The Metabolic Push-Me-Pull-You: HCV core protein, and the even-handed generosity of DM2: Gluconeogenesis and De Novo Lipogenesis, Oh My!

One of the features of HCV which I will probably return to again and again in this work-in-progress is that it represents a useful model of metabolic syndrome and DM2. HCV-infected hepatocytes run simultaneous gluconeogenesis and de novo lipogenesis, showing disordered insulin signalling (not just insulin resistance, which would decrease lipogenesis). The mechanisms involved (metabolic and immunologic) are being studied in detail and seem to have a lot to tell us about non-viral DM2. I would suggest that they are more relevant than drug-damage models of insulin deficiency or resistance; the interference is more subtle, the adjustments are clearly the results of adaptive processes, are highly effective, and are only mildly cytotoxic.

At the very least, HCV presents another angle from which to look at these problems.

Steatosis and insulin resistance in hepatitis C: a way out for the virus?

The hepatitis C virus (HCV) induces lipid accumulation in vitro and in vivo. The pathogenesis of steatosis is due to both viral and host factors. Viral steatosis is mostly reported in patients with genotype 3a, whereas metabolic steatosis is often associated with genotype 1 and metabolic syndrome. Several molecular mechanisms responsible for steatosis have been associated with the HCV core protein, which is able to induce gene expression and activity of sterol regulatory element binding protein 1 (SREBP1) and peroxisome proliferator-activated receptor γ (PPARγ), increasing the transcription of genes involved in hepatic fatty acid synthesis. Steatosis has been also implicated in viral replication. In infected cells, HCV core protein is targeted to lipid droplets which serve as intracellular storage organelles. These studies have shown that lipid droplets are essential for virus assembly. Thus, HCV promotes steatosis as an efficient mechanism for stable viral replication. Chronic HCV infection can also induce insulin resistance.

(The hepatitis C virus has evolved to be transmitted from infected cells on the lipid transport system; therefore it “wants” infected cells to maximise triglycerides and release of (HCV-carrying) VLDL; the higher the TAG, the more HCV virions in serum and the greater the chance of infection from blood-to-blood  encounters with the host; also, the virus can ensure that it stays a step ahead of the host’s immune defences by regularly infecting naïve cells.)

Del Campo and Romero Gomez, the authors of this paper, are experimenting with the statins (mainly fluvastatin) as anti-HCV agents. A sensible deduction from the evidence, but quite possibly not as sensible as carbohydrate restriction and intermittent fasting. However I don’t doubt that it is easier to get funding for a drug trial than for a trial of an antiviral Atkins-type diet. Yup, I can see how that suggestion might go down at the funding board.

A suggestion might be that the virus is inducing IR at the gluconeogenesis end, while promoting those genes that normally respond to insulin at the lipogenesis end of hepatocyte metabolism.

But what is the advantage of promoting gluconeogenesis and elevated blood glucose?

HCV can perhaps replicate more effectively if TAGs are not being used as a fuel, i.e., if carbohydrate is the cell’s main energy substrate. If viral manipulation of cell processes to promote gluconeogenesis results in elevated blood glucose, this will tend to prevent lipids “sponsored” by the virus being oxidised to fuel cell processes.  Gluconeogenesis is underwriting lipogenesis.
(correction: officially, at least, hepatocytes only run on ketoacids (pyruvate and oxaloacetate) from amino acid catabolism. However, another reference (Best and Taylor) implies that these are fasting-state gluconeogenesis substrates, and states that the newborn liver is wholly dependent on sugars and lipids. It seems more likely that any cell uses a mix of energy substrates and that in the case of hepatocytes the preferential usage is ketoacids, if only because other cells cannot metabolize gluconeogenic amino acids. Anyhow, this suggestion can be left in the air for now.)

How do infected hepatocytes (about 2-25% of the total, greater in non-responders, in one study ) manage to produce this two-way excess? We are used to cells that convert glucose (or fructose) or lipids to triglycerides, or glycogen and amino acids (or fructose) to glucose, but how does a cell do both at once? Does the cell take in more substrate than it normally would – a pate de foie gras forced feeding under viral prompting – or does it neglect the many other duties of a hepatocyte and squander the ATP produced through its mitochondrial density on its guest, or both?

[update 10-12-2012: HCV infection can associated with a DROP in tryglycerides; see this recent post. The virus is monopolizing VLDL in the same way it monopolizes cholesterol.]

This even-handed generosity is explained in diabetic research by concurrent insulin sensitivity and insulin resistance.

“[Although] an impairment of insulin receptor signaling to Foxo1 can explain insulin's inability to restrain HGP, one would predict that, if the liver were wholly insulin resistant, triglyceride (TG) synthesis and assembly into ApoB-containing lipoproteins would also be impaired. But the opposite is true in the diabetic liver.

In recent years, the idea that the diabetic liver may harbor a noxious brew of insulin resistance and excessive insulin sensitivity has gained a second wind.”

The HCV toxin that manipulates cell processes is called core protein and yields a number of fractions.

Confusingly, HCV core protein is also an integral part of the viral coat or capsid. Like Batman’s utility belt, it performs an amazing array of functions in its interactions with lipoproteins, mitochondria, immune system pathways, cell surface receptors, and RNA copying mechanisms. Like a pushy talent agent it ruthlessly promotes the interests of its RNA wherever it goes.

HCV core protein (like that of the more benign Hepatitis G virus, GVB-V) shares genomic features with plant oleosins. These proteins are found associated with fatty droplets in grains and seeds, and sesame oleosin is a type1 (IgE) allergen. Presumably oleosin-like properties (unique to these two related viruses) allow the close association with VLDL-LDL that is characteristic of the HCV lifestyle.

Some of the transcription pathways involved are (as we might expect, and to return to our muttons) very close to those implicated in non-viral DM2. HCV core protein promotes gluconeogenesis by increasing activity of nuclear Fox01 transcription factor (Fox01 is to glucose what NF-KappaB is to cytokines) through inhibition of phosphorylation by mitochondrial ROS (inhibition of Mito Complex 1 by HCV core protein).

In Robert Lustig et.al.’s DM2 hyperglycaemia scenario, fructose plays the same role (perhaps decreased phosphorylation is the result of fructose depleting [P1] as in the text book extract I will end with).

 “Hepatic insulin resistance, made worse by elevated fructose concentrations, prevents the phosphorylation of FoXo1, which allows this protein to enter the nucleus and induce the transcription of enzymes that promote gluconeogenesis. “

This is the one paper than anyone curious about Robert Lustig’s ideas should read, especially the sections on dietary fat vs dietary carbohydrate as factors in DM2.

N=1

in 2007 my HCV viral load was 400,000

Earlier this year, after a few months of more-or-less ketogenic diet (25-75g carbs), VL was 26,000

After a month or so of higher carbs (but still low-carb – 50-150g) latest VL was 60,000


Summary 

Trying to get these concepts and references into one smooth flow has been like herding cats, so I will summarize in plain language:

HCV (through its core protein) can promote both insulin resistance (elevating blood glucose) and/or insulin sensitivity (elevating fasting triglycerides). This both mimics and adds to dietary metabolic syndrome, and increases the risk of Type 2 Diabetes. And a diet and lifestyle that encourages DM2 will promote increases in viral load, disease pathology, and resistance to treatment.


Because HCV down-regulates GLUT2 to produce the gluconeogenic effect, reducing glucose uptake of infected hepatocytes, fructose becomes an ideal substrate for both gluconeogenesis and lipogenesis. Fructose consumption is predicted to optimize viral replication.

A carbohydrate-restricted version of the Paleo diet – Paleo-Atkins is a convenient shorthand for this – preferably with time-restricted feeding (16 hour daily fasts and 8 hour feeding windows, or at least no carbohydrate or protein outside of regular mealtimes),  is – or ought to be – the default diet for DM2 and fatty liver.

This diet ought to reduce the HCV viral load (as in my case), improve Hep C pathology (ditto), and improve the response to treatment (we'll see one day, maybe).

Research into HCV core protein effects on gluconeogenic and lipogenic regulation can provide insights into the mysterious aetiology of diabesity. For example,  tending to corroborate the theory that high-fructose diets play an important causative role in metabolic disease.


More on Fructose:

Not everyone metabolizes fructose the usual way, and not everyone who does clears it easily.

Everyone needs to metabolise glucose, and the genes for this have been conserved, but our ancestors sometimes survived for long periods without much fructose exposure, and these genes show more variety.

For example, the higher rate of gout in some South Pacific populations may be the result of adaptation to an ancestral diet low in fructose.

This classic account is from White, Handler and Smith's Principles of Biochemistry (c) 1954, the 1973 edition

Metabolism of Fructose

Although fructose can be phosphorylated in the 6 position at a slow rate by non-specific kinases, most of ingested fructose is phosphorylated in the liver by a fructokinase that specifically directs phosphorylation at the C-1 position of this ketose. No mutase is known that can catalyse conversion of fructose 1-phosphate to fructose 6-phosphate, nor can phosphofructokinase effect synthesis of fructose diphosphate from fructose 1-phosphate. The only pathway available to the latter is made possible by a specific aldose that catalyses the following reaction:

Fructose 1-phosphate ó dihydroxyacetone phosphate + glyceraldehyde

The further metabolism of glyceraldehyde requires reduction by NADH to glycerol, which is then phosphorylated by glycerol kinase, using ATP, and reoxidized by NAD+ to dihydroxyacetone phosphate. The latter then enters the usual glycolytic sequence.

Individuals who lack fructokinase excrete the major portion of ingested fructose in the urine. “Fructose intolerance” is a more serious illness, characterized by genetic lack of the aldose specific for fructose 1-phosphate, which accumulates after fructose ingestion and inhibits diverse enzyme systems. Even normal individuals may experience difficulty with large fructose intake. Although both the kinase and the special aldolase are present in large and equivalent activities, fructose 1-phosphate may accumulate in the liver for some time after a large fructose flux, e.g., after ingestion of a large quantity of sucrose.   

The explanation offered is as follows: Because of the effectiveness of the kinase, both [ATP] and hence [P1] are lowered in liver cells. The P1 inhibition of adenylate deaminase is thus released and inosinic acid [IMP] accumulates; in intact, perfused liver [IMP] increases seven-fold in ten minutes under such circumstances. However, IMP is a powerful inhibitor of the fructose 1-phosphate aldolase (K1 = O.1 mM and Km F-1-P = 0.18 mM), thus delaying further metabolism of this compound.

The presence of fructose and galactose in the intestine, with the amino acids, inhibits intestinal absorption of the latter.

Metabolic effects of androgenic compounds on the sex organs and tissues...include increased fructose production by seminal vesicles and utilization of this sugar by seminal plasma, with concomitant enhancement of the activity of both aldose reductase and ketose reductase.