CN114324860A - Assessment method for correlation between plasmablasts and pancreatic islet immune injury - Google Patents
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Abstract
The invention discloses an evaluation method of correlation between plasmablasts and islet immune injury, which comprises the steps of separating a tissue sample, analyzing and sorting plasmablasts by flow cytometry, co-culturing cells and/or adoptive transferring cells, establishing an insulitis evaluation standard and the like. The evaluation method of the invention can effectively establish the correlation between the plasmablasts and the pancreatic islet immune injury, and provides possibility for early diagnosis of the immune injury of type 1 diabetes.
Description
Technical Field
The invention belongs to the technical field of biology and medicine, and particularly relates to an evaluation method for correlation between immune injury of plasmablasts and pancreatic islets.
Background
Type 1 diabetes is an autoimmune disease in which islet beta cell-specific immune damage results in an absolute deficiency of insulin. Once the patient is diagnosed, exogenous insulin therapy must be initiated as soon as possible to reduce the metabolic load of islet beta cells and reduce the risk of diabetic ketosis or ketoacidosis. The definite diabetes typing diagnosis and early recognition of type 1 diabetes are important for the formulation of a treatment scheme. At present, the type 1 diabetes mellitus lacks of clear diagnosis standards, is mainly judged according to clinical manifestations, has few laboratory indexes for auxiliary diagnosis, and is mainly determined by referring to islet autoantibodies, serum insulin or C peptide. However, the clinical features of type 1 diabetes mellitus people are heterogeneous, and the disease onset age, disease onset characteristics, islet functional state, islet autoantibody type and the like are greatly different, so that diagnosis of some patients is retrospectively, and diagnosis at the early stage of disease onset is very difficult.
The existing detection indexes for assisting clinical diagnosis only comprise islet autoantibodies and islet function determination, and a new method capable of effectively reflecting islet autoimmune damage needs to be searched urgently. Currently, a B lymphocyte subset involved in islet immune injury in type 1 diabetes has been discovered, but the correlation between plasmablasts and islet immune injury is unknown, and the effect of plasmablasts on islet immune injury in type 1 diabetes cannot be assessed.
Disclosure of Invention
Accordingly, the present invention is directed to a method for evaluating the correlation between the immune injury of plasmablasts and pancreatic islets, which can help search the correlation between the immune injury of plasmablasts and pancreatic islets and accurately evaluate the effect of plasmablasts on the immune injury of pancreatic islets of type 1 diabetes.
In order to achieve the purpose, the technical scheme of the invention is realized as follows:
the invention provides a method for evaluating correlation between plasmablasts and islet immune injury, which comprises the following steps:
1) isolating immune cells in pancreatic lymph node tissue and spleen of different stage type 1 diabetic patients or NOD model animals; preferably, the NOD model animals at different stages are NOD model animals raised to 4 weeks, 8 weeks, 12 weeks, 16 weeks, 20 weeks, 24 weeks; preferably, the NOD model animal is a NOD mouse;
2) analyzing immune cells, sorting plasmablasts, and constructing the relationship between the content of the plasmablasts and different stages of a type 1 diabetes patient or an NOD model animal, wherein the relationship is positive;
3) adoptive transfer of plasmablasts or a combination of plasmablasts and T lymphocytes, monitoring the rate and proportion of diabetes, and the condition of insulitis;
and/or co-culturing the T lymphocyte, the plasmablast and the pancreatic island to detect the apoptosis condition of the pancreatic island;
4) and establishing an insulitis evaluation standard, and scoring insulitis according to the infiltration degree of immune cells in the pancreatic islets.
The invention also provides an insulitis scoring standard: 0 minute, complete islets, no immune cell infiltration; 1 minute, only immune cells infiltrate around the islets, or the infiltration area of the immune cells in the islets is less than 25 percent; 2 minutes, the infiltration area of immune cells in the pancreatic islets reaches 25 to 50 percent; 3 minutes, the infiltration area of immune cells in the pancreatic islets reaches 50 to 75 percent; 4 min, the infiltration area of immune cells in the pancreatic islets exceeds 75 percent.
Separating pancreatic lymph nodes and spleens, grinding to obtain a cell suspension, and centrifuging the pancreatic lymph node cell suspension to obtain pancreatic lymph node immune cells; and adding erythrocyte lysate into the spleen cell sediment, and centrifuging after cracking to obtain spleen immune cells.
And the step 2) comprises transferring the immune cells obtained in the step 1) to a flow tube, adding a flow antibody for pre-mixing incubation, adding a dead cell dye for incubation, washing and re-suspending by a Stain buffer, and then using a flow cytometer to sort the required T lymphocytes and plasmablasts.
The step 3) comprises the steps of transferring T lymphocytes and plasmablasts to an NOD/SCID mouse together, monitoring random blood sugar, obtaining pancreatic tissues, slicing, carrying out HE staining and immunohistochemical staining, and collecting pictures.
The ratio of the T lymphocytes to the plasmablasts in the step 3) is 1-10: 10-1, and preferably 20: 2-5 or 2: 1.
The method for detecting islet apoptosis in the step 3) is a TUNEL method.
The T lymphocytes and the plasmablasts are transferred together into a constant-speed bolus injection from the fundus venous plexus; the random blood sugar is monitored by a tail-cutting method for NOD/SCID mice.
The invention also provides an application of the plasmablasts in preparing an islet immune injury agent, wherein the islet immune injury agent optionally further comprises T lymphocytes, and the ratio of the T lymphocytes to the plasmablasts is 1-10: 10-1, preferably 2:1 or 20: 2-5.
Specifically, the invention also provides a method for evaluating the correlation between plasmablasts and islet immune injury, which comprises the following steps:
separating a tissue sample: and (5) separating pancreatic lymph nodes, pancreas and spleen of NOD mice.
Immune cell preparation and islet culture: grinding spleen and pancreatic lymph node of NOD mouse to obtain cell suspension, adding cell lysate for lysis, and centrifuging to obtain immunocyte for use. Adding collagenase to digest pancreas, stopping digestion with serum-containing HBSS solution, selecting islets without fragments under a microscope, and culturing in culture solution.
Flow cytometry analysis: taking immune cells, transferring the immune cells into a flow tube, adding a flow antibody, premixing, incubating in a dark place, adding a dead cell dye, incubating in a dark place, washing and resuspending by a Stain buffer, and analyzing by a flow cytometer.
Preparation of T lymphocytes and plasmablasts: immune cells were taken and aseptically sorted for T lymphocytes and plasmablasts using flow cytometry.
Cell co-culture: the culture was carried out in 3 groups of 20-30 islets/well format using 96-well round bottom cell culture plates, and 2X 10 was added to group 15T lymphocytes and 1X 105One plasmablast was co-cultured with islets of Langerhans, 2 nd group was added with 2X 105Individual T lymphocytes were co-cultured with islets, group 3 islets alone. IL-2 was added at 10ng/mL per well. After cell culture, the TUNEL method detects islet apoptosis. And/or, adoptive transfer of cells: NOD/SCID mice were randomly divided into 3 groups, group 1 each receiving 2X 10 NOD mice6T lymphocytes and 2X 105-5×105Co-transfer of individual plasmablasts, each receiving the same number of T lymphocyte transplants in group 2, and an equal volume of PBS injection in group 3; random blood glucose was monitored in 3 groups of NOD/SCID mice after cell transplantation; NOD/SCID mice were sacrificed after disease onset, and were subjected to HE staining of the pancreas, microscopic examination, and insulitis scoring based on the degree of immune cell infiltration in the islets.
Here, the extraction of immune cells and islet culture are performed on NOD mice in different culture periods, so that gradient control is facilitated, and analysis is facilitated.
Further, the cell CO-culture is specifically performed by gently shaking the culture plate in a circling manner to gather the islets and the immune cells toward the middle, directly contacting the islets and the immune cells, culturing the islets and the immune cells in an incubator with 5% CO2 at 37 ℃ for 72 hours, sucking out the mixture in each well, washing the mixture in an EP tube filled with 1mL of PBS, and depositing the islets at the bottom of the EP tube. The supernatant was carefully aspirated and discarded, washed again, and islet apoptosis was detected by TUNEL.
Furthermore, T lymphocytes and plasmablasts in the adoptive transfer of cells are extracted from NOD mice of 16 weeks old, and the cell transplantation mode is constant-speed bolus injection from the fundus vein cluster; random blood glucose was monitored in NOD/SCID mice by tail-snip.
Further, the insulitis scoring criteria are: 0 minute, complete islets, no immune cell infiltration; 1 minute, only immune cells infiltrate around the islets, or the infiltration area of the immune cells in the islets is less than 25 percent; 2 minutes, the infiltration area of immune cells in the pancreatic islets reaches 25 to 50 percent; 3 minutes, the infiltration area of immune cells in the pancreatic islets reaches 50 to 75 percent; 4 minutes, the infiltration area of immune cells in the pancreatic islets exceeds 75 percent; at least 100 islets were observed per group of NOD/SCID mice for scoring.
In the invention, the insulitis scoring standard can assist in searching the correlation between plasmablasts and islet immune injury, and accurately evaluate the effect of plasmablasts on islet immune injury of type 1 diabetes.
The invention has the following beneficial effects: 1) the assessment method for the correlation between the plasmablasts and the pancreatic islet immune injury can assist in searching the correlation between the plasmablasts and the pancreatic islet immune injury, and accurately assess the effect of the plasmablasts on the pancreatic islet immune injury of type 1 diabetes; 2) the method co-cultures the plasmablasts and the T lymphocytes with the pancreatic islets and adoptively transfers the plasmablasts and the T lymphocytes, so that the synergistic response of the plasmablasts and the T lymphocytes is fully shown, the pancreatic islet immune injury effect is caused, and the possibility is provided for the early diagnosis of the immune injury of the type 1 diabetes; 3) the plasmablast can be used as pancreatic islet immune injury agent.
Drawings
FIG. 1 is a graph showing the proportion of plasmablasts in the spleen of a mouse according to an embodiment of the present invention;
FIG. 2 is a graph showing the proportion of plasmablasts in the lymph nodes of the pancreas of a mouse according to an embodiment of the present invention;
FIG. 3 shows the plasmablast infiltration in the islets of Langerhans of a mouse according to an embodiment of the present invention;
FIG. 4 shows the detection of islet apoptosis in different culture conditions by the TUNEL method according to an embodiment of the present invention;
FIG. 5 is a graph showing the blood glucose changes in 16-week-old NOD/SCID mice according to the present invention;
FIG. 6 is a graph showing the degree of insulitis in 16-week-old NOD/SCID mice of the present invention.
Detailed Description
So that the manner in which the features and aspects of the present invention can be understood in detail, a more particular description of the invention, briefly summarized above, may be had by reference to embodiments, some of which are illustrated in the appended drawings.
Example 1
Immune cell extraction:
1) isolation of NOD mouse pancreatic lymph nodes, pancreas, spleen: NOD mice were sacrificed in batches at 4, 8, 12, 16, 20, and 24 weeks of age. Mice were fixed on an operating plate and the abdomen was moistened with an alcohol cotton ball. The fur and muscle layers are cut layer by layer, and the internal organs are exposed. A pair of oval lymph nodes along the blood vessels on the pancreas were carefully separated, placed in PBS, and placed on ice. Pancreatic tissue was detached from the edge of the intestine and immersed in 4% paraformaldehyde fixing solution. Spleen was separated from pancreas, placed in PBS and placed on ice.
2) Immune cells in spleen and pancreatic lymph node of mouse are extracted: place a disposable sterile screen at a 50mL centrifuge nozzle, place spleen on screen, and carefully grind spleen. The cell suspension in the centrifuge tube was centrifuged, and the supernatant was discarded. Adding 1 Xerythrocyte lysate, and lysing at room temperature for 10 min. After termination of the lysis, centrifugation was carried out and the supernatant was discarded. Washing the cells once, centrifuging, and discarding the supernatant to obtain spleen immune cells at the bottom of the tube.
The immune cells in the pancreatic lymph nodes were extracted from the spleen.
Example 2
Culturing the pancreatic islets:
the mice of 5 weeks old are anesthetized and fixed, the fur and muscle layers are cut, the chest is opened to expose the heart, and care is taken to avoid abdominal bleeding. Find the opening of common duct of pancreas duodenum, ligate with thread. Cutting right auricle, releasing venous blood, finding common bile duct near hepatic portal, intubating, and injecting collagenase. The pancreas was carefully separated and immediately placed on ice. A50 mL plastic centrifuge tube containing pancreatic tissue on ice was supplemented with about 5mL collagenase V and digested at 37 ℃ for 26-28 min. After digestion, immediately placing a 50mL plastic centrifuge tube on ice, and vortexing until the tissue is silt-shaped, for 5 seconds/time, 3 times in total; digestion was stopped by adding 2 volumes of pre-cooled serum-containing HBSS solution. Sieving with 30 mesh sieve, removing undigested tissue mass, washing with HBSS for 3 times, discarding supernatant, adding 5mL Histopaque, blowing, mixing, slowly adding 5mL HBSS along tube wall, and separating the solution into two layers. After centrifugation, the white sandwich layer in the middle of the centrifuge tube was carefully and quickly aspirated by a 1mL pipette, transferred to a petri dish containing serum HBSS, and intact, moderately sized and debris-free islets were selected under a stereomicroscope and placed in a culture solution of RPMI 1640+ 10% FBS + diabody.
Example 3
Immune cell analysis:
spleen and lymph node immune cells from example 1 were transferred to flow tubes. Premixing flow antibodies of mouse anti-B220(PerCP-Cy5.5), anti-CD138(APC), anti-CD44(BB515) and anti-CD3(PE-Cy7), adding the premixed flow antibodies into a flow tube, uniformly mixing the premixed flow antibodies with cells, and incubating the mixed solution at room temperature for 14-20 minutes in a dark place. 1mL of PBS was added, 400g was centrifuged for 5min, and the supernatant was discarded. Then, 1mL of PBS and 1. mu.l of Fixable Viatility Stain (FVS)780 were added, and incubated for 10-15min at room temperature in the absence of light. 1mL of Stain buffer (PBS + 1% FBS) was added, 400g was centrifuged for 5min, and the supernatant was discarded. The cells obtained at the bottom of the tube were resuspended in 300. mu.l Stain buffer (PBS + 1% FBS). The analysis was performed using a BD FACAIra III flow cytometer, and the data obtained were analyzed using FlowJo 10.4 software.
FIG. 1 is a graph showing the proportion of plasmablasts in the spleen of mice according to the present invention, and as shown in FIG. 1, the proportion of plasmablasts in the spleen of NOD mice was low before insulitis occurred, and as the disease progressed, the proportion of plasmablasts in the spleen of 8w, 12w and 16w NOD mice increased, and was significantly higher than that of 4w NOD mice.
FIG. 2 shows the ratio of plasmablasts in the pancreatic lymph nodes of mice according to an embodiment of the present invention, and as shown in FIG. 2, the ratio of plasmablasts in the pancreatic lymph nodes of NOD mice was low before insulitis occurred, and increased significantly as insulitis progressed to 8w, 12w, and 16 w.
FIG. 3 shows the plasmablast infiltration in the islets of mice according to the present invention, as shown in FIG. 3, plasmablast infiltration appeared in the islets of mice with 4w and 16w NOD, and the degree of insulitis of mice with 16w NOD was increased, and the plasmablast infiltration was significantly increased.
The results show that plasmablasts are involved in the islet autoimmune damage and the onset of type 1 diabetes in NOD mice, and the plasmablasts are positively correlated with the islet immune damage degree.
Example 4
Preparation of T lymphocytes and plasmablasts:
immune cells extracted from spleen and pancreatic lymph node of NOD mice 5 weeks old and 16 weeks old were collected by the method of example 1, and T lymphocytes and plasmablasts were aseptically sorted using BD FACAIra III flow cytometer.
Example 5
Cell co-culture:
the culture was carried out using 96-well round-bottomed cell culture plates in 3 groups of 20-30 islets/well, group 1 being the experimental group to which 2X 10 cells were added5T lymphocytes and 1X 105Individual plasmablasts, co-cultured with islets; group 2 addition of 2X 105Co-culturing individual T lymphocytes with islets of Langerhans; group 3 islet culture alone. The group 2 and the group 3 were control groups. IL-2 was added at 10ng/mL per well. Gently shaking the culture plate by drawing circle to make islet and immunocyte gather towards middle, directly contacting, 37 deg.C, 5% C02After 72h incubation in the incubator, the mixture was aspirated from the wells and washed in an EP tube containing 1mL PBS, and the islets settled to the bottom of the EP tube. The supernatant was carefully aspirated and discarded, washed again, and islet apoptosis was detected by TUNEL.
Fig. 4 shows that the TUNEL method detects islet apoptosis under different culture conditions, as shown in fig. 4, by establishing a co-culture system of mouse islets and plasmablasts or/and T lymphocytes in vitro and using the TUNEL kit to detect islet apoptosis, compared with the culture condition without plasmablasts, islet apoptosis is significantly increased when islets are co-cultured with plasmablasts and T lymphocytes, which indicates that plasmablasts and T lymphocytes respond synergistically to cause islet immune injury.
Example 6
Cell adoptive transfer:
t lymphocytes and plasmablasts from NOD mice at 16 weeks of age were transplanted at the following cell sizes. 6 week-old NOD/SCID mice were randomized into 3 groups of 6 mice each. Group 1 was experimental, each receiving 16 weeks old NOD mice 2X 106T lymphocytes and 2X 105-5×105Co-transfer of individual plasmablasts; group 2 each received the same number of T lymphocyte transplants from 20-week-old NOD mice, and group 3 was injected with an equal volume of PBS. The group 2 and the group 3 are control groups.
The transplanting method comprises the following steps: NOD/SCID mice were anesthetized with the inhaled isoflurane method. Fixing the head of the mouse, sucking the transplanted cell suspension by an insulin syringe, slightly obliquely inserting the mouse into the fundus venous plexus along the inner canthus of the mouse, injecting the mouse at a constant speed, and slowly pulling out the syringe after staying for several seconds. Random blood glucose was monitored in 3 groups of NOD/SCID mice by tail-snip after cell transplantation. NOD/SCID mice were sacrificed after disease onset, and were subjected to HE staining of the pancreas, microscopic examination, and insulitis scoring based on the degree of immune cell infiltration in the islets.
Insulitis evaluation criteria (total of individual islet scores/total number of observed islets): 0 minute, complete islets, no immune cell infiltration; 1 minute, only immune cells infiltrate around the islets, or the infiltration area of the immune cells in the islets is less than 25 percent; 2 minutes, the infiltration area of immune cells in the pancreatic islets reaches 25 to 50 percent; 3 minutes, the infiltration area of immune cells in the pancreatic islets reaches 50 to 75 percent; 4 minutes, the infiltration area of immune cells in the pancreatic islets exceeds 75 percent; at least 100 islets were observed per group of NOD/SCID mice for scoring.
In the cell adoptive transfer experiment, the speed and rate of diabetes development in 3 groups of recipient mice were compared by monitoring blood glucose. FIG. 5 shows the blood glucose changes of 16-week-old NOD/SCID mice of the present invention, as shown in FIG. 5, the NOD/SCID mice receiving plasmablasts and T lymphocytes together transferred initially became diabetic at 32 days after transplantation, while the NOD/SCID mice of 2 control groups receiving T lymphocytes or PBS injection alone still remained euglycemic at 80 days after adoptive transfer.
FIG. 6 shows the degree of insulitis of 16-week-old NOD/SCID mice in example of the present invention, and the degree of insulitis of the mice was evaluated by pancreatic HE staining, as shown in FIG. 6, the degree of insulitis of 3 groups of recipient mice showed significant difference, and most of the islets of mice in the group receiving co-transfer of plasmablasts and T lymphocytes were infiltrated with a large amount of immune cells, and the degree of insulitis was more severe than that of the remaining 2 groups. Indicating that plasmablasts are pathogenic cells causing the immune damage of pancreatic islets and the onset of type 1 diabetes.
Here, the experimental results of the step 5) cell co-culture experiment and the step 6) cell adoptive transfer experiment fully show the effect of islet immune injury caused by the synergistic response of plasmablasts and T lymphocytes, and provide possibility for early diagnosis of immune injury of type 1 diabetes.
The NOD mice (non-obese diabetic mice) are female, 4-20 weeks old, SPF-rated; NOD/SCID mice (severe combined immunodeficiency NOD mice) were SPF grade, 6 week old, female.
The method of the invention shows through animal model research that the plasmablast can be used for evaluating the pancreatic islet immune injury of type 1 diabetes, provides evaluation indexes for the pancreatic islet immune injury and the degree thereof, and provides theoretical basis for clinically using the plasmablast for preparing and detecting the reagent of the pancreatic islet immune injury and the medicine for treating the pancreatic islet immune injury.
The above description is only exemplary of the present invention and should not be taken as limiting the scope of the present invention, and any modifications, equivalents, improvements, etc. that are within the spirit and principle of the present invention should be included in the present invention.
Claims (10)
1. A method for assessing the correlation between plasmablasts and pancreatic islet immune damage, comprising the steps of:
1) isolating immune cells in pancreatic lymph node tissue and spleen of different stage type 1 diabetic patients or NOD model animals; preferably, the NOD model animals at different stages are NOD model animals raised to 4 weeks, 8 weeks, 12 weeks, 16 weeks, 20 weeks, 24 weeks; preferably, the NOD model animal is a NOD mouse;
2) analyzing immune cells, sorting plasmablasts, and constructing the relationship between the content of the plasmablasts and different stages of a type 1 diabetes patient or an NOD model animal, wherein the relationship is positive;
3) adoptive transfer of plasmablasts or a combination of plasmablasts and T lymphocytes, monitoring the rate and proportion of diabetes, and the condition of insulitis;
and/or co-culturing the T lymphocyte, the plasmablast and the pancreatic island to detect the apoptosis condition of the pancreatic island;
4) and establishing an insulitis evaluation standard, and scoring insulitis according to the infiltration degree of immune cells in the pancreatic islets.
2. The method of claim 1, wherein the insulitis scoring criteria: 0 minute, complete islets, no immune cell infiltration; 1 minute, only immune cells infiltrate around the islets, or the infiltration area of the immune cells in the islets is less than 25 percent; 2 minutes, the infiltration area of immune cells in the pancreatic islets reaches 25 to 50 percent; 3 minutes, the infiltration area of immune cells in the pancreatic islets reaches 50 to 75 percent; 4 min, the infiltration area of immune cells in the pancreatic islets exceeds 75 percent.
3. The method of claim 1, wherein step 1) comprises isolating pancreatic lymph nodes and spleen, grinding to obtain cell suspension, and centrifuging pancreatic lymph node cell suspension to obtain pancreatic lymph node immune cells; and adding erythrocyte lysate into the spleen cell sediment, and centrifuging after cracking to obtain spleen immune cells.
4. The method as claimed in claim 1, wherein the step 2) comprises transferring the immune cells of the step 1) to a flow tube, adding a flow antibody for pre-mixing incubation, adding a dead cell dye for incubation, washing and re-suspending by a Stain buffer, and then sorting the required T lymphocytes and plasmablasts by using a flow cytometer.
5. The method of claim 1, wherein step 3) comprises co-transferring T lymphocytes and plasmablasts to NOD/SCID mice, monitoring random blood glucose, obtaining pancreatic tissue, sectioning, HE staining and immunohistochemical staining, and collecting images.
6. The method according to claim 1, wherein the ratio of T lymphocytes to plasmablasts in step 3) is 1-10: 10-1, preferably 20: 2-5 or 2: 1.
7. The method according to claim 1, wherein the method for detecting islet apoptosis in step 3) is a TUNEL method.
8. The method of claim 5, wherein the co-transfer of T lymphocytes with plasmablasts is a bolus injection from the fundus venous plexus; the random blood sugar is monitored by a tail-cutting method for NOD/SCID mice.
9. Use of plasmablasts for the preparation of an islet immune-injury agent, optionally further comprising T lymphocytes.
10. The use of the plasmablast of claim 9 for preparing an islet immune-injury agent, wherein the ratio of T lymphocytes to plasmablasts is 1-10: 10-1, preferably 2:1 or 20: 2-5.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2004065181A (en) * | 2002-08-08 | 2004-03-04 | Japan Science & Technology Corp | Diabetes model mouse |
| US20180146649A1 (en) * | 2016-11-29 | 2018-05-31 | Taipei Medical University | Non-obese diabetes mice and its applications |
| CN112933211A (en) * | 2019-12-27 | 2021-06-11 | 艾棣维欣(苏州)生物制药有限公司 | Construction method of new animal model for type 1 diabetes |
-
2021
- 2021-12-07 CN CN202111483867.3A patent/CN114324860A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2004065181A (en) * | 2002-08-08 | 2004-03-04 | Japan Science & Technology Corp | Diabetes model mouse |
| US20180146649A1 (en) * | 2016-11-29 | 2018-05-31 | Taipei Medical University | Non-obese diabetes mice and its applications |
| CN112933211A (en) * | 2019-12-27 | 2021-06-11 | 艾棣维欣(苏州)生物制药有限公司 | Construction method of new animal model for type 1 diabetes |
Non-Patent Citations (3)
| Title |
|---|
| BI YAN等: "1673-P: Plasmablasts Contribute to the Development of Type 1 Diabetes via Enhancing T-Cell Cytotoxicity" * |
| SATORU AKAZAWA等: "Haploinsufficiency of interferon regulatory factor 4 strongly protects against autoimmune diabetes in NOD mice" * |
| 袁宝红 等: "猪骨髓间充质干细胞抑制脂多糖诱导炎症反应改善猪胰岛细胞功能损伤" * |
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