Compound composition for inducing cell migration and application

Abstract

The invention belongs to the technical field of biology, and particularly relates to a compound composition for inducing cell migration by controlling the curvature of a membrane and application. The preparation method comprises the following steps: by taking hemagglutinin protein as a membrane protein aggregation unit, connecting sialic acid lactose and rhodamine B in an acylhydrazone bond manner to form a binding and driving unit (R-SL small molecule induction ligand), so as to form self-assembled molecular tweezers (ATS); the preparation process is simple, and the equipment and labor cost is reduced. The constructed self-assembled molecular tweezers realize the polymerization / depolymerization process of membrane protein through a simple chemical regulation method, so that the membrane curvature is efficiently regulated; cell migration is obviously induced, and wound healing is promoted. The cell level test verifies that when the self-assembly molecular tweezers are used for adjusting the curvature of cells, the self-assembly molecular tweezers can promote the cells to generate pseudopods and induce the neogenesis of blood capillaries, and the self-assembly molecular tweezers have universality, induce cell migration and promote the healing process of wounds. The self-assembled molecular tweezers hydrogel does not pull a wound during replacement, so that the compliance of a patient is improved.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of biotechnology, and particularly relates to a compound combination for inducing cell migration by manipulating membrane curvature and application thereof. BACKGROUND

[0002] In the process of cell phagocytosis, secretion, migration, etc., the cell membrane as a heterogeneous dynamic structure often changes its own morphology to achieve its optimal physiological performance, and its morphology is formed by the direct action of the shape force on the membrane. Membrane curvature is an important parameter in the definition of the shape of cells, organelles and local membrane sub-regions, and is crucial for maintaining the normal function of cells and organelles, including the processes of vesicle generation, cell autophagy, etc. (H. McMahon and Emmanuel Boucrot. “Membrane curvature at a glance.” Journal of Cell Science (2015). 1065 - 1070.). Its formation is a complex process involving multiple mechanisms, including changes in lipid composition, traction of cytoskeleton, hydrophobic insertion of proteins and scaffold formation of protein, etc. (Sun Xia, Hu Junjie. Discussion on the formation mechanism of biological membrane curvature [J]. Acta Biophysica Sinica, 2012, 28(07): 602-610.). High membrane curvature regions are mainly achieved by changing the arrangement of membrane coating proteins. When the coating separates, the curvature is reversible, and the separated proteins will be reintroduced in the next cycle. The cooperation between these entities promotes the formation and maintenance of the basic membrane network, which is crucial for the synthesis of secreted proteins, the activation of signal complexes, the transport of substances within cells, the internalization of substances, and the coordination of complex metabolic processes in cells and organisms. In addition, the stability of membrane curvature is also affected by the properties of the membrane material itself, including normal vector, Gaussian curvature, mean curvature, principal direction vector, spontaneous curvature and effective stiffness, etc.

[0003] The change of membrane curvature is the result of the joint action of membrane proteins, phospholipids and external forces acting on the membrane, mainly generated and regulated by natural membrane plasticity proteins or through external physical stimulation such as optical tweezers and microtubule aspiration technology. Natural membrane curvature generating proteins, such as the membrane shaping protein family with BAR (Bin / amphiphysin / Rvs) domain, are a key class of molecules that regulate cell membrane curvature (Simunovic M, Evergren E, Callan-Jones A, Bassereau P. Curving Cells Inside and Out: Roles of BAR Domain Proteins in Membrane Shaping and Its Cellular Implications. Annu Rev Cell Dev Biol. 2019 Oct 6;35:111-129.). It is known that the dimerization of BAR protein domain can form a banana or a cigar-shaped dimer, and the binding of the dimer with special morphology to the cell membrane can reshape the cell membrane to generate different membrane curvatures. On the other hand, optical tweezers and micro-pipette technology can provide additional tension by applying mechanical stimulation, thereby causing membrane curvature. However, membrane curvature is difficult to regulate, not only because of the limitations of perturbation on the dynamic and complex system of cell membrane, but also because the membrane curvature in many cellular processes is infinitesimal. These greatly limit the application of physical methods in manipulating membrane curvature.

[0004] Biological membranes are essential for basic cell metabolism, but current methods of biological membrane regulation mainly focus on the polymerization of membrane plasticity proteins and physical manipulation. Both methods have non-negligible drawbacks. On the one hand, the regulation of membrane plasticity protein polymerization greatly limits the selection of protein aggregation units. On the other hand, physical methods such as optical tweezers and micro-pipettes usually require very expensive equipment, and the operation of such equipment is difficult, greatly increasing the use cost of physical manipulation methods. At the same time, physical operation can cause mechanical damage to the cell sample, affecting the metabolism and activity of the cell. Therefore, a feasible method is urgently needed to regulate membrane curvature, realize the polymerization / depolymerization process of membrane proteins, and efficiently regulate membrane curvature.

[0005] Wound repair is a hot research topic in the field of medicine today, because the skin is the largest and most exposed organ of the human body, which is extremely vulnerable to injury (Amini Nik Saeid, Cambridge Elizabeth, Yu Winston, Guo Anne, Whetstone Heather, Nadesan Puviindran, Poon Raymond, Hinz Boris, Alman Benjamin A. β-Catenin–regulated myeloid cell adhesion and migration determine wound healing[J]. Journal of Clinical Investigation, 2014 (6).), when some damage causes the loss of integrity of living tissue, at this time it will become a wound (Roosterman Dirk, Goerge Tobias, Schneider Stefan W, Bunnett Nigel W, Steinhoff Martin. Neuronal control of skin function: the skin as a neuroimmunoendocrine organ. [J]. Physiological reviews, 2006.). According to the nature of the wound, it can be divided into acute and chronic wounds. Acute wounds usually need to heal within 8 to 12 weeks. The main cause of acute wounds is mechanical damage caused by external factors, such as abrasions and tears caused by friction between the skin and hard surfaces. Mechanical damage also includes penetrating wounds caused by cuts and gunshots, and surgical wounds caused by surgery such as tumor resection. Burns and chemical injuries caused by radiation, electricity, corrosive chemicals and heat sources are also included in the category of acute injuries. Chronic wounds heal slowly, usually do not heal within 12 weeks, and are often caused by repeated tissue damage (Gurtner Geoffrey C, Werner Sabine, Barrandon Yann, Longaker Michael T. Wound repair and regeneration. [J]. Nature, 2008.), which often leads to severe tissue necrosis of bones, joints and nerves.These non-healing or slow healing wounds are caused by repeated trauma to the wound site or underlying physiological conditions, such as diabetes, persistent infection, poor underlying care, and other patient-related factors (Nancy Broderick. Understanding chronic wound healing [J]. The Nurse Practitioner, 2009.).

[0006] Normal wound healing includes four classic stages: hemostasis, inflammation, proliferation, and remodeling (R. Li, K. Liu, X. Huang, et al. Bioactive Materials Promote Wound Healing through Modulation of Cell Behaviors. Adv Sci (Weinh) 2022, 9(10): e2105152.). After the formation of the wound, platelets release growth factors to recruit leukocytes to the damaged area, triggering the inflammatory stage of wound healing (Robson M C, Steed D L, Franz M G. Wound healing: Biologic features and approaches to maximize healing trajectories [J]. Curr Probl Surg, 2001, 38(2): 72-140.). In the late inflammatory stage, monocytes differentiate into macrophages, starting the proliferation stage of wound healing, promoting the migration, proliferation, and differentiation of wound repair cells (including endothelial cells, fibroblasts, and keratinocytes), which are responsible for processes such as angiogenesis, granulation tissue formation, extracellular matrix deposition, wound contraction, and re-epithelialization (da Silva L, Carvalho E, Cruz M T. Role of neuropeptides in skin inflammation and its involvement in diabetic wound healing [J]. Expert Opin Biol Ther, 2010, 10(10): 1427-1439; Kim S K, Lee KJ, Hahm J R, et al. Clinical significance of the presence of autonomic and vestibular dysfunction in diabetic patients with peripheral neuropathy [J]. Diabetes Metab J, 2012, 36(1): 64.). In the remodeling stage of wound healing, the cells involved in the proliferation stage undergo apoptosis, leaving a mature, essentially avascular, and less cellular environment.

[0007] However, when the healing process is disrupted by microorganisms or some underlying pathological mechanisms, the wound fails to heal and develops into a chronic wound (Q. Zheng, C. Chen, Y. Liu, et al. Metal Nanoparticles: Advanced and Promising Technology in Diabetic Wound Therapy. Int J Nanomedicine 2024,19: 965-992.). Furthermore, the secretion of pro-angiogenesis-related cytokines is reduced, further hindering angiogenesis, which maintains cell growth, supplies nutrients, and provides growth factors (GFs) (S. Sreekumar, MS Kiran. Localized trans-browning and pro-angiogenesis inductive self-assembled collagen resveratrol bio-matrix for tissue repair and regeneration in obese conditions. Int J Biol Macromol 2024: 130322.). Disorders in one or more stages of wound repair can affect the overall rate and quality of wound healing. If skin wounds fail to heal for a long time, leading to excessive exudate and necrosis of healthy skin tissue around the wound, bacterial infection can occur, causing wound ulceration or even sepsis and other serious consequences, ultimately affecting the body's health. Therefore, accelerating wound healing by inducing cell migration can help the body recover health quickly. Summary of the Invention

[0008] This invention discovers that sialyl lactose and rhodamine B are linked via acylhydrazone bonds to form a binding and driving unit, namely the R-SL small molecule inducible ligand. Hemagglutinin protein, as a membrane protein aggregation unit, specifically recognizes sialyl lactose on the R-SL small molecule inducible ligand to form "self-assembled molecular tweezers" (ATs). These "self-assembled molecular tweezers" achieve the polymerization / depolymerization process of membrane proteins through chemical regulation, thereby efficiently controlling cell membrane curvature, enabling cells to form pseudopodia, enhancing cell migration ability, and promoting wound healing. Based on this, this invention was completed.

[0009] In a first aspect, the present invention provides an R-SL small molecule inducible ligand, wherein the R-SL small molecule inducible ligand comprises sialyl lactose and rhodamine B; wherein the sialyl lactose and rhodamine B are linked by an acylhydrazone bond; wherein the sialyl lactose on the R-SL small molecule inducible ligand specifically recognizes hemagglutinin protein; and wherein the structural formula of the R-SL small molecule inducible ligand is shown in formula (I): Equation (Ⅰ).

[0010] Secondly, the present invention provides a "self-assembled molecular tweezers" assembly, the "self-assembled molecular tweezers" assembly comprising an R-SL small molecule inducible ligand and a hemagglutinin protein; wherein, the R-SL small molecule inducible ligand comprises sialyl lactose and rhodamine B, the sialyl lactose and rhodamine B being linked by an acylhydrazone bond; the sialyl lactose on the R-SL small molecule inducible ligand specifically recognizes the hemagglutinin protein; the structural formula of the R-SL small molecule inducible ligand is shown in formula (I): Equation (Ⅰ).

[0011] Thirdly, the present invention provides the application of the "self-assembled molecular tweezers" assembly as described in the second aspect in the preparation of drugs that induce cell migration.

[0012] Furthermore, the "self-assembled molecular tweezers" includes an R-SL small molecule inducible ligand and a hemagglutinin protein, wherein the R-SL small molecule inducible ligand comprises sialyl lactose and rhodamine B; the sialyl lactose and rhodamine B are linked by an acylhydrazone bond; the sialyl lactose on the R-SL small molecule inducible ligand specifically recognizes the hemagglutinin protein; the structural formula of the R-SL small molecule inducible ligand is shown in formula (I): Equation (Ⅰ).

[0013] Furthermore, the induction of cell migration includes promoting wound healing.

[0014] Furthermore, the wound includes both acute and chronic wounds.

[0015] Furthermore, the wound includes, but is not limited to, one or more of the following: cuts, tears, abrasions, burns, bruises, punctures, ulcers, and radiation injuries.

[0016] In one embodiment of this application, the wound-healing drug induces cell pseudopodia formation and cell migration by specifically recognizing sialic acid lactose on the R-SL small molecule induced ligand with endogenous hemagglutinin protein, thereby promoting wound healing.

[0017] Fourthly, the present invention provides a pharmaceutical composition for inducing cell migration, the pharmaceutical composition comprising the active ingredients of the "self-assembly molecular tweezers" combination described in the second aspect and other pharmaceutically acceptable excipients for inducing cell migration, and pharmaceutically acceptable excipients thereof.

[0018] Furthermore, the induction of cell migration includes promoting wound healing.

[0019] Furthermore, the wound includes both acute and chronic wounds.

[0020] Furthermore, the wound includes, but is not limited to, one or more of the following: cuts, tears, abrasions, burns, bruises, punctures, ulcers, and radiation injuries.

[0021] In one embodiment of this application, the wound-healing drug induces cell pseudopodia generation and cell migration by specifically recognizing sialic acid lactose on the R-SL small molecule ligand with endogenous hemagglutinin protein, thereby promoting wound healing.

[0022] Furthermore, the drug can be formulated into various dosage forms with commonly used pharmaceutical excipients.

[0023] Furthermore, the "self-assembled molecular tweezers" includes an R-SL small molecule inducible ligand and a hemagglutinin protein; wherein the R-SL small molecule inducible ligand includes sialyl lactose and rhodamine B, which are linked by an acylhydrazone bond; the sialyl lactose on the R-SL small molecule inducible ligand specifically recognizes the hemagglutinin protein; the structural formula of the R-SL small molecule inducible ligand is shown in formula (I): Equation (Ⅰ).

[0024] Fifthly, the present invention provides a hydrogel comprising the "self-assembled molecular tweezers" assembly described in the second aspect and a hydrogel precursor solution; the hydrogel precursor solution is prepared from an organic solvent and a natural polymer, and the hydrogel is formed by covalent cross-linking of the organic solvent.

[0025] Furthermore, the "self-assembled molecular tweezers" includes an R-SL small molecule inducible ligand and a hemagglutinin protein; wherein the R-SL small molecule inducible ligand includes sialyl lactose and rhodamine B, which are linked by an acylhydrazone bond; the sialyl lactose on the R-SL small molecule inducible ligand specifically recognizes the hemagglutinin protein; the structural formula of the R-SL small molecule inducible ligand is shown in formula (I): Equation (Ⅰ).

[0026] Furthermore, the organic solvent is selected from one or more of glycidyl methacrylate, acryloyl chloride, N-hydroxysuccinimide acrylate, maleic anhydride, acrylic anhydride and / or methacrylic anhydride (MA); preferably methacrylic anhydride (MA).

[0027] Furthermore, the natural polymer is selected from one or more of polyethylene glycol, chitosan, cellulose and its derivatives, hyaluronic acid, silk fibroin, alginate, collagen and / or gelatin; preferably gelatin.

[0028] In a sixth aspect, the present invention provides the use of the hydrogel as described in the fifth aspect in the preparation of products that induce cell migration.

[0029] Furthermore, the induction of cell migration includes promoting wound healing.

[0030] Furthermore, the wound includes both acute and chronic wounds.

[0031] Furthermore, the wound includes, but is not limited to, one or more of the following: cuts, tears, abrasions, burns, bruises, punctures, ulcers, and radiation injuries.

[0032] Furthermore, the hydrogel comprises a "self-assembled molecular tweezers" assembly and a hydrogel precursor solution, the hydrogel precursor solution being prepared from an organic solvent and a natural polymer, and the hydrogel being formed through covalent cross-linking of the organic solvent.

[0033] Furthermore, the "self-assembled molecular tweezers" comprises an R-SL small molecule inducible ligand and a hemagglutinin protein, wherein the R-SL small molecule inducible ligand comprises sialyl lactose and rhodamine B; the sialyl lactose and rhodamine B are linked by an acylhydrazone bond; the sialyl lactose on the R-SL small molecule inducible ligand specifically recognizes the hemagglutinin protein; the structural formula of the R-SL small molecule inducible ligand is shown in Formula I: Equation (Ⅰ).

[0034] Furthermore, the organic solvent is selected from one or more of glycidyl methacrylate, acryloyl chloride, N-hydroxysuccinimide acrylate, maleic anhydride, acrylic anhydride and / or methacrylic anhydride (MA); preferably methacrylic anhydride (MA).

[0035] Furthermore, the natural polymer is selected from one or more of polyethylene glycol, chitosan, cellulose and its derivatives, hyaluronic acid, silk fibroin, alginate, collagen and / or gelatin; preferably gelatin.

[0036] Furthermore, the products include pharmaceuticals and medical devices.

[0037] Furthermore, the medical supplies and medical devices include injectable surgical fillers, bionanomaterials, drug delivery systems, and tissue repair and cell scaffolds.

[0038] Beneficial effects This application uses hemagglutinin protein as the membrane protein aggregation unit, and links sialic acid lactose and rhodamine B through acylhydrazone bonds to form a binding and driving unit (R-SL small molecule inducible ligand), forming "self-assembled molecular tweezers" (ATs). The preparation process of these "self-assembled molecular tweezers" is simple, and the operation is simple when regulating cell membrane curvature and inducing cell migration, reducing equipment and labor costs.

[0039] The "self-assembling molecular tweezers" constructed in this application achieve the polymerization / depolymerization process of membrane proteins through a simple chemical regulation method, thereby efficiently regulating membrane curvature; significantly inducing cell migration and promoting wound healing.

[0040] This application has been verified through cell-level experiments that the "self-assembling molecular tweezers" promote the generation of pseudopodia, enhance cell migration ability, induce capillary regeneration, and further advance the wound healing process when regulating cell curvature.

[0041] The self-assembled molecular tweezers constructed in this application have universality in inducing cell migration, promoting the migration of both cancer cells and non-cancer cells, and promoting wound healing.

[0042] The self-assembled molecular tweezers hydrogel constructed in this application can induce cell migration and promote wound healing. It does not pull on the wound when replaced, thereby improving patient compliance. Attached Figure Description

[0043] Figure 1 To determine the reaction route for preparing R-SL small molecule induced ligands.

[0044] Figure 2 The images are for microscopic characterization, and the spectra are for evidence of rhodamine molecular aggregation.

[0045] Note: a is a transmission electron microscopy image of liposomes after incubation with hemagglutinin protein, small molecule inducible ligand, and "self-assembled molecular tweezers"; b is a microscopic image of liposomes after co-incubation with hemagglutinin protein, small molecule inducible ligand, and "self-assembled molecular tweezers"; c is a molecular aggregation spectrum of rhodamine; Liposome is liposome; HA is hemagglutinin protein; R-SL is small molecule inducible ligand; ATs is "self-assembled molecular tweezers".

[0046] Figure 3 Laser confocal microscopy characterization results of induced pseudopodia in HepG2 cells induced by "self-assembling molecular tweezers".

[0047] Note: HepG-2 refers to HepG2 cells; ATs stands for "self-assembling molecular tweezers".

[0048] Figure 4 Laser confocal microscopy characterization results of induced pseudopodia in BPH-1 and BV2 cells induced by "self-assembling molecular tweezers".

[0049] Note: A is a laser confocal image of induced pseudopodia in BPH-1 cells; B is a laser confocal image of induced pseudopodia in BV2 cells; BPH-1 refers to BPH-1 cells; BV2 refers to BV2 cells; ATs stands for "self-assembling molecular tweezers".

[0050] Figure 5 Fluorescence microscopy results showing the promoting effect of small molecule inducible ligands on the generation of induced pseudopodia in HepG2 cells overexpressing hemagglutinin.

[0051] Note: HepG-2 refers to HepG2 cells; ATs stands for "self-assembling molecular tweezers".

[0052] Figure 6 Microscopic results showing the promoting effect of small molecule inducible ligands on the generation of induced pseudopodia in BPH-1 and BV2 cells overexpressing hemagglutinin.

[0053] Note: A is a laser confocal image of induced pseudopodia in BPH-1 cells; B is a laser confocal image of induced pseudopodia in BV2 cells; BPH-1 refers to BPH-1 cells; BV2 refers to BV2 cells; ATs stands for "self-assembling molecular tweezers".

[0054] Figure 7 This study aimed to test the ability of HepG2 cells treated with "self-assembling molecular tweezers" to induce pseudopodia to promote migration.

[0055] Note: A is the crystal violet staining result for Transwell cell migration; B is the OD value of crystal violet staining; HepG2 is HepG2 cells; HA is hemagglutinin; R-SL is small molecule inducible ligand; ATs is "self-assembling molecular tweezers".

[0056] Figure 8 This test examines the migration-promoting ability of BPH-1 and BV2-induced pseudopodia after treatment with "self-assembled molecular tweezers".

[0057] Note: A and C are the results of crystal violet staining for Transwell cell migration; B and D are the OD values ​​of crystal violet staining; BPH-1 is BPH-1 cell; BV2 is BV2 cell; HA is hemagglutinin; R-SL is small molecule inducible ligand; ATs is "self-assembling molecular tweezers".

[0058] Figure 9 This study aimed to investigate the promoting effect of small molecule inducible ligands on the formation of induced pseudopodia in L929 cells overexpressing hemagglutinin.

[0059] Note: ab represents fluorescence microscopy results; cd represents Transwell results indicating cell migration.

[0060] Figure 10 This demonstrates the promoting effect of "self-assembled molecular tweezers" on angiogenesis in HUVECs.

[0061] Note: a is the microscopic observation result of the formation of vascular structures in HUVEC cells after treatment with "self-assembled molecular tweezers"; b and c are the analysis of the number of nodes and tube length of vascular structures formed by HUVEC cells after treatment with "self-assembled molecular tweezers".

[0062] Figure 11 It is a hydrogel for "self-assembling molecular tweezers".

[0063] Figure 12 The effect of wound healing on SD rats.

[0064] Note: a is a schematic diagram of the verification experiment on the promoting effect of "self-assembled molecular forceps" on wound healing in SD rats; b is a photograph of wounds (8 mm) treated with GelMA hydrogel (gel group) or loaded with AT gel (gel-AT group) and untreated (control group) on days 0, 3, 7, 10 and 14; c is the wound bed closure status of each group within 14 days; d is the wound healing rate on days 3, 7, 10 and 14; e is the H&E staining result of wound tissue on day 14; f is the Masson staining result of wound tissue on day 14. Detailed Implementation

[0065] The specific embodiments of the present invention will be further described below. It should be noted that these descriptions are for the purpose of aiding understanding the present invention, but do not constitute a limitation thereof. Furthermore, the technical features involved in the embodiments described below can be combined with each other as long as they do not conflict with each other.

[0066] Unless otherwise specified, the test methods in the following embodiments are conventional methods, and the test materials used in the following embodiments are all available through conventional commercial channels.

[0067] Terminology Explanation "Pharmaceutical acceptable" ingredients are substances that are suitable for use in humans and / or mammals without excessive adverse side effects (such as toxicity, irritation, and allergic reactions), i.e., substances with a reasonable benefit / risk ratio.

[0068] "Pharmaceutically acceptable carrier" refers to a carrier used for the administration of therapeutic agents.

[0069] BAR protein domains: BAR (Bin / Rvs) domains are found in more than 35 proteins encoded in the human genome. These proteins play roles in various cellular processes, such as endocytosis (i.e., endocytokines, sorting linkers, and Rvs) and actin reorganization (i.e., RhoGAP and RhoGEF). The BAR domain of Arfaptin has been shown to bind Rac1. The BAR domain of Rvs is approximately 210 amino acids long and contains a coiled helix consisting of three extended α-helices. The functional BAR dimer is a banana-shaped bundle with six helices, positively charged at its tip and concave side, mediating phospholipid binding. The concave side's curvature conforms to a circular membrane with a diameter of approximately 220 angstroms. While all BAR domains are highly likely to bind curved lipids, some BAR domains can actually induce membrane curvature. Therefore, it can be predicted that BAR domains may recruit target proteins to curved membranes during vesicle formation or structurally assist in inducing membrane curvature.

[0070] Optical tweezers: A parallel laser beam focused by a microscope objective creates a micrometer-scale spot. The larger the numerical aperture of the objective, the smaller the focused spot (reaching hundreds of nanometers), and the greater its electric field gradient. For dielectric particles, the strongly focused spot acts as a three-dimensional optical potential trap, confining the particle to its point of lowest potential energy. If the particle deviates from this point, it experiences a restoring force pointing back to it. The strongly focused laser beam creates a "trap" for dielectric particles, trapping them within it. Moving the focused spot causes the particle to move with it. Such a strongly focused spot allows for micromanipulation of particles, including trapping, moving, and rotating them, much like a pair of "tweezers," hence the name optical tweezers.

[0071] Micropipette aspiration technique (MAT) is a technique used to study the mechanical and viscoelastic properties of cells by measuring the dynamics of cell deformation under negative pressure. It is one of the earliest techniques used to measure cellular mechanical properties. The advantages of micropipette aspiration technique lie in its ability to directly observe the occurrence of molecularly mediated cell adhesion events, and the controllability of each adhesion event. This allows for precise control over when two cells begin to contact and separate, as well as the magnitude of the force applied to the contact. Furthermore, by combining it with probabilistic dynamics theory of small systems, kinetic force spectrum theory, and mechanochemical coupling theory, it is possible to obtain more comprehensive information reflecting the reaction kinetics, including positive and negative reaction rates and affinity of intermolecular interactions under no external force, as well as fracture force spectrum and lifetime spectrum.

[0072] Acylhydrazone (Hyd) bond: A common type of bond formed by a covalent bond between a nitrogen atom and a carbon atom, it is a stable, polar covalent bond in organic compounds. Its structural characteristic is that the nitrogen atom shares an electron pair with one or more carbon atoms, forming a double bond. The Hyd bond is usually located at the center of the acylhydrazone compound, connecting the carbon atom containing the C=O (acyl group) and the nitrogen atom. The general structure of the acylhydrazone bond can be represented as RC(=O)-N-R', where R and R' can be hydrogen atoms or carbon chains.

[0073] Sialyllactose is a human milk oligosaccharide, one of the most abundant sialylated oligosaccharides among human milk oligosaccharides (HMOs). 6'-sialyllactose (6'-SL) and 3'-sialyllactose (3'-SL) are the main forms of sialyllactose, both of which have antibacterial activity and immunomodulatory effects. Sialyllactose can specifically recognize hemagglutinin protein.

[0074] Hemagglutinin (HA): Hemagglutinin (HA) or hemagglutinin (BE) is an antigenic glycoprotein located on the surface of influenza viruses, responsible for binding the virus to infected cells. Hemagglutinin refers to erythroglobulin, which is columnar and binds to receptors on the surface of erythrocytes in animals such as humans, birds, pigs, and guinea pigs, causing clotting, hence the name hemagglutinin, also called erythrocyte hemagglutinin. Two proteins exist on the surface of the virus: hemagglutinin and neuraminidase. In the highly pathogenic avian influenza virus H5N1, "H" refers to the former and "N" to the latter. After hydrolysis, hemagglutinin protein separates into light and heavy chains. The heavy chain can bind to sialic acid receptors on the host cell membrane, while the light chain assists in the fusion of the viral envelope with the host cell membrane. Hemagglutinin plays a crucial role in the introduction of the virus into host cells, possesses immunogenicity, and anti-hemagglutinin antibodies can neutralize influenza viruses. Rhodamine B is a synthetic red basic fluorescent dye, commonly known as pollen red. This dye is frequently used as a fluorescent reagent and is widely applied in colored glass, mining, and steel industries; it is a common basic industrial dye.

[0075] Dipalmitate phosphatidylcholine (DPPC): A zwitterionic glycerophospholipid commonly used to form lipid monolayers, bilayers, and liposomes for a variety of applications. It has been used to form protein liposomes for implanting gamma-glutamyl transpeptidase into the human erythrocyte membrane.

[0076] Dioleoyl lecithin (DOPC): A commonly used phospholipid molecule, it is prepared by the condensation of 1,2-dioleoyl lecithin and 1,2-n-dodecanool. It is typically used alone or in combination with other components in the generation of micelles, liposomes, and other types of artificial membranes.

[0077] Fluorescent labeling of lysosomal liposomes (NBD-PE): NBD-PE is a fluorescent labeling of lysosomal liposomes. It consists of a single-tailed (16:0) phospholipid derivative and an NBD fluorophore conjugated to the head group, making it an effective lipid fluorescent probe.

[0078] HepG 2 cells: Human liver cancer cells were isolated in 1975 from tumor tissue of a 15-year-old Caucasian male with liver cancer. These cells reportedly do not carry the hepatitis B virus genome but express a range of proteins, including alpha-fetoprotein, albumin, α-2-macroglobulin, α-1-antitrypsin, transferrin, α-1-antichymotrypsin, haptoglobin, ceruloplasmin, plasminogen, complement C4 and C3 activators, fibrinogen, α-1-acid glycoprotein, α-2-HS-glycoprotein, β-lipoprotein, and retinol-binding protein; they also express insulin receptors and insulin-like growth factor II receptors, and exhibit 3-hydroxy-3-methylglutaryl-CoA and hepatic triglyceride lipase activity. Further data indicate that these cells are non-tumorigenic in immunosuppressed SD rats but can form tumors in semi-solid culture media, making them suitable for large-scale culture systems. They have been widely used in cytotoxicology experiments and tumor biology research.

[0079] BHP-1 cells: Selected from human prostate hyperplasia cells, they are a transformed cell line.

[0080] BV2 cells are a microglia cell line derived from SD rats that are widely used in neuroscience research. This immortalized cell line can serve as an in vitro model for studying neurodegenerative diseases and related cellular conditions and processes, such as neuroinflammation. Furthermore, BV2 cells are considered an alternative model system for primary microglia.

[0081] Matrigel: Basement membrane matrix extracted from EHSSD rat tumors rich in extracellular matrix proteins. Its main components include laminin, type IV collagen, nestin, heparin sulfate glycoprotein, as well as growth factors and matrix metalloproteinases. Matrigel matrix can effectively promote the adhesion and differentiation of normal and transformed adherent-dependent epithelial cells and other cell types, including neurons, supporting cells, chicken lens, vascular endothelial cells, and hepatocytes.

[0082] Masson staining, also known as Masson staining, is a classic method for staining connective tissue. It is one of the main methods for displaying fibers in tissue and is an authoritative and classic technique for staining collagen fibers. The staining principle is related to the size of the anionic dye molecules and their penetration into the tissue. The size of the molecules is reflected by their molecular weight; small molecular weights easily penetrate dense tissues with low permeability, while large molecular weights can only enter loose tissues with high permeability. However, pale green or aniline blue dyes have very large molecular weights. Therefore, after Masson staining, muscle fibers appear red, while collagen fibers appear green or blue, primarily used to distinguish between collagen fibers and muscle fibers.

[0083] Example 1: Preparation of R-SL small molecule inducible ligands Test methods R-SL Small Molecule Induced Ligand Synthesis Rhodamine B was hydrolyzed with hydrazine hydrate to prepare rhodamine hydrazide (RhB-Hydrazide). The rhodamine hydrazide was then dissolved in anhydrous methanol and stirred under a nitrogen atmosphere. Sialyllactose (2,3) and yttrium trifluoromethanesulfonate were added to the rhodamine hydrazide solution. This mixture was then concentrated, and the crude product was purified by column chromatography for separation.

[0084] Test results like Figure 1 As shown, the small molecule inducible ligand R-SL, which appears as a pink powder, was isolated.

[0085] Example 2: Induction of Phospholipid Membrane Curvature in Artificial Liposomes by "Self-Assembled Molecular Tweezers" Test methods Artificial liposome preparation To mimic the cell membrane, hemagglutinin protein is inserted into the membrane.

[0086] Dipalmitoylphosphatidylcholine (DPPC): dioleoyllecithin (DOPC): NBD-PE (fluorescent labeling for lysosomal liposomes) were dissolved in chloroform solution at a molar ratio of 4:1:0.1. The mixture was dried under a nitrogen atmosphere. Finally, the dried lipid membrane was hydrated with PBS buffer. The resulting artificial liposomes had a final concentration of 20 mg / mL.

[0087] Add the hemagglutinin protein solution to the Alexa Fluor™ 405 NHS dye solution; Ultrafiltration is used to remove unbound dye molecules; Blue fluorescently labeled hemagglutinin protein and small molecule ligands were added to the liposome system at a concentration ratio of 1:10 and co-incubated. Transmission electron microscopy and laser confocal microscopy were used for image analysis, and ultraviolet spectroscopy and circular dichroism spectroscopy were used to characterize the aggregation behavior of small molecule induced ligands.

[0088] Test results like Figure 2 As shown, liposomes can only generate membrane tubular structures in systems where both hemagglutinin and small molecule inducible ligands are present. This is because the aggregation of small molecule inducible ligands induces the aggregation of hemagglutinin, thereby affecting the rearrangement of phospholipid molecules bound to and surrounding the hemagglutinin. The rearrangement of phospholipid molecules leads to changes in the local membrane curvature, ultimately generating membrane tubular structures. Therefore, this application refers to this method, which can induce changes in membrane curvature, as the "self-assembly molecular tweezers" method.

[0089] Example 3: Induction of HepG2 (cancer cell) cell membrane curvature in a live cell system using "self-assembled molecular tweezers". Test methods Add 5 mL of FBS and 500 µL of double antibiotics to 45 mL of 1640 medium; When the HepG 2 cell confluence rate reached 80%, the culture medium was removed, the cells were washed with PBS, digested, and centrifuged for counting. With 5×10 4 Seeds were placed in Confocal glass dishes at a density of 10 cells / well. After the cells adhered, the original culture medium was discarded and 1640 culture medium was added to the Confocal dishes. Individual samples of hemagglutinin, small molecule inducible ligands, and self-assembled molecular tweezers (hemagglutinin + small molecule inducible ligands) were prepared to the required working concentrations: HA at 4.2 μM and R-SL at 42 μM.

[0090] After co-culturing, the cells were fixed with paraformaldehyde, and the cytoskeleton was stained with Alexa Fluor 488 phalloidin fluorescent dye. The images were then taken and recorded under a microscope.

[0091] Test results like Figure 3 As shown, the control group HepG2 cells exhibited a typical cobblestone shape, while in the cell experiment group co-incubated with "self-assembling molecular tweezers", the local curvature of the cell membrane changed significantly, and the cells showed a long / multiple pseudopodia morphology, which further promoted cell migration.

[0092] Example 4: Induction of BPH-1 (glial cell) cell membrane curvature in a live cell system using "self-assembled molecular tweezers". Test methods Add 5 mL of FBS and 500 µL of double antibiotics to 45 mL of 1640 medium; When the BPH-1 cell fusion rate reaches 80%, remove the culture medium, wash with PBS, digest the cells, and centrifuge to count them. With 5×10 4 Seeds were placed in Confocal glass dishes at a density of 10 cells / well. After the cells adhered, the original culture medium was discarded and 1640 culture medium was added to the Confocal dishes. Individual samples of hemagglutinin, small molecule inducible ligands, and self-assembled molecular tweezers (hemagglutinin + small molecule inducible ligands) were prepared to the required working concentrations: HA at 4.2 μM and R-SL at 42 μM. After co-culturing, the cells were fixed with paraformaldehyde, and the cytoskeleton was stained with Alexa Fluor 488 phalloidin fluorescent dye. The images were then taken and recorded under a microscope.

[0093] Test results like Figure 4 As shown, the control group BPH-1 cells exhibited a typical cobblestone shape, while after co-incubation with "self-assembling molecular tweezers", they showed morphological features of multiple induced pseudopodia, which further promoted cell migration.

[0094] Example 5: Induction of BV2 (glial cell) cell membrane curvature in a living cell system using "self-assembled molecular tweezers". Test methods Add 5 mL of FBS and 500 µL of double antibiotics to 45 mL of 1640 medium; When the BV2 cell fusion rate reaches 80%, remove the culture medium, wash with PBS, digest the cells, and centrifuge to count them. With 5×10 4 Seeds were placed in Confocal glass dishes at a density of 10 cells / well. After the cells adhered, the original culture medium was discarded and 1640 culture medium was added to the Confocal dishes. Individual samples of hemagglutinin, small molecule inducible ligands, and self-assembled molecular tweezers (hemagglutinin + small molecule inducible ligands) were prepared to the required working concentrations: HA at 4.2 μM and R-SL at 42 μM. After co-culturing, the cells were fixed with paraformaldehyde, and the cytoskeleton was stained with Alexa Fluor 488 phalloidin fluorescent dye. The images were then taken and recorded under a microscope.

[0095] Test results like Figure 4 As shown, the control group BV2 cells exhibited a typical cobblestone shape, while after co-incubation with "self-assembling molecular tweezers", they showed morphological features of multiple induced pseudopodia, which further promoted cell migration.

[0096] Example 6: Induction of cell membrane curvature by endogenous expression of hemagglutinin protein forming "self-assembled molecular tweezers" plasmid extraction (1) Inoculate the bacterial strain carrying hemagglutinin protein plasmid (GFP reporter gene, Amp resistance) into 10-15 mL of LB liquid medium containing 100 μg / mL of the corresponding antibiotic, and culture in a shaker at 37°C for 12-16 hours to amplify the plasmid. (2) Centrifuge the cultured system from (1) at 3,000-5,000g for 10 minutes and collect 10-15 ml of bacterial culture; (3) Discard the culture medium in the bacterial solution obtained in (2), and invert it onto absorbent paper and gently pat to absorb the residual liquid. Use a mixture of buffer P1 / RNase A with 500 µL added beforehand to vortex and resuspend the bacteria. After resuspension, no cell clumps should be visible. (4) Transfer the resuspension from (3) to a 2 mL centrifuge tube, add 500 µL buffer P2, gently invert 8 to 10 times, and let stand at room temperature for 2 minutes. Invert and mix occasionally, gently invert and mix, do not vortex, otherwise it will cause genomic DNA breakage and contamination. When the bacterial culture volume exceeds 10 mL, the lysis buffer will be very viscous and difficult to mix. After full lysis, the solution becomes viscous and clear. If necessary, you can slowly invert and mix until the lysis buffer becomes clear, but this step should not take more than 4 minutes. Add 700 µL of buffer P3 and immediately invert to mix 15 to 20 times. After adding buffer P3, invert to mix immediately to prevent precipitate aggregation that could affect the neutralization effect. The mixing process must be gentle. When the amount of bacterial culture is large, neutralization may be more difficult. Increase the number of inversions until the solution is completely neutralized. Centrifuge the above solution for 10 minutes (≥13,000 g). Place the HiPure DNA Mini Column III column into the collection tube, transfer half the volume of supernatant into the column, and centrifuge at 13,000 g for 30-60 seconds; discard the eluent, put the column back into the collection tube, transfer the remaining supernatant into the column, and centrifuge at 13,000 g for 30-60 seconds; discard the eluent, put the column back into the collection tube, add 500 µL of buffer PW1 into the column, and centrifuge at 13,000 g for 30-60 seconds; discard the eluent, put the column back into the collection tube, add 600 µL of buffer PW2 into the column, and centrifuge at 13,000 g for 30-60 seconds; discard the eluent, put the column back into the collection tube, add 600 µL of buffer PW2 into the column, and centrifuge at 13,000 g for 30-60 seconds. Discard the filtrate, put the column back into the collection tube, centrifuge at 13,000 g for 2 minutes to dry the column; put the column into a sterile 1.5 mL centrifuge tube. Add 75 ~ 100 µL of Elution Buffer or sterile water to the center of the membrane of the column, let stand for 2 minutes, and centrifuge at 12,000 g for 1 minute; After centrifugation, discard the column, take 2 μL of sample, determine the concentration of the newly extracted plasmid using Nanodrop, label it, and store the plasmid for later use.

[0097] plasmid transfection When HepG 2 cells reach 70%~90% confluence, use Lipofectamine 3000 for plasmid transfection; dilute Lipofectamine 3000™ reagent (2 tubes) with serum-free DMEM medium and mix gently and thoroughly. DNA was diluted with serum-free DMEM medium to prepare a DNA premix, and then P3000 reagent was added and mixed well. Dilute Lipofectamine 3000™ reagent and add it to the DNA diluted in step 2) (1:1), then incubate (at room temperature for 5 minutes). Add the DNA-liposome complex to the cell culture dish and incubate HepG2 cells at 37 °C for 48 hours; Successful transfection was determined by observing the presence of green fluorescence in HepG 2 cells under a fluorescence microscope.

[0098] Small molecule induced ligands and changes in cell membrane curvature due to overexpression of hemagglutinin protein HepG2 After discarding the original culture medium, add 1 ml of 1640 complete culture medium containing small molecule inducible ligand (final concentration 42 μm) and continue culturing for 18 hours. The culture was terminated, the cells were washed with PBS, and then HepG2 cells were fixed with 4% paraformaldehyde.

[0099] result like Figure 5 As shown, the results indicate that the cell membrane curvature of HepG2 cells can only be induced in the presence of "self-assembled molecular tweezers"; this suggests that "self-assembled molecular tweezers" induces cells to generate elongated inducible pseudopodia structures, resulting in an overall multi-pseudopodia state, which further promotes cell migration.

[0100] Example 7: Universality of "Self-Assembled Molecular Tweezers" for Manipulating the Membrane Curvature of BPH-1 and BV2 Cells BPH-1 and BV2 cells overexpressing hemagglutinin were co-incubated with a certain concentration of the small molecule inducible ligand R-SL for observation.

[0101] like Figure 6As shown, through the interaction of small molecule-induced ligands with endogenously expressed hemagglutinin protein on the cell membrane, a "self-assembled molecular tweezer" is formed during incubation. After the formation of the "self-assembled molecular tweezer," both BPH-1 and BV2 cells exhibit a transformation from a smooth, cobblestone-like structure to a morphology with multiple pseudopodia, thereby regulating cell membrane curvature. This demonstrates that it is a highly efficient chemical manipulation platform that can be successfully applied in different cell systems (cancer cells and non-cancer cells).

[0102] Example 8: Detection of pseudopodia migration ability induced by "self-assembled molecular tweezers" in HepG2 cells HepG2 cells overexpressing hemagglutinin were co-incubated with the small molecule inducible ligand R-SL, followed by a Transwell assay to assess the migration ability of the induced pseudopodia.

[0103] like Figure 7 As shown, under the induction of "self-assembled molecular tweezers", more crystal violet staining was found, and the OD value was 1.5, which was higher than that of other groups. This indicates that the pseudopodia structure induced by "self-assembled molecular tweezers" can significantly promote cell migration ability; it shows that it can perform normal biological functions and is not a simple artificial non-functional pseudopodia structure.

[0104] Example 9: Detection of pseudopodia migration ability induced by "self-assembled molecular tweezers" in BPH-1 and BV2 cells. BPH-1 and BV2 cells overexpressing hemagglutinin were co-incubated with the small molecule inducible ligand R-SL for 18 h, followed by a Transwell assay to assess the migration ability of the induced pseudopodia.

[0105] The test results show that... Figure 8 As shown, through the interaction of a small molecule inducible ligand with endogenously expressed hemagglutinin protein on the cell membrane, a "self-assembled molecular tweezer" is formed during incubation. After the formation of the "self-assembled molecular tweezer," both BPH-1 and BV2 cells exhibit a transformation from a smooth, cobblestone-like structure to one with multiple pseudopodia. Under the induction of the "self-assembled molecular tweezer," crystal violet staining was increased. Quantitative analysis using OD values ​​showed that the OD values ​​of this group were all 0.8, higher than other groups, indicating that the pseudopodia structure induced by the "self-assembled molecular tweezer" significantly promoted cell migration. The "self-assembled molecular tweezer" can induce the production of inducible pseudopodia with clear biological functions in different cell lines (cancer cells and non-cancer cells), demonstrating the good universality of this method.

[0106] Example 10: Promotion of pseudopodia formation and migration ability of self-assembled molecular tweezers in SD rat epithelial fibroblasts (L929) Following the conditions of Example 7, L929 cells (SD rat epithelial fibroblasts) overexpressing hemagglutinin were co-incubated with the small molecule inducible ligand R-SL in the "self-assembly molecular tweezers" for 12 h for confocal imaging and Transwell assays to evaluate the ability of L929 cells induced by the "self-assembly molecular tweezers" strategy to generate pseudopodia and migrate.

[0107] Experimental results showed that the morphology of treated L929 cells changed significantly, exhibiting a multi-pseudopodia morphology. After induction with "self-assembly molecular tweezers," L929 cells produced obvious pseudopodia structures (such as...). Figure 9 As shown in the figure), and under the induction of "self-assembling molecular tweezers", more crystal violet staining was observed. Quantitative analysis was performed using OD values, and the OD value of this group was 1.8, which was higher than that of other groups, indicating that these newly formed pseudopodia structures have a significant ability to promote cell migration (e.g., Figure 9 (As shown in c-9d), this conclusion is verified by the Transwell results in the figure. In summary, the cell morphology of multipodia and the significantly enhanced migration ability contribute to faster wound healing.

[0108] Example 11: Promotion of angiogenesis by "self-assembled molecular forceps" on human umbilical vein endothelial cells (HUVECs) Test methods The day before the experiment, place Matrigel in an ice box and put it in a 4°C freezer to allow the gel to melt slowly overnight (Note: also prepare some pre-cooled 4°C pipette tips and 96-well plates). Add 50 μL of pre-cooled Matrigel to the well plate and place it in a 37°C, 5% CO2 incubator for 30 minutes to allow the substrate to polymerize. HUVEC cells were spaced at 1 x 10 cells per well. 5 Inoculate the gel substrate at a density of 1000; The study was divided into four groups: control group, hemagglutinin protein group, R-SL ligand group, and "self-assembled molecular tweezers" group, with replicates set for each group. After culturing cells in an incubator at 37 °C and 5% CO2 for 24 hours, the formation of the vascular network in HUVEC cells was photographed. Calculate and count the number of branch points and parameters such as branch length for each domain.

[0109] Test results The results showed that after 8 hours of co-culture, the treated group exhibited greater formation of reticular structures, with 100 meshes and a vascular length of 25,000 μm / flied, compared to 63 and 16,000 μm / flied in the control group, respectively. This indicates that the "self-assembling molecular forceps" treatment induces HUVEC cells to generate more vascular structures. These newly formed vascular structures deliver nutrients, immune cells, and oxygen to the wound, providing energy and promoting rapid wound healing (e.g., ...). Figure 10 (As shown).

[0110] Example 12 Preparation of hydrogels containing "self-assembled molecular tweezers" 10 g of gelatin was completely dissolved in 100 mL of deionized water at 50 °C. Then, 8 mL of methylpropionic anhydride (MA) was added dropwise to the gelatin solution, and the mixture was stirred vigorously (200 rpm) and reacted at 50 °C for 1 hour. Finally, the dialyzed gelatin methacryloyl solution (GelMA) was lyophilized for 1 week, and the powder was stored at -80 °C for future use.

[0111] In the experimental group, the "self-assembled molecular tweezers" and hydrogel precursor solution were thoroughly mixed, and then crosslinked by ultraviolet irradiation to obtain the shaped hydrogel material (e.g. Figure 11 (As shown).

[0112] Example 13 Wound healing in SD rats The two components of the "self-assembled molecular forceps," "hemagglutinin protein" and "R-SL," are delivered to the wound surface via a conventional hydrogel, which this application names Gel-AT. This hydrogel is used to demonstrate the efficacy of the "self-assembled molecular forceps" in animal wound healing. First, wound model mice were created using standard methods and divided into a Control group (wounds without further treatment), a GelMA group (wounds treated with the hydrogel component alone), and a Gel-AT group (wounds treated with the hydrogel-loaded "self-assembled molecular forceps"). The experimental procedure is as follows: Figure 12 As shown in a.

[0113] Test methods Select suitable SD rats for the experiment, anesthetize the SD rats with isoflurane (for three weeks), fix the rats on the operating table, perform hair removal on the back, and disinfect with iodine. Cut the skin along the rat's dorsal markings to create full-thickness skin defects: Use sterile ophthalmic scissors to cut the skin along the rat's dorsal markings to create two circular (~8 mm in diameter) full-thickness skin defects; The rats were divided into three groups: Control group, GelMA group, and GelMA + drug group; each group contained several rats, which were separated into different cages and fed normally. The rats' activities were observed daily to prevent the dressing from falling off. Observations were conducted on days 3, 5, 7, 10, and 14; samples were taken after 14 days and subjected to HE and Masson staining for observation.

[0114] Test results The test results are as follows Figure 12 As shown, the Gel-AT treatment group significantly accelerated wound healing, with a wound healing rate exceeding that of the control group and GelMA (…). Figure 12 Quantitative analysis showed that the wound healing rates in the Gel-AT group were 62% and 79% on days 7 and 10, respectively, significantly higher than other groups (control group: 15% and 61%, GelMA group: 38% and 60%). Notably, this group showed near-complete healing (94.8%) on day 14, demonstrating superior wound healing promotion. Figure 12 d). Simultaneously, samples were taken from the wound area and H&E staining was performed (…). Figure 12 e), the results showed that the Gel-AT group exhibited better re-epithelialization compared to the control group and the gel group (indicated by black arrows). Masson staining ( Figure 12 f) shows that collagen deposition is uniform in the AT-treated group (blue area), providing strong support for cell migration.

[0115] The results above demonstrate that the "self-assembled molecular forceps" constructed in this application can significantly promote the wound healing process. Furthermore, through cell-level experiments, this application suggests that the "self-assembled molecular forceps" primarily advances the wound healing process by promoting the healing of L929 epithelial cells and inducing capillary regeneration.

Claims

1. An R-SL small molecule inducible ligand comprising sialyllactose and rhodamine B; the sialyllactose and rhodamine B are linked by an acylhydrazone bond; the sialyllactose on the R-SL small molecule inducible ligand specifically recognizes hemagglutinin protein; the structure of the R-SL small molecule inducible ligand is shown as formula (I): Formula (I).

2. A "self-assembling molecular tongs" combination comprising an R-SL small molecule inducing ligand and a hemagglutinin protein; wherein, The R-SL small molecule inducible ligand comprises sialyllactose and rhodamine B, and the sialyllactose and rhodamine B are linked by an acylhydrazone bond; the sialyllactose on the R-SL small molecule inducible ligand specifically recognizes hemagglutinin protein; the structure of the R-SL small molecule inducible ligand is shown as formula (I): Formula (I).

3. Use of the "self-assembling molecular tongs" of claim 2; said "self-assembling molecular tongs" comprising R-SL small molecule inducing ligand and hemagglutinin protein, for the preparation of a medicament for inducing cell migration, wherein, The R-SL small molecule inducible ligand comprises sialyllactose and rhodamine B; the sialyllactose and rhodamine B are linked by an acylhydrazone bond; the sialyllactose on the R-SL small molecule inducible ligand specifically recognizes hemagglutinin protein; the structure of the R-SL small molecule inducible ligand is shown as formula (I): Formula (I).

4. The use of claim 3, wherein the induced cell migration promotes wound healing; the wound includes acute wound and chronic wound; the drug for inducing cell migration specifically recognizes endogenous hemagglutinin protein through the sialyllactose on the R-SL small molecule inducible ligand, induces cell pseudopod generation, induces cell migration, and promotes wound healing.

5. A pharmaceutical composition for inducing cell migration, comprising the "self-assembling molecular tongs" combination of claim 2 and an active ingredient of another pharmaceutical for inducing cell migration, and pharmaceutically acceptable adjuvants thereof; the "self-assembling molecular tongs" comprising an R-SL small molecule inducing ligand and a hemagglutinin protein, wherein, The R-SL small molecule inducible ligand comprises sialyllactose and rhodamine B; the sialyllactose and rhodamine B are linked by an acylhydrazone bond; the sialyllactose on the R-SL small molecule inducible ligand specifically recognizes hemagglutinin protein; the structure of the R-SL small molecule inducible ligand is shown as formula (I): Formula (I).

6. The pharmaceutical composition of claim 5, wherein the induced cell migration promotes wound healing; the wound includes acute wound and chronic wound; the drug for inducing cell migration specifically recognizes endogenous hemagglutinin protein through the sialyllactose on the R-SL small molecule inducible ligand, induces cell pseudopod generation, cell migration, and promotes wound healing.

7. A hydrogel comprising the "self-assembling molecular pincer" compound of claim 2 and a hydrogel precursor solution; the hydrogel precursor solution is prepared from an organic solvent and a natural polymer, the hydrogel is formed by covalent bond cross-linking of the organic solvent; the "self-assembling molecular pincer" comprises an R-SL small molecule induced ligand and a hemagglutinin protein, wherein, The R-SL small molecule inducible ligand comprises sialyllactose and rhodamine B; the sialyllactose and rhodamine B are linked by an acylhydrazone bond; the sialyllactose on the R-SL small molecule inducible ligand specifically recognizes hemagglutinin protein; the structure of the R-SL small molecule inducible ligand is shown as formula (I): Formula (I).

8. Use of the hydrogel according to claim 7 in the preparation of a product for inducing cell migration; said inducing cell migration comprises promoting wound healing; said hydrogel comprises "self-assembling molecular tongs" compounds and a hydrogel precursor solution prepared from an organic solvent and a natural polymer, said hydrogel being formed by covalent bond cross-linking of the organic solvent; said "self-assembling molecular tongs" comprise R-SL small molecule induced ligands and hemagglutinin proteins, wherein, The R-SL small molecule inducible ligand comprises sialyllactose and rhodamine B; the sialyllactose and rhodamine B are linked by an acylhydrazone bond; the sialyllactose on the R-SL small molecule inducible ligand specifically recognizes hemagglutinin protein; the structure of the R-SL small molecule inducible ligand is shown as formula (I): Formula (I).

9. Use according to claim 8, wherein the organic solvent is selected from one or more of glycidyl methacrylate, acryloyl chloride, acryloyl chloride, acrylic acid-N- hydroxysuccinimide ester, maleic anhydride, acrylic anhydride and / or methacrylic anhydride (MA); preferably methacrylic anhydride (MA); and wherein the natural polymer is selected from one or more of polyethylene glycol, chitosan, cellulose and derivatives thereof, hyaluronic acid, silk fibroin, alginate, collagen and / or gelatin; preferably gelatin.

10. Use according to claim 8, wherein the product is selected from medical supplies and medical devices; and wherein the medical supplies and medical devices are selected from injectable surgical fillers, bio-nanomaterials, drug delivery systems and tissue repair and cell scaffolds.

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