Creatine supplementation prevents fatty liver in rats fed choline-deficient diet: a burden of one-carbon and fatty acid metabolism

Aim: To examine the effects of creatine (Cr) supplementation on liver fat accumulation in rats fed a choline-deficient diet.

Methods: Twenty-four rats were divided into 3 groups of 8 based on 4 weeks of feeding an AIN-93 control diet (C), a choline-deficient diet (CDD) or a CDD supplemented with 2% Cr. The CDD diet was AIN-93 without choline.

Results: The CDD significantly increased plasma homocysteine and TNFα concentration, as well as ALT activity. In liver, the CDD enhanced concentrations of total fat (55%), cholesterol (25%), triglycerides (87%), MDA (30%), TNFα (241%) and decreased SAM concentrations (25%) and the SAM/SAH ratio (33%). Cr supplementation prevented all these metabolic changes, except for hepatic SAM and the SAM/SAH ratio. However, no changes in PEMT gene expression or liver phosphatidylcholine levels were observed among the three experimental groups, and there were no changes in hepatic triglyceride transfer protein (MTP) mRNA level. On the contrary, Cr supplementation normalized expression of the transcription factors PPARα and PPARγ that were altered by the CDD. Further, the downstream targets and fatty acids metabolism genes, UCP2, LCAD and CPT1a, were also normalized in the Cr group as compared to CDD-fed rats.

Conclusion: Cr supplementation prevented fat liver accumulation and hepatic injures in rats fed with a CDD for 4 weeks. Our results demonstrated that one-carbon metabolism may have a small role in mitigating hepatic fat accumulation by Cr supplementation. The modulation of key genes related to fatty acid oxidation pathway suggests a new mechanism by which Cr prevents liver fat accumulation.


Alex’s Notes: Creatine synthesis within the liver consumes a considerable amount of our universal methyl donor, S-adenosylmethionine (SAM). The amino acids glycine and arginine are combined to form ornithine and guanidoacetate by the enzyme AGAT. Guanidoacetate then requires methylation from SAM using the enzyme GAMT to produce creatine.It has been suggested that the production of creatine accounts for up to 40% of the SAM used within the body. I’m trying to simplify the biochemistry as much as possible, but it is something that must be understood in order to appreciate yet another reason that creatine supplementation is beneficial.

In recent years, it has been proposed that the progression of fatty liver is associated with impaired methylation. Specifically, it has been attributed to a reduced availability of SAM and increased homocysteine levels. As we can see in this image here, SAM is formed from the amino acid methionine, which is obtained from the diet or resynthesized from homocysteine through folate and vitamin B12, or through betaine with the use of the BHMT enzyme.Because betaine is synthesized from choline, it follows that a choline-deficient diet (CDD) impairs remethylation via BHMT and, thus, decreases the availability of SAM.

Creatine supplementation (CR) has previously demonstrated ability to prevent fatty liver in rats fed a high-fat diet, and thus the current study aimed to clarify if this was related to methylation and SAM availability. As such, 24 male Wistar rats were randomly assigned to one of three groups: control, CDD, and CDD + CR, which was accomplished by adding 2% creatine monohydrate to the diet. After four weeks, the rats were decapitated and analyzed. This dose of CR was equivalent to about 0.74 g/kg/day in a human, which would be about 60g for a 180 lb individual and far above doses used even for loading.

I don’t want to say that dose is unrealistic because bulk monohydrate powder is cheap and plentiful, but you can be judge, as my point was merely to emphasize this is not a standard amount used by most persons. Anyway, there were no differences in weight, body fat, or food intake in the groups, and surprisingly no differences in liver weight either. Nonetheless, the CDD resulted in significant increases of total liver fat (55%), triglycerides (87%), cholesterol (25%), and plasma ALT compared to controls. Yet, CR not only completely prevented these increases, but actually reduced liver triglycerides to levels below the control group (not statistically significant though).

As would be expected from our homocysteine (Hcy) cycle image above, CDD also resulted in a significant increase in plasma Hcy concentrations (51%), and decreased hepatic SAM concentrations (25%). CR supplementation did prevent increases in Hcy, but failed to prevent the decline in SAM. This is interesting in light of the 80% reduction of AGAT activity (enzyme involved in creatine synthesis). In fact, the activity of GAMT (second enzyme in creatine synthesis that requires SAM) actually increased non-significantly above control values. Out of all the enzymes tested, the results suggested that methylation has a limited role in fatty liver and that CR’s role is limited to BHMT and GNMT.

The CDD also increased hepatic damage, inflammation, and oxidative stress as evidenced by the increase in plasma ALT (50%), TNF-α (241%), MDA (30%), and a decrease in liver GSH (31%). CR completely prevented all these alterations.

What does this all mean?

The CDD impaired methionine metabolism by decreasing hepatic SAM availability and reduced BHMT and GNMT gene expression. In other words, a choline deficient diet reduces the remethylation of Hcy to methionine by BHMT and restricts available methyl groups for SAM formation and subsequent methylations. However, creatine supplementation was able to suppress its own synthesis within the liver as shown by the 80% reduction of AGAT activity, which led to a reduction of Hcy as expected. Despite this reduction, CR was unable to restore hepatic SAM concentrations.

So the takeaway is thus that creatine monohydrate supplementation of a choline deficient diet prevents fatty liver through mechanisms that go beyond methylation and one-carbon metabolism.


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