The new insight that molecular genetics has cast on the subject of PDH-E1 deficiency, is enabling us to begin resolving some of the associated thorny problems. Knowing about the X linkage of the E1α has allowed us to put into perspective the equal number of females with PDH-E1 deficiency. Affected females have one normal E1α gene and one that carries the mutation, but even random lyonization, giving 50 percent overall activity, where the mutant allele gives no activity may produce an adversely affected individual. In the central nervous system, it is likely that such affected females will lose cells that have low viability because they express a null allele. With affected males, the situation is not quite as clear. Here the activity in skin fibroblasts of the PDH complex might be expected to represent that found in the brain, the organ where most pathology is experienced. For instance, we had two males, one with 1.6 percent residual activity and the other with 10 percent activity (refer to the HSC Web site www.sickkids.on.ca) who died of neonatal lactic acidosis. On the other hand, we have two surviving males with ataxia at 17 percent and 39 percent, while we had one with 30 percent who died at 13 months with psychomotor retardation and brain atrophy. Another male at 27 percent residual activity of the PDH complex merely suffered from exercise intolerance. Clearly some of these activities show little correlation between clinical phenotype and residual activity. In an affected male, we documented activities of 0.8, 4.5, 5.2, 13.2, 18.8, and 22.4 percent of normal activity of the PDH complex for kidney, liver, brain, heart, skeletal muscle, and skin fibroblasts, respectively. As we accumulate data, we begin to see that there is a differential turnover of the PDH complex in different tissues, which accounts for much of the loss of activity.116 In some males with defects affecting the C-terminus, the CRM+ve material correlates with residual activity, suggesting that if the complex could be stabilized in such cases, it would have close to normal activity. Clearly there is much to learn about the expression of normal and mutant forms of the E1α protein in tissues.
Defects in the E2 (MIM *245348) and X (MIM *245349) Components of the Pyruvate Dehydrogenase Complex
Seven patients have been described in whom there is good evidence of a defect in protein X or the E2 dihydrolipoyl transacetylase. In a patient with 24 percent PDH complex residual activity, who presented at 2 weeks of age with hyperammonemia and lactic acidosis, and who eventually died at 3.5 years of age with profound psychomotor retardation, there was only 32 percent of measurable transacetylase while E1 and E3 activities were normal.118 This patient had a normal amount of E1α, E1α, and E3 proteins, absent E2, and reduced amounts of protein X. Six patients with an absent protein X on fibroblast Western blots have been described.23,118-121 These patients had residual PDH complex activities of 12 to 20 percent in skin fibroblasts, and presented with psychomotor retardation usually accompanied by Leigh disease and development of spastic quadriplegia.23,118-121 Three of these patients were shown to possess homozygous deletions of 4 bp, 59 bp, and 85 bp, respectively, in the cDNA for X-protein.23,24 These results support the observations that in the bovine PDH complex, when the X protein is removed, only 15 percent residual activity is retained.123 Further evidence defines a definitive role for the X protein in the binding of the E3 subunit to the complex.25,123
A male born to nonconsanguineous Japanese parents presented at 3 years of age with an ataxic gait but normal mental development. By 5.5 years of age he was significantly worse with loss of fine and gross motor coordination and lesions observable in the putamen and basal ganglia. His lactic acid was elevated between 2.5 and 9 mM, although his residual fibroblast activity was 55 percent.118 An extra band just below protein X was consistently observed in immunoblots of his skin fibroblasts with anti-PDH antibody; E2, E3, Eα, and Eβ seemed to be present in normal amounts. It was concluded that this extra band was a mutant form of protein X, but subsequent analysis showed it to be an E2 band.
Lipoamide Dehydrogenase Deficiency (MIM *246900): Combined Α-Keto Acid Dehydrogenase Complex Deficiency
Combined deficiency of the α-keto acid dehydrogenase complexes is a comparatively rare entity, there being few well-documented cases in the world literature.124-129,134-136,194 Surprisingly, none of the affected children presented at birth, but they developed lactic acidemia that became troublesome at a few months of age. Because the data indicate that the α-keto acid dehydrogenase complexes include only the E3 component in common,27 it is appropriate that the E3 component, otherwise known as lipoamide dehydrogenase, be deficient in most cases of the combined defect. That all complexes are deficient is consistent with the observation of elevated pyruvate, lactate, α-ketoglutarate, and branched chain amino acids levels in blood samples from these patients.124,125,127,129 The branched chain amino acids are not elevated to the extent seen in classic maple syrup urine disease. Urine organic acids typically show elevated lactate, pyruvate, α-hydroxybutyrate, α-hydroxyisovalerate, and α-ketoglutarate.126,127,130 In one case, α-ketoisocaproic acid was present in addition to these other acids.129 When postmortem examination of the brain was carried out,124-126 myelin loss and cavitation were found in discrete areas of the basal ganglia, thalamus, and brain stem; the cerebral cortex appeared to be free of pathology.
The diagnosis was made initially by measuring activity of the α-keto acid dehydrogenase complexes in tissues or in fibroblasts.124-126 In seven of nine known cases, the combined defect in the complexes is definitely the result of a defect in lipoamide dehydrogenase,124-126,128,134-136 which was between 0 and 20 percent of the activity found in controls. In another two cases, despite the α-keto acid dehydrogenases being deficient to the extent of 25 percent of normal, lipoamide dehydrogenase activity was 59 and 63 percent of normal.127,131 This anomalous situation could be explained if it is postulated that the abnormal protein can carry out the lipoamide dehydrogenase reaction, but the ability of these E3 proteins to interact with the E2 transacetylases is affected by the mutation.127
Hinman et al.132 described two patients who died of Leigh disease in which abnormalities were described in the ability of their lipoamide dehydrogenase to restore activity to E3-depleted PDH-complex preparations. The fibroblasts from these patients had 60 percent of normal PDH complex, even when activated with dichloroacetate.132 Two mutations in the lipoamide dehydrogenase cDNA, K37E and P453L, were described in a patient with 6 percent E3 activity but 10 to 30 percent of PDH-complex activity, who died after a ketoacidotic episode at 12 months of age.133 Two further patients were characterized at the molecular level by the same group. A patient who had a combination of A1173G/del455-457 died at 5 years with microcephaly and psychomotor retardation. Enzyme activity for lipoamide dehydrogenase was 3 percent and PDH complex was 31 percent in fibroblasts.134 A second patient who died at 28 months with delayed development, hypotonia, and Leigh disease had a combination of insA105/G1533A resulting in one transcript with R460G and one null allele due to the frameshift in the mitochondrial leader sequence.135 A milder version of the disease has been described with normal cognitive function but some motor impairment at 5 years of age.136 InsA105 has also been described in two patients with a similar presentation.137 A patient with prominent hepatocellular disease was also described by the same group with 12 percent residual lipoamide dehydrogenase activity and normal cognitive function.138
Pyruvate Dehydrogenase Phosphatase Deficiency
In a small number of cases with congenital lactic acidemia, a defect was demonstrated in the enzyme which activated the PDH complex by removing phosphate groups from E1α serine residues. Three cases, two reported by Sorbi and Blass139 and one by De Vivo et al.,140 had a clinical picture typical of Leigh disease, while a fourth patient described by Robinson and Sherwood141 died at age 6 months after a course of unremitting lactic acidosis. Two of the cases140 showed poor reactivation of the PDH complex after inactivation by incubation with ATP in postmortem tissues. The other cases139 showed a normal activity of the complex in fibroblasts in the native state, but no activation could be demonstrated after incubation of the fibroblasts with dichloroacetate. None of the methods used in these studies was a direct assay of the activity of pyruvate dehydrogenase phosphatase activity. A method was described by Wicking et al.48 for measuring PDH phosphatase activity in fibroblast by the release of32 P from added32 P-labeled PDH phosphate. This is a more direct method of measurement that should be capable of detecting abnormal activity of this enzyme.
Pyruvate carboxylase is a biotin-containing protein of subunit molecular weight Mr = 125,000, each active enzyme molecule consisting of four tightly bound identical subunits. Each subunit has one molecule of covalently bound biotin and possesses binding sites for pyruvate, ATP, HCO3, and acetyl CoA.142-144 The enzyme is almost totally dependent on the presence of acetyl CoA as an allosteric activator for activity. As the first enzyme in the gluconeogenic pathway, it is activated in conditions where fatty acids are mobilized and acetyl-CoA is generated.145
Pyruvate carboxylase is widely viewed as the major regulatory enzyme and the flux-generating step in the pathway of gluconeogenesis,145 being regulated by the relative acetyl-CoA/CoA and ATP/ADP ratios in liver mitochondria.146 This enzyme is always intramitochondrial and has its highest activity in liver and kidney, where its role in gluconeogenesis is important. It is found in lesser amounts in other tissues such as brain, muscle, adipocytes, and fibroblasts where its function is believed to be anaplerotic.147-149 In these tissues, it plays a role in the maintenance of 4-carbon intermediates in the citric acid cycle.150
The full-length cDNA has now been elucidated for both the yeast151 and mammalian152,153 pyruvate carboxylase. The partial human gene has been mapped to the long arm of chromosome 11 at 11q13154, and the coding region of the human gene consists of 19 exons covering 16 kb of sequence.156 The rat gene structure has been elucidated and found to have 19 exons in the coding sequence, with two alternative 5′ untranslated sequences in front of the initial ATG. The two alternative 5′UTRs are generated from different promoters in such a way that heavy expression of PC takes place in liver and kidney, while a lower level of expression is maintained in other tissues157 (Fig. 100-11).
The generation of alternative transcripts from the pyruvate carboxylase gene. Two types of transcript are generated: Class I starting at exon 1D and Class II starting at 1B in a tissue-specific fashion.
Human Deficiency (MIM 266150).
The known instability of pyruvate carboxylase in suboptimal conditions of preservation and storage plagued the early attempts to define the nature of human pyruvate carboxylase deficiency.155 Many reports were based on measurements of enzyme activity in liver biopsy or postmortem liver specimens, and although some of these cases were undoubtedly bona fide cases of pyruvate carboxylase deficiency, some almost certainly were not. Early reports associated pyruvate carboxylase deficiency with subacute necrotizing encephalomyelopathy (Leigh disease).158 In a series of nine patients with Leigh disease, skin fibroblast pyruvate carboxylase was examined and found to be normal.158 In five patients, the diagnosis of Leigh disease was confirmed at autopsy, and in eight the urine inhibitor of thiamine triphosphate synthesis was present. Hommes et al.155 reviewed a number of cases where diagnosis of pyruvate carboxylase deficiency was made on liver tissue obtained by biopsy or at postmortem. The majority of these determinations are reported as single measurements and many of them have low or undetectable activity. In one case,159 detailed measurements of the kinetics of pyruvate carboxylase activity were made; it was found that a low Km component of the enzyme was missing. In the partially purified human enzyme, there are two kinetic components, one with a low Km and one with a high Km for pyruvate.144 This type of defect is difficult to test for and may not be detected in single-measurement assay systems for enzyme activity.
The demonstration that pyruvate carboxylase activity could be measured in cultured skin fibroblasts led to the accurate definition of the clinical sequelae of pyruvate carboxylase deficiency.148,149,160 There appear to be three distinct groups of patients who have been identified using the highly reproducible cultured skin fibroblast assay (Table 100-3). This initially became evident because the patients presenting with pyruvate carboxylase deficiency in North America (Group A) had a simple presentation of lactic acidemia and psychomotor retardation,161,162 while the patients presenting in France and the United Kingdom (Group B) had a more complex biochemical presentation with lactic acidemia, hyperammonemia, citrullinemia, and hyperlysinemia.161,163-165 Those in Group B all presented in the neonatal period and died before 3 months of age as compared with the longer survival of Group A. Group B patients also demonstrated a redox disturbance, the cytosolic compartment being more reduced, as evidenced from an increased ratio of blood lactate to pyruvate, and the mitochondrial compartment being more oxidized, as judged by a higher ratio of blood acetoacetate to β-hydroxybutyrate. In later reports, there are some Group A patients who have a blood lactate to pyruvate ratio above the normal range. Both Group A and Group B patients exhibit hyperalaninemia and hyperprolinemia and have less than 5 percent pyruvate carboxylase activity in their fibroblasts as compared to controls.160-173 Proximal renal tubular acidosis was present in 3 Group A cases, and 12 of 18 Group A cases were full-blooded North American Indian children, 8 were from either the Manitoba or Ontario Ojibwa, 2 were from the Saskatchewan Cree, and 2 were from the Micmac in Nova Scotia.174 The one unifying factor about these tribal communities is that they belong to the same linguistic group of Algonquian-speaking Indians.174 Thus, there was strong epidemiologic and anthropologic evidence for a founder effect in relation to pyruvate carboxylase deficiency in these Amerindian populations.
Table 100-3: The Presentation of Human Pyruvate Carboxylase (PC) Deficiency |Favorite Table|Download (.pdf) Table 100-3: The Presentation of Human Pyruvate Carboxylase (PC) Deficiency
| ||Group A (18 Cases) ||Group B (11 Cases) ||Group C |
|Origin ||12 Amerindian, 1 Canadian with Italian parents, 4 U.S. Caucasian, 1 Japanese ||2 Canadian (1 with Egyptian parents), 4 French, 2 U.K., 1 West German, 1 Saudi Arabian, 1 Swede ||1 U.S. Caucasian |
|Presentation ||Metabolic acidosis ||Metabolic acidosis ||Acute metabolic acidosis |
| ||Delayed neurologic development ||Hepatomegaly || |
|Age of presentation ||Birth to 5 months ||Neonatal ||14 months |
|Survival ||2 survive to 5 years; severe mental retardation ||All dead within 3 months ||7 years survival with normal development |
|Biochemical ||α-Ketoglutarate in urine ||α-Ketoglutarate in urine ||α-Ketoglutarate in urine |
| ||Lactic acidosis mild with severe attacks on infection ||Lactic acidosis chronically severe || |
|Pyruvate carboxylase activity ||<5% control ||<5% control ||<5% control |
|Lactate/pyruvate ratio ||Normal ||Elevated (x5) ||— |
|Acetoacetate/3HOB ratio ||Normal? ||Elevated (x5) ||— |
|Blood ammonia ||Normal ||Elevated (x5) ||Normal |
|Alanine ||Elevated ||Elevated ||Elevated |
|Citrulline ||Normal ||Elevated (x5) ||Normal |
|Lysine ||Normal ||Elevated ||Elevated |
|Proline ||Elevated ||Elevated ||Elevated |
|PC[3H] biotin protein (125 kDa) ||Present ||Absent in 6/10 cases ||— |
|PC immunoreactive protein ||Present ||Absent in 6/10 cases ||Yes |
|PC mRNA ||Present ||Absent in 4/10 cases ||Present |
| ||Refs. 159–165 +5 unpublished cases* ||Refs. 167–172 +3 unpublished cases* ||Ref. 174 |
A third distinct variety of pyruvate carboxylase deficiency was described by Van Coster et al.175 This child frequently presented with metabolic acidosis as an infant, but between these episodes seemed to be well. Her skin fibroblast pyruvate carboxylase level was measured at 1.8 percent of control values, which could account for her increased levels of blood lactate. Although many of her blood parameters were the same as the Group A cases of pyruvate carboxylase deficiency, she developed normally and did not become psychomotor retarded. At 7 years she may have a learning deficit in mathematics and slight dysarthria but is otherwise healthy. She is a large child (>95 percentile) for height and weight, who is treated with oral fluids and bicarbonate at the onset of febrile illness.175
The biotin-containing enzymes present in cultured skin fibroblasts can be visualized either by [3H]biotin labeling (Fig. 100-12) or by [35S]streptavidin blotting. In both cases, the cell proteins are separated by sodium dodecylsulfate (SDS) polyacrylamide gel electrophoresis.163,164,166 When this was done with fibroblasts from patients with pyruvate carboxylase deficiency, it was found that the patients with Group A or C presentation showed the same 125-kDa band corresponding to the subunit of pyruvate carboxylase. However, many of the patients in Group B showed no band for pyruvate carboxylase, but did show normal bands for the α subunit of propionyl CoA carboxylase (73 kDa) and the α subunit of methyl crotonyl CoA carboxylase (75 kDa).161,164 Immunoprecipitation of labeled protein from cells preincubated with [35S]methionine also showed absence of the 125-kDa protein in some of the Group B patients when antipyruvate carboxylase antiserum was used.161,164 Finally, Northern blotting with a cloned cDNA probe for pyruvate carboxylase showed absent mRNA in four of six Group B patients who lacked a demonstrable pyruvate carboxylase protein.164
Incorporation of biotin into carboxylase protein. The mRNA for carboxylase produces a translated apocarboxylase, which must incorporate activated biotin in the form of biotinyl-AMP with the release of 5′ AMP. The resulting holocarboxylase protein has biotin covalently attached at a lysine residue.
In trying to correlate the biochemical symptoms of those patients with the physical and molecular facts about pyruvate carboxylase that arise from the study of the patient's skin fibroblasts, it is evident that in patient Groups A and C, all patients produce a biotin-containing 125-kDa pyruvate carboxylase protein that has little activity. In Group B, there are two groups of patients who are different in molecular terms: those who produce a 125-kDa biotin-containing pyruvate carboxylase protein and those who do not. However, all Group B patients are more severely affected, with death in infancy, and they have a complex biochemical disturbance affecting pyruvate metabolism, the urea cycle, and intracellular redox states.174 What then determines whether pyruvate carboxylase deficiency presents with the simple phenotype of lactic acidemia or with the complex phenotype of lactic acidemia, citrullinemia, and hyperammonemia? Because we know that the pyruvate carboxylase gene is not expressed in all cases in which the presentation is the complex Group B phenotype, we can associate this phenotype with total absence of activity. Thus, we can hypothesize that the other patients in Group B who have expression of pyruvate carboxylase protein must also have total absence of activity to attain this phenotype. Where there is expression of the enzyme with the milder Groups A and C phenotypes, we must assume that there is enough residual activity in the mutant pyruvate carboxylase to ameliorate the most severe symptoms of the deficiency, but in the case of Group A, not enough to prevent lactic acidemia and psychomotor retardation.
That all patients with pyruvate carboxylase deficiency develop lactic acidemia is almost certainly due to a failure of the Cori cycle and, in times of starvation, also to a failure of gluconeogenesis itself. Patients in both groups have been documented with hypoglycemic episodes, although this is not a major problem.164 The Group B patients with the complex presentation show features that are suggestive of depletion of intracellular aspartate and oxaloacetate.168-170 In the urea cycle, aspartate is the second nitrogen donor, and low levels would cause the accumulation of both citrulline and ammonia (Fig. 100-13). Aspartate is also an essential component of the shuttle system that is responsible for the transport of reducing equivalents from the cytosol to the mitochondria.176 Electrogenic ejection of aspartate from mitochondria is necessary to maintain the typically very oxidized NAD+/NADH ratio in the cytosol as opposed to the very reduced NAD>+/NADH ratio in the mitochondrial compartment. A lack of aspartate would result in the cytosol being more reduced and the mitochondria being more oxidized, exactly the situation seen in the Group B patients with the complex phenotype. Thus, it would seem that pyruvate carboxylase activity in the Group B patients is so low that it cannot sustain oxaloacetate and aspartate levels. The few tissue measurements of pyruvate carboxylase activity that have been done in liver have yielded 6.2, 17.2, and 0.0 percent of control values for three Group A patients and 0.3 and 0.3 percent for two Group B patients.149,158,160,168
The role of pyruvate carboxylase in the generation of aspartate for use in the urea cycle and the reoxidation of NADH. See text for explanation.
The pathology of pyruvate carboxylase deficiency has been documented in one Group A and two Group B cases, and is primarily present in liver and brain. Hepatomegaly, which is seen frequently in this defect, seems to be due to lipid droplet accumulation in hepatocytes.148,158,160,174 In one Group A case, hyperplasia of hepatocyte endoplasmic reticulum was seen,148 and the architecture of liver mitochondria had an abnormal appearance with increased matrix density, increased matrix granule size, and dilatation of intracrystal space.173 The central nervous system pathology common to both Group A and B patients consists of very poor myelination and paucity of neurons in the cerebral cortex, gliosis, ventricular enlargement, thinning of the corpus callosum, and proliferation of astrocytes.149,168,173 In both Group B patients that came to autopsy, there was additional damage in the form of cavitated infarcts or cysts present in the cerebral cortex.168,173 In one case, microscopic examination of the kidney showed diffuse vacuolation of the kidney tubules.148 The appearance of cerebral cortex in pyruvate carboxylase deficiency suggests that myelination is not taking place, that neuronal death is occurring, and that a virtual developmental arrest of the brain is the net result. Thus, pyruvate carboxylase is obviously essential for normal brain development, and this is related to its anaplerotic role in metabolism. It has been documented that pyruvate carboxylase activity is plentiful in astrocytes but is low or absent in neurons.177 Current evidence suggests that neurotransmitter pools are replenished in part by de novo synthesis within the presynaptic terminals and that glutamine derived metabolically from astrocytes appears to be a major metabolic precursor of the transmitter pools for both glutamate and γ-aminobutyric acid (GABA). The anaplerotic formation of glutamine is thought to occur using pyruvate carboxylation as the initial step, and this can only take place in astrocytes. Thus, in an individual lacking pyruvate carboxylase activity, the neurons lose their ability to be replenished with glutamine from astrocytes, and depletion of 4- and 5-carbon intermediates in the neuron may result in neuronal death. To compound this problem, the lack of the anaplerotic function of pyruvate carboxylase in myelin lipid synthesis at the same time leads to poor or absent myelin formation. The absence of these two essential roles of pyruvate carboxylase in the anaplerotic processes of brain metabolism undoubtedly is a major contributor to the pathology seen in the central nervous system in this disorder. Group A children who survive with this defect are grossly mentally retarded and often have accompanying seizure activity. This lack of a key anaplerotic enzyme may also be the cause of abnormalities seen in kidney and of accumulation of lipid in Type 1 skeletal muscle fibers, in addition to the urea cycle and redox abnormalities previously mentioned. Presumably, the patient placed in Group C has enough activity to maintain normal neurologic function.175
Mutations Present in the Pyruvate Carboxylase Gene.
Investigation of the Amerindian form of pyruvate carboxylase deficiency has shown that in 13 patients tested, a c.1828G→A missense mutation was homozygous, these individuals all being either Ojibwa or Cree in origin.156 A c.2229G→T transversion was present in two Micmac brothers, showing that the deficiency in this group was not due to a founding effect.156 In some communities of the Ojibwa, the carrier rate for this mutation was as high as 1 in 10 individuals.156 The two mutations caused an Ala to Thr change at residue 610 and a Met to Ile change at residue 743 respectively, both of these being in conserved regions possibly involved in pyruvate binding.156
Heterozygote Detection and Prenatal Diagnosis.
The activity of pyruvate carboxylase in the cultured skin fibroblasts from normal individuals varies over quite a wide range. For this reason, although it may be possible to identify heterozygotes for this defect within a family, it is not possible to do this in the general population.162,178 To add to the confusion, there is a report of a patient with a 50 percent deficiency of pyruvate carboxylase in skin fibroblasts, who had severe chronic lactic acidemia.179 This patient had a kinetic defect in the enzyme that was perhaps similar to a case described earlier.159 This type of defect awaits a clearer definition in both enzymatic and clinical terms.
Prenatal diagnosis of an affected child with pyruvate carboxylase deficiency was reported on two occasions. Both families had already had a child who died with the Group B complex presentation of the defect.180,181 In one case, the absence of [3H]biotin-labeled protein was demonstrated in amniocytes.181
Phosphoenolpyruvate Carboxykinase Deficiency
A defect in this enzyme is rarely reported as a cause of lactic acidemia in childhood. The enzyme exists in two compartments in two distinct isoenzymic forms, and for this reason, the diagnosis of a suspected deficiency of either one of the two isoenzymes is difficult.
Recently, the human enzyme cDNA sequence was established for both isoforms, which have 78 percent identity at the amino acid level.182 Three cases of phosphoenolpyruvate carboxykinase (PEPCK) deficiency have been documented in which the assay of activity was carried out with a liver homogenate.183,184 In another group of cases, the cytosolic PEPCK in liver was measured and found to be deficient185 (MIM 261680). In two cases, the defect was defined by measurement of PEPCK in cultured skin fibroblasts.74,149 Because it has been shown that the majority of PEPCK present in fibroblasts is mitochondrial in origin, the detection of 15 and 16 percent74,149 of normal activity in these cases was suggestive of deficiency of mitochondrial PEPCK (MIM 261650). This was confirmed by the demonstration of 6 percent of normal activity in the mitochondria from fibroblasts of one patient.74
Both of the children described with mitochondrial PEPCK deficiency had lactic acidemia, hypoglycemia, hypotonia, hepatomegaly, and failure to thrive.72,186,187 One patient had more severe symptoms, and, in addition, had peripheral edema, disordered liver function, and episodes of unexplained pyrexia; she died at age 6 months.187 The other patient with mitochondrial PEPCK deficiency had survived to age 10 years with some continuing muscular weakness and hypotonia. These latter symptoms are most likely due to a lack of mitochondrial PEPCK in muscle, where it is thought to play an essential role in the regulation of the pool size of 4-carbon intermediates.150 A later publication described the sib of one of these PEPCK-deficient cases who had the same clinical presentation but normal PEPCK activity in leukocytes and fibroblasts.188 This obviously makes it unlikely that PEPCK was the primary deficiency in this family.188
The cytosolic form of PEPCK is subject to induction and repression, being induced by catabolic states and repressed by anabolic states.189 There is a strong suggestion in the cases described by Vidnes and Sovik185 that the hypoglycemia and low cystolic PEPCK seen in this group of neonates was a result of hyperinsulinism, a condition that would repress cytosolic PEPCK expression in the liver. Hepatomegaly and hypoglycemia were present in two infants described by Hommes et al.184 with 5 and 10 percent of control PEPCK activity. Localization of the site of the PEPCK was not attempted in postmortem liver samples from these children, both having succumbed to uncontrollable hypoglycemic episodes. Interestingly, one of these children had the inexplicable hypertriglyceridemia and hypercholesterolemia that was evident in the child we described.72,186,187