CN115715798A - Cytokine for diabetic wound repair and application thereof - Google Patents

Abstract

A cytokine for promoting the migration of fibroblast and its application in preparing the medicines and medical devices for promoting the repair of diabetic wound are disclosed, which features that the cytokine is used as its active component.

Description

Cytokine for diabetic wound repair and application thereof

Technical Field

The invention relates to an application of protein, in particular to a cytokine which has the effect of wound surface repair as an active ingredient and an application thereof in pharmacy and medical equipment containing medicine.

Background

The skin is an important barrier structure of the human body, which protects internal tissue organs from external mechanical damage, and plays a key role in resisting external adverse stimuli such as microbial infection, ultraviolet radiation and extreme temperature. However, because of the specific location of the skin, it is more vulnerable, and there are over ten million new cases of skin defects worldwide each year, however, the existing treatments still have limited effects on skin repair, especially on the complete structure and functional recovery of the refractory wound surface. The wound surface which cannot be effectively repaired seriously affects the physiological function and the life quality of the patient, and the social medical burden is increased. How to effectively promote wound healing and skin regeneration is always a frontier focus problem which needs to be confronted together globally.

The skin, which is the largest organ in the human body area, is innervated and regulated as well as other organs. The skin nervous system can activate various exogenous or endogenous stimuli, and generate different regulation and control effects according to specific stimuli, thereby realizing basic physiological functions such as sensation and thermoregulation. The peripheral nervous system also nourishes and regulates the self-renewal of skin structures by secreting neuropeptides in the local microenvironment, participating in the homeostatic regulation of the normal skin microenvironment.

Clinical observations and research results show that the nervous system is also involved in regulating and controlling various tissue regeneration processes including the skin, and traumatic or pathological loss of nervous system function is often accompanied by various tissue regeneration difficulties. Patients with impaired sensory and autonomic innervation, such as: patients with spina bifida and spinal cord injuries, often with wounds below the level of spinal cord injury, present difficulty healing. Pressure sores in diabetic patients are believed to be caused by long-term compression and impaired blood circulation following neuropathy.

Diabetic wounds are one of the common diabetic complications, with chronic and difficult-to-heal characteristics. Approximately 25% of diabetic patients suffer from Diabetic Foot Ulcers (DFUs), which are also one of the most common types of difficult to heal wounds. CN106540241A discloses a method for treating diabetic foot by using fibroblast growth factor,spraying recombinant bovine basic fibroblast growth factor (ALFGF) onto wound surface at a dose of 150AU/cm ² The dressing change is given 1 time according to the condition of the wound surface for 1-3 days, and 2 groups continuously observe for 4 weeks and then evaluate the curative effect. CN111450312A discloses a hydrogel dressing for effectively promoting wound repair and regeneration, which is a heparin-bFGF hydrogel dressing prepared from heparin and bFGF, so that the heparin and the bFGF can be stably and slowly released within a period of more than 10 days to promote wound healing of chronic wound ulcer.

Dermal and/or epidermal 3D scaffold analogs have been used clinically for 30 years in wound repair, including some patients with diabetic wounds. However, due to the lack of ordered cellular organization, these inorganic materials still have a great difference from real skin, and cannot completely replace normal skin to complete daily physiological functions. Whether allograft or patient autologous tissue transplantation, the donor material deficiency still faces. Therefore, improving the self-repair ability of cells in the in-situ area of the wound surface is still one of the treatment strategies. Although some growth factors can promote cell proliferation, such as: PDGF, which has been approved by the FDA for the treatment of diabetic wounds, still has limited ultimate efficacy in clinical applications. This suggests that strategies that simply increase cell proliferation are not sufficient to solve the complex diabetic wound treatment problem.

Thus, current diabetic wound therapy still lacks a rapid and effective treatment regimen, and patients often need to undergo multiple surgical interventions. Because of limited treatment effect, the diabetic refractory wound surface is also a main reason of non-traumatic amputation, and great burden is caused to patients and society.

Transforming growth factor beta (TGF- β) is one of the key signaling pathways for tissue regeneration, and its effects can be complicated and diverse due to the specific environment and the molecules involved. The cellular origin of the ligand molecules is also an important factor affecting function, dermal-derived TGF- β has the effect of inhibiting proliferation of adjacent epithelial cells, whereas keratinocyte-expressed TGF- β inhibits fibroblast-secreted TGF- β. Although the TGF-. Beta.pathway has long been recognized as a valuable target for therapeutic action, the therapeutic approaches developed for it have remained short of expectations due to the lack of mechanistic studies.

Disclosure of Invention

The invention aims to provide a cytokine which is used as an active ingredient for promoting the repair of diabetic wounds.

The invention also aims to provide a cytokine which is used as an active ingredient for promoting the migration of fibroblasts and facilitating the repair of diabetic wounds.

The invention also aims to provide an application of the cell factor, namely the deep part of the dermis layer, in the preparation of a medicament for treating diabetic wounds in the early stage of wound repair.

It is still another object of the present invention to provide a use of a cytokine for the preparation of a drug-containing medical device.

A cytokine, transforming growth factor beta 3 (TGF-beta 3), which is ubiquitous in mammals, such as: but are not limited to: human, monkey and mouse, etc., with high homology in molecular sequence, such as: SEQ ID No 1 and SEQ ID No 2.

In the present invention, transforming growth factor beta 3 includes proteins or polypeptides having homology with the natural sequence. These proteins or polypeptides can be artificially edited and produced by genetic engineering means, and these functional domains that bind to downstream signaling molecules can be covalently bound using small molecules or polymers that are part of Bioconjugate Chemistry (Bioconjugate Chemistry) and have produced related applications such as: ADC drugs, acylation or pegylation, and the like.

In a specific embodiment, the cytokine is an artificial transforming growth factor β 3 with a homology of greater than 98%.

In another specific embodiment, the cytokine is an artificial transforming growth factor β 3 with a homology of greater than 95%.

In another specific embodiment, the cytokine is an artificial transforming growth factor β 3 with a homology greater than 93%.

In another specific embodiment, the cytokine is an artificial transforming growth factor β 3 with a homology greater than 90%.

In another specific embodiment, the cytokine is an artificial transforming growth factor β 3 with a homology greater than 85%.

In another specific embodiment, the cytokine is an artificial transforming growth factor β 3 with a homology greater than 80%.

In another specific embodiment, the cytokine is an artificial transforming growth factor β 3 with a homology of greater than 75%.

In another specific embodiment, the cytokine is an artificial transforming growth factor β 3 with a homology of greater than 70%.

In the invention, the relevance of TGF-beta 3 and the diabetic wound is found and established by single cell transcription analysis and a mouse disease model, and the migration of fibroblasts can be promoted. The application of TGF-beta 3 to the diabetic skin (dermis) can obviously promote the repair of diabetic wounds and has high specificity.

The cytokine of the present invention is used as a drug when it acts on an in vitro cell or an in vivo cell to promote migration of fibroblasts.

When the cytokine of the present invention acts on in vitro cells or in vivo cells to achieve the purpose of repairing diabetic wounds, it is understood that the cytokine of the present invention is used as a drug.

TGF-beta 3 shown in the invention is mixed with other auxiliary materials to prepare the medicine (preparation) for repairing the diabetic wound.

The pharmaceutical excipients can be those conventionally used in various preparations, such as: but are not limited to, isotonic agents, buffers, flavoring agents, excipients, fillers, binders, disintegrating agents, lubricants, and the like; it may also be selected for use in accordance with the substance, such as: the auxiliary materials can effectively improve the stability and solubility of the compounds contained in the composition or change the release rate, absorption rate and the like of the compounds, thereby improving the metabolism of various compounds in organisms and further enhancing the administration effect of the composition.

In aqueous injection, the adjuvant generally comprises isotonic agent, buffer, necessary emulsifier (such as Tween-80, pluronic and Poloxamer), solubilizer, bacteriostatic agent, etc. In addition, the pharmaceutical composition also comprises other pharmaceutically acceptable pharmaceutical excipients, such as: antioxidants, pH modifiers, analgesics, and the like.

The adjuvants used for preparing oral liquid preparation generally include solvent, and necessary correctant, bacteriostat, emulsifier and colorant, etc.

The excipients used for the preparation of tablets generally include fillers (e.g., starch, powdered sugar, dextrin, lactose, compressible starch, microcrystalline cellulose, calcium sulfate, calcium hydrogen phosphate, mannitol, etc.), binders (e.g., ethanol, starch slurry, sodium carboxymethylcellulose, hydroxypropylcellulose, methylcellulose, ethylcellulose, hydroxypropylmethylcellulose, gelatin solution, sucrose solution, and an aqueous or alcoholic solution of polyvinylpyrrolidone, etc.), disintegrants (e.g., dry starch, sodium carboxymethyl starch, low-substituted hydroxypropylcellulose, crosslinked polyvinylpyrrolidone, and crosslinked sodium carboxymethylcellulose), and lubricants (e.g., magnesium stearate, colloidal silica, talc, hydrogenated vegetable oil, polyethylene glycol 4,000, polyethylene glycol 6,000, magnesium lauryl sulfate, etc.), and the like.

The adjuvants used for preparing emulsion are generally water, oil (such as fatty acid), emulsifier, and necessary antiseptic and correctant.

The excipients used to make granules are similar to tablets, but differ in the granulation process. Mixing the obtained granule with glidant, and encapsulating to obtain capsule.

Various adjuvants and compounds can be made into dosage forms beneficial for drug delivery (such as: but not limited to aqueous solution injection, powder injection, pill, powder, tablet, patch, suppository, emulsion, cream, gel, granule, capsule, aerosol, spray, powder spray, sustained release agent, controlled release agent, etc. In addition, specific administration purposes or modes may be achieved, such as: sustained release administration, controlled release administration, pulse administration, and the like, and used auxiliary materials such as: but are not limited to gelatin, albumin, chitosan, polyether and polyester-based polymer materials, such as: but are not limited to, polyethylene glycol, polyurethane, polycarbonate, copolymers thereof, and the like. The main indications of so-called "facilitated administration" are: but not only improving the treatment effect, improving the bioavailability, reducing the toxic and side effects, improving the patient compliance and the like.

TGF-. Beta.3 of the present invention is combined with other excipients, such as: chemical coupling to further improve the drug effect of the compound, reduce the toxic effect, prolong the administration period and the like. These adjuvants are generally polymers such as: polyesters, polyethers, and polyamides.

Medicated medical devices made by combining a drug with a medical device have also become common, such as: a dressing comprising TGF-beta 3. The TGF-beta 3 of the invention is also loaded or coated on a bracket material as an active component and is used for preparing medical appliances for repairing diabetic wounds. Common scaffold materials are such as: PLA, PLGA, and metals, among others. And mixing with a biocompatible degradable material to prepare the micro-needle and the micro-needle array thereof, or loading the micro-needle array in the metal micro-needle to prepare the micro-needle chip. When the micro-needle penetrates into the skin, TGF-beta 3 is released in the dermis layer, and the capability of healing the diabetic wound is improved.

The TGF-beta 3 is verified to be in the wound microenvironment and present the distribution characteristics of tissues, is mainly distributed in the deep layer of the dermis, has direct correlation with the diabetic wound repair and has exact diabetic wound repair function. Further inhibition experiments show that exogenous TGF-beta 3 should be given at the early stage of diabetic wound repair, so that the early repair of the diabetic wound is facilitated. The application of TGF-beta 3 in diabetic wound repair provided by the invention fills the blank that the expected effect of diabetic wound repair treatment is difficult to achieve.

Drawings

FIG. 1 is a graph showing the trend of analyzing the change in the Expression level of transforming growth factor 3 (TGF-. Beta.3) in skin tissue cells using biological information, in which each point represents a cell, the ordinate represents the Relative Expression level (Relative Expression) of Tgfb3 transcription level, the abscissa represents Pseudo-time (Pseudo-time) of the cell, the origin of the abscissa is the seventh day of wound surface, and cells in different states (State) are distinguished by different colors;

FIG. 2 is a graph showing the immunofluorescent staining results of skin tissue sections at the fifth day after the mouse skin wound was generated; wherein the fluorescent signal is indicative of a transforming growth factor 3 (TGF-beta 3) protein profile. Here, DAPI staining was used to indicate the nucleus, and the primary antibody to TGF-. Beta.3 was detected as a rabbit polyclonal antibody.

FIG. 3 is a graph showing the results of immunofluorescence staining of skin tissue sections of foot ulcers in diabetic patients; wherein, the fluorescent signal indicates the protein distribution condition of transforming growth factor (TGF-beta 3), DAPI staining is used for indicating cell nucleus, and the primary antibody for detecting TGF-beta 3 is rabbit source polyclonal antibody.

FIG. 4 is a graph showing the effect of TGF-. Beta.3 addition on skin structural cells using cultured fibroblast NIH-3T 3; wherein, the Ctrl cultured cells in the control group do not have any special treatment, and the TGF-beta 3 is added into the culture medium in the cells in the experimental group (TGF beta); FIG. 4A is a graph of cells observed by a scratch test after 18 hours (18 hr) of treatment; FIG. 4C is a graph showing the ratio of the area not covered by cells to the original area (% Closure/Star _ Wound) after 18hr of cell migration in different treatment groups; FIG. 4B is a graph showing the evaluation of cell proliferation 24 hours after the treatment of cells, in which the cells cultured in the control group (Ctrl) were not subjected to any special treatment, the cells in the experimental group (TGF. Beta.) were subjected to addition of TGF-. Beta.3 to the medium, and FIG. 4D is a statistical graph showing the ratio (% EdU + cell) of the number of the proliferated cells (EdU-positive cells) to the total number of the cells in the treated cells;

FIG. 5A is a flow chart of an experiment using a small molecule inhibitor to inhibit TGF-beta 3 function at different time points in a mouse model;

FIG. 5B is a photograph showing wound healing observed in mice of the blank control group (Ctrl), the early intervention group (-TGF-. Beta.e) and the late intervention group (TGF-. Beta.l) after 14 days (D14).

FIG. 5C is a statistical chart of wound healing observed in mice in the blank control group (Ctrl), early intervention group (-TGF-. Beta.e), and late intervention group (TGF-. Beta.l) after 14 days (D14); wherein the ordinate values represent the percentage of the actual observed Wound relative to the initial Wound (D0 Wound area%);

FIG. 5D 14 days (D14) after section observation the staining pattern of mouse skin wound tissue for blank control group (Ctrl), early intervention group (-TGF-. Beta.e) and late intervention group (TGF-. Beta.l), observed with HE staining;

FIG. 5E section after day 14 (D14) is observed to obtain statistical plots of skin wounds of mice in the blank control group (Ctrl), early intervention group (-TGF-. Beta.e) and late intervention group (TGF-. Beta.l), and the epidermal layer thickness (Epithelium thickness) of the wound area measured in micrometers (μm) at day 14 in different groups of mice.

Fig. 6A is a statistical view of staining of tissue sections after the seventh day of topical injection of exogenous TGF- β 3 to diabetic skin wounds in diabetic mice, in which the ordinate values represent Wound width (Wound width) observed on the tissue sections, the unit of measurement in micrometers (μm), i.e., the tissue structure of the wounds of Diabetic Mice (DM) injected with physiological saline only, and the treatment group of deep skin injection of transforming growth factor solution (DM + T) around the wounds. The graph B is a statistical chart of skin healing conditions (DM + T) observed in a control group (DM) and a treatment group (DM + T) on day 14 of diabetic skin wounds of diabetic mice locally injected with exogenous transforming growth factor, and the numerical value of the ordinate in the chart indicates the percentage (D0 Wounded area%) of the actually observed wounds relative to the initial wounds.

Detailed Description

The technical scheme of the invention is described in detail in the following with reference to the accompanying drawings. Although the present invention has been described in detail with reference to the preferred embodiments, it will be understood by those skilled in the art that various changes may be made and equivalents may be substituted without departing from the true spirit and scope of the present invention, which is defined in the claims.

Other reagents used in the examples of the present invention were purchased from Bilun sky, unless otherwise specified.

The mice used in this example were 6-8 week old wild type C57BL/6J mice, and were bred under standard conditions.

The bioinformatic analysis in this example was derived from a single cell transcriptional expression profile integrated from a common database. The single cells are all derived from cells separated from the wound skin of the mouse. Sequencing the single cells by using monocle software according to the sampling time point and the state of the cells (abscissa, the leftmost end of the coordinate axis is the fourth day of wound surface), and displaying the relative expression quantity (ordinate) of the transcription growth factor of each cell

The mouse wound models used in this example all used peripherally-fixed full-thickness skin defect models. The specific implementation method comprises the following steps: after mice were anesthetized with isoflurane and shaved clean, the used scissors sterilization produced a full-thickness excision wound of 6mm in diameter on dorsal skin. Tightly sewing a silica gel ring on the outward side of the silica gel ring on the skin around the wound, sticking a layer of hydrophobic film, and finally wrapping the wound surface of the mouse by using a medical bandage.

Inhibition experiments for TGF-. Beta.3. A local injection of 150. Mu.l of SB431542 (final concentration 5mg/ml, from seleck) saline solution was injected at the wound site according to the grouping. The injection needle penetrates into the dermal layer of the wound surface, and the early-stage inhibition group is injected on the day of the wound surface, while the late-stage injection group is injected on the seventh day of the wound surface.

For the TGF-beta 3 drug treatment experiments, 200 microliters of TGF-beta 3 (1 g/ml, purchased from Boster) or placebo saline was injected immediately after the resection wound model was generated.

The process of taking the mouse skin tissue strictly follows the ethics of experimental animals. After euthanizing the mice, the skin (about 1.5 cm in diameter) of the wound and surrounding area was harvested with a scalpel for subsequent analysis.

The fibroblast lines used in the experiments were NIH3T3 cells, 5% CO at 37 ℃% ₂ The culture is carried out in a constant temperature and humidity sterile incubator. The medium used for the culture was high-glucose DMEM medium supplemented with 10% fetal bovine serum (Gibco) and 1% antibiotic-antifungal drug (Gibco). To avoid mycoplasma contamination, a portion of the cells were removed for cytoplasmic DAPI staining at each passage of the cells. The culture medium is replaced every 2 days in the cell culture process until the cell healing degree reaches 85-90 percent, and cell passage is carried out.

In cell migration experiments, NIH3T3 cells were seeded in 6-well culture plates in complete medium to form almost 100% confluent cell monolayers. 16 hours before the experiment, the original cell culture was replaced with high-glucose DMEM without fetal bovine serum. For the formation of a scratch, a single layer of NIH3T3 cells was scraped with a 200 microliter pipette tip to form a straight scratch. Cell debris and fluid were gently aspirated and medium containing transforming growth factor (0.1 g/ml) was added. Photographic recordings were made at hours 0 and 18 of the scratch test. The migration rate calculation method comprises the following steps: % scratch closure = ((0 hr scratch area-18 hr scratch area) × 100)/0 hr scratch area.

For the measurement of cell proliferation, NIH3T3 cells were seeded in 12-well plates at a cell culture density of about 30% of the bottom of the confluent plate, and a transforming growth factor was added to the aforementioned conventional medium to a concentration of 0.1g/ml. Then, 10. Mu.M of 5-ethyl-2' -deoxyuridine (EdU) was added to the medium 2 hours before observation. The EdU staining kit used here is BeyoClick ^TM The EdU kit and instructions for handling the cells. And taking a picture to record a dyeing result.

For morphological observation of tissues, skin tissues of human or mouse were fixed overnight with 4% paraformaldehyde solution, followed by dehydration with 20% sucrose. The Tissue was then embedded in Tissue-Tek o.c.t. (Sakura Finetek, 4583) and the embedding medium was frozen in a negative twenty degree environment. The tissue was then cryosectioned at a slice thickness of 10-15 microns.

For pathological section observation of skin tissues, hematoxylin and eosin staining kits were used. The staining process of the skin sections was performed according to the standard procedure of the kit. The stained tissue sections were mounted with resin.

For fluorescent staining observation of skin tissue, skin sections were incubated in antigen retrieval solution (Beyotime, P0088) in a water bath at 95 ℃ for 25 minutes followed by gentle washing of the samples with PBS. Sections were blocked for 1 hour in staining buffer (2% BSA +0.05% Triton in PBS), diluted in staining buffer for transforming growth factor primary antibody (rabbit derived antibody, abcam, ab 15537) and incubated overnight at 4 ℃. After 3-5 PBS washes, the cells were incubated in staining buffer containing secondary antibody for 1 hour at room temperature.

For immunofluorescence of cultured cells, the cell attachment sheet was fixed in 4% paraformaldehyde solution at room temperature for 15 minutes. The cell samples were then subjected to cell permeabilization using 0.25% Triton x-100 in PBS for approximately 10 minutes. The cellular immunofluorescence staining process is consistent with the tissue staining procedure.

Example 1

The trend of the change in the expression level of transforming growth factor 3 (TGF-. Beta.3) in skin tissue cells was analyzed using the bioinformatics, and the results are shown in FIG. 1. The graphic result shows that the expression level of the transforming growth factor 3 is higher on the seventh day of wound repair, and the expression level is gradually reduced in the later stage. The secretion time of transforming growth factor 3 is mainly in the early stage of wound repair.

Example 2

The fifth day after the mouse skin wound was generated, the immunofluorescent staining of the skin tissue sections was performed, and the results are shown in fig. 2.

The staining results showed that the protein of transforming growth factor 3 is distributed mainly in the deep layer of the dermis, which indicates that transforming growth factor is indeed present in the microenvironment of the wound surface and presents the distribution characteristics of the tissue.

Example 3

Immunofluorescent staining of skin tissue sections of foot ulcers in diabetic patients resulted in the results shown in figure 4.

The staining results showed that no protein of transforming growth factor 3 was detected in the deep dermal region of the diabetic wound. This indicates abnormal secretion of transforming growth factor 3 in diabetic wounds.

Example 4

The effect on skin structure cells was observed in TGF-. Beta.3 co-culture in fibroblasts NIH-3T3, and the results are shown in FIG. 4.

As can be seen in FIGS. 4A and 4C, the ratio of the area covered by the cells to the original area was significantly lower after 18 hours of TGF- β 3 administration than the control group, i.e., the cells migrated faster in the TGF- β 3 treated experimental group, indicating that TGF- β 3 promoted the migration of fibroblasts.

As can be seen in FIGS. 4B and 4D, the ratio of proliferating cells (EdU positive cells) to total cell number was significantly higher after 18 hours of TGF- β 3 administration than the control group, i.e., the cells of the TGF- β 3 treated experimental group proliferated faster than the control group, indicating that TGF- β 3 promoted proliferation of fibroblasts.

Example 5

The inhibition of TGF-. Beta.3 function at various time points was observed using small molecule inhibitors, and the relevant experimental procedure is shown in FIG. 5A. Day 0 (D0) is the starting time point of wound formation, a wound with a diameter of 6mm was constructed on the back of the skin of a 6-8 week-old mouse, a blank control group (Ctrl) was not treated at all, and the experimental mice were divided into an early intervention group (-TGF- β e) and a late intervention group (TGF- β l), and D7 and D14 represent days 7 and 14 after wound formation, respectively.

Wound healing after 14 days in mice in the blank control group (Ctrl), early intervention group (-TGF-. Beta.e) and late intervention group (TGF-. Beta.l) is shown in FIGS. 5B and 5C, and staining of sections is shown in FIGS. 5D and 5E. According to the results, after early intervention and inhibition are carried out on TGF-beta 3, the wound healing is obviously lower than that of a control group and a late intervention group, and the early repair effect of TGF-beta 3 on the diabetic wound is more obvious. Thickness observations of epithelial cells also demonstrate that, following early intervention and inhibition of TGF- β 3, repair of epithelial structures is inhibited, with a thickness significantly lower than that of the control group. TGF- β 3 is therefore of direct relevance for diabetic wound repair.

Example 6

The results of 7 days and 14 days after the exogenous TGF-beta 3 is locally injected into the diabetic skin wound of the diabetic mouse are observed. The staining statistics after 7 days of dosing are shown in fig. 6A, and the wound healing statistics are shown in fig. 6B.

When the diabetic mouse is applied with exogenous TGF-beta 3 on the diabetic skin wound surface for the seventh day, compared with a DM control group of normal saline, the area of the wound surface tissue is obviously reduced, which indicates that TGF-beta 3 can promote the repair of the diabetic wound surface. Similarly, compared with a DM control group of normal saline, the percentage of the wound surface relative to the initial wound surface is obviously reduced after the fourteenth day of TGF-beta 3 administration, and the positive correlation that TGF-beta 3 can promote the repair of the diabetic wound surface is also proved, so that the TGF-beta 3 has an exact repair effect on the diabetic wound surface.

Claims (10)

1. An application of transforming growth factor beta 3 as an active ingredient in preparing a medicament for promoting the migration of fibroblasts.

2. An application of transforming growth factor beta 3 as active component in preparing the medicine for treating diabetic wound is disclosed.

3. The use according to claim 2, wherein said transforming growth factor β 3 is derived from a mammal.

4. The use according to claim 2, wherein said transforming growth factor β 3 is derived from a human.

5. The use according to claim 2, wherein said transforming growth factor β 3 further comprises a protein having structural homology to a native protein.

6. The use according to claim 2, wherein said transforming growth factor β 3 further comprises a polypeptide homologous to the transforming growth factor β 3 functional domain that triggers binding of a downstream signaling molecule.

7. An application of transforming growth factor beta 3 as active component in preparing the medical appliance containing medicine for treating diabetic wound is disclosed.

8. The use according to claim 7, wherein the medical device is a medicated dressing.

9. The use according to claim 7, wherein the medical device is a microneedle.

10. The use according to claim 7, wherein the medical device is a stent.

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