Use of creg1 protein in the preparation of a medicament or reagent for inhibiting the aging of hematopoietic stem cells

Abstract

The application discloses application of CREG1 protein in preparation of medicines or reagents for inhibiting hematopoietic stem cell aging, and belongs to the field of biological medicines. The application first finds that the CREG1 protein has the function of inhibiting hematopoietic stem cell aging (including inhibiting hematopoietic stem cell proliferation abnormality, maintaining hematopoietic stem cell resting state, reducing hematopoietic stem cell DNA damage, and weakening hematopoietic stem cell myeloid differentiation tendency). Therefore, the CREG1 protein has the prospect of being developed into the medicines or reagents for inhibiting hematopoietic stem cell aging, and also has the prospect of being developed into the medicines for improving the immune function decline, anemia, bone marrow hypofunction or blood system diseases caused by hematopoietic stem cell aging.

Description

Technical Field

[0001] This invention belongs to the field of biomedicine and relates to new uses of known proteins, specifically the application of CREG1 protein in the preparation of drugs or reagents that inhibit the aging of hematopoietic stem cells. Background Technology

[0002] Hematopoietic stem cells (HSCs) are a vital cell population that maintains lifelong hematopoiesis, possessing self-renewal and multi-directional differentiation capabilities. With age or exposure to environmental stressors, HSCs gradually undergo functional decline, manifested as reduced self-renewal capacity, altered differentiation bias, and weakened regenerative capacity. This process is known as hematopoietic stem cell senescence.

[0003] Hematopoietic stem cell aging is not only closely related to the decline of the body's immune function, but also to a variety of age-related diseases, such as anemia, bone marrow dysfunction, and hematological disorders. Therefore, finding drugs that can delay or inhibit hematopoietic stem cell aging is of great significance for the treatment of related diseases.

[0004] CREG (Cellular Repressor of E1A-Stimulated Genes) is a highly conserved secreted glycoprotein that plays a crucial role in cell differentiation, proliferation regulation, and tissue homeostasis. According to literature reports, CREG1 enhances p16 by transcriptionally regulating cell cycle-related genes. INK4a Induced cellular senescence (Reference: Moolmuang B, Tainsky MA. CREG1 enhances p16) INK4a - induced cellular senescence. Cell Cycle. 2011).

[0005] There are currently a lack of research reports on the effects of CREG protein on hematopoietic stem cell aging. Summary of the Invention

[0006] The first objective of this invention is to provide the application of CREG1 protein in the preparation of drugs that inhibit hematopoietic stem cell aging; the second objective is to provide the application of CREG1 protein in the preparation of reagents that inhibit hematopoietic stem cell aging; the third objective is to provide the application of CREG1 protein in the preparation of drugs that improve immune function decline, anemia, bone marrow dysfunction or hematological diseases caused by hematopoietic stem cell aging; the fourth objective is to provide a pharmaceutical composition that inhibits hematopoietic stem cell aging; and the fifth objective is to provide a cell culture reagent or cell cryopreservation reagent for inhibiting hematopoietic stem cell aging.

[0007] The above-mentioned objective of this invention is achieved through the following technical solution: Application of CREG1 protein in the preparation of drugs that inhibit the aging of hematopoietic stem cells.

[0008] In one specific embodiment, the inhibition of hematopoietic stem cell senescence includes one or more of the following: inhibiting abnormal proliferation of hematopoietic stem cells, maintaining the quiescent state of hematopoietic stem cells, reducing DNA damage to hematopoietic stem cells, and weakening the myeloid differentiation tendency of hematopoietic stem cells.

[0009] In one specific embodiment, the CREG1 protein is a recombinant CREG1 protein.

[0010] Application of CREG1 protein in the preparation of reagents to inhibit hematopoietic stem cell senescence.

[0011] In one specific embodiment, the inhibition of hematopoietic stem cell senescence includes one or more of the following: inhibiting abnormal proliferation of hematopoietic stem cells, maintaining the quiescent state of hematopoietic stem cells, reducing DNA damage to hematopoietic stem cells, and weakening the myeloid differentiation tendency of hematopoietic stem cells.

[0012] In one specific embodiment, the CREG1 protein is a recombinant CREG1 protein.

[0013] Application of CREG1 protein in the preparation of drugs to improve immune function decline, anemia, bone marrow dysfunction or hematological diseases caused by hematopoietic stem cell aging.

[0014] In one specific embodiment, the CREG1 protein is a recombinant CREG1 protein.

[0015] A pharmaceutical composition for inhibiting the aging of hematopoietic stem cells, comprising the active ingredient CREG1 protein, and further comprising a pharmaceutically acceptable carrier, diluent, or excipient.

[0016] A cell culture reagent or cell cryopreservation reagent for inhibiting hematopoietic stem cell senescence, comprising the active ingredient CREG1 protein, which inhibits hematopoietic stem cell senescence.

[0017] Beneficial effects: This invention is the first to discover that the CREG1 protein has the effect of inhibiting hematopoietic stem cell senescence (including inhibiting abnormal hematopoietic stem cell proliferation, maintaining the resting state of hematopoietic stem cells, reducing DNA damage to hematopoietic stem cells, and weakening the myeloid differentiation tendency of hematopoietic stem cells). Therefore, the CREG1 protein has the potential to be developed into a drug or reagent to inhibit hematopoietic stem cell senescence, and also has the potential to be developed into a drug to improve immune function decline, anemia, bone marrow dysfunction, or hematological diseases caused by hematopoietic stem cell senescence. Attached Figure Description

[0018] Figure 1The expression level of CREG1 protein in the bone marrow of aging mice was significantly decreased. In the figure, A is the result of Western blot detection of CREG1 protein in mouse bone marrow tissue, and B is a statistical graph of the relative expression level of CREG1 protein.

[0019] Figure 2 The study showed that treatment of senescent hematopoietic stem cells with recombinant CREG1 protein could inhibit their proliferation, maintain quiescence, reduce DNA damage, and weaken myeloid differentiation tendency. Among them, A is the experimental flowchart, B is the statistical chart of the number of HSCs recovered per well, C is the statistical chart of cell cycle distribution ratio, D is the statistical chart of the proportion of γH2AX positive cells, and E is the statistical chart of the proportion of CD41 positive cells.

[0020] Figure 3 The study showed that treatment of aging mice with recombinant CREG1 protein improved their bone marrow hematopoietic stem cell-related aging phenotype. In the figure, A is the experimental flowchart, B is the statistical graph of the number of HSCs in the bone marrow, C is the statistical graph of the number of short-term hematopoietic stem cells in the bone marrow, D is the statistical graph of the proportion of γH2AX positive cells, and E and F are the statistical graphs of the proportion of HSCs with polar distribution of Cdc42 and α-Tubulin, respectively.

[0021] Figure 4 The results of the competitive transplantation experiment are shown below. A is a flowchart of the competitive transplantation experiment, B is a statistical graph of the number of donor-derived cells in peripheral blood, C-E are graphs showing the changes in the chimerism rate of donor-derived cells in peripheral blood at different time points after transplantation, F is a statistical graph of the number of donor-derived cells in bone marrow, and G is a statistical graph of the composition of donor-derived myeloid cells, B cells, and T cells in the bone marrow of recipient mice. Detailed Implementation

[0022] The substantive content of the present invention will be described in detail below with reference to specific embodiments. However, those skilled in the art should know that the scope of protection of the present invention should not be limited to these specific embodiments.

[0023] Example 1: The expression level of CREG1 protein in the bone marrow of aging mice was significantly decreased. I. Experimental Materials The experimental animals were male C57BL / 6J mice, purchased from Hunan Slack Jingda Experimental Animal Co., Ltd. The mice were housed in an SPF-grade animal facility at a temperature controlled at 22±2°C and a relative humidity of 45%–70%, with 12-hour light-dark cycles and free access to food and water. The young group consisted of mice aged 2 months, and the aged group consisted of mice aged 24 months.

[0024] Bone marrow total protein lysis was performed using a mixture of RIPA lysis buffer and protease inhibitor. The primary antibodies used in Western blot analysis included anti-CREG1 antibody (Proteintech, 12220-1-AP) and anti-β-actin antibody (Cell Signaling Technology, #4970); the secondary antibody was HRP-labeled anti-rabbit IgG secondary antibody (Cell Signaling Technology, #7074). Chemiluminescence assays used ECL luminescent substrates.

[0025] II. Test Methods 1. Extraction of protein from mouse bone marrow tissue After euthanizing the mice, the femur and tibia were separated, and attached muscle and connective tissue were removed. The bone surface was washed with PBS and placed in a culture dish containing PBS. The epiphyseal end was cut off, and PBS was drawn into a syringe and inserted into the medullary cavity from one end. The bone marrow tissue was repeatedly flushed out into an EP tube. The tube was centrifuged at 4°C and 1000 rpm for 5 min, and the supernatant was discarded. An appropriate amount of RIPA lysis buffer was added to the collected bone marrow tissue or cell pellet (approximately 100–200 μL per 10 mg of tissue, adjusted according to the sample volume). The pellet was homogenized thoroughly on ice using a homogenizer or electric grinder until no obvious tissue clumps remained.

[0026] Place the homogenate on ice for 20–30 min to lyse thoroughly, shaking every 5–10 min during this period. Centrifuge at 4°C and 12,000–14,000 rpm for 15–20 min. Carefully aspirate the supernatant and transfer it to a new pre-chilled EP tube; this is the total protein extract.

[0027] 2. Expression of CREG1 in mouse bone marrow using Western blot method Prepare a separating gel and a 5% stacking gel of appropriate concentration. Add 20 μg of protein sample to each well. Electrophoresis on the stacking gel at a constant voltage of 80 V for approximately 30 min. After bromophenol blue enters the separating gel, adjust the voltage to 120 V and continue electrophoresis until the bromophenol blue reaches the bottom of the gel. After electrophoresis, construct a transfer membrane sandwich in the following order: sponge – filter paper – gel – PVDF membrane – filter paper – sponge. Transfer the membrane at a constant current of 250–300 mA in an ice bath for 90 min. After transfer, place the PVDF membrane in blocking buffer and block on a shaker at room temperature for 1 h. Incubate the membrane with diluted primary antibody overnight on a shaker at 4 °C. Western blot analysis was performed using 1:800 anti-CREG1 (Proteintech, Cat No. 12220-1-AP) antibody and 1:1000 anti-β-actin (Cell Signalling, USA) antibody as primary antibodies, and horseradish peroxidase-labeled goat anti-mouse (or anti-rabbit) antibody (Cell Signalling, USA) as secondary antibodies. The bands were visualized using an ECL kit (Amersham, USA). ImageJ 1.51 software was used to measure the grayscale values ​​of the bands and perform statistical analysis.

[0028] III. Test Results Western blot results showed that ( Figure 1 Compared with young mice, the expression level of CREG1 protein in the bone marrow tissue of aged mice was significantly reduced; gray-scale quantitative analysis showed that the relative expression level of CREG1 in the aged group was significantly lower than that in the young group. These results indicate that decreased CREG1 protein expression is associated with the aging state of the hematopoietic system, providing an experimental basis for further evaluation of the role of CREG1 protein in the regulation of hematopoietic stem cell aging.

[0029] Example 2: Treatment of senescent hematopoietic stem cells (HSCs) with recombinant CREG1 protein can inhibit their proliferation, maintain their resting state, reduce DNA damage, and weaken their myeloid differentiation tendency. I. Experimental Materials The experimental animals were 24-month-old male C57BL / 6J mice. Bone marrow mononuclear cells were used to sort senescent hematopoietic stem cells. Bone marrow mononuclear cells were pre-enriched with magnetic beads using Miltenyi Biotec's LineageCell Depletion Kit (mouse 130-090-858) and CD117 MicroBeads (mouse 130-091-224). - CD117 +This refers to the hematopoietic stem cell population obtained through sorting; the recombinant mouse CREG1 protein used is a commercially available recombinant protein, namely NovoProtein's Recombinant mouse CREG1 (C-His) (CM14); in vitro culture was performed using Zhongshan Jinqiao hematopoietic stem cell culture medium (DLM15506). Subsequent sorting and identification can be performed using flow cytometry antibodies including Lineage cocktail (BioLegend, 133303), Sca-1 (BioLegend, 108113), c-Kit (BioLegend, 105812), CD150 (BioLegend, 115925), and CD48 (BioLegend, 103403). Cell cycle analysis was performed using Ki-67 staining reagent (BD Biosciences, 556026) and Hoechst 33342 (Thermo Fisher Scientific, H3570); DNA damage was detected using γH2AX antibody (Cell Signaling Technology, #9718); and differentiation bias was detected using BD Biosciences' BD Pharmingen™ Purified Rat Anti-Mouse CD41 antibody (553847).

[0030] II. Test Methods 1. Isolation and identification of hematopoietic stem cells Single-cell suspensions were prepared from the bone marrow of 24-month-old wild-type mice. After filtration through a 40 μm cell filter, the cells were pre-enriched with magnetic beads using the Miltenyi Biotec Lineage Cell Depletion Kit (mouse, 130-090-858) and mouse CD117 MicroBeads (mouse, 130-091-224). Flow cytometry was then used for further sorting and identification to obtain hematopoietic stem cell populations for experimental use.

[0031] 2. CREG1 processing method The senescent hematopoietic stem cells obtained above were seeded into an in vitro culture system and cultured using hematopoietic stem cell culture medium (DLM15506) produced by Zhongshan Jinqiao Company. Recombinant mouse CREG1 protein (Cat. No.: CM14) from NovoProtein was added to the basal culture conditions for treatment. The final concentrations of CREG1 protein were set at 50 ng / mL, 100 ng / mL, and 200 ng / mL, respectively, and treated continuously for 5 days. A blank control group without CREG1 protein was also included. All cells were cultured at 37℃, 5% CO2, and 5% O2. After treatment, cells from each group were collected for subsequent analysis of cell proliferation, resting state, DNA damage, and myeloid differentiation tendency.

[0032] 3. Detection of cell proliferation and resting state The proliferation and resting status of hematopoietic stem cells after CREG1 treatment were evaluated using methods such as BrdU / EdU incorporation assay, Ki67 staining, and cell cycle analysis. For the BrdU / EdU incorporation assay, BrdU or EdU reagent was added to the cell culture system after treatment, and cells were collected after incubation for 4 hours. Fixation, permeabilization, and staining (either colorimetric or fluorescent) were performed according to the instructions of the corresponding detection kit. Flow cytometry was used to detect the proportion of BrdU or EdU-positive cells to evaluate cell proliferation levels. For Ki67 detection, treated hematopoietic stem cells were collected, fixed, and permeabilized before staining with Ki67 antibody. The proportion of Ki67-positive cells was analyzed by flow cytometry to evaluate cell proliferation status. Cell resting status was evaluated using cell cycle analysis. Treated cells were collected, fixed, and stained with DNA dye. Flow cytometry analysis was performed in conjunction with Ki67 expression. Cells that were Ki67-negative and had a DNA content of 2N were defined as G0 phase cells. The effect of CREG1 on the resting maintenance ability of hematopoietic stem cells was assessed by comparing the proportion of G0 phase cells in each group.

[0033] 4. DNA damage detection The level of DNA damage in hematopoietic stem cells after CREG1 treatment was evaluated using γ-H2AX assay. After treatment, hematopoietic stem cells were washed with PBS, fixed with 4% paraformaldehyde at room temperature for 10 min, then permeabilized with PBS containing 0.5% Triton X-100 for 10 min, and then blocked with blocking solution containing 3% bovine serum albumin at room temperature for 1 h. For flow cytometry, after fixation, permeabilization, and blocking, cells were incubated with Alexa Fluor 647 anti-H2A.X Phospho (Ser139) Antibody in the dark. After staining, the cells were washed, and the proportion of γ-H2AX positive cells was detected by flow cytometry to evaluate the level of DNA damage. For immunofluorescence, after fixation, permeabilization, and blocking, cells were incubated overnight at 4°C with the corresponding primary antibody, then washed three times with PBST, and then incubated with fluorescently labeled secondary antibody. After staining, imaging was performed using a confocal microscope (ZEISS LSM 880), and the difference in γ-H2AX positive signal between the treatment group and the control group was compared to assess the effect of CREG1 on DNA damage in hematopoietic stem cells.

[0034] 5. Analysis of myeloid differentiation tendency Flow cytometry was used to detect changes in the myeloid differentiation tendency of hematopoietic stem cells after CREG1 treatment. After culturing the treated HSCs for 48 h, the cells were collected, stained with PE anti-mouse CD41, and then analyzed by flow cytometry. The trend of hematopoietic stem cells towards myeloid differentiation was evaluated by comparing the proportion of CD41 positive cells in HSCs in the CREG1-treated group and the untreated control group.

[0035] III. Test Results Figure 2 The results showed that the abnormal proliferation of senescent hematopoietic stem cells was inhibited after treatment with recombinant CREG1 protein, and cell cycle analysis indicated an increased proportion of G0 phase resting cells. Simultaneously, the proportion of γH2AX-positive cells decreased, suggesting a reduction in DNA damage; and the proportion of CD41-positive cells decreased, indicating a weakening of myeloid-related differentiation bias. These results indicate that exogenous CREG1 protein can significantly improve the abnormal phenotype of senescent hematopoietic stem cells and has an inhibitory effect on hematopoietic stem cell senescence.

[0036] Example 3: After treatment of aged mice with recombinant CREG1 protein, the number of HSCs in the bone marrow of aged mice was reduced, DNA damage was significantly decreased, and cell polarity was restored.

[0037] I. Experimental Materials The aging mice were derived from the same source as in Example 1, and the recombinant protein and related flow cytometry antibodies were from the same source as in Example 2. The polarity-labeling staining antibodies were anti-Cdc42 (Santa Cruz, sc-8401) and anti-alpha-Tubulin Antibody (Cellsignalling Technology, #2144). II. Test Methods 1. Mouse treatment and grouping The aged mice were divided into two groups: Experimental group (recombinant CREG1 protein treatment group): The treatment was administered by intraperitoneal injection of CREG1 protein (1.5 mg / kg) daily for 7 consecutive days.

[0038] Control group (saline or carrier group): Intraperitoneal injection of saline or corresponding carrier solution was administered. Injections were given continuously for 7 days, and the treatment cycle could be increased or adjusted as needed for the experiment.

[0039] 2. Isolation and identification of hematopoietic stem cells The specific steps are the same as in Example 2.

[0040] 3. HSCs quantity detection HSCs Count: The sorted LSK cell population was counted by flow cytometry, and the absolute number of HSCs in the bone marrow of mice in the treatment and control groups was compared. Data Analysis: The number of LSK cells in different groups was counted to analyze whether recombinant CREG1 protein could significantly increase the number of HSCs in the bone marrow of aged mice.

[0041] 4. DNA damage analysis The specific steps are the same as in Example 2.

[0042] 5. Cell polarity analysis Polarity labeling staining: Immunofluorescence staining was performed using anti-CDC42 (1:200) or anti-alpha-Tubulin Antibody (1:500). Imaging was performed using a confocal microscope (ZEISS LSM 880). The effectiveness of CREG1 protein in restoring the polarity characteristics of HSCs was assessed based on the expression levels of CDC42 and Tubulin.

[0043] III. Test Results Figure 3 The results showed that, compared with the control group, treatment with recombinant CREG1 protein improved the abnormal accumulation of hematopoietic stem cell-related populations in the bone marrow of aged mice, reduced DNA damage levels, and partially restored abnormal cell polarity. These results suggest that CREG1 protein can improve the aging-related phenotypes of hematopoietic stem cells in aged bone marrow at the whole animal level.

[0044] Example 4: Competitive transplantation experiments confirmed that recombinant CREG1-treated aged HSCs significantly enhanced hematopoietic reconstitution capacity and increased lymphoid differentiation ratio in recipient mice. I. Experimental Materials Donor cells were derived from 24-month-old C57BL / 6J (CD45.2 background) aged mice, and were used for transplantation after in vitro CREG1 treatment or control treatment. Competitive cells were derived from bone marrow cells from CD45.1 background mice. Recipient mice had a CD45.1 / CD45.2 background and received tail vein transplantation after being irradiated with a lethal dose in fractions. Peripheral blood chimerism and lineage analysis were performed using flow cytometry against CD45.1 (BioLegend, 110714), CD45.2 (BioLegend, 109821), CD3ε (BioLegend, 100305), B220 (BioLegend, 103207), Gr-1 (BioLegend, 108415), and CD11b (BioLegend, 101225). The chimerism rate of bone marrow LT-HSC at the transplant endpoint was analyzed using flow cytometry panel F: CD45.1 (BioLegend, 110714), CD45.2 (BioLegend, 109821), Lineage cocktail (BioLegend, 133303), Sca-1 (BioLegend, 108113), c-Kit (BioLegend, 105825), CD150 (BioLegend, 115904), and CD48 (BioLegend, 103431).

[0045] II. Test Methods 1. Donor cell preparation (1) Separation of aged HSCs The specific steps are the same as in Example 2.

[0046] (2) Isolation and treatment of aged HSCs The specific steps are the same as in Example 2.

[0047] 2. Preparation of competing cells From competitive donor mice ( In the background, bone marrow mononuclear cells (BMMNCs) or whole bone marrow cells are isolated, counted, and used for later use. Competitive cells require no special treatment and are used for transplantation by mixing with experimental / control HSCs in a fixed ratio.

[0048] 3. Pretreatment of recipient mice Recipient mice were irradiated with a lethal dose 4–6 hours before transplantation. Irradiation was performed using X-rays (total dose 9.0–9.5 Gy) to deplete the endogenous hematopoietic system. After irradiation, mice were placed in a sterile environment and given antibiotics (such as sulfamethoxazole / trimethoprim) in their drinking water to prevent infection.

[0049] 4. Competitive transplantation Treated aged HSCs were mixed with competing cells at a predetermined ratio (common ratios are 1:1 or 1:2, depending on the number of HSCs). Recipient mice were injected via tail vein with the mixed cell suspension (100–200 μL) within 4–6 hours after irradiation. Eight–10 recipient mice were transplanted per group to ensure statistical power.

[0050] 5. Monitoring of hematopoietic reconstitution Blood samples were collected periodically after transplantation (e.g., at 4, 8, 12, and 16 weeks) via the tail vein to detect the chimerism rate and differentiation of donor-derived cells in the peripheral blood of recipient mice. (1) Detection of chimerism rate Peripheral blood was collected, and erythrocytes were lysed before flow cytometry analysis. CD45.1 and CD45.2 antibodies were used to differentiate between elderly donor HSCs. ) and competing cells ( The number and proportion of cells from which the source is located.

[0051] (2) Multilineage differentiation analysis Simultaneously detect myeloid ( B lymphatic system ( ), T lymphatic system ( Cells of various lineages, etc. The proportion of cells.

[0052] 6. Endpoint Analysis Sixteen weeks post-transplantation, recipient mice were sacrificed, and bone marrow was collected to analyze the chimerism rate of LSK cells and long-term HSCs (CD48⁻CD150⁺). The proportion of CD45.2⁺ cells in various lineages, including myeloid (Gr-1⁺ / Mac-1⁺), B lymphoid (B220⁺), and T lymphoid (CD3⁺), was also measured.

[0053] III. Test Results Competitive transplantation results showed ( Figure 4 Compared with the untreated control group, aged HSCs treated with recombinant CREG1 protein exhibited higher hematopoietic reconstitution capacity in recipient mice, increased donor-derived cell chimerism in peripheral blood and bone marrow, and a trend of increased lymphoid differentiation. These results indicate that CREG1 protein not only improves the aging phenotype of aged HSCs in vitro and in vivo, but also enhances their long-term functional hematopoietic reconstitution capacity.

[0054] Example 5: A pharmaceutical composition for inhibiting hematopoietic stem cell senescence A pharmaceutical composition for inhibiting the aging of hematopoietic stem cells, comprising the active ingredient CREG1 protein, and further comprising a pharmaceutically acceptable carrier, diluent, or excipient.

[0055] Example 6: A cell culture reagent for inhibiting hematopoietic stem cell senescence A cell culture reagent for inhibiting hematopoietic stem cell senescence contains the active ingredient CREG1 protein, which inhibits hematopoietic stem cell senescence. This cell culture reagent can be used for in vitro culture of hematopoietic stem cells to inhibit senescence of hematopoietic stem cells during culture.

[0056] Example 7: A cell cryopreservation reagent for inhibiting hematopoietic stem cell senescence A cell cryopreservation reagent for inhibiting hematopoietic stem cell senescence contains the active ingredient CREG1 protein, which inhibits hematopoietic stem cell senescence. This cell culture reagent can be used for hematopoietic stem cell cryopreservation to inhibit the senescence of hematopoietic stem cells during cryopreservation.

[0057] In summary, this invention is the first to discover that the CREG1 protein has the effect of inhibiting hematopoietic stem cell senescence (including inhibiting abnormal hematopoietic stem cell proliferation, maintaining the quiescent state of hematopoietic stem cells, reducing DNA damage to hematopoietic stem cells, and weakening the myeloid differentiation tendency of hematopoietic stem cells). Therefore, the CREG1 protein has the potential to be developed into a drug or reagent to inhibit hematopoietic stem cell senescence, and also has the potential to be developed into a drug to improve immune function decline, anemia, bone marrow dysfunction, or hematological diseases caused by hematopoietic stem cell senescence.

[0058] The purpose of the above embodiments is to specifically illustrate the substantive content of the present invention, but those skilled in the art should know that the scope of protection of the present invention should not be limited to the specific embodiments.

Claims

1. Application of CREG1 protein in the preparation of drugs that inhibit the aging of hematopoietic stem cells.

2. The application according to claim 1, characterized in that: The inhibition of hematopoietic stem cell aging includes one or more of the following: inhibiting abnormal proliferation of hematopoietic stem cells, maintaining the resting state of hematopoietic stem cells, reducing DNA damage to hematopoietic stem cells, and weakening the myeloid differentiation tendency of hematopoietic stem cells.

3. The application according to claim 1 or 2, characterized in that: The CREG1 protein is a recombinant CREG1 protein.

4. Application of CREG1 protein in the preparation of reagents to inhibit hematopoietic stem cell aging.

5. The application according to claim 4, characterized in that: The inhibition of hematopoietic stem cell aging includes one or more of the following: inhibiting abnormal proliferation of hematopoietic stem cells, maintaining the resting state of hematopoietic stem cells, reducing DNA damage to hematopoietic stem cells, and weakening the myeloid differentiation tendency of hematopoietic stem cells.

6. The application according to claim 4 or 5, characterized in that: The CREG1 protein is a recombinant CREG1 protein.

7. Application of CREG1 protein in the preparation of drugs to improve immune function decline, anemia, bone marrow dysfunction or hematological diseases caused by hematopoietic stem cell aging.

8. The application according to claim 7, characterized in that: The CREG1 protein is a recombinant CREG1 protein.

9. A pharmaceutical composition for inhibiting the aging of hematopoietic stem cells, characterized in that: It contains the active ingredient CREG1 protein, as well as pharmaceutically acceptable carriers, diluents, or excipients.

10. A cell culture reagent or cell cryopreservation reagent for inhibiting the senescence of hematopoietic stem cells, characterized in that: It contains CREG1 protein, an active ingredient that inhibits the aging of hematopoietic stem cells.

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