1α,25-dihydroxyvitamin D3 promotes osteogenic activity and downregulates proinflammatory cytokine expression in human periodontal ligament cells.

BACKGROUND AND OBJECTIVE: The aim of this study was to assess the impact of 1α,25-dihydroxyvitamin D3 (vitamin D3) on osteogenic and inflammatory properties of human periodontal ligament (PDL) cells and investigate underlying mechanisms. MATERIAL AND METHODS: Human PDL cells, obtained from four subjects, were stimulated with vitamin D3 for 4-48 h. The bone markers osteopontin and osteocalcin and proinflammatory cytokine/chemokine expression was determined by quantitative real-time polymerase chain reaction and enzyme-linked immunosorbent assay. Cytokine and chemokine expression was determined after stimulation with the inflammation promoter lipopolysaccharide (LPS) in the presence or absence of vitamin D3. Alkaline phosphatase activity was assessed using p-nitrophenylphosphate substrate. RESULTS: Treatment with 30 ng/mL of vitamin D3, corresponding to an optimal plasma concentration of vitamin D, for 24 h had no effect on PDL cell number and morphology but increased PDL cell osteopontin and osteocalcin mRNA expression by about 70 and 40%, respectively, and, moreover, treatment with vitamin D3 for 48 h enhanced PDL cell alkaline phosphatase activity by about two times showing that vitamin D3 exerts pro-osteogenic effects in human PDL cells. Stimulation with LPS (1 µg/mL) for 4 h increased PDL cell interleukin (IL)-6 cytokine and chemokine ligand 1 (CXCL1) chemokine mRNA expression several fold. The LPS-induced increase in IL-6 and CXCL1 transcripts was attenuated by vitamin D3 (30 ng/mL). Treatment with vitamin D3 (3-300 ng/mL) for 24 h reduced the LPS-evoked increase in PDL cell IL-6 protein by about 50%. Vitamin D3 (30 ng/mL) had no effect on LPS-induced IL-1ß and MCP-1 mRNA expression. CONCLUSIONS: Vitamin D3 promotes osteogenic differentiation but also downregulates inflammation promoter-induced IL-6 cytokine and CXCL1 chemokine expression in human PDL cells, suggesting that vitamin D3 both stimulates bone regeneration and antagonizes inflammation in human periodontal tissue.

Human endogenous peptide p33 inhibits detrimental effects of LL-37 on osteoblast viability.

BACKGROUND AND OBJECTIVE: High levels of the antimicrobial peptide, LL-37, are detected in gingival crevicular fluid from patients with chronic periodontitis. LL-37 not only shows antimicrobial activity but also affects host-cell viability. The objective of the present study was to identify endogenous mechanisms that antagonize the detrimental effects of LL-37 on osteoblast viability, focusing on the human peptide p33 expressed on the surface of various cell types. MATERIAL AND METHODS: Human osteoblast-like MG63 cells and human hFOB1.19 osteoblasts were treated with or without LL-37 in the presence or absence of p33. Recombinant human p33 was expressed in an Escherichia coli expression system. Lactate dehydrogenase (LDH) was assessed using an enzymatic spectrophotometric assay. DNA synthesis was determined by measuring [(3) H]-thymidine incorporation. Cell number was assessed by counting cells in a Bürker chamber. Intracellular Ca(2+) was monitored by recording Fluo 4-AM fluorescence using a laser scanning confocal microscope. Cellular expression of p33 was determined by western blotting. RESULTS: LL-37 caused a concentration-dependent release of LDH from human osteoblasts, showing a half-maximal response value (EC50 ) of 4 µm and a rapid and sustained rise in the intracellular Ca(2+) concentration of osteoblasts, suggesting that LL-37 forms pores in the cell membrane. p33 (10 µm) inhibited the LL-37-induced LDH release and LL-37-evoked rise in intracellular Ca(2+) concentration, suggesting that p33 prevents LL-37-induced permeabilization of the cell membrane. Moreover, p33 blocked LL-37-induced attenuation of osteoblast numbers. Also, mucin antagonized, at concentrations representative for nonstimulated whole saliva, LL-37-evoked LDH release, whilst cationic endogenous polyamines had no impact on LL-37-induced LDH release from osteoblasts. CONCLUSIONS: The endogenous peptide p33 prevents LL-37-induced reduction of human osteoblast viability. Importantly, this mechanism may protect the osteoblasts from LL-37-induced cell damage in patients suffering from chronic periodontitis associated with high levels of LL-37 locally.

Group B streptococcus in prosthetic hip and knee joint-associated infections.

The incidence of invasive group B streptococcus (GBS) infections in non-pregnant adults is increasing. Little is known about GBS in periprosthetic joint infections (PJIs). We aimed to analyse the clinical presentation of GBS PJI and its treatment in association with the outcome. The characteristics of 36 GBS PJIs collected from 10 centres were investigated. In 34 episodes, follow-up examination of ≥ 2 years was available, allowing treatment and outcome analysis. Most infections (75%) occurred ≥ 3 months after implantation. Most patients (91%) had at least one comorbidity; 69% presented with acute symptoms and 83% with damaged periprosthetic soft tissue. In 20 of 34 episodes debridement and retention of implant was attempted, but in five of these the prosthesis was ultimately removed. Hence, in 19 (56%) episodes, the implant was removed, including 14 immediate removals. In four episodes the removal was permanent. Penicillin derivatives and clindamycin were the most common antimicrobials administered (68%). In 94% the infection was cured, and in 82% functional mobility preserved. Debridement with implant retention was successful if the duration of symptoms was short, the prosthesis stable, and the tissue damage minor (10/10 vs 3/10 episodes, P = 0.003). Surgery that complied with a published algorithm was associated with a favourable outcome (P = 0.049).

Dextran sulfate sodium-induced colitis generates a transient thymic involution--impact on thymocyte subsets.

One of the most widely used animal models for inflammatory bowel disease (IBD) is the dextran sulfate sodium (DSS)-induced colitis. We have previously reported that 5 days administration of DSS in C57Bl/6J mice induces a colonic inflammation that progresses into chronicity after DSS removal, whereas in BALB/cJ mice the inflammation resolves within 4 weeks post-DSS. Here we show that both thymic size and thymocyte numbers dramatically decreased in the acute phase of inflammation in C57Bl/6 mice, 7 days after DSS withdrawal. Mature, CD4(+) and CD8(+) single positive (SP) CD69(lo) CD62L(hi) thymocytes were enriched in these mice, accompanied by a major decrease in the number of immature double positive (DP) thymocytes. However, the different maturation stages within the DP thymocyte subset were unchanged between healthy and inflamed C57Bl/6J mice. Interestingly, as the inflammation progressed into the chronic phase, the thymus recovered and 2 weeks after the acute inflammatory phase all the thymic parameters investigated in this study were restored to normal. In contrast, BALB/cJ mice only develop mild thymic alterations. Nevertheless, we found that within the double negative (DN) thymocytes an increased frequency and also total numbers of CD44(+) CD25(-) (DN1) cells correlated with the severity of colitis, and that the frequency of CD44(-) CD25(-) (DN4) thymocytes decreased proportionally in the acute phase in BALB/cJ mice. Our observations suggest that the thymic effects are intimately connected to the intestinal inflammatory response in colitis regardless of the inflammatory stimuli.

Periods of cluster headache induced by nitrate therapy and spontaneous remission of angina pectoris during active clusters.

Glyceryl trinitrate (GTN) is known to induce single extra attacks of cluster headache (CH) during active cluster periods, most probably via actions of nitric oxide (NO). Induction of whole periods of CH by organic nitrates has, however, attracted little attention in the literature. We report on eight patients with episodic CH and coexistent effort-induced angina pectoris. Cases 1-6 had been free of their headaches for many years but got recurrence of CH within a few weeks after the administration of long-acting organic nitrates (isosorbide-dinitrate, isosorbide-5-mononitrate or slow-release GTN) aimed at treating their chest pains. These nitrate-induced headache periods were more severe and had a longer duration than the previous spontaneous ones. Furthermore, one of the subjects and two additional cases experienced a marked reduction of their anginal attacks during successive CH periods. Exercise time to effort-induced angina was increased in all three patients and one of them revealed a markedly elevated threshold for eliciting ischaemic cardiac symptoms by standardized physical exercise on a cycle ergometer. We hypothesize whether extra CH periods elicited by sustained nitrate therapy and remission of angina pectoris during active clusters are caused by central mechanisms involving inhibition of sympathetic tone and effects on both cranial vessels and cardiac functions.

Age at onset and sex ratio in cluster headache: observations over three decades.

Five hundred and fifty-four patients with episodic cluster headache (ECH) and chronic cluster headache (CCH) were examined between 1963 and 1997. Mean age at onset was significantly higher in women with CCH compared with women with ECH and in men with ECH or CCH. In women with CCH age at onset was evenly distributed from 10 to 69 years, whereas in men with CCH and in both sexes with ECH, there was a peak when they were in their 20s. In women with ECH a second peak of onset occurred in their 50s. Although not statistically significant, primary CCH started later in women (mean 50.8 years) than secondary CCH (mean 35.5 years). There was a significant variation in the male : female ratio with respect to age at onset, being largest between 30 and 49 years of age (ECH 7.2 : 1; CCH 11.0 : 1) and lowest after 50 (ECH 2.3 : 1; CCH 0.6 : 1). During the observation period of more than 30 years there was a trend towards a decreasing male preponderance; the male : female ratio was significantly higher among patients with onset before rather than after 1970. The proportion of episodic vs. chronic CH did not change during the study period. The nature of the sex- and age-related pattern of cluster headache onset remains to be elucidated but mechanisms associated with sex hormone regulation, perhaps of hypothalamic origin, may be involved, as well as environmental factors related to lifestyle.

Genetic and environmental influences on recurrent headaches in eight to nine-year-old twins.

The major objective of the present study was to estimate genetic and environmental influences on recurrent headaches in prepubescent twins. A nationwide cohort of 8 to 9-year-old Swedish twins (n = 1,480) was screened for nonsymptomatic and recurrent headaches through a questionnaire mailed to their parents (75% response frequency). Among positives, 79% of headaches were classified as migraine or tension-type headache in close accordance with the International Headache Society criteria. The prevalences of migraine and nonmigrainous headaches were 2.4% and 11.3%, respectively, and without significant differences between the sexes or zygosity types. Inheritance on liability to recurrent headaches was estimated to 70% (a2 = 0.70, 95% CI = 0.54-0.82) for boys and girls but the genetic effect was found to be uncorrelated between the sexes. We conclude that genetic and individual specific effects are important for recurrent headaches of migrainous and nonmigrainous types in prepubescent children, and that different genetic etiologies might exist for boys and girls.

Exposure to motor vehicle exhaust and childhood cancer.

OBJECTIVES: A case-referent study was conducted to test the hypothesis that exposure to motor vehicle exhaust increases the risk of childhood cancer. METHODS: Data from a study of residential magnetic field exposure and childhood cancer were used. From a population of 127000 children living within 300 m of transmission lines in Sweden, 142 cases of childhood cancer were identified, including 39 cases of leukemia and 33 cases of central nervous system tumor. Approximately 4 referents per case were selected at random from the study base. The nitrogen dioxide content of the outdoor air was estimated as an indicator of motor vehicle exhaust. The applied methods give the 99th percentile of the nitrogen dioxide content of the outdoor air for 1-h averages over 1 year. RESULTS: A relative risk estimate of 2.7 [95% confidence interval (95% CI) 0.9-8.5] was found for total cancer at exposure levels of > or = 50 microg/m3, related to those with < or = 39 microg/m3. At > or = 80 microg/m3, the relative risk was estimated at 3.8 (95% CI 1.2-12.1). Elevated, but imprecise risk estimates were found for leukemia and central nervous system tumors. CONCLUSIONS: The results indicate an association between childhood cancer and motor vehicle exhaust, although the number of cases was small. These findings and the results of previous studies suggest that further studies of the association between motor vehicle exhaust and childhood cancer are warranted.

Structural and metabolic requirements for activators of the peroxisome proliferator-activated receptor.

Fatty acids have recently been demonstrated to activate peroxisome proliferator-activated receptors (PPARs) but specific structural requirements of fatty acids to produce this response have not yet been determined. Importantly, it has hitherto not been possible to show specific binding of these compounds to PPAR. To test whether a common PPAR binding metabolite might be formed, we tested the effects of long-chain omega-3 polyunsaturated fatty acids, differentially beta-oxidizable fatty acids and inhibitors of fatty acid metabolism. We determined the activation of a reporter gene by a chimaeric receptor encompassing the DNA binding domain of the glucocorticoid receptor and the ligand binding domain of PPAR. The omega-3 unsaturated fatty acids were slightly more potent PPAR activators in vitro than saturated fatty acids. The peroxisomal proliferation-inducing, non-beta-oxidizable, tetradecylthioacetic acid activated PPAR to the same extent as the strong peroxisomal proliferator WY 14,643, whereas the homologous beta-oxidizable tetradecylthiopropionic acid was only as potent as a non-substituted fatty acid. Cyclooxygenase inhibitors, radical scavengers or cytochrome P450 inhibitors did not affect activation of PPAR. In conclusion, beta-oxidation is apparently not required for the formation of the PPAR-activating molecule and this moiety might be a fatty acid, its ester with CoA, or a further derivative of the activated fatty acid prior to beta-oxidation of the acyl-CoA ester. These data should aid understanding of signal transduction via PPAR and the identification of a receptor ligand.

Increased expression of the female-predominant cytochrome P4502C12 in liver nodules from male Wistar rats.

Nodules and hepatomas from male and female rats treated according to the resistant hepatocyte (RH) model were analyzed with respect to expression of the male-predominant cytochrome P4502C11 (P450(16 alpha)) and the female-predominant cytochrome P4502C12 (P450(15 beta)) at the transcriptional and at the mRNA and protein levels. In male nodules isolated 8 and 11 to 12 months after initiation and in hepatomas, the expression of P450(16 alpha) mRNA was 3- to 11-fold lower than in surrounding liver, whereas a 2- to 8-fold higher expression of P450(15 beta) was observed compared with surrounding tissue. These alterations in P450 mRNA expression were reflected by similar changes at the protein level. Nuclear transcription of the cytochrome P450(16 alpha) gene was lower in male nodules than in surrounding liver whereas transcription of the P450(15 beta) gene was higher in the nodules. In nodules and hepatomas from female rats no significant differences in either mRNA expression or protein level of either P450(16 alpha) or P450(15 beta) were seen. The present study indicates that liver nodules are, to some extent, withdrawn from the normal endocrine regulation of rat liver function. Furthermore, the observed increase in a specific cytochrome P450 species (P450(15 beta)) in male liver nodules contradicts the previous suggestion of a general downregulation of this enzyme family as a characteristic of the nodular phenotype.

Sex-differentiated deoxycholic acid promotion of rat liver carcinogenesis is under pituitary control.

The influence of continuous growth hormone (GH) infusion and of implantation of ectopic pituitary grafts (PG) to male rats on the sex differences (male greater than female) in efficiency of promotion with dietary deoxycholic acid (DCA; 0.5% w/w) was studied in the livers of diethylnitrosamine (DEN)-initiated Wistar rats. For comparison, liver regeneration after partial hepatectomy (PH) was examined in differently treated animals. The endpoints examined included the number and size of enzyme-altered foci, hepatic c-myc expression and liver weight gain. The area per focus was 2- to 3-fold larger in initiated, DCA-treated males than in the corresponding PG-bearing males, GH-treated males and in females. The expression of the c-myc gene was increased approximately 2-fold in initiated and promoted males, while the increase in females was very small. In both groups of hormone-treated males the expression was at the same level as in females, significantly lower than in the corresponding DEN/DCA-treated males. Liver weight gain in response to PH in initiated as well as uninitiated rats of both sexes was significantly stimulated by DCA. No sex differences or effects of PG on regeneration could be discerned. In conclusion, a sex difference, regulated by a pituitary influence, in focal growth and in c-myc expression was observed during DCA promotion of DEN-initiated rats. This might indicate a correlation between the GH-regulated, proliferation-associated c-myc gene and focal growth during sex differentiated promotion in rat liver. Furthermore, the lack of sex differences in liver weight gain in response to PH during DCA treatment suggests that selective mitoinhibition is not involved, and might in this model indicate hormone-dependent selective stimulation of focal growth as a mechanism for tumor promotion.