**8. Research directions and priorities**

The research on vitamin K2 and cardiovascular health has come a long way, going from observational studies of populations and warfarin studies, to developing biomarkers, finding effective dosage schedules, and begin carrying out RCT studies examining disease endpoints. Very few disease endpoints have been reported at this time, but several should be completed in the next few years. Diseases are multifaceted, so the solutions should likewise be multifaceted. Trying to reverse a complex disease with a single nutrient has generally been unsuccessful. It is likely that the best success will be found when vitamin K2 is used in the context of a whole food plant-based diet that contains some dietary source of vitamin K2, along with supplements to reverse disease damage when appropriate.

[2] Price WA. Nutrition and physical degeneration. 6th edition. New Canaan, Conn: Keats

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183

[3] Masterjohn C. On the trail of the elusive X-factor: Vitamin K2 revealed. Wise traditions in food, farming and the healing arts [Internet]. 2007 [cited 2016 May 17]. Available from: http://www.westonaprice.org/health-topics/abcs-of-nutrition/on-the-trail-of-

[4] Jie K-SG, Bots ML, Vermeer C, Witteman JCM, Grobbee DE. Vitamin K intake and osteocalcin levels in women with and without aortic atherosclerosis: a population-

[5] Schurgers LJ, Geleijnse JM, Grobbee DE, Pols HAP, Hofman A, Witteman JCM, et al. Nutritional intake of vitamins K1 (Phylloquinone) and K2 (Menaquinone) in the

[6] Geleijnse JM, Vermeer C, Grobbee DE, Schurgers LJ, Knapen MHJ, van der Meer IM, et al. Dietary intake of menaquinone is associated with a reduced risk of coronary heart

[7] Gast GCM, de Roos NM, Sluijs I, Bots ML, Beulens JWJ, Geleijnse JM, et al. A high menaquinone intake reduces the incidence of coronary heart disease. Nutr Metab

[8] Erkkilä AT, Booth SL, Hu FB, Jacques PF, Manson JE, Rexrode KM, et al. Phylloquinone intake as a marker for coronary heart disease risk but not stroke in women. Eur J Clin

[9] Erkkilä AT, Booth SL, Hu FB, Jacques PF, Lichtenstein AH. Phylloquinone intake and risk of cardiovascular diseases in men. Nutr Metab Cardiovasc Dis NMCD. 2007 Jan;

[10] Maas AHEM, van der Schouw YT, Beijerinck D, Deurenberg JJM, Mali WPTM, Grobbee DE, et al. Vitamin K intake and calcifications in breast arteries. Maturitas. 2007 Mar

[11] Beulens JWJ, van der A DL, Grobbee DE, Sluijs I, Spijkerman AMW, van der Schouw YT. Dietary phylloquinone and menaquinones intakes and risk of type 2 diabetes.

[12] Beulens JWJ, Bots ML, Atsma F, Bartelink M-LEL, Prokop M, Geleijnse JM, et al. High dietary menaquinone intake is associated with reduced coronary calcification. Athe-

[13] Juanola-Falgarona M, Salas-Salvadó J, Martínez-González MÁ, Corella D, Estruch R, Ros E, et al. Dietary intake of vitamin K is inversely associated with mortality risk. J

the-elusive-x-factor-a-sixty-two-year-old-mystery-finally-solved/

based study. Atherosclerosis. 1995 Jul 1;116(1):117–23.

Netherlands. J Nutr Environ Med. 1999 Jan;9(2):115–22.

Cardiovasc Dis NMCD. 2009 Sep;19(7):504–10.

Diabetes Care. 2010 Aug 1;33(8):1699–705.

rosclerosis. 2009 Apr;203(2):489–93.

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17(1):58–62.

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disease: the Rotterdam study. J Nutr. 2004 Nov;134(11):3100–5.

Pub; 2003. 524 p.

One of the major benefits seen by Dr. Weston Price in the 1930s was the prevention and reversal of dental caries using vitamin K2 [2]. This could be a very fruitful area of research that is untapped at this point. Mental health effects of vitamin K2 are likely as well, as the brain contains a significant concentration of K2. Varicose veins in general and prostate health in particular should be studied in light of the K2 research presented here in this chapter. Mechanisms of the function of MGP should be worked out along the way as well. Identification of the function of other vitamin K–dependent proteins is still lacking. There is a gamma-carboxylation–rich protein (GRP) that is quite small yet has 16 carboxylation sites and is highly expressed in cartilage. Its exact function is still unknown. So, there is work that can be done at the molecular level as well as at the public health level in furthering our understanding of vitamin K2.

#### **Conflict of Interest**

Michael Donaldson is a research scientist at the Hallelujah Acres Foundation for investigations pertaining to the Hallelujah Diet. Funding for this research has been provided by Hallelujah Acres, Inc.

#### **Author details**

Michael S. Donaldson\*

Address all correspondence to: mdonaldson@myhdiet.com

Hallelujah Acres, Zillah, WA, USA

#### **References**

[1] Dam H. The antihaemorrhagic vitamin of the chick. Biochem J. 1935 Jun;29(6):1273–85.

[2] Price WA. Nutrition and physical degeneration. 6th edition. New Canaan, Conn: Keats Pub; 2003. 524 p.

**8. Research directions and priorities**

182 Vitamin K2 - Vital for Health and Wellbeing

vitamin K2.

Acres, Inc.

**Conflict of Interest**

**Author details**

**References**

Michael S. Donaldson\*

Hallelujah Acres, Zillah, WA, USA

supplements to reverse disease damage when appropriate.

Address all correspondence to: mdonaldson@myhdiet.com

The research on vitamin K2 and cardiovascular health has come a long way, going from observational studies of populations and warfarin studies, to developing biomarkers, finding effective dosage schedules, and begin carrying out RCT studies examining disease endpoints. Very few disease endpoints have been reported at this time, but several should be completed in the next few years. Diseases are multifaceted, so the solutions should likewise be multifaceted. Trying to reverse a complex disease with a single nutrient has generally been unsuccessful. It is likely that the best success will be found when vitamin K2 is used in the context of a whole food plant-based diet that contains some dietary source of vitamin K2, along with

One of the major benefits seen by Dr. Weston Price in the 1930s was the prevention and reversal of dental caries using vitamin K2 [2]. This could be a very fruitful area of research that is untapped at this point. Mental health effects of vitamin K2 are likely as well, as the brain contains a significant concentration of K2. Varicose veins in general and prostate health in particular should be studied in light of the K2 research presented here in this chapter. Mechanisms of the function of MGP should be worked out along the way as well. Identification of the function of other vitamin K–dependent proteins is still lacking. There is a gamma-carboxylation–rich protein (GRP) that is quite small yet has 16 carboxylation sites and is highly expressed in cartilage. Its exact function is still unknown. So, there is work that can be done at the molecular level as well as at the public health level in furthering our understanding of

Michael Donaldson is a research scientist at the Hallelujah Acres Foundation for investigations pertaining to the Hallelujah Diet. Funding for this research has been provided by Hallelujah

[1] Dam H. The antihaemorrhagic vitamin of the chick. Biochem J. 1935 Jun;29(6):1273–85.


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[26] Han KH, O'Neill WC. Increased peripheral arterial calcification in patients receiving

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http://dx.doi.org/10.5772/63413

185

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[62] Ford ES, Bergmann MM, Kröger J, Schienkiewitz A, Weikert C, Boeing H. Healthy living is the best revenge: findings from the European prospective investigation into cancer and nutrition-potsdam study. Arch Intern Med. 2009 Aug 10;169(15):1355–62.

**Chapter 10**

**Provisional chapter**

**Menaquinone‐4 Enhances Steroidogenesis in Testis**

**Menaquinone‐4 Enhances Steroidogenesis in Testis**

**Derived Tumor Cells Via the Elevation of cAMP Level**

Naturally existing vitamin K consists of vitamins K1 and K2. Menaquinone‐4 (MK‐4), an analog of vitamin K2 and a product of vitamin K1 metabolism, can be detected in several organs, including the testis; however, the function of MK‐4 in these tissues has not been well characterized. Recent studies have suggested that vitamin K is involved in enhancing protein kinase A (PKA) activity in several cell types, thus regulating numerous PKA‐dependent biological processes. To highlight the effect of vitamin K, we focused on its role in the steroidogenic pathway. Experiments on vitamin K–deficient rats revealed a reduced expression of genes involved in the biosynthesis of cholesterol and steroid hormones in the testis. Moreover, compared with control animals, rats fed on MK‐4 diet presented significantly higher testosterone levels in the plasma and testis. These results suggest that vitamin K is involved in the steroidogenic pathway in the testis. Testosterone levels were found to increase in a dose‐dependent manner also in cell‐based experiments upon addition of MK‐4, but such an effect was not observed in vitamin K1 levels. Furthermore, the effect of MK‐4 on testosterone production was abolished by the specific PKA inhibitor H89, thus confirming the regulatory role of MK‐ 4 on PKA activation. Here, we describe how MK‐4 modulates PKA activation by enhancing intracellular 3′,5′‐cyclic adenosine monophosphate (cAMP) levels in testis‐ derived I‐10 cells. The presented evidence supports the role of MK‐4 in cAMP/PKA

**Keywords:** menaquinone‐4, steroidogenesis, cAMP, protein kinase A, Leydig cells

Naturally existing vitamin K comprises vitamin K1 (phylloquinone) and vitamin K2 (mena‐ quinone). Menaquinone‐4 (MK‐4), an analog of vitamin K2, contains a 2‐methyl‐1,4‐naphtho‐

and reproduction in any medium, provided the original work is properly cited.

© 2017 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution,

© 2017 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

**Derived Tumor Cells Via the Elevation of cAMP Level**

Hsin‐Jung Ho, Hitoshi Shirakawa and Michio Komai

Hsin‐Jung Ho, Hitoshi Shirakawa and Michio Komai

Additional information is available at the end of the chapter

Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/63982

signaling and steroidogenesis.

**1. Introduction**

**Abstract**


#### **Menaquinone‐4 Enhances Steroidogenesis in Testis Derived Tumor Cells Via the Elevation of cAMP Level Menaquinone‐4 Enhances Steroidogenesis in Testis Derived Tumor Cells Via the Elevation of cAMP Level**

Hsin‐Jung Ho, Hitoshi Shirakawa and Michio Komai Hsin‐Jung Ho, Hitoshi Shirakawa and Michio Komai

Additional information is available at the end of the chapter Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/63982

#### **Abstract**

[62] Ford ES, Bergmann MM, Kröger J, Schienkiewitz A, Weikert C, Boeing H. Healthy living is the best revenge: findings from the European prospective investigation into cancer

[63] Zhang Y, Tuomilehto J, Jousilahti P, Wang Y, Antikainen R, Hu G. Lifestyle factors on the risks of ischemic and hemorrhagic stroke. Arch Intern Med. 2011 Nov 14;171(20):

[64] Nimptsch K, Rohrmann S, Kaaks R, Linseisen J. Dietary vitamin K intake in relation to cancer incidence and mortality: results from the Heidelberg cohort of the European prospective investigation into cancer and nutrition (EPIC-Heidelberg). Am J Clin Nutr.

and nutrition-potsdam study. Arch Intern Med. 2009 Aug 10;169(15):1355–62.

1811–8.

188 Vitamin K2 - Vital for Health and Wellbeing

2010 Mar 24;91(5):1348–1358.

Naturally existing vitamin K consists of vitamins K1 and K2. Menaquinone‐4 (MK‐4), an analog of vitamin K2 and a product of vitamin K1 metabolism, can be detected in several organs, including the testis; however, the function of MK‐4 in these tissues has not been well characterized. Recent studies have suggested that vitamin K is involved in enhancing protein kinase A (PKA) activity in several cell types, thus regulating numerous PKA‐dependent biological processes. To highlight the effect of vitamin K, we focused on its role in the steroidogenic pathway. Experiments on vitamin K–deficient rats revealed a reduced expression of genes involved in the biosynthesis of cholesterol and steroid hormones in the testis. Moreover, compared with control animals, rats fed on MK‐4 diet presented significantly higher testosterone levels in the plasma and testis. These results suggest that vitamin K is involved in the steroidogenic pathway in the testis. Testosterone levels were found to increase in a dose‐dependent manner also in cell‐based experiments upon addition of MK‐4, but such an effect was not observed in vitamin K1 levels. Furthermore, the effect of MK‐4 on testosterone production was abolished by the specific PKA inhibitor H89, thus confirming the regulatory role of MK‐ 4 on PKA activation. Here, we describe how MK‐4 modulates PKA activation by enhancing intracellular 3′,5′‐cyclic adenosine monophosphate (cAMP) levels in testis‐ derived I‐10 cells. The presented evidence supports the role of MK‐4 in cAMP/PKA signaling and steroidogenesis.

**Keywords:** menaquinone‐4, steroidogenesis, cAMP, protein kinase A, Leydig cells
