O-GlcNAcylation and Its Role in Diseases

Figure 1.  Overview of the Hexosamine Biosynthetic Pathway (HBP) and O-GlcNAcylation.
The HBP integrates four metabolism pathways, including
carbohydrate (glucose), amino acid (glutamine), lipid (Acetyl-CoA) and nucleotide (UTP).

O-linked-N-acetylglucosaminylation (O-GlcNAcylation) is a type of glycosylation that occurs when a monosaccharide, O-GlcNAc, is added onto serine or threonine residues of nuclear or cytoplasmic proteins by O-GlcNAc transferase (OGT) and which can be reversibly removed by O-GlcNAcase (OGA). 

Dysregulation of O-linked β-D-N-acetylglucosamine (O-GlcNAc) has been implicated in many pathologies including Alzheimer's disease (AD), cancer, diabetes, and neurodegenerative disorders.^[1]

Figure 2: Dynamic Competition between glycosylation and phosphorylation of proteins.
A: Competition between OGT and kinase for the serine or threonine functional group of a protein.
B: Adjacent-site occupancy where O-GlcNAc and O-phosphatase occur next to each other and
can influence the turnover or function of proteins reciprocally.
The G circle represents an N-acetylglucosamine group, and
the P circle represents a phosphate group. Figure adapted from Hart.^[3]

OGT vs OGA

Since its discovery in the early 1980s, O-GlcNAc addition (O-GlcNAcylation) to serine/threonine residues has been found to be a key post-translational modification of proteins in the nucleus, cytosol and mitochondria. 

As a highly dynamic process, O-GlcNAc rapidly cycles onto serine/threonine residues of target proteins in a fashion analogous to phosphorylation (see Figure 2). 

While there are roughly 500 kinases and 150 phosphatases that regulate protein phosphorylation in humans, there are only 2 enzymes that regulate the cycling of O-GlcNAc:

• O-GlcNAc transferase (OGT) 
• Catalyzes the addition of O-GlcNAc to serine/threonine residues
• OGT utilizes UDP-GlcNAc as the donor sugar for sugar transfer.
• O-GlcNAcase (OGA) 
• Catalyzes the removal of O-GlcNAc

The chemical reaction of OGT can be written as:

1. UDP-N-acetyl-D-glucosamine + [protein]-L-serine → UDP + [protein]-3-O-(N-acetyl-D-glucosaminyl)-L-serine
2. UDP-N-acetyl-D-glucosamine + [protein]-L-threonine → UDP + [protein]-3-O-(N-acetyl-D-glucosaminyl)-L-threonine

Using the below pharmacological compounds, you can control the level of protein O-GlcNAcylation:

• To decrease it, by administering DON and Azaserine
• To increase it by using OGA blocker
• Such as Glucosamine, PUGNAc, NButGT
• NButGT
  • 1,2-dideoxy-2′-propyl-α-d-glucopyranoso-[2,1-D]-Δ2′-thiazoline 
• Elevated O-GlcNAc has been associated with diabetes.

NButGT (more-specific) vs PUGNAc (less-specific)

The regulation of OGT is directly involved in diabetes. OGT and O-GlcNAc-modified protein levels are increased in the pancreatic islets of diabetic rats.^[6] Overexpression of OGT in liver, muscle and fat tissues causes insulin resistance.^[7,8] 

Using OGA inhibitors (i.e., PUGNAc and NButGT) can acutely increases the levels of O-GlcNAcylation.  Scientists have noticed that even both inhibitors can induce elevated O-GlcNAcylation, there is a discrepancy found:

• Treatment with PUGNAc leads to insulin resistance
• Treatment with NButGT doesn't lead to insulin resistance

This discrepancy might be resulted from the off-target effects (e.g., on lysosomal hexosaminidases) of the less-specific PUGNAc, suggesting the necessity of choosing more specific and selective inhibitors to explore the roles of OGA. 

Caution should also be made in terms of inhibitor usage, since effects of the inhibitors are, generally, in a dose- and time-dependent manner, which may cause significant differences in protein O-GlcNAcylation levels in a tissue-specific manner and thus the related diabetic phenotypes.

O-GlcNAcylation–Based Treatments as Potential Interventions for AD

O-GlcNAcylation is notably decreased in Alzheimer’s disease (AD) brain. Necroptosis is activated in AD brain and is positively correlated with neuroinflammation and tau pathology. 

In one study,^[5] scientists found that NButGT—a specific inhibitor of OGA, reduces Aβ production by lowering γ-secretase activity. Moreover, NButGT 

• Reduces Aβ production 
• By lowering γ-secretase activity
• Attenuates
• Accumulation of Aβ
• Neuroinflammation
• Memory impairment 

in the 5XFAD mice. 

This is the first study to show the relationship between Aβ generation and O-GlcNAcylation in vivo. These results suggest that O-GlcNAcylation may be a suitable therapeutic target for the treatment of AD.

O-GlcNAcylation Can Regulate Many Hallmarks of Cancer

Elevated HBP and O-GlcNAcylation has been reported in nearly all cancers examined and can regulate many “hallmarks of cancer”, including growth, survival, metabolism, angiogenesis, and metastasis.^[12]

Up to this point, nearly all evidence suggests that the HBP helps fuel cancer cell metabolism, growth, survival, and spread. Further research should elucidate whether the HBP plays a role in cancer initiation and maintenance, heterogeneity, and regulation of the tumor microenvironment, including immune surveillance.^[13]

References

1. Hart, Gerald W.; Slawson, Chad; Ramirez-Correa, Genaro; Lagerlof, Olof (2011-06-07). "Cross Talk Between O-GlcNAcylation and Phosphorylation: Roles in Signaling, Transcription, and Chronic Disease". Annual Review of Biochemistry. 80: 825–858.
2. Haltiwanger, RS; Holt, GD; Hart, GW (1990-02-15). "Enzymatic Addition of O-GlcNAc to Nuclear and Cytoplasmic Proteins. Identification of a Uridine diphospho-N-acetylglucosamine:peptide beta-N-acetylglucosaminyltransferase". Journal of Biological Chemistry. 265 (5): 2563–8.
3. Hart GW, Housley MP, Slawson C (April 2007). "Cycling of O-linked beta-N-acetylglucosamine on nucleocytoplasmic proteins". Nature. 446 (7139): 1017–22.
4. Macauley, MS; Bubb, AK; Martinez-Fleites, C; Davies, GJ; Vocadlo, DJ (2008-12-12). "Elevation of Global O-GlcNAc Levels in 3T3-L1 Adipocytes by Selective Inhibition of O-GlcNAcase Does Not Induce Insulin Resistance". The Journal of Biological Chemistry. 283 (50): 34687–95.
5. O-GLCNACYLATION AMELIORATES THE PATHOLOGICAL MANIFESTATIONS OF ALZHEIMER’S DISEASE BY INHIBITING NECROPTOSIS
6. Akimoto Y, Hart GW, Wells L, et al. Elevation of the post-translational modification of proteins by O-linked N-acetylglucosamine leads to deterioration of the glucose-stimulated insulin secretion in the pancreas of diabetic Goto-Kakizaki rats. Glycobiology. 2007;17:127–140.
7. McClain DA, Lubas WA, Cooksey RC, et al. Altered glycan-dependent signaling induces insulin resistance and hyper-leptinemia. Proc Natl Acad Sci USA. 2002;99:10695–10699.
8. Yang X, Ongusaha P, Miles P, et al. Phosphoinositide signaling links O-GlcNAc transferase to insulin resistance. Nature. 2008;451:964–970.
9. Macauley MS, Bubb AK, Martinez-Fleites C, et al. Elevation of global O-GlcNAc levels in 3T3-L1 adipocytes by selective inhibition of O-GlcNAcase does not induce insulin resistance. J Biol Chem. 2008;283:34687–34695.
10. Macauley MS, He Y, Gloster TM, et al. Inhibition of O-GlcNAcase using a potent and cell-permeable inhibitor does not induce insulin resistance in 3T3-L1 adipocytes. Chem Biol. 2010;17:937–948.
11. Macauley MS, Shan X, Yuzwa SA, et al. Elevation of global O-GlcNAc in rodents using a selective O-GlcNAcase inhibitor does not cause insulin resistance or perturb glucohomeostasis. Chem Biol. 2010;17:949–958.
12. Ferrer CM, Sodi VL, Reginato MJ. O-GlcNAcylation in cancer biology: linking metabolism and signaling. J Mol Biol. 2016;428:3282–94.
13. Fueling the fire: emerging role of the hexosamine biosynthetic pathway in cancer

Eat More Phytochemically Rich “Hyperfoods” to Fight COVID-19

In [1], scientists use a machine learning approach to identify potential bioactive anti-COVID-19 molecules in foods.

Top-Ranked Hyperfoods to Fight COVID-19

Using a network-based machine learning method, scientists have shown that certain plant-based foods such as:

• Berries
• Cruciferous vegetables
• Apples
• Citrus fruits
• Onions
• Garlic
• Beans

are most enriched in terms of the diversity and relative abundance of bioactive molecules targeting the SARS-CoV-2-human interactome.

For instance, blackcurrant is the top-ranked food source, which has the below phytochemical profile:

• alpha-tocopherol
• arachidonic acid
• catechol
• cinnamaldehyde
• coumarin
• kaempferol 
• quercetin 
• quercetin 3-rutinoside 
• myricetin 
• n-butanoate 
• daidzein 
• genistein 
• procyanidin 
• selenomethionine 
• trans-resveratrol 
• trehalose 

Figure 1.  The contained profiles of compounds within specific foods,
with predicted effectiveness in targeting SARS-CoV-2-host interactome networks (source: [1]). 

The Whole is Greater than the Sum of its Parts

Scientists surmises that it's more plausible that consumption of whole foods, with their associated phytochemicals en masse may provide greater health benefits, due to molecular additive or synergistic effects. It therefore follows that the antiviral properties of a given food will be governed by two key factors:

1. The additive, antagonistic and synergistic actions of their individual components
2. The way in which these simultaneously modulate different intracellular pathways involved in SARS-CoV-2 pathogenesis

Based on these assumptions, they have constructed a food map with the theoretical anti-COVID-19 capacity of each ingredient ranked according to an “enrichment score” derived from the diversity and relative levels of candidate compounds with antiviral properties (Figure 1).

Table 1. Top-ranked phytochemically rich food sources (i.e., Hyperfoods)

Common Name	Enrichment Score
Blackcurrant'	28.2
Broccoli|Brussel sprouts|Cabbage|Cauliflower|Common cabbage|Kohlrabi|Savoy cabbage|White cabbage'	25.7
Vaccinium (Blueberry, Cranberry, Huckleberry)'	25.5
Common bean|Green bean|Yellow wax bean'	24.6
Apple'	24.6
Strawberry'	24.2
Rubus (Blackberry, Raspberry)'	23.5
Sweet orange'	22.5
Carob'	22.3
Garden onion'	22.1
Lettuce|Romaine lettuce'	21.0
Garlic|Soft-necked garlic'	20.9
European plum'	20.8
Lemon'	20.7
Parsley'	20.6
Common grape'	20.5
Pear'	20.1
Almond'	19.7
Prunus (Cherry, Plum)'	19.4
Sweet cherry'	19.3
Apricot'	18.8
Carrot|Wild carrot'	18.6
Pistachio'	18.6
Sweet potato'	18.3
Soy bean'	18.1
American cranberry'	18.1
Mango'	17.9
Dock'	17.9
Cherry tomato|Garden tomato|Garden tomato (var.)'	17.8
Broad bean'	17.8
Peach'	17.6
Banana'	17.3
Chinese cabbage|Turnip'	17.3
Green bell pepper|Italian sweet red pepper|Orange bell pepper|Pepper (C. annuum)|Pepper (C. frutescens)|Red bell pepper|Yellow bell pepper'	17.2
Chicory'	17.1
Lentils'	17.0
Black elderberry'	16.9
Dill'	16.9
Wild celery'	16.6
Pitanga'	16.5
Sour cherry'	16.3
Potato'	16.3
Pineapple'	16.3
Capers'	16.2
Spinach'	16.2
Papaya'	16.2
Horseradish tree'	16.2
Common walnut'	16.1
Cucumber'	16.0
Date'	15.8
Sweet bay'	15.6
Rosemary'	15.6
Rape|Swede'	15.5
Mung bean'	15.4
Common beet|Red beetroot'	15.4
New Zealand spinach'	15.2
Wild leek'	15.1
Chinese mustard'	14.9
Chives'	14.9
Common hazelnut'	14.9
Gooseberry'	14.9
Olive'	14.6
Alfalfa'	14.6
Muskmelon'	14.5
Chickpea'	14.5
Pecan nut'	14.3
Globe artichoke'	13.9
Common thyme'	13.9
Taro'	13.7
Asparagus'	13.6
Ginger'	13.5
Black-eyed pea|Cowpea|Yardlong bean'	13.4
Black mulberry'	13.3
Peppermint'	12.8
Garden cress'	12.6
Sacred lotus'	12.6
Rocket salad (ssp.)'	12.5
Allium (Onion)'	12.5
Barley'	12.4
Fig'	12.4
Avocado'	12.3
Garden rhubarb'	12.1
Fennel'	12.1
Kumquat'	12.1
Pomegranate'	12.1
Towel gourd'	12.1
Common pea'	12.0
Malabar spinach'	12.0
Mandarin orange (Clementine, Tangerine)'	12.0
Watermelon'	11.9
Coriander'	11.8
Eggplant'	11.7
Radish'	11.6
Turmeric'	11.5
Purslane'	11.4
Common buckwheat'	11.3
Okra'	11.3
Parsnip'	11.3
Quince'	11.2
Endive'	11.2
Pummelo'	11.2
Saffron'	10.8
Sweet marjoram'	10.8
Cucurbita (Gourd)'	10.4
Lime'	10.1
Sweet basil'	10.0
Colorado pinyon'	9.9
Custard apple'	9.8
Kiwi'	9.0
Common mushroom'	9.0
Brazil nut'	9.0
Yam'	9.0
Soursop'	8.0
Macadamia nut'	8.0
Scarlet bean'	8.0
Lima bean'	8.0

References

1. Network machine learning maps phytochemically rich “Hyperfoods” to fight COVID-19
2. Pineapple is good for helping to boost immune system

Undenatured Type II Collagen (UC-II®) for Joint Health

Denatured type II collagen (UC-II) appears to exert joint-health benefits by oral tolerance, based on pre-clinical research. ^[1]

Figure 1.  Gut-associated lymphoid tissue (GALT) are MALT located in gut
Image Source: DOI: 10.1183/13993003.01701-2015

Oral Tolerance

Oral tolerance is an immune process the body uses to distinguish between 

• Innocuous compounds and
• e.g., dietary proteins, intestinal bacteria
• Harmful foreign invaders

It takes place in the gut-associated lymphoid tissue (GALT). The GALT is mostly made up of mesenteric lymph nodes and patches of lymphoid tissue neighboring the small intestine (Peyer’s patches). ^[2]

Peyer’s patches take in and screen compounds from the gut lumen and, depending on the compound, switch the body’s immune response on or off. 

Gut-associated lymphoid tissue (GALT) is a component of the mucosa-associated lymphoid tissue (MALT) which works in the immune system to protect the body from invasion in the gut. GALT makes up about 70% of the immune system by weight; compromised GALT may significantly affect the strength of the immune system as a whole.

One of the best things you can do to improve oral tolerance is to support a rich diversity of gut bacteria. These bacteria produce short chain fatty acids (SCFA), which help dampen inflammation and manage autoimmune diseases. A limited diet can reduce gut bacteria diversity.

Figure 2.  Synovial fluid (By Madhero88)

What's Collagens

Cartilage — the smooth tissue at the end of bones that cushions the joints, allowing them to glide together smoothly — cannot regenerate itself. The key structural macromolecules of the cartilage tissue in the mammals are:

• Collagen 
• Proteoglycans

Glucosamine, hyaluronic acid, and chondroitin sulfate are vital basic natural constituents of cartilage and synovial fluid. 

Collagens are extracellular matrix molecules used by the cells for structural integrity and a range of further functions. 

Denatured type II collagen, by contrast, lacks these essential structural components.

Figure 3.  Schematic diagram of the proposed mode of action for type II collagen (UC-II) 

How Intaking UC-II Can Help Joint-Health

The interaction between gut-associated lymphoid tissue in the duodenum and epitopes of orally administered undenatured type II collagen facilitates oral tolerance to the antigen and stems systemic T-cell attack on joint cartilage.^[2]

UC-II® contains active epitopes that are able to interact with Peyer’s patches and induce oral tolerance.  A possible mechanism of action for UC-II activity is briefly summarized below (see Figure 3):

1. Transforms naive T-cells into Treg 
• When consumed, UC-II® is believed to be taken up by the Peyer’s patches, where it activates immune cells. It transforms naive T-cells into T regulatory (Treg) cells that specifically target type II collagen. 
2. Treg cells then migrate through the circulation
3. Encounters type II collagen in joint cartilage
1. When they recognize type II collagen in joint cartilage, Treg cells secrete anti-inflammatory mediators (cytokines), including the transforming growth factor-beta (TGF-beta), interleukin 4 (IL-4) and interleukin 10 (IL-10). 
2. This action helps reduce joint inflammation and promotes cartilage repair. 

This process initiates anti-inflammatory and cartilage protective pathways that prevent the immune system from injuring its joint cartilage while promoting cartilage repair and regeneration. 

Summary of cytokines and their functions

Cytokine	Family	Main sources	Function
IL-1β	IL-1	Macrophages, monocytes	Pro-inflammation, proliferation, apoptosis, differentiation
IL-4	IL-4	Th-cells	Anti-inflammation, T-cell and B-cell proliferation, B-cell differentiation
IL-6	IL-6	Macrophages, T-cells, adipocyte	Pro-inflammation, differentiation, cytokine production
IL-8	CXC	Macrophages, epithelial cells, endothelial cells	Pro-inflammation, chemotaxis, angiogenesis
IL-10	IL-10	Monocytes, T-cells, B-cells	Anti-inflammation, inhibition of the pro-inflammatory cytokines
IL-12	IL-12	Dendritic cells, macrophages, neutrophils	Pro-inflammation, cell differentiation, activates NK cell
IL-11	IL-6	Fibroblasts, neurons, epithelial cells	Anti-inflammation, differentiation, induces acute phase protein
TNF-α	TNF	Macrophages, NK cells, CD4+lymphocytes, adipocyte	Pro-inflammation, cytokine production, cell proliferation, apoptosis, anti-infection
IFN-γ	INF	T-cells, NK cells, NKT cells	Pro-inflammation, innate, adaptive immunity anti-viral
GM-CSF	IL-4	T-cells, macrophages, fibroblasts	Pro-inflammation, macrophage activation, increase neutrophil and monocyte function
TGF-β	TGF	Macrophages, T cells	Anti-inflammation, inhibition of pro-inflammatory cytokine production

Preclinical Studies

Preclinical studies support oral tolerance as the mode of action of UC-II® undenatured type II collagen and confirm that the undenatured form of type II collagen is critical for joint-health benefits: 

• In an animal model (mouse) of RA, only UC-II protected against joint damage, an action attributed to oral tolerance.^[4]
• In an animal model (rat) of RA, UC-II provided symptom relief, an action attributed to oral tolerance and modulating inflammatory pathways.^[3]
• In a cell study, Treg cells specific for type II collagen secreted anti-inflammatory cytokines, which play a chief role in the cells’ ability to induce oral tolerance.^[5]
• In a cell study with human chondrocytes (cells that make up cartilage), the anti-inflammatory action of IL-10 protects against damage from tumor necrosis factor-alpha (TNF-α), a pro-inflammatory mediator elevated in osteoarthritis.^[6] 
• Clinically validated lab assays confirm active epitopes in UC-II® undenatured type II collagen resist digestion and retain the undenatured 3D-structure needed to interact with Peyer’s patches and induce oral tolerance.^[2]

References

1. Undenatured Type II Collagen (UC-II) in Joint Health and Disease: A Review on the Current Knowledge of Companion Animals
2. Bagchi D., Misner B., Bagchi M., Kothari S.C., Downs B.W., Fafard R.D., Preuss H.G. Effects of orally administered undenatured type II collagen against arthritic inflammatory diseases: A mechanistic exploration. Int. J. Clin. Pharmacol. Res. 2002;22:101–110.
3. Tong T., Zhao W., Wu Y.-Q., Chang Y., Wang Q.-T., Zhang L.-L., Wei W. Chicken type II collagen induced immune balance of main subtype of helper T cells in mesenteric lymph node lymphocytes in rats with collagen-induced arthritis. Inflamm. Res. Off. J. Eur. Histamine Res.
4. Nagler-Anderson C., Bober L.A., Robinson M.E., Siskind G.W., Thorbecke G.J. Suppression of type II collagen-induced arthritis by intragastric administration of soluble type II collagen. Proc. Natl. Acad. Sci. USA. 1986;83:7443–7446. Soc. 2010;59:369–377.
5. Asnagli H., Martire D., Belmonte N., Quentin J., Bastian H., Boucard-Jourdin M., Fall P.B., Mausset-Bonnefont A.-L., Mantello-Moreau A., Rouquier S., et al. Type 1 regulatory T cells specific for collagen type II as an efficient cell-based therapy in arthritis. Arthritis Res. Ther. 2014;16:R115.
6. Müller R.D., John T., Kohl B., Oberholzer A., Gust T., Hostmann A., Hellmuth M., Laface D., Hutchins B., Laube G., et al. IL-10 overexpression differentially affects cartilage matrix gene expression in response to TNF-alpha in human articular chondrocytes in vitro. Cytokine. 2008;44:377–385.
7. Mucosa Associated Lymphoid Tissues (MALT)

Melatonin Controlling the Balance between Th17 & Treg Cells

Both Treg dysfunction and hyperactive responder T-cell proliferation contribute to disease. In this article, we will discuss the importance of regulating between Th17 and Treg cells.

Figure 1.  IL‐6  mediates Th17/Treg balance.
IL‐6 induces Th17 differentiation from naïve T cells together with TGF‐β.
On the other hand, IL‐6 inhibits Treg differentiation induced by TGF‐β.

IL-6: Regulator of Treg/Th17 Balance

Inflammatory T cells are thought to be central to the pathology of autoimmune diseases. Especially, the recently identified T‐cell subset, the Th17 cells, shows pro‐inflammatory functions and plays a critical role in various autoimmune disorders.

Recent studies have demonstrated that IL-6 has a very important role in regulating the balance between (see Figure 1)

• Th17 cells (T Helper 17 Cells)
• Is a key player in the pathogenesis of autoimmune diseases and protection against bacterial infections
• Both melatonin and active form of vitamin D (1,25-Dihydroxyvitamin D3) are  inhibitors for Th17 differentiation
• Treg (Regulatory T cells)
• Functions to restrain excessive effector T-cell responses

Dysregulation or overproduction of IL-6 leads to autoimmune diseases such as multiple sclerosis (MS) and rheumatoid arthritis (RA), in which Th17 cells are considered to be the primary cause of pathology. 

IL-6 is also central to the pathogenesis of both haemophagocytic lymphohistiocytosis (HLH) and cytokine release syndrome (CRS) and, several studies had shown a correlation between IL-6 levels and adverse outcomes in patients with COVID-19.^[13]

Given the critical role of IL-6 in altering the balance between Treg and Th17 cells, controlling IL-6 activities (i.e., by Tocilizumab an anti–interleukin-6 receptor monoclonal antibody) is potentially an effective approach in the treatment of various autoimmune and inflammatory diseases. 

Figure 2. Schematic representation of the host immune response
against microbial pathogens (Source: 12).

What's Treg?

Regulatory T cells (Tregs) are involved in shutting down immune responses after they have successfully eliminated invading organisms, and also in preventing autoimmunity:

• Come in many forms
• With the most well-understood being those that express CD4, CD25, and FOXP3 (CD4+CD25+ regulatory T cells). 
• These "Tregs" are different from helper T cells.
• Act to suppress activation of the immune system 
• Thereby maintain immunological homeostasis and tolerance to self-antigens (self Ags)
• Contribute to maintaining self-tolerance by down-regulating immune response to self and non-self Ags in an Ag-nonspecific manner.
• Elimination/reduction of CD4+CD25+ Tregs relieves this general suppression, thereby not only enhancing immune responses to non-self antigens (Ags), but also eliciting autoimmune responses to certain self-Ags. 
• Abnormality of this T cell-mediated mechanism of peripheral tolerance can be a possible cause of various autoimmune diseases
• Defects in FOXP3+ Tregs can elicit type 1 diabetes in most individuals regardless of other genetic or environmental influences, thus pointing to a key role for these cells in maintaining islet-specific tolerance.
• Therapies that increase the number or functional capacity of FOXP3+ Tregs can lead to prevention or cure of disease in preclinical models of autoimmunity, including type 1 diabetes
• Suppress Th2 cell function
• Treg from atopic individuals are defective in suppressing Th2 cell function compared with those from non-atopic individuals.

Figure 3: TCR recognition of MHC complexes. When a TCR binds an antigen-MHC complex displayed by a sick or infected cell, the T cell can induce cell death called apoptosis (top). In order for mature, antigen-recognizing T cells to develop without being self-reactive and causing autoimmunity, T cells must go through both positive and negative selection. In positive selection, T cells in the thymus that bind moderately to MHC complexes receive survival signals (middle). However, T cells whose TCRs bind too strongly to MHC complexes, and will likely be self-reactive, are killed in the process of negative selection (bottom).

Treg Selection/Activation

The process of Treg selection is determined by the affinity of interaction with the self-peptide MHC complex. Selection of a T cell to become a Treg is a “Goldilocks” process - i.e. not too high, not too low, but just right.^[9] A T cell that receives (see Figure 3):

• A very strong signals 
• It will undergo apoptotic death (i.e., negative selection)
• A weak signal 
• It will survive and be selected to become an effector cell (i.e., positive selection)
• An intermediate signal
• It will become a regulatory cell

Due to the stochastic nature of the process of T cell activation, all T cell populations with a given TCR will end up with a mixture of Teff and Treg – the relative proportions determined by the affinities of the T cell for the self-peptide-MHC.

Th17 

T helper 17 cells (Th17) are a subset of pro-inflammatory T helper cells defined by their production of interleukin 17 (IL-17). They are also characterized by:

• Th17s are developmentally distinct from Th1 and Th2 lineages (See Figure 2)
• Providing protection against viral infections, and they are also associated with the development of autoimmune diseases because of the recruitment of cells in the granulocyte lineage, especially neutrophils.
• The signals that cause Th17s to differentiate actually inhibit Treg differentiation.
• IL-6 induces the development of Th17 cells from naïve T cells together with TGF-beta; in contrast, IL-6 inhibits TGF-beta-induced Treg differentiation (See Figure 1). 
• Playing an important role in maintaining mucosal barriers and contributing to pathogen clearance at mucosal surfaces; such protective and non-pathogenic Th17 cells have been termed as Treg17 cells.[2]
• The loss of Th17 cell populations at mucosal surfaces has been linked to chronic inflammation and microbial translocation.
• Th17 cell differentiation
• TGF‐β, IL‐6, IL‐1β, and IL‐23 promote Th17 cell differentiation.
• Both melatonin and active form of vitamin D (1,25-Dihydroxyvitamin D3)  inhibit the differentiation of Th17 cells.
• mTOR signaling, which integrates input from insulin, growth factors, and amino acids, while sensing cellular nutrient and energy levels, also regulates Th17 cell differentiation and IL‐17 gene expression.

Figure 4.  Melatonin had an anti-senescent effect in cADMSCs by inhibiting ERS through activation of rhythmic expression of NRF2, activating the ERAD pathway, and inhibiting the NF-κB pathway (Source: [15]).  NF-κB is a major transcription factor that regulates genes responsible for both the innate and adaptive immune response.  Through a cascade of phosphorylation events, the kinase complex is activated and NF-κB is able to enter the nucleus to upregulate genes involved in T-cell development, maturation, and proliferation.

Melatonin 

Based its wide distribution in cells of mammals, it is not surprising that melatonin affects a variety of molecular pathways including those for sleep, circadian rhythms, sexual behavior, immune function, apoptosis, proliferation, metastasis, and angiogenesis oxidative stress.

Melatonin is reported to exert biorhythm regulation, anti-oxidation,^[16] and anti-senescence effects^[15]  in various animal and cell models.

Melatonin significantly influences T-cell-mediated immune responses, which are crucial to protect mammals against cancers and infections, but are associated with pathogenesis of many autoimmune diseases. 

Through modulation in T-cell responses, melatonin exerts beneficial effects in various inflammatory diseases, such as 

• Type 1 diabetes
• In which effector T cells destroy beta cells while Treg counter this assault
• Systemic lupus erythematosus
• Multiple sclerosis
• Melatonin alleviates the clinical symptoms of EAE in mice 
• EAE in mice is a well‐known animal model for MS because there are numerous pathological and histological similarities shared by EAE and MS. 
• The secretion of melatonin is inversely correlated with clinical relapses of Multiple sclerosis (MS) in humans in an epidemiological study
• COVID-19
• In one study, it confirmed the effectiveness of melatonin in treating mild to moderate outpatients with COVID-19.^[14]  

Mechanistically, melatonin 

• Limits peripheral and central Th1/Th17 responses 
• By inhibiting the differentiation of Th17 cells
• Limits the T effector memory population
• Elevates responses of Treg cells

References

1. Melatonin signaling in T cells: Functions and applications (important)
2. Role of Endogenous Melatonin in the Regulation of Th17/Treg Balance during Pregnancy
3. Immunologic self-tolerance maintained by activated T cells expressing IL-2 receptor alpha-chains (CD25). Breakdown of a single mechanism of self-tolerance causes various autoimmune diseases.
4. The Type of Responder T-Cell Has a Significant Impact in a Human In Vitro Suppression Assay
5. Critical stoichiometric ratio of CD4+ CD25+ FoxP3+ regulatory T cells and CD4+ CD25− responder T cells influence immunosuppression in patients with B-cell acute lymphoblastic leukaemia
6. Study on ratio imbalance of peripheral blood Th17/Treg cells in patients with rheumatoid arthritis
7. Ratio balance of Th17 and Treg cells in peripheral blood of patients with chronic lymphocytic leukemia
8. A bovine whey protein extract can induce the generation of regulatory T cells and shows potential to alleviate asthma symptoms in a murine asthma model
9. Li M (August 2016). "T Cell Receptor Signaling in the Control of Regulatory T Cell Differentiation and Function". Nature Reviews Immunology. 16 (4): 220–233.
10. IL-6: regulator of Treg/Th17 balance
11. Recognition of self-peptide–MHC complexes by autoimmune T-cell receptors
12. Intracellular Pathogens: Host Immunity and Microbial Persistence Strategies
13. Coomes, E. A. & Haghbayan, H. Interleukin-6 in Covid-19: a systematic review and meta-analysis. Rev. Med. Virol. 30, e2141 (2020).
14. A Pilot Study on Controlling Coronavirus Disease 2019 (COVID-19) Inflammation Using Melatonin Supplement
15. Melatonin prevents senescence of canine adipose-derived mesenchymal stem cells through activating NRF2 and inhibiting ER stress
16. Reiter RJ, Tan DX, Rosales-Corral S, Galano A, Zhou XJ, Xu B. Mitochondria: central organelles for melatonin’s antioxidant and anti-aging actions. Molecules. 2018; 23:509.
17. Bruns DR, Drake JC, Biela LM, Peelor FF 3rd, Miller BF, Hamilton KL. Nrf2 signaling and the slowed aging phenotype: evidence from long-lived models. Oxid Med Cell Longev. 2015; 2015:732596.
18. Hiebert P, Wietecha MS, Cangkrama M, Haertel E, Mavrogonatou E, Stumpe M, Steenbock H, Grossi S, Beer HD, Angel P, Brinckmann J, Kletsas D, Dengjel J, Werner S. Nrf2-mediated fibroblast reprogramming drives cellular senescence by targeting the matrisome. Dev Cell. 2018; 46:145–161.
• This suggests that time-controlled activation of NRF2 may be critical for homeostasis in multicellular organism.
19. Antigenic drift: Understanding COVID-19 (good)