A new paper has been published in the International Journal of Food Sciences and Nutrition titled “Dietary A1 beta-casein affects gastrointestinal transit time, dipeptidyl peptidase-4 activity, and inflammatory status relative to A2 b-casein in Wistar rats”
The key findings are:
1. A1 beta casein slows down transit of food through the digestive system relative to A2 beta-casein and this is an opioid effect.
2. A1 beta-casein induces a pro- inflammatory effect in the colon which is also an opioid effect.
3. A1 beta casein relative to A2 beta-casein causes up-regulation of the enzyme DPP4 in the small intestine and this is apparently a non-opioid effect.
4. In contrast to the A1 beta-casein, there is no evidence of opioid effects from the A2 beta-casein in relation to either food transit times or pro-inflammatory effects.
The above mentioned differences between the A1 and A2 beta-casein are all statistically significant, with considerable effect sizes, and using normal scientific criteria can be considered proven in relation to rats.
The research trials were undertaken by AgResearch (NZ) which is a Crown Research Institute (i.e. it is Government owned). The research was jointly funded by A2 Corporation and the New Zealand Government. The paper itself was a collaborative effort between AgResearch and A2 Corporation.
I have had direct involvement in this paper and I am a listed author along with three AgResearch scientists and A2 Corporation’s Chief Scientist. I was brought in to assist with the statistical analyses of the raw data provided by AgResearch, together with the consequent interpretation thereof, and to assist in writing the paper.
The research protocol
The research protocol was for the rats to be fed milk that contained beta-casein of the A1 or A2 type, with the level of the specific beta-casein unaltered from what was naturally in the milk of that type. Some 40% of the calories in the diet were from milk.
Half of the rats were dosed with naloxone to counteract any opioid effects. This was done because it is well understood that A1 beta-casein releases the peptide (protein fragment) beta casomorphin-7 (BCM7). Recent work from France, and published in the American Journal of Clinical Nutrition in 2013 (Boutrou et al 2013) has confirmed that BCM7 is released from ordinary milk at levels consistent with biological effects. Prior work published in the Journal of Food Chemistry (de Noni 2008) had confirmed that this comes either totally or almost totally from A1 beta-casein.
It is widely known that casomorphins attach to opioid receptors in the digestive system and thereby slow down the transit of food. This is actually the same effect that heavy doses of codeine have in humans, leading to constipation. Accordingly, the effect of A1 beta-casein slowing down the transit of food through the digestive system was hypothesised prior to the trial. However, it had not previously been specifically and directly shown that this effect could be observed from milk containing A1 beta-casein and so this needed to be confirmed.
The trial not only found such differences but it was evident that the differences were considerable and statistically significant. The differences disappeared when the rats were fed naloxone which further confirmed that the effect was indeed opioid in character. In contrast, the naloxone had no significant effect in rats fed the A2 diet indicating no measurable opioid effect.
The second effect was in the colon. The rats fed A1 showed an increase of about 65% in the inflammatory marker myeloperoxidase (MPO) and this was statistically significant (p=.02). This finding was essentially confirmatory, in that similar findings were reported in 2013 in the European Journal of Nutrition with mice (Ul Haq et al). MPO is produced within a particular type of white blood cells (neutrophils) and hence this is an immune response.
The finding of a statistically highly significant (p=.002) 40% increase in the enzyme dipeptidyl peptidase- 4 (DPP4) is particularly intriguing but also not surprising. This is because DPP4 is the only known enzyme capable of metabolising BCM7. Polish scientists (Wasilewska et al 2011) had earlier shown that in normal children high immuno-reactive BCM7 levels in the blood are associated with increased levels of DPP4, whereas in a sub group of children who had both particularly high BCM7 and also suffered from a respiratory acute life threatening event (ALTE), the DPP4 levels were particularly low. The implications of that earlier work which was published in the journal Neuropeptides are that DPP4 is normally up-regulated by BCM7 as a defensive mechanism to metabolise the BCM7, but that children who lack this response are at increased risk of sudden infant death syndrome through respiratory depression from BCM7.
The DPP4 results in the current trial are the first time that a causative and statistically significant effect in a trial situation has been established, and this complements very nicely the epidemiological association (also statistically significant) found in humans by Wasilewska et al.
However, the most exciting implications of the up regulation of DPP4 lie in a totally different field. This is because high levels of DPP4 affect glucose metabolism. Indeed there is a modern class of drugs for Type 2 diabetes known both as ‘gliptins’ and ‘DPP4 inhibitors’ which work specifically by reducing DPP4. And yet here we find that A1 beta-casein has increased DPP4 relative to A2 beta-casein. Confirmatory work in humans, particularly those at risk or currently suffering from Type 2 diabetes, is now of great importance.
The fact that no statistically significant opioid effects of A2 beta-casein were evident is somewhat of a side issue but also important.
The overall implications
The overall implications of this paper are profound.
1. In regard to digestive transit, it has indeed now been shown that A1 beta-casein does indeed slow down transit in rats, and it is well understood that the same opioid receptors exist in the human digestive system. Indeed we already knew that other opioids have this effect in humans. This finding of slower food transit provides strong support for the existing observational evidence that A1 is associated with digestive discomfort, bloating, and constipation relative to A2. Slower transit naturally means more opportunity for food fermentation.
2. The statistically significant increase in the inflammatory marker MPO has now been evidenced in two different trials with rats and mice. It is extremely unlikely that both could be by chance. So we can say with great confidence that at least in both rats and mice A1 beta-casein is pro inflammatory. This has important implications for both sub clinical inflammation in humans and potentially for inflammatory bowel syndrome, and once again complements observational evidence in humans.
3. The up-regulation of DPP4 in response to A1 beta-casein is both logical and exciting. Does the replacement of A1 beta-casein with A2 beta-casein provide a natural means of managing some of the baffling aspects of Type 2 diabetes?
The counter perspective
This paper will be seen as threatening by the mainstream dairy industry and so it is inevitable they will attack it. Their key argument is likely to be that the work was done with rodents rather than humans.
The counter to the counter perspective
Obviously, it would be better to do this work with humans but there are some very real constraints. To state the obvious, we cannot do a lot of this work with live humans. However, rodents do have the same fundamental digestive system as humans, they do have the same range of enzymes and they do have comparable immune systems. I am also aware that some relevant human data from a clinical trial will soon be forthcoming.
A real strength of this paper is that there is prior evidence in humans consistent with the effects shown here. There were no surprises. There was prior knowledge that opioids affect the transit of food in both rodents and humans. So this proves in a trial situation with rats, and specifically for A1 beta-casein, what had previously been observed in general for opioids across multiple species.
There was also prior evidence from human babies that DPP4 is up-regulated by the BCM7 released from A1 beta-casein. So here again what has been observed in humans (through a statistically significant association) has now been proven in a trial with rodents.
The evidence for pro-inflammatory effects in the colon is also confirmatory relative to prior findings in 2013 with mice that MPO levels increase in the presence of A1 beta-casein. These findings are also going to underpin some forthcoming clinical evidence relating to digestive effects in humans.
In summary, previously observed observational differences in human digestive function between A1 and A2 beta-casein have now been proven with rats under trial conditions.
Barnett MPG, McNabb WC, Roy NC, Woodford KB, Clarke AJ. 2014. Effects of dietary A1 and A2 beta-casein on gastrointestinal transit time, DPP-4 activity and inflammatory status, in Wistar rats. International Journal of Food Sciences and Nutrition, Early Online: 1–8. Informa UK Ltd. DOI: 10.3109/09637486.2014.898260
We compared the gastrointestinal effects of milk-based diets in which the β-casein component was either the A1 or A2 type in male Wistar rats fed the experimental diets for 36 or 84 h. Gastrointestinal transit time was significantly greater in the A1 group, as measured by titanium dioxide recovery in the last 24 h of feeding. Co-administration of naloxone decreased gastrointestinal transit time in the A1 diet group but not in the A2 diet group. Colonic myeloperoxidase and jejunal dipeptidyl peptidase (DPP)-4 activities were greater in the A1 group than in the A2 group. Naloxone attenuated the increase in myeloperoxidase activity but not that in DPP-4 activity in the A1 group. Naloxone did not affect myeloperoxidase activity or DPP-4 activity in the A2 group. These results confirm that A1 β-casein consumption has direct effects on gastrointestinal function via opioid-dependent (gastrointestinal transit and myeloperoxidase activity) and opioid-independent (DPP-4 activity) pathways.
See the A1 and A2 milk category on this website for further posts. (https://keithwoodford.wordpress.com/category/a1-and-a2-milk)
The references specifically referred to in this post are
1. Boutrou R, Gaudichon C, Dupont D, Jardin J, Airinei G, Marsset-Baglieri A, et al. Sequential release of milk protein-derived bioactive peptides in the jejunum in healthy humans. Am J Clin Nutr 97: 1314-23.
2. De Noni I. 2008. Release of b-casomorphins 5 and 7 during simulated gastro-intestinal digestion of bovine beta-casein variants and milk-based infant formulas. Food Chemistry 110: 897-903.
3. Ul Haq MR, Kapila R, Sharma R, Saliganti V, Kapila S. 2013. Comparative evaluation of cow β-casein variants (A1/A2) consumption on Th2-mediated inflammatory response in mouse gut. Eur J Nutr; e-pub ahead of print 29 October 2013; doi: 10.1007/s00394-013-0606-7
4. Wasilewska J, Sienkiewicz-Szlapka E, Kuzbida E, Jarmolowska B, Kaczmarski M, Kostyra E. 2011. The exogenous opioid peptides and DPPIV serum activity in infants with apnoea expressed as apparent life threatening events (ALTE). Neuropeptides 2011; 45: 189-95.
For a more detailed list of references go to:
Keith Woodford consults in a professional capacity for A2 Corporation and other agri-food companies as an independent adviser. However he does not represent any of these companies and he retains total independence in everything that he writes.