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A Time Course Analysis of the Effects of Arachidin 1 and Arachidin 3 on a Rotavirus‐Infected Human Intestinal Cell Line
Author(s) -
Witcher Caleb Michael,
Lockwood Hannah Nicole,
NapierJameson Rebekah,
Mattila Macie N,
Strange Essence B,
Taylor Josephine,
Clack Beatrice,
MedinaBolivar Fabricio,
Ball Judith M
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.938.4
Subject(s) - rotavirus , virus , virology , cytoplasm , cell culture , electron microscope , diarrhea , biology , microbiology and biotechnology , chemistry , medicine , pathology , genetics , physics , optics
Rotavirus (RV) causes severe gastroenteritis in infants and children worldwide. To prevent RV induced diarrhea, research efforts are focusing on new vaccines and novel therapeutic approaches. Previously, our laboratory has demonstrated that arachidin 1 (A1) and arachidin 3 (A3) have significant effects on the amount of infectious RV particles released from a human intestinal cell line, HT29.f8, at 18 hours post infection (hpi). Morphometric analyses have suggested changes in programed cell death pathways. The objective of this study was to observe the effects of A1 and A3 on RV infected HT29.f8 cells at 12, 14, 16, 18 hpi. A time course morphometric analysis was performed to visualize the effects of the arachidins on cellular ultrastructure and RV particles using transmission electron microscopy (TEM) and scanning electron microscopy (SEM). Additionally, changes in ratios of nucleus to cytoplasm during the time course were determined with/without RV and arachidin treatments. RV particles (with/without envelope) in infected cells (with and without the arachidins) were measured and statistically compared to determine if the observed virus particles represented different populations. To quantify the infectious virus particles released from the RV infected cells, plaque forming assays were performed. Additionally, tunable resistive pulse sensing technology (TRPS) using the qNano system from Izon was used to quantify total infectious virus particles and determine the size distribution of the released RV particles from HT29.f8 cells with/without the arachidins during the time points of collection. qRT‐PCR was performed to further identify changes in the expression of key apoptotic and autophagic genes, and western blots assays were used to confirm changes in the expressed proteins that are essential for the two cell death pathways. The data suggests that the arachidins have a significant role in influencing HT29.f8 cells to enter apoptotic or autophagic cell death pathways, and inhibit the maturation and production of progeny RV populations. The plaque assays confirmed a change in the amount of infectious virus particles produced with the addition of thee arachidins. This implies that A1 and A3 arrest RV particles at the enveloped stage of development. Thus, these small molecules have the potential to be developed as anti‐RV therapeutic agents. Support or Funding Information This work was supported by: 1) The Office of Research and Sponsored Programs at Stephen F. Austin State University (RPS # 107552‐26112‐150); 2) The NSF‐EPSCoR (grant# EPS‐0701890; Center for Plant Powered Production‐P3).