Photoperiod Affects Leptin Action on the Choroid Plexus in Ewes Challenged with Lipopolysaccharide-Study on the mRNA Level.

The ovine choroid plexus (ChP) expresses the long isoform of the leptin receptor, which makes this structure a potential target for leptin action. In sheep, leptin concentration in plasma is higher during long days (LD) than short days (SD). This study evaluates the influence a of photoperiod on leptin impact on the gene expression of Toll-like receptor 4 (TLR4), proinflammatory cytokines (IL1B, IL6), their receptors (IL1R1, IL1R2, ILRN, IL6R, IL6ST) and inflammasome components necessary for pro-IL-1ß activation (NLRP3, PYCARD, CASP1), chemokine (CCL2), leptin receptor isoforms (LEPRa, LEPRb) and a suppressor of cytokine signalling (SOCS3) in the ChP of ewes treated or not with lipopolysaccharide (LPS). Studies were conducted on adult female sheep divided into four groups (n = 6 in each): control, leptin (20 µg/kg), LPS (400 ng/kg), and LPS and leptin injected under SD and LD photoperiods. The leptin alone did not affect the gene expression but in co-treatment with LPS increased (p < 0.05) IL1B but only during SD, and SOCS3, IL1R2, IL1RN, IL6ST and CCL2 only during LD, and decreased (p < 0.05) the IL1R1 expression only during SD photoperiod. This indicates that the immunomodulatory action of leptin on the ChP is manifested only under the LPS challenge and is photoperiodically dependent.

Neostigmine Attenuates Proinflammatory Cytokine Expression in Preoptic Area but Not Choroid Plexus during Lipopolysaccharide-Induced Systemic Inflammation.

The study was designed to examine whether the administration of neostigmine (0.5 mg/animal), a peripheral inhibitor of acetylcholinesterase (AChE), during an immune/inflammatory challenge provoked by intravenous injection of bacterial endotoxin-lipopolysaccharide (LPS; 400 ng/kg)-attenuates the synthesis of proinflammatory cytokines in the ovine preoptic area (POA), the hypothalamic structure playing an essential role in the control of the reproduction process, and in the choroid plexus (CP), a multifunctional organ sited at the interface between the blood and cerebrospinal fluid in the ewe. Neostigmine suppressed (p < 0.05) LPS-stimulated synthesis of cytokines such as interleukin- (IL-) 1ß, IL-6, and tumor necrosis factor (TNF) α in the POA, and this effect was similar to that induced by the treatment with systemic AChE inhibitor-donepezil (2.5 mg/animal). On the other hand, both AChE inhibitors did not influence the gene expression of these cytokines and their corresponding receptors in the CP. It was found that this structure seems to not express the neuronal acetylcholine (ACh) receptor subunit alpha-7, required for anti-inflammatory action of ACh. The mechanism of action involves inhibition of the proinflammatory cytokine synthesis on the periphery as well as inhibition of their de novo synthesis rather in brain microvessels and not in the CP. In conclusion, it is suggested that the AChE inhibitors incapable of reaching brain parenchyma might be used in the treatment of neuroinflammatory processes induced by peripheral inflammation.

Plasma and cerebrospinal fluid interleukin-1ß during lipopolysaccharide-induced systemic inflammation in ewes implanted or not with slow-release melatonin.

BACKGROUND: Interleukin-1ß (IL-1ß) is important mediator of inflammatory-induced suppression of reproductive axis at the hypothalamic level. At the beginning of inflammation, the main source of cytokines in the cerebrospinal fluid (CSF) is peripheral circulation, while over time, cytokines produced in the brain are more important. Melatonin has been shown to decrease pro-inflammatory cytokines concentration in the brain. In ewes, melatonin is used to advance the onset of a breading season. Little is known about CSF concentration of IL-1ß in ewes and its correlation with plasma during inflammation as well as melatonin action on the concentration of IL-1ß in blood plasma and the CSF, and brain barriers permeability in early stage of lipopolysaccharide (LPS)-induced inflammation. METHODS: Systemic inflammation was induced through LPS administration in melatonin- and sham-implanted ewes. Blood and CSF samples were collected before and after LPS administration and IL-1ß and albumin concentration were measured. To assess the functions of brain barriers albumin quotient (QAlb) was used. Expression of IL-1ß (Il1B) and its receptor type I (Il1r1) and type II (Il1r2) and matrix metalloproteinase (Mmp) 3 and 9 was evaluated in the choroid plexus (CP). RESULTS: Before LPS administration, IL-1ß was on the level of 62.0 ± 29.7 pg/mL and 66.4 ± 32.1 pg/mL in plasma and 26.2 ± 5.4 pg/mL and 21.3 ± 8.7 pg/mL in the CSF in sham- and melatonin-implanted group, respectively. Following LPS it increased to 159.3 ± 53.1 pg/mL and 197.8 ± 42.8 pg/mL in plasma and 129.8 ± 54.2 pg/mL and 139.6 ± 51.5 pg/mL in the CSF. No correlations was found between plasma and CSF IL-1ß concentration after LPS in both groups. The QAlb calculated before LPS and 6 h after was similar in all groups. Melatonin did not affected mRNA expression of Il1B, Il1r1 and Il1r2 in the CP. The mRNA expression of Mmp3 and Mmp9 was not detected. CONCLUSIONS: The lack of correlation between plasma and CSF IL-1ß concentration indicates that at the beginning of inflammation the local synthesis of IL-1ß in the CP is an important source of IL-1ß in the CSF. Melatonin from slow-release implants does not affect IL-1ß concentration in plasma and CSF in early stage of systemic inflammation.

Profile of toll-like receptor mRNA expression in the choroid plexus in adult ewes.

The blood-cerebrospinal fluid barrier (BCSFB) located in the epithelial cells of the choroid plexus (CP) forms the interface between the cerebrospinal fluid (CSF) and pathogen components circulating in the blood. The CP is also implicated in the passage of peripheral immune signals and circulation of immune cells into the central nervous system. Toll-like receptors (TLRs) are patternrecognition receptors that play a crucial role in the recognition of pathogens and triggering of the innate immune response. In sheep, ten members of the TLR family have been identified and cloned. We used real-time PCR analyses to examine the profiles of TLR mRNA expression in the CP of cerebral ventricles in healthy adult ewes. The transcripts for all ten TLRs except TLR8 were present; however, we observed a high variation in the degree of expression of the TLR5 and TLR1 genes (coefficient of variation: 61% and 46%, respectively) as well as a moderate variation in the expression of the TLR4 (34%), TLR2 (27%) and TLR6 (26%) genes. The TLR9, TLR7, TLR3 and TLR10 genes were the four receptors with relatively invariable expression levels (coefficient of variation: 7%, 8%, 16% and 17%, respectively) across the six adult ewes. The concentration of cortisol in blood collected prior to sacrificing the ewes ranged from 0.18 to 78.9 ng/ml. There was no correlation between cortisol concentration and mRNA expression of any of the examined TLRs. These data suggest that the CP has the potential to sense the presence of many bacterial and viral components and mediate responses for the elimination of invading microorganisms, thereby protecting the brain.

[Familial occurrence of granulocyte disorders in two children with recurrent otitis media]. / Rodzinne wystepowanie zaburzen funkcji granulocyta u dwojga dzieci z nawracajacym zapaleniem uszu.

UNLABELLED: Releasing of oxygen reactive species in activated granulocytes play an important role in the intracellular killing of bacteria. The respiratory burst is accompanied by chemiluminescence (chl). In vitro, granulocyte chl can be stimulated by nFLMP (bacterial peptide) or opsonised zymosan (OZ) after their binding to chemotactic surface receptors or FcR and CR, respectively. Chl can be induced also by non receptor way by PMA. We have found that blood granulocytes taken from children with recurrent upper respiratory tract infections demonstrated significantly decreased ability to oxidative metabolism after stimulation by n-FMLP. Inability of granulocytes to respond to only one of three above mentioned stimulators can suggests deficit of this enzyme isoform which is necessary for intracellular signal transduction. THE AIM of this study was to assess the activity of peripheral blood granulocytes in two children from two families, suffering from recurrent otitis media. MATERIALS AND METHODS: Study was performed in the 12 month old boy and 13 month old sick girl and in the children's parents. Chemiluminescence was measured in three blood samples, before and after n-FMLP, OZ and PMA stimulation. The light emission was measured in scintillation counter in presence of luminol. The study was performed after 2-3 weeks after treatment of otitis media. RESULTS: Both children had reduced response with FMLP and PMA and additionally, the girl had decreased response after OZ. Both boy's parents had decreased response only with n-FMLP. The diseased girl's father demonstrated a lack of response after all three stimulators. CONCLUSION: 1. The study of chemiluminescence stimulated with the three known stimulators increased the chance of recognising disturbances in granulocyte function. 2. Recognising wrong function child's granulocytes should be an indication for parents' study.