Amino Acid Turnover, Protein Metabolism, and Nitrogen Balance in Acute Kidney Injury

Nitrogen, Amino Acids, and Protein Balance

Nitrogen is the fundamental component of amino acids (AAs), and AAs form the molecular structure of proteins. Proteins are polymers of AAs linked by peptide bonds. They are major functional substrates in cells and tissues and are essential for body growth, maintenance, and recovery. Protein metabolism also generates calories, about 4 kcal/g, similar to carbohydrates. Protein degradation by enzymatic reactions releases nitrogen. Nitrogen is lost in body secretions and excreted in sweat, feces, and urine, the latter most notably as urea nitrogen that accounts for 85% to 90% of urinary nitrogen loss. The remaining urinary nitrogen is lost as creatinine and ammonia that facilitates hydrogen ion excretion in renal tubular acid-base handling, and as a trace protein.

In a healthy adult human with metabolic equilibrium, stable protein intake and synthesis should be balanced by protein degradation and loss to maintain tissue integrity. This should allow stable body muscle mass and anthropometry . In simplistic terms, nitrogen balance, which infers the difference of nitrogen intake and nitrogen loss, can be measured in nutritional intake and excretory products to reflect overall protein balance, given physiologic understanding that most proteins and AAs are 15.6% nitrogen by weight. These sound ideal in theory but are subjected to various assumptions and inaccuracies in quantification (especially with insensible body losses and gut bacterial turnover, etc.). See Table 74.1 for further illustration of this concept.

Different types of AAs may exert varying significance. Tissue protein is synthesized from an intracellular pool of 21 AAs, of which 9 are termed essential (EAAs), because these are from dietary sources and not synthesized de novo. Estimated daily requirements of individual EAAs in steady state are inferred from respective nitrogen balance or isotope studies. The other nonessential AAs (NEAAs) are interconvertible and synthesized in the body or from other dietary AAs ( Table 74.2 ). The reader may refer to more detailed biochemistry reviews on protein metabolism.

Protein and Muscle Turnover

Increase in muscle activity stimulates the expression of insulin-like growth factor-1 (IGF-1), which is expressed ubiquitously in muscles and mitosis-competent cells. IGF-1 induces downstream autocrine and paracrine activities, including phosphoinositide 3-kinase (PI3K), downstream Akt (protein kinase B), and mTOR (mammalian target of rapamycin) phosphorylation-activations, to promote protein synthesis and muscle hypertrophy. Protein synthesis is an energy-dependent process, with three phases, controlled in part by three groups of proteins: initiation (controlled by eukaryotic initiation factors, or EIFs), elongation (eukaryotic elongation factors, EEFs), and termination (eukaryotic release factors, ERFs). After termination, proteins undergo tertiary and quaternary structure development and folding.

The reverse happens when Akt is downregulated via dephosphorylation, PTEN (phosphatase and tensin homolog), and SHIP (SH2-domain-containing inositol 5′-phosphatase). Breakdown is mediated through the ubiquitin-proteasome pathway, where proteins are ubiquitinated/conjugated and undergo proteasome-led recognition and degradation by proteasome. Forkhead box group O-1 (FOXO1) translocation into the nucleus increases transcription of ubiquitin ligases (the most common is muscle ring finger protein-1 [MURF-1] and Atrogin-1 or Muscle Atrogen F-box protein 1 [MAFBx], and other newly discovered ones. These ligases bind ubiquitin to proteins, forming an ubiquitin chain, targeting said proteins for proteolysis and AA release by the 20S proteasome. The ubiquitin-proteasome pathway is adenosine triphosphate (ATP) dependent, and in the setting of unstable patients other pathways may be activated such as the autophagic-lysosomal pathway. Myostatin, a member of transforming growth factor-β (TGF-β) superfamily, also works on downstream deregulation of Akt-mTOR pathways, as does nuclear factor-κβ, which may modulate the tumor necrosis superfamily receptor activation. Fig. 74.1 illustrates these interactions in maintaining protein metabolism and facilitating muscle turnover.

Protein Energy Wasting and Muscle Wasting in Critical Illness and Acute Kidney Injury

The inflammatory milieu during critical illness results in decreased protein synthesis, shifting the balance toward net protein catabolism. This is common across a range of factors associated with critical illness, including sepsis, trauma, and burns, and compounded by various comorbidities often present in an aging population. Immobilization further contributes to the impaired stimulus for muscle protein synthesis. Acquired insulin resistance secondary to impaired GLUT-4 membrane translocation contributes to hyperglycemia and inhibits protein synthesis. Protein catabolism is driven by systemic inflammatory response syndrome, oxidative stress, and catabolic hormones, including catecholamines and glucocorticoids. Muscle protein degradation occurs along with increased hepatic gluconeogenesis and ureagenesis from AAs, as promoted by cortisol and glucagon, respectively, with stimulated hepatic diversion to production of acute phase reactants. The markedly reduced systemic concentrations of most AAs seen in sepsis may suggest enhanced hepatic extraction to fuel this process.

Such catabolic effect of critical illness and multiorgan dysfunction probably overwhelms that from AKI alone. AKI and tubular injury augment the inflammatory stress. Proximal renal tubular epithelial cells secrete inflammatory cytokines such as tumor necrosis factor alpha (TNF-α) and interleukins, which potentially are upregulated in AKI. Metabolic acidosis (from acute illness and/or AKI) and hyperparathyroidism (with more prolonged AKI) worsen the catabolism, as inferred from studies in chronic kidney disease (CKD). Anorexia and reduced nutrient intake before and during early hospitalization are worsened because of volume intake restriction in oligoanuric AKI and gastrointestinal intolerance during acute phase of critical illness. In patients who receive RRT, early generation regenerated cellulose (cuprophan) membranes are less biocompatible than modern synthetic membranes (e.g., polyamide, polysulfone, polyacrylonitrile), and the former invokes more complement activation with blood-membrane contact, which may aggravate protein catabolism. The term protein energy wasting (PEW), introduced by the International Society of Renal Nutrition and Metabolism in 2008 to describe this phenomenon in CKD, but similarly applicable in AKI, aptly describes the above condition ( Fig. 74.2 ).

Protein catabolism in critical illness with or without AKI translates to early and severe skeletal muscle wasting in patients. This was demonstrated elegantly in an observational study, which reported pronounced reduction in muscle bulk by ultrasonographic measurements, with consistent histologic features in muscle fibers and depressed protein to DNA ratio, over the first week of ICU care. Muscle protein synthesis was depressed to levels seen in healthy fasting state initially, rising to rates comparable with healthy fed state by 1 week, but yet remained in net catabolism . The degree of muscle wasting was significantly worse in patients with multiorgan failure (vs. single organ). Older age, metabolic acidosis, and lower ratio of PaO ₂ to FiO ₂ were associated with greater loss of muscle mass. Interestingly, no clear single molecular mechanism based on physiologic understanding, correlated with muscle loss and proteolysis, suggesting incomplete understanding of molecular signaling in muscle protein turnover in critical illness.

Amino Acid and Nitrogen Balance in Acute Kidney Injury and Renal Replacement Therapy