Establishment and characterization of a novel cell line from midgut tissue of Helicoverpa armigera (Lepidoptera: Noctuidae)

Abstract

The midgut of lepidopteran larvae serves as a target for many pathogens such as Bacillus thuringiensis (Bt). Cell lines originating from midgut tissues will be very helpful tools in many research fields. However, to date, no Bt-susceptible midgut-derived cell lines are available. Here, we reported that a novel cell line, designated as HNU-Ha-MG1, was established from midgut tissue of the fourth instar larvae of Helicoverpa armigera. This cell line grew well in Grace's insect cell culture medium supplemented with 10-15% fetal bovine serum. The shape of the most cells was round or polygonal, and some tended to aggregate to form multiple cell masses. The size of the cells was 13.8 ± 1.8 μm in diameter, and the maximum density reached (2.40 ± 0.15) × 10(6) cells/ml. The population doubling time during logarithmic growth phase was 58.6 ± 7.0 h at 28°C. The number of chromosomes was about 90-130, which exhibited typical chromosome characteristics of lepidopteran cell lines. The patterns of random amplified polymorphic DNA of the cell line were different from those of Sl-HP and Hi5 cell lines which were frequently used in our laboratory. 20-Hydroxyecdysterone induced apoptosis in a very small part of cells at 2 μg/ml but did not affect expression of autophagy-related protein 8 (Atg8) and its lipidation at 36 h post-treatment. The cell line was permissive to Autographa californica nuclear polyhedrosis virus (AcMNPV) and H. armigera nuclear polyhedrosis virus (HaSNPV). This cell line was found to be susceptible to activated Cry1C at the final concentration of 0.5-1.0 μg/ml but not to the activated Cry1Ac.

Can the mammalian organoid technology be applied to the insect gut?

“…Unfortunately, robust growth of midgut insect cells is typically accompanied by cellular de-differentiation, with as its most conspicuous failure the inability to organize as a tight epithelium that separates two physiological compartments. 16,17,18 By contrast, in vitro models of the mammalian intestine are much more advanced with respect to cell type diversity, substrata resembling underlying connective tissue and devices to provide mechanical forces and stresses. 19 The discrepancy begs the question whether the huge gap in knowledge and sophistication between mammalian intestinal and insect midgut in vitro models is a mere consequence of the much greater research efforts invested in engineering appropriate intestinal in vitro models in mammals, or reflects a fundamental difference between mammals and insects (or vertebrates and invertebrates) in the regulation of tissue and organ function and structure.…”

“…Midgut in vitro models should resemble the physiology of the midgut epithelium as faithfully as possible for research on insecticide uptake and mode of action. Unfortunately, robust growth of midgut insect cells is typically accompanied by cellular de‐differentiation, with as its most conspicuous failure the inability to organize as a tight epithelium that separates two physiological compartments 16,17,18 . By contrast, in vitro models of the mammalian intestine are much more advanced with respect to cell type diversity, substrata resembling underlying connective tissue and devices to provide mechanical forces and stresses 19 .…”

Cell lines from diamondback moth exhibiting differential susceptibility to baculovirus infection and expressing midgut genes

“…Continuous cell lines are valuable tools for studying fundamental biological mechanisms that underlie complex interwoven interactions in an organism (Shao et al ., ; Arif & Pavlik, ; Zhang et al ., ; Lynn, ). To study specific mechanisms in an insect using cell lines as an in vitro system, it is desirable to have homologous cell lines of the insect and have the cell lines retaining the required biological characteristics for the study (Chen et al ., ; Li et al ., ). Continuous cell lines from P. xylostella have been established and reported, but studies of these cell lines were limited to examination of their susceptibility to baculovirus infection and sensitivity to toxins from Bt (Lee & Hou, ; Kariuki & Mcintosh, ; Kariuki et al ., ).…”

Susceptibility of a Cloned Cell Line from Helicoverpa armigera to Homologous Nucleopolyhedrovirus

“…Insect cell lines derived from the same species but from different tissues tend to differ in their growth characteristics. The population doubling time of B-5 cell line was shorter than that of other H. armigera cell lines that have been established from different tissues, NIV-HA-1195 (hemocyte)[36] and HNU-Ha-MG1 (midgut)[37] cell lines. The population doubling time for NIV-HA-1195 and HNU-Ha-MG1 cell lines were 60 h and 58.6 h, respectively.…”