Immunohistochemical and electron microscopic study of extrinsic hepatic reinnervation following orthotopic liver transplantation in rats.

BACKGROUND/AIMS: Because little has been known about the morphological and functional consequences of liver transplantation on hepatic autonomic nerves, we examined the time-course of extrinsic hepatic innervation at the level of the porta hepatis of liver allografts. METHODS: Orthotopic liver transplantation was performed using male Lewis rats. Crosscut tissue specimens were obtained postoperatively for up to 6 months from the porta hepatis of transplanted livers, and processed for immunohistochemical staining for protein gene product 9.5 (PGP 9.5) and growth-associated protein 43 (GAP-43), and for transmission electron microscopy (TEM). RESULTS: Extrinsic nerve fibers at the porta hepatis stained positively for PGP 9.5 throughout the entire study period. In contrast, the immunoreactivity of GAP-43 was negative at postoperative day (POD) 1 and 2. GAP-43-positive nerves were first observed to appear in the porta hepatis at POD 3. The immunoreactivity of GAP-43 remained positive thereafter until 3 months post-OLT, and became negative in all the specimens at 4 months post-OLT. Transmission electron microscopy demonstrated a small number of regenerating axons existing among many degenerating axons at POD 3. At 3 months post-OLT, most regenerating axons had been fully ensheathed by the cytoplasm of Schwann cells, although their density remained at a lower level compared with normal. CONCLUSION: The results of this study suggest that liver allografts become extrinsically reinnervated, with the regenerating axons reaching the hepatic hilus 3 days after transplantation. The process of extrinsic hepatic reinnervation is considered to almost terminate 4 months after transplantation in rats.

Regeneration of the hepatic nerves following surgical denervation of the liver in dogs.

This study was conducted to examine the regeneration process of hepatic nerves following surgical hepatic denervation in dogs. A denervation model was surgically created by removing all visible nerves around the hepatoduodenal ligament along with the peri-hepatic tissues. The hepatic nerves were examined on the hepatic specimens taken at 1 week, 1 month and 3 months post-denervation by means of immunohistochemical staining, and also electron microscopy. At 1 week post-denervation, the extrinsic hepatic nerves were observed not to have regenerated. However, at 1 month post-denervation, GAP-43-positive nerves were identified and regenerating axons were shown to be present on electron microscopic observation. The immunoreactivity for anti-GAP-43 antibody was not shown any longer at 3 months post-denervation, and the regenerated nerve axons were shown to be similar to those at pre-denervation on ultrastructural study. Results of the present study suggested that regeneration of the extrinsic hepatic nerves began to appear about 1 month after the hepatic denervation, and was completed by 3 months post-denervation.

Expressed sequence tags from cultured cells of rice (Oryza sativa L.) under stressed conditions: analysis of transcripts of genes engaged in ATP-generating pathways.

Large-scale sequencing of randomly selected cDNA clones was used to isolate numerous genes in rice (Oryza sativa L.). Total RNA used for cDNA synthesis was prepared from suspension-cultured cells of rice grown under stressed conditions, such as in saline or nitrogen-starvation conditions. A total of 780 cDNA clones were partially sequenced and about 15% could be identified as putative genes. In the library constructed under saline conditions, we identified several genes associated with signal transduction, such as protein kinase and small GTP-binding protein genes. Many stress-related genes were isolated from both the saline and nitrogen-starvation libraries. These results indicate that stress treatment of suspension-cultured cells makes it possible to efficiently isolate various types of plant genes. To examine the usefulness of such tagged cDNAs for the study of gene expression in a specific metabolic pathway, we analyzed mRNA levels of genes engaged in the ATP-generating pathways in cultured cells of rice under different stresses, such as 20% sucrose, salt stress, cold stress and nitrogen-starvation stress. The results suggest that the coordinated induction of several genes in key steps under stressed conditions may be essential for activation of the entire energy-producing pathway to maintain homeostasis in rice cells. Expressed sequence tags identified by random cDNA sequencing provide the opportunity to generate a transcript map of rice genes.

[Electron microscopic study on the degeneration and regeneration of the rat liver nerve fibers].

With advances in liver transplantation, attention has been directed to the regeneration of hepatic nerves. In this study, examination is made of ultrastructural degeneration and regeneration of hepatic nerves following severance at a point 8 mm proximal to the portal fissure in the rat liver. Immediately after the operation, distal nerve axons swelled and showed the disappearance of microtubules and neurofilaments. Newly formed axons, small in diameter and containing many microtubules and neurofilaments, could been seen at 2 days after the operation among those that had degenerated. The regenerated axons were surrounded completely by Schwann cells. The Schwann cell processes became shorter at days and regenerated axons were situated in the space between Schwann cell and the basement membrane. Degenerated axons could no longer be seen 14 days after the operation, and regenerating axons increasing in diameter filled the nerve fibers. At 21-30 days, Schwann cell processes gradually extended to enclose regenerating axons, nerve fiber regeneration was complete 42-56 days after the operation. The manner of nerve regeneration was consistent with the results of a previous immunohistochemical study using anti GAP-43 (growth associated protein-43) antibody.