Zielsetzung: In einer Population im Westen der nigerianischen Stadt Kaduna wurden seit 20-30 Jahren vermehrt Kinder mit einer deformierenden Knochenerkrankung registriert. Ziel der Studie war, eine Diagnose zu stellen und Risikofaktoren für die Erkrankung zu identifizieren. Studiendesign: 26 Familien aus 20 Dörfern wurden in die Studie einbezogen. In einer nicht-randomisierten Fall-Kontroll-Studie wurden 53 erkrankte Kinder mit 48 gesunden sowie 16 fraglich erkrankten Geschwistern anhand ihrer Ergebnisse aus Anamnese, klinischer Untersuchung und Laborchemie miteinander verglichen. Ebenfalls wurden Daten von 24 Vätern und 36 Müttern ausgewertet. Weitere Untersuchungen umfassten Ernährung, Anthropometrie, Umweltfaktoren und Genetik der teilnehmenden Familien. Ergebnisse: Die betroffenen Kinder wiesen deutliche Rachitissymptome auf, bei allen lag eine Kalzium-defiziente Rachitis vor. Zwischen den Laborergebnissen von Fall- und Kontrollgruppe bestanden signifikante Unterschiede, nicht jedoch zwischen der Gruppe der fraglichen Fälle und der Kontrollgruppe. In der Fallgruppe waren die Serumspiegel von Kalzium und 25-Vit. D signifikant niedriger, die Serumspiegel von 1,25-Vit. D, ALP und PTH signifikant höher als in der Kontrollgruppe. Bei den Eltern zeigten die Mütter insbesondere in der Stillzeit signifikant niedrigere Kalzium- und signifikant höhere 1,25- Vit. D- und PTH-Serumspiegel als die Väter. Als Ursache für den Kalziummangel der Studienteilnehmer konnte eine kalziumarme und phytatreiche Diät der Familien identifiziert werden. Hinweise auf einen gesunkenen Lebensstandard und eine Abnahme der Bodenqualität erklären die in den letzten Jahrzehnten stark gestiegene Prävalenz der Erkrankung. Bei weitgehend gleichen Ernährungs- und Umweltfaktoren innerhalb einer Familie konnten keine individuellen Faktoren identifiziert werden, die bei einzelnen Familienmitgliedern zum Ausbruch der Erkrankung führten. Trotz einzelner Hinweise auf eine mögliche genetische Prädisposition war kein einheitliches Vererbungsmuster in den Stammbäumen der Familien erkennbar. Schlussfolgerung: Neben dem Hauptfaktor einer kalziumarmen Ernährung müssen weitere Faktoren für eine Kalzium-defiziente Rachitis vorliegen. Mehrere Hinweise deuten auf eine multifaktorielle Genese der Erkrankung hin. Die noch offenstehenden Fragen sollten durch weitere Studien geklärt werden, um die richtigen Maßnahmen für Prävention und Therapie zu treffen. / OBJECTIVES: In the last 20-30 years, an increasing number of children with deformed bones have been registered in a region west of the city Kaduna in Nigeria. The aim of the study is to find a diagnosis and identify risk factors. METHODS: Twenty-six families from 20 villages participated in our non-randomised case-controlled study. Fifty-three affected children were compared with 48 healthy siblings. Additional to the latter group, 16 other individuals were also included, whose health status was questionable. Data from their anamnesis, clinical examination and laboratory determinations were collected. The laboratory results from 24 fathers and 36 mothers were also analysed. Further investigations included the nutrition, anthropology, genetics and environmental factors of participating families. RESULTS: Affected children showed clear clinical signs of rickets, the calcium-deficient form being common to all. While cases and controls showed significant differences in their laboratory results, questionable cases showed no difference compared to their healthy siblings. Compared to controls, cases had significantly lower serum levels of 25-OH-Vitamin D (p < 0,001), calcium (p < 0,001) and phosphate (p = 0,001), but significantly higher serum levels of 1,25-OH-Vitamin D (p = 0,007), alkaline phosphatase (p < 0,001) and parathyroid hormone (p < 0,001). Mothers had significantly lower serum levels of calcium (p = 0,016), but significantly higher levels of 1,25-OH-Vitamin D (p = 0,001) and parathyroid hormone (p = 0,001) than fathers. Breastfeeding mothers had significantly higher serum levels of 1,25-OH-Vitamin D (p < 0,001) and alkaline phosphatase (p = 0,005) compared to non-breastfeeding mothers. A nutrition with low intakes of calcium and high intakes of phytates explains the lack of available calcium for the participants. Evidence for a decreased living standard and a deteriorated soil quality explains the rise in the prevalence of the disease. With widely similar nutrition and environmental factors within a family, no risk factors leading to the disease in single family members could be identified. Despite some hints for a possible genetic predisposition, the family trees of participating families showed no consistent inheritance pattern. CONCLUSIONS: Besides a calcium-deprived nutrition as a main factorcontributing to calcium deficiency rickets, our study shows that other factors are also involved. Here, information suggest a multifactorial genesis of the disease. Further studies are needed to clarify the outstanding questions and to implement improved measures for prevention and therapy.

A dissertation submitted to the Faculty of Science, University of the Witwatersrand, Johannesburg, in fulfilment of the requirements for the degree of Master of Science in Chemistry. University of the Witwatersrand, Johannesburg. May 2016 / In this research project, the morphogenesis and polymorphism of calcium carbonate (CaCO3) and its subsequent use as a template in the fabrication of hollow carbon spheres (HCS) is reported. A series of ratios (i.e. 5:0, 5:1, 5:2, 5:3, 5:4, 5:5, and 0:5) of binary solvent mixtures consisting of polar aprotic (dimethylformamide and dimethyl sulfoxide) and polar protic (methanol, ethanol, isopropanol, and 2-butanol) solvents, with 10% PEG200 as a crystal modifier, were used to influence the morphogenesis and polymorphism of precipitated CaCO3 (PCC). The PCC products were characterised using scanning electron microscopy (SEM), powder X-ray diffraction (PXRD), and laser Raman spectroscopy. An increase in the ratio of the polar protic solvent (methanol, ethanol, isopropanol, and 2-butanol) relative to the polar aprotic solvent (DMF & DMSO) within the binary solvent mixture favored the formation of vaterite particles of different morphologies, while an increase in the ratio of polar aprotic solvent (DMF & DMSO) within the binary solvent mixture favored the precipitation of rhombohedral calcite crystals. Time-resolved ex situ PXRD and SEM measurements revealed that the nucleation and phase transformation of the CaCO3 under polar protic and aprotic solvents followed the dissolution-reprecipitation mechanism. The major phase transformation occurred within 3 hours after mixing the precursor solutions. The effect of poly (4-styrenesulfonic acid) (PSSA) as an additive in the crystallisation of CaCO3 at different temperatures (i.e. 30, 40, 75, and 100 °C) and different crystallisation times (3, 6, 12, and 24 hrs) was investigated. The as-synthesised CaCO3 products were subjected to: SEM, laser Raman spectroscopy, PXRD, thermogravimetric analysis (TGA), and transmission electron microscopy (TEM). The crystallisation of CaCO3 in the presence of PSSA resulted in the self-assembly of vaterite particles into spherical bulk crystals. Varying the crystallisation temperature led to different particle attachment (CPA) pathways, which in turn resulted in bulk crystal morphologies that varied. Changes in the crystallisation temperature were found to not have changed the polymorphism of the precipitated CaCO3 due to the kinetic stabilisation effects of PSSA, instead hollow vaterite spheres formed at 75 and 100 °C. The possibility to synthesise HCS using CaCO3 as a template under chemical vapour deposition (CVD) at different temperatures (i.e. 600, 700 and 800 °C) was, for the first time, demonstrated. The evolution of CO2(g) from the decomposition of the template during CVD resulted in the formation of a rough surface topography on the carbon shell of the HCS. This surface roughness increased with the increase in the reaction temperature due to the increased rate of CaCO3 decomposition. The structural integrity of the spherical template was not affected by the CO2(g) evolution during carbonisation at all the reaction temperatures. The as-synthesised HCS at 600, 700, and 800 °C gave specific BET surface areas of: 193, 55, and 51 m2/g, respectively. / MT2016

by Warren, Tak-keung Lee. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1990. / Bibliography: leaves 140-154. / Acknowledgements / List of Abbreviations / Summary / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- Calcium Nutrition and Bone Health --- p.1 / Chapter 1.2 --- Calcium Requirements in Children --- p.4 / Chapter 1.3 --- Concern for Calcium Intakes in Hong Kong Chinese --- p.8 / Chapter 1.3.1 --- In Adults --- p.8 / Chapter 1.3.2 --- In Children --- p.9 / Chapter 1.4 --- Aims of the Study --- p.11 / Chapter Chapter 2 --- Calcium Homeostasis --- p.12 / Chapter 2.1 --- Calcium in Body Skeleton --- p.12 / Chapter 2 .2. --- Hormonal Regulation of Calcium Concentration in the Extracellular Fluid Compartment --- p.13 / Chapter 2.3 --- Calcium Absorption --- p.14 / Chapter 2.3.1 --- Calcium Transport Across the Intestine --- p.14 / Chapter 2.4 --- Effects of Age on Calcium Metabolism --- p.17 / Chapter 2.5 --- Effects of Nutrients on Calcium Bioavailability --- p.18 / Chapter 2.5.1 --- Calcium Nutritional Status --- p.19 / Chapter 2.5.2 --- Vitamin D Nutritional Status --- p.20 / Chapter 2.5.3 --- Protein --- p.21 / Chapter 2.5.4 --- Phosphorous and Calcium to Phosphorus Ratio --- p.22 / Chapter 2.5.5 --- Sodium --- p.22 / Chapter 2.5.6 --- Lactose --- p.23 / Chapter 2.5.7 --- Glucose and Glucose Polymers --- p.24 / Chapter 2.5.8 --- Phytate --- p.24 / Chapter 2.5.9 --- Oxalate --- p.25 / Chapter Chapter 3 --- Methods of Evaluating Calcium Nutritional Status And Bone Mineral Content --- p.27 / Chapter 3.1 --- Methods of Evaluating Calcium Nutrition Status --- p.27 / Chapter 3.1.1 --- Blood Biochemistry --- p.27 / Chapter 3.1.2 --- Total Body Neutron Activation --- p.27 / Chapter 3.1.3 --- Metabolic Balance Study --- p.28 / Chapter 3.1.4 --- Dietary Assessment --- p.29 / Chapter 3.2 --- Methods of Evaluating Bone Mineral Content --- p.29 / Chapter 3.2.1 --- In Vivo Measurement of Bone Mineral Content --- p.29 / Chapter 3.3 --- Review on Adopted Methods --- p.31 / Chapter 3.3.1 --- Single Photon Absorptiometry (SPA) --- p.31 / Chapter 3.3.2 --- Review on Methods in Dietary Assessment --- p.36 / Chapter 3.3.3 --- Dietary Assessment Methods Adopted in the Present Study --- p.49 / Chapter 3.3.4 --- Food Composition Tables --- p.51 / Chapter Chapter 4 --- "Subjects, Materials and Methods" --- p.54 / Chapter 4.1 --- Subjects --- p.54 / Chapter 4.2 --- Weight and Height Measurement --- p.57 / Chapter 4.3 --- Dietary Assessment --- p.58 / Chapter 4.3.1 --- Dietary History Record --- p.59 / Chapter 4.3.2 --- Food Frequency --- p.62 / Chapter 4.3.3 --- 24-Hour Recall --- p.64 / Chapter 4.3.4 --- Estimation of the Amount of Food --- p.64 / Chapter 4.3.5 --- Nutrient Analysis --- p.65 / Chapter 4.4 --- Measurement of Bone Mineral Mass by Single Photon Absorptiometry (SPA) --- p.66 / Chapter 4.4.1 --- The Instrument --- p.66 / Chapter 4.4.2 --- Calibration of the Instrument --- p.68 / Chapter 4.4.3 --- Subject Positioning --- p.68 / Chapter 4.4.4 --- Setting Up Pre-Scan Parameters --- p.70 / Chapter 4.4.5 --- Bone Scanning: Search Scan and Measure Scan --- p.73 / Chapter 4.4.6 --- Evaluation of the Accuracy and Precision of Bone Mineral Content Measurement --- p.73 / Chapter 4.4.7 --- Radiation Exposure --- p.75 / Chapter Chapter 5 --- Results --- p.77 / Chapter 5.1 --- Sample Size --- p.77 / Chapter 5.2 --- Representative of the Sample --- p.77 / Chapter 5.3 --- Weight and Height --- p.78 / Chapter 5.4 --- Validity of the Food Composition Table --- p.78 / Chapter 5.5 --- Calcium Intake of Children at Five --- p.78 / Chapter 5.6 --- "Bone Mineral Content (BMC), Bone Mineral Density (BMD), and Bone Width (BW) of 128 Hong Kong Children at Five" --- p.80 / Chapter 5.7 --- Comparisons of Bone Mineral Content (BMC) in Children with High and Low Calcium Intake --- p.81 / Chapter 5.8 --- "Relationships between Bone Mineral Density (BMD), Body Size and Dietary Intakes of Calcium, Protein and Energy" --- p.85 / Chapter 5.9 --- "Inter-correlations between Bone Mass Measurements, Body Size and Dietary Intakes in Hong Kong Children" --- p.87 / Chapter 5.10 --- Planning for Further Investigation --- p.88 / Chapter Chapter 6 --- "A Study of Calcium Intake and Bone Mineral Content of Children at Five Years Old in Jiangmen, Guangdong, China" --- p.107 / Chapter 6.1 --- Introduction --- p.107 / Chapter 6.2 --- "Subjects, Materials and Methods" --- p.108 / Chapter 6.2.1 --- Subject Selection --- p.108 / Chapter 6.2.2 --- Weight and Height Measurements --- p.109 / Chapter 6.2.3 --- Dietary Assessment --- p.109 / Chapter 6.2.4 --- Measurement of Bone Mineral Content --- p.111 / Chapter 6.2.5 --- Results --- p.112 / Chapter Chapter 7 --- Discussions --- p.120 / Chapter 7.1 --- Reliability of Dietary Calcium Assessment --- p.120 / Chapter 7.2 --- Reliability of Bone Mineral Measurement --- p.121 / Chapter 7.3 --- Representative of Studied Children in Hong Kong --- p.122 / Chapter 7.4 --- Implications of the Study --- p.123 / Chapter 7.4.1 --- Calcium Intake and Bone Mineral Content in Hong Kong Chinese Pre-school Children --- p.123 / Chapter 7.4.2 --- Calcium Intake and Bone Mineral Content of Jiangmen Pre-school Children --- p.131 / Chapter 7.5 --- The Use of Bone Mineral Density (BMD) As A Parameter in Representing Bone Mineral Mass --- p.134 / Chapter 7.6 --- A Need for Further Studies --- p.135 / Chapter 7.7 --- Conclusions --- p.136 / References --- p.140 / Chapter Appendix I --- Dietary Record Form Used in Hong Kong Study --- p.155 / Chapter Appendix II --- Dietary Questionnaire Used in Jiangmen Study --- p.160 / Chapter Appendix III --- Validity of the Compiled Food Composition Table --- p.165 / List of Figures / List of Tables

Graduation date: 1982

Recent research has shown an association between low dietary calcium intake and obesity in adults as well as overweight in young children; however, this relationship has not been investigated in adolescents. The purpose of this study was to examine the relationship between inadequate calcium intake and overweight in adolescents. The hypothesis of this study was that there is a negative correlation between dietary calcium intake and overweight in adolescents. The study population consisted of middle school and high school students (n = 102) in a local school district. The gender and ethnic distributions of the sample were as follows: 74% female, 26% male, 63% Caucasian, 16% African-American, 12% Hispanic, and 8% other. Dietary calcium and energy intakes were assessed using a previously validated calcium-focused food frequency questionnaire (FFQ) for youths. Calcium intake was also assessed using a single question on daily milk consumption. The FFQ was administered by trained interviewers to groups of three to five students. Body fat was assessed using body mass index for age (BMI-for-age) and sum of triceps and subscapular skinfolds (STS). The mean reported calcium intake was 1,972 ± 912 mg/day, and mean reported energy intake was 3,421 ± 1,710 kcals/day. Reported calcium intake from the FFQ was inflated since approximately 75% reported drinking less than three glasses of milk a day. According to BMI-for-age, 29% were classified as at risk of overweight or overweight. Using STS, 39% were classified as overweight. Chi-square analysis using either method of dietary calcium intake and either method of overweight assessment did not show dependence between categories of calcium intake and level of weight or body fat. This study failed to show a relationship between dietary calcium intake and risk of overweight or overweight in adolescents.

The divalent Ca2+ metal ion acts as a universal second messenger in virtually all eukaryotic cells from fungi to plants to mammals. In mammals, Ca2+ signaling is vital to a variety of physiological processes including fertilization, cell proliferation, secretion, and muscular contraction. In electrochemically non-excitable tissues, the release of Ca2+ from intracellular stores such as the endoplasmic reticulum is tightly regulated by the inositol 1,4,5-trisphosphate receptor (IP3R). The IP3R Ca2+ release channel is activated by the binding of the small molecule inositol 1,4,5-trisphosphate (IP3) in response to extracellular stimuli such as hormones, growth factors, and neurotransmitters. The conformational changes accompanying IP3 binding were investigated using a biophysical approach. A specific focus of this work is to decipher how signals of ligand binding are transmitted from the N-terminal IP3-binding core to the C-terminal channel domain. To such end, biophysical studies of the ligand-induced conformational changes within the N-terminal domain of IP3R (a.a. 1 – 604) were performed. The results implicated the presence of two flexible linkers which join stably folded domains. This prompted the proposal of a model in which an equilibrium mixture of conformational substrates containing compact and more extended structures co-exist. Determinants within the N- and C-terminal regions of IP3R have previously been reported to be critical to channel function. Employing nuclear magnetic resonance (NMR) as well as biochemical methods, an intermolecular interaction between the S4-S5 linker, the cytoplasmic loop between the fourth and fifth transmembrane helices of IP3R, and the suppressor domain was identified. The determination of the crystal structure of the suppressor domain from isoform type 3 IP3R (IP3R3SUP) allowed us to map the residues involved in this interaction to one face of the molecule. The characterization of this interaction provides insight into the N- and C-terminal determinants essential to the IP3R channel gating mechanism.

The divalent Ca2+ metal ion acts as a universal second messenger in virtually all eukaryotic cells from fungi to plants to mammals. In mammals, Ca2+ signaling is vital to a variety of physiological processes including fertilization, cell proliferation, secretion, and muscular contraction. In electrochemically non-excitable tissues, the release of Ca2+ from intracellular stores such as the endoplasmic reticulum is tightly regulated by the inositol 1,4,5-trisphosphate receptor (IP3R). The IP3R Ca2+ release channel is activated by the binding of the small molecule inositol 1,4,5-trisphosphate (IP3) in response to extracellular stimuli such as hormones, growth factors, and neurotransmitters. The conformational changes accompanying IP3 binding were investigated using a biophysical approach. A specific focus of this work is to decipher how signals of ligand binding are transmitted from the N-terminal IP3-binding core to the C-terminal channel domain. To such end, biophysical studies of the ligand-induced conformational changes within the N-terminal domain of IP3R (a.a. 1 – 604) were performed. The results implicated the presence of two flexible linkers which join stably folded domains. This prompted the proposal of a model in which an equilibrium mixture of conformational substrates containing compact and more extended structures co-exist. Determinants within the N- and C-terminal regions of IP3R have previously been reported to be critical to channel function. Employing nuclear magnetic resonance (NMR) as well as biochemical methods, an intermolecular interaction between the S4-S5 linker, the cytoplasmic loop between the fourth and fifth transmembrane helices of IP3R, and the suppressor domain was identified. The determination of the crystal structure of the suppressor domain from isoform type 3 IP3R (IP3R3SUP) allowed us to map the residues involved in this interaction to one face of the molecule. The characterization of this interaction provides insight into the N- and C-terminal determinants essential to the IP3R channel gating mechanism.

In excitable tissues, voltage-gated calcium channel activity is critical in the linkage of electrical stimuli to physiological responses, and so modulation of calcium channels therefore has significant implications. The exact mechanisms of calcium channel modulation and membrane expression, however, remain elusive. Previous work suggests that the calcium channel β subunit (Cavβ) modulates the expression and biophysical properties of the pore-forming α1 subunit. Previous research has shown that the core domains of Cavβ subunits are highly conserved, although alternative splicing in the highly variable N-terminus and HOOK regions is commonly observed in invertebrates, teleost fish and mammals. Splicing in these regions can produce unique isoforms that differentially modulate the membrane trafficking and gating properties of high voltage-activated calcium channels. With this in mind, two novel isoforms of an invertebrate Cavβ subunit have been identified and cloned from the pond snail Lymnaea stagnalis, which contain a novel N-terminus not previously identified. In addition, one of these novel isoforms excludes an optional, short exon in the HOOK region of LCavβ. Intron sequencing and amino acid alignments of the variable N-terminal and HOOK regions with mammalian and fish homologs have revealed that the genomic structure of Cavβ subuinits is conserved, despite the divergence in sequence and function between genes and splice isoforms. It was determined that the previously characterized LCavβ isoform, as well as the two new isoforms, can act to fine-tune calcium channel activity by modulating the membrane expression, voltage-dependencies of activation and inactivation and gating kinetics of invertebrate homologs of L-type (LCav1) and neuronal (LCav2) calcium channels. It is hoped that broadening our knowledge of simplified invertebrate calcium channels, like those found in Lymnaea, may advance our understanding the workings of our own highly elaborate and dynamic calcium channel complexes, and the nervous system as a whole.

In excitable tissues, voltage-gated calcium channel activity is critical in the linkage of electrical stimuli to physiological responses, and so modulation of calcium channels therefore has significant implications. The exact mechanisms of calcium channel modulation and membrane expression, however, remain elusive. Previous work suggests that the calcium channel β subunit (Cavβ) modulates the expression and biophysical properties of the pore-forming α1 subunit. Previous research has shown that the core domains of Cavβ subunits are highly conserved, although alternative splicing in the highly variable N-terminus and HOOK regions is commonly observed in invertebrates, teleost fish and mammals. Splicing in these regions can produce unique isoforms that differentially modulate the membrane trafficking and gating properties of high voltage-activated calcium channels. With this in mind, two novel isoforms of an invertebrate Cavβ subunit have been identified and cloned from the pond snail Lymnaea stagnalis, which contain a novel N-terminus not previously identified. In addition, one of these novel isoforms excludes an optional, short exon in the HOOK region of LCavβ. Intron sequencing and amino acid alignments of the variable N-terminal and HOOK regions with mammalian and fish homologs have revealed that the genomic structure of Cavβ subuinits is conserved, despite the divergence in sequence and function between genes and splice isoforms. It was determined that the previously characterized LCavβ isoform, as well as the two new isoforms, can act to fine-tune calcium channel activity by modulating the membrane expression, voltage-dependencies of activation and inactivation and gating kinetics of invertebrate homologs of L-type (LCav1) and neuronal (LCav2) calcium channels. It is hoped that broadening our knowledge of simplified invertebrate calcium channels, like those found in Lymnaea, may advance our understanding the workings of our own highly elaborate and dynamic calcium channel complexes, and the nervous system as a whole.

Functional foods go beyond simply supplying nutrients and are increasingly becoming a focus in the prevention and treatment of disease; however, the benefits of biofortified crops to human nutrition have not been well demonstrated. Modern breeding, molecular genetics, and biotechnology are currently focusing on how to improve the nutritional content in foods. Potatoes, carrots, and lettuce are popular vegetables eaten today and are targets in developing nutrient dense crops (biofortification). Biofortification of vegetables to increase calcium (Ca) in the diet has had promising results. Here we describe the current standing of nutrient biofortification of crops. Ca distribution within the plant cell moderates critical functions from signaling to growth and development and can affect overall plant vigor. The endoplasmic reticulum (ER) located Ca-ATPase ACA2 (Arabidopsis Ca-ATPase, isoform 2) is thought to play a role in intracellular calcium homeostasis. In yeast studies; a truncated pump (Δ80- ACA2) lacking the N-terminal region is about 10-fold more active than the full-length ACA2 pump. Single point mutations have been shown to increase activity of ACA2 in yeast as well. Previously in our lab, human feeding studies demonstrated that increased Ca accumulation and bioavailability in transgenic plants accompanied increased activity of the deregulated vacuolar Ca / H antiporter CAX1 (Cation Exchanger 1) termed sCAX1. In this study, transgenic tobacco plants expressing deregulated Ca transporters are compared. The phenotypes of deregulated vacuolar localized CAX and the ER localized ACA2 are compared. These results suggest deregulation of ACA2 may provide an additional tool to utilize in altering the calcium accumulation in agriculturally important crops.