Flow cytometric analysis of apoptosis in mouse thymocytes by galectin-3

Kim, Tae-jung; Woo, Hee-jong

Korean Journal of Veterinary Research 1999;39(6):1112-1118.

Tae-jung Kim, Hee-jong Woo

Laboratory of Immunology, Department of Microbiology, College of Veterinary Medicine, Seoul National University

유세포 분석기를 이용한 galectin-3에 의해 유도된 흉선세포의 apoptosis 분석

김태중, 우희종

서울대학교 수의과대학

Abstract

Galectin-3 plays an important role in cell development, differentiation and cancer metastasis, including cell-cell/extracellular matrix (ECM) interactions and is supposed to have an effect of apoptosis on T-cells in thymic clonal selection. In this study, to know the effect of galectin-3 on thymocyte development, we used recombinant human galectin-3 (rHgal-3) from Escherichia coli, JM105, which was inserted with human gal-3 gene-transformed plasmid vector (prGal-3) to express human galectin-3. Expressed rHgal-3 was confirmed by western blot using the culture supernant of hybridoma (M3/38) producing monoclonal antibody to human galectin-3. Sepharose gel affinity chromatography was used to purify the expressed rHgal-3. Thymocytes and hepatocytes from 6-week-old male BALB/c mice were incubated with rHgal-3 and showed marked increase of apoptotic population on analysis using flow cytometry with 7-AAD in a dosedependent manner. However, rHgal-3 failed to induce apoptosis on hepatocytes. Interestingly, this apoptotic effect was not inhibited by lactose, a specific lectin domain inhibitor. From these results, we concluded that extrinsic -3 induces apoptosis on mouse thymocytes, and galectin-3 may have an apoptotic effect on T-cells in thymic clonal selection.

Key Words: galectin-3, apoptosis, thyrnocytes, negative selection, FACS