WO2018157773A1 - 一种促进创伤后组织修复与再生的修复肽及其应用 - Google Patents

一种促进创伤后组织修复与再生的修复肽及其应用 Download PDF

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WO2018157773A1
WO2018157773A1 PCT/CN2018/077232 CN2018077232W WO2018157773A1 WO 2018157773 A1 WO2018157773 A1 WO 2018157773A1 CN 2018077232 W CN2018077232 W CN 2018077232W WO 2018157773 A1 WO2018157773 A1 WO 2018157773A1
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repair
peptide
regeneration
promoting
post
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PCT/CN2018/077232
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French (fr)
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陈小佳
洪岸
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暨南大学
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Priority claimed from CN201810135019.5A external-priority patent/CN108503690B/zh
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Priority to US16/483,996 priority Critical patent/US11260101B2/en
Publication of WO2018157773A1 publication Critical patent/WO2018157773A1/zh

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/04—Linear peptides containing only normal peptide links
    • C07K7/06—Linear peptides containing only normal peptide links having 5 to 11 amino acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
    • A61K8/64—Proteins; Peptides; Derivatives or degradation products thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A61P17/02—Drugs for dermatological disorders for treating wounds, ulcers, burns, scars, keloids, or the like
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/04—Linear peptides containing only normal peptide links
    • C07K7/08—Linear peptides containing only normal peptide links having 12 to 20 amino acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

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  • the invention belongs to the field of biomedicine, and particularly relates to a repair peptide for promoting post-traumatic tissue repair and regeneration and an application thereof, which can promote tissue repair and cell proliferation in a wound site, in particular, tissue regeneration in skin damage and nerve damage. Repair application.
  • Epidermal cells such as mucous membranes of the respiratory tract, digestive tract and genitourinary tract
  • lymphocytes hematopoietic cells, etc.
  • Other cells such as vascular endothelial cells, periosteal cells, and various glandular organs such as liver, pancreas, endocrine glands, sweat glands, sebaceous glands, and renal tubular epithelial cells belong to the sub-level cells with strong regenerative power.
  • tissue regeneration There are many factors that affect tissue regeneration. In addition to the regenerative capacity of tissues and cells, there are several factors: (1) the extent and extent of tissue damage: the longer the repair and regeneration, the more difficult it is to have a large range of cell necrosis. The replacement of the same number of cells requires healthy cells to produce new cells, and the nascent cells regenerate new cells, which takes a long time. (2) Age factor: Children and adolescents have strong regenerative ability and rapid wound healing; the elderly have weak tissue regeneration ability and slow healing. (3) Nutritional status: Adequate protein, vitamin C, E and minerals Calcium and zinc supply can promote the repair of damage, and vice versa will delay the repair of various injuries.
  • Drug effects Some drugs such as adrenocortical hormone and pituitary adrenocorticotropic hormone can inhibit inflammation, but it is not conducive to the body to eliminate wound infection, but also inhibit granulation tissue growth and collagen synthesis, accelerate collagen decomposition; and anticancer drugs Cytotoxic drugs can also delay healing.
  • Blood supply After tissue damage, local capillary damage, or other factors lead to hardening of the blood vessels, etc., will lead to insufficient blood supply, resulting in tissue malnutrition, hindering healing.
  • Innervation Tissues that lose innervation lose their ability to regenerate.
  • the skin is the largest area of the human body and most directly contacts the outside world, and is also the most vulnerable to external stimuli.
  • the healing of normal wounds is essential for maintaining the body's steady state, and abnormal healing such as unhealed or delayed due to large-scale burns, chronic wound healing caused by other diseases, and abnormal healing such as pathological scar formation, etc. Not only affects the appearance, but also causes structural and functional obstacles, often causing heavy psychological pressure on the patient, seriously affecting the patient's physical and mental health. Therefore, how to quickly achieve the best healing effect in the treatment of skin wounds has been a hot topic of research.
  • the healing process of the skin is a complex, coordinated and orderly process of tissue repair and regeneration, which is a process in which a variety of cells and cytokines work together.
  • the process can be roughly divided into inflammatory reaction phase, proliferative phase and remodeling phase. There are no strict boundaries between the three phases, and a large number of cells and cytokines participate in the healing process.
  • Cells involved in healing include keratinocytes, vascular endothelial cells, fibroblasts, neutrophils, monocytes/macrophages, and mast cells; cytokines involved in healing include Fibroblast Growth Factor (FGF).
  • FGF Fibroblast Growth Factor
  • VEGF Vascular endothelial growth factor
  • PDGF Platelet derived growth factor
  • TGF- ⁇ Tumor necrosis factor- ⁇
  • TGF Transforming growth factor
  • IGF Insulin-like growth factors
  • Cytokines secrete cells to regulate wound healing, and cytokines in turn affect the function of cells. Abnormalities in any one of the links may cause abnormal skin healing results, and the skin healing process is affected by many factors (age, nutritional status, endocrine changes). , drugs, local blood circulation, infection, ionizing radiation and systemic diseases, etc., therefore, the mechanism of skin healing is still not completely clear.
  • the nervous system is a functional regulation system that plays a leading role in the human body.
  • the functions and various physiological processes of various organs and systems of the human body are connected, interacted, and closely coordinated under the direct or indirect regulation of the nervous system, thereby achieving and maintaining normal life activities.
  • the human nervous system can feel the changes of the external environment, accept changes in the internal and external environment, and constantly and thoroughly adjust various functions in the body to adapt the body to changes in the internal and external environment.
  • the nervous system consists of the central part and its peripheral part.
  • the central part includes the brain and the spinal cord, which are located in the cranial cavity and the spinal canal respectively. The two are closely related in structure and function to form the central nervous system.
  • the peripheral portion includes 12 pairs of cranial nerves and 31 pairs of spinal nerves, which constitute the peripheral nervous system. Peripheral nerves are distributed throughout the body, linking the brain and spinal cord to other organs throughout the body, allowing the central nervous system to sense both internal and external changes (transportation of sensory information through afferent nerves) and regulation of various functions in the body (through efferent nerves) Communicate adjustment instructions) to ensure the integrity of the human body and its adaptation to the environment.
  • the central nervous system composed of the human brain and spinal cord lacks self-regeneration and repair ability, and there is no effective treatment for cell death, tissue destruction and permanent loss of nerve function due to its damage.
  • Peripheral nerve injury is divided into open injury and non-open injury.
  • the former is generally associated with open injury of soft tissue, causing partial or complete truncation of the nerve; the latter is complicated by blunt non-open injury of soft tissue, causing contusion, compression or stretch of the nerve trunk, which occurs small in the nerve. Hemorrhage and edema, myelin edema and degeneration.
  • the results of peripheral nerve injury are mainly manifested as neurological paralysis in the clinic, and sensory disturbance, dyskinesia and muscle atrophy occur in the area innervated by the nerve.
  • Nerve growth factor enhancer 1.
  • Principal potassium is a cognitive enhancer for the treatment of mild to moderate Alzheimer's disease (AD). Increasing neurotrophic growth factor levels in damaged or degenerated neurons to enhance neuronal function, stimulate axon growth, and improve memory is the first drug to enter phase III clinical trials to enhance nerve regeneration.
  • acetyl L-carnitine is a cholinergic agonist, can actively pass the blood-brain barrier, in the neurodegenerative and aging model can protect the central and peripheral nervous synapses, improve the level of nerve growth factor, improve the elderly
  • Mouse cognitive deficits are currently undergoing Phase III clinical trials in the United States.
  • neurotrophic drugs 1, gangliosides (GM1), promote nerve remodeling (including nerve cell survival, axonal elongation and synaptic growth), in cell differentiation, development, nerve tissue repair, neuronal plasticity Other aspects play an important role.
  • GM1 gangliosides
  • brain protein hydrolysate a new drug to improve brain metabolism, easy to enter the brain nerve cells through the blood-brain barrier, affecting its protein synthesis and respiratory chain, enhance the antioxidant capacity of brain cells, protect the nervous system from toxic substances , delay brain cell death and promote its survival.
  • Cytidine disodium triphosphate injection which is a nucleotide drug that not only promotes protein synthesis, but also regulates and promotes the synthesis and construction of membrane structures of nerve cells, glial cells and vascular wall cells.
  • Oxacetam a new type of central nervous system drug that promotes learning and memory, selectively acts on the cerebral cortex and hippocampus to activate, protect or promote the recovery of nerve cells.
  • the primary object of the present invention is to provide a small molecule polypeptide which promotes repair and regeneration of tissue after trauma, and is named as a repair peptide.
  • the present invention provides a small molecule polypeptide consisting of 4-15 amino acid residues, which is a plurality of line peptides and cyclic peptides derived from the sequence GPAINVET, which are temporarily named as repair peptides, and the polypeptide sequence is obtained.
  • the phage peptide library (7-peptide library) was used as a target for the extramembranous segment of fibroblast growth factor receptor 2 (FGFR2), which was obtained by multiple rounds of screening and identification. So far, no literature has reported that the sequence polypeptide can be used for skin wound repair and nerve damage repair.
  • Another object of the present invention is to provide the use of the above-described repair peptide for the repair and regeneration of skin wounds and nerve damage.
  • An object of the present invention is to provide a repair peptide which promotes repair and regeneration of post-traumatic tissue, which contains four or more linear peptides or cyclic peptides of 4 to 15 amino acids in length of the GPANVET core sequence.
  • the R1 peptide sequence of the repair peptide is GPAINVET
  • the R2 peptide sequence is KVKVGPANVKVKV,
  • the R3 peptide sequence is GPANVEKEKEK
  • the R4 peptide sequence is CGPANVETC.
  • the R1 peptide contains only the core sequence GPAINVET; the R2 peptide contains 5 consecutive amino acids GPAINV of the core sequence; the R3 peptide contains 6 consecutive amino acids GPANVE of the core sequence; the R4 peptide contains all 7 amino acids of the core sequence, and
  • the disulfide bond of a cysteine forms a cyclic peptide.
  • repair peptides of the present invention can be obtained by chemical synthesis.
  • the product is preferably a medical product, a skin care product or a cosmetic.
  • the type of formulation of the product is preferably, but not limited to, a solution, lyophilizate, emulsion, cream, gel, mask or dressing, and the like.
  • the present invention is an experiment on animal models of mouse fibroblast balb/c 3T3, human vascular endothelial cell HUVEC, chicken embryo allantoic membrane, mouse and rat.
  • the repair peptide provided by the present invention is similar to the conventionally used recombinant bFGF (including 155 amino acids), and can also bind to the membrane receptor FGFR2 to promote skin wound repair and nerve damage repair and regeneration, but
  • the molecular weight is smaller, and the chemical synthesis method is used instead of the genetic engineering method, so the required production cost and the use cost are lower under the conditions of the same molar concentration, so it is a good substitute for the wound repair and cell regeneration of the recombinant bFGF. And proliferation products.
  • the recombinant bFGF is harsh in storage and transportation conditions in vitro, and is easily degraded, resulting in low accumulation of therapeutic sites and difficulty in sustained action, thus limiting and affecting the development of its use and related dosage forms to some extent, and the repair provided by the present invention.
  • the peptide Compared with growth factors such as bFGF, NGF and BDNF, the peptide has a greatly reduced degree of degradation because it has only 4 to 15 amino acids, and the experimental results disclosed in the present invention prove that the repair peptide can continue to act on the treatment site, so that it is technically Advancement.
  • Figure 1 is a graph showing the results of the ITC method for detecting the affinity of the repair peptides R1 and bFGF to FGFR2.
  • Figure 2 is a graph showing the results of CCK-8 assay for detecting the proliferative effects of the repair peptides R1, R2, R3, R4 and bFGF on mouse fibroblast Balb/c 3T3.
  • Figure 3 is a graph showing the results of CCK-8 assay for pro-cell proliferation of human umbilical vein endothelial cells (HUVEC) by repair peptides R1, R2, R3, R4 and bFGF.
  • HAVEC human umbilical vein endothelial cells
  • Figure 4 is a graph showing the results of repair peptide R1 promoting angiogenesis in chicken embryo chorioallantoic membrane.
  • Fig. 5 is a graph showing the results of promoting repair of angiogenesis in chicken embryo chorioallantoic membrane at the same concentration by the repair peptides R1, R2, R3, and R4.
  • Figure 6 is a graph showing the results of experimental repair of rat skin wound healing at the same concentration of repair peptides R1, R2, R3, and R4.
  • Figure 7 is a schematic diagram showing the establishment of a rat dorsal root nerve injury model and drug treatment.
  • Figure 8 is a retreat threshold for mechanical stimulation of rats measured after repair of dorsal root ganglion lesions by repair peptides R1, R2, R3, and R4: the ordinate indicates the threshold of the right hand than the number of left hand lifts. A larger value indicates that the more times the claw is lifted, the more sensitive it is to mechanical pain stimulation, and the better the sensory function is restored.
  • the sham was labeled as sham operation group, PBS was treated with PBS solution after trauma, and R1, R2, R3 and R4 were treated with different repair peptides. The results showed that repair peptide can improve the response to mechanical pain after injury in rats. , indicating a significant repair effect on dorsal root ganglion injury.
  • Figure 9 is a recovery test of heat pain stimulation in rats after repair of dorsal root ganglion injury with repair peptides R1, R2, R3, and R4: the ordinate indicates the enthalpy of the right hand than the left hand. The smaller the value, the shorter the time required to lift the claw, the more sensitive it is to the heat pain stimulus, and the better the sensory function recovery.
  • the sham was labeled as sham operation group, PBS was treated with PBS solution after trauma, and R1, R2, R3 and R4 were treated with different repair peptides. The results showed that repair peptide can improve the response to heat pain after injury in rats. , indicating a significant repair effect on dorsal root ganglion injury.
  • Figure 10 shows the detection of neuronal axon growth by indirect immunofluorescence 2 weeks after detection of dorsal root ganglion pinch injury using the repair peptide.
  • the antibody and fluorescent secondary antibody of marker protein NF200 were used.
  • NF200 is a marker protein of nerve fibers.
  • Figure 11 shows the detection of neuronal axon growth by indirect immunofluorescence 2 weeks after detection of dorsal root ganglion pinch injury using the repair peptide.
  • the antibody and fluorescent secondary antibody of the marker protein Laminin were used.
  • Laminin is a marker protein of peripheral nerve fibers.
  • Figure 12 shows the detection of neuronal axon growth by indirect immunofluorescence 2 weeks after detection of dorsal root ganglion pinch injury using the repair peptide.
  • the antibody and fluorescent secondary antibody of the marker protein CGRP were used.
  • CGRP proteins are associated with pain receptors that transmit pain.
  • the affinity detected by isothermal microcalorimeter is shown in Fig. 1.
  • K represents an affinity constant
  • Kd represents a dissociation constant
  • K is larger, and Kd is smaller, indicating a stronger affinity).
  • HUVEC cells were harvested, 96 plates per well were plated, and after starvation for 24 hours, the repair peptides R1, R2, R3, R4 and bFGF were taken and treated at 5 ⁇ doubling concentration for 24 hours, followed by CCK-8 reagent. Processing, the microplate reader detects the OD value of each well. As shown in Fig. 3, in the lower concentration group ( ⁇ 0.16 ⁇ M), the repair peptides R1, R2, R3, R4 and bFGF proliferated, but in the high concentration group, the bFGF group showed inhibition of cell proliferation, but the repair peptide R1, R2, R3, and R4 still have proliferative activity. The results suggest that the above repair peptides can promote the growth of vascular endothelial cells, and the effect is sustained.
  • repair peptide R1 can promote the growth of blood vessels in the chicken embryo chorioallantoic membrane
  • the fertilized egg cultured for 5 days is taken, the egg head is turned up, the chicken embryo urinary sac film is exposed, the silica gel ring is placed in the middle, and 10 ⁇ L of 25 ⁇ doubling concentration of R1 is added to the silica gel ring, and the clean filter paper is closed to open the opening.
  • the PBS solution treatment group was a blank control.
  • the chick embryo chorioallantoic membrane was peeled off, observed under a stereoscopic microscope, photographed, and image ImageJ plus was used to analyze the image.
  • the capillaries in the chicken chorioallantoic membrane increased significantly and increased with increasing dose. It is indicated that R1 significantly promotes the formation of blood vessels.
  • repair peptides R1, R2, R3, R4 promote the detection of angiogenesis in chicken embryo chorioallantoic membrane
  • the fertilized egg cultured for 5 days was taken, the egg head was turned up, and the chicken embryo chorioallantoic membrane was exposed.
  • 100 ⁇ L of the repair peptides R1, R2, R3 and R4 at a concentration of 100 ng/mL were added thereto, and a clean filter paper was used to close the opening.
  • the PBS solution treatment group was a blank control.
  • the chick embryo chorioallantoic membrane was peeled off, observed under a stereoscopic microscope, photographed, and image ImageJ plus was used to analyze the image. As shown in Fig.
  • This model was used to examine the effect of drugs on the recovery of sensory function in peripheral nerves of rats.
  • the specific method is as shown in Fig. 7, and the spinal cord C5 to T1 segments of the rat are exposed by a surgical method. Use the tweezers to clamp the right dorsal root at the same strength, so that the dorsal root is damaged but does not break. After that, the test drug was injected into the attachment of the rat every day for 3 weeks and 21 days.
  • the mechanical pain stimulation test is a classical method widely used to examine the sensory function of neuropathic pain animals.
  • the rat stands on an elevated platform with a wide gauge wire mesh. Von-Frey fibers (which are very precise calibration wires) are inserted from below and pass through the mesh to puncture the lower surface of the front paw. On the threshold, the animal quickly prys its claws off the fibers.
  • the mechanical withdrawal threshold is defined as the minimum gauge line stimulus that causes the withdrawal response. Each foot (left front and right front) was measured 5 times, and the number of claw lifts was recorded.
  • the ordinate indicates the enthalpy of the number of times the right hand is lifted compared to the left hand. A larger value indicates that the more times the claw is lifted, the more sensitive it is to mechanical pain stimulation, and the better the sensory function is restored.
  • the sham was labeled as sham operation group, PBS was treated with PBS solution after trauma, and R1, R2, R3 and R4 were treated with different repair peptides. The results showed that repair peptide can improve the response to mechanical pain after injury in rats. , indicating a significant repair effect on dorsal root ganglion injury.
  • the rats were placed in a clear plastic chamber (18 x 29 x 13 cm, 2.2 mm thick). The animals were placed in the environment 20 minutes prior to testing. The heat source is then placed under the glass plate below the right front or left front paw. The claw retracts the automatic activation timer with a 0.1 second delay. Three measurements were taken at 15 minute intervals and the average paw withdrawal time was recorded.
  • the ordinate indicates the ⁇ value of the right hand than the left hand lifting time.
  • the sham was labeled as sham operation group, PBS was treated with PBS solution after trauma, and R1, R2, R3 and R4 were treated with different repair peptides. The results showed that repair peptide can improve the response to heat pain after injury in rats. , indicating a significant repair effect on dorsal root ganglion injury.
  • the rats enter the deep anesthesia state, open the chest cavity, expose the heart, gently cut the right atrial appendage, and insert the needle from the quasi-apical position to the left ventricle.
  • the peristaltic pump was turned on, firstly perfused with PBS, and then perfused with 4% PFA fixative until the liver color of the rat was pink and the body was stiff.
  • C5-T1 spinal cord tissue and dorsal root were excised with surgical instruments, fixed in 4% PFA for 24 h, overnight at 15% sucrose, and then overnight at 30% sucrose. After being embedded in OCT, the frozen section can be frozen, the thickness is 15 ⁇ m, and only the section of the tissue of the C7 section is taken for the next experiment.
  • the prepared tissue was cryosectioned, and the liquid outside the specimen was gently wiped off with a filter paper, and the tissue was circled with an immunohistochemical pen, and then blocked with 10% sputum serum in a wet box for 1 hour at room temperature. Then, the blocking solution outside the specimen was gently wiped off with a filter paper, and the primary antibody (1:100) diluted with 10% sputum serum was added dropwise, and incubated overnight at 4 degrees in a wet box. The primary antibody was discarded and the PBS solution was washed 3 times for 5 min each time.
  • the liquid outside the specimen was wiped off with a filter paper, and the fluorescently labeled secondary antibody diluted with 10% sputum serum was added dropwise, and then incubated in a wet box at room temperature for 2 hours.
  • the secondary antibody was aspirated and the PBS solution was lightly 3 times for 5 min each time.
  • an anti-fluorescence quencher containing DAPI dye solution was added dropwise, and then sealed with a clean cover glass and stored at 4 degrees in the dark.
  • the antibodies were: NF200 (purchased from abcam, article number ab40796), Laminin (purchased from sigma, product number L9393), CGRP (purchased from sigma, product number C8198), counterstained with fluorescent secondary antibody (purchased from invitrogen) Observe and photograph under a fluorescence microscope.
  • NF200 is a marker protein of nerve fibers, which can observe the growth of axons of neurons
  • Laminin is a marker protein of peripheral nerve fibers, which can observe the growth of axons stimulated by drugs
  • CGRP is related to pain receptors that transmit pain.
  • the labeled sham was a sham operation group
  • PBS was treated with PBS solution after trauma
  • R1, R2, R3, and R4 were groups to be tested treated with different repair peptides.
  • the results are shown in Figures 10 to 12. It can be seen from the experimental test that the repair peptide treatment group can significantly promote the growth of synapses and promote the repair of damaged dorsal root nerves.

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Abstract

提供一种促进创伤后组织修复与再生的修复肽及其应用，所述修复肽为含有GPANVET核心序列的连续4个及其以上的4-15个氨基酸长度的线性肽或环肽。

Description

一种促进创伤后组织修复与再生的修复肽及其应用 技术领域
本发明属于生物医药领域，特别涉及一种促进创伤后组织修复与再生的修复肽及其应用，它能促进创伤部位组织修复与细胞的增殖，特别是在皮肤损伤、神经损伤方面的组织再生和修复方面的应用。
背景技术
许多因素会造成细胞和组织的损伤。损伤后组织部位及其附近存活的健康细胞不断进行分裂和增殖，以取代死亡细胞和修复受损组织，生物体的这种生理机能称为再生和修复。
正常情况下，人体不同器官的修复和再生能力有所不同。表皮细胞(如呼吸道、消化管和泌尿生殖器的粘膜被覆上皮)、淋巴细胞、造血细胞等属于再生力强的细胞，这些种类的细胞每时每刻都在进行衰老与新生，具有应对损伤的强大再生修复能力，因此属于再生能力较强的细胞。而其他如血管内皮细胞、骨膜细胞、各种腺体器官的细胞如肝、胰、内分泌腺、汗腺、皮脂腺及肾小管上皮细胞等则属于次一级的有较强再生力的细胞，当受到损伤时，也能表现出一定的再生能力。具体反映在局部组织当这些正常的细胞被破坏后，由残留的同类细胞分裂、补充；而如果该局部的细胞完全被破坏，细胞全部坏死，则该部位就不能被修复。另外，还有再生力微弱或无再生力的细胞，如中枢神经细胞和神经节细胞再生很弱，遭损坏后极难恢复原有功能；心肌细胞再生能力极弱，损毁后均由纤维结缔组织代替，很难恢复原有的结构和功能。
影响组织再生的因素有很多。除了组织和细胞自身的再生能力外，还有如下几种因素：(1)组织受损伤的程度和范围：越大则修复和再生的时间越长，因为大范围的细胞坏死后，难以有相当数量的同类细胞代替，需要健康的细胞产生新细胞，新生细胞再生成新细胞，需要的时间就要漫长一些。(2)年龄因素：儿童和青少年组织再生能力强，创伤愈合快；老年人组织再生能力弱，愈合慢。(3)营养状况：充足的蛋白质、维生素C、E和矿物质钙、锌供应能够促进损伤的修复，反之则会延缓各种损伤的修复。(4)药物影响：有些药物如肾上腺皮质激素和垂体促肾上腺皮质激素能抑制炎症，但不利于机体消除伤口感 染，还能抑制肉芽组织生长和胶原合成，加速胶原分解；还有抗癌药中的细胞毒药物也可延缓愈合。(5)血液供应：组织损伤后，由于局部毛细血管也受到破坏，或者其他因素导致血管硬化等，都会导致血液供应不足而导致组织营养不良，妨碍愈合。(6)神经支配：失去神经支配的组织会丧失再生能力。
从上可知，机体的组织受到损失后，除却无法外界干预的如年龄等客观因素外，从增加受损部位的营养供给、采用药物干预其促进存活细胞增殖、促进毛细血管的新生和神经细胞的再生等，可以加快组织的修复。
以下从目前组织修复的三大应用领域：皮肤、神经和骨组织的创伤修复情况进行更为详细的介绍：
一、皮肤组织的创伤修复
皮肤是人体面积最大的且最直接与外界接触组织，同时也是最容易受到外界刺激而损伤的器官。正常创伤的愈合对维持机体稳态至关重要，而异常愈合如大面积烧烫伤造成的不愈或者迟愈、其他疾病导致的慢性创面愈合、以及不正常的愈合如病理性瘢痕的形成等等不仅影响了美观，更会造成结构和功能的障碍，往往对患者造成沉重的心理压力，严重影响着患者的身心健康。因此，如何在皮肤创伤治疗中快速达到最佳愈合效果，一直是人们研究的热点。
皮肤的愈合过程是一个复杂的、协调有序的组织修复再生过程，是多种细胞及细胞因子共同作用完成的过程。该过程大致可以分为炎症反应期、增生期和重塑期，三个期相互交叉并无严格的界线，且均有大量的细胞及细胞因子参与愈合过程。参与愈合的细胞包括角质细胞、血管内皮细胞、成纤维细胞、中性粒细胞、单核/巨噬细胞和肥大细胞等；参与愈合的细胞因子包括成纤维细胞生长因子(Fibroblast Growth Factor，FGF)、血管内皮生长因子(Vascular endothelial growth factor，VEGF)、血小板衍生因子(Platelet derived growth factor，PDGF)、肿瘤坏死因子-α(Tumor necrosis factor-α，TNF-α)、转化生长因子(Transforming growth factor，TGF)、胰岛素样生长因子(Insulin-like growth factors，IGF)和多种凝血因子等。细胞分泌细胞因子调节创伤愈合，细胞因子又反过来影响细胞的功能，其中任何一个环节的异常都可能造成皮肤愈合结果的异常，而且皮肤愈合过程受到的影响因素众多(年龄、营养状况、内分泌变化、药物、局部血液循环、感染、电离辐射和系统性疾病等)，因此，皮肤愈合的机制目前仍不完全明确。
目前临床上，皮肤创伤修复主要针对急性皮肤软组织创伤、慢性皮肤软组织创伤来进行不同的用药和处理。基于我国传统中医药的优势，在创伤修复领 域涌现了许多单方如康复新液，复方则由少则四、五味，多则二三十味中药进行配伍制备而来，例如治疗烧烫伤的“湿润烧伤膏”、“京万红”等。现代医药开发特别是基因工程技术介入的生物医药开发后，促细胞生长的重组生长因子的生产也是创伤修复药物开发的生力军，比如已经在临床上应用于创伤、烧烫伤等的生物技术药物--外用重组碱性成纤维细胞生长因子(basic Fibroblast Growth Factor，bFGF)，就是我国第二个基因工程I类新药(注：碱性成纤维细胞生长因子又称为成纤维细胞生长因子-2，FGF2)。
与皮肤修复的多肽药物开发方面，2014年中科院昆明动物研究所报道了来源于蜥蜴的一条强效的皮肤修复肽Tylotoin，含12个氨基酸残基，表现出强烈的促创伤愈合活性，可以促进角质上皮和成纤维细胞的增殖和迁移，还能促进TGF-b和IL-6的生成。也有中国发明专利公开了从华南雨蛙、红瘰疣螈、无指盘臭蛙中分别提取出了能促进皮肤修复的多肽，分别为16肽、13肽、24和11肽。
二、神经损伤的创伤修复
神经系统是人体内起主导作用的功能调节系统。人体各器官、系统的功能和各种生理过程在神经系统的直接或间接调节控制下，互相联系、相互影响、密切配合，从而实现和维持正常的生命活动。同时，人体的神经系统能感受到外部环境的变化，接受内外环境的变化信息，对体内各种功能不断进行迅速而完善的调整，使人体适应体内外环境的变化。
神经系统由中枢部分及其外周部分所组成。中枢部分包括脑和脊髓，分别位于颅腔和椎管内，两者在结构和功能上紧密联系，组成中枢神经系统。外周部分包括12对脑神经和31对脊神经，它们组成外周神经系统。外周神经分布于全身，把脑和脊髓与全身其他器官联系起来，使中枢神经系统既能感受内外环境的变化(通过传入神经传输感觉信息)，又能调节体内各种功能(通过传出神经传达调节指令)，以保证人体的完整统一及其对环境的适应。
人类大脑和脊髓组成的中枢神经系统缺乏自我再生和修复能力,因其损伤导致的细胞死亡，组织破坏及神经功能永久性缺失，目前尚无有效的治疗手段。
外周神经损伤分开放性损伤和非开放性损伤。前者一般伴发于软组织的开放性损伤，引起神经的部分截断或全截断；后者并发于软组织的钝性非开放性损伤，引起神经干的挫伤、压迫或牵张，在神经内发生小的溢血和水肿，髓鞘水肿和变性。外周神经损伤的结果在临床上主要表现为神经麻痹，受该神经支配的区域出现感觉障碍，运动障碍和肌肉萎缩等。
在临床上一般神经损伤的药物有：一、神经生长因子增强剂：1、普立宁钾，是一种认知增强剂，用于治疗轻、中度阿尔茨海默病(AD)，通过提高受损或退化神经元的神经营养生长因子水平来增强神经细胞功能，刺激轴突生长，改善记忆能力，是首个进入Ⅲ期临床试验用于增强神经再生的药物。2、乙酰L-肉碱：是一种胆碱能激动药，能主动通过血脑屏障，在神经退行性及衰老模型中可保护中枢及周围神经突触，提高神经生长因子水平，改善老年大鼠认知缺陷，目前正在美国进行Ⅲ期临床试验。二、神经营养类药物有；1、神经节苷脂(GM1)，促进神经重构(包括神经细胞生存、轴突延长和突触生长)，在细胞分化、发育、神经组织修复、神经元可塑性等方面起重要作用。2、脑蛋白水解物，为一种改善脑代谢的新药，易透过血脑屏障进入大脑神经细胞，影响其蛋白质合成及呼吸链，增强脑细胞抗氧化能力，保护神经系统免受有毒物质侵害，延缓脑细胞死亡，促进其存活。3、三磷酸胞苷二钠注射液，这是一种核苷酸类药物，不仅可促进蛋白质合成，还可调节和促进神经细胞、神经胶质细胞及血管壁细胞膜性结构的合成与构建，能对抗由兴奋性氨基酸、自由基引起的神经细胞损伤，从而具有支持存活、增强活性、延缓死亡，提高细胞抗损伤和修复能力，促进神经轴突再生长，并改善支配血管的神经功能，起到抗血管硬化作用。4、奥拉西坦，它是一类能促进学习记忆能力的新型中枢神经系统药物，能选择性地作用于大脑皮层和海马，激活、保护或促进神经细胞的功能恢复。
发明内容
为了克服现有技术的缺点与不足，本发明的首要目的在于提供一种促进创伤后组织修复与再生的小分子多肽，命名为修复肽。本发明提供的是一种4～15个氨基酸残基组成的小分子多肽，该种多肽是基于序列GPANVET而衍生的若干线肽和环肽，均暂命名为修复肽，该多肽序列的获得是通过噬菌体肽库(7肽库)，以成纤维细胞生长因子受体2(FGFR2)的膜外段为靶点，多轮筛选并鉴定而获得的。目前为止尚无文献报道该序列多肽能用于皮肤创伤修复和神经损伤修复。
本发明的另一目的在于提供上述修复肽的在皮肤创伤和神经损伤修复和再生方面的应用。
本发明的目的通过下述技术方案实现：
本发明的目的是提供一种促进创伤后组织修复与再生的修复肽，该修复肽含有GPANVET核心序列的连续4个及其以上的4～15个氨基酸长度的线性肽 或环肽。
具体的，所述的修复肽的R1肽序列为GPANVET，
R2肽序列为KVKVGPANVKVKV，
R3肽序列为GPANVEKEKEK，
R4肽序列为CGPANVETC。
其中R1肽仅含有核心序列GPANVET；R2肽含有核心序列的5个连续氨基酸GPANV；R3肽含有核心序列的6个连续的氨基酸GPANVE；R4肽含有核心序列的全部7个氨基酸，且通过两端两个半胱氨酸的二硫键形成环肽。
本发明所述的这些修复肽均可以采用化学合成的方法获得。
本发明的目的是提供修复肽在制备皮肤损伤、神经损伤方面的组织再生和修复方面的产品中的应用，特别是在制备皮肤创伤、烧烫伤、慢性创面愈合、皮肤细胞修复和新生，以及脑外伤、脑中风、脑水肿以及脑缺氧等造成的中枢神经损伤，和外周神经损伤造成的感觉障碍、运动障碍和营养障碍的神经修复和再生的产品中的应用。
所述的产品优选为医用产品、护肤品或化妆品。
所述的产品的制剂类型优选但不局限为溶液、冻干剂、乳剂、霜剂、凝胶、面膜或敷料等。
本发明的内容为在小鼠成纤维细胞balb/c 3T3、人血管内皮细胞HUVEC、鸡胚尿囊膜、小鼠和大鼠的动物模型上的实验。
本发明相对于现有技术具有如下的优点及效果：
本发明提供的修复肽是与目前类似的产品外用重组bFGF(包括155个氨基酸)相比，同样能跟膜受体FGFR2结合发挥作用，促皮肤创伤修复和神经损伤修复和再生的效果相当，但分子量更小，而且采用化学合成方法而非基因工程方法，故在同等摩尔浓度的使用条件下，所需的生产成本和使用成本更低，所以是很好的替代重组bFGF的创伤修复和细胞再生和增殖产品。而且重组bFGF在体外保存和运输条件较为苛刻，易降解，导致治疗部位的积累低，难以持续作用，因此在某些程度上限制和影响了其使用和相关剂型的开发，而本发明提供的修复肽相对bFGF、NGF和BDNF等生长因子，因为仅有4到15个氨基酸，故降解程度大大降低，而且本发明公开的实验结果证明，该类修复肽能持续作用于治疗部位，因此具有技术上的先进性。
附图说明
图1是ITC法检测修复肽R1和bFGF与FGFR2的亲和力的结果图。
图2是CCK-8法检测修复肽R1、R2、R3、R4和bFGF对小鼠成纤维细胞Balb/c 3T3的促细胞增殖作用的结果图。
图3是CCK-8法检测修复肽R1、R2、R3、R4和bFGF对人脐静脉血管内皮细胞(HUVEC)的促细胞增殖作用的结果图。
图4是修复肽R1促进鸡胚尿囊膜中血管生成的结果图。
图5是修复肽R1、R2、R3、R4在同等浓度下促进促进鸡胚尿囊膜中血管生成的结果图。
图6是修复肽R1、R2、R3、R4在同等浓度下促进大鼠皮肤创伤愈合的实验结果图
图7是大鼠背根神经损伤模型的建立及药物治疗的示意图。
图8是修复肽R1、R2、R3、R4修复背根神经节损伤后的测量大鼠对机械刺激的撤退阈值：纵坐标表示右手比左手抬爪次数的阙值。值越大表示抬爪次数越多，对机械痛刺激越敏感，感觉功能恢复越好。标记sham为假手术组，PBS为创伤后用PBS溶液处理组，R1、R2、R3、R4为采用不同修复肽处理的待测组，结果可知修复肽能提高大鼠损伤后对机械痛的反应，说明对背根神经节损伤有显著的修复作用。
图9是修复肽R1、R2、R3、R4修复背根神经节损伤后的大鼠对热痛刺激恢复实验：纵坐标表示右手比左手抬爪时间的阙值。值越小表示抬爪所需时间越短，对热痛刺激越敏感，感觉功能恢复越好。标记sham为假手术组，PBS为创伤后用PBS溶液处理组，R1、R2、R3、R4为采用不同修复肽处理的待测组，结果可知修复肽能提高大鼠损伤后对热痛的反应，说明对背根神经节损伤有显著的修复作用。
图10是检测背根神经节夹伤后，采用修复肽处理后2周，间接免疫荧光法检测神经元轴突的生长情况，图示采用的是标志蛋白NF200的抗体和荧光二抗。NF200为神经纤维的标志蛋白。
图11是检测背根神经节夹伤后，采用修复肽处理后2周，间接免疫荧光法检测神经元轴突的生长情况，图示采用的是标志蛋白Laminin的抗体和荧光二抗。Laminin为周围神经纤维的标志蛋白。
图12是检测背根神经节夹伤后，采用修复肽处理后2周，间接免疫荧光法检测神经元轴突的生长情况，图示采用的是标志蛋白CGRP的抗体和荧光二抗。 CGRP蛋白则与传导疼痛的疼痛感受器有关。
具体实施方式
下面结合实施例及附图对本发明作进一步详细的描述，但本发明的实施方式不限于此。
下列实施例中所使用的实验方法如无特殊说明，均为常规方法，实施例中所使用的实验材料、试剂等如无特殊说明，均为商业途径获取。应理解，这些实施例仅用于说明本发明而不是用于限制本发明的范围。
实施例1
1、ITC法检测修复肽R1与FGFR2的亲和力
采用等温微量量热仪(ITC)检测的亲和力，结果如图1所示，修复肽R1与FGFR2的亲和常数K＝7.35E8±2.10E8M -1，解离常数Kd≈1.37nM，而bFGF与FGFR2的亲和常数为K＝1.62E5±2.50E4M -1，解离常数Kd≈6.17μM。结果显示修复肽R1与FGFR2的亲和力远远高于bFGF与FGFR2的亲和力，前者是后者的约4500倍。(K表示亲和常数，Kd表示解离常数，K越大，Kd越小，表示亲和力越强)。
2、CCK-8法检测修复肽R1、R2、R3、R4和bFGF对成纤维细胞Balb/c 3T3的促增殖作用
取Balb/c 3T3细胞，按每孔4000个细胞铺96板，贴壁后饥饿24小时后，取修复肽R1、R2、R3、R4和bFGF，按5×倍增浓度处理24小时，之后采用CCK-8试剂处理，酶标仪检测各个孔OD值。如图2所示，在较低浓度组，修复肽R1、R2、R3、R4与bFGF促增殖活性相当，而高浓度组，bFGF组出现细胞增殖抑制的现象，但修复肽R1、R2、R3、R4依然有促增殖活性。结果提示，上述修复肽与bFGF都能促进成纤维细胞增殖方面，且没有出现高浓度抑制增殖的现象，说明其作用持续性好。
3、CCK-8法检测修复肽R1、R2、R3、R4和bFGF对人脐静脉血管内皮细胞(HUVEC)的促增殖作用
取HUVEC细胞，按每孔4000个细胞铺96板，贴壁后饥饿24小时后，取修复肽R1、R2、R3、R4和bFGF，按5×倍增浓度处理24小时，之后采用CCK-8试剂处理，酶标仪检测各个孔OD值。如图3所示，在较低浓度组(≦0.16μM)， 修复肽R1、R2、R3、R4与bFGF促增殖活性相当，而高浓度组，bFGF组出现细胞增殖抑制的现象，但修复肽R1、R2、R3、R4依然有促增殖活性。结果提示，上述修复肽均能促进血管内皮细胞生长，且作用持续性好。
4、修复肽R1能促进鸡胚尿囊膜中血管的生长
取培养5天的受精鸡蛋，大头朝上去蛋壳，暴露鸡胚尿囊膜，放置硅胶圈于正中，将25×倍增浓度的R1各10μL滴加于硅胶圈中，干净的滤纸封闭开口，设立PBS溶液处理组为空白对照。再培养3天，之后剥取鸡胚尿囊膜，体视镜下观察，拍照，软件ImageJ plus对图像进行分析处理。如图4所示，加入R1后，鸡胚尿囊膜中的毛细血管明显增多，随着剂量增加而增加。说明R1明显促进了血管的生成。
5、修复肽R1、R2、R3、R4促进促进鸡胚尿囊膜中血管生成的检测
取培养5天的受精鸡蛋，大头朝上去蛋壳，暴露鸡胚尿囊膜，将100ng/mL浓度的修复肽R1、R2、R3、R4各100μL滴加其上，干净的滤纸封闭开口，设立PBS溶液处理组为空白对照。再培养3天，之后剥取鸡胚尿囊膜，体视镜下观察，拍照，软件ImageJ plus对图像进行分析处理。如图5所示，分别加入修复肽R1、R2、R3和R4后，鸡胚尿囊膜中的毛细血管明显增多，随着剂量增加而增加。说明上述修复肽可以促进血管的生成。
6、修复肽R1、R2、R3、R4促进大鼠皮肤创伤愈合的检测
取Balb/c小鼠，在1％的戊巴比妥钠腹腔注射麻醉下(100mg/kg)，背部脱毛并消毒，1％碘伏及75％酒精常规消毒。对称两侧各切1cm长刀口，消毒外科手术缝合两针，再将修复肽的R1、R2、R3和R4肽各10μL滴加于伤口处，伤口开放不覆盖棉纱，设立PBS溶液处理组为对照组。之后每日滴加一次，同时观察伤口愈合情况。如图6所示，加入修复肽的R1、R2、R3和R4肽后，愈合速度增快。说明上述修复肽明显促进了皮肤创伤的愈合，并促进愈合部位血管的生成。
7、大鼠背根神经损伤模型的建立
该模型用于检测药物对大鼠外周神经感觉功能恢复的影响。
具体方法如图7所示，通过外科手术的方法将大鼠脊髓C5到T1节段暴露 出来。用镊子在同等力度下夹右侧背根，使其背根损伤但不断裂。之后每天在大鼠损伤处附件注射待测药物，连续观察3周21天。
8、修复肽R1、R2、R3、R4修复背根神经节损伤后的测量大鼠对机械刺激的反应
机械痛刺激试验是一种广泛用于检查神经性疼痛动物的感觉功能的经典方法。
具体方法：大鼠站立在高架平台上，其中表面是宽规格的金属丝网。Von-Frey纤维(是非常精密的校准金属线)从下面插入，穿过网孔，戳穿前爪的下表面。在门槛上，动物快速地将它的爪子从纤维上甩开。机械性退缩阈值被定义为引起退缩反应的最小规格线刺激。每只脚(左前和右前)分别测量5次，记录抬爪的次数。
如图8所示，纵坐标表示右手比左手抬爪次数的阙值。值越大表示抬爪次数越多，对机械痛刺激越敏感，感觉功能恢复越好。标记sham为假手术组，PBS为创伤后用PBS溶液处理组，R1、R2、R3、R4为采用不同修复肽处理的待测组，结果可知修复肽能提高大鼠损伤后对机械痛的反应，说明对背根神经节损伤有显著的修复作用。
9、修复肽R1、R2、R3、R4修复背根神经节损伤后的大鼠对热痛刺激恢复实验
将大鼠置于透明塑料室(18×29×13cm，2.2mm厚)中。在测试前20分钟时放入使动物适应环境。然后将热源放置在右前或左前爪下方的玻璃板下。爪撤回自动激活计时器，有0.1秒的延迟。以15分钟的间隔进行3次测量并记录平均缩爪时间。
如图9所示，纵坐标表示右手比左手抬爪时间的阙值。值越小表示抬爪所需时间越短，对热痛刺激越敏感，感觉功能恢复越好。标记sham为假手术组，PBS为创伤后用PBS溶液处理组，R1、R2、R3、R4为采用不同修复肽处理的待测组，结果可知修复肽能提高大鼠损伤后对热痛的反应，说明对背根神经节损伤有显著的修复作用。
10、间接免疫荧光法检测神经元轴突的生长情况
到待测时间点(2周或4周)，待大鼠进入深度麻醉状态后，打开胸腔，暴 露心脏，轻轻剪破右心耳，从准心尖位置进针至左心室。打开蠕动泵，先以PBS灌注，再以4％PFA固定液灌注，至大鼠肝脏颜色为桃黄色、身体僵直。
用手术器械剥离出C5-T1脊髓组织及背根，在4％PFA中后固定24h，在15％蔗糖中4度过夜，再在30％蔗糖中4度过夜。用OCT包埋后即可进行冰冻切片，厚度15μm，只收C7节段的组织的切片进行下一步实验。
取制作好的组织冷冻切片，用滤纸轻轻擦去标本外的液体，用免疫组化笔圈住组织后，用10％驴血清在湿盒中室温封闭1小时。接着用滤纸轻轻擦去标本外的封闭液，滴加用10％驴血清稀释的一抗(1：100)，于湿盒中4度孵育过夜。弃去一抗，PBS溶液清洗3次，每次5min。用滤纸擦去标本外的液体，滴加用10％驴血清稀释的荧光标记二抗后，置于湿盒中避光室温孵育2h。吸去二抗，PBS溶液轻3次，每次5min。用滤纸擦去标本外的液体后，滴加含有DAPI染液的抗荧光淬灭剂，然后用干净的盖玻片封片，并于4度避光处保存。
抗体分别为：NF200(购自abcam公司，货号ab40796)，Laminin(购自sigma公司，货号L9393)，CGRP(购自sigma公司，货号C8198)，用荧光二抗(购自invitrogen公司)进行复染，荧光显微镜下观察并拍照。
其中：NF200为神经纤维的标志蛋白，可观察神经元轴突的生长情况；Laminin为周围神经纤维的标志蛋白，可观察药物刺激轴突生长的情况；CGRP则与传导疼痛的疼痛感受器有关。
标记sham为假手术组，PBS为创伤后用PBS溶液处理组，R1、R2、R3、R4为采用不同修复肽处理的待测组。结果如图10～12所示，通过实验检测可知，修复肽处理组能明显促进神经突触的生长，促进受损背根神经的修复。
上述实施例为本发明较佳的实施方式，但本发明的实施方式并不受上述实施例的限制，其他的任何未背离本发明的精神实质与原理下所作的改变、修饰、替代、组合、简化，均应为等效的置换方式，都包含在本发明的保护范围之内。

Claims (10)

  1. 一种促进创伤后组织修复与再生的修复肽，其特征在于：
    所述修复肽为含有GPANVET核心序列的连续4个及其以上的4～15个氨基酸长度的线性肽或环肽。
  2. 根据权利要求1所述的促进创伤后组织修复与再生的修复肽，其特征在于：
    所述的修复肽的T1肽序列为GPANVET的线性肽。
  3. 根据权利要求1所述的促进创伤后组织修复与再生的修复肽，其特征在于：
    所述的修复肽的T2肽序列为KVKVGPANVKVKV的线性肽。
  4. 根据权利要求1所述的促进创伤后组织修复与再生的修复肽，其特征在于：
    所述的修复肽的T3肽序列为GPANVEKEKEK的线性肽。
  5. 根据权利要求1所述的促进创伤后组织修复与再生的修复肽，其特征在于：
    所述的修复肽的T4肽序列为CGPANVETC的环肽。
  6. 权利要求1～5任一项所述的促进创伤后组织修复与再生的修复肽在制备皮肤损伤、神经损伤方面的组织再生和修复方面的产品中的应用。
  7. 根据权利要求6所述的应用，其特征在于：
    所述的促进创伤后组织修复与再生的修复肽在制备皮肤创伤、烧烫伤、慢性创面愈合、皮肤细胞修复和新生、神经损伤修复和再生的产品中的应用。
  8. 根据权利要求6或7所述的应用，其特征在于：
    所述的产品为医用产品、护肤品或化妆品。
  9. 根据权利要求6或7所述的应用，其特征在于：
    所述的产品的制剂类型为溶液、冻干制剂、乳剂、霜剂、凝胶、面膜或敷料。
  10. 根据权利要求8所述的应用，其特征在于：
    所述的产品的制剂类型为溶液、冻干制剂、乳剂、霜剂、凝胶、面膜或敷料。
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