When illness abounds, many turn to vitamins and supplements to boost their immune systems. Along this line, a few observational studies and a larger systematic review have linked vitamin D deficiencies with either a higher risk of testing positive for SARS-CoV-2, the virus that causes COVID-19, or becoming severely ill from the disease.^1-4 Aside from its vital role in bone health, vitamin D offers a compelling reason to be helpful in the context of COVID-19 by functioning to increase antimicrobial peptide expression and anti-inflammatory effects that support immune defense and acting directly on angiotensin-converting enzyme 2 (ACE2), which serves as the cell-surface entry receptor for SARS-CoV-2.

Vitamin D Regulation of Innate and Adaptive Immunity

Vitamin D₃ is obtained from certain dietary sources and through the action of UVB light on 7-dehydro cholesterol in skin. Hydroxylation of vitamin D₃ in the liver produces 25-hydroxy vitamin D₃, the principal circulating form of vitamin D, which is an intermediate in the pathway leading to the production of 1,25-dihydroxy vitamin D₃, the most receptor-active form of the vitamin. Antigen-presenting cells (APCs) and epithelial cells can also synthesize 1,25-dihydroxy vitamin D₃ through 1α-hydroxylase/CYP27B1, which is regulated by immune stimulus. For example, human macrophages stimulated through toll-like receptor recognition of M. tuberculosis induce CYP27B1 and vitamin D receptor expression.⁵ 1,25-dihydroxy vitamin D₃ signals through the vitamin D receptor (VDR), which is widely expressed in epithelial tissues and immune cells (Figure 1).

Figure 1. Innate and acquired immune responses are influenced by vitamin D availability.

Ligand binding induces the VDR to dimerize with retinoid X receptors (RXRs) and promotes DNA binding to vitamin D response elements to directly induce the expression of cathelicidin antimicrobial peptide and β-defensin-2 genes.⁶ Cathelicidin induces a variety of proinflammatory cytokines, stimulates the chemotaxis of neutrophils, monocytes, macrophages, and T cells into the site of infection, and promotes the clearance of respiratory pathogens by inducing apoptosis and autophagy of infected epithelial cells. β-Defensin-2 stimulates the expression of antiviral cytokines and chemokines involved in the recruitment of monocytes, macrophages, natural killer cells, neutrophils, and T cells. 1,25-dihydroxy Vitamin D₃ can induce expression of the nucleotide-binding oligomerization domain-containing protein 2, which enhances β-defensin-2 expression.

Another property of 1,25-dihydroxy vitamin D₃ is the promotion of autophagy, wherein viral particles are packaged for lysosomal degradation and subsequent antigen presentation for adaptive antiviral immune responses. The mechanisms through which vitamin D promotes autophagy involves downregulating the mTOR pathway and promoting Beclin 1 and the class III PI3K complex.⁷ Vitamin D may also stimulate the formation of autophagosomes to promote viral clearance indirectly through induction of cathelicidin expression.⁸

1,25-dihydroxy Vitamin D₃ also exerts inhibitory and anti-inflammatory actions on the adaptive immune system, decreasing the maturation of dendritic cells and suppressing T helper cell type 1 responses.⁹ Dendritic cells treated with 1,25-dihydroxy vitamin D₃ can also induce regulatory T cells, which may have a role in preventing a cytokine storm caused by a viral infection.¹⁰

Vitamin D and ACE2

ACE2, a part of the renin-angiotensin system, is the major entry point into cells for SARS-CoV-2 (Figure 2). Once the virus attaches to human ACE2 through its spike glycoprotein, it reduces the expression of ACE2, which leads to excessive angiotensin (Ang) II production. Through its interaction with the Ang II type 1 (AT₁) receptor, Ang II can stimulate growth and proliferation of lung fibroblasts, upregulate the expression of TGF-β, cause lung inflammation through increases in reactive oxygen species and proinflammatory cytokines, and induce alveolar epithelial cell apoptosis, all of which lead to lung injury. When ACE2 is readily available, it catalyzes the cleavage of Ang II into Ang (1-7), which signals through the Mas receptor. Ang (1-7) signaling counteracts the proinflammatory and profibrotic Ang II activity at the AT₁ receptor. 

Figure 2. Potential interactions of vitamin D with the renin-angiotensin system.

1,25-dihydroxy Vitamin D₃ has been shown to induce ACE2/Ang (1-7)/Mas receptor axis activity while inhibiting ACE/Ang II/AT₁ activity.¹¹ It also acts as a negative modulator of the renin-angiotensin system by inhibiting renin expression and generation. Thus, 1,25-dihydroxy vitamin D₃ may provide protection against lung injury.

Clinical Evidence

While many observational studies demonstrate a trend for sufficient levels of vitamin D prior to infection being important to immune health, they cannot prove that vitamin D protects people against infection. In fact, a meta-analysis of 31 peer-reviewed observational studies has found no statistically significant association with COVID-19 or health outcomes.¹² Randomized clinical trials are better suited to determine whether vitamin D affects COVID-19 risk. However, the largest randomized, placebo-controlled trial of vitamin D administration in hospitalized patients with respiratory infection and critical illness due to COVID-19 published to date did not support routine administration of vitamin D in subjects with moderate to severe COVID-19.¹³ The COVID-19 and High-Dose Vitamin D Supplementation Trial (COVIT-TRIAL) has recruited participants to test the effect of vitamin D supplementation on the prognosis of COVID-19 in high-risk older patients.¹⁴ Several other clinical trials are also in preparation to test the effect of vitamin D supplements during or in the prevention of COVID-19.¹⁵

While there is evidence that vitamin D may have positive effects on immune function, specific antiviral effects remain unproven. Moreover, because the value of vitamin D has been defined largely through studies on bone health, there are still big gaps in the understanding of what a normal level should be for proper immune function. From 2010-2012, the Correction of Vitamin D Deficiency in Critically Ill Patients (VITdAL-ICU) study was conducted as a multicenter, randomized clinical trial to test the effect of vitamin D₃ administration (540,000 IU) versus placebo in 475 critically ill patients with a vitamin D deficiency (≤ 20 ng/ml 25-hydroxy vitamin D₃) recruited from medical or neurological intensive care units and cardiothoracic surgery or mixed-surgery units.¹⁶ Among patients with vitamin D deficiency who were critically ill, administration of high-dose vitamin D₃ compared with placebo did not improve hospital length of stay, hospital mortality, or 6-month mortality. The Vitamin D to Improve Outcomes by Leveraging Early Treatment (VIOLET) trial conducted from 2017-2018 sought to examine the effect of the same dose of vitamin D₃ versus placebo on 90-day mortality in 3,000 critically ill patients with vitamin D deficiency but was stopped early after enrollment of 1,360 patients demonstrated very low likelihood for benefit.¹⁷ If there is indeed a benefit for COVID-19, much work will need to be done to determine the ideal dosage to reach peak advantage.

Cayman offers several products to study the underlying mechanisms of the protective effects of vitamin D, including vitamins D₂, D₃, D₄, and analogs (some of which are isotopically labeled), assay kits, metabolites, precursors, and a vitamin D receptor antagonist.

Vitamin D & Analogs

Vitamin D₂
Vitamin D₃
Vitamin D₄
Calcipotriol (hydrate)
BXL-628
EB 1089
22-Oxacalcitriol
25-hydroxy Vitamin D₃ 3,3'-aminopropyl ether
5,6-trans-Vitamin D₃

Vitamin D Assay Kits

Vitamin D ELISA Kit
Human Vitamin D Receptor Reporter Assay System

Vitamin D Metabolites

1,24-dihydroxy Vitamin D₃ (hydrate)
Calcitriol
3-epi-25-hydroxy Vitamin D₃
1-hydroxy Vitamin D
25-hydroxy Vitamin D₂
1,25-dihydroxy Vitamin D₂
Paricalcitol

Vitamin D Precursors

Cholesterol
Ergosterol
7-dehydro Cholesterol
Previtamin D₃

Vitamin D Receptor Antagonist

ZK 159222

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References

1. Meltzer, D.O., Best, T.J., Zhang, H., et al. Association of vitamin D status and other clinical characteristics with COVID-19 test results. JAMA Netw. Open 3(9), e2019722 (2020).

2. Meltzer, D.O., Best, T.J., Zhang, H., et al. Association of vitamin D levels, race/ethnicity, and clinical characteristics with COVID-19 test results. JAMA Netw. Open 4(3), e214117 (2021).

3. Pereira, M., Damascena, A.D., Azevedo, L.M.G., et al. Vitamin D deficiency aggravates COVID-19: Systematic review and meta-analysis. Crit. Rev. Food Sci. Nutr. (2020).

4. Ricci, A., Pagliuca, A., D'Ascanio, M., et al. Circulating vitamin D levels status and clinical prognostic indices in COVID-19 patients. Respir. Res. 22(1), 76 (2021). 

5. Liu, P.T., Stenger, S., Li, H., et al. Toll-like receptor triggering of a vitamin D-mediated human antimicrobial response. Science 311(5768), 1770-1773 (2006).

6. Wang, T.-T., Nestel, F.P., Bourdeau, V., et al. Cutting edge: 1,25-Dihydroxyvitamin D₃ is a direct inducer of antimicrobial peptide gene expression. J. Immunol. 173(5), 2909-2912 (2004).

7. Wang, J. Beclin 1 bridges autophagy, apoptosis, and differentiation. Autophagy 4(7), 947-948 (2008).

8. Yuk, J.-M., Shin, D.-M., Lee, H.-M., et al. Vitamin D3 induces autophagy in human monocytes/macrophages via cathelicidin. Cell Host Microbe 6(3), 231-243 (2009).

9. van Etten, E. and Mathieu, C. Immunoregulation by 1,25-dihydroxyvitamin D₃: Basic concepts. J. Steroid Biochem. Mol. Biol. 97(1-2), 93-101 (2005).

10. Bilezikian, J.P., Bikle, D., Hewison, M., et al. Mechanisms in endocrinology: Vitamin D and COVID-19. Eur. J. Endocrinol. 183(5), R133-R147 (2020).

11. Mahdavi, A.M. A brief review of interplay between vitamin D and angiotensin-converting enzyme 2: Implications for a potential treatment for COVID-19. Rev. Med. Virol. 30(5), e2119 (2020).

12. Bassatne, A., Basbous, M., Chakhtoura, M., et al. The link between COVID-19 and VItamin D (VIVID): A systematic review and meta-analysis. Metabolism 119, 154753 (2021). 

13. Murai, I.H., Fernandes, A.L., Sales, L.P., et al. Effect of a single high dose of Vitamin D₃ on hospital length of stay in patients with moderate to severe COVID-19: A randomized clinical trial. JAMA 325(11), 1053-1060 (2021). 

14. Annweiler, C., Beaudenon, M., Gautier, J., et al. COvid-19 and high-dose VITamin D supplementation TRIAL in high-risk older patients (COVIT-TRIAL): Study protocol for a randomized controlled trial. Trials 21(1), 1031 (2020).

15. Teymoori-Rad, M. and Marashi, S.M. Vitamin D and Covid-19: From potential therapeutic effects to unanswered questions. Rev. Med. Virol. 31(2), e2159 (2021).

16. Amrein, K., Schnedl, C., Holl, A., et al. Effect of high-dose vitamin D₃ on hospital length of stay in critically ill patients with vitamin D deficiency: The VITdAL-ICU randomized clinical trial. JAMA 312(15), 1520-1530 (2014).

17. National Heart, Lung, and Blood Institute PETAL Clinical Trials Network, Ginde, A.A., Brower, R.G., Caterino, J.M., et al. Early high-dose vitamin D₃ for critically ill, vitamin D-deficient patients. N. Engl. J. Med. 381(26), 2529-2540 (2019).