Application of CCR5 inhibitor in promotion of severe acute pancreatitis tissue repair

Abstract

The present application relates to the use of an Axl agonist, a Mertk agonist, a CCR5 inhibitor or a CD22 inhibitor in the prevention or treatment of severe acute pancreatitis (SAP).

Description

Technical Field

[0001] This invention relates to the fields of biotechnology and pharmaceutical technology, specifically to the application of CCR5 inhibitors in the prevention and treatment of severe acute pancreatitis. Background Technology

[0002] Severe acute pancreatitis (SAP), accompanied by systemic inflammatory response syndrome and multiple organ failure, has a mortality rate as high as 30%, and currently lacks specific drug treatment. SAP patients often experience local complications during the recovery period, leading to significantly prolonged recovery time and increased hospital stays. The progression and prognosis of the disease are closely related to the degree of tissue damage and repair capacity. Previous studies have shown that SAP is characterized by rapid and large-scale cell death. Research has also found that almost all known cell death pathways are clearly associated with the severity of SAP, including classic apoptosis and necrosis pathways, as well as newly discovered pathways such as necroptosis, pyroptosis, and ferroptosis. Most studies have focused on key regulatory factors in pancreatic acinar or ductal cells, revealing the strict regulatory mechanisms of cell death pathway activation in these cells, thus providing potential targets for early treatment of SAP.

[0003] However, the narrow treatment window for SAP necessitates the exploration of a drug that limits tissue damage and promotes tissue repair in the later stages of this deadly disease.

[0004] A key characteristic of pancreatitis is the infiltration of numerous immune cells, such as neutrophils, macrophages, dendritic cells, and T / B cells. During acute pancreatitis (AP), this inflammatory cell infiltration can directly act on pancreatic acinar cells, participating in tissue damage and repair processes by releasing inflammatory cytokines. Recent research shows that macrophage phenotypes undergo dynamic changes during the injury and repair phases of AP. In the repair phase, the shift of macrophages to an M2 polarization state promotes pancreatic tissue regeneration, while activated macrophages can also construct a specific immune microenvironment for tissue repair after pancreatic injury, thereby aiding in the regeneration of damaged acinar cells. The study by Manohar et al. further revealed the crucial role of macrophages in mediating the severity of pancreatitis and tissue repair after SAP. However, the regulatory factors or sensors expressed by these immune cells within the microenvironment remain largely unknown. How they coordinate signals activated by cell death pathways, particularly those involving apoptotic cells, to sense and respond to tissue damage and repair requires further investigation for practical applications. Summary of the Invention

[0005] To achieve the above objectives, this study found that the TAM receptor tyrosine kinase expressed in macrophages or neutrophils consists of three homologous receptor tyrosine kinases: TYRO3, AXL, and MERTK. Among them, AXL and MERTK (rather than receptors such as Tyro3 that sense apoptosis signals) are selectively expressed in pancreatic macrophages, and the expression levels of AXL and MERTK in pancreatic macrophages are correlated with the degree of SAP tissue damage and repair. Single-cell transcriptome analysis showed enhanced ligand-receptor interactions between Cxcr2+ neutrophils and CD163+ macrophages via the CCL4-CCR5 axis. Inhibition of CCR5 may promote pancreatic tissue repair after SAP treatment, providing a broader therapeutic prospect and potential drugs for SAP treatment.

[0006] Specifically, this application provides the following technical solutions: 1. Use of Axl agonists, Mertk agonists, CCR5 inhibitors, or CD22 inhibitors in the prevention or treatment of severe acute pancreatitis (SAP) or sepsis.

[0007] 2. Use of Axl agonists, Mertk agonists, CCR5 inhibitors, or CD22 inhibitors in the preparation of medicaments or formulations for the prevention or treatment of severe acute pancreatitis (SAP) or sepsis.

[0008] 3. The use according to item 2, wherein the prevention or treatment of severe acute pancreatitis (SAP) includes prevention or treatment of severe acute pancreatitis (SAP) during the tissue damage phase, repair phase, and postoperative phase.

[0009] 4. Use of Axl agonists, Mertk agonists, CCR5 inhibitors, or CD22 inhibitors in the preparation of drugs or formulations for promoting the repair of pancreatic tissue damage.

[0010] 5. Use of Axl agonists, Mertk agonists, CCR5 inhibitors, or CD22 inhibitors in the preparation of drugs or formulations for promoting regeneration of pancreatic acinar cells after damage.

[0011] 6. Use of Axl agonists, Mertk agonists, CCR5 inhibitors, or CD22 inhibitors in the preparation of medicaments or formulations for the treatment or relief of pancreatic inflammation.

[0012] 7. The use according to any one of items 4 to 6, wherein the pancreatic tissue damage or pancreatic acinar cell damage or pancreatic inflammation is caused by severe acute pancreatitis (SAP).

[0013] 8. The use according to any one of items 1 to 7, wherein the CCR5 inhibitor is selected from Maraviroc, and / or the CD22 inhibitor is selected from InVivoMab anti-mouse CD22.

[0014] 9. A pharmaceutical composition comprising one or more of an Axl agonist, a Mertk agonist, a CCR5 inhibitor, or a CD22 inhibitor, preferably for the prevention or treatment of severe acute pancreatitis (SAP); or preferably for promoting the repair of pancreatic tissue damage; or preferably for promoting the regeneration of damaged pancreatic acinar cells; or preferably for relieving or treating pancreatic inflammation; or preferably for the prevention or treatment of sepsis.

[0015] 10. The pharmaceutical composition according to claim 9, wherein the pharmaceutical composition is in the form of an injection, tablet, drops, powder, spray, gel, ointment or lozenge.

[0016] 11. The pharmaceutical composition according to item 9 or 10, wherein the CCR5 inhibitor is selected from Maraviroc, and / or the CD22 inhibitor is selected from InVivoMab anti-mouse CD22.

[0017] 12. A method for preventing or treating severe acute pancreatitis (SAP), comprising administering to a subject an effective dose of any one or more of an Axl agonist, a Mertk agonist, a CCR5 inhibitor, or a CD22 inhibitor.

[0018] 13. A method for preventing or treating sepsis, comprising administering to a subject an effective dose of any one or more of an Axl agonist, a Mertk agonist, a CCR5 inhibitor, or a CD22 inhibitor.

[0019] 14. The method of item 12 or 13, wherein the CCR5 inhibitor is selected from Maraviroc, and / or the CD22 inhibitor is selected from InVivoMab anti-mouse CD22.

[0020] 15. Use of M2 macrophages in the prevention or treatment of severe acute pancreatitis (SAP) or sepsis.

[0021] 16. Use of M2 macrophages in the preparation of drugs or cell preparations for the prevention or treatment of severe acute pancreatitis (SAP) or sepsis.

[0022] 17. The use according to item 16, wherein the drug or cell preparation is a drug or cell preparation for adoptive cell therapy.

[0023] 18. The use according to items 15-17, wherein the M2 macrophage is an IL-4 or IL-13 induced macrophage. Attached Figure Description

[0024] Figure 1 A is a UMAP diagram showing the results of RNA sequencing analysis.

[0025] Figure 1 B is a bubble chart showing the percentage of various positive cells.

[0026] Figure 1 C is a bubble chart showing the percentage of positive cells in each group.

[0027] Figure 1 D is a schematic diagram of the model of damage and repair induced by rain frog peptides in severe pancreatitis (SAP).

[0028] Figure 1 E represents the expression levels of AXL and MERTK in macrophages as shown by a representative histogram.

[0029] Figure 1 F represents the quantitative analysis results of macrophage fluorescence intensity.

[0030] Figure 2 A represents a hematoxylin and eosin stained pancreatic section from AXL and MERTK-deficient mice (n=5).

[0031] Figure 2 B represents the histological score of the pancreas.

[0032] Figure 2 C and 2D are representative immunohistochemical images of CD45 and cleaved Caspase-3 in pancreatic tissue. Data are presented as mean ± standard error (SEM). P-values ​​were calculated using a two-tailed unpaired Student's t-test. *P < 0.05, ns, no statistically significant difference.

[0033] Figure 3 A is a schematic diagram of the SAP experimental procedure induced by sodium taurocholate (NaT).

[0034] Figure 3 B represents a representative hematoxylin and eosin stained pancreatic section (n=5) from AXL and MERTK-deficient mice in the NaT-induced model.

[0035] Figure 3 C represents the pancreatic histological score in the NaT-induced model.

[0036] Figure 4 A is a schematic diagram of pancreatic macrophage sorting after pancreatitis induction.

[0037] Figure 4 B and 4C represent the expression of pro-inflammatory genes (Il1b, Il6, and Tnf) and pro-repair genes (Mrc1 and Retnla) in pancreatic macrophages, respectively, using RT-qPCR analysis (n=5 per group).

[0038] Figure 4 D and 4E represent the flow cytometry analysis results of pancreatic leukocytes in mice after SAP (n=5 per group). Representative flow cytometry images show the expression of inducible nitric oxide synthase (iNOS) (left image of 4D) and arginase (left image of 4E) in CD45+CD11b+F4 / 80+ macrophages, with the right images showing the quantitative results. Data are expressed as mean ± standard error. P-values ​​were calculated using a two-tailed unpaired Student's t-test. *P<0.05, ns indicates no significant difference.

[0039] Figure 5 A is the experimental flowchart for adoptive transfer of macrophages.

[0040] Figure 5 B is a schematic diagram of the experimental protocol for isolating bone marrow-derived macrophages (BMDM) and inducing IL-4 polarized macrophages.

[0041] Figure 5 C is a representative flow cytometry plot showing the expression of arginase and inducible nitric oxide synthase (iNOS) in BMDM.

[0042] Figure 5 D represents representative images and histological scores of pancreatic tissue sections stained with hematoxylin and eosin (n=6 per group).

[0043] Figure 5 E and 5F are representative immunohistochemical images of CD45 and lysed Caspase-3 (n=6) in pancreatic tissue, respectively. The right-hand image shows the quantitative results. Data are presented as mean ± standard error (SEM). P-values ​​were calculated using a two-tailed unpaired Student's t-test. *P<0.05.

[0044] Figure 6 A is a schematic diagram of the experimental procedure for scRNA-seq analysis of mouse pancreatic CD45+ cells (2 samples per group, each sample from 3 mice).

[0045] Figure 6 B is a UMAP diagram classified by cell type.

[0046] Figure 6C shows a representative flow cytometry image (left) and quantitative results (right) of neutrophils (CD45+CD11b+Ly6G+) at a specified time point.

[0047] Figure 6 D is a bubble diagram showing ligand-receptor interactions between Cxcr2+ neutrophils and other myeloid cell types.

[0048] Figure 6 E represents the number of RNA points for each ligand or receptor in RNA in situ hybridization detection.

[0049] Figure 6 F shows representative images of CCL4_CCR5 ligand-receptor pairs (left) and quantitative results of colocalized RNA sites (right). Data are mean ± standard error. P-values ​​were calculated using a two-tailed unpaired Student's t-test. *P < 0.05, ns, not statistically significant.

[0050] Figure 7 A represents the expression of a specific marker gene as shown in the violin diagram.

[0051] Figure 7 B is a combined UMAP diagram showing the relative proportions of cell types in each group (right) with coloring.

[0052] Figure 7 C represents the quantitative results of flow cytometry analysis of macrophages (CD45+CD11b+F4 / 80+), monocytes (CD45+CD11b+Ly6G+Ly6C+), and dendritic cells (CD45+CD11c+F4 / 80-MHCII+). Cxcr2+ neutrophils are marked with dashed rectangles.

[0053] Figure 7 D and 7E are representative images (left) of the PTPRC_CD22 and IL1B_ADRB2 ligand-receptor pairs and the quantitative results of colocalized RNA sites (right), respectively. Data are presented as mean ± standard error (SEM). P-values ​​were calculated using a two-tailed unpaired Student's t-test. *P < 0.05, ns, no significant difference.

[0054] Figure 8 A is a schematic diagram of treatment with the CCR5 inhibitor maraviro after pancreatitis induction.

[0055] Figure 8 B is a representative image of a pancreatic tissue section stained with hematoxylin and eosin. Figure 8 C represents the histological score (n=3 per group).

[0056] Figure 8D and 8E are representative immunohistochemical images (left) and quantitative results (right) of CD45, CD86, ARG1, and LY6B.2.

[0057] Figure 9 A is a schematic diagram of a CD22 inhibitor dosing regimen following pancreatitis induction.

[0058] Figure 9 B is a representative image (20X) of pancreatic tissue stained with HE.

[0059] Figure 9 C represents the pancreatic histopathological score. * indicates P < 0.05 compared to the normal control group; # indicates P < 0.05 compared to SAP. Detailed Implementation

[0060] The following specific embodiments illustrate and explain the implementation of the present invention in detail, but the following content should not be construed as limiting the present invention in any way.

[0061] Severe acute pancreatitis (SAP) refers to acute pancreatitis accompanied by organ failure or local complications such as necrosis, abscesses, or pseudocysts. Acute pancreatitis itself refers to an acute chemical inflammation of the pancreas and surrounding tissues caused by the pancreas's own digestive enzymes. Patients with severe acute pancreatitis often experience systemic inflammatory response syndrome and multiple organ failure, characterized by rapid and large-scale cell death; their condition is critical, and the mortality rate is relatively high.

[0062] Sepsis is a systemic inflammatory response syndrome caused by infection, commonly seen in patients with severe trauma or infectious diseases. Causes include infections from bacteria, fungi, viruses, and parasites, leading to an inflammatory response and imbalance in immune regulation. Severe sepsis can also result in organ dysfunction and circulatory disorders.

[0063] Axl and Mertk are both tyrosine kinases belonging to the TAM homologous receptor family. These receptors are activated by binding to specific ligands—growth arrest-specific protein 6 (GAS6) and protein S (PROS1)—and by interacting with phosphatidylserine (PtdSer) on the outer membrane of apoptotic cells, participating in the clearance of these cells—a mechanism known as phagocytic clearance. AXL and MERTK can mediate apoptotic cell clearance, and when synergistically acting with IL-4 or IL-13, they can promote tissue repair and remodeling in the intestine following worm-induced lung or colitis. Monocyte-derived macrophages, through AXL and MERTK-mediated erythrocyte phagocytosis and hematoma clearance mechanisms, can promote neurological function recovery after cerebral hemorrhage. Recent studies show that macrophages recognize different types of apoptotic cells through AXL and MERTK, thereby determining the functional differentiation of these cells in liver injury repair. Some studies have shown that AXL and MERTK have a negative regulatory role in inflammatory responses. This application found that AXL and MERTK (rather than other receptors that sense apoptosis signals such as Tyro3) are selectively expressed in pancreatic macrophages, and the expression levels of AXL and MERTK in pancreatic macrophages are correlated with the degree of SAP tissue damage and repair.

[0064] This application further revealed through single-cell transcriptome analysis that enhanced ligand-receptor interactions exist between Cxcr2+ neutrophils and CD163+ macrophages via the CCL4-CCR5 axis. Therefore, inhibiting CCR5 can also promote pancreatic tissue repair after SAP treatment. PTPRC-CD22 is another ligand-receptor pair between Cxcr2+ neutrophils and other immune cells, and inhibiting CD22 may have similar effects to inhibiting CCR5. This also provides a promising approach with a broader therapeutic window for interventions that promote pancreatic tissue repair and recovery.

[0065] Based on this, one aspect of this application provides the use of an Axl agonist, a Mertk agonist, a CCR5 inhibitor, or a CD22 inhibitor in the prevention or treatment of severe acute pancreatitis (SAP) or sepsis.

[0066] Furthermore, this application provides the use of Axl agonists, Mertk agonists, CCR5 inhibitors, or CD22 inhibitors in the preparation of medicaments or reagents for the prevention or treatment of severe acute pancreatitis (SAP) or sepsis.

[0067] In a specific implementation, the prevention or treatment of severe acute pancreatitis (SAP) includes prevention or treatment during the tissue damage phase, repair phase, and postoperative phase of severe acute pancreatitis (SAP).

[0068] In some preferred embodiments, the technical solution of this application is used in the recovery phase of severe acute pancreatitis (SAP).

[0069] The defining characteristics of SAP patients are pancreatic necrosis and persistent organ failure, which are two determinants of disease severity and prognosis. SAP is often accompanied by more local complications after surgery or during the recovery period, resulting in significantly prolonged recovery time and hospital stays.

[0070] Furthermore, this application provides the use of Axl agonists, Mertk agonists, CCR5 inhibitors, or CD22 inhibitors in the preparation of medicaments or formulations for promoting the repair of pancreatic tissue damage.

[0071] Furthermore, this application provides the use of an Axl agonist, a Mertk agonist, a CCR5 inhibitor, or a CD22 inhibitor in the preparation of a medicament or formulation for promoting the regeneration of pancreatic acinar cells after damage.

[0072] Furthermore, this application provides the use of an Axl agonist, a Mertk agonist, a CCR5 inhibitor, or a CD22 inhibitor in the preparation of a medicament or formulation for treating or alleviating pancreatic inflammation.

[0073] In some specific embodiments, the pancreatic tissue damage or pancreatic acinar cell damage followed by regeneration or pancreatic inflammation is caused by severe acute pancreatitis (SAP).

[0074] In some preferred embodiments of this application, the Axl agonist, Mertk agonist, CCR5 inhibitor, or CD22 inhibitor can be used for pancreatic acinar cell regeneration, pancreatic tissue damage repair, or inflammation relief in patients with severe acute pancreatitis (SAP) during the postoperative, tissue damage, or tissue repair phases.

[0075] In some preferred embodiments of this application, the Axl agonist, Mertk agonist, CCR5 inhibitor, or CD22 inhibitor can be used for pancreatic tissue repair and improvement of complications during the recovery period of severe acute pancreatitis (SAP).

[0076] In some embodiments of this application, the use of the Axl agonist, Mertk agonist, CCR5 inhibitor, or CD22 inhibitor for the prevention or treatment of severe acute pancreatitis (SAP) is achieved by selectively phagocytizing apoptotic neutrophils.

[0077] In some embodiments of this application, the use of the Axl agonist, Mertk agonist, CCR5 inhibitor, or CD22 inhibitor for the prevention or treatment of severe acute pancreatitis (SAP) is achieved by inducing macrophages to convert to a reparative macrophage phenotype.

[0078] Furthermore, another aspect of this application provides the use of repair macrophages, specifically M2 macrophages, in the prevention or treatment of severe acute pancreatitis (SAP) or sepsis.

[0079] This application also provides the use of M2 macrophages in the preparation of drugs or cell preparations for the prevention or treatment of severe acute pancreatitis (SAP) or sepsis.

[0080] Macrophages are classified into two subsets based on their function and inflammatory secretion levels: M1 (pro-inflammatory phenotype) and M2 (pro-repair phenotype). M1 macrophages are primarily activated by LPS and IFNγ, secreting pro-inflammatory factors such as IFN-γ and TNF-α. M2 macrophages are typically activated by anti-inflammatory cytokines (such as IL-4 and IL-13) or immune complexes, producing anti-inflammatory cytokines such as IL-10 and TGF-β, which participate in tissue repair, promote angiogenesis, and suppress inflammation. They are associated with Th2 immune responses and contribute to humoral immunity.

[0081] In this specific embodiment, M2 macrophages or drugs or cell preparations made from them can be used for adoptive cell therapy.

[0082] The phenotypic transformation of reparative macrophages is crucial for constructing a key microenvironment supporting pancreatic tissue repair following severe acute pancreatitis (SAP). This application reveals that AXL and MERTK macrophages promote tissue repair by phagocytizing apoptotic cells, a mechanism that induces the formation of a reparative macrophage phenotype in various diseases, including cerebral hemorrhage, pulmonary helminth infection, and colitis. AXL and MERTK macrophages selectively phagocytose apoptotic neutrophils (and a small number of apoptotic T cells), thereby inducing macrophages to exhibit different phenotypic characteristics. We found that in the SAP model, myeloid-specific knockout of Axl and Mertk leads to insufficient phagocytic function of apoptotic neutrophils (but does not affect other immune cells), which in turn hinders the phenotypic transformation of pancreatic macrophages to a reparative state. Therefore, adoptive transfer of M2 macrophages can restore pancreatic regenerative capacity.

[0083] This study confirms that Axl and Mertk are selectively and inducibly expressed on pancreatic macrophages and play a crucial role in tissue repair after SAP. Specific loss of Axl and Mertk may lead to the accumulation of apoptotic neutrophils and hinder the phenotypic transition of macrophages to a pro-repair state, thereby impairing pancreatic tissue repair. Therefore, the pancreatic tissue repair process after SAP can be improved by administering Axl agonists, Mertk agonists, or adoptive transfer of M2 macrophages.

[0084] In some specific embodiments of this application, CCR5 inhibitors can accelerate pancreatic tissue repair and promote inflammation resolution after severe acute pancreatitis, especially in cases of macrophage AXL and MERTK deficiency.

[0085] In some specific embodiments, the CCR5 inhibitor is selected from Maraviroc. This FDA-approved drug can significantly shorten the conversion cycle in the treatment of acute pancreatitis.

[0086] In some specific embodiments, the CD22 inhibitor is selected from InVivoMab anti-mouse CD22.

[0087] In some specific implementations, the impaired pancreatic tissue repair capacity caused by the lack of Axl and Mertk can be compensated for by adoptive transfer.

[0088] In some specific implementations, adoptive transfer of macrophages can be used to treat severe acute pancreatitis. Studies have shown that adoptive transfer or engineered macrophages have potential value in reducing excessive inflammation in the heart, liver, and lungs and promoting tissue repair.

[0089] Furthermore, this application provides a pharmaceutical composition comprising one or more of an Axl agonist, a Mertk agonist, a CCR5 inhibitor, or a CD22 inhibitor.

[0090] In some specific embodiments, the pharmaceutical composition may also include a pharmaceutically acceptable carrier or excipient.

[0091] In some specific embodiments, the pharmaceutically acceptable carrier or excipient includes binders, buffers, thickeners, diluents, stabilizers, antioxidants, solubilizers, lubricants, disintegrants, preservatives, colorants, flavorings, emulsifiers, etc.

[0092] In some specific embodiments, the pharmaceutical composition is in the form of an injection, tablet, drops, powder, spray, gel, ointment, or lozenge.

[0093] In some specific embodiments, the pharmaceutical composition is used for the prevention or treatment of severe acute pancreatitis (SAP).

[0094] In some preferred embodiments, the pharmaceutical composition is used to promote the repair of pancreatic tissue damage in severe acute pancreatitis (SAP).

[0095] In some preferred embodiments, the pharmaceutical composition is used to promote the regeneration of damaged pancreatic acinar cells in severe acute pancreatitis (SAP).

[0096] In some preferred embodiments, the pharmaceutical composition is used to promote the relief of inflammation in severe acute pancreatitis (SAP).

[0097] In some specific embodiments, the pharmaceutical composition is used for the prevention or treatment of sepsis.

[0098] In a specific implementation, the total dose of any one or more of the Axl agonist, Mertk agonist, CCR5 inhibitor, or CD22 inhibitor is 1-100 mg / kg of the subject, preferably 5-25 mg / kg of the subject.

[0099] In a specific implementation, the method includes administering the drug orally, by injection, or by topical application to the subject.

[0100] The effective dose described in this application refers to an amount sufficient to achieve the desired therapeutic outcome or to affect undesirable symptoms but generally insufficient to cause harmful side effects, and this amount can be easily determined by those skilled in the art. In specific embodiments, it can be determined by a clinician based on the subject's age, symptoms, route of administration, etc.

[0101] This application reveals the specific expression of AXL and MERTK in pancreatic macrophages and their role in pancreatic tissue repair after SAP (Surgical Apnea-Potential). Axl and MERTK can improve the clearance of apoptotic neutrophils and induce a pro-repair phenotype in pancreatic macrophages. Furthermore, the interaction between Cxcr2+ neutrophils and CD163+ macrophages mediated by the CCL4-CCR5 axis is significantly enhanced. Therefore, the use of CCR5 inhibitors also has clinical application potential in promoting pancreatic tissue repair.

[0102] Example Unless otherwise specified, all other materials, reagents, etc. used in the following embodiments of this application are commercially available.

[0103] The main experimental materials and methods used in the embodiments of this application are as follows.

[0104] animal : C57BL / 6J mice (catalog number SM-001) were purchased from the Shanghai Model Organisms Center, China. Based on the C57BL / 6J genetic background, mice with conditional deletion of AXL and MERTK in myeloid-specific macrophages (Axl...) were constructed. Ly-sMΔ Mertk LysMΔ (Provided by Dr. Carla V. Rothlin, Yale University School of Medicine). As a control group, littermate Floxed mice (Axlf / fMertkf / f) were used to validate Axl. Ly-sMΔ MertkLysMΔ Genetic background of the mice. All experimental mice were housed in a 12-hour light / dark cycle environment with free access to standard feed. The experimental mice used in this study were all 8-12 weeks old, and the ratio of male to female mice was balanced in each experiment. All animal studies were approved by the animal ethics committees of Shanghai General Hospital (2019-A019-01) and Peking Union Medical College Hospital (XHDW-2023-154).

[0105] Induction and treatment of severe acute pancreatitis : A hyperstimulated pancreatitis model was induced by intraperitoneal injection of 100 μg / kg spirulina protein (HY-A0190, MedChemExpress) for 10 hours daily for two consecutive days. Biliary acute pancreatitis was induced by retrograde bile duct perfusion with sodium taurocholate (NaT, 2.5%, 2 μL / g). Mice were humanely sacrificed at designated time points. In some experiments, immediately after the last injection of spirulina protein into the severe acute pancreatitis model, 50 mg / kg maraviro (HY-13004, MedChemExpress) or a solvent was administered intraperitoneally.

[0106] Histology, immunohistochemistry and immunofluorescence : Pancreatic tissue fixed with 4% formalin was embedded in paraffin. The tissue was then sectioned into 4-micrometer thick sections and stained with hematoxylin and eosin (H&E) according to standard procedures. Histological scoring of the pancreatic tissue was based on the literature (Aberrantinnate immune activation following tissue injury impairs pancreatic regeneration, Alexandra E Folias et al.). PloS oneThe method described in 9, e102125 (2014) was followed. Specifically, two experienced pathologists randomly selected 10 images from each slide in a blinded manner and scored them from 0 to 5 points, assessing lobular integrity, acinar dedifferentiation, and immune cell infiltration. In the immunohistochemical experiments, pancreatic slides were dewaxed and hydrated before antigen retrieval was performed using EDTA solution (catalog number E673004, Sangon Biotech Co., Ltd.). The sections were then co-incubated with lysed caspase-3 antibody (catalog number 9661, Cell Signaling Technology), CD45 antibody (catalog number 70257, Cell Signaling Technology), αSMA (ab5694, Abcam), Col1a (72026, Cell Signaling Technology), arginase-1 (93668s, Cell Signaling Technology), CD86 (19589s, Cell Signaling Technology), and Ly6B.2 (MCA771GA, BioRad), visualized using an immunohistochemistry kit (PK4001, Vector Laboratories), and imaged using a microscope (DFC550, Leica).

[0107] Quantitative reverse transcription polymerase chain reaction : Total RNA was purified from pancreatic tissue or tissue-derived macrophages using Trizol reagent (catalog number 15596018CN, Ingenium Biotech). Subsequently, RNA (500 ng) was reverse transcribed into cDNA using the PrimeScript™ RT Master Mix kit (catalog number RR036A). Reverse transcription polymerase chain reaction (RT-PCR) was performed using the TB Green® Premix Ex Taq™ kit (catalog number RR420A) from Takara Bio Inc., via a QuantStudio 7 Flex real-time quantitative PCR system (Applied Biosystems, Inc.). mRNA expression levels were calculated using the relative CT method (2-ΔΔCT) and normalized with Rplp0 as an internal control. Primer sequences are detailed in Table 1.

[0108] Table 1. Mouse gene primer sequences used for RT-qPCR

[0109] Pancreatic leukocyte separation : Pancreatic tissue was minced and digested for 20 minutes at 37°C in digestion buffer containing 2 mg / mL type IV collagenase (C5138, Sigma Aldrich). The tissue fragments were resuspended in 40 mL of ice-cold stop buffer containing 10% fetal bovine serum, filtered through a 70 μm cell sieve, centrifuged, and lysed with 1 mL of ACK lysis buffer (A10492-01, ThermoFisher Scientific) for 1 minute. The remaining leukocytes were washed twice with FACS buffer in PBS containing 2% FBS for single-cell RNA sequencing analysis or flow cytometry.

[0110] Single-cell RNA sequencing (scRNA-seq) : CD45+ cells were sorted from pancreatic leukocytes using the Invitrogen Magni-Sort™ Mouse CD45 Positive Selection Kit (catalog number 8802-6865). Dead cells were removed using a dead cell removal kit (catalog number 130-090-101, Miltenyi Biotec). Single cells were encapsulated in droplet emulsions using GemCode Technology (10×Genomics) according to the manufacturer's instructions. cDNA was reverse transcribed, barcoded, and amplified to construct a library using the Chromium Next GEM Single Cell 3' Kit v3.1 (catalog number 1000268, 10×Genomics). Paired-end sequencing of the cDNA library was performed on an Illumina Nova 6000 PE150 sequencing platform.

[0111] Raw reads were demultiplexed using Cell Ranger v7.0.1 and aligned with the mm10 reference genome using the STAR alignment tool. A gene counting matrix was generated based on the alignment results. Ribosomal genes were screened from the detected gene matrix, and low-quality cells were removed based on the following criteria: fewer than 800 or more than 6000 genes, more than 30,000 unique molecular identifiers (UMIs), and mitochondrial genes accounting for more than 10%. The remaining data were standardized and logarithmically transformed. Normalization and logarithmic transformation were performed using the scanpy package (SCANPY: Large-Scale Single-Cell Gene Expression Data Analysis) in Python 3. Highly variable genes were identified using this software, and principal component analysis (PCA) was performed using default parameters. Two-dimensional visualization was performed using uniform manifold approximation and projection (UMAP), and cell clustering was identified using the "Louvain" algorithm. Cell markers for each cluster were determined by screening using the "Ranked Genomes" algorithm, and cell types were annotated based on the highly expressed genes in each cluster. After calculating the number of each cell type in each sample, differential proportion analysis was used to detect the changes in cell population proportions between the Axlf / f Mertkf / f and AxlLysMΔ MertkLysMΔ groups. Ligand-receptor interaction analysis was performed using the CellphoneDB database (version 4.1.0, https: / / www.cellphonedb.org / ). Heatmaps and scatter plots were generated using R packages with default parameter settings for visualization. The raw data has been submitted to the NCBI Sequence Reading Archive (SRA). Database ID: PRJNA1147010.

[0112] Flow cytometry and fluorescence-activated cell sorting : Pancreatic leukocytes or bone marrow-derived macrophages (BMDMs) were incubated with an Fc blocker (catalog number 553142, BD Biosciences) in FACS buffer for 10 minutes. For surface staining, cells were incubated with the following antibodies: Pacific Blue-labeled CD45 antibody (clone S18009F, Biolegend), FITC-labeled CD11b antibody (clone M1 / 70, eBioscience), PE-F4 / 80 or PE / Cy7-F4 / 80 fluorescent markers (clone BM8, Biolegend), PE / Cy7-AXL antibody (clone MAXL8DS, Biolegend), and APC / Cy7-MERTK (clone DS5MMER, Biolegend). For intracellular staining, cells were fixed and permeabilized using the BD Cytofix / Cytoperm kit (554714, BD Bioscience), followed by staining with PE-iNOS (cloned CXNFT, eBioscience) and APC-arginase (cloned AlexF5, eBioscience). Cells were collected using a BD LSRFortessa flow cytometer (BD Biosciences) and analyzed using FlowJo software (version 10.8.1, Tree Star). To obtain pancreatic macrophages, CD45+CD11b+ F4 / 80+ cells were sorted using the BD FACSAria III sorting system (BD Biosciences).

[0113] Isolation of BMDM and adoptive transfer of IL-4 polarized macrophages : Bone marrow-derived macrophages (BMDMs) were isolated from and differentiated from bone marrow cells. Specifically, bone marrow cells were washed from the femur and tibia of mice with ice-cold PBS. Cells were subjected to density gradient centrifugation in a Percoll (40501ES60, Yeosen) and then co-cultured in DMEM (11965092, Gibco) with 20 ng / ml mouse M-CSF (96-315-02-10, Peprotech) for 6 days to prepare BMDMs. BMDMs were then treated with 50 ng / ml IL-4 (96-214-14-20, Peprotech) for 24 hours to differentiate into M2 macrophages. For adoptive transfer of macrophages, 2 × 10⁶ IL-4 was administered intravenously immediately after the last injection of cholecystokinin in mice with severe acute pancreatitis. 6 One M2 macrophage.

[0114] RNA in situ hybridization and quantification : RNA in situ hybridization experiments were performed on formalin-fixed paraffin-embedded tissues using the RNAscope Multiplex Fluorescence Kit v2 (catalog number 323100, Advanced CellDiagnostics [ACD]), strictly following the manufacturer's instructions. The specific procedure was as follows: First, the slides were placed in a hybridization oven and treated sequentially with hydrogen peroxide solution, target retrieval reagent, and proteinase IV; then, they were incubated with mouse probes provided by ACD (including Ccl4, Ccr5, Ptprc, Cd22, Il1b, and Adrb2). After hybridization, signal amplification was achieved using a six-step amplification reaction buffer. Finally, images were acquired at 60x magnification using a Nikon AXR microscope and analyzed using the open-source software CellProfiler 54. We referenced the following: (Colocalization of different influenza viral RNA segments in the cytoplasm filament viral budding as shown by single-molecule sensitivity FISH analysis, Yi-ying Chou et al.) PLoS Pathog The method described in 9, e1003358 (2013) designed a CellProfiler analysis workflow to quantify the number of RNA spots and their colocalization in each fluorescence channel. Colocalization analysis was performed using the RelateObjects module of CellProfiler to count the number of double-positive RNA spots in each image.

[0115] Statistical analysis : Data are expressed as mean ± SEM. All statistical analyses were performed using Prism 8.0 (GraphPad software). Unpaired two-tailed Student's t-tests were used for comparisons between two groups. For comparisons of multiple groups, one-way ANOVA followed by Tukey's post-hoc test was performed.

[0116] In experiments with multiple independent variables, comparisons were evaluated using two-way ANOVA and Tukey's multiple comparison test. A p-value < 0.05 was considered statistically significant. Data analysis and plotting were performed using Prism 8 (GraphPad software). Sample size was not pre-determined using statistical methods. No experimental data were excluded.

[0117] Main reagents: The CCR5 inhibitor used was Maraviroc, purchased from MedChemExpress, catalog number HY-13004.

[0118] The CD22 inhibitor used is InVivoMab anti-mouse CD22, purchased from Bioxcell, catalog number BE0011.

[0119] Example 1: Correlation study of AXL and MERTK expression levels with tissue damage and repair during severe acute pancreatitis Receptors that sense apoptotic cell signals (including TAM receptors Tyro3, ​​AXL, and MERTK) are mainly expressed in innate immune cells, especially macrophages and dendritic cells (DCs). To investigate the expression patterns of these cells in pancreatic immune cells, single-cell RNA sequencing was used to sort pancreatic CD45+ cells at baseline (without pancreatitis) and on day 4 after pancreatitis induction. Unsupervised clustering analysis based on the Seurat algorithm, combined with established immune cell-specific markers, identified six major immune cell types: B cells, dendritic cells, macrophages, natural killer (NK) cells, neutrophils, and T cells (e.g., B cells, dendritic cells, macrophages, natural killer (NK) cells, neutrophils, and T cells). Figure 1 (As shown in A). Axl and Mertk were selectively expressed on pancreatic macrophages, while Tyro3, ​​Timd4, Adgrb1, and Itgb3 showed extremely low expression on both pancreatic macrophages and infiltrating immune cells. Furthermore, Cd36 and Itgav did not exhibit cell-specific expression patterns—they were expressed not only on pancreatic macrophages but also on dendritic cells (DCs) and / or NK cells (e.g., Figure 1 (As shown in B). Further observation revealed that after inducing pancreatitis, the expression levels of Axl and Mertk in macrophages were significantly upregulated, while their levels were not detected in other immune cells (e.g., Figure 1 (as shown in C).

[0120] Furthermore, the dynamic changes of Axl and Mertk in pancreatic macrophages during SAP were detected by flow cytometry (experimental procedure as follows). Figure 1 (As shown in D). The results showed that the expression levels of Axl and Mertk were upregulated on day 2, gradually decreased from day 4 to day 14, and returned to baseline levels on day 14 (as shown in D). Figure 1 (As shown in E and 1F). These results suggest that Axl and Mertk in macrophages may play an important role in mediating tissue damage and repair during SAP.

[0121] Example 2: Effects of Axl and Mertk on pancreatic tissue repair To determine the specific roles of AXL and MERTK in tissue damage and repair within macrophages, we obtained AXL and MERTK-deficient mouse strains from Dr. Carla Rothlin of Yale School of Medicine. Given that AXL and MERTK are primarily expressed in pancreatic macrophages as Axl and Mertk (Axl... LysMΔ Mertk LysMΔ The study of Axl genes showed that the pancreatic macrophage response to pancreatitis is primarily supplemented by circulating monocytes in the bone marrow. f / f Mertk f / f Mice were crossed with the Cre lineage (LysM-Cre) targeting myeloid mononuclear cells to successfully construct a mouse model mouse with myeloid cell-specific deletions of Axl and Mertk.

[0122] A cyanin-induced SAP model was induced by hourly injections over two consecutive days. Pancreatic damage was assessed on day 2, and tissue repair was assessed on days 4, 6, 8, and 14. The study found that myeloid cell-specific knockout of Axl and Mertk (Axl...) LysMΔ Mertk LysM Δ On day 2, it had no significant effect on pancreatic injury, but histological assessments, including lobular integrity, acinar dedifferentiation, and inflammatory infiltration, revealed that the knockout mutation significantly delayed the tissue repair process following cyanin-induced pancreatitis (e.g., Figure 2 (As shown in A and 2B).

[0123] To rule out the influence of cyanin-induced experimental pancreatitis on Axl LysMΔ Mertk LysMΔ To investigate the mouse model-specific effects, we established a separate SAP model using retrograde biliary-pancreatic infusion of sodium taurocholate. Specifically, the degree of tissue damage was assessed one day after induction of pancreatitis, and tissue repair was assessed four days later (experimental procedure as follows). Figure 3 (As shown in A). Similarly, the experimental results also showed that in NaT-induced pancreatitis, myeloid cell-specific knockout of the Axl and Mertk genes had no effect on pancreatic damage on day 1, but significantly delayed the pancreatic tissue repair process on day 4 (e.g., Figure 3 (As shown in B and 3C). These data collectively demonstrate that Axl and Mertk, derived from circulating monocytes, play a crucial role in pancreatic tissue repair after SAP.

[0124] Example 3: Effects of Axl and Mertk on apoptotic cells Subsequently, immunostaining was performed on CD45 and caspase-3. The results showed that after myeloid-specific knockout of AXL and MERTK, persistent immune cell infiltration and apoptotic cell aggregation were observed in the pancreas at all test time points (e.g., Figure 2 (As shown in C and 2D). This finding suggests that myeloid cell-specific knockout of AXL and MERTK leads to an abnormal accumulation of apoptotic immune cells in the pancreas, thereby persistently triggering an inflammatory response during SAP and resulting in unhealed tissue damage.

[0125] Example 4: Effects of Axl and Mertk on the expression levels of inflammation-related factors To determine the effects of AXL and MERTK on the inflammation-related gene signatures of reparative macrophages in SAP, we analyzed data from AXL... LysMΔ Mertk LysMΔ CD45+CD11b+ F4 / 80+ macrophages were sorted from the pancreas of WT mice, and changes in the expression of pro-inflammatory, anti-inflammatory, and pro-repair genes were detected during the first week of SAP (experimental procedure as follows). Figure 4 (As shown in A).

[0126] The experiment revealed that pancreatitis initially induces upregulation of pro-inflammatory genes (such as Il1b, Il6, and Tnf) starting from day 2, followed by a gradual decrease in their levels, approaching baseline by day 8. However, specific knockout of Axl and Mertk genes in myeloid cells significantly enhanced the upregulation of related pro-inflammatory genes, a trend that began on day 2 and persisted at all detection time points (e.g., day 2). Figure 4 (As shown in B). On the other hand, pancreatitis induces downregulation of repair-related genes (including Mrc1 and Retnla) in sorted macrophages, a phenomenon that also begins on day 2, gradually increases, and reaches baseline levels on day 8. However, after myeloid-specific knockout of the Axl and Mertk genes, the downregulation of these repair genes is more pronounced on day 2, and the process of these genes returning to baseline levels on day 8 is significantly inhibited (e.g., ...). Figure 4 (as shown in C).

[0127] Example 5: Effects of Axl and Mertk on Phenotypic Transformation of Macrophages Studies have shown that macrophage phenotypic transformation is crucial for creating a supportive microenvironment, which is essential for pancreatic tissue repair after SAP. Therefore, flow cytometry was used to further analyze changes in macrophage phenotype during SAP.

[0128] The results showed that, although Axl LysMΔ Mertk LysMΔ The number of iNOS+ (M1) macrophages in mice was reduced on day 2 (e.g. Figure 4(As shown in D), but there was no difference in the number of M1 macrophages in the two groups of pancreas, indicating that AXL and MERTK have limited roles in mediating the M1 macrophage phenotype. On the other hand, Axl LysMΔ Mertk LysMΔ The number of arginase+ (M2) macrophages in mice was upregulated on day 2, and the tissue repair transition began on day 4, but remained lower than that in the wild-type control group (e.g., Figure 4 (As shown in E), this highlights the crucial roles of AXL and MERTK in promoting the phenotypic transformation of M2 macrophages. Similar results for Arg-1 gene expression were also observed in sorted macrophages. Taken together, these data suggest that myeloid cell-specific knockout of Axl and Mertk, due to insufficient phagocytic function, hinders the phenotypic transformation of macrophages to the pancreatic repair phenotype (M2 type), ultimately leading to impaired pancreatic tissue repair, although no effect was shown on the degree of tissue damage.

[0129] Example 6: The effect of reparative macrophages on pancreatic regeneration capacity To confirm the effect of reparative macrophages on pancreatic tissue repair after complete SAP induction, we used IL-4 polarized macrophages in vitro in Axl... LysMΔ Mertk LysMΔ Adoptive transfer experiments were conducted in mice. Specifically, bone marrow-derived macrophages were isolated and stimulated in vitro with IL-4 for 24 hours to prepare arginase-positive (M2) macrophages for adoptive transfer (experimental procedure as follows). Figure 5 (As shown in A and 5B).

[0130] Cell purity was confirmed by flow cytometry; over 90% of IL-4-stimulated cells were arginase-positive (e.g., ...). Figure 5 (As shown in C). Histological experiments revealed that, compared with Axl and Mertk-deficient mice that had not received adoptive transfer, the pancreatic tissue of Axl and Mertk-deficient mice that received adoptive transfer showed significantly better results on day 4, as assessed by histopathological scores for lobular integrity, acinar dedifferentiation, and inflammatory infiltration (e.g., Axl and Mertk-deficient mice that received adoptive transfer). Figure 5 (As shown in D). Furthermore, CD45+ immunostaining revealed that adoptive transfer using arginase-rich macrophages significantly reduced pancreatic immune cell infiltration (e.g., ...). Figure 5 (As shown in E). The number of apoptotic cells detected by caspase-3 was also significantly reduced (e.g., Figure 5 (As shown in F). These data combined indicate that adoptive transfer of in vitro polarized arginase+ macrophages can effectively restore pancreatic tissue repair dysfunction, and its mechanism of action is to enhance the clearance of apoptotic cells within the pancreas.

[0131] Example 7: Effects of Axl and Mertk on the interaction between Cxcr2+ neutrophils and CD163+ macrophages Myeloid cell-specific loss of Axl and Mertk enhances the interaction between Cxcr2+ neutrophils and CD163+ macrophages via the CCL4-CCR5 axis. Effective repair of tissue damage depends on the coordinated interaction between the local immune microenvironment and the tissue regeneration response. Given that adoptive transfer of IL-4-polarized macrophages in vitro can restore the damaged pancreas's repair capacity by enhancing the clearance of apoptotic neutrophils, the interactions among pancreatic infiltrating immune cells (especially neutrophils and macrophages) were further investigated. Specifically, on day 4 after inducing SAP culture, adoptive transfer of IL-4-polarized macrophages from Axl... LysMΔ Mertk LysMΔ CD45+ immune cells were sorted from the pancreas of wild-type littermates for single-cell RNA sequencing (e.g., Figure 6 As shown in A). Dimensionality reduction was performed using uniform manifold approximation and projection (UMAP) analysis and unsupervised clustering. Combined with established immune cell standard markers and SingleR technology, a total of 17 cell types were identified, including B cells, dendritic cells, monocytes, macrophages, NK cells, neutrophils, and T cells (e.g., B cells, NK cells, neutrophils, and T cells). Figure 6 (as shown in B).

[0132] Expression analysis of specific marker genes confirmed the specificity of each marker for different cell populations (e.g., Figure 7 (As shown in A). Differential proportion analysis showed that among the 17 cell populations, only Cxcr2+ neutrophils were significantly higher in Axl. LysMΔ Mertk LysMΔ Significantly elevated in mice (e.g.) Figure 7 (as shown in B).

[0133] Flow cytometry was used to further examine the dynamic changes of infiltrating immune cells in the pancreas during the first week after SAP surgery. The results showed that only on day 4, Axl... LysMΔ Mertk LysMΔ A significant increase in the number of pancreatic infiltrating neutrophils was observed in mice, while no significant differences were observed in the number of immune cells such as monocytes, macrophages, and dendritic cells at other time points (e.g., ...). Figure 6 (As shown in C and 7C). This indicates that the orderly clearance mechanism of pancreatic infiltrating neutrophils plays a crucial role in tissue repair during SAP. Future research may be needed to elucidate the heterogeneity and functional characteristics of pancreatic neutrophils during SAP, thereby better understanding their role in mediating the dynamic process of pancreatic tissue damage and repair.

[0134] Next, the CellPhone DB235 database was used to analyze the interactions between Cxcr2+ neutrophils and other innate immune cell subsets, including monocytes, dendritic cells, and macrophages. Notably, the interactions between Cxcr2+ neutrophils and two macrophage clusters (CD163+ Mrc1+ / high macrophages) were more significant, as evidenced by the enrichment of ligand-receptor pairs (…). Figure 6 D) can be confirmed. Furthermore, Axl LysMΔ Mertk LysMΔ The predicted interaction patterns were consistent with those of its littermate control group. Among the predicted ligand-receptor pairs, three potentially valuable pairings were selected: CCL4-CCR5, IL1B-ADRB2, and PTPR-CCD22, which were further validated using in situ RNA hybridization. Experimental data showed that the number of RNA spots in CCL4-CCR5 was significantly higher than that in PTPRC, CD22, IL1B, and ADRB2 (e.g., ...). Figure 6 Colocalization analysis shows that in Axl (as shown in E). LysMΔ Mertk LysMΔ In mice, the number of CCL4-CCR5 double positive spots was significantly increased, while the number of PTPRC_CD22 or IL1B_ADRβ2 double positive spots did not change significantly (e.g. Figure 6 (as shown in F, 7D, and 7E).

[0135] The comprehensive analysis results indicate that after myeloid cell-specific knockout of Axl and Mertk, the signaling pathway interaction between CXCR2+ neutrophils and CD163+ Mrc1+ / high macrophages was significantly enhanced, and this process is likely achieved through the CCL4-CCR5 axis.

[0136] Example 8: Effects of CCR5 inhibition on pancreatic tissue repair and inflammation Given the experimental results above, which show that the enhanced crosstalk between Cxcr2+ neutrophils and CD163+ macrophages is achieved through the CCL4-CCR5 axis, it is speculated that impaired pancreatic tissue repair caused by myeloid cell-specific Axl and Mertk defects may be compensated for by inhibiting CCR5 to disrupt the enhanced interaction between these immune cells. Therefore, the therapeutic effect of CCR5 inhibitors on pancreatic tissue repair after SAP surgery was further tested.

[0137] Maraviroc is an FDA-approved selective CCR5 antagonist. Following SAP induction, it was administered daily via intraperitoneal injection (50 mg / kg), and pancreatic tissue repair was assessed on day 4 of induction (experimental procedures as follows). Figure 8(As shown in A). Pancreatic histopathological scoring showed that CCR5 inhibitors could promote the growth of wild-type mice and Axl. LysMΔ Mertk LysMΔ Repair of pancreatic tissue in mice (e.g.) Figure 8 (As shown in B and 8C). Immunostaining with CD45 revealed a significant reduction in pancreatic immune cell infiltration in both groups of mice treated with CCR5 antagonists. More importantly, under CCR5 inhibitor treatment, the number of CD86+ macrophages (indicating M1 macrophages) in the pancreas of WT mice remained unchanged, while Axl... LysMΔ Mertk LysMΔ These cells were significantly reduced in mice; meanwhile, arginase-rich macrophages were increased in WT mice, while in Axl mice... LysMΔ Mertk LysMΔ In mice, recovery was observed. Immunostaining with Ly6B.2 revealed that Axl... LysMΔ Mertk LysMΔ Increased neutrophil infiltration in the mouse pancreas, but after treatment with a CCR5 antagonist, Axl... LysMΔ Mertk LysMΔ This cell was significantly reduced in WT mice (e.g.) Figure 8 (As shown in D and 8E). These data suggest that during SAP, macrophage subsets expressing CCR5 are likely to be reprogrammed into a pro-repair phenotype, with or without Axl and Mertk.

[0138] Example 9: Effects of CD22 inhibitors on pancreatic tissue damage repair Previous observations suggest that the interaction between Cxcr2+ neutrophils and Mrc1+ macrophages induced by myeloid-specific knockout of Axl and Mertk may be mediated by the PTPRC-CD22 axis. Furthermore, studies have shown that CD22 gene knockout or drug inhibition primarily promotes the clearance of myelin fragments, Aβ oligomers, and α-synuclein fibrils by microglia by reactivating macrophage phagocytic function, and reprograms microglia to a homeostatic transcriptional state and improves cognitive function in aged mice. Therefore, it is hypothesized that intervening in these enhanced interactions between immune cells with CD22 inhibitors can compensate for the impaired pancreatic tissue repair caused by myeloid-specific Axl and Mertk gene knockout. Further experimental verification is needed.

[0139] Specific experimental methods: An 8-week-old C57 wild-type male mouse model of acute pancreatitis (AP) was induced by two days of injection of cerulein (CER) (dose 100 μg / kg, 10 times daily, 1 hour apart) to test the therapeutic effect of CD22 inhibitors on pancreatic tissue repair after AP. Daily intraperitoneal injections of FDA-approved InVivoMab anti-mouse CD22 (300 μg / mouse) were initiated immediately after AP induction, and pancreatic tissue repair was assessed on day 4 post-induction (experimental procedure as follows). Figure 9 As shown in Figure A). Pancreatic histopathological scoring (including lobar integrity, acinar dedifferentiation, and inflammatory infiltration) showed that CD22 inhibitors promoted pancreatic tissue repair in WT mice (e.g., Figure 9 (As shown in B and 9C).

[0140] The above results indicate that CD22 inhibitors promote tissue repair and inflammation resolution following severe acute pancreatitis.

[0141] The foregoing has merely illustrated the principles of this application. It should be understood that the scope of this application is not intended to be limited to the exemplary aspects described herein, but should include all currently known and future-developed equivalents. Furthermore, it should be noted that several improvements and modifications can be made without departing from the technical principles of this application, and these improvements and modifications should also be considered within the scope of this application.

Claims

1. Use of Axl agonists, Mertk agonists, CCR5 inhibitors, or CD22 inhibitors in the prevention or treatment of severe acute pancreatitis (SAP) or sepsis.

2. Use of Axl agonists, Mertk agonists, CCR5 inhibitors, or CD22 inhibitors in the preparation of medicaments or formulations for the prevention or treatment of severe acute pancreatitis (SAP) or sepsis.

3. The use according to claim 2, wherein, The prevention or treatment of severe acute pancreatitis (SAP) includes prevention or treatment during the tissue damage phase, repair phase, and postoperative phase of severe acute pancreatitis (SAP).

4. Use of Axl agonists, Mertk agonists, CCR5 inhibitors, or CD22 inhibitors in the preparation of drugs or formulations for promoting the repair of pancreatic tissue damage.

5. Use of Axl agonists, Mertk agonists, CCR5 inhibitors, or CD22 inhibitors in the preparation of drugs or formulations for promoting regeneration of pancreatic acinar cells after damage.

6. Use of Axl agonists, Mertk agonists, CCR5 inhibitors, or CD22 inhibitors in the preparation of medicaments or formulations for the treatment or relief of pancreatic inflammation.

7. A pharmaceutical composition comprising one or more of an Axl agonist, a Mertk agonist, a CCR5 inhibitor, or a CD22 inhibitor, preferably for the prevention or treatment of severe acute pancreatitis (SAP); or preferably for promoting the repair of pancreatic tissue damage; or preferably for promoting the regeneration of damaged pancreatic acinar cells; or preferably for relieving or treating pancreatic inflammation; or preferably for the prevention or treatment of sepsis.

8. A method for preventing or treating severe acute pancreatitis (SAP), comprising administering to a subject an effective dose of any one or more of an Axl agonist, a Mertk agonist, a CCR5 inhibitor, or a CD22 inhibitor.

9. Use of M2 macrophages in the prevention or treatment of severe acute pancreatitis (SAP) or sepsis.

10. Use of M2 macrophages in the preparation of drugs or cell preparations for the prevention or treatment of severe acute pancreatitis (SAP) or sepsis.

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