CN117838827B - 一种治疗慢性心力衰竭的中药组合物及制备方法 - Google Patents
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Abstract
本发明涉及一种治疗慢性心力衰竭的中药组合物及制备方法,所述中药组合物由以下重量份组成的中药材制成:仙茅8‑12份、仙灵脾8‑12份、黑附片4‑8份、茯苓10‑15份、白术8‑12份、生姜8‑12份、白芍8‑12份、巴戟天8‑12份、甘松4‑8份、大腹皮12‑18份、猪苓8‑12份、泽泻8‑12份、桂枝8‑12份、炒麦芽10‑15份和焦山楂10‑15份。本发明的中药组合物具有温阳利水,补肾运脾强心功效,能够调节能量代谢、改善心室重构及电重构、治疗由阳虚水泛引起的慢性心力衰竭,有效率高,为慢性心力衰竭的治疗提供了可靠的治疗方案。
Description
技术领域
本发明涉及中药技术领域,具体涉及一种治疗慢性心力衰竭的中药组合物及制备方法。
背景技术
深入研究心力衰竭的发病机制及相关治疗,对于降低心力衰竭发病率具有重要意义。中医认识和治疗慢性心肌衰竭历史悠久,认为本病属于“心悸”、“喘证”、“心痹”、“心水”、“水肿”、“痰饮”等病范畴。心力衰竭后心阳渐微,日久损及肾阳、脾阳,致心、脾、肾阳皆虚,温煦、气化、运化、推动作用失司,痰瘀水饮交阻为患,进一步损耗心阳,反复恶性循环。因此,温阳利水,补肾运脾、强心是治疗慢性心力衰竭的根本。例如中国专利公告号CN102423347B的中药即主要以温阳利水的治疗原则为主,且其中的鹿角片价格昂贵,药材不易获取。然而,心力衰竭的基础是心肌能量代谢重构和心室重构,电重构与二者相互促进、相互恶化,加速心衰进程,故心肌能量代谢重构、心室重构和电重构治疗是心力衰竭治疗的关键。但目前临床运用的药物没有能够在治疗心衰的同时具备改善心肌能量代谢重构、心室重构和电重构的功效。
发明内容
(一)要解决的技术问题
鉴于现有技术的上述缺点、不足,本发明提供一种治疗慢性心力衰竭的中药组合物,具有温阳利水,补肾运脾强心功效,能够调节能量代谢、改善心室重构及电重构、治疗由阳虚水泛引起的慢性心力衰竭,有效率高,其为慢性心力衰竭的治疗提供了可靠的治疗方案。
(二)技术方案
第一方面,本发明提供一种治疗慢性心力衰竭的中药组合物,其由以下重量份组成的中药材制成:仙茅8-12份、仙灵脾8-12份、黑附片4-8份、茯苓10-15份、白术8-12份、生姜8-12份、白芍8-12份、巴戟天8-12份、甘松4-8份、大腹皮12-18份、猪苓8-12份、泽泻8-12份、桂枝8-12份、炒麦芽10-15份和焦山楂10-15份。
优选地,所述中药材的组成为:仙茅10份、仙灵脾10份、黑附片6份、茯苓15份、白术10份、生姜9份、白芍10份、巴戟天10份、甘松6份、大腹皮15份、猪苓10份、泽泻10份、桂枝10份、炒麦芽15份、焦山楂10份;或者,
所述中药材的组成为:仙茅8份、仙灵脾8份、黑附片4份、茯苓10份、白术8份、生姜8份、白芍8份、巴戟天8份、甘松4份、大腹皮12份、猪苓8份、泽泻8份、桂枝8份、炒麦芽10份、焦山楂15份;或者,
所述中药材的组成为:仙茅12份、仙灵脾12份、黑附片8份、茯苓15份、白术12份、生姜12份、白芍12份、巴戟天12份、甘松6份、大腹皮12份、猪苓12份、泽泻12份、桂枝12份、炒麦芽15份、焦山楂12份;
所述中药材的组成为:仙茅9份、仙灵脾9份、黑附片5份、茯苓14份、白术9份、生姜9份、白芍9份、巴戟天9份、甘松8份、大腹皮15份、猪苓9份、泽泻9份、桂枝9份、炒麦芽15份、焦山楂15份。
根据本发明的较佳实施例,所述中药组合物为胶囊剂、中药散剂、片剂、口服液、中药药丸或水煎剂。
第二方面,本发明提供一种治疗慢性心力衰竭的中药组合物的制备方法,所述中药组合物为胶囊剂、中药散剂、中药药丸或水煎剂;其制备方法如下:
所述胶囊剂的制备方法为:按照上述重量份称量各中药材,经清洁和切碎或切段后,先将黑附片放入砂锅中并加水,煎煮30-50min,再将其他中药材放入砂锅进行煎煮、过滤,收获滤液,将滤液继续熬煮浓缩至膏状,干燥、灭菌、研碎,过100-200目筛后,灌装胶囊。
所述中药散剂的制备方法为:按照上述重量份称量各中药材,经清洁和切碎或切段后,先将黑附片放入砂锅中并加水,煎煮30-50min,再将仙茅、仙灵脾、茯苓、白术、甘松、生姜、白芍、巴戟天、大腹皮、猪苓、泽泻、桂枝放入砂锅进行煎煮、过滤,收获滤液,将滤液继续熬煮浓缩至膏状;然后将炒麦芽和焦山楂粉碎后过筛,得到粉末;将膏状物与粉末混合后,进行制粒、烘干、灭菌,即得到中药散剂。
所述中药药丸的制备方法为:按照上述重量份称量各中药材,经清洁和切碎或切段后,先将黑附片放入砂锅中并加水,煎煮30-50min,再将其他中药材放入砂锅进行煎煮、过滤,收获滤液,将滤液继续熬煮浓缩至相对密度为1.1-1.2的稀浸膏,混入蜜丸,搅拌均匀后制成中药药丸;中药材总质量与蜜丸质量比为1:1-2。
所述水煎剂的制备方法为:按照上述重量份称量各中药材,经清洁和切碎或切段后,先将黑附片放入砂锅中并加水,煎煮30-50min,再将其他中药材放入砂锅中,第一次煎煮15-20min,倒出1/2-2/3药液;再加温水,进行第二次煎煮,倒出1/2-2/3药液,将两次药液混合,即为所述水煎剂。
根据本发明的较佳实施例,在以上各实施例的煎煮过程中,加水量为没过全部中药材3-5cm。对吸水量大的药材,可先吸饱水后,再放入砂锅中煎煮。
本发明中各中药材的预处理方法可参见《中国药典2010年版》,经预处理后的药材用于本发明的中药组合物的制备。
在本发明的中药组合物中,黑附片又叫做黑顺片,是中药附子采摘以后经过炮制,然后切片所制成的一种药材,颜色变成黑色。黑附片的功效主要是温补肾阳,温阳利水,回阳救逆,用于肾阳不足所导致的一系列问题。炒麦芽是通过特殊手法炮制过的麦芽,而生麦芽没有经过炮制。麦芽经过炒制后,消食导滞的效力更强。焦山楂是通过一定加工方式做成的山楂的一种品种(中火加热至外表焦褐色),功效与山楂不同,山楂具有扩血管、强心、降血压和降脂、抗衰老防癌的功效,焦山楂主治秋季腹泻,除消食导滞外,长于消食止泻。焦山楂较生山楂性味温和,产生的胃酸较少,副反应弱,适于体弱胃酸相对较多者。
(三)有益效果
与现有技术相比,本发明的主要技术效果在于:本申请的发明人考虑到心衰久病,多脾肾两虚,先天后天俱不足,因此本发明的中药组合物以二仙即“仙茅与仙灵脾”为君药,配以焦二仙即“焦山楂与炒麦芽”为臣药,附子,生姜,茯苓,白术,泽泻,猪苓为佐助药,白芍为佐制药,桂枝为使药。其中,君药仙茅与仙灵脾温肾助阳而散寒,配合以巴戟天补益肾气共同达到扶助先天的功效;臣药焦山楂与炒麦芽消食理气健脾,调养后天;附子温振肾阳以助心阳,加以茯苓淡渗、白术苦燥以健脾化水,泽泻、猪苓利湿泻浊,配以生姜温中化水,培土制水,以治中州,并加白芍酸敛反佐,防止利水伤阴,共助君臣之功,以制水气凌心之逆;更以桂枝为使,化气活血,引药入心,气血通调。前述组方实现了脾肾同治,先天及后天同步调养的功效。
此外,由于心衰久病,多夹杂气郁;因此在前述组方基础上,发明人又添加甘松醒脾解郁、大腹皮理气利水,以达气血水同调的目的,共奏温阳利水,补肾运脾强心的功效。
附图说明
图1为本发明中药组合物(水煎剂)对各组心衰大鼠心肌炎症及细胞肥大及纤维化程度影响的HE染色图(注:放大倍数为200×)。
图2为显示中药组合物(水煎剂)对各组心衰大鼠心肌细胞凋亡的改善作用的TUNEL染色图(注:放大倍数为400×)。
图3为显示中药组合物(水煎剂)对各组心衰大鼠心肌凋亡代表性蛋白BaX,Bcl-2,Caspase3、Caspase8、Caspase9的抑制作用的免疫组化图。
图4为本发明中药组合物(水煎剂)对各组心衰大鼠电信号传导的关键通路p38-MAPK信号通路及关键蛋白Cx43表达的影响。
图5为本发明中药组合物(水煎剂)对各组心衰大鼠线粒体自噬活性的影响。
图6为本发明中药组合物(水煎剂)对各组心衰大鼠心肌组织PINK1/parkin通路相关蛋白表达的影响。
具体实施方式
为了更好的解释本发明,以便于理解,下面结合附图,通过具体实施方式,对本发明作详细描述。
本发明所提供的一种治疗慢性心力衰竭的中药组合物,其由以下重量份组成的中药材制成:仙茅8-12份、仙灵脾8-12份、黑附片4-8份、茯苓10-15份、白术8-12份、生姜8-12份、白芍8-12份、巴戟天8-12份、甘松4-8份、大腹皮12-18份、猪苓8-12份、泽泻8-12份、桂枝8-12份、炒麦芽10-15份和焦山楂10-15份。
其中仙茅、仙灵脾共同组成君药。仙茅、仙灵脾可组成二仙,二仙温肾助阳而散寒,配合巴戟天为佐药补益肾气,达到共扶先天之效。本发明的中药组合物以焦山楂与炒麦芽为臣药,焦山楂与炒麦芽组成焦二仙,消食理气健脾,调养后天;由此实现脾肾同治和先天后天并调。以附子,生姜,茯苓,白术,泽泻,猪苓为佐助药,桂枝为佐使药。附子温振肾阳以助心阳,加以茯苓淡渗、白术苦燥以健脾化水,泽泻、猪苓利湿泻浊,配以生姜温中化水,培土制水,以治中州。白芍为佐制药,白芍酸敛反佐,防止利水伤阴,共助君臣之功,以制水气凌心之逆。更以桂枝为使药,化气活血,引药入心,气血通调。
本发明的药方中,白术、生姜、黑附片、白芍和茯苓组成温阳利水的经典药方“真武汤”的近似药方。茯苓、泽泻、猪苓、桂枝、白术组成淡渗利湿的经典药方“五苓散”的近似药方,这两种近似药方共同达到壮肾中阳,化气利水、健脾祛湿的功效。最后辅以甘松醒脾解郁、大腹皮理气利水,以达气血水同调,共奏温阳利水,补肾运脾强心的功效。
以上各组方在本发明中药组合物中所起作用如下:
仙灵脾,即淫羊藿,性辛,味温。入肝、肾经。补肾助阳,袪风湿利水。同时仙灵脾可抑制离体右心房肌的肌力和心率,同时有抗血小板聚集、降低红细胞聚集性及降低全血粘度作用。配仙茅,称为二仙,能补命门而兴阳道。而黑附片辛热,入心、脾、肾经,可温通心脉,补助肾阳,更助先天之用,如配与茯苓、白术,更增温阳利水之功,获得具与《伤寒论》中“真武汤”类似的温阳利水功效,产生强心以及扩张外周血管的作用,增强心肌收缩。再配与巴戟天,更补益肾气,同时有促肾上腺皮质激素作用,可以刺激造血,保护心肌。而心衰久病,穷必及肾,上药共举扶助先天之功,少阴心肾相系,共同温养心肾,以全其功。
山楂味酸而甘,消食力佳,为消化食积停滞常用要药,麦芽可促进食物的消化,尤能消米面食积。在临床上用于食积不化、脘闷腹胀,可与山楂配伍,以调养后天,而炒麦芽同时可以促进胃蛋白酶的分泌,焦山楂同时具有强心、降脂的作用,并且可以扩张外周血管,以降压。
桂枝味辛、甘,性温。入心、肺、膀胱经。可温通经脉,通阳化气。如《本经疏证》述:“和营、通阳、利水、下气、行气、补中,为桂枝六大功效。”常合茯苓、白术等组成淡渗利湿经典药方五苓散的近似药方,擅长治疗对阴寒遏阻阳气,津液不能输布导致水湿痰饮停聚形成相关病症,如心衰心阳不足水饮偏盛的心悸,同时可以扩张血管,增加冠脉循环,同时扩张外周血管,增强血液循环,还有保护心缺血再灌注损伤,同时又有利尿的作用。
茯苓味甘、淡,性平。归心、肺、脾、肾经。功能利水渗湿,健脾宁心,《本草纲目》曰:“茯苓气味淡而渗,其性上行,生津液,开腠理,滋水源而下降,利小便,故张洁古谓其属阳,浮而升,言其性也;东垣谓其为阳中之阴,降而下,言其功也。”同时具有增强造血、利尿的功能。常配与白术、猪苓、泽泻以增强利水之功,以利水宁心。
生姜味辛,性微温。入肺、脾、胃经。有温中合和胃,散寒痰饮之用,如治疗心衰导致的胃肠道不适,其中生姜醇可以增肌心脏收缩力和延长心脏收缩的持续时间。
白芍味苦、酸,性微寒。养血敛阴,防止过于利水而伤阴,同时又可以活血利水,如《本经》曰:“主邪气腹痛,除血痹,破坚积,寒热疝瘕,止痛,利小便,益气。”同时能显著增加小鼠心肌营养血流量,有升高血压和增强心力作用。
甘松温而不热,甘而不滞,其气芳香,能开脾郁;其性温通,能行气止痛,善治疗心腹疼痛,同时能保护心肌细胞及降血压作用,配与大腹皮更能行气利水疏滞、宽中除胀而增强止痛效果。
本发明的中药组合物,即本申请水煎剂,从性味分析,药性偏温,诸药相配,气、水、血同调,全方君臣佐使,配伍得当,厘然有度,补泻兼施,攻实不伤正,补虚而不助邪,共奏温补脾肾,补阳利水,兼顾护阴津之功,与心衰形成的机理十分契合。实验证明,所述中药组合物,在慢性心衰疾病的治疗之中,能够抑制慢性心力衰竭心肌细胞纤维化,减轻心肌细胞凋亡,增强心肌收缩力,改善心室重构和电生理重构,调节能量代谢,进而延缓心力衰竭进程。
实施例1.1
本实施例为一种治疗慢性心力衰竭的中药水煎剂,其制备方法如下:
(1)称取仙茅10g、仙灵脾10g、黑附片6g、茯苓15 g、白术10g、生姜9g、白芍10g、巴戟天10g、甘松6g、大腹皮15g、猪苓10g、泽泻10g、桂枝10g、炒麦芽15g、焦山楂10g。
(2)先将黑附片放入砂锅中并加水,煎煮40min,再将其他中药材放入砂锅中,第一次煎煮20min,倒出1/2药液;再加倒出药液等体积的温水,进行第二次煎煮,倒出2/3药液,将两次药液混合,即为水煎剂,将水煎剂用两层纱布过滤,去除小药渣,得到清澈透亮的水煎药液。
实施例1.2
本实施例为一种治疗慢性心力衰竭的中药水煎剂,其制备方法如下:
称取仙茅12g、仙灵脾9g、黑附片7g、茯苓14 g、白术11g、生姜10g、白芍10g、巴戟天10g、甘松7g、大腹皮14g、猪苓10g、泽泻10g、桂枝10g、炒麦芽12g、焦山楂12g。按照实施例1.1的方法制备,得到清澈透亮的水煎药液。
实施例1.3
本实施例为一种治疗慢性心力衰竭的中药水煎剂,其制备方法如下:
称取仙茅10g、仙灵脾10g、黑附片8g、茯苓13g、白术12g、生姜12g、白芍12g、巴戟天12g、甘松8g、大腹皮12g、猪苓12g、泽泻11g、桂枝11g、炒麦芽13g、焦山楂13g。按照实施例1.1的方法制备,得到清澈透亮的水煎药液。
对比例1
本对比例是在实施例1.1基础上去除“仙茅”药材,其余组份和制备方法与实施例1.1相同,制得一种水煎药液。
对比例2
本对比例是在实施例1.1基础上去除“炒麦芽和焦山楂”,其余组份和制备方法与实施例1.1相同,制得一种水煎药液。
使用本实施例1.1、实施例1.2以及实施例1.3的水煎剂对模型心衰大鼠进行处理,研究不同中药组方制备的水煎剂对心衰大鼠心功能的影响,实验方法如下:
(1)实验材料:实验动物:6-7周龄,SPF级SD大鼠,雄性,共48只,体重240-260g,购于湖南斯莱克景达实验动物有限公司(使用许可证SYXK(赣)2017-0004,许可证SCXK(湘)2019-0004)。将它们安置在恒定的室温(25℃)下,湿度(55±3)℃。12h昼夜循环,动物自由进食饮水。实验动物使用经江西中医药大学实验动物伦理委员会批准(JZLLSC20210059)并按实验动物使用的3R原则给予人道的关怀。实验药物:抗心衰中药颗粒剂水溶液(采用实施例1.1制备的水煎剂);阳性对照药:卡托普利。
(2)造模方法:将大鼠固定在自制鼠板上,进行腹腔内注射阿霉素,每次2.5 mg/(kg·只),2周内注射6次,累积剂量15mg/(kg·只),造模周期为4周。以大鼠一般状态、HWI等心功能指标和血清及心肌组织中的相关因子评价药物对慢性心衰的治疗作用。
(3)实验分组:造模后喂养4周,采用随机数字表法将存活的大鼠分为6组,每组7只,分组如下:
。
(4)心脏彩超测定:
本实验小动物超声影像系统Vevo2100型由江西中医药大学动物科技中心实验室提供。用异氟烷吸入麻醉,应用MS250超声探头进行心脏超声影像分析,在M模式下取左室长轴切面进行测量,并计算大鼠左心室射血分数(LVEF);短轴缩短率(LVFS)。
(5)样本制备及计算左心室质量指数(LVMI):
取材前称体质量,大鼠麻醉后置于鼠板固定,用10ml注射器抽取腹主动脉血,静置30min后离心机离心(10000g,10min),离心后用移液器吸取上层血清,将其放置于-80℃冰箱保存以备接下来的ELISA实验检测。开胸取大鼠心脏,注意取材过程中心脏的完整性,取出的心脏用在4℃预冷的生理盐水洗净后用滤纸吸干水分,用手术剪沿房室沟剪掉左、右心房,剪掉主、肺动脉,紧贴室间隔剪下右心室游离壁,剩余部分即为左心室,称取左心室质量,计算左心室质量指数(LVMI=大鼠左心室质量/大鼠体质量)。经横切面,在左室心肌结扎部位以下处用手术刀切取几块组织,注意避免过度牵拉等机械损伤,用4%多聚甲醛固定切取的心肌组织。剩余的左室心肌梗死边缘组织放入冻存管内,置于液氮罐中冷冻保存以备Western blot蛋白提取检测。
(6)心脏组织HE染色:
大鼠左室心肌组织固定后常规脱水,石蜡包埋,将包埋好的组织样本置于-20℃冷冻数分钟后切4-5um超薄片;切片后常规脱蜡→自来水洗2min→苏木素液染色5min→自来水洗1min→1%盐酸酒精分化3s→自来水洗1min→0.2%氨水返蓝30s→自来水洗1min→0.5%伊红液染色3min→自来水洗(速洗)→乙醇梯度脱水→二甲苯I、Ⅱ分别脱水3min;封片后在显微镜下观察,使用Lecia Applaction Stiue图像系统采集分析样本相关部位。
(7)TUNEL法检测:
将蜡块切成厚度为5μm的切片后脱蜡处理→100%乙醇浸泡5min后再梯度乙醇水化→0.85%NaCl溶液中浸洗5min→PBS洗5min后固定15min→PBS浸洗2次,每次5min→加100μL浓度为20μg/ml的蛋白酶K溶液,室温孵育10min→PBS洗5min后固定5min→PBS浸洗5min→加入100μL的平衡缓冲液→室温平衡5min。
平衡时进行标记,将核苷酸混合物在冰上解冻,配制足量的rTdT酶孵育缓冲液,每个切片需要50μL。将此孵育缓冲液加到切片组织处,覆盖一个塑料薄膜。切片放在湿盒中于37℃暗处孵育1h(50μL孵育缓冲液配方:45μL平衡缓冲液+5μL核苷酸混合物+1μL rTdT酶,配好后避光4℃放置); 将20×氯化钠-柠檬酸钠缓冲液(20×SSC)溶液1:10稀释至2×SSC。标记好的切片从37℃中取出,移除塑料薄膜,在2×SSC溶液中室温浸泡15 min。PBS洗3次,每次5min。
封片后利用荧光显微镜观察荧光。紫外光激发DAPI染的细胞核为蓝色,凋亡的细胞被蓝光激发呈现为绿色。切片4℃避光保存。应用Image Pro Plus6.0图像分析光密度值分析计算凋亡率。
(8)免疫组织化学染色法检测心肌凋亡代表性蛋白Bax,Bcl-2,Caspase3表达:
将蜡块切成厚度为5μm的切片,脱蜡→100%乙醇浸泡5min→梯度乙醇水化→抗原修复→3% H2O2室温孵育10min以消除内源性过氧化氢酶活性→PBS洗4次,每次3min→3%BSA溶液室温封闭15min(暗盒操作)→倾去封闭液,勿洗→滴加适当比例稀释的一抗(使用封闭液稀释)4℃ 杂交过夜(不超过12h,暗盒操作)→PBS洗3min×3次→加生物素标记的二抗37℃孵育15 min(暗盒操作)→PBS 洗3min×3次→加辣根酶标记链霉卵白素工作液37℃孵育15 min(暗盒操作)→PBS洗3min×3次→DAB 显色,显微镜下观察显色情况,反应部位产生棕色沉淀物→显色充分后去离子水充分冲洗。一般显色时间2-10min→室温下苏木素溶液复染10min,ddH2O洗,0.25%氨水返蓝,去离子水冲洗→分别在70%、95%、100%乙醇中浸泡3min,二甲苯Ⅰ与二甲苯Ⅱ中各浸泡3min→中性树胶封片。显微镜400倍视野下随机采取5个视野,应用Image Pro Plus6.0图像分析软件进行结果分析。
(9)统计分析:计量数据进行统计描述,多组数据分析采用单因素方差分析检验,方差齐时采用最小显著差(LSD)法;方差不齐时选用Tamhaneˊs T2法,以P<0.05为差异具有统计学意义,P<0.01为差异具有极显著统计学意义,P>0.05表明两组之间对比无统计学上的差异。
(10)实验结果:实验结果如表1和图1-图6所示。
与空白对照组(JSS)相比,模型组(MX) EF 显著降低(p<0.01),具有统计学意义,且 EF 平均值<45%;与模型组相比,卡托普利组(LST) EF 上升不明显(p>0.05),对比例1组(DB1)和对比例2组(DB2)组的EF值有所上升(p<0.05),具有统计学意义;实施例1.2组(ZWTD)、实施例1.3组(ZWTZ)和实施例1.1组(ZWTG)的EF则显著上升(p<0.01),具有统计学意义,EF 平均值≥53.8%。实施例1.3组、实施例1.1组对 EF 值得改善明显优于卡托普利组(p<0.01);因此与对比例1组和对比例2组相比,实施例1.1-1.3对 EF 值的改善更显著(p<0.05),实施例1.1-1.3组的组间比较差异无统计学意义(p>0.05)。其中,实施例1.1处理组效果最显著。
与空白对照组相比,模型组(MX) FS 显著降低(P<0.01),具有统计学意义;与模型组相比,卡托普利组(LST) FS 只有小幅上升,差异无统计学意义(P>0.05);实施例1.1-1.3组 FS 上升较明显(P<0.01),具有统计学意义;对比例1组、对比例2组 FS 也有一定上升(P<0.05),但FS上升幅度高于LST组但低于实施例1.1-1.3组。其中,实施例1.3处理组效果最显著。
模型组与空白对照组相比 LVIDs 显著升高(P<0.01),具有统计学意义;与模型组(MX)相比,卡托普利组及对比例1组、对比例2组LVIDs有所降低(P<0.05),具有统计学意义,实施例1.1-1.3组的LVIDs显著降低(P<0.01),具有统计学意义。与对比例1组、对比例2组相比,实施例1.1-1.3组对 LVIDs 值的改善更显著(p<0.05)。其中,实施例1.2处理组效果最显著。
模型组与空白对照组相比 LVIDd 显著升高(P<0.01),具有统计学意义;与模型组比,实施例1.1-1.3组 LVIDd 显著降低(P<0.01)具有统计学意义,且对模型组LVIDd降低幅度大于卡托普利组(LST组)和对比例1、及对比例2组。其中,实施例1.3组降低效果最显著。
综上所述,本申请水煎剂能够改善慢性心力衰竭大鼠的一般状况,改善心功能及心肌结构:抑制 EF 的降低及 LVIDs 的增加,抑制心肌纤维化,改善心肌细胞肿胀,改善心肌细胞内线粒体的超微结构;而缺少二仙或二焦的配方难以达到本发明实施例1.1-1.3的功效。
表1:本发明的中药水煎剂对各组心衰大鼠心功能的影响
。
注:JSS:空白对照组;MX:模型组;LST:阳性对照组;ZWTD:实施例1.2组;ZWTZ:实施例1.3组;ZWTG:抗心衰中药颗粒剂水溶液组(实施例1.1组);DB1:对比例1组;DB2:对比例2组;LVIDs:左心室收缩期内径;LVIDd:左心室舒张末期内径;EF:射血分数;FS:左室短轴缩短率。
如图1所示,HE染色显示:空白对照组心肌细胞排列整齐,细胞无明显肿胀,无明显的心肌纤维化改变,组织内未见明显的血管充血,无明显的嗜酸性细胞;模型组心肌细胞排列紊乱,部分细胞出现波浪样改变,嗜酸性细胞增多,小片状心肌间质纤维化改变;卡托普利组及实施例1.2组、实施例1.3组、实施例1.1组细胞排列较模型组整齐,可见嗜酸性细胞增多、心肌波浪样变,但程度较模型组有所减轻,可见散在的心肌间质纤维化改变,纤维化面积较模型组明显减少,从而抑制心肌纤维化。其中实施例1.2组和实施例1.1相较于模型组的改善作用最明显。
如图2所示,正常心肌细胞细胞核发出蓝色荧光,而凋亡心肌细胞的细胞核发出绿色荧光。与空白对照组比较,模型组大鼠的心肌细胞凋亡率显著升高(P<0.01),说明大鼠心衰发生发展过程中存在着过度的心肌细胞凋亡现象;与模型组比较,实施例1.2组、实施例1.3组、实施例1.1组及阳性药对照组(卡托普利组)凋亡率下降(P<0.01),提示经本申请水煎剂或卡托普利干预后凋亡现象改善。其中,实施例1.1的凋亡现象少于实施例1.2组和实施例1.3组,与阳性对照组更接近。
如图3所示,与空白对照组比较,模型组Bcl-2表达显著下降Bax、Caspase3、Caspase8、Caspase9表达显著升高。与模型组相比,实施例1.2组Bcl-2表达增强,实施例1.3组、实施例1.1组及阳性对照组Bcl-2表达增强显著。实施例1.2组、实施例1.3组、实施例1.1组及阳性药对照组Bax、Caspase3、Caspase8、Caspase9蛋白表达降低,其中实施例1.3组Bcl-2、Bax、Caspase3、Caspase8、Caspase9蛋白表达与空白对照组无明显差异,提示本申请水煎剂可有效抑制心衰大鼠过度凋亡,实施例1.3组抗凋亡效果最佳。由此可见,本申请水煎剂能通过抑制心肌细胞凋亡减轻心衰过程中的心室肥厚,增强心肌收缩力,进而延缓心衰发生发展。
如图4的A和B显示:与模型组相比,实施例1.2组、实施例1.3组、实施例1.1组心肌细胞Cx43、p38-MAPK、MMP2、MMP9、p53、p-Cx43蛋白表达水平程下降趋势,而TIMP1表达水平程上升趋势,N:空白对照组;M:模型组;S:阳性对照组;D:实施例1.2组;Z:实施例1.3组;G:实施例1.1组。图4的C显示:实施例1.2组、实施例1.3组、实施例1.1组心肌细胞Cx43、p38-MAPK、MMP2、MMP9的mRNA转录水平程下降趋势,而TIMP1-mRNA转录水平程上升趋势(P<0.01);图4的D显示:实施例1.2组、实施例1.3组、实施例1.1组细胞内Cx43荧光信号整体相对减弱,Cx43紊乱情况有所改善;图4的E显示:β-半乳糖苷酶对各组细胞进行染色,衰老的细胞被染成深蓝色,而实施例1.2组、实施例1.3组、实施例1.1组对比模型组蓝色较浅,卡托普利组相比于模型组亦有较明显改善。心衰过程中,心室重构是引起心衰的最主要的病理基础,而电重构与心室的结构重构有密切关系。本申请水煎剂具有抗心衰作用,其作用涉及MAPK/Cx43通路的蛋白途径,即心室结构重构与电重构相互作用。
如图5所示为各组大鼠电镜超微结构图。箭头1所示为自噬小体;箭头3所示为线粒体髓样变性;线粒体髓样变性是线粒体自噬的前期过程;箭头2所示为溶酶体嵴被破坏;箭头5所示为线粒体与溶酶体结合过程;箭头4表示线粒体嵴破坏。其中,空白对照组(JSS)心肌组织整体结构基本正常,心肌肌原纤维排列规则,线粒体结构具有完整性,线粒体双层外膜结构清晰,线粒体内膜向内折叠形成线粒体嵴,线粒体嵴连续、排列整齐,线粒体基质饱满,可见自噬小体,未见溶酶体;模型组(MX)心肌组织整体结构异常,心肌肌原纤维排列不整齐,线粒体数量增加,线粒体变大变圆,基质变浅,双层外膜结构清晰,线粒体嵴疏松断裂,变短变少,可见自噬小体、线粒体髓样变性及溶酶体;卡托普利组(LST)心肌组织整体结构异常,肌原纤维排列不规则,线粒体结构异常,线粒体数量与空白对照组相比明显增加,与模型组相比减少,线粒体变大变圆,基质变浅,线粒体嵴疏松断裂、变短变少,可见线粒体髓样变性;实施例1.2组(ZWTD)心肌组织结构轻度异常,心肌肌原纤维排列较整齐,线粒体呈椭圆形,双层外膜结构清晰,未见明显的线粒体嵴改变,可见自噬小体;实施例1.3组(ZWTZ)轻度异常,线粒体结构未见明显异常,基质饱满,未见明显嵴断裂及变短变少,可见自噬小体及线粒体与溶酶体融合过程;实施例1.1组(ZWTG)心肌组织结构轻度异常,线粒体双层外膜结构清晰,线粒体嵴断裂,可见自噬小体及线粒体髓样变性。
参见图6所示,为大鼠 PINK1/parkin 通路蛋白表达图。与空白对照组相比,模型组(MX) LC3Ⅱ、LC3Ⅰ、beclin1、p-PINK1、p-parkin 蛋白表达均显著降低(p<0.01),具有统计学意义,P62蛋白表达显著上升(p<0.01),具有统计学意义,PINK1蛋白、parkin蛋白变化不明显。与模型组相比,卡托普利组(LST)及实施例1.2组(ZWTD)、实施例1.3组(ZWTZ)、实施例1.1组(ZWTG)中的LC3Ⅱ、beclin1、p-PINK1、 p-parkin的蛋白含量显著增加(P<0.01),具有统计学意义,且实施例1.2组、实施例1.3组、实施例1.1组的 P62蛋白显著降低(P<0.01),具有统计学意义,PINK1蛋白、parkin蛋白变化不明显。
以上本实验结果提示,本申请水煎剂可能是通过激活 PINK1/parkin 信号通路介导的线粒体自噬,从而提高受损线粒体内的物质利用,改善心肌的能量供应,改善慢性心力衰竭大鼠的心功能及心肌结构,抑制慢性心力衰竭病程的进展,从而治疗慢性心力衰竭。
实施例2
本实施例为一种治疗慢性心力衰竭的中药散剂,其制备方法如下:
(1)称取仙茅8g、仙灵脾8g、黑附片4g、茯苓10g、白术8g、生姜8片、白芍8g、巴戟天8g、甘松4g、大腹皮12g、猪苓8g、泽泻8g、桂枝8g、炒麦芽10g、焦山楂15g。
(2)先将黑附片放入砂锅中并加水,煎煮40min,再将仙茅、仙灵脾、茯苓、白术、甘松、生姜、白芍、巴戟天、大腹皮、猪苓、泽泻、桂枝放入砂锅进行煎煮、过滤,收获滤液,将滤液继续熬煮浓缩至膏状;然后将炒麦芽和焦山楂粉碎后过筛,得到粉末;将膏状物与粉末混合后,与1倍量的淀粉混合,进行制粒、烘干、灭菌,即得到中药散剂。
实施例3
本实施例为一种治疗慢性心力衰竭的中药药丸,其制备方法如下:
(1)称取仙茅12g、仙灵脾12g、黑附片8g、茯苓15g、白术12g、生姜12片、白芍12g、巴戟天12g、甘松6g、大腹皮12g、猪苓12g、泽泻12g、桂枝12g、炒麦芽15g、焦山楂12g。
(2)准备与上述中药材总质量1:1的蜜丸。先将黑附片放入砂锅中并加水,煎煮40min,再将其他中药材放入砂锅进行煎煮、过滤,收获滤液,将滤液继续熬煮浓缩至相对密度为1.15的稀浸膏,混入蜜丸,搅拌均匀后制成中药药丸。
实施例4
本实施例为一种治疗慢性心力衰竭的中药药丸,其制备方法如下:
(1)称取仙茅9g、仙灵脾9g、黑附片5g、茯苓14g、白术9g、生姜9片、白芍9g、巴戟天9g、甘松8g、大腹皮15g、猪苓9g、泽泻9g、桂枝9g、炒麦芽15g、焦山楂15g。
(2)准备与上述中药材总质量1:2的蜜丸。先将黑附片放入砂锅中并加水,煎煮40min,再将其他中药材放入砂锅进行煎煮、过滤,收获滤液,将滤液继续熬煮浓缩至相对密度为1.2的稀浸膏,混入蜜丸,搅拌均匀后制成中药药丸。
实施例5
本实施例为一种治疗慢性心力衰竭的胶囊,其制备方法如下:
(1)称取仙茅8g、仙灵脾8g、黑附片4g、茯苓10g、白术8g、生姜9片、白芍8g、巴戟天8g、甘松6g、大腹皮10g、猪苓12g、泽泻12g、桂枝12g、炒麦芽9g、焦山楂15g。
(2)先将黑附片放入砂锅中并加水,煎煮35min,再将其他中药材放入砂锅进行煎煮、过滤,收获滤液,将滤液继续熬煮浓缩至膏状,干燥、灭菌、研碎,过200目筛后,灌装胶囊。
实施例6
本实施例为一种治疗慢性心力衰竭的中药水煎剂,其制备方法如下:
(1)称取仙茅10g、仙灵脾8g、黑附片6g、茯苓12g、白术8g、生姜9片、白芍8g、巴戟天12g、甘松6g、大腹皮9g、猪苓12g、泽泻9g、桂枝9g、炒麦芽9g、焦山楂12g。
(2)先将黑附片放入砂锅中并加水,煎煮40min,再将其他中药材放入砂锅中,第一次煎煮20min,倒出1/2药液;再加倒出药液等体积的温水,进行第二次煎煮,倒出2/3药液,将两次药液混合,即为水煎剂,将水煎剂用两层纱布过滤,去除小药渣,得到清澈透亮的水煎药液。
临床疗效的典型案例
案例1:患者邱某,男性,91岁,因“胸闷气喘10年余,伴双下肢水肿1周”就诊,症见:患者胸闷气喘,动则加重,双下肢重度浮肿,按之凹陷,手足不温,纳差,小便频数、量少,大便稍溏。舌暗红,苔白腻,脉沉细。为阳虚水泛所引起的心力衰竭,较为严重,治以温阳利水,补肾运脾强心,采用实施例1.1的中药水煎剂,每日2次,每次300mL,治疗半年后随访,患者胸闷气喘好转,双下肢无水肿,病情稳定。由于患者病情较重,还需随访。
案例2:患者郭某,女性,74岁,因“胸闷气喘3年,加重1月”就诊,症见:患者胸闷气喘明显,活动后加重,全身乏力,双下肢凹陷性水肿,手足不温,纳差,尿少,大便稍稀。舌淡暗,苔薄腻,脉弱。为阳虚水泛所引起的心力衰竭,治以温阳利水,补肾运脾强心,采用实施例2中药散剂,每日2次,每次30g温水冲服,治疗5月后随访,患者胸闷气喘好转,双下肢无水肿,病情稳定。
案例3:患者徐某,男性,66岁,因“反复胸闷气喘7年余,伴下肢水肿1月”就诊,症见:患者胸闷气喘,活动后明显,休息稍缓解,下肢轻度浮肿,按之凹陷,四肢不温,纳差,小便清长,大便稀,不成形。舌淡红,边有齿痕,苔润,脉沉细。为阳虚水泛所引起的心力衰竭,治以温阳利水,补肾运脾强心,采用实施例3的中药药丸,每日2次,每次1颗(15g/丸),治疗4月后随访,患者胸闷气喘好转,双下肢无水肿,病情稳定。
案例4:患者陈某,女性,87岁,因“反复胸闷气短21年余,再发伴气喘7天”就诊,症见:患者胸闷气喘明显,休息稍缓解,全身疲劳乏力,畏寒怕冷,下肢轻度水肿,纳差,小便频,大便稀。舌淡,苔白润,脉沉细。为阳虚水泛所引起的心力衰竭,治以温阳利水,补肾运脾强心,采用实施例4所述中药药丸,每日2次,每次1颗(15g/丸),治疗2月后随访,患者胸闷气喘好转,双下肢无水肿,病情稳定。
案例5:患者李某,男性,82岁,因“胸闷伴双下肢水肿3天”就诊,症见:患者胸闷、气促发作,休息可缓解,伴乏力双下肢明显水肿,按之凹陷不起,纳欠佳,小便清长,大便平。舌淡,苔白,脉滑。为阳虚水泛所引起的心力衰竭,治以温阳利水,补肾运脾强心,采用实施例5中药胶囊,每日3次,每次3颗,治疗3月后随访,患者胸闷气喘好转,双下肢无水肿,病情稳定。
案例6:患者殷某,女性,71岁,因“反复胸闷1年余,再发伴气喘3天”就诊,症见:患者胸闷气喘,夜间端坐呼吸,活动后明显,略乏力,双下肢中度凹陷性水肿,四肢冷,进食后胃脘不适,尿频量少,大便质稀。舌淡暗,脉结代。为阳虚水泛所引起的心力衰竭,治以温阳利水,补肾运脾强心,采用实施例6的中药水煎剂,每日2次,每次300mL,治疗1月后随访,患者胸闷气喘好转,双下肢无水肿,病情稳定。
案例7:患者余某,男性,88岁,因“反复胸闷气喘伴下肢水肿1年余,加重3天”就诊,症见:患者胸闷气喘,动则加剧,休息后缓解,乏力,双下肢轻度水肿,按之凹陷,手足不温,纳差,夜尿频,大便溏。舌暗,苔白腻,脉沉缓。为阳虚水泛所引起的心力衰竭,治以温阳利水,补肾运脾强心,采用实施例6中药水煎剂,每日2次,每次300mL,治疗一周后患者胸闷气喘好转,双下肢无水肿,病情稳定,无复发。
案例8:患者朱某,男性,75岁,因“反复胸闷气喘10年余,加重伴下肢水肿半月”就诊,症见:患者胸闷气喘,活动加剧,自觉活动耐量下降,双下肢凹陷性水肿明显,纳差,小便短少,大便溏。舌淡红,苔白厚腻,脉滑。为阳虚水泛所引起的心力衰竭,治以温阳利水,补肾运脾强心,采用实施例1.2中药水煎剂,每日2次,每次300mL,治疗1年后随访,患者胸闷气喘好转,双下肢无水肿,病情稳定。
案例9:患者龚某,女性,87岁,因“反复胸闷气喘4月余,加重3天”就诊,症见:患者胸闷气喘,夜间阵发性呼吸困难,活动耐量下降,双下肢稍凹陷性水肿,纳差,小便频,大便质稀,不成形。舌淡红,齿痕明显,苔白腻,脉弦滑。为阳虚水泛所引起的心力衰竭,治以温阳利水,补肾运脾强心,采用实施例3的中药药丸,每日2次,每次1颗(15g/丸),治疗3月后随访,患者胸闷气喘好转,双下肢无水肿,病情稳定。
最后应说明的是:以上各实施例仅用以说明本发明的技术方案,而非对其限制;尽管参照前述各实施例对本发明进行了详细的说明,本领域的普通技术人员应当理解:其依然可以对前述各实施例所记载的技术方案进行修改,或者对其中部分或者全部技术特征进行等同替换;而这些修改或者替换,并不使相应技术方案的本质脱离本发明各实施例技术方案的范围。
Claims (5)
1.一种治疗慢性心力衰竭的中药组合物,其特征在于,其由以下重量份组成的中药材制成:仙茅8-12份、仙灵脾8-12份、黑附片4-8份、茯苓10-15份、白术8-12份、生姜8-12份、白芍8-12份、巴戟天8-12份、甘松4-8份、大腹皮12-18份、猪苓8-12份、泽泻8-12份、桂枝8-12份、炒麦芽10-15份和焦山楂10-15份。
2.根据权利要求1所述的中药组合物,其特征在于,所述中药材的组成为:仙茅10份、仙灵脾10份、黑附片6份、茯苓15份、白术10份、生姜9份、白芍10份、巴戟天10份、甘松6份、大腹皮15份、猪苓10份、泽泻10份、桂枝10份、炒麦芽15份、焦山楂10份;或者,
所述中药材的组成为:仙茅8份、仙灵脾8份、黑附片4份、茯苓10份、白术8份、生姜8份、白芍8份、巴戟天8份、甘松4份、大腹皮12份、猪苓8份、泽泻8份、桂枝8份、炒麦芽10份、焦山楂15份;或者,
所述中药材的组成为:仙茅12份、仙灵脾12份、黑附片8份、茯苓15份、白术12份、生姜12份、白芍12份、巴戟天12份、甘松6份、大腹皮12份、猪苓12份、泽泻12份、桂枝12份、炒麦芽15份、焦山楂12份;
所述中药材的组成为:仙茅9份、仙灵脾9份、黑附片5份、茯苓14份、白术9份、生姜9份、白芍9份、巴戟天9份、甘松8份、大腹皮15份、猪苓9份、泽泻9份、桂枝9份、炒麦芽15份、焦山楂15份。
3.根据权利要求1所述的中药组合物,其特征在于,所述中药组合物为胶囊剂、中药散剂、片剂、口服液、中药药丸或水煎剂。
4.一种权利要求1或2或3所述的治疗慢性心力衰竭的中药组合物的制备方法,其特征在于,所述中药组合物为胶囊剂、中药散剂、中药药丸或水煎剂;制备方法如下:
所述胶囊剂的制备方法为:按照上述重量份称量各中药材,经清洁和切碎或切段后,先将黑附片放入砂锅中并加水,煎煮30-50min,再将其他中药材放入砂锅进行煎煮、过滤,收获滤液,将滤液继续熬煮浓缩至膏状,干燥、灭菌、研碎,过100-200目筛后,灌装胶囊;
所述中药散剂的制备方法为:按照上述重量份称量各中药材,经清洁和切碎或切段后,先将黑附片放入砂锅中并加水,煎煮30-50min,再将仙茅、仙灵脾、茯苓、白术、甘松、生姜、白芍、巴戟天、大腹皮、猪苓、泽泻、桂枝放入砂锅进行煎煮、过滤,收获滤液,将滤液继续熬煮浓缩至膏状;然后将炒麦芽和焦山楂粉碎后过筛,得到粉末;将膏状物与粉末混合后,进行制粒、烘干、灭菌,即得到中药散剂;
所述中药药丸的制备方法为:按照上述重量份称量各中药材,经清洁和切碎或切段后,先将黑附片放入砂锅中并加水,煎煮30-50min,再将其他中药材放入砂锅进行煎煮、过滤,收获滤液,将滤液继续熬煮浓缩至相对密度为1.1-1.2的稀浸膏,混入蜜丸,搅拌均匀后制成中药药丸;中药材总质量与蜜丸质量比为1:1-2;
所述水煎剂的制备方法为:按照上述重量份称量各中药材,经清洁和切碎或切段后,先将黑附片放入砂锅中并加水,煎煮30-50min,再将其他中药材放入砂锅中,第一次煎煮15-20min,倒出1/2-2/3药液;再加温水,进行第二次煎煮,倒出1/2-2/3药液,将两次药液混合,即为所述水煎剂。
5.根据权利要求4所述的中药组合物的制备方法,其特征在于,在煎煮过程中,加水量为没过全部中药材3-5cm。
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