Antiproliferative mechanisms of cytoplasmic fraction and arginine deiminase from Lactococcus lactis ssp. lactis on a human stomach cancer cell line

Abstract

Lactic acid bacteria are known to have antitumor activity, but the underlying mechanisms remain unclear. Our previous study have demonstrated that cytoplasmic fraction of Lactococcus lactis ssp. lactis (L.lac CF) inhibited the proliferation of SNUC2A human colon cancer cell line (SNUC2A cells), through induction of S phase accumulation. The present study investigated if the antiproliferative mechanisms of L.lac CF on SNU-1 human stomach cancer cell line (SNU-1 cells) linked to the induction of cell cycle arrest and apoptosis, and to find potential antiproliferative compounds in L.lac CF. The proliferation of SNU-1 cells was inhibited by the treatment of L.lac CF in a time- and dose-dependent. In addition, L.lac CF showed a significantly more relevant G0/G1 cell cycle arrest, which associated increase of p21 and reduction of cyclin D1 and phosphorylation level of Rb. Furthermore, L.lac CF induced apoptosis of SNU-1 cells, but not SNU-C2A cells, as evidenced by nuclei condensation, increase of sub-G1 peak, and DNA fragmentation. In particular, p53 and bcl-2 appears to play a critical role in the apoptosis of SNU-1 cells. We further investigated if arginine deiminase (ADI) is involved in the antiproliferative activity of L.lac CF, since arginine is known as essential amino acid for growing cells. Surprisingly, only L.lac CF, which showed significantly strong antiproliferative activity and ADI activity among tested six LAB. The concentration of L.lac CF showed linear relationship with both ADI activity and antiproliferative activity. Furthermore, addition of Larginine recover the antiproliferative activity of L.lac CF. We purified ADI from L.lac CF by subsequent fractionations, and the IC50 value of final ADI fraction was 2 µg/ml. The ADI fraction also exerted apoptosis as evidenced by nuclei condensation in cells. These results, taken together, indicate that the antiproliferative activity of L.lac CF on SNU-1 cells may be contributable to induction of G0/G1 cell cycle arrest and apoptosis, particularly the latter one by ADI.