HS6101不同時(shí)間給藥對(duì)環(huán)磷酰胺所致小鼠造血功能損傷的恢復(fù)作用

邢爽，申星，熊國林，柳曉蘭，楊萌，代常亮，劉曉宇，余祖胤

HS6101不同時(shí)間給藥對(duì)環(huán)磷酰胺所致小鼠造血功能損傷的恢復(fù)作用

邢爽，申星，熊國林，柳曉蘭，楊萌，代常亮，劉曉宇，余祖胤

目的 探討HS6101(6101)不同時(shí)間給藥對(duì)環(huán)磷酰胺(CTX)致小鼠造血功能損傷的恢復(fù)作用。方法 103 只5～6周齡雄性ICR小鼠，分為CTX對(duì)照組、6101預(yù)防組、6101治療組、6101預(yù)防+治療組共4組。CTX損傷小鼠建模方法為腹腔注射CTX 100mg/kg，1次/d，連續(xù)注射3d。6101給藥劑量為27μg/只，單次皮下注射0.2ml。6101預(yù)防組給藥時(shí)間為CTX首次注射前1h，6101治療組給藥時(shí)間為CTX末次注射后1h，6101預(yù)防+治療組給藥時(shí)間為CTX首次注射前1h+CTX末次注射后1h，CTX對(duì)照組注射生理鹽水0.2ml/只。分別于CTX首次注射前1d和注射后3、5、7、9、11、13、15、17d取尾靜脈血10μl檢測(cè)血常規(guī)，比較6101各給藥組與CTX對(duì)照組間的差異。另取30只雄性ICR小鼠，在上述分組基礎(chǔ)上增加正常對(duì)照組，6只/組，建模與給藥方法同上，各組分別于CTX首次注射后4、9d處死3只小鼠，取左側(cè)股骨分離單個(gè)核細(xì)胞進(jìn)行骨髓造血祖細(xì)胞集落培養(yǎng)，取右側(cè)股骨做常規(guī)病理組織學(xué)觀察。結(jié)果 腹腔注射CTX后小鼠外周白細(xì)胞、中性粒細(xì)胞、淋巴細(xì)胞、紅細(xì)胞和血小板數(shù)均迅速減少。6101預(yù)防組小鼠外周血白細(xì)胞、中性粒細(xì)胞和淋巴細(xì)胞數(shù)最低值明顯高于CTX對(duì)照組，且其第3天的檢測(cè)值高于6101治療組及6101預(yù)防+治療組；6101預(yù)防組外周血紅細(xì)胞在第3、5、7天均明顯高于CTX對(duì)照組，第3天高于6101治療組和6101預(yù)防+治療組；6101預(yù)防組血小板數(shù)第3天時(shí)明顯高于其他3組，而6101治療組與6101預(yù)防+治療組則明顯低于CTX對(duì)照組，血小板開始恢復(fù)后6101預(yù)防給藥促恢復(fù)作用更明顯。第4天和第9天的骨髓造血細(xì)胞集落培養(yǎng)結(jié)果顯示，6101預(yù)防組、治療組和預(yù)防+治療組的粒巨噬系、巨核系、爆式紅系和紅系細(xì)胞集落數(shù)均高于CTX組；第4天骨髓病理組織學(xué)觀察結(jié)果也表明6101各給藥組小鼠骨髓組織結(jié)構(gòu)優(yōu)于CTX對(duì)照組。實(shí)驗(yàn)中6101治療組和預(yù)防+治療組小鼠出現(xiàn)死亡(分別為3/18、9/18)，可能與其早期血小板數(shù)顯著降低有關(guān)，提示6101不宜在CTX注射后給藥。結(jié)論 6101于CTX注射前1h給藥可明顯促進(jìn)CTX所致ICR小鼠造血功能損傷的恢復(fù)。

HS6101；脂肽化合物；環(huán)磷酰胺；小鼠；造血系統(tǒng)

細(xì)胞保護(hù)劑對(duì)保障腫瘤患者順利接受放化療具有重要作用，但在應(yīng)用時(shí)大多具有很強(qiáng)的給藥時(shí)效性。例如，氨磷汀(WR-2721)作為廣譜細(xì)胞保護(hù)劑能夠減低順鉑、環(huán)磷酰胺等化療藥物的毒性[1]，但需在化療前15～30min靜脈注射給藥[2]。近年來，脂肽類Toll樣受體激動(dòng)劑類的細(xì)胞保護(hù)劑受到高度關(guān)注，但其效應(yīng)亦明顯受給藥時(shí)間影響[3-4]。HS6101是人工合成的作用于Toll樣受體的一種脂肽，與CBLB613屬同一類細(xì)胞保護(hù)劑[4-5]，能明顯減輕射線引起的小鼠及恒河猴的造血細(xì)胞損傷，本課題組前期觀察到它對(duì)化療藥物損傷的小鼠造血細(xì)胞有保護(hù)作用，但給藥時(shí)間對(duì)其效應(yīng)的影響尚不明了。因此，本研究觀察了HS6101不同時(shí)間給藥對(duì)化療藥物環(huán)磷酰胺所致ICR小鼠造血功能損傷恢復(fù)的影響，旨在為確定HS6101的臨床給藥時(shí)間提供實(shí)驗(yàn)依據(jù)。

1 材料與方法

1.1 藥物與儀器 環(huán)磷酰胺(CTX)粉針劑為山西普德藥業(yè)有限公司生產(chǎn)(國藥準(zhǔn)字號(hào)H14023686，批號(hào)04130201)。HS6101(6101)白色凍干粉劑為浙江海正藥業(yè)股份有限公司研制(批號(hào)20120903)。小鼠骨髓細(xì)胞集落培養(yǎng)基M3434購自加拿大Stem Cell公司。Celltac E MEK-7222K型血細(xì)胞自動(dòng)分析儀購自日本Nihon Kohden公司。

1.2 動(dòng)物及分組 雄性5～6周齡SPF級(jí)ICR小鼠103只，體重20.0±2.0g，購自斯貝福(北京)實(shí)驗(yàn)動(dòng)物科技有限公司，動(dòng)物質(zhì)量合格證號(hào)SCXK-(京)2011-0004。動(dòng)物常規(guī)飼養(yǎng)于軍事醫(yī)學(xué)科學(xué)院實(shí)驗(yàn)動(dòng)物中心SPF級(jí)小鼠實(shí)驗(yàn)室內(nèi)，動(dòng)物飼養(yǎng)設(shè)施合格證號(hào)SYXK-(軍)2007-004，適應(yīng)性飼養(yǎng)3d后進(jìn)行實(shí)驗(yàn)。另30只供骨髓細(xì)胞培養(yǎng)與骨髓病理切片觀察的動(dòng)物分籠單獨(dú)飼養(yǎng)。實(shí)驗(yàn)動(dòng)物分為CTX對(duì)照組、6101預(yù)防組(-1h)、6101治療組(+1h)與6101預(yù)防+治療(–1h+1h)組(表1)。骨髓細(xì)胞培養(yǎng)與骨髓病理切片用實(shí)驗(yàn)動(dòng)物除上述4組外，增加正常對(duì)照組，每組6只。

1.3 化療藥物損傷小鼠模型制備 采用CTX 100mg/kg，1次/d，連續(xù)3d腹腔注射建立CTX損傷小鼠模型。

1.4 HS6101給藥方法 用1ml無菌注射用水將西林瓶封裝(2mg/支)的凍干粉劑HS6101溶解成2.0mg/ml儲(chǔ)存液，再以無菌生理鹽水將其稀釋至135μg/ml，每只小鼠皮下注射0.2ml，各組給藥時(shí)間見表1。CTX對(duì)照組每只小鼠注射0.2ml生理鹽水。

1.5 觀察指標(biāo) 每天2次觀察小鼠的一般狀況。血細(xì)胞計(jì)數(shù)：分別于注射CTX前1d及CTX首次注射后3、5、7、9、11、13、15、17d取尾靜脈血10μl檢測(cè)血常規(guī)。骨髓造血祖細(xì)胞集落培養(yǎng)：首次注射CTX后4、9d每組各取3只小鼠頸椎脫臼處死，無菌條件下取左側(cè)股骨，RPMI 1640培養(yǎng)液沖洗并分離骨髓單個(gè)核細(xì)胞進(jìn)行體外造血祖細(xì)胞集落培養(yǎng)，觀察骨髓有核細(xì)胞形成造血祖細(xì)胞集落的能力。骨髓病理切片：首次注射CTX后4、9d，每組各取3只小鼠頸椎脫臼處死，取右側(cè)股骨于Helly's液中固定24h后取出，再于4%甲醛溶液中固定1周，脫鈣后行石蠟切片，常規(guī)伊紅-蘇木素(HE)染色，光學(xué)顯微鏡下觀察骨髓組織病理學(xué)改變。

表1 實(shí)驗(yàn)動(dòng)物分組Tab.1 Group assignment of the experimental animals

1.6 統(tǒng)計(jì)學(xué)處理 采用SPSS 19.0進(jìn)行統(tǒng)計(jì)分析。計(jì)量資料以?±s表示，外周血常規(guī)數(shù)據(jù)采用重復(fù)測(cè)量的方差分析，造血細(xì)胞集落數(shù)的組間比較采用單因素k水平的方差分析(One-way ANOVA)，兩組間的比較采用LSD-t或Dunnet t檢驗(yàn)。P<0.05為差異有統(tǒng)計(jì)學(xué)意義。

2 結(jié) 果

2.1 實(shí)驗(yàn)鼠一般狀況 6101給藥后1d起6101治療組和6101預(yù)防+治療組動(dòng)物精神狀態(tài)即較CTX對(duì)照組、6101預(yù)防組變差，活動(dòng)減少，眼睛有分泌物且睜眼困難，第3天時(shí)癥狀緩解，基本恢復(fù)，且從第5天起出現(xiàn)動(dòng)物死亡，20d觀察期內(nèi)死發(fā)生亡率分別為16.7%(3/18)和50.0%(9/18)，CTX對(duì)照組及6101預(yù)防組動(dòng)物全部存活。

2.2 外周血白細(xì)胞、中性粒細(xì)胞和淋巴細(xì)胞數(shù)的變化 ICR小鼠連續(xù)3d腹腔注射CTX后外周血白細(xì)胞數(shù)迅速下降，CTX對(duì)照組、6101預(yù)防組、6101治療組、6101預(yù)防+治療組均于CTX首次注射后3d降至最低，分別為CTX注射前的36%、72%、30%、38%，但6101預(yù)防組明顯高于其他3組(P<0.01)，第5天各組白細(xì)胞數(shù)開始明顯升高，6101各給藥組均高于CTX對(duì)照組(P<0.05)；7d后白細(xì)胞數(shù)各組間差異無統(tǒng)計(jì)學(xué)意義(P>0.05)；第9天檢測(cè)值“反跳”達(dá)峰值，分別為CTX注射前的336%、261%、265%和284%，第17天檢測(cè)值基本回落至CTX注射前水平。6101對(duì)CTX損傷ICR小鼠外周血中性粒細(xì)胞數(shù)的影響與白細(xì)胞數(shù)基本一致。小鼠腹腔注射CTX后外周血淋巴細(xì)胞數(shù)迅速下降，第3天降至最低，CTX對(duì)照組、6101預(yù)防組、6101治療組、6101預(yù)防+治療組分別為CTX注射前的36%、66%、21%、43%，6101預(yù)防組明顯高于其他3組(P<0.05)，第9天升高達(dá)峰值，分別為290%、180%、250%和215%，之后回落波動(dòng)于CTX注射前水平(圖1)。

圖1 6101不同時(shí)間給藥對(duì)CTX損傷小鼠外周血白細(xì)胞、中性粒細(xì)胞和淋巴細(xì)胞數(shù)的影響Fig.1 Effect of the delivery time of 6101 on the peripheral blood leukocyte, neutrophil and lymphocyte count of CTX injured mice

2.3 外周血紅細(xì)胞數(shù)變化 6101預(yù)防組紅細(xì)胞數(shù)在注射CTX后第3、5、7天升高，第11天開始回落至CTX注射前水平，第3、5、7天紅細(xì)胞數(shù)均明顯高于CTX對(duì)照組(P<0.05)，第3天高于6101治療組和6101預(yù)防+治療組，第7天CTX對(duì)照組、6101預(yù)防組、6101治療組、6101預(yù)防+治療組紅細(xì)胞數(shù)檢測(cè)值分別為CTX注射前的86%、107%、113%和120%，6101各給藥組明顯高于CTX對(duì)照組(P<0.05，圖2)。

2.4 外周血血小板數(shù)變化 腹腔注射CTX后小鼠外周血小板數(shù)迅速降低，注射CTX后第3天，6101預(yù)防組血小板數(shù)明顯高于其他3組，而6101治療組和6101預(yù)防+治療組則明顯低于CTX對(duì)照組。注射CTX后1周左右血小板數(shù)開始恢復(fù)，6101預(yù)防組第7、9天的血小板數(shù)檢測(cè)值低于CTX對(duì)照組，但是第13和15天則明顯高于同期CTX對(duì)照組；6101治療組第9、11天檢測(cè)值低于CTX組，第15天高于CTX對(duì)照組，而且第11天和13天的檢測(cè)值明顯低于6101預(yù)防組；6101預(yù)防+治療組在第7、11天的血小板檢測(cè)值低于同期CTX對(duì)照組，第13和15天則高于同期CTX對(duì)照組。第17天時(shí)4組血小板數(shù)相當(dāng)，基本恢復(fù)至正常水平(表2)。

圖2 6101不同時(shí)間給藥對(duì)CTX損傷小鼠外周血紅細(xì)胞數(shù)的影響Fig. 2 Effect of the delivery time of 6101 on the peripheral blood erythrocyte count of CTX injured mice

表2 6101不同時(shí)間給藥的CTX損傷ICR小鼠外周血小板數(shù)(±s)Tab.2 Platelet count in peripheral blood of CTX injured ICR mice with different time of 6101 treatment (±s)

表2 6101不同時(shí)間給藥的CTX損傷ICR小鼠外周血小板數(shù)(±s)Tab.2 Platelet count in peripheral blood of CTX injured ICR mice with different time of 6101 treatment (±s)

(1)P<0.05, (2)P<0.01 compared with CTX group; (3)P<0.05, (4) P<0.01 compared with 6101-1h group

?

2.5 骨髓造血細(xì)胞集落數(shù)變化 CTX首次注射后4、9d取小鼠股骨骨髓細(xì)胞做體外集落形成實(shí)驗(yàn)，計(jì)數(shù)每組各系造血祖細(xì)胞集落數(shù)。CTX首次注射后第4天，CTX對(duì)照組小鼠骨髓造血干細(xì)胞形成粒巨噬細(xì)胞集落(GM-CFU)、巨核細(xì)胞集落(MKCFU)、爆式紅系(BFU-E)、紅細(xì)胞集落(CFU-E)和混合系集落(Mix-CFU)數(shù)量均明顯低于正常鼠，而6101預(yù)防組、治療組、預(yù)防+治療組的造血細(xì)胞集落形成能力均明顯改善，特別是對(duì)GM-CFU、MKCFU、BFU-E、CFU-E的生成有明顯刺激作用。盡管6101治療組小鼠在給藥后8d內(nèi)死亡未能做培養(yǎng)，但第9天時(shí)6101預(yù)防組和6101預(yù)防+治療組的GMCFU、MK-CFU、CFU-E和BFU-E集落形成數(shù)均高于CTX組，6101預(yù)防+治療組的GM-CFU、MK-CFU和BFU-E集落形成數(shù)與正常小鼠相當(dāng)(圖3)。

圖3 6101不同時(shí)間給藥對(duì)CTX損傷小鼠骨髓細(xì)胞集落數(shù)的影響Fig.3 Effect of the delivery time of 6101 on bone marrow cells colony-forming unit (CFU) of CTX injured mice

2.6 骨髓病理組織切片觀察 分別在CTX首次注射后第4、9天處死小鼠，取股骨做病理組織切片，光學(xué)顯微鏡下觀察發(fā)現(xiàn)，第4天時(shí)CTX對(duì)照組小鼠骨髓細(xì)胞數(shù)量中等，類型多樣，細(xì)胞密度局部適中，部分骨髓細(xì)胞呈條索狀排列，散在空腔內(nèi)無骨髓細(xì)胞，且血竇內(nèi)充滿紅細(xì)胞。與CTX對(duì)照組比較，6101預(yù)防組小鼠骨髓細(xì)胞數(shù)量明顯增多，細(xì)胞密度大，脂肪和空隙較少，血竇內(nèi)紅細(xì)胞較多。6101治療組小鼠骨髓細(xì)胞數(shù)量、密度和組織結(jié)構(gòu)基本同6101預(yù)防組。與CTX對(duì)照組比較，6101預(yù)防+治療組小鼠骨髓細(xì)胞數(shù)量很多，密度非常大，巨核細(xì)胞數(shù)量似較少，細(xì)胞間空隙很小(圖4)。對(duì)骨髓病理切片進(jìn)行綜合評(píng)分，以正常小鼠骨髓為滿分(10分)，CTX對(duì)照組、6101預(yù)防組、6101治療組和6101預(yù)防+治療組的評(píng)分分別為6、8、8、9分，6101預(yù)防+治療組優(yōu)于其他各組。第9天時(shí)，CTX對(duì)照組小鼠骨髓細(xì)胞數(shù)量很多，密度非常高，除局部幾處稍大空隙外，骨髓細(xì)胞間空隙很小，脂肪細(xì)胞和血竇紅細(xì)胞少。與CTX對(duì)照組比較，6101預(yù)防組小鼠骨髓細(xì)胞數(shù)量很多，密度非常高，細(xì)胞間空隙很少，脂肪細(xì)胞和血竇紅細(xì)胞少。與CTX對(duì)照組比較，6101預(yù)防+治療組小鼠骨髓細(xì)胞數(shù)量很多，密度非常高，細(xì)胞間空隙很少，脂肪細(xì)胞和血竇紅細(xì)胞少。CTX注射后后第9天各組骨髓細(xì)胞超常增多，已無法對(duì)病理切片進(jìn)行綜合評(píng)分。

圖4 CTX首次注射后4d小鼠骨髓病理組織切片(HE染色×100)Fig.4 Bone marrow histopathology of ICR mice on day 4 after the first administration of CTX (HE staining ×100)

3 討 論

HS6101是人工合成的含9肽氨基酸殘基的脂肽化合物，屬于Toll樣受體激動(dòng)劑，與CBLB613類似，在小鼠體內(nèi)可刺激誘導(dǎo)生成IL-1β、IL-6、IL-10、IL-12、KGF、G-CSF、GM-CSF、TNF-1α等細(xì)胞因子，產(chǎn)生顯著的輻射防護(hù)作用[4-5]，但目前尚未見HS6101促進(jìn)化療后造血功能恢復(fù)的相關(guān)文獻(xiàn)報(bào)道。鑒于HS6101能刺激體內(nèi)上述細(xì)胞因子的生成，推測(cè)其可能促進(jìn)化療后骨髓造血功能的恢復(fù)。在前期工作中，我們觀察到HS6101預(yù)防給藥對(duì)CTX引起的小鼠造血細(xì)胞損傷有改善作用。由于腫瘤患者常常會(huì)因病情、體質(zhì)狀況或其他原因在化療后才能接受輔助治療，或者需要在化療前后均接受輔助治療，為了明確給藥時(shí)間對(duì)HS6101作用效果的影響，本實(shí)驗(yàn)重點(diǎn)觀察了HS6101在首次CTX化療前1h給藥(-1h)、末次CTX后1h給藥(+1h)以及CTX前后均給藥(-1h+1h)對(duì)CTX損傷小鼠造血功能的影響。

本實(shí)驗(yàn)結(jié)果表明，HS6101于首次注射CTX前1h單次皮下注射給藥對(duì)CTX所致小鼠造血系統(tǒng)損傷有較好的預(yù)防作用，可明顯改善化療藥物引起的外周血白細(xì)胞、中性粒細(xì)胞、血小板、淋巴細(xì)胞和紅細(xì)胞減少，并在一定程度上促進(jìn)外周血白細(xì)胞與淋巴細(xì)胞數(shù)的恢復(fù)。此外，還能減輕CTX引起的造血干細(xì)胞增殖分化能力下降并促進(jìn)其恢復(fù)，而且對(duì)骨髓造血組織的結(jié)構(gòu)和造血細(xì)胞均有改善和保護(hù)作用，觀察期內(nèi)該組動(dòng)物全部存活。6101于造模后1h單次注射或分別在造模前1h與造模后1h各1次注射給藥對(duì)CTX引起的外周血細(xì)胞減少無明顯改善作用，但當(dāng)造血開始恢復(fù)后，則可在一定程度上促進(jìn)小鼠白細(xì)胞、中性粒細(xì)胞、紅細(xì)胞和淋巴細(xì)胞數(shù)的恢復(fù)，這與它能夠減輕CTX引起的小鼠造血干細(xì)胞增殖分化能力下降并改善骨髓造血組織結(jié)構(gòu)、促進(jìn)造血干細(xì)胞分化有關(guān)。然而，6101于造模后1h單次注射或分別在造模前1h與造模后1h各1次注射給藥對(duì)血小板的恢復(fù)均無促進(jìn)作用，相反，似乎有加劇早期血小板減少的趨勢(shì)，該現(xiàn)象值得關(guān)注并有待進(jìn)一步驗(yàn)證。血小板嚴(yán)重減少會(huì)導(dǎo)致死亡，6101治療組和6101預(yù)防+治療組早期血小板數(shù)明顯低于CTX對(duì)照組和6101預(yù)防組可能是前兩組動(dòng)物出現(xiàn)死亡、而后兩組動(dòng)物全部存活的主要原因，該結(jié)果提示HS6101不宜于CTX損傷后給藥。綜合分析本實(shí)驗(yàn)結(jié)果，我們認(rèn)為HS6101于首次化療前1h給藥促進(jìn)CTX損傷ICR小鼠造血功能恢復(fù)的效果優(yōu)于其他兩個(gè)時(shí)間點(diǎn)給藥。此外，G-CSF等細(xì)胞因子[6-8]或其他輔助治療藥物[9-11]需多次注射或數(shù)十天口服給藥，而HS6101僅需單次皮下注射給藥，具有明顯的優(yōu)勢(shì)。

化療藥物引起的白細(xì)胞和血小板減少等并發(fā)癥一直是導(dǎo)致腫瘤患者難以順利足量完成化療的主要原因，而本研究結(jié)果顯示，HS6101在首次注射CTX 前1h單次皮下注射給藥可明顯促進(jìn)CTX損傷ICR小鼠造血功能恢復(fù)，為腫瘤化療患者的輔助治療提供了新的選擇。

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Effect of the administration time of HS6101 on hematopoietic recovery in ICR mice injured by cyclophosphamide

XING Shuang1, SHEN Xing1, XIONG Guo-lin1, LIU Xiao-lan1, YANG Meng1, DAI Chang-liang2, LIU Xiao-yu2, YU Zuyin1*1Institute of Radiation Medicine, Academy of Military Medical Sciences, Beijing 100850, China
2Zhejiang Hisun Pharmaceutical Co., Ltd, Taizhou, Zhejiang 318000, China
*

, E-mail: yuzy79@163.com
This work was supported by the Military Key Technology Research and Development Program in the Major Drug Discovery of PLA (2010ZX09401)

ObjectiveTo explore the effect of the administration time of HS6101 on hematopoietic recovery in ICR mice injured by cyclophosphamide (CTX).MethodsOne hundred and three male ICR mice were divided into 4 groups: CTX control, HS6101 prevention, HS6101 treatment, and HS6101 prevention+treatment groups. CTX was intraperitoneally injected into the ICR mice at a dose of 100mg/(kg.d) for three consecutive days to establish a chemotherapeutics-injured model. HS6101 at a dose of 27μg/mouse in 0.2ml was subcutaneously injected into the mice 1h before the first administration of CTX in HS6101-preventiongroup, 1h after the last administration of CTX in HS6101 treatment group, and both at 1h before the first administration and 1h after the last administration of CTX in HS6101 prevention + treatment group. Physiological saline was subcutaneously injected into the mice in CTX control group (0.2ml/mouse). 10μl peripheral blood was collected from the caudal vein for WBC, neutrophil lymphocyte, RBC and platelet counts on day -1, 3, 5, 7, 9, 11, 13, 15, 17 with the MEK-7222K cell analyzer, and the cell count was compared between HS6101 treatment mice and CTX control mice. Another 30 male ICR mice were used for bone marrow colony forming unit (CFU) assay and bone marrow histopathological examination, and they were assigned into normal control, CTX control, HS6101 prevention, HS6101treatment and HS6101 prevention + treatment groups (each n=6). On the day 4 and day 9 after CTX injection, mice were sacrificed and bone marrow cells were collected from the left femur for mononuclear cell (MNC) isolation. 1×104MNCs were planted in 1.0ml mouse CFU culture medium M3434 and cultured in incubator with the temperature of 37℃, and 5% CO2for 7 days. After that, granulocyte macrophage-colony-forming unit (GM-CFU), megakaryocyte colony forming unit (MK-CFU), mixture-colony-forming unit (Mix-CFU), burst-forming unit-erythroid (BFU-E) and colony-forming unit-erythroid (CFU-E) were counted. Then the right femur was taken for histopathology examination.ResultsAfter CTX injection, counts of WBC, neutrophils, lymphocytes, RBC and platelets of all the mice decreased rapidly. However, the nadirs of WBC, neutrophils and lymphocytes counts in HS6101 prevention group were higher than those in CTX control group, and the counts on day 3 were higher than those in HS6101 treatment group and HS6101 prevention+ treatment group. On day 3, RBC count in HS6101 prevention group was the highest. It was higher on day 5 and day 7 than that of mice in CTX group. In addition, the platelet count in HS6101 prevention group was also the highest on day 3, although that in HS6101 treatment group and 6101 prevention + treatment group was lower than CTX control group. Bone marrow colony forming unit assay showed that the counts of GM-CFU, MK-CFU, BFU-E and CFU-E in all the HS6101 treatment mice were significantly higher than those in CTX control mice. On day 4, histopathological examination of bone marrow from HS6101-treated mice displayed more intact architecture compared with CTX control mice. Three of eighteen (3/18) mice died in HS6101 treatment group, and nine of eighteen (9/18) died in HS6101 prevention + treatment group, suggesting that HS6101 should not be administered after CTX injection.ConclusionAdministration of HS6101 at 1h before giving CTX could significantly promote hematopoietic recovery in ICR mice injured by CTX.

HS6101; lipopeptides; cyclophosphamide; mice; hematopoietic system

R322.2

A

0577-7402(2015)04-0303-06

10.11855/j.issn.0577-7402.2015.04.10

2014-07-11；

2015-02-22)

(責(zé)任編輯：李恩江)

軍隊(duì)“重大新藥創(chuàng)制”科技重大專項(xiàng)課題(2010ZX09401)

邢爽，理學(xué)博士，助理研究員。主要從事急性放射病實(shí)驗(yàn)治療方面的研究

100850 北京 軍事醫(yī)學(xué)科學(xué)院放射與輻射醫(yī)學(xué)研究所(邢爽、申星、熊國林、柳曉蘭、楊萌、余祖胤)；318000 浙江臺(tái)州 浙江海正藥業(yè)股份有限公司(代常亮、劉曉宇)

余祖胤，E-mail：yuzy79@163.com