Polypeptide lx with activity of promoting repair of chronic non-healing skin wounds and application thereof

Abstract

The application discloses a polypeptide LX with activity of promoting repair of chronic non-healing skin wounds and an application thereof. An amino acid sequence of the polypeptide LX is RAVFCEIFKRC, and the application of the polypeptide LX is an application in preparation of a medicine for promoting repair of skin wounds. The application provides a new polypeptide LX, and the polypeptide LX has excellent activity of promoting skin tissue regeneration, has good repairing effects on deep second-degree scald and chronic skin wounds of diabetes, and after 16 days of treatment of chronic skin wounds of a type 2 diabetes mouse by using the polypeptide LX, a skin wound repair rate reaches 98.4%, and is significantly higher than that of rh-bFGF (89.8%). The polypeptide LX has only 11 amino acids, is easy to synthesize and low in cost, provides a new way for development of medicines for repairing chronic skin wounds, and also provides a new direction for development of medicines for treating skin wounds.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to a polypeptide LX with activity in promoting the repair of deep second-degree burns and chronic skin wounds in diabetic patients, and its applications. Background Technology

[0002] Despite significant efforts in treating chronic skin wounds, their management remains a major clinical challenge, with their increasing incidence imposing a substantial psychological and economic burden on patients and healthcare systems worldwide. It is reported that up to 6 million people in the United States alone are affected by chronic, non-healing wounds. The skin, the largest organ in the human body, accounting for approximately 15% of an adult's total body weight, is a crucial barrier responsible for vital physiological functions such as resisting pathogen invasion, preventing moisture loss, and protecting against UV damage. When external environmental factors (including heat, mechanical, physical, chemical, and biological damage) disrupt the integrity of the skin's structure, its primary function as a defense mechanism is impaired. The skin regeneration process of a normal wound primarily involves four overlapping but distinct phases: hemostasis, inflammation, cell proliferation, and matrix remodeling. Wounds that take more than 12 weeks to heal are defined as chronic, non-healing wounds, often associated with factors such as poor blood circulation, edema, infection, obesity, and autoimmune diseases like diabetes. For example, in the healing process of chronic, non-healing skin wounds in diabetic patients, damaged blood vessels in the skin tissue hinder oxygen supply, leading to hypoxia and impaired angiogenesis (endothelial cells), re-epithelialization (keratinocytes and fibroblasts), and extracellular matrix formation. Furthermore, the continuous infiltration of inflammatory cells (such as neutrophils and macrophages) into the wound bed produces excessive inflammatory mediators and reactive oxygen species (ROS), damaging wound tissue. Excessive recruitment of high-oxygen-consuming inflammatory cells exacerbates hypoxia, creating a vicious cycle that delays wound healing. Due to the complex pathophysiological mechanisms involved in diabetic skin ulcers, routine treatments for diabetic foot ulcers approved by the US Food and Drug Administration (FDA) are insufficient to meet the needs of wound healing. Therefore, seeking an effective strategy to promote the healing of chronic, non-healing diabetic wounds is crucial for restoring normal physiological function, restoring homeostasis, and alleviating the psychological and economic burden on patients and society.

[0003] In recent years, stem cell therapy, tissue engineering technology, and hyperbaric oxygen therapy have provided novel treatment strategies for the repair of chronic, difficult-to-heal skin wounds. However, these approaches suffer from drawbacks such as high cost and long treatment cycles, hindering their widespread adoption. Drug therapy continues to play a crucial role in skin wound treatment. Common clinical wound repair drugs include small molecule compounds, growth factors (rh-bFGF, EGF), animal and plant-derived drugs, and low-adhesion dressings. While these drugs exhibit promising effects, they also suffer from numerous limitations, such as difficulty in synthesis, high synthesis costs, demanding storage conditions, instability of active ingredients, and a tendency to induce scarring, making it difficult to meet clinical needs for skin wound treatment. Therefore, the discovery of novel and highly effective lead molecules for promoting skin tissue regeneration and the exploration of their molecular mechanisms for promoting wound repair are particularly important and necessary.

[0004] Compared to small molecule compound drugs, peptide drugs exhibit stronger activity, lower toxicity, higher safety, higher development success rate, and stronger targeting. Furthermore, the development of peptide drugs has a broad commercial market and has gradually become a "potential stock" that cannot be ignored in the field of drug development.

[0005] The present invention aims to provide a polypeptide that exhibits good skin wound repair activity in both deep second-degree burns and diabetic chronic skin trauma. Summary of the Invention

[0006] The first objective of this invention is to provide a polypeptide LX with skin wound repair activity, and the second objective of this invention is to provide the application of said polypeptide LX.

[0007] The first objective of this invention is achieved by the polypeptide LX having the amino acid sequence shown in SEQ ID No. 1: RAVFCEIFKRC, and its structural formula is as follows: Figure 1 As shown.

[0008] The second objective of this invention is achieved by using the polypeptide LX in the preparation of a drug that promotes skin wound repair. The skin wound is a deep second-degree burn or a chronic skin wound caused by diabetes.

[0009] The beneficial effects of this invention are as follows: This invention provides a novel polypeptide LX, which exhibits excellent skin tissue regeneration activity and good repair effects on deep second-degree burns and chronic skin wounds caused by diabetes. After treating type II diabetic mice with chronic skin wounds for 16 days, the skin wound repair rate reached 98.4%, significantly higher than rh-bFGF (89.8%). This invention's polypeptide LX contains only 11 amino acids, is easy to synthesize, and has low cost, providing a new avenue for the development of drugs for the repair of chronic skin wounds and a new direction for the development of drugs for the treatment of skin wounds. Attached Figure Description

[0010] Figure 1 This is a primary structure diagram of the polypeptide LX of the present invention;

[0011] Figure 2 This is a diagram showing the activity of the peptide LX of this invention in promoting the repair of deep second-degree burns in mice; where A represents the repair of deep second-degree burn wounds in mice.

[0012] Complex variation diagram, B is a quantitative diagram of the repair of deep second-degree burn wounds in mice;

[0013] Figure 3 This diagram illustrates the regenerative activity of the polypeptide LX of this invention in promoting deep second-degree burn skin tissue; where A represents a deep second-degree burn in a mouse.

[0014] HE staining images of skin tissue in the affected area on day 14. B is a quantitative map of the thickness of regenerated epidermis in the skin tissue of the wound area of ​​mice on day 14 after burns. C is a quantitative map of the thickness of regenerated granulation tissue in the skin tissue of the wound area of ​​mice on day 14 after burns.

[0015] Figure 4 This is a diagram showing the skin wound repair activity of the polypeptide LX of this invention on type II diabetic mice; where A represents the skin wound repair activity of type II diabetic mice.

[0016] Image B shows the changes in the repair of the entire dermal wound in mice, where B is a quantitative image of the repair of the entire dermal wound in type II diabetic mice. Detailed Implementation

[0017] The present invention will now be described in further detail with reference to the accompanying drawings and embodiments, but this does not limit the present invention in any way. Any modifications or improvements made based on the teachings of the present invention shall fall within the protection scope of the present invention.

[0018] This invention discloses a polypeptide LX with skin wound repair activity, the amino acid sequence of which is: RAVFCEIFKRC.

[0019] The present invention also provides the application of the polypeptide LX in the preparation of drugs that promote skin wound repair.

[0020] The skin trauma is a deep second-degree burn or a chronic skin trauma caused by diabetes.

[0021] The present invention further provides a pharmaceutical composition comprising the polypeptide LX as an active agent and its pharmaceutically effective carrier.

[0022] Example 1: Synthesis of polypeptide LX

[0023] The amino acid sequence of the skin-promoting chronic wound repair active polypeptide LX is RAVFCEIFKRC. The skin-promoting wound repair active polypeptide LX is prepared by conventional chemical synthesis or gene expression.

[0024] The active peptide LX for promoting skin wound repair in this invention was synthesized by Wuhan Baiyixin Biotechnology Co., Ltd.

[0025] Example 2: Detection of the skin wound repair activity of peptide LX

[0026] I. Detection of the tissue repair activity of peptide LX in deep second-degree burns

[0027] Experimental method: Deep second-degree burn experiment in mice

[0028] 1. Male Kunming mice (22-30 g) were anesthetized with 1% sodium pentobarbital (0.1 mL / 20 g). After removing the fur from their backs, two 10 mm diameter circular copper clips were applied to the mouse's back skin for 20 seconds to create two deep second-degree burn wounds. After modeling, the mice were randomly divided into three groups of six mice each: a blank control group (vehicle, PBS), a positive control group (positive, rh-bFGF, 100 ng / mL), and a peptide group (LX, 1 nM). Different drugs were applied to the wound area twice daily at a dose of 20 µL. To observe changes in wound healing, the wound was photographed and recorded on days 0, 4, 8, and 14 post-surgery to assess the therapeutic effect of peptide LX. Furthermore, on day 14 post-surgery, tissue from the mouse back wounds was collected for histopathological analysis to reveal the regeneration of skin wound repair tissue. ImageJ software was used to quantify the wound area and the thickness of epidermal and granulation tissue regeneration at different treatment days. The wound healing rate of mouse backs was calculated using the following formula and statistical analysis was performed using GraphPad Prism8 software. Three independent replicate experiments were conducted on mice with deep second-degree burns, and six mice were included as parallel controls in each experiment for different drug treatment groups. Wound healing rate (%) = (Initial wound area (0 days) - Wound area at different treatment days (4, 8, 14 days)) / Initial wound area (0 days) × 100%

[0029] 2. To evaluate the regeneration of mouse wound tissue after different treatments, H&E staining analysis was performed. Skin tissue was cut into 5 μm thick sections, dewaxed, graded hydration, and then subjected to H&E staining. Wound tissue regeneration was recorded using an optical microscope at the same magnification. Changes in the epidermis, new tissue formation, and epithelialization of different pathological tissues were calculated using ImageJ software, and the data were statistically analyzed using Prism software.

[0030] Result: From Figure 2 It can be seen that the polypeptide LX of this invention can significantly promote the repair of full-thickness skin wounds on the back of mice. After 14 days of treatment, the wound repair activity of LX is similar to that of the positive control rh-bFGF, and the wound repair rate is close to 100%. Figure 3 HE staining results showed that the peptide LX of the present invention significantly promoted the regeneration of wound epidermis in mice and significantly accelerated re-epithelialization. In addition, it also promoted the regeneration of granulation tissue in the wound area.

[0031] II. Detection of the skin wound repair activity of peptide LX in type II diabetic mice

[0032] Experimental Methods: Full-Cortical Injury Experiment in Type II Diabetic Mice

[0033] After feeding mice a high-fat diet for one week, a type 2 diabetes model was established in C57BL / 6 mice by intraperitoneal injection of streptozotocin (STZ) at a concentration of 30 mg / mL, at a dose of 30 mg / kg of mouse body weight. C57BL / 6 mice were anesthetized with 1% sodium pentobarbital (0.1 mL / 20 g), and the fur on their backs was removed. Two full-thickness wounds with a diameter of 10 mm were created on both sides of the mouse's back using a biopsy puncture device. Mice were randomly divided into four groups: a normal control group (PBS), a diabetic control group (PBS), a positive control group (rh-bFGF, 100 ng / ml), and a peptide group (LX, 1 nM). The medication was administered twice daily at 20 µL each time, and wound changes were photographed and recorded on days 0, 4, 8, and 16 post-surgery.

[0034] Result: From Figure 4 It was found that the peptide LX of this invention can significantly promote the repair of full-thickness skin wounds on the back of mice. After 8 days of treatment, the wound repair activity of LX reached 58.5%, which was significantly higher than that of the blank control (47.4%) and rh-bFGF (41.7%). Notably, after 16 days of LX treatment, the skin wound repair rate reached 98.4%, which was significantly higher than that of rh-bFGF (89.8%), demonstrating good therapeutic activity and research and development potential.

[0035] In summary, the wound repair peptide LX provided by this invention has strong activity in promoting the repair of deep second-degree burns and chronic skin wounds in type II diabetes, and has certain clinical application value.

Claims

1. A polypeptide LX with activity promoting the repair of chronic, difficult-to-heal skin wounds, characterized in that, The amino acid sequence of the polypeptide LX is RAVFCEIFKRC, and its structural formula is as follows: 。 2. The use of the polypeptide LX of claim 1 in the preparation of a drug that promotes skin wound repair.

3. The application according to claim 2, characterized in that, The skin trauma is a deep second-degree burn or a chronic skin trauma caused by diabetes.

4. A pharmaceutical composition comprising the polypeptide LX of claim 1 as an active agent and its pharmaceutically effective carrier.

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