Use of lactoferrin and compositions thereof in promoting gut development

Abstract

The application provides application of lactoferrin and a composition thereof in promoting intestinal development. The lactoferrin of the application can up-regulate the transcription level of a phospholipid globulin polysaccharide coding gene gpc1, a hyaluronic acid synthase 1 coding gene has1, a hyaluronic acid synthase 2 coding gene has2, a hyaluronic acid synthase 3 coding gene has3, an exosin glycosyltransferase 1 coding gene ext1 and an exosin glycosyltransferase 2 coding gene ext2, thereby promoting development of a protein skeleton of a glycocalyx layer, synthesis of hyaluronic acid, synthesis of heparan sulfate, further promoting development of the glycocalyx layer, and achieving the effects of perfecting an intestinal barrier, regulating intestinal flora and improving intestinal health.

Description

Technical Field

[0001] This invention relates to the field of biotechnology, and more particularly to the application of lactoferrin and its compositions in promoting intestinal development. Background Technology

[0002] In the physiological structure of the intestine, the glycocalyx is a specialized and intricate structure covering the surface of intestinal epithelial cells, primarily composed of glycans and proteins. A well-developed intestinal glycocalyx plays a crucial role in intestinal health, preventing pathogen adhesion and invasion, reducing the risk of intestinal infection, and providing specific binding sites for symbiotic microorganisms in the intestine. This helps maintain the balance and stability of the intestinal microbiota, which is essential for the normal functioning of the intestinal barrier and the homeostasis of the overall intestinal ecosystem. For example, during neonatal intestinal development, a well-developed glycocalyx provides binding sites for symbiotic microorganisms, prevents pathogen adhesion, and acts as a barrier against luminal toxins and enzymes.

[0003] Currently, although gut health has received considerable attention and numerous products on the market exist to improve gut function, such as probiotic preparations, prebiotic products, and some gut repair drugs, products specifically targeting the development of the intestinal epithelial glycocalyx are extremely scarce. Most existing gut health products indirectly affect gut health by regulating the quantity and types of gut microbiota, with unclear direct effects on this important structure. The few products that involve intestinal barrier repair mainly focus on strengthening the tight junctions between intestinal epithelial cells, lacking effective means to promote glycocalyx development. Therefore, developing a product that can effectively promote intestinal glycocalyx development is significantly necessary and has broad market prospects, playing a crucial role in improving gut health maintenance. Summary of the Invention

[0004] This invention provides an application of lactoferrin in promoting the development of the intestinal glycocalyx layer;

[0005] The promotion of intestinal glycocalyx development includes:

[0006] Upregulate the transcriptional level of gpc1, the gene encoding phosphoglobulin; and / or,

[0007] Upregulate the transcriptional level of the hyaluronic acid synthase 1 encoding gene has1; and / or,

[0008] Upregulate the transcriptional level of the hyaluronic acid synthase 2 encoding gene has2; and / or,

[0009] Upregulate the transcriptional level of the hyaluronic acid synthase 3 encoding gene has3; and / or,

[0010] Upregulate the transcriptional level of ext1, the gene encoding exochrome glycosyltransferase 1; and / or,

[0011] Upregulate the transcriptional level of ext2, the gene encoding exochrome glycosyltransferase 2; and / or,

[0012] Promotes the development of the protein backbone of the intestinal glycocalyx; and / or,

[0013] Promotes hyaluronic acid synthesis in the glycocalyx layer of the intestine; and / or,

[0014] Promotes the synthesis of heparan sulfate in the glycocalyx layer of the intestine.

[0015] In the applications described above, the concentration of lactoferrin used is 0.05-1.5 mg / mL.

[0016] This invention provides the application of a composition of lactoferrin and casein phosphopeptide in promoting the development of the intestinal glycocalyx, wherein the lactoferrin is the lactoferrin described above.

[0017] As described above, the application concentration of lactoferrin is 0.05-1.5 mg / mL; and / or,

[0018] The application concentration of casein phosphopeptide is 0.05-1.5 mg / mL.

[0019] In the application described above, the concentration ratio of lactoferrin to casein phosphopeptide is (1-10):1.

[0020] The applications described above, wherein promoting the development of the intestinal glycocalyx layer include:

[0021] Upregulate the transcriptional level of gpc1, the gene encoding phosphoglobulin; and / or,

[0022] Upregulate the transcriptional level of the hyaluronic acid synthase 1 encoding gene has1; and / or,

[0023] Upregulate the transcriptional level of the hyaluronic acid synthase 2 encoding gene has2; and / or,

[0024] Upregulate the transcriptional level of the hyaluronic acid synthase 3 encoding gene has3; and / or,

[0025] Upregulate the transcriptional level of ext1, the gene encoding exochrome glycosyltransferase 1; and / or,

[0026] Upregulate the transcriptional level of ext2, the gene encoding exochrome glycosyltransferase 2; and / or,

[0027] Promotes the development of the protein backbone of the intestinal glycocalyx; and / or,

[0028] Promotes hyaluronic acid synthesis in the glycocalyx layer of the intestine; and / or,

[0029] Promotes the synthesis of heparan sulfate in the glycocalyx layer of the intestine.

[0030] This invention provides the use of a composition in the preparation of products related to promoting the development of the intestinal glycocalyx, wherein the composition comprises lactoferrin and optionally casein phosphopeptide.

[0031] As described above, the product includes at least one of health supplements and pharmaceuticals;

[0032] Health products include at least one of the following: health foods, functional foods, functional beverages, functional powders, functional capsules, and functional powders, which improve the intestinal barrier, regulate intestinal flora, or improve intestinal health.

[0033] The drugs include at least one of the following: prevention or treatment of gastrointestinal infections caused by glycocalyx damage; prevention or treatment of inflammatory bowel disease caused by glycocalyx damage; prevention or treatment of intestinal barrier impairment caused by glycocalyx damage; and prevention or treatment of intestinal flora imbalance caused by glycocalyx damage.

[0034] In the applications described above, the product also includes a carrier and / or physiologically acceptable excipients;

[0035] The carrier includes at least one of microcapsules, microspheres, nanoparticles, and liposomes;

[0036] Physiologically acceptable excipients include at least one of the following: fillers, flavoring agents, diluents, wetting agents, dispersants, binders, disintegrants, lubricants, color, flavor and aroma modifiers, solvents, solubilizers, cosolvents, emulsifiers, antioxidants, metal complexing agents, inert gases, preservatives, local analgesics, pH adjusters, and isotonic or isotropic modifiers.

[0037] Through long-term research and extensive experiments, this invention has discovered that lactoferrin, when used in the body, can upregulate the transcriptional level of the gene encoding phosphoglobulin gpc1, thereby promoting the development of the glycocalyx protein backbone; it can upregulate the transcriptional levels of the genes encoding hyaluronic acid synthase 1 (has1), hyaluronic acid synthase 2 (has2), and hyaluronic acid synthase 3 (has3), thereby promoting the synthesis of hyaluronic acid in the glycocalyx; and it can also upregulate the transcriptional levels of the genes encoding exochrome glycosyltransferase 1 (ext1) and exochrome glycosyltransferase 2 (ext2), thereby promoting the synthesis of heparan sulfate in the glycocalyx. Therefore, lactoferrin can promote the development of the glycocalyx by promoting the production of important components in the intestinal glycocalyx (glycocalyx protein backbone, hyaluronic acid, and heparan sulfate), and further improve the intestinal barrier, regulate the intestinal flora, and enhance intestinal health. The novel applications of lactoferrin provided by this invention not only fill existing market gaps and overcome the limitations of existing products, but also offer new approaches to regulating gut microbiota, improving gut health, preventing or treating gastrointestinal infections caused by glycocalyx damage, preventing or treating inflammatory bowel disease caused by glycocalyx damage, preventing or treating intestinal barrier damage caused by glycocalyx damage, and preventing or treating gut microbiota dysbiosis caused by glycocalyx damage. This invention has significant necessity and broad market prospects, and is of great importance for improving gut health maintenance. Attached Figure Description

[0038] Figure 1 This is a diagram showing the relative transcriptional level of the gpc1 gene in one embodiment of the present invention;

[0039] Figure 2 This is a diagram showing the relative transcriptional level of the has1 gene in one embodiment of the present invention;

[0040] Figure 3 This is a diagram showing the relative transcriptional level of the has2 gene in one embodiment of the present invention;

[0041] Figure 4 This is a diagram showing the relative transcriptional level of the has3 gene in one embodiment of the present invention;

[0042] Figure 5 This is a diagram showing the relative transcriptional level of the ext1 gene in one embodiment of the present invention;

[0043] Figure 6 This is a diagram showing the relative transcriptional level of the ext2 gene in one embodiment of the present invention;

[0044] Figure 7 This is a diagram showing the relative transcriptional levels of the lactoferrin and casein phosphopeptide group gpc1 gene in one embodiment of the present invention.

[0045] Figure 8This is a diagram showing the relative transcriptional levels of the has1 gene in the lactoferrin and casein phosphopeptide groups in one embodiment of the present invention.

[0046] Figure 9 This is a diagram showing the relative transcriptional levels of the has2 gene in the lactoferrin and casein phosphopeptide groups in one embodiment of the present invention.

[0047] Figure 10 This is a diagram showing the relative transcriptional levels of the has3 gene in the lactoferrin and casein phosphopeptide groups in one embodiment of the present invention.

[0048] Figure 11 This is a diagram showing the relative transcriptional levels of the lactoferrin and casein phosphopeptide ext1 gene in one embodiment of the present invention.

[0049] Figure 12 This is a diagram showing the relative transcriptional levels of the lactoferrin and casein phosphopeptide ext2 genes in one embodiment of the present invention.

[0050] Figures 1-12 In the text, A, AB, B, BC, and C are letter markers for significance analysis. The same letter indicates no significant difference, while different letters indicate a significant difference. The letters A, AB, B, BC, and CA represent the means from highest to lowest. Detailed Implementation

[0051] To enable those skilled in the art to better understand the present invention, the present invention will be further described in detail below. The specific embodiments listed below are merely descriptions of the principles and features of the present invention, and the examples are only for explaining the present invention and are not intended to limit the scope of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0052] The intestinal glycocalyx is a complex network structure covering the surface of intestinal epithelial cells. It serves as a core hub for the intestinal physical barrier and functional regulation, primarily composed of mucins, proteoglycans, glycoproteins, glycolipids, and free polysaccharides assembled through covalent or non-covalent interactions to form a dynamic functional layer rich in carbohydrates. Among these, phosphoglobulin 1 (Glypican 1) is a key membrane-bound proteoglycan in the glycocalyx. Its core protein is anchored to the intestinal epithelial cell membrane surface via glycosylphosphatidylinositol (GPI), and covalently linked to glycosaminoglycan chains such as heparan sulfate. As a protein backbone component of the glycocalyx, Glypican 1 not only maintains the spatial stability of the glycocalyx through its own structure, but its heparan sulfate chains also bind water, ions, and small molecule nutrients through charge interactions, participating in the osmotic regulation and substance exchange functions of the glycocalyx. Heparan sulfate (HS) is a linear sulfated polysaccharide. Besides being a glycan component of phosphoglobulin-1, it can also exist in the glycocalyx layer by binding with other proteoglycans. Its highly sulfated structure allows it to specifically recognize and bind to adhesion proteins of intestinal pathogens (such as E. coli and rotavirus), preventing direct contact between pathogens and intestinal epithelial cells. Simultaneously, it can regulate the recruitment and activation of intestinal immune cells (such as macrophages and dendritic cells), making it a crucial executor of the glycocalyx layer's immune defense function. Hyaluronic acid (HA) is another important glycosaminoglycan. Unlike HS, it usually exists in the glycocalyx layer in free form or bound to hyaluronic acid-binding proteins. It is highly hydrophilic and can increase the thickness and elasticity of the glycocalyx layer by absorbing water and swelling, enhancing its buffering capacity against mechanical damage (such as intestinal peristalsis friction). HA can also regulate cell proliferation and differentiation by binding to CD44 receptors on the surface of intestinal epithelial cells, promoting the repair and renewal of the glycocalyx layer.

[0053] Based on the above research, in order to develop a product that can effectively promote the development of the intestinal glycocalyx, this invention, through long-term research and extensive experiments, has discovered that lactoferrin (LF) can upregulate the transcriptional level of the gene encoding phospholipoglobulin gpc1, thereby promoting the development of the glycocalyx protein backbone; lactoferrin can upregulate the transcriptional levels of the genes encoding hyaluronic acid synthase 1 (has1), hyaluronic acid synthase 2 (has2), and hyaluronic acid synthase 3 (has3), thereby promoting the synthesis of hyaluronic acid in the glycocalyx; lactoferrin can also upregulate the transcriptional levels of the genes encoding exochrome glycosyltransferase 1 (ext1) and exochrome glycosyltransferase 2 (ext2), thereby promoting the synthesis of heparan sulfate in the glycocalyx. In short, lactoferrin can promote the development of the intestinal glycocalyx by promoting the development of important components (glycocalyx protein backbone, hyaluronic acid, and heparan sulfate), and further achieve the effects of improving the intestinal barrier, regulating the intestinal flora, and improving intestinal health.

[0054] Therefore, the first aspect of the present invention provides the application of lactoferrin in promoting the development of the intestinal glycocalyx layer.

[0055] The novel applications of lactoferrin provided by this invention not only fill existing market gaps and overcome the limitations of existing products, but also offer new approaches to regulating gut microbiota, improving gut health, preventing or treating gastrointestinal infections caused by glycocalyx damage, preventing or treating inflammatory bowel disease caused by glycocalyx damage, preventing or treating intestinal barrier damage caused by glycocalyx damage, and preventing or treating gut microbiota dysbiosis caused by glycocalyx damage. This invention has significant necessity and broad market prospects, and is of great importance for improving gut health maintenance.

[0056] In the above technical solution, when the application concentration of lactoferrin is 0.05-1.5 mg / mL, it can better promote the development of the glycocalyx layer. For example, the application concentration of lactoferrin can be 0.05 mg / mL, 0.15 mg / mL, 0.25 mg / mL, 0.35 mg / mL, 0.45 mg / mL, 0.55 mg / mL, 0.65 mg / mL, 0.75 mg / mL, 0.85 mg / mL, 0.95 mg / mL, 1.00 mg / mL, 1.10 mg / mL, 1.20 mg / mL, 1.30 mg / mL, 1.40 mg / mL, 1.50 mg / mL, or any value between any two of the above ranges.

[0057] In the above technical solution, promoting the development of the intestinal glycocalyx layer includes at least one of the following: upregulating the transcription level of the gene gpc1 encoding phosphoglobulin glycan, upregulating the transcription level of the gene has1 encoding hyaluronic acid synthase 1, upregulating the transcription level of the gene has2 encoding hyaluronic acid synthase 2, upregulating the transcription level of the gene has3 encoding hyaluronic acid synthase 3, upregulating the transcription level of the gene ext1 encoding exoprotein glycosyltransferase 1, and upregulating the transcription level of the gene ext2 encoding exoprotein glycosyltransferase 2.

[0058] Experimental results showed that intervention with lactoferrin could upregulate the transcriptional levels of the following genes in human colon adenocarcinoma cells Caco-2: gpc1 (encoding phosphoglobulin glycan), has1 (encoding hyaluronic acid synthase 1), has2 (encoding hyaluronic acid synthase 2), has3 (encoding hyaluronic acid synthase 3), ext1 (encoding exochrome glycosyltransferase 1), and ext2 (encoding exochrome glycosyltransferase 2), thereby promoting the development of the intestinal glycocalyx.

[0059] Understandably, lactoferrin can be used to promote the development of the intestinal glycocalyx layer, and can also be used to prepare agonists that promote the transcription of the gene encoding phosphoglobulin gpc1, the gene encoding hyaluronic acid synthase 1 has1, the gene encoding hyaluronic acid synthase 2 has2, the gene encoding hyaluronic acid synthase 3 has3, the gene encoding exochrome glycosyltransferase 1 ext1, and the gene encoding exochrome glycosyltransferase 2 ext2.

[0060] In the above technical solutions, promoting the development of the intestinal glycocalyx layer also includes at least one of the following: promoting the development of the protein backbone of the intestinal glycocalyx layer, promoting the synthesis of hyaluronic acid in the intestinal glycocalyx layer, and promoting the synthesis of heparan sulfate in the intestinal glycocalyx layer.

[0061] Experiments have verified that lactoferrin intervention can upregulate the transcriptional level of the gene encoding phospholipoglobulin 1 (gpc1) in human colon adenocarcinoma cells Caco-2, thereby promoting the development of the intestinal glycocalyx protein backbone and thus the development of the intestinal glycocalyx. Lactoferrin intervention can also upregulate the transcriptional levels of the genes encoding hyaluronic acid synthase 1 (has1), hyaluronic acid synthase 2 (has2), and hyaluronic acid synthase 3 (has3) in human colon adenocarcinoma cells Caco-2, thereby promoting the synthesis of hyaluronic acid in the intestinal glycocalyx and thus promoting the development of the intestinal glycocalyx. Furthermore, lactoferrin intervention can upregulate the transcriptional levels of the genes encoding exokinin glycosyltransferase 1 (ext1) and exokinin glycosyltransferase 2 (ext2) in human colon adenocarcinoma cells Caco-2, thereby promoting the synthesis of heparan sulfate in the intestinal glycocalyx and thus promoting the development of the intestinal glycocalyx.

[0062] Understandably, lactoferrin can be used in the preparation of products that promote the development of the intestinal glycocalyx, as well as in the preparation of agonists that promote the development of the protein backbone of the intestinal glycocalyx, agonists that promote the synthesis of hyaluronic acid in the intestinal glycocalyx, and agonists that promote the synthesis of heparan sulfate in the intestinal glycocalyx.

[0063] Based on the above research, the present invention also found that the simultaneous use of lactoferrin and casein phosphopeptide has a better effect on promoting the development of the intestinal glycocalyx. Therefore, the second aspect of the present invention provides the application of a composition of lactoferrin and casein phosphopeptide in promoting the development of the intestinal glycocalyx, wherein the lactoferrin is the lactoferrin described above.

[0064] Experiments have shown that lactoferrin and casein phosphopeptide (CPP) have a synergistic effect. When used simultaneously, they significantly upregulate the transcription level of the gene encoding phosphoglobulin glucan, gpc1, thereby promoting the development of the glycocalyx protein backbone. Furthermore, they upregulate the transcription levels of the genes encoding hyaluronic acid synthase 1 (has1), hyaluronic acid synthase 2 (has2), and hyaluronic acid synthase 3 (has3), thus promoting hyaluronic acid synthesis in the glycocalyx. Simultaneous use also significantly upregulates the transcription levels of the genes encoding exochrome glycosyltransferase 1 (ext1) and exochrome glycosyltransferase 2 (ext2), thereby promoting heparan sulfate synthesis in the glycocalyx. The combined use of lactoferrin and casein phosphopeptide can further promote the development of the glycocalyx by promoting the production of key components (glycocalyx protein backbone, hyaluronic acid, and heparan sulfate), effectively strengthening the intestinal barrier, regulating gut microbiota, and improving intestinal health.

[0065] In the above technical solution, when the application concentration of lactoferrin is 0.05-1.5 mg / mL, it can better promote the development of the glycocalyx layer. For example, the application concentration of lactoferrin can be 0.05 mg / mL, 0.15 mg / mL, 0.25 mg / mL, 0.35 mg / mL, 0.45 mg / mL, 0.55 mg / mL, 0.65 mg / mL, 0.75 mg / mL, 0.85 mg / mL, 0.95 mg / mL, 1.00 mg / mL, 1.10 mg / mL, 1.20 mg / mL, 1.30 mg / mL, 1.40 mg / mL, 1.50 mg / mL, or any value between any two of the above ranges.

[0066] In the above technical solution, when the application concentration of casein phosphopeptide is 0.05-1.5 mg / mL, it can better match lactoferrin and synergistically promote the development of the glycocalyx layer. For example, the application concentration of casein phosphopeptide can be 0.05 mg / mL, 0.15 mg / mL, 0.25 mg / mL, 0.35 mg / mL, 0.45 mg / mL, 0.55 mg / mL, 0.65 mg / mL, 0.75 mg / mL, 0.85 mg / mL, 0.95 mg / mL, 1.00 mg / mL, 1.10 mg / mL, 1.20 mg / mL, 1.30 mg / mL, 1.40 mg / mL, 1.50 mg / mL, or any value between any two of the above ranges.

[0067] In the above technical solution, the concentration ratio of lactoferrin to casein phosphopeptide is (1-10):1, which can better exert the synergistic effect of lactoferrin and casein phosphopeptide, thereby further promoting the development of the glycocalyx layer. For example, the concentration ratio of lactoferrin to casein phosphopeptide can be 1:1, 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, etc.

[0068] In the above technical solution, promoting the development of the intestinal glycocalyx layer includes at least one of the following: upregulating the transcription level of the gene gpc1 encoding phosphoglobulin glycan, upregulating the transcription level of the gene has1 encoding hyaluronic acid synthase 1, upregulating the transcription level of the gene has2 encoding hyaluronic acid synthase 2, upregulating the transcription level of the gene has3 encoding hyaluronic acid synthase 3, upregulating the transcription level of the gene ext1 encoding exoprotein glycosyltransferase 1, and upregulating the transcription level of the gene ext2 encoding exoprotein glycosyltransferase 2.

[0069] Experiments have shown that the combined intervention of lactoferrin and casein phosphopeptide can upregulate the transcriptional levels of the following genes in human colon adenocarcinoma cells Caco-2: gpc1 (encoding phosphoglobulin glycan), has1 (encoding hyaluronic acid synthase 1), has2 (encoding hyaluronic acid synthase 2), has3 (encoding hyaluronic acid synthase 3), ext1 (encoding exochrome glycosyltransferase 1), and ext2 (encoding exochrome glycosyltransferase 2), thereby promoting the development of the intestinal glycocalyx.

[0070] Understandably, the combination of lactoferrin and casein phosphopeptide can be used to promote the development of the intestinal glycocalyx layer, and can also be used to prepare agonists that promote the transcription of the gene encoding phosphoglobulin gpc1, the gene encoding hyaluronic acid synthase 1 has1, the gene encoding hyaluronic acid synthase 2 has2, the gene encoding hyaluronic acid synthase 3 has3, the gene encoding exoprotein glycosyltransferase 1 ext1, and the gene encoding exoprotein glycosyltransferase 2 ext2.

[0071] In the above technical solutions, promoting the development of the intestinal glycocalyx layer also includes at least one of the following: promoting the development of the protein backbone of the intestinal glycocalyx layer, promoting the synthesis of hyaluronic acid in the intestinal glycocalyx layer, and promoting the synthesis of heparan sulfate in the intestinal glycocalyx layer.

[0072] Experiments have verified that the combined intervention of lactoferrin and casein phosphopeptide can upregulate the transcriptional level of the gene encoding phospholipoglobulin g1 in human colon adenocarcinoma cells Caco-2, thereby promoting the development of the intestinal glycocalyx protein backbone and thus the development of the intestinal glycocalyx. The combined intervention of lactoferrin and casein phosphopeptide can also upregulate the transcriptional levels of the genes encoding hyaluronic acid synthase 1 (has1), hyaluronic acid synthase 2 (has2), and hyaluronic acid synthase 3 (has3) in human colon adenocarcinoma cells Caco-2, thereby promoting the synthesis of hyaluronic acid in the intestinal glycocalyx and thus promoting the development of the intestinal glycocalyx. Furthermore, the combined intervention of lactoferrin and casein phosphopeptide can upregulate the transcriptional levels of the genes encoding exokinin glycosyltransferase 1 (ext1) and exokinin glycosyltransferase 2 (ext2) in human colon adenocarcinoma cells Caco-2, thereby promoting the synthesis of heparan sulfate in the intestinal glycocalyx and thus promoting the development of the intestinal glycocalyx.

[0073] Understandably, the combination of lactoferrin and casein phosphopeptide can be used to promote the development of the intestinal glycocalyx, and can also be used to prepare agonists that promote the development of the protein backbone of the intestinal glycocalyx, agonists that promote the synthesis of hyaluronic acid in the intestinal glycocalyx, and agonists that promote the synthesis of heparan sulfate in the intestinal glycocalyx.

[0074] A third aspect of the present invention provides the use of a composition in the preparation of products related to promoting the development of the intestinal glycocalyx, wherein the composition comprises lactoferrin and optionally casein phosphopeptide.

[0075] In the above technical solutions, the product includes at least one of health products and medicines.

[0076] Furthermore, health supplements include at least one of the following: functional beverages, functional powders, functional capsules, and functional powders, which enhance the intestinal barrier, regulate gut microbiota, or improve gut health.

[0077] Furthermore, the drug includes at least one of the following: prevention or treatment of gastrointestinal infection caused by glycocalyx damage; prevention or treatment of inflammatory bowel disease caused by glycocalyx damage; prevention or treatment of intestinal barrier impairment caused by glycocalyx damage; and prevention or treatment of intestinal flora imbalance caused by glycocalyx damage.

[0078] The aforementioned products also include carriers and / or physiologically acceptable excipients.

[0079] The carrier includes at least one of microcapsules, microspheres, nanoparticles, and liposomes.

[0080] Physiologically acceptable excipients include at least one of the following: fillers, flavoring agents, diluents, wetting agents, dispersants, binders, disintegrants, lubricants, color, flavor and aroma modifiers, solvents, solubilizers, cosolvents, emulsifiers, antioxidants, metal complexing agents, inert gases, preservatives, local analgesics, pH adjusters, and isotonic or isotropic modifiers.

[0081] In actual use, the above products can be administered via at least one of the following methods: oral administration, intravenous injection, intraperitoneal injection, intramuscular injection, subcutaneous injection, sublingual administration, nasal administration, and transdermal administration.

[0082] The technical solution of this application will be further explained below with reference to specific embodiments. Experimental methods without specific conditions are generally performed under conventional conditions or as recommended by the manufacturer. Unless otherwise specified, all reagents used are commercially available or obtained through public channels.

[0083] In the following examples, the culture conditions for human colon adenocarcinoma cells Caco-2 were as follows: Caco-2 cells were cultured using MEM complete medium (containing 20% ​​fetal bovine serum) produced by Nanjing Senbeijia Biotechnology Co., Ltd., and seeded in 25cm... 2 The cell culture flasks were placed in a carbon dioxide incubator at 37°C, 5% carbon dioxide, and 95% relative humidity. The culture medium was changed every 1-2 days. When the cell confluence reached 80%-90%, the cells were digested with 0.25% trypsin digestion solution and passaged at a volume ratio of 1:3.

[0084] Example 1: Experimental Grouping

[0085] The density of Caco-2 cells was adjusted to 1×10⁻⁶. 5 pcs / cm 2Caco-2 cells were seeded in 24-well plates and grouped and treated according to the groups and intervention substances in Table 1. Specifically, the control group received no intervention and was cultured in MEM complete medium for 36 h; the low-dose lactoferrin group (0.084 mg / mL) was treated by adding lactoferrin to MEM complete medium to a final concentration of 0.084 mg / mL and culturing for 36 h; the medium-dose lactoferrin group (0.294 mg / mL) was treated by adding lactoferrin to MEM complete medium to a final concentration of 0.294 mg / mL and culturing for 36 h; the high-dose lactoferrin group (1.029 mg / mL) was treated by adding lactoferrin to MEM complete medium to a final concentration of 1.029 mg / mL and culturing for 36 h; and the lactoferrin and casein phosphopeptides (CPP) group was treated by adding lactoferrin to MEM complete medium to a final concentration of 1.029 mg / mL and casein phosphopeptides to 0.1029 mg / mL and culturing for 36 h.

[0086] Table 1

[0087]

[0088] Example 2: Real-time quantitative PCR (qPCR) experiment

[0089] Total RNA was extracted and reverse transcribed from cells cultured for 36 h in Example 1. The relative transcription levels of genes related to glycocalyx development in Caco-2 cells were detected by qPCR. The qPCR primers are shown in Table 2. Genes related to glycocalyx development include: glypican 1 (gpc1), hyaluronan synthase 1 (has1), hyaluronan synthase 2 (has2), hyaluronan synthase 3 (has3), exostosin glycosyltransferase 1 (ext1), and exostosin glycosyltransferase 2 (ext2).

[0090] Table 2

[0091]

[0092] Example 3: Effect of lactoferrin on the protein backbone of the glycocalyx layer

[0093] The glypican 1 (gpc1) gene is the gene encoding the proteoglycan GPC1. GPC1 is the protein backbone of the glycocalyx and an important carrier of glycosaminoglycan chains, including heparan sulfate (HS) and hyaluronic acid (HA).

[0094] The relative transcription level of the gpc1 gene obtained by qPCR in Example 2 is shown. Figure 1 . Figure 1 The results showed that the gpc1 gene transcription level in the low-dose lactoferrin group (0.084 mg / mL) was slightly higher than that in the control group, while the gpc1 gene transcription levels in the medium-dose lactoferrin group (0.294 mg / mL) and the high-dose lactoferrin group (1.029 mg / mL) were significantly higher than those in the control group (p<0.05). These results indicate that lactoferrin can upregulate gpc1 transcription levels, thereby promoting the development of the glycocalyx protein backbone and thus promoting the development of the glycocalyx layer.

[0095] Example 4: Effect of lactoferrin on hyaluronic acid synthase in the glycocalyx layer

[0096] The genes encoding hyaluronan synthase 1 (HAS1), hyaluronan synthase 2 (HAS2), and hyaluronan synthase 3 (HAS3) are the coding genes for hyaluronan synthase (HAS). HAS is crucial for the synthesis of hyaluronan (HA), a major glycosaminoglycan component of the glycocalyx and a highly viscous component of the intestinal mucus layer, responsible for tissue repair, stability, and anti-inflammatory effects. Compared to HAS1 and HAS2, HAS3 can synthesize low molecular weight HA, which is essential for the development of intestinal stem cells; therefore, the presence of HAS3 is particularly important.

[0097] The relative transcription level of the has1 gene obtained by qPCR in Example 2 is shown. Figure 2 The relative transcription level of the has2 gene is visible. Figure 3 The relative transcription level of the has3 gene is visible. Figure 4 . Figure 2 The results showed that the has1 gene transcription level in the low-dose lactoferrin group (0.084 mg / mL) was higher than that in the control group, while the has1 gene transcription level in the medium-dose lactoferrin group (0.294 mg / mL) and the high-dose lactoferrin group (1.029 mg / mL) was significantly higher than that in the control group (p<0.05). Figure 3The results showed that the has2 gene transcription level in the low-dose lactoferrin group (0.084 mg / mL) was slightly higher than that in the control group, the has2 gene transcription level in the medium-dose lactoferrin group (0.294 mg / mL) was higher than that in the control group, and the has2 gene transcription level in the high-dose lactoferrin group (1.029 mg / mL) was significantly higher than that in the control group (p<0.05). Figure 4 The results showed that the transcriptional levels of the has3 gene in the low-dose (0.084 mg / mL), medium-dose (0.294 mg / mL), and high-dose (1.029 mg / mL) lactoferrin groups were significantly higher than those in the control group (p<0.05). These results indicate that lactoferrin can upregulate the transcriptional levels of has1, has2, and has3, thereby promoting HA synthesis in the glycocalyx layer and thus promoting its development.

[0098] Example 5: Effect of lactoferrin on heparan sulfate in the glycocalyx layer

[0099] The exostosin glycosyltransferase 1 (ext1) and exostosin glycosyltransferase 2 (ext2) genes are involved in the elongation of the heparan sulfate (HS) chain and are responsible for integrating the HS chain with nucleotide sugars in the Golgi apparatus. HS is the main glycosaminoglycan component in the glycocalyx, which supports organogenesis, growth factor signaling, and bacterial adhesion.

[0100] The relative transcription level of the ext1 gene obtained by qPCR in Example 2 is shown. Figure 5 The relative transcription level of the ext2 gene is visible. Figure 6 . Figure 5 The results showed that the ext1 gene transcription level in the low-dose lactoferrin group (0.084 mg / mL) was slightly higher than that in the control group, while the ext1 gene transcription level in the medium-dose lactoferrin group (0.294 mg / mL) and the high-dose lactoferrin group (1.029 mg / mL) was significantly higher than that in the control group (p<0.05). Figure 6The results showed that the transcriptional levels of the ext2 gene in the medium-dose (0.294 mg / mL) and high-dose (1.029 mg / mL) lactoferrin groups were slightly higher than those in the control group, while the transcriptional level of the ext2 gene in the low-dose (0.084 mg / mL) group was significantly higher than that in the control group. These results indicate that lactoferrin can upregulate the transcriptional levels of ext1 and ext2, thereby promoting the synthesis of HS in the glycocalyx layer and thus promoting the development of the glycocalyx layer.

[0101] Example 6: Effects of lactoferrin and casein phosphopeptides on the protein backbone of the glycocalyx layer

[0102] The relative transcription levels of the gpc1 gene in the lactoferrin and casein phosphopeptide group (cpp+lactoferrin) obtained by qPCR in Example 2 are shown. Figure 7 .

[0103] Figure 7 The results showed that the relative transcriptional level of Gpc1 in the lactoferrin and casein phosphopeptide group (CPP + lactoferrin) was significantly upregulated compared with the control group and the high-dose lactoferrin group (1.029 mg / mL) (p<0.05). These results indicate that the combination of lactoferrin and casein phosphopeptide can significantly upregulate Gpc1 transcriptional levels, further promoting the development of the glycocalyx protein backbone, thereby promoting glycocalyx development; and that lactoferrin and casein phosphopeptide have a synergistic effect in promoting glycocalyx development.

[0104] Example 7: Effects of lactoferrin and casein phosphopeptides on hyaluronic acid synthase in the glycocalyx layer

[0105] The relative transcription levels of the lactoferrin and casein phosphopeptide group (cpp+lactoferrin) has1 gene obtained by qPCR in Example 2 are shown. Figure 8 The relative transcription level of the has2 gene is visible. Figure 9 The relative transcription level of the has3 gene is visible. Figure 10 .

[0106] Figure 8 The results showed that the relative transcriptional level of has1 in the lactoferrin and casein phosphopeptide group (cpp+lactoferrin) was significantly upregulated compared with the blank control group (p<0.05), but there was no significant difference compared with the high-dose lactoferrin group (1.029 mg / mL). Figure 9The results showed that the relative transcriptional level of has2 in the lactoferrin and casein phosphopeptide group (cpp+lactoferrin) was significantly upregulated compared with the blank control group (p<0.05), but there was no significant difference compared with the high-dose lactoferrin group (1.029 mg / mL). Figure 10 The results showed that the relative transcriptional level of has3 in the lactoferrin and casein phosphopeptide group (cpp + lactoferrin) was significantly upregulated compared with the control group (p<0.05), but there was no significant difference compared with the high-dose lactoferrin group (1.029 mg / mL). These results indicate that the combination of lactoferrin and casein phosphopeptide can upregulate the transcriptional levels of has1, has2, and has3, thereby promoting HA synthesis in the glycocalyx and thus promoting glycocalyx development; and its promoting effect is similar to that of the high-dose lactoferrin group alone.

[0107] Example 8: Effects of lactoferrin and casein phosphopeptides on heparan sulfate in glycocalyx layer

[0108] The relative transcription levels of the lactoferrin and casein phosphopeptide (cpp+lactoferrin) ext1 gene obtained by qPCR in Example 2 are shown. Figure 11 The relative transcription level of the ext2 gene is visible. Figure 12 .

[0109] Figure 11 The results showed that the relative transcriptional level of ext1 in the lactoferrin and casein phosphopeptide group (cpp+lactoferrin) was significantly upregulated compared with the blank control group (control) and the high-dose lactoferrin group (1.029 mg / mL) (p<0.05). Figure 12 The results showed that the relative transcriptional level of ext2 in the lactoferrin and casein phosphopeptide group (cpp + lactoferrin) was not significantly different from that in the control group and the high-dose lactoferrin group (1.029 mg / mL). These results indicate that the combination of lactoferrin and casein phosphopeptide can upregulate the transcriptional level of ext1, thereby promoting the synthesis of HS in the glycocalyx and thus promoting the development of the glycocalyx.

[0110] In summary, co-culturing Caco-2 cells with lactoferrin before they adhere to the culture vessel can simulate the state of intestinal growth and maturation. Under this cellular state, lactoferrin significantly promotes the development of the glycocalyx layer in Caco-2 cells. Furthermore, lactoferrin can be used in combination with casein phosphopeptides to synergistically promote the development of the glycocalyx layer skeleton and the synthesis of heparan sulfate. These results indicate that supplementing with specific doses of lactoferrin or a combination of lactoferrin and casein phosphopeptides in specific proportions during the intestinal maturation stage can significantly promote the development of the intestinal glycocalyx layer, thereby promoting overall healthy intestinal development.

[0111] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. Application of lactoferrin in the preparation of health products that promote the development of the intestinal glycocalyx layer; The promotion of intestinal glycocalyx development includes: Upregulates the transcriptional level of gpc1, the gene encoding phosphoglobulin glycan; And / or, Upregulate the transcriptional level of the hyaluronic acid synthase 1 encoding gene has1; and / or, Upregulate the transcriptional level of the hyaluronic acid synthase 2 encoding gene has2; and / or, Upregulate the transcriptional level of the hyaluronic acid synthase 3 encoding gene has3; and / or, Upregulate the transcriptional level of ext1, the gene encoding exochrome glycosyltransferase 1; and / or, Upregulate the transcriptional level of ext2, the gene encoding exochrome glycosyltransferase 2; and / or, Promotes the development of the protein backbone of the intestinal glycocalyx; and / or, Promotes hyaluronic acid synthesis in the glycocalyx layer of the intestine; and / or, Promotes the synthesis of heparan sulfate in the glycocalyx layer of the intestine.

2. The application according to claim 1, characterized in that, The application concentration of the lactoferrin is 0.05-1.5 mg / mL.

3. Application of the combination of lactoferrin and casein phosphopeptide in the preparation of health products related to promoting the development of the intestinal glycocalyx layer.

4. The application according to claim 3, characterized in that, The application concentration of the lactoferrin is 0.05-1.5 mg / mL; and / or, The application concentration of the casein phosphopeptide is 0.05-1.5 mg / mL.

5. The application according to claim 3 or 4, characterized in that, The concentration ratio of lactoferrin to casein phosphopeptide is (1-10):

1.

6. The application according to claim 3 or 4, characterized in that, The promotion of intestinal glycocalyx development includes: Upregulate the transcriptional level of gpc1, the gene encoding phosphoglobulin; and / or, Upregulate the transcriptional level of the hyaluronic acid synthase 1 encoding gene has1; and / or, Upregulate the transcriptional level of the hyaluronic acid synthase 2 encoding gene has2; and / or, Upregulate the transcriptional level of the hyaluronic acid synthase 3 encoding gene has3; and / or, Upregulate the transcriptional level of ext1, the gene encoding exochrome glycosyltransferase 1; and / or, Upregulate the transcriptional level of ext2, the gene encoding exochrome glycosyltransferase 2; and / or, Promotes the development of the protein backbone of the intestinal glycocalyx; and / or, Promotes hyaluronic acid synthesis in the glycocalyx layer of the intestine; and / or, Promotes the synthesis of heparan sulfate in the glycocalyx layer of the intestine.

7. The application according to claim 1, characterized in that, The health products include at least one of the functional foods that improve the intestinal barrier, regulate the intestinal flora, or improve intestinal health.

8. The application according to claim 7, characterized in that, The health products also include carriers and / or physiologically acceptable excipients; The carrier includes at least one of microcapsules, microspheres, nanoparticles, and liposomes; Physiologically acceptable excipients include at least one of the following: fillers, diluents, wetting agents, dispersants, binders, disintegrants, lubricants, flavor and color modifiers, solvents, solubilizers, cosolvents, emulsifiers, antioxidants, metal complexing agents, inert gases, preservatives, pH adjusters, and isotonic or isotropic modifiers.

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