|Blackburn, Michael - Mike|
Submitted to: In Vitro Cellular And Developmental Biology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/5/2003
Publication Date: 7/5/2003
Citation: Smagghe, J., Elsen, K., Loeb, M.J., Gelman, D.B., Blackburn, M.B. 2003. Effect of a fat body extract on larval midgut cells and growth of lepidoptera. In Vitro Cellular And Developmental Biology. Vol. 39 pg. 8-12 Interpretive Summary: Caterpillars of the Gypsy Moth, Lymantria dispar, and the cotton leafroller, Spodoptera littoralis cause considerable damage to shade trees and to vegetable crops. All of the plant matter that they eat enters that portion of the digestive tract know as the midgut, where it is digested and absorbed into the body to sustain the life of the caterpillars. Therefore, an agent that will disrupt the midgut will be important to pest control. We had found that an extract of pupal fat body tissue (FBX) is needed for midgut cells to multiply in tissue culture if a small amount of insect molting hormone is included in the growing medium. When the extract is fed or injected into the caterpillars, they stop feeding and die. In this work we found that FBX causes midgut cells to lose their normal organization and to multiply greatly, disrupting gut function. In tissue culture, midgut stem cells from the Gypsy Moth multiply in response to FBX and its purified active ingredient, but will also multiply in response to larger amounts of the insect molting hormones as well. It is possible that these compounds work together in the intact animal. Understanding this process may lead to a novel method for treating unwanted species of caterpillars in the field. However, at this point, only scientists will be able to use this information in order to develop practical methods for use.
Technical Abstract: An extract of fat body (FBX) prepared from green fat body tissue from newly ecdysed pupae of Manduca sexta must be added to cultures with a very low (1 pg/ l) titer of insect molting hormone (20-hydroxyecdysone, 20E) in order to induce midgut stem cells to multiply in vitro. However, FBX fed or injected into insect pests Lymantria dispar or Spodoptera littoralis larvae caused death. In this work, we showed that the cells in the midguts of these larval species receiving FBX had lost normal organization, hypertrophied, and formed disorganized masses of cells. In addition, the external cuticle of S. littorialis doubled, indicating that an abnormal form of metamorphosis was brought about by the FBX. Normal molting could be induced by approximately 1000 times more 20E than present in the FBX, indicating that the 20E derived from the FBX was far from sufficient to induce molting in S. littoralis larvae. Midgut stem cells from L. dispar, the only cell type able to undergo mitosis in midgut, were cultured in vitro and incubated with 20E, ecdysone, the ecdysteroid agonist, RH-2485 as well as varying quantities of FBX as well as the purified active principal in FBX, the multiplication protein (MP) that induces midgut stem cell proliferation in vitro. The cells multiplied in response to all of these agents. FBX induced multiplication of stem cells in a dose dependent manner. MP was needed at a lower dose than the other agents except for the non-degradable 20E agonist, RH-2485. It is probable that MP is synergistic with molting hormone in vivo. Studies are under way to elucidate the true mechanism of action.