MARESZ K.#, FLORCZYK D.^, KOZIEL J.^, BADMAEV V.*
*NattoPharma ASA , #The International Science and Health Foundation, ^Jagiellonian University, Krakow
Corresponding author: email@example.com
Keywords: Vitamin K2, inflammation, human peripheral blood monocytes
Markers of low-grade inflammation positively correlate with the endothelial damage, atheroma formation and cardiovascular disease. In addition, the circulating proinflammatory markers tend to increase with aging. The long term, 3 year supplementation of menaquinone-7 (180 mcg/day) in postmenopausal healthy women, resulted in a statistically significant improvement in functional measurements of arterial stiffness (carotid distensibility, carotid compliance, pulse wave velocity), a hallmark of age-related cardiovascular decline. Based on the cumulative literature data, it is possible that the role of menaquinone-7 in the reported clinical improvement of the cardiovascular health might be due to its potential immunoregulatory and anti-inflammatory roles.
The goal and methods:
The goal of the following study was to test a hypothetical role of vitamin K2 in modulation of the immune and inflammatory biomarkers and functions in vitro. High purity natural vitamin K2, 98% menaquinone-7 (MenaQ7®Crystals) was evaluated in vitro, for its potential to inhibit production of pro-inflammatory marker TNF-alpha by human peripheral blood monocytes (PBMC). The K2 was dissolved in ethanol and evaluated in tissue culture of PBMC in four concentrations:0,1uM, 1 uM, 10 uM and 100 uM. The peripheral blood monocytes (PBMC) were obtained from a sample of peripheral blood of healthy individuals in a gradient isolation method. The level of TNF-alpha production by PBMC was evaluated with or without vitamin K2 pre-incubation, up to 30 hours, in the presence of three different TLR ligands such as LPS, PAM and MALP.
Vitamin K2 added to the cell culture of PBMC at the same time as the TLR agonists, did not influence the production of pro-inflammatory mediators. The PBMC produced the same amount of TNF-alpha with the concurrent LPS, MALP and PAM treatments in the presence or absence of vitamin K2 in up to 100uM concentration. Next step was to evaluate the potential modulation of the immune and inflammatory biomarkers with vitamin K2, when the cells were pretreated with vitamin K2 prior to treatment with TLR agonists.
It was found that vitamin K2 in concentration below the 10uM and short time pretreatment, up to 6 hours, did not inhibit significantly the production of TNF-alpha after the TLR activation.
However, the 30 hours pretreatment of PBMC with at least 10uM of vitamin K2 effectively inhibited the proinflammatory function of PBMC, and the effect was dose dependent.
The 10uM of vitamin K2 after 30 hours of PBMC pretreatment, resulted in 20%,statistically significant, inhibition of the TNF-alpha production after LPS activation (p<0.05), and 43% inhibition after MALP activation (p<0.001). PAM activation was inhibited by 20% after 10uM Vitamin K2, however this inhibition wasn’t statistically significant.
The response to 100uM of vitamin K2 was more powerful quantitatively and qualitatively (inhibition of all three TLRs) in comparison to the 10uM dose. The inhibition of TNF alpha production by LPS was 63% and by PAM 30% , which was statistically significantly higher (p<0.001), when compared to 43% inhibition after MALP activation (p<0.05).
We plan to test additional biomarkers of immune and inflammatory response in the presence of vitamin K2 in vitro. This work may elucidate the anti-inflammatory mechanism of vitamin K2 and establish the potential biomarker targets in clinical testing of vitamin K2 role in the cardiovascular health.
Vitamin K2 is able to modulate the immune and inflammatory functions in the dose response inhibition of TNF-alpha production by the healthy human PBMC in vitro.
Acknowledgements: The present study was supported by Nattopharma ASA (Oslo, Norway). None of the authors had any possible conflicts of interest.