CN111888324B - Injectable rare earth-DNA photo-thermal hydrogel, and preparation method and application thereof - Google Patents

Injectable rare earth-DNA photo-thermal hydrogel, and preparation method and application thereof Download PDF

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CN111888324B
CN111888324B CN202010607711.0A CN202010607711A CN111888324B CN 111888324 B CN111888324 B CN 111888324B CN 202010607711 A CN202010607711 A CN 202010607711A CN 111888324 B CN111888324 B CN 111888324B
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ucnp
rare earth
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photothermal
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CN111888324A (en
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刘凯
刘斌
王帆
张洪杰
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Changchun Institute of Applied Chemistry of CAS
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Changchun Institute of Applied Chemistry of CAS
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0019—Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K41/00—Medicinal preparations obtained by treating materials with wave energy or particle radiation ; Therapies using these preparations
    • A61K41/0052—Thermotherapy; Hyperthermia; Magnetic induction; Induction heating therapy
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/26—Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/06—Ointments; Bases therefor; Other semi-solid forms, e.g. creams, sticks, gels
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Abstract

The invention provides an injectable rare earth-DNA photo-thermal hydrogel, which comprises the following components: DNA molecules with negative charges and rare earth-based nano materials with positive charges; the rare earth-based nano material with positive charges is UCNP-Au. The DNA molecule and the rare earth-based nano material form a hydrogel material with injectability, high-efficiency photothermal treatment effect and biocompatibility under the action of physical winding, hydrogen bonds and electrostatic force. Experimental results show that the photothermal efficiency of the injectable rare earth-DNA photothermal hydrogel can reach 42.67%. In addition, tumor suppression rates are increased, thereby reducing the chances of tumor recurrence and metastasis. By means of injection beside tumor, the toxic side effect of the medicine on normal tissues in the treatment process is reduced. And moreover, the degradation of the photo-thermal hydrogel in any time period can be realized by regulating and controlling the proportion of the DNA molecules and the rare earth-based nano material. The invention has simple and quick operation and low tumor recurrence and metastasis probability.

Description

Injectable rare earth-DNA photo-thermal hydrogel, and preparation method and application thereof
Technical Field
The invention relates to the technical field of self-assembly of natural biological macromolecules and emerging rare earth nanocomposite materials, in particular to injectable rare earth-DNA photo-thermal hydrogel and a preparation method and application thereof.
Background
Bladder urothelial cancer (UBC) is the most common malignancy in the urinary system, with non-muscle invasive types being most common. Transurethral cystectomy (TURBT) is the leading treatment for this type of bladder cancer due to its simplicity and ease of operation. However, due to incomplete resection, a high rate of tumor recurrence is likely to occur after surgery, often requiring regular intravesical infusion therapy with chemotherapeutic drugs. In addition, frequent application of chemotherapeutic drugs not only causes great side effects to human body, but also results in increased drug resistance, thus being extremely unfavorable for the treatment of bladder cancer. Therefore, there is a great need to develop new therapeutic strategies for UBC.
Near Infrared (NIR) photoresponsive materials have gained widespread interest in the area of cancer photothermal therapy (PTT) due to their minimal invasiveness and large depth of penetration. Under the traditional tail vein injection treatment mode, the accumulated concentration of the nano material at the tumor part is low, and the photo-thermal treatment effect of the tumor part is greatly weakened. In addition, the photothermal material generates certain toxic and side effects on normal tissues and organs when circulating in vivo. In particular, some nanomaterials pose a long-term risk to the organism due to their non-degradability. In addition, photothermal agents present major difficulties in treating UBC due to the complex fluid environment within the urothelial bladder.
In order to overcome the limitations, the development of a novel photothermal material for targeted therapy of UBC and reduction of tumor recurrence and metastasis rate have great clinical significance.
Disclosure of Invention
In view of the above, the technical problem to be solved by the present invention is to provide an injectable rare earth-DNA photothermal hydrogel, which has high photothermal conversion efficiency, and can generate active oxygen during irradiation process, so that tumors can be substantially eliminated by one treatment, and thus, hydrogel degradation in any time period can be realized, the probability of tumor recurrence and metastasis is reduced to a certain extent, and meanwhile, the injectable rare earth-DNA photothermal hydrogel has good biocompatibility.
The invention provides an injectable rare earth-DNA photo-thermal hydrogel, which comprises the following components:
DNA molecules with negative charges and rare earth-based nano materials with positive charges;
the rare earth-based nano material with positive charges is UCNP-Au.
Preferably, the negatively charged DNA molecule is salmon sperm DNA.
Preferably, the mass fraction ratio of the DNA molecules with negative charges to the rare earth-based nano material with positive charges is (15-20): 1.
the invention provides a preparation method of injectable rare earth-DNA photo-thermal hydrogel, which comprises the following steps:
A) dissolving DNA with negative charges in deionized water, and dispersing to obtain smDNA aqueous solution;
B) heating the smDNA aqueous solution and then annealing to obtain DNA sol;
C) and (3) crosslinking the UCNP-Au and the DNA sol to obtain the rare earth-DNA photo-thermal hydrogel.
Preferably, the UCNP-Au is prepared by the following method:
preparing UCNP nanoparticles;
mixing UCNP nano-particles with hydrochloric acid and an organic solvent, then mixing with polyethyleneimine, stirring, and then washing to obtain UCNP @ PEI;
and mixing the UCNP @ PEI and the chloroauric acid solution, and heating under the condition that the pH value is 7 to obtain the UCNP-Au.
Preferably, the negatively charged DNA molecule is salmon sperm DNA.
Preferably, the concentration of the DNA molecules with negative charges in the rare earth-DNA photo-thermal hydrogel is 3-4 w%, and the concentration of UCNP-Au is 0.2 w%;
the mass fraction ratio of the DNA molecules with negative charges to the rare earth-based nano material with positive charges is (15-20): 1.
preferably, the heating temperature in the step B) is 85-95 ℃; the heating time is 2-3 min.
The rare earth-DNA photothermal hydrogel provided by the invention or prepared by the preparation method provided by any one of the above technical schemes is applied to preparation of a medicine for treating tumors.
Preferably, the drug is a photosensitizer, and the light is 808nm laser; the dosage form of the medicine is injection.
Compared with the prior art, the invention provides an injectable rare earth-DNA photothermal hydrogel, which comprises the following components: DNA molecules with negative charges and rare earth-based nano materials with positive charges; the rare earth-based nano material with positive charges is UCNP-Au. The DNA molecule and the rare earth-based nano material form a hydrogel material with injectability, high-efficiency photothermal treatment effect and biocompatibility under the action of physical winding, hydrogen bonds and electrostatic force. Experimental results show that compared with the traditional inorganic photothermal material, the injectable rare earth-DNA photothermal hydrogel has the photothermal efficiency of 42.67%. In addition, the protective effect of the heat shock protein on tumor cells can be weakened by utilizing the high temperature accumulated by the hydrogel, and compared with a pure nano material, the one-time tumor inhibition rate is increased, so that the recurrence and metastasis rate of the tumor is reduced. The toxic and side effects of the medicine on normal tissues in the treatment process can be reduced by a tumor-side injection means which is simpler and more convenient to operate. And moreover, the degradation of the photo-thermal hydrogel in any time period can be realized by regulating and controlling the proportion of the DNA molecules and the rare earth-based nano material. Compared with the existing clinical tumor non-operative treatment means, the invention has simple and rapid operation in the treatment process and low tumor recurrence and metastasis probability.
Drawings
FIG. 1 is a schematic diagram illustrating the synthesis of injectable rare earth-DNA photothermal hydrogel in examples 1 and 2 of the present invention;
FIG. 2 is the shear rheological results of injectable rare earth-DNA photothermal hydrogel of example 1 of the present invention;
FIG. 3 shows the shear rheological results of injectable rare earth-DNA photothermal hydrogel of example 2 of the present invention;
FIG. 4 shows the in vitro photothermal conversion effect of the injectable rare earth-DNA photothermal hydrogel of example 1 of the present invention;
FIG. 5 shows that the upconversion-gold hybrid nanomaterial in example 1 of the present invention generates oxygen radicals under irradiation of light, thereby having a photodynamic effect;
FIG. 6 is the anti-mouse bladder cancer tumor status of injectable rare earth-DNA photothermal hydrogel of example 1 of the present invention;
FIG. 7 shows the H & E results of the tumor sections after laser irradiation of the injectable rare earth-DNA photothermal hydrogel of example 1.
Detailed Description
The invention provides an injectable rare earth-DNA photo-thermal hydrogel, a preparation method and application thereof, and a person skilled in the art can realize the injection by appropriately improving process parameters by referring to the content. It is expressly intended that all such similar substitutes and modifications apparent to those skilled in the art are deemed to be within the scope of the invention. While the methods and applications of this invention have been described in terms of preferred embodiments, it will be apparent to those of ordinary skill in the art that variations and modifications in the methods and applications described herein, as well as other suitable variations and combinations, may be made to implement and use the techniques of this invention without departing from the spirit and scope of the invention.
The invention provides an injectable rare earth-DNA photo-thermal hydrogel, which comprises the following components:
DNA molecules with negative charges and rare earth-based nano materials with positive charges;
the rare earth-based nano material with positive charges is UCNP-Au.
The injectable rare earth-DNA photothermal hydrogel provided by the invention comprises DNA with negative charges.
The DNA molecule with negative charge of the invention is salmon sperm DNA.
The specific extraction method of salmon sperm DNA is not limited in the invention, and the method is well known to those skilled in the art; preferably, salmon sperm DNA is double-stranded DNA with long fragment (2000bp), and may contain sodium salt, and is lyophilized before use.
The injectable rare earth-DNA photo-thermal hydrogel provided by the invention comprises a positively charged rare earth-based nano material. The rare earth-based nano material with positive charges is UCNP-Au.
The UCNP-Au is prepared by the following method:
preparing UCNP nanoparticles;
mixing UCNP nano-particles with hydrochloric acid and an organic solvent, then mixing with polyethyleneimine, stirring, and then washing to obtain UCNP @ PEI;
and mixing the UCNP @ PEI and the chloroauric acid solution, and heating under the condition that the pH value is 7 to obtain the UCNP-Au.
The UCNP-Au of the invention is firstly prepared into UCNP nano-particles.
The method for preparing the UCNP nanoparticles according to the present invention is not limited, and those skilled in the art are familiar with the method.
The preferable concrete is as follows: YCl3、YbCl3、ErCl3Mixing oleic acid and octadecene, stirring, heating to 140 deg.C under nitrogen protectionMaintaining the temperature of 160 ℃ below zero for 30-40 min, and then reducing the temperature to 40-50 ℃ to obtain a first mixed solution;
taking NH4Dissolving F and NaOH in methanol, ultrasonically mixing uniformly, mixing with the first mixed solution, stirring for 1-2 h at 50 ℃, then evaporating to remove the solvent, and washing to obtain a solution A;
mixing YCl3、NdCl3Mixing oleic acid and octadecene, stirring, heating to 160 ℃ under the protection of nitrogen, maintaining for 30-40 min, and then cooling to 40-50 ℃ to obtain a second mixed solution;
taking NH4Dissolving F and NaOH in methanol, ultrasonically mixing uniformly, heating and stirring with the second mixed solution and the solution A, heating to evaporate the solvent, heating to 250-300 ℃ under the protection of nitrogen, and maintaining for 1-1.5 hours. Cooling to room temperature, and washing to obtain the UCNP nano-particles.
The washing is preferably carried out for 2-3 times by using ethanol and water.
And then mixing the UCNP nano particles with hydrochloric acid and an organic solvent, then mixing with polyethyleneimine, stirring, and washing to obtain UCNP @ PEI. Namely: the surface oleic acid molecule is replaced by Polyethyleneimine (PEI) by ligand exchange.
The method specifically comprises the following steps: dispersing UCNP nano particles in a cyclohexane solvent, centrifuging, adding ethanol and hydrochloric acid, carrying out ultrasonic treatment for 20-30 min, then washing, and dispersing in water; mixed with polyethyleneimine dispersed in water, stirred overnight, and the product washed to give UCNP @ PEI. The washing is preferably carried out for 2-3 times by using ethanol and water.
And mixing the UCNP @ PEI and the chloroauric acid solution, and heating under the condition that the pH value is 7 to obtain the UCNP-Au. Namely: and growing gold nanoparticles with ultra-small nanometer size on the surface of the gold nanoparticles in situ by oxidation reduction to obtain UCNP-Au.
According to the invention, the surfactant PEI is used as a connecting bridge of UCNP and Au, compared with common silicon dioxide, the distance between UCNP and Au is shortened, the light transmission efficiency is enhanced, and the photo-thermal conversion capability is further improved.
Dissolving chloroauric acid in water, titrating with NaOH solution until the pH value is 7, then dripping UCNP @ PEI solution, heating and stirring to 60-70 ℃, and maintaining for 4-5 min. Washing with water and ethanol for 3 times to obtain UCNP-Au.
The mass fraction ratio of the DNA molecule with negative charge to the rare earth-based nano material with positive charge is (15-20): 1; specifically, the ratio can be 15:1, 16:1, 17:1, 18:1, 19:1 or 20: 1; or a point value between any two of the above values.
According to the invention, natural biological macromolecules are used for the first time to construct a hydrogel material with photo-thermal performance, and compared with traditional inorganic and organic photo-thermal materials, the biological macromolecules are used as a support of a nano material, and the tumor treatment effect of the photo-thermal hydrogel is improved by increasing the concentration of photo-thermal nanoparticles in unit volume and the biocompatibility of the whole material.
The invention can realize the degradation of the photo-thermal hydrogel in any time period by regulating the proportion of DNA molecules and rare earth-based nano materials. Can prolong and shorten the treatment period according to the treatment needs and avoid the generation of drug resistance caused by multiple times of administration.
The invention provides a preparation method of injectable rare earth-DNA photo-thermal hydrogel, which comprises the following steps:
A) dissolving DNA with negative charges in deionized water, and dispersing to obtain smDNA aqueous solution;
B) heating the smDNA aqueous solution and then annealing to obtain DNA sol;
C) and (3) crosslinking the UCNP-Au and the DNA sol to obtain the rare earth-DNA photo-thermal hydrogel.
The preparation method of the injectable rare earth-DNA photo-thermal hydrogel provided by the invention comprises the steps of dissolving DNA with negative charges in deionized water, and dispersing to obtain smDNA aqueous solution.
The DNA molecule with negative charge of the invention is salmon sperm DNA.
The specific extraction method of salmon sperm DNA is not limited in the invention, and the method is well known to those skilled in the art; preferably, salmon sperm DNA is double-stranded DNA with long fragment (2000bp), and may contain sodium salt, and is lyophilized before use.
The negatively charged DNA was dissolved in deionized water, degassed, and allowed to stand overnight.
Heating the smDNA aqueous solution and then annealing to obtain DNA sol; the heating temperature is preferably 85-95 ℃; more preferably 90 ℃; the heating time is 2-3 min;
the prepared salmon sperm DNA sol (smDNA) is heated and then rapidly annealed to room temperature. The cross-linking between long DNA chains is improved by the action of forming new hydrogen bonds between single DNA chains through mismatching
The pretreated smDNA is used as a support of the nano material, and the tumor treatment effect of the photo-thermal hydrogel is improved by increasing the photo-thermal conversion efficiency and the overall biocompatibility of the material.
UCNP-Au is provided. The UCNP-Au is prepared by the following method:
preparing UCNP nanoparticles;
mixing UCNP nano-particles with hydrochloric acid and an organic solvent, then mixing with polyethyleneimine, stirring, and then washing to obtain UCNP @ PEI;
and mixing the UCNP @ PEI and the chloroauric acid solution, and heating under the condition that the pH value is 7 to obtain the UCNP-Au.
The present invention has been described above for the UCNP-Au clearly, and is not described herein again.
And carrying out secondary crosslinking on the UCNP-Au and the DNA sol by using Van der Waals force to obtain the rare earth-DNA photo-thermal hydrogel.
Adding UCNP-Au aqueous solution under the condition that the DNA solution still keeps fluidity, uniformly mixing, and realizing secondary crosslinking by using electrostatic force between the UCNP-Au aqueous solution and the UCNP-Au aqueous solution to obtain the final product rare earth-DNA hydrogel.
The DNA molecule with negative charge and the rare earth-based nano material with positive charge form hydrogel mainly through physical winding, hydrogen bonds and electrostatic acting force.
Adding UCNP-Au after smDNA pretreatment, mixing uniformly and storing at 4 DEG C
According to the invention, the concentration of DNA molecules with negative charges in the rare earth-DNA photo-thermal hydrogel is 3-4 w%, and the concentration of UCNP-Au is 0.2 w%.
The mass fraction ratio of the DNA molecules with negative charges to the rare earth-based nano material with positive charges is (15-20): 1; preferably (16-19): 1; specifically, the ratio can be 15:1, 16:1, 17:1, 18:1, 19:1 or 20: 1; or a point value between any two of the above values.
The rare earth-DNA photothermal hydrogel provided by the invention or prepared by the preparation method provided by any one of the above technical schemes is applied to preparation of a medicine for treating tumors.
Wherein the medicine is a photosensitizer, and the light is 808nm laser; the dosage form of the medicine is injection.
Namely, the hydrogel of the invention utilizes heat (the temperature is up to 70-90 ℃ in three minutes under the low-power laser irradiation, the temperature is 20-40 ℃ higher than that of a common inorganic and organic photothermal material) and active oxygen (ROS) generated by 808nm laser irradiation to kill the tumor by dissolving cell membranes and denaturing protein.
The UCNP-Au converts near infrared light penetrating deeper into tissue into visible light that can be absorbed by Au by UCNP, and then converts light into heat and generates Reactive Oxygen Species (ROS) using photothermal conversion and photodynamic properties of Au.
The photothermal hydrogel provided by the invention has injectability, and can be used for treating tumors in situ by a simpler and more convenient operation method. Compared with conventional drug treatment, the toxic and side effects of the drug on normal tissues in the treatment process are reduced.
By utilizing the photo-thermal conversion and the photodynamic property of Au, light can be converted into heat and active oxygen can be generated. In one aspect, cell membranes are lysed and proteins denatured by high temperature; on the other hand, ROS is used for directly killing cell membranes, organelles and functional biomolecules in cells. Realizing synergistic high-efficiency treatment.
The invention provides an injectable rare earth-DNA photo-thermal hydrogel, which comprises the following components: DNA molecules with negative charges and rare earth-based nano materials with positive charges; the rare earth-based nano material with positive charges is UCNP-Au. The DNA molecule and the rare earth-based nano material form a hydrogel material with injectability, high-efficiency photothermal treatment effect and biocompatibility under the action of physical winding, hydrogen bonds and electrostatic force. Experimental results show that compared with the traditional inorganic photothermal material, the injectable rare earth-DNA photothermal hydrogel has the photothermal efficiency of 42.67%. In addition, the protective effect of the heat shock protein on tumor cells can be weakened by utilizing the high temperature accumulated by the hydrogel, and compared with a pure nano material, the one-time tumor inhibition rate is increased, so that the recurrence and metastasis rate of the tumor is reduced. The toxic and side effects of the medicine on normal tissues in the treatment process can be reduced by a tumor-side injection means which is simpler and more convenient to operate. And moreover, the degradation of the photo-thermal hydrogel in any time period can be realized by regulating and controlling the proportion of the DNA molecules and the rare earth-based nano material. Compared with the existing clinical tumor non-operative treatment means, the invention has simple and rapid operation in the treatment process and low tumor recurrence and metastasis probability.
In order to further illustrate the present invention, the following will describe in detail an injectable rare earth-DNA photothermal hydrogel, its preparation method and application in combination with the examples.
Example 1
The invention provides a hydrogel which has good biocompatibility, can be injected, has obvious photo-thermal conversion effect and has a photodynamic effect, and is prepared from a DNA molecule with negative charge, namely salmon sperm DNA and UCNP-Au nano particles with a large number of positive charges, and the preparation process comprises the following steps:
1. synthesizing UCNP-Au nanoparticles with a large number of positive charges:
the first step is as follows: preparing a shell-core hexagonal phase-rare earth doped fluorescence up-conversion nano material (beta OA-UCNP) by a high-temperature oil solvent method. The method comprises the following specific steps: YCl3152.3mg, YbCl355.88mg and ErCl35.47mg were added to a 100mL round bottom flask, followed by 6mL oleic acid and 18mL octadecene, with constant stirring. After the air is exhausted, the temperature is raised to 160 ℃ under the protection of nitrogen and maintained for 40min, and then the temperature is reduced to 50 ℃. Take 149mg NH4F and 100mg NaOH are dissolved in 10mL of methanol, and the mixture is added into the solution after ultrasonic mixing and stirred for 2h at 50 ℃. The solution was heated to 100 ℃ to distill off the methanol, heated to 300 ℃ under nitrogen protection, and maintained for 1 h. The temperature is reduced to room temperature, the ethanol/cyclohexane solution is alternately washed for 3 times, and finally the solution A is dispersed in 10mL of cyclohexane. Getting YCl3156.3mg, NdCl350.1mg was added to a 100mL round bottom flask followed by 6mL oleic acid and 18Octadecene ml, continue stirring. After the air is exhausted, the temperature is raised to 160 ℃ under the protection of nitrogen and maintained for 40min, and then the temperature is reduced to 50 ℃. Take 149mg NH4F and 100mg NaOH are dissolved in 10mL of methanol, and after ultrasonic mixing, the mixture is added into the solution, and then 10mL of the solution A is added, and the mixture is stirred for 2 hours at 50 ℃. The solution is heated to 100 ℃ to evaporate the methanol and the cyclohexane, and is heated to 300 ℃ under the protection of nitrogen for 1 hour. The temperature is reduced to room temperature, the ethanol/cyclohexane solution is washed for 3 times alternately, and finally the obtained up-conversion nanoparticles (UCNPs) are dispersed in 10mL of cyclohexane.
The second step is that: the surface oleic acid molecule is replaced by Polyethyleneimine (PEI) by ligand exchange. The method comprises the following specific steps: 10mL of UCNPs cyclohexane solution was centrifuged, and 10mL of ethanol and 10mL of hydrochloric acid (1M) were added to the pellet and sonicated together for 30 min. After washing with water and ethanol for 3 times, the mixture was dispersed in 10mL of deionized water. 100mg of polyethyleneimine was dissolved in 10mL of deionized water, and then added to the above solution, followed by stirring overnight. The final product was washed 3 times with ethanol and water, and the UCNP @ PEI was dispersed in 10mL deionized water for use.
The third step: and then growing gold nanoparticles with ultra-small nanometer sizes on the surface of the gold nanoparticles in situ through oxidation reduction to obtain UCNP-Au. The method comprises the following specific steps: 13mg of chloroauric acid was dissolved in 5mL of deionized water, titrated to pH 7 with 0.1M NaOH solution, then dropped into 10mL of UCNP @ PEI solution, heated and stirred to 70 ℃ for 5 min. Washing with water and ethanol for 3 times to obtain UCNP-Au.
In the invention, the beta-phase OA-UCNP is doped by rare earth elements such as Y, Yb, Nd, Er and the like, and the Nd ions absorb near-infrared laser with the wavelength of 808nm, and transmit energy to the Er ions to emit green and red visible light. After the PEI is coated on the surface of the UCNP through ligand exchange, the nano material is ensured to have positive charges in an aqueous solution due to the high-density amino groups. The method is characterized in that ultra-small nanometer Au nanoparticles (less than or equal to 2nm) are grown on the surface of UCNP @ PEI in situ by using the redox effect, and the size of UCNP-Au is about 36nm as shown by a transmission electron microscope. UCNP-Au can be uniformly dispersed in an aqueous solution (hydrated particle size 247.9 nm). Each 1mg of the UCNP-Au hybrid material measured by an inductively coupled plasma emission spectrometer contained Au47.2. mu.g.
2. Preparation of rare earth-DNA hydrogel
The first step is as follows: the negatively charged DNA molecule, salmon sperm DNA (2000bp), was dispersed in an aqueous solution, vacuolated, and allowed to stand overnight. Viscous DNA sols are obtained due to physical entanglement between long DNA strands.
The second step is that: the prepared salmon sperm DNA sol (smDNA) was heated at 90 ℃ for 2 minutes, and then rapidly annealed to room temperature. Because new hydrogen bonds are formed between the DNA single chains through mismatching, the effect is to promote the cross-linking between the DNA long chains.
The third step: adding UCNP-Au aqueous solution under the condition that the DNA solution still keeps fluidity, uniformly mixing, and realizing secondary crosslinking by using electrostatic force between the UCNP-Au aqueous solution and the DNA solution to obtain the final product rare earth-DNA hydrogel (DNA4 w%, UCNP-Au0.2w%).
The above preparation process can be referred to FIG. 1. In example 1, the mass fraction ratio of salmon sperm DNA (2000bp) to UCNP-Au nanoparticles was 20: 1.
example 2
Example 2 the procedure of example 1 was similar, except that the mass fraction ratio of salmon sperm DNA (2000bp) to UCNP-Au nanoparticles was 15: 1.
example 3
To further illustrate the above embodiments, the photothermal conversion, photodynamic, in vivo antitumor properties of the injectable rare earth-DNA photothermal hydrogel provided by the present invention will be described in detail below with reference to the accompanying drawings, but the present invention should not be construed as being limited to the scope of the present invention.
FIG. 2 shows the shear rheological results of the injectable rare earth-DNA photothermal hydrogel of example 1, in which the viscosity of the gel decreases with increasing shear frequency, which is a necessary factor for ensuring the injectability of the gel. The hydrogel properties were fully demonstrated by the fact that G' was always close to, but greater than, G "at all shear frequencies.
FIG. 3 shows the shear rheological results of the injectable rare earth-DNA photothermal hydrogel of example 2.
FIG. 4 shows the photothermal conversion results of the injectable rare earth-DNA photothermal hydrogel of example 1. In vitro experiments show that the hydrogel has higher photothermal conversion efficiency (42.67% vs 32.44%) compared with the pure nanoparticles.
Fig. 5 shows the photodynamic effect of example 1. Green fluorescence represents the production of Reactive Oxygen Species (ROS) within the cell, which can directly kill cell membranes, organelles, and functional biomolecules within the cell.
FIG. 6 shows the tumor-bearing (human bladder cancer) test in mice of example 1. 100 μ L of rare earth-DNA photothermal hydrogel (smDNA 4W%, UCNP-Au0.2w%) was injected peritumorally into BABL/c nude mice, followed by a single laser irradiation (808nm laser, 1.0W. cm)-2For 3 minutes). It can be seen that the tumor body gradually shrinks and disappears after the treatment, and no recurrence is observed at the end of the observation period.
Fig. 7 is the H & E staining results of major organs, and it can be seen that the photothermal hydrogel did not cause significant damage to major organs of mice.
The foregoing is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, various modifications and decorations can be made without departing from the principle of the present invention, and these modifications and decorations should also be regarded as the protection scope of the present invention.

Claims (7)

1. An injectable rare earth-DNA photothermal hydrogel, comprising:
DNA molecules with negative charges and rare earth-based nano materials with positive charges;
the rare earth-based nano material with positive charges is UCNP-Au;
the UCNP-Au is prepared by the following method:
preparing UCNP nanoparticles;
mixing UCNP nano-particles with hydrochloric acid and an organic solvent, then mixing with polyethyleneimine, stirring, and then washing to obtain UCNP @ PEI;
mixing the UCNP @ PEI and a chloroauric acid solution, and heating under the condition that the pH value is 7 to obtain UCNP-Au;
the DNA molecule with negative charge is salmon sperm DNA;
the mass fraction ratio of the DNA molecules with negative charges to the rare earth-based nano material with positive charges is (15-20): 1.
2. a preparation method of injectable rare earth-DNA photo-thermal hydrogel is characterized by comprising the following steps:
A) dissolving DNA with negative charges in deionized water, and dispersing to obtain smDNA aqueous solution;
B) heating the smDNA aqueous solution and then annealing to obtain DNA sol;
C) crosslinking the UCNP-Au and the DNA sol to obtain rare earth-DNA photo-thermal hydrogel;
the UCNP-Au is prepared by the following method:
preparing UCNP nanoparticles;
mixing UCNP nano-particles with hydrochloric acid and an organic solvent, then mixing with polyethyleneimine, stirring, and then washing to obtain UCNP @ PEI;
and mixing the UCNP @ PEI and the chloroauric acid solution, and heating under the condition that the pH value is 7 to obtain the UCNP-Au.
3. The method of claim 2, wherein the negatively charged DNA molecule is salmon sperm DNA.
4. The preparation method of claim 2, wherein the concentration of the negatively charged DNA molecules in the rare earth-DNA photothermal hydrogel is 3-4 w%, and the concentration of UCNP-Au is 0.2 w%;
the mass fraction ratio of the DNA molecules with negative charges to the UCNP-Au is (15-20): 1.
5. the preparation method according to claim 2, wherein the heating temperature in the step B) is 85-95 ℃; the heating time is 2-3 min.
6. Use of the rare earth-DNA photothermal hydrogel of claim 1 or the rare earth-DNA photothermal hydrogel prepared by the preparation method of any one of claims 2 to 5 in the preparation of a medicament for treating tumors.
7. The use of claim 6, wherein the drug is a photosensitizer and the light is 808nm laser light; the dosage form of the medicine is injection.
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