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In this study, we developed a column-switching high-performance liquid chromatography (HPLC) method with fluorescence detection for the analysis of vitamin K. Column-switching is accomplished by changing the direction of flow using a switching valve with a set time program. Using this method, three vitamin K, phylloquinone (PK), menaquinone-4 (MK-4), and menaquinone-7 (MK-7), were separated and identified with high sensitivity, and impurities were eliminated. This method was used to determine the vitamin K content in meat, fish meat, snails, bivalves, sea urchins, seaweeds, vegetables, tea, soy products, milk products, and supplements. The results showed that chicken showed the highest content of MK-4 (15.35 ± 0.35 μg/100 g), matcha showed the highest content of PK (3069.66±80.10 μg/100 g), and dried natto showed the highest content of MK-7 (3997.57±79.42 μg/100 g). This method can also be used to analyze vitamin K in supplements and pharmaceuticals. The results of this study revealed that different manufacturers add different types of vitamin K to their commercial supplements and infant formulas. The developed method provides highly reproducible and quantitative results and allows for the rapid analysis of the three vitamin K types. Thus, the method developed in this study may aid the sequential analysis of vitamin K in different samples to assess food nutrients. [ABSTRACT FROM AUTHOR]

Objective To investigate whether menaquinone-4 （MK-4） can exert a protective effect against carbon tetrachloride （CCl4）-induced acute liver injury （ALI） in mice by alleviating ferroptosis. Methods After adaptive feeding， adult male ICR mice， aged 8 weeks， were divided into Control group， MK-4 group， CCl4 model group （6-hour， 12-hour， and 24-hour）， and MK-4+CCl4 group （6-hour， 12-hour， and 24-hour）， with 6 mice in each group. The mice in the Control group were given intraperitoneal injection of an equal dose of corn oil； the mice in the MK-4 group were given intraperitoneal injection of 40 mg/kg MK-4 solution， followed by an equal dose of corn oil after 1 hour； the mice in the MK-4+CCl4 group （6-hour， 12-hour， and 24-hour） were given intraperitoneal injection of 40 mg/kg MK-4 solution， and after 1 hour， the mice in this group and the CCl4 model group （6-hour， 12-hour， and 24-hour） were given intraperitoneal injection of 0.3 mL/kg CCl4 solution， with samples collected at 6， 12， and 24 hours. HE staining was used to observe the pathological changes of mouse liver； Prussian blue staining was used to observe iron accumulation in liver tissue； a biochemical analyzer was used to measure the serum levels of aspartate aminotransferase （AST） and alanine aminotransferase （ALT）； related kits were used to measure the levels of tissue iron content and the oxidative stress indices malondialdehyde （MDA） and glutathione （GSH） in liver homogenate； RT-PCR was used to measure the expression levels of ferroptosis marker genes （acyl-CoA synthetase long-chain family member 4 ［ACSL4］， prostaglandin-endoperoxide synthase 2 ［PTGS2］， and glutathione peroxidase 4 ［GPX4］） and iron metabolism-related genes （hemojuvelin ［HJV］， transferrin receptor 1 ［TFR1］， and ferroportin ［FPN］）， and Western blot was used to measure the protein expression level of GPX4. A one-way analysis of variance was used for comparison of continuous data between multiple groups， and the least significant difference t-test was used for further comparison between two groups. Results In the aging study， compared with the Control group， the CCl4 model group （6-hour， 12-hour， and 24-hour） had significant increases in liver weight coefficient and the serum levels of ALT and AST （all P<0.05）， and HE staining also showed that liver injury gradually aggravated over time. Meanwhile， compared with the CCl4 model group （6-hour， 12-hour， and 24-hour）， the MK-4+CCl4 （12-hour） group had significant reductions in liver weight coefficient and the serum levels of ALT and AST （all P<0.05）， with a reduction in the necrotic area of liver tissue， and therefore， 12-hour mouse tissue samples were used for detection in the following study. Compared with the Control group， the CCl4 group had a significant increase in MDA and a significant reduction in GSH （both P<0.05）， and compared with the CCl4 group， the MK-4+CCl4 group had a significant reduction in MDA and a significant increase in GSH （both P<0.05）. Compared with the Control group， the CCl4 group had significant increases in the key ferroptosis indices ASCL4 and PTGS2 and a significant reduction in GPX4 （all P<0.05）； compared with the CCl4 group， the MK-4+CCl4 group had significant reductions in the mRNA expression levels of ASCL4 and PTGS2 and a significant increase in the mRNA expression level of GPX4 （all P<0.05）. Western blotting showed that compared with the Control group， the CCl4 group had a significant reduction in the protein expression level of GPX4 （P<0.05）， and compared with the CCl4 group， the MK-4+CCl4 group had a significant increase in the protein expression level of GPX4 （P<0.05）. Prussian blue staining showed that compared with the Control group， the CCl4 group had a significant increase in iron accumulation； after MK-4 intervention， compared with the CCl4 group， the MK-4+CCl4 group had a significant reduction in iron accumulation. As for the measurement of iron metabolism genes in mouse liver， compared with the Control group， the CCl4 group had a significant increase in iron content， significant reductions in the mRNA expression levels of FPN and HJV， and a significant increase in the mRNA expression level of TFR1 （all P<0.05）； after protection with MK-4， there was a significant reduction in iron content， significant increases in the mRNA expression levels of FPN and HJV， and a significant reduction in the mRNA expression level of TFR1 （all P<0.05）. Conclusion MK-4 intervention in advance can alleviate CCl4-induced ALI in mice， possibly by inhibiting ferroptosis and improving the expression of iron metabolism-related genes in mouse liver. [ABSTRACT FROM AUTHOR]

Long-term usage of bisphosphonates, especially zoledronic acid (ZA), induces osteogenesis disorders and medication-related osteonecrosis of the jaw (MRONJ) in patients, thereby contributing to the destruction of bone remodeling and the continuous progression of osteonecrosis. Menaquinone-4 (MK-4), a specific vitamin K 2 isoform converted by the mevalonate (MVA) pathway in vivo , exerts the promotion of bone formation, whereas ZA administration suppresses this pathway and results in endogenous MK-4 deficiency. However, no study has evaluated whether exogenous MK-4 supplementation can prevent ZA-induced MRONJ. Here we showed that MK-4 pretreatment partially ameliorated mucosal nonunion and bone sequestration among ZA-treated MRONJ mouse models. Moreover, MK-4 promoted bone regeneration and inhibited osteoblast apoptosis in vivo. Consistently, MK-4 downregulated ZA-induced osteoblast apoptosis in MC3T3-E1 cells and suppressed the levels of cellular metabolic stresses, including oxidative stress, endoplasmic reticulum stress, mitochondrial dysfunction, and DNA damage, which were accompanied by elevated sirtuin 1 (SIRT1) expression. Notably, EX527, an inhibitor of the SIRT1 signaling pathway, abolished the inhibitory effects of MK-4 on ZA-induced cell metabolic stresses and osteoblast damage. Combined with experimental evidences from MRONJ mouse models and MC3T3-E1 cells, our findings suggested that MK-4 prevents ZA-induced MRONJ by inhibiting osteoblast apoptosis through suppression of cellular metabolic stresses in a SIRT1-dependent manner. The results provide a novel translational direction for the clinical application of MK-4 for preventing MRONJ. [Display omitted] • Menaquinone-4 prevents medication-related osteonecrosis of the jaw caused by zoledronic acid. • Menaquinone-4 alleviates zoledronic acid-induced osteoblast apoptosis and osteogenesis obstacles. • Medication-related osteonecrosis of the jaw possesses multiple cellular metabolic stresses in osteoblasts. • Menaquinone-4 relieves oxidative stress, ER stress, mitochondrial dysfunction, and DNA damage via sirtuin 1 signaling. [ABSTRACT FROM AUTHOR]

The effect of vitamin K on bovine endometrial epithelial cells has not been thoroughly investigated. The objective of this study was to examine the effect of the biologically active form of vitamin K, menaquinone-4, on gene expression in bovine endometrial epithelial cells. First, we examined the mRNA and protein expression levels of UBIAD1, a menaquinone-4 biosynthetic enzyme. Second, we screened for potential target genes of menaquinone-4 in bovine endometrial epithelial cells using RNA-sequencing. We found 50 differentially expressed genes; 42 were upregulated, and 8 were downregulated. Among them, a dose-dependent response to menaquinone-4 was observed for the top three upregulated (TRIB3 , IL6 , and TNFAIP3) and downregulated (CDC6 , ORC1 , and RRM2) genes. It has been suggested that these genes play important roles in reproductive events. In addition, GDF15 and VEGFA , which are important for cellular functions as they are commonly involved in pathways, such as positive regulation of cell communication, cell differentiation, and positive regulation of MAPK cascade, were upregulated in endometrial epithelial cells by menaquinone-4 treatment. To the best of our knowledge, this is the first study showing the expression of UBIAD1 in the bovine uterus. Moreover, the study determined menaquinone-4 target genes in bovine endometrial epithelial cells, which may positively affect pregnancy with alteration of gene expression in cattle uterus. • UBIAD1, a menaquinone-4 (MK-4) biosynthetic enzyme, is expressed in bovine uterine tissues. • A total of 50 potential MK-4 target genes were found by RNA-sequencing. • No cytotoxicity was identified with MK-4 treatment. • GDF15 and VEGFA were upregulated in the uterine endometrial epithelial cells by MK-4 treatment. [ABSTRACT FROM AUTHOR]

Osteoporosis is a progressive metabolic disease characterized by decreased bone mineral density and increased fracture risk. Previous studies have shown that higher intake of vitamin K (VK) correlates with a reduced risk of osteoporosis. However, the effect of menaquinone-4 (MK-4), a specific form of VK, still remains obscure. Therefore, in this study, we investigated the effects of MK-4 on osteoclast differentiation by differentiating RAW 264.7 cells into osteoclasts with the help of receptor activator of nuclear factor-kappa B ligand (RANKL), assessed the mRNA expression of osteoclast-specific genes, and studied the effects of MK-4 in vivo in ovariectomized mice, a postmenopausal osteoporosis murine model. MK-4 inhibited osteoclast differentiation, decreased the mRNA expression of nuclear factor of activated T cells c1 (NFATc1), osteoclast-associated receptor (OSCAR), and cathepsin K (CTSK), and inhibited bone loss in ovariectomized mice. The findings strongly suggest that MK-4 is a therapeutic alternative for postmenopausal osteoporosis. [ABSTRACT FROM AUTHOR]

BackgroundThe relationship between vitamin intake and depression has attracted increasing attention. However, several studies examining such relationship among populations at different age groups have produced inconsistent findings. This study was aimed to investigate the cross-sectional association between vitamin K intake and depressive symptoms in US adults.MethodsWe used the data from a nationally representative sample of 11,687 adults from the 2013 to 2018 National Health and Nutrition Examination Survey (NHANES). Vitamin K intake was assessed by the 24-h dietary recall at the first day. Depressive symptoms were assessed using the 9-item Patient Health Questionnaire (PHQ-9). Logistic regression and generalized additive model were used to examine the association between vitamin K intake and depressive symptoms.ResultsThe weighted prevalence of depressive symptoms was 10.2% (8.0% in men and 12.0% in women). We observed a significant inverse linear relationship between vitamin K intake and depressive symptoms in models adjusted for age, sex, race/ethnicity, marital status, educational status, family poverty income ratio (PIR), home status, body mass index (BMI), smoking status, physical activity, sleep disorders, hypertension, hyperlipidemia, and diabetes. The odds ratios (OR) (95% CI) for the highest compared with the lowest quartile of vitamin K intake was 0.68 (95% CI: 0.52, 0.89, p-trend < 0.05). The association was similar in subgroups stratified by age, sex, race/ethnicity, marital status, educational status, PIR, home status, BMI, smoking status, physical activity, sleep disorders, hypertension, hyperlipidemia, and diabetes.ConclusionVitamin K intake was inversely and independently associated with the odds of depressive symptoms in the US adults. Prospective studies are warranted to confirm our findings.

Background: In Japan, three doses of vitamin K are administered to neonates as prophylactic regimens against vitamin K deficiency bleeding (VKDB). In this study, we aimed to evaluate the efficacy of this prophylactic vitamin K regimen using the hepaplastin test (HPT) performed one, two weeks, and one month after birth. The secondary aim of this study was to evaluate the utility of HPT screening in healthy neonates., Method: This study included a retrospective analysis of HPT values in neonates born between June 2009 and February 2018 using the prophylactic regimen implemented in 2011., Results: The study group included 6075 neonates, of whom 274 (4.5%) had a low HPT value (<40%) at the time of discharge (approximately one week after birth). Follow-up HPT was performed in 118 neonates at two weeks, with a low HPT value persisting in 11 neonates (9%). There was no effect of breast or formula milk on HPT values, and all neonates achieved an HPT value >40% at one month, regardless of whether vitamin K supplementation was provided at two weeks. None of the infants had underlying diseases that led to secondary VKDB., Conclusion: Healthy newborns maintained adequate HPT values with the triple-dose vitamin K administration, regardless of the feeding method. Therefore, HPT screening might not be essential for healthy neonates., Competing Interests: Human subjects: Consent for treatment and open access publication was obtained or waived by all participants in this study. The Institutional Review Board of Fuji City General Hospital issued approval #179. All procedures followed were in accordance with the ethical standards of the responsible committee on human experimentation (institutional and national) and with the Helsinki Declaration of 1964 and later versions. Ethical aspects of this study were reviewed and approved by the Institutional Review Board of Fuji City General Hospital in 2018 (#179). Animal subjects: All authors have confirmed that this study did not involve animal subjects or tissue. Conflicts of interest: In compliance with the ICMJE uniform disclosure form, all authors declare the following: Payment/services info: All authors have declared that no financial support was received from any organization for the submitted work. Financial relationships: All authors have declared that they have no financial relationships at present or within the previous three years with any organizations that might have an interest in the submitted work. Other relationships: All authors have declared that there are no other relationships or activities that could appear to have influenced the submitted work., (Copyright © 2024, Matsuoka et al.)

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Background: Vitamin K is a term that comprises a family of structurally related quinones, phylloquinone (PK) and the menaquinones (MKn), that share a common naphthoquinone ring but vary in sidechain length (n) and saturation. Dietary PK is a biosynthetic precursor to tissue menaquinone-4 (MK4), but little is known about the absorption and metabolism of dietary MKn.Objective: To characterize the absorption and metabolism of dietary MKn relative to PK.Methods: In the 4-week diet study, 10-week-old male and female C57BL/6 mice were pair-fed a vitamin K deficient diet (control) or a diet supplemented with 5.0 μmol/kg total PK, MK4, and/or MK9 (separately and in combination). In the 1-week stable isotope study, 12-week-old mice were pair-fed diets containing 2.2 μmol/kg PK (unlabeled control), 2H7PK, 13C11MK4, 2H7MK7, or 2H7MK9. Vitamin K tissue content was quantified by HPLC and/or LC-MS, and concentrations were compared by sex and diet group using 2-factor ANOVA.Results: Regardless of the form(s) of vitamin K provided in the diet, tissue MK4 concentrations did not differ across equimolar supplemented groups in the kidney, adipose, reproductive organ, bone, or pancreas in either males or females in the diet study (all P values > 0.05). Isotopic labeling confirmed the naphthoquinone ring of MK4 in tissues originated from the administered dietary PK or MKn. Despite equimolar supplementation, accumulation of the administered dietary form differed across diet groups in small intestinal segments (all P values < 0.002) and the liver (P < 0.001). Female mice had greater total vitamin K than males in every tissue examined (P < 0.05).Conclusions: Dietary PK, MK4, MK7, and MK9 all served as precursors to tissue MK4 in mice. This study expands our understanding of vitamin K metabolism and supports a common conversion mechanism of all dietary vitamin K forms to MK4. Further investigation of the metabolism and physiological roles of MK4 that may be independent of classical vitamin K function is warranted. [ABSTRACT FROM AUTHOR]

The autosomal dominant disorder Schnyder corneal dystrophy (SCD) is caused by mutations in UbiA prenyltransferase domain-containing protein-1 (UBIAD1), which uses geranylgeranyl pyrophosphate (GGpp) to synthesize the vitamin K2 subtype menaquinone-4 (MK-4). SCD is characterized by opacification of the cornea, owing to aberrant build-up of cholesterol in the tissue. We previously discovered that sterols stimulate association of UBIAD1 with ER-localized HMG-CoA reductase, which catalyzes a rate-limiting step in the synthesis of cholesterol and nonsterol isoprenoids, including GGpp. Binding to UBIAD1 inhibits sterol-accelerated ER-associated degradation (ERAD) of reductase and permits continued synthesis of GGpp in cholesterol-replete cells. GGpp disrupts UBIAD1-reductase binding and thereby allows for maximal ERAD of reductase as well as ER-to-Golgi translocation of UBIAD1. SCD-associated UBIAD1 is refractory to GGpp-mediated dissociation from reductase and remains sequestered in the ER to inhibit ERAD. Here, we report development of a biochemical assay for UBIAD1-mediated synthesis of MK-4 in isolated membranes and intact cells. Using this assay, we compared enzymatic activity of WT UBIAD1 with that of SCD-associated variants. Our studies revealed that SCD-associated UBIAD1 exhibited reduced MK-4 synthetic activity, which may result from its reduced affinity for GGpp. Sequestration in the ER protects SCD-associated UBIAD1 from autophagy and allows intracellular accumulation of the mutant protein, which amplifies the inhibitory effect on reductase ERAD. These findings have important implications not only for the understanding of SCD etiology but also for the efficacy of cholesterol-lowering statin therapy, which becomes limited, in part, because of UBIAD1-mediated inhibition of reductase ERAD.

The effect of dietary vitamin K3 (VK3) on ruminant animals is not fully investigated. The aim of this study was to examine the effects of dietary VK3 on lactation performance, rumen characteristics, and VK1 and menaquinone (MK, or VK2) dynamics in the rumen, plasma, and milk of dairy cows. Eight Holstein dairy cows in late lactation periods were used in two crossover trials including a control (nontreatment) and a 50 or 200 mg/day (d) VK3 supplementation group. After 14 days, plasma, ruminal fluid, and milk were sampled and their VK1 and MKs contents were measured using fluorescence-high-performance liquid chromatography (HPLC). Milk production was unchanged after feeding 50 mg/day VK3 but marginally decreased after feeding 200 mg/day VK3. The molar ratio of propionate in ruminal fluid was significantly increased on feeding 200 mg/day VK3. Additionally, MK-4 concentrations significantly increased in both plasma and milk after VK3 feeding (50 and 200 mg/day). In ruminal fluid, MK-4 concentrations increased after 200 mg/day VK3 feeding. These results suggest that VK3 may be a good source of MK-4, the biologically active form of VK, in Holstein dairy cows during their late lactation periods. This study provides a basis for understanding the physiological role of VK in dairy cows. [ABSTRACT FROM AUTHOR]

Objective: The vitamin K family has a wide range of effects in the body, including the central nervous system. Menaquinone-4 (MK-4), a form of vitamin K2, is converted from phylloquinone (PK), which is the main source of dietary vitamin K and is the main form of vitamin K in the brain. We conducted this study to investigate the serum concentration of MK-4 and the correlations between MK-4 and developmental quotients in children with autism spectrum disorder (ASD).Methods: We selected 731 children with ASD who were diagnosed for the first time. During the same period, 332 neurotypical children who underwent regular physical examinations in our outpatient department were selected as the TD group. We investigated the general situation of children, including gender and age. Children in ASD group were assessed for autistic symptoms and development quotients, including Autism Behavior Checklist (ABC), Childhood Autism Rating Scale (CARS), ADOS-2, and Griffiths Development Scales-Chinese Language Edition (GDS-C). Both groups of children were tested for serum menaquinone-4. We compared serum menaquinone-4 levels of ASD group and TD group. We then conducted a correlation analysis between the level of menaquinone-4 and the developmental quotient of children with ASD.Results: The results of this study indicate that the serum concentration of MK-4 in children with ASD is lower than that in children with typical development (t = −2.702, P = 0.007). The serum concentration of MK-4 is related to the developmental quotients of several subscales in ASD children, and this correlation is more obvious in males.Conclusion: we conclude that MK-4 is present in lower concentrations in children with ASD, which may affect cognition and developmental quotients. The role of MK-4 in ASD needs to be further explored.

Vitamin K possesses efficacy as a topical dermatological agent. However, vitamin K is phototoxic and susceptible to photodegradation. Herein, we investigated the mechanisms underlying the phototoxicity of phylloquinone (PK, vitamin K 1) and menaquinone-4 (MK-4, vitamin K 2) under ultraviolet A (UVA) irradiation using various reactive oxygen species (ROS) scavengers. This resulted in the production of superoxide anion radicals via type I and singlet oxygen via type II photodynamic reactions, which were quenched by the ROS scavengers: superoxide dismutase and sodium azide (NaN 3). In HaCaT cells, MK-4 and PK induced the production of intracellular ROS, particularly hydrogen peroxide, in response to UVA irradiation. Furthermore, the addition of catalase successfully decreased maximum ROS levels by approximately 30%. NaN 3 and catalase decreased the maximum reduction in cell viability induced by UVA-irradiated PK and MK-4 in cell viability by approximately 2–7-fold. Additionally, ROS scavengers had no effect on the photodegradation of PK or MK-4 at 373 nm. Therefore, the phototoxicities of PK and MK-4 were attributed to the generation of singlet oxygen and hydrogen peroxide, underscoring the importance of photoshielding in circumventing phototoxicity. • UVA irradiated vitamin K induces typeIand typeIIphototoxic reactions. • The phototoxicity of vitamin K on HaCaT cells was suppressed by NaN 3 and catalase. • The phototoxicity of vitamin K attributed to the generation of singlet oxygen and hydrogen peroxide. • The ROS generated during the photochemical reaction had no impact on the photodegradation of vitamin K. [ABSTRACT FROM AUTHOR]

The effects of vitamin K (VK) on immune cells in ruminants are yet to be fully investigated. The objective of this study was to examine the effects of VK on peripheral blood mononuclear cells (PBMC) in Holstein dairy cows. A cell proliferation assay was performed to evaluate the effect of menaquinone-4 (MK-4, the biologically active form of VK) on immune response of PBMC. The proliferation of PBMC stimulated by MK-4 was significantly higher than that of nonstimulated controls. The expression of T cell-related genes in PBMC, stimulated with MK-4, was assessed by quantitative PCR. No significant changes were observed in the mRNA expression levels of both CD4 and CD8 as helper T cell and cytotoxic T cell markers, respectively. The present study demonstrated that MK-4 positively influenced cow PBMC proliferation and suggested the possibility of bovine-specific immune cell activation. The present study lays a foundation for understanding the physiological role of VK in cattle. [ABSTRACT FROM AUTHOR]

There is variability in the reported effects of compounds on osteoblasts arising from differences in experimental design and choice of cell type/origin. This makes it difficult to discern a compound's action outside its original study and compare efficacy between compounds. Here, we investigated five compounds frequently reported as anabolic for osteoblasts (17β-estradiol (oestrogen), icariin, lactoferrin, lithium chloride, and menaquinone-4 (MK-4)) on human mesenchymal progenitors to assess their potential for bone tissue engineering with the aim of identifying a potential alternative to expensive recombinant growth factors such as bone morphogenetic protein 2 (BMP-2). Experiments were performed using the same culture conditions to allow direct comparison. The concentrations of compounds spanned two orders of magnitude to encompass the reported efficacious range and were applied continuously for 22 days. The effects on the proliferation (resazurin reduction and DNA quantification), osteogenic differentiation (alkaline phosphatase (ALP) activity), and mineralised matrix deposition (calcium and collagen quantification) were assessed. Of these compounds, only 10 µM MK-4 stimulated a significant anabolic response with 50% greater calcium deposition. Oestrogen and icariin had no significant effects, with the exception of 1 µM icariin, which increased the metabolic activity on days 8 and 22. 1000 µg/mL of lactoferrin and 10 mM lithium chloride both significantly reduced the mineralised matrix deposition in comparison to the vehicle control, despite the ALP activity being higher in lithium chloride-treated cells at day 15. This demonstrates that MK-4 is the most powerful stimulant of bone formation in hES-MPs of the compounds investigated, highlighting its potential in bone tissue engineering as a method of promoting bone formation, as well as its prospective use as an osteoporosis treatment. [ABSTRACT FROM AUTHOR]

Dietary vitamin K 1 (phylloquinone: PK) and menaquinone (MK-n) are converted to menadione (MD) in the small intestine and then translocated to various tissues where they are converted to vitamin K 2 (menaquinone-4: MK-4) by UbiA prenyltransferase domain containing protein 1 (UBIAD1). MK-4 is effective in bone formation and is used to treat osteoporosis in Japan. UBIAD1 is expressed in bone and osteoblasts and shows conversion to MK-4, but the role of UBIAD1 in osteogenesis is unknown. In this study, we investigated the function of UBIAD1 in osteogenesis using a tamoxifen-dependent UBIAD1-deficient mouse model. When UBIAD1 deficiency was induced from the first week of life, the femur was significantly shortened, and bone mineral density (BMD) was reduced. In addition, the expression of bone and chondrocyte matrix proteins and chondrocyte differentiation factors was significantly decreased. In primary cultured chondrocytes, chondrocyte differentiation was significantly reduced by UBIAD1 deficiency. These results suggest that UBIAD1 is an important factor for the regulation of chondrocyte proliferation and differentiation during osteogenesis. • Vitamin Ks are converted to menaquinone-4 in the body. • The vitamin K-converting enzyme is UBIAD1. • Vitamin K is important for bone formation. • UBIAD1 has important functions in bone formation, especially in cartilage formation. [ABSTRACT FROM AUTHOR]

The content and composition of dietary supplements is of great interest due to their increasing consumption and variety of available brand offered in the market. Accurate determination of vitamins is important for the improvement of dietary supplement quality and nutrition assessments. In this regard, the simultaneous determination of vitamin D3 (calcitriol—CT and cholecalciferol—CHL) and K2 (menaquinone-4—MK-4 and menaquinone-7—MK-7) in dietary supplements was developed by using ultra-high-pressure liquid chromatography (UHPLC). The overall runtime per sample was above 35 min, with the retention times of 2.40, 6.59, 7.06, and 32.6 min for vitamin D3 (CT and CHL) and vitamin K2 (MK-4 and MK-7), respectively. The limits of detection and limits of quantification for the target nutritional compounds ranged between 0.04–0.05 µg/mL, respectively. The validation results indicated that the method had reasonable linearity (R2 ≥ 0.9990), good recovery (>82%), satisfactory intra-day precision (≤1.9%) and inter-day precision (≤3.5%), and high selectivity and specificity. The validated UHPLC method was demonstrated to be precise, accurate, and robust for the simultaneous determination of vitamins D3 (CT and CHL) and K2 (MK-4 and MK-7) in dietary supplements.

Objective: The present study is aimed to develop and validate an Ultra-Fast Liquid Chromatography-Diode Array Detector (UFLC-DAD) method for the quantification of vitamin K2 as Menaquinone-4 (MK-4) in rabbit plasma. Method: Standard solutions and spiked plasma samples of MK-4 and Internal Standard (IS) were prepared from primary stock solutions of 1mg/ml concentration in ethanol each. Protein precipitation was carried out for the MK-4 and IS extraction from plasma spiked samples. Chromatographic separation was employed using Isopropyl Alcohol and Acetonitrile (50:50 v/v) as mobile phase and a C-18 column with 1ml/min flow rate and a run time of 10 mins. Detection was carried out in the range 190-600 nm with 269 nm set as a reference wavelength. Result: The retention times of MK-4 and IS were at 5.5 ± 0.5 mins and 8 ± 0.5 mins respectively. Calibration curve for MK-4 was found to be linear in the range of 0.374 to 6 µg/ml with an R2 value of 0.9934. The % RSD for accuracy was <15%, inter and intraday precisions were <10%. The samples were found to be stable throughout the study. Conclusion: This method can be applied to the estimation of MK-4 in rabbit plasma using UFLC-DAD. [ABSTRACT FROM AUTHOR]

Purpose: The aim of this study was to evaluate the effect of combining human parathyroid hormone (1-34) (PTH1-34; PTH) and menaquinone-4 (MK-4) on calvarial bone defect repair in osteopenic rats.Methods: Fourteen week olds were subject to craniotomy for the establishment of osteopenic animal models fed through a chronically low-protein diet. After that, critical calvarial defect model was established and all rats were randomly divided into four groups: sham, MK-4, PTH, and PTH + MK-4. The animals received MK-4 (30 mg/kg/day), PTH1-34 (60 μg/kg, three times a week), or PTH1-34 (60 μg/kg, three times a week) plus MK-4 (30 mg/kg/day) for 8 weeks, respectively. Serum γ-carboxylated osteocalcin (Gla-OC) levels, histological and immunofluorescent labeling were employed to evaluate the bone formation and mineralization in calvarial bone defect. In addition, Microfil perfusion, immunohistochemical, and micro-CT suggested enhanced angiogenesis and bone formation in calvarial bone healing.Results: In this study, treatment with either PTH1-34 or MK-4 promoted bone formation and vascular formation in calvarial bone defects compared with the sham group. In addition, combined treatment of PTH1-34 plus MK-4 increased serum level of Gla-OC, improved vascular number and vascular density, and enhanced bone formation in calvarial bone defect in osteopenic conditions as compared with monotherapy.Conclusions: In summary, this study indicated that PTH1-34 plus MK-4 combination therapy accelerated bone formation and angiogenesis in calvarial bone defects in presence of osteopenia. [ABSTRACT FROM AUTHOR]

The absence of vitamin E from the diet can lead to cardiovascular disease, cancer, cataracts, and premature aging. Vitamin K deficiency can lead to bleeding disorders. These fat-soluble vitamins are important nutritional factors that can be determined in different methods in vegetables. In this work, the simultaneous determination of α-tocopherol, α-tocopheryl acetate, phylloquinone, and menaquinone-4 by gas chromatography–mass spectrometry (GC–MS) has been optimized using both direct injection and solid phase microextraction (SPME). Three different sample pre-treatment approaches based on: (A) solid–liquid–liquid–liquid extraction (SLE–LLE), (B) SLE, and (C) SPME were then applied to extract the target analytes from vegetables samples using menaquinone as internal standard. All the procedures allowed the determination of the target analytes in onion, carrot, celery, and curly kale samples. Similar results were obtained with the three different approaches, even if the one based on SPME offers the best performance, together with a reduced use of solvent, time consumption, and experimental complexity, which makes it the preferable option for industrial applications.

Initially discovered for its role in blood coagulation, there is now convincing evidence that vitamin K (VK) has important actions in the nervous system. In brain, VK is present in the form of menaquinone-4 (MK-4), a byproduct of the main dietary source, phylloquinone. It contributes to the biological activation of various proteins (i.e., Gas6), and participates in the synthesis of sphingolipids, a class of lipids widely present in brain cell membranes with important cell signaling functions. In a previous study, we reported that lifetime consumption of a low VK diet resulted in mild cognitive impairment in aged rats, a finding associated with an alteration of the sphingolipid profile. To confirm the role of VK as it relates to sphingolipids, cognition, and behavior outside the context of aging, we conducted a study of acute VK deficiency using a pharmacological model of VK deficiency in brain. In this procedure, rats (8 weeks) are maintained on a ratio of warfarin (a VK antagonist) to VK whereby coagulation is maintained while inducing VK deficiency in extrahepatic tissues. After 10 weeks of treatment, rats who were subjected to the warfarin plus phylloquinone protocol (WVK) exhibited longer latencies in the Morris water maze test as well as lower locomotor activity and exploratory behavior in the open field test, when compared to control rats. The WVK treatment resulted in a dramatic decrease in MK-4 level in all brain regions despite the presence of high local concentrations of phylloquinone, which suggests an inhibition of the biosynthetic MK-4 pathway in the presence of warfarin. Additionally, WVK treatment affected sphingolipid concentrations in key brain regions, notably those of the ganglioside family. Finally, brain MK-4 was correlated with performances in the open field test. This study confirms the modulatory role of VK in cognition and behavior and the implication of sphingolipids, notably those of the ganglioside family.

Recently, new therapeutics have been developed for hepatocellular carcinoma (HCC). However, the overall survival rate of HCC patients is still unsatisfactory; one of the reasons for this is the high frequency of recurrence after radical treatment. Consequently, to improve prognosis, it will be important to develop a novel anti-tumor agent that is especially effective against HCC recurrence. For clinical application, long-term safety, together with high anti-tumor efficacy, is desirable. Recent studies have proposed menahydroquinone-4 1,4-bis-N,N-dimethylglycinate hydrochloride (MKH-DMG), a prodrug of menahydroquinone-4 (MKH), as a promising candidate for HCC treatment including the inhibition of recurrence; MKH-DMG has been shown to achieve good selective accumulation of MKH in tumor cells, resulting in satisfactory inhibition of cell proliferation in des-γ-carboxyl prothrombin (DCP)-positive and DCP-negative HCC cell lines. In a spleen-liver metastasis mouse model, MKH-DMG has been demonstrated to have anti-proliferation and anti-metastatic effects in vivo. The characteristics of MKH-DMG as a novel anti-HCC agent are presented in this review article.

The objective of this study was to investigate the effects of human parathyroid hormone (1-34) (PTH1-34; PTH) plus menaquinone-4 (vitamin K2; MK) on the osseous integration of hydroxyapatite (HA)-coated implants in osteoporotic rats. Ovariectomized female Sprague-Dawley rats were used for the study. Twelve weeks after bilateral ovariectomy, HA-coated titanium implants were inserted bilaterally in the femoral medullary canal of the remaining 40 ovariectomized rats. All animals were then randomly assigned to four groups: Control, MK, PTH and PTH + MK. The rats from groups MK, PTH and PTH + MK received vitamin K2 (30 mg/kg/day), PTH1-34 (60 μg/kg, three times a week), or both for 12 weeks. Thereafter, serum levels of γ-carboxylated osteocalcin (Gla-OC) were quantitated by ELISA and the bilateral femurs of rats were harvested for evaluation. The combination of PTH and MK clearly increased the serum levels of Gla-OC (a specific marker for bone formation) compared to PTH or MK alone. The results of our study indicated that all treated groups had increased new bone formation around the surface of implants and increased push-out force compared to Control. In addition, PTH + MK treatment showed the strongest effects in histological, micro-computed tomography and biomechanical tests. In summary, our results confirm that treatment with PTH1-34 and MK together may have a therapeutic advantage over PTH or MK monotherapy on bone healing around HA-coated implants in osteoporotic rats. [ABSTRACT FROM AUTHOR]

Vascular calcification (VC) caused by chronic kidney disease (CKD)-mineral and bone disorder is a common complication of CKD. Recent studies have demonstrated that menaquinone-4 (MK-4) is negativly associated with VC in patients with CKD. Furthermore, we have previously shown that runt-related transcription factor 2 (Runx2) is important in the phenotypic transformation process of rat vascular smooth muscle cells (VSMCs), which is the key step for the development of VC. The present study investigated the influence of MK-4 on the phenotypic transformation process of rat VSMCs in order to illustrate its role in the process of VC. Calcification assays were perfomed to access the calcified degree of rat VSMCs. Additionally, the genes and proteins related to phenotypic transformation were measured by reverse transcription-polymerase chain reaction and western blotting methods. It was revealed that calcium deposition in the cells was evidently increased with an addition of β-glycerophosphate (β-GP) and could be completely prevented by co-incubation with MK-4 in a dose-dependent manner. Furthermore, the expression of Runx2 in the β-GP-induced VSMCs was inhibited by MK-4. It was also revealed that the expression of SMAD1 and bone morphogenetic protein (BMP)-2 were decreased in the β-GP-induced VSMCs treated with MK-4 in a dose-dependent manner; however, the expression of SMAD7 was increased in the β-GP-induced VSMCs treated with MK-4 in a dose-dependent manner. These observations suggest that MK-4 reduces mineralization by regulating the BMP-2 signaling pathway in order to attenuate the expression of Runx2. [ABSTRACT FROM AUTHOR]

Initially discovered for its role in blood coagulation, there is now convincing evidence that vitamin K (VK) has important actions in the nervous system. In brain, VK is present in the form of menaquinone-4 (MK-4), a byproduct of the main dietary source, phylloquinone. It contributes to the biological activation of various proteins (i.e., Gas6), and participates in the synthesis of sphingolipids, a class of lipids widely present in brain cell membranes with important cell signaling functions. In a previous study, we reported that lifetime consumption of a low VK diet resulted in mild cognitive impairment in aged rats, a finding associated with an alteration of the sphingolipid profile. To confirm the role of VK as it relates to sphingolipids, cognition, and behavior outside the context of aging, we conducted a study of acute VK deficiency using a pharmacological model of VK deficiency in brain. In this procedure, rats (8 weeks) are maintained on a ratio of warfarin (a VK antagonist) to VK whereby coagulation is maintained while inducing VK deficiency in extrahepatic tissues. After 10 weeks of treatment, rats who were subjected to the warfarin plus phylloquinone protocol (WVK) exhibited longer latencies in the Morris water maze test as well as lower locomotor activity and exploratory behavior in the open field test, when compared to control rats. The WVK treatment resulted in a dramatic decrease in MK-4 level in all brain regions despite the presence of high local concentrations of phylloquinone, which suggests an inhibition of the biosynthetic MK-4 pathway in the presence of warfarin. Additionally, WVK treatment affected sphingolipid concentrations in key brain regions, notably those of the ganglioside family. Finally, brain MK-4 was correlated with performances in the open field test. This study confirms the modulatory role of VK in cognition and behavior and the implication of sphingolipids, notably those of the ganglioside family. [ABSTRACT FROM AUTHOR]

There is variability in the reported effects of compounds on osteoblasts arising from differences in experimental design and choice of cell type/origin. This makes it difficult to discern a compound’s action outside its original study and compare efficacy between compounds. Here, we investigated five compounds frequently reported as anabolic for osteoblasts (17β-estradiol (oestrogen), icariin, lactoferrin, lithium chloride, and menaquinone-4 (MK-4)) on human mesenchymal progenitors to assess their potential for bone tissue engineering with the aim of identifying a potential alternative to expensive recombinant growth factors such as bone morphogenetic protein 2 (BMP-2). Experiments were performed using the same culture conditions to allow direct comparison. The concentrations of compounds spanned two orders of magnitude to encompass the reported efficacious range and were applied continuously for 22 days. The effects on the proliferation (resazurin reduction and DNA quantification), osteogenic differentiation (alkaline phosphatase (ALP) activity), and mineralised matrix deposition (calcium and collagen quantification) were assessed. Of these compounds, only 10 µM MK-4 stimulated a significant anabolic response with 50% greater calcium deposition. Oestrogen and icariin had no significant effects, with the exception of 1 µM icariin, which increased the metabolic activity on days 8 and 22. 1000 µg/mL of lactoferrin and 10 mM lithium chloride both significantly reduced the mineralised matrix deposition in comparison to the vehicle control, despite the ALP activity being higher in lithium chloride-treated cells at day 15. This demonstrates that MK-4 is the most powerful stimulant of bone formation in hES-MPs of the compounds investigated, highlighting its potential in bone tissue engineering as a method of promoting bone formation, as well as its prospective use as an osteoporosis treatment.

Vitamin K is an essential cofactor of γ-glutamylcarboxylase as related to blood coagulation and bone formation. Menaquinone-4, one of the vitamin K homologues, is biosynthesized in the body and has various biological activities such as being a ligand for steroid and xenobiotic receptors, protection of neuronal cells from oxidative stress, and so on. From this background, we focused on the role of menaquinone in the differentiation activity of progenitor cells into neuronal cells and we synthesized novel vitamin K derivatives with modification of the ω-terminal side chain. We report here new vitamin K analogues, which introduced an alkylated phenyl group at the ω-terminal side chain. These compounds exhibited potent differentiation activity as compared to control. [ABSTRACT FROM AUTHOR]

The aim of the present study was to determine the molecular basis of the beneficial effects of genistein and/or menaquinone-4 (MK-4) on bone quality. Initially, 1 μM genistein was applied to MC3T3-E1 cells for 24 h and the upregulated mRNAs that were detected by microarray were selected for further examination by reverse transcription-quantitative-polymerase chain reaction. Among them, alterations were observed in the level of GATA-binding protein 6 (GATA6), Notch gene homolog 2 (NOTCH2), Wnt family member 5A (WNT5A), bone γ-carboxyglutamate protein (BGLAP), chondroadherin (CHAD), dipeptidyl peptidase 4 (DPP4), ectonucleotide pyrophosphatase/phosphodiesterase 2 (ENPP2), alkaline phosphatase (ALP) 3 and ATPase phospholipid-transporting 11A (ATP11A) in response to treatment with 0.1 μM 17-β-estradiol, 1 μM genistein, and/or 1 μM MK-4. GATA6, NOTCH2 and WNT5A are considered to be associated with osteoclast, but not osteoblast, function; however, increases in osteoblastic mRNAs, including BGLAP and CHAD, were observed in each of the treatment groups at 48 h. Immunocytochemical analysis confirmed an increase in CHAD and DPP4 proteins following the administration of genistein + MK-4. Furthermore, genistein + MK-4 led to alterations in cell morphology to spindle or oval shapes, and increased the intensity of ALP staining. Although the level of ALP mRNA was not consistently altered in response to the treatments, a marked increase in ALP activity was observed following 96 h treatment with genistein + MK-4. Therefore, the simultaneous intake of genistein and MK-4 appears to be beneficial for the maintenance of bone quality. [ABSTRACT FROM AUTHOR]

K vitamins are well known as essential cofactors for hepatic γ-carboxylation of coagulation factors, but their potential role in chronic diseases including cancer is understudied. K2, the most abundant form of vitamin K in tissues, exerts anti-cancer effects via diverse mechanisms which are not completely understood. Our studies were prompted by previous work demonstrating that the K2 precursor menadione synergized with 1,25 dihydroxyvitamin D3 (1,25(OH) 2 D 3) to inhibit growth of MCF7 luminal breast cancer cells. Here we assessed whether K2 modified the anti-cancer effects of 1,25(OH) 2 D 3 in triple negative breast cancer (TNBC) cell models. We examined the independent and combined effects of these vitamins on morphology, cell viability, mammosphere formation, cell cycle, apoptosis and protein expression in three TNBC cell models (MDA-MB-453, SUM159PT, Hs578T). We found that all three TNBC cell lines expressed low levels of the vitamin D receptor (VDR) and were modestly growth inhibited by 1,25(OH) 2 D 3 in association with cell cycle arrest in G0/G1. Induction of differentiated morphology by 1,25(OH) 2 D 3 was observed in two of the cell lines (MDA-MB-453, Hs578T). Treatment with K2 alone reduced viability of MDA-MB-453 and SUM159PT cells but not Hs578T cells. Co-treatment with 1,25(OH) 2 D 3 and K2 significantly reduced viable cell number relative to either treatment alone in Hs578T and SUM159PT cells. The combination treatment induced G0/G1 arrest in MDA-MB-453 cells, Hs578T and SUM159PT cells. Combination treatment altered mammosphere size and morphology in a cell specific manner. Of particular interest, treatment with K2 increased VDR expression in SUM159PT cells suggesting that the synergistic effects in these cells may be secondary to increased sensitivity to 1,25(OH) 2 D 3. The phenotypic effects of K2 in TNBC cells did not correlate with γ-carboxylation suggesting non-canonical actions. In summary, 1,25(OH) 2 D 3 and K2 exert tumor suppressive effects in TNBC cells, inducing cell cycle arrest leading to differentiation and/or apoptosis depending on the specific cell line. Further mechanistic studies to clarify common and unique targets of these two fat soluble vitamins in TNBC are warranted. • Vitamin K2 (menaquinone-4) exerts tumor suppressive effects in three triple negative breast cancer cell lines in vitro. • Co-treatment of 1,25(OH) 2 D 3 with K2 produces additive or synergistic effects depending on cell line. • Cell cycle arrest, differentiated morphology and/or loss of cell viability were induced by combination treatment. • 1,25(OH) 2 D 3 plus K2, individually and in combination, alter mammosphere morphology, self-renewal and stem cell. [ABSTRACT FROM AUTHOR]

Objective: The aim of this study was to determine the biological effects of natural dietary intake of vitamin K as phylloquinone (K1) and menaquinone-4 (MK-4) and a control diet also containing menadione (K3) on levels of K1 and total MK-4 (menaquinone-4) and menaquinone-4-2,3-epoxide (MK-4O)) in liver and pancreas, and on femur bending resistance in a fast-growing animal model. Design: Chickens were fed four wheat-based diets from day 11 to day 22 after hatching. The diets contained different combinations of fat sources: rapeseed oil, animal rendered fat, soybean oil and hydrogenated soybean oil. Concentration of K1 in the three experimental diets was 120 ng/g whereas MK-4 levels were 23, 52 and 63 ng/g respectively. The control diet contained 157 ng K1/g, 75 ng MK-4/g and 2.250 ng K3/g. Results: Growth rates and femur strength confirmed adequate supply of nutrients and vitamin K in the test groups. There were no significant differences in femur bending resistance among the test groups, but these were higher than the control. K1, MK-4 and MK-4O were found in liver. In pancreas, mainly MK-4O was found with small amounts of MK-4, but none had content of K1. In the test groups the hepatic levels of MK-4 and MK-4O reflected the dietary intake of MK-4. Conclusion: The chickens were in good health with good bone resistance without supplements of K3 in the feed, but at least a natural content of 23 ng MK-4/g feed. Liver and pancreas appears to use MK-4 in different ways.

Hepatocellular carcinoma (HCC) shows poor prognosis owing to its very frequent recurrence even after curative treatment. Thus, an effective and safe long-term chemopreventive agent is strongly in demand. Menahydroquinone-4 (MKH) is an active form of menaquinone-4 (MK-4, vitamin K2) that is involved in the synthesis of vitamin K-dependent proteins in the liver. We hypothesized that efficient delivery of MKH might be critical to regulate HCC proliferation. The discovery of a suitable prodrug targeting HCC in terms of delivery and activation could reduce the clinical dose of MK-4 and maximize efficacy and safety. We previously showed that MKH dimethylglycinate (MKH-DMG) enables effective delivery of MKH into HCC cells and exhibits strong antitumor effects compared with MK-4. In this study, we prepared anionic MKH hemi-succinate (MKH-SUC) and non-ionic MKH acetate (MKH-ACT), in addition to cationic MKH-DMG, and evaluated MKH delivery profiles and antitumor effects in vitro. MKH-SUC showed the highest uptake and the most efficient release of MKH among the examined compounds and exhibited rapid and strong antitumor effects. These results indicate that MKH-SUC might have a good potential as an MKH delivery system for HCC that overcomes the limitations of MK-4 as a clinical chemopreventive agent.

• Vitamin K 2 derivatives with a ketone group at the C-1′ position of the side chain were synthesized. • Conversion amount of the vitamin K 2 derivatives to MK-4 was less than that of natural vitamin K 2 homologues. • Conversion amount of the vitamin K 1 derivative with only a ketone group at the C-1′ position to MK-4 was almost same as that of natural vitamin K 1. Prior to being utilized in the body, dietary vitamin K homologues are converted to menaquinone-4 (MK-4) by cleavage of the side chain part followed by prenylation. We predicted that the prenylation would occur due to the electron deficiency at the C-1′ position of the allyl moiety. Therefore, as an alternative method to make the C-1′ position electron-deficient, a new vitamin K derivative was synthesized by introducing a ketone group, and its conversion to MK-4 was investigated. Introduction of a ketone group at the C-1′ position of the side chain of 2′,3′-dihydrophylloquinone, which is known to resist conversion to MK-4, induced conversion to the MK-4 analog to a degree similar to that of natural phylloquinone. Thus, the electron deficiency at the C-1′ position is important for the conversion to MK-4. [Display omitted] [ABSTRACT FROM AUTHOR]

Alkaline phosphatase (ALP) hydrolyzes several monophosphate esters into inorganic acid and alcohol. In humans, 4 kinds of ALP isozymes have been identified: tissue-nonspecific ALP, intestinal ALP, placental ALP, and germ cell ALP. Intestinal ALP is expressed at a high concentration in the brush border membrane of intestinal epithelial cells and is known to be affected by several kinds of nutrients, such as lipids, but the physiological function of intestinal ALP has remained elusive. Vitamin K is an essential cofactor for the posttranslational carboxylation of glutamate residues into γ-carboxy glutamate. Menaquinone-4 (MK-4) with 4 isoprene units, vitamin K 2 , has been shown to induce bone-type ALP activity and osteoblastogenesis in human bone marrow cells. In this study, we investigated the effects of MK-4 on the level of ALP activity and expression of ALP messenger RNA in the human colon carcinoma cell line Caco-2, which is known to differentiate into small intestinal epithelial cells in vitro . After treatment with MK-4, there were significant increases in the ALP activities of Caco-2 cells. Inhibitor and thermal inactivation experiments demonstrated that the increased ALP had properties of intestinal-type ALP. Semiquantitative reverse transcription–polymerase chain reaction analysis revealed that expressions of human intestinal ALP and sucrase-isomaltase, which are intestinal differentiation markers, were highly enhanced in Caco-2 cells by MK-4. This is the first report concerning ALP messenger RNA expression induced by vitamin K 2 in Caco-2 cells. Further studies on the physiological functions of human intestinal ALP will provide useful data on the novel effects of vitamin K. [ABSTRACT FROM AUTHOR]

Background: There has been limited characterization of biological variables that impact vitamin K metabolism. This gap in knowledge can limit the translation of data obtained from preclinical animal studies to future human studies.Objective: The purpose of this study was to determine the effects of diet, sex, and housing on serum, tissue, and fecal vitamin K concentrations and gene expression in C57BL6 mice during dietary vitamin K manipulation.Methods: C57BL6 4-mo-old male and female mice were randomly assigned to conventional or suspended-wire cages and fed control [1400 ± 80 μg phylloquinone (PK)/kg] or deficient (31 ± 0.45 μg PK/kg) diets for 28 d in a factorial design. PK and menaquinone (MK) 4 plasma and tissue concentrations were measured by HPLC. Long-chain MKs were measured in all matrices by LC-atmospheric pressure chemical ionization-mass spectrometry. Gene expression was quantified by reverse transcriptase-polymerase chain reaction in the liver, brain, kidney, pancreas, and adipose tissue.Results: Male and female mice responded differently to dietary manipulation in a tissue-dependent manner. In mice fed the control diet, females had ∼3-fold more MK4 in the brain and mesenteric adipose tissue than did males and 100% greater PK concentrations in the liver, kidney, and mesenteric adipose tissue than did males. In mice fed the deficient diet, kidney MK4 concentrations were ∼4-fold greater in females than in males, and there were no differences in other tissues. Males and females differed in the expression of vitamin K expoxide reductase complex 1 (Vkorc1) in mesenteric adipose tissue and the pancreas and ubiA domain-containing protein 1 (Ubiad1) in the kidney and brain. There was no effect of housing on serum, tissue, or fecal concentrations of any vitamin K form.Conclusions: Vitamin K concentrations and expression of key metabolic enzymes differ between male and female mice and in response to the dietary PK concentration. Identifying factors that may impact study design and outcomes of interest is critical to optimize study parameters examining vitamin K metabolism in animal models. [ABSTRACT FROM AUTHOR]

Human milk is the only source of vitamin K for exclusively breastfed neonates. This vitamin is crucial both for blood coagulation (vitamin K 1 ) and for the normal neurological and skeletal development of the foetus and newborn (vitamin K 2 ). Since vitamin K is ubiquitous in foods, deficiency is not common in adults, but plasma levels and hepatic storage are very low at birth. In light of the importance of this valuable micronutrient, a non-invasive method for verifying that exclusively breastfed infants are receiving an adequate supply of the vitamin is clearly a topic of great significance. In spite of this, the determination of the several vitamin K homologues in human milk has still not been completely elucidated. This paper presents an HPLC–MS/MS method for the simultaneous determination of phylloquinone, menaquinone-4 (MK-4), and menaquinone-7 (MK-7) in human milk after a simple and effective isolation procedure. Overnight cold saponification and extraction of the analytes with hexane provided yields above 75%. This procedure, combined with high performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS), made it possible to achieve limits of detection (LODs) below 0.8 ng/mL. After a complete validation study, the method was applied to measure vitamin K congeners in several human milk samples; results showed vitamin K 1 concentrations comparable with those reported in the literature. In addition, this is the first study performed for the determination of MK-4 and MK-7 in the maternal milk of Italian women. [ABSTRACT FROM AUTHOR]

Whether through the vitamin K-dependent proteins or the individual K vitamers, vitamin K (VK) is associated with a number of age-related conditions (e.g., osteoporosis, atherosclerosis, insulin resistance, cognitive decline). In light of this, we investigated the influence of lifetime dietary VK exposure on the tissue distribution of phylloquinone (K1) and menaquinone-4 (MK-4) vitamers in 3-, 12- and 22-month-old male and female rats fed different K1 diets since weaning or subjected to a 40% calorie restricted diet (CR) since adulthood. Dietary K1 intakes around the minimal amount required for normal blood coagulation had no significant influence on body weights of both male and female rats at different life stages. Tissue contents of the K vitamers differed according to organs, were generally higher in females than in males, and increased with K1 intake. The MK-4/total VK ratios tended to be increased in old age possibly reflecting an increased physiological demand for MK-4 during aging. Our study also confirmed the greater susceptibility of male rats to low VK containing diet, notably at a younger age. Despite lifelong higher K1 intakes per unit body weight, tissue K1 and MK-4 contents at 20 months were generally lower in CR rats compared to their ad libitum (AL) counterparts. Whether the lower tissue MK-4 content is the result of lower synthesis from K1 or greater tissue utilization remains to be determined. However, the more youthful coagulation profile observed in old CR rats (vs. AL rats) tends to support the notion that CR is associated with greater utilization of the K vitamers to sustain physiological functions. [ABSTRACT FROM AUTHOR]

Background: The relationship between vitamin intake and depression has attracted increasing attention. However, several studies examining such relationship among populations at different age groups have produced inconsistent findings. This study was aimed to investigate the cross-sectional association between vitamin K intake and depressive symptoms in US adults., Methods: We used the data from a nationally representative sample of 11,687 adults from the 2013 to 2018 National Health and Nutrition Examination Survey (NHANES). Vitamin K intake was assessed by the 24-h dietary recall at the first day. Depressive symptoms were assessed using the 9-item Patient Health Questionnaire (PHQ-9). Logistic regression and generalized additive model were used to examine the association between vitamin K intake and depressive symptoms., Results: The weighted prevalence of depressive symptoms was 10.2% (8.0% in men and 12.0% in women). We observed a significant inverse linear relationship between vitamin K intake and depressive symptoms in models adjusted for age, sex, race/ethnicity, marital status, educational status, family poverty income ratio (PIR), home status, body mass index (BMI), smoking status, physical activity, sleep disorders, hypertension, hyperlipidemia, and diabetes. The odds ratios (OR) (95% CI) for the highest compared with the lowest quartile of vitamin K intake was 0.68 (95% CI: 0.52, 0.89, p -trend < 0.05). The association was similar in subgroups stratified by age, sex, race/ethnicity, marital status, educational status, PIR, home status, BMI, smoking status, physical activity, sleep disorders, hypertension, hyperlipidemia, and diabetes., Conclusion: Vitamin K intake was inversely and independently associated with the odds of depressive symptoms in the US adults. Prospective studies are warranted to confirm our findings., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Zhang, Tan, Xi, Yang, Zhang, Li, Chen and Zuo.)

Objectives Decreased concentration of menaquinone-4 (MK-4) seems to be an important risk factor of vascular calcification in haemodialysis (HD) patients. Optimal dietary intake, as well as serum MK-4 reference range, in HD has not been determined, yet. The aim of the present study was to assess daily vitamin K 1 and MK-4 intakes and their relation to serum MK-4 concentration in HD patients. Design and methods Daily vitamin K 1 and MK-4, micro- and macronutrients and energy intakes were assessed using 3-day food diary completed by patients and serum MK-4 concentration was measured by HPLC [limit of quantification (LOQ): 0.055 ng/mL] in 85 HD patients (51 males) and 22 apparently healthy subjects. Results Daily MK-4 intake was significantly lower (by 29%) among HD, while K 1 consumption was similar in both groups. Daily MK-4 intake was associated with fat and protein consumption in HD (r = 0.43, p < 0.001 and r = 0.33, p = 0.004, respectively). In HD serum MK-4 concentration was more frequently below LOQ (in 41% HD and 5% controls, p < 0.001) and in those HD with quantifiable values was lower than in the controls (by 42%). The correlations between MK-4 concentrations and both MK-4 and K 1 daily intakes were weaker in HD (r = 0.38 and r = 0.30 respectively) than in the control group (r = 0.47 and r = 0.45, respectively). In multiple regression analysis the variability of serum MK-4 concentrations in HD patients was explained by its daily intake. Conclusions Decreased serum MK-4 concentration in HD patients is caused by lower dietary MK-4 intake, mainly due to diminished meat consumption, and in addition, probably reduced K 1 conversion. [ABSTRACT FROM AUTHOR]

Introduction Warfarin is characterized by a large inter-individual variability in dosage requirement. This study aimed to analyze the contribution of the CYP4F2 genetic polymorphism and plasma vitamin K concentration on the warfarin pharmacodynamics in patients and to clarify the plasma vitamin K concentration affecting warfarin sensitivity index in rats. Materials and Methods Genetic analyses of selected genes were performed and plasma concentrations of warfarin, vitamin K1 (VK1) and menaquinone-4 (MK-4) were measured in 217 Japanese patients. We also assessed the association of plasma VK1 and MK-4 concentrations with the warfarin sensitivity index (INR/Cp) in rats. Results Patients with the CYP4F2 (rs2108622) TT genotype had significantly higher plasma VK1 and MK-4 concentrations than those with CC and CT genotypes. The multiple linear regression model including VKORC1, CYP4F2, and CYP2C9 genetic variants, age, and weight could explain 42% of the variability in warfarin dosage. The contribution of CYP4F2 polymorphism was estimated to be 2.2%. In contrast, plasma VK1 and MK-4 concentrations were not significantly associated with warfarin dosage in patients. Nevertheless, we were able to demonstrate that the warfarin sensitivity index was attenuated and negatively correlated with plasma VK1 concentration by the oral administration of VK1 in rats, as it resulted in a higher VK1 concentration than that in patients. Conclusions The plasma VK1 and MK-4 concentrations are significantly influenced by CYP4F2 genetic polymorphism but not associated with warfarin therapy at the observed concentration in Japanese patients. The CYP4F2 polymorphism is poorly associated with inter-individual variability of warfarin dosage requirement. [ABSTRACT FROM AUTHOR]

Background: Menaquinone-4 is a type of vitamin K that has a physiological function in maintaining bone quality via γ-carboxylation of osteocalcin. However, little is known about the beneficial effect of intake of dosages below 1500 µg/day. Findings: Fifteen healthy males aged 25.0 years (median) participated in a non-placebo-controlled dose-examination study. They received menaquinone-4 daily for 5 weeks at 0, 300, 600, 900, and 1500 µg/day in weeks 1, 2, 3, 4, and 5, respectively. Compared with baseline, serum γ-carboxylated osteocalcin levels were significantly greater at an intake of 900 µg/day or more; serum undercarboxylated osteocalcin levels and the ratio of serum undercarboxylated osteocalcin to γ-carboxylated osteocalcin were significantly lower than baseline at doses of 600 µg/day or more. Conclusions: This preliminary graded-dose study suggested that menaquinone-4 supplementation at 600 µg/day or more is likely to be important in terms of vitamin K requirements for bone health. [ABSTRACT FROM AUTHOR]

Highlights: [•] High-selectivity LC–MS analysis of K1, K1O, and MK-4 in human serum and plasma. [•] Method performance allows applying a simple and cost-effective extraction procedure. [•] “Endogenous lipid suppressors” were frozen and removed during LLE. [•] Blood samples from fasting warfarinised patients did not show deficiency of K1. [Copyright &y& Elsevier]