1. What are Cytokines

Cytokines are soluble proteins or glycoproteins produced by cells. Cytokines act as chemical communicators between cells; they are also called messenger proteins.

Most are secreted by different cells and move in the surrounding of the cells and roam freely in the blood stream and tissue but some can be expressed on the cell membrane and others are held in reservoirs in the extracellular matrix (see glossary). Cytokines don’t enter the cell, but deliver the information via special receptors on the cell surface. They bind to specific receptors on the surface of target cells, which are coupled to intracellular signal transduction and second messenger pathways.

After docking to their receptors, special biological reactions are triggered prompting the cell to proliferate, to produce other cytokines (messengers) or antibodies etc.

Figure: The cell receives the information from cytokine-activated receptor to produce another messenger

Individual cells produce and receive different cytokines resulting in a busy information exchange.

Figure: Cells have receptors for different cytokines and are also able to produce different cytokines.

Cytokine is a collective term, which includes Interleukins* (cytokines produced by Leukocytes like Interleukin-1 to Interleukin- 33), Growth factors (like TGF-beta, VEGF and others) and Interferons (alpha, beta and gamma).

(* inter=between; leukin=from leukocyte)

During an inflammation, a huge amount of cytokines is produced in the affected tissue – much more than being attached to the receptors.

Up to now, about 120 different cytokines have been discovered with very different tasks. Two groups of them are involved in inflammatory processes:

1. Pro-inflammatory cytokines: They help to build up an inflammatory process. Main cytokines in this group are called interleukin-1 (IL-1), tumour necrosis factor (TNF) or interleukin-6 (IL-6).

2. Anti-inflammatory cytokines: for instance interleukin-4 (IL-4), interleukin 10 (IL-10), vascular endothelial factor (VEGF) and transforming growth factor-beta (TGF-beta) deliver information which help to stop the inflammation and initiate healing processes such as vessel and tissue-building and scarring.

What happens with the excess of cytokines?

In a healthy organism the concentration of cytokines is very low and some cytokines are not detectable with our determination methods (ELISA, etc.). During a dysfunction or during an inflammation, cells produce a huge amount of different cytokines.

This overproduction could be explained to ascertain that the information arrives at the right time and the right place.

But what happens with the high amounts of overproduced cytokines? This information potential is a risk for the organism to bring information to the wrong place and time. There exists a sneaky mechanism to dispose these unnecessary cytokines – and for this sneaky mechanism “enzymes” are necessary.

This important process will be described in the following chapter: “The connection between cytokines, inflammation and enzyme therapy” and is one of the most important chapters to understand the effect of enzyme therapy.

2. What is an acute Inflammation?

Aulus Cornelius Celsus first described this process 2000 years ago as: reddening, swelling, heat and pain which end up with healing and scarring.

If you have ever had an insect bite, a sprained ankle, a sore throat, or bad sunburn, you know what an acute inflammation is. Inflammation is the answer, or a bunch of answers to injuries and/or infections caused by foreign substances or by trauma– No matter, if the injuries or the infection is big or small.

Overall, the reaction of our body to an inflammation is a defined process, which is separated into the following steps:

• Occurrence of the inflammation

1. Activated blood platelets (also called Thrombocytes) cluster and close the blood vessels to prevent further loss of blood. This happens in wounds as well as internal injuries.

2. Only clustered thrombocytes secrete Cytokines to attract immune-competent cells of the blood like Granulocytes, Monocytes and Dendritic cells.

3. Some of these cytokines instruct Endothelial cells (cells of the vascular wall) to open up the usually tight connection between the cells. Liquid flows out of the vessel into the surrounding tissue, producing a swelling (oedema). Finally, endothelial cells are activated to growth. [This is necessary to build new blood vessel in the healing wound to secure supply with nutrients and oxygen once the wound is healed].

Figure: Platelets (thrombocytes) close the injured blood vessel with a “plug” by clustering; thereafter they deliver cytokines (messengers), which attract immune-competent cells.

4. Scavenger cells (Macrophages, Dendritic cells) start to clean up – they eat (phagocyte) the foreign substances or organisms (viruses, bacteria, fungi) and offer digested parts to a special kind of blood cells, the T-cells. This procedure is called “Antigen presentation”. These T-cells become activated and instruct B-cells (also immune cell of the blood) to produce antibodies against these foreign materials.

Figure: Scavenger cells phagocyte bacteria and offer these digested proteins to T-cells. So activated T-cells inform the B-cells to produce antibodies.

5. The new antibodies attaches to the invading material like bacteria in the wound. Bacteria attached by antibodies scavenger cells (Macrophages and Dendritic cells) recognize, phagocyte and kill the bacteria easer and quicker. These phagocyte cells also remove the remnants of the dead body cells destroyed by the injury.

The next phase of this process is as important; the inflammation has to come to an end!!

• Ending of the inflammation

1. A cytokine called VEGF (Vascular endothelial growth factor) activates healthy endothelial cells (cells of the vascular walls, see glossary) to proliferate.

2. These new cells form tubes and are capable to supply the regenerated tissue with blood (which brings food and oxygen). The swelling disappears.

3. Immune cells (T-cells, Macrophages, Dendritic cells) switch to production of a different kind of cytokines which inform their surrounding that the most challenging part is over, cleaning up is in progress and there is no need for further pro-inflammatory activity.

4. A few cells are busy with building huge amounts of extra cellular matrix – padding material between the cells- so that the wound closes.

Figure: the cells are surrounded by extracellular matrix, proteins with the function of an extracellular skeleton.

The duration of an inflammatory process depends on extent and kind of injury and/or infection.

3. What is a chronic inflammation?

Why are we getting chronic diseases?

The answer is astonishingly simple: In certain situations an acute inflammation becomes chronic.

To put it bluntly: Imagine a healing wound that is opened again and again. How will the cells react? Some will produce pro-inflammatory cytokines; others will produce anti-inflammatory cytokines, because everything was just beginning to heal – before the wound was opened again. If this procedure is repeated several times, the whole process will become deregulated – the wound will not heal, the inflammation becomes chronic.

Figure 5: Normally acute inflammation will lead to a complete cure (A), while repetitive boost of an inflammation becomes chronic – with all consequences

Reason for Chronification of acute inflammations

Bacteria:

You can imagine how e.g. a bacterial infection can lead to a chronic inflammation: First, the infection will be battled with internal help (acute inflammation) and external help (antibiotics) and everything is eased out. If a few bacteria stay hidden away and start to multiply, the whole process will repeat and repeat and repeat, until it gains its own momentum.

Chronic urinary tract infection, chronic otitis media and chronic inflammation of Para nasal sinuses (chronic sinusitis) or periodontitis are examples for such procedures.

Virus:

Viral infections with Hepatitis or HIV induce the chronic inflammatory processes. There is no medication that can kill a Virus if the immune system cannot get rid of it and the body lives with constant infection.

Chronic intoxication by e.g. alcohol, asbestos, siliceous:

Consumption of alcohol kills some liver cells (hepatocytes). Some alcohol now and then will not harm the liver. An inflammation may appear locally and vanish fast. Daily consumption of large quantities for a long time causes inflammatory-processes in short intervals – setup and degradation interlock and inflammation becomes chronic. We then call this a chronic inflammation of the liver – leading to a hardening of the liver (alcoholic cirrhosis), which often leads to a liver tumour. Inhalation of asbestos or siliceous over time induces chronic lung inflammations with a high incidence to develop lung tumours.

Autoimmune diseases:

When an organism fails to recognize some of its own constituent as part of itself it results in an attack against its own cells and tissues. Autoimmune diseases therefore are chronic. Prominent examples include:

• Ankylosing spondilitis a chronic, painful, progressive inflammatory arthritis primarily affecting spine and sacroiliac joints, causing eventual fusion of the spine.

• Diabetes mellitus (type 1) is the result of an autoimmune attack on the islet cells of the pancreas

• Multiple sclerosis is a disorder of the central nervous system (brain and spinal cord) characterized by decreased nerve function due to myelin loss and secondary axonal damage

• Rheumatoid arthritis is an autoimmune disorder that triggers the body’s immune system to attack the bone joints

• Sjögren’s syndrome is an autoimmune disorder in which immune cells attack and destroy the exocrine glands that produce tears and saliva

During an Inflammation we find an overproduction of cytokines, temporarily in acute inflammation and continuous overproduction in chronic inflammation.

1. Internal endogenous, body-produced enzymes (proteases):

During an inflammation, different enzymes are produced in the area of the event to support healing processes, to destroy dead cells, to activate cell proliferation etc. Normally there are locally more enzymes produced than required.

Some examples of endogenous produced enzymes are Elastase, Collagenase, and Metalloproteinase (also called MMP or metalloendopeptidases). They play an important role in embryonic development, wound healing, inflammation, chronic inflammation and tumor metastasis – generally in processes including matrix degradation.

Consequently, after an inflammation they have to be neutralized so that they can’t do any harm to other parts of the body; the same is true for cytokines: locally overproduction is necessary, but systemically they will do harm.

2. Exogenous enzymes (proteases):

There are different exogenous enzymes from our diet, from honey, fruits, vegetables, uncooked meat, and buyable Enzyme preparation reach our blood.

2. How does our body associate with these enzymes?

You remember:

• During inflammation enzymes are produced.

• By nutrition enzymes are absorbed and reach the blood.

• By Enzyme-Therapy, also enzymes are absorbed and reach the blood.

3. The function of antiproteases

The blood contains proteins that catch enzymes – these molecules are called antiproteases.

The two of the most important antiproteases are:

• alpha1antitrypsin: which binds and destroys enzymes produced in the body: e.g. Elastase, trypsin, chymotrypsin

• alpha2macroglobulin: has the function to bind and dispose both external enzymes (absorbed nutritionally, e.g. Bromelain, Papain, Subtilisin, Serrapeptase, Ficin or Nattokinase) and internal enzymes (e.g. Trypsin, Chymotrypsin, Elastase, Thrombin etc.).

The important differences between these 2 antiproteases lay in their capacity to bind enzymes:

• Alpha1antitrypsin binds enzymes produced in the body only.

• After binding to alpha1antitrypsin, the enzyme is inactivated

• Alpha2macroglobulin binds also enzymes, which reach the blood after ingestion.

• After binding to alpha2macroglobulin, the enzyme stays active until destruction in the cell.

• Alpha2macroglobulin binds cytokines when activated by enzymes

• Alpha1antitrypsin don’t bind cytokines, its combination with trypsin or chymotrypsin is disturbed in the cell after binding to LRP

The consequence of these findings:

We need more exogenous enzymes during inflammation to activate alpha-2-macroglobulin for disposal of overproduced cytokines.

In our diet, we absorb enzymes, but not enough, so we have to use other possibilities like enzyme therapy.

Conclusion: activated a2Macroglobulin but not other antiproteases like a2antitrypsin, has the capacity to bind cytokines. This binding is fatal for the cytokines, because in a short period of time, the complex a2M+enzyme+cytokine is removed from blood and metabolized in endothelial cells.

Enzyme therapy supports the disposal of pro- and anti-inflammatory cytokines during overproduction.

There is a further function of enzymes that has not been well studied to date:

When the enzyme, for example Bromelain is absorbed, it reaches the blood in an active form. Bromelain binds to a2Macroglobulin and Bromelain stay active until the moment where a2M binds to the LRP-receptor on the endothelial cell. During this time – arrival in blood and disposal by LRP – enzymes are active and it could be that some changes, which are observed in enzyme therapy, are based on this circumstances:

1. Reduction of some adhesion proteins on the cell surface after enzyme therapy

2. Reduction of circulating antibodies in blood after enzyme therapy.

Summary:

Proteolytic enzymes (Proteases) are important molecules produced continuously by our body. During embryogenesis, healing processes, tumor metastasis and rebuilding processes enzymes are necessary. On the other hand, there are also very sophisticated mechanisms to protect the organism from too much, unwanted enzymes. This function is covered by antiproteases. Only one of these antiproteases, called alpha2macroglobulin (a2M) has the function to bind not only the body’s own proteases but also enzymes absorbed from the gut.

Binding an enzyme, alpha2macroglobulin (a2M) then is activated, which means that a2M binds also cytokines. No other antiproteases in human blood have this function.

Because the complex of a2M+enzyme+cytokine is quickly removed and (metabolized) destroyed by cells in the blood vessels, this mechanism is a simple and sophisticated way to remove overproduced cytokines from the organism.

The enzyme therapy supports this mechanism.