Introduction: Bone health in sarcoidosis can be altered due to inflammation, bone involvement or treatment. Aim: To investigate change in BMD over time in Danish patients with sarcoidosis. Methods: In 117 patients with sarcoidosis, BMD was measured twice with 12 months interval. T-scores and change in scores were compared between glucocorticoid treated and non-treated patients. Results: Mean age of the patients was 47 years, 57% were men, 35% were treated with prednisolone and 16% with bisphosphonate. BMD increased over time, Table 1. No difference in mean change of T-score was observed between glucocorticoid treatment groups. Hip: -0.02 (-0.20; 0.17, p= 0.8642) and columna (col): -0.03 (-0.22; 0.16, p= 0.7619). For columna the increase was associated with male gender: 0.28 (0.01; 0.49, p= 0.0022) and bisphosphonate treatment: 0.25 (0.01; 0.49, p=0.0418). T - and Z-scores as means with 95% CI (). Glu: Glucocorticoid treatment. Conclusion: BMD increased over 12 months without association with glucocorticoid treatment. For columna the increase was associated with male gender and bisphosphonate treatment.

Objective: To examine longitudinal changes in serum cobalamins, transcobalamin (TC) and haptocorrin (HC) during lactation and to investigate the influence of vitamin B 12 supplementation on these parameters. Design: A 9-month follow-up study. Subjects and methods: Lactating mothers ( N =89) including 23 supplemented with vitamin B 12 (1–18 μ g/daily), 41 partly supplemented and 25 not supplemented. Blood samples collected 3 weeks (baseline) and 4 and 9 months post-partum were analysed for cobalamins, TC and HC. Both the total concentration and the cobalamin-saturated form (holo) of TC and HC were analysed. Results: No significant differences were observed in serum cobalamins or its binding proteins related to supplementation with vitamin B 12 or the duration of lactation. Serum cobalamins remained unchanged from 3 weeks to 9 months post-partum. Total TC (holoTC) (median±s.e. pmol/l) decreased between 3 weeks (710±23 (85±12)) and 9 months (602±21 (76±11)) ( P <0.0001 ( P =0.0002)), whereas total HC (holoHC) increased from (422±11 (300±9)) at 4 months to (455±13 (317±10)) to 9 months post-partum ( P <0.0001 ( P <0.0001)). Conclusion: We report a decrease in TC and an increase in HC during a 9-month period post-partum. No differences were observed between the vitamin B 12 -supplemented and the unsupplemented groups. Thus, supplementation with vitamin B 12 has no impact on the circulating level of serum cobalamins or its binding proteins in a Danish population of lactating mothers.

BACKGROUND Recommended daily intakes of vitamin B-12 vary between 2 and 6 microg. OBJECTIVE The objective was to examine the associations between vitamin B-12 intake and markers of vitamin B-12 deficiency. DESIGN We studied 98 Danish postmenopausal women aged 41-75 y. Serum cobalamin, transcobalamin (TC) saturated with vitamin B-12 (holo-TC), TC saturation (holo-TC/total TC), methylmalonic acid (MMA), and total homocysteine (tHcy) were measured to assess vitamin B-12 status. Dietary intakes of vitamin B-12 were determined from 7-d weighed food records. Gastric pH measurement and the alkali-challenge test were performed with the use of Heidelberg radiotelemetric capsules. RESULTS The total intake of vitamin B-12 ranged from 1.2 to 23.9 microg/d. All vitamin B-12-related variables, except gastric pH, correlated significantly with total vitamin B-12 intake. Those taking supplements (54%) had higher circulating concentrations of cobalamin and TC saturation and lower concentrations of MMA and tHcy than did those not taking supplements. All subjects were divided into quintiles according to their total vitamin B-12 intake. For all the variables analyzed, the curves appeared to level off at a daily vitamin B-12 intake of approximately 6 microg; the median (and 25th-75th percentiles) for cobalamin was 380 (270-480) pmol/L, for holo-TC was 119 (92-162) pmol/L, for MMA was 0.12 (0.14-0.17) micromol/L, and for tHcy was 9.75 (8.3-11.40) micromol/L (n = 58). CONCLUSION A daily vitamin B-12 intake of 6 microg appeared to be sufficient to correct all the vitamin B-12-related variables measured in the postmenopausal Danish women in this study.

To investigate the relation between lactation and markers of folate and vitamin B12 (B12) deficiency in women with and without vitamin supplementation. A 9-month follow-up study. Blood samples from 91 women, who gave birth to a single healthy child, were collected 3 weeks, 4 and 9 months postpartum and analysed for circulating level of homocysteine (tHcy), methylmalonic acid (MMA), folate and B12. The participants were categorized as exclusively, partly or not breast-feeding dependent on the degree of lactation 4 months postpartum. During follow-up, lifestyle factors were recorded by structured interviews. Among 72 exclusively breast-feeding women, the median (10–90% percentile) tHcy was 5.8 (3.1–8.3) μmol/l 3 weeks postpartum, 6.1 (4.1–10.3) μmol/l 4 months postpartum and 5.3 (3.6–8.7) μmol/l 9 months postpartum. At 9 months postpartum, none of the women breast-fed exclusively. No significant change occurred in the concentration of B12 and folate. Exclusively breast-feeding women without vitamin supplementation had higher median tHcy than supplemented exclusively breast-feeding women 4 and 9 months postpartum (7.0 vs 5.4 μmol/l (P<0.001) and 5.8 vs 4.5 μmol/l (P=0.003), respectively). Six women had increased (>15 μmol/l) tHcy; four of these were unsupplemented and exclusively breast-feeding. We found no overall indication of depletion of the folate and B12 stores during the lactation period in this population. However, folate-supplemented women had lower tHcy and higher folate levels, suggesting a beneficial effect of supplementation with folate throughout lactation. The Biomedical Laboratory Scientist Education and Research Fund and LEO Pharma Research Foundation supported this study. DPC Scandinavia, Denmark donated reagents for the folate and tHcy analysis.

Background-A causal relation between hyperhomocysteinemia (HHcy) and accelerated atherosclerosis has been established in apolipoprotein E-deficient (apoE(-/-)) mice. Although several cellular stress mechanisms have been proposed to explain the atherogenic effects of HHcy, including oxidative stress, endoplasmic reticulum (ER) stress, and inflammation, their association with atherogenesis has not been completely elucidated.Methods and Results-ApoE(-/-) mice were fed a control or a high-methionine (HM) diet for 4 ( early lesion group) or 18 ( advanced lesion group) weeks to induce HHcy. Total plasma homocysteine levels and atherosclerotic lesion size were significantly increased in early and advanced lesion groups fed the HM diet compared with control groups. Markers of ER stress (GRP78/94, phospho-PERK), oxidative stress (HSP70), and inflammation (phospho-IkappaB-alpha) were assessed by immunohistochemical staining of these atherosclerotic lesions. GRP78/94, HSP70, and phospho-IkappaB-alpha immunostaining were significantly increased in the advanced lesion group fed the HM diet compared with the control group. HSP47, an ER-resident molecular chaperone involved in collagen folding and secretion, was also increased in advanced lesions of mice fed the HM diet. GRP78/94 and HSP47 were predominantly localized to the smooth muscle cell-rich fibrous cap, whereas HSP70 and phospho-IkappaB-alpha were observed in the lipid-rich necrotic core. Increased HSP70 and phospho-IkappaB-alpha immunostaining in advanced lesions of mice fed the HM diet are consistent with enhanced carotid artery dihydroethidium staining. Interestingly, GRP78/94 and phospho-PERK were markedly increased in macrophage foam cells from early lesions of mice fed the control or the HM diet.Conclusions-Multiple cellular stress pathways, including ER stress, are associated with atherosclerotic lesion development in apoE(-/-) mice.

It has been demonstrated that hyperhomocysteinemia (HHcy) accelerates atherosclerosis in apolipoprotein E-deficient (apoE−/−) mice. In this study, vitamin-defined chow diets were used to induce HHcy in apoE−/− mice in an attempt to identify possible pathogenic pathways. Six-week-old female apoE−/− mice were divided into seven groups: vitamin-defined purified chow diet alone (control), or same diet supplemented with either d,l-homocysteine (↑Hcy) or l-homocystine (↑Hcy–Hcy), or diet high in l-methionine (↑Met), or diet high in B-vitamins (↑vitamin), or diets deficient in folate (↓folate) or vitamin B6 (↓B6). Eighteen weeks later, plasma total homocysteine (tHcy), lipids and atherosclerotic plaque burden (aortic root, aortic arch, and brachiocephalic trunk) were measured. tHcy levels were similar in the ↑vitamin, ↓folate, ↓B6 and control groups (9.2–10.1 μmol/l, NS), but elevated mildly in the ↑Hcy–Hcy group (16.1 μmol/l) and moderately in the ↑Met and ↑Hcy groups (53.6 and 51.5 μmol/l, respectively). Mice in the latter two groups had significantly more atherosclerosis in the aortic root. Although B vitamin-supplementation failed to lower tHcy levels, mice had less atherosclerosis in the aortic arch. In summary, dietary methionine and homocysteine, but not homocystine, enhanced the development of atherosclerosis. Supplementation with B vitamins appeared to confer homocysteine-independent protection against atherosclerosis. These results suggest that (1) there may be a threshold level below which homocysteine is not atherogenic; (2) the atherogenic effect of HHcy may be mediated via an intracellular pathway; and/or (3) the anti-atherogenic effect of B vitamins in normohomocysteinemic mice is independent of tHcy levels.

That physical exercise stimulates pituitary GH secretion has been known for forty years, but the underlying mechanisms as well as the physiological significance remain elusive. We have previously shown that the concomitant increase in core temperature is essential for the exercise-induced GH release, inasmuch as exercise performed at 4 C results in a suppression of GH secretion, whereas passive heating constitutes a potent stimulus for GH release. Moreover, studies in normal subjects show that GH stimulates sweat production and evaporative heat loss during heat exposure with and without exercise, whereas GH-deficiency is associated with reduced sweat secretion and increased heat storage during similar conditions. The neurotransmitters involved in GH secretion during exercise remain uncertain; we therefore investigated the putative role of ghrelin, which is a gut-derived endogenous ligand for the GHS receptor. We measured circulating ghrelin levels before during and after submaximal aerobic exercise in healthy subjects and GH-deficient patients. The circulating ghrelin levels were unchanged during and after exercise in all subjects. Growth hormone stimulates lipolysis and lipid oxidation during basal and fasting conditions and we recently investigated whether GH also regulates substrate metabolism during exercise. The design involved GH-deficient patients studied during exercise with and without GH administration as compared to untreated healthy subjects. Growth hormone predominantly stimulated the turnover of free fatty acids in the recovery phase after exercise. Conclusions: 1) the increase in GH release during exercise is associated with the concomitant increase in body temperature, 2) GH stimulates sweat secretion and heat evaporation during exercise, which seems to be of distinct physiological significance, 3) ghrelin is not involved in exercise-induced GH release, 4) the impact of GH on substrate metabolism during exercise includes increased FFA turnover.

In humans, maternal hypercholesterolemia during pregnancy promotes microscopical fatty streaks in the children. The mechanism is unknown. Fatty streaks are clinically silent, and many of them regress and never develop into advanced atherosclerosis. The aim of this study was to investigate whether hypercholesterolemia in pregnant mice induced more advanced atherosclerosis in their adult progeny. Hypercholesterolemic (HC) apolipoprotein E knockout (apoE(-/-)) female mice were mated with normocholesterolemic (NC) wild-type (apoE(+/+)) males and vice versa. All parents were almost identical genetically except for apoE. Therefore, all progeny became genetically identical and heterozygous apoE(+/-). They were born of either HC (i.e. apoE(-/-)) or NC (i.e. apoE(+/+)) mothers. The progeny were killed 6 months after birth and the amount of atherosclerosis in the aortic root was assessed. Females developed more atherosclerosis than males (P<0.001) but, regardless of sex, maternal hypercholesterolemia during pregnancy had no influence on the amount of atherosclerosis in adult progeny. Males of HC mothers had lower plasma cholesterol levels than males of NC mothers. Thus, in mice, maternal hypercholesterolemia during pregnancy does not promote the development of advanced atherosclerosis in their adult progeny.

Elevated plasma levels of total homocysteine (hcy) have been associated with an increased occurrence of arterial thrombosis. In the present study, we investigated the influence of hyperhomocysteinaemia on platelet aggregation and arterial thrombus formation in vivo. Fifty-one rats were included in the study, of which 29 received hcy in the drinking water for 4 weeks. Blood samples were withdrawn for measurement of platelet count and mean platelet volume. Platelet aggregation response in platelet-rich plasma following adenosine diphosphate or collagen stimulation were examined. In vivo thrombus formation was investigated by transillumination and videotape recording of the rat femoral artery after a thrombogenic injury was established. Off-line videotape analysis using computer-assisted planimetry permitted quantification of the thrombus area, and area versus time curves were obtained. In the intervention group receiving hcy, total hcy in plasma increased two-fold to 14.3 μmol/l, as compared with 7.3 μmol/l in the control group (P < 0.001). The platelet count and mean platelet volume did not differ between the two groups. In vivo thrombus formation expressed as the area under the curve or maximum thrombus area was not found to be altered in the presence of an increased homocysteine level, neither was adenosine diphosphate-induced platelet aggregation. However, collagen-induced platelet aggregation significantly decreased in the hcy group (P = 0.02). Pro-thrombotic effects of isolated mild hyperhomocysteinaemia are not supported by the present study in rats.

Hyperhomocysteinemia is an independent risk factor for atherothrombosis. However, causality is unproven, and it remains unknown whether hyperhomocysteinemia promotes atherosclerosis, plaque rupture, and/or thrombosis. We evaluated the short- and long-term effects of hyperhomocysteinemia on plaque size and structure in 99 atherosclerosis-prone apolipoprotein E-deficient mice. Hyperhomocysteinemia was induced by methionine (Met) or homocysteine (HcyH) supplementation: low Met (+11 g Met/kg food), high Met (+33 g Met/kg food), low HcyH (0.9 g HcyH/L drinking water), and high HcyH (1.8 g HcyH/L drinking water). Met and HcyH supplementation significantly raised plasma total homocysteine levels by 4- to 16-fold above those observed in mice fed a control diet (up to 146.1 μmol/L). Compared with controls, aortic root plaque size was significantly larger in supplemented groups after 3 months (56% and 173% larger in high-Met and high-HcyH, respectively) but not after 12 months. Hyperhomocysteinemia was associated with an increase in the amount of collagen in plaques after both 3 and 12 months. Mechanical testing of the tail tendons revealed no weakening of collagen after 12 months of hyperhomocysteinemia. Many plaques in both control and supplemented mice appeared rupture prone morphologically, but all aortic root plaques and all but 1 coronary plaque had an intact surface without rupture or thrombosis. Thus, diet-induced hyperhomocysteinemia promotes early atherosclerosis and plaque fibrosis but does not, even in the long term, weaken collagen or induce plaque rupture.

BACKGROUND:Red wine polyphenols and ethanol reduce fatty streak formation (early atherosclerosis) in various animal models. These experimental results support the observation that alcoholic beverages protect against myocardial infarction in humans. However, fatty streaks may not reflect the pathology of mature and clinically relevant atherosclerosis. The present study examined the effects of red wine polyphenols and ethanol on mature atherosclerosis in apolipoprotein E-deficient mice.METHODS AND RESULTS:Eighty-four 7-week-old mice were randomized to receive water, red wine (diluted to 6% ethanol v/v), 6% ethanol v/v, or red wine powder in water. All mice were fed a normal chow diet. At 26 weeks of age, the mice were killed. HDL cholesterol was raised 12.0% (95% CI, 4.0% to 20.0%) and 9.2% (95% CI, 1.5% to 16.9%) by red wine and ethanol, respectively. At the end of study, all mice exhibited advanced atherosclerosis in the aortic bulb, whereas less mature atherosclerosis predominated in the brachiocephalic trunk. The amount of atherosclerosis in the aortic bulb and the brachiocephalic trunk were similar in all groups (P:=0.92 and P:=0.14, respectively). To evaluate whether ethanol or red wine polyphenols were protective by stabilizing atherosclerotic plaques rather than reducing their size, we measured the percentage of collagen-poor areas in left coronary sinus plaques as a morphological criterion of plaque stability. The percentage of collagen-poor areas did not differ between groups (P:=0.71).CONCLUSIONS:Neither ethanol nor red wine polyphenols reduced mature atherosclerosis or changed the content of collagen in plaques in apolipoprotein E-deficient mice.

To investigate the mechanisms behind the water- and sodium-retaining effects of growth hormone (GH), we studied the effect of GH on 1) water and sodium homeostasis, 2) the renin-angiotensin-aldosterone system (RAAS), and 3) lithium clearance (C(Li)) with and without concomitant prostaglandin (PG) synthesis inhibition with ibuprofen. GH administration for 6 days induced a significant increase in plasma renin, which was abolished by coadministration of ibuprofen (mU x l(-1) x 24 h(-1): control: 22.4 +/- 4.3; GH: 37.7 +/- 8.8; ibuprofen: 15.2 +/- 3.0; GH + ibuprofen: 19.7 +/- 2.5; ANOVA: P < 0.01). Comparable increments in extracellular volume were seen after 6-day treatment with GH alone and in combination with ibuprofen [liters: control, 19.57 +/- 0.92; GH, 20.80 +/- 1.00 (ANOVA: P < 0.0005); ibuprofen, 19.38 +/- 0.90; GH + ibuprofen, 21.63 +/- 1.37 (ANOVA: P < 0.0005)]. Treatment with GH increased C(Li) and changed the tubular handling of sodium and water. The absolute distal sodium reabsorption was increased, and this was only partially counterbalanced by decreased reabsorption in the proximal tubules. The data demonstrate that GH-induced activation of the RAAS can be blocked by concomitant PG synthesis inhibition and that the tubular effects of GH include increased distal nephron sodium and water reabsorption.

Sinusitis is commonly occurring in patients infected with Human Immunodeficiency Virus I (HIV), but the occurrence and etiology have not been established. The purpose of this study was prospectively to determine the occurrence, site and type of paranasal sinus abnormalities seen on MRI in HIV-infected patients with fever, to relate the abnormalities to clinical and immunological parameters, and to determine the microbiological agents found in the sinus aspirates. MRI was performed in 54 HIV-infected patients with 70 evaluable episodes of fever. Patients receiving antibiotics were excluded. Bactrim was permitted, when given as prophylaxis of Pneumocystis carinii pneumonia. If abnormalities were found on MRI, sinus aspiration was performed and the aspirate investigated. MRI abnormalities were found in 54.3% of the patients with a significantly higher occurrence of pathological changes in AIDS patients compared with HIV-infected without AIDS. In approximately 2/3 of the aspirates a probable, etiologic agent was found. However, 1/3 of these agents were atypical such as cytomegalovirus and mycobacteriae; in one patient Non-Hodgkin's lymphoma was found. The high occurrence of sinusitis in HIV-infected patients and the atypical findings in the sinus aspirates stress the importance of searching for sinusitis and the etiology to ensure the correct treatment.

The widespread use of oral glucose in the treatment of hypoglycemia is mainly empirically based, and little is known about the time lag and subsequent magnitude of effects following its administration. To define the systemic impact and time course of effects following oral glucose during hypoglycemia, we investigated 7 healthy young men twice. On both occasions, a 6-hour hyperinsulinemic (1.5 mU/kg/min)-hypoglycemic clamp was performed to ensure similar plasma glucose profiles during a stepwise decrease toward a nadir less than 50 mg/100 mL after 3 hours. On the first occasion, subjects ingested 40 g glucose and 4 g 3-ortho-methylglucose ([3-OMG] to trace glucose absorption) dissolved in 400 mL tap water after 3.5 hours. The second examination was identical except for the omission of 40 g oral glucose, and glucose levels were clamped at hypoglycemic concentrations similar to those recorded on the first examination. Plasma glucose curves were superimposable, and all participants reached a nadir less than 50 mg/100 mL. Similar increases in growth hormone (GH) and glucagon were observed in both situations. The glucose infusion rates (GIRs) were lower after oral glucose, with the difference starting after 5 to 10 minutes, being statistically significant after 20 minutes, and reaching a maximum of 8.5 +/- 1.6 mg/kg/min after 40 minutes. Circulating 3-OMG increased after 20 minutes. In both situations, infusion of insulin resulted in insulin levels of approximately 150 microU/mL and a suppression of C-peptide levels from 2.0 to 1.1 nmol/L (P < .01). After glucose ingestion, both serum C-peptide and glucagon-like peptide-1 (GLP-1) increased (C-peptide from 1.1 +/- 0.05 to 1.4 +/- 0.05 nmol/L and GLP-1 from 3.2 +/- 0.8 to 18.1 +/- 3.3 pmol/L), in contrast to the situation without oral glucose (P < .05). Isotopically determined glucose turnover was similar. In conclusion, our data suggest that oral glucose affects systemic glucose metabolism rapidly after 5 to 10 minutes. Quantitatively, the immediate impact is relatively small, with the gross impact observed after approximately 40 minutes. Future studies aiming to identify therapeutic oral agents with prompt effect seem warranted.

OBJECTIVE It is generally accepted that there is no clinically significant circadian variation in total insulin-like growth factor (IGF)-I or total IGF-II in healthy subjects. In contrast there is a significant nocturnal decrease in free IGF-I in healthy subjects, corresponding to the nocturnal increase in IGF binding protein-1. In this study we have investigated the circadian variation in circulating free IGF-I and IGF-II in patients with acromegaly and patients with adult onset growth hormone deficiency.PATIENTS Seven acromegalic patients were studied with and without treatment with a slow-release formulation of octreotide. Seven GH-deficient patients were studied without GH replacement. In addition 5 of the GH-deficient patients were studied during GH replacement.DESIGN Serum samples were obtained every hour for 24 h. Free IGF-I and IGF-II were measured every 2nd hour. Total IGF-I and IGF-II were measured every 2nd hour (acromegalic patients) or every 4th hour (GH deficient patients). IGF binding protein (IGFBP)-1 was measured every 2nd hour (acromegalic patients) or every hour (GH deficient patients).RESULTS In the untreated acromegalic patients there was a significant nocturnal decrease in free IGF-I, but not free IGF-II, before treatment. During treatment there was a significant nocturnal decrease in both free IGF-I and free IGF-II. Peak values of free IGF-I were 112% and 75% above trough (treatment and withdrawal, respectively). In the GH-deficient patients there were no significant circadian variations in free IGF-I or free IGF-II in either of the two occasions. In contrast, there was a significant circadian variation of total IGF-I after adjustment for changes in plasma volume in both treated and untreated acromegaly and GH deficiency in all cases with a peak between 0300 h and 0400 h. The nocturnal increase in total IGF-I ranged from 20% to 35%.CONCLUSIONS A significant circadian variation in free IGF-I and IGF-II was demonstrated in acromegalic patients. In contrast no significant circadian variation in free IGF-I and IGF-II was found in GH-deficient patients. Part of the variations may be due to poorly understood variations in IGF-I release. It is not clear whether and to what extent the observed circadian changes in free and total IGF-I are involved in circadian changes in IGF-I bioactivity.

BACKGROUND There is a need for systematic evaluation of methods before their release to the market. We addressed this problem in novel homocysteine assays as part of an European Demonstration Project involving six centers in four countries. METHODS Two immunological methods for measurement of plasma total homocysteine (P-tHcy), the fluorescence polarization immunoassay (FPIA) and the enzyme immunoassay (EIA), were compared with two comparison methods, HPLC and gas chromatography-mass spectrometry (GC-MS). All laboratories performed the following procedures: (a) familiarization; (b) determination of linearity and precision by analyzing five plasma samples with interrelated concentrations for 20 days; (c) correlation using patients' samples; and (d) assessment of long-term performance. RESULTS Both immunological methods were linear for P-tHcy between 5 and 45 micromol/L. The intralaboratory imprecision (CV) was <5% for FPIA and <9% for EIA used with a sample processor. The bias was -2% to 3% for FPIA and 2-4% for EIA used with a sample processor. CONCLUSIONS The immunological methods provide results with little bias compared with HPLC and GC-MS. The imprecision of the assays must be considered in the context of their intended use(s).