Trima Accel의 Leukoreduction Chamber 내 단핵세포의 성상분석과 수지상세포의 분화유도
Analysis of Characteristics of Mononuclear Cells Remaining in the Leukoreduction System Chamber of Trima Accel and Their Differentiation Into Dendritic Cells
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배경 : Trima Accelⓡ의 leukoreduction system (LRS) chamber 내 세포의 성상을 전혈유래 백혈구연층과 비교, 분석함으로 써 연구용으로 활용 가능성을 확인하고 수지상세포로의 분화실 험을 실시하였다. 방법 : 건강한 성인 26명으로부터 Trima Accelⓡ을 이용하여 혈소판 성분채집을 실시한 후 LRS chamber를 분리하였다. LRS chamber 내 T 세포, B 세포, NK 세포, CD14+ 세포의 구성성 분을 유세포분석기를 이용하여 분석하였으며, CD34+ 세포 수 를 측정하였다. Magnetic cell sorting (MACSⓡ) Seperation (Miltenyi Biotec Inc., USA)로 CD14+ 세포를 분리한 후 수지 상세포의 성숙을 유도하였다. 결과 : LRS chamber 내의 총 백혈구 수는 10.8×108 (범위 7.7-18.0×108)였으며, 각각의 백혈구가 차지하는 비율의 중간 값(범위)은 CD4+ 세포 29.6% (18.7-37.6), CD8+ T 세포 27.7% (19.2-40.0), B 세포 5.5% (2.2-12.1), NK 세포 15.7% (13.7- 19.9), CD14+ 세포가 12.4% (8.6-32.3)였다. LRS chamber와 400 mL 전혈유래 백혈구연층의 총 백혈구 수는 전혈유래 백혈 구연층에서 유의하게 많았으나, 림프구와 단핵구 수는 두 그룹 간에 통계학적인 차이는 없었다. CD4+ 세포 수와 B 세포 수가 두 그룹 간에 통계학적으로 유의한 차이를 보였고, 전혈유래 백 혈구연층에서 더 많았다(P<0.05). LRS chamber 내의 CD34+ 세포 수 중간값(범위)은 0.9×106 (0.2-2.6 ×106)이었다. 또한 LRS CHAMBER에서 분리한 CD14+ 세포는 7일 동안 사이토카 인 첨가 후 배양하여 수지상세포로 분화 및 성숙함을 확인하였다. 결론 : Trima Accelⓡ의 LRS chamber 내의 말초혈액 단핵 세포는 생존력과 기능성을 가진 세포들로서 연구용 소재로 매 우 유용하다고 판단되었다.
Abstract
Background : We investigated the characteristics of the mononuclear cells remaining in the leukoreduction system (LRS) chambers of Trima Accelⓡ in comparison with those of standard buffy coat cells, and evaluated their potential for differentiation into dendritic cells. Methods : Twenty-six LRS chambers of Trima Accelⓡ were collected after platelet pheresis from healthy adults. Flow cytometric analysis for T, B, NK, and CD14+ cells was performed and the number of CD34+ cells was counted. Differentiation and maturation into dendritic cells were induced using CD14+ cells seperated via Magnetic cell sorting (MACSⓡ) Seperation (Miltenyi Biotec Inc., USA). Results : Total white blood cell (WBC) count in LRS chambers was 10.8×108 (range 7.7-18.0× 108). The median values (range) of proportions of each cells were CD4+ T cell 29.6% (18.7-37.6), CD8+ T cell 27.7% (19.2-40.0), B cell 5.5% (2.2-12.1), NK cell 15.7% (13.7-19.9), and CD14+ cells 12.4% (8.6-32.3) respectively. Although total WBC count was significantly higher in the buffy coat (whole blood of 400 mL) than the LRS chambers, the numbers of lymphocytes and monocytes were not statistically different. The numbers of B cells and CD4+ cells were significantly higher in the buffy coat than the LRS chambers (P<0.05). The median value (range) of CD34+ cells obtained from the LRS chambers was 0.9×106 (0.2-2.6×106). After 7 days of cytokine-supplemented culture, the CD14+ cells were successfully differentiated into dendritic cells. Conclusions : The mononuclear cells in LRS chambers of Trima Accelⓡ are an excellent alternative source of viable and functional human blood cells, which can be used for research purposes.
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