Two boys with profound fasting hypoglycaemia and no ketone bodies

The first impression is that LF appears to have a similar problem to TFP, who was unable to metabolism long-chain fatty acids, and therefore became profoundly hypoglycaemic on fasting (see the exercise on "Muscle weakness and hypoketotic coma on fasting"). However, in this case we have no evidence of muscle weakness. While it is possible the LF cannot oxidise fatty acids, it is also possible that he is able to oxidise them, but cannot synthesise ketone bodies from the resultant acetyl CoA.

In order to determine whether his problem was in the oxidation of fatty acids, or the synthesis of ketone bodies from acetyl CoA, fibroblasts were cultured and incubated with [3H]fatty acids. The results show nmol [3H]water released per hour per mg protein for two replicate assays on LF's fibroblasts and mean ± sd (and range in parentheses) for fibroblasts from 20 control subjects.

(From data reported by Morris AAM, et al, Pediatr Res 44:3 392-6, 1998)

There is obviously no problem with LF's ability to oxidise fatty acids, so the problem must lie in the synthesis of ketone bodies from acetyl CoA.

The pathway of ketone body synthesis is shown below:

Hydroxymethylglutaryl CoA (HMG CoA) is an intermediate in the catabolism of the amino acid leucine. After an 11 hour fast, LF was given a dose of 200 mg /kg body weight leucine, and blood samples were taken for measurement of beta-hydroxybutyrate. The results are shown on the right:

LF can obviously make beta-hydroxybutyrate from leucine, which means that he has adequate activity of HMG CoA lyase. Since he cannot make beta-hydroxybutyrate from fatty acids, this suggests that his problem is lack of either beta-ketothiolase or HMG CoA synthase. You do not know this at present, but beta-ketothiolase is also involved in the catabolism of ketone bodies, so it is most likely that he lacks HMG CoA synthase.

Both LF and the control subject are fasting. The control subject has a normal ability to synthesise ketone bodies from acetyl CoA, and as his fast continues so he synthesises more to provide a metabolic fuel for muscle, so as to spare glucose for the brain and red blood cells.

By contrast, LF cannot synthesise ketone bodies from acetyl CoA. However, the beta-hydroxybutyrate that he has synthesised from leucine is metabolised by muscle - this explains why his plasma concentration of beta-hydroxybutyrate falls again after the peak produced from leucine has been metabolised.

In the fasting state there is more catabolism of tissue proteins than replacement synthesis. This is partly because of the high ATP cost of protein synthesis and partly because of the use of the amino acids released from tissue proteins being used for gluconeogenesis or as metabolic fuel. The most likely source of beta-hydroxybutyrate will be leucine liberated by the catabolism of tissue proteins. It is also possible that LF has some residual activity of HMG CoA synthase, so that he can synthesise a small amount of ketone bodies from acetyl CoA, but nowhere near enough to meet his energy needs in fasting.

You will see from the pathway of ketone body synthesis above that acetoacetate undergoes a non-enzymic decarboxylation to acetone. Acetone is poorly metabolised (if at all). Therefore formation of acetone represents a loss of (potentially valuable) metabolic fuel in the fasting state and starvation. Beta-hydroxybutyrate is stable, so reducing acetoacetate to beta-hydroxybutyrate minimises this loss.

Although acetone is metabolically more or less useless, clinically it is extremely useful, because it has a distinct smell, and can be detected on the breath. This provides a rapid way of telling that an unconscious person is ketotic, before blood or urine samples are tested for ketone bodies.

The presence of methylglutaconic and hydroxymethylglutaric acids in his urine (and also plasma) suggests that the most likely cause of his problems is lack of HMG CoA lyase. As hydroxymethylglutaryl CoA and methylglutaconyl CoA accumulate in liver, there will be a lack of CoA for other pathways. In order to release free CoA, some of these organic acids will be esterified to carnitine, and some of the hydroxymethylglutaryl CoA and methylglutaconyl coA will be hydrolysed to release the free acids and CoA.

As we saw in the case of LF, HMG CoA lyase is also the final step in synthesis of acetoacetate and beta-hydroxybutyrate from acetyl CoA, so he will be unable to synthesise ketone bodies to provide an alternative metabolic fuel for muscle and other tissues.

During his stay in hospital it was noticed that after a meal he frequently hyperventilated and became acidotic. He was given a test dose of 100 mg /kg body weight leucine after a 10 hour fast. As expected, there was no increase in his plasma concentration of ketone bodies, and he became severely acidotic, requiring an intravenous infusion of bicarbonate.

The metabolic acidosis is obviously due to an inability to metabolise leucine, because of the lack of HMG CoA lyase. This suggests that a low protein diet, and especially one with a very restricted amount of leucine, will be beneficial.

In addition, since he is unable to synthesise ketone bodies in the fasting state, he should avoid prolonged fasting. At times of stress (e.g. when he has an infection) it might be advisable to feed him a glucose polymer solution by naso-gastric tube through the night.

He was maintained on a low protein, leucine-restricted diet, and apart from a few metabolic crises when he has had an infection, he has remained well for the last four years.

Each degree C of fever represents a 13% increase in basal metabolic rate, so that after a normal overnight fast he will be more hypoglycaemic, and would normally be more dependent on ketone body synthesis, than usual.

It was considered possible that in addition to the failure to synthesised ketone bodies from acetyl CoA in the fasting state, his very profound hypoglycaemia might be the result of a defect in gluconeogenesis, or perhaps inhibition of one or another of the key enzymes of gluconeogenesis by HMG.

One obvious experiment would be to give him a test dose of alanine after an overnight fast. If there is no defect in, or inhibition of, gluconeogenesis, then his plasma glucose should rise, since alanine is one of the main gluconeogenic amino acids. This was done, and the result was that his plasma glucose rose after the test dose of alanine, suggesting that he does not suffer from any impairment of gluconeogenesis.

While he was in hospital he was given a slow intravenous infusion of beta-hydroxybutyrate during an overnight fast. In the morning his plasma concentration of glucose was significantly higher than after an overnight fast without beta-hydroxybutyrate infusion.

It is not possible to synthesis glucose from ketone bodies.

The fact that intravenous infusion of beta-hydroxybutyrate led to a higher fasting plasma glucose concentration must mean that providing beta-hydroxybutyrate as an alternative fuel for use by muscle and other tissues spared glucose for use by the brain and red blood cells, so that he was less hypoglycaemic. If muscle and other tissues are provided with ketone bodies they will not use glucose.

Key points from this exercise:

• Acetoacetate is synthesised by reaction of 3 mol of acetyl CoA to form acetoacetyl CoA, then hydroxymethylglutaryl CoA (HMG CoA), followed by cleavage to yield acetyl CoA and acetoacetate.
• Acetoacetate is chemically unstable, and undergoes non-enzymic decarboxylation to acetone, which is poorly metabolised (if at all). Therefore much of the acetoacetate synthesised in the liver id reduced to beta-hydroxybutyrate before export for use by muscle and other tissues as a metabolic fuel.
• Genetic defects of HMG CoA synthase lead to profound hypoglycaemia on prolonged fasting, with very low concentrations of ketone bodies. Small amounts of acetoacetate and beta-hydroxybutyrate are formed from the amino acid leucine.
• Genetic defects of HMG CoA lyase lead to profound hypoglycaemia on prolonged fasting, with no detectable ketone bodies. There is also metabolic acidosis due to accumulation of intermediates of leucine catabolism; HMG CoA is a normal intermediate in leucine metabolism.