Toll-like receptor 4 gene polymorphism 1196 C/T does not influence the risk of neuropsychiatric systemic lupus erythematosus in Polish population--a preliminary report.

OBJECTIVE: Recent data indicate that Toll-like receptors (TLRs) participate in various neuropathologic conditions, including ictogenesis, myelin disruptions associated with chronic alcohol abuse, behavioral and cognitive dysfunctions associated with alcohol-induced neuroinflammatory damage, and activation of microglia to reduce amyloid ß deposits. As seizures and depression are well known neuropsychiatric syndromes in systemic lupus erythematosus (SLE) the aim of the study was to investigate whether TLR4 gene polymorphism 1196C/T (rs4986791, Thr399Ile) was a candidate for susceptibility of development of neuropsychiatric systemic lupus erythematosus (NPSLE). METHODS: The study covered 60 patients with SLE and 100 healthy individuals. TLR4 1196C/T genotyping was performed by real-time polymerase chain reaction with the SimpleProbe. RESULTS: The SLE group comprised 86.7% of patients with wild-type homozygotes CC and 13.3% heterozygotes CT and no homozygotes TT. The control group consisted of 85% wild-type homozygotes CC, 15% heterozygotes CT and no homozygotes TT. The frequencies of genotype and allele distribution in SLE patients did not differ significantly from those of the control subjects. The probability of describing the possible risk of SLE imputed to genotype did not significantly differ in comparison with the healthy individuals (p = 0.77, odds ratio = 0.87, 95% confidence interval 0.34-2.19). A significant genotype association of genotype CC with arthritis was found in SLE patients (p = 0.02). It was further confirmed by a significant association of a dominant allele C with arthritis (p = 0.02). No association between CC and CT genotypes of TLR4 1196C/T and NPSLE was found. Allele distribution of TLR4 1196C/T also was not associated with NPSLE. No other significant differences were found in genotype and allele frequencies regarding clinical manifestation of SLE patients. CONCLUSION: In the Polish population of SLE patients, 1196C/T polymorphism of TLR4 gene does not increase the risk of development of NPSLE; however, genotype CC and a dominant allele C is associated with arthritis in the course of SLE.

Bone turnover rate in postmenopausal women: bimodal distribution?

There is considerable evidence that elevated bone turnover is an independent form of low bone mineral density (BMD) risk factor of osteoporotic fractures. The aim of our study was to test whether a group of postmenopausal women could be divided into subgroups of high and low bone turnover rate using different pairs of bone turnover markers (one resorption, one formation). Cluster analysis was used to obtain high and low bone turnover subgroups within the study group. A magnitude of difference in lumbar spine BMD (expressed as Z-score) between high- and low-turnover groups was used as a criterion of division success. According to this criterion, the division obtained with a urinary type I collagen crosslinked N-telopeptide/bone alkaline phosphatase pair of markers appeared to be the most significant. This method of separation of two subgroups was highly concordant with the division based on the upper thresholds of the normal values for those markers found for the premenopausal women. It seems that the observed existence of high-and low-turnover subject clusters is not an incidental phenomenon, because the effects obtained for the whole study group were further confirmed by the consistent results of cluster analysis, performed separately for two randomly selected subgroups (A and B) from the study group. The results obtained appear to support the view that bone turnover rate in postmenopausal women is distributed in the bimodal fashion. This finding seems to justify further investigations of more elaborated models, enabling clinicians to individually classify their patients as low- or high-turnover cases with higher efficiency, as in the case of cutoff values for single markers.

[Genetic factors in osteoporosis]. / Czynniki genetyczne w osteoporozie.

Osteoporosis is a widespread disease affecting more than 1/4th of the female and 1/10th of the male population. It is characterised by a low bone mass, which in turn leads to osteoporotic fractures. Bone mass can be described as bone mineral density (BMD). BMD in human population is subject to quite significant interpersonal variability, for 75 to 80% of which, the genetic factors are responsible. To investigate the dependence of BMD on genetic factors, a possible links between the BMD and the natural polymorphism of so called "candidate genes" are checked. The first candidate gene to be investigated was the gene coding for the receptor of the active form of vitamin D. A statistical linkage between the naturally occurring polymorphism of that gene and the BMD was found by many research centres. It was found that certain polymorphic variants of that gene are linked to higher BMD's than the other ones. This trend existed in different human races and various age groups in many countries including Poland. A batch of negative results which appeared in some papers can be explained either by high calcium consumption in the given population, or the existence of another gene affecting bone metabolism and closely coupled to the vitamin D receptor gene. Other investigated and promising genes are the genes coding for other receptors (e.g. oestrogen), regulatory proteins (e.g. IL-6) and structural proteins (e.g. type I collagen).

Persistent hypercalciuria and elevated 25-hydroxyvitamin D3 in children with infantile hypercalcaemia.

The aim of the study was to characterize abnormalities of calcium-phosphate and vitamin D3 metabolism in children with a past history of "mild" Lightwood-type idiopathic infantile hypercalcaemia. Seventeen seemingly healthy children aged 2-12 years, with long-term idiopathic hypercalcaemic syndrome since infancy were studied. Two reference groups were also included (vitamin D3 intoxication/healthy and Williams groups). Despite a long-term milk-restricted diet and a restricted vitamin D3 intake, urinary calcium excretion in the study group was 0.117 +/- 0.07 mumol/kg per 24 h. Compared with the reference groups (0.047 +/- 0.029 and 0.067 +/- 0.06 mumol/kg per 24 h, P < 0.05), there was significant hypercalciuria in the children with idiopathic hypercalcaemia since infancy. Serum concentrations of 25-hydroxyvitamin D3 in the study group were also elevated compared with the reference groups (57.4 +/- 15.5 vs. 34.6 +/- 9.3 and 22.7 +/- 10.5 ng/ml). 1,25-Dihydroxyvitamin D3 levels were at the upper limit of normal (45.9 +/- 13.1 vs. 35.0 +/- 8.1 and 30.0 +/- 13.7 pg/ml). Non-progressive, clinically silent nephrocalcinosis was visible on ultrasound examinations. The disturbances of vitamin D3 and calcium-phosphate metabolism persistent in the normocalcaemic phase of idiopathic infantile hypercalcaemia may be a primary metabolic defect of the condition. The mechanisms leading to elevation of metabolites of 1,25-dihydroxy- and 25-hydroxyvitamin D3 and the relationship between this and persistent hypercalciuria and nephrocalcinosis need pathophysiological explanation.

Pyridinium crosslinks of collagen as a marker of bone resorption rates in children and adolescents: normal values and clinical application.

The aim of our study was to establish normal values of urinary pyridinoline (Pyr) and deoxypyridinoline (DPyr) excretion for children aged 3-18 years, examine the biological variability of the marker, and assess its clinical value for pediatric patients with growth hormone deficiency. Pyr and DPyr was measured in first void urine samples from 692 healthy subjects (340 boys, 352 girls) by high-performance liquid chromatography. At sampling, age, body height, and weight was recorded for all individuals. Short-term variability in crosslinks excretion was examined in four healthy children. The clinical value of the marker was studied in seven patients with growth hormone (GH) deficiency. In childhood, crosslinks excretion exceeded normal adult values by about fivefold and declined during puberty. In the age range of 13-18 years, gender-related differences in Pyr and DPyr levels were observed, presumably resulting from the earlier onset of puberty in girls. Urinary levels of Pyr and DPyr were highly correlated both in males and females. Pyr/DPyr ratio was significantly higher in adolescents than children, suggesting enhanced release of Pyr from extraosseous sources. In both genders, neither age nor anthropometric variables showed a linear effect on crosslinks excretion. The range of within-subject, short-term variability in urinary Pyr and DPyr was relatively high (CV: 6%-21%), indicating that single measurements of crosslinks excretion may not adequately reflect bone resorption rates in children. Pyr and DPyr levels were significantly lower in GH-deficient patients and normalized during human growth hormone (hGH) therapy. Significant correlations between growth velocity (GV) and crosslinks levels were found, but individual prediction of GV increment during hGH treatment may be inaccurate. Pyr/DPyr ratio was not related to GV. It is concluded that measurement of urinary Pyr and DPyr excretion in children may be a valuable tool to assess bone resorption rates in population-based studies. In individual patients, however, only qualitative evaluation of disease severity and response to treatment seems justified.

[Evaluation of bone density using the DEXA method and ultrasonography in women after menopause during treatment with vitamin D]. / Ocena gestosci masy kostnej metoda DEXA i ultradzwiekowa u kobiet po menopauzie w przebiegu leczenia witamina D.

A new group of drugs-oestrogen agonists-antagonists (tamoxifen and raloxifen) is discussed. In view of their potentially favourable effects on the bone and lipids and lacking unfavourable action on the uterus and mammary gland, after successful completion of clinical studies, these preparations may be possibly important for the prophylaxis and treatment of osteoporosis.

[Pyridinoline and deoxypyridinoline excretion in urine of healthy children--dependence on age]. / Wydalanie pirydynoliny i dezoksypirydynoliny w moczu dzieci zdrowych--zaleznosc od wieku.

Pyridinoline (Pyr) and deoxypyridinoline (DPyr) are crosslinking compounds of bone collagen. Their urinary excretion is considered to be the first sensitive and specific marker of bone resorption in a number of metabolic bone diseases in adults. Application of crosslinks measurements to evaluate bone turnover rate in pediatric patients is so far limited because of lack of reference values. Therefore, the aim of our study was to determine urinary excretion of Pyr and DPyr in healthy children aged 3-18 yrs, and to evaluate the possible relationship between the levels of both compounds and body height, weight, BMC, and BMD. Pyr and DPyr levels were determined in first void urine samples obtained from 249 children (124 boys, 125 girls). Urine aliquots were hydrolysed, Pyr and DPyr extracted on CF1 cellulose, and analysed by HPLC with fluorimetric detection. Bone mineral content (BMC) and density (BMD) were measured with Lunar DPX-L apparatus in 205 children (104 boys, 101 girls) from the same population, aged over 5.5 yrs. In prepubertal children, a tendency towards lowering of urinary Pyr and DPyr levels with advancing age was shown. At puberty, urinary excretion of both crosslinks markedly decreased. This phenomenon was observed at various calendar age in girls as compared to boys, reflecting sex-dependent differences. Significant negative correlation (p < 0.0001) between urinary Pyr and DPyr levels and calendar age, body height and weight, BMC and BMD, were also found. The obtained results suggest that references values for Pyr and DPyr excretion in growing children should be related to calendar age, sex, and--in case of adolescents--phase of puberty.

[Changes in vitamin D metabolism in children following partial intestinal resection]. / Veränderungen des Vitamin D-Stoffwechsels bei Kindern nach partieller Darmresektion.

Six children (five after partial intestinal resection, one after colectomy) were studied with respect to absorption and metabolism of vitamin D. The results showed decreased 25-OH-D3 serum concentrations below 20 ng/ml (in two patients even below 10 ng/ml), possibly due to disturbed enterohepatic circulation of vitamin D metabolites.

Simplified HPLC method for quantitation of vitamin D in blood serum.

A single-step high performance liquid chromatography (HPLC) method for vitamin D quantitation in blood serum is described. Methanol/hexane extraction, silica Sep-Pak purification, differential dissolution, filtration, and the use of a precolumn in place of an injector sample loop allow for using only one (reversed phase) HPLC step instead of the usual two. Detection limit with the HPLC system used was about 1 micrograms/L of serum. Recovery of vitamin D3 added to serum ranged from 90% to 109%. Intra and interassay coefficient of variation values were 8.1% and 15.4%, respectively.

Endogenous, cutaneous vitamin D synthesis stimulation as an effective way of improving the vitamin D status in children with hepatobiliary malfunctions.

Repetitive (5 days) ultraviolet treatment was used to improve vitamin D status of 26 pediatric patients suffering from different hepatobiliary disorders. The treatment resulted in a significant improvement of patients' vitamin D and 25-hydroxyvitamin D levels regardless of their disease and other factors such as increased blood bilirubin content or jaundice. Presence of a dependence between the increase in 25-hydroxyvitamin D concentrations after treatment and its original levels was confirmed.

Application of the rat thymus receptor in a specific, sensitive and simplified assay of 1,25-dihydroxyvitamin D in blood serum.

A simple method has been employed to prepare crude nuclear extract from rat thymus, using hypertonic buffer after previous treatment with hypotonic buffer. The preparation is free from serum vitamin D-binding protein and contains a 3.7 S receptor molecule, which specifically binds 1,25-dihydroxyvitamin D-3 (1,25-(OH)2D3). The receptor is of high affinity (KD = 0.85 X 10(-11) M at O degrees C) and low capacity (260-460 fmol/g tissue). The Scatchard analysis of ligand binding results in a concave downward curve. The Hill analysis of the same data gives good linear fitting (r = +0.971) with the Hill coefficient nH = 1.63. These facts indicate positive cooperativity between two ligand binding sites of the rat thymus 1,25-(OH)2D3 receptor. The preparation was used in a competitive protein binding assay of 1,25-(OH)2D in serum extracts, purified on Sep-Pak C18 followed by silica Sep-Pak cartridges. The method was sensitive to 0.5 pg/tube (2.0 ng/l) when 1 ml of serum was extracted. Intra- and interassay coefficients of variation were 9% and 14%, respectively. The serum 1,25-(OH)2D concentration estimated in 33 children (mean age 6.5 +/- 3 years) was 46.6 +/- 18.4 ng/l (mean +/- S.D.).

Rapid method for measuring physiological concentrations of 25-hydroxyvitamin D levels in blood serum.

A new single-step Extrelut column procedure for obtaining the 25-hydroxyvitamin D extract, suitable for a direct radiocompetitive test, is described. Separation of 0.1 ml of serum, on a small Extrelut column and radioassay allows a rapid, sensitive and accurate determination of 25-hydroxyvitamin D concentration. The detection limit of the determination was 2 nmol/l (0.8 microgram/l) of serum. The recovery of 250HD3 added to serum was 99-108%. Intra-assay and interassay CV values were 10 and 14%, respectively. Although the method does not permit separation of dihydroxylated metabolites of vitamin D from 25-hydroxyvitamin D, it is useful in clinical practice.

Insolation effect on vitamin D supply in children during sanatorium treatment.

Serum 25/OH/D3 levels were determined in 58 children aged 5-7 years staying from April 1 to September 30 1981 in Kolobrzeg in a sanatorium. A comparison of isolation conditions in that time period with the serum values of 25/OH/D3 showed that the supply of vitamin D was sufficient only in the period from June to September. In the remaining months no correlation was observed between the values of solar radiation and vitamin D levels. During the study and for three months before it exogenous vitamin D administration was stopped.