Product Name 3D Human Pancreatic MicrovascularCatalog Number EP007Product Format 6 , 12, and 24 wellStorage 37° CGENERAL INFORMATIONHuman pancreas development starts between 26 and 35 days post conception with theemergence of dorsal and ventral buds from the foregut epithelium. At 6 weeks of gestation(equivalent to 4 weeks post conception) the two buds fuse and become a single organformed by stratified epithelium embedded in mesenchyme. The stratified epithelium willgive rise to both the exocrine and endocrine compartments of the definitive pancreas.One important physiological regulator of development and normal function of theendocrine cells of the pancreas is the microcirculation through specialized sinusoidalcapillaries that irrigate the islets of Langerhans. The endothelial cells of these capillariesare highly fenestrated to facilitate the exchange of signals. The dense network ensures thateach endocrine cell (glucagon-producing α-cell, insulin-producing β-cell, somatostatinproducing δ-cell, ghrelin-producing ϵ-cell and pancreatic polypeptide-producing PP-cell) isin close proximity to the circulation. It makes up a considerable part of the islets and it isresponsible for critical communication via blood signals between the endocrine andexocrine pancreas and also between the different cell types that populate the islets. Aftertransplantation of islets to the pancreas, angiogenesis is key to restoring proper functionOur 3D Human Pancreatic Microvascular Endothelial Angiogenesis model is constructusing GFP- Tagged human pancreatic microvascular endothelial cells are co-cultured withRFP- Tagged human Supporting cells. GFP positive lymphatic capillary like tubuleformation can be monitored in real time under fluorescence microscope throughout thewhole process of the experiment
Advantages:1) Cells used in the 3D model are all human cells; results obtained are more relevant tohuman situations rather than those data from animal models, i.e. CAM et al.2) The whole angiogenesis process can be monitored (from cell inoculation to the endof experiment), therefore, more crucial information can be acquired at multiple timepoints from a single experiment.3) No need to perform post-experimental staining for endothelial markers, this isparticularly important, if those markers are changed in experimental conditionsinvolved in the studies.The 3D Human Pancreatic Microvascular Endothelial Angiogenesis contains all of thematerials necessary to perform multiple angiogenesis assays in 6, 12, or 24 well formats.The 3D model is designed that the testing materials, i.e. compounds, conditioned media, ortissue explants, can be added into the system at any time, ranging from the onset ofvasculogenesis to advanced angiogenesis. The resulting effect on tubule formation (tubularlength, number of branches et al) can be monitored throughout the whole process underinverted fluorescence microscope.Reagents and Materials Provided:(1) 1 x vial of mixture of Human Pancreatic Microvascular Endothelial Angiogenesis ECsand RFP-tagged supporting cells (-80°C or liquid N2)(2) 1 x 24-well Alpha Coat Solution coated plate (Room temperature, for 2 months)(3) 1 x 500ml of Endo-Growth Medium (4°C)
Protocols: Day 11. Pre warm Endo-Growth Medium to 37ºC in a water bath2. Accurately pipette 24ml Endo-Growth Medium into a 50ml Falcon tube;3. Rapidly thaw the vial of cryopreserved cells in a 37ºC water bath;4. Transfer all cells gently into 24ml pre warmed Endo-growth medium;5. Mix well the cells gently using a serological pipette;6. Add 1.0ml of cell suspension to each well of the pre coated 24-well plate.7. Make sure the cells are evenly dispersed in the wells.8. Place the plate in an incubator (37ºC, 5% CO2 and humidified).Day 29. Take the plate from the incubator and examine cells under inverted fluorescencemicroscopy (GFP Human Pancreatic Microvascular Endothelial Angiogenesis shouldsparsely and evenly distributed among RFP positive human supporting cells).10. Wash the cells one with 2 ml of PBS11. Add 2.0ml of fresh Endo-Growth medium (control) or Experimental media (Endo-Growthmedium, plus pro- or anti-angiogenic regents according to customer’s needs).12. Place the plate back into the incubator. Day 4, 6, 8, 10, 12, and 14……13. Replace the medium every 2 days until the end of the experiments.