The fabrication of mono-domain highly ordered nanoporous alumina on a wafer scale by a guided electric field.

This paper describes the formation of mono-domain highly ordered nanoporous alumina on the scale of a 2 inch diameter silicon wafer by anodization of aluminium evaporated on a patterned SiO(2) mask on a silicon substrate. The position of the ordered pores correlates with holes in the SiO(2) mask, which guide the electric field during anodization and initiates pore nucleation. The technique is suitable for the production of ordered nanoporous alumina on a wafer scale and overcomes the time, cost and scale limitations of existing processes.

Effect of theophylline on calcium metabolism and circulating vitamin D metabolites.

Theophylline has been shown to induce the hepatic microsomal enzyme system. These same enzymes increase the metabolism of vitamin D and 25-hydroxyvitamin D when induced by chronic barbiturate or phenytoin administration. To assess the long-term effects of theophylline on vitamin D and calcium metabolism, young rats were treated for 4 weeks with constant subcutaneous theophylline infusions. Theophylline-treated animals had a significantly increased urinary calcium excretion (p less than 0.0001), a significantly decreased total body calcium per gram body weight (p less than 0.05), and significantly decreased serum 25-hydroxy-vitamin D concentrations (p less than 0.002) when compared to control animals. These alterations in the concentration of 25-hydroxyvitamin D may impair the ability to increase 1,25-dihydroxyvitamin D-dependent intestinal calcium absorption to compensate for excessive urinary calcium losses. These data suggest that theophylline promotes skeletal calcium loss, and its use may be a risk factor for the development of osteopenia in humans.

A 1,25-dihydroxyvitamin D3 receptor-like protein in mammalian and avian liver nuclei.

The actions of 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] are thought to be mediated through receptor proteins which have been described in a variety of avian and mammalian tissues, but not in the liver. To determine if a binding protein for 1,25-(OH)2D3 is present in this tissue, rat liver was homogenized in a low ionic strength buffer containing 10 mM Tris (pH 7.4), 2.2 m sucrose, 3 mM calcium chloride, 0.2% Triton X-100, and 0.04% Trasylol (sucrose buffer) and centrifuged over a 10-ml cushion of sucrose buffer at 61,000 x g for 80 min at 4 C. The resultant nuclear pellet was extracted in a 26 mM Tris (pH 7.4) buffer containing 0.3 M potassium chloride, 5 mM dithiothreitol, 1 mM EDTA, and 10 mM sodium molybdate. Saturable 1,25-(OH)2D3 binding was identified in high salt extracts of rat liver nuclei and was eliminated by treatment with trypsin. This liver binding protein cosediments on high salt 5-20% sucrose density gradients with the 1,25-(OH)2D3 receptor protein from intestine and is distinct from the 6.OS tissue binding protein for 25-hydroxyvitamin D3. Perfusion of rat liver with PBS to remove receptor-positive blood cells before isolation of the nuclei did not change 1,25-(OH)2D3 binding. The nuclear protein bound 1,25-(OH)2D3 more avidly than either 24,25-(OH)2 D3 or 25-hydroxyvitamin D3. Saturation analysis of 1,25-(OH)2D3 binding revealed an apparent equilibrium dissociation constant of 20.6 +/- 2.2 pM (mean +/- SEM) at 4 C and a maximum binding capacity of 49.0 +/- 14.6 fmol/extract from 1.0 mg DNA. The 1,25-(OH)2D3-binding binding protein was present in liver nuclei isolated from mice, rabbits, and chicks and in nuclei isolated from cultured rat hepatocytes. The ligand specificity, sedimentation coefficient, limited binding capacity, trypsin sensitivity, and nuclear location of the hepatic 1,25-(OH)2D3-binding protein are similar to those of 1,25-(OH)2D3 receptors described in other tissues and suggest that the liver may be a target organ for [1,25-(OH)2D3] action.

Estrogen regulation of the nuclear 1,25-dihydroxyvitamin D3 receptor in rat liver and kidney.

We previously identified a receptor protein for 1,25-dihydroxyvitamin D3 in rat liver nuclei. The present studies were undertaken to investigate the ontogenesis of the hepatic nuclear vitamin D receptor (nVDR) and the estrogen regulation of this receptor in the liver, small intestine, and kidneys. The hepatic nVDR was significantly elevated in adult female rats compared to prepubertal female rats, while in male rats, this increase was not observed. Oophorectomized rats contained significantly less hepatic nVDR than did intact female rats. Administration of estradiol to castrated male or oophorectomized rats increased the hepatic nVDR. Further studies demonstrated that the increase in the hepatic nVDR was observed only after 2 weeks of estradiol treatment and was positively correlated with circulating estradiol concentrations. Castration of male rats did not alter the hepatic nVDR compared to intact male rats nor did testosterone administration to castrated male rats for 4 weeks change the hepatic nVDR concentration. Unlike the liver, intact female rats contained significantly less renal nVDR than did kidneys from intact male or castrated male rats. Estradiol administration to oophorectomized rats significantly decreased the renal nVDR. Renal nVDR concentrations correlated inversely with the serum concentration of estradiol. Castration of male rats had no effect on the renal nVDR. Intestinal nVDR concentrations were unaffected by castration of male rats or by treatment of castrated male rats with estrogen for up to 4 weeks. These results indicate that estradiol increases the nVDR in liver, decreases the nVDR in kidney and does not change the nVDR in the intestine. Physiological concentrations of testosterone do not regulate the nVDR in these tissues. Estradiol regulation of this receptor is organ specific and, therefore, conclusions about the regulation of the nVDR in one tissue cannot be extrapolated to other tissues.

Hospitalizations for fractures after renal transplantation in the United States.

PURPOSE: To investigate the incidence, risk factors, and associated mortality of fractures in renal transplant recipients. METHODS: Retrospective registry study of 33,479 patients in the United States Renal Data System (USRDS) who received kidney transplants between 1 July 1994 and 30 June 1997. Associations with hospitalizations for a primary discharge diagnosis of fractures (all causes) were assessed. RESULTS: Renal transplant recipients had an adjusted incidence ratio for fractures of 4.59 (95% confidence interval 3.29 to 6.31). In multivariate analysis, recipients with prevalent fractures, as well as recipients who were Caucasian, women, in the lower quartiles of recipient weight (<95.9 kg), had end stage renal disease caused by diabetes, and had prolonged pretransplant dialysis were at increased risk for hospitalization because of fractures after transplantation. Recipients hospitalized for hip fractures had decreased all-cause survival (hazard ratio for mortality 1.60, 95% CI 1.13 to 2.26) in Cox Regression analysis. CONCLUSIONS: In the early post-transplant course (<3 years), renal transplant recipients had a greater incidence of fractures than the general population, which were associated with decreased patient survival. Preventive efforts should focus on recipients with the risk factors identified in this analysis, most of which can be easily obtained through history and physical examination.

The effects of lactation on bone mineral content in healthy postpartum women.

Bone mineral contents were estimated by dual photon absorptiometry of the lumbar spine (L2-L4) and single photon absorptiometry of the mid- and distal radius in 19 healthy women on their second postpartum day and at 6 months postpartum. All bone mineral measurements were performed by one technician, and the single and dual photon absorptiometry results were read by one observer. Daily oral calcium intakes were estimated from dietary histories obtained by a dietitian. Twelve women who breast-fed exclusively throughout the first 6 months postpartum were compared with seven formula-feeding women who did not breast-feed or who breast-fed for less than 3 months postpartum. No differences were found in age, parity, height, weight, or daily calcium intake between the breast- and formula-feeding women. Breast-feeding women had a significant decrease (averaging 6.5%) in bone mineral of the lumbar spine at 6 months postpartum as compared with 2 days postpartum (1.14 +/- 0.03 versus 1.22 +/- 0.03 g/cm2, mean +/- SEM; P less than .001), whereas no significant change occurred in the formula-feeding women at 6 months (1.24 +/- 0.03 versus 1.26 +/- 0.04 g/cm2). At 6 months postpartum, the breast-feeding women had a significantly lower mean bone mineral content of the lumbar spine than did formula-feeding women (P less than .05). No significant changes were noted in bone mineral content of the mid- or distal radius in either group of women during the period of evaluation. We conclude that during the first 6 months postpartum, breast-feeding is associated with bone mineral loss from the lumbar spine, but not from the mid- or distal radius.

Androgen deprivation in men with prostate cancer is associated with an increased rate of bone loss.

The objective of this work was to determine the effect of androgen deprivation therapy (ADT) on rates of bone mineral density (BMD) loss in men with prostate cancer. It was a prospective study comparing men receiving ADT to age matched controls for 2 y. Subjects received a history, physical exam, bone mineral density measurement, and laboratory evaluation every 6 months. Thirty-nine subjects receiving continuous ADT for prostate cancer (subjects) were compared to 39 age-matched controls not receiving ADT (controls). Twenty-three subjects and 30 controls completed the study through 24 months. Men in the ADT group demonstrated greater rates of bone mineral density loss than men in the control group at every site except the lumbar spine. Twenty-four month per cent of bone mineral density loss is presented as mean+/-standard error (s.e.). At the distal forearm, the ADT group value was -9.4%+/-1.0% and -4.4%+/-0.3% for controls (P<0.0005). The ADT group femoral neck values were -1.9%+/-0.7% and 0.6%+/-0.5% in the control group (P=0.0016). The ADT group total hip value was -1.5%+/-1.0% and 0.8%+/-0.5% in the control group (P=0.0018). The ADT group trochanter value was -2.0%+/-1.3% and -0.1%+/-0.5% in the control group (P=0.0019). The ADT group lumbar spine value was -0.2%+/-0.8 % and 1.1%+/-0.6% in the control group (P=0.079). Our data demonstrate greater rates of bone mineral density loss in men receiving androgen deprivation therapy for prostate cancer.

Influence of clinical characteristics and parameters associated with thyroid hormone therapy on the bone mineral density of women treated with thyroid hormone.

Reports of reduced bone mineral density (BMD) in patients receiving long-term replacement and suppression therapy with L-thyroxine have generated considerable interest and controversy. A substantial literature has evolved, with interpretation of conflicting results obscured by a variety of confounding factors. We examined the BMD measurements of 202 white women who were taking thyroid hormone to determine the contribution to BMD of a number of clinical characteristics and parameters associated with thyroid hormone therapy. Measurements of BMD (N = 335 over 2.6 +/- 1.6 years) of the spine (L2-L4) were performed in 195 subjects. The BMD of three sites of the hip was measured (N = 247 over 1.8 +/- 1.1 years) in 157 subjects. The BMD of the proximal radius was also measured (N = 172 over 1.8 +/- 1.2 years) in 124 subjects. Increasing age and a history of previous thyrotoxicosis had a deleterious effect on spine BMD. Body mass index (BMI) was positively correlated with spine BMD. Dose of thyroid hormone, duration of therapy, type of underlying thyroid disease, history of thyroidectomy, or serum-free thyroxine index did not influence either the initial BMD or the change in spine BMD over time. In the hip, age correlated with a decrease, and BMI with an increase in BMD. A history of previous thyrotoxicosis was associated with a decrease in hip BMD at all three sites (0.05 < p < 0.10). No other clinical parameters significantly influenced either the initial BMD or the change in hip BMD over time. Increasing age and dose of thyroid hormone, and a prior history of thyrotoxicosis had a deleterious effect on the BMD at the proximal radius. In summary, thyroid hormone therapy was not associated with a significant effect on BMD of the spine or hip, but a decreased BMD of the proximal radius was related to both previous thyrotoxicosis and to dose of thyroid hormone.

Artificial pancreas as an aid during insulinoma resection.

Blood glucose control by an artificial pancreas facilitated the intraoperative management of two patients with insulin-secreting pancreatic islet cell tumors (one had a large adenoma, the other diffuse islet-cell hyperplasia). The artificial pancreas is a glucose-controlled insulin and dextrose infusion system uniquely suited for the control of blood glucose concentrations during surgery. The artificial pancreas produced a stable glycemic baseline before surgery, prevented hypoglycemia during palpation of the pancreas and manipulation of the tumor, and indirectly amplified small decreases in the blood sugar level with large increases in the rate of dextrose infusion. The advantages and limitations of continuous glucose monitoring during surgery and the use of this instrument in locating occult insulin-secreting tumors are reviewed.

Decreased turnover of soluble liver proteins in mice with alloxan-induced diabetes.

Mice with alloxan-induced diabetes were used as a model to assess whether the synthesis and/or degradation of soluble liver proteins in general is affected in vivo by the diabetic state. Protein turnover was measured 2-3 wk after diabetes was induced. Degradation of liver cytosol proteins was decreased in diabetic mice as measured by the loss of protein radiolabeled with [14C]bicarbonate. The incorporation of radiolabeled amino acids into protein was also decreased in diabetic mice. When [3H]leucine was administered as the precursor for protein synthesis, the radiospecific activity of leucine derived from leucyl-tRNA in livers was similar in control and diabetic mice. Thus, the rate of protein synthesis appears to be decreased. There was no indication that diabetes affected the turnover of long- or short-lived proteins differentially. The activities of several cellular proteinases were unaffected or slightly decreased in livers of diabetic mice. These data indicate that protein turnover is decreased in this chronic form of diabetes.

In vitro dissolution of calcium carbonate preparations.

Calcium supplements are widely used for the treatment of osteoporosis. The bioavailability of these preparations is unknown. Because poor tablet dissolution accounts for a majority of drug bioavailability problems, we determined the in vitro dissolution at 30, 60, and 90 minutes of 27 commercially available calcium carbonate supplements using the method of the U.S. Pharmacopoiea. At 30 minutes, five preparations (18%) were more than 75% dissolved, four (15%) between 33 and 74%, and the remaining 18 (67%) were less than 33% dissolved. After 90 minutes, 17 (63%) of the preparations were less than 50% dissolved. Dissolution correlated negatively with the weight of filler (noncalcium carbonate material in the tablet) (rs = -0.51, P less than 0.01) but not with tablet hardness or cost. Similar to previous studies, we also found no correlation of dissolution with the stated calcium content, chemical source of calcium carbonate (oyster shell or chemical precipitate), or retail source. We conclude that there is a wide range of in vitro dissolution among the calcium carbonate preparations tested, and that the filler is an important determinant of the dissolution of these tablets. These results raise concern about the bioavailability of the calcium in these preparations and may have important implications for the therapeutic use of the various calcium carbonate supplements.

Normal rabbit intestinal cytosol as a source of binding protein for the 1,25-dihydroxyvitamin D3 assay.

Cytosol prepared from small intestine of vitamin D-sufficient rabbits contains a specific high-affinity binding protein for 1,25-dihydroxyvitamin D3 (1,25(OH)2D3). This binding protein sediments at 3.0-3.5 S in sucrose density gradients containing 0.3 M KCl. Scatchard analysis using intestinal cytosol demonstrated a Kd of 0.05 nM and a maximum binding capacity of 92 fmol/mg cytosol protein for 1,25(OH)2D3 at 4 degrees C. Competitive binding studies with various metabolites of vitamin D showed a relative binding affinity of this protein for 1,25(OH)2D3 greater than 25-hydroxyvitamin D3 greater than vitamin D3. With 200 micrograms of rabbit intestinal cytosol protein, as little as 1.0-2.5 pg of 1,25(OH)2D3 reproducibly displaced the tracer sterol from the binding protein. Analyses of human plasma 1,25(OH)2D3 content yielded values consistent with published results. The vitamin D-replete rabbit provides a convenient, plentiful, and inexpensive source of binding protein for 1,25(OH)2D3 assays.

Ontogenesis of the rabbit intestinal receptor for 1,25-dihydroxyvitamin D3--evidence for increased receptor content during late suckling and lactating periods.

The 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) intestinal receptor was not detected in term fetal rabbits. This receptor was present at 2 weeks postpartum and its concentration reached a maximum at 4 weeks of age, and declined to adult levels by 10 weeks postpartum. The 1,25(OH)2D3 intestinal receptor concentration was elevated at 2 weeks postpartum in lactating rabbits, but returned to normal adult concentrations by 4 weeks postpartum. In rabbits of various ages, only minor changes in the equilibrium dissociation constant of this receptor were observed. These data indicate that increasing the small intestine 1,25(OH)2D3 receptor concentration is one mechanism by which the rabbit adapts to periods of increased calcium demand.