Ria-Ann R. Lomeda

Zinc deficiency suppresses matrix mineralization and retards osteogenesis transiently with catch-up possibly through Runx 2 modulation

A characteristic sign of zinc deficiency is retarded skeletal growth, but the role of zinc in osteoblasts is not well understood. Two major events for bone formation include osteoblast differentiation by bone marker gene expression, which is mainly regulated by bone-specific transcription factor Runx2 and extracellular matrix (ECM) mineralization by Ca deposits for bone nodule formation. We investigated whether zinc deficiency down-regulates bone marker gene transcription and whether this might occur through modulation of Runx2. We also investigated whether zinc deficiency decreases ECM mineralization in osteoblastic MC3T3-E1 cells. In the presence of 5 mumol/L TPEN as zinc chelator, zinc deficiency (ZnD: 1 micromol Zn/L) decreased bone marker gene (collagen type I, osteopontin, alkaline phosphatase, osteoclacin and parathyroid hormone receptor) expression, as compared to normal osteogenic medium (OSM) or zinc adequate medium (ZnA: 15 micromol/L) (P<0.05) both at 5 days (proliferation) and 15 days (matrix maturation). Decreased bone marker gene transcription by zinc deficiency could be caused by decreased nuclear Runx2 protein (P=0.05) and transcript (P<0.05) levels in ZnD. Furthermore, within the first 24 h of differentiation when Runx2 expression is induced, maximal Runx2 mRNA and nuclear protein levels were delayed in ZnD compared to OSM and ZnA. ECM Ca deposition was also lower in ZnD, which was also indirectly confirmed by detection of decreased cellular (synthesized) and medium (secreted) ALP activity as well as matrix ALP activity. Taken together, zinc deficiency attenuated osteogenic activity by decreasing bone marker gene transcription through reduced and delayed Runx2 expression and by decreasing ECM mineralization through inhibition of ALP activity in osteoblasts. Decreased and delayed bone marker gene, Runx2 expression and ECM mineralization in osteoblasts by zinc deficiency can be a potential explanation for the retarded skeletal growth which is the major zinc deficiency syndrome.

Marginal zinc deficiency in rats decreases leptin expression independently of food intake and corticotrophin-releasing hormone in relation to food intake.

Zn deficiency reduces food intake and growth rate in rodents. To determine the relationship between Zn deficiency and the regulation of food intake, we evaluated leptin gene expression in epididymal white adipose tissue (eWAT), and hypothalamic corticotropin-releasing hormone (hCRH) and hypothalamic neuropeptide Y (hNPY) of rats Zn-deficient only to show reduced food intake and growth rate but not food intake cycling. Growing male Sprague-Dawley rats (240 g) were randomly assigned to one of four dietary groups: Zn-adequate (ZA; 30 mg/kg diet), Zn-deficient (ZD; 3 mg/kg diet), pair-fed with ZD (PF; 30 mg/kg diet) and Zn-sufficient (ZS; 50 mg/kg diet) (n 8), and were fed for 3 weeks. Food intake and body weight were measured, as were blood mononuclear cells and pancreas Zn levels. eWAT leptin, hCRH and hNPY mRNA levels were determined. Food intake was decreased by about 10 % in ZD and PF rats compared to ZA and ZS rats. Growth and eWAT leptin mRNA levels were unaffected in PF rats but were significantly (P < 0.05) decreased in ZD rats. However, hNPY showed a tendency to increase, and hCRH significantly (P < 0.05) decreased, in both ZD and PF rats. These results suggest that while leptin gene expression may be directly affected by Zn, hNPY and hCRH are likely responding to reduced food intake caused by Zn deficiency.

Zinc deficiency negatively affects alkaline phosphatase and the concentration of Ca, Mg and P in rats

Zn is an essential nutrient that is required in humans and animals for many physiological functions, including immune and antioxidant function, growth, and reproduction. The present study evaluated whether Zn deficiency would negatively affect bone-related enzyme, ALP, and other bone-related minerals (Ca, P and Mg) in rats. Thirty Sprague Dawley rats were assigned to one of the three different Zn dietary groups, such as Zn adequate (ZA, 35 mg/kg), pair fed (PF, 35 mg/kg), Zn deficient (ZD, 1 mg/kg) diet, and fed for 10 weeks. Food intake and body weight were measured daily and weekly, respectively. ALP was measured by spectrophotometry and mineral contents were measured by inductively coupled plasma-mass spectrophotometer (ICP-MS). Zn deficient rats showed decreased food intake and body weight compared with Zn adequate rats (p<0.05). Zn deficiency reduced ALP activity in blood (RBC, plasma) and the tissues (liver, kidney and small intestine) (p<0.05). Also, Zn deficiency reduced mineral concentrations in rat tissues (Ca for muscle and liver, and Mg for muscle and liver) (p<0.05). The study results imply the requirement of proper Zn nurture for maintaining bone growth and formation.

Zinc deprivation inhibits extracellular matrix calcification through decreased synthesis of matrix proteins in osteoblasts

SCOPE Zinc is implicated as an activator for bone formation, however, its influence on bone calcification has not been reported. This study examined how zinc regulates the bone matrix calcification in osteoblasts. METHODS AND RESULTS Two osteoblastic MC3T3-E1 cell subclones (SC 4 and SC 24 as high and low osteogenic differentiation, respectively) were cultured in normal osteogenic (OSM), Zinc deficient (Zn-, 1 μM), or adequate (Zn+, 15 μM) media up to 20 days. Cells (SC 4) were also supplemented with (50 μg/mL) or no ascorbic acid (AA) in combination with Zinc treatment. Zn- decreased collagen synthesis and matrix accumulation. Although AA is essential for collagen formation, its supplementation could not compensate for Zinc deficiency-induced detrimental effects on extracellular matrix mineralization. Zn- also decreased the medium and cell layer alkaline phosphatase ALP activity. This decreased ALP activity might cause the decrease of Pi accumulation in response to Zn-, as measured by von Kossa staining. Ca deposition in cell layers, measured by Alizarin red S staining, was also decreased by Zn(-) . CONCLUSION Our findings suggest that zinc deprivation inhibits extracellular matrix calcification in osteoblasts by decreasing the synthesis and activity of matrix proteins, type I collagen and ALP, and decreasing Ca and Pi accumulation. Therefore zinc deficiency can be considered as risk factor for poor extracellular matrix calcification.

The decreased molar ratio of phytate:zinc improved zinc nutriture in South Koreans for the past 30 years (1969-1998)

For the assessment of representative and longitudinal Zn nutriture in South Koreans, Zn, phytate and Ca intakes were determined using four consecutive years of food consumption data taken from Korean National Nutrition Survey Report (KNNSR) every 10 years during 1969-1998. The nutrient intake data are presented for large city and rural areas. Zn intake of South Koreans in both large city and rural areas was low during 1969-1988 having values between 4.5-5.6 mg/d, after then increased to 7.4 (91% Estimated Average Requirements for Koreans, EAR = 8.1 mg/d) and 6.7 mg/d (74% EAR) in 1998 in large city and rural areas, respectively. In 1968, Zn intake was unexpectedly higher in rural areas due to higher grain consumption, but since then until 1988 Zn intake was decreased and increased back in 1998. Food sources for Zn have shifted from plants to a variety of animal products. Phytate intake of South Koreans during 1969-1978 was high mainly due to the consumption of grains and soy products which are major phytate sources, but decreased in 1998. The molar ratios of phytate:Zn and millimmolar ratio of phytate×Ca:Zn were decreased due to the decreased phytate intake in South Koreans, which implies higher zinc bioavailability. The study results suggest that Zn nutriture has improved by increased dietary Zn intakes and the decreased molar ratio of phytate:Zn in South Koreans in both large city and rural areas.