Pyruvate dehydrogenase phosphatase1 mRNA expression is divergently and dynamically regulated between rat cerebral cortex, hippocampus and thalamus after traumatic brain injury: a potential biomarker of TBI-induced hyper- and hypo-glycaemia and neuronal vulnerability.

Cerebral pyruvate depletion and lactate acidosis are common metabolic characteristics of patients with traumatic brain injury (TBI) and are associated with poor prognosis. Pyruvate dehydrogenase (PDH) is the rate-limiting enzyme coupling glycolysis to mitochondrial tricarboxylic acid (TCA) cycle. Brain PDH activity is regulated by its phosphorylation status and other effectors. Phosphorylation of PDH E1α1 subunit by PDH kinase inhibits PDH activity while dephosphorylation of phosphorylated PDHE1α1 by PDH phosphatase (PDP1) restores PDH activity. In situ hybridization showed that PDP1 mRNA is highly expressed in the cerebral cortex, hippocampus and thalamus of rat. Controlled cortical impact (CCI) induced a significant increase in PDP1 mRNA expression in ipsilateral cerebral cortex at 4 h (P<0.05) and 24 h post CCI (P<0.01) that returned to basal level 72 h post CCI. PDP1 mRNA level increased transiently in ipsilateral hippocampal dentate gyrus and CA1-3 subfields 4 h post CCI (P<0.01) but decreased significantly 24 h and 72 h (P<0.01) post CCI, coinciding with a marked increase in neuronal apoptosis in ipsilateral hippocampus 24 h post CCI. PDP1 mRNA expression in thalamus and other subcortical regions decreased persistently post CCI. Contralateral CCI and craniotomy showed similar effects on PDP1 mRNA expression as ipsilateral CCI. Because GFAP mRNA expression was induced in brain regions where PDP1 expression was altered, further study should determine the potential relationship between astrocyte activation, PDP1 alteration, and pyruvate metabolism following TBI.

Cloning, expression, and properties of the regulatory subunit of bovine pyruvate dehydrogenase phosphatase.

cDNA encoding the regulatory subunit of bovine mitochondrial pyruvate dehydrogenase phosphatase (PDPr) has been cloned. Overlapping cDNA fragments were generated by the polymerase chain reaction from bovine genomic DNA and from cDNA synthesized from bovine poly(A)+ RNA and total RNA. The complete cDNA (2885 base pairs) contains an open reading frame of 2634 nucleotides encoding a putative presequence of 31 amino acid residues and a mature protein of 847 residues with a calculated Mr of 95,656. This value is in agreement with the molecular mass of native PDPr (95,800 +/- 200 Da) determined by matrix-assisted laser desorption-ionization mass spectrometry. The mature form of PDPr was expressed in Escherichia coli as a maltose-binding protein fusion, and the recombinant protein was purified to near homogeneity. It exhibited properties characteristic of the native PDPr, including recognition by antibodies against native bovine PDPr, ability to decrease the sensitivity of the catalytic subunit to Mg2+, and reversal of this inhibitory effect by the polyamine spermine. A BLAST search of protein data bases revealed that PDPr is distantly related to the mitochondrial flavoprotein dimethylglycine dehydrogenase, which functions in choline degradation.

Pyruvate dehydrogenase phosphatase deficiency: a cause of congenital chronic lactic acidosis in infancy.

A male child presented on the first day of life with metabolic acidosis with elevated blood lactate (15 mM), pyruvate (0.4 mM), and free fatty acid (1.3 mM) levels and a blood pH of 7.16. The severity of the acidosis was diminished by intravenous administration of glucose in large doses and by bicarbonate. On two occasions, when the acidosis was particularly severe, peritoneal dialysis using an acetate buffer was required. Restriction of the dietary intake of saturated fatty acids or treatment with nicotinic acid also appeared to diminish the severity of acidosis. No improvement was achieved by the administration of thiamine or biotin. Tissues taken at postmortem showed normal activity of gluconeogenic enzymes and pyruvate dehydrogenase. The activity of pyruvate dehydrogenase in tissue homogenates preincubated with ATP was reduced by 60-75% both in liver of the patient and of the controls because of the inactivation of the enzyme by pyruvate dehydrogenase kinase. Addition of Ca++ and Mg++ to the inactivated enzyme caused a prompt return of the activity to normal in controls but not in the patient. This defect, which was apparent in muscle and liver but not in brain, we attribute to a markedly reduced activity of pyruvate dehydrogenase phosphatase in the patient.

Activation of pyruvate dehydrogenase in adipose tissue by insulin. Evidence for an effect of insulin on pyruvate dehydrogenase phosphate phosphatase.

1. The mechanism by which insulin activates pyruvate dehydrogenase in rat epididymal adipose tissue was further investigated. 2. When crude extracts, prepared from tissue segments previously exposed to insulin (2m-i.u/ml) for 2min, were supplemented with Mg-2+, Ca-2+, glucose and hexokinase and incubated at 30 degrees C, they displayed an enhanced rate of increase in pyruvate dehydrogenase activity compared with control extracts. 3. When similar extracts were instead supplemented with fluoride, ADP, creatine phosphate and creatine kinase, the rate of decrease in pyruvate dehydrogenase activity observed during incubation at 30 degrees C was unaffected by insulin treatment. 4. It is suggested that insulin increases the fraction of pyruvate dehydrogenase present in the tissue in the active dephospho form by increasing the activity of pyruvate dehydrogenase phosphate phosphatase.