The biological activity of serum bacterial lipopolysaccharides associates with disease activity and likelihood of achieving remission in patients with rheumatoid arthritis.

BACKGROUND: Dysbiotic intestinal and oral microbiota have been implicated in the pathogenesis of rheumatoid arthritis (RA), but the mechanisms how microbiota could impact disease activity have remained elusive. The aim of this study was to assess the association of the biological activity of serum lipopolysaccharides (LPS) with disease activity and likelihood of achieving remission in RA patients. METHODS: We measured Toll-like receptor (TLR) 4-stimulating activity of sera of 58 RA patients with a reporter cell line engineered to produce secreted alkaline phosphatase in response to TLR4 stimulation. Levels of LPS-binding protein, CD14, and CD163 were determined by ELISA assays. RESULTS: The patient serum-induced TLR4 activation (biological activity of LPS) was significantly associated with inflammatory parameters and body mass index at baseline and at 12 months and with disease activity (DAS28-CRP, p<0.001) at 12 months. Importantly, baseline LPS bioactivity correlated with disease activity (p=0.031) and, in 28 early RA patients, the likelihood of achieving remission at 12 months (p=0.009). The level of LPS bioactivity was similar at baseline and 12-month visits, suggesting that LPS bioactivity is an independent patient-related factor. Neutralization of LPS in serum by polymyxin B abrogated the TLR4 signaling, suggesting that LPS was the major contributor to TLR4 activation. CONCLUSION: We describe a novel approach to study the biological activity of serum LPS and their impact in diseases. The results suggest that LPS contribute to the inflammatory burden and disease activity on patients with RA and that serum-induced TLR4 activation assays can serve as an independent prognostic factor. A graphical summary of the conclusions of the study.

Pharmacokinetics of quercetin from quercetin aglycone and rutin in healthy volunteers.

BACKGROUND: Quercetin is a flavonoid with a wide range of biological activities. It mainly occurs in plants as glycosides, such as rutin (quercetin rutinoside) in tea. Quercetin and rutin are used in many countries as vasoprotectants and are ingredients of numerous multivitamin preparations and herbal remedies. OBJECTIVES: The primary objective was to characterise and compare the absorption and the pharmacokinetics of quercetin from quercetin aglycone and rutin. A secondary objective was to investigate which forms of quercetin are present in plasma. METHODS: In this double blind, diet-controlled, two-period cross-over study, 16 healthy volunteers received three different doses of quercetin and rutin orally. The doses corresponded to 8 mg, 20 mg and 50 mg quercetin aglycone. Blood samples were obtained between 0 h and 32 h post-dose. RESULTS: The overall kinetic behaviour of quercetin differed remarkably after ingestion of quercetin aglycone or rutin. The mean area under the plasma concentration-time curve from 0 h to 32 h [AUC(0-32)] and maximum plasma concentration (Cmax) values of the two treatments were similar. However, time to reach Cmax (tmax) was significantly shorter after the quercetin aglycone treatment than after the rutin treatment (1.9, 2.7 and 4.8 versus 6.5, 7.4 and 7.5 h, for doses 1, 2 and 3, respectively). Also, the absorption of quercetin from quercetin aglycone was predictable and inter-individual variation was small. In contrast, after ingestion of rutin, inter-individual variations in AUC(0-32) and Cmax values were considerable and seemed to be associated with gender and use of oral contraceptives. Quercetin and rutin were found in plasma as glucuronides and/or sulfates of quercetin and as unconjugated quercetin aglycone, but no rutin was detected. CONCLUSIONS: In clinical trials, studying the effects of quercetin from rutin, bioavailability must be taken into consideration and plasma quercetin concentrations monitored. Whether our results apply to other glycosidic drugs as well, especially other rutosides, should be investigated.

Catechol estrogens as inhibitors of leukotriene synthesis.

Estrogens have a beneficial effect on atherosclerosis and osteoporosis after menopause, but their exact mechanism of action is still unknown. The aim of the present study was to investigate the effects of estradiol and its metabolites catechol estrogens on arachidonic acid metabolism in vitro. Estradiol had no effect on arachidonic acid metabolism up to 33 microM in A23187-stimulated human whole blood. All catechol estrogens (2-hydroxyestradiol, 2-hydroxyestrone, 4-hydroxyestradiol and 4-hydroxyestrone) had similar kinds of actions on arachidonic acid metabolism, being over ten times more potent inhibitors of leukotriene synthesis (IC50 values 0.044-0.16 microM) than thromboxane (IC50 values 0.99-2.1 microM) and prostaglandin E2 synthesis (IC50 values 0.84-5.5 microM). It is suggested that some of the protective actions of estrogens--e.g., on atherosclerosis and osteoporosis--may be related to the inhibition of leukotriene synthesis by catechol estrogens.

Changes in inflammation-related blood constituents of mastitic cows.

The following blood constituents were measured in the blood and/or serum from healthy and mastitic cows: lipid peroxides (LPO), alkaline phosphatase (AP), lactate dehydrogenase (LDH), urea, glutathione peroxidase (GSH-Px), prostaglandin E2 (PGE2), cyclic AMP (cAMP), electrolytes (Na, K), white blood cells (WBC), haemoglobin (Hb), haematocrit, eosinophils, and electrical conductivity. The levels of these blood variables, excluding GSH-Px, erythrocytes variables and conductivity, were higher in mastitic than in healthy animals. In the blood of the mastitic animals, the AP and LPO levels were 44 and 38% higher than in the healthy animals. The substantial decline in the GSH-Px levels in mastitic animals (P < 0.05) may be related to changes in lipid peroxidation and PG formation and possibly to oxidative stress caused by the host defences. The eosinophil levels were positively correlated with those of PGE2 and cAMP. The possible implications of these findings for the diagnosis of mastitis are discussed.

Catecholamines inhibit leukotriene formation and decrease leukotriene/prostaglandin ratio.

Adrenaline, noradrenaline, isoprenaline, and to a lesser extent dopamine inhibit the release of leukotriene (LT) B2 from calcium ionophore-stimulated human polymorphonuclear leukocytes, while the release of prostaglandin (PG) E2 is proportionally elevated. The inactivity of salbutamol, a noncatechol adrenergic beta 2-receptor agonist, and the inability of propranolol to antagonize the effects of adrenaline, suggest the mediation through beta-receptor independent mechanisms. Neither are alpha-1-receptors involved, as prazosin, a specific antagonist, fails to inhibit the reaction. As the principles for biochemical regulation of LT- and PG-production are met by catecholamines in several tissues, the mechanism is considered to be of general physiological importance. Catecholamines may function as coenzymes/antioxidants which, by altering the redox state of the enzyme iron or heme, decrease the LT/PG ratio thus protecting the organism against tissue anaphylaxis and other LT-related pathophysiology.

A controlled study with taltrimide and sodium valproate: valproate effective in partial epilepsy.

Taltrimide was compared with valproate and placebo in 17 patients with intractable epilepsy being on carbamazepine monotherapy. Taltrimide (400 mg/day), valproate (1000 mg/day) or placebo were added to the treatment for periods of 3 months using a randomized cross-over design. Serum carbamazepine concentrations remained within the therapeutic range throughout the trial. Thirteen patients completed the study. In partial epilepsy of 7 the seizure frequency was reduced by 27% during valproate (p less than 0.05), compared with placebo, while no improvement was found during taltrimide. In 6 with primary generalized epilepsy, the number of seizures was reduced by 49% during taltrimide and by 38% during valproate, but neither effect was significant, compared with placebo. Headache was reported by 3 patients while on taltrimide. One with hypersensitivity history developed petecchiae and nasal bleeding during taltrimide and, therefore, the treatment was stopped. The three other interruptions were independent of taltrimide. Thus, the only statistically significant effect in this study was that of valproate in partial epilepsy.

Role of plasma fatty acids, prostaglandins and antioxidant balance in bovine mastitis.

Plasma levels of free fatty acids (FFA) were measured in cows with mastitis. FFA levels are known to be elevated in milk of affected cows, and similar increases were now observed in blood plasma. Of the long-chain polyunsaturated fatty acids which serve as precursors for prostaglandins (PGs), leukotrienes (LT) and related eicosanoids, arachidonic acid (C20: 4 omega-6) was elevated by 156 per cent and C20: 3 (consisting of both omega-3 and omega-6) 153 per cent; eicosapentaenoic acid (20: 5 omega-3), not present in the plasma of healthy animals, was detected in blood of mastitic animals. Arachidonic acid correlated positively with the plasma levels of PGs. Total plasma phospholipids were increased as well, and they were positively correlated with the levels of linoleic and arachidonic acids. Changes in FFA levels correlated positively with parameters of inflammation, like bovine serum albumin and serum N-acetyl-beta-D-glucosaminidase, but negatively with the antioxidant status of the animals (vitamin E, glutathione peroxidase). As PGs and LTs are mediators of inflammation, and products of lipid peroxidation, the increase of plasma FFA could be related to tissue damage caused by oxygen free radicals. Similar processes are, however, involved also in host defence, e.g. in oxidative killing of pathogens.

Free fatty acids and lipid peroxidation in normal and mastitic bovine milk.

Milk levels of free fatty acids (FFA) and prostaglandins (PGs) were determined in healthy cows and in those with mastitis and compared with some parameters commonly used for diagnosing mastitis. A modified gas chromatography-mass spectrometry method was used. In mastitis there was a substantial overall increase in FFA levels in milk. Specific attention was paid to the fate of the long-chain polyunsaturated fatty acids, known as precursors for the production of PGs, leukotrienes and other eicosanoids. Both the omega-6 and omega-3 lines of these lipids were increased in mastitis, but only in proportion to other FFA. The milk levels of PGs (PGF2, PGF2 alpha, thromboxane B2) were elevated in the same proportion. The possible importance of lipid peroxidation, inflammatory reaction and host defence are evaluated in relation to tissue destruction and oxidative killing of pathogens. It may be possible to manipulate the intake and fate of FFA in the organism by dietary intake, antioxidants and proper storing of feed. It is concluded that in studies concerning FFA special attention should be given to those long-chain fatty acids that are precursors for the formation of eicosanoids.

Inflammation-related changes in cyclic AMP and cyclic GMP in bovine mastitis.

Cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) concentrations in milk and plasma samples from healthy and mastitic cows were determined by radioimmunoassay and compared with prostaglandins (PGE2, PGF2 alpha and thromboxane B2 [TXB2]), phospholipids and other relevant parameters in milk and blood. The concentrations of cAMP were about five times higher in plasma (p less than 0.01) than in milk, whereas the cGMP concentration in milk was three times higher (p less than 0.01) than that in plasma in both healthy and diseased animals. In mastitic milk, the cAMP and cGMP concentrations were 19% and 65% and in blood plasma 13% and 84% higher respectively than in healthy animals. In milk, cyclic nucleotide concentrations correlated with the markedly elevated cell count and also with the prostaglandin concentration and pH. In blood, cAMP correlated positively with phospholipids and cGMP with reduced glutathione (GSH). These changes are considered to be important in the disease process and, in particular, the increase in cGMP deserves further study.

Evidence for the presence of cytochrome P-450 in mastitic bovine mammary gland.

The content of cytochrome P-450 in mastitic and healthy lactating bovine mammary gland was analyzed. Evidence for the existence of cytochrome P-450 in mastitic tissues was found. The amount of cytochrome P-450 formed in the assay ranged from 0.017 to 0.031 nmole/mg protein. The highest concentrations were found in severely inflamed samples, whereas and traces or no cytochrome P-450 were found in healthy tissues. The significance of cytochrome P-450 in mastitic mammary gland is discussed in relation to host defence involving oxidative killing of pathogens.

Milk and blood levels of silicon and selenium status in bovine mastitis.

Milk and blood levels of silicon, selenium and the selenoenzyme glutathione peroxidase (GSH-Px) were measured in 20 healthy and 21 mastitic cows. In milk samples from healthy quarters the mean silicon concentration was 0.81 and in affected ones 0.39 ppm. In serum the mean silicon values were 1.63 and 1.02 ppm respectively. The selenium status was not altered but the level of erythrocyte GSH-Px was lowered in mastitic animals. Silicon is known to have marked effects on free radical formation, lipid peroxidation and macrophage activity. Its possible role in infection and inflammation is evaluated. Some of the functions of silicon may resemble those of selenium. The possibility of lowered levels of silicon and of the selenoenzyme in mastitis calls for experimentation with dietary or pharmaceutical supplementation of these trace elements.

Milk prostaglandins and electrical conductivity in bovine mastitis.

Prostaglandin (PG) levels in milk samples from healthy and mastitic cows were determined by radioimmunoassay. In composite milk the PG levels were rather high both in healthy and mastitic samples, and the only significant difference was in thromboxane B2 (TXB). In quarter milk samples classified according to the degree of mastitis by use of somatic cell counts, PGE2 was 40, PGF2 alpha 15, and TXB2 44 per cent higher respectively in affected samples. PG levels were in good correlation with somatic cell counts (r = 0.63-0.68, p less than 0.01) and electrical conductivity (r = 0.36-0.52, p less than 0.01), two established criteria for diagnosis of mastitis. PGE2 also correlated with protein, TXB2 with fat content. PGF 2 alpha was in a negative correlation with milk yield. The good correlation of PGs with somatic cell counts and electrical conductivity suggests that PGs might be used as markers of mastitic inflammation.

Possible roles of vitamin E and glutathione metabolism in bovine mastitis.

Plasma and milk levels of vitamin E were determined in mastitic and healthy cows and compared with erythrocyte GSH and GSH-peroxidase, selenium, silicon, prostaglandins and parameters commonly used for diagnosing mastitis. In mastitis both the plasma and milk levels of vitamin E were significantly lowered. In the milk vitamin E correlated negatively with electrical conductivity and PGE2. In the blood vitamin E was in a positive correlation with erythrocyte GSH. The role of lipid peroxidation in relation to the inflammatory and immunological reactions of mastitis is discussed.

Prostaglandins and glutathione peroxidase in bovine mastitis.

Prostaglandin (PG) levels in milk and plasma samples from mastitic cows were determined by radioimmunoassay and compared with erythrocyte glutathione peroxidase (GSH-Px) and other relevant parameters in milk and blood. The overall levels of PGE2, PGF2 alpha and thromboxane B2 (TXB2) in milk were two to four times higher than in blood plasma both in healthy and diseased animals (P less than 0.01). In mastitic milk the PG levels were 24 to 55 per cent and in blood plasma 41 to 95 per cent higher than in healthy animals. The changes were significant and largest for the PGF2 alpha values. In milk, the PG concentrations correlated with the markedly elevated cell count (r = 0.49 to 0.57), and TXB2 values also correlated with milk yield. In blood, PGE2 and PGF2 alpha correlated positively with serum albumin, and PGE2 also correlated with glutathione (GSH). PGE2 and PGF2 alpha correlated negatively with GSH-Px and gamma-glutamyl transferase. The substantial decline in GSH-Px in mastitic animals (P less than 0.01) may be related to changes in lipid peroxidation and PG formation. The possible implications of these findings in the treatment of mastitis are discussed.

Relevance of prostaglandins in migraine.

Prostaglandins (PG), particularly PGE, may be linked to the pathophysiology of migraine in several important ways. PGE1 may "simulate" a migraine attack in healthy volunteers. PGE may be elevated in patients with migraine. In animal experiments and in human infusions, PGEs cause vasodilation and hyperalgesia, both typical reactions of inflammation. The view that vascular headache is an "inflammatory reaction" allows the best concept concerning the local role of PGs and the effectiveness of PG-inhibitors in the treatment of migraine. The local role of PGs may provide a common denominator in several hormonal, neural and other influences on vessels. The common triggers of a migraine attack like menstruation, alcohol and stress influence the PG-system and even the dietary reactions, hormonal influences, sleep and reserpine have some connections with the PG-system. A local role for PGs does not diminish the importance of other pathophysiological mechanisms operating during an attack. On the contrary, PGs may fill in gaps in our understanding of how the overt pain of attacks is produced.