Monday, February 17, 2014

Protein Power - Study Suggests: 40%+ is Where True Magic Happens. Plus: If Protein is King, Whey is the Emperor

It's not just about more protein it's about significantly more protein and - possibly - also about whey!
Just to make sure: Yes, I know the study I am about to discuss in today's SuppVersity article is a rodent study - a rodent study by researchers from the University College Cork and the University College Dublin (Mc Allan. 2014).  And yes, I know that you ain't no fury little mouse or rat...

... but I do also know that the beneficial metabolic effects of high protein intakes appear to be even more, not less pronounced in human beings and will thus not mention 500x that the assumption that we'd see similar benefits in men and women would obviously require experimental confirmation... alright?

Now that we are clear, dear non-dams and non-bucks...

... you are probably already drooling at the sought of reading yet another "high protein is good for you" study. Don't worry I am not going keep you on the tenderhooks longer than absolutely necessary. What is necessary, though is a very brief summary of the study design, which was designed to elucidate the effects macronutrient quality and composition on energy balance and the gut microbia - probably two of the hottest topics in today's discussions on the health and fitness bulletin boards of this world.
You can learn more about protein intake at the SuppVersity

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Spread or waste your protein?

Protein requ. of athletes

High EAA intra-workout fat loss

Fast vs. slow protein

Too much ado about protein?
As the Irish researcher point out, their goal was to investigate how changes to protein quality (casein versus whey protein isolate; WPI) and the protein to carbohydrate (P/C) ratio within a high fat diet (HFD) impacts on the aforementioned parameters. The questions the experiment was supposed to answer were thus:
  • Protein Quality Would adding whey protein on top of an obesogenic high fat rodent diet yield to a different weight and microbiota response than casein protein?
  • Protein Quantity ➲ Would diets with 20%, 30% or 40% of the total energy intake from protein have different effects on body weight and microbiota in the rodents?
In view of the fact that casein is the standard protein in many of the high fat diets that are used in experiments like this, the study would thus also be able to give us an idea of whether or not the use of the slow-releasing IGF-1 boosting dairy protein contributes to the obesogenic effects.

As it turned out, the analysis of issues related to question #2, i.e. "Would diets with 20%, 30% or 40% of the total energy intake from protein have different effects on body weight and microbiota in the rodents?" did produce the more intriguing results, though.
Figure 1: Weight gain, fat and lean mass, as well as energy intake and respiratory exchange ratio (RER) after 21-weeks on diets with different amounts of whey protein in them (McAllan. 2014),
If you take a look at the data in Figure 1, it's easy to see that (a) in comparison to casein (data from casein experiment not shown, because it was not discussed in detail | maybe there will be a follow up paper!?), WPI at a similar energy content normalised energy intake, increased lean mass and caused a trend towards a reduction in fat mass (P= 0.08). You may find that surprising, but it's actually been known for quite some time now that whey buffers many of the ill-health effects of high fat diets in rodents.
Figure 2: Adipose tissue mRNA expression of selected genes (McAllan. 2014)
Although the addition of whey protein did not alter the oxygen consumption or locomotor activity, it was able to ...
  • reduce the plasma leptin and liver triacylglycerold levels, and...
  • attenuate the reduction in adipose FASN mRNA 
in HFD-fed mice (compared to what the researchers observed in rodents on the casein chow). Moreover, a high throughput sequence-based analysis of faecal microbial populations revealed that the
"[...]microbiota in the HFD-20% WPI group clustering closely with HFD controls, although WPI specifically increased Lactobacillaceae/Lactobacillus and decreased Clostridiaceae / Clostridiumin HFD-fed mice." (McAllan. 2014)
To understand the potential implications of these changes we will have to take a closer look at the recent evidence linking Clostridiaceae and Lactobacillaceae to the diabesity epidemic:
  • Lactobacillus reuteri has anti-breast-cancer effects as well  (Lakritz. 2014).
    certain types of clostridiaceae are characteristic for obesity prone animals; their transplanation to normal mice will make them similarly vurnerable to the obesogenic effects of HFDs (Duca. 2014); similar differences, i.e. higher levels of clostridiaceae in obese individuals, have been observed in human studies, as well (Ferrer. 2013)
  • lactobacilli, above all those of the reuteri type, have recently been used in several studies for their anti-obesogenic (Million. 2013a, b), anti-autoimmune (Forsberg. 2013), anti-caries (Stensson. 2013), anti-helicobacter plyori (Francavilla. 2013), pro-vitamin-D (Jones. 2013), and a whole host of other beneficial effects; for other types of lacutobacilli researchers have observed that they exert similar anti-obesity effects that may be mediated by the intestinal productino of the anti-obesity isomer of CLA, i.e. trans‐10, cis‐12‐conjugated linoleic acid (Lee. 2007)
If we look at the previously listed metabolic effects, these changes in the make-up of the gut microbiome obviously correspond with the remarkable health improvements that occured in the whey-fed rodents.

High protein, low carb - What does it do?

Table 1: Plasma amino acid levels (mmol/L); blue bars to the right indicate sign. inter-group difference (McAllan. 2014)
Contrary to what you'd expect based on appetite increasing effects researchers ascribe to high protein diets (Weigle. 2005), the increase in protein-to-carbohydrate ratio (P/C) did not lead to measurable reductions in energy intake, but ....
"[...]the highest ratio [40% of the total energy intake from protein] reduced HFD-induced weight gain, fat mass and plasma triacylglycerol, non-esterified fatty acids, glucose and leptin levels, while it increased lean mass and oxygen consumption." (McAllan. 2014)
As the scientists point out, similar effects were observed on adipose mRNA expression, where the highest ratio of protein to carbohydrates reduced HFD-associated expression of UCP-2 (a protein that has the fat stores eat themselves up), the inflammatory marker TNF-alpha and CD68 a gylcoprotein that messes with LDL cholesterol.

On the other hand, the (really) high protein diet increased the diet-associated expression of the b3-adrenergic recepto (b3-AR), lipoprotein lipase, a water soluble enzyme that hydrolyzes triglycerides in lipoproteins, such as those found in chylomicrons and very low-density lipoproteins (VLDL), as well as the expression of insulin receptors and the glucose transporters GLUT4 - all of which should be old acquaintances of loyal SuppVersity readers.
Bottom line: The beneficial metabolic effects the addition of 40% whey protein isolate to a highly obesogenic baseline diet produced in the study at hand are remarkable and highly specific. "Specific", in that they don't occur with "an increase in protein intake".

Figure 3: Body weight development over the 21-week study period. The 40% whey diet clearly sticks out (McAllan. 2014)
In other words, the anti-obesogenic, anti-diabetic and anti-hyperlipidemic effects occurred not in response to "any type and amount of additional protein" that was added on top of what can be considered a model of a high fat version of the Western Diet. The previously discussed benefits were observed only, when this protein was whey protein and comprised a whopping 40% of the total energy intake of the rodents. The casein-based diets, as well as diets with lower amounts of whey protein isolate were ineffective, or - as you can see in Figure 3 - they "clustered together and away from the 40% WPI group", whose body weight - and this unquestionably quite remarkable - was hardly different from that of those 10 mice who were fed a regular, low fat diet for the whole 21-week study period.

References: 
  • Duca, Frank A., et al. "Replication of obesity and associated signaling pathways through transfer of microbiota from obese prone rat." Diabetes (2014): DB_131526.
  • Forsberg, Anna, et al. "Pre‐and post‐natal Lactobacillus reuteri supplementation decreases allergen responsiveness in infancy." Clinical & Experimental Allergy 43.4 (2013): 434-442.
  • Jones, Mitchell L., Christopher J. Martoni, and Satya Prakash. "Oral Supplementation With Probiotic L. reuteri NCIMB 30242 Increases Mean Circulating 25-Hydroxyvitamin D: A Post Hoc Analysis of a Randomized Controlled Trial." The Journal of Clinical Endocrinology & Metabolism 98.7 (2013): 2944-2951.
  • Lakritz, Jessica R., et al. "Beneficial bacteria stimulate host immune cells to counteract dietary and genetic predisposition to mammary cancer in mice." International Journal of Cancer (2014).
  • Lee, K., et al. "Antiobesity effect of trans‐10, cis‐12‐conjugated linoleic acid‐producing Lactobacillus plantarum PL62 on diet‐induced obese mice." Journal of applied microbiology 103.4 (2007): 1140-1146.
  • McAllan, Liam, et al. "Protein Quality and the Protein to Carbohydrate Ratio within a High Fat Diet Influences Energy Balance and the Gut Microbiota In C57BL/6J Mice." PLOS ONE 9.2 (2014): e88904.
  • Million, M., et al. "Correlation between body mass index and gut concentrations of Lactobacillus reuteri, Bifidobacterium animalis, Methanobrevibacter smithii and Escherichia coli." International Journal of Obesity (2013a).
  • Million, Matthieu, and Didier Raoult. "The role of the manipulation of the gut microbiota in obesity." Current infectious disease reports 15.1 (2013b): 25-30.
  • Stensson, Malin, et al. "Oral Administration of Lactobacillus reuteri during the First Year of Life Reduces Caries Prevalence in the Primary Dentition at 9 Years of Age." Caries research 48.2 (2013): 111-117.
  • Weigle, David S., et al. "A high-protein diet induces sustained reductions in appetite, ad libitum caloric intake, and body weight despite compensatory changes in diurnal plasma leptin and ghrelin concentrations." The American journal of clinical nutrition 82.1 (2005): 41-48.