CN-122005943-A - Vascularized organoid scaffold, islet organoid, and preparation method and application thereof

CN122005943ACN 122005943 ACN122005943 ACN 122005943ACN-122005943-A

Abstract

The invention discloses a vascularized organoid stent, an islet organoid, a preparation method and application thereof, wherein the preparation method of the vascularized organoid stent comprises the steps of dissolving a gel-forming phase material into a photoinitiator solution to obtain a gel-forming phase solution, and preserving the gel-forming phase solution for later use; the method comprises the steps of stirring and dissolving a sacrificial phase material in water to obtain a sacrificial phase solution, preserving heat for later use, enabling the sacrificial phase material to be incompatible with a gel-forming phase material, fully and uniformly mixing the gel-forming phase solution and the sacrificial phase solution according to a proportion to obtain a double-water phase mixed solution, preparing the double-water phase mixed solution into a preset shape, irradiating the double-water phase solution with ultraviolet light to obtain double-water phase hydrogel after the gel-forming phase is fully crosslinked, cleaning the double-water phase hydrogel, removing the sacrificial phase material which does not participate in crosslinking, and forming mutually communicated pore channels to obtain the vascularized organoid stent. The vascularized organoid stent prepared by the invention has communicated pore canal, adjustable porosity and aperture, and provides sufficient space structure for subsequent angiogenesis.

Inventors

WANG SHUFANG
WANG JIE
WANG YUKANG
YI XIN

Assignees

南开大学

Dates

Publication Date: 20260512
Application Date: 20251231
Priority Date: 20250624

Claims (6)

1. A method for preparing a vascularized islet organoid, comprising the steps of: step 1, taking a gel-forming phase material and a photoinitiator to be uniformly dispersed under a shading condition to obtain a gel-forming phase solution, re-suspending HUVECs and islet cells by the gel-forming phase solution, diluting a cell suspension, and preserving heat for later use; Step 2, taking a sacrificial phase material, stirring and dissolving the sacrificial phase material in water to obtain a sacrificial phase solution, re-suspending HUVECs by the sacrificial phase solution, diluting the HUVECs into cell suspension, and preserving the cell suspension for later use, wherein the sacrificial phase material and the gel-forming phase material are incompatible; Step 3, fully and uniformly mixing the cell suspensions obtained in the step 1 and the step 2 according to the volume to obtain a double-aqueous-phase cell suspension, and preparing the double-aqueous-phase cell suspension into a preset shape; And 4, preparing a double-aqueous-phase cell suspension with a preset shape by using ultraviolet light to irradiate the step 3, cleaning a non-crosslinked sacrificial phase solution after the gel-forming phase is completely crosslinked to obtain a double-aqueous-phase porous hydrogel containing cells, and culturing the double-aqueous-phase hydrogel containing cells at 25-40 ℃ to obtain the vascularized islet organoid.
2. The method for preparing a vascularized islet organoid according to claim 1, wherein the cell suspension of step 1 further comprises a vegf mimetic peptide, preferably in a ratio of 60 to 100 μg/1 mL of vegf mimetic peptide to gel-forming phase solution.
3. The method for preparing the vascularized islet organoid according to claim 1, wherein the gel-forming phase material in step 1 is GelMA or HAMA, the sacrificial phase material in step 2 is Dextran, PEG or PEO, and the volume ratio of gel-forming phase solution to sacrificial phase solution in step 3 is (2-4): 1; In the step 2, the sacrificial phase material is Dextran, PEG or PEO, the concentration of the sacrificial phase solution is 2% -20%, and when the sacrificial phase material is Dextran, the molecular weight of the Dextran is 150% -500 kDa.
4. The method of claim 1, wherein the gel-forming phase solution prepared in step 1 is added with decellularized matrix pECM of porcine pancreas to support islet cell growth.
5. The vascularized islet organoids prepared by the method of preparation of claim 1.
6. Use of the vascularized islet organoids of claim 5 in a drug screening model or in the preparation of a graft.

Description

Vascularized organoid scaffold, islet organoid, and preparation method and application thereof Technical Field The invention relates to the technical field of biological materials and biomedicine, in particular to a vascularized organoid stent, an islet organoid, a preparation method and application thereof. Background Diabetes is a hyperglycemic metabolic disease caused by a defect in insulin secretion or impaired biological action thereof, and islets are endocrine parts of the pancreas, mainly composed of an irregular cell population dispersed in the pancreas, and can regulate blood sugar by secreting insulin and glucagon, maintaining metabolic stability of the body. At present, a convenient and efficient method for controlling blood sugar and treating diabetes is not found, and the appearance of islet organoids provides new hopes for treating diabetes. The islet organs contain rich vascular network structures, play a vital role in conveying oxygen and nutrient substances to the islet organs, and in a signal path, the blood vessels can convey hormone from endocrine glands to target organs, sense signal stimulation and regulate physiological functions, and meanwhile, the islet organs are also a main way of drug delivery, have an irreplaceable effect in aspects of disease diagnosis and treatment and the like, and are vital in maintaining vital activities. Therefore, in the in vitro research of islet organoids, the construction of a vascular network has important significance, firstly, through vascular cooperative work, high-efficiency transmission and substance exchange of nutrient substances can be ensured, blood circulation and physiological function stability are maintained, islet cells can be cultured in vitro for a long time under the action of sufficient nutrient substances and oxygen, organoid tissue functions are improved, and secondly, vascularized organoids can be used for simulating islet-related diseases and are used as a model for drug screening, so that the development of accurate medical treatment is promoted. However, the prior art has not proposed a perfect and effective construction strategy of vascularized islet organoids, so how to realize the preparation of vascularized islet organoids in vitro is a technical problem to be solved in the research and treatment of diabetes. CN 116622618A discloses a vascularized islet organoid and its preparation method and application, firstly, the patent designs that the primary islet cells need to be subjected to pretreatment before encapsulating islet cells, the primary islet cells are co-cultured with stem cells and endothelial cells for 4-6 days to enable the cells to be aggregated into compact cell balls, the culture period is long, each islet cell mass needs to be separated, the operation is complex, secondly, single cell masses need to be collected before encapsulating, the loss is large, and finally, the survival time of the primary islet cells is limited when in vitro culturing, and the primary islet cells are difficult to maintain activity for a long time after being subjected to pretreatment which takes 4-6 days in the patent, and then the primary islet cells are encapsulated. The patent does not need to separate primary islets singly, and the culture medium number is large, so that the later-stage organoid transplantation is convenient. Disclosure of Invention The invention aims at providing a preparation method of a vascularized organoid stent aiming at the problem that an effective vascularized islet organoid construction strategy is not perfected in the prior art. It is another object of the present invention to provide a vascularized organoid scaffold. The invention also provides a vascularized islet organoid. The invention also provides a preparation method of the vascularized islet organoids. The invention also provides application of the vascularized islet organoid in drug screening or implant of islet related diseases. The technical scheme adopted for realizing the purpose of the invention is as follows: A method of preparing a vascularizable organoid scaffold comprising the steps of: step 1, taking a gel-forming phase material and a photoinitiator to be uniformly dispersed under a shading condition to obtain a gel-forming phase solution, and preserving heat at 25-40 ℃ for later use; step 2, taking a sacrificial phase material, stirring and dissolving the sacrificial phase material in water to obtain a sacrificial phase solution, and preserving the sacrificial phase solution at 25-40 ℃ for standby, wherein the sacrificial phase material is incompatible with the gel-forming phase material in the step 1; Step 3, fully and uniformly mixing the gel-forming phase solution prepared in the step 1 and the sacrificial phase solution prepared in the step 2 according to a proportion to obtain a double-aqueous-phase mixed solution, and preparing the double-aqueous-phase mixed solution into a preset shape; And 4, irradiating the aqueous two-phase