WO2025019942A1 - Vasculature-on-a-chip systems for mimicking and monitoring gas embolism - Google Patents

Abstract

A system for determining the presence of a gas bubble in a blood sample. The system has a microfluidic chip comprising a plurality of interconnected microfluidic channels arranged in a vascular-like configuration to mimic a given part of a vascular system, with a first inlet for receiving the blood sample and a second inlet for injecting the gas into a flow of the fluid to create the gas bubble in the microfluidic channels. The system has a circulation system comprising a pump, a reservoir containing the blood sample and in communication with the pump, and a conduit in communication with the pump and/or the reservoir, the conduit directing the blood sample to the first inlet. The system also has an imaging system configured to capture images of the microfluidic channels of the microfluidic chip, and a detection system configured to determine the presence and location of the gas bubble.

Description

VASCULATURE-ON-A-CHIP SYSTEMS FOR MIMICKING AND MONITORING GAS EMBOLISM

CROSS-REFERENCE TO A RELATED APPLICATION

[0001] This disclosure claims priority from U.S. Patent Application No. 63/528,418 filed on July 23, 2023 and U.S. Patent Application No. 63/673,026 filed on July 18, 2024, the entire content of each of which is incorporated herein by reference.

TECHNICAL FIELD

[0002] The present disclosure relates to the field of in vitro models of emboli (presence of air or an air bubble) in a vasculature-like structure.

BACKGROUND OF THE ART

[0003] Gas embolism may occur when gas bubbles are present in the circulation, resulting in blocked blood flow and blocked oxygen transfer to critical organs, e.g., the brain. While generally considered rare, the presence of gas emboli, which can result from sudden decompression or many invasive medical procedures, often has devastating outcomes, with high mortality (up to 33%) and severe neurological sequelae (up to 35%). Gas emboli in the vasculature are characterized by: (i) unpredictable occurrence due to the poor understanding of their formation at the microscopic level, (ii) relative rarity or underreporting, (iii) variable medical causes and responses, and (iv) high incidence of patient mortality and morbidity, making traditional experimental research difficult, and nearly impossible to conduct ethical clinical trials. The formation of gas emboli is caused initially by a physical process which introduces gas in the bloodstream, the gas bubble takes shape with a slower physico-biochemical processes during which a membrane (made of bloodstream components) forms around the gas producing the gas bubble. The gas bubble then blocks the bloodstream and can cause O2 depletion, and cell tissue or organ death. A following cascade of deleterious effects could also include, in the short term, the damage to vascular endothelium and blood components due to the friction at the gas-tissue interface. [0004] There are many conditions associated with the presence of air in the vasculature, such as decompression sickness (DCS). DCS is a type of decompression illness that arises from the sudden decrease in pressure around the body, leading to the formation of gas bubbles in the bloodstream and tissues. These bubbles typically form via heterogeneous nucleation or diffusion from supersaturated tissues overburdened with gas content. Gas embolism can also occur during surgeries, and may result from stagnant gas bubbles obstructing microvascular blood flow. Despite the serious implications of such conditions, the mechanisms of bubble formation and its dynamic halting behavior in microvasculature remain poorly understood.

[0005] Presently, the causal links between macroscopic conditions, especially iatrogenic (i.e., medical), and the occurrence of a gas embolism are obscure, making the gathering of physical fact-based evidence difficult. Facts-based evidence of gas embolism formation is indeed extremely difficult, and often impossible, to obtain because:

(i) the onset of gas embolism is very rapid, within seconds to minutes, wherein arterial embolism can be fatal in 2 minutes if blockage to the brain occurs;

(ii) gas embolism can occur deep inside the human body, making it difficult to observe and monitor its onset without access to very specialized facilities, e.g., magnetic resonance imaging (MRI), computed tomography (CT) scan, ultrasound, etc.; and/or

(iii) gas embolism often occurs in the field, or in non-specialized medical facilities.

[0006] Presently medical decisions regarding the treatment of gas embolism with hyperbaric oxygen therapy (HBOT) are typically based on neurological assessments of the patients. Improvements are desired for the treatment of medical conditions arising from gas emboli. Therefore, it would be desirable to have models that represent DCS and/or gas emboli more accurately, which would allow treatment options to be tested.

SUMMARY

[0007] In one aspect, there is provided a system for determining the presence of a gas bubble in a blood sample, the system comprising: [0008] a microfluidic chip comprising a plurality of interconnected microfluidic channels arranged in a vascular-like configuration to mimic a given part of a vascular system, the microfluidic chip comprising a first inlet for receiving the blood sample and a second inlet for injecting the gas into a flow of the fluid to create the gas bubble in the flow within the microfluidic channels;

[0009] a circulation system comprising a pump, a reservoir containing the blood sample and in communication with the pump, and a conduit in communication with the pump and/or the reservoir, the conduit directing the blood sample to the first inlet; and

[0010] an imaging system configured to capture images of the microfluidic channels of the microfluidic chip, and a detection system configured to determine the presence and location of the gas bubble based on the captured images of the imaging system.

[0011] In some embodiments, interconnected microfluidic channels form a plurality of bifurcations.

[0012] In some embodiments, the microfluidic chip further comprising a third inlet for providing a drug.

[0013] In some embodiments, the system further comprises a housing to seal the microfluidic chip and create a pressurized environment for the microfluidic chip.

[0014] In some embodiments, the housing is a micro-hyperbaric chamber.

[0015] In a further aspect, there is provided a method of determining the effectiveness of a treatment agent to reduce the volume of a gas bubble in a vascular system, the method comprising:

[0016] flowing a blood sample in a microfluidic chip comprising a plurality of microfluidic channels interconnected and arranged in a vascular-like configuration to mimic a given part of the vascular system, the microfluidic chip comprising a first inlet for receiving the blood sample, a second inlet for injecting gas, a third inlet for inject the treatment agent, and an outlet; [0017] injecting the gas in the second inlet to create the gas bubble in the blood sample within microfluidic channels;

[0018] injecting the treatment agent in the third inlet; and

[0019] imaging the microfluidic chip; and

[0020] based on the image, determining one or more of: a location of the gas bubble in the microfluidic channels; a volume of the gas bubble; and a change in volume of the gas bubble.

[0021] In some embodiments, the blood sample is selected from patient blood or artificial blood.

[0022] In yet a further aspect, there is provided a method of determining a parameter at which the formation of a gas bubble occurs in a vascular system, the method comprising:

[0023] flowing a blood sample in a microfluidic chip comprising a plurality of microfluidic channels interconnected and arranged in a vascular-like configuration to mimic a given part of the vascular system; and

[0024] subjecting the microfluidic chip to a pressure variation;

[0025] imaging the microfluidic chip; and

[0026] based on the imaging, determining the parameter at which formation of a gas bubble occurs in the blood sample within the microfluidic channels.

[0027] In some embodiments, the pressure variation comprises subjecting the microfluidic chip to an increase in pressure followed by a decrease in pressure.

[0028] In some embodiments, the pressure variation is applied by placing the microfluidic chip in an environment that increases the pressure and then decreases the pressure. [0029] In some embodiments, the pressure variation is performed by compressing the microfluidic chip.

[0030] In some embodiments, the parameter is one of or a combination of time and pressure.

[0031] In another aspect, there is provided a method of performing hyperbaric oxygen therapy for a patient in need thereof having a gas bubble in their bloodstream, the method comprising:

[0032] subjecting the patient to a given program of oxygen and pressure in a hyperbaric chamber;

[0033] concurrently with the subjecting, flowing a blood of the patient in a microfluidic chip placed inside a second hyper baric chamber with the same oxygen and pressure as the hyperbaric chamber, the microfluidic chip comprising a plurality of microfluidic channels interconnected and arranged in a vascular configuration that mimics the vascular system of the patient and comprising a micro gas bubble in the microchannels mimicking the gas bubble in the patient’s bloodstream; and

[0034] monitoring the micro gas bubble in the microchannels and ceasing the hyperbaric oxygen therapy within the hyperbaric chamber when the micro gas bubble in the microfluidic chip is eliminated.

[0035] In some embodiments, the method further comprising before placing the patient in the hyperbaric chamber, performing a magnetic resonance imaging on the patient to determine the volume of the gas bubble.

[0036] In some embodiments, the microfluidic chip is selected among an array of microfluidic chips based on having the vascular configuration that best mimics the vascular system of the patient where the gas bubble is present.

[0037] In yet another aspect, there is provided a method of treating a gas embolism in a patient in need thereof, the method comprising administering to the patient’s bloodstream a therapeutically effective amount of a pharmaceutical composition comprising a surfactant and a pharmaceutically acceptable excipient.

[0038] In some embodiments, the pharmaceutical composition further comprises perfluorocarbons.

[0039] In an additional aspect, there is provided the use of a pharmaceutical composition comprising a surfactant and perfluorocarbons for treating a gas embolism in a subject need thereof.

[0040] In yet an additional aspect, there is provided a method of monitoring the formation of bubbles in a surgical procedure, the method comprising flowing the patient’s blood in a microfluidic chip comprising a plurality of interconnected microfluidic channels arranged in a vascular-like configuration to mimic a given part of a vascular system where the surgical procedure is performed, and monitoring the formation of gas bubbles in the microfluidic channels.

[0041] In a further aspect, there is provided a method of determining the formation of gas bubbles in a vascular system, the method comprising:

[0042] applying a compressive force a microfluidic chip to compress the microfluidic chip, the microfluidic chip comprising a plurality of microfluidic channels interconnected and arranged in a vascular-like configuration to mimic a given part of the vascular system;

[0043] releasing the compressive force on the microfluidic chip;

[0044] after the releasing, flowing a blood sample in the microfluidic channels of the microfluidic chip; and

[0045] imaging the microfluidic chip to determine whether gas bubbles have formed in the microfluidic channels.

[0046] In still a further aspect, there is provided a method of determining the formation of gas bubbles in an individual with a wearable device, the method comprising: [0047] flowing a blood sample in a microfluidic chip inside the wearable device, the microfluidic chip comprising a plurality of interconnected microfluidic channels arranged in a vascular-like configuration to mimic a given part of the vascular system; and

[0048] imaging the microfluidic channels to determine when a gas bubble forms in the microfluidic channels; and

[0049] emitting an alert signal to the wearer when the gas bubble is detected in the wearable device.

[0050] Many further features and combinations thereof concerning the present improvements will appear to those skilled in the art following a reading of the instant disclosure.

DESCRIPTION OF THE DRAWINGS

[0051] FIG. 1 is a schematic showing a system for determining the presence of a gas bubble according to embodiments of the present disclosure.

[0052] FIG. 2A is a schematic of a microfluidic device with two inlets having channels in a Y-junction configuration at an angle of 90°, a drug inlet and an outlet.

[0053] FIG. 2B is a schematic of a microfluidic device with two inlets having channels in a T-junction configuration at an angle of 90°, a drug inlet and an outlet.

[0054] FIG. 2C is a schematic of a microfluidic device with two inlets having channels in a junction configuration at an angle of 75°, a drug inlet and an outlet.

[0055] FIG. 2D is a schematic of a microfluidic device with two inlets having channels in a junction configuration at an angle of 60°, a drug inlet and an outlet.

[0056] FIG. 2E is a schematic of a microfluidic device with two inlets having channels in a junction configuration at an angle of 30°, a drug inlet and an outlet.

[0057] FIG. 2F is a schematic of a microfluidic device with two inlets having channels in a junction configuration at an angle of 15°, a drug inlet and an outlet. [0058] FIG. 3A is a schematic of a microfluidic device with two inlets having channels in a Y-junction configuration at an angle of 90°, a biomimetic microvasculature on chip (BMOC) network with bifurcation angles of 90°, a drug inlet and an outlet.

[0059] FIG. 3B is a schematic of a microfluidic device with two inlets having channels in a Y-junction configuration at an angle of 90°, a BMOC network with bifurcation angles of 60°, a drug delivery inlet and an outlet.

[0060] FIG. 3C is a schematic of a microfluidic device with two inlets having channels in a Y-junction configuration at an angle of 90°, a BMOC network with bifurcation angles of 30°, a drug delivery inlet and an outlet.

[0061] FIG. 4A is a schematic of a BMOC network design with bifurcation angles of 76°.

[0062] FIG. 4B is a schematic of a microfluidic device with two inlets having channels in a Y-junction configuration at an angle of 90°, a BMOC network design with bifurcation angles of 76° as per Fig. 4A, a drug delivery inlet and three outlets.

[0063] FIG. 5A is a schematic of a BMOC network design with bifurcation angles of 24°.

[0064] FIG. 5B is a microfluidic device with two inlets having channels in a Y-junction configuration at an angle of 90°, a BMOC network design with bifurcation angles of 24° as per Fig. 5A, a drug delivery inlet and an outlet.

[0065] FIG. 6A is a microscopy image of Y- and T-junction channels.

[0066] FIG. 6B is a microscopy image of a BMOC network showing bifurcation angles of 90°, 60°, and 30° as shown in FIG. 3A, FIG. 3B, and FIG. 3C, respectively.

[0067] FIG. 6C is a microscopy image of BMOC networks as shown in FIG. 4A and FIG. 5A.

[0068] FIG. 7A is a schematic of a microfluidic system with three injection pumps, an imaging apparatus and a collection reservoir. [0069] FIG. 7B is a photograph of the microfluidic system of Fig. 7A.

[0070] FIG. 7C is a schematic representation of the detection system according to some embodiments.

[0071] FIG. 8A is a schematic showing a BMOC designed according to Murray’s law of bifurcation having a parent diameter of 100 pm, a daughter diameter of 16 pm and bifurcation angles of 76°.

[0072] FIG. 8B is a schematic of a microfluidic device with two inlets having channels in a Y-junction configuration, a BMOC as shown in FIG. 8A, and an outlet.

[0073] FIG. 9 is a microscopic image showing a BMOC designed according to FIG. 8A.

[0074] FIG. 10A is a microscopic image showing gas bubbles’ evolution inside a bifurcated network without surfactant. Visualization utilized an inverted confocal microscope paired with a high-speed camera. Air pressure = 340 mbar, synthetic blood flow rate = 400 pl/h.

[0075] FIG. 10B is a microscopic image showing gas bubbles’ evolution inside a bifurcated network with surfactant. Visualization utilized an inverted confocal microscope paired with a high-speed camera. Air pressure = 340 mbar, synthetic blood flow rate = 400 pl/h.

[0076] FIG. 11 A is a histogram depicting the area of bubbles within the BMOC without surfactant under 300 mbar of synthetic blood pressure and 260 mbar of air pressure.

[0077] FIG. 11 B is a histogram depicting the area of bubbles within the BMOC without surfactant under 400 mbar of synthetic blood pressure and 340 mbar of air pressure.

[0078] FIG. 11C is a histogram depicting the area of bubbles within the BMOC without surfactant under 600 mbar of synthetic blood pressure and 500 mbar of air pressure.

[0079] FIG. 11 D is a histogram depicting the area of bubbles within the BMOC with surfactant under 300 mbar of synthetic blood pressure and 260 mbar of air pressure. [0080] FIG. 11E is a histogram depicting the area of bubbles within the BMOC with surfactant under 400 mbar of synthetic blood pressure and 340 mbar of air pressure.

[0081] FIG. 11 F is a histogram depicting the area of bubbles within the BMOC with surfactant under 600 mbar of synthetic blood pressure and 500 mbar of air pressure.

[0082] FIG. 12A is a histogram depicting the maximum Feret diameter of bubbles within the BMOC without surfactant under 300 mbar of synthetic blood pressure and 260 mbar of air pressure.

[0083] FIG. 12B is a histogram depicting the maximum Feret diameter of bubbles within the BMOC without surfactant under 400 mbar of synthetic blood pressure and 340 mbar of air pressure.

[0084] FIG. 12C is a histogram depicting the maximum Feret diameter of bubbles within the BMOC without surfactant under 600 mbar of synthetic blood pressure and 500 mbar of air pressure.

[0085] FIG. 12D is a histogram depicting the maximum Feret diameter of bubbles within the BMOC with surfactant under 300 mbar of synthetic blood pressure and 260 mbar of air pressure.

[0086] FIG. 12E is a histogram depicting the maximum Feret diameter of bubbles within the BMOC with surfactant under 400 mbar of synthetic blood pressure and 340 mbar of air pressure.

[0087] FIG. 12F is a histogram depicting the maximum Feret diameter of bubbles within the BMOC with surfactant under 600 mbar of synthetic blood pressure and 500 mbar of air pressure.

[0088] FIG. 13A is a schematic of a microfluidic device with a main channel having an inlet and an outlet, and two side chambers with their respective inlets.

[0089] FIG. 13B is a schematic of a microfluidic device with a main channel having an inlet and an outlet, and one side chamber with its respective inlet. [0090] FIG. 14A is a microscopy image of two microfluidic devices as shown in FIG. 13A and FIG. 13B.

[0091] FIG. 14B is a microscopy image of the microfluidic devices of FIG. 14A showing the formation of gas bubbles in the main channel upon compression of the side chamber(s).

[0092] FIG. 15A is a microscopy image of the microfluidic device of FIG. 13A showing the formation of gas bubbles in the main channel upon gas insufflation in the two side chambers.

[0093] FIG. 15B is a microscopy image of the microfluidic device of FIG. 14A showing the formation of gas bubbles in the main channel upon water compression in the side chamber.

[0094] FIG. 16A is a schematic of a microfluidic device with a main channel having multiple side chambers in proximity to two side channels, each channel having an inlet and an outlet.

[0095] FIG. 16B is a schematic of the microfluidic device of FIG. 16A having various chamber designs.

[0096] FIG. 17A is a microscopy image of the chamber designs of FIG. 16A.

[0097] FIG. 17B is a microscopy image showing bubble formation in a microfluidic channel when pressure is applied to each of the chamber designs of FIG. 16A.

[0098] FIG. 18 is a schematic of a microfluidic system comprising the microfluidic devices of FIG. 13A, FIG. 13B or FIG. 16A, two pumps, an imaging system, and a computer.

[0099] FIG. 19 is a schematic of a microfluidic device having a main channel surrounding at least one alveolar sac configuration arising from a second channel, each of the channels having an inlet and an outlet. [0100] FIG. 20A is a schematic of a multilayer microfluidic device having a main channel layer with microvasculature chambers being surrounded by a second layer with alveolar sacs arising from a second channel, and a membrane layer, wherein each of the channels have an inlet and an outlet.

[0101] FIG. 20B is a diagram of the membrane interfacing the microvasculature chambers and the alveolar sac configurations of FIG. 20A.

[0102] FIG. 21 A is a schematic of a BMOC having a systemic microvasculature covered by a thick polydimethylsiloxane (PDMS) layer connected to a pulmonary microvasculature covered by a thin PDMS layer.

[0103] FIG. 21B is a scanning electron microscopy image of the thin PDMS layerfrom FIG. 21A.

[0104] FIG. 22A is a photograph of a mini hyperbaric system for benchtop experimental simulation of gas emboli containing a microfluidic device.

[0105] FIG. 22B is a schematic representation showing an exploded view of a microscale hyperbaric system containing a microfluidic device.

[0106] FIG. 23A is a schematic representation showing an exploded view of a microscale hyperbaric system containing a microfluidic device including exemplary photograph of components (battery, peristaltic pump and blood reservoir).

[0107] FIG. 23B is a photograph of the microscale hyperbaric system of FIG. 23A.

[0108] FIG. 24A is a schematic of a microfluidic device designed at biologically relevant diameters and geometries.

[0109] FIG. 24B is a schematic of a microfluidic device with the same geometry as FIG. 5A but with different dimensions (certain channel widths increased).

[0110] FIG. 24C is a schematic of a microfluidic device with the same geometry as FIG. 5B but with different dimensions (certain channel widths increased). [0111] FIG. 25 is a microscopy image of the microfluidic devices from FIG. 5A, FIG. 5B, and FIG. 5C.

[0112] FIG. 26A is a graph showing the volumes of generated bubbles in tissue compression over time using microfluidic devices undergoing compression at 1 bar for 2 hours.

[0113] FIG. 26B is a graph showing the volumes of generated bubbles in tissue compression over time using microfluidic devices undergoing compression at 1 bar for 4 hours.

[0114] FIG. 26C is a graph showing the volumes of generated bubbles in tissue compression over time using microfluidic devices undergoing compression at 1 bar for 6 hours.

[0115] FIG. 26D is a graph showing the volumes of generated bubbles in tissue compression overtime using microfluidic devices undergoing compression at 1 bar for 12 hours.

[0116] FIG. 27A is a graph showing the volumes of generated bubbles in tissue compression over time using microfluidic devices undergoing compression at 3 bar for 2 hours.

[0117] FIG. 27B is a graph showing the volumes of generated bubbles in tissue compression over time using microfluidic devices undergoing compression at 3 bar for 4 hours.

[0118] FIG. 27C is a graph showing the volumes of generated bubbles in tissue compression over time using microfluidic devices undergoing compression at 3 bar for 6 hours.

[0119] FIG. 27D is a graph showing the volumes of generated bubbles in tissue compression overtime using microfluidic devices undergoing compression at 3 bar for 12 hours. [0120] FIG. 28A is a graph showing the volumes of generated bubbles in diver model compression over time using microfluidic devices undergoing compression at 1 bar for 2 hours.

[0121] FIG. 28B is a graph showing the volumes of generated bubbles in diver model compression over time using microfluidic devices undergoing compression at 1 bar for 4 hours.

[0122] FIG. 28C is a graph showing the volumes of generated bubbles in diver model compression over time using microfluidic devices undergoing compression at 1 bar for 6 hours.

[0123] FIG. 28D is a graph showing the volumes of generated bubbles in diver model compression overtime using microfluidic devices undergoing compression at 1 bar for 12 hours.

[0124] FIG. 29A is a graph showing the volumes of generated bubbles in diver model compression over time using microfluidic devices undergoing compression at 3 bar for 2 hours.

[0125] FIG. 29B is a graph showing the volumes of generated bubbles in diver model compression over time using microfluidic devices undergoing compression at 3 bar for 4 hours.

[0126] FIG. 29C is a graph showing the volumes of generated bubbles in diver model compression over time using microfluidic devices undergoing compression at 3 bar for 6 hours.

[0127] FIG. 29D is a graph showing the volumes of generated bubbles in diver model compression overtime using microfluidic devices undergoing compression at 3 bar for 12 hours.

DETAILED DESCRIPTION [0128] The present disclosure addresses the modeling challenge for gas emboli, with regards to the formation of air bubbles and the treatment of air bubbles. The goal of the present models are to mimic gas emboli in vitro using microfluidic systems that replicate the blood flow in vasculature-on-a-chip models, which allow real-time monitoring. It has been presently found that microfluidic chips can advantageously be used to simulate a particular geometry of a vascular system.

[0129] A system is therefore disclosed that includes a microfluidic chip (vasculature- on-a-chip model), a pump for flowing a liquid in the microfluidic chip and an imaging apparatus, which allows for the simulation of blood flow in vitro. This system can be used for a number of possible purposes, including for example drug discovery research for treating gas emboli, for the calibration of invasive medical procedures to avoid or limit the occurrence of gas embolism, forthe monitoring of the evolution of gas emboli in hyperbaric chamber therapy, and for monitoring the formation of air bubbles when the system is subjected to different conditions such as pressure variations.

[0130] The present systems leverage the mimicking and miniaturization of the biological environment using polydimethylsiloxane (PDMS) based microfluidic chips. This allows the study of the formation and evolution of gas bubbles in real time and in vitro. The additional advantage of the proposed systems is the ability to perform an imagingbased, quantitative study using onboard imaging modalities that are rapidly adaptable to the need, and the system in use. Overall, the systems and the workflows described herein are versatile for studying different types of global and local gas emboli.

[0131] Fig. 1 illustrates the system 1 of the present disclosure according to some embodiments. The system of the present disclosure is for determining the presence of a gas bubble in a blood sample. There is provided a microfluidic chip 100 comprising a plurality of interconnected microfluidic channels arranged in a vascular-like configuration to mimic a given part of a vascular system. The microfluidic chip has an inlet for receiving the blood sample. A circulation system 10 is used for circulating the blood sample in the microfluidic chip. The circulation system 10 has a pump 11 and a reservoir 12 containing the blood sample and a conduit 13 in communication with the pump 11 and the reservoir 12 to direct the blood sample to the inlet of the microfluidic chip 100. The system has an imaging system 20 configured to capture images of the microfluidic channels of the microfluidic chip and a detection system 21 configured to determine the presence and location of the gas bubbles based on the captures images of the imaging system. The imaging system 20 and detection system 21 are electrically coupled or electrically connected.

[0132] The microfluidic networks in the microfluidic chip are provided in a vascular- like configuration that mimics the microvasculature which can be of different complexity. The vasculature of animals (e.g., humans), including the microvasculature, comes in different geometrical complexities, with varying lengths, bifurcations angles, and branching architecture. The vascular-like configuration of the microfluidic chip can be designed to replicate a given part of the vasculature in an animal (e.g. human). The vascular system has been well studied and the different dimensions of the arteries and veins is known. For example, during a surgery, the vascular section of the patient that is at risk of developing gas bubbles is known and accordingly a geometry of the vascular- like configuration can be pre-fabricated to generally replicate the relevant bifurcation angle(s) and other features of the vasculature. It is contemplated herein, that the vascular- like configuration of the microfluidic chip can be fabricated on a case-by-case basis for personalized modelling for a given part of the vascular system of an individual. However, there are many similarities in terms of physical dimensions across vascular systems, and for a more efficient approach, there is also contemplated that microfluidic chips with generic vascular-like configurations are pre-fabricated and then based on the situation the microfluidic chip that best fits the situation to model is selected. From a point of care perspective, it is possible that a multitude of microfluidic chips with different vascular-like configurations are available to the medical practitioner to select from. The vascular-like configuration can be complex or simple.

[0133] Figs. 2A-2F illustrate certain simple geometries (single bifurcation). Two inlets 101 , for a blood sample, are provided which lead to a bifurcation at different angles, for example 90, 75, 60, 30, and 15 degrees as illustrated in the figures. However, the angle can be modified to be any suitable value that mimics a particular bifurcation of the vascular system. There is optionally provided a drug inlet 102 when the microfluidic chip is used for drug screening and discovery for the treatment of gas bubbles in the bloodstream. The gas can be introduced in the microfluidic channels through one of the inlets 101 or through a dedicated gas inlet. The volume of gas introduced to create the gas bubble aims to replicate the size of a gas bubble in the bloodstream relative to the size of the microfluidic channels. In otherwords, the size and volume of the gas bubble created in the microfluidic channels follows the same scale down process as the microfluidic channels. In the embodiments, of Figs. 2A-2F (dimensions in micrometers) a single outlet 103 is shown for each microfluidic chip, however, multiple outlets are also possible, particularly when the geometry of the microfluidic channels is complex.The embodiments of Figs. 2A-2F use planar representations of complex arterial and venous blood flow systems with complex bifurcation architecture. These arterial and venous bifurcation geometries are based on Murray's design law of conservation, utilizing the squared and cubed mathematical models for modelling fluid flow through natural vasculature channels in animals such as humans and pigs.

[0134] Figs. 3A-3C (dimensions in micrometers) provide embodiments where an additional complexity feature is added in the vascular-like configuration of the microfluidic chip. Namely, a biomimetic microvasculature on chip (BMOC) network 104 is added. The BMOC is a geometry that mimics the progressive shrinking in size of blood vessels which occurs in the body to allow for organs to extract nutrients and oxygen from the blood. Indeed, the vascular system gets progressively small until a capillary bed is formed. Capillaries generally have a thickness of a single cell and allow for exchanges to occur between organs and the blood (e.g. intake of oxygen). Figs. 3A-3C provide a simplistic representation of a BMOC as a diamond shape. The BMOC (i.e. inside the diamond shape represented in Figs. 3A-3C) can include bifurcations of various angles to mimic a given part of the vascular system.

[0135] As an example of a BMOC, a microfluidic device was designed to mimic the microvasculature of alveoli with a horizontally placed alveolar membrane-like gasexchange barrier Fig. 4A. The microfluidic chip containing the alveolar BMOC 104 is shown in Fig. 4B (dimensions in micrometers). This microfluidic chip can be used as a model for systemic gas embolism for the lungs. The alveoli-lung membrane barrier was fabricated to mimic the pores on the alveoli surface for gas exchange. This is only one example, many different geometries of the vascular system can be fabricated to mimic various parts of the vascular system. For example, the vascular system of the brain, the liver, the pancreas, or any other organ can be fabricated.

[0136] Figs. 5A and 5B (dimensions in micrometers) show respectively a different BMOC and a microfluidic chip comprising same. The geometry of the BMOC of Fig. 4A is based on Murray’s law whereas the geometry of the BMOC of Fig. 5A is based on the power squared law.

[0137] The geometries described above have been fabricated, and microscopy images of the bifurcations are showed in Figs. 6A-6C. Specifically, Fig. 6A shows the Y and T bifurcations, Fig. 6B shows 90, 60 and 30 degree bifurcation angles and Fig. 6C shows the BMOC of Fig. 5A (top) and Fig. 4A (bottom). PDMS was the material used for the microfluidic chip. PDMS is transparent and rubber-like. The transparency of PDMS enables optical imaging and the rubber-like characteristic mimics tissues in the body. The concentration of the buffer when preparing PDMS and the curing time can be changed to tune the properties of the PDMS. This can be done to have a different hardness of PDMS in order to mimic different tissues depending on the targeted vascular region to mimic. Although PDMS is a preferred embodiment, there are alternatives to PDMS and the present disclosure is not limited to PDMS.

[0138] An exemplary embodiment of the system of the present disclosure is illustrated in Fig. 7A and a photograph thereof is shown in Fig. 7B. The system has three pumps 106a, 106b, 106c respectively for gas, a blood sample and a drug. Each pump is fluidly connected to the appropriate inlet on the microfluidic chip 100. An imaging apparatus 107 is positioned to image the flow of the blood sample and the gas in the microfluidic channels of the microfluidic chip. The blood sample can be collected in a reservoir 108. The microfluidic chip 100 can for example have a BMOC as per Fig. 8A with angles at 76 degrees and a structure as per Fig. 8B (dimensions in micrometers).

[0139] It should be noted that the embodiment of Fig. 7A is only an exemplary embodiment. The drug inlet (and corresponding drug pump 106c) is an optional feature that is only present when the purpose of the system is to screen or discover drugs for treating the presence of gas bubbles in a bloodstream. Furthermore, in some embodiment, the gas pump 106b and gas inlet are excluded when the purpose of the system is to study the onset of a gas bubble due to changes in the environment (e.g. pressurization / depressurization). Moreover, the collection reservoir 108 is an optional feature. Indeed, in some embodiments, the blood sample can be recycled to the pump. As will be described in further details herein below, some embodiments of the system require operating the flow of the blood sample for long periods of time (e.g. multiple hours or even indefinitely). In such embodiments, instead of a collection reservoir 108, there is provided a system to recycle the blood sample to the pump such that it is provided again at the inlet of the microfluidic chip.

[0140] The term “blood sample” as used herein is defined as a fluid that is blood or mimics the physical properties of blood (e.g. viscosity, density, turbidity, etc.), which may also be referred to herein as “artificial blood” or “synthetic blood”. The blood sample is for example an aqueous composition designed to mimic the viscosity and density of blood. Protein and lipid additions can be made to the aqueous composition to tune its physical characteristics. In other embodiments, the blood sample can be blood. In particular, for a more accurate representation of the vascular system of an individual, the individual’s own blood can be used as the blood sample. The blood sample can also be donor blood. Because the present system utilizes a microfluidic chip, the volume of blood sample (such as blood) required is small. The blood viscosity is generally dependent on the content of red blood cells (hematocrit) and plasma viscosity. The shear thinning property of blood is mainly attributed to red blood cell (RBC) rheological properties. Accordingly, different individuals with different contents of RBC will have different blood viscosities. The blood sample of the present disclosure can be adapted to have an increased or decreased viscosity to best mimic a particular situation. In a clinical setting, when possible, it is preferable that the blood sample is blood, particularly blood from the patient with gas in their bloodstream.

[0141] As illustrated in Fig. 7 A, multiple pumps can be provided in the present system. However, only one pump is necessary in the simplest embodiment and the additional pumps are optional. There are many pumps appropriate for use in conjunction with microfluidic chips, such as mechanical pumps, pressure pumps or syringe pumps. In preferred embodiments, the pump is a peristaltic pump to mimic the pulsative nature of blood flow in the body. In some embodiments, the physical processes in the microvasculature may be mimicked using pressure pumps and associated accessories, such as solution reservoirs and syringes, pressure sensors for mimicking, controlling, and observing the blood flow and other physical parameters in the mimicked microvasculature. In certain embodiments, to mimic the flow conditions in the microvasculature and test the impact of different drugs on fluid behavior like flow properties, bubble retention, and collapse in the system, the system may use multiple pressure pumps (operated by pressure) or pulseless syringe pumps, which can be systematically connected to the microfluidic devices and observed for the genesis, evolution, and treatment of gas emboli.

[0142] The imaging apparatus of the system is shown as a camera in Fig. 7 A. There are alternatives to the camera as optical imaging and there are also alternatives to optical imaging. For example, instead of a camera a microscope can be used. Alternatives to optical imaging include but are not limited to ultrasound imaging and magnetic resonance imaging. It should be noted that optical imaging is the simplest to implement and is the non-limitative preferred embodiment.

[0143] In one embodiment, the imaging apparatus is a multimodal, adaptable imaging apparatus for acquiring real-time imaging data. This can be a cell phone camera or a microscope imaging. Different light sources and operational modalities may be used for monitoring the genesis, treatment efficiencies, and impact of therapeutics on gas embolism. Studying the gas embolism phenomenon with images in vivo is daunting, and in vast majority of situations impossible in real-time during experiments. This is because the presence of gas bubbles in the bloodstream often leads to death. The present model allows for studies on the presence of gas in the bloodstream which was not previously possible, specifically studies relating to the embolism phenomenon with image-based quantitative analysis.

[0144] The detection system is for example as illustrated in Fig. 7C. The detection system 21 (which may also be referred to herein as a computing device) can, for example, be any suitable type of computing device, including a desktop or laptop computer, a portable computing device, for instance a smartphone or the like, a dedicated electronic device, for instance a device configured for performing gene sequencing, or the like. The computing device 21 comprises a processing unit 22 and a memory 24 which has stored therein computer-executable instructions 26. The memory 24 may also store other relevant information for the implementation of the present method. The processing unit 22 may comprise any suitable devices configured to implement the functionality of the computing device 21 such that instructions 26, when executed by the detection system 21 or other programmable apparatus, may cause the image processing of the captures images to yield an output that determines a characteristic of the gas bubble (e.g. presence, size of the bubble, position or location of the gas bubble). The processing unit 22 may comprise, for example, any type of general-purpose microprocessor or microcontroller, a digital signal processing (DSP) processor, a central processing unit (CPU), an integrated circuit, a field programmable gate array (FPGA), a reconfigurable processor, other suitably programmed or programmable logic circuits, custom-designed analog and/or digital circuits, or any combination thereof.

[0145] The memory 24 may comprise any suitable known or other machine-readable storage medium. The memory 24 may comprise non-transitory computer readable storage medium, for example, but not limited to, an electronic, magnetic, optical, electromagnetic, infrared, or semiconductor system, apparatus, or device, or any suitable combination of the foregoing. The memory 24 may include a suitable combination of any type of computer memory that is located either internally or externally to device, for example random-access memory (RAM), read-only memory (ROM), compact disc read-only memory (CDROM), electro-optical memory, magneto-optical memory, erasable programmable read-only memory (EPROM), and electrically-erasable programmable read-only memory (EEPROM), Ferroelectric RAM (FRAM) or the like. The memory 24 may comprise any storage means (e.g., devices) suitable for retrievably storing machine-readable instructions 26 executable by processing unit 22.

[0146] Computer-executable instructions may be in many forms, including program modules, executed by one or more computers or other devices. Generally, program modules include routines, programs, objects, components, data structures, and the like, that perform particular tasks or implement particular abstract data types. Typically the functionality of the program modules may be combined or distributed as desired in various embodiments. [0147] Image processing algorithms are well established and available. In preferred embodiments, the imaging system captures an optical image of the microfluidic channels. The resulting image can be analyzed automatically by an algorithm that knows or is trained for finding gas in the microfluidic channel (e.g. absence of color, lighter color, darker color, or any other visual indicator that differentiates the gas bubble from the flowing blood sample). The image analysis is capable of therefore identifying the characteristics of the bubble. The detection system can comprise an interface in which the results of the analysis are displayed. The display can for example show the live image of the microfluidic device and also have on the screen the size of the bubble.

[0148] The system described herein allows to study which blood vessel junctions and their dimensions (channel widths, lengths), bifurcation angles have an increased risk of blockage to blood flow by air bubbles; what are the physical conditions that favor blockages and if any drugs can be used to assuage the bubble formation or its evolution to fatal gas embolism; and what are the causes of gas embolism during gas insufflation. The miniaturized setup and the associated microfluidic designs described herein provide opportunities to study gas embolism and prevention strategies. Researchers can investigate the effects of different angles of air introduction (iatrogenic gas embolism) and bifurcation of mimicked blood vessels of different widths to observe how bubbles behave in simple and complex microchannels. The design adaptability enhances the understanding of gas embolism dynamics and evaluates drug effectiveness under different conditions.

[0149] The present system is a cost-effective alternative to in vivo investigation methods for evaluating the occurrence of gas embolism and the effectiveness of different drugs. By using bio-mimicked microfluidic devices, one can simulate and observe the evolution of bubbles in a controlled environment, reducing the need for expensive animal studies.

[0150] By providing an alternative to animal testing and invasive procedures on humans, the invention contributes to the reduction of ethical concerns associated with traditional research methods. This can be particularly beneficial when studying the occurrence and treatment of gas embolisms, where direct experimentation on living organisms may be challenging or undesirable. Incidentally, the food and drug administration does not require mandatorily animal studies for drug discovery.

[0151] As explained above, in some embodiments, the present system is used for performing drug discovery or screening. The system has a microfluidics chip, a pump and an imaging apparatus. The microfluidics chip contains a network of channels that mimics a vascular system. The channels generally aim to mimic the bifurcation angles of blood vessels (venous or arterial) or pulmonary gas exchange (alveoli). The pump is associated with a reservoir housing, connecting tubes and channels, and measurement devices for determining the pressure of the blood sample (blood sample and gases in the case of a gas exchange system). The imaging system can comprise a microscope for capturing the image and an image processing device which can be a mobile phone, a computer or any other suitable processor. The present drug screening system can significantly streamline the drug development process by quickly identifying promising candidates and eliminating ineffective ones, saving time and resources, and potentially accelerating the availability of effective treatments.

[0152] The present system can also be used to calibrate invasive medical procedures by creating a local pressurization and depressurization to test the limits of the blood vessels in the model system performing applying them in surgery. The ability to vary local pressures within a microfluidic setup to study the impact of multiple periods and extents of compression and decompression on intravascular bubble formation thus allows to adapt surgical procedure to attempt to avoid bubble formation.

[0153] The geometry of the microfluidic channels can be used to create a local increase in pressure using fluids or trapped gases which will lead to the breakage of the walls of the mimicked blood vessels and initiation of the gas bubbles due to local variation of pressure. A piston like system can be used to mechanically create a pressurization and depressurization in the microfluidic channels. A laser with similar specifications as a surgical laser can be used to evaluate its effect on the blood flow.

[0154] These microfluidic designs provide the ability to manipulate and control the pressure exerted within the device through the use of different geometric networks. This concept recreates the effects of indentation on a microscale level by incorporating the ability to use different 'tips' on a single platform. The induced pressure is under a broad range of applications and a higher level of control with real-time imaging and visualization of the onset of bubble formation.

[0155] The system therefore enables the study of optimum preventive measures and design features to optimize surgical tools under conditions that resemble those of different medical procedures and surgical interventions.

[0156] Most incidences related to iatrogenic gas embolism are reported in (a) procedures where surgical incisions are superior to the heart at distances greater than the central venous pressure, and (b) direct air injection where the lethal dose varies from 3 to 5 mL/kg for healthy individuals. However, there are no data related to the onset of bubble formation in these instances. The present systems enable the ability to utilize microchannels for local gas and/or liquid pressurization in a controllable manner. In fact, during laparoscopic surgeries, air or carbon dioxide is insufflated into the peritoneal cavity via a Veress needle that is inserted without visual guidance. This procedure can be simulated in the proposed microfluidic structures, where bubbles are exogenously generated in vascularized channels facing a 'micro-indentation channel' at different levels of pressurization. The onset of gas bubbles during surgery (insufflation) can be monitored in some embodiments with the present system. The patient’s blood can be flown in microfluidic channels mimicking the vascular geometry where the surgery is occurring. The onset of bubbles can be observed in the microfluidic channels to adjust the surgical procedure and attempt to avoid any further bubble formation.

[0157] There is also provided the use of the present system for monitoring hyperbaric therapy using the microfluidic chip as described above along with the blood flow system and imaging system additionally with an enclosure allowing the controlled pressurization of the system described. When optical imaging is performed, the enclosure must have transparent walls, or at the very least a window to allow the visualization of the gas emboli.

[0158] One of the reasons why the described systems are suitable models for gas embolism is because gas embolism is, in the first instance a purely physical process, with biochemical, and biological processes, 'kicking in' later, in matter of minutes to hours, and even days. Furthermore, the major target of hyperbaric therapy is the dissolution of the gas bubbles, again mostly a physical process, which admittedly impact gravely on biochemical and biological systems. Consequently, the present methods, systems and models focus on the initial stages of gas embolism, thus there is no need for complicated tissue engineering constructs.

[0159] Generally the hyperbaric setup is operable in a high pressure range whereas there is a need for clear imaging during the hyperbaric conditions. Pyrex was determined as a preferred material for the hyperbaric chamber material because it can withstand 1000 psi, which is at least two times higher than the existing operation limits of the in-market clear pressure vessels made of organic polymers, another downside of using organic polymers is the poor imaging abilities and translucence and high background fluorescence. Also, several microscope-on-stage isolation chambers do not tolerate high- pressure variations, significantly reducing experimental scalability at extreme pressures. In the present model, the pressure range around 150 psi can be expected (e.g. situation of a swimmer at a depth of 100 m).

[0160] In addition to its use in the context of decompression sickness, the present mini-hyperbaric chamber can be used to study different phenomena in microfluidic systems in which the variation of the global pressure is a crucial parameter. For instance, microfluidic devices, commonly known as lab-on-a-chip, are used to manipulate and analyze small volumes of biological samples. The pressure applied within these devices can affect the behavior of cells, such as cell deformation, flow dynamics, and fluid mixing. The mini-hyperbaric chamber can be a platform for this investigation. In addition, in bioreactors or cell culture systems, pressure can influence the growth and behavior of cells. For example, high-pressure bioreactors can enhance the production of specific metabolites or proteins, while low-pressure conditions may be necessary for specific cell types to thrive. Further, the demonstrated type of pressure devices can be useful also for growing extremophiles that require high pressure to be cultured in the lab.

[0161] The present hyperbaric chamber model system can be used to monitor the treatment of a patient undergoing hyperbaric oxygen therapy (HBOT). An important disadvantage of classical HBOT is that it is not possible to image the patient’s bloodstream inside the hyperbaric chamber because imaging apparatus are fire hazards due to the use of hyper oxygenic conditions. The present system can be used to model the patient’s situation and in parallel perform a hyperbaric treatment to determine when the gas bubble is eliminated.

[0162] There are multiple strategies to mimic the patient’s bloodstream on the microfluidic chip. Before entering the hyperbaric chamber, the patient can be subjected to MRI imaging to determine the location and size of the gas bubbles in their bloodstream. Based on this information, the microfluidic chip can be selected or fabricated to have a vascular-like configuration that aims to mimic the vascular region affected by gas bubbles in the patient. Since the size of the bubbles are known for the MRI imaging, the bubbles can be reproduced in the microfluidic channels. In other cases the size and location of the bubbles is generally known and there is no need to image the patient. This is generally the case when the bubbles are caused during a surgery or during an ultrasound imaging. Then, the microfluidic chip is placed in a mini hyperbaric chamber and subjected to the same conditions as the patient in the hyperbaric chamber. When the gas bubble is eliminated in the microfluidic chip, then that means that the patient’s bubbles have also most likely been eliminated. The monitoring of HBOT is important because it is often the case that medical practitioners will take the patient out of the chamber due to discomfort but the bubbles have not been eliminated. This monitoring thus increases patient survival.

[0163] A different shape and size of the Pyrex housing may be used to optimize the configuration and overall size of the mini-hyperbaric chamber. The encasing for the Pyrex container and the camera or imaging setup may be designed in different configurations for different applications and requirements. A different image-capturing technic or imaging camera may be used to improve the extraction of data from the sample placed inside the mini hyperbaric chamber. The enclosure dimensions are variable because they are dependent on the type of device that has to be sealed.

[0164] Pressure vessels are used simulate and study microscopic level fluid dynamics and biological systems. In some embodiments, pressure vessels are placed inside the Pyrex mini hyperbaric chambers. The traditional tabletop clear pressure vessel has limitations in its pressure range and imaging clarity due to the use of polyplex polymers as construction materials. The polyplex polymers lead to complicated assembly involving metal rods which further hinders imaging with handheld cameras. In contrast, the mini- hyperbaric chamber in the present disclosure is made of a Pyrex body and plastic caps, and offers clear imaging, including microscopic details (resolution around 10 pm). Additionally, the current systems enables fluid operations within the Pyrex chamber, allowing simultaneous pressurization and depressurization of a microfluidic device, which is an advantage of the present systems. The ability to provide real time imaging within the min hyperbaric chamber also allows to visualize and record the onset of bubble formation and its subsequent evolution as a result of local pressure modifications.

[0165] The mini hyperbaric chamber is a portable pressurizing and depressurizing microenvironment that includes all the elements needed for continuous blood circulation at the microscopic scale. The ability to provide constant monitoring during all stages of local pressure variations is an advantage.

[0166] Accordingly, in some embodiments, the microfluidic chip is provided without a direct entry of gas in the microfluidic channels and the purpose is to determine the onset and formation of gas bubble in the microfluidic channels. The bubble formation can be caused by a pressurization / depressurization protocol, by compression of the chip. Such a system can be incorporated in a wearable device (such as a watch or wrist wearable device) for divers or soldiers to take to the field. The microfluidic chip can have one or a plurality of vascular-like geometries that are known to be at risk for bubble formation and a continuous flow of blood sample (e.g. the wearer’s own blood) is flowed in the system. An alert notification can be emitted when a bubble is formed in order to alert the wearer. This can help a diver properly time their return to the surface for example. The compression on the chip can be performed to mimic the diver’s situation. For example, the PDMS is compressed before any blood sample is flown and the compression is released when the blood sample is flown. As the compression is releasing there is a change in pressure and potential gas entry similar to the dissolution of nitrogen in the body of the diver at different pressures which can create gas bubble.

[0167] It has also been presently found that a surfactant can be used to reduce or eliminate a gas bubble in the bloodstream. The term “surfactant” as used herein refers to a chemical substance that can reduce the surface tension of a fluid in which it is dissolved. Surfactants may act at the liquid-liquid, liquid-gas or liquid-solid interfaces to reduce surface tension. For example, pulmonary surfactants are found in the fluid lining of the airliquid interface of an alveolus, preventing its collapse during respiration. In the case of a gas embolism, surfactants can act to reduce the surface tension at the embolus-liquid interface, allowing their potential use for the medical treatment of gas emboli. Surfactant may be any of detergents, emulsifiers, wetting agents, foaming agents, additives or dispersants. Surfactants may be an amphiphilic molecule, specifically an amphiphilic molecule having a hydrophilic head region and a hydrophobic tail region. Surfactants may be classified by their head region properties being non-ionic, anionic, cationic, or zwitterionic.

[0168] Medically relevant surfactants may be used in drug formulations for the treatment of gas emboli, which may include surfactants from any of carbomer copolymers, carbomer interpolymers, cholesterols, ethylene glycol stearates, emulsifying waxes, fatty acids, glyceryl distearates, glyceryl monooleates, glyceryl monolinoleates, glycerin monostearates, lanolin alcohols, lecithins, mono/diglycerides, poloxamers, polyoxylethylene 50 stearates, polyoxyl 10 oleyl ether, polyoxyl 20 cetostearyl ether, polyoxyl 35 castor oil, polyoxyl 40 hydrogenated castor oils, polyoxyl 40 stearate, polyoxyl lauryl ethers, polyoxyl stearyl ethers, polysorbates (e.g., 20, 40, 60, 80), poloxamer (184, 188), propylene glycol monostearates, sodium cetostearyl sulfates, sodium lauryl sulfates, sorbitan monolaurates, sorbitan sesquioleates, sorbitan monooleates, sorbitan monostearate, sorbitan trioleates, sodium stearates, stearic acid, steroid acids, fatty acid esters, and phospholipids.

EXAMPLE

EXAMPLE 1 : Predicting gas embolism events through dynamic analysis of bubble motion within bio-mimicked microvasculature.

[0169] Gas embolism risks during medical surgeries result from stagnant gas bubbles obstructing microvascular blood flow. Despite fatal and sometimes chronic impacts, the mechanisms of bubble formation and its dynamic halting behavior in microvasculature remain poorly understood. The present disclosure pertains to devices and methods for examining the evolution of introduced bubbles in bifurcating microvasculature. For studying the dynamics of the air bubbles in a microvasculature, air bubbles are introduced directly into the microvasculature to mimic real-world scenarios, such as those occurring during surgical procedures. The present disclosure further pertains to devices and methods exploring fact-based therapeutic intervention for the prevention and treatment of gas embolism. This includes examining their interaction with gas embolism dynamics and their effectiveness in countering bubble-related halts, providing insights into innovative treatments for gas embolism.

[0170] A biomimetic microvasculature on chip (BMOC) may be used to investigate bubbles’ evolution and evaluate alternative therapies. The chip is designed with biologically relevant dimensions and follows Murray's law of bifurcation for examining bubble dynamics by simulating microvasculature geometry, blood composition, and air pressures. Initially, the parent diameter is set at 100 pm, while the daughters reach a final diameter of 16 pm, chosen with consideration of arteriole and venule dimensions associated with gas embolism risk (Fig. 9).

[0171] This involves assessing pharmaceutical interventions for the removal and reabsorption of bubbles. The proposed medications utilize efficient surfactants to lower the surface tension between gas and liquid, along with perfluorocarbons (PFCs) known fortheir high gas solubility due to their molecular structure. This approach aims to prevent the formation of stagnant bubbles, reducing their size (Figs. 10A-10B) or aid in the therapeutic procedure during oxygen therapy for patients who have experienced gas embolism.

[0172] The dynamic analysis of bubble motion is presented through histograms depicting bubble area and maximum Feret diameters within the microvasculature under two scenarios: the injection of synthetic blood and the injection of a surfactant (Poloxamer). Figs. 11A-11 F and 12A-12F illustrate these scenarios. The presence of the surfactant significantly reduces the bubble area and Feret diameters, thereby decreasing the risk of gas embolism.

[0173] The angles within the microvasculature contribute to bubble fragmentation, allowing smaller bubbles to pass through with less pressure drop. The utilization of surfactants amplifies the likelihood of bubble introduction at reduced air pressures within the microvasculature. Conversely, the presence of surfactants diminishes the entrapment of bubbles within the network under conditions of moderate to high air pressures.

Example 2: Iatrogenic gas embolism simulation system.

[0174] Microfluidic chips were designed as per Fig. 13A and 13B. The microfluidic chip as per Fig. 13A has two pressure chambers 105 that surround the microfluidic channel in order to create a gas insufflation situation (see Fig. 15A). Fig. 13B has a single pressure chamber 105 that applies a water compression in the micro indentation channels. This pressure chamber was designed with different geometries (see Figs. 15B and 17A- 17B). The onset of bubble formation was observed by microscopy (Figs. 14A-14B). Different geometries, dimensions and configurations were explored (Figs. 16A-16B) and the system as a whole is shown in Fig. 18, with two pumps 106, a microscope 107 and a computer 109 for processing the image from the microscopy 107. The size of the main vascularized channel was varied between these three different diameters: 20 pm, 30 pm and 40 pm and was is surrounded by 200 pm alveolar chambers on each side at a separation of 10 pm (see Fig. 13A). The other main vascularized channel was varied with six different diameters : 20 pm, 30 pm, 50 pm, 80 pm, 250 pm and 500 pm and was facing a 1 mm² cavity at a separation of 50 pm (see Fig. 13B). In the mixed configuration (Figs. 16A-16B), six diameters of a vascularized channel were used 20 pm, 30 pm, 50 pm, 80 pm, 120 pm and 180 pm and they were facing four different cavities at a separation of 50 pm. The pump used was a pulseless syringe pumps. The liquid used was blood which was housed in the reservoir syringe.

Example 3: Hyperbaric chamber system.

[0175] A mini hyperbaric chamber for recreating and studying the genesis of global gas embolism was created. The mini hyperbaric chamber miniaturizes an actual hyperbaric unit in hospitals, to the extent that the apparatus can be mounted on a microscope stage or imaged using a standalone imaging setup like a mobile camera. The clear transparent properties for imaging and visualization at a microscopic scale provide an additional advantage that does not exist with existing clear pressure vessels. The operating pressure range (up to 150 psi) of the miniaturized hyperbaric setup is at least twice that of the existing clear pressure vessels (60 psi). The demonstrated device can handle pressure generated using different gas combinations. Different inlet and outlet tubes controlled by manual stoppers help pressurize and depressurize the chamber while simultaneously operating fluids into the model device placed inside the chamber. The model microfluidic device placed inside the mini-hyperbaric chamber can be imaged using an onboard mobile camera to study the impact of pressure variation. Pyrex clear bottles were used as the chamber housing.

[0176] The micro-hyperbaric chamber was designed and fabricated with a peristaltic pump, a blood reservoir, and a battery to allow for the circulation of blood within a biomimetic system undergoing a series of pressure variations.

[0177] A planar biomimetic system representing the human alveolar system with microchannels allowing gas and blood exchange across a cylindrical-shaped barrier (see Figs. 19 and 20A-20B). This barrier follows a similar architecture as the alveolar sacs, with dimensions resembling the actual microvasculature.

[0178] A multilayer biomimetic system that allows for the 3D study of gas exchange across the alveolar and vascular networks was also fabricated (see Fig. 21A-21 B). The first layer consists of a simulation of the bronchiole, alveolar duct and alveolar sac, while the second layer is comprised of a capillary network with simple bifurcations. Both layers are separated by a vertical air-blood barrier at varying thicknesses and pore distributions.

[0179] The microvascular designs were created using Fusion 360 software and printed on a mylar sheet to produce a photomask, which was manufactured by CAD/Art Services, Inc. A layer of SU-8 negative photoresist 2050 with a thickness of 40 pm from Kayaku Advanced Materials, Inc. was coated onto 4-inch silicon wafers obtained from Alpha Nanotech Inc., using a spin coater (Laurell Technologies Corporation, model WS- 650-23 B). The silicon wafer, photomask alignment, and UV exposure processes were carried out using an OAI Hydralign Series 200 mask aligner system. The spin coater was used to apply the photoresist layer at a speed of 3600 rpm. Subsequently, the spin-coated silicon wafer was baked at temperatures of 65 and 95 degrees Celsius for 3 and 6 minutes, respectively. Afterwards, the photomask was positioned on the spin-coated silicon wafer, and the combined structure was exposed to UV light for 55 seconds at an intensity of 3.1 mW cm-2. Following the UV exposure, the post-baking procedure was performed at 65 and 95 degrees Celsius for 1 and 6 minutes, respectively. The unexposed section of the photoresist layer was dissolved using SU-8 developer from Kayaku Advanced Materials, Inc., with a development time of 1 minute. The thickness of the developed photoresist layer was assessed using a Bruker Dektak XT contact profilometer. Finally, the silicon wafer underwent a 24-hour surface treatment with chlorotrimethylsilane to enhance the bonding between the mineral and organic components at the interface.

[0180] To fabricate the microfluidic channels, a mixture of polydimethylsiloxane (PDMS) and curing agent (Sylgard Silicone elastomer 184, Dow Corning Corp., Midland, Ml.) was combined in a weight ratio of 10:1 . The resulting PDMS mixture was poured onto the fabricated master and subjected to degassing to eliminate any trapped air bubbles. The curing process took place overnight in an oven at a temperature of 60 degrees Celsius. Once the curing was complete, the PDMS containing the microchannel networks was separated from the silicon wafer, and holes with a diameter of 1 .5 mm were created to serve as inlets and outlets.

[0181] In order to replicate the pulmonary microvasculature, it was necessary to seal the microchannels with a membrane to achieve the accurate rate of gas transport exchanges in the alveoli microvasculature (Figs. 21 A-21 B). Forthis purpose, a waterproof and breathable film commonly utilized in wound bandages (P+P Custom Medical Supplies Inc.) can be employed to fabricate the membrane. According to reports from the company, this transparent film exhibits oxygen permeability similar to that of human skin. With its adhesive properties, the film effectively seals the microchannels, enabling the preparation of sealed microfluidic devices for use in experimental procedures.

[0182] The mini-hyperbaric chamberwas set up as shown in Fig. 22A. The microfluidic chip “a” is positioned inside of a Pyrex bottle “d”. A phone “c” is used as the imaging system and is positioned on top of the microfluidic chip to image the microfluidic channels. The pressure pump “b1 ” has a reservoir of blood “b2” and fluid connections “b3” to provide the blood to the microfluidic chip “a”. The hyperbaric chamber is connected via fluid connections “b3” to a gas inlet “b4” which is monitored by a manometer. There is also contemplated the combination of the elements of Fig. 22A inside a single apparatus. An exploded view of such an apparatus is shown in Figs. 22B and 23A. Such an apparatus was also fabricated and is shown in Fig. 23B.

Example 4: Investigating the genesis and treatment of decompression sickness in micro-hyperbaric systems.

[0183] Decompression sickness (DCS) is a type of decompression illness that arises from the sudden decrease in pressure around the human body, leading to the formation of gas bubbles in the bloodstream and tissues. These bubbles form via either heterogeneous nucleation, or diffusion from supersaturated tissues overburdened with gas content.

[0184] Hyperbaric oxygen therapy (HBOT) is the primary treatment technique that involves breathing pure oxygen in a pressurized chamber. HBOT is administered according to therapeutic compression tables, often established by military or naval protocols, providing detailed guidelines on the duration and pressure levels required.

[0185] Inert gases dissolve in biological tissue, under high-pressure conditions, at concentrations determined by their solubility. The solubility depends on the nature of the gas, the composition of the tissue (the proportion of aqueous versus lipid components), the temperature, and the partial pressure. Nitrogen is five times more soluble in fatty tissues than in blood components. If the rate of pressure reduction exceeds the rate of gas elimination, the dissolved gases will form bubbles by either emerging from solution or diffusing into the blood vessels (tissue supersaturation). Decompression theory explores the complex interplay of factors that include gas solubility, partial pressure, diffusion, and bubble mechanics within living tissues.

[0186] Three microfluidic devices are designed at biologically relevant diameters and geometries, adhering to Murray’s law of bifurcation (Figs. 24A-24C).

[0187] A synthetic blood solution was prepared and stored in a custom-designed and 3D-printed blood reservoir. This reservoir fed the working fluid to a peristaltic pump, chosen for its ability to simulate the pulsatile nature of blood flow.

[0188] A custom-built motor hat was mounted directly on top of the pump, powered by four lithium-ion 18650 batteries.

[0189] A 2L clear vessel was used to establish a controllable environment with gas intake from either a nitrogen tank or an air nozzle, introduced at controlled pressures of 1 bar and 3 bars. [0190] Still images were acquired using an inverted fluorescence confocal microscope and analyzed using a custom-developed MATLAB code (Figs. 25, 23A and 23B).

[0191] Microscale hyperbaric systems offer a novel approach to understanding and exploring decompression illness in a controlled and miniaturized setting. These systems provide precise control over pressure and decompression rates, enabling the replication of conditions that induce DCS in real life.

[0192] PDMS-based devices were used to simulate human tissues undergoing compression with and without the synthetic blood solution at 1 bar and 3 bars. The compression periods varied between 2 hours, 4 hours, 6 hours, and 12 hours. The volumes of generated bubbles were calculated using the MATLAB script and combined for both types of gases at 0 min, 10 min and 30 min post-return to standard temperature and pressure conditions (Figs. 26A-26D, 27A-27D, 28A-28D, and 29A-29D).

[0193] It was found that: 1 . Longer compression periods were associated with larger bubble volumes. 2. The risk of developing the largest and most lethal bubbles was at its peak 30 minutes after decompression, indicating that sudden removal from high pressures had lasting effects on the system. 3. Compression at 1 bar appeared to generate numerous small bubbles, but a few emerged with significantly larger volumes than the maximum volumes generated after compression at 3 bars. 4. The significant difference in calculated bubble volumes between compressed tissues and compressed systems highlights the impact of nucleation in the manifestation and progression of DCS.

[0194] While supersaturation causes dissolved gases to exceed their solubility limit and form bubbles, nucleation accelerates this process. These experiments are essential for understanding the risk factors associated with decompression sickness and for optimizing decompression protocols and gas mixtures to enhance diver safety.

Claims

WHAT IS CLAIMED IS:

1 . A system for determining a presence of a gas bubble in a blood sample, the system comprising: a microfluidic chip comprising a plurality of interconnected microfluidic channels arranged in a vascular-like configuration to mimic a given part of a vascular system, the microfluidic chip comprising a first inlet for receiving the blood sample and a second inlet for injecting gas into a flow of the blood sample to create the gas bubble in the flow within the microfluidic channels; a circulation system comprising a pump, a reservoir containing the blood sample and in communication with the pump, and a conduit in communication with the pump and/or the reservoir, the conduit directing the blood sample to the first inlet; and an imaging system configured to capture images of the microfluidic channels of the microfluidic chip, and a detection system configured to determine the presence and location of the gas bubble based on the images captured by the imaging system.

2. The system of claim 1 , wherein interconnected microfluidic channels form a plurality of bifurcations.

3. The system of claim 1 , wherein the microfluidic chip further comprising a third inlet for providing a drug.

4. The system of claim any one of claims 1 to 3, further comprising a housing to seal the microfluidic chip and create a pressurized environment forthe microfluidic chip.

5. The system of claim 4, wherein the housing is a micro-hyperbaric chamber.

6. A method of determining effectiveness of a treatment agent to reduce a volume of a gas bubble in a vascular system, the method comprising: flowing a blood sample in a microfluidic chip comprising a plurality of microfluidic channels interconnected and arranged in a vascular-like configuration to mimic a given part of the vascular system, the microfluidic chip comprising a first inlet for receiving the blood sample, a second inlet for injecting gas, a third inlet for inject the treatment agent, and an outlet; injecting the gas in the second inlet to create the gas bubble in the blood sample within microfluidic channels; injecting the treatment agent in the third inlet; and imaging the microfluidic chip; and based on the image, determining one or more of: a location of the gas bubble in the microfluidic channels; a volume of the gas bubble; and a change in volume of the gas bubble.

7. The method of claim 6, wherein the blood sample is selected from patient blood or artificial blood.

8. A method of determining a parameter at which formation of a gas bubble occurs in a vascular system, the method comprising: flowing a blood sample in a microfluidic chip comprising a plurality of microfluidic channels interconnected and arranged in a vascular-like configuration to mimic a given part of the vascular system; and subjecting the microfluidic chip to a pressure variation; imaging the microfluidic chip; and based on the imaging, determining the parameter at which formation of a gas bubble occurs in the blood sample within the microfluidic channels.

9. The method of claim 8, wherein the pressure variation comprises subjecting the microfluidic chip to an increase in pressure followed by a decrease in pressure.

10. The method of claim 9, wherein the pressure variation is applied by placing the microfluidic chip in an environment that increases the pressure and then decreases the pressure.

11. The method of claim 8, wherein the pressure variation is performed by compressing the microfluidic chip.

12. The method of any one of claims 8 to 11 , wherein the parameter is one of, or a combination of, time and pressure.

13. A method of performing hyperbaric oxygen therapy for a patient in need thereof having a gas bubble in their bloodstream, the method comprising: subjecting the patient to a given program of oxygen and pressure in a hyperbaric chamber; concurrently with the subjecting, flowing a blood of the patient in a microfluidic chip placed inside a second hyper baric chamber having oxygen and pressure corresponding to that of the hyperbaric chamber, the microfluidic chip comprising a plurality of microfluidic channels interconnected and arranged in a vascular configuration that mimics a vascular system of the patient and comprising a micro gas bubble in the microfluidic channels mimicking the gas bubble in the bloodstream of the patient; and monitoring the micro gas bubble in the microfluidic channels and ceasing the hyperbaric oxygen therapy within the hyperbaric chamber when the micro gas bubble in the microfluidic chip is eliminated.

14. The method of claim 13, further comprising before placing the patient in the hyperbaric chamber, performing a magnetic resonance imaging on the patient to determine a volume of the gas bubble.

15. The method of claim 13 or 14, wherein the microfluidic chip is selected among an array of microfluidic chips based on having the vascular configuration that best mimics the vascular system of the patient where the gas bubble is present.

16. A method of treating a gas embolism in a patient in need thereof, the method comprising administering to a bloodstream of the patient a therapeutically effective amount of a pharmaceutical composition comprising a surfactant and a pharmaceutically acceptable excipient.

17. The method of claim 15, wherein the pharmaceutical composition further comprises perfluorocarbons.

18. Use of a pharmaceutical composition comprising a surfactant and perfluorocarbons for treating a gas embolism in a subject need thereof.

19. A method of monitoring formation of bubbles in a surgical procedure, the method comprising flowing blood of a patient through a microfluidic chip comprising a plurality of interconnected microfluidic channels arranged in a vascular-like configuration to mimic a given part of a vascular system where the surgical procedure is performed, and monitoring the formation of gas bubbles in the microfluidic channels.

20. A method of detecting formation of gas bubbles in a vascular system, the method comprising: applying a compressive force a microfluidic chip to compress the microfluidic chip, the microfluidic chip comprising a plurality of microfluidic channels interconnected and arranged in a vascular-like configuration to mimic a given part of the vascular system; releasing the compressive force on the microfluidic chip; after the releasing, flowing a blood sample in the microfluidic channels of the microfluidic chip; and imaging the microfluidic chip to determine whether gas bubbles have formed in the microfluidic channels.

1. A method of detecting formation of gas bubbles in an individual with a wearable device, the method comprising: flowing a blood sample in a microfluidic chip inside the wearable device, the microfluidic chip comprising a plurality of interconnected microfluidic channels arranged in a vascular-like configuration to mimic a given part of a vascular system; and imaging the microfluidic channels to determine when a gas bubble forms in the microfluidic channels; and emitting an alert signal to the individual when the gas bubbles are detected in the wearable device.