Differences in preterm and term milk fatty acid compositions may be caused by the different hormonal milieu of early parturition

Introduction

The hormonal milieus of pregnancy and lactation are driving forces of nutrient fluxes supporting infant growth and development. The decrease of insulin sensitivity with compensatory hyperinsulinemia with advancing gestation, causes adipose tissue lipolysis and hepatic de novo lipogenesis (DNL).

Subjects and methods

We compared fatty acid (FA) contents and FA-indices for enzyme activities between preterm (28–36 weeks) and term (37–42) milks, and between colostrum (2–5 days), transitional (6–15) and mature (16–56) milks. We interpreted FA differences between preterm and term milks, and their changes with lactation, in terms of the well known decrease of insulin sensitivity during gestation and its subsequent postpartum restoration, respectively.

Results

Compared with term colostrum, preterm colostrum contained higher indices of DNL in the breast (DNL-breast) and medium chain saturated-FA (MCSAFA), and lower DNL-liver and monounsaturated-FA (MUFA). Preterm milk also had higher docosahexaenoic acid (DHA) in colostrum and transitional milk and higher arachidonic acid (AA) in mature milk. Most preterm-term differences vanished with advancing lactation. In both preterm and term milks, DNL-breast and MCSAFA increased with advancing lactation, while DNL-liver, MUFA, long chain SAFA and AA decreased. DHA decreased in term milk. MUFA was inversely related to MCSAFA in all samples, correlated inversely with PUFA in colostrum and transitional milks, but positively in mature milk. MCSAFA correlated inversely with PUFA in mature milk.

Conclusion

Higher maternal insulin sensitivity at preterm birth may be the cause of lower MUFA (a proxy for DNL-liver) and higher MCSAFA (a proxy for DNL-breast) in preterm colostrum, compared with term colostrum. Restoring insulin sensitivity after delivery may be an important driving force for milk FA-changes in early lactation.     

Gestational age dependent changes of the fetal brain, liver and adipose tissue fatty acid compositions in a population with high fish intakes

IntroductionThere are no data on the intrauterine fatty acid (FA) compositions of brain, liver and adipose tissue of infants born to women with high fish intakes.Subjects and methodsWe analyzed the brain (n=18), liver (n=14) and adipose tissue (n=11) FA compositions of 20 stillborn infants with different gestational ages (range 8–38 weeks) born to Tanzanian women with low linoleic acid (LA) intakes and high intakes of docosahexaenoic (DHA) and arachidonic (AA) acids from local fish.Results and discussionWith advancing gestation, brain saturated-FA (SAFA; in g/100 g FA), polyunsaturated-FA (PUFA), DHA, 20:3ω6, 22:4ω6 and 22:5ω6 increased, while monounsaturated-FA (MUFA), 20:3ω9, 22:3ω9 and AA decreased. Decreasing brain AA might be caused by increasing AA-metabolism to 20:3ω6, 22:4ω6 and 22:5ω6. In the liver, SAFA, PUFA and LA increased, while MUFA decreased with gestation. The steep increase of (mostly de novo synthesized) SAFA in adipose tissue coincided with relative decreases of MUFA, PUFA, DHA, LA and AA with advancing gestation. Compared to Western infants, the currently studied African infants had higher DHA, lower AA, and a higher DHA/AA-ratio in brain and adipose tissue, while the LA content of adipose tissue was lower.ConclusionThe low LA and high DHA and AA intakes by the mothers of these infants might support optimal α-linolenic (ALA) vs. LA competition for Δ5D and Δ6D-activities and DHA vs. AA antagonism. Conversely, the Western diet, characterized by high LA and lower DHA and AA intakes, might disturb these evolutionary conserved mechanisms aiming at an optimal ω3/ω6-balance.     

Modulation of blood oxylipin levels by long-chain omega-3 fatty acid supplementation in hyper- and normolipidemic men

IntroductionLong chain omega-3 polyunsaturated fatty acids (LC n-3 PUFA) such as EPA and DHA have been shown to possess beneficial health effects, and it is believed that many of their effects are mediated by their oxygenated products (oxylipins). Recently, we have shown that serum levels of several hydroxy, epoxy, and dihydroxy FAs are dependent on the individual status of the parent FAs in a cohort of normo- and hyperlipidemic subjects. So far, the effect of an increased dietary LC n-3 PUFA intake on hydroxy, epoxy, and dihydroxy FA levels has not been investigated in subjects with mild combined hyperlipidemia.Subjects and methodsIn the present study, we compared oxylipin patterns of 10 hyperlipidemic (cholesterol >200 mg/dl; triglyceride >150 mg/ml) and 10 normolipidemic men in response to twelve weeks of LC n-3 PUFA intake (1.14 g DHA and 1.56 g EPA). Levels of 44 free hydroxy, epoxy and dihydroxy FAs were analyzed in serum by LC-MS. Additionally, oxylipin levels were compared with their parent PUFA levels in erythrocyte membranes; a biomarker for the individual PUFA status.ResultsDifferences in the oxylipin pattern between normo- and hyperlipidemic subjects were minor before and after treatment. In all subjects, levels of EPA-derived oxylipins (170–4800 pM) were considerably elevated after LC n-3 PUFA intake (150–1400%), the increase of DHA-derived oxylipins (360–3900 pM) was less pronounced (30–130%). The relative change of EPA in erythrocyte membranes is strongly correlated (r≥0.5; p<0.05) with the relative change of corresponding epoxy and dihydroxy FA serum levels. The effect on arachidonic acid (AA)-derived oxylipin levels (140–27,100 pM) was inconsistent.Discussion and conclusionsThe dietary LC PUFA composition has a direct influence on the endogenous oxylipin profile, including several highly biological active EPA- and DHA-derived lipid mediators. The shift in oxylipin pattern appears to be dependent on the initial LC PUFA status particularly for EPA. The finding that also levels of other oxylipins derived from ALA, LA or AA are modified by LC n-3 PUFA intake might suggest that at least some of the effects of EPA and DHA could be mediated by a shift in the entire oxylipin profile.     

Intrauterine, postpartum and adult relationships between arachidonic acid (AA) and docosahexaenoic acid (DHA)

Erythrocyte (RBC) fatty acid compositions from populations with stable dietary habits but large variations in RBC-arachidonic (AA) and RBC-docosahexaenoic acid (DHA) provided us with insight into relationships between DHA and AA. It also enabled us to estimate the maternal RBC-DHA (mRBC-DHA) status that corresponded with no decrease in mRBC-DHA during pregnancy, or in infant (i) RBC-DHA or mRBC-DHA during the first 3 months postpartum (DHA-equilibrium) while exclusively breastfeeding. At delivery, iRBC-AA is uniformly high and independent of mRBC-AA. Infants born to mothers with low RBC-DHA exhibit higher, but infants born to mothers with high RBC-DHA exhibit lower RBC-DHA than their mothers. This switch from ‘biomagnification’ into ‘bioattenuation’ occurs at 6 g% mRBC-DHA. At 6 g%, mRBC-DHA is stable throughout pregnancy, corresponds with postpartum infant DHA-equilibrium of 6 and 0.4 g% DHA in mature milk, but results in postpartum depletion of mRBC-DHA to 5 g%. Postpartum maternal DHA-equilibrium is reached at 8 g% mRBC-DHA, corresponding with 1 g% DHA in mature milk and 7 g% iRBC-DHA at delivery that increases to 8 g% during lactation. This 8 g% RBC-DHA concurs with the lowest risks of cardiovascular and psychiatric diseases in adults. RBC-data from 1866 infants, males and (non-)pregnant females indicated AA vs. DHA synergism at low RBC-DHA, but antagonism at high RBC-DHA. These data, together with high intakes of AA and DHA from our Paleolithic diet, suggest that bioattenuation of DHA during pregnancy and postnatal antagonism between AA and DHA are the physiological standard for humans across the life cycle.     

Different roles of zinc plus arachidonic acid on insulin sensitivity between high fructose- and high fat-fed rats

AimsThis study was to determine the effects of zinc plus arachidonic acid (ZA) treatment on the insulin action in the specific ZA target organs using hyperinsulinemic euglycemic clamp method.Main methods18 Sprague–Dawley rats weighing ~ 130 g were divided into 3 groups of 6 rats and treated them with 1) normal rat chow, 2) high fructose (60.0%) diet only, or 3) the same fructose diet plus drinking water containing 10 mg zinc plus 50 mg arachidonic acid (AA)/L. In a separate study, male Wistar rats weighing ~ 250 g were fed normal rat chow (n = 4) or high fat (66.5%) diet with drinking water containing zero (n = 9) or 10 mg AA plus 20 mg zinc /L (n = 9). After 4 week treatment, insulin action was assessed using the hyperinsulinemic eguglycemic clamp technique.Key findingsHigh fructose feeding impaired suppression of hepatic glucose output by insulin compared to controls during the clamp procedure (4.39 vs. 2.35 mg/kg/min; p < 0.05). However, ZA treatment in high fructose-fed rats showed a significant improvement of hepatic insulin sensitivity compared to non-treatment controls (4.39 vs. 2.18 mg/kg/min; p < 0.05). Glucose infusion rates in Wistar rats maintained on a high fat diet (HFD) were significantly lower compared to control rats (22.8 ± 1.3 vs. 31.9 ± 1.4 mg/kg/min; p < 0.05). ZA treatment significantly improved (~ 43%) peripheral tissue insulin sensitivity in HFD fed animals (26.7 ± 1.3 [n = 9] vs. 22.8 ± 1.3 mg/kg/min; p < 0.05).SignificanceThese data demonstrate that ZA treatment is effective in improving glucose utilization in hyperglycemic rats receiving either a high-fructose or a high-fat diet.     

Dietary supplementation with maize distillers dried grains with solubles during late gestation and lactation: Effects on sow and litter performance,and on colostrum and milk composition

The objectives of this study were to determine the effects of maize distillers dried grains with solubles (DDGS) during late gestation and lactation on sow and piglet performance, and on colostrum and milk composition. Thirty-six second- and third-parity (2.43 parity) sows (Yorkshire) were allotted to 1 of 3 groups and fed diets containing 0 (control), 200 or 400 g DDGS/kg during the last 20 d of gestation and throughout a 21 d of lactation. Experimental diets contained 12.9 MJ metabolizable energy/kg and 9.7 g lysine/kg. The colostrum and milk samples were obtained on d 0 (farrowing) and d 21 (weaning). There were no differences (P>0.05) in the sows’ average gestation lengths, weaning-to-estrus interval, average daily feed intake, and the lactation backfat and body weight change between dietary treatments. There were no dietary effects (P>0.05) of DDGS on the numbers of total, born alive piglets, average birth weights, piglets per litter at weaning or piglets average daily gain during lactation. No differences (P>0.05) were observed in total solids, protein, fat and lactose among the sows fed the DDGS diets compared with the control. The composition of total solids and protein of sows colostrum and milk were higher at farrowing (d 0) than at weaning (d 21) (P<0.001). However, the fat and lactose content of sows colostrum and milk were increased (P<0.001) from d 0 (farrowing) to d 21 (weaning). In conclusion, the results suggest that 400 g DDGS/kg (87 g lysine/kg) supplemented with 5.2 g lysine/kg included in late gestation and lactation diets is sufficient to replace all the dietary soybean meal without significantly affecting sow and litter performance or colostrum and milk composition.     

Effect of dietary selenite on development and intestinal absorption in offspring rats

AimThe present study aims to compare selenium (Se) status in offspring rats born to selenium-deficient and selenium supplemented dams and to analyse Se's influence on intestinal parameters and the intestinal absorption of selenomethionine (Se-Met).Main methodsMale and female Wistar rats (150–200 g) were randomised in: control (C) (0.1 ppm Se), Se-deficient (SD) (0.01 ppm Se) and Se-supplemented (SS) (0.5 ppm Se) groups; and were mated to obtain their offspring. Se levels in serum, urine and faeces in offspring and in mothers' milk were measured by graphite-furnace atomic absorption spectrometry. Duodenal transport studies in offspring were performed using an in vivo perfusion of different Se-Met concentrations (2, 5, 10, 25, 75 and 150 μM).Key findingA Se-deficient diet provoked a decrease in the offspring's body weight and intestinal parameters, while the supplemented diet increased these values. Serum Se levels were similar between Se-deficient and control offspring because the urinary excretion of Se was smaller to compensate for Se homeostasis. Intestinal Se-Met absorption obeys the Michaellis–Menten equation with lower apparent constant (K_m) and maximal velocity (V_max) in the SD group. However, the C and SS groups presented similar K_m and different V_max. The V_max showed greater values in the following order of rank: SS > C > SD groups.SignificanceSelenium intake deficiencies in offspring lead to the development of compensatory mechanisms in order to normalise serum selenium levels. These mechanisms, however, do not permit normal body development; nor do they regulate intestinal parameters and Se-Met transport.     

Inducible COX-2 dominates over COX-1 in prostacyclin biosynthesis: mechanisms of COX-2 inhibitor risk to heart disease

AimOur aim is to understand the molecular mechanisms of the selective nonsteroidal anti-inflammatory drugs (NSAID), cyclooxygenase-2 (COX-2) inhibitors', higher “priority” to reduce synthesis of the vascular protector, prostacyclin (PGI2), compared to that of nonselective NSAIDs.Main methodsCOX-1 or COX-2 was co-expressed with PGI2 synthase (PGIS) in COS-7 cells. The Km and initial velocity (½t Vmax) of the coupling reaction between COX-1 and COX-2 to PGIS were established. The experiment was further confirmed by a kinetics study using hybrid enzymes linking COX-1 or COX-2 to PGIS. Finally, COX-1 or COX-2 and PGIS were respectively fused to red (RFP) and cyanic (CFP) fluorescence proteins, and co-expressed in cells. The distances between COXs and PGIS were compared by FRET.Key findingsThe Km for converting arachidonic acid (AA) to PGI2 by COX-2 coupled to PGIS is ~ 2.0 μM; however, it was 3-fold more (~ 6.0 μM) for COX-1 coupled to PGIS. The Km and ½t Vmax for COX-2 linked to PGIS were ~ 2.0 μM and 20 s, respectively, which were 2–5 folds faster than that of COX-1 linked to PGIS. The FRET study found that the distance between COX-2-RFP and PGIS–CFP is shorter than that between COX-1-RFP and PGIS–CFP.SignificanceThe study provided strong evidence suggesting that the low Km, faster ½t Vmax, and closer distance are the basis for COX-2 dominance over COX-1 (coupled to PGIS) in PGI2 synthesis, and further demonstrated the mechanisms of selective COX-2 inhibitors with higher potential to reduce synthesis of the vascular protector, PGI2.     

The delta 6 desaturase knock out mouse reveals that immunomodulatory effects of essential n-6 and n-3 polyunsaturated fatty acids are both independent of and dependent upon conversion

Typically fatty acids (FA) exert differential immunomodulatory effects with n-3 [α-linolenic acid (ALA), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)] and n-6 [linoleic acid (LA) and arachidonic acid (AA)] exerting anti- and pro-inflammatory effects, respectively. This over-simplified interpretation is confounded by a failure to account for conversion of the parent FA (LA and ALA) to longer-chain bioactive products (AA and EPA/DHA, respectively), thereby precluding discernment of the immunomodulatory potential of specific FA. Therefore, we utilized the Δ6-desaturase model, wherein knockout mice (D6KO) lack the Fads2 gene encoding for the rate-limiting enzyme that initiates FA metabolism, thereby providing a model to determine specific FA immunomodulatory effects. Wild-type (WT) and D6KO mice were fed one of four isocaloric diets differing in FA source (9 weeks): corn oil (LA-enriched), arachidonic acid single cell oil (AA-enriched), flaxseed oil (ALA-enriched) or menhaden fish oil (EPA/DHA-enriched). Splenic mononuclear cell cytokine production in response to lipopolysaccharide (LPS), T-cell receptor (TCR) and anti-CD40 stimulation was determined. Following LPS stimulation, AA was more bioactive compared to LA, by increasing inflammatory cytokine production of IL-6 (1.2-fold) and TNFα (1.3-fold). Further, LPS-stimulated IFNγ production in LA-fed D6KO mice was reduced 5-fold compared to LA-fed WT mice, indicating that conversion of LA to AA was necessary for cytokine production. Conversely, ALA exerted an independent immunomodulatory effect from EPA/DHA and all n-3 FA increased LPS-stimulated IL-10 production versus LA and AA. These data definitively identify specific immunomodulatory effects of individual FA and challenge the simplified view of the immunomodulatory effects of n-3 and n-6 FA.     

Cryopreserved mobilized autologous blood progenitors stored for more than 2 years successfully support blood count recovery after high-dose chemotherapy

Background aimsThe ability of hematopoietic progenitor cells–apheresis (HPC-A) that have been stored for many years after cryopreservation to reconstitute hematopoiesis following high-dose chemo/radiotherapy has not been well-documented.MethodsIn this retrospective study, eight Canadian centers contributed data from 53 autologous stem cell transplants (ASCT) performed using HPC-A that had undergone long-term storage (>2 years, range 2–7 years) and 120 ASCT using HPC-A stored for <6 months (short-term storage).ResultsThe doses of nucleated and CD34⁺ cells per kilogram recipient weight were similar between the short- (mean ± SD, 4.7 ± 4.9 × 10⁸ and 6.8 ± 4.3 × 10⁶, respectively) and long- (4.0 ± 4.9 × 10⁸ and 6.1 ± 3.4 × 10⁶, respectively) term storage groups. The median days to neutrophils (absolute neutrophil count; ANC) >0.5 × 10⁹/L (median 11 days for both short- and long-term storage) and platelets >20 × 10⁹/L (median 12 and 11 for short- and long-term storage, respectively) post-ASCT were not significantly different between the two groups. When ASCT performed with <5 × 10⁶/kg CD34⁺ cells was compared there was also no difference in ANC or platelet recovery (median 12 days for both after short-term storage, and 12 and 11 days, respectively, after long-term storage). Fourteen HPC-A products stored for >5 years also showed similar count recoveries as the entire long-term storage group (median 11 days for both ANC and platelets).ConclusionsCryopreserved HPC-A can be stored for at least 5 years with no apparent loss in their ability to support hematopoietic reconstitution after high-dose chemotherapy.     

Association between interleukin 15 receptor, alpha (IL15RA) polymorphism and Korean patients with ossification of the posterior longitudinal ligament

ObjectivesRecently, a number of evidences have been reported concerning the genetic factor involved in the development of ossification of the posterior longitudinal ligament (OPLL). The purpose of this study was to investigate single nucleotide polymorphisms (SNPs) of the interleukin 15 receptor, alpha (IL15RA) gene as a risk factor in Korean patients with OPLL.DesignTo investigate the genetic association, two coding SNPs (rs2296139, Thr73Thr; rs2228059, Asn182Thr) in IL15RA were genotyped in 166 OPLL patients and 230 control subjects. SNPStats, SNPAnalyzer, and Helixtree programs were used for association analysis.ResultsIn the present study, we found the association between a missense SNP (rs2228059) and the risk of OPLL in codominant (p = 0.0028, OR = 1.58, 95% CI = 1.17–2.14), dominant (p = 0.0071, OR = 1.82, 95% CI = 1.17–2.82), and recessive models (p = 0.036, OR = 1.79, 95% CI = 1.04–3.09). The frequency of rs2228059 allele was significantly associated with the susceptibility of OPLL (p = 0.0043, OR = 1.52, 95% CI = 1.14–2.02). After Bonferroni correction, the missense SNP (rs2228059, Asn182Thr) still had significant correlations (p = 0.0056 in codominant model; p = 0.0142 in dominant model; p = 0.0086 in allele analysis). Haplotype variation in IL15RA was associated with OPLL (global haplotype test, p = 0.025).ConclusionsThese results suggest that IL15RA polymorphism may be associated with the susceptibility of OPLL in Korean population.     

Gender-specific fatty acid profiles in platelet phosphatidyl-choline and -ethanolamine

Previous studies suggested that women synthesise docosahexaenoic acid (DHA) more efficiently from their precursors than men. This study investigated the relationship between diet, platelet phospholipids fatty acids and gender. Dietary intake and platelet phosphatidyl-choline (PC) and phosphatidylethanolamine (PE) fatty acids were determined in Caucasian 40 men and 34 women. Absolute and %energy intakes of arachidonic acid (AA), eicosapentaenoic acid (EPA), and DHA, and the ratios of total n-6/n-3 PUFA and linoleic/alpha-linolenic acids did not differ between the sexes. However, women had higher DHA in PC (1.19 vs 1.05 wt%, p<0.05) and PE (3.62 vs 3.21 wt%, p<0.05) than men. Also EPA (1.10 vs 0.93 wt%, p<0.05) was higher in women's PE. Conversely, men had elevated AA and total n-6 fatty acids in PC. The higher platelet DHA levels and lower platelet AA/EPA and AA/DHA ratios in women of child-bearing age compared with men, may lead to less platelet aggregation and vaso-occlusion.     

Milk in the island of Chole [Tanzania] is high in lauric, myristic, arachidonic and docosahexaenoic acids, and low in linoleic acid reconstructed diet of infants born to our ancestors living in tropical coastal regions

BACKGROUND: We need information on the diet on which our genes evolved. OBJECTIVE: We studied the milk fatty acid [FA] composition of mothers living in the island of Chole [Tanzania, Indian Ocean]. These mothers have high intakes of boiled marine fish and coconut, and consume plenty amount of fruits and vegetables. DESIGN: The outcome was compared with three fish-eating tribes living along Tanzanian freshwater lakes [Kerewe, Nyakius, Nyiramba], four tribes living in the Tanzanian inland [Hadzabe, Maasai, Sonjo, Iraqw] and our milk FA database. RESULTS: Milk from Chole contained high levels of 12:0 [20.17 g%], 14:0 [21.19], 12:0/14:0 ratio [0.92 g/g], arachidonic acid [AA, 0.50 g%] and docosahexaenoic acid [DHA, 0.73], but low levels of linoleic acid [LA, 4.23]. The combination of a high medium chain fatty acid [MCFA; 相似文献   

NADPH-d and Fos reactivity in the rat spinal cord following experimental spinal cord injury and embryonic neural stem cell transplantation

AimsThe aim of this study is to determine the role of nitric oxide (NO) in neuropathic pain and the effect of embryonic neural stem cell (ENSC) transplantation on NO content in rat spinal cord neurons following spinal cord injury (SCI).Main methodsNinety adult male Sprague–Dawley rats were divided into 3 groups (n = 30, each): control (laminectomy), SCI (hemisection at T12–T13 segments) and SCI + ENSC. Each group was further divided into sub-groups (n = 5 each) based on the treatment substance (L-NAME, 75 mg/kg/i.p.; l-arginine, 225 mg/kg/i.p.; physiological saline, SF) and duration (2 h for acute and 28 days for chronic groups). Pain was assessed by tail flick and Randall–Selitto tests. Fos immunohistochemistry and NADPH-d histochemistry were performed in segments 2 cm rostral and caudal to SCI.Key findingsTail-flick latency time increased in both acute and chronic L-NAME groups and increased in acute and decreased in chronic l-arginine groups. The number of Fos (+) neurons decreased in acute and chronic L-NAME and decreased in acute l-arginine groups. Following ENSC, Fos (+) neurons did not change in acute L-NAME but decreased in the chronic L-NAME groups, and decreased in both acute and chronic l-arginine groups. NADPH-d (+) neurons decreased in acute L-NAME and increased in l-arginine groups with and without ENSC transplantation.SignificanceThis study confirms the role of NO in neuropathic pain and shows an improvement following ENSC transplantation in the acute phase, observed as a decrease in Fos(+) and NADPH-d (+) neurons in spinal cord segments rostral and caudal to injury.     

Casein-derived tripeptide Ile-Pro-Pro improves angiotensin-(1-7)- and bradykinin-induced rat mesenteric artery relaxation

AimsMilk casein-derived bioactive tripeptides isoleucine–proline–proline (Ile–Pro–Pro) and valine–proline–proline (Val–Pro–Pro) lower blood pressure in animal models of hypertension and humans. In some studies, their angiotensin-converting enzyme (ACE)-inhibitory effect has been demonstrated. Besides classical ACE-angiotensin II-AT₁-receptor pathway (ACE-Ang II- AT₁), the significance of ACE2-angiotensin-(1–7)-Mas-receptor (ACE2-Ang-(1–7)-Mas) axis in the blood pressure regulation has now been acknowledged. The present study was aimed to further evaluate the renin–angiotensin system (RAS)-related vascular effects of Ile–Pro–Pro in vitro using rat mesenteric arteries.Main methodsSuperior mesenteric arteries of spontaneously hypertensive rat (SHR) were isolated, cut into rings and mounted in standard organ bath chambers. Endothelium-intact arterial rings were incubated in Krebs solution either with Ile–Pro–Pro, proline–proline (Pro–Pro), isoleucine (Ile), proline (Pro) or captopril for 6 h at + 37 °C and vascular reactivity was measured.Key findingsIn the presence of AT₁-antagonist valsartan, Ang II induced vasodilatation, which was more pronounced in the arteries incubated with Ile–Pro–Pro (P < 0.05) compared to the other compounds. Ang-(1–7)-induced vasodilatation was augmented by Ile–Pro–Pro or Pro (P < 0.001 vs. control). Mas-receptor antagonist A-779 did not alter the responses. Ile–Pro–Pro and Pro augmented also bradykinin-induced relaxations (P < 0.001 vs. control). Control arteries and arteries incubated with captopril showed only slight relaxations at higher bradykinin concentrations.SignificanceCasein-derived tripeptide Ile–Pro–Pro and amino acid Pro enhance the vasodilatory effect of Ang-(1–7) and bradykinin. The role of ACE2-Ang–(1–7)-Mas axis in the modulation of vascular tone by these compounds seems probable.     

Higher serum bone alkaline phosphatase as a predictor of mortality in male hemodialysis patients

AimsHigher serum alkaline phosphatase predicts lower mortality in chronic kidney disease and hemodialysis patients without liver dysfunction because it reflects high bone turnover. The purpose of our study was to compare the significance of serum bone alkaline phosphatase (BAP) with that of other bone markers in prediction of all-cause mortality(ACM) in male hemodialysis patients.Main methodsThe study was performed for 5 years. Serum BAP, intact osteocalcin (iOC), ß-CrossLaps (CTX), and intact parathyroid hormone (iPTH) were measured in 196 male hemodialysis patients without radiographic fracture. Their day-to-day variation during 5 consecutive days and diurnal variation were determined in 13 healthy males.Key findingsThe patients were divided into higher and lower groups based on serum levels of bone markers(mean ± SD: iPTH 218.6 ± 214.5 pg/ml, BAP 23.6 ± 12.2U/L, iOC 42.8 ± 45.2 ng/ml, CTX 1.71 ± 1.23 nmol/L BCE). In Kaplan–Meier analysis, the higher BAP group had significantly higher ACM than the lower BAP group (P = 0.013), whereas mortality did not differ between the higher and lower groups in other markers. Cox regression hazard analysis identified higher log BAP as a significant independent predictor [hazard ratio(HR) 8.32(95%CI:1.18–58.98)] for ACM after adjustment for various factors including pre-existing cardiovasucular disease, presence of DM. The significant association of mortality with serum BAP alone, in contrast with other markers including CTX [HR0.64 (95%CI:0.16–2.47)], iOC [HR0.97(95%CI:0.36–2.64)], iPTH [HR0.84(95%CI:0.44–1.60)],it may be due to the narrower day-to-day variation and the absence of diurnal variation in serum BAP compared to other markers.SignificanceHigher serum BAP may be a predictor of ACM in male hemodialysis patients.     

CDP-choline-induced contractions in the mouse gastric fundus through purinoceptors and Rho/Rho-kinase signalling

AimsThis study aimed to investigate the effects of cytidine-5′-diphosphocholine (CDP-choline), an endogenous lipid precursor, on the reactivity of the mouse gastric fundus and to determine the mechanism(s) mediating its effects.Main methodsPossible contractile effect of CDP-choline (10^? 5–10^? 2 M) was investigated in the absence and presence of a muscarinic receptor antagonist, atropine (3 × 10^? 6 M), an acetylcholine esterase inhibitor, physostigmine (10^? 6 M), a Na⁺ channel blocker, tetrodotoxin (TTX, 3 × 10^? 6 M), a Rho-kinase inhibitor, Y-27632 (10^? 5 M), a purinoceptor antagonist, suramin (2 × 10^? 4 M), a nitric oxide synthase inhibitor, N^G-nitro-L-arginine (L-NA, 3 × 10^? 4 M), a Ca²⁺ channel blocker, nifedipine (10^? 6 M), an α₇ nicotinic receptor antagonist, methyllycaconitine citrate (MLA, 10^? 6 M) and a G protein (G_i/o) inhibitor, pertussis toxin (PTX, 2 μg/ml). The metabolites of CDP-choline, namely choline (10^? 4–10^? 2 M), cytidine 5′-triphosphate (CTP, 10^? 5–10^? 2 M), cytidine (10^? 5–10^? 2 M) and cytidine monophosphate (CMP, 10^? 3–10^? 2 M) were also tested. Besides, phosphorylation of MYPT1, which indicates Rho-kinase activity, was also detected.Key findingsCDP-choline produced contractions in a concentration-dependent manner. The contractions were not affected by atropine, physostigmine, TTX, PTX, MLA or L-NA. However, Y-27632, suramin or nifedipine partly reduced these contractions. CDP-choline increased phosphorylation of MYPT1. Among CDP-choline metabolites, cytidine had no contractile effects. However, choline induced considerable contractions, which were sensitive to atropine. CMP and CTP had also contractile activity, comparable to that of CDP-choline.SignificanceThese results suggest that CDP-choline produced contraction through, at least in part, purinoceptors and Rho/Rho-kinase signalling in the mouse gastric fundus.     

Characterization of anticancer properties of 2,6-diisopropylphenol–docosahexaenoate and analogues in breast cancer cells

The present study describes the characterization and evaluation of novel anticancer conjugates, 2,6-diisopropylphenol–docosahexaenoate (PP–DHA), and its analogues including 2,4-diisopropylphenol–docosahexaenoate (DIPP–DHA), 2-isopropylphenol–docosahexaenoate (IPP–DHA), 2-cyclohexanephenol-docosahexaenoate (CHP–DHA) and phenol–docosahexaenoate (P–DHA) on breast cancer cell lines. Representative breast cancer cell lines, based on estrogen α receptor (ER) and oncogene Her-2 expression, were used and include MDA-MB-231 (ER-negative, Her-2-negative), MCF-7 (ER-positive, Her-2-negative) AU565 (ER-negative, Her-2-positive) and MDA-MB-361 (ER-positive, Her-2-positive). The PP–DHA conjugate significantly inhibited cell growth and induced cell loss in the breast cancer cell lines similarly; however, this conjugate was not effective against normal mammary epithelial cells. The effect of various conjugates were in PP–DHA > IPP–DHA > DIPP–DHA > CHP–DHA >> P–DHA order. PP–DHA and IPP–DHA conjugates were stable in human and mouse serum. Furthermore, the non-hydrolyzable amide-linked conjugate analogues affected breast cancer cells in a manner similar to that of the ester-linked conjugates. This suggests that ester-linked PP–DHA and IPP–DHA conjugates were stable during treatment to breast cancer cells due to structural hindrance. PP–DHA did not affect PPARα or PPARγ activities but its anticancer effects appear to be mediated in part though the inhibition of histone deacetylase (HDAC) activity. Further experiments are needed to confirm their molecular target and to test the effectiveness of these compounds in an in vivo model for their anticancer properties. In conclusion, these results suggest that the novel PP–DHA and IPP–DHA conjugates and their amide derivatives may be useful for the treatment of breast cancer.     

Aclidinium bromide, a novel long-acting muscarinic antagonist for COPD with improved preclinical renal and urinary safety profile

AimsAclidinium bromide is a novel, long-acting, inhaled muscarinic antagonist currently in registration phase for the treatment of chronic obstructive pulmonary disease. Since urinary difficulty and retention have been reported for anticholinergic agents such as tiotropium and ipratropium, it is important to examine the preclinical urinary and renal safety profile of aclidinium.Main methodsThe effect of aclidinium on urine and electrolyte excretion, renal function and voiding cystometry was analysed in conscious water-loaded Wistar rats (10–1000 μg/kg, s.c.), anaesthetised Beagle dogs (1000 μg/kg, i.v.) and anaesthetised guinea pigs (3–100 μg/kg, intratracheally), respectively. Aclidinium plasma levels were determined in an independent study. Active comparators were tiotropium (all studies) and ipratropium (cystometry only).Key findingsAclidinium 1000 μg/kg had no effect on urine excretion in rats, in contrast to tiotropium 100 μg/kg which significantly decreased this parameter (p < 0.05). Aclidinium 1000 μg/kg also had no effect on renal function in Beagle dogs. In guinea pigs, aclidinium 3–100 μg/kg had no effect on urinary bladder function, whereas tiotropium and ipratropium 100 μg/kg decreased the peak micturition pressure (p < 0.05), increased the volume of urine retained in the bladder (p < 0.01) and showed a trend to decrease the volume of urine excreted.SignificanceAclidinium had no significant effect on urinary and renal function in the animal models studied. These results, together with the rapid plasma clearance of aclidinium reported previously, suggest a lower propensity to induce urinary retention in humans than tiotropium and ipratropium.     

Field evaluation of the MM3-SERO ELISA for detection of anti-Fasciola IgG antibodies in milk samples from individual cows and bulk milk tanks

We carried out a field evaluation of the MM3-SERO ELISA for the diagnosis of Fasciola hepatica infection, by analysing serum and milk samples from individual cows and samples from bulk milk tanks. The diagnostic performance of the assay was assessed with serum samples from all 257 cows in eight fluke-free herds, and 240 cows with natural fasciolosis (diagnosed in vivo and/or post-mortem). Assay performance for individual milk samples was determined by analysis of paired serum and milk samples from 947 lactating cows from 33 F. hepatica-infected farms. The diagnostic usefulness of the assay for bulk tank milk was evaluated by analysis of bulk milk from infected (33) and non-infected (35) farms. For serum samples, the sensitivity, specificity and diagnostic accuracy of the assay were respectively 99.2% (95% CI: 97.0%–99.9%), 100% (95% CI: 98.6%–100%) and 0.997 (95% CI: 0.987–1.000). The only two infected animals in which serum antibodies were not detected had very low parasitic burdens (with only 2 and 3 flukes observed). The performance of the MM3 SERO ELISA for individual milk samples was similar to that for serum samples, and the stepwise linear regression revealed a strong correlation between the results for the milk samples and the serum samples (R² = 0.84; p < 0.001). The agreement between results obtained with pairs of serum and milk samples was very high: there was matching classification in 96% (910/947) of paired samples (kappa = 0.92; p < 0.001). Individual milk samples may therefore be used, instead of serum samples, in the MM3-SERO ELISA, for reliable detection of seropositive cows. Testing bulk tank milk samples enabled detection of infected herds, even when the within-herd prevalence of infection was as low as 12%. We conclude that the MM3-SERO ELISA is a sensitive and highly specific test for serodiagnosis of bovine fasciolosis, and can be used with individual samples of either serum or milk. Use of the assay with bulk milk samples enables estimation of the within-herd prevalence of infection.