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Calcitriol modifies tight junctions, improves barrier function, and reduces TNF‐α‐induced barrier leak in the human lung‐derived epithelial cell culture model, 16HBE 14o‐

Rybakovsky et al., Physiological Reports, doi:10.14814/phy2.15592
Apr 2023  
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Vitamin D for COVID-19
8th treatment shown to reduce risk in October 2020
*, now known with p < 0.00000000001 from 120 studies, recognized in 9 countries.
No treatment is 100% effective. Protocols combine complementary and synergistic treatments. * >10% efficacy in meta analysis with ≥3 clinical studies.
4,200+ studies for 70+ treatments.
In Vitro study showing that calcitriol improved barrier function in human airway epithelial cells. Authors note that this mechanism could explain in part the efficacy of vitamin D seen for COVID-19 and other airway diseases.
21 preclinical studies support the efficacy of vitamin D for COVID-19:
Vitamin D has been identified by the European Food Safety Authority (EFSA) as having sufficient evidence for a causal relationship between intake and optimal immune system function20-23. Vitamin D inhibits SARS-CoV-2 replication in vitro10,17, mitigates lung inflammation, damage, and lethality in mice with an MHV-3 model for β-CoV respiratory infections10,17, reduces SARS-CoV-2 replication in nasal epithelial cells via increased type I interferon expression13, downregulates proinflammatory cytokines IL-1β and TNF-α in SARS-CoV-2 spike protein-stimulated cells9, attenuates nucleocapsid protein-induced hyperinflammation by inactivating the NLRP3 inflammasome through the VDR-BRCC3 signaling pathway14, may be neuroprotective by protecting the blood-brain barrier, reducing neuroinflammation, and via immunomodulatory effects24, minimizes platelet aggregation mediated by SARS-CoV-2 spike protein via inhibiting integrin αIIbβ3 outside-in signaling8, and improves regulatory immune cell levels and control of proinflammatory cytokines in severe COVID-1925. Symptomatic COVID-19 is associated with a lower frequency of natural killer (NK) cells and vitamin D has been shown to improve NK cell activity26,27.
Rybakovsky et al., 11 Apr 2023, peer-reviewed, 7 authors. Contact:
In Vitro studies are an important part of preclinical research, however results may be very different in vivo.
This PaperVitamin DAll
Calcitriol modifies tight junctions, improves barrier function, and reduces TNF‐α‐induced barrier leak in the human lung‐derived epithelial cell culture model, 16HBE 14o‐
Elizabeth Rybakovsky, Katherine M Diguilio, Mary Carmen Valenzano, Sophie Geagan, Kaithlyn Pham, Ronald N Harty, James M Mullin
Physiological Reports, doi:10.14814/phy2.15592
Using the 16HBE 14o-human airway epithelial cell culture model, calcitriol (Vitamin D) was shown to improve barrier function by two independent metrics -increased transepithelial electrical resistance (TER) and reduced transepithelial diffusion of 14 C-D-mannitol (J m ). Both effects were concentration dependent and active out to 168 h post-treatment. Barrier improvement associated with changes in the abundance of specific tight junctional (TJ) proteins in detergent-soluble fractions, most notably decreased claudin-2. TNF-αinduced compromise of barrier function could be attenuated by calcitriol with a concentration dependence similar to that observed for improvement of control barrier function. TNFαinduced increases in claudin-2 were partially reversed by calcitriol. The ERK 1,2 inhibitor, U0126, itself improved 16HBE barrier function indicating MAPK pathway regulation of 16HBE barrier function. Calcitriol's action was additive to the effect of U0126 in reducing TNF-α -induced barrier compromise, suggesting that calcitriol may be acting through a non-ERK pathway in its blunting of TNF-α -induced barrier compromise. This was supported by calcitriol being without effect on pERK levels elevated by the action of TNF-α. Lack of effect of TNF-α on the death marker, caspase-3, and the inability of calcitriol to decrease the elevated LC3B II level caused by TNF-α, suggest that calcitriol's barrier improvement does not involve a cell death pathway. Calcitriol's improvement of control barrier function was not additive to barrier improvement induced by retinoic acid (Vitamin A). Calcitriol improvement and protection of airway barrier function could in part explain Vitamin D's reported clinical efficacy in COVID-19 and other airway diseases.
AUTHOR CONTRIBUTIONS ACKNOWLEDGMENTS The authors are very grateful to Ms. Terri Olshefski and Ms. Elene Mironidis of the Editorial Office of the Lankenau Institute for Medical Research for their work in formatting and editing our manuscript for publication. The assistance of Ms. Elizabeth Newberry in proofreading is also gratefully acknowledged. We are very thankful to Dr. Mazen Hassan and Ms. Kari Heller for their help in obtaining needed reference material. FUNDING INFORMATION Financial support for this research came in part from a research grant from the Sharpe-Strumia Research Foundation (JMM) and NIH grant AI139392 (RNH). ETHICAL STATEMENT This Study used neither animal nor human subjects. All science was conducted in an ethical manner.
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