CN112575073A - c-Kit作为成瘾诊断及监测标志物的应用 - Google Patents
c-Kit作为成瘾诊断及监测标志物的应用 Download PDFInfo
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Abstract
本发明公开了c‑Kit作为成瘾诊断标志物的应用，属于医药技术领域。本发明采用免疫组化、免疫荧光等分子检测技术及靶向分子成像技术，证明急性吗啡给药可明显增强大鼠伏隔核区域c‑kit蛋白磷酸化表达。同时，采用定量PCR技术检测发现与未成瘾大鼠相比，吗啡成瘾大鼠外周血c‑kit mRNA表达量明显升高。最后采用评价药物成瘾的小鼠条件位置偏爱动物模型，发现c‑kit抑制剂甲磺酸伊马替尼可有效预防吗啡成瘾形成及其复吸。因此c‑Kit可作为药物成瘾诊断标志物用于制备诊断药物成瘾，治疗及复吸预防后监测生物标志物，以此开发相应的蛋白和核酸等相关产品。
Description
技术领域
本发明属于医药技术领域，具体涉及c-Kit作为成瘾诊断及监测标志物的应用。
背景技术
物质成瘾和行为成瘾是全球严重公共卫生问题，机制不清，缺乏有效诊断和治疗措施。对于物质成瘾，尤其在禁毒戒毒实践中，唾液、尿液等样品是毒品检测中最常规的样品类型。由于吸食毒品后的中间代谢物在体液中存在的周期较短、毒品代谢产物变化以及定性检测方法灵敏性受到限制，使社会上一些非法毒品滥用者通过选择性临时停吸的方法进行规避检查，致使无法准确判定。同样，毛发检测虽然可以提供被检测人员的长期用药信息，但毛发护理等操作措施降低头发中毒品的保留，致使检测率降低。最近，美国国家卫生研究院(National Institutes of Health，NIH)宣布△FosB蛋白可作为一种诊断生物标记，其表达水平可用于诊断人类中的成瘾，但需要脑部活检，临床应用受到限制。因此寻找特异性强、灵敏度高、稳定性好、操作方便的生物学指标对成瘾诊断和检测，以及治疗后监测具有重要的意义。
c-Kit基因位于人染色体4q11-12，属于原癌基因，编码的c-Kit受体是Ⅲ型酪氨酸激酶受体蛋白家族成员之一。作为c-Kit受体的配体，干细胞因子(stem cell factor，SCF)与c-Kit结合后可激活下游多种信号转导通路，包括PI3K/Akt、Ras/ERK、PLC-γ/PKC等在药物奖赏效应和记忆中起着重要作用的信号通路。申请人已经发现(CN 105974131A)大鼠吗啡成瘾后，伏隔核核部c-kit的磷酸化活性显著升高，且c-kit抑制剂-伊马替尼及其类似物(CN 106074555A)通过抑制c-kit磷酸化改善吗啡成瘾症状。最近研究表明，外泌体(exosomes)作为一种细胞主动分泌到胞外的直径为30～100nm间的微小囊泡，大量存在于血液、尿液、唾液等体液中，含有蛋白质、脂质、遗传物质(如mRNA、miRNA、LncRNA等)等物质，外泌体稳定性极高，含量也很丰富；同样，神经元可分泌外泌体，而且中枢神经系统(CNS)外泌体能自由穿过人体血脑屏障(BBB)进入外周循环，外周循环中的外泌体也能通过BBB进入颅内发挥其作用。外泌体释放到循环血液中，反映释放细胞的功能状态，各种内容物未遭破坏并发挥相应的生理作用，这些特点使得外泌体可作为疾病诊断的循环生物标志物。因此，检测外泌体来源的c-Kit分子内容物状态，可望为物质依赖和行为依赖诊断、治疗后监测带来新的契机。
发明内容
针对现有技术存在的问题，本发明目的之一在于提供c-Kit相关活性产物作为物质和行为成瘾诊断与治疗后监测生物标志物的应用。
本发明的目的之二在于提供一种通过示踪或检测及监测c-kit基因或c-kit蛋白产物活性来反映物质或行为成瘾状态的产品。
本发明主要通过以下技术方案来体现：
本发明通过免疫组化和免疫荧光，分子生物学检测技术及分子靶向成像技术验证急性吗啡给药与治疗后以及其他成瘾状态大鼠的伏隔核等脑部区域c-kit磷酸化水平的变化；进一步通过实时定量PCR技术分析吗啡成瘾大鼠外周血浆中c-kit mRNA表达量与药物成瘾的相关性；最后通过小鼠条件性位置偏爱(Conditioned Place Preference,CPP)模型，探究c-kit抑制剂-伊马替尼系统给药对小鼠吗啡CPP成瘾形成及再巩固的影响，结果表明c-kit活性产物能够作为判定药物成瘾病理状态的诊断标志物，在禁毒戒毒实践和其他成瘾治疗中具有重要的应用价值。通过以上检测，提供检测c-kit相关表达产物在制备成瘾诊断与治疗后监测产品中的应用。所述产品包括试纸、试剂盒、芯片、高通量测序平台或影像学检测等各种用于检测c-kit活性的产品。这些产品基于包括反转录PCR、荧光实时定量PCR、免疫检测、原位杂交、芯片、高通量测序平台或脑功能磁共振、组学等各种方法来实现检测c-kit基因、RNA、蛋白活性的目的。
本发明所述的成瘾物质是指麻醉药品和精神药物等。其中麻醉药品可分为阿片类、可卡因类和大麻类；精神药物分为镇静催眠药、抗焦虑药、中枢兴奋剂及致幻剂等；其他的也包括酒精、烟草和挥发性有机溶剂等；成瘾行为是指食物成瘾、网瘾、赌博成瘾等行为。
与现有成瘾诊断与治疗检测技术相比，本发明可以获得包括以下技术效果：
本发明发现了一种诊断药物成瘾的分子标志物，使用该分子标志物可以在药物成瘾发生的早期即可作为判断，防止吸毒人员对自身或社会造成更大的危害；同时可检测成瘾动态病理变化过程，用于治疗后监测。
附图说明
图1为免疫组化结果图；A为急性吗啡大鼠脑区c-kit表达图；B为甲磺酸伊马替尼给药后急性吗啡大鼠脑区c-kit表达图。
图2为大鼠近红外二区荧光脑部活体成像监测分析图。
图3为吗啡成瘾大鼠外周血浆外泌体内c-kit mRNA表达水平检测结果；A为给药实验流程图；B为血浆外泌体内c-kit mRNA分析检测结果图。
图4为甲磺酸伊马替尼对小鼠吗啡成瘾形成的影响结果图；A为给药实验流程图；B为甲磺酸伊马替尼对CPP分值的影响。
图5为甲磺酸伊马替尼对小鼠吗啡成瘾后心理渴求的影响结果图；A为给药实验流程图；B为甲磺酸伊马替尼对CPP分值的影响。
具体实施方式
下述实施例都是示意性的，仅对本发明作进一步详细的说明，构成本发明一部分，但限制本发明的范围。
下述实施例所采用的阿片类药物为吗啡，其它阿片类药物与吗啡具有相似的作用机制，吗啡具有广泛的代表性，所属领域技术人员可以在其他的阿片类药物中重现类似的研究结果。其他成瘾性物质有可卡因、酒精、尼古丁等，成瘾性行为有食物成瘾和赌博成瘾等。下述实施例中所用的材料、试剂等，如无特殊说明，均可从商业途径得到。
实施例1：急性吗啡给药对大鼠脑区c-Kit表达的影响
一、材料
药品：吗啡(青海制药厂)，甲磺酸伊马替尼(Selleck Chemicals)。
实验动物：SPF级SD雄性大鼠，体重180-220g，购自三峡大学动物实验中心，动物合格证号为NO.42010200001637，生产许可证号：SCXK(鄂)2017-0012。鼠饲料，购于武汉大学实验动物中心。
二、实验方法
(1)免疫组织化学检测成瘾相关脑区c-Kit活性变化及其信号转导通路激活情况
实验大鼠分为溶剂组(生理盐水+生理盐水组、生理盐水+甲磺酸伊马替尼组)和吗啡组(吗啡+生理盐水组、吗啡+甲磺酸伊马替尼组)(n＝5)，分别经腹腔给药给予大鼠1mL/kg溶剂或30mg/kg甲磺酸伊马替尼，30min后分别经皮下给药给予大鼠1mL/kg生理盐水或10mg/kg吗啡。1h后麻醉灌流，脑组织用4％多聚甲醛溶液固定，在不同浓度酒精溶液中脱水、二甲苯透明后，包埋于石蜡中，然后切成3μm厚的切片。将组织脱蜡，并在0.01M枸橼酸钠缓冲液(pH6.0)中煮沸10-15min，并在冷水中冷却降温至室温。然后进行免疫组化和免疫荧光共定位多标检测成瘾激活相关脑区c-Kit活性变化及其信号转导通路激活情况及甲磺酸伊马替尼的阻断作用，以确定成瘾激活c-Kit特异脑区以及甲磺酸伊马替尼的预防作用。
(2)靶向分子影像学技术检测成瘾相关脑区c-Kit活性动态变化
NIR-II荧光成像分析：随机将实验大鼠分为溶剂组、急性吗啡组和急性吗啡+甲磺酸伊马替尼组(n＝5)，分别经腹腔给药给予大鼠1mL/kg生理盐水或30mg/kg甲磺酸伊马替尼，30min后，分别经皮下给药给予实验大鼠1mL/kg生理盐水或10mg/kg吗啡，随后尾部静脉给药给予实验大鼠100μg PEG1000-c-Kit荧光探针，在NIR-II荧光成像指导下动态用近红外二区成像相机监测拍摄大鼠分别在2h、6h、8h、12h后脑部区域动态变化，以此来表示c-Kit蛋白磷酸化水平变化。12h检测完后，大鼠深度麻醉并断头取脑，进一步脑区定位并清晰观测脑部c-Kit活性变化。以确定成瘾激活c-Kit特异脑区以及甲磺酸伊马替尼治疗后c-Kit活性动态变化监测效果。
三、实验结果
免疫组化结果见图1，与正常大鼠相比，大鼠急性吗啡给药1h后伏隔核区域c-Kit蛋白磷酸化水平明显升高，前额皮层、海马等区域无明显变化(图1A)；大鼠给予甲磺酸伊马替尼后，伏隔核、前额皮层、海马等脑部区域c-Kit蛋白磷酸化活性明显下降(图1B)。NIR荧光成像检测也发现急性吗啡给药大鼠脑部亮度越来越亮，探针富集强度在6h后达到高峰，与生理盐水组形成明显对比，说明急性吗啡给药可明显增强大鼠脑部c-Kit磷酸化活性，并在注射c-Kit抑制剂甲磺酸伊马替尼后大鼠脑部亮度减弱，吗啡激活的c-Kit蛋白磷酸化活性得到抑制(图2)。
实施例2：吗啡成瘾大鼠血浆外泌体内c-Kit mRNA表达水平变化
一、材料
药品：吗啡(青海制药厂)。
实验动物：SPF级SD雄性大鼠，体重180-220g，购自三峡大学动物实验中心，动物合格证号为NO.42010200001637，生产许可证号：SCXK(鄂)2017-0012。鼠饲料，购于武汉大学实验动物中心。
二、实验方法
随机将实验大鼠分成两组，溶剂组和吗啡组(n＝5)，给药实验流程如图3A所示，吗啡组连续6天分别经皮下给药给予实验大鼠5、10、15、20、25、25mg/kg吗啡，生理盐水组每日皮下注射1mL/kg生理盐水，并于24h后，大鼠摘眼球取血4mL。
分离血浆：采血后将全血转移至洁净的1.5mL离心管(含有140μL的50mM EDTA-Na2)，立即在4℃条件下1500rpm离心15min，提取上清；随后再次以3000rpm离心10min，提取上清。
外泌体提取：
(1)在各个样本离心的1mL上清液中加入4mL 4℃预冷的PBS溶液进行稀释，再加入1mL BPS试剂(Blood PureExo Solution)充分混匀；
(2)静置2h后在4℃以10000rpm离心60min，弃去上清液，沉淀中富含外泌体颗粒。取400ul PBS溶液均匀吹打离心沉淀物，并移至新的1.5mL离心管，再于4℃以下以12000rpm离心2min，保留富含外泌体颗粒的上清液；
(3)将上清液放入EPF柱(Exosome Purafication Filter)中，于4℃以3000rpm离心10min，此时收集的EPF柱管底的液体即为纯化后的外泌体颗粒。
外泌体RNA提取：
(1)往上述提取的外泌体中加入200μL氯仿及1000μL Trizol裂解液，剧烈震荡15s，室温静置8min，再于4℃以12000rpm离心15min；
(2)取上层水相移至新的1.5mL Eppendorf管，加入等体积的异丙醇小心翻转混匀，并在4℃冰箱静置沉淀10min，再于4℃以12000rpm离心10min，保留底部白色沉淀；
(3)加入1mL经DEPC处理过的水新鲜配制的75％乙醇洗涤沉淀，于4℃以10000rpm离心5min，再于4℃以10000rpm离心5min，并保留沉淀，并干燥5min；
(4)向该干燥后沉淀中加入20μL Nuclease-free water溶解RNA，得到外泌体RNA溶液，并用紫外分析测定所抽提RNA的浓度。
外泌体RNA逆转录：
(1)以20μL反应体系为例，按下表所列加入各组分，配置成混合液与RNA溶液混匀，于42℃温育5min；
反转录PCR体系：
| 试剂 | 使用量(u1) |
| 5XPrime Script Buffer(for Real Time) | 4 |
| Prime Script RT Enzyme Mix I | 1 |
| RT Primer Mix | 1 |
| Total RNA/对照样本 | 10 |
| RNase Free dH2O | 4 |
| Total | 20ul |
(2)加入15U的AMV反转录酶，混匀后于37℃反应15min，随后于85℃煮样5s，使AMV反转录酶失活终止反应，冰浴5min，此时获得第一链cDNA，4℃保存。
实时定量PCR扩增：
(1)以上述cDNA为模板，使用Premix Ex TaqTM试剂盒(TakaRa)，在ABI steponeplus实时荧光定量PCR仪上进行操作。
其中，c-Kit mRNA上游引物为5’-cgcagcttccttatga ccac-3’(SEQ ID NO.1)，下游引物为5’-agtggcctcaactaccttcc-3’(SEQ ID NO.2)；
荧光定量检测内参基因GAPDH的mRNA表达量的上游引物为5’-ttcaacggcacagtcaagg-3’，下游引物为5’-ctcagcaccagcatcacc-3’。
(2)反应体系为：
| 试剂 | 体积(μL) |
| 模板cDNA | 2 |
| 上游引物 | 0.8 |
| 下游引物 | 0.8 |
| SYBR Premix Ex TaqII(2×) | 10 |
| ROX Reference Dye(50×) | 0.4 |
| 去离子水 | 6 |
| 总体积 | 20 |
(3)荧光PCR扩增程序为：
(4)每个样品均重复三次取平均值，以保证定量的精确度。
统计分析：以GAPDH为内参基因，对c-Kit mRNA进行归一化处理，以确保在相等数量的样本中比较c-Kit mRNA的表达量，c-Kit mRNA相对表达量＝2-△△Ct，其中△△Ct＝吗啡成瘾大鼠血浆外泌体内c-Kit mRNA平均△Ct值-(吗啡成瘾大鼠血浆外泌体内c-Kit mRNACt值-吗啡成瘾大鼠血浆外泌体内GAPDH基因Ct值)，得出c-Kit mRNA表达量相对于内参基因表达量的变化倍数。
三、实验结果
结果见图3B，与未成瘾大鼠相比，吗啡成瘾大鼠后外周血浆内c-Kit mRNA表达水平明显升高，差异具有统计学意义(p<0.05)。
实施例3：甲磺酸伊马替尼对小鼠吗啡成瘾形成的影响
一、材料
药品：吗啡(青海制药厂)；甲磺酸伊马替尼(Selleck Chemicals)。
实验动物：SPF级昆明品系雄性小鼠，体重18-22g，购自三峡大学动物实验中心，动物合格证号为NO.42010200001676，生产许可证号：SCXK(鄂)2017-0012。鼠饲料，购于武汉大学实验动物中心。
实验仪器：条件性位置偏爱仪(中国医学科学院药物研究所研制)：实验采用计算机自动控制。装置由三箱构成的条件性位置偏爱箱：两个侧室和一个中间室。三室由可移动的隔板分开，内外均为黑色。其中A箱和B箱位于中间箱的两侧，大小相同，A箱侧壁上有9盏能发出黄光二极管构成的正方形，底板为不锈钢钢条，B箱底板为不锈钢网格。大鼠在各箱的停留时间和出入次数可通过数据传送到计算机，自动收集记录行为学资料。
二、实验方法
动物分组与处理：小鼠随机分为四组，分别为对照组：①生理盐水+溶剂组、②生理盐水+甲磺酸伊马替尼给药组和实验组：③吗啡+溶剂组、④吗啡+甲磺酸伊马替尼给药组(n＝10)。
吗啡CPP模型建立
基础值测试：第1天，拿掉隔板，开放三箱的通道，启动计算机上CPP程序，小鼠由中间室放入，任其在三箱中自由活动15分钟，电脑同步记录其在各室中停留的时间。根据测试结果进行淘汰，分组，并区分各小鼠的伴药侧与非伴药侧。
条件性位置偏爱训练：训练示意图如图4A所示，第2至9天，封闭三箱间的通道。第2、4、6、8天，实验组皮下注射吗啡(15mg/kg)并放入伴药侧45分钟，其中实验组③在注射吗啡前30分钟给予腹腔注射生理盐水(1mL/kg)，实验组④给予腹腔注射甲磺酸伊马替尼(45mg/kg)；对照组皮下注射生理盐水(1mL/kg)并放入非伴药侧45分钟，其中对照组①在注射吗啡前30分钟给予腹腔注射生理盐水(1mL/kg)，对照组②给予腹腔注射甲磺酸伊马替尼(45mg/kg)。第3、5、7、9天，实验组及对照组小鼠均皮下注射生理盐水，实验组放入非伴药侧，对照组放入伴药侧，均为45分钟。每只小鼠的伴药侧是固定的，小鼠在每天训练完后被放回饲养笼。
吗啡CPP测试：第10天，CPP测试，与基础值测试阶段相似。开放三箱间的通道，不予任何注射，启动计算机上CPP程序，小鼠由中间室放入，任其在箱中自由活动15分钟，电脑同步记录其在各室中停留的时间。偏爱分数(CPP score)被定义为伴药室所呆时间与非伴药室所呆时间的差值。将小鼠在伴药箱中CPP后测值与前侧值比较确定小鼠是否形成CPP。
三、实验结果
结果见图4B，与未注射甲磺酸伊马替尼小鼠相比，注射甲磺酸伊马替尼后小鼠条件位置偏爱无法正常形成，说明甲磺酸伊马替尼可通过抑制c-Kit磷酸化活性预防吗啡成瘾。
实施例4：甲磺酸伊马替尼对吗啡成瘾小鼠潜伏心理渴求的影响
一、材料
药品：吗啡(青海制药厂)；甲磺酸伊马替尼(Selleck Chemicals)。
实验动物：SPF级昆明品系雄性小鼠，体重18-22g，购自三峡大学动物实验中心，动物合格证号为NO.42010200001676，生产许可证号：SCXK(鄂)2017-0012。鼠饲料，购于武汉大学实验动物中心。
实验仪器：条件性位置偏爱仪(中国医学科学院药物研宄所研制)：实验采用计算机自动控制。装置由三箱构成的条件性位置偏爱箱：两个侧室和一个中间室。三室由可移动的隔板分开，内外均为黑色。其中A箱和B箱位于中间箱的两侧，大小相同，A箱侧壁上有9盏能发出黄光二极管构成的正方形，底板为不锈钢钢条，B箱底板为不锈钢网格。小鼠在各箱的停留时间和出入次数可通过数据传送到计算机，自动收集记录行为学资料。
二、实验方法
动物分组与处理：小鼠随机分为四组，分别为生理盐水+溶剂组、生理盐水+磺酸伊马替尼给药组和吗啡+溶剂组、吗啡+甲磺酸伊马替尼给药组。给药实验流程如图5A所示。
(1)吗啡CPP模型建立
基础值测试：第1天，拿掉隔板，开放三箱的通道，启动计算机上CPP程序，小鼠由中间室放入，任其在三箱中自由活动15分钟，电脑同步记录其在各室中停留的时间。根据测试结果进行淘汰，分组，并区分各小鼠的伴药侧与非伴药侧。
条件性位置偏爱训练：第2至9天，封闭三箱间的通道。第2、4、6、8天，实验组皮下注射吗啡(10mg/kg)并放入伴药侧45分钟；对照组皮下注射生理盐水(1mL/kg)并放入非伴药侧45分钟。第3、5、7、9天，实验组及对照组小鼠均皮下注射生理盐水，实验组放入非伴药侧，对照组放入伴药侧，均为45分钟。每只小鼠的伴药侧是固定的，小鼠在每天训练完后被放回饲养笼。
吗啡CPP测试：第10天，CPP测试，与基础值测试阶段相似。开放三箱间的通道，不予任何注射，启动计算机上CPP程序，小鼠由中间室放入，任其在箱中自由活动15分钟，电脑同步记录其在各室中停留的时间。偏爱分数(CPP score)被定义为伴药室所呆时间与非伴药室所呆时间的差值。将小鼠在伴药箱中CPP后测值与前侧值比较确定小鼠是否形成CPP。
(2)环境线索诱发觅药行为模型的建立
在实验的第10天，每组小鼠分别腹腔给药甲磺酸伊马替尼(45mg/kg)与生理盐水(1mL/kg)，30min后，将各组小鼠分别暴露于伴药箱，停留15min，再回到笼养环境中。
(3)吗啡CPP重新测试
第12和18天，测试小鼠对伴药箱的偏爱程度，与基础值测试阶段相似，中间第13-17天，不做任何处理。
(4)吗啡CPP的点燃
第17天，利用小剂量的吗啡(5mg/kg，i.p.)进行点燃。吗啡注射后5分钟，将小鼠放入中间箱，开始15分钟的CPP值测试。
三、实验结果
结果见图5B，小鼠条件位置偏爱形成后，环境线索再暴露前给予生理盐水和甲磺酸伊马替尼后检测CPP Score，发现给予生理盐水小鼠条件位置偏爱依然存在：而给予甲磺酸伊马替尼小鼠CPP Score明显降低，抑制给药环境引起的心理渴求，且1周后不被点燃。给药组与对照组相比差异具有显著性，说明c-Kit被抑制后可明显改善吗啡成瘾症状。
结论：急性吗啡给药后可通过免疫组化和NIR-II荧光成像可明显看到增强大鼠脑部伏隔核区域c-Kit蛋白磷酸化表达水平，并且大鼠吗啡成瘾后外周血浆内c-Kit mRNA表达水平也明显增加，甲磺酸伊马替尼可明显抑制c-Kit活性预防和治疗成瘾，并防治复燃；当c-Kit磷酸化活性被抑制后，小鼠吗啡成瘾无法形成，且吗啡戒断后复吸也被明显抑制。上述结果说明c-Kit可以作为成瘾的诊断生物标志物，并且通过甲磺酸伊马替尼抑制c-Kit蛋白磷酸化活性后，抑制吗啡引起的成瘾记忆形成、记忆再巩固及戒断后复吸，说明成瘾所致c-Kit活化后其相关产物包括蛋白、核酸等可作为诊断成瘾和监测其治疗效果的诊断标志物，在未来戒毒禁毒及其他类型的成瘾治疗中具有重要的意义。
上述实施例为本发明较佳的实施方式，但本发明的实施方式并不受上述实施例的限制，其他的任何未背离本发明的精神实质与原理下所作的改变、修饰、替代、组合、简化，均应为等效的置换方式，都包含在本发明的保护范围之内。
Claims (6)
1.c-Kit在制备物质成瘾或行为成瘾诊断、治疗后监测的产品中的应用。
2.一种通过示踪或检测及监测成瘾患者体内c-kit各种RNA、DNA或c-kit蛋白活性、相关代谢产物来反映物质或行为成瘾状态的产品。
3.根据权利要求1所述的应用或权利要求2所述的产品，其特征在于：所述的物质包括麻醉药品、精神药物、酒精、烟草、挥发性有机溶剂等，其中麻醉药品可分为阿片类、可卡因类和大麻类等药品；精神药物分为镇静催眠药、抗焦虑药、中枢兴奋剂及致幻剂等；所述的行为包括网瘾、赌博成瘾等各种行为。
4.根据权利要求1所述的应用或权利要求2所述的产品，其特征在于：所述的产品包括试纸、试剂盒、芯片、高通量测序平台或影像学等在体内或体外诊断产品。
5.根据权利要求1所述的应用或权利要求2所述的产品，其特征在于：所述的产品能够通过检测样本中c-kit基因、RNA和蛋白的表达或其相关代谢产物来诊断和监测物质成瘾和行为成瘾；所述样本包括但不局限于血液、尿液、唾液等体液和组织。
6.根据权利要求1所述的应用或权利要求2所述的产品，其特征在于：所述的产品基于包括反转录PCR、荧光实时定量PCR、免疫检测、原位杂交、芯片、高通量测序平台或脑功能磁共振、组学等各种方法来实现检测c-kit基因、RNA、蛋白活性的目的。
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2023102991A1 (zh) * | 2021-12-06 | 2023-06-15 | 中国科学院深圳先进技术研究院 | 一种成瘾药物戒断后复吸相关调控基因及其筛选方法 |
| CN116497031A (zh) * | 2023-06-02 | 2023-07-28 | 徐州医科大学 | 一种缓解小鼠吗啡成瘾行为的siRNA及其应用 |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2003072106A2 (en) * | 2002-02-27 | 2003-09-04 | Ab Science | Use of tyrosine kinase inhibitors for treating substance use disorders |
| CN105974131A (zh) * | 2016-06-16 | 2016-09-28 | 武汉大学 | c-Kit作为药物成瘾治疗靶点的应用 |
| CN106074555A (zh) * | 2016-06-16 | 2016-11-09 | 武汉大学 | 伊马替尼及其衍生物在制备治疗药物成瘾的药物中的新用途 |
-
2019
- 2019-09-30 CN CN201910939677.4A patent/CN112575073B/zh active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2003072106A2 (en) * | 2002-02-27 | 2003-09-04 | Ab Science | Use of tyrosine kinase inhibitors for treating substance use disorders |
| CN105974131A (zh) * | 2016-06-16 | 2016-09-28 | 武汉大学 | c-Kit作为药物成瘾治疗靶点的应用 |
| CN106074555A (zh) * | 2016-06-16 | 2016-11-09 | 武汉大学 | 伊马替尼及其衍生物在制备治疗药物成瘾的药物中的新用途 |
Non-Patent Citations (5)
| Title |
|---|
| 中国癌症基金会等: "《中国肿瘤临床年鉴 2015》", 30 September 2016, 中国协和医科大学出版社 * |
| 朱世敏等: "SCF/c-kit受体作为物质成瘾预防和治疗潜在新靶点的研究", 《第九届药物毒理学年会——新时代·新技术·新策略·新健康论文集》 * |
| 李艳琴等: "伊马替尼及其衍生物对物质依赖戒断后心理渴求的防治作用及机制研究", 《第十四届全国药物依赖性学术会议暨国际精神疾病研讨会论文摘要汇编》 * |
| 田森等: "外泌体microRNAs在恶性血液系统疾病中的研究进展", 《解剖学杂志》 * |
| 陈萍萍: "SCF/c-kit信号通路在阿片类药物依赖心理渴求中的作用及机制", 《中国优秀硕士学位论文全文数据库 医药卫生科技辑》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2023102991A1 (zh) * | 2021-12-06 | 2023-06-15 | 中国科学院深圳先进技术研究院 | 一种成瘾药物戒断后复吸相关调控基因及其筛选方法 |
| CN116497031A (zh) * | 2023-06-02 | 2023-07-28 | 徐州医科大学 | 一种缓解小鼠吗啡成瘾行为的siRNA及其应用 |
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