Application of nano-composite particles in preparation of medicine for regulating ribosome activity

Abstract

The invention discloses application of nano-composite particles in preparation of a medicine for regulating ribosome activity. The bionic magnetoelectric nanoparticles capable of forming the biological effect of immune cells are prepared, under the application of an external magnetic field, the bionic magnetoelectric nanoparticles can target the immune cells and promote polarization and proliferation of the immune cells after being coated with a cell membrane, and experiments verify that the process is regulated and controlled by a transcription factor TAF9B. By adjusting the electrostatic interaction of ribosomes, the nanocomposite optimizes protein synthesis and metabolic adaptability.

Description

Technical Field

[0001] The invention belongs to the field of biomedicine, and in particular relates to the use of nanocomposite particles in preparing drugs for regulating ribosome activity. Background Art

[0002] CD4 + T helper cells can polarize into Th1 and Th2 subsets under various conditions, and the conversion from Th1 to Th2 is thought to have clinical benefits for patients with diseases in some cases. However, there is no particularly effective therapeutic strategy to achieve a stable conversion of Th1 to Th2 in vivo.

[0003] Furthermore, protein synthesis is an energy- and resource-demanding process, and thus, changes in T cell energy production are intrinsically linked to proteome remodeling. However, whether electroactive biomaterials can influence ribosomal electrostatic interactions and thereby modulate T cell function remains largely unexplored. Summary of the invention

[0004] In order to solve at least some of the technical problems in the prior art, the present invention has developed a new method of combining cell coating with a core-shell nanocomposite material with magnetoelectric response properties to target immune cells. Under external stimulation, the composite material can promote Th2 cell polarization and enhance immune cell proliferation, and it has been experimentally verified that this process is regulated by the transcription factor TAF9B. Subsequent analysis revealed the co-localization of the composite material with ribosome components, and found that it accelerates protein synthesis by regulating ribosome electrostatic interactions. Specifically, the present invention includes the following contents.

[0005] In a first aspect of the present invention, there is provided use of nanocomposite particles in the preparation of drugs for regulating ribosome activity, wherein the nanocomposite particles include a piezoelectric material layer and a magnetically responsive core structure, and the regulation includes promoting translation or protein biosynthesis.

[0006] In certain embodiments, according to the use of the present invention, the piezoelectric material layer is prepared from an inorganic piezoelectric material and / or an organic polymer piezoelectric material, and the magnetically responsive core structure is prepared from a magnetic material.

[0007] In certain embodiments, according to the use of the present invention, the particle size of the nanocomposite particles is 10-500 nm.

[0008] According to a second aspect of the present invention, a pharmaceutical composition is provided, comprising nanocomposite particles and a pharmaceutically acceptable carrier, wherein the nanocomposite particles include a piezoelectric material layer and a magnetically responsive core structure.

[0009] In certain embodiments, according to the pharmaceutical composition of the present invention, the pharmaceutically acceptable carrier comprises a membrane-forming lipid.

[0010] In certain embodiments, the pharmaceutical composition according to the present invention further comprises a ligand molecule that binds to a target cell.

[0011] The third aspect of the present invention provides a method for regulating cell activity in vitro, comprising the steps of allowing nanocomposite particles to enter cells and subjecting them to magnetic field treatment, wherein the nanocomposite particles comprise a piezoelectric material layer and a magnetically responsive core structure.

[0012] In certain embodiments, according to the method for regulating cell activity in vitro of the present invention, the regulating cell activity comprises at least one of the following situations:

[0013] (1) Regulate ribosome electrostatic interactions;

[0014] (2) regulating the local electrostatic potential within the ribosome exit tunnel;

[0015] (3) Regulate protein biosynthesis;

[0016] (4) regulating the amount or expression of transcription factors;

[0017] (5) Regulate Th1 / Th2 balance.

[0018] In certain embodiments, according to the method for regulating cell activity in vitro of the present invention, the concentration of the nanocomposite particles is less than 100 μg / mL.

[0019] In certain embodiments, according to the method for regulating cell activity in vitro of the present invention, the nanocomposite particles are coated with a membrane material and contacted with cells, so that the nanocomposite particles enter the cells.

[0020] The present invention prepares a bionic magnetoelectric nanoparticle that can shape the biological effects of immune cells, especially T cells. Under the application of an external magnetic field, the bionic magnetoelectric nanoparticle can target CD4 after being coated with the cell membrane. + T cells and promoted Th2 cell polarization and proliferation. By modulating ribosomal electrostatic interactions, the nanocomposite optimized protein synthesis and metabolic fitness. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 Preparation and characterization of DC@CFO / BFO nanoparticles.

[0022] Figure 2 Shown by CD11c + MHC II +Fluorescence activated cell sorting of cells to collect cell membranes (a); surface potential of DC@CFO / BFO and CFO / BFO nanoparticles (b); glycoprotein content on DC@CFO / BFO nanocomposites (c).

[0023] Figure 3 The in vivo targeting and safety evaluation of DC@CFO / BFO nanoparticles are shown. Figures a and b show the in vivo targeting and safety evaluation of DC@CFO / BFO nanoparticles by combining FITC-coupled CFO / BFO nanoparticles or DC@CFO / BFO nanocomposites with mouse naive CD4 + T cells were incubated and FITC+ cells were detected by flow cytometry. c FITC-conjugated CFO / BFO nanoparticles or DC@CFO / BFO nanocomposites were incubated with mouse spleen cells and FITC was detected by flow cytometry. + cell.

[0024] Figure 4 Figure 2 shows that DC@CFO / BFO nanocomposites promote Th2 cell polarization and proliferation under external magnetic field stimulation. Figure 2 shows the activation of naive T cells (CD4 + CD25 - CD62L hi CD44 lo Flow cytometric analysis of IFNγ, IL4, IL17A, FOXP3 and CD25 staining in a ) (to assess differentiation efficiency) and cultured under various polarization conditions with or without DC@CFO / BFO nanocomposites and magnetic field treatment, followed by restimulation with PMA and ionomycin. f, g show activation of CD4 with antibodies against CD3 / CD28 + T cells and induced them to differentiate into Th2 cells, and then these cells were treated with DC@CFO / BFO nanocomposites in the presence or absence of magnetic field stimulation to perform CD4 + CFSE dilution staining and flow cytometric analysis of Ki-67 expression in T cells.

[0025] Figure 5 a represents CD4 + PCA analysis of T cell transcripts. b CD4 + T cell gene expression heat map.

[0026] Figure 6 a is the CD4 + PCA analysis of T cell metabolites. b is the volcano plot analysis of the metabolites of the DC@CFO / BFO+M group and the DC@CFO / BFO group. c and d are the CD4 +Representative metabolite levels of T cells.

[0027] Figure 7 The results show that DC@CFO / BFO nanoparticles enhance protein translation in Th2 cells under magnetic field stimulation. a is a CD4 + Imaging of T cells. RPS3 and RPL7 were used to label ribosomes. Lyso-Tracker Red was used to track lysosomes. Scale bar is 5 μm. b and c show activation of CD4 T cells with antibodies against CD3 / CD28. + T cells were induced to differentiate into Th2 cells, treated with DC@CFO / BFO nanocomposites and stimulated by magnetic field. Puromycin-labeled proteins in T cells were identified by immunoblotting analysis. d and e show activation of CD4 with antibodies against CD3 / CD28 + T cells were induced to differentiate into Th2 cells, treated with DC@CFO / BFO nanocomposites and stimulated by magnetic field. Multimeric profile analysis of T cells was performed. f is a volcano plot analysis of pairwise comparison of quantitative proteomics analysis between DC@CFO / BFO+M group and DC@CFO / BFO group. g shows activation of CD4 with antibodies against CD3 / CD28 + T cells and induced to differentiate into Th2 cells were treated with DC@CFO / BFO nanocomposites and subjected to magnetic field stimulation. The theoretical isoelectric point (pI) and hydrophilicity of upregulated proteins, downregulated proteins, and insignificant proteins are shown, respectively.

[0028] Figure 8 a shows the CD4 expression in different treatment groups under Th2 differentiation conditions + PCA analysis of T cell proteome. b shows the CD4 + Proteome heatmap of T cells under Th2 differentiation conditions.

[0029] Fig. 9 Figure 4 shows the role of TAF9B in mediating magnetoelectric nanoparticle-mediated T cell responses. a shows activation of CD4 with antibodies against CD3 / CD28. + T cells and induced them to differentiate into Th2 cells, and then these cells were treated with DC@CFO / BFO nanocomposites in the presence or absence of magnetic field stimulation. +GSEA of genes expressed in T cells. b is the mRNA level of TAF9B in each fraction derived from the multimer spectrum detected by quantitative real-time PCR. c is the mRNA level of wild-type TAF9B and mutant TAF9B in each fraction derived from the multimer spectrum detected by quantitative real-time PCR. d shows the activation of naive CD4 + T cells were transfected with retrovirus containing TAF9B siRNA. T cells were then cultured under Th2 polarization conditions with or without DC@CFO / BFO nanocomposites and magnetic field treatment, and then restimulated with PMA and ionomycin. CD4 + IL4 + e shows the activation of naive CD4 + T cells were transfected with retrovirus containing TAF9B siRNA. T cells were then cultured under Th1 polarization conditions with or without DC@CFO / BFO nanocomposites and magnetic field treatment, and then restimulated with PMA and ionomycin. CD4 + IFNγ + f shows the activation of naive CD4 + T cells were transfected with retrovirus containing TAF9B siRNA. T cells were then cultured under Th2 polarization conditions with or without DC@CFO / BFO nanocomposites and magnetic field treatment. The expression level of Ki-67 in each group was detected by flow cytometry.

[0030] Fig.10 a is the expression level of TAF9B in the specified immune cell subsets analyzed by The Human Protein Atlas database. b is the quantitative proteomics analysis of CD4 + c shows the protein level of TAF9B in T cells. + Immunoblot analysis of TAF9B expression in T cells. d shows CD4 + T cells were activated with plate-bound anti-CD3 and anti-CD28 antibodies and transfected with retrovirus containing TAF9B siRNA. T cells were then cultured under Th17 polarizing conditions with or without the addition of DC@CFO / BFO nanocomposites and magnetic field treatment, followed by restimulation with PMA and ionomycin. Flow cytometry analysis of CD4 + Il17A + e shows the cell ratio CD4 + T cells were activated with plate-bound anti-CD3+anti-CD28 antibodies and transfected with retrovirus containing TAF9B siRNA. T cells were then cultured under Treg polarization conditions and treated or not with DC@CFO / BFO nanocomposites and magnetic field stimulation, and the proportion of iTregs in each group was analyzed by flow cytometry. DETAILED DESCRIPTION

[0031] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but should be understood as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0032] It should be understood that the terms described in the present invention are only for describing special embodiments and are not intended to limit the present invention. In addition, for the numerical range in the present invention, it should be understood that the upper and lower limits of the scope and each intermediate value therebetween are specifically disclosed. Each smaller range between the intermediate value in any stated value or stated range and any other stated value or intermediate value in the described range is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the scope.

[0033] Unless otherwise indicated, all technical and scientific terms used herein have the same meanings as commonly understood by those skilled in the art to which the invention belongs. Although the present invention describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials related to the documents. In the event of a conflict with any incorporated document, the content of this specification shall prevail.

[0034] use

[0035] One aspect of the present invention provides the use of nanocomposite particles in the preparation of drugs for regulating ribosome activity, wherein the nanocomposite particles include a piezoelectric material layer and a magnetically responsive core structure, and the regulation includes promoting translation or protein biosynthesis.

[0036] Herein, "magnetic response" means that the core-shell structure in the nanoparticles of the present invention can produce relative displacement under the action of an external magnetic field, wherein the core structure of the nanoparticles will stretch or shorten along the magnetization direction when magnetized in the magnetic field. "Piezoelectric performance" means that the shell structure of the nanoparticles of the present invention can convert the mechanical energy generated by the core structure into electrical energy.

[0037] In a preferred embodiment, the magnetic field strength of the external magnetic field is 0.01-300 mT. The magnetic field strength is also preferably 0.1-200 mT, further preferably 0.5-100 mT, more preferably 0.5-50 mT, and most preferably 0.5-10 mT, such as 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10 mT. In a preferred embodiment, the magnetic field is applied at the above magnetic field strength once a day, 15-30 minutes each time, for 3 consecutive days.

[0038] In the present invention, the shell structure with piezoelectric properties is prepared from inorganic piezoelectric materials and / or organic piezoelectric materials. In a preferred embodiment, the shell structure with piezoelectric properties is prepared from inorganic piezoelectric materials. In a more preferred embodiment, the inorganic piezoelectric material is selected from at least one of ferrites, niobic acids, titanates, silicates and aluminates. In a most preferred embodiment, the shell structure with piezoelectric properties is prepared from ferrite materials. In a specific embodiment, the shell structure with piezoelectric properties is bismuth ferrite.

[0039] In the present invention, the core structure with magnetic response performance is prepared from a magnetic material, wherein the magnetic material has the following structure: a Fe 1-a O b , wherein N includes at least one of Co, Mn, Zn and Ni, a is a number from 0.25 to 0.75, and b is 3. In a specific embodiment, the core structure having magnetic response performance is cobalt ferrite.

[0040] It is understood that the nanocomposite particles of the present invention may or may not contain elements doped therein. When containing elements doped therein, the elements doped therein include, but are not limited to, at least one of carbon, nitrogen, phosphorus, sulfur, silicon, aluminum, iron, titanium, nickel, manganese, copper, silver and zinc, thereby improving the piezoelectric properties and / or magnetic response properties of the nanocomposite particles.

[0041] In a specific embodiment, the nanocomposite particles are cobalt ferrite-bismuth ferrite nanoparticles.

[0042] In the present invention, the average particle size of the cobalt ferrite-bismuth ferrite nanoparticles is 10-500nm, preferably 50-400nm, also preferably 100-300nm, further preferably 150-300nm, for example 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300nm.

[0043] The present invention does not particularly limit the preparation of nanocomposite particles. The shell structure with piezoelectric properties and the core structure with magnetic response properties can be prepared by methods known in the art. It is understood that any doping elements or other layer structures (such as graphene layers, etc.) can be contained between the shell structure and the core structure, the outer layer of the shell structure, and the core structure to improve the piezoelectric properties and / or magnetic response properties of the nanoparticles. The specific preparation method of the nanocomposite particles can refer to the steps disclosed in CN117919206A, the contents of which are incorporated herein by reference in their entirety.

[0044] In a preferred embodiment, the nanocomposite particles further include a membrane material coating the nanocomposite particles. "Coating", "coating" and "modification" are used interchangeably, and refer to a composite material having highly controllable biological functions obtained by coating natural or biomimetic cell membrane materials on the surface of synthetic nanocomposite particles. The membrane material can be an artificially synthesized cell membrane or a naturally occurring cell membrane, which is not particularly limited. Preferably, the cell membrane material used in the present invention is a cell membrane separated from immune cells (such as but not limited to the cell membranes of dendritic cells, B lymphocytes and macrophages). In a specific embodiment, the cell membrane is separated from dendritic cells.

[0045] In order to make the nanocomposite particles have good targeting, the outer surface of the membrane material may contain protein molecules. In a preferred embodiment, the protein molecules include at least one binding molecule that can bind to an immune cell, preferably a T cell surface. In the present invention, the T cell surface contains at least one of the following antigens that can bind to the binding molecule: CD1, CD2, CD3, CD5, CD7, CD8, CD16, CD25, CD26, CD27, CD28, CD30, CD38, CD39, CD40L, CD44, CD45, CD62L, CD69, CD73, CD80, CD83, CD86, CD95, CD103, CD119, CD126, CD150, CD153, CD154, CD161, CD183, CD223, CD254, CD275, CD45RA, CXCR3, CXCR5, FasL, IL18R1, CTLA-4, OX40, GIT R, LAG3, ICOS, PD-1, leu-12, TCR, TLR1, TLR2, TLR3, TLR4, TLR6, NKG2D, CCR, CCR1, CCR2, CCR4, CCR6 and CCR7.

[0046] In a preferred embodiment, the protein molecule is capable of targeting CD4 + MHC II complex of T cells.

[0047] In a preferred embodiment, the purpose of the present invention is achieved by administering a therapeutically effective amount of nanocomposite particles and a magnetic field of 0.01-300 mT to a subject in need. The term "subject" used in the present invention refers to any animal (such as a mammal), including but not limited to humans, non-human primates, rodents, and the like who are about to receive a specific treatment. Generally, "subject" and "patient" are used interchangeably in the present invention and both refer to the subject.

[0048] As used herein, the terms "administer", "apply", or "administering" are used interchangeably and refer to implanting, absorbing, ingesting, injecting, inhaling, or otherwise introducing nanocomposite particles or compositions containing the same.

[0049] The term "effective amount" as used herein refers to the amount of a drug or agent that induces a biological or pharmaceutical response in a tissue, system, animal or human being, such as that sought by a researcher or clinician. The administration method of the magnetostrictive piezoelectric nanoparticles of the present invention or a pharmaceutical composition containing the same is not particularly limited, and representative administration methods include but are not limited to: intramuscular injection, intravenous injection, intravenous drip or intraperitoneal injection.

[0050] Since the nanocomposite particles of the present invention can regulate the activity of immune cells, especially the biological functions of T cells, the nanocomposite particles of the present invention can be used to prepare drugs for treating or improving T cell-mediated immune system diseases, including but not limited to, for example, gastrointestinal inflammation and / or autoimmune arthritis caused by T cells.

[0051] Pharmaceutical composition

[0052] One aspect of the present invention provides a pharmaceutical composition comprising nanocomposite particles and a pharmaceutically acceptable carrier, wherein the nanocomposite particles include a piezoelectric material layer and a magnetically responsive core structure. In a preferred embodiment, the pharmaceutically acceptable carrier includes a film-forming lipid. Further preferably, the film-forming lipid includes a ligand molecule that binds to a target cell. The ligand molecule can bind to an immune cell, preferably an antigen on the surface of a T cell. In the present invention, the surface of the T cell contains at least one of the following antigens that can bind to the ligand molecule: CD1, CD2, CD3, CD5, CD7, CD8, CD16, CD25, CD26, CD27, CD28, CD30, CD38, CD39, CD40L, CD44, CD45, CD62L, CD69, CD73, CD80, CD83, CD86, CD95, CD103, CD119, CD126, CD150, CD153, CD154, CD161, CD183, CD223, CD254, CD275, CD45RA, CXCR3, CXCR5, FasL, IL18R1, CTLA-4, OX40, GIT R, LAG3, ICOS, PD-1, leu-12, TCR, TLR1, TLR2, TLR3, TLR4, TLR6, NKG2D, CCR, CCR1, CCR2, CCR4, CCR6 and CCR7.

[0053] Methods for regulating cell activity in vitro

[0054] One aspect of the present invention provides a method for regulating cell activity in vitro, comprising the steps of allowing nanocomposite particles to enter cells and subjecting them to magnetic field treatment, wherein the nanocomposite particles comprise a piezoelectric material layer and a magnetically responsive core structure.

[0055] In a preferred embodiment, the method comprises:

[0056] a. preparing nanocomposite particles including a piezoelectric material layer and a magnetically responsive core structure;

[0057] b. contacting the nanocomposite particles with cells in the presence of a magnetic field of 0.01-300 mT.

[0058] In step a of the present invention, cobalt ferrite CoFe can be synthesized by hydrothermal method and sol-gel method. 2 O 4 For the core, bismuth ferrite BiFeO 3 For preparing nanocomposite particles with shells, the specific parameters and reaction conditions of the sol-gel method are not particularly limited and can be adjusted according to actual needs.

[0059] In step b of the present invention, the cells refer to immune cells, especially CD4+ T cells. CD4 + T cells can contain the nanocomposite particles by endocytosis. In a preferred embodiment, in order to target CD4 + T cells, the nanocomposite particles are further coated with cell membranes using cell membrane coating technology, wherein the outer surface of the cell membrane contains protein molecules. Those skilled in the art are familiar with how to make the outer surface of the cell membrane contain or express proteins that can target and bind to CD4 + Protein molecules that are antigens on the surface of T cells.

[0060] In a preferred embodiment, the membrane material is continuously passed through a 300-500nm, preferably 320-480nm, also preferably 340-460nm, more preferably 360-440nm, for example 360, 370, 380, 390, 400, 410, 420, 430, 440nm porous membrane (for example, a polycarbonate porous membrane), and then extruded through a 100-300nm, preferably 120-280nm, also preferably 140-260nm, more preferably 160-240nm, for example 160, 170, 180, 190, 200, 210, 220, 230, 240nm porous membrane to the nanoparticles obtained in step (1) to obtain cell membrane-coated nanoparticles.

[0061] In the present invention, the source of the membrane structure is not particularly limited, as long as its surface contains protein molecules that can bind to the surface of T cells. Examples of membrane structures include, but are not limited to, naturally occurring biological membranes or artificially synthesized membrane structures. In a preferred embodiment, the biological membrane is a cell membrane separated from immune cells, examples of which include, but are not limited to, a cell membrane separated from dendritic cells, B lymphocytes or macrophages. In a preferred embodiment, the biological membrane is a cell membrane of dendritic cells.

[0062] In the method of the present invention, the concentration of the nanocomposite particles is less than 100 μg / mL, preferably less than 90 μg / mL, such as 10-90 μg / mL, or 10-80 μg / mL, such as 10, 20, 30, 40, 50, 60, 70, 80 μg / mL.

[0063] In the present invention, the regulation of cell activity includes at least one of the following situations:

[0064] (1) Regulate ribosome electrostatic interactions;

[0065] (2) regulating the local electrostatic potential within the ribosome exit tunnel;

[0066] (3) regulate (especially promote or accelerate) protein biosynthesis;

[0067] (4) regulating (particularly increasing or upregulating) the amount or expression of a transcription factor (TAF9B);

[0068] (5) Regulate the balance of Th1 / Th2 cells, especially promote the differentiation of T cells into Th2 cell subsets or increase the amount of Th2 cells or promote the proliferation of Th2 cells.

[0069] In the present invention, ribosome electrostatic interaction, local electrostatic potential in the ribosome exit tunnel, the amount of protein, the amount or expression level of transcription factors, and the amount of Th1 / Th2 cells can be determined by any method known in the art, without particular limitation. The determination method includes, but is not limited to, chemiluminescent immunoassay, enzyme-linked immunosorbent assay, real-time fluorescence quantitative PCR, gene chip technology, flow cytometry, or immunoblotting.

[0070] Example

[0071] 1. Experimental Materials

[0072] C57BL / 6 mice were purchased from Vital River Laboratory Animal Technology. All animals were housed and maintained under specific pathogen-free conditions. All experiments were performed according to protocols approved by the Ethics Committee of Peking University Health Science Center (approval number PUIRB-LA2024005).

[0073] 2. Experimental Methods

[0074] 2.1CoFe 2 O 4 Preparation of nanoparticles

[0075] 0.092 M FeCl 3 , 0.046M CoCl 2 and 0.14 M CTAB were dissolved in deionized water. Then, 6 M NaOH was added to the above solution under mechanical stirring and ultrasonic dispersion, and the resulting solution was transferred to a sealed polytetrafluoroethylene hydrothermal reactor and heated at 200 ° C for 24 hours. The obtained black powder was then washed with deionized water and ethanol and dried at 95 ° C overnight.

[0076] 2.2 Core-shell nanostructured magnetoelectric composite CoFe 2 O 4 -BiFeO 3 Preparation

[0077] 0.011M Bi(NO 3 )·5H 2 O and 0.01 M Fe(NO 3 )·9H 2O was dissolved in ethylene glycol to prepare the BFO precursor. Then 0.15 g of dried CFO nanoparticles were dispersed into 100 mL of the BFO precursor solution and sonicated for 3 h. The solution was then dried overnight, and the dried powder was subsequently annealed at 600 °C for 2.5 h at a heating rate of 10 °C / min. The annealed powder was sonicated for 30 min and filtered through a filter membrane to obtain the CFO / BFO nanocomposite.

[0078] 2.3 Cell membrane-coated nanocomposite particles

[0079] To obtain dendritic cell (DC) membrane-coated CFO / BFO nanocomposites, bone marrow from mouse femur and tibia was first isolated and DC differentiation was induced using 20 ng / ml recombinant GM-CSF and 10 ng / ml recombinant IL4. DCs (CD11c + MHC II + cell).

[0080] The isolated DCs were processed by freezing and thawing in liquid nitrogen for 3 cycles, and the lysates were sonicated for 5 min by a water bath sonicator to collect the cell membranes, which were then continuously extruded through 400 nm and then 200 nm polycarbonate porous membranes to the magnetoelectric core-shell CFO / BFO nanoparticles to obtain DC@CFO / BFO nanocomposite particles.

[0081] 2.4 Characterization

[0082] The surface morphology of the nanoparticles was characterized by scanning electron microscopy, and the nanoparticles and cell membrane-coated nanoparticle composites were observed using high-resolution transmission electron microscopy. The prepared materials were examined by energy dispersive X-ray spectroscopy (EDS) or X-ray diffraction spectroscopy with TEM. Piezoelectric response force microscopy (PFM) measurements were recorded by a commercial atomic force microscope equipped with a ferroelectric test system. Measurements were performed with a vibrating sample magnetometer (VSM) on a Quantum Design PPMS-9DynaCool system.

[0083] 1 mg of nanoparticles was placed in ddHO 2 O or DMSO, respectively, to detect ·OH and ·O 2 - 5,5-Dimethyl-1-pyrroline N-oxide (DMPO) was added to the solution and the reactive species generated from the nanocomposites were detected by electron paramagnetic resonance (EPR) technique. The size and surface potential of the nanoparticles were measured by Zeta Sizer Nano-ZS instrument at room temperature in an aqueous environment.

[0084] 2.5 Immunoblotting analysis

[0085] The cells were lysed using RIPA lysis buffer (Tris (pH 7.4), NaCl, Triton X-100, sodium deoxycholate, SDS) supplemented with a protease inhibitor cocktail and subjected to SDS-PAGE.

[0086] The antibodies used in this example include: anti-CD11b antibody (abcam, ab133357, 1:1000), anti-puromycin antibody (Merck-millipore, MABE341, 1:1000), and anti-TAF9B antibody (Signalwayantibody, 45238, 1:1000).

[0087] 2.6 Sample preparation and flow cytometry

[0088] Immune cells from spleen or lymph nodes were isolated by grinding the tissue in PBS containing FBS and then filtering through a cell strainer.

[0089] For lamina propria mononuclear cell (LPMC) isolation, mouse intestines were removed and cut into pieces. The pieces were then incubated in HBSS. Intestinal epithelial cells (IEC) were removed and passed through a cell strainer. The small pieces of tissue were then placed in digestion solution and slowly spun, and after passing through the strainer, lamina propria lymphocytes were isolated.

[0090] To analyze the surface proteins of DC@CFO / BFO nanocomposites, DC@CFO / BFO nanocomposites were incubated with specific antibodies at room temperature and analyzed by flow cytometry.

[0091] To detect the cellular uptake of nanoparticles, CFO / BFO nanoparticles and DC@CFO / BFO nanocomposites were mixed with the same mass of FITC aqueous solution and stirred. After removing the excess FITC, the FITC-conjugated nanoparticles were incubated with mouse naive T cells or splenocytes, and then FITC was detected by flow cytometry. + cell.

[0092] To detect T cell proliferation, isolated T cells were stained at room temperature. Cell activation and proliferation were then induced and collected for flow cytometry analysis.

[0093] To analyze cell surface marker expression (CD45, CD4, CD8, CD25, B220, CD11b, Ly6G, CD11c, MHC II, F4 / 80), cells were incubated with specific antibodies at room temperature. For intracellular marker staining (IFNγ, IL4, IL17A), cells were treated with a protein transport inhibitor cocktail (eBioscience, 00-4980-03), then fixed and permeabilized, followed by staining with specific antibodies. To detect the expression of nuclear proteins (Ki-67, FOXP3), cells were fixed and permeabilized, and then stained with specific antibodies.

[0094] The following antibodies were used: anti-CD45 antibody (eBioscience, 30-F11), anti-CD4 antibody (BioLegend, GK1.5), anti-CD8 antibody (eBioscience, 53-6.7), anti-CD25 antibody (BioLegend, 3C7), anti-B220 antibody (BioLegend, RA3-6B2), anti-CD11b antibody (BioLegend, M1 / 70), anti-Ly6G antibody (BioLegend, RB6-8C5), anti-CD11c antibody (BioLegend, N418), anti-F4 / 80 antibody (BioLegend, BM8), and anti-MHC antibody (BioLegend, 407). II antibody (BioLegend, IA / IE)(M5 / 114.15.2), anti-IFNγ antibody (BioLegend, XMG1.2), anti-IL4 antibody (BioLegend, 11B11), anti-IL17A antibody (BioLegend, eBio17B7), anti-Ki-67 antibody (BioLegend, 16A8) and anti-FOXP3 antibody (eBioscience, FJK-16s).

[0095] 2.7 Confocal microscopy

[0096] CD4 + T cells were incubated with FITC-labeled CFO / BFO nanoparticles. Cells were fixed and blocked, then stained with specific antibodies and DAPI. Images were acquired using a Nikon TCS A1 microscope. Antibodies against RPS3 (abcam, ab128996, 1:100) and RPL7 (abcam, ab72550, 1:500) were used to label ribosomes. Lyso-Tracker Red (Solarbio, L8010) was used to track lysosomes.

[0097] 2.8 In vitro T cell differentiation

[0098] Isolation of mouse naive CD4 from peripheral lymph nodes by flow cytometry + T cells (CD4 + CD25 - CD62L hi CD44 lo ), and then activated with anti-CD3 antibody and anti-CD28 antibody. For Th1 polarization, anti-IL4 antibody and IL-12 were used. For Th2 polarization, anti-IFN-γ antibody and IL4 were used. For Th17 polarization, IL-6, TGF-β, anti-IFN-γ antibody and anti-IL4 antibody were used.

[0099] For iTreg cell differentiation, TGF-β and IL-2, anti-IFN-γ antibody and anti-IL4 antibody were used. T cells were incubated with 10 μg / mL DC@CFO / BFO nanoparticles and treated with a 1.5 mT magnetic field for 30 minutes every day. After 48 or 72 hours of culture, cells were collected for flow cytometry analysis.

[0100] 2.9 Multimer Spectrum Analysis

[0101] Isolation of CD4 + T cells were incubated with magnetoelectric nanocomposites and magnetic field treatment under Th2 differentiation conditions and then treated with cycloheximide (CHX). The cells were resuspended in hypotonic buffer and passed through a 23-gauge needle three times.

[0102] The resulting homogenate was centrifuged to clarify the lysate. After centrifugation, the supernatant was transferred to a new tube and the absorbance was measured at 260nm. The sample was then diluted with lysis buffer to the same absorbance value of about 15-20 units / mL, and then the sample was loaded into a linear sucrose gradient (10-50%). The sample was then centrifuged. By a piston gradient classifier (BioCompInstruments; connected to an EM-1 type Econo UV monitor (BioRad), to continuously measure at an absorbance wavelength of 254nm. The flowing liquid was collected in sequence, and then quantitative real-time PCR was performed to detect the TAF9B mRNA level in each fraction.

[0103] 2.10 Enzyme-linked immunosorbent assay

[0104] ELISA assays for glycoproteins on cell membranes or nanoparticles were performed according to the manufacturer's instructions (ELISA Laboratories, JYM0538Mo).

[0105] Type 2 collagen (anti-CII) specific antibodies in serum collected from immunized mice were measured by ELISA. Serum samples were added to plates pre-coated with CII, followed by goat antibodies against mouse IgM, IgG1, IgG2a, IgG2b, IgG3, and IgA.

[0106] 2.11 RNA sequencing

[0107] To investigate the role of magnetoelectric nanocomposites in T cell-mediated responses, CD4 + T cells were incubated with magnetoelectric nanocomposites and subjected to magnetic field treatment under Th2 differentiation conditions. Total RNA was then purified. RNA-seq libraries were constructed using a library preparation kit and sequenced on an Illumina platform. Analytical data were obtained by data quality control according to the manufacturer's instructions. Based on the gene model annotation file, the reads were mapped to the reference genome Hisat2 (version 2.0.5). Based on the length of the gene and the read counts aligned to the gene, the reads of the transcript per kilobase fragments per million mapped reads (FPKM) of each gene were calculated. The overlap of the gene set from RNAseq data with the selected data sets (C5.all.v 6.2, H.all.v 6.2) in MSigDB was analyzed using http: / / software.broadinstitute.org / gsea / index.jsp.

[0108] 2.12 Quantitative proteomic analysis

[0109] Isolation of CD4 + T cells were then incubated with the magnetoelectric nanocomposites and an applied magnetic field under Th2 differentiation conditions. 100 μg of protein samples from each subject were analyzed by LC-MS / MS by timsTOF Pro2.

[0110] 2.13 Non-targeted metabolomics analysis

[0111] Isolation of CD4 + T cells were then incubated with magnetoelectric nanocomposites and an applied magnetic field under Th2 differentiation conditions. To extract the metabolites of the T cells, the cell pellets were resuspended in methanol and frozen in liquid nitrogen. These quenched cells were then thawed and vortexed. After centrifugation, the supernatant was freeze-dried under vacuum. The dried samples were incubated in methanol. The samples were then centrifuged and the supernatant was used for LC-MS / MS analysis.

[0112] 3. Experimental results

[0113] 3.1 Preparation and characterization of film-coated magnetoelectric nanoparticles

[0114] This example prepared a specific targeting CD4 + Magnetoelectric nanoparticles that regulate T cell differentiation. Figure 1 The preparation and characterization of DC@CFO / BFO nanoparticles are shown. Figure 1 a is a representative SEM image of CFO / BFO nanoparticles. b is a representative TEM image of CFO / BFO nanoparticles. The surface morphology and core-shell structure of CFO / BFO nanocomposites were confirmed by SEM and TEM characterization. c is a high angle annular dark field (HAADF) and corresponding EDS element mapping image of CFO / BFO nanoparticles. d is the VSM result of CFO / BFO nanocomposites. e is the piezoelectric response amplitude curve and phase curve of CFO / BFO nanocomposites. f and g respectively show the free radicals ·OH and ·O generated by CFO / BFO nanocomposites under an applied magnetic field of 1.5mT. 2 - h is the electron paramagnetic resonance result of DC@CFO / BFO nanocomposite. i is the TEM image of DC@CFO / BFO nanoparticles. j is the immunoblot analysis of surface proteins of DC@CFO / BFO nanoparticles. k is the nano-flow cytometry analysis of CD11c and MHC class II expression levels on the surface of DC@CFO / BFO nanoparticles.

[0115] XRD spectroscopy verified the composition of CFO / BFO nanocomposites and confirmed that CoFe 2 O 4 and BiFeO 3 EDS reveals the uniform distribution of elements in STEM analysis ( Figure 1 c). In addition, VSM analysis demonstrated the excellent responsiveness of CFO / BFO nanocomposites to external magnetic fields, as evaluated by initial magnetization and magnetic saturation intensity ( Figure 1 d). PFM reveals detectable localized piezoelectric response hysteresis loops at random locations of the nanoparticles, characterizing their ferroelectric and magnetoelectric properties ( Figure 1 In addition, the oxides produced in magneto-electrocatalysis were detected using the EPR spectrum of DMPO, and signals were observed in the CFO / BFO nanocomposites under an external magnetic field ( Figure 1 f, g).

[0116] Due to the piezoelectric property of BFO, magnetoelectric CFO / BFO nanocomposites and BFO nanoparticles can generate ·OH and ·O in the presence of external ultrasound. 2 -The characteristic signal of the CFO core was detected, while the nanoparticles alone could not trigger the generation of ROS. These findings therefore elucidate the role of the magnetic field in inducing magnetostrictive strain in the CFO core and initiating potential polarization in the BFO shell, thereby confirming the effective magnetoelectric conversion.

[0117] To achieve CD4 + T cells target specificity, selecting bone marrow-derived dendritic cells (BMDC) and binding to CD4 by controlling their major histocompatibility complex (MHC) II expression. + In brief, mouse bone marrow cells were first isolated and induced to differentiate into dendritic cells using GM-CSF and IL4. CD11c + MHC II + Fluorescence activated cell sorting (FACS) Figure 2 a) and collect the cell membranes. Then, membrane-coated magnetoelectric nanocomposites (DC@CFO / BFO) were prepared by repeated extrusion of CFO / BFO nanoparticles with freshly extracted BMDC membranes ( Figure 1 h). As shown in TEM, a transparent cell membrane layer of about 10 nm was observed on the surface of the membrane-coated CFO / BFO nanoparticles ( Figure 1 i). Dynamic light scattering experiments showed that the size of the DC@CFO / BFO composite material after coating was about 170nm. Then zeta potential measurements showed that the surface charge of the DC@CFO / BFO nanocomposite was less negative than that of CFO / BFO nanoparticles ( Figure 2 b). Subsequent Western blot and nano-flow cytometry analysis confirmed that the DC@CFO / BFO nanocomposites displayed surface markers of DCs, indicating that native BMDC membranes were successfully coated and retained onto the CFO / BFO surface ( Figure 1 In addition, by quantification of glycoproteins, it was found that the glycoprotein content on the DC@CFO / BFO nanocomposites was approximately 90% of that in the free BMDC membrane, indicating that the membrane had the correct orientation when coated onto the nanoparticles ( Figure 2 c).

[0118] In general, this example successfully prepared a dendritic cell membrane-coated magnetoelectric nanocomposite material that exhibited excellent magnetic field responsiveness and magnetostrictive piezoelectric properties.

[0119] 3.2 Biosafety assessment and biodistribution profile

[0120] Using CCK-8 assay, mouse naive CD4 +The cytotoxicity of DC@CFO / BFO nanocomposites with or without magnetic field stimulation was evaluated by T cells, and it was found that DC@CFO / BFO nanocomposites exhibited good biocompatibility at concentrations below 100 μg / mL.

[0121] Next, CD4 + T cell uptake of nanocomposites. FITC-labeled DC@CFO / BFO or CFO / BFO nanoparticles were incubated with naive mouse CD4 + T cells were incubated and the fluorescence signal was assessed by flow cytometry. Figure 3 As shown in a and b, DC membrane coating accelerated the T cell phagocytosis of the nanocomposite, which started at about 2 hours and reached a peak at 8 hours. After 12 hours of co-incubation of the nanocomposite and mouse spleen cells, most of the DC@CFO / BFO nanocomposite was engulfed by CD4 + T cell selection, while most CFO / BFO nanoparticles not coated by DC membranes were taken up by macrophages ( Figure 3 c). Therefore, these data suggest that encapsulation of DC membranes on CFO / BFO nanoparticles increases their targeting specificity and promotes CD4 + Its uptake by T cells.

[0122] To investigate the biodistribution characteristics of the nanocomposites in vivo, FITC-labeled DC@CFO / BFO nanoparticles were administered to C57BL / 6 mice by intravenous injection. In vivo imaging showed that the nanoparticles were enriched in the liver, kidneys, lungs, and spleen and remained in the body for 4 days after injection. In addition, the DC@CFO / BFO nanocomposites had negligible effects on the functional parameters of the liver and kidneys, and no significant abnormalities were observed in major organs by histological analysis. Therefore, the DC@CFO / BFO nanocomposites have excellent safety in vivo.

[0123] 3.3DC@CFO / BFO nanoparticles promote Th2 cell polarization and proliferation under external magnetic field

[0124] In order to further study the DC@CFO / BFO nanocomposites in CD4 + Role in T cell activation and differentiation, activation of CD4 with anti-CD3 and anti-CD28 antibodies under various helper T cell polarization conditions + Flow cytometry analysis showed that the production of IL4 was enhanced, while the production of IFNγ was reduced in T cells treated with the external magnetic field (DC@CFO / BFO nanocomposite), indicating that the combination of magnetoelectric nanocomposite and magnetic field promoted Th2 differentiation and inhibited Th1 differentiation ( Figure 4In addition, T cells produced less IL17A in the DC@CFO / BFO group and the DC@CFO / BFO+M group, and the proportion of induced Treg cells (iTreg) in all groups did not differ significantly ( Figure 4 a, d and e).

[0125] To more precisely characterize the role of magnetoelectric nanocomposites in Th2 polarization, RNA sequencing was performed to analyze the transcriptome of T cells treated with magnetoelectric nanocomposites and magnetic fields under Th2 differentiation conditions. Principal component analysis (PCA) and differentially expressed genes (DEGs) analysis showed that T cells in the DC@CFO / BFO+M group were clearly distinguishable from T cells without magnetoelectric nanoparticles or magnetic field stimulation by their transcriptomes ( Figure 5 Subsequent GO enrichment showed that genes related to “DNA replication”, “T cell differentiation”, and “T cell proliferation” were overexpressed among the significantly upregulated genes in T cells under Th2 cell polarization conditions (vector group and DC@CFO / BFO group) relative to naive T cells, indicating that favorable T cell polarization was induced.

[0126] Subsequently, the overall gene expression changes of T cells treated by DC@CFO / BFO in the presence or absence of a magnetic field were analyzed, and 898 upregulated genes and 1703 downregulated genes were found in T cells after DC@CFO / BFO+M treatment. Compared with T cells in the DC@CFO / BFO group, the upregulated genes were enriched in "Type II immune response" and "Th2 cell differentiation", reflecting the stimulatory effects of DC@CFO / BFO treatment and magnetic field stimulation on Th2 cell polarization. In addition, DC@CFO / BFO nanocomposites promoted signaling pathways related to T cell proliferation when loaded with a magnetic field. Subsequent flow cytometric analysis also confirmed that under Th2 cell polarization conditions, CD4 + T cells responded to DC@CFO / BFO+M treatment and underwent more cell divisions, as detected by CFSE dilution assay and Ki-67 staining ( Figure 4 f, g). Taken together, these findings indicate that treatment with magnetoelectric nanocomposites combined with magnetic field stimulation promoted the induction of Th2 cell polarization and enhanced T cell proliferation in vitro.

[0127] 3.4 DC@CFO / BFO nanoparticles regulate the amino acid metabolism pathway of Th2 cells under magnetic field stimulation

[0128] Rapid T cell proliferation was always associated with optimal metabolic fitness, and genes related to cell metabolism were found to be enriched in T cells from the DC@CFO / BFO+M group. +The metabolic status of T cells was analyzed by non-targeted metabolomics analysis, which revealed extensive changes in T cell metabolic processes induced by DC@CFO / BFO+M ( Figure 6 Differential metabolite analysis showed that the combination of DC@CFO / BFO nanoparticles and magnetic field stimulation enhanced cellular metabolism, resulting in 398 upregulated metabolites and 61 downregulated metabolites ( Figure 6 b). It is noteworthy that amino acid metabolism pathways such as “tyrosine metabolism” and “glycine, serine, and threonine metabolism” act as central hubs in the T cell metabolic network. Representative differential metabolites such as Figure 6 Taken together, these findings suggest that the synergistic effects of magnetoelectric nanocomposite treatment and magnetic field stimulation enhance CD4 + Metabolic adaptability of T cells characterized by enhanced amino acid metabolism.

[0129] 3.5DC@CFO / BFO nanoparticles target ribosomes and enhance protein synthesis in Th2 cells

[0130] CD4 + Transcriptome profiling of T cells revealed enrichment of the “cytoplasmic translation” and “cytoplasmic ribosome” pathways after treatment with DC@CFO / BFO nanocomposites and magnetic field stimulation. The subcellular localization of CFO / BFO was subsequently examined by confocal microscopy. In addition to lysosomal localization, FITC-labeled CFO / BFO also exhibited colocalization with ribosomal components such as RPL7 and RPS3 ( Figure 7 a). The ribosome is a complex molecular machine responsible for protein synthesis in all living cells. It was observed that the DC@CFO / BFO nanocomposite under an external magnetic field resulted in accelerated protein synthesis ( Figure 7 In addition, the multimer profile revealed an increase in multimers in the DC@CFO / BFO+M group, indicating an enhanced translation rate ( Figure 7 d, e).

[0131] To characterize the potential mechanisms of translational regulation mediated by magnetoelectric nanocomposites, quantitative proteomics studies were performed on T cells treated with DC@CFO / BFO nanocomposites and stimulated by an external magnetic field. PCA demonstrated clear differences in the protein expression profiles of the DC@CFO / BFO+M group compared with the other two groups ( Figure 8 Cluster analysis using heatmap visualization indicated that DC@CFO / BFO+M treatment provided heterogeneity with significant changes in initiating protein expression ( Figure 8b). Analysis of differentially expressed proteins (DEPs) revealed 573 upregulated proteins and 306 downregulated proteins in T cells treated with DC@CFO / BFO nanocomposites and subjected to magnetic field stimulation ( Figure 7 f). Further GO enrichment analysis showed that pathways related to “oxidative phosphorylation”, “IL4 production”, and “translation” were activated in the DC@CFO / BFO+M group, which is consistent with the finding that DC@CFO / BFO nanoparticles plus magnetic field promote Th2 cell polarization and optimize T cell metabolic adaptability. Considering the important role of protein physicochemical properties in protein synthesis rate, the isoelectric point (pI) and hydrophilicity of up-regulated proteins, down-regulated proteins, and proteins whose expression was not affected were analyzed separately. Figure 7 As shown in Figure 5, higher values ​​of pI were observed for upregulated proteins compared to downregulated or unaffected proteins after treatment with DC@CFO / BFO nanoparticles and magnetic field stimulation. Electrostatic interactions at the ribosome exit tunnel are associated with nascent protein ejection time, and proteins with positively charged residues at the C-terminus tend to eject more slowly. Therefore, DC@CFO / BFO nanocomposites can directly target cytoplasmic ribosomes and modulate electrostatic interactions at the peptide exit tunnel under magnetic field stimulation, thereby accelerating protein synthesis in Th2 cells.

[0132] 3.6TAF9B mediates Th2 cell regulation of DC@CFO / BFO nanoparticles

[0133] To identify key transcription factors in DC@CFO / BFO+M treatment, gene set enrichment analysis (GSEA) was performed on the transcriptome data ( Fig. 9 By analyzing the expression of TAF9B in various immune cells from the Human Protein Atlas database, it was found that TAF9B was only expressed in T cell subsets ( Fig.10 a). Quantitative proteomics showed that TAF9B was upregulated after treatment with DC@CFO / BFO nanoparticles plus an external magnetic field ( Fig.10 b). Fractions of T cell lysates from the sucrose gradient during multimer profiling were collected and the mRNA level of TAF9B was examined by quantitative PCR. The results showed that the translationally active mRNA of TAF9B (in multimers) was significantly increased after treatment with DC@CFO / BFO composites and loading with a magnetic field ( Fig. 9 b).

[0134] To further verify that DC@CFO / BFO+M treatment modulates electrostatic interactions in the ribosomal exit tunnel, basic amino acids (positively charged amino acids, including arginine and lysine) were replaced with acidic amino acids (aspartic acid) at the C-terminus. Combined with quantitative PCR, multimer mapping showed that DC@CFO / BFO+M treatment showed little effect on the translation rate of the C-terminal mutant of TAF9B ( Fig. 9 c), indicating that the stimulatory effect of the magnetoelectric nanocomposite in regulating protein synthesis depends at least in part on electrostatic interactions at the ribosomal peptide exit tunnel.

[0135] To evaluate the role of TAF9B in T cell polarization and proliferation, we induced CD4 + T cell differentiation Fig.10 c). After treatment with DC@CFO / BFO composites and magnetic field stimulation, knockout of TAF9B significantly inhibited the Th2 polarization induced by DC@CFO / BFO+M ( Fig. 9 d), while the proportion of Th1 cells and Th17 cells increased after TAF9B knockout ( Fig. 9 E and Fig.10 d), the proportion of iTregs had no significant difference ( Fig.10 In addition, Th2 cell proliferation was significantly restricted with the downregulation of TAF9B, as shown by Ki-67 staining ( Fig. 9 f). Therefore, these data demonstrate that TAF9B is an essential transcriptional regulator for DC@CFO / BFO nanoparticle-mediated Th2 cell regulation.

[0136] 4. Discussion

[0137] The study in this example shows that the DC membrane-coated magnetoelectric nanocomposite can regulate Th1 / Th2 cells and promote Th2 cell proliferation, thereby being used to improve T cell-mediated autoimmune responses and maintain host immune homeostasis. +T cells undergo differentiation into various lineages of T helper cells, in which transcription factors play a central role in determining the lineage within a specific cytokine environment. Transcription factor IID (TFIID) components, called TAFs, are essential for pre-initiation complex assembly and RNA polymerase II (Pol II)-mediated transcription initiation. Unlike previously reported TAF9B as a key regulator of apoptosis-related gene expression and control of motor neuron differentiation, the multi-omics analysis of this embodiment determined that TAF9B is a key protein regulated by DC@CFO / BFO nanoparticles and magnetic field stimulation, and is closely related to ribosome interaction and protein structure. Unlike other genes regulated by these stimuli, TAF9B acts as a transcription factor. In order to verify this conclusion, by depleting endogenous TAF9B in T cells, the result was a significant inhibition of Th2 cell polarization. Therefore, TAF9B activation is crucial as the main mechanism of Th2 cell regulation. These findings enhance the understanding of the molecular mechanisms regulating Th2 cell differentiation and highlight the therapeutic potential of targeting TAF9B.

[0138] T cells undergo a major shift in protein synthesis activity during activation, proliferation, and differentiation. Precise regulation of protein synthesis shapes the functional capacity of T cell immune responses. There is evidence that the translation of transcripts encoding ribosomal proteins is regulated during effector T cell differentiation, and translation inhibition is more pronounced in terminal effector cells. The magnetoelectric nanocomposite in this embodiment exhibits colocalization with ribosomal components and optimized amino acid metabolic adaptability. By combining polymer spectra and proteome analysis and the kinetics of protein synthesis, it is revealed that the magnetoelectric nanocomposite regulates the electrostatic interactions of ribosomes and stimulates translation when a magnetic field is applied, thereby promoting Th2 cell polarization and proliferation.

[0139] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit them; although the present invention has been described in detail with reference to the above embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the above embodiments, or replace some of the technical features therein by equivalents. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. Use of nanocomposite particles in the preparation of a drug for regulating ribosome activity, characterized in that: The nanocomposite particle comprises a piezoelectric material layer and a magnetically responsive core structure, and the regulation comprises promoting translation or protein biosynthesis.

2. The use according to claim 1, characterized in that The piezoelectric material layer is prepared from inorganic piezoelectric material and / or organic polymer piezoelectric material, and the magnetic response core structure is prepared from magnetic material.

3. The use according to claim 1, characterized in that The particle size of the nanocomposite particles is 10-500 nm.

4. A pharmaceutical composition, characterized in that The invention comprises a nanocomposite particle and a pharmaceutically acceptable carrier, wherein the nanocomposite particle comprises a piezoelectric material layer and a magnetically responsive core structure.

5. The pharmaceutical composition according to claim 4, characterized in that Pharmaceutically acceptable carriers include membrane-forming lipids.

6. The pharmaceutical composition according to claim 5, characterized in that Further included are ligand molecules that bind to target cells.

7. A method for regulating cell activity in vitro, characterized in that: The method comprises the steps of allowing nanocomposite particles to enter cells and subjecting them to magnetic field treatment, wherein the nanocomposite particles comprise a piezoelectric material layer and a magnetically responsive core structure.

8. The method for regulating cell activity in vitro according to claim 7, characterized in that: The regulating cell activity comprises at least one of the following situations: (1) Regulate ribosome electrostatic interactions; (2) regulating the local electrostatic potential within the ribosome exit tunnel; (3) Regulate protein biosynthesis; (4) regulating the amount or expression of transcription factors; (5) Regulate the balance of Th1 / Th2 cells.

9. The method for regulating cell activity in vitro according to claim 7, characterized in that: The concentration of the nanocomposite particles is less than 100 μg / mL.

10. The method for regulating cell activity in vitro according to claim 7, characterized in that: The nanocomposite particles are coated with a membrane material and contacted with cells, so that the nanocomposite particles enter the cells.

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