Ellis L A, Picciano M F
Department of Nutrition, Pennsylvania State University, University Park 16802, USA.
Endocrinology. 1995 Jun;136(6):2711-20. doi: 10.1210/endo.136.6.7750496.
Human milk from mothers of term (T) and preterm (PT) infants was collected during early (days 2-7), mature (2-16 weeks), or late (> 16 weeks) lactation. PRL-like bioactivity (B) was measured by Nb2 cell proliferation, and PRL immunoreactivity (I) was determined by RIA. PRL activity is reported in PRL equivalents (1 PRL equivalent = 1 ng NIDDK reference material). Milk from early lactation contained significantly greater PRL-like B compared to I (T:B, 132.5 +/- 13.0; I, 83.43 +/- 12; PT:B, 195.8 +/- 56; I, 74.45 +/- 13.7). PRL-like B and I declined as lactation progressed (T mature: B, 41.74 +/- 8.9; I, 27.19 +/- 5.5; T late: B, 17.84 +/- 5.5; I, 27.33 +/- 1.8; PT mature: B, 59.85 +/- 16; I, 45.16 +/- 4.3). Milk PRL B to I ratios were consistently greater than serum B to I ratios during early lactation (milk: T, 1.4 +/- 0.3; PT, 3.6 +/- 1.3; serum: T, 1.0 +/- 0.2; PT, 0.58 +/- 0.12). During early lactation, high PRL-like B was widely distributed among several (n = 4-6) bioactive forms differing in molecular mass [8 to > 66 kilodaltons (kDa)] in T milk, but the majority of B in PT milk was detected in two or three forms. During mature and late lactation, lower PRL-like B was associated with two or three peaks (20 to > 66 kDa). A large fraction of PRL-like B (67%-84%) was associated with the phosphorylated (P-) fraction of human milk. Four immunoreactive forms (24, 30, 32, and 40 kDa) of P-PRL were identified by immunoblot analyses. Alkaline phosphatase treatment converted the 40-kDa immunoreactive P-PRL to 24-kDa PRL, increased the B of the P-fraction by 2-fold, but did not change total PRL I detected. PRL in the Concanavalin-A-retained fraction accounted for 59-69% of PRL in milk based on RIA results. No PRL-like B was detected in the Concanavalin-A-retained fraction of human milk; however, treatment of the glycosylated fraction of milk with peptide-N-glycosidase F increased thymidine incorporation by Nb2 cells 1.67-fold compared to that in controls. The results of this study show that human milk contains considerably greater PRL-like activity than previous reports based on RIA detection. The appearance and regulation of multiple bioactive PRL variants in milk throughout the course of lactation may serve as a mechanism by which milk PRL influences neonatal development.
收集足月（T）和早产（PT）婴儿母亲在早期（第2 - 7天）、成熟（2 - 16周）或晚期（> 16周）哺乳期的母乳。通过Nb2细胞增殖测定催乳素样生物活性（B），通过放射免疫分析（RIA）测定催乳素免疫反应性（I）。催乳素活性以催乳素当量报告（1个催乳素当量 = 1 ng美国国立糖尿病、消化和肾脏疾病研究所参考物质）。与I相比，早期哺乳期的母乳中含有显著更高的催乳素样B（T：B，132.5±13.0；I，83.43±12；PT：B，195.8±56；I，74.45±13.7）。随着哺乳期的进展，催乳素样B和I下降（T成熟：B，41.74±8.9；I，27.19±5.5；T晚期：B，17.84±5.5；I，27.33±1.8；PT成熟：B，59.85±16；I，45.16±4.3）。在早期哺乳期，母乳中催乳素B与I的比值始终高于血清中B与I的比值（母乳：T，1.4±0.3；PT，3.6±1.3；血清：T，1.0±0.2；PT，0.58±0.12）。在早期哺乳期，高催乳素样B广泛分布于几种（n = 4 - 6）分子量不同[8至> 66千道尔顿（kDa）]的生物活性形式中，存在于T组母乳中，但PT组母乳中的大部分B以两三种形式被检测到。在成熟和晚期哺乳期，较低的催乳素样B与两三个峰（20至> 66 kDa）相关。催乳素样B的很大一部分（67% - 84%）与母乳的磷酸化（P -）部分相关。通过免疫印迹分析鉴定出四种免疫反应性形式（24、30、32和40 kDa）的磷酸化催乳素（P - PRL）。碱性磷酸酶处理将40 kDa的免疫反应性P - PRL转化为24 kDa的催乳素，使P部分的B增加2倍，但未改变检测到的总催乳素I。基于RIA结果，伴刀豆球蛋白A保留部分中的催乳素占母乳中催乳素的59% - 69%。在人母乳的伴刀豆球蛋白A保留部分中未检测到催乳素样B；然而，用肽 - N - 糖苷酶F处理母乳的糖基化部分后，与对照组相比，Nb2细胞的胸苷掺入增加了1.67倍。本研究结果表明，人母乳中含有的催乳素样活性比以往基于RIA检测的报告中所显示的要高得多。在哺乳期全过程中，母乳中多种生物活性催乳素变体的出现和调节可能是母乳催乳素影响新生儿发育的一种机制。
