Enhanced polypropylene hollow fiber membrane and application thereof in preparation of in-vivo oxygenation membrane and in-vivo hemodialysis membrane

Abstract

The invention discloses an enhanced polypropylene hollow fiber membrane and application thereof in preparation of an in-vivo oxygenation membrane and an in-vivo hemodialysis membrane, and belongs to the technical field of biological medicine. The preparation method of the hollow fiber membrane comprises the following steps: carrying out forced air drying on polypropylene and SiO2 powder; homogenizing the soybean oil and SiO2 through a high-speed shearing mixer, and carrying out ultrasonic treatment; adding polypropylene powder, homogenizing through a high-speed shearing mixer, adding the obtained spinning suspension into a double-screw extruder, extruding through an annular spinneret plate, and uniformly coating the outer surface of the braided tube with the spinning suspension; and cooling the braided tube coated with the spinning suspension in a coagulating bath, winding the braided tube on a winding roller, soaking the braided tube in dichloromethane, and fully extracting soybean oil to obtain the enhanced polypropylene hollow fiber membrane which is used for preparing an in-vivo oxygenation membrane and has good in-vitro blood compatibility, tissue compatibility and in-vivo biological safety performance. And the material is used for an in-vivo hemodialysis membrane and has good material exchange performance and operation stability.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to an enhanced polypropylene hollow fiber membrane and its application in the preparation of in vivo oxygenation membranes and in vivo hemodialysis membranes. Background Technology

[0002] In clinical practice, hypoxia due to pulmonary failure is common, especially in severe pneumonia. Even with mechanical ventilation, many patients remain critically ill and require life support via vein-to-vein (VV-ECMO) to promote lung recovery or await lung transplantation. VV-ECMO involves drawing the patient's blood from the body, oxygenating it outside the body via an oxygenation membrane, and then reintroducing it into the body via a pump. However, VV-ECMO is prone to complications, such as vascular obstruction due to coagulation disorders, internal organ bleeding, myocardial dysfunction due to uncoordinated contractions, hemolysis and renal insufficiency caused by mechanical damage to red blood cells, and infections. Furthermore, the oxygenation membrane has a short lifespan and requires frequent replacement, increasing costs. The fundamental reason for this is that VV-ECMO alters circulatory mechanics by drawing the patient's blood outside the body, and ex vivo blood is more susceptible to coagulation disorders. Additionally, the pump can damage blood cells. Therefore, finding a disruptive alternative to VV-ECMO is of great significance.

[0003] In 2024, the total number of patients with end-stage renal disease (ESRD) worldwide was approximately 10.502 million, with hemodialysis patients accounting for about 60%-70%. Currently, hemodialysis involves drawing blood from the patient's body, passing it through a dialysis membrane to remove waste products, and then pumping it back into the body. Therefore, it has many complications, such as vascular obstruction due to coagulation disorders, intra-organ bleeding, cardiac dysfunction, hemolysis caused by infection and mechanical damage to red blood cells, and renal insufficiency. The root cause is that the dialysis process alters circulatory dynamics by removing the patient's blood from the body, and blood outside the body is more prone to coagulation disorders. Additionally, the pump can easily damage blood cells. Traditional hemodialysis requires specialized medical facilities, specialized dialysis machines, and professional medical personnel, consuming significant medical resources, being complex, costly, and inconvenient. Therefore, finding a portable, internal hemodialysis technology to replace current external hemodialysis technology is of great significance. Summary of the Invention

[0004] Based on this, the main objective of the present invention is to provide an enhanced polypropylene hollow fiber membrane that has good in vitro blood compatibility, tissue compatibility and in vivo biosafety performance.

[0005] Another objective of this invention is to provide the application of the reinforced polypropylene hollow fiber membrane in the preparation of an intracorporeal oxygenation membrane (ICMO). ICMO involves directly oxygenating blood within the patient's body by directly introducing oxygen into the patient's vena cava. The hypoxic blood in the vena cava is transformed into hyperxic blood through the action of the oxygenation membrane, achieving the purpose of oxygenation. This intracorporeal oxygenation technology is completely different from ECMO technology, not only fundamentally changing the challenges of blood outside the body and the way blood is oxygenated, but also altering hemodynamics, the mechanisms of action of blood cells, and the oxygenation membrane.

[0006] Another objective of this invention is to provide the application of the reinforced polypropylene hollow fiber membrane in the preparation of in vivo hemodialysis membranes. In vivo hemodialysis membranes allow for direct dialysis of blood within the patient's body, which is completely different from extracorporeal dialysis membrane technology. This not only fundamentally changes the challenges of blood removal from the body and the method of hemodialysis, but also alters the dynamics of hemodynamics, blood cells, and the interaction of the dialysis membrane. It requires only a membrane and dialysis fluid, eliminating the need for a dialysis machine. The technology is easy to assemble, portable, low-cost, simple to operate, and has a wide range of applications.

[0007] To achieve the above objectives, the present invention adopts the following technical solution:

[0008] In a first aspect, the present invention provides a method for preparing an enhanced polypropylene hollow fiber membrane, comprising the following steps:

[0009] (1) Polypropylene and SiO2 powder were dried by forced air drying;

[0010] (2) Soybean oil and SiO2 were homogenized by a high-speed shear mixer and then ultrasonically treated for 30 minutes;

[0011] (3) Add polypropylene powder and homogenize it by a high-speed shear mixer to obtain a uniform and stable spinning suspension;

[0012] (4) The spinning suspension is added to a twin-screw extruder and extruded through an annular spinneret to uniformly coat the outer surface of the braided tube.

[0013] (5) The braided tube coated with spinning suspension is cooled in a coagulation bath and wound on a winding roller. It is then soaked in dichloromethane to fully extract soybean oil, thus obtaining an enhanced polypropylene hollow fiber membrane.

[0014] Preferably, in step (1), the mass ratio of polypropylene to SiO2 powder is 10-30:1.5.

[0015] Preferably, in step (2), soybean oil and SiO2 are used in a mass ratio of 68.5-88.5:1.5.

[0016] Preferably, in steps (4) and (5), the parameters of the twin-screw extruder include: spinning temperature of 220°C; coagulation bath of water; air gap of 35 cm; and extrusion rate of 35 r / min. -1 The curling rate is 17.5 m / min. -1 .

[0017] A second aspect of the invention provides the use of reinforced polypropylene hollow fiber membranes in the preparation of in vivo oxygenation membranes (ICMOs).

[0018] A third aspect of the present invention provides the use of reinforced polypropylene hollow fiber membranes in the preparation of in vivo hemodialysis membranes.

[0019] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0020] I. Compared with ECMO membranes, reinforced polypropylene hollow fiber membranes have good characterization properties for use as in vivo oxygenation membranes. They have good oxygen-carbon dioxide exchange performance and operational stability in in vivo and in vitro venous models, and also have good in vitro blood compatibility, tissue compatibility and in vivo biosafety performance, which preliminarily verifies the feasibility of using them as in vivo oxygenation membranes.

[0021] II. Compared with extracorporeal dialysis membranes, reinforced polypropylene hollow fiber membranes have good characterization properties for in vivo hemodialysis membranes. They have good material exchange performance and operational stability in extracorporeal vena cava models and in vivo pig models, which preliminarily verifies their feasibility for use as in vivo hemodialysis membranes. Attached Figure Description

[0022] Figure 1 This is a process flow diagram for preparing the reinforced polypropylene hollow fiber membrane in the example.

[0023] Figure 2 The SEM images of reinforced polypropylene hollow fiber membranes with different contents in the examples are shown below: (a) cross section; (b) view of the separation layer; (c) magnified view of the separation layer surface; (d) canopy view.

[0024] Figure 3 The pore size distribution of reinforced polypropylene hollow fiber membranes with different polypropylene contents is shown in the examples.

[0025] Figure 4 The porosity of the reinforced polypropylene hollow fiber membranes with different polypropylene contents in the examples is shown.

[0026] Figure 5 Examples of reinforced polypropylene hollow fiber membranes with different polypropylene contents: (a) water contact angle; (b) inlet water pressure.

[0027] Figure 6Mechanical properties of reinforced polypropylene hollow fiber membranes with different polypropylene contents in the examples: (a) stress-strain curves; (b) burst strength.

[0028] Figure 7 The diagrams illustrate venous-venous extracorporeal membrane oxygenation (VV-ECMO) and intravenous intracorporeal membrane oxygenation (ICMO) in the embodiments. (a) The principle of VV-ECMO: A blood pump draws blood from the vena cava out of the body, oxygenates it through an extracorporeal membrane oxygenator, and then sends the blood back into the body, i.e., the blood is oxygenated outside the body; (b) The principle of ICMO: An oxygenated membrane is placed directly in the vena cava, and oxygen is directly exchanged with the blood in the vena cava inside the body to oxygenate it, i.e., the blood is oxygenated inside the body.

[0029] Figure 8 The following are schematic diagrams of the ICMO membrane in the in vitro intravenous oxygen / carbon dioxide exchange model in the examples: (a) Schematic diagram of the in vitro intravenous oxygen / carbon dioxide exchange model; (b) Exchange of oxygen / carbon dioxide in the test tube through membrane osmosis; (ch) Oxygen / carbon dioxide exchange experiment in the model; (c) Physical image of the ICMO membrane; (d) Membrane installation position; (e) Position of the inlet and outlet pipes; (f) Experiment when the test tube is filled with Ringer's solution; (g and h) Experiment when the test tube is filled with piglet blood (g is 0% of the oxygen flow rate). mL / s, h is 2mL / s); (il) Long-term operational stability experiment in the model; (i) Water (ion) leakage experiment, after leakage the inlet tube (white arrow) and outlet tube (obvious water droplets, bubbles, black arrow); (j) Plasma leakage experiment, after leakage the inlet tube (white arrow) and outlet tube (obvious yellow foam droplets, bubbles, black arrow); (k) Pore blockage experiment with visible thrombus clumps on the surface, which are not easily removed after shaking, the thrombus clumps block less than 1 / 2 of the membrane surface area; (l) More than 1 / 2 of them are blocked by thrombus.

[0030] Figure 9 The following are schematic diagrams of the ICMO membrane in the piglet ARDS model in the examples: (a) Schematic diagram of the experiment in piglets; (b) Oxygen / carbon dioxide exchange process of ICMO membrane in the vena cava; (c) The process of ICMO membrane being inserted into the vena cava of piglets through the right femoral vein; (d) After ICMO modeling is completed, the two ends of the membrane are connected to the inlet tube (white arrow) and the outlet tube (black arrow) respectively; (e) B-line of the lungs can be seen on chest ultrasound; (f) Ultrasound scan shows that the membrane is located in the vena cava (white arrow); (g) Side view of abdominal plain X-ray, showing the membrane in the vena cava (white arrow) (Iohexol is first injected into the membrane for contrast, and then the contrast agent is expelled by the exhaust method); (h) Anterior and posterior body X-rays with the membrane (white arrow).

[0031] Figure 10The following are oxygen / carbon dioxide exchange curves in in vitro and in vivo models in the examples: (a) During oxygen exchange, the relationship between oxygen flow rate and oxygen partial pressure after ventilation in Ringer's solution or blood in test tubes or blood in the vena cava of piglets; (b) During carbon dioxide exchange, the relationship between carbon dioxide partial pressure before and after ventilation in Ringer's solution or blood in test tubes or blood in the vena cava of piglets.

[0032] Figure 11 This is a schematic diagram of an in vivo hemodialysis membrane model in an in vivo and external venous cavity, as shown in the embodiment.

[0033] Figure 12 This is a schematic diagram of the in vivo hemodialysis membrane in the extracorporeal venous substance exchange model in the embodiment.

[0034] Figure 13 This is a schematic diagram of the in vivo hemodialysis membrane in a piglet renal failure model, as shown in the embodiment.

[0035] Figure 14 The results of in vitro blood compatibility verification in the examples are shown in Figure 1; a is whole blood clotting time; b is plasma recalcification time; c is the quantitative result of relative hemolysis rate; and d is platelet adhesion density.

[0036] Figure 15 The cell SEM images in the example are: a, b, c are glass slides, d, e, f are ECMO membranes, g, h, i are ICMO membranes (a, d, g × 300x, b, e, h × 600x, c, f, i × 1500x).

[0037] Figure 16 The images shown are of live cells (green) and dead cells (red) stained with calcein AM / PI after 3 days of culture in the example; a, b, c are glass slides, d, e, f are ECMO membranes, g, h, i are ICMO membranes (a, d, g are dead cells, b, e, h are live-dead cells, c, f, i are live cells).

[0038] Figure 17 The following are diagrams illustrating the implantation of ECMO and ICMO membranes into the backs of rats in this embodiment: a) before implantation; b) after implantation; c) after implantation; d, e, and f are the ECMO and ICMO membrane specimens: d before implantation, e 2 weeks after implantation, and f 1 month after implantation. Detailed Implementation

[0039] To more fully understand and demonstrate the technical solutions, objectives, and advantages of the present invention, the technical effects produced by the present invention will be further described in detail and completely below with reference to the accompanying drawings and specific embodiments. Obviously, the described embodiments are only a part of the embodiments of the present invention, and not all of them. It should be noted that other embodiments obtained by those skilled in the art without departing from the concept of the present invention are all within the protection scope of the present invention.

[0040] The following embodiments present an in vivo oxygenation membrane (ICMO) technology, which directly oxygenates blood within the patient's body by directly introducing oxygen into the patient's vena cava. The hypoxic blood in the vena cava is transformed into hyperxic blood through the oxygenation membrane, achieving oxygenation. This in vivo oxygenation technology is completely different from current ECMO technology: it not only fundamentally changes the challenges of blood outside the body and the way blood is oxygenated, but also alters hemodynamics, the mechanisms of action of blood cells and the oxygenation membrane, such as… Figure 7 As shown. An in vivo reinforced polypropylene hollow fiber oxygenation membrane was prepared, and its oxygen-carbon dioxide exchange performance and operational stability, as well as its in vitro blood compatibility, tissue compatibility, and in vivo biosafety were verified in in vivo and in vitro venous models.

[0041] The following embodiments present an in vivo hemodialysis membrane technology, which allows for direct dialysis of blood within the patient's body via a dialysis membrane. It requires only a membrane and dialysate, eliminating the need for a dialysis machine. The technology is easy to assemble, portable, low-cost, simple to operate, and has a wide range of applications. This in vivo dialysis membrane technology differs from current extracorporeal dialysis membrane technologies: it not only fundamentally changes the challenges of blood removal from the body and the method of hemodialysis, but also alters hemodynamics, the interaction of blood cells, and the dialysis membrane, such as… Figure 11 As shown, an enhanced polypropylene hollow fiber dialysis membrane for in vivo application was prepared, and its mass exchange performance and operational stability were verified in an in vitro vena cava model and an in vivo pig model.

[0042] Example 1: Preparation of reinforced polypropylene hollow fiber membrane

[0043] 1. Experiment

[0044] 1.1 Materials

[0045] Polypropylene (K8303) was purchased from Yanshan Petrochemical Co., Ltd.; polyester (PET) braided tubing was purchased from Tianjin Boanxin Co., Ltd.; soybean oil was purchased from Yihai Kerry Arawana Holdings Limited; SiO2 was purchased from Shandong Sailike New Materials Co., Ltd.; and dichloromethane was supplied by Macklin.

[0046] 1.2 Preparation of reinforced polypropylene hollow fiber membrane

[0047] like Figure 1The diagram illustrates the spinning process of a reinforced polypropylene hollow fiber membrane combining braided tube reinforcement and thermally induced phase separation (TIPS). First, polypropylene and SiO2 powders are dried in a blower. Second, soybean oil and SiO2 are homogenized using a high-speed shear mixer according to the ratios in Table 1, followed by ultrasonic treatment for 30 minutes. Subsequently, polypropylene powder is added according to the ratios in Table 1 and homogenized again using a high-speed shear mixer to obtain a uniform and stable spinning suspension. The obtained spinning suspension is fed into a twin-screw extruder, the parameters of which are shown in Table 2. The uniform spinning suspension is extruded through an annular spinneret and uniformly coated onto the outer surface of the braided tube. The braided tube coated with the spinning solution is cooled in a coagulation bath and wound on a winding roller. After immersion in dichloromethane to fully extract the soybean oil, a reinforced polypropylene hollow fiber membrane is obtained.

[0048] Table 1: Composition of spinning suspension

[0049] Membrane PP (wt%) <![CDATA[SiO2(wt%)]]> Soybean oil (wt%) PP-10 10 1.5 88.5 PP-15 15 1.5 83.5 PP-20 20 1.5 78.5 PP-25 25 1.5 73.5 PP-30 30 1.5 68.5

[0050] Table 2: Spinning parameters for reinforced polypropylene hollow fiber oxygen film

[0051]

[0052] 1.3 Characterization

[0053] The surface and cross-sectional morphology of the reinforced polypropylene hollow fiber membrane samples were observed using a scanning electron microscope (SEM, GeminiSEM 300, Germany); pore size and distribution were measured using a pore size analyzer (BSD-PB, Beshine Instrument, China). Furthermore, the N2 flux was calculated based on the aforementioned pressure-flow curves; membrane porosity was determined using a wet-dry method; and the static contact angle of the membrane surface was measured using an automatic tilt contact angle measuring instrument (SDC-350H, Dongguan Shengding Precision Instruments Co., Ltd., China). Each sample was measured five times at different locations to obtain the average value. The mechanical properties of the reinforced polypropylene hollow fiber membrane were measured using a tensile testing machine (JBWL-200, Yangzhou Jingbo Precision Machinery Co., Ltd., China). The initial distance and tensile speed were set to 100 mm and 50 mm / min, respectively. -1 The rupture strength of the membrane was tested using a homemade device equipped with a pressure gauge and a nitrogen cylinder. The pressure applied to the membrane was slowly increased until the first rupture occurred; at this point, the pressure was the membrane's rupture strength.

[0054] 2. Results

[0055] 2.1 Membrane morphology and structure

[0056] like Figure 2As shown in Figure a, the interwoven fiber structure of the PET braided tubing is clearly visible. The spinning solution penetrates into the braided tubing and combines with the reinforcing material to form a uniform separation layer. Figure 2 b shows that due to the increase in melt viscosity, the layer thickness increases with the increase in PP content in the spinning solution. For example... Figure 2 As shown in Figure c, with increasing PP content, the spherulitic structure in the cross-section becomes larger and denser. The increased PP content in the spinning solution not only causes the macromolecular chains to intertwine more tightly but also binds the macromolecules together more tightly. The supramolecular structure in the solution increases dramatically with increasing PP content, forming a complex network structure. Furthermore, the direct contact between the outer surface and the coagulation bath (water) results in a rapid cooling rate, which prevents the PP molecules in the spinning solution from aligning themselves across the membrane's cross-section. The supramolecular structure leads to the formation of an irregular, dense layer and porous structure on the membrane surface. Figure 2 As shown in Figure d, the pore size on the membrane surface gradually decreases with increasing PP content. This trend is due to the reduction in the volume of the diluent phase caused by the increased PP content, which inhibits its coarsening, while simultaneously increasing the viscosity of the spinning suspension. Therefore, the pore size on the membrane surface gradually decreases.

[0057] 2.2 Pore size distribution, porosity, and gas flux

[0058] Figure 3 The pore size distribution of the reinforced PP hollow fiber membrane is shown. When the PP content is 10, 15, 20, 25, and 30 wt%, the average pore sizes are 0.36, 0.11, 0.10, 0.09, and 0.08 μm, respectively. With increasing PP content, the pore size distribution of the system gradually decreases, while the viscosity increases significantly. The increased viscosity leads to a decrease in the volume fraction of the soybean oil (diluent) phase during phase separation. The initial droplet size decreases, and the distribution becomes more uniform. Simultaneously, the high viscosity inhibits droplet coarsening, resulting in the formation of small pores and a concentrated distribution.

[0059] like Figure 4 As shown, the porosities were 47.63%, 37.25%, 34.67%, 32.17%, and 29.63% when the PP content was 10, 15, 20, 25, and 30% by weight, respectively. The porosity decreased significantly with increasing PP content. In the TIPS process, the volume fraction of the diluent phase (pore precursor) decreased during phase separation as the diluent content decreased, leading to an increase in pore nucleation density and decreased connectivity. Simultaneously, the viscosity-dominated coarsening inhibition further limited pore expansion, ultimately resulting in a significant decrease in the overall membrane porosity. With increasing PP content, the N2 flux decreased from 327 mL / min to 73 mL / min. -1 cm -2 bar -1 The trend of change is consistent with the pore size distribution and porosity.

[0060] 2.3 Water contact angle and inlet pressure

[0061] like Figure 5 As shown in Figure a, the water contact angles of the prepared membranes at different PP contents were 107°, 115°, 118°, 120°, and 121°. The water contact angle gradually increased with increasing PP content. Figure 5 As shown in b, the feed water pressures of the prepared membranes were 0.03, 0.21, 0.30, 0.42, and 0.45 MPa, respectively. The membrane with a PP content of 10 wt% exhibited poor film-forming performance due to its low content. Large pores permeated the surface of the obtained membrane, allowing water droplets to penetrate the membrane at relatively low pressure. With increasing PP content, the feed water pressure showed a significant improvement due to the increase in water contact angle and the decrease in pore size.

[0062] 2.4 Mechanical properties

[0063] Figure 6 Figure a shows the stress-strain curves of reinforced PP hollow fiber membranes. Tensile strength and strain do not change significantly with PP content. Tensile strength can reach 160 MPa, and strain is approximately 40%, mainly depending on the reinforcement of the PET fiber braided tube. Figure 6 As shown in b, the burst strengths for different PP contents were 0.04, 0.15, 0.18, 0.19, and 0.20 MPa, respectively. The burst strength increased significantly with increasing PP content. During spinning, the PP melt completely penetrated into the braided tube, forming excellent interfacial bonding. Furthermore, higher PP concentrations promoted interchain entanglement and the formation of complex polymer networks, thereby enhancing the burst strength.

[0064] Example 2: Validation of in vivo oxygenation model

[0065] (1) Establishment of an in vitro intravenous oxygen-carbon dioxide exchange model

[0066] An ICMO membrane (i.e., an in vivo reinforced polypropylene hollow fiber oxygen membrane, with a single strand outer diameter of 2.2 mm and an inner diameter of 1.24 mm, prepared in Example 1) was placed inside a 10 cm long test tube. The membrane's inlet tube was connected in sequence to a valve, a pressure gauge, a flow meter, and an oxygen cylinder. The test tube's liquid inlet tube was connected in sequence to a valve and a liquid bag, and the liquid outlet tube was connected in sequence to a valve, a liquid pump, and a liquid bag. Figure 7 Under osmosis, oxygen diffuses through the membrane into the Ringer's solution or blood in the test tube, increasing its oxygen content; conversely, carbon dioxide in the Ringer's solution or blood diffuses back through the membrane under osmosis, causing it to be expelled, thus decreasing the carbon dioxide concentration in the Ringer's solution or blood. This establishes an in vitro venous oxygen-carbon dioxide exchange model. The piglet blood animal experimental protocol used was approved by the Animal Ethics Review Committee of Shanghai First People's Hospital.

[0067] (2) Gas exchange experiment in an in vitro intravenous oxygen-carbon dioxide exchange model

[0068] 2.1 O2 Exchange Experiment

[0069] In an in vitro intravenous oxygen-carbon dioxide exchange model ( Figure 8 In this experiment, Ringer's solution was continuously introduced into the inlet tube of the test tube (flow rate 0.5 mL / s), while O2 (100% oxygen content, flow rates of 0, 2, 4, 6, 8, and 10 mL / s) was introduced through the air inlet tube. After the oxygen passed through the ICMO membrane, the partial pressure difference of O2 in the Ringer's solution flowing into and out of the test tube was measured at 1 minute. Each measurement was taken three times, and the average value was taken. An O2 exchange curve was plotted with the partial pressure difference of O2 in the Ringer's solution as the ordinate and the oxygen flow rate as the abscissa. Blood (heparin anticoagulation, flow rate 0.5 mL / s) was introduced into the inlet tube, and the above experiment was repeated.

[0070] 2.2 CO2 Exchange Experiment

[0071] In the extracorporeal intravenous oxygen-carbon dioxide exchange model, O2 (100% oxygen content, flow rate 2 mL / s) was introduced through the inlet, and Ringer's solution (flow rate 0.5 mL / s, carbon dioxide partial pressure set at 0, 2, 4, 6, 8, 10, ×10) was continuously introduced through the inlet tube. 3 The partial pressure difference of CO2 in Ringer's solution was measured between the inlet and outlet tubes. Each measurement was taken three times, and the average value was calculated. A CO2 exchange curve was plotted with the partial pressure difference of CO2 in the inlet and outlet tubes as the ordinate and the partial pressure of carbon dioxide in the Ringer's solution as the abscissa. Blood (heparin anticoagulation, flow rate 0.5 mL / s) was introduced into the inlet tube of the test tube, and the above experiment was repeated.

[0072] (3) Long-term operational stability experiment in an in vitro intravenous oxygen-carbon dioxide exchange model

[0073] 3.1 Ion Leakage Experiment

[0074] In an in vitro intravenous oxygen-carbon dioxide exchange model, O2 (100% oxygen content, flow rate 2 mL / s) was introduced into the inlet, and the initial gas pressure P0 at the inlet was measured. Ringer's solution was introduced into the liquid inlet (flow rate 0.5 mL / s), and the gas pressure P1 at the inlet was measured. The appearance of water droplets at the inlet and outlet was observed. If water droplets or bubbles appeared, it indicated leakage of water or ions (ions dissolved in water) inside the membrane; at this time, the inlet gas pressure P2 was measured. The time from the start of aeration to the appearance of water droplets was the ion leakage time (h). The above experiment was repeated three times, and the average value was taken.

[0075] 3.2 Plasma Leakage Experiment

[0076] In an extracorporeal intravenous oxygen-carbon dioxide exchange model, O2 (100% oxygen content, flow rate 2 mL / s) was introduced through the inlet, and the initial inlet pressure P0 was measured. Blood (heparin anticoagulated, flow rate 0.5 mL / s) was introduced through the inlet, and the inlet pressure P1 was measured. Every 24 hours, both membranes were removed and gently rinsed three times with physiological saline to remove surface clots. The presence of yellow foamy leakage at the inlet and outlet was observed. The presence of yellow foamy leakage or bubbles indicated plasma leakage within the membrane; at this point, the inlet gas pressure P2 was measured. The time from the start of ventilation to the appearance of plasma was defined as the plasma leakage time (h). The above experiment was repeated three times, and the average value was taken.

[0077] 3.3 Membrane pore blockage experiment

[0078] O2 (100% oxygen content, flow rate 2 mL / s) was introduced through the air inlet, and the initial air pressure P0 was measured. Blood (heparin anticoagulated, flow rate 0.5 mL / s) was introduced through the liquid inlet, and the air pressure P1 was measured. Every 24 hours, the membrane was removed. The membrane surface was not rinsed with saline; it was observed for visible thrombus clusters, which were difficult to remove after shaking. When more than half of the membrane surface area was blocked by thrombus clusters, the inlet gas pressure P2 was measured. The time from the start of ventilation to the membrane surface being half-blocked by thrombus was defined as the membrane pore occlusion time (h). The above experiment was repeated three times, and the average value was taken.

[0079] (4) Experiments with ICMO membrane in an in vitro venous model

[0080] (1) Gas exchange experiment

[0081] 1.1 O2 Exchange Experiment

[0082] In an in vitro intravenous oxygen-carbon dioxide exchange model, the partial pressure of oxygen in Ringer's solution or blood increased with increasing oxygen flow rate, indicating that the ICMO membrane has good oxygen permeability. At the same flow rate, the partial pressure of oxygen in Ringer's solution increased significantly more than in blood, possibly because blood has a stronger oxygen-carrying capacity, with hemoglobin in the blood binding a larger amount of oxygen, while Ringer's solution has a weaker oxygen-carrying capacity and lower oxygen content than blood. It was also found that the partial pressure of oxygen in Ringer's solution was significantly higher than that in blood, possibly because blood is directly obtained from the piglets, while oxygen in Ringer's solution dissolves naturally under standard atmospheric pressure. When the oxygen flow rate was less than 4 mL / s, the partial pressure of oxygen in blood was directly proportional to the flow rate; when the oxygen flow rate was greater than 4 mL / s, the increase in the partial pressure of oxygen in blood tended to plateau. However, the partial pressure of oxygen in Ringer's solution was directly proportional to the flow rate. Figure 10 Although the curves show a significant increase in venous oxygen partial pressure in the in vitro model compared to the in vivo model in pigs, the two models cannot be directly compared due to differences in membrane length, venous blood flow velocity, and total volume.

[0083] 1.2 CO2 Exchange Experiment

[0084] In an extracorporeal intravenous oxygen-carbon dioxide exchange model, as the partial pressure of carbon dioxide in Ringer's fluid or blood increases, the carbon dioxide partial pressure difference in Ringer's fluid or blood also increases, indicating that the ICMO membrane has good carbon dioxide permeability. Under the same partial pressure of carbon dioxide in Ringer's fluid or blood, the carbon dioxide partial pressure difference in Ringer's fluid is significantly greater than that in blood, possibly because some carbon dioxide is bound to hemoglobin in the blood and is not easily released. The carbon dioxide partial pressure difference in blood is directly proportional to the partial pressure of carbon dioxide in the blood; a carbon dioxide partial pressure less than 4 × 10⁻⁶ indicates a lower carbon dioxide partial pressure. 3 At Pa, the decrease in carbon dioxide partial pressure difference is relatively gradual; when the carbon dioxide partial pressure is greater than 4 × 10⁻⁶ Pa, the decrease is relatively gradual. 3 At a pressure of Pa, the decrease in carbon dioxide partial pressure gradient is more steep. This may be because Ringer's solution does not readily dissolve carbon dioxide, while blood has a stronger capacity to dissolve carbon dioxide.

[0085] (2) Long-term operational stability test

[0086] 2.1 Ion Leakage Experiment

[0087] In an extracorporeal intravenous oxygen-carbon dioxide exchange model, Ringer's solution was introduced into the inlet tube. When water droplets appeared at the outlet, air bubbles were observed moving back and forth in the outlet tube, indicating ion leakage in the membrane. At this time, the ICMO membrane pressure fluctuated and became unstable, rising from an initial pressure of 175.7 Pa to 359 kPa, with the ion leakage time being 354.7 hours. Ion leakage leads to abnormal concentrations of potassium, sodium, and other ions in the blood, exacerbating metabolic acid-base imbalances in patients. Ion leakage, combined with the membrane's mechanical damage under the influence of gas pressure and blood flow pressure, results in ion leakage. The Ringer's solution in this experiment contained sodium, potassium, calcium, and chloride ions, so water leakage is equivalent to ion leakage. After water (ions) enters the membrane, the membrane's cross-sectional area decreases significantly, and the ventilation resistance increases significantly. At the same gas flow rate, the inlet pressure increases significantly, and the membrane's gas pressure increases by approximately 100%.

[0088] 2.2 Plasma Leakage Experiment

[0089] When piglet blood was introduced into the inlet tube of the extracorporeal intravenous oxygen-carbon dioxide exchange model, yellow foamy leakage appeared at the outlet. Bubbles could be seen moving back and forth in the outlet tube, indicating that plasma leakage had occurred in the membrane. At this time, the pressure of the ICMO membrane fluctuated and became unstable, rising from the initial pressure of 202 Pa to 442 Pa. The plasma leakage time was 315.9 h.

[0090] Plasma leakage is a major cause of long-term membrane failure, leading to a sharp increase in transmembrane pressure (TMP) and a significant decrease in oxygenation efficiency; therefore, it is a crucial parameter for membrane performance. After plasma enters the membrane, the membrane cross-sectional area decreases significantly, and the ventilation resistance increases significantly. At the same flow rate, the inlet pressure increases significantly, approximately doubling the membrane's gas pressure.

[0091] 2.3 Membrane pore blockage experiment

[0092] In the extracorporeal intravenous oxygen-carbon dioxide exchange model, piglet blood was introduced through the inlet tube. Thrombus clumps and fibrous tissue were observed adhering to the membrane surface and were difficult to remove. When more than half of the membrane surface area was blocked by thrombus clumps, the ICMO membrane pore blockage time increased from the initial pressure of 202 kPa to 234 kPa to 263.7 h, as shown in Tables 3 and 4.

[0093] Table 1: Comparison of characteristics between VV-ECMO and ICMO

[0094]

[0095] Table 2: Long-term operational stability experiment of the in vitro intravenous oxygen / carbon dioxide exchange model (x±s, Pa, h)

[0096]

[0097] Membrane blockage is a significant cause of decreased gas permeability in membrane pores, manifested as a persistent increase in transmembrane pressure and a decline in oxygenation parameters. The main causes are insufficient anticoagulation, such as inadequate heparin dosage or monitoring failure leading to fibrin deposition, low flow rates causing blood stasis, tubing design defects triggering thrombosis, or material defects, such as a rough fiber membrane surface or poor biocompatibility.

[0098] In an extracorporeal intravenous oxygen-carbon dioxide exchange model, thrombus clumps and fibrous tissue were observed adhering to the membrane surface, which were difficult to remove. When more than half of the membrane surface area was blocked by thrombus clumps, the initial pressure differential of the ICMO membrane doubled.

[0099] Clinically, the dynamic pressure of blood pump output in VV-ECMO ranges from 13.3 to 26.6 × 10⁻⁶. 3The pressure of Pa (Pa) acting on the ECMO membrane makes blood cells more prone to rupture and tiny particles in the blood more likely to clog the membrane pores, causing the membrane's filtration efficiency to decrease over time, thus requiring frequent membrane replacement. Since the dynamic pressure in the vena cava is approximately 98 Pa, close to 0, the way the ICMO membrane functions in hemodynamics, blood cells, and oxygenation is fundamentally changed. Therefore, in the model design, the dynamic pressure of the ICMO membrane tube is almost 0 Pa, lower than the oxygen pressure passing through the ICMO membrane. This allows oxygen to pass through the nanopores, preventing blood cells and particles from adhering to the membrane, thus prolonging the membrane's filtration efficiency, reducing blood cell damage, and minimizing the likelihood of tiny particles clogging the membrane pores.

[0100] The ICMO membrane prepared in this embodiment exhibits excellent oxygen permeability and CO2 removal capabilities, and demonstrates good long-term operational stability. In terms of performance, it can completely replace VV-ECMO, providing a direction for the future treatment of patients with severe respiratory failure.

[0101] Example 3: Validation of in vivo hemodialysis membrane application

[0102] 1. Experiments on in vivo dialysis membranes in an in vitro venous substance exchange model

[0103] (1) Establishment of an extracorporeal intravenous substance exchange model

[0104] An in vivo dialysis membrane (an in vivo reinforced polypropylene hollow fiber dialysis membrane, with an outer diameter of 2.2 mm and an inner diameter of 1.24 mm per strand, prepared in Example 1) was placed inside a 50 mL syringe. The membrane inlet was connected in sequence to a valve, a flow meter, and a dialysate bag, while the outlet was connected to a waste bag. Figure 11 The syringe is connected in sequence to the valve and the dialysis fluid bag, and the outlet tube is connected in sequence to the valve, the flow meter, and the outlet fluid bag. Figure 12 a). Under osmosis, toxins or electrolytes in the syringe fluid or blood diffuse through the membrane into the dialysate, reducing their concentration. Figure 12 b). That is, to establish an extracorporeal vena cava material exchange model ( Figure 12 c, d, e).

[0105] (2) Exchange experiments in an in vitro intravenous substance exchange model

[0106] 2.1 Material Exchange Experiment

[0107] In the extracorporeal vena cava substance exchange model, dialysate was continuously infused into the syringe at a flow rate of 0.5 mL / s (Baxter Corporation), while the inlet tubing was continuously infused with 4 L of dialysate (at a flow rate of 0.5 mL / s, containing different concentrations of electrolyte cation Na+) into the syringe. + K + Ca 2+ and anion HCO2 -, lactic acid, HPO4 - The solution contained small molecules of toxins (Cr, BUN, uric acid, triglycerides, cholesterol, etc.), circulated for 4 hours, and the total volume of the final dialysate was calculated to determine the amount of water removed. The concentrations of various substances before and after dialysis were measured. Each measurement was taken 5 times, and the average value was used. A substance exchange curve was plotted with the concentration before dialysis as the ordinate and the concentration after dialysis as the abscissa. Piglet blood (heparin anticoagulated, flow rate 0.5 mL / s) was introduced into the inlet tube, and the above experiment was repeated.

[0108] 2.2 Water Exchange Experiment

[0109] In the extracorporeal vena cava substance exchange model, dialysate was continuously infused into the syringe at flow rates of 0, 0.25, 0.5, 0.75, and 1.0 mL / s (Baxter Corporation), while the flow rate of the inlet tubing was 4 L of dialysate continuously infused into the syringe at a flow rate of 0.5 mL / s (containing fixed electrolyte cation Na+). + K + Ca 2+ and anion HCO2 - , lactic acid, HPO4 - The solution contained small molecules of toxins (Cr, BUN, and uric acid), circulated for 4 hours, and the total volume of the final dialysate was calculated to determine the amount of water removed. Each data point was measured 5 times, and the average value was taken. A water exchange curve was plotted with the flow rate before dialysis as the ordinate and the water loss after dialysis as the abscissa. Piglet blood (heparin anticoagulated, flow rate 0.5 mL / s) was introduced into the inlet tube, and the above experiment was repeated.

[0110] (3) Long-term operational stability experiment in an in vitro venous substance exchange model

[0111] 3.1 Blood plasma leakage experiment

[0112] In the extracorporeal vena cava material exchange model, dialysate was continuously infused into the syringe (flow rate 0.5 mL / s, Baxter), and the inlet tubing flow rate was the same as the continuous infusion of 4 L of blood into the syringe (flow rate 0.5 mL / s, heparin anticoagulation), with cyclic dialysis. The fluid pressure P1 at the inlet was measured (the initial fluid pressure P0 was measured before insertion). Yellow foamy leakage was observed at the dialysate inlet and outlet. If yellow foamy leakage was observed, it indicated plasma leakage inside the membrane, at which point the inlet pressure P2 was measured. The time from the start of dialysis to the appearance of plasma was defined as the plasma leakage time (h). The above experiment was repeated 5 times, and the average value was taken.

[0113] 3.2 Membrane pore blockage experiment

[0114] In the extracorporeal vena cava material exchange model, dialysate (0.5 mL / s, Baxter) was continuously infused into the syringe, while the inlet tubing was continuously infused with 4 L of blood (0.5 mL / s, heparin anticoagulation) in a cyclic dialysis process. The inlet fluid pressure P1 was measured (the initial fluid pressure P0 was measured before insertion). Changes in the inlet fluid pressure were observed during dialysis. If the transmembrane pressure continuously increased (exceeding the baseline value by more than 30%), it indicated increased resistance to fluid exchange due to membrane pore blockage. When the inlet fluid pressure suddenly increased by 1 / 3, the measured value was P2, indicating membrane pore blockage. The membrane was removed; the membrane surface was not rinsed with saline, and thrombus clumps were observed. These thrombi were difficult to remove after shaking. When more than half of the membrane surface area was blocked by thrombus clumps, the time from the start of dialysis to when half of the membrane surface was blocked by thrombi was defined as the membrane pore blockage time (h). The above experiment was repeated 5 times, and the average value was taken.

[0115] 2. Gas exchange experiment using in vivo dialysis membranes in a piglet model of renal failure.

[0116] (1) Establishment of a piglet renal failure model

[0117] Ten healthy piglets, regardless of sex, aged 3-4 months, and weighing 35.5-40.1 kg, were selected. An adenine + high-phosphorus diet was used (daily adenine gavage (150-300 mg / kg) combined with a high-phosphorus diet for 8 weeks). Successful establishment of a piglet renal failure model was indicated by a serum creatinine (Scr) ≥ 2 times baseline. Figure 13 As shown.

[0118] After successful modeling, piglets were given basic anesthesia with propofol (10 mg / kg, IM), followed by anesthesia with 3% sodium pentobarbital (20-30 mg / kg, IV), and fixed on the operating table. Endotracheal intubation and mechanical ventilation were then initiated.

[0119] Locate the root of the right femoral vein, disinfect and drape, and after routine heparin anticoagulation, perform a puncture at a point approximately 2 cm from the root of the right femoral vein. Insert the dialysis membrane into the inferior vena cava (to a depth of 50 cm), connecting the inlet and outlet tubing of the dialysate to both ends of the membrane, and secure it in place. Apply the membrane and set it aside for later use. This animal experimental protocol strictly adheres to all ethical guidelines for laboratory animals at Shanghai First People's Hospital.

[0120] Observation indicators: After modeling, observe the piglets' pupil size, respiration, heart rate, and other general conditions. Perform hematological tests, including complete blood count, biochemistry, coagulation analysis, myocardial injury markers, and blood gas analysis. Use ultrasound to scan the lungs to observe for consolidation and the presence of the dialysis catheter in the vena cava; perform X-ray examinations of the lungs and abdomen to assess pulmonary consolidation and the presence of the dialysis catheter in the vena cava.

[0121] (2) Material exchange experiment of in vivo dialysis membrane in a pig model of renal failure

[0122] 2.1 Material Exchange Experiment

[0123] Dialysis fluid was continuously introduced through the inlet (flow rate 0.5 mL / s, Baxter Corporation), and dialysis was performed for 4 hours. The concentrations of various substances before and after dialysis (including the fixed electrolyte cation Na) were measured. + K + Ca 2+ and anion HCO2 - , lactic acid, HPO4 - (Toxins, small molecules Cr, BUN, and uric acid). Each data point was measured five times, and the average value was taken. A mass exchange curve was plotted with the concentration before dialysis as the ordinate and the concentration after dialysis as the abscissa.

[0124] 2.2 Water Exchange Experiment

[0125] Dialysis fluid (flow rates 0, 0.25, 0.5, 0.75, 1.0 mL / s, Baxter) was continuously introduced through the inlet. Dialysis was performed for 4 hours, and the total volume of the dialysate was calculated to determine the amount of water removed. Each data point was measured 5 times, and the average value was taken. A water exchange curve was plotted with the flow rate before dialysis as the ordinate and the water loss after dialysis as the abscissa.

[0126] (3) Long-term operational stability experiment of in vivo dialysis membrane in a piglet renal failure model

[0127] 3.1 Blood plasma leakage experiment

[0128] In a piglet renal failure model, a continuous flow of dialysate (heparin anticoagulation, flow rate 0.5 mL / s, Baxter) was used to perform dialysis on the in-vivo dialysis membrane. The inlet pressure P1 was measured (the initial pressure P0 was measured before insertion). The presence of yellow foamy leakage was observed at the dialysate inlet and outlet. If yellow foamy leakage was observed, it indicated plasma leakage within the membrane; at this point, the inlet pressure P2 was measured. The time from the start of dialysis to the appearance of plasma was defined as the plasma leakage time (h). The experiment was repeated 5 times, and the average value was taken.

[0129] 3.2 Membrane pore blockage experiment

[0130] In a piglet renal failure model, a continuous infusion of dialysate (heparin anticoagulation, flow rate 0.5 mL / s, Baxter) was performed through an in vivo dialysis membrane. The inlet fluid pressure P1 was measured (the initial fluid pressure P0 was measured before insertion). Changes in the inlet fluid pressure were observed during dialysis. A sustained increase in transmembrane pressure (exceeding the baseline value by more than 30%) indicated increased resistance to fluid exchange due to membrane pore blockage. When the inlet fluid pressure suddenly increased by 1 / 3, the measured value was P2, indicating membrane pore blockage. The time from the start of ventilation to pore blockage was defined as the pore blockage time (h).

[0131] 3. Statistical Analysis

[0132] Experimental data were analyzed using Origin 9.5 software. Results are expressed as mean ± standard deviation. One-way ANOVA was used to determine statistical significance, and Tukey's test was employed. P < 0.05 was indicated by *, and P < 0.01 by **. P < 0.05 indicated a statistically significant difference.

[0133] 4. Results

[0134] 4.1 Experiments with in vivo dialysis membranes in an in vitro venous model

[0135] (1) Exchange experiments in an in vitro intravenous substance exchange model

[0136] Material exchange experiment

[0137] In an extracorporeal venous substance exchange model, as electrolytes (K+) in the dialysate or blood are exchanged... + Na + Ca 2+ and HCO3 - HPO4 - and Lactate - The increased concentrations of small molecule toxins (creatinine, urea nitrogen, and uric acid) and lipids (triglycerides and cholesterol) led to a greater decrease in the concentration of substances being dialyzed, indicating that the in vivo dialysis membrane has good dialysis capacity. At the same concentration, the dialysate showed a significantly lower concentration of substances than the blood, possibly because blood carries more substances and has a stronger buffering capacity. Although the curves show a significant increase in venous oxygen partial pressure in the in vitro model compared to the in vivo pig model, the differences in membrane length, total blood volume, and interstitial fluid, along with the continuous production of small molecule toxins in the pigs, make a direct comparison between the two models impossible.

[0138] Water exchange experiment

[0139] In the extracorporeal venous mass exchange model, the amount of water removed from both the dialysate and blood increases with increasing dialysate flow rate, indicating that a faster dialysate flow rate effectively removes water from both the dialysate and blood. At the same dialysate flow rate, the amount of water in the dialysate is significantly less than that in the blood, possibly due to the higher colloidal molecular tension in the blood.

[0140] (2) Long-term operational stability test

[0141] Blood plasma leakage experiment

[0142] When piglet blood was introduced into the inlet tube of the extracorporeal intravenous material exchange model, the appearance of yellow foamy leakage at the outlet indicated that plasma leakage had occurred in the membrane. At this time, the fluid pressure of the dialysis membrane fluctuated and became unstable, rising from an initial fluid pressure of 202 Pa to 442 Pa. The plasma leakage time was 315.9 h (Table 5).

[0143] Ion leakage leads to abnormal concentrations of potassium and sodium ions in the blood, exacerbating metabolic acid-base imbalances in patients. Ion leakage also causes mechanical damage to the membrane under the influence of gas pressure and blood flow pressure. Plasma leakage is a major cause of long-term membrane failure, leading to a sharp increase in transmembrane pressure (TMP) and a significant decrease in dialysis efficiency; therefore, it is an important parameter of membrane performance. Generally, ion leakage occurs earlier than plasma leakage because plasma leakage only occurs after a ruptured pore and the formation of a larger pore. However, ion leakage is difficult to diagnose, so only plasma leakage is studied.

[0144] Membrane pore occlusion experiment

[0145] In an extracorporeal intravenous material exchange model, piglet blood was introduced through the inlet tube. When the in vivo dialysis membrane pressure increased from the initial 202 kPa to 234 kPa, thrombus clumps and fibrous tissue were observed adhering to the membrane surface, which were difficult to remove. When more than half of the membrane surface area was blocked by thrombus clumps, the membrane pore blockage time was 263.7 h (Table 6).

[0146] Table 5: Comparison of characteristics between extracorporeal hemodialysis and in vivo hemodialysis

[0147]

[0148] Table 6: Long-term operational stability experiment of the in vitro intravenous oxygen / carbon dioxide exchange model

[0149]

[0150] Membrane pore occlusion experiment

[0151] Membrane obstruction is a significant cause of decreased membrane permeability, manifested as a persistent increase in transmembrane pressure. The main causes are inadequate anticoagulation, such as insufficient heparin dosage or monitoring failure leading to fibrin deposition; low flow rates causing blood stasis; tubing design defects inducing thrombosis; or material defects, such as a rough fibrous membrane surface or poor biocompatibility. In in vitro venous exchange models, thrombus clumps and fibrous tissue are visible adhering to the membrane surface, which are difficult to remove. When more than half of the membrane surface area is obstructed by thrombus clumps, the initial fluid differential pressure of the in vivo dialysis membrane doubles.

[0152] This embodiment demonstrates that the in vivo dialysis membrane possesses excellent electrolyte and small molecule toxin dialysis capabilities as well as drainage capacity, exhibiting good long-term operational stability. In terms of performance, it can completely replace extracorporeal hemodialysis, providing a direction for the future treatment of patients with renal failure.

[0153] 4.2 Gas exchange experiment using in vivo dialysis membrane in a piglet model of renal failure

[0154] (1) Status of experimental animal models

[0155] The piglets are in good condition, and 10 piglets completed the experiment. Abdominal X-ray showed that the dialysis membrane in the vena cava was in place, without kinking, bending, or abdominal perforation.

[0156] (2) Material exchange experiment

[0157] Material exchange experiment

[0158] In the extracorporeal intravenous substance exchange model, the decrease in dialysis concentration increases with the increase in the concentration of electrolytes, small molecule toxins, and lipids in the dialysate or blood; at the same concentration, the decrease in Ringer's solution is more significant than that in blood.

[0159] Water exchange experiment

[0160] In the extracorporeal venous substance exchange model, as the dialysate flow rate increases, the amount of water reduced in the dialysate or blood increases; at the same dialysate flow rate, the amount of water reduced in the dialysate is significantly less than that in the blood.

[0161] (3) Long-term operational stability test

[0162] Blood plasma leakage experiment

[0163] In the piglet renal failure model, when yellow foamy exudate appeared at the outlet, it indicated that plasma leakage had occurred in the membrane; at this time, the fluid pressure of the dialysis membrane fluctuated and became unstable, rising from the initial fluid pressure of 202 Pa to 442 Pa, and the plasma leakage time was 315.9 h (Table 5).

[0164] Membrane pore occlusion experiment

[0165] In a piglet model of renal failure, when the in vivo dialysis membrane pressure increased from an initial fluid pressure of 202 kPa to 234 kPa, thrombus clumps and fibrous tissue were observed adhering to the membrane surface, which were difficult to remove. When more than half of the membrane surface area was blocked by thrombus clumps, the membrane pore blockage time was 263.7 h (Table 6).

[0166] In the piglet renal failure model, X-ray examination showed that the dialysis catheter was in place, indicating successful model establishment. As the blood electrolytes (K+) in the piglets increased... + Na + Ca 2+ and HCO3 - HPO4 - and Lactate - The increased concentrations of small molecule toxins (creatinine, urea nitrogen, and uric acid) and lipids (triglycerides and cholesterol) after dialysis resulted in a greater decrease in the concentration of these substances in the piglet's blood, indicating that the in vivo dialysis membrane has good dialysis capacity. In the extracorporeal venous exchange model, the amount of water expelled from the piglet's blood increased with the increase of dialysate flow rate, indicating that a faster dialysate flow rate can effectively remove water from both the dialysate and the blood. The shorter plasma leakage time and membrane pore blockage time in piglets compared to in vitro operation during long-term operation may be related to the complex environment of piglets.

[0167] Example 4: Evaluation of in vitro blood compatibility test

[0168] This embodiment evaluates the in vitro blood compatibility and anticoagulant properties of slides, ECMO samples, and hollow fiber membrane samples through in vitro blood compatibility tests. Whole blood clotting time, recalcification time, relative hemolysis rate, and platelet adhesion were measured. The animal experimental protocol using New Zealand white rabbits was approved by the Animal Ethics Review Committee of Shanghai First People's Hospital.

[0169] (1) Whole blood clotting time determination

[0170] Physiological saline (negative control), glass slides (positive control), ECMO sample membrane, and hollow fiber membrane sample membrane (5 parallel samples per group) were added to 24-well plates. Activation was performed by mixing an appropriate amount of 3.8% sodium citrate rabbit blood and 0.1 mol / L CaCl2 solution at a volume ratio of 10:1. 100 μL of the activated rabbit blood was added to each of the 24-well plates containing the samples. The hemoglobin content of the supernatant in the sample wells corresponding to preheated distilled water at 37°C was measured. The sample solutions were centrifuged in EP tubes (2 × 10⁻⁶). 3 (r / min, 10 min), 200 μL of supernatant was placed in a 96-well plate and the OD value of each sample was measured at 540 nm at 5, 15, 25, 35 and 45 min. The whole blood coagulation time curve was plotted with OD value as the ordinate and time as the abscissa.

[0171] like Figure 14 As shown in Figure a, the absorbance of the supernatant was measured at 540 nm after whole blood culture. Compared with the slide catheter group, the ECMO sample group and the hollow fiber membrane sample group showed the highest absorbance at any time point. Higher absorbance indicates a higher hemoglobin (Hb) content in the supernatant, suggesting more free blood cells, fewer coagulated blood cells, and a slower clotting rate. In summary, under the same experimental conditions, the control group (slide group) had a shorter clotting time. Therefore, ECMO samples and hollow fiber membranes can reduce blood cell adhesion and exhibit excellent anticoagulant properties.

[0172] (2) Determination of recalcification time

[0173] ECMO and hollow fiber membrane samples (5 parallel samples per group) were added to 96-well plates and exposed to PPP and TCPs containing and without CaCl2 as positive and negative controls, respectively. Rabbit blood containing 3.8% sodium citrate was also used. 3 Centrifuge at r / min for 5 min, take 100 μL of supernatant plasma and add it to each well. Then, except for the control well, add 100 μL of 0.025 mol / L CaCl2 solution to each well. After mixing, immediately place the wells in an ELISA reader and measure the OD value of each sample at 405 nm at 5, 10, 15, 20, 25 and 30 min. Plot the plasma recalcification time curve with OD value as the ordinate and time as the abscissa.

[0174] Plasma recalcification is a method for measuring the intrinsic coagulation system. Recalcification time refers to the time required for plasma to recalcify after the calcium source has been removed and then replenished with calcium. 2+ The time required for plasma coagulation was then measured. The plasma recalcification time was compared between the ECMO sample group, hollow fiber membrane sample group, positive control group, and negative control group. The positive control group showed the fastest thrombus formation time, at 3 minutes, followed by an inflection point. The ECMO sample group and hollow fiber membrane sample group maintained low absorbance throughout, close to the negative control group, without showing an inflection point. These results indicate that the ECMO sample group and hollow fiber membrane sample group had less activation of intrinsic coagulation. For the plasma recalcification time assay, exposure to PPP and TCPs containing and without CaCl2 served as positive and negative controls, respectively (n=5, *p<0.05, **p<0.01). Figure 14 As shown in b.

[0175] (3) Relative hemolysis rate determination

[0176] Venous blood (from the left marginal ear vein of a healthy New Zealand white rabbit) was collected using a 2.7 mL plastic vacuum blood collection tube (Becton Dickinson, USA) and mixed with 3.8% sodium citrate at a ratio of 9:1 (v / v) to prepare fresh anticoagulated blood. This anticoagulated blood was then diluted with physiological saline at a ratio of 4:5 (volume ratio) to prepare diluted blood for later use. ECMO membrane and hollow fiber membrane samples (sterilized as described above) were rinsed three times with deionized water, cut into semi-circular pieces (10 mm in diameter), and placed in centrifuge tubes (15 mL). 10 mL of physiological saline was added to each tube. 10 mL of ultrapure water was used as a positive control, and 10 mL of PBS buffer solution was used as a negative control. All centrifuge tubes were placed in a 37°C water bath and incubated for 30 minutes. Then, 0.2 mL of diluted rabbit blood was added, and the mixture was gently shaken and incubated for another 2 hours. Then, centrifuge (1200 rpm, 5 min) and collect the supernatant in a cuvette. Measure the absorbance (OD value) using a spectrophotometer (wavelength 545 nm). The hemolysis rate is calculated as follows (Formula (2)):

[0177] Hemolysis rate = [OD] sample -OD water (positive) ] / [OD PBS (negative) -OD water (positive) ]×100% Formula (2)

[0178] The international standard for hemolysis rate of biomaterials should not exceed 5%. Hemolysis rate analysis showed that the ECMO sample group had a significantly reduced hemolysis rate of 3.249%; the hollow fiber membrane sample exhibited the best blood compatibility among all samples, with a hemolysis rate of only 2.864%. Both of these materials had hemolysis rates below 5%, classifying them as blood-safe materials. Ultrapure water and 0.9% physiological saline (NS) were used as the positive and negative groups, respectively (n=5, *p<0.05, **p<0.01). Figure 14 As shown in c.

[0179] (4) Platelet adhesion test

[0180] Venous blood (from the left marginal ear vein of a healthy New Zealand white rabbit) was collected using a plastic vacuum blood collection tube (2.7 mL, Becton Dickinson, USA) and mixed with 3.8% sodium citrate at a ratio of 9:1 (v / v). Platelet-rich plasma (PRP, 2 × 10⁻⁶) was obtained by centrifugation (1200 rpm, 10 min). 7 / mL, Plt). The platelet adhesion assay was performed as follows: Three types of 10mm diameter discs—physiological saline (negative control), glass slide membrane (positive control), ECMO sample membrane, and hollow fiber membrane—were sterilized by immersing in 75% ethanol for 15 minutes, followed by repeated rinsing with deionized water (5 times). The samples were then added to 24-well cell culture plates, with 500μL of PRP added to each well. The plates were incubated at 37°C with gentle shaking for 2 hours. Afterward, the plates were gently washed 10 times with PBS buffer to completely remove any unadhered platelets and other components. Finally, the samples were fixed in 4% paraformaldehyde solution at 4°C for 4 hours, followed by dehydration using a gradient ethanol solution method (30%, 50%, 70%, 80%, 90%, 95%, 100%), with each immersion lasting 15 minutes. After dehydration and drying in a fume hood, the samples were sputter-coated with gold, and the adhesion of platelets to the membrane surface was observed using scanning electron microscopy. In addition, a lactate dehydrogenase (LDH) kit was used to quantitatively test platelet adhesion levels.

[0181] After co-incubation with platelet-rich rabbit plasma for 2 hours, platelet adhesion density on the inner surface of the catheter was determined using a lactate dehydrogenase (LDH) kit. The platelet deposition rates on the inner surface of the ECMO and hollow fiber membrane catheters were 252 and 197 platelets / mm², respectively. 2 The platelet deposition on the slide was 1638 platelets / mm². 2 Compared to glass slides, ECMO samples and hollow fiber membrane samples showed fewer adsorbed platelets, indicating that these samples not only significantly reduced platelet deposition but also effectively inhibited platelet activation and transformation. The anti-platelet aggregation effect of the hollow fiber membrane sample material surface (n=5, *p<0.05, **p<0.01) was also significant. Figure 14 As shown in d.

[0182] The results showed that hollow fiber membranes have good anticoagulant properties in vitro.

[0183] Example 5: Evaluation of cell culture compatibility

[0184] Human umbilical vein endothelial cells (HUVECs) were cultured in a specially prepared high-glucose medium (DMEM), 10% fetal bovine serum, and 1% penicillin / streptomycin. Before cell seeding, ECMO membranes and hollow fiber membranes were cut into 10 mm diameter discs and placed one disc into a 24-well plate, then covered with a sterile stainless steel ring. Each sample was sterilized in 75% ethanol for 12 hours, washed three times with PBS, irradiated with ultraviolet light for 2 hours, and then cultured overnight in growth medium. The HUVECs were cultured at 2.0 × 10⁻⁶ cells per well. 4Inoculate at a density of 10 cells / well and change the culture medium every 2 days.

[0185] HUVECs viability was assessed using a cell counting kit (CCK-8). Cells were cultured on glass slides, ECMO membranes, and hollow fiber membranes for 1, 3, and 5 days, respectively. At the designated culture time points, the wells were removed from the CO2 incubator, and a series of procedures were performed to remove the culture medium. The samples in the wells were washed with PBS, and 360 μL of DMEM medium and 40 μL of CCK-8 solution were added sequentially to the fluorescent plates. After incubation in a CO2 incubator for 1 hour, the optical density (OD) was measured at 450 nm using a microplate reader.

[0186] Before SEM imaging of the cells, cultured HUVECs were fixed with 4% paraformaldehyde and dehydrated on day 3 using a gradient of ethanol (30%, 50%, 70%, 80%, 90%, 95%, 100%). Morphological analysis was performed using a scanning electron microscope (SEM, PhenomXL, Netherlands) at an accelerating voltage of 10 mV and an accelerating voltage of 5 mA for 35 seconds. Figure 15 As shown.

[0187] In addition, live cells (green) and dead cells (red) were stained with calcein AM / PI after 3 days of culture, and the stained samples were immediately observed under a TS100 fluorescence microscope (Nikon, Japan). Figure 16 As shown.

[0188] As implantable medical materials, catheter coatings, both before and after modification, must ensure no significant damage to human vascular endothelial cells. Therefore, the CCK-8 assay was used to detect the cytotoxicity of the catheter materials. The same amount of HUVECs (hyperendothelial cells) were seeded onto each sample. The adhesion ability of ECMO membranes and hollow fiber membranes to HUVECs was similar, indicating that the modified coating had no significant impact on the biocompatibility of the catheter membrane material. Regarding proliferation activity, on days 1, 3, and 5, the proliferation activities of ECMO membranes and hollow fiber membranes were similar, with little difference and no statistical significance (P<0.05), indicating that both ECMO membranes and hollow fiber membranes have good proliferative activity for cells and no toxic side effects.

[0189] On day 3, live / dead staining of HUVECs cells showed that a small number of dead cells were present on both the ECMO membrane and the hollow fiber membrane, a common phenomenon during cell metabolism. A large number of cells survived on both membrane materials, and they were evenly dispersed with roughly the same cell number and density. The modified coating did not significantly affect the growth of host cells on the catheter membrane material, classifying it as a safe biomedical material.

[0190] Example 6: In vivo biocompatibility of hollow fiber membrane tubes

[0191] To evaluate the biocompatibility of the materials in vivo, a 1 cm diameter wound was created on the back of rats. A sterilized ECMO sample and a 2.5 cm long hollow fiber membrane were then embedded in the wound tissue. Samples were collected and photographed on days 14 and 28 post-embedding to observe the growth of the ECMO sample and hollow fiber membrane on the surrounding tissue. The animal experimental protocol for SD rats was approved by the Animal Ethics Review Committee of Shanghai First People's Hospital.

[0192] Two different catheter materials, ECMO samples and hollow fiber membranes, were subcutaneously embedded in SD rats, with the embedding sites located on both sides of the rat's spine on the back. Then, the two sterilized materials were embedded in wound sites for testing and evaluation. Figure 17 As shown.

[0193] Surgical Procedure Observation: All surgical procedures were performed strictly according to aseptic techniques. Based on the weight of the SD rats, intraperitoneal anesthesia was administered. Five minutes after anesthesia, the rats were fully anesthetized. The catheter material was then embedded in their backs according to the designated location, and the wound was sutured. Throughout the entire surgical process, the rats' vital signs remained normal. To prevent hypothermia, non-surgical areas were covered with absorbent cotton for warmth during surgery, while the laboratory temperature was maintained at 25°C. Approximately 3 hours after anesthesia, the rats began to awaken, exhibiting slight head twisting and forelimb movement. They crawled slowly, but their hind limbs were weak. At this time, the rats' vital signs were normal, and some rats urinated and defecated. Approximately 30 minutes after awakening, they began to walk unsteadily. One hour later, they began to forage and drink water, and could squat on their hind limbs, occasionally walking. A noticeable bulge was visible at the suture site, with a small amount of blood seeping from the slightly red wound, but no significant bleeding was observed.

[0194] Observations on days 3 and 7 post-surgery: SD rats exhibited poorer behavior in drinking, foraging, and sleeping compared to before surgery, and did not bite the wound on their backs. Stimulation of the rats showed significantly slower responses compared to the un-operated state, and mild redness and swelling were observed at the wound site. Gentle touch on the back revealed a slight bulge at the sample implantation site, and light pressure on the wound revealed a small amount of tissue fluid exudation. No obvious encapsulation was observed around the catheter after sample collection.

[0195] Tissue samples were taken 14 and 28 days post-operation: the slides were found to be wrapped in a layer of dark black fibrous tissue. Both the ECMO sample catheter and the hollow fiber membrane catheter had formed a thin, translucent fibrous membrane with a high-density blood network on their surfaces. Compared with before implantation, the appearance and morphology of the membrane had not changed significantly, and only a slight inflammatory reaction was observed around the implantation site, with no obvious granulation tissue proliferation. In contrast, the hollow fiber membrane catheter had a sparser and thinner wrapping material than the ECMO sample catheter, with more visible blood network.

[0196] After embedding ECMO membranes and hollow fiber membrane tubes into the backs of rats, the rats still exhibited behavioral abnormalities such as drinking, foraging, and sleeping on the third day post-surgery, showing sluggishness compared to pre-surgery levels. Inflammation was also observed in the wounds, suggesting possible infection. Tissue samples taken 14 and 28 days post-surgery revealed red fibrous tissue encapsulating the ECMO membranes and hollow fiber membrane tubes. The tissue covering the hollow fiber membranes was relatively lighter in color, indicating that both ECMO membranes and hollow fiber membrane tubes experienced prolonged, persistent inflammation after implantation, forming a thick biofilm on the surface. This demonstrates good biocompatibility and potential for future applications.

Claims

1. A method for producing an enhanced polypropylene hollow fiber membrane, characterized by, The method comprises the following steps: (1) polypropylene and SiO2 powder are air-dried; (2) soybean oil and SiO2 are homogenized by a high-speed shearing mixer, and then ultrasonically treated for 30 minutes; (3) polypropylene powder is added and homogenized by a high-speed shearing mixer to obtain a uniform and stable spinning suspension; (4) the spinning suspension is added into a double-screw extruder, extruded through an annular spinneret, and uniformly coated on the outer surface of a braided tube; (5) the braided tube coated with the spinning suspension is cooled in a coagulation bath, wound on a winding roller, and immersed in dichloromethane to sufficiently extract soybean oil, thereby obtaining a reinforced polypropylene hollow fiber membrane.

2. The method for producing the reinforced polypropylene hollow fiber membrane according to claim 1, characterized by, In step (1), the mass ratio of polypropylene and SiO2 powder is 10-30:1.

5.

3. The method of producing the reinforced polypropylene hollow fiber membrane according to claim 1, wherein, In step (2), the mass ratio of soybean oil and SiO2 is 68.5-88.5:1.

5.

4. The method of producing the reinforced polypropylene hollow fiber membrane according to claim 1, wherein In steps (4) and (5), the parameters of the twin-screw extruder include: spinning temperature of 220°C; coagulation bath of water; air gap of 35 cm; extrusion rate of 35 r·min -1 ; and crimping rate of 17.5 m·min -1 .

5. An enhanced polypropylene hollow fiber membrane, characterized by, The reinforced polypropylene hollow fiber membrane is prepared by the method of claim 1.

6. Use of the reinforced polypropylene hollow fiber membrane of claim 5 in the preparation of an in-vivo oxygenation membrane.

7. Use of the reinforced polypropylene hollow fiber membrane of claim 5 in the preparation of an in-vivo hemodialysis membrane.

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