Impact of mutations within the [Fe-S] cluster or the lipoic acid biosynthesis pathways on mitochondrial protein expression profiles in fibroblasts from patients.

Lipoic acid (LA) is the cofactor of the E2 subunit of mitochondrial ketoacid dehydrogenases and plays a major role in oxidative decarboxylation. De novo LA biosynthesis is dependent on LIAS activity together with LIPT1 and LIPT2. LIAS is an iron­sulfur (Fe-S) cluster-containing mitochondrial protein, like mitochondrial aconitase (mt-aco) and some subunits of respiratory chain (RC) complexes I, II and III. All of them harbor at least one [Fe-S] cluster and their activity is dependent on the mitochondrial [Fe-S] cluster (ISC) assembly machinery. Disorders in the ISC machinery affect numerous Fe-S proteins and lead to a heterogeneous group of diseases with a wide variety of clinical symptoms and combined enzymatic defects. Here, we present the biochemical profiles of several key mitochondrial [Fe-S]-containing proteins in fibroblasts from 13 patients carrying mutations in genes encoding proteins involved in either the lipoic acid (LIPT1 and LIPT2) or mitochondrial ISC biogenesis (FDX1L, ISCA2, IBA57, NFU1, BOLA3) pathway. Ten of them are new patients described for the first time. We confirm that the fibroblast is a good cellular model to study these deficiencies, except for patients presenting mutations in FDX1L and a muscular clinical phenotype. We find that oxidative phosphorylation can be affected by LA defects in LIPT1 and LIPT2 patients due to excessive oxidative stress or to another mechanism connecting LA and respiratory chain activity. We confirm that NFU1, BOLA3, ISCA2 and IBA57 operate in the maturation of [4Fe-4S] clusters and not in [2Fe-2S] protein maturation. Our work suggests a functional difference between IBA57 and other proteins involved in maturation of [Fe-S] proteins. IBA57 seems to require BOLA3, NFU1 and ISCA2 for its stability and NFU1 requires BOLA3. Finally, our study establishes different biochemical profiles for patients according to their mutated protein.

Early and late complications after liver transplantation for propionic acidemia in children: a two centers study.

Propionic acidemia (PA) is a severe metabolic disorder with cardiac and neurologic complications and a poor quality of life. Liver transplantation (LT) was thus proposed in PA to increase enzyme activity. We studied retrospectively LT in PA in two European centers. Twelve patients underwent 17 LTs between 1991 and 2013. They developed severe, unusual and unexpected complications, with high mortality (58%). When present, the cardiomyopathy resolved and no acute metabolic decompensation occurred allowing dietary relaxation. Renal failure was present in half of the patients before LT and worsened in all of them. We suggest that cardiac and renal functions should be assessed before LT and monitored closely afterward. A renal sparing immunosuppression should be used. We speculate that some complications may be related to accumulated toxicity of the disease and that earlier LT could prevent some of these consequences. As kidney transplantation has been performed successfully in methylmalonic acidemia, a metabolic disease in the same biochemical pathway, the choice of the organ to transplant could be further discussed.

Renal transplantation in 4 patients with methylmalonic aciduria: a cell therapy for metabolic disease.

INTRODUCTION: Patients with methylmalonic acidemia (MMA) may develop many complications despite medical treatment, in particular, severe central nervous system damage and chronic kidney disease (CKD). A kidney transplant may partially correct the metabolic dysfunctions. Liver, kidney and combined liver-kidney transplantations have been advocated but no guidelines are available to identify the most suitable organ to transplant. PATIENTS AND METHODS: Four patients with MMA (mut° phenotype) received a kidney graft because of repeated metabolic decompensations, with progression to CKD in 3 patients (end-stage kidney disease in two patients and CKD stage III in one patient with an estimated glomerular filtration rate [eGFR] of 40ml/min/1.73m(2)) but normal renal function in one (eGFR of 93ml/min/1.73m(2)) before transplantation. RESULTS: The medium age at transplantation was 7.9y (5-10.2) and the median follow-up was 2.8years (1.8-4.6). Renal transplantation improved the relevant metabolic parameters in 4/4 patients and renal function in the patients with CKD. Plasma and urinary MMA levels immediately decreased and remained normal or subnormal (mean values of plasma MMA before transplantation 1530µmol/L versus 240µmol/L after transplantation, and mean values of urine MMA before transplantation 4700mmol/mol creatinine versus 2300mmol/mol creatinine after transplantation). No further acute metabolic decompensation was observed and protein-intake was increased from 0.60 to 0.83g/Kg/day. One patient transplanted at age 9.7years developed a hepatoblastoma at age 11years with subsequent neurological complications and eventually died. The three other patients are alive. Two of them remained neurologically stable. The 3rd patient who displayed choreoathetosis transiently improved his neurological condition immediately after transplantation and then remained stable. CONCLUSION: Kidney transplantation represents an interesting alternative therapeutic option in methylmalonic aciduria, for renal complications but also as a "cellular therapy" that may significantly reduce metabolic decompensations and hospitalizations. However, further neurological impairment remains possible.

Treatment of acute decompensation of maple syrup urine disease in adult patients with a new parenteral amino-acid mixture.

BACKGROUND: Acute decompensation of maple syrup urine disease (MSUD) is usually treated by enteral feeding with an amino-acid mixture without leucine (Leu), valine or isoleucine. However, its administration is ineffective in cases of gastric intolerance and some adult patients refuse enteral feeding via a nasogastric tube. We developed a new parenteral amino-acid mixture for patients with MSUD. METHODS: Seventeen decompensation episodes in four adult patients with MSUD treated with a parenteral amino-acid mixture (group P) were compared to 18 previous episodes in the same patients treated by enteral feeding (group E). RESULTS: The mean Leu concentration at presentation was similar in the groups P and E (1196.9 µmol/L and 1212.2 µmol/L, respectively). The mean decrease in the Leu concentration during the first 3 days of hospitalisation was significantly higher in group P than group E (p = 0.0026); there were no side effects. The mean duration of hospitalisation was similar (4 vs. 4.5 days, p = NS). No patient in group P deteriorated whereas one patient in group E required dialysis. CONCLUSION: This new parenteral amino-acid mixture is safe and allows efficient Leu concentration decrease during acute MSUD decompensation episodes in adults. Its use avoids the need for nasogastric tube insertion.

Gestational age-related reference values for amniotic fluid organic acids.

BACKGROUND: Normative data for amniotic fluid (AF) levels of organic acids at different gestational ages are lacking. They can provide a useful framework to investigate the accuracy of prenatal diagnosis for organic acidemias. METHODS: We report on the concentration of 21 organic acids in AF obtained by gas chromatography/mass spectrometry between the 12th and 34th weeks of gestation from 92 pregnancies that were not at risk for organic acidurias. RESULTS: We infer normal reference values that can be compared with 134 pregnancies at risk for several metabolic conditions, that is, propionic acidemia, methylmalonic acidemia (methylmalonyl-CoA mutase deficiency or defects in cobalamin metabolism), 4-hydroxybutyric acidemia, glutaric acidemia and pyroglutamic acidemia. CONCLUSION: Most of the metabolites tested did not show conspicuous variations across gestational ages in normal fetuses, with ranges that were consistently similar to available reference values from pooled samples in previous reports. With rare exceptions, knowledge of pathological versus normal values for relevant metabolites leads to clear-cut differentiation of affected versus unaffected fetuses. Nevertheless, it is strongly recommended that mutational analysis and/or additional biochemical approaches complement organic acid analysis for an adequate diagnostic workup.

Management of 35 critically ill hyperammonemic neonates: Role of early administration of metabolite scavengers and continuous hemodialysis.

Neurological involvement is frequent in inherited metabolic disease of the intoxication type. Hyperammonemic coma related to these diseases may cause severe neurological sequelae. Early optimal treatment is mandatory combining metabolite scavengers (MS) and sometimes continuous veno-venous hemodialysis (CVVHD). We aimed to describe the therapeutic management of hyperammonemia in neonates upon diagnosis of their metabolic disease and to compare neonates managed with MS alone or with both MS and CVVHD. We conducted a retrospective study including all neonates admitted for initial hyperammonemia to the pediatric intensive care unit of a Reference Center of Inherited Metabolic Diseases, between 2001 and 2012. The study included 35 neonates. Before admission, MS were initiated for 11 neonates. At admission, the median ammonia levels were 391 µmol/L and were significantly lower in neonates who received MS before admission. At admission, ammonia levels were 644 µmol/L in dialyzed and 283 µmol/L in non-dialyzed neonates. The median time to reach a 50% decrease of the initial ammonia levels was significantly shorter in dialyzed neonates; however, the normalization of ammonia levels was similar between dialyzed and non-dialyzed neonates. Hemodynamic disorders were more frequent in dialyzed neonates. CONCLUSION: MS represent an effective treatment for hyperammonemia and should be available in all pediatric units to avoid the need for CVVHD. Although CVVHD enhances the kinetics of toxic metabolite decrease, it is associated with adverse hemodynamic effects.

Ketone Bodies as a Possible Adjuvant to Ketogenic Diet in PDHc Deficiency but Not in GLUT1 Deficiency.

OBJECTIVE: Ketogenic diet is the first line therapy for neurological symptoms associated with pyruvate dehydrogenase deficiency (PDHD) and intractable seizures in a number of disorders, including GLUT1 deficiency syndrome (GLUT1-DS). Because high-fat diet raises serious compliance issues, we investigated if oral L,D-3-hydroxybutyrate administration could be as effective as ketogenic diet in PDHD and GLUT1-DS. METHODS: We designed a partial or total progressive substitution of KD with L,D-3-hydroxybutyrate in three GLUT1-DS and two PDHD patients. RESULTS: In GLUT1-DS patients, we observed clinical deterioration including increased frequency of seizures and myoclonus. In parallel, ketone bodies in CSF decreased after introducing 3-hydroxybutyrate. By contrast, two patients with PDHD showed clinical improvement as dystonic crises and fatigability decreased under basal metabolic conditions. In one of the two PDHD children, 3-hydroxybutyrate has largely replaced the ketogenic diet, with the latter that is mostly resumed only during febrile illness. Positive direct effects on energy metabolism in PDHD patients were suggested by negative correlation between ketonemia and lactatemia (r 2 = 0.59). Moreover, in cultured PDHc-deficient fibroblasts, the increase of CO2 production after 14C-labeled 3-hydroxybutyrate supplementation was consistent with improved Krebs cycle activity. However, except in one patient, ketonemia tended to be lower with 3-hydroxybutyrate administration compared to ketogenic diet. CONCLUSION: 3-hydroxybutyrate may be an adjuvant treatment to ketogenic diet in PDHD but not in GLUT1-DS under basal metabolic conditions. Nevertheless, ketogenic diet is still necessary in PDHD patients during febrile illness.

Two new cases of serine deficiency disorders treated with l-serine.

OBJECTIVE AND PATIENTS: We report on two new cases of serine deficiency due respectively to 3-phosphoglycerate dehydrogenase (PHGDH) deficiency (Patient 1) and phosphoserine aminotransferase (PSAT1) deficiency (Patient 2), presenting with congenital microcephaly (<3rd centile at birth) and encephalopathy with spasticity. Patient 1 had also intractable seizures. A treatment with oral l-serine was started at age 4.5 years and 3 months respectively. RESULTS: Serine levels were low in plasma and CSF relative to the reference population, for which we confirm recently redefined intervals based on a larger number of samples. l-Serine treatment led in patient 1 to a significant reduction of seizures after one week of treatment and decrease of electroencephalographic abnormalities within one year. In patient 2 treatment with l-serine led to an improvement of spasticity. However for both patients, l-serine failed to improve substantially head circumference (HC) and neurocognitive development. In a couple related to patient's 2 family, dosage of serine was performed on fetal cord blood when the fetus presented severe microcephaly, showing reduced serine levels at 30 weeks of pregnancy. CONCLUSIONS: l-Serine treatment in patients with 2 different serine synthesis defects, led to a significant reduction of seizures and an improvement of spasticity, but failed to improve substantially neurocognitive impairment. Therefore, CSF and plasma serine levels should be measured in all cases of severe microcephaly at birth to screen for serine deficiency, as prompt treatment with l-serine may significantly impact the outcome of the disease. Reduced serine levels in fetal cord blood may also be diagnostic as early as 30 weeks of pregnancy.

Pyruvate carboxylase deficiency: An underestimated cause of lactic acidosis.

Pyruvate carboxylase (PC) is a biotin-containing mitochondrial enzyme that catalyzes the conversion of pyruvate to oxaloacetate, thereby being involved in gluconeogenesis and in energy production through replenishment of the tricarboxylic acid (TCA) cycle with oxaloacetate. PC deficiency is a very rare metabolic disorder. We report on a new patient affected by the moderate form (the American type A). Diagnosis was nearly fortuitous, resulting from the revision of an initial diagnosis of mitochondrial complex IV (C IV) defect. The patient presented with severe lactic acidosis and pronounced ketonuria, associated with lethargy at age 23 months. Intellectual disability was noted at this time. Amino acids in plasma and organic acids in urine did not show patterns of interest for the diagnostic work-up. In skin fibroblasts PC showed no detectable activity whereas biotinidase activity was normal. We had previously reported another patient with the severe form of PC deficiency and we show that she also had secondary C IV deficiency in fibroblasts. Different anaplerotic treatments in vivo and in vitro were tested using fibroblasts of both patients with 2 different types of PC deficiency, type A (patient 1) and type B (patient 2). Neither clinical nor biological effects in vivo and in vitro were observed using citrate, aspartate, oxoglutarate and bezafibrate. In conclusion, this case report suggests that the moderate form of PC deficiency may be underdiagnosed and illustrates the challenges raised by energetic disorders in terms of diagnostic work-up and therapeutical strategy even in a moderate form.

Absence of mutations involving the LIM homeobox domain gene LHX9 in 46,XY gonadal agenesis and dysgenesis.

The etiology of most cases of 46,XY gonadal dysgenesis in the absence of extragenital anomalies is not accounted for by mutations in the genes known to date to be involved in sex determination. We have investigated the possibility that mutations in the gene LHX9, whose murine ortholog causes isolated gonadal agenesis when inactivated, might be responsible for gonadal dysgenesis and agenesis in humans. We isolated a human LHX9 complementary DNA (cDNA), mapped the gene to the long arm of human chromosome 1, and determined its genomic structure. We found that LHX9 is highly conserved between species, sharing in particular over 98% amino acid identity. A mutational screen was performed in a sample of patients with a range of gonadal maldevelopment, including bilateral gonadal agenesis in two sisters with an opposite sex karyotype. We did not detect mutations in the open reading frame of LHX9 in the patients studied. However, the extent of between-species structural conservation suggests that LHX9 deserves further consideration as a determinant of gonadal function in humans.

FISH mapping of the sex-reversal region on human chromosome 9p in two XY females and in primates.

Accumulating evidence suggests that haploinsufficiency of a dosage-sensitive gene(s) in human chromosome 9p24.3 is responsible for the failure of testicular development and feminisation in XY patients with monosomy for 9p. We have used molecular cytogenetic methods to characterise the sex-reversing 9p deletions in two XY females. Fluorescence in situ hybridisation (FISH) with YACs from the critical 9p region containing an evolutionarily conserved sex-determining gene, DMRT1, is a very fast and reliable assay for patient screening. Comparative YAC mapping on great ape and Old and New World monkey chromosomes demonstrated that the critical region was moved from an interstitial position on the ancestral primate chromosome to a very subtelomeric position in chimpanzee and humans by a pericentric inversion(s). Pathological 9p rearrangements may be the consequence of an evolutionary chromosome breakpoint in close proximity to the sex-reversal region.

Adenylate cyclase of catfish hepatocyte membranes: basal properties and sensitivity to catecholamines and glucagon.

Some characteristics of adenylate cyclase of catfish (Ictalurus melas) liver membranes were studied, and the effects of catecholamines and of glucagon were tested. The enzyme has an optimum temperature of 40 degrees C, and a Km for ATP of 0.16 mM at 30 degrees C, and requires Mg2+ for its activity. The enzyme activity is inhibited with a Ca2+ concentration higher than 5 X 10(-5) M, and enhanced with F- higher than 10(-4) M. The response of adenylate cyclase to GTP is biphasic, with a maximum of activity at 10(-5) M GTP. Catecholamines (epinephrine, norepinephrine, isoproterenol, phenylephrine) enhance cyclase activity. Propranolol inhibits the increase in enzyme activity induced by catecholamines, whereas phentolamine is ineffective. This indicates that catecholamines (phenylephrine included) activate adenylate cyclase through a beta-adrenergic mechanism. Glucagon (mammalian) has a smaller effect than epinephrine in increasing the enzyme activity of catfish hepatocyte membranes. This fact is the opposite of that observed for the cyclase activity of rat liver membranes.

Testis determination in mammals: more questions than answers.

In humans, testis development depends on a regulated genetic hierarchy initiated by the Y-linked SRY gene. Failure of testicular determination results in the condition termed 46,XY gonadal dysgenesis (GD). Several components of the testis determining pathway have recently been identified though it has been difficult to articulate a cascade with the known elements of the system. It seems, however, that early gonadal development is the result of a network of interactions instead of the outcome of a linear cascade. Accumulating evidence shows that testis formation in man is sensitive to gene dosage. Haploinsufficiency of SF1, WT1 and SOX9 is responsible for 46,XY gonadal dysgenesis. Besides, data on SRY is consistent with possible dosage anomalies in certain cases of male to female sex reversal. 46,XY GD due to monosomy of distal 9p and 10q might also be associated with an insufficient gene dosage effect. Duplications of the locus DSS can lead to a failure of testicular development and a duplication of the region containing SOX9 has been implicated in XX sex reversal. Transgenic studies in mouse have shown, however, that this mammal is less sensitive to gene dosage than man. Here, we will try to put in place the known pieces of the jigsaw puzzle that is sex determination in mammals, as far as current knowledge obtained from man and animal models allows. We are certain that from this attempt more questions than answers will arise.

Bile salt-induced cytotoxicity and ursodeoxycholate cytoprotection: in-vitro study in perifused rat hepatocytes.

OBJECTIVES: Membrane toxicity induced by hydrophobic bile salts may be important in liver diseases. Administration of ursodeoxycholate reduces serum liver enzymes in chronic liver diseases, but the nature of this effect is still unclear. We aimed at establishing a convenient in-vitro system for investigating the hepatotoxic properties of hydrophobic bile salts and the putative hepatoprotective effect of ursodeoxycholate. METHODS: About 100 mg of freshly isolated rat hepatocytes were suspended on a resin column (Bio-Gel P4 fine) and perifused with different concentrations of bile salts. The effluent was collected at 5-min intervals and assayed for lactate dehydrogenase (LDH), aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activity. Enzyme leakage induced by bile salts was compared with that induced by Triton X-100 (Union Carbide, Danbury, CT, USA) at different concentrations. After perifusion, hepatocytes were collected for electron microscopic observation. RESULTS: Cytotoxicity of individual bile salts, assessed by enzyme release, was time and concentration dependent and corresponded to their hydrophilic-hydrophobic balance. Perifusion with hydrophilic bile salts, cholate and ursodeoxycholate, did not result in a significant enzyme release in concentrations up to 5 mmol/l, whereas hydrophobic bile salts, chenodeoxycholate and deoxycholate, induced significant enzyme leakage even in low concentrations, 0.5 and 0.1 mmol/l, respectively. Addition of ursodeoxycholate significantly reduced the hepatotoxic effect of deoxycholate. This protective effect was evident within minutes. The ultrastructural appearance of hepatocytes exposed to hydrophobic bile salts was very similar to the non-specific cellular lysis observed after exposition to Triton X-100, suggesting that they act mainly in a detergent-like fashion. CONCLUSION: Perifused rat hepatocytes seem a convenient in-vitro system for investigating the hepatotoxic properties of bile salts and hepatoprotective effect of ursodeoxycholate, offering the opportunity to investigate the effects of bile salts under dynamic conditions, mimicking the in-vivo situation, and allowing continuous enzyme release monitoring. Hydrophobic bile salts seem to act mainly in a detergent-like fashion; ursodeoxycholate-related hepatoprotection could be due not only to a dilution effect of toxic bile salts, but also to a direct cytoprotective effect.

Morphological and functional evaluation of isolated rat hepatocytes in three dimensional culture systems.

Various three-dimensional configurations, such as polyester tissue and woven-nonwoven, hydrophilic polyester fabric, either collagen-coated or uncoated, were investigated as potential scaffold for hepatocyte culture, in view of their use in bioreactors for hybrid liver support systems. Attachment, morphology and ultrastructure of primary adult rat hepatocytes were evaluated, as well as urea production and ammonium detoxification during a 24h incubation period in serum-free tissue culture medium. As control, hepatocytes were also plated onto collagen-coated dextran microcarriers and on plastic petri dishes, either collagen-coated or uncoated. In all the three-dimensional cultures, hepatocytes appeared morphologically intact without any statistically significant difference in metabolic activity. Collagen-coating did not influence cell attachment to polyester substrates, whereas woven-nonwoven hydrophilic polyester fabric may offer some potential advantages as three-dimensional system for hepatocyte culture in hybrid liver support systems.

[Vascular complications in injuries of the knee joint (author's transl)]. / Lésions vasculaires dans les traumatismes du genou.

Over a total of 1,377 injuries of the knee joint, vascular complications were seen in 28 cases. Lesions of the popliteal artery or its branches are serious; over the total amount already mentioned, it was necessary to amputate in 12 cases; 12 patients presented the Volkmann syndrome. In cases of fractures of the lower end of femur, upper end of tibia and knee dislocations, the importance of early diagnosis of the acute ischemic syndrome is emphasized. Essential of emergency treatment are recalled.

Some traces of hidden codes.

Recent results in comparative genetics reveal processes of neutral evolution that are reminiscent of neutral codon substitutions, although they operate on novel kinds of sequences and molecular structures. This suggests that in addition to the genetic code, previously unrecognized "degenerate" codes might govern molecular interactions at other levels of biological information. If they exist, such multiple degenerate codes cannot be accounted for by the two concepts usually placed at the heart of molecular biology, i.e. molecular cascades and gene networks. This article reports on some representative examples of putative degenerate codes involving three fundamental levels of biological regulation, i.e. transcription, post-transcriptional regulation and signal transduction. From these examples we suggest that degenerate codes are organized hierarchically and that the concept of neutral evolution can be generalized to all kinds of molecular interactions. In addition, a case of functional evolution is interpreted as the emergence of novel, possibly degenerate codes. Comparative genetics and molecular embryology will be instrumental in testing the existence of multiple degenerate codes and hence, in unraveling the causes of evolution.

[Isolated hepatocytes and their potential therapeutic use]. / Epatociti isolati e loro potenziale uso terapeutico.

Isolated hepatocytes in culture represent an idoneous system for the study of liver physiology and metabolism. Furthermore, they are also widely utilized in pharmacological and toxicological study, in evaluating xenobiotic substance effects on the liver. In this paper, we reviewed the enzymatic methods for liver cell isolation in some mammalian species, as well as the techniques for qualitative and quantitative evaluation of cell number, vitality, purity, morphology and function. Recently, there has been a renewed interest in hepatocyte transplantation and hepatocyte-based liver support systems. From a clinical point of view, isolated hepatocytes could be useful in temporarily substituting an acutely damaged liver, a liver affected by a chronic pathology, or to correct an inherited liver disease carrying a severe metabolic derangement. Early experimental results of allogeneic hepatocyte transplantation, as well as the first clinical trials of bioartificial liver support systems employing xenogeneic hepatocytes are promising and contribute to maintain that interest in liver cell isolation and purification methods.

Acute psychosis in propionic acidemia: 2 case reports.

Propionic acidemia is an inborn deficiency of propionyl-coenzyme A (CoA) carboxylase activity, which leads to mitochondrial accumulation of propionyl-CoA and its by-products. Neurologic complications are frequent, but only a few cases presenting with psychiatric symptoms have been reported so far. We report 2 cases of children with chronic psychiatric symptoms who presented with an acute psychotic episode as teenagers. Both patients had hallucinations, panic and grossly disorganized behavior, for several weeks to several months. They had signs of moderate metabolic decompensation at the beginning of the episode, although the psychiatric symptoms lasted longer than the metabolic imbalance. We propose that these episodes were at least partially imputable to propionic acidemia. Such episodes require psychiatric examination and antipsychotic treatment, which may have to be adapted in case of cardiomyopathy or long QT syndrome.

[Prenatal symptoms and diagnosis of inherited metabolic diseases]. / Maladies héréditaires du métabolisme : signes anténatals et diagnostic biologique.

Inherited metabolic diseases are mostly due to enzyme deficiency in one of numerous metabolic pathways, leading to absence of a compound downstream from and the accumulation of a compound upstream from the deficient metabolite(s). Diseases of intoxication by proteins (aminoacidopathies, organic acidurias, urea cycle defects) and by sugars (galactosemia, fructosemia) usually do not give prenatal symptoms since mothers protect their fetuses from pathological metabolite accumulation. A well-known exception is hypoplasia of corpus callosum, as is sometimes observed in nonketotic hyperglycinemia and sulfite oxidase deficiency. Conversely, women with phenylketonuria "poison" their fetus if they are not treated (spontaneous abortions, intrauterine growth restriction [IUGR], cardiac malformations, and brain disease). Amino acid synthesis defects can lead to prenatal symptoms: microcephaly in serine deficiency (detectable by amino acid analysis in fetal cord blood), and brain malformations in glutamine synthetase deficiency. Impaired folate metabolism is involved in a large fraction of neurodevelopmental defects referred to as spina bifida, yet the underlying genetic component(s) are largely unknown. Energy metabolism diseases caused by defects in the synthesis or utilization of relevant metabolites lead to organ dysfunctions or malformations, but prenatal diagnosis is usually impossible unless genetic analysis can rely on a previously affected child in the family. A somewhat intermediate condition is defects of mitochondrial beta-oxidation of fatty acids, as they may sometimes be symptomatic prenatally (notably the HELLP syndrome or other presentations), and in this case, organic acid and acylcarnitine analysis in amniotic fluid can be informative in the absence of an index case. In contrast, complex molecule diseases commonly give prenatal symptoms that may permit the diagnosis even in the absence of index cases: hydrops fetalis and skeletal anomalies in lysosomal storage diseases, hydrops fetalis in congenital disorders of glycosylation (CDG) and transaldolase deficiency, brain malformations in O-glycosylation defects, brain malformations, kidney cysts and skeletal anomalies in peroxysomal diseases (Zellweger syndrome), syndactyly, genitalia malformations, and IUGR in Smith-Lemli-Opitz (SLO) syndrome. Although many metabolic disorders show biochemical abnormalities during fetal development that are informative for prenatal diagnosis, only a fraction of them are clinically/sonographically symptomatic before birth, thus allowing for prenatal diagnosis in the absence of an index case, i.e., serine deficiency, some fatty acid beta-oxidation defects, transaldolase deficiency, lysosomal diseases, CDG, Zellweger syndrome, and SLO syndrome.