CN115645599B - Thermosensitive gel dressing for wound repair after tumor resection and preparation method thereof - Google Patents
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Abstract
A thermosensitive gel dressing for repairing wound surface after tumor resection comprises CuO 2 Nanoparticle, BSO and chitosan-based thermosensitive gel, cuO 2 Nanoparticle PVP modified CuO 2 The mixing ratio of the chitosan and the chitosan thermosensitive gel is 1:10-1:1. The dressing provided by the invention has the advantages of synergy of various treatment modes, integration of anti-tumor and wound healing promotion, and ensured safety, and can be applied to tumor treatment, sterilization and wound healing promotion.
Description
Technical Field
The invention relates to a material for wound repair, in particular to a gel dressing which has thermosensitive property and is used for wound repair after tumor resection operation.
Background
Malignant tumor tissue has strong oxidation resistance and is easy to relapse and metastasize, surgical excision is the first choice and the best method for treating early tumors, and the treatment mode has defects. If the wound surface is too large after excision, infection can be caused, scars can be left to influence aesthetic feeling, and the probability of tumor recurrence can be increased due to unclean excision. Therefore, there is an urgent need for a multifunctional material that can inhibit tumors and promote wound healing. BSO (L-Buthionine- (S, R) -sulfoximine) is a G-glutamic acid cysteine synthetase (gamma-glutamylcysteine synthetase) inhibitor, which can reduce the level of intracellular glutathione, thereby reducing the antioxidant capacity of cells.
Researchers have found CuO 2 Nanoparticle in tumor subacidity environmentHas a chemical kinetics (CDT) effect (J Am Chem Soc 141, 9937-9945), which releases Cu 2+ Has antibacterial (Kannan, S, mary, et al.2017, 95:928-937.) and wound healing promoting effects (ACS nano.2017,11, (11): 11337-11349).
In addition to improving therapeutic efficiency by multimode combination, increasing the concentration of the drug at the tumor site is another approach. For this reason, researchers have also designed a wide variety of drug carriers, and have been wrapped with BaTiO 3 The injectable thermal gel is used for tumor treatment, and at body temperature, the thermal gel is changed into gel state, so that the purpose of slowly releasing the material in the tumor to improve the treatment effect is achieved (Adv Mater.2020;32 (29): e 2001976), and the thermal gel has better biological safety and slow release function. In addition, the hydrogel has the characteristics of moisture and air permeability and plasticity, so that the hydrogel can be used as a skin surface dressing, and can completely cover a wound to protect the skin.
At present, the CuO is not wrapped by thermal gel 2 Nanoparticles and BSO are reported to be used for both tumor treatment and wound healing.
Disclosure of Invention
The invention aims to provide a thermosensitive gel dressing for wound repair after tumor resection, which is based on chitosan-based thermosensitive gel and is used for wound repair after tumor resection.
A thermosensitive gel dressing for wound repair after tumor resection, comprising: cuO (CuO) 2 Nanoparticles, BSO and chitosan-based thermosensitive gels.
CuO 2 The particle size of the nano particles is 3 nm-25 nm, especially 7 nm-20 nm.
CuO 2 The nanoparticles and BSO may be homogeneously mixed with the thermal gel.
CuO 2 The mixing ratio of the nano particles and the chitosan thermosensitive gel is 1:10-1:1, and the optimal volume ratio is 1:5, so that the nano particles and the chitosan thermosensitive gel have good wetting and ventilation effects, good thermosensitive effect (thermosensitive property of the gel is reduced when the volume ratio of the ketone peroxide to the thermal gel is greater than 1:1) and good tumor treatment effect.
CuO 2 Nanoparticle-based polyvinylpyrrolidone (PVP) -modified CuO 2 The preparation method comprises the following steps: cuCl is added 2 ·2H 2 O and PVP (molecular weight: 40000 Da) are dissolved in water and mixed and stirred uniformly (CuCl) 2 ·2H 2 O and PVP in a weight ratio of 0.013), followed by addition of NaOH (CuCl) 2 ·2H 2 Molar ratio of O to NaOH of 2:1) and H 2 O 2 (CuCl in an amount of 0.01M per 5 mL) 2 ·2H 2 O was added 100 μl). After stirring (e.g., 30 minutes), PVP coated CP nanodots were collected by ultrafiltration and washed multiple times with water. PVP modified CuO obtained 2 And (3) nanoparticles.
The acetic acid solution was mixed with chitosan (4 ml of 0.1M acetic acid was added per 100mg of chitosan), and then beta-GP (concentration to 600 mg/ml) was added to obtain the chitosan-based thermosensitive gel of the present invention.
Will produce CuO 2 The nano particles and BSO are added into the chitosan-based thermosensitive gel, so that the dressing of the invention is obtained.
The technical scheme of the invention has the beneficial effects that:
the thermosensitive gel dressing for wound repair after tumor resection provided by the invention has the advantages that a plurality of treatment modes are coordinated, the anti-tumor and wound healing promotion are integrated, and the multifunctional thermosensitive gel with guaranteed safety can be applied to tumor treatment, sterilization and wound healing promotion. On one hand, the thermal gel can be injected in situ at the tumor part to ensure that the medicine is retained to achieve better effect of killing tumor cells, and on the other hand, the thermal gel can also be used as a skin surface dressing to prevent recrudescence after the tumor is not completely resected in the operation, and meanwhile, the components and the structure of the thermal gel provide a sterile, breathable and moist environment which is beneficial to wound healing.
The dressing preparation method is simple and feasible, is environment-friendly, and can control the particle size of the prepared particles.
Proved by verification, the dressing provided by the invention can achieve better synergistic treatment effects of chemical power, acoustic power and medicines for treating tumor oxidative death.
Drawings
FIG. 1 is a flow chart of the preparation of a nano-drug carrier material according to an embodiment of the present invention.
FIG. 2 is CuO 2 TEM of nanoparticlesA figure;
FIG. 3 is CuO 2 Different mixing ratios of nanoparticles to thermogels are illustrated;
FIG. 4A is CuO 2 A graph of @ Gel cytotoxicity results;
FIG. 4B is a graph of the cytotoxicity results of BSO@gel;
FIG. 4C is CuO 2 A plot of the cytotoxicity results of BSO@gel;
FIG. 4D is CuO 2 @gel, BSO@gel and CuO 2 The cytotoxicity results of each group of BSO@gel under the action of Ultrasound (US) are compared with the result graph;
FIG. 5 is CuO 2 Electron Spin Resonance (ESR) test results plots of nanoparticles in PBS solution;
FIG. 6 is an ultrasonic CuO 2 Electron Spin Resonance (ESR) test results for nanoparticles;
FIG. 7 is CuO 2 A graph of test results of the killing effect of the nano-particles on tumor cells;
FIG. 8 is CuO 2 Electron microscopy of the antibacterial effect of the nanoparticles;
FIG. 9 is CuO 2 A statistical graph of the results of the antibacterial effect of the nanoparticles;
FIG. 10 shows a CuO-containing composition 2 A statistical graph of killing effect of dressing of nano particles on escherichia coli;
FIG. 11 shows a CuO-containing composition 2 A statistical graph of killing effect of dressing of nano particles on golden coccus;
FIG. 12 is CuO 2 The nanometer particles promote the wound healing result graph.
Detailed Description
The technical scheme of the present invention is described in detail below with reference to the accompanying drawings. The embodiments of the present invention are only for illustrating the technical scheme of the present invention and not for limiting the same, and although the present invention has been described in detail with reference to the preferred embodiments, it should be understood by those skilled in the art that modifications and equivalents may be made thereto without departing from the spirit and scope of the technical scheme of the present invention, which is intended to be covered by the scope of the claims of the present invention.
The anti-tumor, antibacterial and healing-promoting experiments and detection methods in this example are as follows:
antitumor: will be 1X 10 6 B16F10 cells were inoculated into four-week-old BALB/c mice until tumors grew to 100mm 2 When left and right, mice were randomly divided into 6 groups of five mice, each group was treated as shown in fig. 7 on days 1, 3, 5 and 7, and the length and width of tumors on days 1, 3, 5, 7 and 9 were recorded and according to the formula: tumor volume was equal to (length x width 2)/2 the number of changes in tumor volume relative to the first day was calculated for each group of mice and plotted.
Antibacterial: after passing the bacteria through different treatments (control group, BSO@gel, cuO) 2 @Gel、CuO 2 +BSO@gel;2 Ultrasound (US) treatment groups: cuO (CuO) 2 @ gel+US and CuO 2 +BSO@gel+US, parameter 1.5W/cm 2 1mhz,50% duty cycle, 4 min) was prepared as SEM samples and photographed.
The density of the bacterial suspension was calculated by the UV-visible spectrum at 600nm, adjusted to 1X 10 6 Colony Forming Units (CFU)/milliliter. 100 μl of the diluted bacteria were spread on the surface of agar plates and treated with different solutions, including 4 non-sonicated groups: control group, BSO@gel, cuO 2 @Gel、CuO 2 +BSO@gel;2 sonication groups: cuO (CuO) 2 @ gel+US and CuO 2 +BSO@gel+US, parameter 1.5W/cm 2 1MHz,50% duty cycle, 4min. After the treatment, the bacterial solution was spread on plates, after incubation for 24 hours in an incubator at 37 ℃, bacterial colony images were taken, the colony count of each agar plate was counted, and bacterial viability was calculated.
Promoting healing: after anesthesia by intraperitoneal injection of 4% chloral hydrate solution at 20mg/kg body weight, a circular full-thickness skin wound of 10mm diameter was created on the back of the mice. 50. Mu.L of 1X 10 is then added 7 cfu/ml staphylococcus aureus suspension is soaked on the wound surface. Then randomly divided into 5 groups (n=5), 5 in each group, namely PBS control group, gel group and CuO 2 +BSO@gel group, cuO 2 +BSO@gel+US group, cuO 2 In +BSO@gel+US group, 100 μl PBS or hydrogels containing different nanoparticles are smeared on the wound every other day, and ultrasound irradiation of 1.5W/cm is added or not added 2 1MHz,50% duty cycle, 3min. The wound size was recorded every other day. Experimental knotMice were sacrificed on day 9 post-beam.
FIG. 1 is a flow chart of the preparation of a nano-drug carrier material according to an embodiment of the present invention. As shown in FIG. 1, cuO can be prepared first 2 Nanoparticles and chitosan-based thermogels, BSO drugs were purchased. And mixing the materials at a low temperature and stirring to obtain the dressing of the embodiment.
Preparation of CuO 2 Nanoparticles: 0.5g PVP (Mw 40000) dissolved in 5mL 0.01M CuCl 2 ·2H 2 O, then 5mL of 0.02M NaOH was added, and after stirring well, 100. Mu. L H was added 2 O 2 After stirring for 30min, the mixture was collected by ultrafiltration tube and washed several times with deionized water. The prepared CuO 2 TEM detection shows that the nanometer particle has spherical shape and homogeneous size, as shown in figure 2.
Preparation of thermal Gel (Gel): 100mg of chitosan was added to 4mL of 0.1M acetic acid and stirred at room temperature until clear. 600mg of beta-GP (beta-glycerophosphate) was dissolved in 1mL of deionized water and filtered through a 0.22 μm filter. The two solutions are placed at 4 ℃ and cooled for 20min, and the beta-GP solution is slowly dripped into the mixed solution of chitosan and acetic acid under stirring.
The proportion of each component in the multifunctional gel can be adjusted according to the treatment requirement, and the recommended proportion is generally as follows if the multifunctional gel is used for intratumoral injection: cuO (CuO) 2 The volume ratio of the hydrogel to the wound dressing is 1:10, and the wound dressing is 1:5 when used for the wound dressing, and the wound dressing needs to be mixed under the low-temperature condition. CuO is prepared from CuO 2 Mixing with hydrogel at different ratio, the gel has fluidity at 4deg.C, gel at 37deg.C, and temperature sensitivity, and the result is shown in figure 3.
The cytotoxicity test of the samples was evaluated using the classical CCK-8 method. First, cells (i.e., B16F10 murine melanoma cells) were treated at 5X 10 3/ The density of wells was inoculated into 96-well plates and then incubated at 37℃with 5% CO 2 CO of humid air 2 The cells were allowed to adhere to the wall by culturing in an incubator for 24 hours. Then CuO with different concentrations (the concentration is based on the mass of copper as a quantitative standard) is used 2 @Gel、BSO@Gel、CuO 2 BSO@gel and pure fresh culture solution are used for replacing culture medium in the adherent cells,incubation was continued for an additional 24 hours. After the incubation was completed, the broth was removed and washed 2 times with fresh broth. Then 100. Mu.L of a serum-free medium containing 10% CCK-8 was added to each well, and the mixture was placed at 37℃with 5% CO 2 CO of humid air 2 Incubate for another 1.5h in the incubator. Absorbance (λ=450 nm) was measured on a microplate reader after gentle shaking. The cytotoxicity index was expressed as a percentage of the cell viability after the sample treatment relative to that of the untreated blank, and the results are shown in fig. 4A, 4B, 4C and 4D, respectively. The results in FIG. 7 show that the present example gives CuO 2 @gel, BSO@gel and CuO 2 The BSO@gel has killing effect on tumor cells, and the killing effect can be improved by ultrasound. The technical means can achieve better synergistic treatment effects of chemical power, acoustic power and medicaments for treating tumor oxidative death.
CuO 2 The nanoparticles were placed in PBS at pH5.5 and US (1.5W/cm 2 1 kHz) for Electron Spin Resonance (ESR) test, see fig. 5. The graph shows that CuO 2 Nanoparticles generate hydroxyl radicals in acidic PBS and ultrasonic triggering in neutral PBS to generate singlet oxygen, which has chemo-dynamic and sonodynamic properties, and thus, can produce therapeutic effects based on chemo-dynamic and sonodynamic. Proved by the verification, the embodiment provides the CuO-containing material 2 The nanoparticle dressing has various effects in anti-tumor, antibacterial and healing promotion, etc., as shown in fig. 7-11. As can be seen from FIG. 7, cuO 2 The +bso@gel+us group showed the smallest value relative to tumor volume, which demonstrated the best inhibition of tumor growth.
In FIG. 8, bacteria undergo various degrees of shrinkage, deformation and membrane breakage after various treatments. CuO (CuO) 2 The +bso@gel+us treatment group bacteria developed the most severe shrinkage, deformation and membrane breakage. In fig. 9, 10 and 11, cuO 2 The +bso@gel+us treatment group had the least number of escherichia coli/gold cocci, i.e. the lowest survival rate of both bacteria.
As can be seen from FIG. 12, cuO 2 +BSO@gel and CuO 2 The wound healing rate of +BSO@gel+US group is the fastest, and the dressing of the embodimentThe material can accelerate wound healing.
Claims (4)
1. A thermosensitive gel dressing for repairing wound surface after tumor resection is characterized by comprising CuO 2 Nanoparticles, BSO and chitosan-based thermosensitive gels, the CuO 2 Nanoparticle PVP modified CuO 2 The mixing ratio of the chitosan-based thermosensitive gel and the chitosan-based thermosensitive gel is 1:5; the PVP molecular weight is 40000Da;
the CuO is 2 The nanoparticle preparation method comprises the following steps:
CuCl is added 2 ·2H 2 O and PVP are dissolved in water, mixed and stirred uniformly, naOH and H are added 2 O 2 Stirring, ultrafiltering, and washing with water for several times to obtain PVP modified CuO 2 A nanoparticle;
CuCl 2 ·2H 2 o and PVP in a weight ratio of 0.013, cuCl 2 ·2H 2 The mol ratio of O to NaOH is 2:1;
CuCl at 0.01M per 5mL 2 ·2H 2 O100. Mu.l of H was added 2 O 2 ；
Adding 4ml of 0.1M acetic acid into every 100mg of chitosan, mixing, and adding beta-GP to obtain the chitosan-based thermosensitive gel;
the beta-GP concentration of the chitosan-based thermosensitive gel is 600mg/ml.
2. The thermosensitive gel dressing for wound repair after tumor resection according to claim 1, characterized in that the CuO 2 The particle size of the nano particles is 3 nm-25 nm.
3. The thermosensitive gel dressing for wound repair after tumor resection according to claim 1, characterized in that the CuO 2 The particle size of the nano particles is 7 nm-20 nm.
4. Thermosensitive gel dressing for wound repair after tumor resection according to claim 1, characterized by its use in the preparation of medical devices.
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| CN103635206A (en) * | 2011-04-01 | 2014-03-12 | 美国佛罗里达大学研究基金会公司 | Thermo-sensitive, mucoadhesive or dermoadhesive, and penetration-enhancing formulations for topical delivery of therapeutics |
| CN113213432A (en) * | 2021-05-14 | 2021-08-06 | 兰州大学 | Nano copper peroxide and preparation method and application thereof |
| US20220135748A1 (en) * | 2020-11-05 | 2022-05-05 | National Taiwan University Of Science And Technology | Hydrogel composition with thermos-sensitive and ionic reversible properties, carrier, method for preparing and method of use thereof |
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| CN103635206A (en) * | 2011-04-01 | 2014-03-12 | 美国佛罗里达大学研究基金会公司 | Thermo-sensitive, mucoadhesive or dermoadhesive, and penetration-enhancing formulations for topical delivery of therapeutics |
| US20220135748A1 (en) * | 2020-11-05 | 2022-05-05 | National Taiwan University Of Science And Technology | Hydrogel composition with thermos-sensitive and ionic reversible properties, carrier, method for preparing and method of use thereof |
| CN113213432A (en) * | 2021-05-14 | 2021-08-06 | 兰州大学 | Nano copper peroxide and preparation method and application thereof |
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