
Active immunization against GRF at 6 months of age delays puberty in beef heifers. The objectives of the present study were to determine whether active immunization against GRF at an earlier age would affect normal onset of puberty and follicular growth and to determine whether these changes were related to alterations in ovarian insulin-like growth factor I (IGF-I) or IGF binding protein (IG-FBP) messenger RNA (mRNA) levels. Heifers were immunized against human serum albumin (HSAi; n = 15) or against GRF conjugated to HSA (GRFi; n = 18) at 3 months of age. A third group of heifers was not immunized (CON; n = 16). Immunization against GRF delayed puberty beyond 13 months of age in 75% of treated heifers. Unilateral ovariectomy at 191 days of age revealed that the delay in puberty was associated with a reduction in the number of large ( > or = 7 mm in diameter) follicles. Large follicles were present in only 22% of GRFi heifers compared to 77% of HSAi heifers. The number of small ( < or = 3 mm in diameter) and medium (4 to 6 mm in diameter) follicles was not affected by GRFi. The percentage of 1- to 3-mm follicles that were atretic was not different between HSAi (65%) and GRFi (62%) heifers. Unilateral ovariectomy had no effect on age at puberty. Immunization against GRF decreased (P < 0.01) concentrations of IGF-I in serum (23 +/- 2 ng/ml) compared to HSAi heifers (109 +/- 11 ng/ml). IGF-I levels in follicular fluid (FFL) of medium and small follicles were also decreased by GRFi from 82 +/- 3 ng/ml in HSAi heifers to 48 +/- 6 ng/ml (P < 0.01). Levels of IGFBP-3 (determined by ligand blot analysis) in serum and FFL of small follicles were decreased by GRFi (P < 0.01). In contrast, IGFBP-2 serum levels were increased from 422 +/- 32 ng/ml in HSAi heifers to 657 +/- 6 ng/ml in GRFi heifers (P < 0.05). Likewise, IGFBP-2 levels in FFL from small and medium follicles were increased from 785 +/- 44 ng/ml to 926 +/- 44 ng/ml (P < 0.05). Ligand blot analysis indicated that IGFBP levels were lower in FFL from large vs. small follicles. The band intensities of IGFBP-4 and -5 were drastically reduced ( > 80%) while the decreases in IGFBP-2 and -3 were less marked ( < 50%). The decreased levels of IGFBP-5 in FFL from large follicles was not associated with an increase in proteolytic fragments detectable by immunoblot analysis. While mRNA transcripts for IGF-I, GH receptor, and IGFBP-2, -3, -4, and -5 were readily detectable in ovarian tissue, GRFi had no effect on ovarian levels of mRNA for each of these proteins. This suggests that the decrease in follicular development associated with GRFi may be related to changes in circulating IGF-I and/or IGFBPs.
6月龄时对生长激素释放因子（GRF）进行主动免疫可延迟肉用小母牛的青春期。本研究的目的是确定更早年龄对GRF进行主动免疫是否会影响青春期的正常开始和卵泡生长，以及确定这些变化是否与卵巢胰岛素样生长因子I（IGF-I）或IGF结合蛋白（IG-FBP）信使核糖核酸（mRNA）水平的改变有关。小母牛在3月龄时对人血清白蛋白（HSAi组；n = 15）或与人血清白蛋白偶联的GRF（GRFi组；n = 18）进行免疫。第三组小母牛不进行免疫（CON组；n = 16）。对GRF进行免疫使75%的受试小母牛青春期延迟至13月龄以后。191日龄时进行单侧卵巢切除显示，青春期延迟与直径大于或等于7毫米的大卵泡数量减少有关。大卵泡仅存在于22%的GRFi组小母牛中，而HSAi组小母牛中这一比例为77%。GRFi对直径小于或等于3毫米的小卵泡和直径4至6毫米的中等卵泡数量没有影响。HSAi组（65%）和GRFi组（62%）小母牛中1至3毫米卵泡的闭锁百分比没有差异。单侧卵巢切除对青春期年龄没有影响。与HSAi组小母牛（109±11纳克/毫升）相比，对GRF进行免疫使血清中IGF-I浓度降低（P < 0.01）（23±2纳克/毫升）。GRFi也使中小卵泡卵泡液（FFL）中的IGF-I水平从HSAi组小母牛的82±3纳克/毫升降至48±6纳克/毫升（P < 0.01）。GRFi使小卵泡血清和FFL中IGFBP-3水平（通过配体印迹分析测定）降低（P < 0.01）。相比之下，IGFBP-2血清水平从HSAi组小母牛的422±32纳克/毫升增加到GRFi组小母牛的657±6纳克/毫升（P < 0.05）。同样，中小卵泡FFL中的IGFBP-2水平从785±44纳克/毫升增加到926±44纳克/毫升（P < 0.05）。配体印迹分析表明，大卵泡FFL中的IGFBP水平低于小卵泡。IGFBP-4和-5的条带强度大幅降低（> 80%），而IGFBP-2和-3的降低不太明显（< 50%）。大卵泡FFL中IGFBP-5水平的降低与免疫印迹分析可检测到的蛋白水解片段增加无关。虽然IGF-I、生长激素受体以及IGFBP-2、-3、-4和-5的mRNA转录本在卵巢组织中易于检测到，但GRFi对这些蛋白质中每一种的卵巢mRNA水平没有影响。这表明与GRFi相关的卵泡发育减少可能与循环IGF-I和/或IGFBPs变化有关。
