Vol 1, No 4 (2014)

Ectopic calcification and bone: a comparison

of the effect of dietary carbohydrates, sugars

and protein

Rachel Nicoll BSc MA

1

_{, John McLaren Howard PhD}

2

_{and Michael Y Henein MD PhD}

1

1 Department of Public Health and Clinical Medicine and Heart Centre

Umea University, Umea, Sweden

2 Acumen Lab, Tiverton, Devon, UK

Introduction

Cardiovascular (CV) calcification presence, extent and

progression has been shown in several studies to correlate

with fracture or low bone mineral density (BMD), particularly in

older men and women

1-3

_{. CV calcification shares some similar}

properties with cortical bone

4

_{and, when severe, can manifest as}

bone formation in both arteries and valves

5,6

_{. CV calcification and}

impaired bone metabolism are particularly prevalent in chronic

kidney disease (CKD), where the condition has become known as

mineral-bone disorder (MBD)

7

_{. The association between bone and}

ectopic calcification is further strengthened by studies showing

that renal calcification can be prevented by ibandronate, which

inhibits bone resorption

8

_{. Although calcification can be associated}

with atherosclerosis (i.e. intimal calcification), the deposition of

hydroxyapatite is primarily medial; its pathophysiology is complex,

involving not only physicochemical factors but also biological

actions in smooth muscle

9

_{. Although a number of studies have}

been carried out on the influence of dietary carbohydrates, sugars

and protein on bone, those relating diet to ectopic calcification are

considerably fewer and the comparison is necessarily restricted.

Carbohydrates and sugars

Ectopic calcification

In a prospective study of premenopausal women, carbohydrate

intake was inversely associated with coronary artery

calcification (CAC) five years after menopause but was not

associated with aortic calcification or carotid artery plaque

10

and a large cross-sectional study showed that whole grain

intake was not associated with two measures of subclinical

atherosclerosis: carotid intima-media thickness (CIMT) and

CAC

11

_{. Although there are no studies considering dietary intake}

of sugars, diabetes mellitus, metabolic syndrome, insulin

resistance and high blood glucose, HbA1C and triglycerides

are known risk factors for the presence of aortic

12

_{, breast}

13

_,

carotid

14

_{and coronary artery calcification}

15,16

_{and may also}

correlate with the aortic calcification score

17

_{or CAC score}

18

_.

The relationship between CAC presence and blood glucose

levels was found particularly among men

19

_{and may be}

non-linear

20

_{, with low insulin levels, as well as hyperinsulinaemia,}

independently predicting CAC presence

21

_{. Similarly in animals, a}

high carbohydrate diet appears to facilitate nephrocalcinosis

22

_,

while starch or any form of sugar can result in increased

incidence of CV or renal calcification, particularly in magnesium

deficiency

23,24

_{. Galactooligosaccharides or a high phytate diet,}

however, can reduce CV and renal calcification

25,26

_{. Likewise,}

in vitro studies have shown that a high glucose medium

enhances calcification of vascular smooth muscle cells (VSMCs)

and increased expression of markers of bone formation,

confirming a cell-mediation process characterized by the

trans-differentiation of VSMCs to osteoblast-like cells

27

_.

Bone

Studies of carbohydrate intake and bone parameters

show mixed results

28,29

_{, with several comparison studies}

of carbohydrates, proteins or fats showing no difference in

markers of bone formation or resorption

30,31

_{. Nevertheless, a}

12 month study found that BMD was significantly lower with

a high carbohydrate diet compared to a high protein diet

32

_.

Abstract

Dietary fibre is generally inversely associated with BMD

33

_{. Type}

2 diabetes and insulin resistance are strongly associated with

impaired bone strength and quality

34

_{, although there may be}

less correlation with BMD

35

_{and bone formation}

36

_{or resorption}

37

_;

some have suggested that it may be hyperinsulinaemia, rather

than hyperglycaemia, that underlies the relationship

34

_{. The effect}

of sugar intake depends upon the type, with sucrose or fructose

intake correlated with increased bone resorption markers

38

_,

adversely affecting mineral homeostasis

39

_{, while higher glucose}

intake decreased bone resorption markers

37

_{. Low lactose intake}

reduced calcium absorption and BMD

40

_{and increased fracture}

risk

41

_{. Prebiotics, such as inulin or fructans, increased mineral}

absorption in humans but results on bone parameters were not

always positive

42,43

_.

In animals, both high

44

_{and low}

45

_{carbohydrate intake can}

have a detrimental effect on bone formation, while higher

dietary fibre from legumes can increase calcium absorption,

bone calcium content and BMD

46

_{. High glucose, sucrose}

and fructose generally result in lower BMD, particularly in

magnesium deficiency, probably through reduced calcium

absorption

47

_{. Lactose again appears to be beneficial for bone}

48

_,

while xylitol can slow bone resorption, increasing BMD, BMC

and biomechanical properties

49

_{. Mannitol and prebiotics}

can improve bone mineral absorption by increasing gut

fermentation, resulting in greater mineral uptake in bone and

improved BMD and bone strength

50

_.

Dietary protein

Ectopic calcification

There have been no human studies investigating protein

intake and CV calcification and in animals they focus almost

exclusively on nephrocalcinosis, with the majority showing that

increased dietary protein reduced renal calcification in rats

with or without CKD with frequent reduction in urinary calcium

excretion

51,52

_{. Likewise, a low protein intake induced more}

severe renal calcification, with decreased glomerular filtration

rate and enhanced urinary albumin loss, suggesting kidney

damage

53

_{. In a study which considered both nephrocalcinosis}

and bone health, krill protein produced lower renal tubular

calcium deposition than casein, but had no effect on BMC

or bone strength

54

_{; although this shows a positive benefit of}

krill protein, this may in part be due to the omega-3 fatty acid

content. A high calcium, high phosphorus or low magnesium

intake can enhance the detrimental effect of low protein intake

on renal calcification

51,53

_{, while a calcium deficiency combined}

with high protein intake also enhanced the rate of ectopic

calcification

54

_.

Bone

Bone is 50% protein and 50% mineral

56

_{and consequently}

dietary protein is essential for bone formation

57

_{but it has}

nevertheless been considered detrimental to bone since

protein induces urinary calcium excretion thought to derive

from bone resorption

58,59

_{. Some recent studies, however,}

suggest that increased calcium excretion may derive from

increased absorption, possibly through elevated gastric acid

production

58,60

_{, higher glomerular filtration rate or decreased}

renal calcium reabsorption

61

_{. This is born out in a 2009}

systematic review and meta-analysis of protein intake studies

and protein supplementation trials, which found a small

positive association with BMD and no detrimental association

with fracture risk

62

_{. Recent studies have generally shown that,}

although there may be little short term effect

63

_{, long term high}

protein intake is positively associated with BMD, BMC and

decreased fracture risk

64

_{and slowed bone turnover}

65

_{. Similarly,}

animal studies have generally shown that while high protein

diets certainly increase urinary calcium excretion, they either

have no effect on bone

66

_{or bone calcium content and bone}

mass is improved

67

_{. In fact, it is generally low protein diets that}

are harmful to bone, particularly after ovariectomy

68,69

_{. There has}

been concern that a high animal/vegetable protein ratio may

be detrimental to bone

70

_{. Despite this, a 2009 meta-analysis}

showed that BMD was around 4% lower in vegetarians than

in omnivores

71

_{, although it was noted that key confounding}

variables had not been taken into account

72

_{. Since then a}

large study found no association between hip fracture and the

animal/plant protein ratio

73

_{, while animal protein was found}

to increase calcium absorption to a greater extent than soy

protein

74

_.

There may be an interaction with calcium status in humans.

Sahni et al found that when calcium intake was <800 mg/d,

there was a three-fold increase in fracture risk with a high

animal protein intake, while in those consuming ≥800 mg/d

calcium those with a high animal protein intake had an 85%

reduced risk of hip fracture

73

_{. Nevertheless, the body is clearly}

attempting to compensate since among postmenopausal

women, a high protein intake increased calcium retention

only in those with a low calcium intake

75

_{. Supplementing both}

protein and calcium significantly reduced bone loss in elderly

hip fracture patients

76

_{, while protein intake was associated}

with BMD increase over three years in subjects aged ≥65

receiving calcium and vitamin D

76,77

_{. A high protein diet in}

calcium repletion can also protect against the bone loss of

weight reduction

78

_{. Age may also play a role; a large study}

showed that increased protein intake was associated with a

decreased risk of osteoporotic hip fracture in subjects aged

50-69 years but not in those aged 70-89, regardless of calcium

intake

79

_{. Nevertheless, other prospective studies of the elderly}

have shown a clear relationship between low protein intake and

Figure 3

: Action of protein and sugars on bone and arteries

Key

IGF-1

Insulin-like growth factor 1

AGEs

Advanced glycation end products

RAGEs Receptor for AGEs

decreased BMD

[80]

_.

Potential mechanisms

There may be several shared mechanisms of action between

the effect of protein and sugars on ectopic calcification and

bone but one of the best studied is insulin-like growth factor

(IGF) 1, which is known to stimulate bone formation and,

together with its receptor, has a protective effect against arterial

calcification

81,82

_{. Protein is known to increase production of}

IGF-1

83

_{, while blood glucose and insulin levels are important}

regulators

84

_{. In vitro studies show that human VSMCs cultured}

in advanced glycation end products (AGEs), which are

significantly increased in diabetes mellitus, induce enhanced

calcification due to both decreased expression of IGF-1 and

upregulation of NF-

_κ

B, which blocks the IGF-1 receptor.

Over-expression of IGF-1 inhibited calcification in VSMCs, while

absence of the IGF-1 receptor negated the effect of AGEs on

calcification.

85

_{Interestingly, the addition of moderate calcium}

to the medium induced the IGF-1 receptor and inhibited

calcification in VSMCs, although high calcium led to inhibition

of the IGF-1 receptor, which increased calcification

86

_{. This may}

help to explain why an imbalance between protein and calcium

is detrimental.

A number of studies have been carried out on AGEs

independently of their effect on IGF-1. When AGEs come into

contact with their receptor (RAGE), intracellular reactive oxygen

species (ROS) are generated and mitogen-activated protein

kinase (MAPK) and nuclear factor kappa-B (NF-kB) signalling

is initiated

[87]

_{. VSMCs cultured with AGEs showed enhanced}

production of ROS, upregulated expression of RAGEs and

increased phosphorylation of p38 MAPK, as well as increased

activity of markers of bone formation: alkaline phosphatase,

osteopontin and osteocalcin

88,89

_{. When an antioxidant or}

anti-RAGE antibody was added to the medium, ROS expression,

p38 MAPK phosphorylation and calcification were reduced

and endogenous antioxidant expression increased, suggesting

that calcification of VSMCs occurs through a RAGE/oxidative

stress pathway via osteoblast-like differentiation of smooth

muscle cells

88,89

_{. A high glucose medium has a similar effect on}

VSMCs

90

_.

The AGE-RAGE interaction is additionally seen in bone

cells, resulting in increased expression of cytokines, growth

factors and adhesion molecules, which influence osteoclasts

and osteoblasts

91

_{and affect the structural and mechanical}

properties of bone

92

_{. In mouse bone cells, AGEs resulted in an}

increased number of resorption pits formed by osteoclasts,

while rat bone particles incubated with glucose were resorbed

to a much greater extent than control bone particles, suggesting

that AGEs enhance osteoclast-induced bone resorption

93

_.

Similarly, the fructose-induced stimulation of receptor activator

of NF-kB ligand (RANKL) significantly increased the number of

osteoclasts and pit formation in rat bone, accompanied by an

increase in ROS, an effect which could be completely abolished

by the antioxidant N-acetylcysteine

94

_{. Among the elderly,}

AGE-modified proteins stimulated monocytes/macrophages to

secrete bone-resorbing cytokines such as interleukin-1 beta,

interleukin-6 and tumour necrosis factor- alpha, with enhanced

net calcium efflux from bone

95

_.

Conclusion

A diet high in carbohydrates appears to increase ectopic

calcification, particularly in magnesium deficiency, although the

effect of carbohydrate and fibre on bone in humans is unclear

but appears to be consistently more detrimental than a high

protein diet. Although there are no human studies of the effect

of intake of sugars on ectopic calcification, diabetes mellitus,

insulin resistance and high blood glucose are risk factors

for both ectopic calcification and bone loss. Dietary sugars

generally increase ectopic calcification in animals, while in vitro

studies show that a high glucose medium significantly enhances

calcification of VSMCs, with increased local markers of bone

formation through action of AGEs which generate ROS. Sugars

generally reduce BMD in humans, although lactose appears

beneficial for bone in calcium deficiency but may increase renal

and CV calcification in animals. Prebiotics can increase mineral

absorption and improve BMD in animals, while also inhibiting

ectopic calcification, but the results on bone in humans are

inconsistent.

Whereas a high carbohydrate diet can increase ectopic

calcification, a diet high in protein may inhibit it, possibly

through induction of IGF-1. Dietary protein intake was generally

inversely correlated with renal calcification in animals with

and without CKD, although there were no human studies of

protein intake and ectopic calcification. In both animals and

humans, there is a modest positive association between protein

intake and BMD and an inverse association with fracture risk;

there appears to be little difference between animal and plant

proteins. There may be a strong synergistic interaction with

calcium, with low calcium and high protein intake significantly

increasing fracture risk and enhancing renal calcification,

while a high protein and calcium intake significantly reduced

fracture risk and inhibited ectopic calcification. Increased

protein is known to increase urinary calcium excretion but

rather than being taken from bone, this seems more likely to

be due to increased intestinal absorption and/or decreased

renal reabsorption, although this may only occur in calcium

deficiency. In general, therefore, a diet high in carbohydrate

and sugars and low in protein is detrimental to both arteries

and bone. This finding accords with our previous review, which

concluded that the dietary fats which are beneficial or harmful

for arteries are also beneficial or harmful for bone.

96

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