Loeb Marcia J, Jaffe Howard
Insect Biocontrol Laboratory, U.S. Department of Agriculture, Bldg 011A, Rm 214, BARC West, Beltsville, MD 20705, USA.
Arch Insect Biochem Physiol. 2002 Jun;50(2):85-96. doi: 10.1002/arch.10033.
Isolated stem cells of Heliothis virescens, cultured in vitro, were induced to differentiate by Midgut Differentiation Factors 3 and 4. These were peptides identified from a chymotrypsin digest of hemolymph taken from newly pupated Lymantria dispar. Partial purification was obtained by filtration through size exclusion filters. The most active preparation was subsequently subjected to a series of 3 Reverse Phase-HPLC procedures. Partial sequences of the peptides were identified via automated Edman degradation as the nanomers EEVVKNAIA-OH (MDF 3) and ITPTSSLAT-OH (MDF 4). These sequences were commercially synthesized. The synthetic compounds proved active in a dose-dependent manner. Stem cells responded to synthetic MDF 3 and MDF 4 as they did to previously identified peptides MDF 1 and 2, which have quite different amino acid sequences. All of the 4 MDFs administered singly induced statistically similar differentiation responses at 2 x 10(-8), 2 x 10(-9), and 2 x 10(-10) M. However, pairs of the 4 MDFs produced even more differentiation, the same response as one alone, no response, or were inhibitory, dependent on the MDF pair and its concentration. The data suggests complicated receptor interactions.
将体外培养的烟芽夜蛾分离干细胞，用中肠分化因子3和4诱导其分化。这些因子是从新化蛹的舞毒蛾血淋巴的胰凝乳蛋白酶消化物中鉴定出的肽。通过尺寸排阻滤器过滤进行部分纯化。随后，将活性最高的制剂进行一系列3次反相高效液相色谱程序。通过自动埃德曼降解法鉴定出这些肽的部分序列为纳米体EEVVKNAIA-OH（MDF 3）和ITPTSSLAT-OH（MDF 4）。这些序列进行了商业合成。合成化合物显示出剂量依赖性活性。干细胞对合成的MDF 3和MDF 4的反应与它们对先前鉴定的具有完全不同氨基酸序列的肽MDF 1和MDF 2的反应相同。单独施用的所有4种MDF在2×10^(-8)、2×10^(-9)和2×10^(-10) M时诱导出统计学上相似的分化反应。然而，4种MDF的组合产生的分化更多，与单独一种产生相同反应、无反应或具有抑制作用，这取决于MDF组合及其浓度。数据表明存在复杂的受体相互作用。
