Inhibitory effects of Bee Venom Herbal Acupuncture(BVA) on collagen induced arthritis in rats : 蜂毒藥鍼이 흰쥐의 Collagen誘發 關節炎중 면역반응에 미치는 影響

국문 초록 (Abstract)

본 연구에서는 봉독약침(BVA)이 콜라겐 2형(CⅡ)으로 유발된 실험적 관절염 흰쥐에 미치는 영향을 검토하였다. CⅡ로 초기 면역시킨 후 14일간 매일 BVA를 투여한 군에서 BVA의 양에 따라 관절염 발생을 억제하였다. 추가 면역 후 14일 동안 200㎕/kg의 BVA를 투여한 실험군에서는 관절염 발병을 효과적으로 억제하였으나 CⅡ로 면역 후 약 40일째의 관절염 흰쥐에서는 BVA가 CⅡ에 대한 면역억제반응에 효과가 인정되지 않았다. 관절염 생성 48일째에 항CⅡ 항체는 현저하게 줄었지만 추가 접종후 측정한 CⅡ에 대한 항체는 변화가 없었다. 한편, BVA 투여는 흰쥐에서 지연형과민반응은 현저하게 억제하였다. BVA 전처리군은 carrageenan 유발된 부종을 2시간 후 BVA 10㎕/kg 에서 23.2%, 3시간 후 15㎕/kg 투여군에서 43.2%로 가장 높은 억제율을 나타내며 현저히 감소시켰다. 대조군으로서 Indomethacin 전처리군은 carrageenan유발 부종을 현저히 감소시켰다 . BVA가 5㎕/kg일 때 anti-CⅡ IgG와 anti-IgG2a의 항체들의 낮은 생성율을 보였다. Saline을 투여한 생쥐군과 비교해 볼 때 14일 동안 하루에 한번씩 200㎕/kg 이상의 BVA를 투여한 군이 CⅡ 면역후 24일이 지난 생쥐의 세포를 in vitro에서 CⅡ로 배양한 결과 lymph node 세포에서의 Il-2와 INF-γ 생산성을 억제하였다. 또한 BVA를 투여한 군은 in vitro에서 CⅡ로 lymph node 세포를 자극한 결과 대식세포, IL-1β, IL-6, TNF-α의 생산을 억제한 반면 ex vivo에서의 비장세포에서는 INF-γ의 분비를 억제하였다. CⅡ로 유도된 흰쥐의 관절염 발생률과 발병률에 대한 BVA의 영향을 평가함에 있어 CⅡ는 3주 간격으로 2번 면역을 시켰으며 BVA는 14일 동안 4가지 다른 방법으로 하루에 한번씩 투여했다. 처음 CⅡ로 면역을 시작한 날부터 매일 200㎍/kg의 BVA를 14일간 투여한 경우 관절염발생, 항체형성, 지연형과민반응을 억제하였으며 추가 면역 후 동시에 BVA로 치료한 경우도 CⅡ에 대한 면역반응과 관절염발생을 억제하였다.

다국어 초록 (Multilingual Abstract)

The effects of Bee Venom herbal acupuncture(BVA), a traditional immunosuppressive Korean herbal acupuncture, on the development of type collagen(C)-induced arthritis(CIA) in rats were studied. Rats were 14 daily i.p. injected with BVA on the day of the primary immunization with C, BVA prevented the development of collagen arthritis in a dose--dependent manner. Also, when rats were i.p. injected with various doses of BVA at a daily dose of 200/kg for 14 days from the day of a booster injection, BVA suppressed the onset of arthritis. Serum antibodies to collagen measured on day 48 were also significantly reduced although no difference was observed in serum antibodies to C measured on the day of a booster injection. DTH was significanly suppressed by treatment of rats with of the BVA. On the other hand, rats which were immunized with C, developed arthritis on approximately 40 days after the primary immunization. Therefore, therapeutic treatment with BVA started 5 weeks after the primary immunization. Its treatment also did not alter the immunological responses to C. Pretreatment by BVA significantly reduced the carrageenan--induced adema in a dose-dependent way to reach a maximal inhibition with the dose 10/kg(23.2%) and after 3h with the dose 15/kg(43.2%). Indomethacin pretreatment induced a significant inhibitory action on carrageenan-induced edema. Lower production of anti-C IgG and IgG2a antibodies were seen in mice given 5/kg of BVA.
As compared with rats treated with saline, BVA i.p. injected at doses of more than 200/kg once a day for 14 days inhibited the ability of inguinal lymph node cells to produce T cell cytokines interleukin-2 and interfcron- when the cells were obtained from rats 24 days after immunization and cultured in vitro with C. Treatment with BVA also inhibited production of macrophage cytokines interleukin-1, IL-6 and tumor necrosis factor- in response to in vitro stimulation of lymph node cells with C. On the other hand, the i.p. injection of BVA suppressed the C-specific secretion of IFN- from splenocytes ex vivo. When the influence of BVA was evaluated on the incidence and development of arthritis in rat CIA. Rats were immunized twice at a 3-week interval with bovine C, with BVA being given i.p. once a day for 14 days with four different regimens. A 14-day course of BVA treatment at a daily dose of 200/kg, which began on the day of the first C immunization, suppressed the development of arthritis, as well as antibody formation and delayed-type hypersensitivit to C. Treatment with BVA, which started on the same day as the booster immunization, also resulted in inhibition of development of arthritis and of immune responses to C.

목차 (Table of Contents)

• Contents = 0
• Introduction = 1
• Materials and methods = 4
• Materials = 4
• Induction of arthritis = 4
• Preparation of LN cell suspension and culture supernatants = 5
• Preparation and culture of splenocytes stimulated by CII = 6
• Arthritis assesment = 6
• Determination of anti-CII IgG and IgG2a antibody = 6
• Measurement of serum anti-CII specific antibodies titer = 7
• Measurement of delayed-type hypersensitivity = 8
• Cytokine assays = 8
• Carrageenan-induced rat edema = 9
• Analytical methods = 9
• Statistical analysis = 9
• Results = 11
• 1. Effect of BVA on the carrageenan-induced arthritis edema = 11
• 2. The effect of BVA i.v on anti-CII IgG and IgG2a antibody production = 14
• 3. Effect of BVA treatment on the established diseases by means of total immunoglobulin in rats = 17
• 4. Effect of i.p. injection of BVA on the prodcution of IFN-？？ from splenocytes = 20
• 5. Reductive effect of i.p. injection of BVA on CII induced arthritis in rats = 22
• 6. Effect of i.p. injection of BVA on serum anti-CII antibody titer = 25
• 7. Inhibition of DTH response to CII by the i.p. injection of BVA in immunmized rats. = 28
• Discussion = 30
• Summary = 35
• References = 37
• 국문초록 = 41