背景:小活络丸具有祛风除湿、活络通痹等功效,用于治疗风寒湿痹、肢体疼痛,麻木拘挛等症.目的:观察小活络丸对小鼠再次免疫应答、特异性免疫(包括细胞免疫和体液免疫)、非特异性免疫(包括补体C3、单核-巨噬细胞系统和红细胞黏附功能)和自由基损伤,以及对疼痛及多种炎症模型的药理作用.设计:随机对照,分层实验.单位:广州市中医中药研究所和广州市中医医院药剂科.材料:选用NIH小鼠和ICR小鼠共628只,鼠龄6～8周.小活络丸(成分:胆南星、制川乌、制草乌、地龙、乳香(制)等;广州陈李济药厂生产,生产批号:19980612).兔抗小鼠IgG和补体C3抗血清试剂盒,由广州市医药卫生研究所药物室提供;超氧化物歧化酶活性和丙二醛含量测定试剂盒,南京建成生物工程研究所产品.方法:实验于1998-09/1999-12在广州市中医中药研究所药理研究室和广州市医药卫生研究所药物研究室动物室完成.①观察小活络丸抑制鸡红细胞诱导小鼠再次免疫应答作用:取ICR小鼠84只,雌雄各半,取出20只作为空白对照组;其余均腹腔注射环磷酰胺0.2 g/kg,1次/d,1 d.第4和12天,共2次腹膜内注射鸡红细胞诱导免疫增强病理模型探讨再次免疫应答.空白对照组(20只)腹腔注射等量生理盐水;动物于加强免疫后,按随机抽签法分为3组:环磷酰胺组20只(灌胃环磷酰胺40mg/kg),小活络丸组21只(灌胃小活络丸混悬液5.54 g/kg),模型组20只(灌胃等量蒸馏水);均1次/d,连续灌胃给药7 d.19 d后分别按单向免疫扩散法测定血清IgG,补体C3含量,采用聚乙二醇沉淀法测定循环免疫复合物含量变化.②观察小活络丸抑制小鼠迟发型超敏反应作用:取ICR小鼠54只,雌雄各半.第1天皮下注射10 g/L 2,4-二硝基氟苯50 μL/只致敏,第4天,将处理后动物按随机抽签法分为3组:泼尼松组(灌胃泼尼松0.01 g/kg),小活络丸组(灌胃小活络丸混悬液5.54 g/kg),模型组(灌胃等量蒸馏水),每组18只.均1次/d,连续灌胃给药7 d.11d后各组小鼠均涂10g/L 2,4-硝基氟苯25 μL/右耳,24 h后计算肿胀度(右、左耳质量差值).③观察小活络丸抑制小鼠红细胞免疫黏附功能:取NIH小鼠36只,雌雄各半,按随机抽签法分为3组:环磷酰胺组(灌胃环磷酰胺20 mg/kg),小活络丸组(灌胃小活络丸5.54 g/kg),空白对照组(灌胃等量蒸馏水),每组12只.均1次/d,连续灌胃给药7 d.7 d后取小鼠眼眶血,计算红细胞C3b受体花环率和红细胞免疫复合物花环率.④观察小活络丸抑制小鼠碳粒廓清作用:取ICR小鼠33只,雌雄不拘,将其按随机抽签法分为3组:泼尼松组(灌胃泼尼松20 mg/kg),小活络丸组(灌胃小活络丸混悬液5.54 g/kg),空白对照组(灌胃等量蒸馏水);均1次/d,连续灌胃7 d.于给药第7天末次给药后2 h,进行碳粒廓清试验,计算廓清指数K(K值=(LogA1-LogA2)/(T2-T1),式中A1和A2分别表示2和10 min所取血样的吸光度,T1和T2表示给碳粒后5和10 min取血时间;以K值高低,评价碳粒廓清作用.⑤观察小活络丸对鸡红细胞免?忖
疫小鼠溶血素、C3和丙二醛含量及超氧化物歧化酶活性的影响:取ICR小鼠80只,雌雄各半,取20只小鼠作为空白对照组,其余均腹腔注射2×108L-1鸡红细胞15 mL/kg作免疫诱导,空白对照组均以等量生理盐水代替鸡红细胞,灌胃蒸馏水,1次/d,连续灌胃7 d.按随机抽签法将免疫处理的动物分为3组:免疫对照组(灌胃给予蒸馏水),环磷酰胺组(灌胃环磷酰胺20 mg/kg),小活络丸组(灌胃小活络丸混悬液5.54g/kg);均1次/d,连续灌胃7 d;于第7天末次给药后2 h,计算IgM型溶血素半数溶血值[(样品吸收度/鸡红细胞半数溶血时的吸收度)×稀释倍数].按相应试剂盒操作说明书方法测定血清C3含量及丙二醛含量和超氧化物歧化酶活性.⑥观察小活络丸抑制小鼠琼脂肉芽组织增生作用:取NIH小鼠59只,皮下注射20g/L琼脂0.5 mL/只,24 h后,按随机抽签法分为3组:双氯芬酸组(灌胃双氯芬酸10 mg/kg),小活络丸组(灌胃给予小活络丸混悬液5.54 g/kg),模型组(灌胃等量蒸馏水);均1次/d,连续灌胃给药7 d;第8天处死小鼠,以每千克体质量琼脂肉芽克数表示抑制增生作用.⑦观察小活络丸抑制小鼠醋酸性扭体反应作用:取NIH小鼠63只,按随机抽签法分为3组:双氯芬酸组(灌胃双氯芬酸50 mg/kg),小活络丸组(灌胃小活络丸混悬液5.54g/kg),模型组(灌胃等量蒸馏水);均1次/d,连续灌胃给药2 d.末次给药后2 h,腹腔注射0.1 mol/L醋酸0.2 mL/只,计数20 min内小鼠的扭体次数.⑧观察小活络丸对二甲苯、巴豆油和角叉菜胶诱发急性渗出性炎症及炎症渗出液中前列腺素E含量的影响:取NIH小鼠219只,双氯芬酸组(灌胃双氯芬酸50 mg/kg),小活络丸组(灌胃小活络丸混悬液5.54 g/kg),模型组(灌胃等量蒸馏水);均1次/d,共2 d.末次给药后2 h,①涂二甲苯25μL/右耳,20 min后;②涂巴豆油25 μL/右耳,4 h后;计算耳肿胀度(右耳片重-左耳片重).③用10 g/L角叉菜胶20 μL/右足皮内注射致炎,3 h后,计算肿胀度(右足和左足间重量差值);并测致炎足渗出液中前列腺素E含量.组间计量资料差异比较采用t(方差不齐用t')检验.主要观察指标:小活络丸对小鼠再次免疫应答、特异性免疫、非特异性免疫和自由基损伤,以及对疼痛及多种炎症模型的药理作用.结果:NIH小鼠和ICR小鼠628只均进入结果分析.①模型组小鼠IgG、循环免疫复合物含量明显高于其他3组(P＜0.01),C3含量明显低于其他3组(P＜0.05～0.01).②泼尼松组和小活络丸组小鼠耳肿胀度明显低于空白对照组(P＜0.01).③空白对照组红细胞C3b受体花环率明显高于其他2组(P＜0.01),红细胞免疫复合物花环率明显低于其他2组(P＜0.01).④泼尼松组和小活络丸组吞噬指数(K值)明显低于空白对照组(P＜0.01).⑤环磷酰胺组和小活络丸组IgM型溶血素半数溶血值和血清丙二醛含量明显低于免疫对照组(P＜0.01),C3含量明显高于免疫对照组(P＜0.01),血清超氧化物歧化酶活力与免疫对照组比较,差异不明显(P＞0.05).⑥双氯芬酸组和小活络丸组小鼠琼脂肉芽组织质量与体质量比值明显低于模型组(P＜0.01).⑦双氯芬酸组和小活络丸组小鼠20 min内扭体次数明显少于模型组(P＜0.01,0.05).⑧双氯芬酸组二甲苯炎症模型和巴豆油炎症模型耳肿胀度、角叉菜胶急性炎症模型足肿胀度、炎症渗出液中前列腺素E含量明显小于或低于模型组(P＜0.05～0.01),小活络丸组炎症渗出液中前列腺素E含量明显低于模型组(P＜0.01).结论:小活络丸既有免疫抑制的药理作用,又有抗增殖性炎症、镇痛、抗氧化药效学效应.
揹景:小活絡汍具有祛風除濕、活絡通痺等功效,用于治療風寒濕痺、肢體疼痛,痳木拘攣等癥.目的:觀察小活絡汍對小鼠再次免疫應答、特異性免疫(包括細胞免疫和體液免疫)、非特異性免疫(包括補體C3、單覈-巨噬細胞繫統和紅細胞黏附功能)和自由基損傷,以及對疼痛及多種炎癥模型的藥理作用.設計:隨機對照,分層實驗.單位:廣州市中醫中藥研究所和廣州市中醫醫院藥劑科.材料:選用NIH小鼠和ICR小鼠共628隻,鼠齡6～8週.小活絡汍(成分:膽南星、製川烏、製草烏、地龍、乳香(製)等;廣州陳李濟藥廠生產,生產批號:19980612).兔抗小鼠IgG和補體C3抗血清試劑盒,由廣州市醫藥衛生研究所藥物室提供;超氧化物歧化酶活性和丙二醛含量測定試劑盒,南京建成生物工程研究所產品.方法:實驗于1998-09/1999-12在廣州市中醫中藥研究所藥理研究室和廣州市醫藥衛生研究所藥物研究室動物室完成.①觀察小活絡汍抑製鷄紅細胞誘導小鼠再次免疫應答作用:取ICR小鼠84隻,雌雄各半,取齣20隻作為空白對照組;其餘均腹腔註射環燐酰胺0.2 g/kg,1次/d,1 d.第4和12天,共2次腹膜內註射鷄紅細胞誘導免疫增彊病理模型探討再次免疫應答.空白對照組(20隻)腹腔註射等量生理鹽水;動物于加彊免疫後,按隨機抽籤法分為3組:環燐酰胺組20隻(灌胃環燐酰胺40mg/kg),小活絡汍組21隻(灌胃小活絡汍混懸液5.54 g/kg),模型組20隻(灌胃等量蒸餾水);均1次/d,連續灌胃給藥7 d.19 d後分彆按單嚮免疫擴散法測定血清IgG,補體C3含量,採用聚乙二醇沉澱法測定循環免疫複閤物含量變化.②觀察小活絡汍抑製小鼠遲髮型超敏反應作用:取ICR小鼠54隻,雌雄各半.第1天皮下註射10 g/L 2,4-二硝基氟苯50 μL/隻緻敏,第4天,將處理後動物按隨機抽籤法分為3組:潑尼鬆組(灌胃潑尼鬆0.01 g/kg),小活絡汍組(灌胃小活絡汍混懸液5.54 g/kg),模型組(灌胃等量蒸餾水),每組18隻.均1次/d,連續灌胃給藥7 d.11d後各組小鼠均塗10g/L 2,4-硝基氟苯25 μL/右耳,24 h後計算腫脹度(右、左耳質量差值).③觀察小活絡汍抑製小鼠紅細胞免疫黏附功能:取NIH小鼠36隻,雌雄各半,按隨機抽籤法分為3組:環燐酰胺組(灌胃環燐酰胺20 mg/kg),小活絡汍組(灌胃小活絡汍5.54 g/kg),空白對照組(灌胃等量蒸餾水),每組12隻.均1次/d,連續灌胃給藥7 d.7 d後取小鼠眼眶血,計算紅細胞C3b受體花環率和紅細胞免疫複閤物花環率.④觀察小活絡汍抑製小鼠碳粒廓清作用:取ICR小鼠33隻,雌雄不拘,將其按隨機抽籤法分為3組:潑尼鬆組(灌胃潑尼鬆20 mg/kg),小活絡汍組(灌胃小活絡汍混懸液5.54 g/kg),空白對照組(灌胃等量蒸餾水);均1次/d,連續灌胃7 d.于給藥第7天末次給藥後2 h,進行碳粒廓清試驗,計算廓清指數K(K值=(LogA1-LogA2)/(T2-T1),式中A1和A2分彆錶示2和10 min所取血樣的吸光度,T1和T2錶示給碳粒後5和10 min取血時間;以K值高低,評價碳粒廓清作用.⑤觀察小活絡汍對鷄紅細胞免?忖
疫小鼠溶血素、C3和丙二醛含量及超氧化物歧化酶活性的影響:取ICR小鼠80隻,雌雄各半,取20隻小鼠作為空白對照組,其餘均腹腔註射2×108L-1鷄紅細胞15 mL/kg作免疫誘導,空白對照組均以等量生理鹽水代替鷄紅細胞,灌胃蒸餾水,1次/d,連續灌胃7 d.按隨機抽籤法將免疫處理的動物分為3組:免疫對照組(灌胃給予蒸餾水),環燐酰胺組(灌胃環燐酰胺20 mg/kg),小活絡汍組(灌胃小活絡汍混懸液5.54g/kg);均1次/d,連續灌胃7 d;于第7天末次給藥後2 h,計算IgM型溶血素半數溶血值[(樣品吸收度/鷄紅細胞半數溶血時的吸收度)×稀釋倍數].按相應試劑盒操作說明書方法測定血清C3含量及丙二醛含量和超氧化物歧化酶活性.⑥觀察小活絡汍抑製小鼠瓊脂肉芽組織增生作用:取NIH小鼠59隻,皮下註射20g/L瓊脂0.5 mL/隻,24 h後,按隨機抽籤法分為3組:雙氯芬痠組(灌胃雙氯芬痠10 mg/kg),小活絡汍組(灌胃給予小活絡汍混懸液5.54 g/kg),模型組(灌胃等量蒸餾水);均1次/d,連續灌胃給藥7 d;第8天處死小鼠,以每韆剋體質量瓊脂肉芽剋數錶示抑製增生作用.⑦觀察小活絡汍抑製小鼠醋痠性扭體反應作用:取NIH小鼠63隻,按隨機抽籤法分為3組:雙氯芬痠組(灌胃雙氯芬痠50 mg/kg),小活絡汍組(灌胃小活絡汍混懸液5.54g/kg),模型組(灌胃等量蒸餾水);均1次/d,連續灌胃給藥2 d.末次給藥後2 h,腹腔註射0.1 mol/L醋痠0.2 mL/隻,計數20 min內小鼠的扭體次數.⑧觀察小活絡汍對二甲苯、巴豆油和角扠菜膠誘髮急性滲齣性炎癥及炎癥滲齣液中前列腺素E含量的影響:取NIH小鼠219隻,雙氯芬痠組(灌胃雙氯芬痠50 mg/kg),小活絡汍組(灌胃小活絡汍混懸液5.54 g/kg),模型組(灌胃等量蒸餾水);均1次/d,共2 d.末次給藥後2 h,①塗二甲苯25μL/右耳,20 min後;②塗巴豆油25 μL/右耳,4 h後;計算耳腫脹度(右耳片重-左耳片重).③用10 g/L角扠菜膠20 μL/右足皮內註射緻炎,3 h後,計算腫脹度(右足和左足間重量差值);併測緻炎足滲齣液中前列腺素E含量.組間計量資料差異比較採用t(方差不齊用t')檢驗.主要觀察指標:小活絡汍對小鼠再次免疫應答、特異性免疫、非特異性免疫和自由基損傷,以及對疼痛及多種炎癥模型的藥理作用.結果:NIH小鼠和ICR小鼠628隻均進入結果分析.①模型組小鼠IgG、循環免疫複閤物含量明顯高于其他3組(P＜0.01),C3含量明顯低于其他3組(P＜0.05～0.01).②潑尼鬆組和小活絡汍組小鼠耳腫脹度明顯低于空白對照組(P＜0.01).③空白對照組紅細胞C3b受體花環率明顯高于其他2組(P＜0.01),紅細胞免疫複閤物花環率明顯低于其他2組(P＜0.01).④潑尼鬆組和小活絡汍組吞噬指數(K值)明顯低于空白對照組(P＜0.01).⑤環燐酰胺組和小活絡汍組IgM型溶血素半數溶血值和血清丙二醛含量明顯低于免疫對照組(P＜0.01),C3含量明顯高于免疫對照組(P＜0.01),血清超氧化物歧化酶活力與免疫對照組比較,差異不明顯(P＞0.05).⑥雙氯芬痠組和小活絡汍組小鼠瓊脂肉芽組織質量與體質量比值明顯低于模型組(P＜0.01).⑦雙氯芬痠組和小活絡汍組小鼠20 min內扭體次數明顯少于模型組(P＜0.01,0.05).⑧雙氯芬痠組二甲苯炎癥模型和巴豆油炎癥模型耳腫脹度、角扠菜膠急性炎癥模型足腫脹度、炎癥滲齣液中前列腺素E含量明顯小于或低于模型組(P＜0.05～0.01),小活絡汍組炎癥滲齣液中前列腺素E含量明顯低于模型組(P＜0.01).結論:小活絡汍既有免疫抑製的藥理作用,又有抗增殖性炎癥、鎮痛、抗氧化藥效學效應.
배경:소활락환구유거풍제습、활락통비등공효,용우치료풍한습비、지체동통,마목구련등증.목적:관찰소활락환대소서재차면역응답、특이성면역(포괄세포면역화체액면역)、비특이성면역(포괄보체C3、단핵-거서세포계통화홍세포점부공능)화자유기손상,이급대동통급다충염증모형적약리작용.설계:수궤대조,분층실험.단위:엄주시중의중약연구소화엄주시중의의원약제과.재료:선용NIH소서화ICR소서공628지,서령6～8주.소활락환(성분:담남성、제천오、제초오、지룡、유향(제)등;엄주진리제약엄생산,생산비호:19980612).토항소서IgG화보체C3항혈청시제합,유엄주시의약위생연구소약물실제공;초양화물기화매활성화병이철함량측정시제합,남경건성생물공정연구소산품.방법:실험우1998-09/1999-12재엄주시중의중약연구소약리연구실화엄주시의약위생연구소약물연구실동물실완성.①관찰소활락환억제계홍세포유도소서재차면역응답작용:취ICR소서84지,자웅각반,취출20지작위공백대조조;기여균복강주사배린선알0.2 g/kg,1차/d,1 d.제4화12천,공2차복막내주사계홍세포유도면역증강병리모형탐토재차면역응답.공백대조조(20지)복강주사등량생리염수;동물우가강면역후,안수궤추첨법분위3조:배린선알조20지(관위배린선알40mg/kg),소활락환조21지(관위소활락환혼현액5.54 g/kg),모형조20지(관위등량증류수);균1차/d,련속관위급약7 d.19 d후분별안단향면역확산법측정혈청IgG,보체C3함량,채용취을이순침정법측정순배면역복합물함량변화.②관찰소활락환억제소서지발형초민반응작용:취ICR소서54지,자웅각반.제1천피하주사10 g/L 2,4-이초기불분50 μL/지치민,제4천,장처리후동물안수궤추첨법분위3조:발니송조(관위발니송0.01 g/kg),소활락환조(관위소활락환혼현액5.54 g/kg),모형조(관위등량증류수),매조18지.균1차/d,련속관위급약7 d.11d후각조소서균도10g/L 2,4-초기불분25 μL/우이,24 h후계산종창도(우、좌이질량차치).③관찰소활락환억제소서홍세포면역점부공능:취NIH소서36지,자웅각반,안수궤추첨법분위3조:배린선알조(관위배린선알20 mg/kg),소활락환조(관위소활락환5.54 g/kg),공백대조조(관위등량증류수),매조12지.균1차/d,련속관위급약7 d.7 d후취소서안광혈,계산홍세포C3b수체화배솔화홍세포면역복합물화배솔.④관찰소활락환억제소서탄립곽청작용:취ICR소서33지,자웅불구,장기안수궤추첨법분위3조:발니송조(관위발니송20 mg/kg),소활락환조(관위소활락환혼현액5.54 g/kg),공백대조조(관위등량증류수);균1차/d,련속관위7 d.우급약제7천말차급약후2 h,진행탄립곽청시험,계산곽청지수K(K치=(LogA1-LogA2)/(T2-T1),식중A1화A2분별표시2화10 min소취혈양적흡광도,T1화T2표시급탄립후5화10 min취혈시간;이K치고저,평개탄립곽청작용.⑤관찰소활락환대계홍세포면?촌
역소서용혈소、C3화병이철함량급초양화물기화매활성적영향:취ICR소서80지,자웅각반,취20지소서작위공백대조조,기여균복강주사2×108L-1계홍세포15 mL/kg작면역유도,공백대조조균이등량생리염수대체계홍세포,관위증류수,1차/d,련속관위7 d.안수궤추첨법장면역처리적동물분위3조:면역대조조(관위급여증류수),배린선알조(관위배린선알20 mg/kg),소활락환조(관위소활락환혼현액5.54g/kg);균1차/d,련속관위7 d;우제7천말차급약후2 h,계산IgM형용혈소반수용혈치[(양품흡수도/계홍세포반수용혈시적흡수도)×희석배수].안상응시제합조작설명서방법측정혈청C3함량급병이철함량화초양화물기화매활성.⑥관찰소활락환억제소서경지육아조직증생작용:취NIH소서59지,피하주사20g/L경지0.5 mL/지,24 h후,안수궤추첨법분위3조:쌍록분산조(관위쌍록분산10 mg/kg),소활락환조(관위급여소활락환혼현액5.54 g/kg),모형조(관위등량증류수);균1차/d,련속관위급약7 d;제8천처사소서,이매천극체질량경지육아극수표시억제증생작용.⑦관찰소활락환억제소서작산성뉴체반응작용:취NIH소서63지,안수궤추첨법분위3조:쌍록분산조(관위쌍록분산50 mg/kg),소활락환조(관위소활락환혼현액5.54g/kg),모형조(관위등량증류수);균1차/d,련속관위급약2 d.말차급약후2 h,복강주사0.1 mol/L작산0.2 mL/지,계수20 min내소서적뉴체차수.⑧관찰소활락환대이갑분、파두유화각차채효유발급성삼출성염증급염증삼출액중전렬선소E함량적영향:취NIH소서219지,쌍록분산조(관위쌍록분산50 mg/kg),소활락환조(관위소활락환혼현액5.54 g/kg),모형조(관위등량증류수);균1차/d,공2 d.말차급약후2 h,①도이갑분25μL/우이,20 min후;②도파두유25 μL/우이,4 h후;계산이종창도(우이편중-좌이편중).③용10 g/L각차채효20 μL/우족피내주사치염,3 h후,계산종창도(우족화좌족간중량차치);병측치염족삼출액중전렬선소E함량.조간계량자료차이비교채용t(방차불제용t')검험.주요관찰지표:소활락환대소서재차면역응답、특이성면역、비특이성면역화자유기손상,이급대동통급다충염증모형적약리작용.결과:NIH소서화ICR소서628지균진입결과분석.①모형조소서IgG、순배면역복합물함량명현고우기타3조(P＜0.01),C3함량명현저우기타3조(P＜0.05～0.01).②발니송조화소활락환조소서이종창도명현저우공백대조조(P＜0.01).③공백대조조홍세포C3b수체화배솔명현고우기타2조(P＜0.01),홍세포면역복합물화배솔명현저우기타2조(P＜0.01).④발니송조화소활락환조탄서지수(K치)명현저우공백대조조(P＜0.01).⑤배린선알조화소활락환조IgM형용혈소반수용혈치화혈청병이철함량명현저우면역대조조(P＜0.01),C3함량명현고우면역대조조(P＜0.01),혈청초양화물기화매활력여면역대조조비교,차이불명현(P＞0.05).⑥쌍록분산조화소활락환조소서경지육아조직질량여체질량비치명현저우모형조(P＜0.01).⑦쌍록분산조화소활락환조소서20 min내뉴체차수명현소우모형조(P＜0.01,0.05).⑧쌍록분산조이갑분염증모형화파두유염증모형이종창도、각차채효급성염증모형족종창도、염증삼출액중전렬선소E함량명현소우혹저우모형조(P＜0.05～0.01),소활락환조염증삼출액중전렬선소E함량명현저우모형조(P＜0.01).결론:소활락환기유면역억제적약리작용,우유항증식성염증、진통、항양화약효학효응.
BACKGROUND: Xiaohuoluo pill can expel pathogenic wind, remove dampness and activate collaterals. It is used for treatment of Bi-syndrome due to wind-cold-dampness, pain and numbness in limbs.OBJECTIVE: To observe the pharmacological effect of Xiaohuoluo pills on secondary immune response, specific immunity (including cellular immunity and humoral immunity), non-specific immunity [including complement 3(C3), mononuclear phagocyte system (MPS) and red blood cell (RBC)adhesion function] and free radical injury as well as pain and many other inflammations in mice.DESIGN: A randomized controlled stratified trial.SETTING: Guangzhou Institute of Traditional Chinese Medicine and Chinese Materia Medica; Department of Pharmacy, Guangzhou Hospital of Traditional Chinese Medicine.MATERIALS: Totally 628 NIH and ICR mice of 6 to 8 weeks were involved in this trial. Xiaohuoluo pills (components: Dannanxing, Zhichuanwu, Zhicaowu, Dilong, Ruxiang and so on; Chenli Pharmaceutical Factory,Guangzhou; Brach No. 19980612) were used in this trial. Rabbit antimouse immunoglobulin G (IgG) and C3 antiserum reagent kit (Guangzhou Institute of Medicine and Health) and reagent kit for measuring the antioxidizing activity of superoxide dismutase (SOD) and the level of malondialdehyde (MDA) (Jiancheng Institute of Bioengineering, Nanjing) were used.METHODS: This trial was carried out in the Guangzhou Institute of Traditional Chinese Medicine and Chinese Materia Medica; Department of Pharmacy, Guangzhou Institute of Medicine and Health during September 1998 to December 1999. ① To observe the suppressive effect of Xiaohuoluo pills on cock red blood cell (CRBC)-induced secondary immune response: Eight-four ICR mice, male and female in half, were selected.Twenty of 84 mice served as blank controls; The other 64 mice were intraperitoneally injected with cyclophos-phamide (CY) of 0.2 g/kgonce. On the 4th and 12th days, CRBC was intraperitoneally injected into the mice twice to induce immunoenhancing pathological models to form secondary immune response. Mice served as blank controls were intraperitoneally injected with the same volume of normal saline; The immunoenhanced mice were assigned into 3 groups by a lot: CY group (n=20, CY, 40 mg/kg, intragastric administration, I.g.), Xiaohuoluo pills (n=21, Xiaohuoluo pills suspension, 5.54 g/kg, I.g.) and model group (n=20, distilled water, the same volume as other groups, I.g.); once a day within 7 successive days. 19 days later, the levels of serum IgG and C3 were measured with single immunodiffusion method, and the level of circulating immune compound (CIC) was measured with polyethylene glycol precipitation method. ② To observe the suppressive effect of Xiaohuoluo pills on delayed type hypersensitivity (DTH): Fifty-four ICR mice, male and female in half, were selected. On the 1st day, the mice were sensitized by subcutaneous injection of 10 g/L 2,4-dinitrofluorobenzen e (DNFB) of 50 μL for each. On the 4th day, the sensitized mice were assigned into 3 groups by a lot: Prednisone group (n=18, prednisone, 0.01 g/kg, I.g.), Xiaohuoluo pills (n=18, Xiaohuoluo pills suspension, 5.54 g/kg, I.g.), model group (n=18, distilled water, the same volume as other groups, I.g.), all once a day within 7 successive days. 11 days later, 10 g/L DNFB of 25μL was spread on the right ear of each mouse in each group. The swelling degree was calculated 24 hours later (The mass difference between right ear and left ear). ③ To observe the suppressive effect of Xiaohuoluo pills on immune adhesion function of RBC of mouse: Thirty-six NIH mice, male and female in half, were selected and assigned into 3 groups by a lot: CY group (n=12, CY, 20 mg/kg,I.g.), Xiaohuoluo pills (n=12, Xiaohuoluo pills suspension, 5.54 g/kg, I.g.)and blank control group (n=12, distilled water, the same volume as other groups, I.g.), once a day within 7 successive days. 7 days later, blood was taken from the orbit of mice for calculating the rosette rate of RBC-C3b receptor and the rosette rate of RBC immune compound. ④ To observe the suppressive effect o20 Mg/kg, I.g.),Xiaohuoluo pills group (Xiaohuoluo pills suspension, 5.54 g/kg, I.g.) , once a day within 7 successive days; IgM-type hemolytic concentration (HC50)was measured at 2 hours after the last administration on the 7th day [ (Sample absorption / Absorption at HC50 of CRBC) ×diluted time]. The levels of serum C3 and MDA and the activity of SOD were measured according to the method from corresponding reagent kit. ⑥ To observe the suppressive effect of Xiaohuoluo pills on agar granulation tissue hyperplasia in mice:Fifty-nine NIH mice were selected and given subcutaneous injection of 20 g/L agar of 0.5 mL for each. 24 hours later, the mice were assigned into 3 groups by a lot: diclofenac group (diclofenac, 10 mg/kg,I.g..), Xiaohuoluo pills group (Xiaohuoluo pills suspension, 5.54 g/kg, I.g.) and model group (distilled water, the same volume as other groups, I.g.), once a day within 7 successive days; On the 8th day, the mice were sacrificed. The hyperplasiainhibiting effect was presented in the form of the mass of agar granulation tissue in one kilogram body mass ⑦ To observe the suppressive effect of Xiaohuoluo pills on acetic distortion reaction: Sixty-three NIH mice were se lected and assigned into 3 groups by a lot: diclofenac group (diclofenac,50 mg/kg, I.g.), Xiaohuoluo pills group (Xiaohuoluo pills suspension, 5.54 g/kg,I.g.) and model group (distilled water, the same volume of other groups, I.g.),once a day within 2 successive days. At 2 hours after the last administration, the mice were given intraperitoneal injection of 0.1 mol/L acetic acid of 0.2 mL for each one. The times of distortion of mice within 20 minutes were counted. ⑧ To observe the effect of Xiaohuoluo pills on the acute exudative inflammation evoked by dimethylbenzene, croton oil and carrageenan, and the level of prostaglandin E in the inflammatory exudates:Totally 219 NIH mice were selected and assigned into 3 groups by a lot:diclofenac group (diclofenac, 50 mg/kg, I.g.), Xiaohuoluo pills group (Xiaohuoluo pills suspension, 5.54 g/kg, I.g.) and model group (distilled water, the same volume of other groups, I.g.) once a day within 2 successive days. At 2 hours after the last administration, dimethylbenzene of 25 μL was spread on the right ear for 20 minutes, or croton oil of 25 μL was also spread on the right ear, 4 hours later, the swelling of right ear was calculated (mass of right ear-mass of left ear). 10 g/L carrageenan of 20 μL was subcutaneously injected into the right foot, 3 hours later, the swelling degree was calculated (The difference of right foot and left foot); and the level of prostaglandin E in the inflammatory exudates was measured. ⑨ t test(t' test for heteroscedasticity) was used for comparing the difference in measurement data among groups.MAIN OUTCOME MEASURES: Pharmacological effect of Xiaohuoluo pills on secondary immune response, specific immunity, non-specific immunity and free radical injury as well as pain and many other inflammations in mice.RESULTS: Totally 628 NIH and ICR mice were involved in result analysis. ① The level of IgG and CIC of mice in the model group was significantly higher than that in the other 3 groups respectively (P ＜ 0.01),while the level of C3 was significantly lower than that in the other 3 groups (P ＜ 0.05 to 0.01). ② The swelling degree of mice in the diclofenac group and Xiaohuoluo pills group was significantly lower than that in the blank control group respectively ( both P ＜ 0.01). ③ The rosette rate of RBC-C3b receptor and RBC immune compound in the blank control group was significantly higher than that in the other 2 gro ups respectively (P ＜ 0.01). ④ The phagocytic index (K value )in the diclofenac group and Xiaohuoluo pills group was significantly lower than that in the blank control group, respectively (both P ＜ 0.01).⑤ IgM-type HC50 and the level of serum MDA of CY group and Xiaohuoluo pills group were obviously lower than those in the immune control group (P ＜ 0.01),while the level of C3 was higher than that of immune control group, there was no significant difference in the activity of serum SOD between CY group or Xiaohuoluo pills group and immune control group (P ＞ 0.05). ⑥The ratio of agar granulation tissue mass to body mass in the diclofenac group or Xiaohuoluo pills group was significantly lower than that in the model group (P ＜ 0.01). ⑦ The times of distortion of mice within 20 minutes in the diclofenac group or Xiaohuoluo pills group were signifi cantly less than those of model group (P ＜ 0.01, 0.05). ⑧The ear swelling degree of dimethylbenzene-induced inflammatory models and croton oil-induced inflammatory models, and foot swelling degree of carrageenan-induced acute inflammatory models as well as the level of prostaglandin E in the inflammatory exudates in the diclofenac group were significantly milder or lower than those in the model group (P ＜ 0.05 to 0.01), and the level of prostaglandin E in the inflammatory exudates in the Xiaohuoluo pills group was significantly lower than that in the model group (P ＜ 0.01).CONCLUSION: Xiaohuoluo pills possess pharmacological effects of immunosuppression, anti-proliferative inflammation, analgesia and antioxidation.
