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44 Gastric cancer cell micro environment modulates the nk cell efficacy in rat splenocytes
  1. A Adithan,
  2. JS John peter,
  3. HS Jeong and
  4. JH Kim
  1. Chonbuk National University, veterinary physiology, Iksan, Republic of Korea

Abstract

Background and aims Natural killer (NK) cells are specialised lymphocytes capable of counteracting pathogens (bacteria, viruses) as well as cancer cells. Unlike T lymphocytes, NK cells do not require antigen-specific recognition to act on target cells. The activation of NK cell requires the action of certain pro-inflammatory cytokines (IL-2, IL-12, IL-18, IL-21).

Methods Gastric cancer cells (SNU-484) were grown in RPMI medium with 10% heat inactivated FBS at a seeding density 1*106 cells/mL for 3 days, supernatant was concentrated 10 fold. Splenocytes were treated with 1% (v/v) SNU-484 supernatant for various periods of time.

Results Flow cytometry (FCM) results suggests that the treatment do not affect the viability of the cells during the study period, further the intracellular levels of NKP30, NKP44, granzyme B, perforin were assessed using Real time-PCR (RT-PCR) and western blot techniques. RT-PCR revealed that NK cell markers were initially down-regulated during 2 days of incubation and increased several folds higher during 5th day when compared to normal control. However, no significant changes were observed in protein expression. SNU-484 cells supernatant treated splenocytes were further analysed for cytolytic activity against K562 cell line as a target with varying (1:6, 1:12, 1:25 and 1:50) target to effector ratio for a period of 24 hour.

Figure 1
Figure 1

Cytolytic activity of splenocytes against K562 cells at 24 h with various effect to target (E:T) ratio. (A), untreated splenocytes; (B), splenocytes were treated with 1% (v/v) SNU 484 supernatant for 1 day prior to the experiment.

Figure 2
Figure 2

(A) Western blot analysis of 1% SNU 484 cell supernatant treated splenocytes for NK cell markers; (B), Normalized expression of various NK cell makers compared to β-action.

Conclusions The results suggest that the treated splenocytes have significantly increased cytolytic activity (49.4%) at the lower effector to target ratio (1:25) when compared to untreated control splenocytes (38.2%). Our results indicate that gastric cancer cell micro-environment can modulate the NK cells efficacy to act against cancer.

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