Molecular and biochemical alterations in tubular epithelial cells of patients with isolated methylmalonic aciduria.

Methylmalonic acidurias (MMAurias) are a group of inherited disorders in the catabolism of branched-chain amino acids, odd-chain fatty acids and cholesterol caused by complete or partial deficiency of methylmalonyl-CoA mutase (mut(0) and mut(-) subtype respectively) and by defects in the metabolism of its cofactor 5'-deoxyadenosylcobalamin (cblA, cblB or cblD variant 2 type). A long-term complication found in patients with mut(0) and cblB variant is chronic tubulointerstitial nephritis. The underlying pathomechanism has remained unknown. We established an in vitro model of tubular epithelial cells from patient urine (hTEC; 9 controls, 5 mut(0), 1 cblB). In all human tubular epithelial cell (hTEC) lines we found specific tubular markers (AQP1, UMOD, AQP2). Patient cells showed disturbance of energy metabolism in glycolysis, mitochondrial respiratory chain and Krebs cycle in concert with increased reactive oxygen species (ROS) formation. Electron micrographs indicated increased autophagosome production and endoplasmic reticulum stress, which was supported by positive acridine orange staining and elevated levels of LC3 II, P62 and pIRE1. Screening mTOR signaling revealed a release of inhibition of autophagy. Patient hTEC produced and secreted elevated amounts of the pro-inflammatory cytokine IL8, which was highly correlated with the acridine orange staining. Summarizing, hTEC of MMAuria patients are characterized by disturbed energy metabolism and ROS production that lead to increased autophagy and IL8 secretion.

Long-term exposure of human proximal tubule cells to hydroxycobalamin[c-lactam] as a possible model to study renal disease in methylmalonic acidurias.

Dysfunction of proximal tubules resulting in tubulointerstitial nephritis and chronic renal failure is a frequent long-term complication of methylmalonic acidurias. However, the underlying pathomechanisms have not yet been extensively studied owing to the lack of suitable in vitro and in vivo models. Application of hydroxycobalamin[c-lactam] has been shown to inhibit the metabolism of hydroxycobalamin and, thereby, to induce methylmalonic aciduria in rats, oligodendrocytes, and rat hepatocytes. Our study characterizes the biochemical and bioenergetic effects of long-term exposure of human proximal tubule cells to hydroxycobalamin[c-lactam], aiming to establish a novel in vitro model for the renal pathogenesis of methylmalonic acidurias. Incubation of human proximal tubule cells with hydroxycobalamin[c-lactam] and propionic acid resulted in a strong, time-dependent intra- and extracellular accumulation of methylmalonic acid. Bioenergetic studies of respiratory chain enzyme complexes revealed an increase of complex II-IV activity after 2 weeks and an increase of complex I and IV activity as well as a decrease of complex II and III activity after 3 weeks of incubation. In addition, human proximal tubule cells displayed reduced glutathione content after the exposure to hydroxycobalamin[c-lactam] and propionic acid.

Pathogenesis of CNS involvement in disorders of amino and organic acid metabolism.

Inherited disorders of amino and organic acid metabolism have a high cumulative frequency, and despite heterogeneous aetiology and varying clinical presentation, the manifestation of neurological disease is common. It has been demonstrated for some of these diseases that accumulating pathological metabolites are directly involved in the manifestation of neurological disease. Various pathomechanisms have been suggested in different in vitro and in vivo models including an impairment of brain energy metabolism, an imbalance of excitatory and inhibitory neurotransmission, altered transport across the blood-brain barrier and between glial cells and neurons, impairment of myelination and disturbed neuronal efflux of metabolic water. This review summarizes recent knowledge on pathomechanisms involved in phenylketonuria, glutaric aciduria type I, succinic semialdehyde dehydrogenase deficiency and aspartoacylase deficiency with examples, highlighting general as well as disease-specific concepts and their putative impact on treatment.

Neurodegeneration and chronic renal failure in methylmalonic aciduria--a pathophysiological approach.

In the last decades the survival of patients with methylmalonic aciduria has been improved. However, the overall outcome of affected patients remains disappointing. The disease course is often complicated by acute life-threatening metabolic crises, which can result in multiple organ failure or even death, resembling primary defects of mitochondrial energy metabolism. Biochemical abnormalities during metabolic derangement, such as metabolic acidosis, ketonaemia/ketonuria, lactic acidosis, hypoglycaemia and hyperammonaemia, suggest mitochondrial dysfunction. In addition, long-term complications such as chronic renal failure and neurological disease are frequently found. Neuropathophysiological studies have focused on various effects caused by accumulation of putatively toxic organic acids, the so-called 'toxic metabolite' hypothesis. In previous studies, methylmalonate (MMA) has been considered as the major neurotoxin in methylmalonic aciduria, whereas more recent studies have highlighted a synergistic inhibition of mitochondrial energy metabolism (pyruvate dehydrogenase complex, tricarboxylic acid cycle, respiratory chain, mitochondrial salvage pathway of deoxyribonucleoside triphosphate (dNTP)) induced by propionyl-CoA, 2-methylcitrate and MMA as the key pathomechanism of inherited disorders of propionate metabolism. Intracerebral accumulation of toxic metabolites ('trapping' hypothesis') is considered a biochemical risk factor for neurodegeneration. Secondary effects of mitochondrial dysfunction, such as oxidative stress and impaired mtDNA homeostasis, contribute to pathogenesis of these disorders. The underlying pathomechanisms of chronic renal insufficiency in methylmalonic acidurias are not yet understood. We hypothesize that renal and cerebral pathomechanisms share some similarities, such as an involvement of dicarboxylic acid transport. This review aims to give a comprehensive overview on recent pathomechanistic concepts for methylmalonic acidurias.

Fluid retention during the first 48 hours as an indicator of burn survival.

The quantity of fluid retained during the first 48 hours of resuscitation has been suggested as an indicator of burn severity and mortality (13). In this study of 82 adult burned patients with more than 20% total body surface burns we found that the net fluid retention during the first 48 hours of resuscitation was a predictor of burn mortality and additionally 230 cc of retained fluid per kilogram of lean body mass in the initial 48 hours postburn was an excellent means for separating survivors from nonsurvivors. Fluid retention as an indicator of burn severity and mortality was compared to other methods of predicting burned patient mortality. Parameters evaluated included the per cent body surface area burned, per cent full thickness burn, presence or absence of inhalation injuries, sex, age, and ultimate outcome. The Abbreviated Burn Severity Index (ABSI) (14) was determined for each patient using these data. A comparison was made between fluid retention data, per cent body surface area burned, and the calculated Abbreviated Burn Severity Index and patient mortality. The power of each variable to predict mortality was evaluated by stepwise regression analysis. From this analysis net fluid retention during the first 48 hours of resuscitation was as accurate as the Abbreviated Burn Severity Index and was a better predictor of mortality than individual components of that Index.

Nutritional indicators of postburn bacteremic sepsis.

The potential of nutritional assessment parameters in predicting sepsis in burn patients was investigated. Sixty-two consecutive patients (mean age 41 years) with an average burn size of 19% were studied. Values were obtained on postburn day 10 for serum albumin, transferrin, nitrogen balance, total lymphocyte count, skin test reactivity, and percentage of ideal body weight. Parameters predictive of imminent septic episode included serum albumin less than 3.0 g/dl (p less than 0.001), total lymphocyte count less than 1500/mm3 (p less than 0.001), anergy (p less than 0.001), and serum transferrin less than 150 mg/dl (p less than 0.001). Nitrogen balance and percentage of ideal body weight were not found to contribute to group discrimination.

Comparison of derived and actual transferrin: a potential source of error in clinical nutritional assessment.

A prospective study was undertaken to evaluate the utility of calculating transferrin from total iron-binding capacity in the nutritional assessment of burned patients. Regression analysis was used to compare total iron-binding capacity with radial immunodiffusion transferrin determinations. The method used for calculating transferrin (0.8 TIBC - 43) is a frequently published conversion formula for deriving transferrin. One hundred twenty-five data sets were obtained from 45 burned patients. Values for derived transferrin ranged from 39 to 235 mg/dl, averaging 121 mg/dl. Actual transferrin averaged 162 mg/dl, ranging from 41 to 320 mg/dl. Forty-eight actual serum transferrin samples were normal (greater than 172 mg/dl) whereas only 17 derived transferrin values were normal. While there is a correlation between total iron-binding capacity and serum transferrin (r = 0.85), to calculate transferrin according to the formula above would have resulted in significant error in the clinical assessment of the patients' nutritional status (p less than 0.001). From our studies, the formula for conversion of total iron-binding capacity to transferrin was found to be (0.68 TIBC + 21). These results suggest that the development of a universal conversion factor is not feasible. Modification of the formula may be necessary at each institution for clinically useful evaluations of serum transferrins are to be derived from iron-binding capacity for use in nutritional assessment.