CN110229752B - Micro negative pressure cell culture device - Google Patents
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- CN110229752B CN110229752B CN201910571338.5A CN201910571338A CN110229752B CN 110229752 B CN110229752 B CN 110229752B CN 201910571338 A CN201910571338 A CN 201910571338A CN 110229752 B CN110229752 B CN 110229752B
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- 238000004113 cell culture Methods 0.000 title claims abstract description 50
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 47
- 230000001105 regulatory effect Effects 0.000 claims abstract description 28
- 238000002347 injection Methods 0.000 claims abstract description 6
- 239000007924 injection Substances 0.000 claims abstract description 6
- 239000007788 liquid Substances 0.000 claims description 6
- 238000007789 sealing Methods 0.000 claims description 6
- 210000004027 cell Anatomy 0.000 abstract description 20
- 230000009193 crawling Effects 0.000 abstract description 10
- 210000002950 fibroblast Anatomy 0.000 abstract description 7
- 210000001339 epidermal cell Anatomy 0.000 abstract description 4
- 230000001737 promoting effect Effects 0.000 abstract description 4
- 230000009286 beneficial effect Effects 0.000 abstract description 3
- 238000000338 in vitro Methods 0.000 abstract description 2
- 230000035755 proliferation Effects 0.000 abstract description 2
- 230000037314 wound repair Effects 0.000 abstract description 2
- 206010052428 Wound Diseases 0.000 description 13
- 208000027418 Wounds and injury Diseases 0.000 description 12
- 238000002474 experimental method Methods 0.000 description 8
- 210000002510 keratinocyte Anatomy 0.000 description 5
- 230000037311 normal skin Effects 0.000 description 5
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 description 4
- 230000000694 effects Effects 0.000 description 4
- 238000010586 diagram Methods 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 210000002615 epidermis Anatomy 0.000 description 3
- 238000009434 installation Methods 0.000 description 3
- 238000000034 method Methods 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- 229910002092 carbon dioxide Inorganic materials 0.000 description 2
- 239000001569 carbon dioxide Substances 0.000 description 2
- 230000002349 favourable effect Effects 0.000 description 2
- 230000003179 granulation Effects 0.000 description 2
- 238000005469 granulation Methods 0.000 description 2
- 238000011081 inoculation Methods 0.000 description 2
- 230000005012 migration Effects 0.000 description 2
- 238000013508 migration Methods 0.000 description 2
- 238000005086 pumping Methods 0.000 description 2
- 210000001519 tissue Anatomy 0.000 description 2
- 206010063560 Excessive granulation tissue Diseases 0.000 description 1
- 206010030113 Oedema Diseases 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 230000001580 bacterial effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 230000000052 comparative effect Effects 0.000 description 1
- 230000019305 fibroblast migration Effects 0.000 description 1
- 210000001126 granulation tissue Anatomy 0.000 description 1
- 230000035876 healing Effects 0.000 description 1
- 238000012606 in vitro cell culture Methods 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 230000004089 microcirculation Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000000399 orthopedic effect Effects 0.000 description 1
- 239000011295 pitch Substances 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 230000008929 regeneration Effects 0.000 description 1
- 238000011069 regeneration method Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 239000000523 sample Substances 0.000 description 1
- 210000003491 skin Anatomy 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 238000001356 surgical procedure Methods 0.000 description 1
- 210000003556 vascular endothelial cell Anatomy 0.000 description 1
- 230000029663 wound healing Effects 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M35/00—Means for application of stress for stimulating the growth of microorganisms or the generation of fermentation or metabolic products; Means for electroporation or cell fusion
- C12M35/04—Mechanical means, e.g. sonic waves, stretching forces, pressure or shear stimuli
Landscapes
- Engineering & Computer Science (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Zoology (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Genetics & Genomics (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- Wood Science & Technology (AREA)
- Sustainable Development (AREA)
- Microbiology (AREA)
- Mechanical Engineering (AREA)
- Biomedical Technology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Cell Biology (AREA)
- Apparatus Associated With Microorganisms And Enzymes (AREA)
Abstract
The invention provides a micro negative pressure cell culture device, which comprises CO 2 Cell incubator, placed in the CO 2 A cell culture device in a cell culture box, a pressure fine regulator and a negative pressure pump. The pressure fine regulator comprises a pressure regulating cavity and a water injection pipeline communicated with the side end, wherein the bottom end of the pressure regulating cavity is provided with a water outlet pipeline, and the top end of the pressure regulating cavity is provided with a pipe joint; the cell culture device is provided with two pipes, one pipe is connected with a negative pressure pump, and the other pipe is arranged on a pipe joint. Experimental study proves that the micro negative pressure cell culture device has a certain promoting effect on cell crawling and proliferation, and the epidermal cells and the fibroblasts cultured in vitro show stronger crawling capability, thereby providing beneficial exploration for clinical wound repair.
Description
Technical Field
The invention belongs to the technical field of experimental devices, and particularly relates to a cell (epidermis, fibroblast, vascular endothelial cell and the like) culture device capable of providing micro negative pressure.
Background
The negative pressure closed drainage technology changes an open wound surface into a closed wound surface, can remove seepage of the wound surface at regular time under the rated negative pressure effect, can relieve edema among tissues, improve microcirculation of the tissues, promote regeneration of capillary vessels and reduce bacterial reproduction, thereby promoting growth of granulation tissues and creating favorable conditions for covering the wound surface. The method has wide application in burn department, surgery and orthopedic wound repair.
The negative pressure value of the current wound negative pressure drainage technology in application is-80 to-120 mmHg, the wound seepage can be effectively managed under the action of the negative pressure value, but no obvious effect is exerted on the growth and crawling of the epidermis, so in clinical practice, when the wound substrate has fresh granulation for growth, the skin grafting operation can be selected to cover the wound or the wound can be repaired by crawling the surrounding normal epidermis.
In clinical practice, after the wound surface has new granulation growth, if a certain micro negative pressure is given, the wound surface of a patient heals more quickly and to a better degree than the wound surface healed by self. However, the minimum negative pressure which can be generated by the negative pressure pump adopted in the prior art is generally-30 mmHg to-50 mmHg, and strictly speaking, the negative pressure pump does not belong to the category of micro negative pressure, so that even if the negative pressure pump is regulated to the minimum, the crawling of the epidermal cells is still little influenced. The negative pressure pump manufacturers have difficulty in further reducing the negative pressure value of the existing machine to a micro negative pressure value (-5 to-10 mmHg) at present due to the limitation of the negative pressure pump probe technology. Therefore, whether the micro negative pressure can promote wound healing or not can not be verified by adopting the current negative pressure closed drainage equipment.
Disclosure of Invention
The invention aims to solve the technical problems and provides a micro negative pressure cell culture device capable of providing a micro negative pressure environment for cell culture. In order to achieve the purpose, the technical scheme adopted by the invention is as follows:
the invention provides a micro negative pressure cell culture deviceThe method has the following technical characteristics: comprising CO 2 Cell incubator placed in the CO 2 A cell culture device in a cell culture box, a pressure fine regulator and a negative pressure pump. The pressure fine regulator comprises a pressure regulating cavity and a water injection pipeline communicated with the side end, wherein the bottom end of the pressure regulating cavity is provided with a water outlet pipeline, and the top end of the pressure regulating cavity is provided with a pipe joint; the cell culture device is provided with two pipes, one pipe is connected with a negative pressure pump, and the other pipe is arranged on a pipe joint.
In the invention, a negative pressure pump is arranged in CO 2 The reason for the cell culture chamber, not being placed outside, is: the negative pressure pump will absorb CO 2 The gas is released in the external environment, the pump is arranged outside, the carbon dioxide gas of the incubator can be consumed, and carbon dioxide needs to be added continuously; is placed in the cell incubator, can relatively maintain CO in the incubator 2 Balance of gas.
Preferably, the micro negative pressure cell culture device provided by the invention further comprises a pressure fine regulator placing frame, wherein the pressure fine regulator placing frame comprises an upper layer and a lower layer, the upper layer is provided with a placing groove with a through hole at the bottom wall, and the lower layer is provided with a push-pull water receiving groove.
The pressure fine adjuster placing frame is also placed in CO 2 In the cell incubator, the pressure fine tuning device is placed in the placing groove, two water outlet pipelines of the pressure fine tuning device penetrate through the through holes and are positioned above the push-pull water receiving groove, so that pressure adjustment is realized at any time, and when water in the water receiving groove is accumulated to a certain degree, the water receiving groove is taken out and poured out.
Preferably, in the micro negative pressure cell culture device provided by the invention, two guide sheets are symmetrically arranged on the inner wall of the lower layer of the pressure fine regulator placing frame, edges which are lapped on the guide sheets are arranged at two ends of the push-pull type water receiving tank, and the width of the water receiving tank is the same as the interval between the two guide sheets. The push-pull type water receiving tank is quite common in the prior art, and the invention only enumerates one of the simplest modes, and realizes the pumping and pouring of the water receiving tank and the water receiving after installation.
Preferably, in the micro negative pressure cell culture device provided by the invention, the cell culture device comprises a circular underframe, a double-layer culture frame arranged on the circular underframe, a transparent sealing cover which is sealed and buckled on the circular underframe, and a lock catch which is arranged on the periphery of the circular underframe and used for locking the transparent sealing cover, wherein two pipes which are respectively connected with the negative pressure pump and the pressure fine regulator are arranged on the circular underframe.
Preferably, in the micro negative pressure cell culture device provided by the invention, a valve is arranged on the water outlet pipeline and is positioned above the push-pull water receiving tank.
Preferably, in the micro negative pressure cell culture device provided by the invention, the front wall of the pressure regulating cavity is provided with scale marks.
Preferably, in the micro negative pressure cell culture device provided by the invention, the negative pressure pump is provided with a display screen, and the negative pressure pump selected by the inventor in actual experiment is a curasys negative pressure pump of CGBIO company.
Effects and effects of the invention
According to the micro negative pressure cell culture device provided by the invention, as the pressure fine regulator is arranged, the micro negative pressure environment of-5 mmHg to-10 mmHg is formed in the cell culture device by regulating the liquid level height in the pressure regulating cavity, and experiments prove that the crawling and proliferation speed of the epidermal cells in the micro negative pressure environment is obviously faster than that of the experimental results in the non-micro negative pressure environment, so that the micro negative pressure cell culture device is favorable for clinically simulating and exploring and promoting the healing of wound surfaces.
In addition, the pressure fine adjuster is simple in structure, low in preparation cost and beneficial to popularization and application in laboratories and clinic.
Drawings
FIG. 1 is a schematic diagram of a micro negative pressure cell culture apparatus according to an embodiment of the present invention;
FIG. 2 is a schematic diagram of a pressure trimmer in accordance with an embodiment of the present invention;
FIG. 3 is a schematic view of a pressure trimmer placement frame in accordance with an embodiment of the present invention;
FIG. 4 is a microscopic result of a normal skin keratinocyte streak test in an example of the present invention;
FIG. 5 is the microscopic results of the fibroblast scratch test in the examples of the present invention.
Detailed Description
The present invention will be described in detail with reference to the following examples and the accompanying drawings. The following examples should not be construed as limiting the scope of the invention.
In the description of the present invention, it should be noted that the azimuth or positional relationship indicated by the terms "upper", "lower", "left", "right", etc. are based on the azimuth or positional relationship shown in the drawings, and are merely for convenience of describing the present invention and simplifying the description, and do not indicate or imply that the apparatus or element referred to must have a specific azimuth, be constructed and operated in a specific azimuth, and thus should not be construed as limiting the present invention.
FIG. 1 is a schematic diagram of a micro negative pressure cell culture apparatus according to an embodiment of the present invention.
As shown in FIG. 1, micro negative pressure cell culture apparatus 100 comprises CO 2 A cell incubator 1 placed in the CO 2 A cell culture device 2, a pressure fine regulator 3, a pressure fine regulator placing frame 4 and a negative pressure pump 5 in the cell culture box.
CO 2 Cell incubator 1 is a common incubator for biological laboratories, and cell incubator 2 is a relatively sealed cell incubator, and comprises a circular underframe, a double-layer incubator 21 arranged on the circular underframe, a transparent sealing cover 22 sealed and buckled on the circular underframe, and a lock catch 23 arranged on the periphery of the circular underframe and used for locking the transparent sealing cover. The circular underframe is provided with two pipes 24 and 25 respectively connected with the negative pressure pump 5 and the pressure fine adjuster 3.
Fig. 2 is a schematic structural view of a pressure fine adjuster according to an embodiment of the present invention.
As shown in fig. 1 and 2, the pressure fine adjuster 3 includes a pressure adjusting chamber 31, a water injection pipe 32 provided at a side end of the pressure adjusting chamber 31, a water outlet pipe 33 provided at a bottom end of the pressure adjusting chamber, and a pipe joint 34 provided at a top end of the pressure adjusting chamber.
The front end of the pressure regulating cavity 31 is provided with scale marks for displaying water level, and the water injection pipeline 32 is used for injecting water into the pressure regulating cavity. The formation of a micro negative pressure environment in the cell culture vessel 2 is related to the liquid level in the pressure regulating chamber, and when the liquid level is regulated, the drainage can be regulated by regulating the valve 331 on the water outlet pipe 33.
Fig. 3 is a schematic view of a pressure fine adjuster placing rack according to an embodiment of the present invention.
As shown in fig. 1 and 3, the pressure trimmer placing frame 4 is also placed in the CO 2 The cell incubator comprises an upper layer and a lower layer, wherein the upper layer is provided with a placement groove 41, and the lower layer is provided with a push-pull water receiving groove 42. The bottom wall of the placement tank 41 has a through hole 411 through which the outlet pipe 33 passes. Two guide pieces 43 are symmetrically arranged on the inner wall of the lower layer, edges 421 which are lapped on the guide pieces are arranged at the two ends of the push-pull water receiving groove, the width of the water receiving groove is the same as the interval between the two guide pieces, and the installation stability of the water receiving groove is improved. The push-pull type water receiving tank is quite common in the prior art, and the invention only enumerates one of the simplest modes, and realizes the pumping and pouring of the water receiving tank and the water receiving after installation.
When the pressure fine-tuning device is used, the pressure fine-tuning device is placed in the placement groove, the water outlet pipeline of the pressure fine-tuning device penetrates through the through hole and is positioned above the push-pull water receiving groove, so that pressure adjustment is realized at any time, and when water in the water receiving groove is accumulated to a certain degree, the water receiving groove is taken out and poured out.
In this embodiment, the negative pressure pump is placed in the CO 2 The negative pressure pump selected by the inventor in the practical experiment is a curasys negative pressure pump of CGBIO company.
The principle of realizing the micro negative pressure environment in the cell culture device by the micro negative pressure cell culture device in the embodiment is as follows: after the negative pressure pump 5 is started, the inside of the cell culture device 2 is in a negative pressure state, and then the CO above the pressure regulating cavity 31 is formed 2 Flows into the cell culture device 2, and the micro negative pressure in the cell culture device 2 is regulated by regulating the liquid level height of the pressure regulating cavity. As the minimum negative pressure value which can be displayed by the negative pressure pump is-5 mmHg to-10 mmHg, through experiments, the device in the embodiment can at least realize micro negative pressure of-5 mmHg to-10 mmHg, even smaller, and only the negative pressure pump can not display.
To verify the beneficial effect of the micro negative pressure cell culture apparatus 100 of the present invention on cell crawling experiments, the inventors conducted comparative experiments. Human normal skin keratinocytes and fibroblasts were used as subjects, and the experimental example adopted the device in this exampleCulturing in a relatively sealed cell culture vessel, wherein the culture is performed at a temperature, humidity, and CO 2 The conditions such as concentration are identical. The experimental results were compared as follows:
the cell crawling ability is observed through a scratch experiment. The intercellular distance was observed at 0, 12 and 24 hours, respectively, and the procedure was repeated 8 times, and then the cell pitches of the control example and the experimental example were compared. The specific measurement data are shown in Table 1, and the microscopic results of the normal skin keratinocytes and fibroblast scratch experiments are shown in FIGS. 4 and 5, respectively.
The results show that the cell spacing between the two groups is not different at the 0 th hour after the inoculation of the normal skin keratinocytes, P is more than 0.05, the cell spacing between the experimental examples at 12 hours and 24 hours is obviously smaller than that of the control example, and P is less than 0.05; the cell spacing of the groups 0 and 12 hours after the fibroblast inoculation is not different, P is more than 0.05, and the cell spacing of the experimental example at 24 hours is obviously smaller than that of the control example, and P is less than 0.05.
TABLE 1 in vitro cell culture scratch test cell spacing Table (X.+ -. S, mm) for control and experimental examples
Research proves that the micro negative pressure cell culture device 100 has a promoting effect on cell crawling, and both the in-vitro cultured epidermal cells and the fibroblasts show stronger crawling capacity. The enhancement of migration capacity for normal skin keratinocytes occurs earlier and begins 12 hours after receiving the negative pressure environment, the migration capacity is enhanced. Whereas for fibroblast migration capacity, enhancement occurs 24 hours after receiving a micro-negative pressure environment.
The foregoing has shown and described the basic principles, principal features and advantages of the invention. It will be understood by those skilled in the art that the present invention is not limited to the embodiments described above, and that the above embodiments and descriptions are merely illustrative of the principles of the present invention, and various changes and modifications may be made without departing from the spirit and scope of the invention, which is defined in the appended claims. The scope of the invention is defined by the appended claims and equivalents thereof.
Claims (6)
1. A micro negative pressure cell culture apparatus, comprising:
CO 2 cell incubator placed in the CO 2 A cell culture device, a pressure fine regulator and a negative pressure pump in a cell culture box,
wherein the cell culture device is a sealed culture device;
the pressure fine adjuster comprises a pressure adjusting cavity and a water injection pipeline communicated with the side end; the water injection pipeline is used for injecting water into the pressure regulating cavity; the front end of the pressure regulating cavity is provided with scale marks for displaying water level, the bottom end of the pressure regulating cavity is provided with a water outlet pipeline, the top end of the pressure regulating cavity is provided with a pipe joint, and a valve is arranged on the water outlet pipeline;
the cell culture device is provided with two pipes, one pipe is connected with the negative pressure pump, the other pipe is arranged on the pipe joint, a micro negative pressure environment is formed in the cell culture device and is related to the liquid level in the pressure regulating cavity, and when the liquid level is regulated, the water discharge can be regulated by regulating a valve on a water outlet pipeline.
2. The micro negative pressure cell culture apparatus of claim 1, further comprising:
the pressure fine regulator placing frame comprises an upper layer and a lower layer, wherein the upper layer is provided with a placing groove with a through hole on the bottom wall, and the lower layer is provided with a push-pull water receiving groove.
3. The micro negative pressure cell culture apparatus according to claim 2, wherein:
the pressure fine regulator comprises a pressure fine regulator placing frame, wherein two guide sheets are symmetrically arranged on the inner wall of the lower layer of the pressure fine regulator placing frame, edges which are lapped on the guide sheets are arranged at two ends of the push-pull type water receiving tank, and the width of the water receiving tank is the same as the interval between the two guide sheets.
4. The micro negative pressure cell culture apparatus of claim 1, wherein:
wherein the cell culture device comprises a round underframe, a double-layer culture frame arranged on the round underframe, a transparent sealing cover which is sealed and buckled on the round underframe, a lock catch which is arranged on the periphery of the round underframe and locks the transparent sealing cover,
and two pipes respectively connected with the negative pressure pump and the pressure fine adjuster are arranged on the round underframe.
5. The micro negative pressure cell culture apparatus according to claim 2, wherein:
wherein, the valve is located above the push-pull water receiving tank.
6. The micro negative pressure cell culture apparatus of claim 1, wherein:
wherein, negative pressure pump has the display screen.
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| CN201910571338.5A CN110229752B (en) | 2019-06-24 | 2019-06-24 | Micro negative pressure cell culture device |
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| CN201910571338.5A CN110229752B (en) | 2019-06-24 | 2019-06-24 | Micro negative pressure cell culture device |
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| CN110229752B true CN110229752B (en) | 2024-03-19 |
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