Characterization of clonal populations of theHeliothis zea cell line IPLB-HZ 1075

Summary A total of 24 clones (HZ 1075/UND-A through X) were initially isolated by dilution plating from the established IPLB-HZ 1075 cell line. Many of the isolates with highly vacuolated cytoplasms eventually died during subculturing. The cloned cell strains differed in their predominant morphology, cell doubling times, and relative ability to support replication of the singly encapsulated nuclear polyhedrosis virus ofHeliothis zea (HzSNPV). The origin of the cloned cell strains was confirmed by comparing their isozyme profiles with those of the parental IPLB-HZ 1075 cell line andH. zea larvae using stains for fumerate hydratase, lactate dehydrogenase (LDH), and malic dehydrogenase (MDH). One dipteran and several lepidopteran cell lines maintained in our lab were also separable using stains for LDH and MDH. This research was supported in part by USDA grant number GAM 8400211, and by a Jesse Jones Faculty Research Award from the University of Notre Dame to M. J. F.     

Enhanced in vitro hair growth at the air-liquid interface: minoxidil preserves the root sheath in cultured whisker follicles

Summary Inasmuch as hair follicles are difficult to maintain in culture, the study of hair biology using cultured hair follicles has met with only limited success. In our attempts to solve the problem of follicle degeneration, we cultured follicles at the air-surface interface on a modified collagen matrix (Gelfoam). In follicles cultured at the air-surface or submerged, we examined follicular morphology, hair shaft growth, sulfotransferase levels, cysteine incorporation, an expression of a tissue inhibitor of metalloproteinase (TIMP), and ultra-high sulfur keratin (UHSK). Follicles cultured at the air-liquid interface produced a 2.7-fold increase in hair growth and maintained an anagen-like morphology. Substrates such as nylon mesh seeded with fibroblasts, Full Thickness Skin?, or 5-μm polycarbonate filter also supported hair growth, whereas Gelfilm, GF-A glass filter, filter paper, or 1-μm polycarbonate filter did not. The UHSK expression was significantly higher in the air-liquid interface cultures compared to the submerged culture. Several potassium channel openers, including minoxidil, a minoxidil analog, and the pinacidil analog (P-1075), all stimulated significant cysteine incorporation in follicles. Minoxidil and its analog specifically preserved the follicular root sheath, in contrast to P-1075 which did not, indicating a difference in the two drug types. The preservation of the root sheath was measured by increased TIMP expression and sulfotransferase activity and indicates that the root sheath is a target tissue for minoxidil. Our results show that follicles cultured at the air-liquid interface maintain a better morphology and produced greater hair growth than follicles cultured on tissue culture plastic.     

Effects of pesticides on division of two lepidopteran cell lines and onAutographa Californica MNPV development in TN368 cells

Summary The concentration of each of 10 pesticides (azinphosmethyl, captan, carbaryl, chlordimedorm, dichlorvos, dimenthoate, fenvalerate, methomyl, methyl parathion, trichlorfon) causing a 50% inhibition (ID₅₀) in cell number relative to an untreated culture for a time period equal to four cell doublings was determined for the TN368 and IPLB-HZ1075 cell lines. The range of ID₅₀ values with either of the cell lines was similar, with captan being most toxic within an ID₅₀ range of 5 to 6 μM/2×10⁵ cells/ml, and methomyl least toxic within a range of 2900 to 3200 μM/2×10⁵ cells/ml. Yet there were significant differences between cell lines in pesticide susceptibility. Fenvalerate, dichlorvos, and chlordimeform were 16, 3, and 1.5 times more toxic, respectively, for TN368 cells than HZ1075 cells, whereas dimethoate and carbaryl were each 2 times more toxic for HZ1075 cells. In general, increasing toxicity paralleled decreasing water solubility, although the order of the pesticides varied somewhat according to the particular cell line and medium. Moreover, there was little aberrant cell morphology in either of the cell cultures during incubation with most of the pesticides at their ID₅₀ levels. Preincubation of TN368 cells with any one of seven different pesticideis before inoculation withAutographa californica MNPV, and subsequent incubation of infected cells in medium plus pesticide, did not significantly suppress polyhedra development except for trichlorfon-incubated cells. In addition, there was a small but consistent variation from control cells in extracellular virus titers assayed from two of five of the pesticide incubations. The titer was consistently depressed with trichlorfon and elevated with fenvalerate, however, further work is required to determine the biological significance of these differences. Primary funding for this research was provided by the Office of Research and Development, U.S. Environmental Protection Agency, Washington, DC, under grant no. R-809453. Additional funding was provided by the Pennsylvania State University College of Agriculture Experiment Station—Project no. 2758. The contents of this publication do not necessarily reflect the views, policies, or recommendations of the Environmental Protection Agency, and the Agency does not necessarily endorse any of the commercial products used in this study. The Pennsylvania State Agricultural Experiment Station. Authorized for publication as Journal Series Paper No. 7432.