A reagent, induction method and application for inducing macrophage transformation in vitro under inflammatory conditions

Abstract

The present invention provides a reagent, an induction method and an application for inducing macrophage transformation in vitro under inflammatory conditions, and belongs to the field of biomedical technology. The present invention provides a reagent for inducing macrophage transformation in vitro under inflammatory conditions, and the reagent has an inhibitory effect on PTBP1. The present invention provides a new method for inducing M1 macrophages into M2 macrophages under inflammatory conditions, which inhibits the gene expression and / or protein expression of PTBP1, changes the polarization phenotype and cytological characteristics of macrophages, reduces the related pro-inflammatory response brought about by M1 macrophages, induces macrophages to polarize to M2 type and exerts anti-inflammatory effects, and promotes the regeneration and repair of inflammatory injuries. The preparation process of the reagent and M2 macrophages of the present invention is simple, the expression is stable, the preparation system is easy to control quality, and it is easy to industrialize.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomedicine, and specifically relates to a reagent, an induction method and an application for inducing macrophage transformation in vitro under inflammatory conditions. Background Art

[0002] Inflammation is a natural defense response of the body to stimuli. Any factor that can cause tissue damage can be the cause of inflammation. Generally speaking, inflammation is protective to the body in nature. During the inflammatory process, a series of local reactions centered on the vascular system limit and eliminate damaging factors, while also promoting the healing of damaged tissues, which is a prerequisite for tissue repair and regeneration. If the integrity of the tissue and the balance in the body are restored during the repair process, the inflammation will automatically subside; however, in some cases, inflammation has potential hazards, and severe inflammatory reactions are often the basis for the onset of some diseases. For example, severe hypersensitivity inflammation can threaten the patient's life. Therefore, it is crucial to use anti-inflammatory drugs to suppress excessive inflammatory reactions.

[0003] A large number of studies have shown that macrophages are the main cells involved in inflammatory responses after disease and injury, and play an important role in the injury and anti-injury process after the occurrence of various diseases. They can monitor pathogens and injuries, participate in antigen phagocytosis and presentation, secrete active substances and immune signaling molecules, and these substances can freely enter and exit the blood-brain barrier, which plays a vital role in maintaining the homeostasis and permeability of the blood-brain barrier. Macrophage polarization is an important pathological process for coordinating inflammation, and can present two classic functional phenotypes: classically activated M1 type and alternatively activated M2 type. Among them, M1 type is induced by IFN-r and LPS, expresses INOS, and secretes TNF-a, IL-1β and other pro-inflammatory factors. It can phagocytose and remove necrotic cells and tissue debris, and promote the production of inflammatory intermediates. When the M1 type is overactivated in large quantities, it will induce or aggravate the damaging lesions of tissue cells. The M2 type is induced by IL-4 or IL-13, expresses ARG1, secretes anti-inflammatory factors such as IL-10 and TGF-β1, plays an anti-inflammatory protective role, promotes angiogenesis and tissue remodeling, inhibits excessive inflammatory response, and promotes the survival of damaged tissue cells. (Activation of corticotropin-releasing factor receptor 1 aggravates dextransodium sulphate-induced colitis in mice by promoting M1 macrophage polarization. 2018). This polarization phenomenon of macrophages is a dynamic pathological process that changes over time. Taking spinal cord injury as an example, in the early stage of injury, some ARG1+ M2 macrophages can be monitored; three days after the injury, INOS+ M1 macrophages gradually increase; on the seventh day after the injury, the number of M2 macrophages gradually decreases, and the injured area is mainly occupied by M1 macrophages; 14 days later, M2 macrophages return to the pre-injury level, while M1 macrophages can persist. This dynamic polarization phenomenon from transient M2 to M1 type marks the transition of macrophages from a short-term "anti-inflammatory" state to a "pro-inflammatory" state. (Reversible differentiation of pro-and anti-inflammatory macrophages. 2012; Repertoire of microglial and macrophager responses after spinal cord injury. 2011). A large number of studies have shown that regulating the polarization of macrophages from pro-inflammatory M1 to anti-inflammatory M2 is the key to reducing the response to injury.Studies have shown that multiple signaling pathways are related to the M1 and M2 transformation of macrophages. There are also numerous regulatory molecules for different signaling pathways, which shut down or activate the corresponding signaling pathways to achieve the transformation of macrophages. The regulatory molecules include miRNA, small molecule proteins and other substances, but they are only reported in scientific research literature. It is unknown whether their actual induction effectiveness in vitro can be used to evaluate the corresponding drug efficacy. Summary of the invention

[0004] The purpose of the present invention is to provide a reagent, an induction method and an application for inducing macrophage transformation in vitro under inflammatory conditions, and to promote macrophage polarization to M2 type by knocking down the expression of PTBP1 under inflammatory conditions, so as to improve the related inflammatory damage of tissue cells.

[0005] The present invention provides a reagent for inducing macrophage transformation in vitro under inflammatory conditions, wherein the reagent has an inhibitory effect on PTBP1;

[0006] The inducing macrophage transformation in vitro includes inducing M1 macrophages into M2 macrophages.

[0007] Preferably, the inhibitory effect includes inhibiting the expression of the coding gene of PTBP1, the expression of mRNA and / or inhibiting the protein expression of PTBP1.

[0008] Preferably, the reagent having an inhibitory effect on PTBP1 includes: at least one of chemically synthesized double-stranded specific interfering RNA, small molecule compounds and antibodies; or at least one of an interfering gene virus, miRNA, antisense oligonucleotide, nucleic acid aptamer and gene editor carrying the interfering gene that interferes with PTBP1 expression.

[0009] The present invention also provides an inducer for inducing M1 macrophages into M2 macrophages under inflammatory conditions, comprising chemically synthesized double-stranded specific interfering RNA, wherein the chemically synthesized double-stranded specific interfering RNA comprises siRNA or shRNA designed using human PTBP1 mRNA as a template.

[0010] Preferably, the sequence of the siRNA is shown as SEQ ID No.1.

[0011] The present invention also provides a method for inducing M1 macrophages to M2 phenotype under inflammatory conditions, comprising inducing macrophages using the above reagent or the above inducer.

[0012] Preferably, when the reagent is a chemically synthesized double-stranded specific infection RNA, the method comprises the following steps: transiently transfecting macrophages with the chemically synthesized double-stranded specific interfering RNA.

[0013] Preferably, the macrophages include peripheral blood macrophages, central nervous system microglia or tissue-specific macrophages.

[0014] The present invention also provides the use of the above reagent or the above inducer in preparing a medicament for antagonizing inflammation.

[0015] Preferably, the inflammation includes inflammation caused by sepsis, nerve damage or neurodegenerative diseases, and bone and joint injuries.

[0016] Beneficial effects: The present invention provides a reagent for inducing macrophage transformation in vitro under inflammatory conditions, and the reagent has an inhibitory effect on PTBP1. The present invention provides a new method for inducing M1 macrophages into M2 macrophages under inflammatory conditions, by inhibiting the gene expression and / or protein expression of PTBP1, changing the polarization phenotype and cytological characteristics of macrophages, reducing the related pro-inflammatory response brought about by M1 macrophages, inducing macrophages to polarize to M2 type and exert anti-inflammatory effects, and promoting the regeneration and repair of inflammatory damage. In the embodiment of the present invention, by transferring siRNA that inhibits PTBP1 into mouse macrophages raw264.7, raw264.7 is promoted to polarize from the pro-inflammatory M1 type to the anti-inflammatory M2 type in an inflammatory environment, thereby enhancing the anti-inflammatory effect of macrophages and promoting damage repair after an inflammatory reaction. The preparation process of the present invention is simple, the expression is stable, the preparation system is easy to control quality, and it is easy to industrialize. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative labor.

[0018] Figure 1 This is the result of western blot detection of PTBP1 expression level in mouse raw264.7 cells after LPS stimulation;

[0019] Figure 2 This is the result of immunofluorescence detection of the average fluorescence intensity of PTBP1 in mouse raw264.7 cells after LPS stimulation;

[0020] Figure 3 This is a diagram showing the results of screening using si-m-PTBP1;

[0021] Figure 4 The relative expression results of macrophage polarization markers (INOS and ARG1) in mouse raw264.7 cells before and after PTBP1 knockdown;

[0022] Figure 5 The relative expression results of macrophage pro-inflammatory cytokines (IL-6, TNF-α) and anti-inflammatory cytokines (IL-10, TGF-β) in mouse raw264.7 cells before and after PTBP1 knockdown;

[0023] Figure 6 This is the result of Western blot detection of the expression levels of proteins related to the MAPK signaling pathway after PTBP1 knockdown. DETAILED DESCRIPTION

[0024] The present invention provides a reagent for inducing macrophage transformation in vitro under inflammatory conditions, wherein the reagent has an inhibitory effect on PTBP1;

[0025] The inducing macrophage transformation in vitro includes inducing M1 macrophages into M2 macrophages.

[0026] The inhibitory effect of the present invention includes inhibiting the expression of PTBP1, and / or inhibiting the biological effect of PTBP1, that is, inhibiting the expression of the coding gene of PTBP1, mRNA expression and / or inhibiting the protein expression of PTBP1. The agent having an inhibitory effect on PTBP1 of the present invention preferably includes: at least one of chemically synthesized double-stranded specific interfering RNA (such as siRNA), small molecule compounds and antibodies; or at least one of a virus carrying an interfering gene that interferes with the expression of PTBP1 (such as shRNA virus), miRNA, antisense oligonucleotide, nucleic acid aptamer and gene editor.

[0027] The macrophages described in the present invention mainly refer to peripheral blood macrophages, but also include central nervous system microglia and various tissue-specific macrophages such as lung macrophages, skin macrophages, etc.

[0028] The present invention also provides an inducer for inducing M1 macrophages to M2 macrophages under inflammatory conditions, comprising a chemically synthesized double-stranded specific interfering RNA, wherein the chemically synthesized double-stranded specific interfering RNA comprises a siRNA or shRNA designed using human PTBP1 mRNA (NM_002819.5) or its transcript variants NM_031990.4, NM_031991.4 and NM_001411140.1 as templates. In the embodiment of the present invention, a siRNA is designed using mouse PTBP1 mRNA as a template, and its sequence is shown in SEQ ID No. 1: ACGATAGCAAGAAGTTCAA.

[0029] The present invention also provides a method for inducing M1 macrophages to M2 phenotype under inflammatory conditions, comprising inducing macrophages using the above reagent or the above inducer.

[0030] The method of the present invention preferably includes the following steps: using chemically synthesized double-stranded specific interfering RNA to transiently transfect macrophages, specifically using the siRNA to inhibit the mRNA transcription of the target gene PTBP1 of M1 macrophages by transient transfection to inhibit the expression of PTBP1 protein, and after the siRNA interference inhibition, the expression of PTBP1 protein is reduced to 50% to 90% of the expression of PTBP1 protein in the control M1 macrophages. The present invention does not specifically limit the method of transient transfection, and it can be carried out using conventional methods in the art.

[0031] The present invention also provides the use of the above reagent or the above inducer in preparing a medicament for antagonizing inflammation.

[0032] By using the method of the present invention, the intracellular RNA splicing protein PTBP1 is used as the target molecule to interfere with and inhibit its protein expression in M1 macrophages, regulate inflammatory pathways such as NF-kb and MAPK, and then inhibit the expression of pro-inflammatory factors such as IL-1β and TNF-a, promote the expression of anti-inflammatory factors such as IL-10 and TGF-β1, and reverse the M1 type to the M2 phenotype. The reagents or inducers involved in the method can be used in anti-inflammatory treatment. The inflammation of the present invention preferably includes inflammation caused by sepsis, nervous system damage or neurodegenerative diseases, and bone and joint damage.

[0033] To further illustrate the present invention, the following is a detailed description of the reagent, induction method and application for inducing macrophage transformation in vitro under inflammatory conditions provided by the present invention in conjunction with the accompanying drawings and examples, but they should not be construed as limiting the scope of protection of the present invention.

[0034] Example 1

[0035] Culture of mouse raw264.7 cells M0

[0036] Culture conditions: DMEM high-glucose medium containing 10% fetal bovine serum and 10% double antibody (penicillin + streptomycin sulfate) (complete medium), 5 mL T25 culture flask.

[0037] 1) Cells can be passaged when they grow to more than 80%

[0038] 2) Discard the original complete medium in the bottle before cell digestion

[0039] 3) Use DMEM high-glucose medium (without fetal bovine serum) / PBS of equal volume or twice the volume for washing. (The purpose here is to clean the metabolites and residual serum on the cell surface.) (raw264.7 is a semi-adherent cell. Note that when adding culture medium / PBS, do not add it toward the bottom of the bottle, but add it gently along the bottle wall.)

[0040] 4) Gently shake the culture flask, discard the culture medium / PBS, and then add an appropriate volume of culture medium / PBS. (Raw264.7 does not require trypsin digestion, the function here is to blow and digest the cells, and the added volume is equal to the volume of the culture system in the culture flask)

[0041] 5) Use a 5 mL pipette to gently aspirate the culture medium / PBS and blow the bottom of the bottle while observing the status of the cells at the bottom of the bottle. When the cells at the bottom of the bottle are visibly suspended in the culture medium / PBS, transfer them into a centrifuge tube.

[0042] 6) Centrifuge at 1000 rpm for 5 min.

[0043] 7) After centrifugation, discard the PBS. Please note that you should not resuspend and discard the cells.

[0044] 8) Add appropriate complete culture medium to resuspend. Generally, the culture process is 1 to 5 times. The cells will be full grown within 2 days. You can resuspend them in 1 mL and add 250 μL to the culture bottle.

[0045] Example 2

[0046] Raw264.7 was seeded into a six-well plate to a density of 30%.

[0047] When the cell density in the T25 bottle converges to 80%, discard the original complete culture medium in the bottle, wash with 1×PBS, shake the culture bottle gently, then discard the PBS, add 3mL 4℃ pre-cooled PBS, use a 5mL pipette to gently aspirate PBS, blow the bottom of the bottle, and observe the state of the cells at the bottom of the bottle. After the cells at the bottom of the bottle are visibly suspended in PBS, transfer them to a centrifuge tube and centrifuge at 1000rpm for 5min. After centrifugation, discard the PBS, add 3mL complete culture medium to the bottle and resuspend it. After the cell pellet is evenly resuspended, aspirate 80μL and add it to each well of the six-well plate. The well plate contains 2mL complete culture medium. Gently shake the well plate to make the cells evenly distributed. The cell density at this time is about 2×10 5 / well, incubate at 37°C, 5% CO2 for 12 hours, and conduct subsequent experiments after the cell density in each well reaches about 30%.

[0048] Example 3

[0049] Determine the most suitable siRNA transfection concentration for RNAi interference of raw264.7

[0050] 1) One day before transfection, 2×10 5 / well of raw264.7 was inoculated into a six-well plate, with 2 mL of complete medium per well. When the cell density reached about 30% confluence 12 hours after inoculation, RNAi interference experiment was performed on raw264.7 (the siRNA used was designed and synthesized by Guangzhou Ruibo Biotechnology Co., Ltd.):

[0051] The target sequence of 001, si-m-Ptbp1 (SEQ ID No. 1), ACGATAGCAAGAAGTTCAA;

[0052] The target sequence of 002, si-m-Ptbp1 (SEQ ID No. 2), GATCATCACGTTCACCAAG;

[0053] The target sequence of 003, si-m-Ptbp1 (SEQ ID No. 3), GTCAGTGCGCATTACACTG;

[0054] 2) Add 200 μL Opti-MEM containing 3 different final concentrations of RNAioligonμcleotides (si-m-PTBP1) of 50 nM, 75 nM, and 100 nM and the corresponding negative control siRNA (NC) to 4 centrifuge tubes, mix gently, and place at room temperature for 5 minutes;

[0055] 3) Add 5 μL Lipofectamine RNAiMAX Reagent to 200 μL Opti-MEM, mix gently, and leave at room temperature for 5 minutes;

[0056] 4) Mix the diluted si-m-PTBP1 / NC of different concentrations with RNAiMAX Reagent: Mix gently and leave at room temperature for 20 minutes to form a complex;

[0057] 5) Discard the cell culture medium and replace it with serum-free or dual-antibody H-DMEM high-glucose medium (1.6 mL / well). Add the above mixture to each well in a circular manner and mix well.

[0058] 6) The well plate was placed in a 5% CO2 incubator, and total protein was extracted after 24 hours. Western blot was performed to determine the siRNA with the best effect.

[0059] Western blot results of PTBP1 si-RNA transfection showed Figure 3As shown in the figure: Among reagents 01, 02, and 03, reagent 01 has the most obvious knockdown effect. Then, si-RNA 01 was used to screen the optimal transfection concentration, and qPCR was used to detect the knockdown efficiency of PTBP1 in three groups (50nM, 75nM, and 100nM). It was found that the knockdown effect of PTBP1 was better as the concentration increased. However, because cell differentiation and death occurred at 100nM, and there was a significant statistical significance at 50nM, 50nM was selected for transfection in subsequent experiments.

[0060] Example 4

[0061] After the cells were grouped, RT-qPCR and Western blot were performed for inflammation induction experiments.

[0062] After determining the siRNA, the raw264.7 cells were inoculated in six-well plates according to the cell density of Example 3, and the mRNA and protein expression analysis was performed in batches. They were divided into NC group (Negative control siRNA treatment for 24 hours and then replaced with complete medium for 24 hours), SI group (si-m-PTBP1 interference for 24 hours and then replaced with complete medium for 24 hours), LPS treatment group (Negative control siRNA + LPS concentration of 500ng / mL cultured for 24 hours), SI+LPS group (si-m-PTBP1 interference for 24 hours and then replaced with LPS culture solution containing 500ng / mL and then cultured for 24 hours).

[0063] After the above grouping, siRNA interference was used to inhibit PTBP1 expression according to the scheme in Example 3, and 2 mL of the specified complete culture medium or culture medium containing LPS was added to continue the culture. At 24 hours, total RNA was extracted and cDNA was synthesized for RT-qPCR detection; total protein was extracted for Western blot detection.

[0064] The results are as follows Figure 1 As shown in the figure, LPS-treated macrophages polarize to the M1 type, which promotes the nuclear translocation of PTBP1 protein. Total proteins were extracted from LPS-untreated macrophages (M0 type) and LPS-treated macrophages (M1 type) for Western blot analysis, and no significant difference was found in the expression of PTBP1 ( Figure 1 Middle A). Further extraction of nuclear proteins from M0 and M1 macrophages for Western blot analysis revealed that after LPS treatment, the amount of PTBP1 protein entering the nucleus of M1 macrophages increased significantly ( Figure 1(B) Cell immunofluorescence results showed that compared with M0 type, the nucleus of M1 type macrophages was significantly enlarged, and the average fluorescence intensity of PTBP1 antigen in the nucleus was significantly enhanced. This indicates that under the conditions of LPS-induced inflammatory response, the cellular distribution of macrophage PTBP1 protein changes, mainly through translocation into the nucleus, and plays a role in macrophage polarization through pathways such as mRNA shearing.

[0065] Example 4

[0066] RT-qPCR steps

[0067] Total RNA was extracted from the raw264.7 cells of mice treated in the above different groups using the QIAzol (Qiagen) kit. RNA was reverse transcribed and synthesized into cDNA using TransScript One-Step gDNA Removal and cDNA Synthesis SuperMix. The reverse transcription system was as follows: 1000 ng of total RNA was added with 1 μL of Lanchored Oligo(dT)18 Primer (0.5 μg / μL), 10 μL of 2×TS-Uni Reaction Mix, 1 μL of gDNA Remover and 1 μL of TransScript-Uni RT / RI Enzyme Mix, and RNase-free water was added to make the total system 20 μL. Reverse transcription was performed in a PCR instrument, and the reaction conditions were set at 42°C for 15 min and 85°C for 5 s.

[0068] By ChamQ TMUniversal SYBR qPCRMasterMix was used for fluorescence quantitative PCR reaction. The system was as follows: 2μL Template cDNA (diluted 1:10, 100-200ng), 10μL 2×ChamQ Universal SYBRqPCRMasterMix, 0.4μL Primer1 (10μM) and 0.4μL Primer2 (10μM) were added, and 7.2μL ddH2O (RNase / DNase free) was added to supplement the total reaction system to 20μL, and qRT-PCR reaction was performed in 7300 Real Time PCR System instrument. The reaction conditions were set as follows: 95℃ pre-denaturation for 30s; 95℃10s, 60℃30s as a PCR cycle reaction, for a total of 40 cycles; melting curve, i.e. 95℃15s, 60℃1min, 95℃15s. Fluorescence quantitative results were analyzed by Applied Biosystem 7300Plus Real-time PCR System software and analyzed by 2 -△△Ct The results were quantitatively analyzed by the method of . The sequences of the PCR primers for different genes used in the reaction are shown in Table 1.

[0069] Table 1 PCR primer sequences

[0070]

[0071] After knocking down the expression of PTBP1 gene in macrophages, the polarization of macrophages from M1 to M2 was significantly promoted and anti-inflammatory effects were exerted. qRT-PCR was performed to verify the macrophage polarization markers INOS and ARG1. The results showed that ( Figure 4 Middle A): After adding LPS, INOS gene expression was upregulated and ARG1 gene expression was significantly downregulated; while after PTBP1 knockdown, INOS gene expression was downregulated and ARG1 gene expression was upregulated. Western blot results ( Figure 4 Middle B) shows that the expression of INOS protein was significantly reduced after PTBP1 knockdown. The above results indicate that PTBP1 knockdown promotes the polarization of macrophages from M1 to M2.

[0072] The secretion of different inflammatory factors is also a manifestation of the different polarization functions of macrophages. Among them, IL-6 and TNF-α are pro-inflammatory cytokines, and IL-10 and TGF-β1 are anti-inflammatory cytokines. RT-qPCR was used to detect the expression of the above cytokines. Figure 5The results showed that the gene expression of TNF-α and IL-6 was significantly upregulated after LPS treatment, but downregulated after PTBP1 knockdown. The changes in the mRNA expression of IL-10 and TGF-β1 were detected, and it was found that the gene expression of IL-10 was upregulated in the PTBP1 knockdown (no LPS) treatment group compared with the NC group (no LPS); IL-10 was significantly upregulated in the PTBP1 knockdown + LPS combined treatment group compared with the NC + LPS group. Similar to the expression changes of IL-10, the expression of TGF-β1 was significantly upregulated in the PTBP1 knockdown (no LPS) or PTBP1 + LPS treatment group compared with the control group. Western blot detection results at the protein level showed that compared with the LPS treatment group alone, the protein expression levels of pro-inflammatory factors TNF-α and IL-6 in the PTBP1 knockdown + LPS co-treatment group were significantly downregulated, while the protein expression of anti-inflammatory factors IL-10 and TGF-β1 was significantly upregulated. The above results indicate that knocking down PTBP1 expression in macrophages reduces the expression of proinflammatory factors and exerts a certain anti-inflammatory effect.

[0073] Example 5

[0074] Western blot was used to detect the expression of PTBP1, INOS, TNF-α, IL-6, TGF-β1, IL-10, p38 and p-p38 proteins

[0075] Take the cells in different groups with good cell status for protein sample extraction, use RIPA lysis buffer to extract total cell protein, (nuclear protein extraction kit) to extract nuclear protein, and BCA protein quantitative analysis kit for protein quantification. Calculate the volume required for 20μg protein according to the measured concentration, mix with 5× SDS loading buffer, boil in boiling water for 5min, and store at -20℃ for Western blot experiment to detect key protein expression. GAPDH / β-actin is used as the internal reference for total protein detection, and H3 is used as the internal reference for nuclear protein detection. INOS uses 8% separation gel; PTBP1, TGF-β1, p38, p-p38 use 10% separation gel; TNF-α, IL-6, IL-10 use 15% separation gel.

[0076] Knocking down the expression of PTBP1 in macrophages inhibits the p38 MAPK signaling pathway in M1 macrophages. The MAPK signaling pathway is a classic inflammatory activation pathway. In M1 macrophages activated by LPS stimulation, the level of phosphorylated p38 protein (p-P38) is significantly upregulated, and the p38 MAPK pathway is in an activated state, promoting the expression of downstream inflammatory factors such as TNF-α and IL-6. Figure 6As shown in the results, after interfering with and inhibiting PTBP1 protein expression, p-P38 protein was significantly downregulated, indicating that inhibiting PTBP1 expression in macrophages under LPS inflammatory conditions effectively reduced the activation of the p38 MAPK signaling pathway, significantly reduced the expression of pro-inflammatory factors, and promoted the M2 polarization of macrophages with anti-inflammatory effects.

[0077] Although the above embodiment describes the present invention in detail, it is only a part of the embodiments of the present invention, not all of the embodiments. People can also obtain other embodiments based on this embodiment without creativity, and these embodiments all fall within the protection scope of the present invention.

Claims

1. An inducing agent for inducing M1 macrophages to M2 macrophages under inflammatory conditions, characterized in that: It comprises chemically synthesized double-stranded specific interfering RNA, wherein the chemically synthesized double-stranded specific interfering RNA comprises siRNA designed with human PTBP1 mRNA as a template, and the sequence of the siRNA is shown as SEQ ID No.1.

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