CN109431967B

CN109431967B - Soluble microneedle for treating psoriatic arthritis

Info

Publication number: CN109431967B
Application number: CN201811590175.7A
Authority: CN
Inventors: 徐月红; 于恺悦; 曹思思; 玉秀明
Original assignee: Sun Yat Sen University
Current assignee: Sun Yat Sen University
Priority date: 2018-12-25
Filing date: 2018-12-25
Publication date: 2021-12-24
Anticipated expiration: 2038-12-25
Also published as: CN109431967A

Abstract

A soluble microneedle for treating psoriatic arthritis, the soluble microneedle comprising a tip layer divided in an insertion direction, an intermediate layer and a base layer, wherein the tip layer comprises a drug for treating arthritis dispersed in a tip matrix, the intermediate layer comprises a drug for treating psoriasis dispersed in an intermediate layer matrix, and the base layer comprises a base matrix. The microneedle has the effect of treating psoriasis and arthritis simultaneously.

Description

Soluble microneedle for treating psoriatic arthritis

Technical Field

The invention relates to the technical field of medicinal preparations, in particular to a soluble microneedle for treating psoriatic arthritis and a preparation method thereof.

Background

Psoriasis is a chronic recurrent dermatosis which takes erythrodermic scales as main clinical manifestations and takes keratinocyte hyperproliferation and massive inflammatory cell infiltration as main pathological characteristics, and the pathogenesis of the psoriasis is mainly abnormal activation of T cells. The medicine for treating psoriasis mainly comprises macrolide immunomodulators, keratolytic agents, coal tar, glucocorticoids and the like. Psoriasis is difficult to treat, adverse reaction of systemic therapy is more, the psoriasis is difficult to treat and easy to occur, and the mind and body of a patient are greatly influenced. And 10% -30% of patients can have chronic inflammatory arthritis related to psoriasis, called as psoriatic arthritis, which is an independent disease simultaneously presenting psoriasis rash and arthritis (the current treatment of arthritis mainly comprises non-steroidal anti-inflammatory drugs, glucocorticoids, antibiotics and the like). Psoriatic arthritis is a chronic inflammatory arthritic condition that affects the insertion sites of skin, joints, tendons, ligaments, and fascia. Gladman, CurrentopinionRheumatology, "Currentcon captisporatoriaritis," 2002,14: 361-.

Because psoriatic arthritis has similar immune disorders as rheumatoid arthritis, current treatments have mainly adopted a similar treatment regimen as rheumatoid arthritis. Psoriatic arthritis has both articular and cutaneous lesions, and therefore a truly effective treatment should achieve relief for both. However, the current clinical therapies cannot achieve the aim of simultaneously relieving the two diseases.

Diclofenac sodium is a nonsteroidal anti-inflammatory analgesic derived from phenylacetic acids, which acts by inhibiting cyclooxygenase activity to block the conversion of arachidonic acid to prostaglandins, and is commonly used in the treatment of rheumatoid arthritis and osteoarthritis. However, in order to achieve the therapeutic effect, diclofenac sodium needs to be injected into the articular cavity to exert the drug effect, and the compliance of patients is poor. Tacrolimus is a macrolide immunomodulator, is effectively applied to the treatment of immune-related skin diseases, including atopic dermatitis (eczema), psoriasis and the like, by inhibiting the dephosphorylation of calcineurin, interfering the expression of cytokines and down-regulating the activation of T cells. However, due to the high hydrophobicity and high molecular weight of tacrolimus, it is limited in percutaneous absorption and skin retention, and is not beneficial to the treatment of local skin diseases.

In conclusion, there is still a lack of drugs clinically available that can achieve therapeutic effects of simultaneously relieving psoriasis and arthritis, and that can improve the problem of patient compliance in current treatments.

Disclosure of Invention

The invention aims to solve the technical problem that in order to overcome the problems in the prior art, the invention provides a soluble microneedle for simultaneously treating psoriasis and arthritis and a preparation method thereof. The soluble microneedle is applied to the affected part, the diclofenac sodium at the needle point can penetrate through the skin to reach the joint cavity to treat arthritis, and the tacrolimus at the middle layer enters the skin along the pores generated by the needle point to treat psoriasis, so that the symptoms of arthritis and dermatitis are simultaneously relieved, and the aim of treating psoriatic arthritis is fulfilled.

The invention provides a soluble microneedle for treating psoriatic arthritis, which consists of a needle point layer, an intermediate layer and a base layer divided in an insertion direction, wherein the needle point layer comprises a medicament for treating the psoriatic arthritis dispersed in a needle point layer matrix, the intermediate layer comprises a medicament for treating the psoriatic arthritis dispersed in an intermediate layer matrix, and the base layer comprises a base layer matrix.

In some embodiments, the agent for treating arthritis is selected from the group consisting of non-steroidal anti-inflammatory drugs, glucocorticoids, and antibiotics, and the agent for treating psoriasis is selected from the group consisting of macrolide immunomodulators, keratolytic agents, coal tar, and glucocorticoids.

In some embodiments, the non-steroidal anti-inflammatory drug is diclofenac sodium. In some embodiments, wherein the macrolide immunomodulator is tacrolimus. In some embodiments, wherein the non-steroidal anti-inflammatory drug is diclofenac sodium and the macrolide immunomodulator is tacrolimus.

In some embodiments, wherein the tip layer matrix and the base layer matrix each comprise: sodium hyaluronate, dextran 40, povidone K17 and water. Preferably, the microneedle tip substrate material consists of the following components in percentage by mass: 10% -30% of sodium hyaluronate (with the molecular weight of 60 KD-100 KD), 4030% -50% of glucan, 1710% -30% of povidone K and the balance of water. Preferably, the microneedle base matrix material consists of the following components in mass ratio: 10% -30% of sodium hyaluronate, 4030% -50% of glucan, 1710% -30% of povidone K and the balance of water.

In some embodiments, wherein the interlayer matrix comprises: sodium hyaluronate, dextran 40, povidone K17, niacinamide and water. Preferably, the microneedle intermediate layer matrix material consists of the following components in percentage by mass: 10% -30% of sodium hyaluronate (with the molecular weight of 60 KD-100 KD), 4030% -50% of glucan, 1710% -30% of povidone K, 5% -25% of nicotinamide and the balance of water.

In some embodiments, wherein the total length of the tip layer and the intermediate layer of the microneedle in the insertion direction is 100-.

The present invention also provides a microneedle array comprising a plurality of microneedles as described above, wherein the base layer of each microneedle is formed in the same plane and is integral.

In another aspect, the present invention provides a method for preparing soluble microneedles for treating psoriatic arthritis, which comprises the following steps:

A. dispersing diclofenac sodium in a microneedle tip substrate at room temperature, injecting the substrate into a microneedle mould, centrifuging, repeatedly reversing the mould to uniformly distribute the substrate, and drying for 2 hours to prepare the microneedle tip carrying the diclofenac sodium;

B. dispersing tacrolimus in a microneedle intermediate layer matrix at room temperature, taking the microneedle mould containing the diclofenac sodium needle points, adding the intermediate layer matrix, centrifuging, repeatedly reversing the mould to enable the matrix to be uniformly distributed, and drying for 2 hours to prepare the microneedle with the needle points carrying the diclofenac sodium and the intermediate layer carrying the tacrolimus;

C. adding the microneedle base matrix into the microneedle mould containing the needle point and the middle layer at room temperature, centrifuging and repeatedly inverting the mould to ensure that the base matrix is uniformly and flatly distributed, and drying at 4 ℃ for 24 hours to prepare the soluble microneedle co-administration system as claimed in claim 1.

The soluble microneedle can provide patient compliance, and when a psoriasis patient is complicated with arthritis, the microneedle is given at the same focus position, the microneedle co-administration system can simultaneously transmit two medicaments for treating the arthritis and the psoriasis, the medicament at the needle point can be diffused to a joint cavity through the skin, the medicament at the middle layer can enter the skin along a pore generated by the needle point, and the two medicaments respectively exert the curative effects of the two medicaments, so that the aim of simultaneously relieving the skin psoriasis symptom and the joint inflammation is fulfilled, and the treatment of the psoriatic arthritis is realized.

Drawings

Fig. 1 is a schematic view of soluble microneedle preparation.

Fig. 2A investigation of mechanical strength of soluble microneedles; fig. 2B soluble microneedle simulated penetration depth (a) and skin penetration depth (B) investigation; fig. 2C is a confocal laser microscope observation of soluble microneedle morphology and drug profile; fig. 2D confocal laser microscopy the 3D reconstructed image of drug distribution after the microneedles penetrated the skin.

Fig. 3 a schematic diagram of arthritis rat modeling (a) and treatment (B).

FIG. 4A is a hematoxylin-eosin stained section of the knee joint of a rat model for the treatment of arthritis at various locations; FIG. 4B shows safranin-fast green stained sections of knee joints of rat model for treatment of arthritis at various sites.

Figure 5A psoriasis lesion area and severity index assessment; is the statistical difference from the modeling group (P<0.05); # is statistically different from the mixed monolayer microneedle set (P<0.05); FIG. 5B evaluation of transdermal water loss; is the statistical difference from the modeling group (P<0.05); # is statistically different from the mixed monolayer microneedle set (P<0.05）。

FIG. 6 evaluation of skin epidermal thickness of rat model treated with psoriasis for each dosing group.

Detailed Description

Example 1 preparation of soluble microneedles

The microneedle mould is made of polydimethylsiloxane, and 144 rows of inverted right rectangular pyramid pores with the depth of about 600 mu m and the bottom diameter of about 300 mu m are arranged on the surface of a square base with the bottom surface of 12mm in length, 12mm in width and 2mm in height. The mold was used in this example and subsequent examples and comparative examples.

As shown in fig. 1, the preparation method of the soluble microneedle is performed according to the following steps:

A. and (2) dispersing 400g of dextran 40 into 1000mL of water, heating to 80 ℃ to fully dissolve the dextran 40, respectively adding 100g of povidone K17 and 100g of hyaluronic acid into the dextran 40 water solution when the dextran 40 water solution is returned to room temperature, and stirring to fully dissolve the dextran 40 water solution to obtain the microneedle tip substrate. Dispersing 16g of diclofenac sodium in the microneedle tip matrix at room temperature, injecting the matrix into a microneedle mould, centrifuging, repeatedly reversing the mould to uniformly distribute the matrix, and drying for 2 hours to prepare the microneedle tip carrying the diclofenac sodium.

B. And (2) dispersing 350g of dextran 40 in 1000mL of water, heating to 80 ℃ to fully dissolve the dextran 40, respectively adding 150g of povidone K17, 200g of nicotinamide and 100g of hyaluronic acid into the dextran 40 aqueous solution when the dextran 40 aqueous solution is returned to room temperature, and stirring to fully dissolve the dextran 40 aqueous solution to obtain the microneedle intermediate layer matrix. At room temperature, 1.8g of tacrolimus was dispersed in the microneedle interlayer matrix to prepare a tacrolimus-loaded interlayer. And (C) taking the microneedle tip with the diclofenac sodium needle tip prepared in the step (A), adding the middle layer loaded with tacrolimus, centrifuging, repeatedly reversing the mold to uniformly distribute the matrix, and drying for 2 hours to prepare the microneedle with the diclofenac sodium needle tip and the tacrolimus loaded in the middle layer.

C. And (2) dispersing 300g of dextran 40 into 1000mL of water, heating to 80 ℃ to fully dissolve the dextran 40, sequentially adding 100g of povidone K17 and 300g of hyaluronic acid into the dextran 40 aqueous solution respectively after the dextran 40 aqueous solution is returned to room temperature, and stirring to fully dissolve the dextran 40 aqueous solution to obtain the microneedle base matrix. And (3) at room temperature, adding the microneedle base substrate into the microneedle mould which is prepared in the step (B) and contains the needlepoint and the middle layer, centrifuging, repeatedly reversing the mould to uniformly and flatly distribute the base substrate, and drying at 4 ℃ for 24 hours to prepare the soluble microneedle.

Comparative example 1 needle-tip diclofenac sodium microneedle

The microneedle tips carrying diclofenac sodium were prepared as described in step a of example 1. Then adding the microneedle interlayer matrix (without tacrolimus) prepared according to the step B in the example 1 to the microneedle tip mould carrying the diclofenac sodium tip, centrifuging and repeatedly reversing the mould to uniformly distribute the matrix, and drying for 2 hours. And finally, adding the microneedle base substrate prepared according to the step C in the example 1 into the microneedle mould containing the diclofenac sodium needlepoint and the blank intermediate layer, centrifuging and repeatedly reversing the mould to ensure that the base substrate is uniformly and flatly distributed, and drying at 4 ℃ for 24 hours to prepare the needlepoint diclofenac sodium microneedle.

Comparative example 2. intermediate layer tacrolimus microneedle

The microneedle tip substrate (without diclofenac sodium) prepared according to example 1 was injected into a microneedle mold, centrifuged and the mold was repeatedly inverted to uniformly distribute the substrate, and dried for 2 hours to prepare a microneedle tip without drug. The tacrolimus-loaded intermediate layer prepared according to the step B in the example 1 was then added to the microneedle mould with the tips of the above microneedles not containing the drug, centrifuged and the mould was repeatedly inverted to uniformly distribute the matrix, and dried for 2 hours. And finally, adding the microneedle base matrix prepared in the step C in the embodiment 1, centrifuging, repeatedly reversing the mold to enable the base matrix to be uniformly and flatly distributed, and drying at 4 ℃ for 24 hours to prepare the intermediate layer tacrolimus microneedle.

Comparative example 3 Mixed monolayer microneedles

And (2) dispersing 350g of dextran 40 in 1000mL of water, heating to 80 ℃ to fully dissolve the dextran 40, respectively adding 150g of povidone K17, 200g of nicotinamide and 100g of hyaluronic acid into the dextran 40 aqueous solution in sequence after the dextran 40 aqueous solution is returned to room temperature, and stirring to fully dissolve to obtain the mixed single-layer microneedle matrix. At room temperature, dispersing 16g of diclofenac sodium and 1.8g of tacrolimus in a microneedle substrate in sequence, injecting the substrate into a microneedle mould, centrifuging and repeatedly reversing the mould to uniformly distribute the substrate, and drying for 2 hours to prepare the microneedle tip carrying the diclofenac sodium and the tacrolimus. Adding a microneedle base matrix into the microneedle mould with the needle point, centrifuging and repeatedly reversing the mould to ensure that the base matrix is uniformly and flatly distributed, and drying for 24 hours at 4 ℃ to prepare the mixed monolayer microneedle.

Example 2 characterization of the soluble microneedles of example 1

Examination of mechanical Strength of microneedles

The microneedle prepared in example 1 was placed on a texture analyzer with the tip facing upward, and the probe of the texture analyzer was set to move downward at a speed of 1mm/s, so that a force-displacement curve could be obtained when the probe pressed the microneedle, thereby reflecting the mechanical strength of the microneedle. As shown in fig. 2A, the soluble microneedle of example 1 has good mechanical strength, which can be 90N or more without breaking the tip.

Investigation of microneedle penetration depth

Taking Parafilm sealing film, tiling 10 layers for simulating skin, placing 10 layers of sealing film on a texture analyzer, fixing a microneedle on a probe, setting the probe to move downwards at the speed of 1mm/s with the needle point facing downwards, setting the force of 30N for pressing the microneedle, observing the number of layers penetrated by the microneedle and the hole rate caused on the sealing film, and calculating the penetration depth of the microneedle by 100 mu m of each layer of sealing film. Meanwhile, the depth of the microneedle penetrating into the skin is observed by using an Optical Coherence Tomography (OCT) technology, the abdominal skin of the SD rat is subjected to unhairing treatment, the microneedle is pressed on the skin with the force of 30N to penetrate into the skin, the depth of the microneedle penetrating into the skin is observed by using the OCT technology, and the laser wavelength of an OCT system is 1310 nm. As can be seen from FIG. 2B, the depth of penetration of the soluble microneedles into the skin can reach 300 μm, which satisfies the requirements of transdermal drug delivery for the treatment of arthritis.

Laser confocal microscope for observing the shape of the microneedle and the distribution of the two drugs after penetrating into the skin

In order to observe the shape of the microneedle and the distribution of the two drugs in the microneedle using a confocal laser microscope, the fluorescent probe was used as a model drug in the experiment. Diclofenac sodium is a hydrophilic drug, so water-soluble rhodamine B (developing red fluorescence) is used as a model drug; tacrolimus is a hydrophobic drug so hydrophobic coumarin 6 (which exhibits green fluorescence) has been used as its model drug. The method of example 1 was used to prepare soluble microneedles using rhodamine B and coumarin 6, respectively, instead of diclofenac sodium and tacrolimus, and the morphology of the microneedles and the distribution of the two model drugs were observed using a laser confocal microscope. As shown in fig. 2C, the total length of the soluble microneedle corresponding to example 1 was 600 μm, the hydrophilic model drug rhodamine B was distributed at the tip of the microneedle, the hydrophobic model drug coumarin 6 was distributed at the intermediate layer, and the two drugs were distributed in a double layer in accordance with the design prediction.

About 350g of SD rat (9-10 weeks old), shaving the hair of the knee part with a shaver, anesthetizing with 20% urethane, penetrating the soluble microneedle carrying the fluorescent model drug into the inner side of the knee, pressing with 30N force for 5min, fixing the microneedle on the skin with 3M medical adhesive tape, removing the microneedle after 2h, dislocating the neck of the rat, taking the skin of the knee part, placing the skin on a glass slide with the stratum corneum facing downwards, observing the distribution of the drug in each layer of the skin by layer scanning with a laser confocal microscope, and reconstructing with 3D to obtain a drug distribution map. As shown in FIG. 2D, the needle tip drug rhodamine B can reach about 250 μm deeper layer of skin, which is beneficial for the drug to enter the joint cavity through the skin to exert drug effect; the intermediate layer medicine coumarin 6 is mainly distributed on the epidermal layer of the skin at a position within about 100 mu m, which indicates that the medicine is mostly remained in the skin and is beneficial to treating diseases in the skin by the medicine, thereby achieving the purpose of simultaneously treating arthritis and psoriasis by the soluble micro-needle.

Example 3 in vitro transdermal assay of soluble microneedles

The experiment was divided into four groups: a bilayer microneedle set (microneedles of example 1), a needle point diclofenac sodium microneedle set (microneedles of comparative example 1), a middle layer tacrolimus microneedle set (microneedles of comparative example 2), and a mixed monolayer microneedle set (microneedles of comparative example 3). After the neck of the male SD rat is cut off, the hair at the knee part of the male SD rat is immediately shaved off, the skin at the abdomen is peeled off, the subcutaneous fat tissue is removed, and the male SD rat is washed clean by physiological saline and stored at the temperature of minus 20 ℃ for standby. Thawing with normal saline before use, and drying surface water with filter paper. The integrity of the rat skin was checked before each experiment. In vitro transdermal experiments rats were held with abdominal skin between the donor and recipient cells using a modified Franz diffusion cell with the stratum corneum facing the donor cell. The effective transdermal area of the diffusion cell is 1.766 cm²The receiving pool volume was 8 mL. Administering soluble microneedles (N = 4) containing diclofenac sodium 300 μ g at the tip and tacrolimus 20 μ g in the middle layer, respectively, in a supply tank, pressing with a force of 30N for 5min, and fixing with 3M medical tape; the receiving solution is normal saline containing 20% ethanol, and an in vitro transdermal experiment is carried out for 24 h. Taking 1mL of receiving solution in 0.5, 1, 2, 4, 6, 9, 12, 16, 20 and 24 hours, and supplementing blank receiving solution with the same volume; filtering the sample with 0.22 μm microporous membrane, and respectively determining the content of diclofenac sodium and tacrolimus according to formula Qn = (C: (C))C _n×V ₀+

)/AAnd calculating the cumulative permeability per unit area. Where Cn denotes the concentration measured at the nth sampling point, Ci denotes the concentration measured at the ith sampling point, and V₀Denotes the volume of the receiving reservoir, V denotes the volume per sampling, A denotes the effective transdermal area, Qn (. mu.g/cm)²) Cumulative flux per unit area. After the in vitro transdermal experiment is ended, taking out the skin of a rat from a diffusion cell, washing the surface of the rat with water, adhering residual medicines on the surface of the rat skin with a 3M medical adhesive tape, shearing the rat skin, adding 1mL of methanol, carrying out ultrasonic extraction for 30min, filtering the supernatant with a 0.22 mu M microporous filter membrane, taking the subsequent filtrate, and measuring the contents of diclofenac sodium and tacrolimus in the filtrate by adopting a high performance liquid chromatography to obtain the skin retention.

Table 1 diclofenac sodium in different soluble microneedles in vitro transdermal results (n = 4)

Remarking: is the statistical difference between the mixed monolayer of the microneedles: (P<0.05）。

Table 2 tacrolimus transdermal in vitro results in different soluble microneedles (n = 4)

As can be seen from Table 1, the diclofenac sodium 24h cumulative permeation amount of the double-layer microneedle and the needlepoint diclofenac sodium microneedle is superior to that of the mixed single-layer microneedle (P<0.05), and no statistical difference exists between the double-layer microneedle and the point diclofenac sodium microneedle, which shows that the external transdermal behavior of the diclofenac sodium with the point of the double-layer microneedle is the same as that of single administration, and is obviously superior to that of a mixed single-layer microneedle. The penetration of each group of micro-needles is obviously higher than that of the skinThe retention of skin indicates that most of the drugs at the needle point can penetrate through the skin to reach the subcutaneous part and reach the joint to exert the drug effect. As can be seen from table 2, the 24h cumulative permeation amount and 24h skin retention of tacrolimus of the double-layer microneedle and the intermediate-layer tacrolimus microneedle are both superior to those of the mixed single-layer microneedle, and there is no statistical difference between the double-layer microneedle and the intermediate-layer tacrolimus microneedle, which indicates that the external transdermal behavior of tacrolimus in the intermediate layer of the double-layer microneedle is the same as that of single administration, and is significantly superior to that of the mixed single-layer microneedle. The skin retention amount and the accumulated permeability of the double-layer microneedle and the middle-layer tacrolimus microneedle are similar, which shows that the microneedle can effectively improve the skin retention amount of the middle-layer drug, is favorable for exerting the drug effect and does not cause the systemic reaction caused by the permeation of a large amount of the drug.

The result of an in vitro transdermal experiment shows that the soluble double-layer microneedle can achieve the purpose of simultaneously delivering a hydrophilic medicament diclofenac sodium at a needle tip and a hydrophobic medicament tacrolimus at a middle layer, the needle tip medicaments penetrate a large amount, the medicament at the middle layer enters the skin along a pore formed by the needle tip, the retention is obviously improved, the construction of a co-administration system is realized, the arthritis and the psoriasis symptoms are simultaneously relieved, and the psoriatic arthritis can be treated.

Example 4 therapeutic Effect of soluble Microtargeting on arthritis

Establishing an arthritis model: in this experiment, about 350g of SD rat (9-10 weeks old) was used as a model animal, a physiological saline mixture containing 2% of lambda carrageenan and 4% of kaolin was used as a modeling agent, and a unilateral modeling follow-up control method was used to establish an arthritis model for the right knee of the rat, and the left knee of the rat was used as a negative control, as shown in FIG. 3. SD rats were anesthetized with 20% urethane, hairs at the knee were shaved with a razor, a 1ml syringe was inserted into the synovial cavity along the ligament at the right knee and 0.1 ml of a molding agent was injected, and the knee was repeatedly bent for 15 min after the injection to uniformly distribute the molding agent in the joint cavity, thereby causing abrasion and swelling of the joint. After molding, the swelling degree of the knee is measured by a vernier caliper to determine whether molding is successful. Rats after molding were randomly divided into four groups (n = 4), which were a molding group, a double-layer microneedle group (microneedles described in example 1), a needle point diclofenac sodium microneedle group (microneedles described in comparative example 1), a middle-layer tacrolimus microneedle group (microneedles described in comparative example 2), and a mixed single-layer microneedle group (microneedles described in comparative example 3), and on the second day after molding, arthritis treatment was performed using the four microneedles, respectively, and no treatment was given to the molding group. The specific administration method is as follows: and (3) pricking microneedles into the inner skin of the knee joint, pressing for 5min by using 30N force, fixing the microneedles by using a 3M medical adhesive tape, taking off the microneedles after 24h, measuring the swelling degree of the knee by using a vernier caliper, and measuring again on the fifth day and the seventh day respectively to determine the treatment effect of the microneedles on the arthritis. Index evaluation: the diameters of the knees of the rats were measured using a vernier caliper on

days

1, 3, 5, and 7, respectively, and the swelling degree of the knees was calculated by the formula (right knee diameter-left knee diameter)/right knee diameter × 100%. The rats were sacrificed on the seventh day of the experiment, and the atrophy degree of the muscles was evaluated by taking gastrocnemius muscle and soleus muscle at the calf part of the rats and using the formula of right calf muscle weight/left calf muscle weight × 100%. Knee joints of rats were isolated and fixed in 4% paraformaldehyde, and paraffin sections (thickness 5 μm) were performed after decalcification treatment, and hematoxylin-eosin staining and safranin-fast green staining were performed, respectively, to observe the conditions of knee joint inflammatory cell infiltration and cartilage erosion.

Table 3 results of soluble microneedle treatment of arthritic knee swelling in rats (n = 4)

Remarking: is the statistical difference from the modeling group (P<0.05); # is statistically different from the middle Tacrolimus microneedle group (P<0.05）；&For the statistical difference from the mixed monolayer microneedle set (P<0.05); @ is the statistical difference between the sample and the needle point diclofenac sodium micro-needle group ((P<0.05）。

The more severe the knee swelling, the more severe the arthritic symptoms, and as can be seen from Table 3, the degree of knee swelling in each group was about 25% in the first day after molding, indicating that the arthritis model was successfully molded in the first day after moldingAfter microneedle administration on the next day, the double-layered microneedles of example 1 significantly reduced the knee swelling, which was superior to that of the other microneedle administration groups and was statistically different (a)P<0.05) while the degree of knee swelling at day seven was still around 20% with the non-dosed stoma set. Wherein the double-layer microneedle has better therapeutic effect than a needle point diclofenac sodium microneedle. The double-layer microneedle has better treatment effect than the mixed single-layer microneedle, and the remarkable advantages of the double-layer structure of the needle point layer and the middle layer are demonstrated.

Table 4 soluble microneedle treatment of arthritic rats hind leg muscular atrophy results (n = 4)

On the side with knee arthritis, the knee fails to support the leg with force due to the inflammatory effect, and atrophy of the calf muscle occurs after a while, so that the smaller the weight ratio of the calf muscle, the more severe the muscular atrophy, and the more severe the arthritis symptoms. As can be seen from Table 4, the weight ratio of calf muscle of the model group is 83.24% + -2.05%, which indicates that the calf muscle has atrophy of about 20%, while the weight ratio of calf muscle of the example 1 after the double-layer microneedle therapy reaches 99.02% + -0.59%, which indicates that almost no muscle atrophy, and indicates that the double-layer microneedle therapy effect is obvious and superior to that of other microneedle administration groups and has statistical differences (the two-layer microneedle therapy effect is superior to that of other microneedle administration groups and the two-layer microneedle therapy effect is obvious) (the two-layer microneedle therapy effect is superior to that of the other microneedle administration groups and has statistical differences: (the invention has the advantages of the inventionP<0.05）。

As can be seen from FIG. 4A, the blank control had smooth and uniform articular cartilage surface, normal chondrocyte morphology, regular arrangement, and no inflammatory cell infiltration in the articular cavity. The articular cartilage of the model building group has rough surface, the arrangement of chondrocytes is disordered, and a large amount of inflammatory cells infiltrate from synovial tissues. Compared with other microneedle treatment groups, the articular cartilage surface is recovered to be smooth after double-layer microneedle treatment, the chondrocytes are regularly arranged and uniformly calcified, and inflammatory cell infiltration does not occur, which shows that the microneedle treatment of the rat model with arthritis in the embodiment 1 has better effect. As shown in FIG. 4B, the joint cartilages can be observed by safranin-fast green staining, the basic dye safranin can be combined with basophilic cartilage to show red color, the acid dye fast green can be combined with acidophilic bone tissue to show green color, namely, the red color in the section is cartilage, and the green color is bone tissue. As can be seen from fig. 4B, the articular cartilage of the model-making group is seriously damaged, and a large area of safranin is lost, but the cartilage safranin staining of the double-layer microneedle treatment group is more uniform and the area distribution is larger than that of other microneedle treatment groups, which is close to that of a blank control, indicating that the double-layer microneedle can effectively alleviate the articular cartilage damage in arthritis and is superior to other groups.

Example 5 therapeutic Effect of soluble Microtargeting on psoriasis

Establishing a psoriasis model: in this experiment, about 200g of SD rat (4-5 weeks old) was used as a model animal, and an imiquimod ointment (containing imiquimod 5%) was used as a model agent to mold the back skin of the rat. SD rats were anesthetized with 20% urethane, and the hair was shaved off on the back with a razor and depilated, with a depilatory area of 2cm × 2 cm. 0.1g of imiquimod cream is evenly smeared on hairless areas, and is continuously administrated for molding for 5 days, so that the back skin of rats has obvious psoriasis symptoms. Rats after 5 days of continuous molding were randomly divided into four groups (n = 4), namely a molding group (no microneedle treatment was given), a double-layer microneedle group (the microneedles described in example 1 were given), a diclofenac sodium microneedle group (the microneedles described in comparative example 1 were given), a middle-layer tacrolimus microneedle group (the microneedles described in comparative example 2 were given), and a mixed single-layer microneedle group (the microneedles described in comparative example 3 were given), and were treated with the four microneedles on the sixth day, respectively, according to the following specific administration methods: the microneedles were pricked into the skin with psoriatic lesions, pressed with 30N force for 5min, then fixed with 3M medical tape, and taken off 24h, i.e. day seven. Index evaluation: observing the skin condition of the administration area by naked eyes on the 1 st, 3 th, 5 th, 7 th, 9 th and 11 th days of the experiment respectively, evaluating the skin damage area and severity index of the psoriasis, and scoring and recording three indexes of erythema, scale and thickening according to the severity respectively, wherein the non-phenomenon is 0 point; slightly 1 point; medium is 2 points; the score is remarkably 3; very significantly 4 points. The total score from 0 to 12 points was used to indicate the severity of psoriasis. And measuring the transdermal water loss value of the skin of the administration area by using a transdermal water loss instrument on

days

0, 1, 3, 5, 7, 9 and 11 respectively, and indicating the condition of the skin barrier function. On day 11 of the experiment, the rats were sacrificed, the skin of the area to be administered was fixed in 4% paraformaldehyde, and hematoxylin-eosin staining was performed after paraffin sectioning (5 μm), and the thickening of the epidermis of the skin was observed.

In the evaluation of the psoriasis skin lesion area and severity index, the higher the score is, the more severe the psoriasis symptoms are, as can be seen from fig. 5A, the score of each group reaches about 9 points after continuous modeling for 5 days, which indicates that the psoriasis model is successfully modeled, and the severity of each group is similar. After microneedle treatment on day six, the score of each microneedle treatment group begins to decrease, and the score of the double-layer microneedle group and the score of the middle-layer tacrolimus microneedle group are lowest on day 11 and are statistically different from those of the model group and the mixed single-layer microneedle group (the score of each microneedle treatment group is not lower than that of the model group and the mixed single-layer microneedle group: (the score of each microneedle treatment group is lower than that of the middle-layer microneedle treatment group and the second microneedle treatment group is lower than that of the model group and the mixed single-layer microneedle treatment group)P<0.05), which shows that the double-layer microneedle has obvious effect on treating psoriasis and has no difference with the tacrolimus microneedle group in the middle layer. In order to further evaluate the recovery of the skin barrier function, the evaluation index introduces the moisture loss through the skin, and the lower the moisture loss is, the better the skin barrier function is, otherwise, the skin barrier function is damaged. As can be seen from FIG. 5B, similar to the scoring index, the water loss through the skin of each group peaked after 5 days of continuous molding, indicating that psoriasis caused severe barrier function breakdown. After the microneedle treatment was performed on day 6, the transdermal water loss value began to decrease, and on day 11, the transdermal water loss values of the double-layer microneedle set and the middle-layer tacrolimus microneedle set were the same as those of the blank control group, which indicates that the skin barrier function was completely recovered after the microneedle treatment of example 1, and the skin barrier function was statistically different from that of the modeling group and that of the mixed single-layer microneedle set (the difference is shown in the following table: (b) ((the statistical difference is included in the present table)P<0.05）。

As shown in FIG. 6A, the skin of the rats in the normal blank control group has a thin epidermis, a compact structure and a normal cell morphology. In the psoriasis model, the hyperkeratosis phenomenon is caused by the hyperproliferation of the keratinocytes, the epidermis is obviously thickened, and the severe inflammatory cell infiltration is caused. To further examine the thickening of the epidermis, the thickness of the epidermis was measured as shown in fig. 6B, and the thickness of the epidermis was smaller than that of the model group in each treatment group after each microneedle treatment, and there was a statistical difference (P<0.05), the skin epidermis thicknesses of the double-layer microneedle group and the middle-layer tacrolimus microneedle group are recovered to the level of a normal blank control group, and the double-layer microneedle group and the middle-layer tacrolimus microneedle group have statistical difference, so that the treatment effect superior to that of other microneedles is shown, and the double-layer microneedle group can effectively relieve psoriasis symptoms and has no difference with the middle-layer tacrolimus microneedle group.

The results of rat model treatment experiments on arthritis and psoriasis are combined, and the double-layer micro-particles have better treatment effects on the arthritis and the psoriasis. When a psoriasis patient has arthritis complications, the double-layer microneedle is fed at the same focus position, the double-layer microneedle co-administration system can simultaneously transmit diclofenac sodium and tacrolimus, the diclofenac sodium at the needle tip can diffuse into a joint cavity through the skin, the tacrolimus at the middle layer can enter the skin along a pore generated by the needle tip, the two medicines respectively exert the curative effects, and the purpose of simultaneously relieving skin psoriasis symptoms and arthritis is achieved, so that the psoriasis arthritis is treated.

The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.

Claims

1. A soluble microneedle for treating psoriatic arthritis, which comprises a needle tip layer, an intermediate layer and a base layer divided in an insertion direction, wherein

The needle tip layer comprises non-steroidal anti-inflammatory drugs dispersed in a needle tip layer matrix,

said intermediate layer comprising a macrolide immunomodulator dispersed in an intermediate layer matrix,

the base layer comprises a base matrix

The total length of the needlepoint layer and the middle layer in the insertion direction is 100-350 mu m, wherein the non-steroidal anti-inflammatory drug of the needlepoint layer diffuses to the joint cavity through the skin, and the macrolide immunomodulator of the middle layer enters the skin along pores generated by needlepoints, so that the psoriasis symptoms and the joint inflammation of the skin are relieved, and the treatment of the psoriatic arthritis is realized.

2. The soluble microneedle of claim 1, wherein the non-steroidal anti-inflammatory drug is diclofenac sodium.

3. The soluble microneedle of claim 1, wherein the macrolide immunomodulator is tacrolimus.

4. The dissolvable microneedle according to claim 1, wherein said non-steroidal anti-inflammatory drug is diclofenac sodium and said macrolide immunomodulator is tacrolimus.

5. The soluble microneedle of claim 1, wherein the tip layer matrix and the base layer matrix each comprise: sodium hyaluronate, dextran 40, povidone K17 and water.

6. The dissolvable microneedle according to claim 1, wherein said intermediate layer substrate comprises: sodium hyaluronate, dextran 40, povidone K17, niacinamide and water.

7. A microneedle array comprising a plurality of microneedles of any one of claims 1-6, wherein the base layer of each microneedle is formed in the same plane and is integral.