Elsevier

Life Sciences

Volume 75, Issue 1, 21 May 2004, Pages 11-19
Life Sciences

Dietary myo-inositol hexaphosphate prevents dystrophic calcifications in soft tissues: a pilot study in Wistar rats

https://doi.org/10.1016/j.lfs.2003.11.030Get rights and content

Abstract

Myo-inositol hexaphosphate (InsP6) is an abundant component of plant seeds. It is also found in significant levels in blood and mammalian tissues, but they are totally dependent on their dietary intake. In the present paper, we describe studies on the effect of InsP6 on a model of dystrophic calcification, which was chemically induced by subcutaneous injection of a 0.1% KMnO4 solution. Male Wistar rats were randomly divided into four groups for treatment over 31 days. A: animals consuming a purified diet in which InsP6 was absent but to which 1% of InsP6 (as sodium salt) was added. In this group, the InsP6 plasma levels (0.393 ± 0.013 μM) were similar to those observed in rats consuming a standard diet. B: animals consuming only the purified diet in which InsP6 was absent. In this case the InsP6 plasma levels decreased (0.026 ± 0.006 μM); C: animals consuming the same purified diet as group B but received daily subcutaneous injections of 50 μg kg-1 etidronate during the last 14 days. In this case the InsP6 plasma levels were also very low (0.025 ± 0.007 μM); D: animals consuming the same diet as group B but a 6% of carob germ (InsP6 rich product) was added. The InsP6 plasma levels (0.363 ± 0.035 μM) were also similar to those observed in rats consuming a standard diet. After 21 days plaque formation was induced. Calcification plaques were allowed to proceed for 10 days, after which the plaque material present was excised, dried and weighed. It was found that the presence of myo-inositol hexaphosphate (phytate) in plasma at normal concentrations (0.3-0.4 μM) clearly inhibited the development of dystrophic calcifications in soft tissues. These results demonstrates that myo-inositol hexaphosphate acts as an inhibitor of calcium salt crystallization.

Introduction

Ectopic calcification is a common disorder associated with soft tissues such as the skin, kidney, tendons and cardiovascular tissues. In arteries, calcifications are correlated with atherosclerotic plaques which increase the risk of myocardial infarction Virmani et al., 2002, Wong et al., 1995. Degenerative calcific aortic stenosis is currently the most common valvular lesion found in clinical cardiology, and it is estimated that 1–2% of the elderly population suffer from this pathology (O'Keefe et al., 1991). All mammalian extracellular fluids are supersaturated with calcium phosphate (hydroxyapatite) and consequently are metastable with respect to this solid. These crystals do not precipitate spontaneously, and under physiological conditions, crystallization takes place only in controlled situations, such as in the formation of bone and teeth. Nevertheless, uncontrolled pathological crystallization may also occur. In fact, indiscriminate crystallization in human fluids does not occur because of regulated thermodynamic (supersaturation) and kinetic factors. Thus, there are three main aspects involved in biological crystallization: supersaturation (thermodynamic factor), the presence of heterogeneous nuclei (crystallization inducers, kinetic factor), and/or crystallization inhibitors (kinetic factor). It is clear that the presence of injured tissue provides heterogeneous nucleants that serve as substrates for initial crystal formation (Valente et al., 1985). On the other hand, the action of the so-called crystallization inhibitors can obstruct or prevent crystal formation, although these processes are poorly understood. Because of their chemical structures, these substances interact with the nucleus or the crystal, provoking important disturbances in their formation and/or development and preventing the crystallization processes. The existence of crystallization inhibitors was first observed in the 1960s. Bliznakov (1965) demonstrated that some molecules reduced the rate of crystal growth of a given substance due to their adsorption on growing surfaces, and when these inhibitory mechanisms were overcome, calcium crystals precipitated and proliferated.

Myo-inositol hexaphosphate (InsP6 or phytate) is an abundant component of plant seeds. In whole grain cereals such as corn, wheat, and rice, it ranges from 1.5% to 6.4%, while defatted and dehulled oilseed meals such as soy, peanut, and sesame meals contain 1.5% or more of the compound. In most seed types the InsP6 is associated with calcium and magnesium ions (the so-called phytin) and is not equally distributed. For example, the endosperm of wheat and rice kernels are almost devoid of InsP6 as it is concentrated in the germ and aleuronic layers of cells of the kernel and in the bran or hull. Corn differs from most cereals as almost 90% of InsP6 is concentrated in the germ portion of the kernel as in the carob germ. The levels found in blood and mammalian tissues clearly depended on their dietary intake Grases et al., 2001a, Grases et al., 2001b. Because we demonstrated that this molecule exhibited a potent capacity as crystallization inhibitor of calcium salts in urine Grases et al., 1996, Grases et al., 1998a, Grases et al., 1998b, we proposed in the present paper to study the effects of dietary InsP6 on artificially provoked dystrophic calcifications in soft tissues. Etidronate, a bisphosphonate class of drug Fleisch, 1997a, Fleisch, 1997b, was included for comparative purposes because of its crystallization-inhibitory effects in both hydroxyapatite and brushite formation Fleisch, 1997b, Grases et al., 2000c.

Section snippets

Animals, diets and experimental design

Twenty-four male Wistar rats of about 300 g from Harlan Iberica s.l. (Barcelona Spain) were acclimatized for 10 days. They were fed normal rat chow ad libitum and had free access to drinking water. Housing was in groups of three rats in Plexiglas cages. Environmental conditions were temperature 21 ± 1 °C, humidity 60 ± 5%, and light-dark periods 12:12 hours. They were randomly assigned into four groups of six rats each. The diets used were 4068.02 Reference Diet (HopeFarms BV, Woerden, The

Statistics

Values in the tables and figures are expressed as means ± (SE). One-way ANOVA was used to calculate significance of differences between groups. Student t-test was used to assess differences of means. The SPSS for the Windows program was used for statistical computations. Values for p are given in Fig. 2 and Table 2, Table 3. A value of p < 0.05 was considered to assess statistical significance.

Results

Blood levels of InsP6 in the four groups of rats are shown in Table 2. Animals consuming during 31 days the purified diet to which a 1% of InsP6 (as sodium salt) was added showed InsP6 blood levels of 0.393 ± 0.013 μM which were similar to those found in rats consuming a standard UAR-A03 diet (Grases et al., 2001e). In animals consuming only the purified diet lacking InsP6, the InsP6 blood levels decreased (0.026 ± 0.006 μM). Animals consuming the same purified diet to which etidronate was

Discussion

Dystrophic mineralization commonly occurs in soft tissues as a consequence of injury, disease, and aging with or without degenerative changes evident in the tissue. Although most soft tissues can undergo calcification, skin, kidney, tendons and cardiovascular tissues appear predisposed to develop this pathology (Anderson and Morris, 1993). Dystrophic calcifications, also termed ectopic calcifications, are typically composed of calcium phosphate salts, such as hydroxyapatite, but can also

Acknowledgements

This work was supported by the Conselleria d'Innovació i Energia del Govern de les Illes Balears and by the project BQU 2003-01659 of the Spanish Ministry of Science and Technology. One of the authors, J.P., expresses his appreciation to the Spanish Ministry of Education, Culture and Sport for a fellowship of the FPU program.

References (35)

  • B Sandstrom et al.

    A high oat-bran intake does not impair zinc absorption in humans when added to a low-fiber animal protein-based diet

    Journal of Nutrition

    (2000)
  • R Virmani et al.

    Pathology of the unstable plaque

    Progress in Cardiovascular Diseases

    (2002)
  • H.C Anderson et al.

    Mineralization

  • G Bliznakov

    Sur le mecanisme de l'action des additifs adsorbants dans la croissance cristalline

  • K.D Demadis et al.

    A crystallographically characterized nine-coordinate calcium-phosphocitrate complex as calcification inhibitor in vivo

    Journal of the American Chemical Society

    (2001)
  • H.A Fleisch

    Bisphosphonates: preclinical aspects and use in osteoporosis

    Annals of Medicine

    (1997)
  • H Fleisch

    Mechanisms of action of the bisphosphonates

    Medicina (B Aires)

    (1997)
  • Cited by (38)

    • Phytic acid: As a natural antioxidant

      2022, Antioxidants Effects in Health: The Bright and the Dark Side
    • Rediscovery of natural compounds acting via multitarget recognition and noncanonical pharmacodynamical actions

      2020, Drug Discovery Today
      Citation Excerpt :

      By contrast, because of its chelating capabilities, InsP6 could also prevent renal stone development by forming insoluble complexes with calcium ions. Thus, the chelating ability of this molecule could be of use against microcalcification deposits in soft tissues [62,63] Classical antiacids display a comparable mechanism to that of InsP6 (i.e., electrostatic neutralization of potential toxic ions diluted in biological fluids).

    • Rice phenolics and other natural products

      2018, Rice: Chemistry and Technology
    • Evaluation of inositol phosphates in urine after topical administration of myo-inositol hexaphosphate to female Wistar rats

      2018, Life Sciences
      Citation Excerpt :

      Myo-inositol hexaphosphate (InsP6, phytate) is abundant in plant seeds [1]. Consumption of InsP6 as a supplement or in the diet may provide important health benefits, such as prevention of renal calculi [2–4], prevention of cardiovascular calcifications and other pathological calcifications [5,6], antioxidant effects [7,8], and prevention of some cancers [9–11] and osteoporosis [12]. The chemistry of inositol phosphates (InsPs) in vivo is complicated because there are multiple forms (InsP1 to InsP6) and multiple isomers of the different forms, due to the dephosphorylation of InsP6 by phosphatases and the de novo synthesis within cells [13–14].

    • Theoretical study of the HAP crystal growth inhibition potency of pyrophosphate, etidronate, citrate and phytate. Deciphered the adsorbed conformation of phytate on the HAP (001) surface

      2017, Applied Surface Science
      Citation Excerpt :

      Due to their chemical structure, phytate is negatively charged at acidic, neutral and basic pH conditions [39], and hence it can bind positively charged calcium ions, preventing pathological calcium depositions, and can bind to already formed HAP crystals, reducing their development [40]. A large number of experimental studies highlighted that phytate ion has a pronounced inhibitory effect on phosphate calcium crystallization, brushite and hydroxyapatite [21,22], and it has been shown that inhibits the formation of kidney stones [22], sialolithiasis [41], dental tartar [42] and soft tissues calcifications, such as cardiovascular calcifications [43–46]. In spite of the large number of reported research and the therapeutic relevance in preventing calcifications of phytate, there is a lack of knowledge of how the molecule binds to the HAP surfaces.

    View all citing articles on Scopus
    View full text