•SLC11A1 gene of goat activity variation is associated with specific nature of the stimulus.•The A16-B8 genotype is the most prominent, with the strongest transcriptional effect of 3′UTR.•The A16-B8 ...genotype was found to be more responsive to MAP exposure.•Within the genomic region of SLC11A1 gene were detected different functional regulatory elements.•85 putative miRNAs were detected to be localized externally or between polymorphic regions A and B.
The study aimed to investigate whether the genetic polymorphisms in the 3′UTR of the caprine SLC11A1 gene are functional, and to assess the role of MAP as a regulatory parameter in gene expression. To this goal we constructed plasmids expressing the Luciferase reporter gene in transient transfections of a mouse (Balb/c) macrophage cell line (RAW264.7), incorporating those polymorphisms that our previous work indicated as more prominent in terms of SLC11A1 expression and responsiveness to MAP infection. Gene expression variation was recorded on the average of the respective measurements after exposure to Mycobacterium avium subsp. paratuberculosis (MAP) combined with microbial antigens and cytokines. In silico analysis of the region under study allowed identification of one cis-acting RNA element, five putative transcriptional regulatory elements and 85 3’end microRNA binding sites. The two polymorphic regions (regions A and B) of the 3’UTR of the caprine SLC11A1 gene were recognized as regulators of its activity, at transcriptional and post-transcriptional level. The GT16 polymorphism at region A, combined with the GT8 polymorphism at region B, results in up-regulation of the SLC11A1 gene. The specific genotype was also found to be more responsive to MAP exposure at a statistically significant level.
Summary
This study investigated the effects of dietary inclusion of soya bean oil combined with fish oil (SFO) on the activities of a) superoxide dismutase (SOD), glutathione reductase (GR), catalase ...(CAT) and glutathione transferase (GST) in blood plasma and b) SOD, GR, CAT and lactoperoxidase (LPO) in the milk of sheep and goats. Furthermore, the oxidative stress indicators for measuring total antioxidant activity and free radical scavenging activity ferric reducing ability of plasma (FRAP) and 2,2′‐azino‐bis(3‐ethylbenzothiazoline‐6‐sulphonic acid) (ABTS) assays and oxidative stress biomarkers malondialdehyde (MDA) and protein carbonyl (PC) were also determined in the blood plasma and milk of the animals. For this purpose, twelve dairy sheep and twelve dairy goats were assigned each to two homogenous subgroups. Treatments in both animal species involved a control diet without added oil and a diet supplemented with 5% soya bean oil and 1% fish oil. The results showed that the inclusion of SFO in the diets of sheep and goats increased significantly the activities of CAT and GR in their blood plasma. The same effect was observed for the activities of GST and FRAP in the blood plasma of goats. Moreover, the fact that the goats had significantly higher average daily PUFA intake (3.62 g/kg BW0.75) compared to sheep (2.51 g/kg BW0.75) resulted in an enhancement in the MDA content in their plasma. A significant increase in CAT activity in the milk in both animal species fed with SFO diets was also found. Finally, due to the higher apparent transfer rate of n‐3 FA from the diet to the milk in sheep, the PC concentrations were found to be enhanced in their plasma and milk. In conclusion, the impact of dietary SFO supplementation on the oxidative status of body and/or on the milk of small ruminants depends not only on the daily PUFA intake, but also on the amount of n‐3 FA that reach their milk.
TMS studies on the CNS effects of benzodiazepines have provided contradictory results. The objective of this study is to describe the effects of lorazepam on silent period (SP) and corticomotor ...excitability. Twelve healthy male subjects (median age 35 years) were studied at baseline, following i.v. lorazepam administration and after reversal of the benzodiazepine effects with i.v. flumazenil. Lorazepam was given at a low-dose in one subject (0.0225 mg/kg bolus + 2 microg/kg/h infusion) and at a high-dose (0.045 mg/kg bolus + 2.6 microg/kg/h infusion) in the rest. Threshold (Thr) was measured at 1% steps. SPs were investigated with two complementary methods. First, SPs were elicited using a wide range of stimulus intensities (SIs) (from 5 to 100% maximum SI at 5% increments). At each SI, four SPs were obtained and the average value of SP duration was used to construct a stimulus/response (S/R) curve of SI versus SP .The resulting S/R curves were then fitted to a Boltzman function, the best-fit values of which were statistically compared for each experimental condition (i.e., baseline vs. lorazepam vs. flumazenil). Second, a large number of SPs (n=100) was elicited during each of the three experimental conditions using blocks of four stimuli with an intensity alternating between MT and 200% MT. This method was employed so as to reveal the dynamic, time-varying effects of lorazepam and flumazenil on SP duration at two stimulus intensity (SI) levels. MEP recruitment curves were constructed at rest and during activation and fitted to a Boltzman function the best-fit values of which were statistically compared for each experimental condition. Lorazepam at a low dose did not affect Thr, SP, or the active MEP recruitment curves. The high dose also had no effect on Thr and the active MEPs whereas the resting MEP recruitment curves were depressed post-lorazepam at the higher range of stimulus intensities. With regard to SP, the Max value of the S/R curve decreased from 251+/-4.6 ms at baseline to 215.2+/-3.1 ms post-lorazepam (P<0.01). V50 also decreased significantly (from 47.92+/-0.9% to 43.73+/-0.81%, P<0.01) whereas there was no significant change regarding slope and SP Thr. The statistical analysis of the SP S/R curves as well as the study of SPs at two SI levels revealed that lorazepam reduced SP duration when high intensity stimuli were used (>60%). In contrast, at low SIs a small increase in SP duration was noted post-drug. Enhancement of GABAergic inhibition by lorazepam results in a reduction of SP duration when high SIs is used. At the lower range of SIs, a small but statistically significant increase in SP duration is observed. The kinetic behavior of this phenomenon as well as the possible underlying mechanisms are discussed.
•In vitro investigation of the impact of the polymorphisms of the 3′ UTR of the SLC11A1 gene to its expression.•The infection induced by the M. avium ssp. paratuberculosis to macrophages causes ...upregulation of the SLC11A1 gene.•The data presented here demonstrate that the homozygous B7 genotype leads to up-regulation of the SLC11A1 and IL-1α genes.•B7/7 genotype indicates increased responsiveness of macrophages to MAP exposure compared to the B8/8 and B7/B8 genotypes.
Johne’s disease or paratuberculosis is a chronic, progressive intestinal disease of ruminants caused by Mycobacterium avium subsp. paratuberculosis (MAP). One of the genes that have been targeted with regard to resistance or sensitivity to paratuberculosis is the SLC11A1 (solute carrier family 11 member A1). Here we extend our previous work to the sequence and structure analysis of the caprine SLC11A1 gene and we assess the functional impact of the most frequent polymorphisms of the 3′ UTR region of the SLC11A1 gene to its expression in goat macrophages exposed in vitro to MAP. The role of these polymorphisms in primary immune response is also investigated with connection to gene expression of two interleukins (IL), one of which pro (IL-1a), and the other anti-inflammatory (IL-10). In order to assess gene response, quantitative detection of the SLC11A1, IL-10 and IL1a mRNA was performed by real time PCR before, and at 1, 3 and 24h after exposure of primary cultures of peripheral blood monocyte-derived macrophages to MAP, collected from 54 goats of the Greek native goat breed. Sequence analysis of the 3′ UTR end of the caprine SLC11A1 gene determined its full length to be 522 bases. Structure analysis confirmed the presence of two microsatellites consisted of a variable number of guanine-thymine repeats (regions A and B). The homozygous B7 genotype B(GTn)7/7 was associated at a statistically significant level with increased expression of the SLC11A1 and IL-1α genes indicating increased in vitro responsiveness and therefore resistance of mononuclear derived macrophages to MAP infection.
Summary
Milk protein synthesis in the mammary gland involves expression of six major milk proteins' genes whose nutritional regulation remains poorly defined. In this study, the effect of long‐term ...under‐ and overfeeding on the expression of as1‐casein: CSN1S1, as2‐casein: CSN1S2, β‐casein: CSN2, κ‐casein: CSN3, α‐lactalbumin: LALBA and β‐lactoglobulin: BLG gene in goat mammary tissue (MT) was examined. Twenty‐four lactating dairy goat, at 90–98 days in milk, were divided into three homogenous subgroups and fed the same ration, for 60 days, in quantities which met 70% (underfeeding), 100% (control) and 130% (overfeeding) of their energy and crude protein requirements. The results showed a significant decrease in mRNA of CSN1S2, CSN2, CSN3 and LALBA genes in the MT of underfed goats compared with the overfed and on the CSN1S1 and BLG gene expressions in the MT of underfed goats compared with the respective control and overfed. CSN2 was the most abundant transcript in goat MT relative to the other milk proteins' genes. Significantly positive correlations were observed between the mRNA levels of caseins' and BLG genes with the milk yield. Moreover, a significant correlation was found between the mRNA levels of CSN1S2 with the milk protein, lactose content and lactose yield and also between the LALBA gene expression with the lactose content and lactose yield respectively. In conclusion, the feeding level and consequently the nutrients availability affected the milk lactose content, protein and lactose yield as well as the milk volume by altering the CSN1S1, CSN1S2, CSN2, CSN3, LALBA and BLG gene expression involved in their metabolic pathways.
Essential oils (EO) from aromatic plants (AP) are used in ruminants’ diets due to several biological activities. The inclusion of AP in ruminants’ diets can be done as dried material, as they are ...harvested once a year. However, storage is one of the main factors that affect the EO content and chemical composition. Thus, the objective of this study was to determine the effect of storage, in the shade under ambient conditions, on the EO content and chemical composition (a) of Origanum vulgare spp. hirtum as a whole plant (OP) and/or (b) as the powdered form after mixing with the concentrate diet. The results showed that storage of the whole OP in the shade for six months has a very low EO loss (5.8%), while grinding and mixing OP with concentrates causes different losses, which depend on the OP/concentrate ratio and the storage period.
The genes of urokinase plasminogen activator (u-PA) and its receptor (u-PAR) are part of the fibrinolytic system which plays an important role in white adipose tissue remodeling. The nucleotides ...sequences of these genes have not been characterized in goats, in contrast to other ruminants (sheep and cow) and were partly isolated in this study. Further to that, taking into account that gene expression is strongly affected by dietary and genetic factors, an experiment was conducted with lactating dairy ewes (n=12) and goats (n=12) with the objective to determine if there are differences between them in u-PA and u-PAR gene activity in their subcutaneous adipose tissue (SUBQ) under the same dietary treatments.
Ewes and goats were given identical diets in a mid-lactation period and another adapted diet in the late lactation period identical between ewes and goats. This late lactation contained less roughage and were fed in a lower amount, and caused – unintendedly – a negative energy balance since animals lost weight in that period.
The results showed that the mRNA expression of u-PAR reduced significantly in the SUBQ of both animal species with Treatment B compared with Treatment A. In addition, the u-PA and u-PAR mRNA expressions were significantly higher in the SUBQ of goats compared to those of sheep, independently from the fed diet and the different days in milk. In conclusion, may be feeding regimen affects the u-PAR mRNA expression while there are significant species differences between sheep and goats in the u-PA and u-PAR mRNA expression of their SUBQ under the same dietary treatments.
Abstract The aim of this study is to investigate the neuroprotective effects of the anticonvulsant topiramate in a new model of traumatic brain injury in rats. A new model of traumatic brain injury, ...based on the weight-drop technique, was developed for the purpose of this study. Seventy-five male Wistar rats weighing 320–470 g were studied. All rats were anesthetized, subsequently submitted to a round craniectomy in the left parietal region and a weight of 50 g was used for the production of a cortical contusion. In study I, 44 rats were randomized in three groups to receive either topiramate 40 mg/kg ( n =13), topiramate 60 mg/kg ( n =14), or water for injection ( n =17) i.p. 30 min after the injury and every 12 h thereafter for 3 days. The rats were tested clinically 24 h, 72 h, 10 days and 20 days after the injury. On day 21 the animals were sacrificed and the brains were removed and prepared for histopathological analysis. In study II, 19 rats were randomized to receive either topiramate 60 mg/kg ( n =10) or water for injection ( n =9) i.p. 30 min after the injury and every 12 h (four doses in total). 48 h after the injury the animals were sacrificed and the brains were rapidly removed and analyzed for water content with the dry-wet weight technique. The animals that received topiramate performed significantly better in neurological tests compared to the animals that received vehicle ten ( P <0.05) and 20 ( P <0.001) days after the injury. There was no difference between the high and the low dose of the drug. Topiramate had no effect on the anatomic volume of the lesion. The animals that received topiramate had a tendency to present with less cerebral edema formation, but the difference was not statistically significant ( P >0.05). These findings suggest that topiramate promotes neurological recovery in rats after traumatic brain injury without affecting the final size of the traumatic lesion and that it might play a role in the reduction of post-traumatic cerebral edema.
The Hemiconvulsions-Hemiplegia-Epilepsy (HHE) syndrome is currently regarded as an extremely rare condition. The etiological and pathophysiological mechanisms underlying this medical rarity as well ...as the optimal therapeutic approaches remain poorly defined and understood. We present the clinical, radiological and electroencephalography (EEG) findings of a patient with the HHE syndrome and describe the response of the continuously present epileptiform abnormalities to transcranial magnetic stimulation (TMS).
A 33-year old male patient was referred to our department for investigation and management of intractable epilepsy. His seizures began at the age of three months when, during the course of a common febrile illness, he developed repetitive clonic seizures involving the left upper and lower limbs, followed by permanent left hemiplegia. After extensive investigations, he was diagnosed with "idiopathic" HHE syndrome. Currently, he suffers from left hemiplegia, severe intellectual impairment Intelligence Quotient (IQ) <30 and asymmetric, bilateral tonic seizures occurring 1-3 times daily despite treatment with valproate, topiramate, lamotrigine, rufinamide, and perampanel. Brain magnetic resonance imaging revealed atrophy of the right hemisphere and serial EEGs disclosed continuous sharp waves, the generators of which were localized by electrical source imaging (ESI) to two distinct sources within the right hemisphere. Repetitive TMS 210 stimuli of 1 Hz at 100 % corticomotor threshold applied with a circular coil over the generators of epileptic discharges (EDs) resulted in a statistically significant decrease of ED counts compared to sham stimulation and the post-verum TMS period.
We present the clinical-laboratory profile and the long-term follow up of a patient with the HHE syndrome. Further, we describe the effects of TMS on EDs. The latter observation raises the possibility that TMS-EEG may be used in select cases with intractable epilepsy as a surrogate marker of responsiveness to more invasive modalities (i.e., cortical stimulation). HIPPOKRATIA 2017, 21(2): 101-104.
The effect of frozen storage and subsequent thawing under microwave heating on fresh bread quality was investigated. Two different flour types were used. Xanthan and guar gum were also added at ‘low’ ...and ‘high’ concentration (0.16 and 0.65
g/100
g flour, respectively). The specific volume, the porosity using a pycnometer, the crust firmness (puncture experiments) and the crumb viscoelastic behaviour (relaxation experiments) were evaluated. After thawing a significant shrinkage was observed, a porosity reduction in all samples, a major softening of the crust in control samples, a viscous crumb in most samples. A more rubbery crust especially in samples containing guar gum was also noticed, which can be ascribed to gums water binding capacity or to gums/water interactions and/or because gums can inhibit gluten-starch interactions influencing the plasticity of the bread. At high gum concentration destabilization effects were observed. Guar gum resulted generally in less desirable, final bread quality considering that it was not so efficient as concerning the maintenance of the gluten matrix after frozen storage, whereas xanthan addition gave better bread properties at ‘low’ concentration.
Differences in respect to the flour type used were mainly observed after microwave heating. Textural attributes could be correlated to structural characteristics of the crumb (porosity and specific volume), indicating their mutual dependence.
