Molecular alterations of bone quality in sequesters of bisphosphonates-related osteonecrosis of the jaws.

UNLABELLED: Compared to healthy bone, the intrinsic bone materials properties in the pre-existing lamellar bone are altered in jaw bone sequesters of bisphosphonates (BP)-related osteonecrosis. INTRODUCTION: The aim of this study was to evaluate the human jaw bone quality, especially intrinsic bone material properties among sequesters of osteonecrosis of the jaw (ONJ) induced by BP. METHODS: Bone sequesters were obtained from 24 patients suffering from ONJ following a BP treatment. Within BP-exposed bone samples, benign-BP and malignant-BP groups were distinguished in relation to the underlying disease: osteoporosis and bone metastases or multiple myeloma, respectively. Healthy cadaveric cortical jaw bone samples were used as controls. The physicochemical parameters of bone samples - mineral/organic ratio, relative proteoglycan content, crystallinity, monohydrogen phosphate content, and type-B carbonate substitution - were evaluated by Raman microspectroscopy. Representative Raman spectral features of bones control and BP-exposed bone sequesters were identified with the Partial-Least-Square Discriminant Analysis (PLS-DA). RESULTS: BP-exposed bone sequesters are characterized by a significant increase of mineral to organic ratio (+12 %) and a significant decrease of relative proteoglycan content (-35 %), thus regulating initial collagen matrix mineral deposition. Structural changes on mineral components are revealed by a significant decrease of both crystallinity (-2 %) and mineral maturation (-41 %) in the BP-exposed bone sequesters compared to healthy bones. These modifications were also observed distinctly in both benign-BP and malignant-BP groups. In addition, a shift of the phosphate ν1 band was highlighted by PLS-DA between bones control and BP-exposed bone sequesters, revealing a disruption of the apatitic phosphate environment in the jaw bone sequesters. CONCLUSIONS: The present data show that jaw bone quality can be altered with an overmineralization and ultrastructural modifications of apatitic mineral in bone sequesters of BP-related ONJ.

Beta-carotene is incorporated or mobilized along with triglycerides in bovine adipose tissue in response to insulin or epinephrine.

Pasture fed cattle ingest substantial amounts of beta-carotene (beta-C). Not all of the carotenoid compound is transformed into vitamin A, but the surplus is deposited in adipose tissue (AT). The mechanisms of beta-C incorporation and mobilization are unknown. Two experiments were conducted using explants from bovine AT cultured in vitro. First, beta-C incorporation by explants from three animals was examined with different beta-C concentrations (0, 1, 5 and 20 microm) and different times of incubation (every 5 h up to 25 h). The data showed a significant increase of beta-C concentration in explants only for 20 microm beta-C. Secondly, effects of insulin and epinephrine on beta-C and triglyceride (TG) contents of explants were studied. Explants from six animals were incubated with either hormone and 0 or 20 microm beta-C for 20 h. Both TG and beta-C contents were affected positively by insulin and negatively by epinephrine. Interestingly, changes in ratios of beta-C/TG (hormone vs. control) were similar (1.7 x 10(-3) and 1.8 x 10(-3)), respectively, for insulin and epinephrine, indicating that beta-C level is directly related to TG content. We also report the presence of mRNA for beta-C 15, 15' oxygenase in bovine AT. The in vitro culture system using explants from bovine AT is a promising model to investigate factors that might affect the accumulation and metabolism of beta-C.

Expression and characterization of a murine enzyme able to cleave beta-carotene. The formation of retinoids.

Because animals are not able to synthesize retinoids de novo, ultimately they must derive them from dietary provitamin A carotenoids through a process known as carotene cleavage. The enzyme responsible for catalyzing carotene cleavage (beta-carotene 15,15'-dioxygenase) has been characterized primarily in rat intestinal scrapings. Using a recently reported cDNA sequence for a carotene cleavage enzyme from Drosophila, we identified a cDNA encoding a mouse homolog of this enzyme. When the cDNA was expressed in either Escherichia coli or Chinese hamster ovary cells, expression conferred upon bacterial and Chinese hamster ovary cell homogenates the ability to cleave beta-carotene to retinal. Several lines of evidence obtained upon kinetic analyses of the recombinant enzyme suggested that carotene cleavage enzyme interacts with other proteins present within cell or tissue homogenates. This was confirmed by pull-down experiments upon incubation of recombinant enzyme with tissue 12,000 x g supernatants. Matrix-assisted laser desorption ionization-mass spectrometry analysis of pulled-down proteins indicates that an atypical testis-specific isoform of lactate dehydrogenase associates with recombinant carotene cleavage enzyme. mRNA transcripts for the carotene cleavage enzyme were detected by reverse transcription-polymerase chain reaction in mouse testes, liver, kidney, and intestine. In situ hybridization studies demonstrated that carotene cleavage enzyme is expressed prominently in maternal tissue surrounding the embryo but not in embryonic tissues at 7.5 and 8.5 days postcoitus. This work offers new insights for understanding the biochemistry of carotene cleavage to retinoids.

Intestinal beta-carotene 15,15'-dioxygenase activity is markedly enhanced in copper-deficient rats fed on high-iron diets and fructose.

The purpose of the present work was to examine effects of the Cu-Fe interaction on intestinal beta-carotene 15,15'-dioxygenase activity when a wide range of dietary Fe (deficiency to excess) was used in relation to Cu status of rats. The effect of dietary carbohydrates was also examined since they play a role in the Cu-Fe interaction in vivo. Weanling male Sprague-Dawley rats (n 72) were divided into twelve dietary groups, which were fed on either low-, normal-, or high-Fe levels (0.9, 9.0, and 90.0 mmol Fe/kg diet respectively) combined with Cu-adequate or -deficient levels (0.94 and 0.09 mmol Cu/kg diet respectively) and with starch or fructose in the diets. The data showed that both Fe concentration and beta-carotene 15,15'-dioxygenase activity in small intestinal mucosa were enhanced with increasing dietary Fe and with Cu deficiency v. Cu adequacy. Dietary fructose did not aggravate the Fe-enhancement, related to Cu deficiency, in the small intestine; however, fructose increased the intestinal dioxygenase activity in rats fed on normal- or high-Fe diets when compared with starch controls. Thus, the highest intestinal dioxygenase activity associated with the lowest hepatic retinol (total) concentration was found in rats fed on the Cu-deficient, high-Fe, fructose-based diet. Finally, a positive linear relationship was found between the dioxygenase activity and Fe concentration in intestinal mucosa. In conclusion, the data indicate that beta-carotene 15,15'-dioxygenase activity requires Fe as cofactor in vivo and the enzyme is modulated by the three dietary components: Cu, Fe, and fructose.

Effects on cholesterol balance and LDL cholesterol in the rat of a soft-ripened cheese containing vegetable oils.

OBJECTIVE: The purpose of this study was to examine effects of a modified soft-ripened cheese containing vegetable oils on cholesterol status, using the rat as the experimental model and the traditional soft-ripened cheese as the control. METHODS: Adult male Wistar rats (approximately 370 g) were divided into two dietary groups (20 rats/group) and fed either the standard diet (STD, containing traditional cheeses made from whole milk) or the experimental diet (EXP, containing modified cheeses made from the combination of skim milk with the following fat mixture: milk fat/oleic acid-enriched sunflower oil/soybean oil mixture). Lipids of the diets came solely from cheeses (14 g/100 g diet); the EXP diet contained (3-fold) less saturated fat, (2-fold) less cholesterol, and (15-fold) more phytosterols than the STD diet. RESULTS: Although serum triglyceride and total cholesterol concentrations were not affected by the type of diet, the EXP diet resulted in a significant reduction of LDL-cholesterol (31%, p < 0.001) and a significant increase of HDL-cholesterol (11%, p < 0.05), compared to the STD diet. Thus, a marked reduction (39%) of serum LDL/HDL cholesterol ratio was observed in the EXP group (p < 0.001). In addition, the two quantitative balances (excreted/ingested) of cholesterol and total neutral sterols (for which phytosterols were excluded) were significantly higher by 183% and 174%, respectively for the EXP group, compared to the STD group (p < 0.05). On another hand, rats fed the EXP diet excreted more cholesterol than they ingested dietary cholesterol (cholesterol balance > 1), indicating that those animals eliminated some endogenous cholesterol in their feces, while the opposite was true for rats fed the STD diet (cholesterol balance < 1). Finally, fecal bile salt concentration was not significantly different between the two dietary groups. CONCLUSIONS: The partial substitution of milk fat by vegetable oils in soft-ripened cheese resulted in a decreased blood LDL/HDL cholesterol ratio and an increased fecal excretion of endogenous cholesterol and neutral sterols and, thus, markedly improved its nutritional qualities. Therefore, the consumption of the described modified cheese may meet the demand of subjects who wish to lower their risk for atherosclerosis and cardiovascular disease.

Enhanced diffraction efficiency of gratings in multilayers.

Computations with the rigorous differential method show that single gratings made by ion implantation have a diffraction efficiency in the +1 transmitted order under TE illumination of only 0.78%. The insertion of such gratings into multilayer dielectric Fabry-Perot cavities leads to an enhancement of the free-space diffraction efficiency. Different designs for the multilayer are considered. An 18.8% efficiency is reached with 11-layer mirrors. This result is obtained by optimization of the thickness of the spacer of the Fabry-Perot cavity that contains the grating and centering of the wavelength of the mirrors. The dependence of optical properties of the structure on the various optogeometrical parameters of the structure is discussed.

Beta-carotene 15,15'-dioxygenase activity is responsive to copper and iron concentrations in rat small intestine.

OBJECTIVE: Previous in vitro studies have suggested that beta-carotene 15,15'-dioxygenase is an iron-dependent enzyme. However, in vivo, it is difficult to alter iron tissue concentration by varying dietary iron because of homeostatic control. On the other hand, an interaction between iron and copper has been shown, i.e., copper-deficiency results in an increase of iron in rat liver. Therefore, we hypothesized that intestinal iron concentration could be increased by copper-deficiency. Our objective was to examine the effects of iron as affected by dietary copper on beta-carotene 15,15'-dioxygenase activity in the small intestine. METHODS: Weanling male Sprague-Dawley rats (40 to 45g) were divided into four dietary groups: two copper-adequate groups (6.0 microg Cu/g diet) and two copper-deficient groups (0.6 microg Cu/g) combined with either normal iron (44 microg Fe/g) or high iron (87 microg Fe/g). Iron and copper concentrations were determined by atomic absorption spectrophotometry and the dioxygenase activity by reverse phase HPLC. RESULTS: Intestinal copper concentration was significantly reduced (40%) by the consumption of the copper-deficient diets, but intestinal iron was not changed by doubling dietary iron in rats fed either copper-adequate or copper-deficient diets. However, as hypothesized, the two copper-deficient groups exhibited higher intestinal iron concentration (> or =137%, p<0.001) than the copper-adequate controls. In addition, intestinal beta-carotene 15,15'-dioxygenase activity was increased by 27% and 106%, respectively, for copper-deficient rats fed either normal or high iron diets, compared to the respective copper-adequate controls (p<0.01). The dioxygenase activity was not significantly affected by dietary iron in either copper-adequate or copper-deficient groups. Finally, the enzyme activity was positively correlated (r=0.67, p<0.0001) with iron concentration and negatively correlated (r=-0.49, p<0.01) with copper concentration in small intestine. CONCLUSIONS: Intestinal beta-carotene 15,15'-dioxygenase may be an iron-dependent enzyme sensitive to copper status in vivo.

beta-carotene 15,15'-dioxygenase activity and cellular retinol-binding protein type II level are enhanced by dietary unsaturated triacylglycerols in rat intestines.

The purpose of this study was to examine effects of dietary triacylglycerols on beta-carotene 15,15'-dioxygenase (EC 1.13.11.21) activity and cellular retinol-binding protein [CRBP (II)] in rats. Six groups of eight rats (7-wk old) were fed one of the following diets: standard (STD; 2.5% soybean oil), saturated (SFA; 15% hydrogenated soybean oil), monounsaturated (MUFA; 15% olive oil), polyunsaturated (PUFA; 15% soybean oil) or clofibrate (CLF; 2.5% soybean oil + 0.2% clofibrate) for 3 wk. The dioxygenase specific activities of the intestinal homogenates in the MUFA and PUFA groups fed the high fat diets were 2.4 times that of the STD group fed a low fat diet (P < 0.01), whereas the activities of the SFA and CLF groups were not significantly different from that of the STD group. The level of CRBP (II) in the intestine of the PUFA group was 1. 3-fold that of the STD group (P < 0.05), whereas there were no significant differences among the other groups. In a second experiment, the dioxygenase activity of rat intestine was followed over 3 wk of feeding the STD and PUFA diets. After the PUFA diet was consumed for 1 d, the activity was enhanced to 2.7 times the baseline level and remained thereafter at that high level, whereas the activity of the STD group remained at the low baseline level. Thus, dietary polyunsaturated triacylglycerols enhanced both beta-carotene 15,15'-dioxygenase activity and CRBP (II) level in rat intestine. These results suggest that the dioxygenase and CRBP (II) are regulated by the same mechanism involving long-chain fatty acids and their metabolites.

Characterization of beta-carotene 15,15'-dioxygenase activity in TC7 clone of human intestinal cell line Caco-2.

The purpose of this study was to identify mammalian cell line(s) which possess intrinsic enzymatic activity of beta-carotene 15, 15'-dioxygenase. This enzyme (EC1.13.11.21) converts beta-carotene to retinal (precursor of retinol and retinoic acid). To assess activity, cellular enzyme preparations were incubated with beta-carotene for 60 min; retinal formed was quantified by HPLC. Activity was not detected in IPEC-1, HepG2, HL60, Wurzburg, or parent Caco-2 cell lines. However, two subclones of Caco-2, PF11 and TC7, possessed activity (2.5 and 14.7 pmol/h.mg, respectively). Using the enzyme preparation of TC7 cells, retinal formation was linear with incubation time and protein concentration; Km and Vm values were 1.6 microM and 23.8 pmol/h.mg, respectively. In addition, when TC7 cells were maintained in serum-free medium, activity was increased 8.2-fold after 19 days of postconfluency. Finally, 48 h incubation with beta-carotene (delivered to TC7 cells in Tween 40) resulted in a 1.7-fold increase of dioxygenase activity and the appearance of vitamin A (9.3 pmol/mg protein). However, retinoic acid was not detected under our experimental conditions. In sum, the TC7 subclone of the Caco-2 cell line possesses beta-carotene 15, 15'-dioxygenase activity and thus can be useful in future investigations of human carotenoid metabolism.

Assay of beta-carotene 15,15'-dioxygenase activity by reverse-phase high-pressure liquid chromatography.

beta-Carotene 15,15'-dioxygenase catalyzes the conversion of beta-carotene into two molecules of retinal. Although this enzyme reaction is the first step in providing animals with vitamin A, there is little knowledge about its regulation in mammals. In order to facilitate studies on this enzyme, we have developed a rapid and simple assay method for the measurement of retinal formation by beta-carotene dioxygenase. All-trans-beta-carotene solubilized in aqueous solution with Tween 40 was incubated with an enzyme preparation of rat tissue at 37 degrees C for 30 min. Then, the reaction was stopped with a formaldehyde treatment followed by addition of acetonitrile. After centrifugation, the supernatant was directly subjected to high-pressure liquid chromatographic analysis of retinal. This assay method did not involve any vigorous extraction or concentration procedure. All-trans- and 13-cis-retinals, major geometric isomers found in the reaction mixture, were coeluted at 7.5 min, but they were well separated from other possible metabolites of beta-carotene such as retinoic acid, retinol, and apocarotenals. Moreover, the recovery of retinal reached more than 93% and the detection limit of standard retinal was 0.2 pmol/0.2 ml of reaction mixture. Enzyme activities of rat tissues were 694, 180, 16, 8, and approx 1 pmol retinal/mg protein/h in the intestine, liver, brain, lung, and kidney homogenates, respectively.

Stoichiometric conversion of all trans-beta-carotene to retinal by pig intestinal extract.

beta-Carotene and other provitamin A carotenoids are major dietary sources of vitamin A for humans and for many animals throughout the world. Two pathways for this oxidative reaction in mammals are: (1) central cleavage by beta-carotene 15,15'-dioxygenase (EC 1.13.11.21) to yield two molecules of retinal per molecule of beta-carotene consumed and (2) eccentric (or random) cleavage via several beta-apo carotenals to yield one molecule of retinal and various smaller fragments per molecule of beta-carotene oxidized. By the use of improved methods to minimize nonenzymatic reactions and to measure isomers of retinal, the mean experimentally measured molar ratio of retinal formed to beta-carotene consumed by pig intestinal preparations (800 and 10,000g supernatants) was 1.88 +/- 0.08, close to the theoretical value of 2.0 for central cleavage. beta-Apo carotenals, retinol, and retinoic acid were detected, if at all, in trace amounts during incubation. Thus, preparations of pig intestinal mucosa, which is considered a good physiologic model for human intestine, clearly convert beta-carotene to retinal, in large part if not solely, by central cleavage. Whether eccentric cleavage plays a greater role in vivo than it does in vitro is still uncertain.

[X-ray signs of foreign body aspiration in children]. / Röntgenzeichen der Fremdkörperaspiration bei Kindern.

Preoperative chest X-rays were taken in both postanterior and partially lateral views of 94 children with foreign-body aspiration. Additional fluoroscopy was employed in 70 patients. In 7% of the cases, the foreign body was radiopaque; in an equal amount of cases, there were no radiological findings. In the remaining results we observed: emphysema as an indirect radiological sign in two-thirds of the cases; in less than one-third, poststenotic atelectasis; pneumonia in 10%; bronchitis in 9%; pneumothorax in 2%.

[Combined treatment of hypopharyngeal carcinoma]. / Die Kombinationsbehandlung des Hypopharynxkarzinoms.

From 1978 through 1984, 120 patients with hypopharyngeal carcinomas were treated cooperatively by the University Hospitals of Erlangen. 120 out of these patients were men and 6 women. 96 patients (80%) suffered from cervical lymph node metastases, 91 (76%) were already in UICC stage IV. There were two cases with stage I, eight cases (7%) with stage II, and 19 cases (16%) with stage III. Among the 116 patients treated by causal therapy, 70 (60%) reached complete remission (CR), 34 (29.5%) partial remission (PR), and 12 (10.5%) were non-responders (NC,P). The cumulative survival at five years was 14% in 120 patients and 24% in 70 patients after CR. None of the patients with PR or NC survived more than two years. 111 patients were irradiated. Group 1: radiotherapy alone (41 patients), group 2: surgery and radiotherapy (47 patients), group 3: chemotherapy and radiotherapy (19 patients). The medium treatment period was 76, 107, and 141 days, respectively. The cumulative survival at five years was 3% in group 1, 28% in group 2, the survival at three years in group 3 was 14%. The results of postoperative radiotherapy, amounting to 36%, were better than the rate of 20% achieved by preoperative radiotherapy and surgery. A survival at five years of 46% was obtained after transoral microlaryngoscopic laser resection and postoperative irradiation, which was therefore superior to conventional operation techniques combined with radiotherapy (16%). Moreover, better functional and cosmetic results were achieved with this method. The prognosis is unfavorably influenced by a high stage, lymph node manifestations, and dedifferentiated tumor histology.

[Reversible functional disorders of the intestinal tract caused by abdominal radiotherapy]. / Reversible Funktionsstörungen des Intestinaltraktes durch abdominelle Strahlentherapie.

In 20 patients with malignancies receiving abdominal radiotherapy, tests for disturbed intestinal functions were performed at the beginning and at the end of the therapy and after 6 to 12 month following radiation. Four noninvasive tests were performed: 75Se-homotaurocholate (75SeHCAT) test for estimation of bile acid malabsorption; Schillings test for quantification of vitamin B12 absorption; H2-breath analysis before and after a test meal containing lactose as a parameter of lactose malabsorption; and 51Cr-EDTA test for estimation of intestinal permeability. Both bile acid and vitamin B12 absorption decreased significantly towards the end of abdominal radiotherapy in more than 50% of patients. Only one patient developed lactose malabsorption. After 6 to 12 month, these abnormalities had completely disappeared. In contrast, small intestinal permeability did not increase during radiotherapy but was significantly elevated 6 to 12 month following treatment as the only indication of chronic injury of small intestinal mucosa. During radiotherapy, a significant correlation existed between the severity of diarrhea and the degree of bile acid malabsorption.

[Sonography of malignant hemangioendothelioma of the liver]. / Sonographie von malignen Hämangioendotheliomen der Leber.

Malignant haemangioendotheliomas of the liver are very rare mesenchymal tumours. Sonographically, they appear as solitary, although more frequently as multicentric, space-occupying growths with blurred and irregularly delineated outlines; the sonographic image shows both echo-dense structures and structures with a low amount of echoes, or which are even echo-free. Short-term sonographic follow-up examinations point to a tendency to rapid growth and varying echogenicity.

[Diagnosis of choledochal cysts. The methodological value of imaging procedures]. / Diagnostik der Choledochuszysten. Methodische Wertigkeit bildgebender Verfahren.

For 10 cases in which we detected cysts in the choledochus ourselves comparing traditional radiological methods (infusion-cholegram, ERC, scintigraphy, barium meal examination, angiography) with recent imaging procedures (ultrasound, CT, NMR) the following sequence of procedures proved to be favorable: Screening methods are ultrasound and infusion-cholegram. CT and NMR furnish good presentations of the intra- and extrahepatic dilatations of the bile duct. ERC still represents the best methods for demonstration of an extrahepatic cyst of the choledochus. Hepato-biliary functional scintigraphy is performed as a supplement. Barium meal examination and coeliacography furnish a small diagnostic contribution only.

[Interdisciplinary diagnosis of choledochus cysts]. / Interdisziplinäre Diagnostik von Choledochuszysten.

Cystic dilatations of the biliary tract are rare anomalies. In the individual case, diagnosis may pose great problems. The relevance of the various methods was assessed in eight patients investigated with different diagnostic methods. In all patients the predominant symptom was uncharacteristic upper abdominal complaints. Two patients showed intermittent jaundice. Among non-invasive methods sonography is preeminent and permits precise demonstration of intra- and extrahepatic biliary duct dilatations. It can be complemented by CAT-scanning. Whereas intravenous cholegraphy does not permit sufficient ascertainment of the diagnosis, ERCP and PTC allow precise demonstration of the anomaly. Functional hepatobiliary scanning is indicated where endoscopic methods are not available. Among the eight patients sonography was decisively relevant in five cases and ERCP in all cases. The remaining functional assessments furnished important additional informations.