Stabilizing feedbacks in glacier-bed erosion.

Glaciers often erode, transport and deposit sediment much more rapidly than nonglacial environments, with implications for the evolution of glaciated mountain belts and their associated sedimentary basins. But modelling such glacial processes is difficult, partly because stabilizing feedbacks similar to those operating in rivers have not been identified for glacial landscapes. Here we combine new and existing data of glacier morphology and the processes governing glacier evolution from diverse settings to reveal such stabilizing feedbacks. We find that the long profiles of beds of highly erosive glaciers tend towards steady-state angles opposed to and slightly more than 50 per cent steeper than the overlying ice-air surface slopes, and that additional subglacial deepening must be enabled by non-glacial processes. Climatic or glaciological perturbations of the ice-air surface slope can have large transient effects on glaciofluvial sediment flux and apparent glacial erosion rate.

Vitamin D plasma binding protein. Turnover and fate in the rabbit.

The metabolic disposition of the plasma binding protein (DBP) for vitamin D and its metabolites was studied in adult rabbits. Apo-DBP was purified from rabbit plasma and enzymatically labeled with radioiodine. The radioiodine-labeled protein retained its ability to bind vitamin D sterols and its physicochemical properties. When 125I-labeled DBP and 131I-labeled rabbit albumin were simultaneously injected intravenously, the 125I was cleared from plasma at a faster rate (t 1/2 = 1.7 d) than 131I (t 1/2 = 5 d) and 125I was present in excess of 131I in kidney, liver, skeletal muscle, heart, lung, intestine, testis, and bone 1 h after injection. In contrast to DBP, 25(OH)D3 was cleared more slowly (t 1/2 = 10.7 d). Compared to albumin, DBP radioactivity appeared earlier and in greater quantity in the urine of catheterized rabbits. Gel filtration analyses of plasma revealed most of the 125I to elute in the position of DBP, with only small amounts in the less than 1,000-dalton region. In contrast, almost all of the urine 125I eluted in this small molecular weight fraction. The molar ratio of DBP to 25(OH)D3 in normal rabbit plasma was 138/1. The extravascular pool of DBP was calculated to be 1.5-2.4 times larger than the intravascular DBP pool, and the molar replacement rate of DBP was 1,350-fold higher than that of 25(OH)D3. The plasma disappearance curves of holo-DBP, prepared either by saturating with 25(OH)D3 or by covalently linking 3 beta-bromoacetoxy-25(OH)D3, were very similar to that of apo-DBP. Neuraminidase treatment of DBP did not alter its plasma survival. These studies indicate that DBP or DBP-25(OH)D3 complex is removed from plasma by a variety of tissues, that the DBP moiety is degraded during this process, and that a significant recirculation of 25(OH)D3 probably occurs. The molar excess of DBP to 25(OH)D3 in plasma, and the relatively rapid turnover of DBP indicate that a high capacity, high affinity, and dynamic transport mechanism for vitamin D sterols exists in rabbit plasma.

1,25-Dihydroxyvitamin D stimulation of specific membrane proteins in chick intestine.

Vitamin D-deficient chicks were injected intracardially with physiological doses of 1,25-dihydroxycholecalciferol (1,25-(OH)2D3) and the formation of intestinal brush-border proteins was followed in vitro. Within 4 h of receiving the hormone the incorporation of radioactive leucine into at least two proteins in the brush-borders was increased. The apparent molecular weights of these proteins were 45 000 and 84 000. The change in the synthesis of these proteins was followed with time and compared with the concomitant changes in intestinal calcium transport. The relationship of these changes is such that there is a strong possibility that the proteins are involved in calcium absorption.

Effect of 1,25-dihydroxyvitamin D on S-adenosylmethionine decarboxylase in chick intestine.

The effect of 1,25-dihydroxyvitamin D on the activity of S-adenosylmethionine decarboxylase in the duodenal mucosa of vitamin D-deficient chicks was investigated. Enzyme activity increased dose-dependently in a biphasic manner with maximal responses at 1 and 6 h, due to an increase in Vmax in both cases. A second dose of 1,25-dihydroxyvitamin D, administered 6 h after the first, resulted in a significant increase in activity 1 h later, confirming the rapidity of the response. This early response was not seen with ornithine decarboxylase. The increase in S-adenosylmethionine decarboxylase activity particularly at 6 h may be due to a rise in cytosolic calcium, since hydrocortisone, an inhibitor of 1,25-dihydroxyvitamin D-stimulated calcium absorption, attenuates this enzyme's activity. Inhibitors of polyamine biosynthesis such as DL-alpha-difluoromethyl ornithine and methylglyoxal bis(guanylhydrazone) had no effect on calcium absorption, but the significance of this in evaluating the importance of 1,25-dihydroxyvitamin D stimulation of S-adenosylmethionine decarboxylase activity needs further investigation.

Some characteristics of new tissue-binding proteins for metabolites of vitamin D other than 1,25-dihydroxyvitamin D.

Protein(s) have been found in a wide range of tissues which have a high affinity for 25-hydroxycholecalciferol. Of the tissues examined only erythrocytes do not have this protein. The properties of the protein have been examined and it has been found that the association constatns range from 2 - 10(9) to 5 - 10(9) M-1 and the sedimentation constants between 5.0 and 6.0 S. It was not possible to distinguish the proteins from the different tissues by their S values, mobility on gel electrophoresis or behaviour on ion-exchange chromatography. These techniques were all used, however, to show that the tissue 25-hydroxycholecalciferol binding protein is distinct from the main plasma binding protein for this steroid and from the intestinal 1,25-dihydroxycholecalciferol-binding protein. A protein has been in the plasma of rachitic animals but not of normals, which is apparently indistinguishable from this new tissue 25-hydroxycholecalciferol-binding protein. The steroid specificity of this new binding protein has been shown to be dependent upon a C-25 hydroxyl group, and an intact conjugated double bond system. Possible functions for this protein have been briefly discussed.

Structure of chick chromosomal genes for calbindin and calretinin.

The chick chromosomal gene for calbindin (the 28,000 Mr intestinal calcium-binding protein) was cloned, and all the exons and flanking regions were sequenced. The promoter region contains typical ATAAA and GGGCGG boxes, the latter being unusual in "non-housekeeping" genes. Three polyadenylation signals are found in the calbindin gene that correspond to the three known mRNAs. Transcription termination is not efficient because homology with consensus sequences found downstream from the polyadenylation signal is weak. There are ten introns, most of which do not fall at homologous positions, neither with respect to the sixfold repeating structure of the calbindin protein, nor with respect to previously sequenced genes for calmodulin and other calcium-binding proteins. The gene for the related protein calretinin was cloned and partially sequenced. The introns are in the same positions in the calretinin and calbindin genes. The introns have apparently been inserted during the divergence of the calcium-binding protein superfamily.

Calbindin D28K expression in transfected mouse NIH3T3 cells.

Calbindin D28K (formerly known as vitamin D-dependent calcium binding protein and referred to here as calbindin) is found in a wide variety of tissues, but only in certain cells within those tissues. Apart from its ability to bind calcium, nothing is known about its function in these cells. To investigate its role we have transfected the chick calbindin cDNA into mouse NIH3T3 fibroblasts and established a new cell line where calbindin is permanently expressed. Immunofluorescence studies show that calbindin is distributed throughout the cytoplasm, and treatment of the cells with cycloheximide shows that it has a relatively long half-life within the cell. Measurements of intracellular calcium concentration using Fura-2 suggest that the presence of calbindin within the cells does not affect the increase in intracellular calcium levels which occurs in response to serum stimulation or the rate at which these return to the basal level, but that it may act as a buffer for the entry of extracellular calcium.

Response of rachitic rat bones to 1,25-dihydroxyvitamin D3: biphasic effects on mineralization and lack of effect on bone resorption.

Rachitic rats, maintained on a diet adequate in Ca and P, were treated daily with varying amounts of 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3]. At low levels (1 ng/day) 1,25-(OH)2D3 sustained a healing response equivalent to that of 25-hydroxyvitamin D3 (100 ng/day) or the parent vitamin. Above 5 ng/day administration of 1,25-(OH)2D3 resulted in an accumulation of osteoid, giving a histological appearance similar to vitamin D deficiency. The effects of 1,25-(OH)2D3 on bone did not correlate with changes in plasma Ca or inorganic phosphorus; doses that were effective in raising bone ash and reducing the amount of osteoid failed to normalize plasma Ca, whilst the amount of sterol required to normalize plasma Ca was excessive in terms of the effect on bone. 1,25-(OH)2D3 did not stimulate any of the histological parameters of bone resorption. We conclude that 1,25-(OH)2D3 can effectively heal the bone lesions of vitamin D deficiency, but that, at high concentrations, the sterol can inhibit mineralization. Furthermore, these results question the accepted role of 1,25-(OH)2D3 as a regulator of bone resorption in vivo.

Differential regulation of calbindin-D28K mRNA in the intestine and eggshell gland of the laying hen.

The effect of shell calcification and 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) on calbindin-D28K (previously known as vitamin D-dependent calcium-binding protein) and calbindin mRNA was investigated in the intestine and eggshell gland (ESG) of juvenile female chicks, laying hens and non-laying female birds with active gonads. Increasing amounts of 1,25-(OH)2D3 were fed to laying hens and juvenile birds treated with oestradiol to develop the ESG. The intestinal concentration of calbindin was increased 30-fold by 1,25-(OH)2D3 in chicks treated with oestradiol and fed a vitamin D-deficient diet. In these same animals, 1,25-(OH)2D3 had no effect on the formation of calbindin mRNA or calbindin in the ESG even though fully viable 1,25-(OH)2D3 receptors are present in this tissue. In laying birds fed adequate amounts of vitamin D3, intestinal, but not ESG, calbindin was increased by the addition of 1,25-(OH)2D3 to the diet. At the onset of egg production the concentrations of calbindin and calbindin mRNA were increased in the intestine and ESG. This increase occurred within the period of calcification of the first egg, through a process unaffected by vitamin D. Calcification of the first egg increased the concentration of calbindin in the ESG by eight- to tenfold, although the concentration of calbindin mRNA was increased by only two- to threefold. These results suggest that the induction of calbindin synthesis by 1,25-(OH)2D3 or by the egg calcification process is associated with an increase in the concentration of calbindin mRNA in the ESG and intestine.(ABSTRACT TRUNCATED AT 250 WORDS)

Immunocytochemical localization of vitamin D-dependent calcium-binding protein in the growing chick lymphoid organs.

Monospecific antiserum against chick duodenal vitamin D-dependent calcium-binding protein (D-CaBP) was used to localize this protein by the peroxidase-antiperoxidase method (PAP) in the thymus, spleen and bursa of Fabricius of normal growing chicks in 20 day old embryos; in normal growing chicks at 1, 2, 3, 4 and 6 weeks of age in chicks fed a rachitogenic diet for 4 weeks. In the normal chick thymus, D-CaBP was localized throughout the cytoplasm and in the nucleus of cortical epithelial reticular cells (ERC) and in Hassal's corpuscles of the medulla. In the normal spleen reticular cells of the marginal zones showed dense deposition of reaction product in the nucleus and throughout the cytoplasm. In the bursa of Fabricius, only a few scattered cells in the medulla showed some staining. Wide variation was encountered in D-CaBP staining in the thymus and spleen of 4 week old chicks from different broods. In 4 week old rachitic chicks, staining in the thymus and in the spleen was generally reduced in intensity and occasionally was entirely absent. The presence of D-CaBP in some reticular cells of the thymus, spleen and bursa of Fabricius, identifies these lymphoid organs as vitamin D3 targets. Thus, 1,25(OH)2D3 may have an important role in some aspects of the immune defence mechanism.

Rat brain synthesizes two 'vitamin D-dependent' calcium-binding proteins.

Two proteins from rat brain reacting against anti-chick intestinal vitamin D-dependent calcium-binding protein were characterized in terms of their mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and their molecular size. The proteins were present in the isolated cytoplasm and were produced following translation of brain mRNA in the rabbit reticulocyte lysate system. Their apparent molecular weight was 29,000 and 27,000 daltons whereas rat kidney contained only one protein cross-reacting with this antiserum and with a molecular weight of 27,000 daltons.

Loss of calbindin-28K immunoreactive neurones from the cortex in Alzheimer-type dementia.

An antibody raised against chick intestinal calbindin D28K was used to study the number and size of calbindin immunoreactive neurones in postmortem human brains from neurologically normal controls and from patients with neuropathologically diagnosed Alzheimer-type dementia (ATD). In the controls, calbindin immunoreactive neurones were observed in all cerebral cortex areas examined including the frontal, temporal and parietal cortices. When compared with the controls, the number and size of calbindin immunoreactive neurones were significantly reduced in the cortices of patients with ATD. These findings suggest that calbindin containing neurones are affected in ATD.

Ultrastructural localization of brain 'vitamin D-dependent' calcium binding proteins.

Rat brain vitamin D-dependent calcium-binding protein (D-CaBP) was assessed for vitamin D dependency, calcium binding and ultrastructural localization within neurons. No evidence of vitamin D dependency could be derived from the experiments on vitamin D-deficient rats. A 95% pure extract of the 27-kDa brain D-CaBP was shown to bind 45Ca on nitrocellulose membrane after sodium dodecyl sulphate-electrophoresis, specifically on the 27-kDa CaBP band. Immunogold staining with electron microscopy allowed detection of D-CaBP into Purkinje cells and climbing fibers of the cerebellum. The immunoreactivity was found to be hyaloplasmic and never membrane-bound. It was present in neuronal soma, neurites and postsynaptic as well as presynaptic terminals. These findings rule out D-CaBP as a possible neurotransmitter and bring further support to the hypothesis that the protein functions as a cytosolic calcium buffer. Immunohistochemical detection of D-CaBP is proposed as a means for morphologic detection of neurons with high calcium metabolism.

Calbindin-immunoreactive cholinergic neurones in the nucleus basalis of Meynert in Alzheimer-type dementia.

An antibody to the calcium binding protein, calbindin D28K (CaBP), was used to study the number and size of CaBP-immunoreactive neurones in the nucleus basalis of Meynert (nbM) of postmortem human brains from neurologically normal controls and from patients with neuropathologically diagnosed Alzheimer-type dementia (ATD). In controls, almost all the large neurones and their processes in the nbM were CaBP immunoreactive. Compared to neurologically normal controls the number of CaBP-immunoreactive neurones in the nbM in patients dying with ATD was significantly reduced and there was a clear loss of the majority of CaBP immunoreactive neurones. The few remaining nbM CaBP immunoreactive neurones in the ATD cases were smaller than those in the neurologically normal controls. Double-staining experiments revealed that many of the nbM CaBP-immunoreactive neurones contained choline acetyltransferase immunoreactivity, so that CaBP is an alternative marker for the nbM cholinergic neurones in the human fore-brain. These findings suggest that a disturbance in calcium homeostasis may be a possible factor contributing to the loss of these cholinergic/CaBP-containing neurones.

Cerebral cortical calbindin D28K and parvalbumin neurones in Down's syndrome.

Anti-calbindin D28K (CaBP) and anti-parvalbumin (PVA) antibodies were used to study the number and size of neurones containing these two calcium binding proteins in post-mortem brains from 7 neurologically normal controls and from 4 elderly patients with clinically diagnosed Down's syndrome (DS) and whose brains contained numerous senile plaques and neurofibrillary tangles. The possible co-existence of these two calcium binding proteins in human cerebral cortex was also examined. In the controls, CaBP immunoreactive neurones were mainly non-pyramidal neurones although some pyramidal neurones were also CaBP immunoreactive. All the PVA immunoreactive neurones were non-pyramidal cells. CaBP and PVA did not apparently co-exist with each other in cortical neurones. When compared with the neurologically normal controls, the number and size of CaBP and PVA immunoreactive neurones were significantly reduced in the cortex of patients with DS. These findings show that CaBP and PVA containing cortical neurones are affected in elderly persons with DS.

Role of nutrition in the development of osteomalacia in the elderly.

A dietary survey has been carried out of elderly people with osteomalacia. The commonest type of osteomalacia was that associated with partial gastrectomy. Although a dietary deficiency of vitamin D has been suggested as a cause of the osteomalacia the intake of this substance was not significantly different from that in controls to explain the development of the disease. It was found, however, that patients with osteomalacia went outside much less than controls and given the contribution of sunlight to meeting vitamin D requirements it is suggested that further consideration should be given to this factor in the aetiology of post-gastrectomy osteomalacia.

Calcium-binding protein immunocytochemistry in organotypic cultures of cerebellum.

Vitamin D-dependent calcium-binding protein is known to be present in specific classes of neurons including Purkinje cells of the rat and chick. Explant cultures of newborn mouse cerebellum consisting of cerebellar cortex and deep nuclear region were fixed at maturity (20 days in vitro). Paraffin sections were reacted with the antiserum to purified chick duodenal calcium-binding protein. The Purkinje cell of these cultures reacted in its entirety--neuronal soma, dendrite, axon and terminals (in the deep nuclear region). The results verified many of the findings of our previous morphological studies. Qualitatively similar results were obtained with cultures maintained either in medium containing serum and embryo-extract or in a defined medium. There is not yet sufficient data to indicate whether this protein in the neurons in culture is dependent upon an exogenous source of vitamin D. This immocytochemical marker should prove useful to identify a specific neuron-type in culture.

Estimation of vitamin D3 and 25-hydroxyvitamin D3 in muscle and adipose tissue of rats and man.

An assay is described based on a high pressure liquid chromatography system for measurement of vitamin D3 and 25-hydroxyvitamin D3 concentration in adipose tissue and muscle. The sensitivity of the assay is less than 1 ng/g of tissue. Neither vitamin D3 nor 25-hydroxyvitamin D3 were found in tissues of vitamin D-deficient rats using this method. 25-Hydroxyvitamin D3 could not be detected in adipose tissue and very little was found in muscle of normal rats. The mean vitamin D3 concentrations in 15 samples of human perirenal adipose tissue was 45.3 ng/g +/- 22.2 (SD) and in 6 samples of axillary tissue 115.6 ng/g +/- 52.4 (SD). Human adipose tissue contains a substantial amount of vitamin D3 and its contribution to the maintenance of vitamin D status is discussed.

A simplified competitive protein-binding assay for 25-hydroxycalciferol.

A single and reliable, competitive, protein-binding assay for plasma 25-hydroxycalciferol (25-OH-D), based on a previously described method which omits the chromatographic step, has been tested without the use of beta-lipoproteins. The accuracy is similar in the absence of beta-lipoproteins and in their presence, the recoveries being respectively 110-136% and 74-85%. The intraassay coefficient of variation is 7.23% in the absence versus 12.56% in the presence of beta-lipoproteins. The sensitivity of the described assay is 0.01 ng per assay tube and 1 mug/1 plasma. The mean plasma 25-OH-D level in eighteen normal Swiss volunteers aged 20 to 40 years is 39.5 +/- S.D. 9.3 mug/l. This level is similar to those found in Western Europe and U.S.A. subjects. Ten alcoholic patients without cirrhosis have significantly lower 25-OH-D plasma levels (13.0 +/- S.D. 8.9 mug/l) than appropriate age controls. This precise and accurate method is convenient for clinical studies since both chromatographic and beta-lipoproteins steps are avoided.