Lipopolysaccharide-induced tyrosine nitration and inactivation of hepatic glutamine synthetase in the rat

Abstract

Glutamine synthetase (GS) in the liver is restricted to a small perivenous hepatocyte population and plays an important role in the scavenging of ammonia that has escaped the periportal urea-synthesizing compartment. We examined the effect of a single intraperitoneal injection of lipopolysaccharide (LPS) in vivo on glutamine synthesis in rat liver. LPS injection induced expression of inducible nitric oxide synthase, which was maximal after 6 to 12 hours but returned toward control levels within 24 hours. Twenty-four hours after LPS injection, an approximately fivefold increase in tyrosine-nitrated proteins in liver was found, and GS protein expression was decreased by approximately 20%, whereas GS activity was lowered by 40% to 50%. GS was found to be tyrosine-nitrated in response to LPS, and immunodepletion of tyrosine-nitrated proteins decreased GS protein by approximately 50% but had no effect on GS activity. Together with the finding via mass spectrometry that peroxynitrite-induced inactivation of purified GS is associated with nitration of the active site tyrosine residue, our data suggest that tyrosine nitration critically contributes to inactivation of the enzyme. In line with GS inactivation, glutamine synthesis from ammonia (0.3 mmol/L) in perfused livers from 24-hour LPS-treated rats was decreased by approximately 50%, whereas urea synthesis was not significantly affected. A t the level of the liver acinus, the pathways of ammonia and glutamine metabolism are embedded in a sophisticated structural and functional organization, with considerable impact for the understanding of the liver in ammonia and pH homeostasis.

Hyperammonemia in gene-targeted mice lacking functional hepatic glutamine synthetase

“…Liver injury can result in a reduction of hepatic GS activity (21,24). Because other potent ammonia detoxicating mechanisms exist in the body, the role of the GS in perivenous liver cells remains controversial.…”

“…For example, LPS-induced liver injury results in PTN of hepatic GS, which inactivates the enzyme (21). The impact of the urea cycle for ammonia metabolism can be seen in children with urea cycle defects, who exhibit hyperammonemia and cognitive symptoms (22).…”

Lipopolysaccharide impairs hepatocyte ureagenesis from ammonia: Involvement of mitochondrial aquaporin‐8

“…Hyperammonemia is observed in experimental rodent models of sepsis, such as cecal ligation and puncture [6] or LPS administration [7] suggesting defective hepatic ammonia detoxification. In fact, LPS has been demonstrated to inhibit hepatic glutamine synthesis through the inactivation of the glutamine synthetase [8]. Nevertheless, studies in isolated perfused livers from septic [25] or LPS-treated [8] rats showed a slightly or not affected ammonia metabolism via ureagenesis.…”

“…In fact, LPS has been demonstrated to inhibit hepatic glutamine synthesis through the inactivation of the glutamine synthetase [8]. Nevertheless, studies in isolated perfused livers from septic [25] or LPS-treated [8] rats showed a slightly or not affected ammonia metabolism via ureagenesis.…”

“…The molecular mechanisms involved in the defective hepatic ammonia detoxification have not been yet fully elucidated. It has been demonstrated that bacterial lipopolysaccharides (LPS) induce inhibition of hepatic ammonia removal via glutamine synthesis [8]; nevertheless, an impairment of ammonia removal via urea synthesis may also be involved. Aquaporin-8 (AQP8) is a member of a family of homologous membrane channel proteins demonstrated to facilitate the diffusional transport of ammonia [9][10][11][12].…”