Regulation of Spermidine/Spermine N 1 -Acetyltransferase and its Involvement in Cellular Proliferation and Development of Acute Pancreatitis

The naturally occurring organic polycations, the polyamines spermidine, spermine and their precursor putrescine, are essential for cellular proliferation and differentiation. Their intracellular level is maintained by strictly regulated metabolic pathways. In order to investigate the functions and metabolism of polyamines, several genetically manipulated rodent lines have been generated. Activation of polyamine catabolism in transgenic rats overexpressing spermidine/spermine Nacetyltransferase (SSAT) under the control of heavy metal-inducible metallothionein I (MT) promoter results in a rapid depletion of spermidine and spermine and leads to the development of severe acute pancreatitis. Previous studies have shown that prophylactic administration of a stable spermidine analog, α-methylspermidine (MeSpd), can prevent the development of zinc-induced acute pancreatitis and restore the delayed liver regeneration after partial hepatectomy of MT-SSAT transgenic rats. In this work, the role of polyamines in other experimental models of pancreatitis, the pathogenesis of polyamine depletion-induced pancreatitis and the therapeutic potential of methylpolyamines were investigated. Activation of polyamine catabolism and depletion of higher polyamines were evident in both cerulein and L-arginine experimental models of pancreatitis, and also in two human pancreatic specimens obtained from patients with acute pancreatitis. Early pathogenesis in MTSSAT transgenic rats involved the activation of cathepsin B and trypsinogen, whereas prior administration of MeSpd inhibited the activation of both proteases. Importantly, the therapeutic administration of MeSpd or α,ω-bismethylspermine (Me2Spm) could dramatically protect MTSSAT transgenic rats from pancreatitis-associated mortality. The ability of stereoisomers of methylpolyamines to protect pancreatic integrity and support cellular growth was also tested. Although only (S,S)-Me2Spm was metabolized to MeSpd, both (R,R)and (S,S)-enantiomers were equally effective in preventing the development of pancreatitis and restoring liver regeneration in MT-SSAT transgenic rats. In vitro, all stereoisomers of both MeSpd and Me2Spm effectively rescued cells from acute cytostasis caused by inhibition of polyamine biosynthesis with α-difluoromethylornithine (DFMO). However, only (S)-MeSpd was able to support growth after prolonged exposure to DFMO. 2D-immunoblot analysis of eukaryotic translation initiation factor 5A (eIF5A) indicated that only (S)-MeSpd could serve as a precursor of hypusine, a unique aminoacid derivative essential for the synthesis of functional eIF5A. The physiological relevance of alternative splicing of SSAT pre-mRNA was also investigated. The alternative splice variant was targeted to a protein synthesis-dependent degradation pathway known as nonsense-mediated mRNA decay. Furthermore, the intracellular polyamine level regulated the balance of the two splice variants: polyamine supplementation favored the generation of the productive variant, subsequently resulting in decreased polyamine levels, whereas polyamine depletion favored the production of the alternative, unproductive variant. Thus, polyamine-regulated unproductive splicing and translation represents a novel posttranscriptional regulation mechanism of SSAT. In conclusion, these findings emphasize the importance of polyamine homeostasis for cellular proliferation and for the integrity and normal function of the liver and the exocrine pancreas, and open up possibilities for novel therapeutic approaches. National Library of Medicine classification: QT 120, QU 61, QU 450, QY 58, WI 702, WI 805 Medical Subject Headings: Acetyltransferases; Animals, Genetically Modified; Disease Models, Animal; Homeostasis; Liver/metabolism; Liver Regeneration; Pancreas/metabolism; Pancreatitis/therapy; Polyamines/metabolism; Putrescine; Rats; Spermidine; Spermidine/analogs & derivatives; Spermine; Spermine/analogs & derivatives; Trypsinogen If it happens, it must be possible. (Unnamed law)