Take home message

• The whey protein beta-lactoglobulin (BLG) has two sides: it causes milk allergy, but it can also calm the overstimulated immune system.
• What face BLG shows depends on the degree of heating of milk or whey, but also whether substances, such as fatty acids, are bound to the whey protein. Especially on the inside of the vase-shaped molecule.

Food allergy

Food allergy, including milk allergy is based on an immune system, which does not respond properly, and milk is not recognised as food. Instead of a food protein, the immune system only ‘sees’ antigens, bits of protein that it has learned to react to with a violent physical reaction, drop in body temperature and even anaphylactic shock. It can also manifest as itchy skin, vomiting, or asthma.

Cows, barns and raw milk

You can have a whole philosophy on whether to drink milk, eat meat, but Western use of cow’s milk is an extension of drinking our own mother’s milk. Both unpocessed cow’s milk and the cows’ barn environment appear to play a crucial role in calming an overreactive immune system, which shows up as food allergy, milk allergy or asthma.

Research from the last 25 years has made it clear again and again, that:

• …consumption of fresh, unprocessed cow’s milk protects against the development of the aforementioned immunological problems;
• …there is something in the barn environment, the barn dust, but also the barn air, that like the cow’s milk itself, gives protection.

In a recent study, dust was collected in cattle pens and adhering chemicals were extracted. The collection site chosen was the cow barn of the American Amish population. These farmers are very attached to their cows and have about the lowest incidence of asthma and allergies in any Western population. Dust from the Amish cow barn was compared with dust in barns with sheep. Researchers also took samples in European cow barns to assess whether they resembled the Amish barns (Dos Santos et al., 2023). Earlier research had shown that the cow barn, but not the sheep barn, gives a reduced asthma incidence in farm children. Even mice that were ‘infected’ with cow barn dust were not found to develop asthma, mice with sheep barn dust, however, did. So, the question was, what else/more/different does a cow give that a sheep cannot?

Two chemicals from the lipocalin family were found 500 to 700 times higher in cow pens. One substance is excreted through cow skin (Odorant Binding Protein = OBP), the other is called ‘Allergen Bos D2’ (Bos taurus = cow). This is not so remarkable, as they are specific substances produced by cows, not sheep. Still, the substances were interesting because cow dust caused no or less asthma.

Other research pointed to a 3^rd lipocalin, namely the beta-lactoglobulin (BLG), which is airborne both through cow’s milk and urine. We reported on this earlier (keyword: cow barn pill).

Lipocalins

Lipocalins function in our body as transporters of numerous substances, such as iron, fatty acids and vitamins. At the cells of the intestinal wall, they dock and then release their contents into the cell. They are a family of small, three-dimensional proteins, which are somewhat ‘vase-shaped’, having a top and bottom, but most importantly, an inside and an outside. In the lipocalin family, you come across many allergens, such as cat hair, birch pollen, but also beta-lactoglobulin (BLG), the main whey protein, present in cow’s milk but not in human milk. Due to their corresponding shape and structure, cross-allergies occur, i.e. you can have an allergic reaction to multiple, similar lipocalins, if your immune system is at least sensitised in the first years of life.

Holo- and apo-lipocalins is of great importance

Crucial to whether a lipocalin becomes an allergy is whether the lipocalin vase is ‘empty’ or ‘filled’. Lipocalins can be loaded with fatty acids: stearic, oleic and linolenic acids (C18 fatty acids saturated and unsaturated). They are hydrophobic substances, substances that do not mix with water, and repel water. They are transported inside the vase in an aqueous environment. Dos Santos et al (2023) found several transport proteins on traditional dairy farms, which can transport non-water-soluble substances from plants, but also fungi, as well as minerals such as iron or zinc. The packets that are transported are called ligands. They are usually stored inside the ‘vase’-shaped holo-lipocalins.

Stable holo-lipocalins are the ligand-loaded lipocalins. They deliver their content to immune receptors, which, all in all, leads to the suppression of the allergic reaction, calming an overstimulated immune system (Roth-Walter et al., 2020). In contrast, empty apo-lipocalins, as well as heat-damaged lipocalins (after pasteurisation of milk or whey), can trigger an allergic response. They are unable to quieten immune cells. In both mice and young humans, consumption of raw milk does not appear to lead to a negative immunological response, but heated cow milk or also shop milk does (Abbring et al., 2019). Heating a high quality of raw milk (biodynamic or Demeter milk) also gives rise to an asthmatic response in mice (Abbring et al., 2017), not the raw biodynamic milk. All in all, then, protection against asthma and allergies revolves around the whey fraction of milk, dust in the barn or barn air enriched with cow urine, which also contains large amounts of BLG. The important lipocalin in milk whey is the BLG, if undamaged and (re)charged, i.e. raw.

The two faces of BLG

One side of BLG is, that as a transport molecule, it passes the acid stomach undigested to then reach the intestinal mucosa of the small intestine. This BLG docks to the mucosa to calm the immune system. How this is possible is related, on the one hand, to the ‘native’/ unchanged protein (holo-), which is loaded with the ligands (iron, fatty acids, retinol). The holo-BLG is healing.

However, BLG also has another side, it is seen as an important protein, which can cause cow’s milk allergy. This is undesirable and should be avoided. However, this BLG is different, empty, an alo-BLG, or a heat-altered whey protein, which has undergone structural change and where new epitopes are formed, as some parts of the BLG turn outwards. Heating temperatures up to about 60^o C are probably still largely reversible for BLG, i.e. there is shape recovery and return to the native state when the temperature drops again. Higher temperatures increasingly lead to irreversible change of the BLG molecule. This is called ‘unfolding’, i.e. the vase is turned inside out. The damage from heating depends on the combination of absolute temperature and time duration. As a side effect, numerous new compounds are formed, such as between denatured BLG and kappa-casein, but also between BLG and the sugars in milk. This BLG is not something you want to have, if your immune system is sensitized in early age.

How is BLG finally digested?

BLG in unheated milk forms a compact and robust protein, which resists well against the low pH of the stomach and degradation by the enzyme trypsin. This contrasts with the cheese proteins, where the important beta-casein is seized directly in metabolism and transforms into 100s of peptides. Rather than a dietary protein (like casein), the function of the BLG is therefore a transport protein, able to deliver numerous substances into the small intestine via the diet, however, if intact. Holo-BLG as a protective package deliverer of substances, which would otherwise have difficulty reaching the small intestine. Once the contents of the BLG are delivered into the first part of the small intestine, eventually the BLG also disintegrates and is broken down into peptides.

Literature

• Abbring, S., Verheijden, K. A., Diks, M. A., Leusink-Muis, A., Hols, G., Baars, T., … & van Esch, B. C. (2017). Raw cow’s milk prevents the development of airway inflammation in a murine house dust mite-induced asthma model. Frontiers in Immunology, 8, 1045.
• Abbring, S., Kusche, D., Roos, T. C., Diks, M. A., Hols, G., Garssen, J., … & van Esch, B. C. (2019). Milk processing increases the allergenicity of cow’s milk—Preclinical evidence supported by a human proof‐of‐concept provocation pilot. Clinical & Experimental Allergy, 49(7), 1013-1025.
• Barbiroli, A., Bonomi, F., Ferranti, P., Fessas, D., Nasi, A., Rasmussen, P., & Iametti, S. (2011). Bound fatty acids modulate the sensitivity of bovine β-lactoglobulin to chemical and physical denaturation. Journal of agricultural and food chemistry, 59(10), 5729-5737.
• Barbiroli, A., Iametti, S., & Bonomi, F. (2022). Beta-lactoglobulin as a model food protein: How to promote, prevent, and exploit its unfolding processes. Molecules, 27(3), 1131.
• Dos Santos, M. M., Pivniouk, V., Rankl, B., Walker, A., Pagani, G., Hertkorn, N., … & Vercelli, D. (2023). Asthma-protective agents in dust from traditional farm environments. Journal of Allergy and Clinical Immunology, 152(3), 610-621
• Roth-Walter, F., Afify, S. M., Pacios, L. F., Blokhuis, B. R., Redegeld, F., Regner, A., … & Hufnagl, K. (2020). Cow milk protein beta-lactoglobulin confers resilience against allergy by targeting complexed iron into immune cells. Journal of Allergy and Clinical Immunology.