I. Brief Introduction of Collagen
Collagen collagen originating from ancient Greece and appearing around 1865, is the most abundant protein in the human body. Collagen is widely distributed in the human connective tissue, such as skin, cartilage and bone, tendons, ligaments, cornea, organ capsule, dura mater, etc. In normal people, 90% of collagen is found in skin and bone. Under the microscope, collagen accounted for 95% of the dermal connective tissue. Normal human bones contain 80% collagen; Tendon tissues contain up to 85% collagen (dry weight after degreasing). The most abundant collagen type Ⅰ, accounts for about 80% ~ 90% of the total collagen.
As the main component of extracellular matrix, collagen not only ACTS as a physical scaffold structure, but also can realize signal transmission between cells and extracellular matrix receptor molecules and participate in the regulation of cell behavior. Collagen can be directly bound to the cell surface receptor, but also indirectly bound to other intermediary molecules, involved in cell surface receptor recognition and signal transduction process. Collagen also plays a very important role in its dependent dynamic cellular activities, such as tissue reconstruction, including morphological processes and wound healing, and plays an important role in immune system regulation, hemostasis and thrombosis.
胶原在体内以多级聚合的方式存在 |
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资料来源：J. K. Mouw, et al., Nature Reviews Molecular Cell Biology 15:771–785 (2014) |
Collagen is structurally composed of three polypeptide chains arranged in the left helix, and three independent collagen peptide chains rely on hydrogen bonds formed between glycine to maintain the structure of three-helix intertwining. Many large molecules of collagen can form a cross-linked structure of fibers side by side, which makes the final product have high mechanical strength. Collagen normal three helical conformation is the basis of its physical and chemical properties and biological activity, make its have high tensile strength, low biodegradability, low antigen activity, stimulating, low cytotoxicity and skeleton as artificial organ or wound dressing promote cell growth, promote cell adhesion, and with new cells and tissues to repair traumatic features, this make collagen a broad scope of biomedical materials, has been used in dressing, artificial organ regenerative medicine, tissue engineering and biological skin care and other fields.
I. Application of collagen
1) Application of collagen in the field of medicine
Since 1976 when collagen products were listed in medical devices for management, approval and marketing in the United States, the application of collagen products and collagen-based biomedical materials in clinical practice has increased day by day in recent years, involving many clinical disciplines.
The triple helical structure of collagen decided the collagen has good biomechanical performance, similar sources of biological properties of collagen and collagen in high similarity between the species determines the collagen has good biocompatibility, collagen material in both as a new organization form of skeleton, before being absorbed or assimilation is absorbed into the host, become a part of the host organizations, and the matrix of surrounding cells have a good interaction, show that the interaction of coordination, and become an integral part of the cells and tissues of normal physiological function.
胶原的三重螺旋结构是其特有的活性构象 |
(Gly-X-Y)n重复序列
图片来源：P. Fratzl (ed.), Collagen: Structure and Mechanics, Springer (2008) |
Wound dressing
Collagen can promote cell growth. When keratinocytes were cultured on collagen, gelatin and hydrolyzed collagen substrate respectively, cell proliferation was observed by microscope after 16 hours of culture. The results showed that collagen significantly promoted the growth of keratinocytes and the cell density was significantly higher than that of other samples. Collagen can not only promote cell division and proliferation, but also maintain cell division function. Collagen can promote the differentiation of corneal epithelial cells, so it can be used as various wound repair dressings. Collagen dressings can take many forms, such as diaphragm, spongy and granular, which can redissolve, absorb the wound exudate, and interact with host extracellular matrix to promote the adhesion, movement, growth and deposition of cells on the new connective tissue. It can induce differentiation, induce chemotaxis of fibroblasts, delay wound contraction and accelerate wound repair. In the past, natural pig skin and amniotic membrane were used in burn treatment, but now more and more use recombinant form of collagen, which is purified into soluble collagen and prepared into a variety of dressings. In addition, fibroblast growth factor, platelet derivative growth factor and epidermal growth factor can be added to the matrix, which can effectively promote wound healing.
Collagen as a wound dressing has good adhesion, suitable for granulation tissue and epithelial cell formation, decrease the wound contracture and antigen reaction, hemostatic performance is good, has the similar leather physical form structure and the advantages of moderate crosslinking and absorbed by the organization, can be used in burns, wounds caused by external forces, the treatment of leg ulcers and bedsores. The main application areas are as follows:
② 止血材料
胶原亦可用于止血材料。血管内皮细胞下层由一系列的细胞外基质蛋白组成，其中的纤维胶原则是诱导血栓形成的主要基质蛋白。当血管壁的内皮细胞被剥离，致使胶原暴露于血液中，流动的血液中的血小板立刻与胶原纤维吸附在一起，发生凝集反应，从而形成血栓。胶原通过激活凝血因子Ⅻ、促进凝血因子Ⅺ和Ⅴ的活化，促进血小板的凝血作用。作为止血剂使用的胶原可以是粉状、片状及海绵状等多种物理形态。
胶原止血剂在治疗细胞组织器官如肝或脾的创伤止血方面效果更明显。由于这些组织一般缺乏结缔组织的支持，在出血、渗出、细胞表面的修复过程中，十分需要纤维蛋白构架维持稳定。在这种情况下，使用胶原基止血剂可减少这些组织中血液的流失。胶原蛋白海绵和微晶胶原止血剂已成功用于粘牙龈手术上颚等部位的止血，还可用于血管嫁接、局部肝切除，脑、胰、肝和肾伤口的止血。在应用中，胶原蛋白快速止血绷带能在10至15秒内止住动脉流血，在急救中特别有效，而常规情况下急救止血除向伤口施压以外没有更有效的方法，该等功效让此类产品作为急救材料，在紧急事件或军事医疗场景具有良好的应用前景。
③ 注射填充材料
去除端肽的未变性胶原具有低抗原性、良好的生物相容性和可生物降解性，有利于成纤维细胞的存活和生长，可用于面部轮廓矫正、皱纹和瘢痕修复等医学美容。注入体内的胶原对宿主细胞有诱导作用，使细胞向注射物内迁移，并进行正常生理代谢，合成自身胶原等细胞外间质成分。随着注射时间的延长，注射物中Ⅰ型和Ⅲ型胶原比例会逐渐接近正常皮肤中胶原的比例，表明成纤维细胞新合成了自身胶原。血液的供应保障了注射物内细胞生存所需的氧气和养分，使无生命的胶原注射物逐步变为有生命的组织。可溶性胶原溶液用于面部整形美容时须具备一定的可塑性和弹性，在常温是凝胶状液体，被注射到体内会转变为半固态的胶体。除了应用于矫正各种皮肤缺陷之外，还可用于食管括约肌、声带的修复、牙周疾病方面的治疗。
④ 药物载体
胶原在不同条件下（如温度、pH、离子强度、浓度、组成和其它高分子作用等），可以发生聚集或组装等行为而呈现多种形态，如水凝胶、膜、微球和微纤等。通过结合抗生素、蛋白类和基因等药物，可以构建出多样化的药物释放体系，从而达到不同的释放要求和治疗效果。另外，通过酶预处理或者化学交联等加工工艺还可实现对其降解行为的调控。早在20世纪80年代，胶原就被用于抗生素药物（如庆大霉素）的局部释放载体，且已实现了商业化。目前，常用的胶原药物载体系统主要包括眼科用的胶原罩、烧伤和创伤科用的胶原海绵、用于基因释放的微球以及透皮释放的控制材料等。
胶原蛋白医用产品的临床应用如下所示：
资料来源：维基百科、FDA、CFDA、中信证券整理
2) Application of collagen in skin care
The skin can be divided into three layers: the epidermis, dermis and subcutaneous tissue. Foreign countries reported the application of collagen in cosmetics in the 1980s, and some well-known brands also launched pure collagen and collagen added skin care products. Compared with hydrolyzed collagen and gelatin, collagen has a small amount of hydrophilic groups, but it has a unique triple helix structure, and its moisturizing effect is second only to glycerol. Natural collagen has the functions of promoting tissue growth and healing, retaining water, whitening and nourishing skin, etc. Therefore, natural collagen is widely used in skin care products and beauty industry.
The hydration of natural collagen is due to its large amount of glycine, hydroxyproline and hydroxylysine. Due to its large amount of hydroxyl, natural collagen has a good moisturizing effect. The triple helix structure of natural collagen can combine with water molecules to produce hydration. Hydroxyproline connects water molecules to form hydration skeleton structure, which enables part of water to bridge between amide group and carboxyl group in the form of mesenchymal water. Such hydration promotes the self-assembly of collagen. The hydroxyl group, carboxyl group and water on the -peptide chain form a lattice-like state. This hydrated structure not only enables collagen to bind more water, but also greatly improves the thermal stability of natural collagen. Collagen peptide and gelatin are hydrolyzed products of collagen, which are a kind of polypeptide mixture. There is no triple helix structure of natural collagen, so they cannot fix and bind too many water molecules. They can only rely on hydrogen bonds to bind a small number of water molecules, but cannot use their structure to bind multi-layer water molecules. Therefore, natural collagen has better biological nutrition functions of water lock, water supply and skin than small molecule collagen peptide.
胶原的水合作用示意图
Note :(a) bare collagen molecule; (b) Collagen binds to a layer of water molecules, relying on hydrogen bonds of hydroxyl and carboxyl groups outside the triple helix; (c) Collagen binds two layers of water molecules, hydrogen bonds form a layer, and the triple helix structure holds part of the water molecules; (c) The three-layer moisture substructure of collagen.
Note: Images from J. Bella, et al., Structure 3 (9):893-906 (1995)
Collagen has a balance between synthesis and degradation in the body. The small molecule collagen peptide diffuses passively into the skin's shallow layer structure, acting as a temporary moisturizer and nurse. The active gel principle with macromolecular structure plays a biological nursing role in the cuticle and dermis of skin by means of physical transdermal absorption and cell-mediated biological transdermal absorption. In terms of physical absorption, active collagen diffuses passively into the skin tissue through the cuticle space and the hair follicle wall space structure. In terms of bioabsorption, active collagen plays a role in cell adhesion, cell signaling and cell growth regulation through its recognition with integrins, discoid domain receptors, glycoproteins and other molecules on the tissue cell surface. These transmembrane protein receptors can recognize and bind specific amino acid sequences in the natural collagen triple helix structure, activate skin keratinocytes and hair follicle immune cells, and play a physiological regulatory role in human skin tissue, thus giving play to its skin nutrition function.
In addition, collagen can also inhibit the production of melanin to achieve the whitening effect. Tyrosine in the skin is catalyzed by copper-containing tyrosinase to produce dopamine, which is then rearranged or REDOX to form melanin-like substances. Collagen can bind to the active center of tyrosinase through tyrosine residue competition, reducing the generation of dopamine and inhibiting the formation of melanin. Or through chelate copper ion, make tyrosinase inactivation, reduce melanin deposition, thus achieve the role of whitening.
External stimuli, physical and chemical factors often cause skin barrier is damaged, the stimulating factors can lead to skin langerhans cells and cutin cell is activated, and allergic skin inflammation or Angle, according to the degree of severity may produce dry, desquamation, erythema, swelling, capillary expansion and other inflammatory response. Natural collagen has the functions of promoting tissue growth and healing, water retention, whitening, nourishing skin and so on. Therefore, it has been widely used in medical treatment and beauty industry in recent years. The main product types developed based on collagen can be summarized as topical collagen skincare stock, collagen facial mask, collagen cleanser and cream, collagen injection and filling beauty needle, etc.
Medical beauty scenarios, medical collagen (at present mainly animal source collagen) injected into the pitting skin defect after filling effect can support not only, also can induce the subject's own tissue reconstruction, generate new tissue will gradually achieving local corrective action together with the surrounding normal skin and plastic surgery injecting collagen in the dermis, can be used to eliminate the deeper wrinkles ruffle.
3) Application of collagen in food
The main application of collagen in the food field is artificial collagen-casings, which are similar to natural casings, with good air permeability and transparency, edible and good taste. Artificial collagen casings also have the advantages of uniform thickness, high tensile strength, strong stability and high amino acid content. Meanwhile, they can save space and cost during transportation and have obvious advantages compared with plastic casings and cellulose casings. Therefore, artificial collagen casings gradually occupy a dominant position in the casings market.
Hydrolyzed collagen can be directly eaten as functional food, such as chewable tablets, protein powder for athletes and enteral nutrition preparations. Animal collagen food has unique water retention, and can improve the taste of food, extend the shelf life and maintain color, mainly as meat, candy, dairy products and other food additives.
