Research On The In Vivo Construction Of Human Brain Organoids In Mice

Due to the complexity and ethics of the human brain,it is difficult to obtain a complete human brain for scientific research under laboratory conditions,which makes it a hot topic to simulate human brain development and verify related diseases in vitro.At present,the production of brain-like organs using human embryonic stem cells and induced pluripotent stem cells can faithfully reproduce the early stages of human embryonic and fetal brain development at the level of cell biology and gene expression.Combined with CRISPR and other gene-editing tools,it is possible to construct artificial models for studying various types of human brain diseases in vitro.However,there are still many problems in obtaining the complete structure and mature function of brain-like organs.When the cortical plates grow to a certain extent,nutrients cannot penetrate the brain-like organs in vitro culture.As the outer cells grow,the inner part of the brain-like body becomes necrotic.What should be more concerned is that,without the support of the vascular system,the degree of neuronal differentiation in brain-like tissue is significantly different from that in normal human brain development.In recent years,with the development of organ-like transplantation technology,many vascularization methods have emerged,but it is difficult to realize in vitro observation of vascularized organs in vivo.In this study,an abdominal imaging system was designed to observe the growth and development of organs in vivo,optimize the culture of organs in vivo and peritoneal vascularization scheme,and characterize the vascularized organs to a certain extent.In this study,a long-term abdominal imaging ring implanted into the abdomen of mice was prepared by using a medical titanium alloy Ti-6Al-4V with good biocompatibility,and the imaging window was bonded by cyanoacrylate.Long-time intraabdominal detection was achieved.The bodyweight of host C57BL/6 mice did not change significantly over more than 2 months.The PEG-treated laparoscope had a lower inflammatory response in mice and a lower mortality rate.The titanium alloy imaging ring has good mechanical strength.Real-time blood flow imaging using fluorescein-labeled dextran using a customized abdominal imaging box combined with a two-photon confocal microscope confirmed that the imaging window can be an effective means of verifying the status of blood vessels in mice.At the same time,sodium alginate(ALG)and hyaluronic acid(HA)were used to prepare a mixed gel.The hydrogel was mixed in a certain proportion to simulate the mechanical properties of human brain tissue.The 3D culture of neural stem cells was successfully realized.In the construction of organoid in vitro,the matrix gel mixed with hydrogel system supported organoid growth and neuronal differentiation.After transplantation in immunodeficient mice,the organs grew well.Astrocytes,early neurons,and other cells were found by section observation,and the existence of vascular structure was verified.The technique of organoid culture using peritoneal membrane was optimized.In this study,an abdominal imaging window constructed by titanium alloy with excellent biocompatibility was used to realize the imaging of abdominal blood flow,and a vascularized organoid was constructed by using the peritoneum.It can be used to culture and characterize the organs in vivo and in vitro and is expected to monitor the growth status of the organs.