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James C Fleet

Publications and source records attributed to James C Fleet.

18 recordsLinked to original sources
PubMed2007

Nucleo-cytoplasmic cycling of the vitamin D receptor in the enterocyte-like cell line, Caco-2.

We examined the effects of 1,25 dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) on the distribution and mobility of the vitamin D receptor (VDR) in the enterocyte-like Caco-2 cell. Confocal microscopy showed that a green fluorescent protein-vitamin D receptor (GFP-VDR) fusion protein is predominantly nuclear (58%) and it does not associate with the apical or basolateral membrane of proliferating or polarized, differentiated cells. In contrast to the previously studied cell types, neither endogenous VDR nor GFP-VDR levels accumulate in the nucleus following 1,25(OH)(2)D(3) treatment (100 nM, 30 min). However, in nuclear photobleaching experiments nuclear GFP-VDR import was significantly increased by 1,25(OH)(2)D(3) during both an early (0-5 min) and later (30-35 min) period (20% per 5 min). Compared to the natural ligand, nuclear import of GFP-VDR was 60% lower in cells treated with the 1,25(OH)(2)D(3) analog, 1-alpha-fluoro-16-ene-20-epi-23-ene-26,27-bishomo-25-hydroxyvitamin D(3) (Ro-26-9228, 5 min, 100 nM). Downstream events like ligand-induced association of VDR with chromatin at 1 h and the accumulation of CYP24 mRNA were significantly lower in Ro-26-9228 treated cells compared to 1,25(OH)(2)D(3) (60 and 95% lower, respectively). Collectively our data are consistent with a role for ligand-induced nuclear VDR import in receptor activation. In addition, ligand-dependent VDR nuclear import appears to be balanced by export, thus accounting for the lack of nuclear VDR accumulation even when VDR import is significantly elevated.

Base Sequence↗
PubMed2007

Transgenic expression of the human Vitamin D receptor (hVDR) in the duodenum of VDR-null mice attenuates the age-dependent decline in calcium absorption.

1,25(OH)(2)D(3) regulates calcium homeostasis through its actions in the intestine, bone, and kidney. These actions are mediated through the VDR. To determine if VDR's actions in the proximal small intestine can sufficiently restore calcium homeostasis, we generated transgenic mice expressing hVDR exclusively in the duodenum of mVDR-null mice by using the adenosine deaminase enhancer (hVDR+/mVDR-null). Unlike wild-type mice, hVDR+/mVDR-null mice express hVDR and VDR target genes only in the proximal small intestine. Despite having functional hVDR in the proximal small intestine, hVDR+/mVDR-null mice were hypocalcaemic when fed a normal rodent diet at weaning, like mVDR-null mice fed the same diet. The hypocalcemia in these mice is prevented if they are given the rescue diet before weaning. However, when 90-day-old rachitic mice were fed a rescue diet, serum calcium improved in hVDR+/mVDR-null mice, but not in mVDR-null mice. In conclusion, transgenic hVDR in the proximal small intestine of hVDR+/mVDR-null mice was transcriptionally active and regulated calcium absorption, but VDR actions elsewhere in the intestine are probably necessary to support adequate calcium homeostasis. In addition, hVDR+/mVDR-null mice responded better to the late rescue diet suggesting that expression of VDR in the proximal small intestine protected the calcium absorbing machinery from age-dependent decline.

Aging↗
PubMed2006

Intestinal resistance to 1,25 dihydroxyvitamin D in mice heterozygous for the vitamin D receptor knockout allele.

We tested the hypothesis that low vitamin D receptor (VDR) level causes intestinal vitamin D resistance and intestinal calcium (Ca) malabsorption. To do so, we examined vitamin D regulated duodenal Ca absorption and gene expression [transient receptor potential channel, vallinoid subfamily member 6 (TRPV6), 24-hydroxylase, calbindin D(9k) (CaBP) mRNA, and CaBP protein] in wild-type mice and mice with reduced tissue VDR levels [i.e. heterozygotes for the VDR gene knockout (HT)]. Induction of 24-hydroxylase mRNA levels by 1,25 dihydroxyvitamin D(3) [1,25(OH)(2) D(3)] injection was significantly reduced in the duodenum and kidney of HT mice in both time-course and dose-response experiments. TRPV6 and CaBP mRNA levels in duodenum were significantly induced after 1,25(OH)(2) D(3) injection, but there was no difference in response between wild-type and HT mice. Feeding a low-calcium diet for 1 wk increased plasma PTH, renal 1alpha-hydroxylase (CYP27B1) mRNA level, and plasma 1,25(OH)(2) D(3), and this response was greater in HT mice (by 88, 55, and 37% higher, respectively). In contrast, duodenal TRPV6 and CaBP mRNA were not higher in HT mice fed the low-calcium diet. However, the response of duodenal Ca absorption and CaBP protein to increasing 1,25(OH)(2) D(3) levels was blunted by 40% in HT mice. Our data show that low VDR levels lead to resistance of intestinal Ca absorption to 1,25(OH)(2) D(3), and this resistance may be due to a role for the VDR (and VDR level) in the translation of CaBP.

Animals↗
PubMed2005

1,25-Dihydroxyvitamin D and 25-hydroxyvitamin D--mediated regulation of TRPV6 (a putative epithelial calcium channel) mRNA expression in Caco-2 cells.

BACKGROUND: TRPV6 is a member of the vanilloid subfamily of transient receptor potential (TRP) proteins and likely functions as an epithelial calcium channel in calcium-transporting organs, such as the intestine, kidney, and placenta. TRPV6 mRNA expression is strongly regulated by 1,25-dihydroxyvitamin D (1,25VD), the active hormonal form of vitamin D, in intestine and in Caco-2 cells, a human colon cancer cell line. AIM OF THE STUDY: The aim of the present study was to characterise the mode of regulation of the 1,25VD-mediated TRPV6 mRNA expression and to test the effect of the precursor of 1,25VD namely 25 hydroxyvitamin D (25 VD) on TRPV6 mRNA expression in Caco-2 cells. METHODS: Caco-2 cells were treated in a 2 x 2 format with 1,25VD and the transcriptional inhibitor actinomycin D (AD, 4 microg/ml), and also with translational inhibitor cycloheximide (CHX, 10 microg/ml) after 14 days in culture and TRPV6 mRNA levels were determined using reverse transcription-real time PCR.TRPV6 mRNA half life studies were performed by inhibiting transcription followed by sampling at various time points for TRPV6 mRNA. Varying concentrations of 25 VD were used to test their effect on TRPV6 mRNA in the presence of 5% FBS and also in the absence of serum (but containing insulin-transferrin-selenium mixture) for 24 h. RESULTS: Treatment with 10(-7) M 1,25VD for 8 h resulted in a 60-fold increase in TRPV6 mRNA and this increase could be completely blocked with AD. Treatment with CHX to inhibit de novo protein synthesis did not prevent the initiation of 1,25VD-induced TRPV6 expression, although it did reduce the extent of TRPV6 mRNA accumulation. We found that TRPV6 mRNA half-life was 8 h in Caco-2 cells and was not altered by 1,25VD treatment. Finally, we observed that treatment with 10(-6) M of the pro-hormone 25 VD for 24 h resulted in a significant increase in TRPV6 expression in Caco-2 cells, which is consistent with the presence of 1alpha-hydroxylase (CYP27B1) expression in Caco-2 cells and a possible autocrine vitamin D signaling pathway in colon cells. CONCLUSIONS: 1,25 dihydroxyvitamin D regulates TRPV6 expression by a process that requires new mRNA and protein synthesis and the point of regulation lies likely at the transcriptional level especially since vitamin D did not increase the half life of TRPV6 mRNA. In addition, the prohormone form of 1,25 dihydroxyvitamin D, i. e. the 25 hydroxyvitamin D, induced TRPV6 mRNA expression in Caco-2 cells.

Caco-2 Cells↗
PubMed2004

Control of differentiation-induced calbindin-D9k gene expression in Caco-2 cells by cdx-2 and HNF-1alpha.

Calbindin D9k (CaBP) is critical for intestinal calcium absorption; its in vivo expression is restricted to differentiated enterocytes of the small intestine. Our goal was to identify factors controlling the transcriptional regulation of this gene in the human intestine. Both the natural gene and a 4600-bp promoter construct were strongly regulated by differentiation (>100-fold) but not by treatment with 1,25(OH)2 vitamin D (<2-fold) in the Caco-2 clone TC7. Deletion-mutation studies revealed that conserved promoter sequences for cdx-2 (at -3158 bp) and hepatocyte nuclear factor (HNF)-1 (at -3131 and at -98 bp) combined to control CaBP expression during differentiation. Other putative response elements were not important for CaBP regulation in TC7 cells (CCAAT enhancer binding protein, pancreatic duodenal homebox-1 (pdx-1), a proximal cdx-2 element). Mutation of the distal HNF-1 site had the greatest impact on CaBP gene expression through disruption of HNF-1alpha binding; both basal and differentiation-mediated CaBP expression was reduced by 80%. In contrast, mutation of the distal cdx-2 element reduced only basal CaBP expression. Whereas a 60% reduction of CaBP mRNA in the duodenum of HNF-1alpha null mice confirmed the physiological importance of HNF-1alpha for CaBP gene regulation, additional studies showed that maximal CaBP expression requires the presence of both HNF-1alpha and cdx-2. Our data suggest that cdx-2 is a permissive factor that influences basal CaBP expression in enterocytes and that HNF-1alpha modulates CaBP gene expression during cellular differentiation.

Animals↗
PubMed2004

Effect of cellular environment on the selective activation of the vitamin D receptor by 1alpha,25-dihydroxyvitamin D3 and its analog 1alpha-fluoro-16-ene-20-epi-23-ene-26,27-bishomo-25-hydroxyvitamin D3 (Ro-26-9228).

The vitamin D analog, 1alpha-fluoro-16-ene-20-epi-23-ene-26,27-bishomo-25-hydroxyvitamin D(3) (Ro-26-9228) is tissue selective, with a gene regulation preference for bone over duodenum in vivo. In the human osteoblast-like cells, hFOB, the vitamin D receptor (VDR)-mediated transcriptional potencies of Ro-26-9228 and 1,25-dihydroxyvitamin D(3) (1,25D(3)) were similar, but in the intestinal cells, Caco-2, transcriptional potency of Ro-26-9228 was 10-50 times lower. We hypothesized that transcriptional activation of the VDR by Ro-26-9228 in the two cell types is regulated differently, and compared VDR extracted from hFOB or Caco-2 cells for their abilities to interact with a p160 coactivator [glucocorticoid receptor-interacting protein (GRIP)] and with retinoid X receptor (RXR) by pull-down assays. 1,25D(3) had similar potencies to induce interactions of VDR from the two cell types with these partners of transcription. In contrast, Ro-26-9228 induced interaction of osteoblastic VDR with RXR and GRIP but did not induce these interactions with VDR from Caco-2 cells. Further studies revealed that in hFOB cells the unoccupied VDR was cytoplasmic and proteasome sensitive, and that ligand treatment caused a rapid accumulation of the VDR in the chromatin. Both cytoplasmic and chromatin-associated ligand-bound VDR from hFOB cells had the abilities to interact with GRIP. In contrast, in Caco-2 cells, unoccupied VDR was localized in both the cytoplasm (70%) and the chromatin (30%). In Caco-2 cells, the cytoplasmic VDR was proteasome resistant, and neither 1,25D(3) nor Ro-26-9228 induced its binding to GRIP. Only a small fraction of the chromatin-associated VDR was proteasome sensitive, and this fraction was distinguishable by a faster electrophoretic mobility. 1,25D(3) induced an accumulation of the proteasome-sensitive VDR in the chromatin of Caco-2 cells and binding to GRIP. Ro-26-9228 failed to induce accumulation of the proteasome-sensitive VDR in the chromatin or binding to GRIP, but a coincubation of Caco-2 cells with the analog and a proteasome inhibitor restored these abilities. These results suggest that Ro-26-9228 has poor ability to promote the accumulation of a proteasome-sensitive, transcriptionally active VDR isoform in Caco-2 cells, whereas it does not have this limitation in hFOB cells.

Caco-2 Cells↗
PubMed2004

Genomic and proteomic approaches for probing the role of vitamin D in health.

Although we have learned a great deal about vitamin D metabolism and function since it first became apparent that this factor was required for bone health, there are still many gaps in our understanding, at both the basic science (eg, the molecular actions and targets of vitamin D) and applied (eg, what "adequate" vitamin D status means) levels. For example, although the identification of extrarenal 1alpha-hydroxylase activity suggests that autocrine/paracrine actions of 1,25-dihydroxyvitamin D complement the classic endocrine actions of the hormone, the practical implications of this finding are only now being explored. In addition, studies showed that 1,25-dihydroxyvitamin D rapidly activates signal transduction pathways in addition to the classic transcriptional activation pathways that require the vitamin D receptor. These new modes of vitamin D action may be crucial to our understanding of both the traditional calcium-regulating actions of vitamin D and the anticancer actions of this essential mediator. Recent developments in genomics and proteomics have provided new opportunities for us to identify molecular targets of vitamin D action. Cancer researchers have demonstrated that these methods have utility for identifying useful biomarkers of disease states. Can these approaches be used to help clarify what constitutes optimal serum concentrations of 25-hydroxyvitamin D? I present an overview of how proteomic and genomic evaluations of cells, animals, and human subjects have been and can be used to improve our understanding of vitamin D biological processes and the role of vitamin D in health.

Biomarkers↗
PubMed2004

Vitamin D requirements: current and future.

The requirements for vitamin D were last set in 1997 by the Food and Nutrition Board of the Institute of Medicine. Intakes were assumed to come from diet, and values were based on achievement of adequate vitamin D status and on observed values to prevent seasonal variations in parathyroid hormone concentrations. Serum 25-hydroxyvitamin D concentrations were considered the best indicator of vitamin D adequacy, because the production of 25-hydroxyvitamin D is not regulated. Normal ranges were obtained from reference populations, but the values varied widely with age and geographic region. Revised recommendations should take into consideration appropriate functional measures for multiple tissues and disease risks. Vitamin D-calcium interdependencies must be taken into account. Higher intakes of both vitamin D and calcium can reduce bone resorption, and higher concentrations of one nutrient might compensate for insufficiency in the other. Better ways to assess vitamin D (and calcium) inputs are needed. Food composition databases are incomplete for both vitamin D and calcium, especially in this era of food fortification, and are complicated by the poor quality control for vitamin D fortification. Upper levels of vitamin D intake were set at 50 microg/d (2000 IU/d) for all ages. Some individuals would require higher levels than these to achieve serum 25-hydroxyvitamin D concentrations for optimal calcium absorption. So much new information on vitamin D and health has been collected since the requirements were set in 1997 that this nutrient is likely the most in need of revised requirements.

Adolescent↗
PubMed2004

Rapid, membrane-initiated actions of 1,25 dihydroxyvitamin D: what are they and what do they mean?

Vitamin D is a conditionally required nutrient traditionally thought to influence physiology as the metabolite 1,25-dihydroxyvitamin D [1,25(OH)(2) D] by binding to the vitamin D receptor (VDR) and stimulating the transcription of genes through direct VDR-DNA interactions. However, over the past 15 y research has demonstrated that 1,25(OH)(2) D, as well as other steroid hormones, can rapidly stimulate ion fluxes and activate protein kinases by transcription-independent mechanisms. This review summarizes recent research on the rapid actions of 1,25(OH)(2) D and identifies questions that remain to be answered in this area.

Animals↗
PubMed2004

1,25 dihydroxycholecalciferol-mediated calcium absorption and gene expression are higher in female than in male mice.

Recent advances in bone and calcium (Ca) metabolism have relied upon genetically modified mice. However, although human studies have identified gender as an important modulator of Ca metabolism, its effect on Ca metabolism has not been examined in mice. Here we examined basal and vitamin D-regulated Ca absorption (in situ ligated loops) and mRNA levels for the apical membrane calcium channel, TRPV6, and the calcium binding protein, calbindin D(9k) (CaBP) mRNA levels (real-time PCR) in duodenum of female and male mice. At 2 mo of age, females fed a 5 g Ca/kg diet had higher Ca absorption (62.3 +/- 4.8 vs. 47 +/- 3.6%) and TRPV6 mRNA levels than males even though plasma 1,25 dihydroxyvitamin D [1,25(OH)(2) D] was not different. In mice fed high (20 g/kg), normal (5 g/kg), or low (0.2 g/kg) Ca diets for 7 d to alter plasma 1,25(OH)(2) D (91 +/- 12, 322 +/- 25, and 587 +/- 43 pmol/L, respectively), the relation between Ca absorption (slope = 0.116 vs. 0.084, P = 0.021) or duodenum TRPV6 mRNA (slope = 0.042 vs. 0.025, P = 0.034) and circulating 1,25(OH)(2) D was steeper in females. After a single 1,25(OH)(2) D injection (200 ng/100 g body weight), peak induction of TRPV6 mRNA was 2-fold greater (at 6 h) and CaBP mRNA was 20% higher in females (at 16 h). Duodenal vitamin D receptor mRNA levels did not differ between genders. Our data indicate that female mice are more sensitive to changes in serum 1,25(OH)(2) D levels than males and that this must be considered when using mice to study calcium and bone biology.

Animals↗
PubMed2004

Short-term low-protein intake does not increase serum parathyroid hormone concentration in humans.

We investigated whether inadequate dietary protein would result in increased serum parathyroid hormone (PTH) concentration, consistent with secondary hyperparathyroidism. Data from 2 controlled feeding studies were utilized. In study 1, 26 healthy women (15 young, 21-46 y, and 11 elderly, 70-81 y) consumed for 12 d each in separate trials 3 levels of protein, 1.00, 0.75, and 0.50 g protein/(kg. d). Blood was drawn from fasting subjects on d 12 of each trial. In study 2, 24 persons (54-80 y) were fed diets with either 1.20 g protein/(kg. d) for 2 wk (HPro, n = 11, 6 men, 5 women) or 1.2 g protein/(kg. d) for 1 wk and then 0.50 g protein/(kg. d) for a 2nd week (IPro, n = 13, 6 men, 7 women). Blood was obtained from fasting subjects after wk 1 and 2. Consistent with altered protein metabolism, urinary total nitrogen excretion and blood urea nitrogen fell progressively with decreasing protein intake in study 1; in study 2, the values decreased from wk 1 to 2 in the IPro group only. Serum intact PTH concentrations did not differ among the 3 protein intakes in study 1, or between the HPro and IPro groups in study 2. These findings do not support the hypothesis that the short-term ingestion of inadequate dietary protein increases serum PTH concentration.

Adult↗
PubMed2003

Gene expression profiling of Caco-2 BBe cells suggests a role for specific signaling pathways during intestinal differentiation.

We examined the pattern of gene expression resulting from spontaneous differentiation of Caco-2 BBe cells to gain insight into the molecular changes necessary for enterocyte differentiation. RNA was prepared from cells harvested at three cell stages: proliferating (50% confluent, 2 days in culture), postproliferative nondifferentiated (8 days), and differentiated (15 days). Gene expression profiles were determined using Affymetrix Human Genome U95A GeneChips. Differentially expressed genes were identified following statistical analysis (i.e., ANOVA, bootstrapping adjustments to P values, false detection rate criterion). We identified 1,150 unique genes as differentially expressed; expression of 48.6% fell and 46% increased from 2 to 15 days, while 5.4% had expression that either peaked or dipped at 8 days. Genes expressed during differentiation included several small-intestine-specific genes involved in nutrient transport/metabolism, e.g., DCT1, hephaestin, folate receptor 1, sucrase-isomaltase, and apolipoproteins CI, CIII, B100, H, and M, indicating that this colonic adenocarcinoma cell line has a hybrid colonocyte/enterocyte phenotype. Patterns of gene expression based upon functional classification suggest a role for cell-cell/cell-matrix interactions, suppression of Wnt signaling, and activation of TGFbeta and phosphatidylinositol 3-kinase pathways during enterocyte differentiation.

Caco-2 Cells↗
PubMed2003

Vitamin D receptor (VDR) knockout mice reveal VDR-independent regulation of intestinal calcium absorption and ECaC2 and calbindin D9k mRNA.

To study the role of calbindin D(9k) (CaBP) and epithelial calcium channel ECaC2 in intestinal calcium (Ca) absorption, vitamin D receptor knockout (KO) and wild-type (WT) mice were fed either 0.5% Ca or a 2.0% Ca rescue diet starting at 21 d of age. Ca absorption and parameters involved in this process were measured at 60 or 90 d of age. Compared with WT, KO mice fed the 0.5% Ca diet had higher plasma parathyroid hormone (PTH) and 1,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)], and lower plasma Ca and insulin-like growth factor-I (IGF-I). Duodenal Ca absorption (% Ca absorbed) in KO mice was reduced 71% relative to WT mice and was associated with 55% lower CaBP mRNA, 47% lower CaBP protein and 95% lower ECaC2 mRNA levels. Compared with WT mice, the percentage of Ca absorbed in KO mice fed the 0.5% Ca diet was inappropriately low for the level of duodenal CaBP. The 2% Ca rescue diet normalized plasma Ca, prevented osteomalacia, increased growth and plasma IGF-I levels, but did not normalize plasma PTH or 1,25(OH)(2)D(3) in KO mice. In addition, the relationship between CaBP protein and the percentage of Ca absorbed was normalized, whereas ECaC2 mRNA fell to near zero. Our data demonstrate that higher CaBP levels do not ensure high rates of duodenal Ca absorption and that transcellular Ca absorption can occur even when ECaC2 gene expression is very low. In addition, our data suggest that the 2% Ca diet promotes a vitamin D receptor-independent anabolic effect on bone formation and calcium absorption, leading to improved calcium balance even in the presence of high PTH levels.

Animals↗
PubMed2003

Calcium transporter 1 and epithelial calcium channel messenger ribonucleic acid are differentially regulated by 1,25 dihydroxyvitamin D3 in the intestine and kidney of mice.

We examined the expression of calcium transporter 1 (CaT1) and epithelial calcium channel (ECaC) mRNA in the duodenum and kidney of mice. Intestinal CaT1 mRNA level increased 30-fold at weaning, coincident with the induction of calbindin-D(9k) expression. In contrast, renal CaT1 and ECaC mRNA expression was equal until weaning when ECaC mRNA is induced and CaT1 mRNA levels fall 70%. Long- and short-term adaptation to changes in dietary calcium (Ca) level and 1,25 dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] injection strongly regulated duodenal calbindin D(9k) and CaT1 mRNA. Following a single dose of 1,25(OH)(2)D(3), induction of CaT1 mRNA occurred rapidly (within 3 h, peak at 6 h of 9.6 +/- 0.8-fold) and preceded the induction of intestinal Ca absorption (significantly increased at 6 h, peak at 9 h). Neither renal CaT1 nor ECaC mRNA were strongly regulated by dietary calcium level or 1,25(OH)(2)D(3) injection. Our data indicate that CaT1 and ECaC mRNA levels are differentially regulated by 1,25(OH)(2)D(3) in kidney and intestine and that there may be a specialized role for CaT1 in kidney in fetal and neonatal development. The rapid induction of intestinal CaT1 mRNA expression by 1,25(OH)(2)D(3), and the marked induction at weaning, suggest that CaT1 is critical for 1,25(OH)(2)D(3)-mediated intestinal Ca absorption.

Animal Feed↗
PubMed2003

Vitamin D receptor alleles, periodontal disease progression, and tooth loss in the VA dental longitudinal study.

BACKGROUND: The relationship between the vitamin D receptor (VDR) genotype and periodontal disease is not known. This study compared periodontal disease progression among polymorphisms of 2 VDR genes in men in the VA Dental Longitudinal Study. METHODS: Subjects were 125 medically healthy, middle-aged men who had serial oral examinations over a mean 23-year period. Probing depth, gingival bleeding on probing, clinical attachment loss (CAL), and alveolar bone loss (ABL) from radiographs were measured at each examination. Progression of periodontal disease was defined as the percentage of teeth per decade that increased ABL by > or = 40%, and the percentage of teeth per decade that developed CAL > or = 5 mm. ApaI and TaqI polymorphisms were determined from buffy coat cells following polymerase chain reaction (PCR) amplification. Mean changes in oral status were adjusted for baseline values of smoking status, number of teeth present, and periodontal status by analysis of covariance. RESULTS: Genotype distributions were 41 AA, 58 Aa, 26 aa; and 53 TT, 46 Tt, 26 tt. The AA genotype showed the highest rates of progression of ABL (5 +/- 1% versus 1 +/- 1% and 2 +/- 1% teeth in Aa and aa, respectively; P = 0.03), CAL (37 +/- 4% versus 17 +/- 4% and 27 +/- 6% teeth; P = 0.004), and tooth loss (2 +/- 0.3 versus 1 +/- 0.3 and 1 +/- 0.4 teeth; P = 0.04). When genotype combinations were examined, progression of ABL, CAL, and tooth loss was highest in the AATT and AATt genotypes. CONCLUSIONS: The ApaI polymorphism of the VDR gene is associated with oral bone loss, clinical attachment loss, and tooth loss in older men. Analysis of the VDR alleles may prove useful for predicting periodontal disease.

Adult↗
PubMed2002

Excentric cleavage products of beta-carotene inhibit estrogen receptor positive and negative breast tumor cell growth in vitro and inhibit activator protein-1-mediated transcriptional activation.

Both retinoids and carotenoids are potentially useful chemopreventive agents. In this study we tested the effect of synthetic excentric cleavage products of beta-carotene on the growth of the MCF-7, Hs578T and MDA-MB-231 human breast cancer cells. The apo-beta-carotenoic acids (beta-apo-CA) beta-apo-14'-, beta-apo-12'-, beta-apo-10'- and beta-apo-8'-CA are structurally similar to all-trans-retinoic acid (atRA) but have different side chain lengths. Nine days of treatment with atRA inhibited MCF-7 and Hs578T cell proliferation in a dose-dependent manner. beta-apo-14'-CA and beta-apo-12'-CA significantly inhibited MCF-7 growth, whereas only beta-apo-14'-CA inhibited Hs578T growth. None of these treatments inhibited the growth of MDA-MB-231 cells. Potential mechanisms of growth inhibition, i.e., regulation of the cell cycle control proteins E2F1 and retinoblastoma protein (RB), and effect on activator protein-1 (AP-1)-mediated gene regulation were examined. beta-apo-14'-CA and atRA inhibited the expression of E2F1 protein in MCF-7 and Hs578T cells. beta-apo-14'-CA, beta-apo-12'-CA and atRA down-regulated RB protein expression in MCF-7 but not in Hs578T cells. The effect of phorbol ester-induced transcriptional activation of a collagenase promoter-reporter gene construct was strongly inhibited by 1 micromol/L beta-apo-14'-CA, atRA (MCF-7, Hs578T) or beta-apo-12'-CA (MCF-7). These effects were due neither to cellular conversion of beta-apo-CA to atRA nor to high affinity binding to the retinoid acid receptors. Thus, beta-apo-CAs were effective inhibitors of breast tumor cell proliferation, possibly mediated through down-regulation of cell cycle regulatory proteins and/or inhibition of AP-1 transcriptional activity. The ability of beta-apo-CA to regulate breast tumor cell growth independently of conversion to atRA suggests that these compounds may have fewer side effects than retinoids and, therefore, have a potential chemotherapeutic value that deserves further examination.

Antioxidants↗
PubMed2002

Vitamin D-inducible calcium transport and gene expression in three Caco-2 cell lines.

The parental cell line (P) of Caco-2 cells and two clones, BBe and TC7, were studied at 11 days postconfluence to test the facilitated diffusion model of vitamin D-mediated intestinal calcium absorption (CaTx). Nuclear vitamin D receptor (nVDR) and calbindin D(9k) (CaBP) were measured by Western blot; 1,25-hydroxyvitamin D(3) 24-hydroxylase (CYP24), CaBP, plasma membrane Ca-ATPase (PMCA), and Ca transport channel (CaT1) mRNA levels were examined by RT-PCR; and net apical-to-basolateral CaTx was examined after treating cells with vehicle or 10 nM calcitriol for 8 (mRNA levels) or 48 h (protein, CaBP mRNA, CaTx). nVDR level was lowest in BBe (38% P value) and directly related to CYP24 induction (TC7 = P, which were 1.56 times greater than BBe). nVDR was inversely related to the vitamin D-induced levels of CaT1 mRNA, CaBP mRNA, PMCA mRNA, and net CaTx, with the highest induction seen in BBe. Basal CaBP mRNA (86 times greater than P) and protein levels were highest in TC7 cells and were not associated with higher net CaTx, suggesting CaBP may not be rate limiting for CaTx in these cells.

Absorption↗