Synthesis of 2-methyl-1,4-naphthoquinones with higher gamma-glutamyl carboxylase activity than MK-4 both in vitro and in vivo.

Vitamin K is the collective term for compounds that share a 2-methyl-1,4-naphthoquinone ring, but differ in the side-chain at the 3-position. We synthesized novel 2-methyl-1,4-naphthoquinone derivatives with different side chain length at the 3-position. Derivatives with C-14 and C-16 tails showed the highest in vitro bioactivity resulting in 2.5 and 2-fold higher carboxylated osteocalcin synthesis in MG63 cells than menaquinone-4 (MK-4, form of vitamin K2). Longer side chain lengths resulted in lower bioactivity. The in vivo vitamin K activity of the C-14 tail derivative was further tested in WKY rats receiving a vitamin K-deficient diet that resulted in a 40% decrease of prothrombin activity. The C-14 tail derivative was able to counteract the effects on vitamin K deficiency induced by the diet and resulted in the complete restoration of prothrombin activity. Compared to naturally occurring forms of vitamin K, synthetic vitamin K derivatives may have higher bioactivity and different pharmacological characteristics that are more favorable for use as supplements or in clinical settings.

Yogurt drink fortified with menaquinone-7 improves vitamin K status in a healthy population.

Population-based studies have shown an inverse association between dietary menaquinones (MK-n, vitamin K2) intake, coronary calcification and CHD risk, suggesting a potential role of vitamin K in vascular health. To date, the effects of increased menaquinone intake on (markers of) vascular health have been investigated using predominantly food supplements. Dairy products contain many essential nutrients and can serve as a good matrix for food fortification in order to support health. We were therefore interested to study the effects of a menaquinone-fortified yogurt drink (menaquinone as menaquinone-7 (MK-7); 28 µg MK-7/yogurt drink) on vitamin K status and markers of vascular health. The yogurt drink was also fortified with n-3 PUFA, vitamin D, vitamin C, Ca and Mg to support vascular and/or general health. Healthy men (n 32) and postmenopausal women (n 28) with a mean age of 56 (sd 5) years received either basic or fortified yogurt drink twice per d for 12 weeks. MK-7 was efficiently absorbed from the fortified yogurt drink. Levels of circulating MK-7 were significantly increased from 0·28 to 1·94 ng/ml. In accordance, intake of the fortified yogurt drink improved vitamin K status, as measured by significant decreases in uncarboxylated osteocalcin and desphospho-uncarboxylated matrix Gla-protein. No effects were, however, seen on markers of inflammation, endothelial dysfunction and lipid metabolism. In summary, consumption of a yogurt drink fortified with low doses of among others MK-7 for 3 months significantly improved vitamin K status in a healthy population.

Inactive Matrix Gla-Protein Is Associated With Arterial Stiffness in an Adult Population-Based Study.

Increased pulse wave velocity (PWV) is a marker of aortic stiffness and an independent predictor of mortality. Matrix Gla-protein (MGP) is a vascular calcification inhibitor that needs vitamin K to be activated. Inactive MGP, known as desphospho-uncarboxylated MGP (dp-ucMGP), can be measured in plasma and has been associated with various cardiovascular markers, cardiovascular outcomes, and mortality. In this study, we hypothesized that high levels of dp-ucMGP are associated with increased PWV. We recruited participants via a multicenter family-based cross-sectional study in Switzerland. Dp-ucMGP was quantified in plasma by sandwich ELISA. Aortic PWV was determined by applanation tonometry using carotid and femoral pulse waveforms. Multiple regression analysis was performed to estimate associations between PWV and dp-ucMGP adjusting for age, renal function, and other cardiovascular risk factors. We included 1001 participants in our analyses (475 men and 526 women). Mean values were 7.87±2.10 m/s for PWV and 0.43±0.20 nmol/L for dp-ucMGP. PWV was positively associated with dp-ucMGP both before and after adjustment for sex, age, body mass index, height, systolic and diastolic blood pressure (BP), heart rate, renal function, low- and high-density lipoprotein, glucose, smoking status, diabetes mellitus, BP and cholesterol lowering drugs, and history of cardiovascular disease (P≤0.01). In conclusion, high levels of dp-ucMGP are independently and positively associated with arterial stiffness after adjustment for common cardiovascular risk factors, renal function, and age. Experimental studies are needed to determine whether vitamin K supplementation slows arterial stiffening by increasing MGP carboxylation.

Menaquinone-7 supplementation improves arterial stiffness in healthy postmenopausal women. A double-blind randomised clinical trial.

Observational data suggest a link between menaquinone (MK, vitamin K2) intake and cardiovascular (CV) health. However, MK intervention trials with vascular endpoints are lacking. We investigated long-term effects of MK-7 (180 µg MenaQ7/day) supplementation on arterial stiffness in a double-blind, placebo-controlled trial. Healthy postmenopausal women (n=244) received either placebo (n=124) or MK-7 (n=120) for three years. Indices of local carotid stiffness (intima-media thickness IMT, Diameter end-diastole and Distension) were measured by echotracking. Regional aortic stiffness (carotid-femoral and carotid-radial Pulse Wave Velocity, cfPWV and crPWV, respectively) was measured using mechanotransducers. Circulating desphospho-uncarboxylated matrix Gla-protein (dp-ucMGP) as well as acute phase markers Interleukin-6 (IL-6), high-sensitive C-reactive protein (hsCRP), tumour necrosis factor-α (TNF-α) and markers for endothelial dysfunction Vascular Cell Adhesion Molecule (VCAM), E-selectin, and Advanced Glycation Endproducts (AGEs) were measured. At baseline dp-ucMGP was associated with IMT, Diameter, cfPWV and with the mean z-scores of acute phase markers (APMscore) and of markers for endothelial dysfunction (EDFscore). After three year MK-7 supplementation cfPWV and the Stiffness Index ßsignificantly decreased in the total group, whereas distension, compliance, distensibility, Young's Modulus, and the local carotid PWV (cPWV) improved in women having a baseline Stiffness Index ß above the median of 10.8. MK-7 decreased dp-ucMGP by 50 % compared to placebo, but did not influence the markers for acute phase and endothelial dysfunction. In conclusion, long-term use of MK-7 supplements improves arterial stiffness in healthy postmenopausal women, especially in women having a high arterial stiffness.

Vitamin K status and mortality after kidney transplantation: a cohort study.

BACKGROUND: Vitamin K modulates calcification by activating calcification inhibitors such as matrix Gla protein (MGP). In kidney transplant recipients, vitamin K insufficiency is common, but implications for long-term outcomes are unclear. STUDY DESIGN: Single-center observational study with a longitudinal design. SETTING & PARTICIPANTS: 518 stable kidney transplant recipients; 56% men; mean age, 51±12 (SD) years; and a median of 6 (IQR, 3-12) years after kidney transplantation. FACTOR: Plasma desphosphorylated-uncarboxylated MGP (dp-ucMGP) levels, reflecting vitamin K status. OUTCOMES: All-cause mortality and transplant failure. RESULTS: At inclusion, median dp-ucMGP level was 1,038 (IQR, 733-1,536) pmol/L, with 473 (91%) patients having vitamin K insufficiency (defined as dp-ucMGP>500pmol/L). During a median follow-up of 9.8 (IQR, 8.5-10.2) years, 152 (29%) patients died and 54 (10%) developed transplant failure. Patients in the highest quartile of dp-ucMGP were at considerably higher mortality risk compared with patients in the lowest quartile (HR, 3.10; 95% CI, 1.87-5.12; P for trend<0.001; P for quartile 1 [Q1] vs Q4<0.001). After adjustment for potential confounders, including kidney function and exclusion of patients treated with a vitamin K antagonist, this association remained significant. Patients in the highest quartile also were at higher risk of developing transplant failure (HR, 2.61; 95% CI, 1.22-5.57; P for trend=0.004; P for Q1 vs Q4=0.01), but this association was lost after adjustment for baseline kidney function (HR, 1.20; 95% CI, 0.52-2.75; P for trend=0.6; P for Q1 vs Q4=0.7). LIMITATIONS: Although MGP exists as various species, only dp-ucMGP was measured. No data were available for vascular calcification as an intermediate end point. CONCLUSIONS: Vitamin K insufficiency, that is, a high circulating level of dp-ucMGP, is highly prevalent in stable kidney transplant recipients and is associated independently with increased risk of mortality. Future studies should address whether vitamin K supplementation may lead to improved outcomes after kidney transplantation.

Association of vitamin K status with adiponectin and body composition in healthy subjects: uncarboxylated osteocalcin is not associated with fat mass and body weight.

Osteocalcin (OC) is a vitamin K-dependent protein found in bone and in circulation. High serum γ-carboxylated OC reflects a high, and high uncarboxylated OC (ucOC) reflects a low vitamin K status. A revolutionary hypothesis is that ucOC acts as a hormone improving glucose handling and reducing fat mass. The objective was to test the logical extrapolation of the ucOC hormone hypothesis to humans that elevated ucOC is associated with higher body weight, BMI and fat mass. In a cross-sectional analysis, the associations of vitamin K status with circulating adiponectin and body composition were investigated in 244 postmenopausal women (study I). The effects of vitamin K treatment on adiponectin, body weight and BMI were investigated in archived samples from forty-two young men and women who received varying doses of menaquinone-7 during 12 weeks (study II) and from a cohort of 164 postmenopausal women who participated in a 3-year placebo-controlled trial on 45 mg menaquinone-4 (MK-4) (study III). No association was found between vitamin K status and circulating adiponectin before or after vitamin K supplementation. A higher carboxylation of OC was significantly correlated with lower body weight, BMI and fat mass of the trunk. Women taking MK-4 maintained their baseline body weight and BMI, whereas women taking placebo showed significant increases in both indices. These findings demonstrate that a high vitamin K status of bone has no effect on circulating adiponectin in healthy people and long-term vitamin K supplementation does not increase weight in healthy postmenopausal women.

Vitamin K-containing dietary supplements: comparison of synthetic vitamin K1 and natto-derived menaquinone-7.

Vitamin K is a cofactor in the production of blood coagulation factors (in the liver), osteocalcin (in bone), and matrix Gla protein (cartilage and vessel wall). Accumulating evidence suggests that for optimal bone and vascular health, relatively high intakes of vitamin K are required. The synthetic short-chain vitamin K(1) is commonly used in food supplements, but recently the natural long-chain menaquinone-7 (MK-7) has also become available as an over-the-counter (OTC) supplement. The purpose of this paper was to compare in healthy volunteers the absorption and efficacy of K(1) and MK-7. Serum vitamin K species were used as a marker for absorption and osteocalcin carboxylation as a marker for activity. Both K(1) and MK-7 were absorbed well, with peak serum concentrations at 4 hours after intake. A major difference between the 2 vitamin K species is the very long half-life time of MK-7, resulting in much more stable serum levels, and accumulation of MK-7 to higher levels (7- to 8-fold) during prolonged intake. MK-7 induced more complete carboxylation of osteocalcin, and hematologists should be aware that preparations supplying 50 mug/d or more of MK-7 may interfere with oral anticoagulant treatment in a clinically relevant way.

Factors affecting bone loss in female endurance athletes: a two-year follow-up study.

BACKGROUND: Low bone mass leading to stress fractures is a well-known and yet unsolved problem among female athletes. PURPOSE: To quantify the rate of bone loss in healthy female athletes and investigate the effects of estrogen and vitamin K supplementation on bone loss. STUDY DESIGN: Prospective cohort study. METHODS: We classified 115 female endurance athletes into amenorrheic, eumenorrheic, or estrogen-supplemented groups and randomized them to receive either placebo or vitamin K(1). The bone mineral densities of the subjects' femoral neck and lumbar spine were measured at baseline and after 2 years. RESULTS: Bone mineral density in the lumbar spine remained constant, but bone density in the femoral neck had decreased significantly after 2 years in all three subgroups. The decrease was higher in amenorrheic (-6.5% +/- 4.0%) than in eumenorrheic (-3.2% +/- 4.1%) and estrogen-supplemented athletes (-3.9% +/- 3.1%). Supplementation with vitamin K did not affect the rate of bone loss. CONCLUSIONS: The rate of bone loss in all three subgroups of female athletes was unexpectedly high; neither estrogen nor vitamin K supplementation prevented bone loss. CLINICAL RELEVANCE: High-intensity training maintained over several years must be regarded in women as a risk factor for osteoporosis, and protocols for optimal treatment should be developed.