This study demonstrates the creation of a dual humanized mouse model by injecting human liver cells and hematopoietic stem progenitor cells into an immunodeficient mouse. The methodology involves surgical techniques to facilitate the integration of human cells into the mouse's biological systems.
Begin with an anesthetized, immunodeficient mouse that lacks various immune cells.
This transgenic mouse is pretreated with an antiviral and a chemotherapeutic agent. The antiviral agent exclusively interacts with viral proteins present in transgenic liver cells, promoting their elimination.
Meanwhile, the chemotherapeutic agent eliminates the actively dividing stem cells in the bone marrow, creating a niche for immune cell generation.
Dissect the mouse and expose the spleen.
Take the cell suspension of human liver cells and hematopoietic stem progenitor cells or HSPCs and inject them into the lower pole of the spleen.
Ligate the spleen above the injection site to restrict cells to the lower pole and facilitate their entry into the mouse circulation.
Close the incision.
The injected liver cells reach the liver tissue and establish human liver cells in the mouse model.
In the bone marrow, migrated HSPCs differentiate into various human immune cells.
This develops a dual humanized mouse model with human liver cells and immune cells.
First, attach one end of a sterile extension tube to a 30-gauge needle, and the other end to a 1-milliliter syringe. Fill the syringe with the suspension of pooled HEPs and HSPCs. Fit the syringe in the notch of a repetitive dispensing pipette, and adjust the dispenser to dispense 10 microliters in each press. Shave each mouse as outlined in the text protocol, and scrub the left side of the body of each mouse with 10% povidone-iodine, followed by 70% isopropyl alcohol.
Using Vannas-type scissors, make a small incision in the skin, muscle, and peritoneum at the left of the peritoneal wall to enter the peritoneal cavity approximately 5 millimeters below the lower edge of the rib cage. Locate the spleen and use forceps to pull it slightly to the operating area for easy access, and insert the 30-gauge needle into the lower pole of the spleen.
Next, unlock the plunger of the dispensing pipette, and dispense 10 microliters of the volume at a time, with a limit of 60 to 80 microliters per spleen. Retract the needle slowly, and clip the spleen with ligating clips using a ligation applier. Then, use cotton-tipped applicators, wetted with sterile PBS, to push the spleen back into the body cavity. Use an interrupted suture pattern to close the muscle layer of the abdominal wall. Accomplish skin closure with an interrupted suture pattern using non-absorbable sutures.
繁體中文
