Published online Feb 27, 2022. doi: 10.4254/wjh.v14.i2.386
Peer-review started: July 31, 2021
First decision: September 29, 2021
Revised: October 12, 2021
Accepted: January 19, 2022
Article in press: January 19, 2022
Published online: February 27, 2022
The role of the hepatic nervous system in liver development remains unclear. We previously created functional human micro-hepatic tissue in mice by co-culturing human hepatic endodermal cells with endothelial and mesenchymal cells. However, they lacked Glisson’s sheath [the portal tract (PT)]. The PT consists of branches of the hepatic artery (HA), portal vein, and intrahepatic bile duct (IHBD), collectively called the portal triad, together with autonomic nerves.
To evaluate the development of the mouse hepatic nervous network in the PT using immunohistochemistry.
Liver samples from C57BL/6J mice were harvested at different developmental time periods, from embryonic day (E) 10.5 to postnatal day (P) 56. Thin sections of the surface cut through the hepatic hilus were examined using protein gene product 9.5 (PGP9.5) and cytokeratin 19 (CK19) antibodies, markers of nerve fibers (NFs), and biliary epithelial cells (BECs), respectively. The numbers of NFs and IHBDs were separately counted in a PT around the hepatic hilus (center) and the peripheral area (periphery) of the liver, comparing the average values between the center and the periphery at each developmental stage. NF-IHBD and NF-HA contacts in a PT were counted, and their relationship was quantified. SRY-related high mobility group-box gene 9 (SOX9), another BEC marker; hepatocyte nuclear factor 4α (HNF4α), a marker of hepatocytes; and Jagged-1, a Notch ligand, were also immunostained to observe the PT development.
HNF4α was expressed in the nucleus, and Jagged-1 was diffusely positive in the primitive liver at E10.5; however, the PGP9.5 and CK19 were negative in the fetal liver. SOX9-positive cells were scattered in the periportal area in the liver at E12.5. The Jagged-1 was mainly expressed in the periportal tissue, and the number of SOX9-positive cells increased at E16.5. SOX9-positive cells constructed the ductal plate and primitive IHBDs mainly at the center, and SOX-9-positive IHBDs partly acquired CK19 positivity at the same period. PGP9.5-positive bodies were first found at E16.5 and HAs were first found at P0 in the periportal tissue of the center. Therefore, primitive PT structures were first constructed at P0 in the center. Along with remodeling of the periportal tissue, the number of CK19-positive IHBDs and PGP9.5-positive NFs gradually increased, and PTs were also formed in the periphery until P5. The numbers of NFs and IHBDs were significantly higher in the center than in the periphery from E16.5 to P5. The numbers of NFs and IHBDs reached the adult level at P28, with decreased differences between the center and periphery. NFs associated more frequently with HAs than IHBDs in PTs at the early phase after birth, after which the number of NF-IHBD contacts gradually increased.
Mouse hepatic NFs first emerge at the center just before birth and extend toward the periphery. The interaction between NFs and IHBDs or HAs plays important roles in the morphogenesis of PT structure.
Core Tip: The portal tract (PT) consists of branches of the hepatic artery (HA), portal vein, intrahepatic bile ducts (IHBD), and autonomic nerves. This study evaluated the mouse hepatic nervous system development using immunohistochemistry. Hepatic nerve fibers (NFs) first emerge at the hepatic hilus just before birth and extend toward the periphery with IHBD in the PT after birth. The hepatic NFs associated more frequently with the HA than the IHBD in the PT after birth. The hepatic NFs may play important roles in the morphogenesis and stabilization of the PT during development of the liver.