Multiple epidemiological studies have shown that low testosterone levels are associated with and predict the future development of type 2 diabetes mellitus and the metabolic syndrome.The aim of our study was to show the influence of testosterone replacement therapy on obesity, HbA1c level, hypertension and dyslipidemia in patients with diabetes mellitus and androgen deficiency.2, were randomized in a placebo-controlled study. They also underwent a routine physical examination and selected by free testosterone examination. We divided patients into two groups: 1) treatment group, where we used diet, physical activity, patient’s antidiabetic therapy and testosterone replacement therapy; 2) placebo group, where we used diet, physical activity, patient’s antidiabetic therapy and placebo.One hundred and twenty-five male patients with diabetes mellitus were screened; 85 subjects aged 41 to 65 years, with BMI from 27.0 to 48.0 kg/mAfter 6 months of treatment we repeated the diagnostic assessments: lipid profile was improved in both groups but in first group it was clinically significant. Free testosterone level increased in all groups, but in group I was clinically significant. HbA1c decreased in both groups, but in group I we obtained the best result. Leptin level after treatment was approximately the same in both groups. Also, blood pressure was reduced in both groups but results were similar.Our study demonstrated that it is possible to break this metabolic vicious circle by raising testosterone levels in diabetic men with androgen deficiency. Re-instituting physiological levels of testosterone, as the study has shown, has an important role in reducing the prevalence of diabetic complications. Over the past few decades, obesity and diabetes mellitus have become a global health challenge. Between 1980 and 2004, the prevalence of obesity increased from 15 to 33% in the United States - a pattern mirrored across the world [et al. [Androgens play an important role not only in sex differentiation and development, but also in regulating the metabolism of glucose, proteins, lipids and some inflammatory factors, all of which might have a great influence on insulin sensitivity. It is well known that reduced levels of total testosterone (TT) in middle-aged or older men may contribute to abdominal obesity, increased insulin resistance (IR), and diabetes mellitus. At least six large prospective clinical trials have suggested that reduction of TT predicted the increasing incidence of T2DM ,6. Low cet al.  recentlyet al. .As it has been previously shown, patients with T2DM have a frequent occurrence of hypogonadotrophic hypogonadism as reflected in low plasma concentrations of testosterone and inappropriately low luteinizing hormone (LH) and follicle-stimulating hormone (FSH) . In viewP =0.006) and free testosterone (P =0.027) levels in men with ED, but there was no significant difference in total testosterone levels. The lower the serum testosterone appears, the greater the severity of ED is suggested [et al. [P =0.0001) after adjustment for age and body mass index (BMI). In addition, hypogonadism in men with T2DM was associated with decreased sexual desire, more symptoms of depression, and lower LH levels.In the European Male Aging Study database of 3,369 men between the ages of 40 and 79 years, 3 sexual symptoms  had a syndromal relationship with decreased testosterone levels . Moreoveuggested . Corona  [et al.  evaluateet al. suggests that a substantial component is mediated through its association with body fat, in particular abdominal visceral adipose tissue. This testosterone-fat relationship is bi-directional, as both weight loss and testosterone therapy increase testosterone levels, reduce fat mass, and decrease IR [Multiple epidemiological studies have shown that low testosterone levels are associated with and predict the future development of T2DM and the metabolic syndrome although this relationship is confounded by the association of total testosterone with SHBG, free testosterone remains associated with measures of IR and T2DM in some, but not all studies. Although the link between low testosterone levels and IR is not solely a consequence of adiposity, the study by Grossmann rease IR .et al. [et al. [P_ = 0.0001) [et al. [et al. [A meta-analysis by Ding et al.  analyzed [et al. ; total t 0.0001) . However [et al.  nor Coro [et al.  analyzedP _ 0.04) [P _ 0.001) [However, a cross-sectional study from the Third National Health and Nutrition Examination Survey (NHANES) group including 1,413 adult men, 101 of which had diabetes, showed that men in the lowest fertile of calculated free testosterone, adjusted for age, ethnicity, and adiposity, were more likely to have prevalent diabetes  . In addi_ 0.001) . Other r_ 0.001) , the Uni_ 0.001)  consiste_ 0.001) -26. In t_ 0.001) .The aim of study was to show the influence of testosterone replacement therapy (TRT) on obesity, HbA1c level, arterial hypertension and dyslipidemia in patients diabetes mellitus with androgen deficiency.2, patient age range 30 to 65 years, positive screening questionnaire for androgen deficiency in males [2 and >48 kg/m2, kidney and liver disease, adenoma of prostate grade II to III, secondary hypogonadism, treatment by testosterone or testosterone stimulation therapy 3 months before screening. Treatment with anti-obesity drugs within 3 months prior to informed consent, uncontrolled hyperglycemia with a glucose level >240 mg/dl (>13.3 mmol/l), any previous (or planned within next 12 months) bariatric surgery (open or laparoscopic) or intervention (gastric sleeve), current treatment with systemic corticosteroids at time of informed consent or pre-planned initiation of such therapy ) is not an exclusion criterion, as this does not cause systemic steroid action). Congestive heart failure of the New York Heart Association (NYHA) class III or IV, acute or chronic metabolic acidosis (present condition in patient history), hereditary galactose intolerance, alcohol or drug abuse within the 3 months prior to informed consent that would interfere with trial participation, acute coronary syndrome ≤6 weeks prior to informed consent, stroke or transient ischemic attack (TIA) ≤3 months prior to screening, change in dose of thyroid hormones and dyslipidemia therapy within 6 weeks before screening.One hundred and twenty-five male patients with diabetes mellitus were tested; subject enrollment was recorded for the period from 2010 until the end of 2013, at the LTD ‘National Institute of Endocrinology’ . The study was approved by the Ethics Committee of this institute. Written informed consent was obtained from all participants. Inclusion criteria were: T2DM, patients with BMI 27.0 to 48.0 kg/min males . Exclusi2) and was expressed in kg/m2. Waist circumference was measured midway between the iliac crest and the lower margin of the 12th rib. Also, for measurement of supine blood pressure subjects were in a supine or semi-recumbent position for a minimum of 5 minutes before the blood pressure measurement; after that blood pressure measurement was recorded to the nearest 2 mmHg mark on the manometer, which was conducted using a calibrated manometer; 2) biochemical measurements were made after a 10 to 12-hour fast: venous blood samples were obtained from 9:00 to 11:00 am and stored at 4°C; HbA1c, liver function tests, lipid profile, free testosterone, leptin, prostate specific antigen (PSA), thyroid stimulating hormone (TSH), FSH, LH; all hormone assays were performed by the fully automated ELISA analyzer Elisys Uno ; 3) validation of a screening questionnaire for androgen deficiency in males :1239–1242); 4) ultrasonography of the abdomen and prostate were performed with a Siemens Acuson Antara ; 5) electrocardiography (ECG).The following analyses were conducted: 1) anthropometric study: height and weight were evaluated in all subjects. BMI was calculated as weight (kg) divided by square of height  a treatment group; 2) a placebo group. In the first group we used diet, physical activity , patient’s antidiabetic therapy and TRT (testosterone undecanoate 250 mg/ml intra-muscularly once every 3 months). In the second group we used diet, physical activity , patient’s antidiabetic therapy and placebo.Eighty-five subjects, with age range 41 to 65 years and BMI from 27.0 to 48.0 kg/mt-test for normally distributed data and Mann–Whitney U-test for non-normally distributed variables. Pearson’s (r) correlation tests were used to determine the relationship between study parameters, depending on variable distribution. P-value <0.05 was considered statistically significant. Statistical analysis was performed using the SPSS 15.0 software package .Shapiro-Wilk test was used to determine normality of data distribution. The results were presented as mean ± standard deviation (SD) and 95% confidence interval (CI 95%) for normally distributed data. Non-normally distributed data were displayed as median with range . Differences between the study group and control group patients were measured using independent sample 2  (Table The patients were randomized in 2 1-sided blinded groups: 1) in the first group we had 42 patients  cholesterol levels decreased, and high-density lipoprotein (HDL) cholesterol increased in both groups but more clinically significant results were shown in the first group Table . Free teSerum testosterone, HbA1c, HDL cholesterol, triglyceride concentrations, BMI, and hypertension improved in both treatment groups after 26 weeks of treatment. We have shown that TRT improves IR and glycemic control in hypogonadal men with diabetes. TRT reduced the HOMA-IR index, indicating an improved fasting insulin sensitivity. This study demonstrates that intramuscular TRT in hypogonadal men with T2DM improves IR, the central biochemical defect associated with these conditions. Although an improvement in IR could be expected to result in better glycemic control, 6 months of treatment demonstrated reduction of HbA1c. Small-scale studies of testosterone treatment in men with metabolic syndrome or T2DM and marginal low or normal testosterone levels showed improvement in glycemic control. The low testosterone levels might have induced a vicious circle, since low testosterone levels induce an increase in visceral obesity and severity of cardiovascular risk factors compared with the metabolic syndrome. Studies have shown that hypogonadism is associated with dyslipidemia, but there are only limited data supporting the effect of testosterone on lipids. Two studies reported a small rise in HDL cholesterol but no effect on TC or LDL cholesterol in hypogonadal men .Furthermore, after TRT, total cholesterol, LDL cholesterol, triglyceride had reduced and HDL cholesterol increased by 16.53 mg/dl, 7.33 mg/dl, 7.29 mg/dl and 15.61 mg/dl respectively, which are consistent with other TRT trials’ results . Men witet al. [It has been shown that lower levels of testosterone in men are associated with higher blood pressure. The first study to demonstrate a favorable effect of testosterone treatment on blood pressure in abdominally obese men was published by Marin et al. . Anotheret al. . The maxFurthermore, leptin also may have some role in this recovery. Leptin is an adipose tissue hormone with so-called ‘double action’: it affects receptors on Leydig cells, lowering testicular testosterone secretion. In addition, leptin suppresses pituitary LH secretion. Hypogonadal diabetic men have increased leptin levels and, as it has been shown, TRT reduces leptin levels in addition to the independent effects of weight loss in lowering leptin levels. A limitation of this study is the relatively small samples of subjects involved that can limit the generalizability of the study.Our study demonstrated that it is possible to break the metabolic vicious circle discussed above by raising testosterone levels in diabetic men with androgen deficiency . In addi


Dense granules were apparent in A. fulgidus cells imaged by cryo electron microscopy (cryoEM) but not so by negative stain electron microscopy. Cryo electron tomography (cryoET) revealed that each cell contains one to several dense granules located near the cell membrane. Energy dispersive X-ray (EDX) spectroscopy and scanning transmission electron microscopy (STEM) show that, surprisingly, each cell contains not just one but often two types of granules with different elemental compositions. One type, named iron sulfide body (ISB), is composed mainly of the elements iron and sulfur plus copper; and the other one, called polyphosphate body (PPB), is composed of phosphorus and oxygen plus magnesium, calcium, and aluminum. PPBs are likely used for energy storage and/or metal sequestration/detoxification. ISBs could result from the reduction of sulfate to sulfide via anaerobic energy harvesting pathways and may be associated with energy and/or metal storage or detoxification. The exceptional ability of these archaeal cells to sequester different elements may have novel bioengineering applications.Inorganic storage granules have long been recognized in bacterial and eukaryotic cells but were only recently identified in archaeal cells. Here, we report the cellular organization and chemical compositions of storage granules in the Euryarchaeon Archaeoglobus fulgidus strain VC16 is a hyperthermophilic, sulfur oxide-reducing, anaerobic archaeon. Belonging to the Archaeoglobales division of the Euryarchaeota, the species is commonly found in marine thermal vents, hot springs, and thermophilic oil field waters. The production of thiosulfate as well as hydrogen sulfide has been implicated in oil and gas souring and in oil pipeline corrosion ). These would occur in the reduction of sulfate (SO42−) to hydrogen sulfide (H2S) during the crucial anaerobic respiration process needed for cell energy production and survival. Conceivably, ferrous thiosulfate or Fe-polysulfide would be formed as intermediates of the sulfate reduction pathway to serve as energy conserving storage materials. The ISB would subsequently be reduced to sulfide (HS−) for energy harvesting in downstream pathway reactions. The biological formation of thiosulfate from sulfite has been reported in several sulfate-reducing bacteria including Desulfovibrio vulgaris, Desulfovibrio desulfuricans, and Thermodesulfobacterium commune, whereby the enzyme systems responsible are not well described [Area scan spectra were obtained through dot mapping by scanning specific areas of additional cells which included one or more inclusion bodies: some of these area scans showed high concentrations of iron and sulfur, not phosphorus and oxygen, within the bodies Figures . Examiniescribed –16. Thesescribed –19. A. fulgidus cells two granules were discovered to be present . The absence of phosphate in culture media caused cells to appear further shrunken . After treating for three generations under the altered conditions, having no phosphate in solution did not allow cells to proliferate . Though possible artifacts associated with negative stain TEM complicate these observations, cryoEM images of cells with reduced phosphate show a clear reduction in size compared to normal concentrations.We further showed that the ability of A. fulgidus VC16 cell located near cell membranes. Notably, the PPBs lack the iron and sulfur elements abundant in the ISBs, while the ISBs lack the phosphate and oxygen abundant in the PPBs. Likewise, the less abundant elements present in the PPBs  are low to absent in the Fe-S bodies and, conversely, the PPBs lack iron. The ratios of the predominant elements seen in PPBs and ISBs are diagnostic: the former exhibits a characteristic oxygen, phosphorus, iron, sulfur ratio of 2 : 1 : 0 : 0, while the latter has an elemental ratio of 0 : 0 : 1 : 2. This elemental analysis also provides a potential approach to high throughput STEM assisted cell screening to characterize inorganic granules in A. fulgidus and in other archaeal and bacterial strains. Further analyses of the variations in granule density and at different stages of development are also now possible.Intracellular granules have been characterized in bacteria, eukaryotes, and archaea utilizing both energy-filtered TEM and energy dispersive X-ray spectroscopy in order to analyze the elemental composition of the inclusion bodies , 20–22.  A. fulgidus strain VC16 are positioned in characteristic membrane-adjacent locations  within Figures –4. These A. fulgidus were mentioned above based on prior PPB studies in the bacteria and eukaryotes over the past thirty years [Potential roles of the PPB granules inty years –19, 25.  A. fulgidus are unknown. Besides potential roles in iron or sulfur-based energy storage, ISB granules may also have roles in metal sequestration and/or detoxification by analogy to the PPBs. A. fulgidus species thrive in highly reducing and metal rich environments. Fluids flowing from hydrothermal vents, for example, from black and white smoker vents, are reported to contain dissolved calcium, copper, zinc, iron, manganese, and strontium in the low to high micromolar range [The role(s) of the newly described ISB granules inar range . Nearby  A. fulgidus cells metabolize sulfate and sulfite as well as thiosulfate [As shown in Figures osulfate , and the A. fulgidus VC16 cryoEM measurements of cell envelope and granule dimensions we can accurately document the cell compartment volumes and surface areas. A spherical cell of one micron in diameter would have an overall cell volume of 0.524 μm3 πr3). Using the following measures of the cell membrane cross section (~37 angstroms thick), the S-layer lattice (~110 angstroms thick), and the periplasmic-like space (~130 angstroms thick) which is sandwiched between and separates the two structures ( A. fulgidus volume is partitioned into 84.3% cytoplasm (0.441 μm3), 2.1% cell membrane (0.011 μm3), 7.2% cell periplasm (0.038 μm3), and 6.4% cell S-layer (0.034 μm3). The intracellular PPB and ISB granules observed in A. fulgidus cells can individually compose up to 1.4% of the cytoplasmic space (~230 nm diameter). Assuming the PPB granule density reported by Toso et al. [ A. fulgidus PPB would store several-hundred-fold more energy in the form of phosphoanhydride bond energy than contained in the cellular ATP pool.From theructures , the A.  E. coli cell envelope ) the analogous A. fulgidus envelope dimensions are remarkably similar (~28 nm thick) [ E. coli OM) and may contribute to cell rigidity and shape [ E. coli periplasmic spaces also differ by about 25% (~13 nm in A. fulgidus versus ~16 nm in E. coli). They provide analogous roles while having remarkably different cell architectures and molecular compositions.Compared to the cross section of a typicalm thick) . Here, tsimilar ~ nm thick Methanospirillum hungatei JF1 [ A. fulgidus granules. The M. hungatei PPBs bodies also differed from A. fulgidus PPBs reported in this study, whereby the M. hungatei bodies contain iron plus calcium rather than magnesium, aluminum, copper, and calcium .Few examples of inorganic storage granules are reported in archaea in contrast to PPB granules in bacteria and eukaryotes –19. We ratei JF1  which co calcium . These d Sulfolobus and Methanosarcina [ Ectothiorhodospira species [ Thiobacillus ferrooxidans, “sulfur globules” were reported to contain an inner core of S7–12 polysulfur plus an outer layer of S19+ polythionates [ A. fulgidus VC16 ISBs is currently unknown. Inspection of negatively stained cell sections of A. fulgidus strain 7324, which is related to A. fulgidus VC16, reveals the presence of electron dense bodies nearby the cell membranes , whhionates . The mol Archaeoglobus fulgidus VC16 are demonstrated for the first time. Each is composed of distinct primary and secondary metals and is likely involved in nutrient and/or energy storage. The presence of polyphosphate bodies in the archaea along with bacteria and Eukarya suggests an ancient origin of these structures. Future studies are needed to explore the biogenesis and physiological uses of these inclusion bodies.The occurrence, location, size, and compositions of two types of intracellular bodies in the thermophilic archaeanA. fulgidus cell ghosts reveal “scars” along cell membrane. Arrows indicate the location of “scars” found in A. fulgidus cell ghosts.Supplementary Figure S1: Negative stained A. fulgidus cells in exponential growth phase showing newly developed granules. A,B) Slices from 3D tomograms reconstructed from cryoET tilt series of A. fulgidus whole cells in growth phase. The insets show the granules present within each cell at higher magnification.Supplementary Figure S2: A. fulgidus development under varying nutritional conditions. A-E) TEM images of (A) a cell grown in normal conditions, (B) a cell grown with excess phosphate, (C) a cell grown with reduced phosphate, (D) a cell grown with reduced lactate, and (E) a cell grown with no phosphate. All samples were negatively stained with 1% UA.Supplementary Figure S3: A. fulgidus cells after three generations of reduced or elevated phosphate conditions. A) Low magnification image of a TEM grid, stained with 1% UA, of a sample grown with no phosphate. Almost no cells are present. B-E) CryoEM images of cells grown in (B) 1 mM phosphate, (C) 2 mM phosphate, (D) 4 mM phosphate , and (E) 9 mM phosphate.Supplementary Figure S4: CryoEM of


Evidence supporting treatment intensification in rheumatoid arthritis (RA) is limited and controversial. We explored outcomes of infliximab dose increases and accounted for pre-existing trough levels in patients with active RA.This study was a retrospective study of 42 RA patients who received increased infliximab following an insufficient response (DAS28 >3.2). Serum concentrations of infliximab and antibodies to infliximab (ATI) and DAS28 and EULAR clinical response parameters were recorded for 1 year. Analyses were performed in three patient groups that were defined by infliximab serum concentration prior to treatment enhancement: no detectable, low (<1.1 μg/mL) or high (≥1.1 μg/mL) drug levels.p = 0.016), but no increase was achieved in the low and no detectable groups. The three study groups exhibited similar disease activity over time, and no improvement was observed in the non-responder EULAR rates.No circulating infliximab was detected in 20 patients (47.6%), but 13 (31%) and 9 (21.4%) patients exhibited low and high levels, respectively. ATI was only detected in patients with no detectable drug levels because the drug interferes with ELISA. DAS28 disease activity globally showed a modest improvement after dose escalation, but this improvement did not persist after 6 and 12 months. Infliximab serum levels increased significantly in the high group (These results suggest that the efficacy of an infliximab dose increase is limited, and the response is independent of the infliximab trough serum concentration that is achieved prior to escalation. The response to tumor necrosis factor (TNF) inhibitor treatment in chronic inflammatory diseases exhibits great therapeutic variability. Failure to respond to anti-TNF therapy may occur at treatment onset, or it may be secondary to an initial improvement , 2. A coEvidence supporting dose intensification in rheumatoid arthritis (RA) is limited and controversial, and few studies relate dose intensification with pre-increase serum trough drug levels –12.The present study retrospectively analyzed the effect of Ifx dose increase in RA non-responders and accounted for previous drug concentrations to further elucidate the utility of proposed algorithms for patients who do not respond to the first anti-TNF therapy.A retrospective observational study was conducted of RA patients included in the Hospital Universitario La Paz Biologics Registry  who started treatment with Ifx as the first TNF-blocking agent. The following inclusion criteria were used: RA patients (older than 18 years) treated from 2005 to 2011 who exhibited an insufficient clinical response defined as DAS28 >3.2, who received an increase in the dose of Ifx, and who had serum sample and clinical assessment data during the first year of treatment. The period of inclusion ended in 2011 because no increase in Ifx dose was used in our clinic after this time. A final observation was carried forward for analysis in patients who stopped Ifx treatment within the first year after dose increase.Ifx treatment could be combined with classic disease-modifying anti-rheumatic drugs (DMARDs) and/or corticosteroids. The research ethics committee of the Hospital Universitario La Paz  approved the study, and informed consent was obtained for the storage and future use of serum samples.Patients were initially treated with 3 mg/kg Ifx intravenously at weeks 0, 2, 6, and 14 and every 8 weeks thereafter. The dose of Ifx was increased via the administration of 4, 5, or 6 mg/kg Ifx or a reduction in the administration interval to 7 or 6 weeks, with a maximum dose of 6 mg/kg every 6 weeks. Dose escalation could also be combined with increasing doses of DMARDs and/or corticosteroids.Medical history, demographic data, anti-citrullinated protein antibodies (ACPA), and rheumatoid factor (RF) were retrospectively retrieved prior to Ifx increase (baseline). DAS28 score, EULAR response, and serum concentrations of Ifx and ATI were also retrieved at baseline (T1), after the first Ifx dose increment (T2), and at 6 months (T3), and 12 months (T4). ACPA were measured using ELISA , and RF was assessed using nephelometry  with cut-off values of 25 and 9 IU/mL. DAS28 was calculated using the erythrocyte sedimentation rate (ESR), and the response to Ifx was evaluated using the European League Against Rheumatism (EULAR) criteria .Serum Ifx concentrations were determined using a capture ELISA as described previously , but a bDescriptive statistics are provided as the mean, SD, median and interquartile range (IQR). A fixed effects analysis of repeated measures was performed, and Bonferroni correction was used for multiple comparisons. Qualitative variables were compared at different time points and between different groups using Fisher’s test, and the Bonferroni test was used for multiple comparisons.A regression mixed model for repeated measurements was performed using group  and time points  as factors to compare DAS28 and delta-DAS28 between groups. Interactions between factors were calculated as fixed effects, and subjects were calculated as random effects. Pair-wise comparisons were calculated using Bonferroni correction. EULAR was analyzed using a generalized linear model with cumulative logit link function. A significance level of 0.05 was used for statistical testing, and all analyses were performed using SAS 9.3 .Patients were assigned to one of three serum Ifx groups: no detectable, low (<1.1 μg/mL) or high (≥1.1 μg/mL). These cut-off levels were based on the observation that serum Ifx >1 mg/L  was assoForty-two patients (37 women) were included in the study. Twenty patients exhibited no detectable free Ifx, 13 patients exhibited Low levels, and 9 patients exhibited High levels. Table The age of disease onset was significantly lower in patients with no detectable drug levels, and the duration of Ifx treatment was significantly longer for patients with High drug levels. Sixteen patients in the no free Ifx group received an increased Ifx dose, and the treatment interval was reduced in eight patients. Eleven and nine patients in the low and high groups, respectively, received an increased Ifx dose, and the treatment interval was reduced in five and four patients, respectively. Both strategies were used simultaneously in some patients, and these numbers are higher. Five patients did not complete the year of treatment because of insufficient clinical response (three patients) and side effects .p < 0.05), but this decrease in DAS28 disappeared at 12 months . The change in DAS28 from baseline (delta-DAS28) demonstrated significant disease worsening (from −0.63 ± 1.18 post-increase to 1.17 ± 1.45 after 12 months (p < 0.001). Figure p = 0.001) and 4.9 ± 1.1 (p = 0.006) in the no detectable and high drug level groups, respectively). No significant change in DAS28 was observed in any of the individual patient groups throughout the study. The decrease in disease activity from the time of post-Ifx dose increase was significant in patients with no detectable Ifx levels after 12 months  and patients with high Ifx levels .Baseline DAS28 for the entire study population Figure A improveEuropean League Against Rheumatism response rates in all three patient groups revealed no significant differences at any time point with 11.1, 16.7, and 12.5% of patients in the no, low, and high groups achieving a good response after the first dose increase, and 44.4, 58.3, and 37.5%, respectively, remained non-responders. Good responders after 12 months of enhanced treatment included 25% of no Ifx patients and 0% of low and high Ifx patients, and 50, 77.8, and 71.4% of no, low, and high patients, respectively, exhibited no response.p = 0.017) but not in the low (p = 0.97) or no (p = 1) groups. No free Ifx was present at 12 months in 4 of 13 patients (30.7%) in the low group despite an increase in Ifx dose. Figure Serum Ifx levels were significantly higher in the high group than the low and no groups at any studied points Figure A. Ifx seThe therapeutic effect of an Ifx dose increase was analyzed in three populations of active non-remitting RA patients who exhibited no detectable, low and high drug levels. No significant improvement in disease activity or responder rate was observed after 1 year of intensified therapy, independently of pre-existing Ifx serum trough levels.Our observations are consistent with anti-TNF failure treatment algorithms , 8 for tAntibodies to infliximab were only detected in patients with no circulating Ifx because of drug interference with the method . The exiOur study has some limitations, such as the retrospective design, the lack of a pre-determined therapeutic protocol to increase Ifx dose, and the lack of a control group, but it reflects the effect of TNF-blocking agents that are used in daily clinical practice. The observational design of this study was also not appropriate for this type of analysis.The enhancing of Ifx therapy is costly and it poses the risk of increased adverse events , which sCP, DP-S, and AB were involved in the study design, patient selection and data collection and analysis. CP wrote the article. AV, LN, DP, and MB were directly involved in patient management. TJ, MC, and AM-F performed the serum assays and data collection. All authors were involved in the interpretation of the data and the critical review of the manuscript.Chamaida Plasencia has received research grants and speaker fees from Pfizer. Dora Pascual-Salcedo has received research grants and speaker fees from Pfizer, AbbVie, Novartis, and MSD. Alejandro Balsa and Emilio Martín-Mola have received unrestricted grants from Pfizer and AbbVie and speaker fees from Pfizer, Janssen, AbbVie, MSD, BMS, UCB, and Roche. The other authors have no competing interests for this research.


Introduction. We report the management of a septic Open Abdomen (OA) patient by the help of negative pressure therapy (NPT) and abdominal reapproximation anchor (ABRA) system in pregnant woman with spontaneous jejunal perforation after emergent cesarean section (C/S) with confounding factor of mild acute pancreatitis (AP). Presentation of Case. A 29-year-old and 34-week pregnant woman with AP underwent C/S. She was arrested after anesthesia induction and responded to cardiopulmonary resuscitation (CPR). There were only ash-colored serosanguinous fluid within abdomen during C/S. After C/S, she was transferred to intensive care unit (ICU) with vasopressor support. On postoperative 1st day, she underwent reoperation due to fecal fluid coming near the drainage. Leakage point could not be identified exactly and operation had to be deliberately abbreviated due to hemodynamic instability. NPT was applied. Two days later source control was provided by conversion of enteroatmospheric fistula (EAF) to jejunostomy. ABRA was added and OA was closed. No hernia developed at 10-month follow-up period. Conclusion. NPT application in septic OA patient may gain time to patient until adequate source control could be achieved. Using ABRA in conjunction with NPT increases the fascial closure rate in infected OA patient. Acute pancreatitis (AP) is a rare event in pregnancy 3/10 000) including a wide range of situations ranging from mild pancreatitis to serious one. Rate of AP is correlated directly with advancing gestational age. Older reviews of AP in pregnancy reported maternal and fetal mortality rates as high as 20 and 50%, respectively [0 000 incThe incidence of bowel injuries is 0.08% in cesarean section (C/S). Patients with a history of abdominal surgery scar are at high risk for intestinal injuries . After CEnterocutaneous fistulas (ECF) are one of the most devastating abdominal complications in intra-abdominal surgery. A newly defined complication is called enteroatmospheric fistula (EAF) which is an enteric fistula in the middle of an Open Abdomen (OA) . The OA We would like to report the management of a septic OA patient by the help of dynamic abdominal closure systems in a 34-week pregnant woman with spontaneous jejunal perforation developing after emergent C/S with her confounding factor of mild acute pancreatitis.μL. Multiple small calculi in different sizes were seen and pancreas was evaluated as edematous suboptimally in abdominal US. The patient was admitted to service to observe and treat for mild AP. The patient underwent C/S emergently for fetal distress 3 days after hospitalization. The patient had a cardiac arrest just after anesthesia induction. She responded to cardiopulmonary resuscitation (CPR) performed approximately for 15 minutes. Fetus was dead on delivery and did not respond to neonatal resuscitation. After C/S, on abdominal exploration, it seemed that there was ash-colored serosanguinous fluid and excessive visceral adhesions. After C/S, the patient was transferred to ICU with vasopressor support. The patient was consulted to us for fecal fluid at the drainage site on postoperative 1st day. At that time her vital parameters under high dose of vasopressor treatment were as follows: BP: 80/45 mmHg, HR: 154 F: 37.9, and RR: 34; her laboratory values were as follows: LDH: 729 U/L, creatinine: 1.6 mg/dL, Alb: 1.61 g/dL, WBC: 2.400 K/uL, CRP: 36.8 mg/dL (0–0.8), procalcitonin: 62.1 ng/mL (<0.5), INR: 1.4, and D-dimer > 1000 ng/mL. In arterial blood gas analysis, pH was 7.24, pCO2 was 34, pO2 was 159, HCO3 was 14, BE was −12, and lac was 9.6. The patient was in septic-shock and metabolic acidosis. The increases in CRP, lactate, and procalcitonin values were shown in A 29-year-old and 34-week pregnant woman was admitted to emergency department with the complaint of abdominal pain, nausea, and vomiting for three days. Epigastric pain radiating to back was present. In her past history, she had been operated on for perforated appendicitis and C/S 10 and 5 years ago, respectively. Her vital parameters were as fallows: blood pressure (BP): 100/50 mmHg, heart rate (HR): 94, and fever: 36.7°C. On her abdominal examination, badly healed midline incision scar from xiphoid to pubis and C/S scar were present. Peritoneal signs and rebound tenderness were positive at epigastric region. In biochemical analysis, LDH: 277 U/L, lipase: 256 U/L, Amylase: 288 U/L, AST: 81 U/L, ALT: 101 U/L, CRP: 7 mg/dL, and in total blood count WBC: 12,000 K/The patient underwent emergent operation from the same Pfannenstiel incision. Enteric influents were seen during exploration and midline incision was added to get better surgical exposure. Since intestines were edematous and fragile and excessive adhesions were present between intra-abdominal visceral tissues, anatomical structures were not identified exactly. During this time, enteric influents were coming from the nonvisible posterior part of adhesive intestinal tissue . LeakageTwo days after NPT application, although source control could not be performed, vital parameters of the patient were better and dose of vasoactive support was decreased. T was 110/65 mmHg, HR was 115, RR was 26, and F was 37.1°C. The improvement of septic parameters of patient was shown in Figures ∗2 as well as 1000 cc intravenous fluid intake a day was ordered. The distal limp of ostomy was also used for enteral nutrition especially for fluid replacement. Additional to oral intake 500 cc %10 dextrose as well as 500 cc %09 NaCl isotonic solution was given daily to the distal limp. By using the distal limp of ostomy, it was understood that the passage of the distal part of the intestine was present. Two months later patient was hospitalized and operated on for closure of the jejunostomy. The distal limp was irrigated with 2000 cc NaCl isotonic solution before the surgery. Only elliptic incision around the stoma was used, and two ends of jejunum were isolated by gentle dissection. Side-to-side jejunojejunostomy anastomosis was performed. No hernia development was observed at 10-month follow-up period ) was used. After debridement and irrigation of OA with warm saline, a perforated silicone sheet was placed over the bowel under the fascia. Then sponge was placed over the silicone sheet. At the end suction tubing was applied. NPT was adjusted in the range of 50–100 mmHg continuously or intermittently [The technique of NPT was as follows: NPT  was as follows: ABRA was applied through the full thickness of the abdominal wall at a distance of approximately 5 cm from the medial fascial margin; a series of midline crossing elastomers are inserted. They are aligned about 3–5 cm apart cross the defect and fixed to button anchors at both sides of OA. The optimal tension was obtained by stretching the elastomers 1.5–2 their tension-free length.In a review of the literature, a total of 20 cases of post-C/S spontaneous cecal perforation were identified. Mortality rates from a cecal perforation range from 30% to 72% . ECF canDuring emergent induction of patient with acute pancreatitis for delivery of death fetus, both these conditions may cause intracellular volume deficiency separately. Arrest on induction may develop due to insufficient preinduction resuscitation as in our case.In severe hemodynamic instability and physiologic derangements definitive repair and definitive closure could not be obtained and restoration of intestinal continuity is postponed to other operations . As far Lots of different methods by combining NPT to other delayed closure methods or a novel device are used for the management of Open Abdomen with EAF , 8. In OWhen NPT was combined with the strategies allowing reapproximation of the fascial edges, high closure rates can be achieved , 10, 15.NPT application in OA patient with EAF may gain time to patient as well as to surgeon until adequate source control could be achieved. Optimal fascial closure of patient with septic OA without mesh could be done by using dynamic abdominal closure methods in conjunction with NPT.


The current fastest frame rate of each single image slice in MR-guided ablation is 1.3 seconds, which means delayed imaging for human at an average reaction time: 0.33 seconds. The delayed imaging greatly limits the accuracy of puncture and ablation, and results in puncture injury or incomplete ablation. To overcome delayed imaging and obtain real-time imaging, the study was performed using a 1.0-T whole-body open configuration MR scanner in the livers of 10 Wuzhishan pigs. A respiratory-triggered liver matrix array was explored to guide and monitor microwave ablation in real-time. We successfully performed the entire ablation procedure under MR real-time guidance at 0.202 s, the fastest frame rate for each single image slice. The puncture time ranged from 23 min to 3 min. For the pigs, the mean puncture time was shorted to 4.75 minutes and the mean ablation time was 11.25 minutes at power 70 W. The mean length and widths were 4.62 ± 0.24 cm and 2.64 ± 0.13 cm, respectively. No complications or ablation related deaths during or after ablation were observed. In the current study, MR is able to guide microwave ablation like ultrasound in real-time guidance showing great potential for the treatment of liver tumors. In popular liver cancer clinical guidelines, ablation therapy is the recommended curative treatment modality. Ultrasound and computerized tomography (CT) are the most common methods for these ablation procedures. However, CT exposes the operator to radiation that cannot be utilized for real-time guidance. Ultrasound provides real-time guidance, but the image quality is worse than CT and magnetic resonance imaging (MRI). In contrast, MRI is characterized by high soft-tissue contrast, sensitivity to thermal signals, and multiplanar capabilitieset al. firstly reported microwave ablation as a feasible tool for MR-guided interstitial hyperthermia at the 8th Annual Meeting of International Society for Magnetic Resonance in Medicine (ISMRM) in 2000, studies on microwave ablation of tumors using MR guidance have been developed. However, image interference generated by radiofrequency or microwave devices in ablation limits the utility of MR. Nevertheless, Morikawa et al. performed MR-guided microwave ablation of liver tumors using a 0.5 T open MRI system in 2002et al. performed MR-guided microwave ablation on 24 patients with liver tumors and shortened the acquisition time for each slice to less than 2 set al. firstly applied respiratory triggering for MR-guided microwave ablation of liver tumors under general anesthesia and overcame the problem of motion artifactset al. performed MR-guided microwave therapy for metastatic liver tumors from breast cancer using open-configuration MR and displayed near-real-time imaging and thermal images on a monitoret al. completed magnetic resonance-guided freehand radio-frequency ablation of malignant liver lesions using an interactive open magnetic resonance scan platform. They successfully reduced the time for each single image slice to 1.3 sSince Morikawa in vivo.To the best of our knowledge, the latest and fastest frame rate of each single image slice for MRI is 1.3 seconds. According to statistic data, the average reaction time for recognition of movement in human is 0.33 secondsQiya Ltd. Co.  donated the microwave generator  and provUnder the supervision of the Division of Laboratory Animal Medicine at our institution, 10 Wuzhishan pigs  weighing 17.5–27.2 kg were studied in this study. All animals received appropriate humane care in compliance with guidelines set by the Laboratory Animal Care and Use Committee and Ethics Committee at our institution. All experimental protocols were approved by both Laboratory Animal Care and Use Committee and Ethics Committee of Sun Yat-sen University Cancer Center.General anesthesia was induced  by injection into the neck muscles of pigs. If anesthesia did not achieved in 10 minutes, an additional dose 3–6 mg/kg would be injected. For the puncture and ablation procedure, 5 mg/kg pentobarbital solution was injected through the venous indwelling needle in the ear every 90 minutes. Xylazine hydrocloride injection  was alternatively used during the procedure for fast anesthesia.A venous indwelling needle in the ear was used for injection of contrast medium or rescue or intravenous anesthesia. Before ablation, pigs were placed on an MRI scan bed under intravenous anesthesia and monitored by an anesthesiologist throughout procedure.The pig was posed in the left lateral position. Imaging markers  were crossed on the abdomen surface. A respiratory-triggered device was bound to the abdominal surface where the maximum motion amplitude of breathing movements was obtained . The resMRI was performed using a 1.0-T whole-body open configuration MR scanner .Differences in blood test results before and after ablation were compared using the In total, 10 Wuzhishan pigs were enrolled in the liver microwave ablation experiments; two males and eight females. The mean age of these animals was 234 days and the mean weight was 21.81 kg. Pig details are shown in During the experiment, we obtained perfect respiratory-triggered liver sequences. The parameters for these sequences were as follows: (1) T1W turbo-field echo (TFE) sequence ; (2) T2W turbo-spin echo (TSE) sequence ; (3) real-time imaging sequence ; and (4) dynamic contrast scan sequence . With these sequences/parameters, we obtained real-time guidance and monitoring of the entire procedure. The shortest time for frame acquisition was 0.202 s. Details of these procedures are shown in The mean size of liver samples was 19.24 cm. The first and second pigs underwent a preliminary test to obtain further information of the equipment and MRI sequences. The time for the puncture procedure was defined from the time when the antenna was inserted into the body to the antenna was posed in the planned position; it was 23 minutes and 15 minutes, respectively. The lengths of the ablated, nearly oval areas were 2.78 cm and 2.83 cm using a power of 60 W, respectively; the widths were 1.65 cm and 1.72 cm, respectively. The other eight pigs were operated based on the preliminary tests. The mean puncture time was shorted to 4.75 minutes and the mean ablation time was 11.25 minutes with a power of 70 W in these animals. The mean ablation length and the widths were 4.62 cm and 2.64 cm, respectively. Details from these pigs are shown in On T1W plain scan images, the antenna represented a long and apparent widening hypointense single line, which was easily distinguished from the normal liver. There was a blooming ball-shaped signal void at the needle tip. On T2W plain scan images, the antenna represented a long hypointense line without surrounding artifacts. The width of the hypointense line was similar to the antenna .In the real-time imaging, vascular showed high signal, the liver was gray signal, gallbladder showed high signal, subcutaneous fat showed high signal. In real-time guided puncture, microwave ablation antenna clearly represented strip of black signal, lower than the liver, same width as ablation antenna. The ablation antenna was moving with breathing exercises. Real-time imaging displayed clear and real-time position of the antenna.In real-time monitoring of the ablation process, the central ablation area showed linear black signal, gradually expanding the scope from ablation antenna. Slightly higher signal area around the black central area with clear boundary representing oval, was gradually expanding. Changes of ablation were clearly showed in the real-time imaging.Instantly after ablation, central ablation area showed long oval, back and low signal around the antenna. Peripheral ablation area showed slightly hyperintense or isointensity in liver with clear or fuzzy boundary. A hypointensity ring was around the boundary on T1WI . AblatioP > 0.05). However, gamma-glutamyl transpeptidase (GGT) and aspartate transaminase (AST) were significantly higher after ablation (P < 0.05). There were no differences in hemoglobin, red blood cell, or albumin counts. Other complications were not monitored. Details of these complications are shown in Of the 10 pigs, no deaths occurred immediately after the ablation procedure; nine pigs were euthanized as planned. Only one pig died of asphyxia during transportation back to the animal facility after ablation. There were no serious complications, and ablation related deaths. Bleeding during the puncture was the most common complication; however, the mean blood loss during the entire procedure was only 6.3 mL. There were no differences in main liver function or renal function before and after ablation .At last, we showed that MR guided microwave liver ablation was safe. There is no serious complications or ablation-related deaths. A small amount of bleeding during the puncture was the most common minor complication. Moreover, there were no differences in liver function, renal function, or routine blood tests before or after ablation .


Effective therapies for the so-called atypical parkinsonian syndrome (APS) such as multiple system atrophy (MSA), progressive supranuclear palsy (PSP), or corticobasal syndrome (CBS) are not available. Dopamine agonists (DA) are not often used in APS because of inefficacy and in a minority of case, their side effects, like dyskinesias, impairment of extrapyramidal symptoms or the appearance of psychosis, and REM sleep behavioral disorders (RBD). Transdermal rotigotine (RTG) is a non-ergot dopamine agonist indicated for use in early and advanced Parkinson’s disease with a good tolerability and safety. Moreover, its action on a wide range of dopamine receptors, D1, D2, D3, unlike other DA, could make it a good option in APS, where a massive dopamine cell loss is documented. In this pilot, observational open-label study we evaluate the efficacy and tolerability of RTG in patients affected by APS. Thirty-two subjects with diagnosis of APS were treated with transdermal RTG. APS diagnosis was: MSA parkinsonian type (MSA-P), MSA cerebellar type (MSA-C), PSP, and CBS. Patients were evaluated by UPDRS-III, neuropsychiatric inventory, mini mental state examination at baseline, and after 6, 12, and 18 months. The titration schedule was maintained very flexible, searching the major clinical effect and the minor possible adverse events (AEs) at each visit. AEs were recorded. APS patients treated with RTG show an overall decrease of UPDRS-III scores without increasing behavioral disturbances. Only three patients were dropped out of the study. Main AEs were hypotension, nausea, vomiting, drowsiness, and tachycardia. The electroencephalographic recording power spectra analysis shows a decrease of theta and an increase of low alpha power. In conclusion, transdermal RTG seems to be effective and well tolerated in APS patients. Atypical parkinsonian syndromes (APS) comprise mostly progressive supranuclear palsy (PSP), corticobasal syndrome (CBS), and multiple system atrophy (MSA) –5. DespiRotigotine (RTG) is a non-ergolinic dopamine agonist administered via a transdermal patch that delivers the drug over a 24-h period. . In six To the best of our knowledge, transdermal RTG has been administered only in patients with idiopathic Parkinson’s disease but not in APS patients. Despite their different neuropathological features, MSA, PSP, and CBS are unanimously recognized as belonging to the nosographical category of the atypical parkinsonisms. Indeed, although MSA is an alpha-synucleinopathy and PSP/CBS are tauopathies, they share as final common pathway the degeneration of the dopaminergic system. The present observational trial has an eminently clinical approach. Indeed, the study was designed to provide a practical therapeutic option to neurologists and physicians in general when facing atypical parkinsonian syndromes. So, the further subdivision of the patients based on the neuropathological substrate was beyond the scope of the study. RTG could be a therapeutic option for APS because its mechanism of action encompasses a wide dopamine receptors spectrum (from D1 to D3), unlike other DA agents. Moreover, RTG patch could have the advantage of a minor adverse events (AEs) rate and the possibility to be administered in patients with swallowing problems. In the present study, RTG was administered to 32 patients with APS, showing an overall good performance on the motor symptoms, without inducing behavioral or extrapyramidal AEs.Thirty-two subjects with diagnosis of atypical parkinsonian disorders were admitted to our National Institute for Research and Cure of Neurodegenerative Disorders Fatebenefratelli (FBF) in Brescia and treated with transdermal RTG. Subjects were evaluated by DVM, a board-certified neurologist and movement disorders specialist who works in the Memory Clinic and Movement Disorders Center. All experimental protocols had been approved by the local ethics committee. Informed consent was obtained from all participants or their caregivers, according to the Code of Ethics of the World Medical Association (Declaration of Helsinki). Table In our subjects, diagnosis was as follows: MSA-P (11), MSA-C (4), PSP (7), and CBS (10). Moreover, all diagnoses were performed exclusively on clinical evaluation. Diagnostic criteria of atypical parkinsonism were inferred from the most diffuse guidelines and clinical criteria –30. BrieAll patients underwent a complete physical and neurological examination. All patients underwent an electroencephalographic recording (EEG), and magnetic resonance imaging (MRI). Only 20 patients underwent single photon emission computed tomography with dopamine transporter scan (SPECT-dat scan). MRI was useful to exclude vascular parkinsonism, whereas SPECT shows diffuse and more symmetric degeneration of nigrostriatal pathways, excluding idiopathic Parkinson’s disease. Patients were assessed by the 56 points unified Parkinson’s disease rating scale (UPDRS) part III , neuropsThe EEG activity was recorded continuously from 19 sites by using electrodes set in an elastic cap  and positioned according to the 10–20 international systems . In order to keep constant the level of vigilance, an operator controlled on-line the subject and the EEG traces, alerting the subject any time there were signs of behavioral and/or EEG drowsiness. The ground electrode was placed in front of Fz. The left and right mastoids served as reference for all electrodes. The recordings were used off-line to re-reference the scalp recordings to the common average. Re-referencing was done prior to the EEG artifact detection and analysis. Data were recorded with a band-pass filter of 0.3–70 Hz, and digitized at a sampling rate of 250 Hz . Electrodes-skin impedance was set below 5 kHz. Horizontal and vertical eye movements were detected by recording the electrooculogram (EOG). The recording lasted 5 min, with subjects with closed eyes. Longer recordings would have reduced the variability of the data, but they would also have increased the possibility of slowing of EEG oscillations due to reduced vigilance and arousal. EEG data were then analyzed and fragmented off-line in consecutive epochs of 2 s, with a frequency resolution of 0.5 Hz. The average number of epochs analyzed was 140, ranging from 130 to 150. The epochs with ocular, muscular, and other types of artifacts were discarded by two skilled electroencephalographists . EEG recAll recordings were obtained in the morning with subjects resting comfortably. Vigilance was continuously monitored in order to avoid drowsiness. A digital FFT-based power spectrum analysis  computed – ranging from 2 to 45 Hz – the power density of EEG rhythms with a 0.5 Hz frequency resolution. Two anchor frequencies were selected according to the literature guidelines, that is, the theta/alpha transition frequency (TF) and the individual alpha frequency (IAF) peak. These anchor frequencies were computed on the power spectra averaged across all recording electrodes. The TF marks the TF between the theta and alpha bands, and represents an estimate of the frequency at which the theta and alpha spectra intersect. TF was computed as the minimum power in the alpha frequency range, since our EEG recordings were performed at rest. The IAF represents the frequency with the maximum power peak within the extended alpha range (5–14 Hz). Based on TF and IAF, we estimated the frequency band range for each subject, as follows: delta from TF-4 to TF- 2, theta from TF-2 to TF, low alpha band from TF to IAF, and high alpha band from IAF to the point of the minimum power after the IAF values. The mean frequency range computed in MCI subjects considered as a whole are: delta 2.9–4.9 Hz; theta 4.9–6.9 Hz; low alpha1 6.9–10.9 Hz; and high alpha2 8.9–11.9 Hz. Finally, in the frequency bands determined on an individual basis, we computed the relative power spectra for each subject. The relative power density for each frequency band was computed as the ratio between the absolute power and the mean power spectra from 2 to 45 Hz. The relative band power at each band was defined as the mean of the relative band power for each frequency bin within that band .post hoc correction, was performed to test the changes in MMSE, UPDRS-III, and NPI considering all the evaluations . Also analysis of EEG results has been performed with ANOVA.One-way analysis of variance (ANOVA), with Newman–Keuls Table At baseline, mean UPDRS-III score was 36.1/56, mean NPI score was 56.4/144, mean MMSE score was 20.3/30. 29/32 (90.7%) patients has reached 6 months of treatment with RTG (mean dose = 3.2 mg/24 h); mean UPDRS-III score was 28.6/56, mean NPI score 50.1/144, mean MMSE score 19.3/30. Three patients were dropped out for AEs. 23/32 (71.8%) patients has reached 12 months of treatment with RTG (mean dose = 3.7 mg/24 h), mean UPDRS-III score was 26.5/56, mean NPI score was 41.2/144, mean MMSE score was 18.1/30. No patient more was dropped out for AEs at 12 months follow-up. 16/32 (50%) patients completed 18 months of treatment with RTG (mean dose = 4.4 mg/24 h) mean UPDRS-III score was 25.5/56, mean NPI score was 35.9/144, mean MMSE score was 15.3/30. No patient was dropped out at 18 months follow-up (Table p < 0.005) was found. In post hoc analysis, all the comparisons with the baseline were statistically significant (p < 0.02).Figure p < 0.03) was found. In post hoc analysis, all the comparisons with the T18 follow-up were statistically significant (p < 0.04). Moreover, the evaluation at T12 was significant as compared to the baseline (p < 0.05).Figure p < 0.05) was found. In post hoc analysis, the comparison between baseline evaluation and the T18 follow-up was statistically significant (p < 0.01).Figure p < 0.001, low alpha p < 0.002, high alpha p = 0.09) follow-up and were maintained at both the 12-month follow-up  and the 18-month follow-up .The power spectra show a significant decrease of theta and increase of low alpha band but no for high alpha power. These results were present at the 6-month  for 48 weeks in 174 patients with possible or probable MSA-Parkinsonism type, in 39 sites in 12 countries, found no significant difference in progression in the total UMSARS score between the verum and placebo groups . A singlwww.allontherapeutics.com). An open-label pilot trial of lithium, an inhibitor of glycogen synthase kinase-3 (GSK-3), in individuals with PSP or CBS  recruited 17 patients and was stopped prematurely because the majority of participants did not tolerate the study drug. A multinational, phase II, double-blind, placebo-controlled trial enrolled 142 patients with PSP, who were treated orally with tideglusib (600 or 800 mg p.d.), also a GSK-3 inhibitor, or placebo for 1 year. There were no significant differences between the high dose, low dose, and either dose groups vs. the placebo group in the primary clinical outcome measures. A subset of 37 patients underwent baseline and 52-week MRI; this substudy demonstrated significantly reduced global brain atrophy in tideglusib-treated patients  (Press release December 18, 2012 by Allon Therapeutics, patients . The effpatients .in vitro studies (Transdermal RTG seems to be effective and well tolerated in patients with APS. Our results show significant improvement in UDPRS-III scores, maintained along the course of the 18 months follow-up. Moreover, only 3 patients were dropped out and 15 patients were affected by transient AEs. A plausible explanation of the results is that RTG transdermal patch has a wider dopaminergic profile action as compared to other non-ergot, prolonged-release DA, allowing a more diffused reply therapy, covering the impotent dopaminergic loss in atypical parkinsonism . Indeed, studies , this as studies , 56.Our results show no positive effect on cognitive status. Anyway, the MMSE score shows that patients were highly cognitively impaired, on average, at the beginning of the study. Further studies, with less initially cognitively compromised patients and grouped for single pathologies, will better clarify this issue.Our results confirm previous works showing that the most significant finding in Parkinson’s disease synchronized oscillatory activity is the prevalence of a rhythmic activity in the range of 4–7 Hz, usually termed theta band. This activity is characteristic of the basal ganglia-thalamocortical circuits, and such activity can be reduced by dopaminergic treatments . The incThis study is a pilot, open-label, observational study with a clear explorative purpose. First of all, we have to remark that small sample size has not made possible the investigation on the single pathology level because of the lack of any statistical power. Of note, the results could not be extended to other conditions such as the primary progressive freezing of gait preceding the PSP, because we have not investigated such phenotype. As a consequence, further randomized, double-blind, placebo-controlled trial studies with a bigger sample size and with more disease-specific scales and assessment for both the daily living functionality and the statistical evaluation are needed to confirm the present results. Anyway, the favorable outcomes could be very useful as a practical guideline to help clinicians and neurologists to find a treatment in atypical parkinsonisms.In our observational, open-label study, RTG appears to be a suitable therapy in APS patients as it has a good tolerability and efficacy profile. The activation of D1–D2–D3 receptor in the caudate–putamen region by RTG patch, compensates for the spread loss of dopaminergic function in these areas and could be responsible for the efficacy of this drug. Anyway, more studies are mandatory to confirm the results.Davide Vito Moretti: on the behalf of all coauthors I declare that appropriate approval and procedures were used concerning human subjects.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The effect of bile acids administration to an experimental mice model of Protoporphyria produced by griseofulvin (Gris) was investigated. The aim was to assess whether porphyrin excretion could be accelerated by bile acids treatment in an attempt to diminish liver damage induced by Gris. Liver damage markers, heme metabolism, and oxidative stress parameters were analyzed in mice treated with Gris and deoxycholic (DXA), dehydrocholic (DHA), chenodeoxycholic, or ursodeoxycholic (URSO). The administration of Gris alone increased the activities of glutathione reductase (GRed), superoxide dismutase (SOD), alkaline phosphatase (AP), gamma glutamyl transpeptidase (GGT), and glutathione-S-transferase (GST), as well as total porphyrins, glutathione (GSH), and cytochrome P450 (CYP) levels in liver. Among the bile acids studied, DXA and DHA increased PROTO IX excretion, DXA also abolished the action of Gris, reducing lipid peroxidation and hepatic GSH and CYP levels, and the activities of GGT, AP, SOD, and GST returned to control values. However, porphyrin accumulation was not prevented by URSO; instead this bile acid reduced ALA-S and the antioxidant defense enzymes system activities. In conclusion, we postulate that DXA acid would be more effective to prevent liver damage induced by Gris. Erythropoietic protoporphyria (EPP) is a hereditary disease due to a decreased activity of ferrochelatase (Fech) EC.4.99.1.1), the last enzyme of heme pathway catalyzing the incorporation of ferrous iron (II) into protoporphyrin IX (PROTO IX) to synthetize heme .1, the l. EPP patPROTO IX is excreted by the hepatobiliary-fecal route and can move to the enterohepatic recirculation. The entrance of PROTO IX to the liver is not saturated; when its excretion through the bile fails to remove the excess of PROTO IX produced by the bone marrow and the liver, this porphyrin accumulates in hepatocytes .Bile acids have choleretic and digestive actions. In humans, bile salts are used as a cathartic to promote bile flow after biliary surgery and to reverse the formation of calculi. In EPP patients bile acids have been used to increase liver porphyrins excretion enhancing cholestasis and in the established cirrhosis to improve both cholestasis and liver function –11.δ-aminolevulinic acid synthetase (ALA-S). To porphyrin accumulation followed cellular damage, inflammatory and necrotic events [The antifungal griseofulvin (Gris) develops in animals a model of EPP  with livc events , resemblc events . Gris alc events .The main difference between human EPP and Gris animal model is that the overproduction of PROTO IX in the latter begins in the liver instead of the bone marrow , 19.Considering the damage produced by Gris in mice liver, the aim of this study was to investigate the effect of administrating deoxycholic (DXA), dehydrocholic (DHA), chenodeoxycholic (CHENO), and ursodeoxycholic (URSO) acids on heme metabolism, antioxidant defense enzymes system, and CYP levels in Gris treated mice, to assess whether they were able to accelerate the excretion of porphyrins through the bilis, in an attempt to diminish liver damage. CF1 weighing 15–17 g at the start of intoxication were used. They were maintained in controlled conditions and allowed free access to food  and water. A 12/12 h light/dark cycle was kept. Animals received human care and were treated in accordance with the guidelines established by the Committee of the Argentine Association of Specialists in Laboratory Animals Care (AADEALC).Male miceAll treatments were performed at the same time of the day. In each experiment, animals were separated into 4 groups of 6 mice each and during two weeks they received the drugs as follows: group I: control diet ; group II: control diet plus Gris ; group III: control diet plus DXA  in the drinking water or CHENO , DHA , or URSO  in food: group IV: control diet plus Gris  and DXA  in the drinking water or CHENO , DHA , or URSO  in food. Animals were starved 16 hours before sacrifice under ether anaesthesia.Blood was obtained by cardiac puncture and plasma was separated by centrifugation. The faeces of each cage were collected during the 16 hours before sacrifice. Liver was removed and immediately processed. Experimental conditions for liver homogenate preparations were different, depending on the parameter assayed. A fraction of the liver was immediately homogenized  in a solution containing 0.9% NaCl, 0.1 mM Tris HCl pH 7.4, and 0.5 mM EDTA, for ALA-S activity determination. The remainder of the organ, previously perfused with sterile ice cold saline, was removed. A fraction was homogenized  in ice cold 0.25 M sucrose. After differential centrifugation of this homogenate, supernatant of 10,000 ×g was used for measuring GSH levels and GST activity. An aliquot of this supernatant was then centrifuged at 105,000 ×g for 60 min. The supernatant obtained was used for measuring SOD activity and the pellet for determining total CYP levels. Another fraction of the perfused liver was homogenized  in 0.05 M sodium phosphate buffer, pH 7.4, and it was used as such for the determination of MDA or it was centrifuged at 10,000 ×g for 10 min; in this supernatant GRed activity was measured. The other fraction of the perfused liver 200–300 mg) was used for quantifying PROTO IX levels. Porphyrins were extracted from liver using the method described by De Matteis and Lim 0–300 mg .λex 400 nm, λem 632 nm). Enzyme units were defined as the amount of enzyme that catalyzes the formation of 1 nmol of product under the standard incubation conditions. One unit of SOD is defined as the amount of enzyme causing 50% inhibition on the rate of NADH oxidation measured in the control. Specific activities were expressed as units/mg protein.ALA-S activity was determined following the method of Marver et al. . Lipid pμm were stained with Haematoxylin and Eosin. Immunohistochemistry for heme oxygenase (HO1) detection was performed using the streptavidin-biotin-peroxidase complex system LSAB (DAKO) as previously reported [Livers were removed and fixed in 10% neutral-buffered formalin. Samples of each hepatic lobule were processed routinely and embedded in paraffin. At least six microtome sections of 3–5 reported .All data represent mean values ± standard deviation of all experiments performed in duplicate. The differences between groups were determined by the analysis of the variance (ANOVA) and the significance level was verified by the Bartlett test. A probability level of 99.9% or 99.5% was considered as significant difference between groups.Histological studies were performed in livers of mice, receiving 0.5% Gris, alone and with the bile acids during 14 days. In all cases, similar alterations were observed . The livThe anti-HO1 immunostaining showed positive mark, mainly in isolated hepatocytes, being abundant in Kupffer's cells in liver of Gris or Gris plus bile acid groups Figures .P < 0.01) this parameter but this enhancement was not reversed by any of the acids assayed  GGT activity (P < 0.01) this enzyme activity; such enhancement also occurred in animals receiving CHENO concomitantly with Gris. This induction was significantly greater than that produced by Gris alone. Administration of DHA did not modify the 40% augmented activity due to Gris. In contrast, DXA and URSO significantly prevented (P < 0.05) the induction of GGT caused by Gris, returning to basal levels (Gris induced 40% (activity . When DHl levels .P < 0.01) produced by Gris  in animals treated with Gris. All the bile acids given alone diminished 40% (P < 0.01) ALA-S, but when they were administered with Gris, no effect was observed with respect to those animals receiving only Gris, except in the case of URSO which restored ALA-S activity to basal levels (ALA-S activity was increased 40% (l levels .P < 0.01) porphyrin enhancement levels induced by Gris, but without returning to basal levels  PROTO IX excretion; treatments with DHA or DXA produced an excretion 3-fold (P < 0.01) higher than that of animals receiving only the antifungal. When Gris was administrated simultaneously with CHENO or URSO, PROTO IX levels in feces were similar to their respective controls . Instead, Gris increased 400% (controls .P < 0.01) MDA levels (P < 0.05) the increase of MDA levels produced by Gris, the treatment with CHENO or DXA returned MDA content to basal levels (P < 0.01)  .P < 0.01) enhancement of GSH levels, alike in mice treated with Gris or with the combination of CHENO–Gris or URSO–Gris. DXA and DHA restored GSH to basal levels  GRed activity. These enhanced levels were not reversed by the administration of CHENO, DHA, or DXA; however addition of URSO decreased to 20% (P < 0.01) GRed activity induced by Gris alone  SOD activity, while no changes were produced by the other bile acids assayed. Gris induced 70% (P < 0.01) SOD activity. When animals received DXA or URSO with Gris, this enzyme activity decreased 70% (P < 0.01) and 50% (P < 0.01), with respect to Gris group, reaching control values  GST activity, without any alteration by the action of the other bile acids studied. Gris induced 30% (P < 0.01) enzyme activity, remaining enhanced after CHENO plus Gris treatment. DHA produced an additional induction (60% P < 0.01) to that induced by Gris. Instead, when DXA or URSO was given to Gris feeding mice, GST activity returned to control values  decrease of CYP levels, while no changes after DXA or CHENO treatments were observed. Gris leads to an increase of 40% (P < 0.01) in hepatic CYP levels, without any modification when Gris was given simultaneously with CHENO or ADX. DHA diminished 130% (P < 0.01) the hepatic CYP induction produced by Gris. Only URSO was able to reverse CYP levels to normal values when it was given concomitantly with Gris (DHA and URSO administration to controls animals produced a 40% (ith Gris .In EPP patients, when hepatocellular damage progresses to a critical stage, PROTO IX accumulation is markedly accelerated due to biliary excretion impairment . EPP patIn animal studies, published results are contradictory; in mice treated with Gris, it was reported that URSO administration may have a cytoprotective and choleretic action on the liver injury induced by Gris , 34 althIn our model, the administration of different bile acids led to several effects. Among the bile acids assayed, only DXA and DHA increased PROTO IX excretion. We have found that URSO did not prevent porphyrins accumulation; however it decreased ALA-S activity, indicating that it is acting on heme biosynthesis. HO1 expression was induced as a consequence of the oxidative stress produced by Gris and PROTO IX accumulation; but this effect was not reversed after bile acids treatment. In our study only URSO and DXA were able to prevent enhancement of GGT and PA induced by Gris, indicating the existence of certain protection against these disturbances, although no histological changes were observed when compared to the group treated only with Gris.It has been reported that URSO possesses antioxidant properties, while hydrophobic bile acids generate ROS acting as prooxidants , 37. PorThe increase of GST activity in mice receiving Gris is indicative of liver damage . It has We have previously observed an increase of hepatic CYP levels in animals treated with Gris alone , but it In conclusion, taking into account that DXA effectively reduced porphyrins accumulation induced by Gris and consequently protected the liver against lipid peroxidation and maintaining GSH levels, GGT and AP activities, and drug metabolizing system in basal levels, we postulate that this bile acid would be the most effective in preventing the liver damage induced by Gris.


Body weight gain in NR gilts was significantly decreased by nutrient restriction. Gilts were considered as anestrus when blood progesterone concentrations lower than 1.0 ng/mL from three consecutive blood samples were recorded. Circulating concentrations of progesterone (< 1.0 ng/mL vs. 2.1 ng/mL) and estradiol (208.6 ng/mL vs. 371.8 ng/mL) were significantly lower in the NR gilts than in the CON gilts. Between 5,360,000 and 5,370,000 sequence reads per sample from the CON and NR gilts’ pituitaries were obtained and mapped to the porcine genome. Analysis of read counts revealed 185 DEGs. Expression of selected genes was validated by the use of quantitative real-time RT-PCR. Bioinformatic analysis identified that the genes identified were enriched in the GO terms “neuroactive ligand-receptor interaction”, “GnRH signaling pathway” and “immune response system”. Our findings provide a new perspective for understanding the nutrient restriction-induced reproductive impairment at the pituitary transcriptional level, and how this is linked to hormone secretion. Moreover, the transcriptomic changes in anestrus gilts associated with nutrient restriction could be a resource for targeted studies of genes and pathways potentially involved in the regulation of reproductive function and animal health.The onset of estrus is a critical sign of female sexual maturity. The pituitary plays a vital role in this process by the secretion of reproductive hormones. To investigate the effects of nutrient restriction on reproductive function and the underlying mechanisms involved, deep RNA sequencing of pituitary gland tissue was carried out to determine the differentially expressed genes (DEGs) between gilts in normal estrus, and gilts in which anestrus was induced by nutrient restriction. Gilts which had gone through two estrus cycles were fed a normal  or nutrient restricted  diet. The NR gilts experienced another three estrus cycles, but did not express estrus symptoms at the anticipated 6 Throughout vertebrates the onset of estrus indicates that the female organism has reached sufficient sexual maturity to reproduce. Studies have shown that reproductive function in pigs is sensitive to the status of energy reserves suggesting that weight loss in humans by extreme dieting methods may have potential negative effects on fertility –3. NutriNutritional status, especially overall energy level, is important for the onset of estrus –3, 5. DeAt the present time, it has become routine to obtain large-scale genetic data at the genomic and transcriptomic levels in pigs due to the development of high-throughput deep sequencing technologies. These approaches include Illumina’s RNA sequencing (RNA-seq) system, which employs whole transcriptome shotgun sequencing wherein mRNA or cDNA is mechanically fragmented, resulting in overlapping short fragments that cover the entire transcriptome. RNA-seq is powerful for unraveling transcriptome complexity, and for the identification of genes, splice variants, non-coding RNAs, and even novel transcriptional units –10.The pig anestrus model is an important tool for the investigation of the influence of nutrition on estrus onset. Identification of differences in gene expression between normal and anestrus gilts could shed light on the underlying mechanisms. Therefore, using nutritionally-induced anestrus gilts, we assessed the hormonal secretion and transcriptomic responses of the pituitary by deep RNA sequencing. The mechanisms behind these nutrition-induced responses were investigated on a molecular basis.ad libitum. The environmental temperature was controlled at 20–24°C. The condition of the animals was monitored and recorded daily.All experimental procedures were approved by the Animal Care and Use Committee of Sichuan Agricultural University. Twenty Landrace × Yorkshire crossbred gilts having experienced two estrus cycles were used in this study. Gilts with similar bodyweight (122 ± 4.5 kg) and age (230 ± 6 days) were allocated to two nutrition allowance levels and either fed a normal diet  or a nutrient restricted diet  . Diets wThe day of the second occurrence of standing heat in gilts was designated as day 0 (d 0). At d 0, the gilts in CON and NR groups were started on their respective feeding regimens. All gilts had fence-line contact with a mature boar to encourage pubertal estrus. Onset of estrus was detected by conducted by back pressure testing and examination of vulval characteristics twice daily, in the morning and afternoon, by an experienced stockperson. Gilts failed to express estrus until the 23rd day of the 6th estrus cycle (d 88) in the NR group. Blood samples (10 mL) were collected from the both groups of gilts every 3 days to determine the concentration of progesterone. When progesterone concentrations were below 1.0 ng/mL for three consecutive days in gilts who did not enter estrus, this was defined as nutritionally induced anestrus .th estrus cycle was observed in the control group) both groups of gilts were anaesthetized with an intravenous injection of Zoletil 50 , and were bled by exsanguination. Brains were immediately removed from the skull and excess tissues were removed. Within a few minutes after slaughter the pituitary gland was removed and flushed with ice-cold PBS, then rapidly frozen in liquid nitrogen. All the samples were maintained at −80°C until RNA analysis were performed.At d 107  were determined using enzyme-linked immunosorbent assay kits , according to the manufacturer’s recommendations. The limits for detection of E2 and P were 0.01 ng/mL, and 1.0 ng/mL, respectively. The intra- and inter-assay coefficients of variation were 6.6% and 7.5% for E2, and 5.3% and 6.4% for P.260:OD280 and denaturing agarose gel electrophoresis. All RNA samples were treated with DNase-I . RNA integrity and concentration were evaluated using an Agilent 2100 Bioanalyzer .Total RNA was extracted with TRIzol reagent  from frozen tissues ground in liquid nitrogen. The quality and purity of RNA samples were checked by spectrophotometry  from the ODhttp://en.genomics.cn/navigation/index.action).For RNA library construction and deep sequencing, equal quantities of RNA from four individual pig pituitaries which were fed a normal diet were pooled, and RNA samples from the four nutrient restricted pig pituitaries were pooled. Approximately 4 μg of RNA from each treatment group were submitted for sequencing. In brief, after RNA extraction, poly A-containing mRNAs were purified using oligo-dT-conjugated magnetic beads and fragmented into small pieces. Using these short fragments as templates, first- and second-strand cDNA were synthesized. These cDNA fragments were subjected to end repair, addition of a single ‘A’ base, and ligation of adapters. Products were subsequently purified and amplified by PCR to construct the final cDNA libraries. Finally, the raw image files from each of the libraries were collected by the Illumina HiSeqTM 2000 sequencing platform in BGI Shenzhen  was employed for sequence analysis. To compare the differentially expressed genes (DEGs) in the two treatments, a rigorous algorithm has been developed and described previously [P-value was used corresponding to a differential gene expression test at statistically significant levels [P-value < 0.005, FDR ≤ 0.001 and an absolute value of log2-fold change ≥1 in sequence counts across libraries.All data is MIAME compliant. For the raw data, we filtered adaptor tags, low quality tags and tags with a copy number of 1 to obtain clean tags. Subsequently, the clean tags were classified according their copy number in the library, and the percentage of total clean tags was calculated and saturation of the library was analyzed. The eviously . The P-vt levels . Genes w-△△Ct method [To confirm the differential expression of genes revealed by RNA-Seq, the expression of genes in the pituitary, including Kiss-1, G protein-coupled receptor 54 (GPR54), FSH, LH, GnRHR and leptin receptor (Ob-R), were measured by qPCR. Complementary DNA (cDNA) was synthesized with random primers . Real-time PCR was used to quantify mRNA expression levels using the Sybr Green Kit . Primers are listed in t method .t test and presented as means ± SEM. Differences were considered significant at P < 0.05. Functional classification of DEGs was conducted using the Gene Ontology atlas. Pathway analysis of DEGs was mainly based on the Kyoto Encyclopedia of Genes and Genomes (KEGG) database [Data from gilts in the normal and nutrient-restricted groups were compared by database .th and 7th estrus (d 88 and d 107). The day of the second occurrence of standing heat was designated as d 0. As shown in P = 0.001). The back fat thickness decreased in the NR group compared with the CON group at d 89 and d 107 .In the present study, nutrient restricted gilts (n = 10) which already had experienced two estrus cycles failed to express estrus at the anticipated time of the 6P < 0.01). This was a reflection of nutritionally induced anestrus [The circulating concentrations of progesterone were determined at d 0 and every 3 days from d 88 to d 107. In the NR gilts progesterone concentrations were lower than 1.0 ng/mL and also beyond the limit of detection from d 88 to d 107. As shown in anestrus .Circulating concentrations of E2 were determined at d 0, d 88, d 91 and d 107. E2 concentrations at d 0, d 88 and d 91 were not affected by nutrient restriction, with the exception of decreased E2 concentration in the NR gilts at d 107 compared with the CON group  ≤ 0.001 and estimated absolute log2-fold change ≥ 1 in the pairwise comparisons . Interestingly, BP enrichment performed by two-sided Fisher’s exact test showed enrichment of processes involving “response to endogenous stimulus”, “response to hormone stimulus”, “humoral immune response mediated by circulating immunoglobulin” and “humoral immune response” as well as several others . In addition, we found that the CON and NR gilts have differences in cellular components, including “extracellular region” and “extracellular region part” .The molecular function (MF), biological process (BP) and cellular component (CC) analysis based on gene ontology (GO) in the 185 identified DEGs are shown in The 185 DEGs could be grouped into 138 pathways based on analysis using the KEGG pathway database. These pathways and the DEGs involved are shown in P < 0.05). The expression of Kiss1, GPR54 and GnRHR were not affected by nutrient restriction.To validate DEGs identified by RNA-seq, we used qRT-PCR to measure the expression of six genes. The genes were selected based on their involvement in neuroactive ligand-receptor interaction and GnRH signaling pathway  or because they are important components of estrus onset . Our results indicated that most of the genes investigated had expression levels which were consistent between RNA-Seq and qRT-PCR assays . There wThe onset of estrus is the hallmark of reproductive maturity, and a sign that the female individual is sufficiently mature to produce offspring. It has been reported that nutritional factors are the major environmental influence on the timing of puberty . Howeverth and 7th estrus). The concentrations of progesterone were below 1.0 ng/mL for the three consecutive blood samples after the point defined as nutritionally induced anestrus [In this study, the gilts were allowed to initially experience two estrus cycles. In the NR group, the nutrition allowance level was 1 kg/d, which is lower than the maintenance nutrient requirement by NRC 1998) . Our resanestrus . It is wanestrus –20. Also98 . Our anestrus . Our preanestrus . The lowSus scrofa UniGene database , growth hormone releasing hormone receptor (GHRHR), growth hormone (GH), prolactin (PRL) were altered by nutrient restriction. These genes not only play a vital role in reproduction and lactation , 22, 23,The GO terms “GnRH signaling pathway”, “oocyte meiosis” and “progesterone-mediated oocyte maturation” have well described roles in reproduction via DEGs including inositol 1,4,5-trisphosphate receptor type 1, calbindin 2 (CALM) and progesterone receptor , 29. In The Kiss1/GPR54 (G-protein-coupled receptor) system is considered as the gate keeper of estrus , 35. ThiIn conclusion, this study provides evidence at the transcriptional level to explain why postpubertal gilts terminate their reproductive functions in a nutrient restricted diet. From the GO and the KEGG analysis, it was also found that nutrient restriction had effects on immunity and growth in the gilts. Based on these results, humans should also be aware of potential risks to reproductive function and even overall health when we are on a diet.S1 FigA, pituitary sample from normally fed gilts (2.86 kg/d); B, pituitary sample from nutrient restricted gilts (1 kg/d).(TIF)Click here for additional data file.S2 FigThe Genes shown in red were upregulated and those shown in green were downregulated in CON relative to NR gilts. See supplementary (TIF)Click here for additional data file.S1 File(ZIP)Click here for additional data file.S1 TableCON denotes normally fed gilts (2.86 kg/d) and NR denotes nutrient restricted gilts (1 kg/d).(XLSX)Click here for additional data file.S2 TableCON denotes normally fed gilts (2.86 kg/d) and NR denotes nutrient restricted gilts (1 kg/d).(XLSX)Click here for additional data file.S3 TableCON denotes normally fed gilts (2.86 kg/d) and NR denotes nutrient restricted gilts (1 kg/d).(XLSX)Click here for additional data file.S4 TableThe Genes shown in red box were upregulated and those shown in green box were downregulated in CON relative to NR gilts in the hyperlink pathway figure. CON denotes normally fed gilts (2.86 kg/d) and NR denotes nutrient restricted gilts (1 kg/d).(DOCX)Click here for additional data file.