Title:A Polylactosamine Specific Lectin from Adenia hondala Induces Apoptosis and Necrosis in Human Epithelial Colon Cancer HT-29 Cells
Volume: 28
Issue: 10
Author(s): Shashikala R. Inamdar*, Narasimhappagari Jagadeesh, Kavita Y. Hiremath, Shivakumar Belur and Mamta Sharma
Affiliation:
- Department of Studies in Biochemistry, Karnatak University, Dharwad-580003,India
Keywords:
AHL, apoptosis, necrosis, HT-29 cells, histochemistry, ROS.
Abstract:
Background: Altered expression of N-glycans such as polylactosamine is observed in
colon cancer. AHL, a polylactosamine specific lectin from Adenia hondala from a medicinal plant
from the Passifloraceae family has been reported earlier.
Objective: The aim of the present study is to study the interaction of AHL with human colon cancer
epithelial HT-29 cells and colon cancer tissues.
Methods: Cell viability was determined by MTT [3-[4, 5- dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium
bromide] assay, while cell surface binding, apoptosis by Annexin-V-PI assay and ROS production
using DCFDA [2’,7’ - dichlorofluorescindiacetate] kit method were analysed by flowcytometry,
immunohistochemistry was performed using biotinylated AHL, protein purification by
affinity chromatography using asialofetuin-coupled Sepharose -4B column.
Results: AHL strongly binds to HT-29 cells with a Mean Fluorescence Intensity of 12.4, which
could be blocked by competing glycoprotein asialofetuin. AHL inhibits HT-29 cell growth in a
dose and time-dependent manner with IC50 of 2.5 μg/mL and differentially binds to human normal
and cancerous tissues. AHL induces apoptosis and slight necrosis in HT-29 cells with an increase
in the early apoptotic population of 25.1 and 36% for 24 h and 48 h respectively and necrotic population
of 1.5 and 4.6% at 24 h and 48 h respectively as revealed by Annexin-V-PI assay. AHL induces
the release of Reactive Oxygen Species in HT-29 cells in a dose-dependent manner.
Conclusion: To the best of knowledge, this is the first report on lectin from Adenia hondala which
is not a RIP with apoptotic and necrotic effects. These findings support the promising potential of
AHL in cancer research.
