CN209584264U - Circumfusion formula biology artificial hepatocyte reactor - Google Patents
Circumfusion formula biology artificial hepatocyte reactor Download PDFInfo
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- 210000004027 cell Anatomy 0.000 abstract description 49
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Abstract
本实用新型公开了一种循环灌注式生物人工肝细胞反应器，包括培养罐、储存罐、导入管、导出管和蠕动泵；培养罐内具有若干个水平设置、呈层状分布的细胞容量调节平板；细胞容量调节平板具有若干个通孔；储存罐包括水平设置的气体交换膜，气体交换膜的边缘与储存罐的内壁无缝相接，液体培养基置于气体交换膜上；储存罐的底面镂空；导入管一端插入储存罐内的液体培养基中，另一端与培养罐的底端连通；蠕动泵设置在导入管上，将储存罐内的液体培养基输送至培养罐中；导出管一端与培养罐的顶端连通，另一端插入储存罐内。本实用新型具有满足临床用细胞量、高交换效率、高细胞黏附率及分布均匀度、低剪切力、高生物相容性等优点。
The utility model discloses a circulating perfusion bioartificial liver cell reactor, which comprises a culture tank, a storage tank, an inlet pipe, an outlet pipe and a peristaltic pump; the culture tank has several horizontally arranged and layered cell capacity adjustments flat plate; the cell capacity adjustment plate has several through holes; the storage tank includes a gas exchange membrane arranged horizontally, the edge of the gas exchange membrane is seamlessly connected with the inner wall of the storage tank, and the liquid medium is placed on the gas exchange membrane; the storage tank The bottom surface is hollowed out; one end of the introduction tube is inserted into the liquid medium in the storage tank, and the other end is connected to the bottom of the culture tank; a peristaltic pump is installed on the introduction tube to transport the liquid medium in the storage tank to the culture tank; the outlet tube One end communicates with the top of the culture tank, and the other end is inserted into the storage tank. The utility model has the advantages of satisfying the amount of clinically used cells, high exchange efficiency, high cell adhesion rate and distribution uniformity, low shear force, high biocompatibility and the like.
Description
技术领域technical field
本实用新型属于医疗机械领域，涉及一种细胞反应器，尤其涉及一种循环灌注式生物人工肝细胞反应器。The utility model belongs to the field of medical machinery and relates to a cell reactor, in particular to a circulating perfusion bioartificial liver cell reactor.
背景技术Background technique
肝功能衰竭患者病死率高，生物人工肝能为患者提供有效的肝功能支持治疗，具有良好的应用前景。生物反应器作为生物人工肝的核心成分，目的是培养数量充足、具有分泌、代谢、解毒等功能正常的细胞，因此决定了生物人工肝的性能。The mortality rate of patients with liver failure is high, and the bioartificial liver can provide effective liver function support therapy for patients, and has a good application prospect. As the core component of the bioartificial liver, the bioreactor is aimed at cultivating a sufficient number of cells with normal functions such as secretion, metabolism, and detoxification, which determines the performance of the bioartificial liver.
生物反应器为肝细胞培养提供适宜的生长代谢环境，也为肝细胞与患者血液相互作用、进行物质交换提供条件。且生物反应器中细胞数量需高于人肝细胞数量15％才有意义，仍是目前研究难点。目前临床应用较多的是灌流床/支架型生物反应器和中空纤维型生物反应器。但其分别存在；细胞活性和功能维持较差、细胞剪切力差易脱落、细胞密度不均匀、生物相容性差、物质交换能力低和微载体颗粒悬浮效果差、物质交换能力低、易阻塞、细胞活性低下等缺点。The bioreactor provides a suitable growth and metabolism environment for the cultivation of hepatocytes, and also provides conditions for the interaction and material exchange between hepatocytes and the patient's blood. And the number of cells in the bioreactor needs to be 15% higher than the number of human hepatocytes to be meaningful, which is still a difficult research point at present. At present, more clinical applications are perfusion bed/stent bioreactor and hollow fiber bioreactor. But they exist separately; poor cell viability and function maintenance, poor cell shear force, easy to fall off, uneven cell density, poor biocompatibility, low material exchange capacity and poor suspension effect of microcarrier particles, low material exchange capacity, easy to block , Low cell viability and other shortcomings.
实用新型内容Utility model content
本实用新型提供一种循环灌注式生物人工肝细胞反应器，以克服现有技术的缺陷。The utility model provides a circulating perfusion bioartificial liver cell reactor to overcome the defects of the prior art.
为实现上述目的，本实用新型提供一种循环灌注式生物人工肝细胞反应器，包括培养罐、储存罐、导入管、导出管和蠕动泵；培养罐内具有若干个水平设置、呈层状分布的细胞容量调节平板；细胞容量调节平板具有若干个通孔；储存罐包括水平设置的气体交换膜，气体交换膜的边缘与储存罐的内壁无缝相接，液体培养基置于气体交换膜上；储存罐的底面镂空；导入管一端插入储存罐内的液体培养基中，另一端与培养罐的底端连通；蠕动泵设置在导入管上，将储存罐内的液体培养基输送至培养罐中；导出管一端与培养罐的顶端连通，另一端插入储存罐内。In order to achieve the above purpose, the utility model provides a circulating perfusion bioartificial liver cell reactor, including a culture tank, a storage tank, an inlet pipe, an outlet pipe and a peristaltic pump; there are several horizontally arranged and layered distributions in the culture tank The cell capacity adjustment plate; the cell capacity adjustment plate has several through holes; the storage tank includes a gas exchange membrane arranged horizontally, the edge of the gas exchange membrane is seamlessly connected with the inner wall of the storage tank, and the liquid medium is placed on the gas exchange membrane ;The bottom surface of the storage tank is hollowed out; one end of the introduction tube is inserted into the liquid medium in the storage tank, and the other end communicates with the bottom of the culture tank; the peristaltic pump is arranged on the introduction tube to transport the liquid culture medium in the storage tank to the culture tank Middle; one end of the outlet tube communicates with the top of the culture tank, and the other end is inserted into the storage tank.
进一步，本实用新型提供一种循环灌注式生物人工肝细胞反应器，还可以具有这样的特征：其中，细胞容量调节平板的上、下表面分别具有上连接体和下连接体，上连接体和下连接体中空、并相通；上连接体与下连接体相匹配，下层的细胞容量调节平板的上连接体与其上层的细胞容量调节平板的下连接体可拆卸固定连接、并相通；上连接体和下连接体的侧壁具有漏液孔。Further, the utility model provides a circulating perfusion bioartificial liver cell reactor, which can also have the following characteristics: wherein, the upper and lower surfaces of the cell capacity adjustment plate have an upper connector and a lower connector respectively, and the upper connector and the lower connector The lower connector is hollow and connected; the upper connector matches the lower connector, and the upper connector of the lower cell capacity adjustment plate is detachably fixedly connected with the lower connector of the upper cell capacity adjustment plate and communicates; the upper connector and the side wall of the lower connecting body has a liquid leakage hole.
进一步，本实用新型提供一种循环灌注式生物人工肝细胞反应器，还可以具有这样的特征：其中，上连接体和下连接体均为圆柱体，设置在细胞容量调节平板的中心；上连接体和下连接体分别具有相匹配的内螺纹和外螺纹；下层的细胞容量调节平板的上连接体与其上层的细胞容量调节平板的下连接体螺纹连接；若干个所示细胞容量调节平板的上连接体和下连接体相互连接形成中心通道；连接的上连接体和下连接体的漏液孔重合。Further, the utility model provides a circulating perfusion bioartificial liver cell reactor, which can also have the following characteristics: wherein, the upper connector and the lower connector are both cylinders, which are arranged in the center of the cell capacity adjustment plate; the upper connector The body and the lower connector have matching internal threads and external threads respectively; the upper connector of the lower cell capacity adjustment plate is screwed to the lower connector of the upper cell capacity adjustment plate; the upper connectors of several shown cell capacity adjustment plates The connecting body and the lower connecting body are connected to each other to form a central channel; the leakage holes of the connected upper connecting body and the lower connecting body coincide.
进一步，本实用新型提供一种循环灌注式生物人工肝细胞反应器，还可以具有这样的特征：其中，底层的细胞容量调节平板的下连接体与导入管的端部相通。Further, the utility model provides a circulating perfusion bioartificial liver cell reactor, which may also have the following feature: wherein, the lower connecting body of the bottom cell capacity adjustment plate communicates with the end of the introduction tube.
进一步，本实用新型提供一种循环灌注式生物人工肝细胞反应器，还可以具有这样的特征：其中，培养罐的底面上具有底面连接体，底面连接体中空，并与导入管相通；底面连接体具有内螺纹；底层的细胞容量调节平板的下连接体与底面连接体螺纹连接。Further, the utility model provides a circulating perfusion bioartificial liver cell reactor, which may also have the following features: wherein, the bottom surface of the culture tank has a bottom connector, the bottom connector is hollow, and communicates with the introduction tube; the bottom connects The body has an internal thread; the lower connecting body of the bottom cell capacity adjustment plate is threadedly connected with the bottom connecting body.
进一步，本实用新型提供一种循环灌注式生物人工肝细胞反应器，还可以具有这样的特征：其中，细胞容量调节平板为聚苯乙烯平板，上表面包覆有赖氨酸层；和/或，气体交换膜为硅橡胶薄膜。Further, the utility model provides a circulating perfusion bioartificial liver cell reactor, which may also have the following characteristics: wherein, the cell capacity adjustment plate is a polystyrene plate, and the upper surface is coated with a lysine layer; and/or, The gas exchange membrane is a silicone rubber membrane.
进一步，本实用新型提供一种循环灌注式生物人工肝细胞反应器，还可以具有这样的特征：其中，储存罐还包括镂空的支撑网，支撑网固定在储存罐内，气体交换膜置于支撑网上；支撑网和气体交换膜设置在储存罐的下部。Furthermore, the utility model provides a circulating perfusion bioartificial liver cell reactor, which may also have the following characteristics: wherein, the storage tank also includes a hollow support net, the support net is fixed in the storage tank, and the gas exchange membrane is placed on the support The net; the support net and the gas exchange membrane are arranged at the lower part of the storage tank.
进一步，本实用新型提供一种循环灌注式生物人工肝细胞反应器，还可以具有这样的特征：其中，细胞容量调节平板上的通孔呈中心对称分布。Further, the utility model provides a circulating perfusion bioartificial liver cell reactor, which may also have the following feature: wherein, the through holes on the cell capacity adjustment plate are distributed centrally and symmetrically.
进一步，本实用新型提供一种循环灌注式生物人工肝细胞反应器，还可以具有这样的特征：其中，培养罐的顶部具有培养罐开口，还包括与培养罐开口相匹配的培养罐盖；导出管的端部穿过培养罐盖，置于培养罐内的顶端；储存罐的顶部具有储存罐开口，还包括与储存罐开口相匹配的储存罐盖；导入管和导出管的分别穿过储存罐盖，伸入储存罐内。Further, the utility model provides a circulating perfusion bioartificial liver cell reactor, which can also have the following features: wherein, the top of the culture tank has a culture tank opening, and also includes a culture tank cover that matches the culture tank opening; The end of the tube passes through the culture tank cover and is placed at the top of the culture tank; the top of the storage tank has a storage tank opening, and also includes a storage tank cover matching the storage tank opening; the inlet tube and the outlet tube respectively pass through the storage tank The jar lid, which protrudes into the storage jar.
本实用新型的有益效果在于：本实用新型提供一种循环灌注式生物人工肝细胞反应器，培养罐内设有若干个细胞容量调节平板，可以增加有效面积、减少反应体积，还可以通过增减细胞容量调节平板的层数来调节所需细胞含量及载体容量。细胞容量调节平板为聚苯乙烯平板，上表面包覆有赖氨酸层，提高生物相容性，提高肝细胞的贴壁粘附率，降低剪切力。其次，培养罐中，液体培养基从培养罐底部漫流，再从顶部溢出，逆向灌流，有利于液体培养基的充分交换，降低液体培养基流动对细胞的影响。此外，本反应器包括单独的储存罐，储存罐容量较大，可确保营养物质的充分循环，只需更换储存罐即可进行培养基更换，克服了之前培养基更换复杂，需涉及培养罐换液的难题。同时，储存罐底部镂空，下部设有支撑网和气体交换膜，无需通过外力、直接充分进行气体交换，有效避免死腔、气泡的产生，降低了生物反应器的成本，缩减了生物反应器的复杂结构。本实用新型具有满足临床用细胞量、高交换效率、高细胞黏附率及分布均匀度、低剪切力、高生物相容性等优点。The beneficial effect of the utility model is that: the utility model provides a circulating perfusion bioartificial liver cell reactor, and several cell capacity adjustment plates are arranged in the culture tank, which can increase the effective area, reduce the reaction volume, and can also increase or decrease Cell capacity adjusts the number of layers of the plate to adjust the required cell content and carrier capacity. The cell capacity adjustment plate is a polystyrene plate, and the upper surface is coated with a lysine layer to improve biocompatibility, increase the adhesion rate of hepatocytes to the wall, and reduce the shear force. Secondly, in the culture tank, the liquid medium overflows from the bottom of the culture tank, and then overflows from the top, reverse perfusion, which is conducive to the full exchange of the liquid medium and reduces the impact of the flow of the liquid medium on the cells. In addition, this reactor includes a separate storage tank with a large capacity to ensure sufficient circulation of nutrients. The medium can be replaced only by replacing the storage tank. Liquid problem. At the same time, the bottom of the storage tank is hollowed out, and the lower part is equipped with a support net and a gas exchange membrane, which can directly and fully perform gas exchange without external force, effectively avoiding the generation of dead space and air bubbles, reducing the cost of the bioreactor and reducing the cost of the bioreactor. complex structure. The utility model has the advantages of satisfying the amount of clinically used cells, high exchange efficiency, high cell adhesion rate and distribution uniformity, low shear force, high biocompatibility and the like.
附图说明Description of drawings
图1是循环灌注式生物人工肝细胞反应器的结构示意图；Fig. 1 is a structural schematic diagram of a circulating perfusion bioartificial liver cell reactor;
图2是细胞容量调节平板的主视图；Fig. 2 is the front view of the cell volume adjustment plate;
图3是细胞容量调节平板的俯视图；Figure 3 is a top view of the cell capacity adjustment plate;
图4是细胞容量调节平板的纵向剖面图；Fig. 4 is a longitudinal sectional view of a cell volume adjustment plate;
图5是培养罐的纵向剖面示意图；Fig. 5 is the longitudinal sectional schematic diagram of cultivation tank;
图6是支撑网的俯视图。Figure 6 is a top view of the support net.
具体实施方式Detailed ways
以下结合附图来说明本实用新型的具体实施方式。The specific embodiment of the utility model is described below in conjunction with accompanying drawing.
如图1所示，本实用新型提供一种循环灌注式生物人工肝细胞反应器，包括培养罐1、储存罐2、导入管3、导出管4和蠕动泵5。As shown in FIG. 1 , the utility model provides a circulating perfusion bioartificial liver cell reactor, which includes a culture tank 1 , a storage tank 2 , an inlet tube 3 , an outlet tube 4 and a peristaltic pump 5 .
培养罐1的顶部具有培养罐开口11，还包括与培养罐开口相匹配的培养罐盖12，培养罐盖12可以密封培养罐开口11。The top of the culture tank 1 has a culture tank opening 11 , and also includes a culture tank cover 12 matching the culture tank opening, and the culture tank cover 12 can seal the culture tank opening 11 .
如图1-4所示，培养罐1内具有若干个水平设置、呈层状分布的细胞容量调节平板13。细胞容量调节平板13为圆盘状，具有若干个通孔 131，通孔131呈中心对称分布。As shown in FIGS. 1-4 , there are several horizontally arranged and layered cell capacity adjustment plates 13 in the culture tank 1 . The cell capacity regulating plate 13 is disc-shaped and has several through holes 131, and the through holes 131 are symmetrically distributed in the center.
细胞容量调节平板13的上、下表面的中心分别具有上连接体132 和下连接体133。上连接体132和下连接体133均为中空的圆柱体，且二者相通。上连接体132和下连接体133分别具有相匹配的内螺纹和外螺纹，下层的细胞容量调节平板13的上连接体132与其上层的细胞容量调节平板13的下连接体133螺纹连接。若干个细胞容量调节平板13依次连接成为整体，且其上连接体132和下连接体133相互连接形成中心通道134，如图5所示。The center of the upper and lower surfaces of the cell capacity adjustment plate 13 has an upper connecting body 132 and a lower connecting body 133 respectively. Both the upper connecting body 132 and the lower connecting body 133 are hollow cylinders, and they communicate with each other. The upper connecting body 132 and the lower connecting body 133 have matching internal threads and external threads respectively, and the upper connecting body 132 of the lower cell capacity adjusting plate 13 is screwed to the lower connecting body 133 of the upper cell capacity adjusting plate 13 . Several cell capacity adjustment plates 13 are sequentially connected to form a whole, and the upper connecting body 132 and the lower connecting body 133 are connected to each other to form a central channel 134, as shown in FIG. 5 .
上连接体132和下连接体133的侧壁具有漏液孔135，连接的上连接体132和下连接体133的漏液孔135重合。The side walls of the upper connecting body 132 and the lower connecting body 133 have liquid leakage holes 135 , and the liquid leakage holes 135 of the connected upper connecting body 132 and lower connecting body 133 overlap.
本实施例中，上连接体和下连接体也可以为其他相匹配的可拆卸固定连接结构。In this embodiment, the upper connecting body and the lower connecting body can also be other matched detachable fixed connecting structures.
细胞容量调节平板13为聚苯乙烯平板，上表面包覆有赖氨酸层。The cell capacity adjustment plate 13 is a polystyrene plate, and the upper surface is coated with a lysine layer.
本实施例中，细胞容量调节平板13的数量可根据使用需求而设制。In this embodiment, the number of cell capacity adjustment plates 13 can be set according to the usage requirements.
培养罐1的底面上具有中空的、圆柱状的底面连接体14。底面连接体14具有内螺纹，与细胞容量调节平板13的下连接体133的外螺纹相匹配。底层的细胞容量调节平板13的下连接体133与底面连接体14螺纹连接、并相通，从而使若干个细胞容量调节平板13整体固定在培养罐 1内。The bottom surface of the culture tank 1 has a hollow, cylindrical bottom surface connecting body 14 . The bottom connecting body 14 has an internal thread, which matches the external thread of the lower connecting body 133 of the cell capacity adjustment plate 13 . The lower connecting body 133 of the bottom cell capacity regulating plate 13 is screwed to the bottom connecting body 14 and communicated, so that several cell capacity regulating plates 13 are integrally fixed in the culture tank 1.
储存罐2的顶部具有储存罐开口21，还包括与储存罐开口21相匹配的储存罐盖22，储存罐盖22可以密封储存罐开口21。The top of the storage tank 2 has a storage tank opening 21 , and also includes a storage tank cover 22 matching the storage tank opening 21 , and the storage tank cover 22 can seal the storage tank opening 21 .
储存罐2包括水平设置在其下部的支撑网23和气体交换膜24。如图6所示，支撑网23为平面网状结构，固定在储存罐2内。气体交换膜 24为硅橡胶薄膜，透气不透液。气体交换膜24膜置于支撑网23上，边缘与储存罐2的内壁无缝相接，液体培养基置于气体交换膜24上。储存罐2的底面镂空，气体交换膜24上的液体培养基可以进行充分气体交换。The storage tank 2 includes a support net 23 and a gas exchange membrane 24 horizontally arranged at its lower portion. As shown in FIG. 6 , the supporting net 23 is a planar mesh structure, and is fixed in the storage tank 2 . The gas exchange membrane 24 is a silicon rubber film, which is air-tight and liquid-tight. The gas exchange membrane 24 is placed on the supporting net 23 , and the edge is seamlessly connected with the inner wall of the storage tank 2 , and the liquid culture medium is placed on the gas exchange membrane 24 . The bottom surface of the storage tank 2 is hollowed out, and the liquid culture medium on the gas exchange membrane 24 can perform sufficient gas exchange.
导入管3一端穿过储存罐盖22、插入储存罐2内的液体培养基中，另一端与培养罐1底端的底面连接体14连通。One end of the introduction pipe 3 passes through the storage tank cover 22 and is inserted into the liquid culture medium in the storage tank 2 , and the other end communicates with the bottom surface connector 14 at the bottom of the culture tank 1 .
导出管4一端穿过培养罐盖12、置于培养罐1内的顶端，另一端穿过储存罐盖22、插入储存罐2内。One end of the outlet pipe 4 passes through the culture tank cover 12 and is placed on the top of the culture tank 1 , and the other end passes through the storage tank cover 22 and is inserted into the storage tank 2 .
蠕动泵5设置在导入管3上，将储存罐2内的液体培养基输送至培养罐1中。The peristaltic pump 5 is arranged on the introduction pipe 3 to transport the liquid culture medium in the storage tank 2 to the culture tank 1 .
使用前，打开培养罐盖12和储存罐盖22，将循环灌注式生物人工肝细胞反应器的去除蠕动泵5之外的剩余整体，置于环氧乙烷气体下灭菌过夜，灭菌完毕后置于超净工作台内备用。Before use, open the culture tank cover 12 and the storage tank cover 22, remove the remaining whole of the circulating perfusion bioartificial liver cell reactor except the peristaltic pump 5, and sterilize it under ethylene oxide gas overnight, and the sterilization is completed Then put it in the ultra-clean workbench for later use.
循环灌注式生物人工肝细胞反应器的使用方法为：The usage method of circulating perfusion bioartificial liver cell reactor is as follows:
步骤一，打开储存罐盖22，将液体培养基注入储存罐2。Step 1, open the storage tank cover 22, and inject the liquid culture medium into the storage tank 2.
步骤二，关闭培养罐盖12和储存罐盖22，打开蠕动泵5，将储存罐 2内的液体培养基抽入导入管3中，至液体培养基充满整个导入管3，即液体培养基输至培养罐1的底端，关闭蠕动泵5。Step 2, close the culture tank cover 12 and the storage tank cover 22, open the peristaltic pump 5, and pump the liquid culture medium in the storage tank 2 into the introduction pipe 3 until the liquid culture medium is full of the entire introduction pipe 3, that is, the liquid culture medium infusion To the bottom of the culture tank 1, turn off the peristaltic pump 5.
步骤三，打开培养罐盖12，将含有细胞的液体培养基由上至下注入培养罐1，待该培养基均匀分布在各层细胞容量调节平板13上表面时，完成初始注入，关闭培养罐盖12。由于细胞只能通过漏液孔135及通孔 131流出，因此可以均匀地分布在各细胞容量调节平板13表面。Step 3, open the culture tank cover 12, pour the liquid medium containing the cells into the culture tank 1 from top to bottom, and when the culture medium is evenly distributed on the upper surface of the cell capacity adjustment plate 13 of each layer, complete the initial injection, and close the culture tank Cover 12. Since the cells can only flow out through the leakage hole 135 and the through hole 131, they can be evenly distributed on the surface of each cell capacity adjustment plate 13.
步骤四，将反应器整体置于37℃环境下，静置2-4h，使肝细胞贴牢固壁于细胞容量调节平板13。Step 4, place the whole reactor in an environment of 37° C., and let it stand still for 2-4 hours, so that the liver cells can be firmly adhered to the cell volume adjustment plate 13 .
步骤五，进行培养基的循环灌注：打开蠕动泵5，储存罐2内的液体培养基通过导入管3，由下至上注入培养罐1，液体培养基上升至培养罐1的顶部时，由导出管4排出至储存罐2。液体培养基从培养罐1的底部逆重力的缓慢向上灌注，可有效降低液体培养基对细胞的剪切力，也可以避免气泡和交换死腔的形成，并可以将通过气体交换膜24的氧气以及营养物质提供给细胞，也可以带走细胞的代谢产物，实现物质的充分交换。Step 5, carry out the circulation perfusion of the culture medium: turn on the peristaltic pump 5, the liquid medium in the storage tank 2 passes through the introduction pipe 3, and pours into the culture tank 1 from bottom to top, when the liquid culture medium rises to the top of the culture tank 1, it is exported Tube 4 discharges to storage tank 2 . The liquid medium is perfused slowly upward against the gravity from the bottom of the culture tank 1, which can effectively reduce the shear force of the liquid medium on the cells, and can also avoid the formation of air bubbles and exchange dead spaces, and can transfer the oxygen passing through the gas exchange membrane 24. As well as providing nutrients to the cells, it can also take away the metabolites of the cells to achieve a full exchange of substances.
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