Preeclampsia is a leading cause of maternal mortality and morbidity worldwide. The neurological complications of preeclampsia and eclampsia are responsible for a major proportion of the morbidity and mortality for women and their infants alike. Hormonal changes during pregnancy and the puerperium carry an increased risk of venous thromboembolism including cerebral venous sinus thrombosis (CVST). Factor 5 leiden (FVL) is a procoagulant mutation associated primarily with venous thrombosis and pregnancy complications. We report a patient with FVL mutation who presented with CVST at 24th week of pregnancy and was diagnosed as HELLP syndrome at 34th week of pregnancy. The pregnancy-related syndromes of preeclampsia, eclampsia, and HELLP syndrome are a group of disorders related to endothelial damage triggered by pregnancy. Preeclampsia is defined as gestational hypertension (blood pressure ±140/90 mmHg) with proteinuria >0.3 g/24 hours. It is seen in about 3–8% of all pregnancies [We report a patient with FVL mutation who presented with CVST at 24th week of pregnancy and was diagnosed as HELLP syndrome at 34th week of pregnancy.A 26-year-old woman, gravida 5, abortus 3, and parity 1, was admitted to us at 24th week of gestation with headache and blurred vision. On ophthalmological examination there was bilateral papilledema and visual field loss was detected. Magnetic resonance imaging of the brain revealed CVST  and antiAt 34th week of gestation, she was hospitalized for high blood pressure, headache, nausea, and vomiting. On admission blood pressure was 220/120 mmHg and laboratory findings were as follows: aspartate transaminase (AST): 227 U/L, alanine transaminase (ALT): 176 U/L, INR: 1.3, white blood cell (WBC): 10.800 U/L, hemoglobin (Hb): 14.1 gr/dL, thrombocyte count (PLT): 153000 U/L, and 3+ proteinuria in the urine analysis. Hematological and urinary parameters were consistent with severe preeclampsia. She underwent cesarean section at 34th week of gestation due to severe preeclampsia. By cesarean section, a baby weighing 1840 g with 1 minute APGAR 9 and 5 minute 10 APGAR scores was delivered.The patient had no significant bleeding during and after cesarean section, but atony-related postpartum hemorrhage developed at 5th postoperative hour. Laboratory investigations showed WBC: 19500 U/L, Hb: 10 gr/dL, PLT: 58000 U/L, AST: 1494 U/L, and ALT: 681 U/L. Due to postpartum hemorrhage, uterine massage was performed and uterotonic drugs were administered. As bleeding continued bakri balloon was inserted. Laboratory tests were WBC: 16,600 U/L, Hb: 8.3 g/dL, PLT: 44000 U/L, AST: 1299 U/L, ALT: 489 U/L, creatinine: 1.48 mg/dL, and LDH: 2779 U/L at 7th postoperative hour. She was operated on again with total hysterectomy and bilateral hypogastric artery ligation. The patient received thirteen units of fresh frozen plasma, 3 units of platelet apheresis, 9 units of whole blood, and 3 units of packed red blood cells. At 4th postoperative day, she was discharged from the intensive care unit and monitoring continued in the service. Laboratory tests and blood pressure returned to normal at 10th postoperative day. Headache and blurred vision did not improve and bilateral papilledema still existed. The patient was started acetazolamide and enoxaparin sodium. A CT venogram showed canalised sinus thrombosis . She wasPregnancy is a hypercoagulable state. The combination of combined oral contraceptive and thrombophilia greatly increases the risk of CVST, particularly in women with hyperhomocysteinemia, FVL, and the prothrombin-gene mutation . In our The field of thrombophilia, the tendency to thrombosis, has been developed rapidly and has been linked to many aspects of pregnancy. Recurrent miscarriage has been associated with thrombophilia. It has been shown that pregnancy complications such as severe preeclampsia, intrauterine growth retardation, abruptio placentae, and stillbirth may be associated with thrombophilia . Our patThrombophilias are inherited or acquired conditions which predispose an individual to thromboembolism. Deficiencies of protein S, protein C, and antithrombin are rare and each of them is found in about 3% of patients with thrombosis. Heterozygosity for the FVL mutation is found in about 5% of the population and the mutation is responsible for 20–30% of venous thromboembolic events , 5. Our The main clinical manifestations of CVST include papilledema 62%), headache (62%), hemiparesis (48%), seizures (31%), and cranial nerve palsy (7%). Patients are managed with heparin followed by warfarin during the succeeding 6 months. Death due to CVST has shown remarkable reduction because of early diagnosis and appropriate anticoagulation [2%, headaPatients with CVST may develop—as well as sometimes present with—chronic intracranial hypertension with headache and papilledema. The priority is prevention of visual function loss; intracranial hypertension should be controlled with acetazolamide and occasionally with repeated lumbar punctures if vision is still threatened. Refractory cases may need a cerebrospinal fluid shunting procedure . Our patThrombophilia and CVST are rarely encountered conditions during pregnancy and augment the risk of life-threatening maternal complications and adverse perinatal outcomes in preeclamptic patients . Therefo


In many resource-limited settings, combination antiretroviral therapy (cART) failure is diagnosed clinically or immunologically. As such, there is a high likelihood that patients may stay on a virologically failing regimen for a substantial period of time. Here, we compared the long-term impact of initiating non-nucleoside reverse transcriptase inhibitor (NNRTI)- versus boosted protease inhibitor (bPI)-based cART in British Columbia (BC), Canada.N=1963, 50%) or bPIs  from 1 January 2000 until 30 June 2013 in BC. At six months, we assessed whether patients virologically failed therapy  >50 copies/mL), and we stratified them based on the pVL at the time of failure ≤500 versus >500 copies/mL. We then followed these patients for another six months and calculated their probability of achieving subsequent viral suppression (pVL <50 copies/mL twice consecutively) and of developing drug resistance. These probabilities were adjusted for fixed and time-varying factors, including cART adherence.We followed prospectively 3925 ART-naïve patients who started NNRTIs  vs. 0.72 (0.61–0.75)). However, if failing NNRTI initiators had a pVL >500 copies/mL, they had a 20% lower probability of suppressing at 12 months than pVL-matched bPI initiators (0.37 (0.29–0.45) vs. 0.46 (0.38–0.54)). In terms of evolving HIV drug resistance, those who failed on NNRTI performed worse than bPI in all scenarios, especially if they failed with a viral load >500 copies/mL.Our results show that patients who virologically failed at six months on NNRTI and continued on the same regimen had a lower probability of subsequently achieving viral suppression and a higher chance of evolving HIV drug resistance. These results suggest that improving access to regular virologic monitoring is critically important, especially if NNRTI-based cART is to remain a preferred choice for first-line therapy in resource-limited settings. HIV treatment options for initial combination antiretroviral therapy (cART) for therapy naïve patients continue to expand, with more than 30 available drugs and co-formulations . The goaNNRTIs have been shown to be highly effective in achieving short-term virologic suppression. However, NNRTIs have a relatively low genetic barrier to the emergence of HIV drug resistance –8. As a Several studies have documented superior efficacy of NNRTIs versus bPIs based on rates of short-term viral suppression ,12–17. HStudy data from eligible patients were extracted from the BC Centre for Excellence in HIV/AIDS (BC-CfE)'s Drug Treatment Program (DTP) monitoring and evaluation system. Since October 1992, the distribution of antiretrovirals in BC has been the responsibility of the DTP. HIV medical care and laboratory monitoring are fully subsidized  to all people living with HIV/AIDS (PLWHA) residents of BC according to BC-CfE's HIV therapeutic guidelines, which have remained generally consistent with those put forward by the International AIDS Society-USA, currently known as the International Antiviral Society-USA (IAS-USA), since 1996 ,19.Eligible participants were cART naïve patients, ≥19 years old, enrolled between 1 January 2000 and 30 June 2013 and followed until no later than 30 June 2014. These patients started cART consisting of two nRTIs as backbone, plus either an NNRTI (efavirenz or nevirapine) or a bPI (ritonavir booster plus either lopinavir or atazanavir). They must also have had a CD4 count and pVL measurement within six months of the initial antiretroviral date. All pVL measurements in BC are centrally done at the St Paul's Hospital virology laboratory. Since the quantification range of pVL assays has evolved over time, for analytical purposes, we truncated our measurements to range from <50 (coded as 49) to >100,000  copies/mL . CD4 celResistant samples were assigned to one of four classes based on a modification of the 2014 IAS-USA list of mutations : 1) lamiMortality data for all causes were obtained through monthly linkages with the BC Vital Statistics Agency. Patients lost to follow-up were censored at the last contact date  prior to any of the following events: 1) if they moved outside of BC; 2) if last contact date was more than 18 months and before 31 December 2012; 3) if last contact date was less than 18 months and after 31 December 2012; 4) if they enrolled in a blinded trial involving receiving placebo medication; or 5) if they had a scheduled treatment interruption.For the purpose of the analyses in this study, patients were included if they did not change cART regimen during the study period, and they were required to have a follow-up of at least six months. Thus, we followed patients since cART initiation, and at six months following cART initiation, we assessed whether patients virologically failed therapy, defined by a pVL >50 copies/mL. Based on the pVL at failure, failing patients were stratified at pVL ≤500 versus >500 copies/mL. We continued to follow those patients who failed at six months for an additional six months, on the same cART regimen, and calculated their probability of achieving subsequent viral suppression defined as a pVL <50 copies/mL twice consecutively (yes or no) and of developing drug resistance to any cART class of medications they were currently taking (yes or no), defined by any combination of the above resistance classes. For analytical purposes, samples with pVL <250 copies/mL were classified as wild type virus. We restricted this analysis to include patients with no resistance prior to starting cART.These probabilities were modeled via a multivariable logistic regression explanatory model, separately for those who started on NNRTI-based and bPI-based cART. Possible explanatory variables for the viral suppression and resistance analyses at 12 months after cART initiation included age (continuous), gender , history of IDU , CD4 cell count at baseline and at six months (continuous), pVL at six months (≤500 or >500 copies/mL), period of cART initiation , adherence level measured between six and twelve months following cART initiation (<95% or ≥95%) and follow-up time (in months). We did not include pVL measured at baseline since it is highly collinear with pVL measured at six months; otherwise, it may bias our coefficient's standard errors. Adherence level, for each period, was defined as the number of days of antiretroviral drugs dispensed divided by the number of days of follow-up (expressed as percent). Estimates of adherence to antiretroviral therapy were based on the different regimen exposures for each patient. Although patients with undetectable pVL less commonly have medications changed, the regimens can change due to adverse effects or other medical and non-medical reasons, and we also accounted for those changes. Measuring adherence to any specific antiretroviral drug as index medication when it can be changed at any point in time makes it difficult for accurate monitoring. For this reason, the measure of adherence that we chose to adopt does not use any specific index medication, and it only measures the overall exposure to any antiretroviral regimen . Althougp-values, was used in the selection of explanatory variables [For building multivariable models, a modified backward stepwise technique, based on Akaike Information Criterion (AIC) and Type III ariables . Goodnesariables . Rates wariables . All anaThe BC-CFE received approval for this study from the University of British Columbia ethics review committee at the St Paul's Hospital, Providence Health Care site (H05–50123). The study complies with the BC's Freedom of Information and Protection of Privacy Act. The study was conducted using anonymized administrative databases, and therefore, informed consent was not required.10 copies/mL (Q1–Q3 4.36–5.00).Among 3925 individuals who started cART between 1 January 2000 and 30 June 2013 and were followed until 30 June 2014, 1963 (50%) individuals started therapy on an NNRTI-based regimen, 1962 (50%) individuals started therapy on a bPI-based regimen, 81% were males, 45% did not have a history of injection drug use at baseline, 31% started cART in 2000 to 2005, 38% in 2006 to 2009 and 31% in 2010 to 2013. At baseline, the median age was 42 years (25th–75th percentiles (Q1–Q3 35–49), median CD4 cell count was 220 (Q1–Q3 130–340) and median pVL was 4.88 logp <0.0001 for the comparison of these two rates). p <0.05).The number of patients who remained on the initial cART regimen within the first 12 months of follow-up was 3359 (86% of 3925). This consisted of 1616 (82% of 1963) in the NNRTI cohort and 1743 (89% of 1962) in the bPI cohort. At six months, we observed that 401  patients in the NNRTI cohort and 601  in the bPI cohort were classified as having virologic failure (i.e. a pVL >50 copies/mL) (p <0.0001)  at six months were more likely to have lower adherence (<95%) and they continued to have lower CD4 cell recovery (<0.0001) .≤500 copies/mL, were 85% lower than those for patients who failed with a pVL >500 copies/mL (0.2 vs. 1.3 cases per 100 person-months), while in the bPI cohort these rates were 67% lower (0.1 vs. 0.3 cases per 100 person-months). Furthermore, note that the resistance rates for those in the bPI cohort were significantly lower than the same rates for the NNRTI cohort for patients with a pVL >500 copies/mL at six months (p<0.0001), and these rates were in the same order of magnitude for both cohorts among patients with a pVL ≤500 copies/mL at six months .p <0.0001). However, if patients on NNRTI-based regimens failed therapy with a pVL >500 copies/mL at six months, they had a 20% lower probability of suppressing at 12 months than those on bPI (0.37 (0.29–0.45) vs. 0.46 (0.38–0.54); p <0.0001). In terms of resistance, those who failed on NNRTI performed worse than bPI in all scenarios, especially if they failed at six months with a pVL >500 copies/mL (0.40 (0.28–0.54) for NNRTI vs. 0.14 (0.09–0.20) for bPI; p <0.0001).Our results illustrate the different impact of remaining on a virologically failing regimen depending on whether it was an NNRTI- or a bPI-based cART. Patients who failed with a high pVL and continued on the first-line NNRTI regimen had a lower probability than those on bPI with regard to the likelihood of subsequently achieving viral suppression, and a higher chance of experiencing evolving HIV drug resistance, in all scenarios. Our results, therefore, suggest that in the absence of pVL monitoring, using NNRTI-based cART regimens as first-line will result in an unintended excess HIV drug resistance prevalence and compromise second-line therapy regimens. Also of concern, the NNRTI-resistant variants have a relatively good fitness and, thus, are likely to be the source of transmitted HIV drug resistance .These results are particularly relevant as we move towards the implementation of the 2015 WHO guidelines recommending universal access to cART for all PLWH regardless of CD4 cell count levels, globally . These gUnfortunately, routine pVL monitoring remains unavailable in many resource-limited settings ,25. ThisThere are some features of this study that are worth mentioning. The study data were based on patient data within a population-based HIV treatment programme with free access to medical care, antiretroviral therapy and laboratory monitoring. Second, estimates of adherence (pharmacy refill record) to antiretroviral therapy were based on the medication exposure of each patient, which is the maximum adherence an individual might have during a period of time. Currently, there is no gold standard to measure adherence to cART. However, this type of adherence measure was found to be highly predictive of disease progression and death, among other key clinical outcomes . Third, Our results showed that NNRTI-based cART had superior virologic short-term effectiveness than bPI-based cART. In contrast, among those patients who failed with a high pVL (≥500 copies/mL) at six months, NNRTI starters were less likely to achieve subsequent virologic suppression and had a higher probability of developing HIV drug resistance. These results highlight the critical and urgent need to secure global access to pVL monitoring.


Both central (CNS) and peripheral nervous system (PNS) complications are frequent and varied in connective tissue diseases. A systematic review was conducted between 1989 and 2014 in the databases Medline, Scopus, and Cochrane Library using the search terms, peripheral and central nervous complications and immunological profiles, to identify studies in specific connective tissue disorders such as rheumatoid arthritis, systemic lupus erythematosus, and primary Sjögren's syndrome. A total of 675 references were identified, of which 118 were selected for detailed analysis and 22 were included in the final review with a total of 2338 participants. Our search focused only on studies upon connective tissue disorders such as rheumatoid arthritis, systemic lupus erythematosus, and primary Sjögren's syndrome associated with seroimmunological data. The reported prevalence of CNS involvement ranges from 9 to 92% across the reported studies. However, the association between CNS and PNS manifestations and seroimmunological profiles remains controversial. Τo date, no laboratory test has been shown as pathognomonic neither for CNS nor for PNS involvement. Connective tissue disorders are chronic inflammatory autoimmune diseases driven by an antibody or T-cell response directed against a self-antigen antibody affecting muscle, joints, and skin such as rheumatoid arthritis (RΑ), systemic lupus erythematosus (SLE), primary Sjögren's syndrome (pSS), and systemic sclerosis –3. Both To our knowledge this is the first systematic review focused in both most common CNS and PNS complications of RA, SLE, and pSS emphasizing the associated immunological features of these specific connective tissue.This is a systematic review of literature, based on the PRISMA guidelines. We search for relevant studies in English in the following databases, from the emergence of the condition to October 2014: Medline (from 1989 to 2014), Scopus (from 1983 to 2012), and Cochrane Library (from 1993 to 2014). The keywords used for the study were the following: “peripheral nervous and central nervous manifestations,” “myositis,” “cranial neuropathy,” “mononeuropathy,” “polyneuropathy,” “myelopathy,” “myelitis,” “multiple sclerosis,” “Neuromyelitis optica spectrum (NMO),” “headaches,” “seizures,” “psychosis,” “depression,” “connective tissue disorders,” “rheumatoid arthritis, systemic lupus erythematosus, primary Sjögren's syndrome,” “Immunological profiles,” “ANA,” “anti-ribosomal P antibodies,” “Antiphospholipid antibodies,” “SSA/SSB,” “RF,” “ACA,” “cryoglobulins,” “A-ds DNA,” and “AQ4 antibody”.Articles not published in English, letters, summaries, dissertations, theses, and case reports were excluded, as well as studies that used children or animal models and studies that were not related to central and peripheral nervous complications of RA, SLE, and pSS. Studies were included only if diagnosis of neurological complications was made by expert neurologists. Only studies with neurological complications primary or secondary to RA, SLE, and PSS with associated immunological features were included. Immunological tests  were included if they were performed at the time of diagnosis and at least yearly during follow-up.The described search identified six hundred seventy-five relevant studies , 107 of We performed qualitative data synthesis, organizing the results by connective tissue type. We did not attempt to perform meta-analysis because of the heterogeneity of the study designs, populations, and results. Using our critical appraisal of individual studies and the body of evidence for each study design, we identified strengths and weaknesses of each study in the discussion. We did not assess publication bias. In 1989, Mellgren et al.  reviewedIn Andonopoulos et al.'s study , 10 of tAmong 33 RA patients, Sivri and Güler-Uysal  found 2 Kasitanon et al.  studied n = 8) and psychosis (n = 29) was 88% and 45%, respectively, compared with only 9% in patients with nonpsychiatric neurologic disease (n = 45). For the entire SLE group, the odds ratio for the association of anti-P antibodies and severe psychiatric manifestations was 7.63 with a 95% confidence interval of 3.61 to 16.14.In Schneebaum et al. study , 19% of Isshi and Hirohata  also anaχ2 = 6.04, P < 0.05). These antibodies are associated with active SLE and CNS involvement particularly in patients without anticardiolipin antibodies.In Tzioufas et al. study , serum wIn Sanna et al. study , 185 SLEP = 0.0028) or with focal NPSLE (P = 0.0008).In Hirohata et al. study , 23 patiIn Závada et al. study , sera ofHanly et al.  identifin = 1), organic brain syndrome (n = 2), cranial nerve disorder (n = 1), and headache (n = 2). Significantly higher levels of both fractions of ACA and IgM ACA and of both fractions of ACA and IgM ACA and antiphospholipid antibodies (repeatedly positive test for serum ACA and/or lupus anticoagulant) were determined in patients with severe disease activity and microangiopathic complications compared with those with less active disease.According to 1997 American College of Rheumatology diagnostic criteria , 15 womeP = 0.017) and PNS mononeuropathy multiplex (P = 0.040).Briani et al.  presenteGono et al.  enrolledAmong 207 SLE patients Florica et al.  showed tP = 0.02). The most common PN observed was peripheral polyneuropathy, with a predominance of the sensory form, present in 76 (36.7%) patients and the sensory-motor variant in 39 (18.8%) patients. Twenty-three (11.1%) patients had a peripheral mononeuropathy and 26 (12.5%) had a cranial neuropathy. Nineteen (9.2%) patients suffered from mononeuritis multiplex and only 11 (5.3%) were diagnosed with chronic inflammatory demyelinating polyradiculoneuropathy (CIDP). There was no difference between PNS features and SLE immune markers (presence of antiphospholipid antibodies).Patients with PNS were more likely to have also CNS involvement (14.2%) compared with patients without PN . This subgroup had more frequent cryoglobulinemia and lymphopenia (P < 0.05) but lower prevalence of anti-Ro/SSA antibodies and hypergammaglobulinemia (P < 0.05). Authors also confirmed cryoglobulinemia as unique predictive factor of PNS disease.In 2014 study by Jamilloux et al. , 420 patIn Spezialetti et al. study , from 77P = 0.007), more frequent CNS involvement , and lower prevalence of ANA , anti-SSA (Ro) , anti-SSB (La) , RF , and hypergammaglobulinemia . In multivariate analysis, nonataxic sensory neuropathy was associated with the presence of CNS involvement  and ANA .In Sène et al. study , patientn = 12), chronic myelopathy (n = 16), or motor neuron disease (n = 1)). The disease mimicked relapsing-remitting multiple sclerosis (MS) in 10 patients and primary progressive MS in 13 patients. CNS symptoms included seizures (n = 7), cognitive dysfunction (n = 9), and encephalopathy (n = 2). Fifty-one patients had PNS involvement: symmetric axonal sensorimotor polyneuropathy with a predominance of sensory symptoms or pure sensory neuropathy occurred most frequently (n = 28), followed by cranial nerve involvement affecting trigeminal, facial, or cochlear nerves (n = 16). Multiple mononeuropathy (n = 7), myositis (n = 2), and polyradiculoneuropathy (n = 1) were also observed. Anti-Ro/SSA or anti-La/SSB antibodies were detected in 21% of patients at the diagnosis of SS, more frequently observed in patients with PNS involvement than in those with CNS involvement (P < 0.01).In Delalande et al. study , fifty-sAlexander et al.  examinedP = 0.59). SSA, SSA, ANA, and dsDNA antibodies were found in these patients but not NMO-IgG.Pittock et al.  detected RA, SLE, and pSS, our results indicate that CNS complications (9–92%) are much more frequent than those of PNS (8–66%) across the various studies . These findings are compatible with previous results in the literature. In this review, the majority of reported studies attributable in SLE CNS manifestations range from 13 to 92% of the events compared to PNS in 8–56%. The most frequent NP syndromes were headache, mood disorders, and seizures which are consistent with previous meta-analysis [In this first systematic review of both CNS and PNS involvement inanalysis , 35. In analysis . In thisanalysis , 38.Overall prevalence is seen to vary widely among the studies and is mainly attributed to a number of factors including bias in selection of patients for study, disease duration, and lack of uniformity in diagnostic criteria. With this in mind, we sought to expand the probability of specific manifestations by using established criteria and double-reading (by a certified neurologist), and excluding uncertain cases.Among the most interesting points of this review is the correlation between CNS/PNS manifestations and immune activation markers . Anti-Ro and anti-La seem to be less frequent in pSS patients with neurological involvement (40%) compared to patients without neurological manifestations (60% of positivity), so, it is necessary for more markers are necessary in pSS to better classify subpopulations of patients with neurological involvement . JamilloSeveral specific autoantibodies are associated with NPSLE . The presence of antiphospholipid antibodies is significantly associated with cerebrovascular disease and cognitive dysfunction, whereas serum anti-ribosomal P antibodies are strongly associated with psychosis and depression in SLE as discussed in this review , 13, 20.However, Andonopoulos et al. failed to find a statistically significant correlation between the observed PNS pathology and any other clinical or immuno/serologic data. In the absence of any obvious reason, it is tempting to ascribe PNS symptoms to vasculitis of the vasa nervorum, but the data provided cannot fully support this hypothesis. The classical severe picture of a patient presenting with mononeuritis multiplex, associated with vasculitis in other organs, as already mentioned, leaves little doubt as to the pathogenesis of the neurologic syndrome. However, no patient has been reported to exhibit additional features of CNS disease that could be clinically detected.Similarly, Sivri and Güler-Uysal found no correlation between mononeuritis and the immunological data in the serum of RA patients , which is in contrast with the finding by other study . This coSeveral limitations which compromised their external validity were identified in the specific studies, mainly by their authors. In terms of reporting complications, methods varied considerably. In this review, incidence was determined from studies of varying sample sizes across multiple age groups. Data collection was retrospective , 29, 32.Notwithstanding, this review is not without its limitations. There are a small number of studies available for analysis. One limitation of this review is that a limited number of databases were searched. Consequently, there is always the possibility that we overlooked studies in other databases published before 1980. In addition, there is always the possibility of publication bias due to underreported negative results and grey literature. However, the use of fewer search limits increases the sensitivity of the search method.Despite the abovementioned limitations, this systematic review provides foundation for further research on the pathophysiology of immune system among the vasculature of the CNS and PNS. It is still a matter of debate whether the CNS and PNS manifestations of RA, SLE, and pSS are direct effect of these diseases or secondary such as side effects of corticosteroids and other immunosuppressive therapies or multiple systemic organ dysfunction . Τo date, no laboratory test, that is, autoantibodies, has been shown as pathognomonic neither for CNS nor for PNS involvement in these connective tissue disorders. Prospective multicenter studies are warrant for further confirmations.


Neospora caninum, Trypanosoma cruzi, Babesia canis, and Hepatozoon canis. However, although cardiac disease due to Leishmania infection has also been documented, the immunopathological features of myocarditis have not been reported so far. The aim of this study was to examine the types of cellular infiltrates and expression of MHC classes I and II in myocardial samples obtained at necropsy from 15 dogs with an established intravitam diagnosis of visceral leishmaniasis. Pathological features of myocardium were characterized by hyaline degeneration of cardiomyocytes, necrosis, and infiltration of mononuclear inflammatory cells consisting of lymphocytes and macrophages, sometimes with perivascular pattern; fibrosis was also present in various degrees. Immunophenotyping of inflammatory cells was performed by immunohistochemistry on cryostat sections obtained from the heart of the infected dogs. The predominant leukocyte population was CD8+ with a fewer number of CD4+ cells. Many cardiomyocytes expressed MHC classes I and II on the sarcolemma. Leishmania amastigote forms were not detected within macrophages or any other cell of the examined samples. Our study provided evidence that myocarditis in canine visceral leishmaniasis might be related to immunological alterations associated with Leishmania infection.Myocarditis associated with infectious diseases may occur in dogs, including those caused by the protozoa Leishmania spp. [ Leishmania infantum (syn: L. chagasi in the New World) [Canine leishmaniasis is a zoonotic disease caused by the protozoan parasitenia spp. . The disnia spp. , 2. The w World) , 4. Evenw World) , 6. CVL w World) . Enlargew World) , 9, wherw World) , 11. Staphylococcus, Streptococcus, Citrobacter, Bartonella, and Borrelia), viruses , fungi , and protozoa  [ L. infantum infection in dogs.Canine myocarditis is a rarely diagnosed disease which can be caused by noninfectious  or infectious agents such as bacteria  –16. DepeBabesia) . The aim Leishmania in southern Italy, were selected for the study. All dogs had an established intravitam diagnosis of leishmaniasis by serological and parasitological methods [Fifteen crossbreed dogs, 8 males and 7 females, aged 7 to 11 years, living in an endemic area for methods . MoreoveEhrlichia canis, Toxoplasma gondii, Babesia canis, Rickettsia rickettsii, Leptospira, Borrelia burgdorferi, and Neospora caninum) and did not show any other clinical signs of heart failure.The dogs were serologically negative for the main infectious agents responsible of myocarditis , were collected from the right atrium, ventricular free walls, and the interventricular septum as described by Rosa et al. . Samplesμm thick) were stained with hematoxylin and eosin (H&E) for histopathological examination to assess a definitive diagnosis of myocarditis.Cryostat sections (5 μm thick), dried at room temperature for 1 hour, fixed in acetone at 4°C for 5 minutes, and then blocked for endogenous peroxidase in 0.3% H2O2 in methanol solution for 20 minutes. Sections were incubated overnight at 4°C with primary antibodies against canine leukocyte antigens diluted in 0.01 M phosphate-buffered saline (PBS), pH 7.2–7.4, as follows:CD3  diluted 1 : 50.CD4  diluted 1 : 50.α, CD8β  diluted 1 : 50.CD8α  diluted 1 : 50.CD79MHC I  diluted 1 : 100.MHC II  diluted 1 : 50.Slides were washed with PBS, then incubated with biotinylated secondary antibody, and labeled with streptavidin biotin for 30 minutes at room temperature, followed by incubation with streptavidin conjugated to horseradish peroxidase . The reaction was revealed by diaminobenzidine treatment  and finally, sections were counterstained with Mayer's haematoxylin. In the negative control sections, the primary antibody was either omitted or replaced with normal serum.Immunohistochemical examination was carried out as previously described . In brieApproximately 20 fields at 20x magnification were evaluated for each section by two independent pathologists  with a concordance rate of 97%. 0 (not detected). 1 (percentage of immunoreactive inflammatory cells per section 1–25%). 2 (percentage of immunoreactive inflammatory cells per section 26–50%). 3 (percentage of immunoreactive inflammatory cells per section >50%).The inflammatory cell immunoreactions were scored as follows:The relationship between the different types of infiltrating immune cells and MHC class I expression was evaluated using Spearman's Rho correlation (Past 3.10 software).Necropsies were performed for diagnostic purposes after receiving the consent of the owner. Each owner consented to the use of tissues for research purposes, according to the internal rules of the Diagnostic Service of Pathology and Animal Health of the University of Naples Federico II. All the procedures were performed for diagnostic purpose; thus the study did not require any consent or ethical approval according to the European Directive 2010/63/EU. L. infantum, showed neither morphologic alterations nor inflammatory cells infiltrates. MHC classes I and II were expressed by endothelial cells of arterioles, venules, and capillaries.Myocardial samples from healthy control dogs, serologically and parasitologically negative for L. infantum amastigotes were detected within macrophages.At H&E stain all affected dogs (100% of studied cases) showed variable numbers of mononuclear cells, represented by lymphocytes and some macrophages, sometimes in a perivascular pattern . In some cases  a nonsuppurative granulomatous myocarditis, characterized by severe interstitial infiltration of mononuclear cells, was identified . Inflammα+ cells rarely detected within the inflammatory infiltrates. Vascular adventitia, endothelial cells, and cellular infiltrates within the myocardium stained intensely for MHC classes I and MHC class II antigens. In addition, many cardiomyocytes had MHC class I .A positive relationship was observed between the CD8 positive cells and MHC class I expression , and the immune response of the host [ L. infantum induces a mixed Th1 and Th2 response in CVL and the control of parasite replication, disease progression, or cure are strictly associated with the balance of these two patterns of immune system reaction [γ-interferon, IL-2, and TNF-α) promoting macrophage anti-Leishmania activity through nitric oxide production that is responsible for parasite killing by apoptosis [ Leishmania amastigotes may also be lysed by CD8+ cytotoxic T lymphocytes even if this mechanism may be suppressed by the presence of high parasitic load [ L. infantum infection and this mechanism could be at the basis of myocardial injury and it was already demonstrated in canine inflammatory myopathy associated with L. infantum infection [The predominant inflammatory infiltrate cell types were CD8+ T lymphocytes and macrophages; macrophages were distinguished from lymphocytes, at light microscopy, by morphological features and they were MCH immunoreactives. However, CD4+ T cells were also found. It is now well accepted that the progression ofthe host , 23. Thereaction . The propoptosis , 25; mortic load , 27. Contic load –32. The nfection . L. infantum infection [ L. infantum infection associated myositis in dogs [Detection of MHC classes I and II expression in cardiomyocytes was a common finding in the majority of samples from infected dogs . Immunohnfection . Our dat in dogs . Further in dogs , 34, 37. L. infantum infection in dog. Our study provides evidence that during leishmaniasis myocarditis can occur with morphological and immunophenotypical pattern superimposable to canine myositis associated with L. infantum infection. Finally, our results, if confirmed on larger scale, could be used to improve therapeutic protocols for the management of dogs affected by leishmaniasis or to address the research towards new drugs useful to modulate immune-system response in order to reduce myocardial inflammation.Our data provided an initial antigenic characterization of infiltrating mononuclear cells and MHC classes I and II expression in myocarditis associated with


Panthera pardus) model system. We apply Bayesian SCR methods to 89 simulated datasets and camera-trapping data from 22 leopards captured 82 times during winter 2010–2011 in Royal Manas National Park, Bhutan. We show that sample interval length from daily, weekly, monthly or quarterly periods did not appreciably affect median abundance or density, but did influence precision. We observed the largest gains in precision when moving from quarterly to shorter intervals. We therefore recommend daily sampling intervals for monitoring rare or elusive species where practicable, but note that monthly or quarterly sample periods can have similar informative value. We further develop a novel application of Bayes factors to select models where multiple ecological factors are integrated into density estimation. Our simulations demonstrate that these methods can help identify the “true” explanatory mechanisms underlying the data. Using this method, we found strong evidence for sex-specific movement distributions in leopards, suggesting that sexual patterns of space-use influence density. This model estimated a density of 10.0 leopards/100 km2 , comparable to contemporary estimates in Asia. These SCR methods provide a guide to monitor and observe the effect of management interventions on leopards and other species of conservation interest.Many large carnivores occupy a wide geographic distribution, and face threats from habitat loss and fragmentation, poaching, prey depletion, and human wildlife-conflicts. Conservation requires robust techniques for estimating population densities and trends, but the elusive nature and low densities of many large carnivores make them difficult to detect. Spatial capture-recapture (SCR) models provide a means for handling imperfect detectability, while linking population estimates to individual movement patterns to provide more accurate estimates than standard approaches. Within this framework, we investigate the effect of different sample interval lengths on density estimates, using simulations and a common leopard ( Across the globe, large carnivores face threats from habitat loss and fragmentation, prey depletion, and poaching . These rTo address the imperfect detectability of large carnivores, researchers have adopted non-invasive, remote sampling techniques, such as automatically triggered camera traps, hair snares or scat surveys that likely reduce animal avoidance of target sites. For many species, these data can identify unique individuals based on genetics or unique markings, facilitating a capture-mark-recapture (CMR) analysis that explicitly incorporates detection probability into the estimate of abundance and/or density. Spatial capture-recapture (SCR) models further refine these techniques, by incorporating the spatial distribution of an individual’s movement inferred from observed captures , 14, whiet al. [et al. [et al. [et al. [Despite these advances, SCR methods raise new questions about sampling design and its relationship to population estimates and precision, particularly as these techniques are applied to new species and systems, including large carnivores. Russell et al. , Sollman [et al.  and Sun  [et al.  examine  [et al.  consider [et al.  address  [et al. , 25, des [et al. , 26. WhiBeyond sampling structure, the ability to incorporate covariates into density estimates raises important questions about model selection, particularly when estimates disagree. While maximum likelihood-based SCR estimation techniques have adopted an Akaike information criterion (AIC) model selection framework, no robust methods have been widely applied in Bayesian approaches to SCR analysis. These limitations in assessing the relative merits of different models restrict the usefulness of these methods, especially for data-poor species like large carnivores.Panthera pardus) from the lower, subtropical foothills of Royal Manas National Park (RMNP), Bhutan. The common leopard occurs from sub-Saharan Africa to the Russian Far East, as well as on the islands of Sri Lanka and Java [Here, we develop two methodological advances that may improve sampling design and analysis of capture-recapture studies for a variety of taxa: (1) we examine the impact of the temporal resolution of the data on SCR model estimates and precision and (2) we implement a Bayes factor approach to model selection with multiple covariates. We address these issues through a simulation study and using a remote camera-trapping data set for the common leopard  and borders India’s Manas Tiger Reserve, forming a trans-boundary conservation landscape. RMNP covers 1057 km2, with elevations from 90 m in the southern foothills to 2900 m in the north (Cervus unicolor), barking deer (Muntiacus muntjak), wild pigs (Sus scrofa), serow (Capricornis thar), goral , Himalayan crestless porcupine (Hystrix brachyuran), langurs (Trachypithecus spp), macaques (Macaca spp.) and several bird species.RMNP is located in the southern foothills of Bhutan  conducted this study in collaboration with the University of Montana. UWICE is a governmental research institute with the mandate to conduct studies in the protected areas of Bhutan. Since we did not handle any animals and used non-invasive, remote camera traps, no animal handling, care nor ethics permits were necessary.2, although international and topographic boundaries made some irregular  to change batteries and memory cards. We identified individuals based on unique pelage patterns and determined the sex of these individuals as possible from sex-specific cues, such as visible genitalia or the presence of young. We performed this identification visually without the aid of any pattern-recognition software. A single author (TT) verified all leopard identifications to minimize any observer effects.et al. [yijk, of individual i in trap j during sampling interval k to the latent distribution of activity centers. We let the observation, yijk, take the value of one for a capture and zero if not captured to produce a capture history for all individuals in all traps over all sampling intervals. We treated multiple detections of an individual in a particular trap during the same sampling interval as a single capture. Individuals could be captured in multiple traps during a sampling interval. We followed the formulation of the observation process used by Gardner et al. [et al. [i.e., 1 ≤ θ ≤ 2, where θ represents the exponent of the kernel from the exponential family of distributions).We estimated the abundance and density of leopards within our study area using a spatial capture-recapture model , 23, 34.et al.  for a car et al.  and Russsex), and (4) a model with sex as a covariate on baseline detectability and the scale of the movement distribution (Sex + σsex). We fit these models using an area that buffered the trapping grid by 10 km, yielding a state space of 1,551 km2. This buffer incorporates individuals with activity centers outside of the trapping grid, but whose movement range extends into the trapping grid, allowing for a robust estimate of density.We fit a suite of 16 models to the data to assess the impact of varying the duration of sampling intervals and different covariates. We used 4 different sampling interval durations, corresponding to daily (91 sampling occasions), weekly (13 sampling occasions), monthly (three sampling occasions) and quarterly (one sampling occasion) sample periods. For each of these durations, we fit four different covariate combinations: (1) a model where detectability depended only on distance between a trap and the individual’s activity center (Distance), (2) a model with sex as a covariate on baseline detectability (Sex), (3) a model with sex as a covariate on the scale of the movement distribution  methods in R , using tThe flexibility of SCR models to incorporate covariates at different levels allows estimates to include added complexity, but also presents ecologists with potential model selection problems when using Bayesian methods. Estimates and precision may vary among different models creating uncertainty regarding the population status and preventing insight into the mechanisms generating patterns of abundance , 23, 40.l for l = 0,…,3, for the data, y. From Bayes’s theorem, the relative posterior probabilities among the models in the candidate set are given byy|Ml) is the likelihood of the data given the model and Pr(Ml) is the prior probability of a model being correct. For our evaluation, we chose an uninformative prior and set Pr(Ml) = 0.25 for all models. Under this assumption, Bayes’s theorem then reduces to the proportion of the total likelihood attributed to a particular model. Gelfand and Dey [f(·) is a probability density function of the same dimension as ωl, the parameters in the model, and ωl(i) is a sample from the posterior distribution for a particular iteration of the MCMC, i = 1,…, c. For our models, c took the value of 7,500, the number of posterior samples. The denominator is the likelihood multiplied by the density at the value of the parameters for any given iteration of the chain. We chose f(·)~t7500, we usedl and M0 [We assessed the support of the additional sex-specific parameters for detectability and scale of the movement distribution using Bayes factors , 45. To l and M0 . The Bay2. Our simulations exposed these individuals to an 8 x 8 trap grid centered within the statespace for 90 (daily) sampling occasions. We generated capture histories using a Binomial observation model with a half-normal hazard rate detection function and sex-specific baseline detection probabilities, i.e. the Sex model. To parameterize the data generation, we used estimates from models fit to our field data. We set σ = 1 unit, λ0, female = 0.05 and βsex = -1.61. For each simulation, we then subsampled the data to have 1 (quarterly), 3 (monthly) and 13 (weekly) sampling occasions. Finally, we fit the Distance, Sex, σsex, and Sex + σsex models to all temporal resolutions of each simulated dataset. To fit these models, we discretized the statespace into an 11.5 x 11.5 grid of points and fixed θ = 1 . We ran all models with 5,000 MCMC iterations, discarding the first 1,000 iterations as burn-in. We use these results to assess the accuracy and precision of model estimates and the fidelity of our Bayesian model selection methods.We conducted a simulation study to corroborate our findings with respect to the effects of sampling interval on estimated density and precision, and test our model selection techniques. We simulated 89 SCR datasets for a population of size N = 250  distributed over an 11 x 11 unit continuous statespace for a density of 1.98 individuals/unitWe captured 22 individual common leopards a total of 82 times between 17 November 2010 and 15 February 2011 in 2,639 camera-trapping nights. The median number of captures per individual was 2 (range: 1–16 captures) and individuals were captured in between one and eight different traps in daily sampling opportunities. We identified six of these individuals as males and the remainder (16 leopards) as female based on photographic evidence of genetalia and/or offspring. We considered camera stations to have operated continuously for the duration of the study, since both cameras at a trap location never malfunctioned simultaneously.2 in the Distance-only model with monthly sampling periods to 15.8 leopards/100 km2 in the sex + σsex model with a single, quarterly sampling interval  and sex-specific covariate on this parameter (βsex). Shorter sampling intervals had lower baseline detection probabilities   . This resex model received 0.971 of the posterior model probability among the four candidate model parameterizations. The Bayes factor comparison to the distance model showed strong support for this model, as it was over 33 times as likely as the distance model and had positive values of log10Bl0 and 2 ln Bl0 (2 (95% CI of 6.25–15.93 leopards/100 km2)  , which cThe simulated datasets represented capture histories for a median of 223 unique individuals (95% CI: 212.0–232.6) recorded a median of 2479 times (95% CI: 2266.0–2752.2) over 90 sampling occasions, 2325 times (95% CI: 2111.8–2569.0) over 13 sampling occasions, 1864 times (95% CI: 1703.2–2053.0) over 3 sampling occasions and 1255 times (95% CI: 1142.8–1360.8) over a single sampling interval. On average, capture events recorded 144 female (95% CI: 138.2–148.0) and 79 male individuals (95% CI: 70.4–86.0).2 with the Distance model, using either daily or weekly sampling periods, to a maximum of 2.35 individuals/unit2 with the Sex + σsex model with a quarterly sampling period   . Precisitervals) .sex models ranged from 0 to 1. However, the Distance and σsex model received relatively little support across all simulations and sampling intervals with the σsex model identified as the top model in 2.2% of simulation replicates with quarterly sampling intervals.The Sex model received the greatest support across all simulations for all sampling intervals. The Bayes factors model selection process identified the Sex model as having the highest posterior probability at a minimum of 60.7% of simulations with a weekly sampling interval and as much as 71.9% of simulations with quarterly sampling intervals . As the We provide two advances in the implementation and use of SCR models through a simulation study and our analysis of the common leopard population in the lower foothills RMNP, Bhutan. First, our investigation of different sample interval lengths provides some guidance to reduce the variability of SCR population estimates, particularly for difficult to detect species, like large carnivores. Second, we apply Bayes factors in a novel context to perform SCR model selection. These results suggest new insights and avenues for future development of these methods within population ecology.Sample interval length affected the precision of density estimates in our SCR analysis of the leopard data and our simulation study. For the common leopard, we found that analyzing our data with daily sample intervals provided the best precision for all models, especially those with covariates ; Fig 2. These results suggest some guidelines for the implementation of SCR models. For camera trapping surveys of elusive species, where samples have accurate time signatures and limited captures take place, we recommend using daily  subdivisions of the data , 46. Whisex model  represented a clear runner-up across the four different sampling intervals. Moreover, our selection methods more frequently identified the Sex + σsex model as most probable with weekly, and to a lesser extent monthly and daily sampling intervals  and was over 33 times more likely than the null, Distance-only model  sampling interval, median density estimates varied from 4.5 in the Distance model to 11.0 in the Sex + σex model . Thus, mly model . We estily model , which idel . Furthermore, our model selection techniques gave additional insight into leopard behavior in our system and improved our density estimate.The support for sex-specific parameters in our leopard example highlights the need to consider relevant behavioral variation in estimating abundance and/or density for large carnivores. Previous leopard radio tracking , 63 and lap zero . Althoug2 (95% CI: 6.25–15.93 leopards/100 km2) was within the reported range of leopard densities, from 1.0–28.9 leopards/100 km2 [et al. [2, similar to our estimate for the Distance model of 4.5 leopards/100 km2 (95% CI: 2.90–6.64). However, they did not consider sex-specific covariates, which we found to increase density in our study. The Bayes factor model selection approach demonstrated strong support for sex-specific movement distributions suggesting the SCR-based estimate of Borah et al. [A comparison of our estimate to other study areas from the Indian sub-continent demonstrates the importance of considering animal ecology in SCR models , 64, 65. [et al.  reportedh et al.  may be bh et al. , 23, 65.et al. [2 (95% CI: 4.54–17.98) to 1.44 leopards/100 km2 (95% CI: 0.41–6.61), corresponding to an increase in tiger density across years. These contrasting changes in density across an entire study area raise the possibility that conspecific interference or avoidance may affect patterns of abundance or movement within a study at finer spatial and/or temporal scales. These interactions could be integrated into SCR analyses by introducing the presence or abundance of other species as spatial covariates or through multi-species models. More broadly, SCR methods have been integrated with concepts of resource selection and ecological distance to provide inference on factors affecting habitat use and animal movements [Looking towards the future, we see a great deal of promise in SCR models incorporating auxiliary ecological information. Our estimate may have been further improved by incorporating a metric of habitat quality , althouget al.  found thovements , 67. TheMany large carnivores, including leopards, have a broad geographic distribution, but increasingly face threats from habitat disturbance, poaching, inadequate prey resources and human-wildlife conflicts. Despite the many risks facing large carnivores, the status of populations across species’ ranges remains poorly understood. SCR methods hold a great deal of promise to help fill this critical gap of knowledge for large carnivores by explicitly incorporating spatial heterogeneity in detections into our estimates of abundance. These methods yield the greatest precision when combined with a high temporal resolution of sampling , although more modest weekly or monthly sampling intervals can be informative, particularly with high capture rates. Moreover, we implement and validate a Bayesian model selection method for use with these models. These methodological advancements may contribute to population assessments, help prioritize conservation efforts and improve understanding of large carnivore ecology as it relates to population biology. These steps towards a more holistic view of conservation may pay dividends for large carnivores and threatened species globally.S1 Filehttp://dx.doi.org/10.5061/dryad.mr1pt). This data is used to fit four spatial capture-recapture models with the SCRbayes package for R . Finally, we conduct model selection with Bayes factors. Users should consult the package help files and additional resources for further information.R code to implement spatial capture recapture models with a novel approach to model selection using the SCRbayes package. This script reads the data for the camera trap locations and leopard capture histories available through the Dryad Database ((DOCX)Click here for additional data file.S2 Filehttps://sites.google.com/site/spatialcapturerecapture/scrbayes-r-package. This code comes as a function (SCR.bf) within the package. Users should consult the package help files and additional resources for further information.This function within the SCRbayes package will operate on standard model output from SCRj.fn to calculate posterior marginal likelihoods and Bayes factors from spatial capture-recapture models implemented in the SCRbayes package. The SCRbayes package can be accessed at: (DOCX)Click here for additional data file.S1 Table0 gives the baseline capture probability at an individual’s activity center per sample interval per camera station. βsex denotes the effect of sex on detection probability on the log scale. The σ parameters describe the scale of an individual’s movement distribution in km, which varies by sex in some models. ψsex estimates the proportion of the population that is male. θ represents the shape parameter of the individual’s movement distribution, where 0.5 is exponential and 1.0 is Gaussian.Median parameter estimates with 95% credibility intervals in parentheses from spatially explicit capture-recapture models of common leopards in Royal Manas National Park during 2010–2011 for all 16 combinations of covariates and data. λ(DOCX)Click here for additional data file.S2 Table0 gives the baseline capture probability at an individual’s activity center per sample interval per camera station. βsex denotes the effect of sex on detection probability on the log scale. The σ parameters describe the scale of an individual’s movement distribution in units, which varies by sex in some models. ψsex estimates the proportion of the population that is male. True values used to generate the simulated were: λ0 = 0.05 , βsex = -1.61, σmale = σfemale = 1.0, ψsex = 0.4.Median of median model parameter estimates with median 95% credibility intervals in parentheses from spatially explicit capture-recapture models fit to all simulated datasets for all 16 combinations of covariates and data. λ(DOCX)Click here for additional data file.


Objective. To systematically review the efficacy and potential immunomodulatory effect of ulinastatin combined with thymosin α1 (UTI) for sepsis. Design. A systematic review and meta-analysis of randomized controlled trials (RCTs). Data Sources. The following databases: PubMed, Embase, and Cochrane Central were searched to identify related clinical trials. The search terms were “ulinastatin”, “thymosin”, and “sepsis”. Results. Six RCTs, 944 septic patients in total, were included in this meta-analysis. The result shows UTI increased the 28-day survival rate of septic patients, odds ratio (OR) = 2.01, 95% CI . After the treatment with UTI, the APACHE II score (four studies) dropped 4.72 further, mean = −4.72, 95% CI  (p < 0.00001). The mean time of ICU stay (four studies) in UTI group decreased 3.03 days further, mean = −3.03  (p = 0.14), and mechanical ventilation time (four studies) decreased 2.05 days, mean = −1.81  (p = 0.002). With the treatment of UTI, CD4+T cells raised 5.13%, mean = 5.13, 95% CI  (p < 0.0001); there was no significant change in CD8+T cells, mean = −0.74  (p = 0.51). Conclusion. According to this meta-analysis, with the treatment of UTI, the short-term survival rate of septic patients was increased and the illness severity was alleviated. ICU stay and mechanical ventilation time were effectively shortened. The beneficial effect of UTI might be due to the potential immunomodulatory effects of these two drugs. Systemic illness caused by microbial invasion of normally sterile parts of the body is referred to as “sepsis,” which is an increasingly common cause of morbidity and mortality, particularly in the elderly, immunocompromised, and critically ill patients , 2. SepsAfter numerous unsuccessful trials of anti-inflammatory agents in patients with sepsis through the years, investigators doubted whether the mortality could be decreased. For although bacteria play a leading role in sepsis, the direct cause of death for patients with sepsis is not the bacteria themselves but rather the persistent paralysis of the immune system and the excessively magnified systemic inflammatory response.α1, calculating that these 2 reagents might prevent the inflammation spreading too fiercely while simultaneously enhancing the body's immunity to avoid immune paralysis. Ulinastatin is a multivalent, Kunitz type serine protease inhibitor that is found in human urine and blood ) OR septic shock[Title/Abstract]) OR infection[Title/Abstract]) OR infectious disease[Title/Abstract])) AND (((ulinastatin) OR trypsin inhibitor) OR thymosin)) AND ((((((RCT[Title/Abstract]) OR randomized controlled trial[Title/Abstract]) OR randomized[Title/Abstract]) OR controlled[Title/Abstract]) OR placebo[Title/Abstract]) OR sham[Title/Abstract])) AND  OR death rate[Title/Abstract]) OR survival rate[Title/Abstract]) OR efficacy[Title/Abstract])” in PubMed and it is adjusted accordingly in other databases. English and Chinese scientific literature databases were thoroughly accessed in the searching process. Emails were sent to corresponding authors to inquire about relevant studies and to pharmaceutical companies to gather information about any ongoing RCTs related to ulinastatin or thymosin α1 for sepsis. When disagreements occurred, they were discussed thoroughly to reach a consensus. The criteria for inclusion were as follows: (1) any RCT concerns ulinastatin combined with thymosin α1 for sepsis; (2) the patients enrolled were of any age and any gender and had a confirmed sepsis; (3) ulinastatin intravenously infused with any dosage and thymosin α1 subcutaneously injected with any dosage.Two authors (Feng Yun Wang and Li Xin Zhou) independently searched and examined the relevant literatures and scanned the title and abstract of retrieved articles to evaluate them for further assessment. Full articles were obtained when the information given in the titles and abstracts implied that the study included an RCT research design intended to investigate the efficacy of ulinastatin and thymosin The quality of the included literatures was assessed by the Cochrane Collaboration tool for assessing risk of bias, which contains the following 5 aspects: sequence generation, allocation concealment, blinding, incomplete outcome data, and selective outcome reporting. The assessment of risk of bias was presented by using a “risk of bias summary figure,” which presents all of the judgments in a cross-tabulation of study by entry. This display of internal validity indicates the weight the writer may give to the results of each study.X = |X2 − X1|, where X represents the mean applied in this meta-analysis; X2 represents endpoint mean; and X1 is the baseline mean of specific variable in included studies. In S2 = S12 + S22 − 2∗R∗S1∗S2, S represents the standard deviation (SD) applied in this meta-analysis; S2 represents the endpoint SD; and S1 represents the baseline SD of specific variable presented in the included studies. R is a constant, usually possessing a value of 0.4 or 0.5 in meta-analysis, as found in the Cochrane Handbook. In this meta-analysis, the value of R is 0.5.Two review authors independently extracted data and the process included the following detailed information: (1) the name of authors, year of publication; (2) intervention measures, including dosage, route, and timing; (3) number of intention-to-treat (ITT) patients in both groups ; (4) endpoints were 28-day survival rate, APACHE II score difference, ICU stay, mechanical ventilation time, CD4+T cell percentage, and CD8+T cell percentage. The data of dichotomous variables such as the number of 28-day survival patients were directly extracted from the included literatures. The meta-analysis data of continuous variables such as APACHE II score difference, CD4+T cell percentage, and CD8+T cell percentage were calculated according to Cochrane Handbook's methodology. For example, χ2 test, where a p value less than 0.1 was considered as the significance set. Values of I2 less than 25% were deemed to have low heterogeneity, and a fixed-effect model for meta-analysis was then used. Value of I2 more than 25% was deemed to have considerable heterogeneity, and a random effects model was then utilized  (p < 0.00001)  .I2 = 88%), so a random-model was applied. The pooling result showed UTI can shorten ICU stay by 3.02 days, mean = −3.03  (p = 0.14)  .p = 0.002)  .p < 0.0001) (p = 0.50)  . CD8+T c = 0.50) .α1 for sepsis. Four out of the six RCTs were in English  (p < 0.00001). On the strength of the above data, we might conclude that UTI treatment could ameliorate the severity of sepsis, which is consistent with its efficacy in promoting 28-day survival rate. Furthermore, the pooling results indicated that UTI treatment could shorten ICU stay by 3.03 days, mean = −3.03  (p = 0.14); however, the result did not reach statistical significance; and it could shorten mechanical ventilation (MV) time by 1.81 days, mean = −1.81  (p = 0.002). The reduction of both ICU stay and MV time could effectively lower the medical cost as well as save plenty of medical resources.With systematic treatment, the APACHE II score dropped dramatically in both UTI group and control group; however, with the treatment of UTI, the APACHE II score further decreased 4.72 when compared with control group, mean = −4.72, 95% CI  (Furthermore, a protective effect of ulinastatin was also observed in relation to other diseases. In major hepatectomy, ulinastatin could protect liver function and improve clinical outcomes, possibly via the inhibition of inflammation and oxidation at an earlier stage . Prophylα1, it was demonstrated that prophylactic usage of it could lower the incidence of infection in patients with nephrotic syndrome [α1 could improve cell-mediated immunity and reduce the occurrence of infection [α1 was used to treat hepatitis B patients in some clinical trials and its efficacy was proved by pathological observation [α1 increased cytokine production, especially IFN-γ and, at a higher-dose, thymosin α1 exhibited better efficacy against HBV, compared with other treatments studied. Evidence shows that thymosin α1 may improve resistance to infection and induce immune tolerance without GVHD in hematopoietic stem cell transplantation patients [With regard to thymosin syndrome . In patinfection . With itervation . The respatients .In this meta-analyses, three of the included RCTs reported data about CD4+T cells and CD8+T cells. The pooling results showed that the percentage of CD4+T cells gradually recovered after the treatment of UTI, which was consistent with former studies that conclude that UTI could improve cell-mediated immunity. The recovering percentage of CD4+T cells is a robust symbol of the ability to restore immune function in clinical situations. Since only three trials were enrolled in the analysis, UTI's potency of regulating immunity still lacks high-level evidence. It is worth mentioning that though UTI was regularly administered for severe sepsis in Asian countries, no severe side effects have been reported so far and no drug-related events were reported in the included RCTs either.α1 at approximately $100/1.6 mg, 3.2 mg/day for 7 days will increase the bill by $1400. Therefore, UTI for septic shock would add a considerable amount to patients' bill if they could not be covered by insurance. Consequently, the high cost might limit the popularity of UTI's application in septic shock. Nevertheless, if UTI could shorten ICU stay and mechanical ventilation time, as the data revealed in this meta-analysis, the extra cost of UTI might be well worth it.Cost-effectiveness not only concerns doctors who are desperately trying to save septic shock patients, it also concerns the families of the patients. Unfortunately, no cost-effectiveness data were revealed by any author in the included studies. To our knowledge, ulinastatin costing approximately $12/50000 U, 600000 U/day for 7 days will increase medical bills by nearly $1000, while thymosin α1 were widely used in the ICU field in Asia, but their efficacy was unclear up until now. While we do not intend to conclude that these two drugs are helpful, we are committed to drawing attention to them. Actually, we are somehow skeptical of the UTI's efficacy, since so far no guidelines exist recommending their application for sepsis. Regionally, UTI were mainly used in Asian countries, which made races of recipients relatively singular in the included trials. The average age of septic patients was usually above 50 years old or more, and drugs misused were either unhelpful or sometimes even harmful.The limitations of this systematic review and its meta-analyses are that all the RCTs were carried out in Asian countries such as Japan, China, South Korea, and India; the subjects were all Asian. Additionally, it is not clear whether the randomization process was adequately concealed/blinded in several RCTs. The lack of investigator blinding is a potential major flaw and source of potential bias that needs to be emphasized. Furthermore, with regard to the meta-analyses' results, only 28-day survival showed low heterogeneity, whereas APACHE II, ICU stay, and mechanical ventilation all showed considerable heterogeneity and random-model was used in the analytic process. That the average illness severity and age differed from trial to trial may be the source of heterogeneity; therefore, more well-designed trials in the near future are a feasible solution to remedy this flaw. Ulinastatin and thymosin In sum, some big international multicenter RCTs are urgently needed to further prove UTI's efficacy. However, by the results of this meta-analysis, we might be able to conclude that with the treatment of UTI, the short-term survival rate might be able to be increased, illness severity might be ameliorated, and ICU stay and mechanical ventilation time might be reduced considerably in patients with sepsis—all of which might help hospitals lower medical costs and save limited medical resources.


Xueshuan Xinmaining Tablet (XXT), the Chinese formula, has long been administered in clinical practice for the treatment of cerebral thrombosis and coronary heart disease. In this study, we aimed to study the effect and the molecular mechanism of activating blood circulation and removing blood stasis. Rat models of cold coagulation blood stasis were induced with ice-water bath and epinephrine to assess the amelioration of blood stasis by XXT. Microarray technique was used to identify gene expression from the model and XXT-treated rats. In addition, Quantitative Real-Time PCR (qPCR) was performed to verify the microarray results. The results showed that XXT had a good therapeutic effect on blood stasis by reducing the whole blood viscosity (WBV), plasma viscosity (PV), increasing PT, APTT and TT, and by inhibiting platelet aggregation. Genes were differentially expressed in rats among the model group and the XXT-pretreated groups. XXT ameliorated blood stasis by regulating the expressions of F13a1, Car1, and Tbxa2r. Yin-Yang of body energy [ Chuanxiong Rhizoma, Salviae Miltiorrhizae Radix et Rhizoma, Hirudo, Bovis Calculus, Moschus, Pubescent Holly Root, Sophorae Flos, Total ginsenoside of ginseng stems and leaves, Borneolum syntheticum, and Bufonis venenum. XXT has been widely used on the cerebral thrombosis and coronary heart disease in China for activating blood circulation and removing blood stasis for more than a decade [ in vitro or in vivo may help explain its mechanism of action [Traditional Chinese medicine (TCM), guided by the theory of traditional Chinese medical science, has been used for over 5,000 years mainly in China and other Asian countries. According to the Chinese medicine theory, body's normal function can be restored by obtaining the balance ofy energy . Chinesey energy . Blood sy energy . Traditiy energy . BSS is y energy . Xueshuaa decade , 8. In sa decade . Analyzif action , 11. In f action . It is rf action –16. Ther Salviae Miltiorrhizae Radix et Rhizoma, Chuanxiong Rhizoma, and Pubescent Holly Root were mixed and extracted with eightfold volume of 60% ethanol for 3 hours every time for 3 times. The filtrates were collected and concentrated. Then the concentrated solution was dried with a vacuum and was ground into fine powder (80 mesh). Sophorae Flos was decocted in 5-fold volume of water (pH 8~9 adjusted with saturated sodium carbonate solution) for 30 min every time for 3 times. The filtrates were combined and concentrated. Then the concentrated solution was dried with a vacuum and was ground into fine powder (80 mesh). Hirudo, Moschus, Bufonis venenum, Bovis Calculus, Borneolum syntheticum, and total ginsenoside of ginseng stems and leaves were, respectively, ground into fine powder (80 mesh). Finally, all the powder was mixed to get the extract of XXT.The extract of Xueshuan Xinmaining Tablet (XXT) was provided by Jilin Huakang Pharmaceutical Co., Ltd. Raw materials were purchased from Guanxian (Sichuan), Zhongjiang (Sichuan), Jilin (Jilin), Luoyang (Henan), Tonghua (Jilin), Wenshan (Yunnan), Shijiazhuang (Hebei), Fusong (Jilin), Meishan (Sichuan), and Zhenjiang (Jiangsu), respectively. All of them were authenticated and verified according to the Chinese pharmacopoeia (2010). The voucher specimens (no. HKYY-20130916-20130925) were deposited in the New Drug Research and Development Laboratory of Jilin University. And the preparation method was described as follows:Some standard compounds were also used in this study. Rutin (Lot number: 100080-200707), ferulic acid (110773-200611), salvianolic acid B (111562-200908), Ginsenoside Rg1 (110703-200726), Ginsenoside Re (110754-200822), Ginsenoside Rb2 (111715-201203), Ginsenoside Rb3 (111686-201203), Ginsenoside Rd (111818-201302), Cryptotanshinone (110852-200806), cholic acid (10078-0013), Cinobufagin (110803-200605), Resibufogenin (0718-9004), and Tanshinone IIA (0766-200011) were purchased from the National Institutes for Food and Drug Control . Ginsenoside Rc was provided by the New Drug Research and Development Laboratory of Jilin University.18 Column . The mobile phase was comprised of acetonitrile (A) and 1% acetic acid in water solvent (B). The gradient mode was as follows: initial 18% A linear gradient to 20% A in 20 min; linear gradient to 30% A in 60 min; linear gradient to 37% A in 90 min; linear gradient to 60% A in 105 min; and linear gradient to 100% A in 140 min. The flow rate was 1.0 mL/min. The components of XXT were identified by the comparison of the retention time from the chromatograms with those known standards. The fingerprints of the mixed standard compounds and the extract of XXT were shown in Figures The fingerprints of the XXT extract were further analyzed with a high performance liquid chromatography (HPLC) system  that consisted of a model 1525 Waters pump, model SEDEX FRANCE 75 Evaporative Light-scattering Detector, and Diamonsil CBuchang Naoxintong (BN) capsule, produced by Shaanxi Buchang Pharmaceutical Co., Ltd., is another traditional Chinese medicine. It has been approved for the treatment of cerebrovascular and cardiovascular diseases for many years . It has  ad libitum). The animal protocols were conducted according to the guide for the administration of laboratory animals  and were approved by the Institutional Animal Care and Use Committee (IACUC) of Jilin University, China.Male Wistar rats weighing 180–220 grams  were purchased one week before the experiment and were habituated to the living and testing environments. The rats were kept under controlled environmental conditions  were as follows: (1) normal control (NC), (2) cold- and epinephrine-induced blood stasis syndrome (BSS), (3) BSS model rats with low dose XXT (0.35 g/kg) treatment, (4) BSS model rats with middle dose XXT (0.70 g/kg) treatment, (5) BSS model rats with high dose XXT (1.40 g/kg) treatment, and (6) BSS model rats with “Buchang Naoxintong (BN)” (0.70 g/kg) treatment as positive control. Blood stasis models were made by placing the rats in ice-cold water (0°C~1°C) for 5 min daily for 7 days and by being administrated with two subcutaneous injections of hypodermic epinephrine (1 mg/kg) at 4-hour intervals at the 8th day [The experimental groups , activated partial thromboplastin time (APTT), prothrombin time (PT) and fibrinogen (FIB), and platelet aggregation were measured according to a previously described method , 24.−1. Plasma viscosity was measured at high shear rate (200 s−1) and low shear rate (50 s−1), respectively. All experiments were completed within 3 h after blood collection.Rats were anesthetized with 10% chloral hydrate (3 mL/kg) at 17 h at the 8th day after the last injection of epinephrine , and bloμL of plasma for 3 min at 37°C, followed by the addition of 60 μL of thrombin agent. APTT was determined by incubating 10 μL of the sample solution and 50 μL APTT-activating agents for 3 min at 37°C, followed by the addition of 50 μL of CaCl2. PT was determined by incubating 40 μL plasma for 3 min at 37°C, followed by the addition of 40 μL thromboplastin agent and 20 μL of the sample. FIB was determined by incubating 10 μL plasma with 90 μL imidazole buffer for 3 min at 37°C, followed by the addition of 50 μL FIB agent and 10 μL of the sample solution.The coagulation function index included thrombin time (TT), activated partial thromboplastin time (APTT), prothrombin time (PT), and fibrinogen (FIB). It was determined by coagulometer  with commercial kits, according to the manufacturers' instructions. To establish the standard curve of TT and thrombin concentration, TT was determined by incubating 60 μL platelet-poor plasma solution, platelet aggregation was induced by the addition of ADP  into 300 μL sample of platelet-rich plasma, and the changes in light transmission were recorded.Platelet-rich plasma was obtained after centrifugation of citrated whole blood at 200 g for 15 min, and then platelet-poor plasma was obtained by centrifugation at 2000 g for another 15 min . The agg t-test was performed by comparing two groups. P values less than 0.05 were considered statistically significant.All quantitative data were given as means ± SD performed using the SPSS 11.5 software package for Windows. Multiple comparisons among groups were performed by one-way analyses of variance (ANOVA). Student'sμg/μL. Agarose gel electrophoresis was used to check the RNA integrity. The RNA samples were stored at −80°C before processing for microarray analysis.Total RNA was extracted from the rat whole blood of each group using Trizol  reagent. Then RNA was concentrated by isopropanol and was purified using the NucleoSpin RNA Clean-up kit  following the manufacturer's protocol. RNA concentration was then determined using Epoch  with A260/A280 ratio between 1.8 and 2.0 and the RNA concentration was adjusted to 1  in vitro transcription using T7 Enzyme Mix, and then the cDNA was purified with RNA Clean-up Kit (MN). For each sample, 5 μg cRNA was inverse-transcribed using CbcScript II and Random Primer and then purified with PCR NucleoSpin Extract II Kit (MN). The cDNA was labeled with KLENOW using Random Primer. The labeled product was purified with PCR NucleoSpin Extract II Kit (MN). The biotinylated DNA was hybridized to 27K Rat Genome Array overnight at 42°C. Following hybridization, arrays were washed and stained. The signal intensity measurement was performed using LuxScan 10K Microarray Scanner . The microarray procedures were carried out according to the manufacture's instruction.The quality of RNA was checked using the Epoch microvolume spectrophotometer system and formaldehyde modified gel electrophoresis. Only high quality RNA (RNA Integrity Number (RIN) > 9.0) was used for microarray experiments. The rat genome oligonucleotide set was generated using 27K Rat Genome Array . Each 27K Rat Genome Array consisted of 26,962 amino acid modified 70-mer probes that represented 22,012 genes and 27,044 transcripts. The cRNA synthesis and labeling were carried out following GeeDomc RNA amplification tag protocol. Total RNA from each sample, along with poly A spikes (labeling control), was converted to double-stranded cDNA. Biotinylated cRNA was synthesized byP < 0.05) induced by blood stasis modeling and XXT treatment were then analyzed using the t-test software package . Finally, at least 2-fold-fluctuation in the normal control group was set as differential expression. A Perl program was used to select the differentially expressed genes that returned to the normal level after XXT treatment. In order to reveal changes in gene expression of biological important subsets, microarray data were subject to hierarchical clustering (Cluster 3.0). These genes were clustered into genes upregulated or downregulated in the blood stasis model and the XXT treatment groups.Raw microarray data were first normalized and the bad spots (misprinted spots) were discarded. Genes with signal intensity (Cy3 or Cy5) over 800 were regarded as the expressed ones. Significant changes in gene expression ( Premix Ex Taq II quantitative augmentation reaction system (Takara). The reaction was incubated in an ABI PRISM 7300 Fast Real-Time PCR System at the condition of SYBR Premix Ex Taq II quantitative augmentation reaction system (Takara). The primer sequences used for Real-Time RT-PCR were shown in Three genes (>2-fold change) that had antithrombotic function were verified by qPCR technique. PrimeScript RT Master Mix  was used to obtain the cDNA. qPCR was performed according to the instructions of SYBRP < 0.01), and the BCN group exhibited a significant decrease in plasma viscosity (P < 0.01) but a less significant one in WBV.The effects of XXT with different dosages on WBV and PV are shown in P < 0.05, P < 0.01) as shown in The effects of XXT on blood coagulation were evaluated by assays of APTT, PT, TT, and FIB content in the plasma. The level of FIB was increased, APTT and TT were shortened, and PT was significantly decreased in the model rats. The levels of PT, APTT, and TT were increased by XXT  and BCN treatment . The platelet aggregation percentage in BCN group was also significantly decreased compared to the model group (P < 0.01).Platelet aggregation rate in the model group was significantly increased. As shown in Data fluctuation by over 2.0-fold was considered to be differential expression. After blood stasis modeling, 435 genes were detected with significant change as compared to the normal control, of which 373 (approximately 85.75%) were upregulated. However, 501 differentially expressed genes, of which 307 genes were upregulated, were detected in the XXT group.We then conducted hierarchical clustering of the differentially expressed genes . The twoPerl analysis was performed for the gene patterns changed by XXT. 13 overexpressed genes in the blood stasis model were downregulated by XXT, and 3 lower-expressed genes were upregulated by XXT . We condP < 0.01) in the blood stasis model but were decreased (P < 0.01) to the normal control level by XXT pretreatment. The result demonstrated that abnormal gene expressions associated with blood stasis development could be restored by XXT pretreatment in rat model.The magnitude and direction of changes in expression of the three genes affected by XXT pretreatment were confirmed by qPCR . The expP < 0.01). And the mechanism was investigated by microarray analysis.Recently, blood stasis syndrome (BSS) is attached to the theory of promoting blood circulation and removing blood stasis (PBCRBS) by scholars in various countries . In AmerResults of microarray technology showed that a multitude of genes was differently expressed in acute blood stasis model rats. These genes, such as gene thrombin receptor-like 2 (F2rl2) and blood coagulation factor XIV (Proc), represent coagulant functions, which are vital in the blood coagulation , 32. To Investigating functional mechanism of complex TCM is a challenging task. In this study, the hemorheological and microarray experiments were conducted to explore the anti-blood stasis effect and related mechanism of XXT. The results demonstrated that XXT could significantly reduce the whole blood viscosity, plasma viscosity, increase PT, APTT, and TT, and inhibit platelet aggregation. Moreover, we also found that peripheral blood mRNA expression was dramatically changed in blood stasis model rats. And prophylactic administration XXT could prevent the abnormal expression of genes.


Metabolic syndrome is characterized by the association of 3 or more risk factors, including: abdominal obesity associated with an excess of abdominal fat, insulin resistance, type 2 diabetes, dyslipidemia and hypertension. Moreover, the prevalence of hypertension and metabolic dysfunctions sharply increases after the menopause. However, the mechanisms involved in these changes are not well understood. Thus, the aim of this study was to assess the effects of fructose overload on cardiovascular autonomic modulation, inflammation and cardiac oxidative stress in an experimental model of hypertension and menopause.Female SHR rats were divided into (n = 8/group): hypertensive (H), hypertensive ovariectomized (HO) and hypertensive ovariectomized undergoing fructose overload (100 g/L in drinking water) (FHO). Arterial pressure (AP) signals were directly recorded. Cardiac autonomic modulation was evaluated by spectral analysis. Oxidative stress was evaluated in cardiac tissue.AP was higher in the FHO group when compared to the other groups. Fructose overload promoted an increase in body and fat weight, triglyceride concentration and a reduction in insulin sensitivity. IL-10 was reduced in the FHO group when compared to the H group. TNF-α was higher in the FHO when compared to all other groups. Lipoperoxidation was higher and glutathione redox balance was reduced in the FHO group when compared to other groups, an indication of increased oxidative stress. A negative correlation was found between IL-10 and adipose tissue.Fructose overload promoted an impairment in cardiac autonomic modulation associated with inflammation and oxidative stress in hypertensive rats undergoing ovarian hormone deprivation. Metabolic syndrome is characterized by the association of 3 or more risk factors, including: abdominal obesity associated with an excess of abdominal fat, insulin resistance, type 2 diabetes, dyslipidemia and hypertension . ChangesHowever, it should be stressed that the prevalence of hypertension and metabolic dysfunctions sharply increases after menopause. The association between autonomic dysfunction and cardiovascular diseases support a link between alterations in the sympathovagal control and increased cardiovascular risk after menopause .Moreover, there seems to be an association between oxidative stress and inflammation, and both would thus play a role in the development of several cardiovascular diseases . However21 day old female SHRs were obtained from the Animals Facilities of the Institute of Cardiology of Rio Grande do Sul. The rats were randomly divided into hypertensive, hypertensive OVX  and hypertensive OVX undergoing fructose overload . All surgical procedures and protocols were approved by the ethics committee of Universidade Nove de Julho (Protocol AN002/12) and were conducted in accordance with the Guide for the Care and Use of Laboratory Animals, issued by the National Institutes of Health.FHO rats received D-fructose (100 g/L) in drinking water for 19 weeks. H and HO animals received standard laboratory chow and water ad libitum. The consumption of chow and water (with or without fructose) was measured weekly.After 10 weeks of fructose overload, the animals were anesthetized (80 mg/kg ketamine and 12 mg/kg xylazine), and the oviduct was sectioned and the ovary removed as described in detail elsewhere –11. DataAt the end of protocol (19 weeks of fructose overload),blood glucose and triglyceride concentrations were measured  after 4-hour fasting , 12. ForAfter metabolic evaluations, rats were anesthetized with ketamine (80 mg/kg) and xylazine (12 mg/kg), and two polyethylene-tipped Tygon cannulas filled with heparinized saline were implanted into the carotid artery and jugular vein for direct measurements of AP and drug administration, respectively. During the experiment, rats received food and water ad libitum; the rats remained conscious while in their cages and were allowed to move freely during the hemodynamic experiments. The arterial cannula was connected to a transducer , and blood pressure signals were recorded for a 30-minute period using a microcomputer equipped with an analog-to-digital converter . The recorded data were analyzed on a beat-to-beat basis to quantify changes in systolic (SAP), diastolic (DAP), and mean arterial pressure (MAP) and heart rate (HR) , 9, 10.Time-domain analysis consisted of calculating mean pulse interval (PI) and SAP, with PI variability and SAP variability measured from its respective time series . For frequency domain analysis, the same time series of PI and SAP were cubic splineY interpolated (250 Hz) and cubic splineY decimated so that they would be equally spaced in time after linear trend removal; power spectral density was obtained through the fast Fourier transformation. Spectral power for low-frequency  and high-frequency  bands were calculated by power spectrum density integration within each frequency bandwidth, using a customized routine  .One day after hemodynamic evaluations the animals were killed by decapitation and the heart (ventricles) was immediately removed, rinsed in saline, and trimmed to remove fat tissue and visible connective tissue. IL-6, IL-10 and TNF-α levels were determined using a commercially available ELISA kit (R&D Systems Inc.), in accordance with the manufacturer's instructions. ELISA was performed in 96-well polystyrene microplates with a specific monoclonal antibody coating. The threshold of sensitivity for the TNF-α , IL-10 and Il-6 assays was 15.0 pg/mL. Absorbance was measured at 540 nm in a microplate reader.The cardiac tissue (~0.5 g) was cut into small pieces, placed in ice-cold buffer, and homogenized in an ultra-Turrax blender with 1 g tissue per 5 mL 150 mMKCl and 20 nM sodium phosphate buffer, pH 7.4. The homogenate was centrifuged at 600 g for 10 min at -26C.For the TBARS assay, trichloroacetic acid  was added to the homogenate to precipitate proteins and to acidify the samples . This miCardiac tissue membrane lipoperoxidation was evaluated by chemiluminescence. The chemiluminescence assay was carried out with an LKB Rack Beta liquid scintillation spectrometer 1215  in the out-of-coincidence mode at room temperature (25°C to 27°C). The supernatants were diluted in 140 mM KCl and 20 mM sodium phosphate buffer, pH 7.4, and added to glass tubes, which were placed in scintillation vials; 3 mM tert-butylhydroperoxide was added, and chemiluminescence was determined up to the maximal level of emission , 16.2O2 absorbance at 240 nm. CAT acitivity was expressed as μmol H2O2 reduced/min/mg protein [The quantification of SOD activity, expressed as U/mg protein, was based on the inhibition of the reaction between O2˙ - and pyrogallol . CAT act protein . GPx act protein .g for 10 min,and 2 M KOH was added to the supernatant. The reaction medium contained 100 mM phosphate buffer (pH 7.2), 2 mM NADPH, 0.2 U/mL glutathione reductase and 70 μM 5,5’-dithiobis (2-nitrobenzoic acid). To determine the GSSG concentration, the supernatant was neutralized with 2 M KOH and inhibited by the addition of 5 μM N-ethylmaleimide. Absorbance was read at 420 nm [To determine GSSG and total glutathione concentration, tissue was homogenized in 2 M perchloric acid and centrifuged at 1000 Data are expressed as mean ± SEM. The Levene Test was used to evaluate data homogeneity. A one-way analysis of variance followed by the Student-Newman-Keuls test was used to compare groups. Pearson correlation analysis was used to identify an association between variables. Significance level was established at P ≤ 0.05.Body weight and adipose tissue weight were higher in FHO group when compared to other groups. Chow consumption was similar among groups . The FHO group, however, presented a statistically significant greater water consumption (82.9 ± 1.4 mL/day) when compared to H group (25.2 ± 0.2 mL/day) and HO group (24.6 ± 3.0 mL/day). The total caloric intake (chow + fructose) was higher in FHO when compared to other groups . Glycemia was similar between groups. Triglycerides levels were increased in the FHO group when compared to all other groups. Insulin sensitive was lower in FHO group when compared to the H group  and total variance VAR of PI were lower in FHO groups when compared to the HO group. Total variance (VAR) of SAP was increased in the HO group when compared to the H group. VAR and LF band of SAP were higher in the FHO group when compared to all other studied groups Table .TNFα was higher in the FHO when compared to all other groups and IL-10 was lower in FHO when compared to the H group. Negative correlation was observed between adiposity tissue and IL-10 among hypertensive groups (r = -0.6)  has found that, prior to 45 years of age, prevalence of hypertension is higher in men than in women. From 45 to 54 years and from 55 to 64 years of age, the percentage of hypertensive men is similar to that of hypertensive women, a period which coincides with the advent of menopause. After 65 years of age, BP levels increase faster in women than in men . MoreoveIn the present study, ovarian hormone deprivation increased both AP and vascular sympathetic modulation in SHR rats. Previous studies from our group have also demonstrated that ovarian hormone deprivation induces BP increase and leads to impaired baroreflex sensitivity in healthy Wistar rats , 10. In Experimentally, fructose overload in drinking water or chow has been used to study metabolic syndrome and promote similar metabolic and autonomic derangements , 22, 23.Moreover, the sum of risk factors promoted an increase in the heart rate, an additional increase in BP, impairment in heart rate variability and systolic arterial pressure variability. Several studies have found cardiac diastolic dysfunction and an increase in BP after fructose overload , 23, 25.Heart rate variability has been used as a tool to assess the role of autonomic nervous system fluctuations in normal healthy individuals and in patients with various cardiovascular and non-cardiovascular disorders . ReductiRegarding inflammation makers, we observed that in this study TNF-α was higher in FHO when compared to other groups while fructose overload promoted a reduction in IL-10. We also found a negative correlation between adipose tissue an IL-10. In fact, adipose tissue is an active endocrine and paracrine organ that releases a large number of cytokines and bioactive mediators, such as leptin, interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) , 29. TheMoreover, decreased antioxidant capacity together with increased lipid peroxidation has been reported in patients with fatty liver, visceral obesity, and metabolic syndrome. There seems to be a correlation between the amount of visceral fat and systemic oxidative markers, indicating that visceral fat is an independent regulator of oxidative changes . RegardiRegarding the limitations of the present study, we did not evaluate non-OVX groups treated with fructose, which would help us determine whether the combination of OVX and high fructose are additive or synergistic. Furthermore, we used a direct acute measurement of BP, which is a well-established and widely accepted method for BP recording in conscious rats; however, this method have some disadvantages when compared to telemetry AP measurements.In conclusion, fructose overload induced an impairment of cardiovascular autonomic modulation associated with increased oxidative stress and inflammatory parameter in hypertensive ovariectomized rats. These data reinforce the role of autonomic changes in immune function modulation , 29, 33,