CN112220965B - Janus porous biological patch for wound repair and preparation method thereof - Google Patents
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Abstract
The invention discloses a Janus porous biological patch for wound repair and a preparation method thereof, wherein a microfluidic single emulsion generating device is utilized to prepare a colloid liquid drop template with a surface coated with hydrophilic silica nanoparticles and hydrophobic magnetic nanoparticles, an internal phase solution is an oil phase in which two particles are dispersed, an external phase solution is a photopolymerization aqueous polyurethane solution, and the two generated colloid liquid drops are assembled in a hexagonal close-packed structure under the action of a magnetic field, and ultraviolet light is solidified and the template is removed. The Janus porous biological patch for wound repair is constructed based on two colloid droplet templates with different surface wettability, so that the Janus porous biological patch has controllable surface wettability, the hydrophobic surface is in direct contact with the wound surface, secondary wounds caused by tissue adhesion can be avoided, and the hydrophilic surface can adsorb exudates at the wound, so that wound repair is accelerated. The preparation method is simple and easy to implement, strong in controllability, good in repeatability and wide in application prospect.
Description
Technical Field
The invention relates to the technical field of biomedical materials, in particular to a Janus porous biological patch for wound repair and a preparation method thereof.
Background
Skin wound surface is very common in life and is also very easy to initiate in operation treatment, so that wound surface repair has attracted great importance in the medical field. An ideal wound repair method is to promote cell adhesion and proliferation, directionally migrate to a wound area, secrete extracellular matrix and restore the original functions of tissues. To achieve these processes, biological patches have been developed. The biological patch is a film patch prepared from one or more biocompatible materials, and can meet the needs of wound repair of different parts of a human body.
In general, an anti-adhesion biological patch made of a hydrophobic material with low surface energy and small surface roughness can effectively isolate a wound area from adjacent tissues, but the promotion effect of the anti-adhesion biological patch on tissue regeneration is not ideal. On the other hand, adherent biological patches such as decellularized tissue matrix, polymeric gauze, etc. have the effect of promoting cell adhesion and infiltration, but at the same time, there are unnecessary wound adhesion, which is liable to cause secondary wounds.
Disclosure of Invention
Aiming at the defects of the prior art, the invention provides a Janus porous biological patch for wound repair and a preparation method thereof.
The invention designs a Janus porous biological patch for wound repair based on surface wettability, which has a Janus structure, and the surface wettability of two sides of the biological patch is quite different, wherein the hydrophobic surface is in direct contact with the wound surface, so that secondary wounds caused by tissue adhesion can be avoided, and the hydrophilic surface can adsorb exudates at the wound, so that wound repair is accelerated. The biological patch is an ideal tissue repair material and has wide clinical application value.
In order to achieve the technical purpose, the invention adopts the following technical scheme:
a Janus porous biological patch for wound repair, the preparation method comprises the following steps:
(1) Constructing a microfluidic single emulsion device: a glass capillary is broken by using a capillary tube drawing instrument, and then the broken glass capillary tube is polished into a tip by using sand paper to serve as an internal phase tube; taking a glass capillary tube with the same size, and polishing one end of the glass capillary tube with sand paper to be smooth to be used as an external phase tube; assembling the inner phase tube and the outer phase tube on a cover glass, keeping the central line horizontally aligned and fixing; connecting and sealing the interface of the two glass capillaries for later use;
(2) Preparing a colloid droplet template: preparing a colloid liquid drop template coated with hydrophilic silica nanoparticles and hydrophobic magnetic nanoparticles by using the microfluidic single emulsion device constructed in the step (1); the internal phase solution is n-hexadecane dispersed with hydrophilic silica nano particles and hydrophobic magnetic nano particles, the external phase solution is a photopolymerization aqueous polyurethane solution added with a surfactant, the internal phase solution is sheared by the external phase solution at the tip of the internal phase tube to generate colloid droplets, and the colloid droplets are automatically layered under the action of a magnetic field and assembled into a colloid droplet template in a hexagonal close-packed structure;
(3) Preparing a Janus porous biological patch for wound repair: taking the colloid liquid drop template prepared in the step (2) as a positive template, carrying out in-situ ultraviolet polymerization on an external phase solution-photopolymerization aqueous polyurethane solution, and removing the positive template to obtain the Janus porous biological patch for wound repair.
The outer diameter of the glass capillary tube adopted in the step (1) is 1mm, the inner diameter is 0.8mm, and the inner phase tube is polished by sand paper until the inner diameter of the tip is 0.4mm; the polished inner phase tube and the polished outer phase tube are ultrasonically cleaned by absolute ethyl alcohol for 2min and naturally air-dried for standby.
Further, in the external phase solution, the surfactant is a mixed solution of sodium dodecyl sulfate and polyether type surfactant F108, and the mass concentration of the mixed solution is 2%m/v.
Further, the curing method of the photopolymerization aqueous polyurethane solution comprises the steps of adding 2%m/v 2-hydroxy-2-methyl propiophenone into the aqueous polyurethane solution, and performing ultraviolet crosslinking curing: wherein the photopolymerized aqueous polyurethane solution is required to be processed in a dark place before photocuring.
Further, the particle size of the hydrophilic silica nanoparticles is 220nm; the particle size of the hydrophobic magnetic nano particles is 240nm;
further, the diameter of the colloidal droplets is 300-500 μm.
Further, the method for removing the colloid droplet template is to soak and clean the colloid droplet template by using normal hexane.
In the step (2), the colloid droplet template coated with the hydrophilic silica nanoparticles and the hydrophobic magnetic nanoparticles automatically separates into two layers under the action of a magnetic field, wherein the lower layer is a single-layer colloid droplet coated with the hydrophobic magnetic nanoparticles, and the upper layer is a double-layer colloid droplet coated with the hydrophilic silica nanoparticles.
Furthermore, the colloid liquid drop template coated with the hydrophilic silica nano particles and the hydrophobic magnetic nano particles on the surface is automatically divided into two layers under the action of a magnetic field, and the proportion of the hydrophilic colloid liquid drops in the hydrophobic colloid liquid drop layer can be adjusted according to the action time and the distance of the magnetic field.
Further, the Janus porous biological patch for wound repair has a hydrophobic surface doped with hydrophilic holes on the lower layer and a hydrophilic surface on the upper layer, wherein the hydrophobic surface is in direct contact with the wound surface, so that secondary wounds caused by tissue adhesion can be avoided, and the hydrophilic surface can adsorb exudates at the wound, thereby accelerating wound repair.
Compared with the prior art, the invention has the beneficial effects that:
1) The preparation method of the Janus porous biological patch for wound repair provided by the invention utilizes a microfluidic single-emulsion droplet generation technology to prepare the colloid droplet template with the surface coated with the hydrophilic silica nanoparticles and the hydrophobic magnetic nanoparticles, and the preparation method is simple and easy to implement, good in controllability and high in accuracy, and can adjust the diameter of droplets on line;
2) The Janus porous biological patch for wound repair provided by the invention has the advantages that the lower layer is a hydrophobic surface doped with hydrophilic holes, and the upper layer is a hydrophilic surface, wherein the hydrophobic surface is in direct contact with the wound surface, so that secondary wounds caused by tissue adhesion can be avoided, and the hydrophilic surface can adsorb exudates at the wound, thereby accelerating wound repair;
3) The Janus porous biological patch for wound repair provided by the invention has the advantages that the whole structure is an inverse opal porous structure, the pore structure is uniform and high in connectivity, the transmission of oxygen and nutrient substances is facilitated, and wastes generated by biological effusion and cell metabolism are discharged, so that wound repair is effectively accelerated.
Drawings
Fig. 1 is a schematic diagram of the preparation of a colloid droplet template of a Janus porous biological patch for wound repair according to the present invention.
Fig. 2 is a schematic representation of the preparation of a Janus porous biological patch for wound repair according to the present invention.
FIG. 3 is a view of the hydrophilic and hydrophobic side of a Janus porous biological patch for wound repair according to the present invention; wherein, figure a is a hydrophilic side electron microscope image, the scale is 5 μm, figure b is a hydrophobic side electron microscope image, and the scale is 2.5 μm.
FIG. 4 shows an application of Janus porous biological patch in wound repair according to an embodiment of the present invention; wherein a is H & E staining results, scale is 1mm, b is Masson staining results, scale is 100 μm.
Detailed Description
In order to better understand the technical solutions of the present invention for those skilled in the art, the following embodiments of the present invention are described in further detail with reference to the accompanying drawings.
The experimental methods used in the examples below, unless otherwise indicated, are conventional methods, and the reagents, methods and apparatus used, unless otherwise indicated, are conventional in the art.
The invention provides a preparation method of Janus porous biological patch for wound repair, which utilizes microfluidic single emulsion droplet generation technology to prepare a colloid droplet template with hydrophilic silica nanoparticles and hydrophobic magnetic nanoparticles coated on the surface, and the preparation method is simple and easy to implement, good in controllability and high in accuracy, and can adjust the diameter of droplets on line.
The Janus porous biological patch for wound repair has the advantages that the lower layer is the hydrophobic surface doped with the hydrophilic holes, the upper layer is the hydrophilic surface, the holes are uniform in structure and high in connectivity, the transmission of oxygen and nutrient substances is facilitated, wastes generated by biological effusion and cell metabolism are discharged, secondary wounds caused by tissue adhesion can be avoided by directly contacting the hydrophobic surface with the wound, and the hydrophilic surface can adsorb exudates at the wound, so that wound repair is accelerated.
The following are examples:
example 1
A Janus porous biological patch with complete anisotropic wettability for wound repair, which is prepared by the following method:
(1) Construction of microfluidic single emulsion devices
A glass capillary (the outer diameter is 1mm, the inner diameter is 0.8 mm) is broken by using a capillary tube drawing instrument, and then the broken glass capillary tip is polished by sand paper until the inner diameter is 0.4mm for standby, wherein the glass capillary is an inner phase tube; and (3) taking a glass capillary tube with the same size, polishing one end of the glass capillary tube with sand paper to be smooth, taking the glass capillary tube as an outer phase tube, ultrasonically cleaning an inner phase tube and the outer phase tube with absolute ethyl alcohol for 2min, and naturally air-drying for later use. And assembling the inner phase tube and the outer phase tube on a cover glass, keeping the central lines horizontally aligned, fixing by using AB glue, connecting the joint parts of the two glass capillaries by using a flat-mouth needle head, and finally sealing by using AB glue, and using after the glue is dried.
(2) Preparation of colloidal drop templates
N-hexadecane containing 7% hydrophobic magnetic nanoparticles (m/v) was prepared as the internal phase solution 1. The aqueous solution containing 7% hydrophilic silica nanoparticles (m/v) was replaced with absolute ethanol, repeated 3 times to ensure complete removal of water, then the solution was dispersed into n-hexadecane, placed in an oven, and left overnight at 40 ℃ to remove the absolute ethanol, obtaining n-hexadecane solution containing 7% hydrophilic silica nanoparticles as the internal phase solution 2. An aqueous polyurethane solution containing 2% sodium dodecyl sulfate and polyether surfactant F108 (m/v) was prepared, and then 2% of photoinitiator 2-hydroxy-2-methyl propiophenone (m/v) was added as an external phase solution. Taking two microfluidic single emulsion devices, wherein an inner phase pipe of one device is filled with an inner phase solution 1, an outer phase pipe of the other device is filled with an outer phase solution 2, the outer phase pipe of the other device is filled with an outer phase solution, and colloid droplets with different sizes are obtained by adjusting the flow rates of the inner phase solution and the outer phase solution of the two devices.
(3) Preparation of Janus porous biological patch for wound repair
And (3) automatically and completely layering the colloid liquid drops prepared in the step (2) under the action of a magnetic field, assembling the colloid liquid drops in a hexagonal close-packed structure to form a colloid liquid drop positive template, performing in-situ ultraviolet polymerization on an external phase solution, namely, performing photopolymerization on the aqueous polyurethane solution, and then cleaning n-hexadecane by using n-hexane to remove the positive template to obtain the Janus porous biological patch for wound repair.
(4) Characterization of Janus porous biological patch for wound repair
And the Janus porous biological patch structure for wound repair is observed and characterized under a split microscope and a scanning electron microscope, and the diameter of the holes of the Janus porous biological patch for wound repair is measured to be about 350 mu m, so that the connectivity among the holes is good.
Example 2
A Janus porous biological patch with partial anisotropic wettability for wound repair, which is prepared by the following method:
(1) Construction of microfluidic single emulsion devices
A glass capillary (the outer diameter is 1mm, the inner diameter is 0.8 mm) is broken by using a capillary tube drawing instrument, and then the broken glass capillary tip is polished by sand paper until the inner diameter is 0.4mm for standby, wherein the glass capillary is an inner phase tube; and (3) taking a glass capillary tube with the same size, polishing one end of the glass capillary tube with sand paper to be smooth, taking the glass capillary tube as an outer phase tube, ultrasonically cleaning an inner phase tube and the outer phase tube with absolute ethyl alcohol for 2min, and naturally air-drying for later use. And assembling the inner phase tube and the outer phase tube on a cover glass, keeping the central lines horizontally aligned, fixing by using AB glue, connecting the joint parts of the two glass capillaries by using a flat-mouth needle head, and finally sealing by using AB glue, and using after the glue is dried.
(2) Preparation of colloidal drop templates
N-hexadecane containing 7% hydrophobic magnetic nanoparticles (m/v) was prepared as the internal phase solution 1. The aqueous solution containing 7% hydrophilic silica nanoparticles (m/v) was replaced with absolute ethanol, repeated 3 times to ensure complete removal of water, then the solution was dispersed into n-hexadecane, placed in an oven, and left overnight at 40 ℃ to remove the absolute ethanol, obtaining n-hexadecane solution containing 7% hydrophilic silica nanoparticles as the internal phase solution 2. An aqueous polyurethane solution containing 2% sodium dodecyl sulfate and polyether surfactant F108 (m/v) was prepared, and then 2% of photoinitiator 2-hydroxy-2-methyl propiophenone (m/v) was added as an external phase solution. Taking two microfluidic single emulsion devices, wherein an inner phase pipe of one device is filled with an inner phase solution 1, an outer phase pipe of the other device is filled with an outer phase solution 2, the outer phase pipe of the other device is filled with an outer phase solution, and colloid droplets with different sizes are obtained by adjusting the flow rates of the inner phase solution and the outer phase solution of the two devices.
(3) Preparation of Janus porous biological patch for wound repair
And (3) automatically layering the colloid liquid drops prepared in the step (2) under the action of a magnetic field and assembling the colloid liquid drops in a hexagonal close-packed structure to form a colloid liquid drop positive template, in the process, adjusting the proportion of the hydrophilic colloid liquid drops in a hydrophobic colloid liquid drop layer according to the action time and the distance of the magnetic field, then polymerizing an external phase solution-photopolymerization aqueous polyurethane solution in situ by ultraviolet light, and then cleaning n-hexadecane by using n-hexane to remove the positive template to obtain the Janus porous biological patch for wound surface repair.
(4) Characterization of Janus porous biological patch for wound repair
The Janus porous biological patch structure for wound repair is observed and characterized under a split microscope and a scanning electron microscope, the diameter of the holes of the Janus porous biological patch for wound repair is measured to be about 350 mu m, a certain proportion of hydrophilic holes are dispersed on the hydrophobic side, and the connectivity among the holes is good.
Example 3
Janus porous biological patch was used in the rat wound model:
taking the Janus porous biological patch for wound repair prepared in example 2 as an example, randomly dividing 10 male SD rats with the weight of about 200g into 2 groups, namely a control group and an experimental group, respectively, 5 rats per group, then performing anesthesia by injecting 10% (w/v) chloral hydrate solution into the abdominal cavity, establishing a wound model with the diameter of 1cm on the back of the rats, wherein the control group does not receive any treatment, the experimental group is attached with the Janus porous biological patch prepared in example 2, the hydrophobic side is directly contacted with the wound, and after 9 days of treatment, taking down tissue samples of the wound and performing Masson and H & E staining;
the results are shown in FIG. 4, and H & E staining results show that the thickness of the new granulation tissue of the experimental group is significantly higher than that of the control group, indicating that the tissue regeneration condition of the experimental group is better than that of the control group. From the Masson staining image result, the collagen deposition amount of the experimental group is obviously larger than that of the control group, which shows that the Janus porous biological patch for wound repair can effectively promote wound collagen regeneration. The above results show that the Janus porous biological patch for wound repair can effectively promote wound repair, wherein the lower layer of the Janus porous biological patch is a hydrophobic surface doped with hydrophilic holes, the upper layer of the Janus porous biological patch is a hydrophilic surface, the hole structure is uniform and high in connectivity, the transmission of oxygen and nutrient substances is facilitated, wastes generated by biological effusion and cell metabolism are discharged, in addition, secondary wounds caused by tissue adhesion can be avoided by direct contact between the hydrophobic surface and the wound, and the hydrophilic surface can adsorb exudates at the wound, so that wound repair is accelerated.
The present invention is not limited to the preferred embodiments, and any simple modification, equivalent replacement, and improvement made to the above embodiments by those skilled in the art without departing from the technical scope of the present invention, will fall within the scope of the present invention.
Claims (7)
1. A Janus porous biological patch for wound repair, characterized in that: the preparation method comprises the following steps:
(1) Constructing a microfluidic single emulsion device: a glass capillary is broken by using a capillary tube drawing instrument, and then the broken glass capillary tube is polished into a tip by using sand paper to serve as an internal phase tube; taking a glass capillary tube with the same size, and polishing one end of the glass capillary tube with sand paper to be smooth to be used as an external phase tube; assembling the inner phase tube and the outer phase tube on a cover glass, keeping the central line horizontally aligned and fixing; connecting and sealing the interface of the two glass capillaries for later use;
(2) Preparing a colloid droplet template: preparing a colloid liquid drop template coated with hydrophilic silica nanoparticles and hydrophobic magnetic nanoparticles by using the microfluidic single emulsion device constructed in the step (1); the internal phase solution is n-hexadecane dispersed with hydrophilic silica nano particles and hydrophobic magnetic nano particles, the external phase solution is a photopolymerization aqueous polyurethane solution added with a surfactant, the internal phase solution is sheared by the external phase solution at the tip of the internal phase tube to generate colloid droplets, and the colloid droplets are automatically layered under the action of a magnetic field and assembled into a colloid droplet template in a hexagonal close-packed structure;
(3) Preparing a Janus porous biological patch for wound repair: taking the colloid droplet template prepared in the step (2) as a positive template, carrying out in-situ ultraviolet polymerization on an external phase solution-photopolymerization aqueous polyurethane solution, and removing the positive template to obtain the Janus porous biological patch for wound repair;
in the step (2), a colloid droplet template coated with hydrophilic silica nanoparticles and hydrophobic magnetic nanoparticles on the surface is automatically divided into two layers under the action of a magnetic field, wherein the lower layer is a single-layer colloid droplet coated with the hydrophobic magnetic nanoparticles, and the upper layer is a double-layer colloid droplet coated with the hydrophilic silica nanoparticles;
the colloid liquid drop template with the surface coated with the hydrophilic silica nano particles and the hydrophobic magnetic nano particles is automatically divided into two layers under the action of a magnetic field, and the proportion of the hydrophilic colloid liquid drops in the hydrophobic colloid liquid drop layer can be adjusted according to the action time and the distance of the magnetic field;
the Janus porous biological patch for wound repair has a hydrophobic surface doped with hydrophilic holes on the lower layer and a hydrophilic surface on the upper layer, wherein the hydrophobic surface is in direct contact with the wound surface, so that secondary wounds caused by tissue adhesion can be avoided, and the hydrophilic surface can adsorb exudates at the wound, thereby accelerating wound repair.
2. The Janus porous biological patch for wound repair of claim 1, wherein: the outer diameter of the glass capillary tube adopted in the step (1) is 1mm, the inner diameter is 0.8mm, and the inner phase tube is sanded to the inner diameter of the tip end of the glass capillary tube of 0.4mm; the polished inner phase tube and the polished outer phase tube are ultrasonically cleaned by absolute ethyl alcohol for 2min and naturally air-dried for standby.
3. The Janus porous biological patch for wound repair of claim 1, wherein: in the external phase solution, the surfactant is a mixed solution of sodium dodecyl sulfate and polyether type surfactant F108, and the mass concentration of the mixed solution is 2% m/v.
4. The Janus porous biological patch for wound repair of claim 1, wherein: the curing method of the photopolymerization aqueous polyurethane solution comprises the steps of adding 2% m/v of 2-hydroxy-2-methyl propiophenone into the solution, and performing ultraviolet crosslinking curing: wherein the photopolymerized aqueous polyurethane solution is required to be processed in a dark place before photocuring.
5. The Janus porous biological patch for wound repair of claim 1, wherein: the particle size of the hydrophilic silica nano particles is 220nm; the particle size of the hydrophobic magnetic nano particle is 240 nm.
6. The Janus porous biological patch for wound repair of claim 1, wherein: the diameter of the colloid liquid drop is 300-500 mu m.
7. The Janus porous biological patch for wound repair of claim 1, wherein: the method for removing the colloid drop template is to soak and clean the colloid drop template by using normal hexane.
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| CN106117458A (en) * | 2016-07-06 | 2016-11-16 | 东南大学 | Amphiphilic Janus colloidal crystal microsphere and preparation method thereof, application |
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| CN111407924B (en) * | 2020-04-17 | 2020-10-27 | 南京鼓楼医院 | Composite patch with anisotropic surface and preparation method and application thereof |
| CN111410766B (en) * | 2020-04-27 | 2021-01-01 | 南京鼓楼医院 | Preparation method of hierarchical-structure inverse opal porous biological scaffold |
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