Bionic breathing type exposure system for inhalant

Abstract

The utility model belongs to the field of in-vitro inhalation and exposure, and particularly relates to a bionic breathing type exposure system for inhales. The system comprises a breathing power device, a mouth branch and a nose branch, the mouth branch and the nose branch are connected with the breathing power device, the downstream of the mouth branch and the downstream of the nose branch converge to form a throat structure and then are connected with a trachea branch, and the trachea branch comprises a bronchial tree with a plurality of branches. The mouth branch and / or the nose branch are / is connected with a diverter valve used for adjusting the diverter flow of the mouth branch and the nose branch; at least one of the branches is connected with a lung unit exposure device; the lung unit exposure device comprises a microporous membrane for cell growth, a gas chamber and a liquid chamber, wherein the gas chamber and the liquid chamber are located on the two sides of the microporous membrane respectively. According to the utility model, the grading and shunting environment of a bronchial tree where cells are located can be accurately simulated, the respiratory physiological microenvironment where the cells are located can be reduced, and the exposure dose of the cells can be accurately evaluated, so that a more accurate cell exposure experiment result can be obtained.

Description

Technical Field

[0001] This utility model belongs to the field of external inhalation exposure, specifically relating to a biomimetic respiratory exposure system for inhaled substances. Background Technology

[0002] Studies on pulmonary respiratory toxicology and biological effects in fields such as environmental pollution, occupational hazards, bioaerosols, inhaled preparations, industrial chemicals, pesticides, cosmetics and disinfectants cannot be separated from in vitro inhalation exposure studies. Traditional in vitro inhalation exposure studies mostly use liquid-liquid interface exposure methods. This method cannot objectively and in real time simulate the gas-liquid interface environment of lung tissue, the periodic respiratory state and the real physical state of the inhaled substance, resulting in differences between test results and actual results.

[0003] To more realistically simulate the toxicological or biological effects of inhaled substances through inhalation or special inhalation in vitro, current research has focused on full-smoke exposure systems, such as Vitrocell, and aerosol exposure systems based on biomimetic breathing, which have been developed in recent years.

[0004] The Vitrocell-based full-smoke exposure system consists of a smoking machine, a smoke delivery system, and an exposure chamber with an exposed gas-liquid interface. It is primarily used for smoke exposure, specifically for simulating specific inhalations in exposure experiments. During smoke exposure experiments, synthetic air at a certain flow rate is continuously introduced into the gas-liquid interface exposure chamber to simulate inhalation, while synthetic gas at a certain flow rate is continuously pumped out to simulate exhalation. A smoking machine is used to draw smoke to simulate smoke generation, which is then diluted by the simulated inhaled synthetic air and introduced into the exposure chamber to simulate specific inhalations. While this principle of exposure experimentation can simulate the gas-liquid interface environment of lung tissue, it cannot simulate periodic respiratory states or changes in O2 and CO2 concentrations in inhaled and exhaled gases. It also cannot reflect the transmission and distribution of inhaled / exhaled gases by the bronchial tree within the body, nor the resulting changes in exposure dose and hydrodynamics. The biomimetic level of the exposure experiment still needs improvement.

[0005] Chinese invention patent application CN 115232733 A, published on October 25, 2022, discloses an aerosol exposure system capable of biomimetic breathing, including a breathing power device. The breathing power device is connected to a biomimetic oral cavity (i.e., oral branch) and a biomimetic nasal cavity (i.e., nasal branch) arranged in parallel. Downstream of the biomimetic oral cavity and biomimetic nasal cavity are connected a biomimetic pharynx and a biomimetic tracheobronchial tree (i.e., tracheal branch). The tracheobronchial tree includes a first lung unit simulating the left lung and a second and third lung units simulating the right lung. An intrapulmonary culture unit is provided in the second lung unit. The lower openings of the first, second, and third lung units are connected to a carbon dioxide supply device to simulate the increase in CO2 content when exhaled.

[0006] The aforementioned aerosol exposure system further improves the biomimetic nature of respiratory exposure devices, simulating changes in CO2 content during inhalation and exhalation, and reflecting the expansion and contraction states of lung units under constant exposure pressure. However, the following problems still exist: First, it does not provide specific devices and methods for distributing different flow rates via the mouth and nose. Therefore, when studying inhaled substances of different volumes, resistances, and compositions (such as cigarette smoke) via the mouth and nose, it cannot simulate the real inhalation and exhalation patterns of distributing different volumes via the mouth and nose. Second, although the system provides a method for constructing the respiratory tract, it does not provide the distribution ratio of inhalation and exhalation at different levels of the bronchial tree, which makes it difficult to scientifically... This study explores effective exposure doses at different bronchial tree levels and branches. While some experiments and computer modeling have provided information on shunting patterns at different bronchial tree levels, the modeling equipment and data used in these experiments differ from the actual bronchial trees and exposure units used in human bodies and systems. Therefore, exploring exposure doses at different branches of the bronchial tree under this system's exposure mode is not particularly meaningful. Furthermore, it lacks specific cell culture and exposure devices and methods adapted to different bronchial tree levels, supporting cell culture, exposure, and CO2 introduction. This makes it impossible to scientifically characterize the true physiological microenvironment of the cells. Therefore, reliable and definitive experimental protocols and exposure doses cannot be provided. Utility Model Content

[0007] The purpose of this invention is to provide a biomimetic respiratory exposure system for inhaled substances, in order to solve the problems that existing exposure systems cannot accurately characterize the respiratory physiological microenvironment of cells, cannot accurately simulate the oral-nasal shunt problem, cannot provide the exposure dose for different branches of the bronchial tree at levels 0-3, and further cannot accurately evaluate the exposure dose.

[0008] To achieve the above objectives, the technical solution adopted by this utility model is as follows:

[0009] A biomimetic respiratory exposure system for inhaled substances includes a respiratory power device and oral and nasal branches connected to the respiratory power device. The downstream branches of the oral and nasal branches merge to form a pharyngeal structure and then connect to a tracheal branch. The tracheal branch includes a bronchial tree with multiple branches. A shunt valve for adjusting the flow rate of the oral and / or nasal branches is connected to the oral and / or nasal branches. At least one of the multiple branches is connected to a lung unit exposure device adapted to the corresponding branch shunt ratio or adapted to a target shunt ratio. The lung unit exposure device includes a microporous membrane for cell growth and a gas chamber and a liquid chamber located on both sides of the microporous membrane. The liquid chamber is equipped with a CO2 exhalation device and a fine baffle to refine the CO2 exhaled by the CO2 exhalation device.

[0010] This invention is an improved version. The shunt valves on the oral and nasal branches precisely control the shunt flow rate of inhaled gas / aerosols via the mouth and nose. The lung unit exposure device, adapted to the corresponding branch shunt ratio or target shunt ratio, accurately evaluates the exposure dose of different levels of bronchial tree branches. The refined baffle provides uniform, dense, and fine CO2 to meet the physiological environment of exhaled CO2. This system realistically recreates the actual respiratory physiological microenvironment of cells. Combined with precise oral and nasal shunts and the shunt ratio of the lung unit exposure device, it can accurately assess the cell exposure dose, thereby obtaining more accurate cell exposure experimental results.

[0011] Preferably, the flow diversion valve has a flow diversion ratio of 100% mouth branch, 100% nose branch, or mixed flow diversion, wherein the mixed flow diversion is a flow diversion of mouth branch and nose branch according to a set ratio.

[0012] Preferably, the diverter valve is a rotary diverter valve, a pressure-tube diverter valve, a relative diverter valve, or a pneumatic diverter valve. These diverter valves mainly adjust the airflow distribution ratio of the mouth branch and the nose branch by changing the ratio of the inner diameter cross-sectional area of ​​the airflow paths of the mouth branch and the nose branch, or by adjusting the suction resistance of the airflow paths of the mouth branch and the nose branch.

[0013] Preferably, the branch shunt ratio of the multiple branches of the bronchial tree is determined by measuring the flow rate, and the target shunt ratio deviates from the branch shunt ratio to simulate different signs; a lung unit exposure device with a matching gas chamber volume is selected according to the branch shunt ratio and the target shunt ratio, and the gas chamber volume increases or decreases with respiration.

[0014] Preferably, the lung unit exposure device includes a culture unit, which includes the microporous membrane, a gas chamber, and a liquid chamber. The top wall of the gas chamber is made of a soft elastic material, which expands when inhaled and collapses when exhaled, so as to cause the volume of the gas chamber to change with respiration.

[0015] More preferably, the lung unit exposure device includes a constant temperature water bath, and the culture unit is disposed in the constant temperature water bath. Attached Figure Description

[0016] Figure 1 This is a schematic diagram of the structure of a biomimetic respiratory exposure system for inhaled substances according to an embodiment of the present invention;

[0017] Figure 2 for Figure 1 Schematic diagram of the main components of the central lung unit exposure device;

[0018] Figure 3 This is a schematic diagram of the structure of a biomimetic respiratory exposure system for inhaled substances according to another embodiment of the present invention;

[0019] Among them, 1-Respiratory power device; 2-Oral branch; 3-Nasal branch; 4-Tracheal branch; 5-Lung unit exposure device; 6-CO2 exhalation device; 7-Computer controller; 8-Artificial lip; 9-First solenoid valve; 10-Diverter valve; 11-First flow sensor; 12-Inhaled material generator; 13-Second solenoid valve; 14-Upper pharyngeal tube; 15-Lower pharyngeal tube; 150-Third solenoid valve; 16-Bronchial tree; 17-Left branch unit; 18-Right branch unit; 19-Bionic alveoli; 20-Constant temperature water bath; 21-Culturation unit Unit; 22-Cultivation tank; 23-Sealing cap; 24-Cell chamber; 25-Microporous membrane; 26-Respiratory tubing; 27-First connecting tube; 28-Second flow sensor; 29-CO2 exhalation tubing; 30-Refined baffle; 31-Tank body; 32-Tank cover; 33-Inlet pipe; 34-Outlet pipe; 35-Water bath; 36-CO2 exhalation power unit; 37-Gas storage chamber; 38-CO2 cylinder; 39-Synthetic air cylinder; 40-CO2 outlet pipe; 41-Second connecting tube; 42-Fourth solenoid valve; 43-Fifth solenoid valve. Detailed Implementation

[0020] The technical concept of this utility model is to construct a multi-level, multi-branch terminal bronchial tree, clarify the airflow ratio of different branch terminals, and connect different branch terminals either to alveoli that match the airflow ratio or to a lung unit exposure device that can simulate the air-liquid interface environment of cells. Combined with oral and nasal diversion, it can realistically restore the different exposure environments of cells, accurately measure the exposure dose, and thus conduct exposure evaluation more realistically and accurately.

[0021] Furthermore, by refining the design of the baffles and constant-temperature water bath in the lung unit exposure device, the process of cells in the body being in a constant-temperature environment and gas passing smoothly and evenly through the air-blood barrier (i.e. the gas-liquid interface for cell growth) was fully reproduced, further improving the biomimicry of the breathing process of the device.

[0022] The application of the above-mentioned biomimetic breathing exposure system in biomimetic breathing exposure experiments mainly includes: seeding and culturing cells on the microporous membrane of the lung unit exposure device to construct a cell layer at the gas-liquid interface, and then conducting an exposure experiment in a breathing mode. The exposure dose is determined based on the ratio of oral and nasal branch shunting and the proportion of airflow entering the lung unit exposure device, and the exposure evaluation is performed at the exposure dose.

[0023] When the above-mentioned biomimetic breathing exposure system is applied in biomimetic breathing exposure experiments, the exposure dose is determined by combining the inspiratory volume, the ratio of oral and nasal branch shunting, the airflow ratio of the terminal branches of the bronchial tree connected to the lung unit exposure device, and the intake volume into the lung unit exposure device. For the first time, the exposure dose of cells under biomimetic breathing state has been obtained, creating a basic condition for accurate cell exposure evaluation.

[0024] Preferably, the exposure dose is calculated using the following formula:

[0025] Exposure dose (oral) = Number of inhaled orally inhaled substances (N) 吸入物口 × Diverter valve retention coefficient S 保 × Percentage of airflow entering the exposure chamber V 进装置 / V 吸 ;

[0026] Inhaled substance flow rate: Exposure dose (mL) = Inhaled substance flow rate V 口 / 鼻特殊吸 × Diverter valve retention coefficient S 保 × Percentage of airflow entering the exposure chamber V 进装置 / V 吸 .

[0027] Preferably, the diversion valve retention coefficient S is determined based on the loss of the inhaled material through the diversion valve. 保 The retention coefficient S of the diversion valve 保 The range is 0% to 100%.

[0028] Preferably, the breathing pattern is selected from natural breathing, special breathing containing the inhaled substance to be studied, and a combination of natural breathing and the special breathing.

[0029] The implementation process of this utility model will be described in detail below with reference to specific embodiments.

[0030] The specific embodiments of the biomimetic respiratory exposure system for inhaled substances of this utility model are as follows:

[0031] Example 1

[0032] The biomimetic respiratory exposure system for inhaled substances in this embodiment includes a respiratory power device 1, an oral branch 2, a nasal branch 3, a tracheal branch 4, a lung unit exposure device 5, a CO2 exhalation device 6, and a computer controller 7.

[0033] The respiratory power device 1 provides respiratory power to the oral branch 2, nasal branch 3, and tracheal branch 4, specifically in the form of an electric syringe. The movement of the syringe is controlled by a computer controller 7. The syringe movement includes pulling the piston outward to allow gas / aerosol to enter the syringe and pushing the piston inward to allow gas / aerosol to exit the syringe, thus providing power for the inhalation and exhalation processes. The computer controller 7 can set the changes in the gas / aerosol flow waveform (respiratory wave, bell wave, square wave, sine wave, triangle wave, etc.), the push / pull time, and the push / pull interval. Specifically, the syringe's volume range is 0-1000mL with an accuracy of 0.05-5mL, the frequency is 0-100 times / min, the push / pull time is 0-60s with an accuracy of 0.01s, and the push / pull interval is 0-999min with an accuracy of 0.01s.

[0034] Oral branch 2 primarily simulates the oral inhalation breathing pathway. Figure 1 It includes an oral inhalation tubing and an artificial lip 8 connected to the inlet of the oral inhalation tubing. The oral inhalation tubing is equipped with a first solenoid valve 9, a diverter valve 10, and a first flow sensor 11. A generator 12 for inhaled substances, such as a cigarette, can be connected to the artificial lip 8.

[0035] Nasal branch 3 primarily simulates the nasal inhalation breathing pathway. Figure 1 It includes a nasal inhalation tubing, and a second solenoid valve 13 is installed on the nasal inhalation tubing.

[0036] The downstream branches of oral branch 2 and nasal branch 3 converge to form the pharyngeal structure, which then connects to tracheal branch 4. The pharyngeal structure consists of the upper pharyngeal tube 14 and the lower pharyngeal tube 15. Oral branch 2 and nasal branch 3 are both connected to one end of the upper pharyngeal tube 14, and the other end of the upper pharyngeal tube 14 connects to the respiratory power device 1 and the lower pharyngeal tube 15. Tracheal branch 4 simulates the connection between the lower respiratory tract and the lungs, meaning that gas or aerosols are inhaled through the mouth and nose and enter tracheal branch 4, where gas exchange is completed in the lungs. The oral branch 2, nasal branch 3, tracheal branch 4, upper pharyngeal tube 14, and lower pharyngeal tube 15 completely simulate the upper and lower respiratory tracts of the human body, creating favorable conditions for accurate simulation of in vivo exposure. A third solenoid valve 150 is installed on the lower pharyngeal tube 15.

[0037] The bronchial tree 16 of tracheal branch 4 includes a left branch unit 17 and a right branch unit 18. Figure 1In the bronchial tree 16, both the left branch unit 17 and the right branch unit 18 contain two levels of branches. Thus, after adding the main branches that form the left and right branch units 17 and 18, the bronchial tree 16 forms a three-level branch structure, more closely resembling the hierarchical structure of the bronchial tree 16 in vivo. The bronchial tree 16 can be constructed using computed tomography imaging and 3D printing technology. The left branch unit 17 has 10 terminal branches, and the right branch unit 18 has 8 terminal branches. Of all the terminal branches, one connects to the lung unit exposure device 5, and the remaining 17 connect to the biomimetic alveoli 19. The biomimetic alveoli 19 are spherical and can expand and contract (i.e., inflate during air intake and deflate during air exhaust). The proportion of their expandable volume matches the airflow distribution ratio of each terminal branch of the bronchial tree 16, ensuring consistent air pressure when each lung unit exposure device expands and guaranteeing that the airflow distribution ratio does not change with the proportional expansion of each lung unit.

[0038] The airflow distribution ratio of each final branch in the left branch unit 17 is denoted as Ln, and the airflow distribution ratio of each final branch in the right branch unit 18 is denoted as Rn. The specific airflow distribution ratios of Ln and Rn can be determined by flow measurement. That is, a flow sensor is connected to the end of each branch, and the flow ratio measured by the flow sensor at the end of each branch at different stages is examined under the same total air intake to determine the airflow distribution ratio of different branches. See Table 1 below for details. The distribution ratios for stages 2, 1, and 0 are the sum of those for each branch in the corresponding 3-stage bronchial tree.

[0039] Table 1. Airflow distribution ratio of each terminal branch of the bronchial tree.

[0040]

[0041]

[0042] By combining the aforementioned settings of the biomimetic alveoli 19 with the shunt ratio of the terminal branches of the bronchial tree 16, the increase of the biomimetic alveoli 19 does not cause a change in the shunt ratio of the terminal branches of the bronchial tree 16. This better simulates the transmission, distribution, and shunting of inhaled substances in a healthy human body, further improving the quantification and biomimetic simulation of airflow through the bronchial tree 16. When studying different pathological signs (such as right lung atelectasis), the shunt ratio of the terminal branches of the bronchial tree 16 can also be adjusted by using the biomimetic alveoli 19. This ensures that the increase of the biomimetic alveoli 19 causes a change in the shunt ratio of the terminal branches of the bronchial tree 16, better simulating the transmission, distribution, and shunting of inhaled substances in a human body with specific pathological signs, further improving the quantification and biomimetic simulation of airflow through the bronchial tree 16.

[0043] The lung unit exposure device 5 simulates the gas exchange process carried out by alveolar cells during respiration. It typically carries epithelial cells and can simulate the gas-liquid interface environment of the epithelial cells and the expansion and contraction of the alveoli themselves during respiration. Specifically, such as... Figure 2 As shown, the lung unit exposure device 5 includes a constant temperature water bath 20 and a culture unit 21 installed in the constant temperature water bath 20. The culture unit 21 includes a culture tank 22 and a sealing cover 23 for sealing the culture tank 22. Cell chambers 24 are disposed within the culture tank 22; the cell chambers 24 can be commercially available Transwell chambers, and the bottom wall of the cell chambers 24 is a microporous membrane 25 for cell growth. The culture tank 22 and the sealing cover 23 form a closed inner cavity of the culture unit. The lower side of the microporous membrane 25 is in contact with the culture medium, and the upper side is in contact with air. The cells growing on the microporous membrane 25 are in a gas-liquid interface environment. The sealing cover 23 is a hemispherical shell made of an elastic material, allowing the inner cavity of the culture unit to expand and contract. A breathing tube 26 is disposed on the sealing cover 23, and the breathing tube 26 is connected to the terminal branch of the bronchial tree 16 via a first connecting tube 27. A second flow sensor 28 is disposed on the first connecting tube 27.

[0044] A CO2 exhalation conduit 29 is connected to the liquid side of the microporous membrane 25 in the culture tank 22. The CO2 exhalation conduit 29 is connected to the CO2 exhalation device 6 to exhale a certain concentration of CO2 from the liquid side onto the cell layer on the microporous membrane 25, simulating a high concentration of exhaled CO2. A refining baffle 30 is also provided on the liquid side of the microporous membrane 25 in the culture tank 22. Both the refining baffle 30 and the microporous membrane 25 are arranged horizontally, with the microporous membrane 25 located within the surface area of ​​the refining baffle 30. The refining baffle 30 refines the CO2 exhaled from the CO2 exhalation conduit 29, generating uniform, dense, and fine CO2 bubbles to simulate the gas exchange process of the blood-air barrier and prevent larger CO2 clouds directly exhaled from the CO2 exhalation conduit 29 from damaging the cells or disrupting the gas-liquid interface environment of the cells.

[0045] The constant temperature water bath 20 is used to simulate the constant temperature environment of the human body. It includes an open tank body 31 and a tank cover 32 that seals with the tank body 31. The tank body 31 is connected to a water inlet pipe 33 and a water outlet pipe 34. The water inlet pipe 33 and the water outlet pipe 34 are connected to a water bath machine 35 to provide water at a constant temperature of 37°C. In addition, the sealing cover 23 of the culture unit 21 is set on the tank cover 32, and the sealing cover 23 of the culture unit 21 and the culture tank 22 are provided with matching sealing structures (such as sealing grooves and sealing rings). When the tank body 31 and the tank cover 32 are closed and sealed, the sealing cover 23 of the culture unit 21 and the culture tank 22 are also sealed simultaneously.

[0046] The CO2 exhalation device 6 generates a certain concentration of CO2 and exhales it through the CO2 exhalation tubing 29. The CO2 exhalation device 6 includes a CO2 exhalation power unit 36, a gas storage chamber 37, a CO2 cylinder 38, and a synthetic air cylinder 39. The CO2 cylinder 38 and synthetic air cylinder 39 are connected to the gas storage chamber 37. A CO2 outlet pipe 40 is connected to the gas storage chamber 37 and is connected to the CO2 exhalation power unit 36. The CO2 exhalation power unit 36 ​​is connected to the CO2 exhalation tubing 29 via a second connecting pipe 41. The CO2 exhalation power unit 36 ​​uses an electric syringe, the same type used in the breathing power unit 1. The CO2 in the gas storage chamber 37 that meets the requirements enters the electric syringe through the CO2 outlet pipe 40, and then, by pushing the piston of the electric syringe, the CO2 is pushed into the CO2 exhalation tubing 29.

[0047] The CO2 exhalation device 6 is connected to the computer controller 7, on which the concentration of CO2 generated can be set. The CO2 cylinder 38 and the synthetic air cylinder 39 can supply the corresponding gas into the gas storage chamber 37 according to computer instructions. A gas sensor on the gas storage chamber 37 provides feedback on the CO2 concentration. When the CO2 concentration in the gas storage chamber 37 reaches the set concentration, the gas intake stops. A fourth solenoid valve 42 and a fifth solenoid valve 43 are respectively installed on the CO2 outlet pipe 40 and the second connecting pipe 41, which can be used in conjunction with the CO2 exhalation power device 36 to match the required breathing rhythm.

[0048] In other implementations, both CO2 cylinder 38 and synthetic air cylinder 39 can be standard gas cylinders with specific CO2 and O2 concentrations, connected to the gas storage chamber 37. The gas storage chamber is in a vacuum state. By setting the CO2 concentration (consistent with the CO2 concentration in the standard gas cylinders) and pressure, gas from these two standard gas cylinders enters the gas storage chamber 37. When the CO2 concentration and pressure reach the desired levels, the gas storage chamber 37 contains CO2 and O2 at the same concentration as the standard gas. Then, according to a set exhalation rhythm, a certain concentration of CO2 and O2 is exhaled into the exposure unit.

[0049] The first, second, third, fourth, and fifth solenoid valves can be pressure-operated valves, and their opening and closing are controlled by the computer controller 7. The first and second flow sensors can be in-line flow sensors, with both ends connected to the pipeline under test to obtain the flow rate through the pipeline, facilitating the calculation of the exposure dose. The first flow sensor 11 is used to obtain the gas flow rate V entering through the mouth. 口 The second flow sensor 28 is used to acquire the inhaled gas flow rate V entering the lung unit exposure device 5. 肺 The sensing signals from the first and second flow sensors are received by the computer controller 7, which can display the flow waveform curve and obtain the flow value on the computer.

[0050] Example 2

[0051] The biomimetic respiratory exposure system for inhaled substances in this embodiment, such as Figure 3 As shown, it includes an oral branch 2, a nasal branch 3, a tracheal branch 4, a respiratory power device 1, a lung unit exposure device 5, and a CO2 exhalation device 6. The implementation of the tracheal branch 4, the respiratory power device 1, the lung unit exposure device 5, and the CO2 exhalation device 6 is the same as in Example 1. The implementation of the oral branch 2 and the nasal branch 3 will be described below.

[0052] Figure 3 In the middle section, the oral branch 2 includes an oral inhalation tubing and an artificial lip 8. A first solenoid valve 9 and a first flow sensor 11 are installed on the oral inhalation tubing. A generator 12 for the inhaled substance, such as a cigarette, can be connected to the artificial lip 8. The nasal branch 3 includes a nasal inhalation tubing, with a second solenoid valve 13 and a diverter valve 10 installed on it. The diverter valve 10 is a fixed-flow diverter valve.

[0053] The above-mentioned diversion valve 10 is mainly set to adjust the diversion ratio of the oral branch 2 and the nasal branch 3. Under the condition of a certain inhalation volume (for example, the single inhalation volume of the breathing power device 1 is controlled to be 500mL), the mouth or nose can be adjusted to be the main inhalation, thereby simulating different inhalation states.

[0054] The diverter valve 10 can be a non-fixed flow diverter valve or a fixed flow diverter valve. In different implementation scenarios, it can be installed on the mouth branch 2 and the nose branch 3 to achieve inhalation and exhalation control under different diversion conditions via the mouth, nose, and mouth + nose. The diverter valve 10 controls the ratio of airflow through the mouth and nose tubes by controlling the cross-sectional area ratio of the mouth and nose tubes and / or by using the complexity of the internal path to cause changes in suction resistance.

[0055] When the gas or aerosol diverted through the mouth or nose is unlikely to clog the diversion line, the diversion valve 10 can be installed on the mouth or nose branch line. When the gas or aerosol diverted through the mouth or nose is likely to clog the diversion line, the diversion valve 10 must be installed on the branch line where the easily clogged gas or aerosol is not inhaled. When all three mouth and nose branch lines divert gases or aerosols that are likely to clog the diversion line, a fixed flow diversion valve should be used on both mouth and nose branch lines.

[0056] Furthermore, when the cigarette smoke is diverted via mouth and nose, the cigarette is connected to the mouth branch 2 pipe, and the nose branch 3 pipe is connected to the atmosphere. When a non-constant flow diversion valve is connected to the mouth branch 2 pipe, the retention coefficient S of different types of non-constant flow diversion valves... 保 The results are shown in Table 2 below. A fixed flow diverter valve can also be used. Since the internal structure of the fixed flow diverter valve is a straight-through pipe, or when it is not connected to a gas / aerosol that is prone to clogging, the loss can be ignored, and the retention coefficient S... 保 It is calculated as 100%.

[0057] Table 2 Retention coefficient S for different non-constant flow split valves 保

[0058] Non-constant flow diverter valve <![CDATA[Retention coefficient S 保 (%)]]> Complex paths within metal scale 15.04 Metal Rotation 32.27 Plastic Rotation 81.36 Plastic crimp tube 54.61

[0059] The shunt valve can be selected based on the required oral / nasal shunt and aerosol blockage. The lung unit exposure device 5 and bronchial tree 16 are selected based on the number of exposure units connected and the bronchial tree 16 exposure level to be studied. The selection and matching of the lung unit exposure device and the bronchial tree branch levels generally follows these principles: When the distribution ratio at the end of bronchial tree 16 does not change, each branch end should be connected to an exposure device and / or a biomimetic lung unit with the same gas distribution ratio as the bronchial tree; when the distribution ratio at the end of bronchial tree 16 changes, each branch end should be connected to an exposure device and / or a biomimetic lung unit with the target gas distribution ratio; when the airflow entering the exposure device does not cause cell damage, the gas chamber volume of the exposure device should be less than 1.6 times the airflow volume; when investigating cell damage caused by the airflow entering the exposure device, the gas chamber volume of the exposure device is selected based on the cell survival rate induced by preliminary experiments.

[0060] The biomimetic respiratory exposure system described in the above embodiments can be used to conduct exposure experiments under different respiratory modes and with different vital signs. Cells and culture media can be collected from the exposure device, or the cells in the chamber can be further incubated or processed, depending on the purpose, thereby providing an exposure evaluation. The following describes typical respiratory modes based on the above-described biomimetic respiratory exposure system:

[0061] (1) One breathing pattern

[0062] a. Gas / aerosol enters the respiratory power unit 1 through oral branch 2 and / or nasal branch 3: Set the respiratory waveform and inhalation flow rate V. 吸 mL, inhalation time S 吸 s, select the airflow distribution coefficient for the oral cavity and nasal cavity (e.g., oral V) 口吸 mL, via nasal V 鼻吸 For the diverter valve corresponding to mL), when the oral and nasal flow rates are greater than 0, the oral and nasal connecting solenoid valves (first and second solenoid valves) are opened during this stage, the breathing power device 1 is in intake mode, and the next stage waiting time S is entered. a等 s.

[0063] b1 and b2 are synchronized.

[0064] b1. Gas / aerosol enters the bronchial tree 16 via the respiratory power device 1 and then enters the lung unit exposure device 5: Set the respiratory waveform and push the flow rate V. 推 mL, push time S 推At this stage, the solenoid valve (third solenoid valve) of the tubing connected to the bronchial tree 16 lung unit exposure device 5 opens, the syringe is in push-air mode, and the flow rate into the lung unit exposure device 5 is monitored by the flow sensor and the computer controller 7. The inflow rate is V. 进装置 mL, proceed to the next stage waiting time S b1等 s.

[0065] b2. CO2 Intake and Exhalation Power Device 36: Sets the CO2 concentration in the gas storage chamber 37, sets the breathing waveform, and sets the CO2 inflow rate V. CO2进 mL, entry time S CO2进 During this stage, the passage connecting the gas storage chamber 37 to the CO2 exhalation power device 36 is opened, the syringe is in air intake mode, and the process proceeds to the next stage, waiting time S. b2等 s.

[0066] c1 and c2 are synchronized.

[0067] c1. CO2 enters the lung unit exposure device 5: Set the respiratory waveform, exhaled CO2 flow rate V CO2呼 mL, exhalation time S CO2呼 During this stage, the passage connecting the CO2 exhalation power device 36 to the CO2 exhalation tubing 29 is opened, the syringe is in push mode, and the process proceeds to the next stage, waiting time S. c1等 s.

[0068] c2. Lung unit exposure device 5 gas / aerosol is exhaled through bronchial tree 16 into respiratory power device 1: set respiratory waveform, total exhaled flow rate V of bronchial tree 16. 支呼 mL, the exhaled flow rate from the exposure device is V 装置呼 mL, exhalation time S 装置呼 s, the lung unit exposure device 5 is connected to the exhalation tubing of the bronchial tree 16, the tubing between the bronchial tree 16 and the respiratory power device 1 is opened at this stage, the syringe is in air intake mode, and the next stage waiting time S is entered. c2等 s.

[0069] d. Gas / aerosol in respiratory power device 1 is exhaled through the mouth and / or nose: set the respiratory waveform, exhalation flow rate V 呼出 mL, exhalation time S 呼 s, select the airflow distribution coefficient for the oral cavity and nasal cavity (e.g., oral V) 口吸 mL, via nasal V 鼻吸 When the flow rate via the mouth or nose is greater than 0, the solenoid valve connecting the mouth and nose is set to open during this stage, the syringe is in air-propelling mode, and the process proceeds to the next set of actions after a waiting time of 5 seconds. 组1等循 s.

[0070] Among them, V 吸 mL = V 口特殊吸 mL+V口吸 mL+V 鼻吸 mL≥V 推 mL≥V 进装置 mL; S 推 s≥S CO2进 s;S b1等 s+S 推 s = S CO2进 s+S b2等 s;V CO2进 mL≥V CO2呼 mL; S CO2呼 =S c1等 S c1等 =S c2等 V 装置呼 mL = V CO2呼 mL+V 进装置 mL;V 支呼 mL≥V 呼出 mL≥V 吸 mL+V CO2呼 mL.

[0071] (2) Special inhalation breathing pattern via oral route (special inhalation such as cigarette smoke inhalation)

[0072] a. Gas / aerosol enters the respiratory power unit 1 via oral branch 2 and / or nasal branch 3: Breathing waveform and inhalation flow rate V are set. 吸 mL, inhalation time S 吸 s, select the airflow distribution coefficient for the oral cavity and nasal cavity (e.g., oral V) 口特殊吸 mL, V 口吸 mL, via nasal V 鼻吸 For the corresponding flow divider valve (mL), when the oral and nasal flow rates are greater than 0, the oral and nasal connecting solenoid valve is opened during this stage, the breathing power unit 1 is in intake mode, and the next stage waiting time S is entered. a等 s.

[0073] b1 and b2 are synchronized.

[0074] The rest are the same as (1) group of breathing patterns b1, b2, c1, c2, d.

[0075] Among them, V 吸 mL = V 口特殊吸 mL+V 口吸 mL+V 鼻吸 mL≥V 推 mL≥V 进装置 mL; S 推 s≥S CO2进 s;S b1等 s+S 推 s = S CO2进 s+S b2等 s;VCO2进 mL≥V CO2呼 mL; S CO2呼 s = S c1等 s;S c1等 s = S c2等 s;V 装置呼 mL = V CO2呼 mL+V 进装置 mL;V 支呼 mL≥V 呼出 mL≥V 吸 mL+V CO2呼 mL.

[0076] (3) Special nasal inhalation breathing mode (special inhalation such as drug inhalation)

[0077] a. Gas / aerosol enters the respiratory power unit 1 via oral branch 2 and / or nasal branch 3: Breathing waveform and inhalation flow rate V are set. 吸 mL, inhalation time S 吸 s, select the airflow distribution coefficient for the oral cavity and nasal cavity (e.g., oral V) 口吸 mL, via nasal V 鼻特殊吸 mL, V 鼻吸 For the corresponding flow divider valve (mL), when the oral and nasal flow rates are greater than 0, the oral and nasal connecting solenoid valve is opened during this stage, the breathing power unit 1 is in intake mode, and the next stage waiting time S is entered. a等 s.

[0078] b1 and b2 are synchronized.

[0079] The rest are the same as (1) group of breathing patterns b1, b2, c1, c2, d.

[0080] Among them, V 吸 mL = V 鼻特殊吸 mL+V 口吸 mL+V 鼻吸 mL≥V 推 mL≥V 进装置 mL; S 推 s≥S CO2进 s;S b1等 s+S 推 s = S CO2进 s+S b2等 s;V CO2进 mL≥V CO2呼 mL; S CO2呼 s = S c1等 s;S c1等 s = S c2等 s;V 装置呼 mL = V CO2呼 mL+V 进装置 mL;V 支呼 mL≥V呼出 mL≥V 吸 mL+V CO2呼 mL.

[0081] (4) Special inhalation breathing modes via mouth and nose (special inhalation such as drug inhalation)

[0082] a. Gas / aerosol enters the respiratory power unit 1 via oral branch 2 and / or nasal branch 3: Breathing waveform and inhalation flow rate V are set. 吸 mL, inhalation time S 吸 s, select the airflow distribution coefficient for the oral cavity and nasal cavity (e.g., oral V) 口特殊吸 mL, V 口吸 mL, via nasal V 鼻特殊吸 mL, V 鼻吸 For the corresponding flow divider valve (mL), when the oral and nasal flow rates are greater than 0, the oral and nasal connecting solenoid valve is opened during this stage, the breathing power unit 1 is in intake mode, and the next stage waiting time S is entered. a等 s.

[0083] b1 and b2 are synchronized.

[0084] The rest are the same as (1) group of breathing patterns b1, b2, c1, c2, d.

[0085] Among them, V 吸 mL = V 口特殊吸 mL+V 鼻特殊吸 mL+V 口吸 mL+V 鼻吸 mL≥V 推 mL≥V 进装置 mL; S 推 s≥S CO2进 s;S b1等 s+S 推 s = S CO2进 s+S b2等 s;V CO2进 mL≥V CO2呼 mL; S CO2呼 s = S c1等 s;S c1等 s = S c2等 s;V 装置呼 mL = V CO2呼 mL+V 进装置 mL;V 支呼 mL≥V 呼出 mL≥V 吸 mL+V CO2呼 mL.

[0086] (5) Multiple sets of breathing or special inhalation actions in sequence (special inhalation such as cigarette smoke inhalation)

[0087] After configuring the specific parameters of the breathing or special inhalation actions involved, match them according to the time intervals of special inhalation and breathing, and execute multiple sets in sequence according to the number of special inhalations and breathing. These sets are then set in series. For example, a single smoking action combined with multiple natural breathing actions can be combined into one set, and multiple sets can be executed continuously to simulate the overall exposure process during human smoking.

[0088] The exposure dose for the above exposure experiments can be calculated as follows:

[0089] (1) Timing based on the number of inhaled substances: Exposure dose (oral) = Number of inhaled substances (N) 吸入物口 ) × Diverter valve retention coefficient (S) 保 ) × Percentage of airflow entering the exposure chamber (V) 进装置 / V 吸 )=Number of inhalation ports (N 吸入物口 ) × Diverter valve retention coefficient (S) 保 ) × Percentage of airflow entering the exposure chamber (V) 吸 ×R n or V 吸 ×L n ).

[0090] (2) The inhaled substance is measured by flow rate: Exposure dose (mL) = Inhaled substance flow rate (V) 口 / 鼻特殊吸 ) × Diverter valve retention coefficient (S) 保 ) × Percentage of airflow entering the exposure chamber (V) 进装置 / V 吸 ) = number of inhalation ports (V 口 / 鼻特殊吸 ) × Diverter valve retention coefficient (S) 保 ) × Percentage of airflow entering the exposure chamber (V) 吸 ×R n or V 吸 ×L n ).

[0091] (3) The exposure dose (mg) is calculated based on the mass of the inhaled substance: Exposure dose (mg) = mass per mouthful (mg) 每口 ) × Exposure dose (or) = Mass per milliliter (M) 每毫升 ) × exposure dose (mL).

[0092] The application of the biomimetic breathing exposure system based on Embodiment 2 above in biomimetic breathing exposure experiments is described in detail below:

[0093] 1) Mode Settings

[0094] Select the special oral inhalation mode. Based on the special inhalation method of 35mL of smoke being inhaled orally and the possibility of smoke blockage in the tubing, connect the nasal branch 3 tubing to the diversion valve 10 to adjust the oral and nasal flow. Set the following mode parameters based on the breathing parameters (500mL inhalation, 520mL exhalation, 1s breathing wait, 2s inhalation and 2s exhalation times) and the smoking parameters (0, 84, and 168 puffs respectively, 35mL inhalation volume, 2s inhalation time, 55s inhalation interval):

[0095] (1) One breathing pattern

[0096] a. Gas / aerosol enters the respiratory power unit 1 via the mouth and / or nose: Breathing waveform is set, inhalation flow rate V. 吸 mL (500 mL), inhalation time S 吸 s(2s), select the airflow distribution coefficients for the oral cavity and nasal cavity (e.g., oral V 口吸 0mL, via nasal V 鼻吸 (500mL), when the flow rate via the mouth and nose is greater than 0, the solenoid valve connecting the mouth and nose is opened during this stage, the syringe is in air intake mode, and the process proceeds to the next stage waiting time S. a等 s(1s).

[0097] b1 and b2 are synchronized.

[0098] b1. Gas / aerosol enters the bronchial tree 16 via the respiratory power device 1 and then enters the lung unit exposure device 5: Set the respiratory waveform and push the flow rate V. 推 mL (500mL), push time S 推 At s(2s), the solenoid valve of the tubing connecting the bronchial tree 16 to the lung unit exposure device 5 opens during this stage, the syringe is in push-air mode, and the flow rate into the lung unit exposure device 5 is V. 进装置 mL (20 mL), proceed to the next stage waiting time S b1等 s(1s).

[0099] b2. CO2 enters the CO2 exhalation power device 36: The CO2 concentration (4%) in the gas storage chamber 37 is pre-set, and during this stage, CO2 of the set concentration is mixed in the gas storage chamber 37. The breathing waveform is set, and the CO2 flow rate V... CO2进 mL (20 mL), entry time S CO2进 During the s(2s) phase, the passage connecting the gas storage chamber 37 to the CO2 exhalation power device 36 is opened, the syringe is in air intake mode, and the process proceeds to the next stage, waiting time S. b2等 s(1s).

[0100] c1 and c2 are synchronized.

[0101] c1. CO2 enters the lung unit exposure device 5: Set the respiratory waveform, exhaled CO2 flow rate V CO2呼 mL (20 mL), exhalation time S CO2呼 During the s(2s) phase, the passage connecting the CO2 exhalation power device 36 to the CO2 exhalation tubing 29 is opened, the syringe is in push mode, and the process proceeds to the next phase, waiting time S. c1等 s(1s).

[0102] c2. Lung unit exposure device 5: Gas / aerosol exhaled through bronchial tree 16 into the syringe: Set respiratory waveform, bronchial tree exhalation flow rate V 支呼 mL (520 mL), the exhaled flow rate from the exposure device is V 装置呼 mL (40 mL), exhalation time S 装置呼 s(2s), the lung unit exposure device 5 connects to the exhalation tubing of the bronchial tree 16. The tubing connecting the bronchial tree 16 and the lung unit exposure device 5 is opened during this stage, the syringe is in air intake mode, and the next stage waiting time S is entered. c2等 s(1s).

[0103] d. Gas / aerosol in respiratory power device 1 is exhaled through the mouth and / or nose: set the respiratory waveform, exhalation flow rate V 呼出 mL (520 mL), exhalation time S 呼 s(2s), select the airflow distribution coefficients for the oral cavity and nasal cavity (e.g., oral V 口吸 mL, via nasal V 鼻吸 When the flow rate via the mouth or nose is greater than 0, the solenoid valve connecting the mouth and nose is set to open during this stage, the syringe is in air-propelling mode, and the process proceeds to the next set of actions after a waiting time of 5 seconds. 组1等循 s(1s).

[0104] (2) Breathing pattern of a single smoke

[0105] a. Flue gas enters through inhalation branch 2 pipe, and air enters breathing power unit 1 through nasal branch 3 pipe: Breathing waveform is set, inhalation flow rate V... 吸 mL (500 mL), inhalation time S 吸 s(2s), select the airflow distribution coefficients for the oral cavity and nasal cavity (e.g., oral V 吸烟 mL (35 mL), V 口吸 0mL, via nasal V 鼻吸 (465mL), when the flow rate via the mouth and nose is greater than 0, the solenoid valve connecting the mouth and nose is opened during this stage, the syringe is in air intake mode, and the next stage waiting time S is initiated. a等 s(1s).

[0106] b1 and b2 are synchronized.

[0107] The rest are the same as (1) group of breathing patterns b1, b2, c1, c2, d.

[0108] (3) A series of natural breathing or smoking breathing movements in sequence

[0109] One cycle consists of 10 natural breathing movements followed by one special smoking action. When exposed to 0 puffs of smoke, an unlit cigarette is inserted into the mouth, and the cycle is repeated 168 times; when exposed to 84 puffs of smoke, a lit cigarette is inserted into the mouth, and the cycle is repeated 84 times, followed by an unlit cigarette; when exposed to 168 puffs of smoke, a lit cigarette is inserted into the mouth, and the cycle is repeated 168 times.

[0110] (2) Device connection: according to Figure 2 Connect the devices and prepare the cell chamber 24 to be exposed. Connect the inhaled material generator 12 (cigarette) to the oral branch 2 and the fixed flow proportional valve to the nasal branch 3. The total inhalation volume from the oral branch 2 and nasal branch 3 is 500 mL, and the oral shunt flow rate is 35 mL. Select the third-order bronchial tree 16 and connect it to one lung unit exposure device 5. Connect the rest to the biomimetic alveoli 19. The cell chamber 24 in the lung unit exposure device 5 is a 6-well Transwell chamber.

[0111] (3) Exposure experiment and sample collection: The exposure experiment was carried out according to the above settings. After the exposure experiment was completed, the cells in the lung unit exposure device 5 were collected and incubated for 24 hours before the CCK-8 experiment was performed.

[0112] (4) Exposure assessment

[0113] Exposure dose is calculated per mouthful:

[0114] Exposure dose (oral) = Number of inhaled substances (N) 吸入物口 ) × Diverter valve retention coefficient (S) 保 ) × Percentage of airflow entering the exposure chamber (V) 进装置 / V 吸 )=Number of inhalation ports (N 吸入物口 ) × Diverter valve retention coefficient (100) × Airflow ratio entering the exposure chamber (20mL / 500mL) = 0.04 × Number of inhaled substances inhalation ports (N) 吸入物口 If the exposure dose of the device is 0 puffs, 3.36 puffs, and 6.72 puffs for 0 puffs, 84 puffs, and 168 puffs, respectively, then the exposure dose of the device is 0 puffs, 3.36 puffs, and 6.72 puffs for 0 puffs, 84 puffs, and 168 puffs, respectively.

[0115] The survival rates of Beas-2b cells under different exposure doses are shown in Table 3 below.

[0116] Table 3. Beas-2b cell survival rate under different exposure doses.

[0117] Number of exposed flue gas outlets (outlets) Cell viability (%) 0 100 3.36 92±3.1％ 6.72 77±6.8％

[0118] As shown in Table 3, the survival rate of Beas-2b cells decreased with the continuous increase of the number of cigarettes smoked (puffs), which is consistent with the previous research results.

[0119] As can be seen from the above description, the biomimetic respiratory exposure system for inhaled substances of this invention effectively simulates the respiratory physiological microenvironment of cells and scientifically characterizes the shunting of oral and nasal passages as well as the shunting of the third-order bronchial tree 16, thereby ensuring the scientific rigor and accuracy of inhaled substance-based exposure studies.

Claims

1. A biomimetic respiratory exposure system for inhaled substances, comprising a respiratory power device and oral and nasal branches connected to the respiratory power device, wherein the downstream branches of the oral and nasal branches merge to form a pharyngeal structure and then connect to a tracheal branch, wherein the tracheal branch includes a bronchial tree with multiple branches, characterized in that, The oral and / or nasal branches are connected to shunt valves for adjusting the flow rate of the oral and nasal branches; at least one of the multiple branches is connected to a lung unit exposure device adapted to the corresponding branch shunt ratio or adapted to the target shunt ratio; the lung unit exposure device includes a microporous membrane for cell growth and a gas chamber and a liquid chamber located on both sides of the microporous membrane, respectively; the liquid chamber is equipped with a CO2 exhalation device and a fine baffle for refining the CO2 exhaled by the CO2 exhalation device.

2. The biomimetic respiratory exposure system for inhaled substances as described in claim 1, characterized in that, The flow divider valve can be configured to divide the flow into 100% orifice branch, 100% nasal branch, or a mixed flow divider, wherein the mixed flow divider is configured to divide the flow into orifice branch and nasal branch in a set ratio.

3. The biomimetic respiratory exposure system for inhaled substances as described in claim 2, characterized in that, The diverter valve is a rotary diverter valve, a pressure pipe diverter valve, a relative diverter valve, or a pneumatic diverter valve.

4. The biomimetic respiratory exposure system for inhaled substances as described in claim 1, characterized in that, The bronchial tree's multiple branches are shunted by measuring flow rate, and the target shunting ratio deviates from the branch shunting ratio to simulate different signs; a lung unit exposure device with a matching gas chamber volume is selected based on the branch shunting ratio and the target shunting ratio, and the gas chamber volume increases or decreases with respiration.

5. The biomimetic respiratory exposure system for inhaled substances as described in claim 1, characterized in that, The lung unit exposure device includes a culture unit, which includes the microporous membrane, a gas chamber, and a liquid chamber. The top wall of the gas chamber is made of a soft elastic material, which expands when inhaled and collapses when exhaled, so as to cause the volume of the gas chamber to change with respiration.

6. The biomimetic respiratory exposure system for inhaled substances as described in claim 5, characterized in that, The lung unit exposure device includes a constant temperature water bath, and the culture unit is disposed in the constant temperature water bath.

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