Hodges J K, van Aarde R J, Heistermann M, Hoppen H O
Department of Reproductive Biology, German Primate Centre, Göttingen.
J Reprod Fertil. 1994 Sep;102(1):163-8. doi: 10.1530/jrf.0.1020163.
The aim of this study was to examine the progestin content and biosynthetic potential of the corpus luteum of the African elephant (Loxodonta africana). Luteal tissue was collected from nonpregnant and early, mid- and late pregnant elephants (n = 2 per group) shot in the Kruger National Park. Pieces of individual corpora lutea (2-3 per animal; 23 in total) were stored directly in ethanol before hormone analysis. Matching tissue pieces were incubated for 2 h with [3H]pregnenolone (2 x 10(5) c.p.m.), after which tissue plus medium were also stored in ethanol. Progesterone and 17 alpha-hydroxyprogesterone immunoreactivity in tissue extracts were determined by enzymeimmunoassay and radioimmunoassay, respectively, before and after reverse phase HPLC. Progesterone immunoreactivity predominated over that of 17 alpha-hydroxyprogesterone in all corpora lutea examined but concentrations of both hormones were very low (73-374 ng g-1 and 3-93 ng g-1, respectively after HPLC). There were no obvious differences in hormone concentrations in corpora lutea from animals at different reproductive stages. Progesterone and 17 alpha-hydroxyprogesterone immunoreactivity assayed before HPLC was 10-30 times higher than that measured after chromatographic separation. HPLC consistently revealed two large immunoreactive peaks associated with relatively nonpolar compounds, which together accounted for most (at least 75%) of all progesterone immunoreactivity measured. Large amounts of radioactivity with the same retention times as these peaks were also detected after HPLC in samples incubated with [3H]pregnenolone. Analysis of conversion products from four corpus luteum incubations indicated that between 52% and 84% of [3H]pregnenolone had been converted; 19-33% was accounted for by progesterone, and 12-50% by the two substances represented by the unidentified peaks. Subsequent GCMS analysis identified the two immunoreactive peaks as 5 alpha-pregnane-3 alpha-ol-20-one and 5 alpha-pregnane-3,20-dione (5 alpha-dihydroprogesterone). These results indicate that the major progestins contained within and biosynthesized by corpora lutea of African elephants are 5 alpha-reduced metabolites, and that progesterone and 17 alpha-hydroxyprogesterone are quantitatively of minor importance.
本研究的目的是检测非洲象（Loxodonta africana）黄体的孕激素含量和生物合成潜力。从在克鲁格国家公园被射杀的未怀孕以及怀孕早期、中期和晚期的大象身上采集黄体组织（每组2头）。将单个黄体切成小块（每头动物2 - 3块；共23块），在进行激素分析前直接保存在乙醇中。将配对的组织块与[³H]孕烯醇酮（2×10⁵ 计数每分钟）一起孵育2小时，之后组织和培养基也保存在乙醇中。在反相高效液相色谱（HPLC）前后，分别通过酶免疫测定法和放射免疫测定法测定组织提取物中孕酮和17α - 羟孕酮的免疫反应性。在所检测的所有黄体中，孕酮的免疫反应性高于17α - 羟孕酮，但两种激素的浓度都非常低（HPLC后分别为73 - 374 ng g⁻¹ 和3 - 93 ng g⁻¹）。处于不同生殖阶段动物的黄体中激素浓度没有明显差异。HPLC前测定的孕酮和17α - 羟孕酮免疫反应性比色谱分离后测定的值高10 - 30倍。HPLC始终显示出两个与相对非极性化合物相关的大免疫反应峰，它们共同占所测所有孕酮免疫反应性的大部分（至少75%）。在用[³H]孕烯醇酮孵育的样品中，HPLC后也检测到与这些峰保留时间相同的大量放射性。对四次黄体孵育的转化产物分析表明，52%至84%的[³H]孕烯醇酮已被转化；19% - 33%转化为孕酮，12% - 50%转化为未鉴定峰所代表的两种物质。随后的气相色谱 - 质谱联用（GCMS）分析确定这两个免疫反应峰为5α - 孕烷 - 3α - 醇 - 20 - 酮和5α - 孕烷 - 3,20 - 二酮（5α - 二氢孕酮）。这些结果表明，非洲象黄体中所含及生物合成的主要孕激素是5α - 还原代谢产物，而孕酮和17α - 羟孕酮在数量上不太重要。
