Conventional spirometric parameters have shown poor correlation with symptoms and health status of chronic obstructive pulmonary disease (COPD). While it is well-known that the pattern of the expiratory flow-volume curve (EFVC) represents ventilatory dysfunction, little attempts have been made to derive quantitative parameters by analyzing the curve. In this study, we aimed to derive useful parameters from EFVC via graphic analysis and tried to validate them in patients with COPD.Using Graphical Analysis 3.4 Vernier Software, we derived from the EFVC such parameters as area of obstruction (Ao), area of triangle (AT), area of rectangle (AR) and ratio of volume at 75 and 25 % peak expiratory flow (PEF) (0.25/0.75 V). For validation, we reviewed clinical and spirometric data of 61 COPD patients from Seoul National University Airway Registry (SNUAR) and Korean obstructive Lung Disease (KOLD) cohorts.p = 0.037). Six-minute walking distance (6MWD) highly correlated with Ao/AR  and Ao/PEF  whereas neither FEV1 nor FEV1/FVC had significant correlation with 6MWD.Of all parameters, only RV/TLC significantly correlated with scores from St. George’s Respiratory Questionnaire (SGRQ)  contains supplementary material, which is available to authorized users. These conventional spirometric parameters, such as FEV1 or FVC, are currently accepted standards in grading the severity of airflow limitation; when FEV1/FVC is less than 0.70, diagnosis of COPD is made.Chronic obstructive lung disease (COPD) is defined as having airflow limitation, measured as forced expiratory volume in one second  [1 and hospital readmission in such patients is also reported [However, these parameters cannot exactly determine the health status of patients with COPD. For example, FEVe (SGRQ) ; little reported . Therefo1 or FEV1/FVC [Exercise capacity, which can be simply measured by 6-min walking distance (6MWD), is reflective of activities of daily living and functional status of patients with COPD but shows poor correlation with FEVFEV1/FVC , 6.While it is well-known that the concave shape of expiratory flow-volume curve (EFVC) is suggestive of the presence of underlying small airway obstruction, few attempts have been made to quantify the concave area and correlate it with clinical indices in patients with COPD –9. In thThe following study was approved by the institutional review board of Seoul National University Bundang Hospital (B-1108/134-004 and B-0508-023-009). Informed written consent for participation in the study was obtained from all participants.For validation of new graphic parameters, we analyzed the clinical and baseline pre-bronchodilator best spirometric data of 61 COPD patients from the Seoul National University Airway Registry (SNUAR) and the Korean Obstructive Lung Disease (KOLD) cohort . 40 patiSpirometry was performed using a Vmax 22 instrument . Lung volumes were measured by body plethysmography . Diffusing capacity for carbon monoxide (DLco) was measured by the single-breath method using a Vmax229D (Sensor-Medics). All pulmonary function tests were performed as recommended by the American Thoracic Society and European Respiratory Society.For graphic analysis of the EFVC, we used Graphical Analysis 3.4 Vernier Software program. Using the program, we first integrated the area under the curve. Figure Figure As Ao approaches to 1, the concavity of EFVC becomes more severe. As Ao gets closer to 0, the EFVC becomes less concave. Xp denotes the lung volume at PEFR.Because the baseline pulmonary function depends on the patients’ heights and other factors, we needed to define Ao in ratio rather than raw values. In order to calibrate and adjust for such individual differences, we divided Au by At. Using a raw value such as Au may result in wrong direction and inappropriate interpretation of result  were defined as follows; we used the reference equation derived from the Korea National Health and Nutrition Examinations Survey IV to calculate predicted FVC in Korean COPD patients , 12.\docThe ratio of lung volumes at 75 and 25 % of PEFR (0.25/0.75 V) were also calculated.Statistical analysis was performed using the statistics software, PASW Statistics for Windows, Version 18.0, Chicago: SPSS Inc. The association between clinical and spirometric parameters was determined by using Pearson correlation analysis.2. Mean FEV1 was 65 % and FEV1/FVC 0.48.The demographic, clinical, and conventional spirometric characteristics are shown in Table p = 0.037). Inspiratory capacity (IC), as percent predicted, showed positive correlation with 6MWD.Among 61 subjects, the pre-bronchodilator EFVC data for three subjects were missing, so they were excluded in the correlation analysis. Table The results for new parameters derived from EFVC are given in Table p = 0.005) and Ao/PEF . Neither FEV1 nor FEV1/FVC had significant correlation with 6MWD.Among clinico-physiological parameters, 6MWD highly correlated with Ao/AR  curve of 34 stable COPD patients at resting state and found that the area under the curve divided by the surrounding rectangular area, the rectangular area ratio (RAR), is indicative of concavity of SEFV curve; FEVhe curve . Ma et ahe curve . While Rhe curve . Assessi1 ≥ 80 %).The authors acknowledge that the present study has several limitations. First, the size of study sample was too small to fully validate our new parameters. Second, we analyzed the pre-bronchodilator best expiratory curves rather than the post-bronchodilator curves, which could have affected the curvature. Third, most patients were in moderate to severe stage of COPD, GOLD II-III, and we did not include the patients with mild COPD (FEVIn this study, we found that Ao/AR and Ao/PEF are potentially useful parameters which correlate well with exercising capacity in patients with COPD. Yet, further studies are warranted to validate the new parameters in a sample of sufficient size and also in SEFV curves.


To the Editor: Although the provisionally named influenza D virus was first isolated as an influenza C–like virus from pigs with respiratory illness in Oklahoma in 2011  to test a convenience sample of 28 serum samples from healthy animals in a herd of female cattle in the Ibaraki Prefecture in central Japan. Two viruses with heterologous antigenicities, D/swine/Oklahoma/1334/2011 (D/OK) and D/bovine/Nebraska/9–5/2012 (D/NE)  for D/NE were the same as or only 2 times lower than those for D/OK in all seroconversion samples, unlike the results for the seropositive samples in the first testing . This rehttp://www.clustal.org) and MEGA version 6.0  successfully amplified the full genome sequence (GenBank accession nos. LC128433 for D/bovine/Ibaraki/7768/2016), which was aligned to the influenza D virus sequence. However, we could not isolate infectious virus by using sensitive cells (sion 6.0 . AlthougIn summary, a cattle herd in Japan had influenza D virus infection. Although this study is a case report with a small number of samples, the observation shows a potential role for influenza D virus in the bovine respiratory disease complex.Detection of hemagglutination-inhibition antibodies to influenza D virus in a herd of cows in Ibaraki Prefecture, Japan


We aimed to determine the association between nonalcoholic fatty liver disease (NAFLD) and diabetes risk among body size phenotypes which was based on cross-classification of body mass index (BMI) categories  and metabolic status .We conducted a cross-sectional analysis using a cohort of 10,761 apparently healthy Chinese adults who underwent comprehensive health checkups including abdominal ultrasonography. Subjects were classified as metabolically at-risk by having any two of the following, consistent with the Adult Treatment Panel-III metabolic syndrome definition: (1) systolic/diastolic blood pressure ≥130/85 mmHg, (2) triglycerides ≥1.7 mmol/L, (3) fasting blood glucose ≥5.6 mmol/L, (4) HDL-cholesterol ≥1.0/1.3 mmol/L for men/women.2) and overweight/obese (BMI ≥23 kg/m2) group were 2.10 (1.85–3.93) and 1.85 (1.35–2.53), respectively. Among participants with metabolically at-risk, the significant association between NAFLD and diabetes was lost, regardless of obesity status. There were only 27.1 % subjects with the presence of the three factors  occurring together, while the three factors occurring together was common (56.16 %) in diabetic individuals. The multivariate-adjusted ORs for diabetes were 1.1 (0.61–1.98) for overweight/obesity, 2.23 (1.05–5.14) for NAFLD, and 8.04 (5.0–12.09) for metabolically at-risk. The OR for the presence of all the three factors occurring together was 23.22 (13.96–38.63).Among participants without metabolically at-risk, multivariate-adjusted odds ratios (ORs) for diabetes from NAFLD compared with those without NAFLD in the normal-weight (BMI <23 kg/mNAFLD was associated with diabetes risk among participants without metabolically at-risk. The clustering of overweight/obesity, NAFLD, and metabolically at-risk is common in diabetic subjects and strikingly and markedly increases the diabetes risk. Along with the increasing westernization of diet, physical inactivity, and the obesity epidemic , the worHence, we aim to evaluate the association of NAFLD with CVD risk factors and diabetes risk among body size phenotypes.The study participants were Chinese employees and retired workers aged 20–100 years from the Wuhan Iron and Steel Company (WISCO), which is one of the largest iron and steel companies in China. In WISCO, the Industrial Safety and Health Law requires employees and retired workers to receive periodic health evaluations at the WISCO General Hospital . The present cohort included all employees and retired workers who received a comprehensive health examination  at the Healthcare system, WISCO general Hospital, between June 2008 and December 2010 .All subjects were asked to complete a standard questionnaire that gathered information on age, sex, cigarette smoking and alcohol consumption habits, histories of current and previous illness, and medical treatment. We excluded 4992 participants from this study, comprising 1493 participants who were taking medications for hypertension, diabetes, dyslipidemia, or hyperuricemia, 1271 with alcohol consumption in amounts >70 g/week for women (73) and >140 g/week for men (1198), 857 participants with hepatitis B surface antigen (HBsAg) positivity, and 1758 missing information on age, sex, anthropometric assessment, components of metabolic syndrome, test results for HBsAg, or liver ultrasound scans. As some individuals met more than one exclusion criteria, the remaining available 10,761 participants (6901 men and 3860 women) were included in our data analysis. The fact that men accounted for 64.1 % of total participants was in consistent with the proportion of male employees at WISCO. According to the Private Information Protection Law, information that might identify subjects was safeguarded by the Health Examination Center. This study was approved by the institutional review board of WISCO general Hospital. Because we only retrospectively accessed a de-identified database for purposes of analysis, informed consent requirement was exempted by the institutional review board.2), and overweight/obesity (BMI ≥23 kg/m2). Participants’ seated BP was measured twice for every 5 min on the right arm after 5 min of rest by trained nurses with a sphygmomanometer. The mean of the two readings was used in data analysis.Anthropometric measurements, including weight, height, and blood pressure (BP) were measured following standardized protocols from the World Health Organization (WHO). Weight was measured with the participants wearing light clothing and height was measured without shoes. BMI was calculated as weight (in kilograms) divided by the square of height (in meters). According to the WHO criteria for Asians , subjectOvernight fasting (at least 8 h) blood samples were collected from the antecubital vein of each individual. Biochemical measurements, including assessment of fasting blood glucose (FBG), total cholesterol (TC), triglycerides (TG), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), alanine aminotransferase (ALT), and hepatitis viral antigen/antibody, were measured enzymatically on an autoanalyzer . All the blood measurements were followed the same protocol.Ultrasound tests were performed by trained sonographers using a high-resolution, real-time scanner . One experienced radiologists used standard criteria in evaluating the images for the presence or absence of hepatic fat .Subjects were classified as metabolically at-risk by having any two of the following, consistent with the Adult Treatment Panel-III (ATP III) metabolic syndrome definition  and withAccording to the 2014 American Diabetes Association criteria , diabeteAll statistical analyses were conducted using SPSS software . Variables were presented as means and standard errors (SE) for continuous variables and as percentages for categorical variables. Based on cross-classification of four body size phenotypes and the NAFLD status, participants were categorized into eight mutually exclusive groups . Differences in characteristics among these groups were verified by analysis of covariance or Cochran–Mantel–Haenszel Chi square statistics as appropriate. A post hoc (Scheffé method) multiple comparison was used to establish the differences between the groups, when necessary. Furthermore, a logistic regression model was used to estimate crude and adjusted odds ratios (OR) and corresponding 95 % confidence intervals (CI) for the presence of NAFLD compared with the absence of NAFLD within each body size phenotype. The three models were as follows: Model 1 was an unadjusted model. Model 2 was adjusted for age, and sex. Model 3 was additionally adjusted for TG/HDL-C (which was a good indicator of insulin resistance).2. Similar processes were repeated for NAFLD and metabolic at-risk.Venn diagram was constructed as a visual display of NAFLD and overweight/obesity based on BMI ≥23 kg/mA logistic regression model was used to examine the joint associations of body size phenotypes and the NAFLD status with diabetes risk to determine whether the associations of body size phenotypes with diabetes differ based on the presence of NAFLD. Significance was accepted at a two-tailed P < 0.05.The prevalence of NAFLD in the MHNW, MHO, MANW, and MAO phenotype was 8.79, 31.72, 50.98, and 76.19 %, respectively.Table 2. Among those subjects (6699) with either NAFLD or overweight/obesity, considerable participants (35.54 %) had overweight/obesity alone compared with 7.99 % with NAFLD alone. Moderate magnitude of overlap existed between NAFLD and overweight/obesity (56.47 %)  Fig. a. When aSimilarly, we further assessed overlap between NAFLD and metabolically at-risk. Low magnitude of overlap existed between NAFLD and metabolically at-risk 39.91 %)  cluster together in both the whole population and the diabetic population. We found a significant dissociation between these three factors in the whole population, with only 27.1 % subjects with the presence of all the three factors occurring together. The simultaneous presence of the three factors was common (56.16 %) in diabetic population, suggesting the frequency and importance of the clustering of these three factors for diabetes. Furthermore, we have examined the combined and separate associations of NAFLD, overweight/obesity, and metabolically at-risk with diabetes risk. We found that NAFLD and metabolically at-risk both independently associated with diabetes risk, with stronger associations seen for metabolically at-risk. We also found that the occurrence of all three factors together had the strongest association with diabetes.Accumulating evidence shows that NAFLD is strongly associated with CVD risk factors , 26. HowThe concomitant presence of overweight/obesity, NAFLD, and metabolically at-risk is common , 34. HowThe dissociation between the three factors results in different combinations of the three factors, conferring differential odds of diabetes and different macrovascular complication phenotypes in diabetic patients , as eachAlthough we observed greater ORs for diabetes associated with metabolically at-risk compared with NAFLD, being NAFLD was associated with poorer lipid and glucose profiles within the same metabolic status. Among both the normal-weight group and overweight/obese group, the most favorable lipid and glucose profiles were found in subjects with metabolically health/without NAFLD. In addition, there was a marked increase in odds of diabetes in subjects with metabolically at-risk/with NAFLD, regardless of obesity status. The fact that metabolically at-risk and NAFLD have independent effects on each other and have synergistic effects on diabetes risk suggests that targeted specific approaches to preventing visceral fat or ectopic fat accumulation or to reversing the visceral fat or ectopic fat accumulation may have a considerable impact on decreasing diabetes risk. The presence of all the three factors occurring together, which was the most frequent combination of the three factors among diabetic subjects, had the strongest association with diabetes, indicating that an optimal prevention strategy for diabetes should emphasize amelioration of the effects of each of the three factors.Several prospective cohort studies suggest that MHO phenotype might be associated with a nonsignificant or significant increased risk of diabetes incidence , 44–47. There are several limitations. The cross-sectional design implies that no causal relationship can be drawn. We only had data on Chinese adults comprised of selected populations  with a preponderance of men, thus, extrapolating results to the Chinese general population or to other racial or ethnic population should be interpreted cautiously. Since NAFLD was diagnosed by ultrasonography, which is a reasonably accurate technique for detecting modest amounts of liver fat (>30 % liver fat infiltration), participants with minor amounts of fatty infiltration might not have been captured. That NAFLD was not confirmed by liver biopsy was also a limitation. A standard 75-g oral glucose tolerance test was not performed, which might suggest an underestimation of the diabetes prevalence.The association of NAFLD with diabetes is more pronounced in normal-weight individuals than in overweight/obese individuals among participants without metabolic at-risk. There is a significant dissociation of the three factors  in the whole population, while the three factors commonly occur together in diabetic individuals. Of the three factors, overweight/obesity had the weakest association with diabetes and metabolically at-risk had the strongest association. The presence of all three risk factors occurring together strikingly and markedly increases the diabetes risk. NAFLD should be considered as a more meaningful predictor of diabetes in MHO individuals.


P < 0.001). Biochemical complications and mortality rate of the animals were significantly higher in the HA + 18 mL/Kg ILE group. ILE reversed sings of HA neurotoxicity; however, synergistic effect of high dose of ILE and HA increased complications and mortality.There are many reports on the effect of intravenous lipid emulsion (ILE) as an antidote in drugs related toxicities. We determined the effects of ILE on neurotoxicity of haloperidol (HA), a highly lipophilic antipsychotic, as a model of antipsychotics poisoning. We used six groups of five male rabbits. Two groups received distilled water intravenously followed by infusions of either 18 mL/kg of normal saline or ILE 20%, after 30 minutes. The third group received 18 mL/kg of normal saline after HA (2.6 mg/kg) administration. The three other groups received ILE 20% solution  following HA injection. Catalepsy scores, temperature, pupil size, and mortality rate were measured at 0, 0.5, 1, 2, 3, 4, 8, and 24 hours after HA administration began. Blood and tissue samples were taken from all animals at 24 hours or at death time for biochemical, cell count, and pathological studies. ILE reversed cataleptic scores, miotic pupils, and hypothermia of HA intoxication much faster than normal saline ( Intravenous lipid emulsion (ILE) has been known as a novel antidote against lipophilic xenobiotics intoxication in animal studies and some case reports –4. The nAcute poisoning with antipsychotic drugs is an important cause of referring intoxicated patients to the hospital , 7. HaloIn this study, we tried to determine whether ILE would have beneficial neurologic effects in the setting of HA toxicity.Thirty male New Zealand rabbits with body weight between 2500 and 3000 g were used. They were housed in individual cages with 12-hour light/dark cycle and easy access to food and water. The experiments were approved by the Ethics Committee for Animal of MUMS .Haloperidol ampoules, 5 mg/mL,  were used and each 1 mL of HA, 5 mg, was diluted up to 5 mL with aseptic distilled water (ADW). ILE 20%  and aseptic normal saline (NS) were also administered intravenously. (a) Negative control group (NC): they received ADW and then 18 mL/kg of NS. (b) ILE 0 group: they received ADW and 18 mL/kg ILE 20%. (c) NS group: 18 mL/kg of NS was administered after receiving HA infusion.  These 3 groups received ILE 20% solution with 6, 12, and 18 mL/kg doses , respectively, following HA administration.Thirty rabbits were divided into six groups as follows.These doses had been selected based upon previous studies –12.A 24 G venues cannula was inserted into marginal ear vein of rabbits in aseptic conditions. Subsequently, either diluted HA or an equal volume of ADW was infused intravenously within 15 minutes. HA was administrated at a dose of 2.6 mg/kg body weight of rabbits . Zero time was considered when HA or ADW infusions began. ILE 20% or NS were intravenously infused, in an infusion rate of 4 mL/min, at 30 min. At the end of the infusion, the cannulas were removed. All studies started at 08:00 am.Pupils size, catalepsy score, body temperature, and mortality rate of rabbits were evaluated before starting drug or solution administration at baseline (time 0) and through 24 hours at 0.5 hour (just before infusion of ILE 2% or NS), 1, 2, 3, 4, 8, and 24 hours.The pupils size were assessed in millimeter based on the diameter of right eye pupil, when the rabbits were placed on laboratory table in an identical location and at equal direction under similar brightness of light at all evolution times.Body temperatures of animals were evaluated at all evolution times by a rectal thermometer according to the method previously reported in the literature .The animal right foreleg was brought round behind its back. The latency of returning the malpositioned forelimb to normal state was measured in seconds. Sixteen seconds was considered as the cutoff time to protect the animal limb against possible injuries.The experimenter abducted, internal rotated, and extended the right forelimb of the animal in abnormal position. The latency of returning the limb to its normal position was recorded. The cutoff time was considered 60 seconds.The rabbit was placed in a nearly upright position with its forepaws over a step. The latency of the animal being able to maintain this posture was recorded. The cutoff time was 4 min (240 seconds).Catalepsy of animals was assessed by Banerjee et al. method with some modification . Three sAt each run, every part of catalepsy scores was measured twice and the means of data were recorded.Blood samples were drawn from all animals after 24 hours or at the time of death if they died sooner than 24 h. One milliliter of whole blood was drawn into a test tube containing anticoagulant (citrate) and the cells blood count was measured by cell counter . Biochemical tests , aspartate transaminase (AST), alkaline phosphates (ALP), blood urea nitrogen (BUN), serum creatinine (Cr), amylase, and lipase) were performed by autoanalyzer  on 0.3 milliliter of animals' blood serums.Autopsy was performed immediately after sacrifice or at death time if they died sooner than 24 h. Tissues from various organs, right hemisphere of the brain, right lung, and right kidney were fixed in 10% buffered formalin and embedded in paraffin. Right hemispheres of brain were divided into four sections: frontal lobe, parietal and basal ganglion region, occipital lobe, and cerebellum and medulla. Fragments of the various organs were sectioned at 70 Wm by vibratome and stained with hematoxylin and eosin. The tissue samples were evaluated by a single pathologist. All tissues and blood samples were coded and all experiments were blindly performed.Further sections of the brain were processed in TUNEL procedure to detect evidence of apoptotic cells. Sections were stained according to the manufacturer instructions . In brief, sections were dried onto poly-l-lysine-coated slides, treated with proteinase K (20 mg/mL), and incubated with TdT enzyme. Percentages of apoptotic nuclei were calculated by counting them being distributed within 100 nuclei.t-test was performed as post-test if the P value was less than 0.05. Kruskal-Wallis ANOVA test were used if necessary. Dunn test was set as posttest only if the result was positive; the P value was less than 0.05. Fisher's exact test was used to evaluate differences in mortality rate of different groups. The alpha level was set at 0.05.Statistical analysis of all variables was performed by SPSS 11.5. Two-way repeated-measures ANOVA was used to evaluate differences in continuous numerical variables of different groups. Tukey P < 0.01) (All HA-treated rabbits suffered from hypothermia ( < 0.01) . Rectal  < 0.01) . The NS P < 0.0001)  . By the  0.0001) .P < 0.05 and P < 0.001, resp.) in the second hour. At the third hour all HA-intoxicated animals which received ILE  had significantly lower cataleptic scores than NS group; however, their mean scores were higher than those in NC group   .P < 0.0001), but they did not reach NC group score (P < 0.0001). There was no significant difference between mean catalepsy score of NS group in second and third hours (55.28 ± 2.57 and 55.0 ± 2.0 seconds). All HA-treated animals were cataleptic at 8th hour; however, the means of scores of ILE groups were significantly lower than the NS score (P = 0.0022).The mean cataleptic scores of HA + ILE treated animals  were reduced at 3rd hour in comparison with NS group (P < 0.0001) (P < 0.0001), although, there were significant differences between the mean score of ILE groups (time of standing of ILE 12 > ILE 6 > ILE 18) (P < 0.05). At 4th hour, the NS group was cataleptic but none of the ILE groups were.After administration of HA, the standing ability of animals was significantly reduced ( 0.0001) . This laP = 0.023, Fisher's exact test).All animals were alive at 24th hour  except 2 rabbits of ILE 18 groups  . There was no statistically significant difference between means of BUN and Cr in ILE 6 and ILE 12 and control groups . In addition, the means of ALP in ILE 6 and ILE 18 groups were raised, but the means of ALP of NC, ILE 0, and ILE 12 groups were the same .HA injection could raise the mean of ALP in NS group in comparison with NC (P < 0.05). The mean level of serum AST in NS group was also increased but was not statistically significant compared to NC group (The serum AST of ILE 18 group rose significantly (NC group .P < 0.05). However, they were not statistically higher than mean serums ALT in NS group   count in NC, ILE 0, and NS groups; however, means of WBC count in ILE 6, ILE 12, and ILE18 were significantly reduced in comparison with NC group  . There wThere were 2–5% apoptotic cells in different sections of rabbits' brains that were not statistically significant. The renal pathology of one rabbit in NS group showed acute tubular necrosis pattern. The others had no similar findings. The prevalence of acute tubular necrosis was not statistically significant between different groups. Pathological evaluation revealed inflammatory cell infiltration through the lung tissue of rabbits, especially in ILE 0 and ILE 18 groups Figures  and 8.There are some reports about the effect of ILE on xenobiotics-induced central nervous system (CNS) toxicity . For insHA is a typical potent neuroleptic that is an antagonist of D2 receptors . It induIt seems that there are some other explanations for antidotal effect of ILE against HA-induced catalepsy. There are some reports that coadministration of voltage calcium channel blockers with HA could increase the catalepsy score of rats and decrease the onset of catalepsy by reducing the neurotransmitter dopamine in striatal region , 24. HowIntravenous administration of HA-induced hypothermia in all rabbits and ILE 20% restored the reduced temperature to normal values sooner than normal saline. Lin has suggested that hypothermia-induced HA may be due to a decrease in metabolic heat production as well as an increase in ear blood flow . Boschi HA-induced miosis was reversed by ILE 20% more rapidly than normal saline. The miosis that resulted from HA administration is more related to peripheral effect of HA than central effects. HA-induced miosis is not mediated by blocking alpha-adrenergic receptors of radial muscles or stimulating cholinergic receptors of circular muscles of iris . Thus, IRising BUN and Cr in rabbits happened in NS group and one rabbit of this group suffered acute tubular necrosis. Dopamine causes renal vasodilation which is attenuated by haloperidol . RabbitsMinor elevations of serum aminotransferases commonly happen secondary to HA therapy , and theHA administration has the possibility of drug-induced leucopenia/neutropenia, especially in patients with preexisting low WBC or coadministration of compounds which could induce leucopenia , 43. AdmWe found no difference between amylase and lipase of all groups. Although, daily injection of 2 mg/kg of HA into mice has significantly raised the protein content and the volume of the basal pancreatic juice . InfusioOur histopathological examinations showed the dose dependent lung infiltration due to fat emboli. Lung injuries secondary to ILE infusion are complicated. It seems that two important factors are involved in this topic, underling pulmonary condition and lipid load. Some case series revealed lowered blood oxygen content, increased shunting, and pulmonary vasoconstriction after ILE infusion in patients with respiratory failure and/or sepsis , 51, andOn the other hand, part of this variation is related to the infusion rate and concentration of ILE that result in lipid load . JacobovHA-intoxicated animals, which received 18 mL/kg of ILE, had the highest mortality rate and they also showed worse laboratory tests. Based on our results, the best dose of ILE, as antidote for neurological manifestation of HA toxicity in rabbits, is 12 mL/kg, while Harvey and Cave recommended the 6 mL/kg of ILE as antidote for hemodynamic and cardiac toxicity of local anesthetic drugs . This diIn conclusion, ILE could reverse HA-induced neurotoxicity by lipid sink phenomena or direct effect of ILE. However, dose determination and safety of ILE in neurotoxicity need additional research.


Semecarpus anacardium (Linn.) (abbreviated as SF).Diabetes mellitus is a global health problem and constantly increasing day by day. The number of diabetic people in world is expected to rise to 366 million in 2030. The available drugs for diabetes, insulin or oral hypoglycemic agents have one or more side effects and search for new antidiabetic drugs with minimal or no side effects from medicinal plants is a challenging for us. The present study was undertaken to investigate the antidiabetic and antioxidant activity of The antidiabetic activity was determined by using alloxan-induced diabetic rats. After 15 days of treatment, serum biochemical parameters such as TC, TG, LDL, HDL, SGOT and SGPT were estimated. The survival rate, body weight, organ weight, liver glycogen and blood parameters (RBC and Hb) were also measured. The antioxidant activity was measured by DPPH free radical scavenging assay. Phytochemical screening, total phenolic and total flavonoid content were determined by using standard methods.50 values of SA extract and ascorbic acid were found 72.24 μg/ml and 17.81 μg/ml, respectively. Phytochemical screening showed the presence of steroids, triterpenoids, flavonoids, glycosides, saponins, and tannins that were contribute to biological activity.The results showed that the survival rate was 100% in rats of Group SA 400. The effect of extract on blood glucose level in Groups SA 100, SA 200 and SA 400 were dose-dependent throughout the treatment period. No significant changes in organ weight to body weight ratio were observed, liver weights significantly improved in Groups SA 200 and SA 400. The bark extract exhibited significant (p < 0.05) anti-diabetic activity with lowering TC, TG, LDL level dose-dependently and protected liver which may be partially explained by attenuation of SGOT and SGPT levels and increases liver glycogen. The percentage of Hb and RBC counts were negatively correlated with the doses of extracts. In DPPH scavenging assay, ICS. anacardium possess strong anti-diabetic and antioxidant potentials and support traditional medicinal use for the treatment of diabetes mellitus and good source for natural antioxidants.These results indicated that stem barks of  Diabetes mellitus (DM) is a metabolic syndrome with multiple etiology, is characterized by chronic hyperglycemia together with disturbances in carbohydrate, protein and fat metabolism results from a decrease in circulating concentration of insulin (insulin deficiency), a decrease in the response of peripheral tissues to insulin (insulin resistance) or both . DM is tPatients with diabetes have lipid disorders and an increased risk of coronary heart disease, peripheral vascular disease and cerebrovascular disease ,8. HyperIn addition to hyperglycemia, systemic or local elevations in insulin may contribute to aberrant lipid metabolism and vascular wall function. Imperfect normalization of glucose metabolism by replacement insulin therapy may alter the concentrations and compositions of potentially atherogenic lipoproteins ,12. ChanSemecarpus anacardium (L.) is commonly known as Beula (Bangla), Bhallataka (Sanskrit), or Marking nut tree (English), belongs to the family Anacardiaceae [S. anacardium stem barks on glucose, GOT, GPT, total cholesterol (TC) triglyceride (TG), LDL and HDL in the blood and glycogen content in liver on alloxan induced-diabetic rats and antioxidant activity by DPPH scavenging assay.rdiaceae . In tradrdiaceae . Previourdiaceae , anti-arrdiaceae  as well rdiaceae . Phytochrdiaceae -23. The rdiaceae , hypochordiaceae , antitumrdiaceae , antioxirdiaceae  and hepardiaceae . It alsordiaceae -31. TherSemecarpus anacardium (Linn.) was collected from Madhupur forests, Tangail, April in 2009 and the plant authenticity was confirmed from the Bangladesh National Herbarium, Dhaka.Fresh stem bark of the plant The collected stem barks were washed and sun dried under shadow for several days. The dried stem barks were powdered in an electrical grinder after overnight drying in an oven below 50°C. The powdered plant barks were extracted with 95% ethanol at room temperature. The bottle were kept at room temperature and allowed to stand for several 7–10 days with occasional shaking and stirring. The extracts thus obtained were filtered through cotton and then through filter paper (Whatman Fitter Paper No. 1). The filtrate was defatted with petroleum ether for several times. Then, the defatted liquor was allowed to evaporate using rotary evaporator at temperature 40-45°C. Finally, a highly concentrated ethanol extract were obtained and kept in desiccators to dry to give a solid mass (1.67%).The standard drug, Metformin hydrochloride was the generous gift samples from Chemico Laboratories Ltd. Alloxan monohydrate was purchased from Sisco Research Laboratories Pvt. Ltd., Mumbai, India. Blood samples analyzed for blood glucose content by using BioLand G-423 glucose test meter . Serum TC and TG concentrations were analyzed by measuring absorbance by UV-spectrophotometer , using wet reagent diagnostic kits . Serum LDL and HDL Cholesterol measured by blood analyzer  using wet reagent diagnostic kits (Centronic GmbH Germany & Crescent Diagnostic Kits). SGOT and SGPT level were measured by using Crescent Diagnostic Kits and Human Gesellschaft fur Biochemical mbH Germany according to manufacturer’s protocol. DPPH , TCA (trichloroacetic acid), ferric chloride, Gallic acid and Quercetin were obtained from Sigma Chemical Co. USA. Ascorbic acid was obtained from SD Fine Chem. Ltd., Biosar, India. Folin-ciocalteu reagent was purchased from Merck, Germany. All chemicals and solvents were of reagent grade.Nine-week-old male Long Evans rats (150-180 g) purchased from ICDDRB, Dhaka, Bangladesh and were housed in animals cages under standard environmental conditions . The rats were fed with standard pellet diet obtained from ICDDRB, Dhaka and water ad libitum. The animals used in this study were cared in accordance with the guidelines on animal experimentation of our institute. The experimental procedures involving animals were conducted in accordance with the guidelines of Institute of Biological Sciences, University of Rajshahi, Bangladesh. The study protocol was approved by Institutional Animal, Medical Ethics, Biosafety and Biosecurity Committee (IAMEBBC) at the Institute of Biological Sciences, University of Rajshahi, Bangladesh.After fasting 16 h, diabetes was induced into rats by in intra-peritoneal injection (i. p.) of alloxan monohydrade (120 mg/kg), dissolved in saline . After 72 h, plasma glucose levels were measured by glucometer using a blood sample from tail-vein of rat. Rat with blood sugar level higher than 11.5-13.5 mmol/l are considered as moderate diabetic. Age-matched normal healthy rat were used as normal control.I.Normal Control II.Diabetic Control III.Diabetic Standard, IV.Diabetic + Extract 100 mg/kg V.Diabetic + Extract 200 mg/kg VI.Diabetic + Extract 400 mg/kg In the experiment, a total of 30 long Evans rats were divided into following six groups for the oral administration of extracts/drugs or vehicle.S. anacardium at doses of 100, 200 and 400 mg/kg body weight once daily, for 15 days using intrgastric tube. Group III received metformin (150 mg/kg body weight), while Group II serves as diabetic control (received vehicle 0.5% MC). The blood samples were analyzed for blood glucose content by Glucometer at 0th, 5th 10th and 15th days of treatment.The animals of Group IV, Group V and Group VI received bark extract of The body weights of rats of each group were measured on 0, 3, 6, 9, 12 and 15 days during the treatment period. At the end of experiments, all the rats were anesthetized with diethyl ether, chest were opened and blood samples were withdrawn directly by heparinised syringes from aorta of heart and stored in test tube containing anticoagulant (EDTA). Then liver, kidney, pancreases, heart, and lung were removed and cleaned of the surrounding tissues. The organ weights (OW) were measured immediately and the ratio of organ weights to body weight ratio (O/B) were calculated and parts of them were stored in 10% formalin and in a refrigerator at −20°C for histopathology and biochemical estimations respectively.The blood samples were centrifuged at 4000 rpm for 20 minutes to separate serum by using an Ultra-Centrifuge Machine  and the serum was preserved at −20°C to examine Total Cholesterol (TC), Triglyceride (TG), LDL-cholesterol (LDL), HDL-cholesterol (HDL), SGOT and SGPT concentrations by UV Spectrophotometric method  using wet reagent diagnostic kits according to manufacturer’s protocol (Centronic GmbH Germany & Crescent Diagnostic Kits).et. al [The liver glycogen content was determined according to the method described by Tarnoky K. et. al . BrieflyThe scavenging effects of samples for DPPH  free radical were monitored according to the method of Yen . AccuratThe percentage (%) of scavenging of the DPPH free radical was measured by using the following equation:0 = absorbance of the controlWhere, A1 = absorbance of the extract/ standardA50 was calculated.Then the percentage (%) of inhibition was plotted against log concentration and from the graph ICThe phytochemical tests have been performed by the standard methods  -36.Total phenol content in extract was determined by Folin-Ciocalteu reagent . ExtractThe total flavonoid content was determined using a method previously described by Kumaran K . 1 ml ofData were expressed as mean ± Standard Error of Mean (SEM). Statistical comparison were performed by one-way (ANOVA) followed by Dunnett’s Multiple Comparison Test and the values were considered as statistically significant when p values were less than 0.05 (p < 0.05). Statistical calculations and the graph were prepared using GraphPad Prism Software version 5.0 .The survival rate among the groups of rats with SA extract were 40%, 60% and 80% of 5 rats each in Group DC, Group SA 100 and Group SA 200 respectively, after 15 days treatment period. None of the rats died in Group DS, Group SA 400 and Group NC. The 15 days survival rate was significantly higher in Group SA 400 compared to Group DC   group. Treatment of diabetic rats with metformin and experimental groups significantly (p < 0.05) improved the level of glycogen content compared to DC group as shown in Figure The effects of different doses of extract on SGOT and SGPT levels were shown in Table Although no significant differences were observed between Groups NC, DC and DS, treatment with extracts reduced Hb % and RBC counts in rats. The effects were negatively correlated with the doses of extract Table .Table 650 value. The result of the DPPH scavenging activity of SA extract is shown in Figure 50 values of 72.24 μg/ml compared to ascorbic acid with IC50 17.81 μg/ml.The free radical scavenging activity of SA extract has been evaluated by using the DPPH free radical. The antioxidant quality of an extract is determined by the ICSemecarpus anacardium revealed the presence of flavonoid, steroid, glycoside, saponin, tannins, and triterpinoid . As shown in Figure The amount of total flavonoids determined by spectrophotometer is shown in Figure Diabetes is becoming the third “killer” of mankind after cancer and cardiovascular diseases, because of its high prevalence, morbidity and mortality . The chrEmblica officinalis and honey as reported by Rajendran et. al [S. anacardium reduced blood sugar significantly in glucose loaded hyperglycemic rats and produced more intense hypoglycemia in alloxan induced diabetic rats [The present investigation revealed that none of the rats died in Group DS, Group SA 400 and Group NC. The survival rate was significantly higher in Group SA 400 than that in Group DC (p < 0.05). The nuts showed no mortality when given in combinations with n et. al . The blotic rats .S. anacardium bark exhibited significant antihyperglycemic and antihyperlipidemic activity in alloxan-induced diabetic rats, and the activity was comparable to that of the standard antidiabetic drug, Metformin HCl .No significant changes in the body weight and organ weight to body weight ratio were observed among the treatment groups. Although the weight of liver significantly decreased, the weight of heart, kidney, lung and pancreas did not changed after 15 days of treatment. Treatment with extract improved liver weight significantly in Group SA 200 and SA 400. Oral Administration of extract significantly (p < 0.05) reduced the total cholesterol, triglycerides and LDL-CH levels and increased HDL-CH in Groups SA 100, SA 200 and SA 400 and all the effects were dose-dependent. Among the treatment groups Group SA400 significantly improved dyslipidemia. The ethanol extract of the plant et. al with nuts of the same plant in cholesterol fed rabbits [A significant elevation of glycogen content was observed in all the treatment groups. However, the effect was most prominent in Group SA 400 rats. Similar observations were obtained by Sharma  rabbits . On the  rabbits . However rabbits . The steS. anacardium stem bark showed the presence of steroids, triterpenoids, flavonoids, glycosides, saponins and tannins. Antioxidant activity is correlated with the total phenolic content and total flavonoids content in the extract that were likely to contribute to the radical scavenging activity. It is known that only flavonoids of a certain structure and particularly hydroxyl position in the molecule determine antioxidant properties, in general these properties depend on the ability to donate hydrogen or electron to a free radical.The effect of antioxidants on DPPH radicals is thought to be due to their hydrogen donating ability . RadicalThe present investigation established the pharmacological evidence to support the folklore claim and the plant barks have antidiabetic and antioxidant activity. However, further investigations were warranted to isolate bioactive compounds from stem bark, to observe their antidiabetic and antioxidant effects and to find out the possible mechanism action for their beneficial effects both.S. anacardium possessed highest phenolic and flavonoid compound and exhibuted strong antidiabetic and antioxidant activities, which were comparable to the commercial antidibetic drug metformine and antioxidant ascorbic acid. This seems that the Semecarpus anacardium extract can be used as natural antidiabetic and antioxidant agent.The present study indicated that the stem barks of the plant


Through education, a social group transmits accumulated knowledge, skills, customs, and values to its members. So far, to the best of our knowledge, the association between educational attainment and neural correlates of emotion processing has been left unexplored. In a retrospective analysis of The Netherlands Study of Depression and Anxiety (NESDA) functional magnetic resonance imaging (fMRI) study, we compared two groups of fourteen healthy volunteers with intermediate and high educational attainment, matched for age and gender. The data concerned event-related fMRI of brain activation during perception of facial emotional expressions. The region of interest (ROI) analysis showed stronger right amygdala activation to facial expressions in participants with lower relative to higher educational attainment (HE). The psychophysiological interaction analysis revealed that participants with HE exhibited stronger right amygdala—right insula connectivity during perception of emotional and neutral facial expressions. This exploratory study suggests the relevance of educational attainment on the neural mechanism of facial expressions processing. A large body of evidence substantiates the general consensus that individual differences, cognitive performance, and cerebral structure and function are modulated by environmental aspects such as cultural exposure, occupational attainment, and cognitive enrichment factors , insula and anterior cingulate cortex  and those with intermediate-to-LE. We defined the groups of subjects with HE and with LE based on the mean of the variable “Years of education” presented in Table For this retrospective analysis, data of participants were selected from the functional magnetic resonance imaging (fMRI) part of the multicenter longitudinal cohort study NESDA. A detailed description of the fMRI NESDA study including participants and selection criteria is presented elsewhere  group. As age  corresponding to 13 to 18 years of education.Out of the 58 healthy participants included in the main NESDA study  = 2300 ms, echo time (TE) = 28.0 ms (TE = 30 ms at AMC and LUMC), flip angle 90°, matrix size 64 × 64 (AMC and LUMC: 96 × 96), interleaved slice acquisition, 39 slices (35 at AMC and LUMC), 3 × 3 (AMD and LUMC: 2.29 × 2.29) mm in-plane resolution, and 3 mm slice thickness). Anatomical images included a sagittal 3D gradient-echo T1-weighted sequence .Images were acquired on a Philips 3T MR-system located at the Neuroimaging Center UMCG, AMC, LUMC using a SENSE-8 channel coil at UMCG and LUMC, and SENSE-6 head channel coil at AMC. For each participant a series of echo planar imaging (EPI) volumes—sensitive to the blood oxygenation level dependent (BOLD) effect—were obtained, entailing a T2Functional imaging data were preprocessed and analyzed using the statistical parametric mapping software package (SPM5) implemented in Matlab 6.5 (The MathWorks Inc.). A detailed description of the preprocessing was previously presented  as between subjects factor, and centers as factor of nuisance. Significance threshold was set to p < 0.05 Family-Wise Error (FWE) corrected for multiple comparisons for the main effect. Region of interest (ROI) analyses were conducted to increase statistical sensitivity of a group effect on the brain areas involved in emotional processing with an initial threshold of p < 0.001. Small volume correction (SVC) with a threshold of pFWE < 0.05 was applied within a priori defined regions: fusiform gyrus, amygdala, mPFC, ACC and insula using anatomical masks created with Wake Forest University PickAtlas  of the amygdala time series and a vector coding for the task effect . In the second level analysis, we included the contrast images for the PPI effect which, during each facial expression, indicated a positive functional connectivity of amygdala with the rest of the brain. Two separate repeated measurement analyses were conducted, one for the left and one for the right amygdala. Facial expressions were defined as within subject factor, and group as between subjects factor. Scan sites were defined as nuisance factors. The ROI analyses were conducted employing ACC, insula, fusiform, and mPFC masks defined with WFU PickAtlas using and initial threshold of p < 0.001 uncorrected and p < 0.05 FWE SVC for the ROI area. Outside these regions of interest the FWE correction was applied.In an additional analysis we tested for an effect of educational attainment on the functional connectivity between amygdala—ACC, amygdala—insula, amygdala—fusiform and amygdala—mPFC. For this purpose a Psycho-Physiological Interaction (PPI) analysis  = 0.279, p > 0.05 (2-tailed)), MADRS score (t(26) = 0.717, p > 0.05 (2-tailed)), BAI (t(26) = 1.337, p > 0.05 (2-tailed)), gender (χ2(1)= 0.144, p > 0.05 (2-tailed)) or on handedness (Fisher’s Exact test p = 0.222). Also, no significant correlation was found between the participant’s age and years of education (r(12) = −0.018). Finally, no group effect was found on the RT to facial expressions  = 0.015, p = 0.904).Characteristics of the samples are displayed in Table Z = 6.31 and right: , Z = 7.50), right middle occipital gyrus , left amygdala , right middle frontal gyrus  activation at pFWE < 0.05 (whole-brain correction). Right amygdala  activation to facial expressions across all participants was observed at a slightly more lenient threshold of p < 0.05 false discovery rate.Facial expressions (>scrambled) elicited increased bilateral fusiform gyrus  and no group difference.We also checked that our baseline does not elicit an effect on any of the areas found activated in response to facial expressions of emotion. Thus, we conducted an independent t test, controlling for centers and, across both groups, we found a main effect of scramble face perception, in right middle occipital gyrus . Participants with HE showed a stronger right amygdala—right insula (, p < 0.05, SVC, Figure Regarding amygdala functional connectivity we found an effect of group on right amygdala—right insula connectivity (42, 3, 9], , 3, 9, ZThe present study investigated the possible modulatory role of educational attainment on the neural circuitry involved in processing of facial expressions. The results support the hypothesis of a putatively formational effect of education, that may affect emotion processing in general. We found that at the neural level, amygdala response to facial expressions was negatively modulated by educational attainment. Additionally, we found that the connectivity of the right amygdalar complex with insula was also associated with educational attainment in a positive manner. Our study provides—to the best of our knowledge—the first neuropsychological evidence of the modulatory association of education on perception of facial expressions.Our findings are in line with previous studies on amygdala response to facial perception Todorov, . We founMorris et al.  reportedAnother outcome of our analyses was that, relative to LE participants, subjects with HE displayed a stronger amygdala—insula functional coupling during perception of facial expressions. As in the previous analysis, no group by facial expressions of emotion interaction was found indicating a general effect of educational attainment on amygdala connectivity. Insula is implicated in “generation of affective state in response to emotive stimuli” (Phillips et al., It may be of interest for further studies to explore the effect of educational attainment on emotional perception intensity and accuracy, and also on cognitive processes. As the lack of additional data on socioeconomic or environmental factors is a limitation of the present study, these formative effects could be considered in future studies.The present study suggests that educational attainment modulates both the neural response and neural network involved in processing facial expressions. In particular, it shows that HE may be associated with reduced amygdala responses to facial expressions and stronger amygdala—insula coupling during face perception. As emphasized by Duarte et al. , it may The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The aim of this study was to compare the effectiveness of velvet antler (VA) from different sections for promoting longitudinal bone growth in growing rats. VA was divided into upper (VAU), middle (VAM), and basal sections (VAB). An in vivo study was performed to examine the effect on longitudinal bone growth in adolescent rats. In addition, in vitro osteogenic activities were examined using osteoblastic MG-63 cells. VA promoted longitudinal bone growth and height of the growth plate in adolescent rats. Bone morphogenetic protein-2 (BMP-2) in growth plate of VA group was highly expressed compared with control. The anabolic effect of VA on bone was further supported by in vitro study. VA enhanced the proliferation, differentiation, and mineralization of MG-63 cells. The mRNA expressions of osteogenic genes such as collagen, alkaline phosphatase, and osteocalcin were increased by VA treatment. These effects of in vivo and in vitro study were decreased from upper to basal sections of VA. In conclusion, VA treatment promotes longitudinal bone growth in growing rats through enhanced BMP-2 expression, osteogenic activities, and bone matrix gene expressions. In addition, present study provides evidence for the regional differences in the effectiveness of velvet antler for longitudinal bone growth. Longitudinal bone growth, which determines the size and shape of the body frame, proceeds at a rate distinct from that of muscle and other tissues and is controlled by specific mechanisms. During childhood and adolescence, the long bones grow at the ends of the bones which occur at the growth plate by endochondral ossification of epiphyseal plate . Growth The current methods for increasing final height include treatment with a growth hormone alone or in combination with an analog of gonadotropin-releasing hormone . HoweverVelvet antler (VA), the immature antler of male deer, is traditionally used for thousands of years in Asian countries, such as Korea, China, Taiwan, and Mongolia. It is currently estimated that the global production of velvet antlers is near to 1300 tons/year, which is still rapidly growing to meet the requirements of medicinal markets . As a tyA whole VA stick is usually divided into four portions, with the value decreasing from the tip to the base. Chemical analyses of VA revealed that there are regional differences in chemical composition: the contents of proteins and lipids decrease downward from the tip to the base, while those of ash, calcium, and collagen increase  suggestiThe objective of the present study was to compare the effect of VA from different sections on longitudinal bone growth of adolescent rats and elucidate the underlying mechanisms for the effect.VA, obtained from farmed Elk deer 75 days after casting, was kindly provided by the Animal Genetic Resources Station . VA was divided into upper (VAU), middle (VAM), and basal (VAB) sections . The crin = 8); vehicle  and upper (VAU), middle (VAM), and basal (VAB) sections of antler extract (100 mg/kg) were orally administered daily via stomach tube for 5 consecutive days. Treatment dose was decided from the preliminary study. On the sixth day, all rats were sacrificed under chloral hydrate anesthesia for tissue analysis.Longitudinal bone growth was determined in 3-week-old male Sprague-Dawley rats . The experimental procedures were performed in accordance with protocols approved by the Institutional Animal Care and Use Committee, Semyung University (smecae 15-09-01). The animals were housed under controlled temperature and lighting conditions. The rats were randomly divided into four groups . Bone growth was measured by measuring the gap between fluorescent line formed by calcein and the epiphyseal end line of the growth plate at three different locations using a fluorescent microscope , and the averages were obtained.To measure the effect on longitudinal bone growth rate, calcein was used as a fluorescence marker to label the bone line on the surface of the tibia. Calcein plays the role of fluorescent dye under ultraviolet illumination. Calcein  was injected intraperitoneally 24 h before sacrifice. The dissected tibias were fixed in 0.1 M phosphate buffered formalin for 2 h, decalcified, and dehydrated by immersing in 30% sucrose for 1 day at 4°C. Dehydrated bone was sectioned longitudinally at sagittal sections of proximal part with a thickness of 40 μm using a sliding microtome as described above and stained with cresyl violet. The growth plate height was measured at three different locations and the averages were obtained.Cresyl violet staining (Sigma-Aldrich) was used to stain the chondrocytes in the growth plate of the samples. Each tibia sample was sectioned longitudinally at a thickness of 40 For the detection of bone morphogenetic protein-2 (BMP-2) in the growth plate, the dehydrated tibia sections were incubated overnight in 1% Triton X-100 containing goat BMP-2 antibody  at room temperature. Then, sections were incubated with anti-goat antibody  for 60 min and stained with 0.05% 3,3-diaminobenzidine  containing 0.03% hydrogen peroxide.2 atmosphere in Dulbecco's Modified Eagle's Medium  containing 10% heat-inactivated FBS  and 100 U/mL penicillin/streptomycin (Sigma-Aldrich). Bone marrow cells prepared from the femur of male ICR mice were cultured in α-modified minimal essential medium  containing 10% heat-inactivated FBS. After 3 days of culture, floating cells were removed and attached cells were used as osteoclast precursors.Human osteoblast-like MG-63 cells were obtained from the American Type Culture Collection . Cells were cultured at 37°C in a 5% COμg/mL) and incubated for 24 h. Subsequently, BrdU was added to each well and reincubated for 2 h. After 14 h of incubation at 37°C, labeling media was removed, cells were fixed, and the cells with BrdU label in the DNA were located with peroxidase-conjugated anti-BrdU antibody solution. Then bound anti-BrdU with substrate was colorimetrically measured with a microplate reader  at 450 nm.The effects of VA from different sections on proliferation of MG-63 cell were determined by a colorimetric immunoassay kit  which is based on quantitating bromodeoxyuridine (BrdU) incorporation into the newly synthesized DNA of replicating cells. MG-63 cells were seeded  in 96-well plates containing DMEM with 10% FBS and allowed to adhere overnight. The media were replaced with DMEM containing VA samples ; VA samples (50 and 100 μg/mL) were added to the cells and incubated for another 24 h. The 50 and 100 μg/mL concentrations were chosen since the maximum effect and plateau had been gained under those two concentrations in cell proliferation assay results. The cells were then rinsed with PBS and lysed in 300 μL of 0.2% of Triton X-100. The cell lysates were centrifuged at 13,000 rpm for 5 min. The supernatant of the lysate was used for the measurement of ALP activity by measuring the release of p-nitrophenol from p-nitrophenylphosphate at pH 9.8.Alkaline phosphatase (ALP) activity was measured using enzymatic assay. MG-63 cells, incubated in DMEM, were seeded in 12-well plates  and incubated for 24 h. The medium was changed to osteogenic medium (DMEM with 10 mM μg/mL) were added to each well and incubated for 7 days. Cells were fixed with Bouin's fluid and stained with 0.1% Sirius red  in a saturated aqueous solution of picric acid for 30 min. Stained dye was dissolved and the absorbance was measured at 540 nm.Collagen synthesis was measured using picrosirius red method . MG-63 cThe formation of calcium phosphate was determined by alizarin red-S assay . MG-63 cμg/mL) was added to each well and incubated for 24 h. Total RNA was extracted using RNeasy® Protect Mini Kit , and cDNA was synthesized from mRNA using QuantiTect® Reverse Transcription Kit (Qiagen). Real-time PCR was performed using QuantiTect™ SYBR® Green PCR Kit (Qiagen) according to the manufacturer's protocol. The PCR primer sequences are shown in Real-time PCR was performed to analyze relative gene expression. MG-63 cells were seeded in 6-well plates  and allowed to adhere overnight. The medium was changed to osteogenic medium; VA sample  and M-CSF (30 ng/mL) in the presence of VA samples. Osteoclasts were identified by staining tartrate-resistant acid phosphatase (TRAP), a marker enzyme of osteoclasts , using cP values of less than 0.05 (P < 0.05) indicated significant differences. All in vitro experiments were performed with triplicate independent samples.The data were expressed as mean ± SD. One-way ANOVA followed by Tukey's multiple comparison test was performed for statistical analysis (GraphPad Prism ver. 6), and μm/day, and administration of VAU significantly increased the longitudinal bone growth to 718.6 ± 48.6 μm/day (P < 0.05). VAU, VAM, and VAB caused 16.4%, 9.6%, and 2.7% increase in longitudinal bone growth, respectively, compared with control, suggesting that the effect decreases downward from upper section to the base. Although VAM caused slight increase in longitudinal bone growth, statistical significance was not observed. The present study provides first evidence for the effectiveness as well as regional differences of VA on longitudinal bone growth in adolescent rats.To determine the growth per day, instead of the total growth over a period, exact methods for measurement are necessary. Recently, the use of tetracycline or calcein as intravital markers of the growth process to label mineralizing bone in the rat has been reported , 18. In Recently, Tseng et al.  comparedThe growth plate, which is located at the distal end of the bone, is the main location where longitudinal bone growth occurs owing to the stimulation of chondrocyte proliferation. The growth plate consists of four distinctive histological zones beginning with the resting zone and extending through the proliferative, hypertrophic, and ossification zones . The topμm, and administration of 100 mg/kg VAU and VAM significantly increased the height of the growth plate to 567.4 ± 23.4 μm and 566.2 ± 20.6 μm, respectively . MG-63 cell proliferation reached peak value at 50 μg/mL VAU treatment exhibiting 119% of the basal value, while VAM treatment reached peak at 100 μg/mL exhibiting 111% of the basal value. VAB at the highest concentration (100 μg/mL) significantly (P < 0.05) increased cell proliferation compared to the control exhibiting 106% of the basal value. Effects of VAU and VAM on increasing cell proliferation were significantly higher than those of control at 50 and 100 μg/mL concentration. Therefore, the effect of VA on ALP activity in MG-63 cells was further determined at 50 and 100 μg/mL concentration. VAU treatment significantly increased the ALP activity, an early-stage osteoblasts differentiation marker, at 100 μg/mL concentration  and VAM (6.2%) treatment compared with control, and the increased collagen synthesis was significantly higher than VAB. Cellular calcium deposition was examined using Alizarin red-S staining. Significant increases in mineralization were found with VAU and VAM treatment compared to the control and VAB group. Mineralization was significantly increased to 118% and 107% of the control by VAU and VAM treatment, respectively. The anabolic effect of VA on bone was further supported by the findings in this study demonstrating that VA enhanced the proliferation, differentiation, and mineralization of osteoblastic cells. Furthermore, the effects of VAU were greater than the effect of VAM, suggesting that the effects were decreased from the upper section to the base section. Lee et al. [Bone regeneration is regulated by a fine balance of biochemical and cellular events that ultimately stimulate osteoblasts to produce new tissue, in particular, new extracellular matrix composed mainly of collagen. The collagen matrix is then mineralized via ALP activity, which induces formation of calcium phosphate crystal seeds. Bellows et al.  reportede et al.  reported COL, ALP, OCN, and OPN in MG-63 cells were measured by real-time PCR . The effects of VAU, VAM, and VAB on COL mRNA expressions were increased 7.7-, 6.0-, and 5.8-fold, respectively, while those of OCN expressions were increased 22.5-, 20.2-, and 15.0-fold, respectively. The expressions of ALP mRNA were significantly increased with VAU (3.7-fold) and VAM (3.5-fold) treatment compared with control and VAB group (1.2-fold). OPN mRNA expressions were not altered by all sections of VA. These data suggest that VA may play role at early stage as well as late stage of osteoblast differentiation, and VA-induced COL, ALP, and OCN gene expression could accelerate mineralization, subsequently leading to an increase in the extent of calcium deposition. Furthermore, the effects on osteogenic gene expressions were decreased from upper section to basal section of VA, and the effect of VAM was similar to VAU. Lee et al. [It is known that preosteoblastic cells produce proteins of the extracellular matrix including collagen at first and then to successively produce alkaline phosphatase (ALP) and osteopontin/osteocalcin during mineralization phase . Accorditime PCR . All sece et al.  reported Bacillus-fermented VA extract has been reported to inhibit osteoclast differentiation [ Cordyceps militaris, contain more sialic acid than nonfermented VA, and the stimulatory effects on osteoblastic cell proliferation and ALP production were increased by fermentation. Therefore, the effect of nonfermented VA in the present study may not be strong enough to induce osteoclast differentiation.Osteoclasts are the only cell type capable of resorbing mineralized bone. To examine the effect of VA on osteoclast differentiation, bone marrow-derived osteoclast precursor cells were cultured in osteoclastogenic media with different parts of VA extract. TRAP-positive multinuclear osteoclasts were generated in response to stimulation from M-CSF and RANKL. The results showed that the addition of VA did not affect osteoclast formation (data not shown).ntiation  which dontiation  suggesteIn conclusion, results of the present study suggest that VA promotes longitudinal bone growth in adolescent rats through, at least in part, MG-63 cell osteogenesis, and the effect was decreased downward from upper section to basal section. VA stimulated the proliferation, differentiation, and mineralization of osteoblasts through upregulation of osteogenic gene expressions. These results support the stimulating nature of VA toward the function of osteoblastic cells. Although the molecular mechanisms underlying osteoblast differentiation remain to be defined, the results of the present study suggest that BMP-2, which plays an important role in regulating osteoblast differentiation and subsequent bone formation, might participate in these mechanisms. Furthermore, the present study provides strong evidence for the regional differences in the effectiveness of VA in longitudinal bone growth. However, further studies are needed to elucidate the bioactive chemical constituents associated with these effects.The present study has some limitations. Only a single dose of VA was used in the in vivo study. The dose for traditional use of VA is usually 1 g/day, taken all at once or divided throughout the day. Therefore, the dose of 100 mg/kg/day given to the experimental rats in this study was equivalent to the traditional human dosing regimen of 1 g/day based on body surface area conversion, where animal dose is equal to human equivalent dose × 6.2, assuming that an adult person's weight is 60 kg .


Measurement of serum β-hCG concentration commonly used to diagnose tubal ectopic pregnancy (EP) and follow up patients treated conservatively. The aim of this study was to determine the predictive value of maternal serum β-hCG concentration in ruptured tubal ectopic pregnancy to help physicians identify those women who are at greatest risk.This is a cross-sectional study conducted on all women with a diagnosis of tubal ectopic pregnancy who were treated in Alzahra Hospital, in Rasht, from March 2002 to February 2011. The data was collected for each woman from medical records and included age, parity, gravidia, gestational age, primary level of serum β-hCG, rupture status, past history of pelvic inflammation disease, EP, abortion, and intrauterine contraceptive device use. Women with tubal rupture were compared to those without rupture. Statistical analysis was conducted by SPSS 19 for Windows. A total of 247 cases of tubal ectopic pregnancy were recorded during the study period. One hundred and ninety seven (79.8%) were cases with unruptured EP and 50 patients (20.2 %) were cases with ruptured EP. The mean level of β-hCG was significantly higher in patients with ruptured EP compared to patients with unruptured EP (p=0.03). Logistic regression analysis revealed that >1750 IU/ml of β-hCG levels  was the significant risk factors for tubal rupture. Higher β-hCG levels seem to be significant risk factors for rupture of a tubal EP. Ectopic pregnancy (EP) occurs when fertilized ovum implants outside endometrial cavity and the incidence is approximately 2% of all pregnancies , 2. The Hemorrhage from rupture of tubal ectopic pregnancy is still the leading cause of pregnancy related maternal death in the first trimester and accounts for 10-15% of all pregnancy related deaths, despite improved diagnostic methods leading to earlier detection and treatment   -8. Some 00 IU/L) . Thus in present study, we aimed to determine the predictive value of maternal serum β-hCG concentration for rupture in patient with tubal ectopic pregnancy.This was a cross-sectional study conducted on all women with tubal ectopic pregnancy who were treated in the Al-Zahra Hospital in Rasht, after approving by the Ethical Committee of Guilan University of Medical Sciences, from March 2002 to February 2010. Patient with heteropic pregnancy was excluded from study. Ectopic pregnancy was confirmed by positive β-hCG and transvaginal ultrasonography findings in all patients. The ultrasonographic findings that designate the ectopic mass included gestational sac like anechoic mass surrounded by peripheral hyperechogenic halo that does not contain a viable embryo and located in the paraovarian region, solid or complex irregularly boarded mass which resembles a haematosalpinx or pelvic haematoma around the paraovarian region, and ectopic gestational. Moreover, gestational age was calculated according to the last menstrual period (LMP) at the time of admission. The information collected for all woman from medical records included; age, parity, gravidity, gestational age, serum β-hCG concentration at the time of admission in hospital, rupture status, past history of pelvic inflammatory disease (PID), previous EP, number of normal pregnancy, abortion history, and the use of intrauterine contraceptive device (IUD).Ruptured EP for patients was defined according to physical examinations and ultrasonographic finding . Finally, the study group was divided into two subgroups: ruptured EP and unruptured EP.Statistical analysist-test and Mann-Whitney-Wilcoxon test for independent samples. Pearson’s chi-square and Fisher’s exact test were also applied for comparison of groups where appropriate. Receiver operating characteristic (ROC) curve analysis was used to find cut-off point for β-hCG variables. The sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) were calculated. Logistic regression analysis was used to identify predictors for tubal rupture in EP. The Statistical Package for the Social Sciences, version 18.0, SPSS Inc, Chicago, Illinois, USA were used for the data analyses. P<0.05 was considered to indicate statistical significance.The results are expressed as mean±SD for continuous variables or percentage and number for categorical variables, as appropriate. Comparison of maternal, obstetric history and maternal serum β-hCG concentrations among two subgroups was performed by Student’s A total of 249 cases of tubal ectopic pregnancy were recorded during the study period. Two patients were excluded for heteropic pregnancy. Of these, 197 patients (79.8%) were cases with unruptured and 50 (20.2%) were cases with ruptured tubal ectopic pregnancies. Age was a statistically significant factor between women with unruptured and ruptured tubal ectopic pregnancies (p<0.0001). The mean gestational age was significantly lower and mean level of β-hCG was significantly higher in the ruptured group compared with the unruptured group . Ectopic pregnancy was occurred in the right tubal in 109 patients (55.33%) of unruptured groups and in left tubal in 27 patients (60%) of ruptured groups. No significant associations existed regarding parity, gravidity, the number of previous normal pregnancies, past history of PID, previous EP, abortion history, and IUD use . The ROCIn the noninvasive management of a tubal pregnancy, the clinician should focus on selecting those patients in whom the risk of tubal rupture was higher. The occurrence of tubal rupture in EP ranges from 18.0% to 64.5% as reported in previous large population-based studies. The high rupture rates may be caused by lack of diagnostic tools such as transvaginal ultrasound and β-hCG measurements or to delayed diagnosis , 11. Theet al study, prevalence of women with rupture EP was 29.5% and the mean age of these patients was more significantly compared to unruptured EP . Therefore, middle-age is important risk factors for tubal rupture. In Berlingieri tured EP . In cont. Thereforegnancy . It seemet al likely suggested that the probability of predicting risk of tubal rupture or active bleeding is over 10% when gestational age is >10 weeks and serum β-hCG levels >8500 IU/m. However, the serum β-hCG cut-off level (>1300 IU/ml) in Mol et al study was lower than our study    . This fiy groups . Furthery groups , 19. It y groups , 12.In conclusion, based on our results, β-hCG concentration is the best predictive factor for rupture in tubal ectopic pregnancies, which may contribute to the choice of treatment approaches for women who desire future pregnancy. It is recommended to accomplish more studies to determine the level of serum β-hCG on which risk of rupture is more significantly.


Psoroptes cuniculi.Limited available sequence information has greatly impeded population genetics, phylogenetics and systematics studies in the subclass Acari (mites and ticks). Mitochondrial (mt) DNA is well known to provide genetic markers for investigations in these areas, but complete mt genomic data have been lacking for many Acari species. Herein, we present the complete mt genome of the scab mite P. cuniculi was collected from a naturally infected New Zealand white rabbit from China and identified by morphological criteria. The complete mt genome of P. cuniculi was amplified by PCR and then sequenced. The relationships of this scab mite with selected members of the Acari were assessed by phylogenetic analysis of concatenated amino acid sequence datasets by Bayesian inference (BI), maximum likelihood (ML) and maximum parsimony (MP).P. cuniculi is the same as those of Dermatophagoides farinae (Pyroglyphidae) and Aleuroglyphus ovatus (Acaridae), but distinct from those of Steganacarus magnus (Steganacaridae) and Panonychus citri (Tetranychidae). Phylogenetic analyses using concatenated amino acid sequences of 12 protein-coding genes, with three different computational algorithms , showed the division of subclass Acari into two superorders, supported the monophylies of the both superorders Parasitiformes and Acariformes; and the three orders Ixodida and Mesostigmata and Astigmata, but rejected the monophyly of the order Prostigmata.This mt genome  is circular and consists of 37 genes, including 13 genes for proteins, 22 genes for tRNA, 2 genes for rRNA. The gene arrangement in mt genome of P. cuniculi represents the first mt genome of any member of the family Psoroptidae. Analysis of mt genome sequences in the present study has provided new insights into the phylogenetic relationships among several major lineages of Acari species.The mt genome of  The metazoan mitochondrial (mt) genome possess a circular double strand DNA that varies in size from 15 to 20 kb, generally encoding for 36–37 genes: 12–13 protein-coding genes (PCGs), two ribosomal RNAs (rRNA) genes and 22 transfer RNAs (tRNA) -10. In set al.[et al.[et al.[et al. [et al. [et al.[et al.[Currently, the Acari encompasses over 55,000 species (ticks and mites) and forms the largest group among the class Arachnida . Many spet al. inferredl.[et al. and Burgl.[et al. inferred.[et al.  and Burg [et al.  showed s. [et al. showed sl.[et al. showed tP. cuniculi in the family Psoroptidae of the order Astigmata. P. cuniculi is a common worldwide parasite of rabbits, causing considerable economic loss due to weight loss, decreased food consumption, vestibular dysfunction, meningitis and death [P. cuniculi and 50 other Acari species that have been sequenced to date.In the present study, we sequenced the complete mt genome of the scab mite nd death . Also, wP. cuniculi was collected, was handled in accordance with good animal practices required by the Animal Ethics Procedures and Guidelines of the People’s Republic of China.This study was approved by the Animal Ethics Committee of The Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences (Permit code. LVRIAEC2013-006). The New Zealand white rabbit, from which P. cuniculi samples were collected from a naturally infected New Zealand white rabbit in Sichuan Province, China. The adult mites were isolated from ear cerumens of the rabbit, according to the previous descriptions [P. cuniculi was conducted by morphological criteria and site of predilection [SV Genomic DNA Purification System, Promega). In brief, P. cuniculi collected from the same host were ground in a mortar and subsequently suspended in 275 μL lysis buffer . After vortexing, the sample was incubated at 55°C for 16–18 h, and the DNA was extracted by shaking with 250 μL Wizard SV Lysis Buffer. After centrifugation, the DNA was washed, eluted and stored.riptions . The preilection , then fiP. cuniculi as shown in Table cox1 was amplified using the primers COIF/COIR [Dermatophagoides farinae and D. pteronyssinus[Taq buffer (Mg2+ Plus), 10 pmol of each primer, 1 U Ex Taq polymerase (Takara), 1 μL of DNA sample in a thermocycler  under the following conditions: 94°C for 5 min ; then followed by 35 cycles of 94°C for 30 s (denaturation), 36 ~ 60°C for 30 s , and 68°C for 1 ~ 4.5 min (extension) according to the product length; with a final extension step at 68°C for 10 min. Each PCR reaction yielded a single band as detected in a 1% (W/V) agarose gel upon ethidium-bromide staining (not shown). PCR products were subsequently sent to Invitrogen Biotechnology Company  for sequencing using a primer-walking strategy.Nine pairs of PCR primers were used to amplify overlapping segments of the complete mt genome of OIF/COIR . The othonyssinus,23. All D. farinae available using the computer program MAFFT 7.122 [UCN, tRNA-Ile, tRNA-Pro, tRNA-Val and tRNA-Ala) were identified by recognizing potential secondary structures and anticodon sequences by eye, and two rRNA genes were predicted by comparison with those of other mites previously reported [Sequences were assembled manually and aligned against the complete mt genome sequences of FT 7.122  to identFT 7.122 , and aliFT 7.122  and the FT 7.122  were usereported ,23.Centruroides limpidus (GenBank accession number NC_006896) as an outgroup [For comparative purposes, amino acid sequences predicted from published mt genomes of 50 Acari species were also included in the present analysis, using the scorpion outgroup . The 12 outgroup  using thoutgroup  based onoutgroup ; the firoutgroup  using thoutgroup , with inP. cuniculi was 14,247 bp in length , 22 tRNA genes, two rRNA genes and one D-loop region  and seven tRNA genes  encoded on the minority strand (N-strand), all other genes were encoded on the majority strand (J-strand)  and Aleuroglyphus ovatus (Acaridae), but distinct from those of Steganacarus magnus (Steganacaridae) and Panonychus citri (Tetranychidae). The nucleotide composition of the complete mt genome of P. cuniculi is A = 29.4%, T = 41.7%, G = 16.8% and C = 12.1%, with a typically high A + T content of 71.1% within the range of values found in D. farinae (71.4%) and D. pteronyssinus (72.6%)  use ATA, 6 genes  use ATG, 2 genes  use ATT and 1 gene (nad2) uses TTG as start codon, respectively  use TAA and 2 genes (atp8 and nad6) use TAG as termination codon, respectively , however, some authors later demonstrated that there are two distinct groups: Acariformes and Parasitiformes -37, and et al.  confirme. [et al. tested t. [et al., however. [et al.. However. [et al..et al. (2007) showed that the order Astigmata is located on the outside of the order Oribatida, usually as sister-group of the order Endeostigmata [et al.[The superorder Parasitiformes is composed of four orders: Opilioacarida, Holothyrida, Ixodida, and Mesostigmata, and the superorder Acariformes is divided into two suborders: Sarcoptiformes  and Trombidiformes (Prostigmata or Actinedida) . Domes estigmata . Howevera [et al. speculatSteganacarus magnus, from the Steganacaridae was included in the present study. No species from the superorder Opilioacariformes and order Holothyrida was included in our analyses. Therefore, expanding taxon sampling from these lineages of Acari is clearly the next step for phylogenetic studies of Acari using mtDNA. In addition, phylogenetic analysis in the present study was based only on mtDNA sequences, so we believe it is still necessary to employ nuclear genomic sequences to provide additional evidence for phylogenetic analyses and genome evolution of the Acari.In the present study, our results provide strong support that families Ixodidae and Argasidae are sister taxa, consistent with those of previous studies ,42. HereP. cuniculi of animal health significance, and represents the first mt genome study of any member of the family Psoroptidae. Phylogenetic analyses of the mt genome sequences of the P. cuniculi species, in combination with 50 other Acari species, support the monophylies of the superorders Parasitiformes and Acariformes; the orders Ixodida, Mesostigmata and Astigmata, however, reject the monophyly of the order Prostigmata. Our results indicate the need to test hypotheses of Acari phylogeny with more taxa and different molecular markers.The present study determined the mt genome of the mite species The authors declare that they have no competing interests.XBG, GHL, HQS, TYL and GYY performed the experiments, analyzed the data and drafted parts of the manuscript. XQZ conceived and funded the study, revised and edited the manuscript. All authors read and approved the final manuscript.


The aim of this study was to evaluate the incidence and determinants of AKI in a large cohort of cardiac arrest patients.We reviewed all patients admitted, for at least 48 hours, to our Dept. of Intensive Care after CA between January 2008 and October 2012. AKI was defined as oligo-anuria (daily urine output <0.5 ml/kg/h) and/or an increase in serum creatinine . Demographics, comorbidities, CA details, and ICU interventions were recorded. Neurological outcome was assessed at 3 months using the Cerebral Performance Category scale .A total of 199 patients were included, 85 (43%) of whom developed AKI during the ICU stay. Independent predictors of AKI development were older age, chronic renal disease, higher dose of epinephrine, in-hospital CA, presence of shock during the ICU stay, a low creatinine clearance (CrCl) on admission and a high cumulative fluid balance at 48 hours. Patients with AKI had higher hospital mortality , but AKI was not an independent predictor of poor 3-month neurological outcome.AKI occurred in more than 40% of patients after CA. These patients had more severe hemodynamic impairment and needed more aggressive ICU therapy; however the development of AKI did not influence neurological recovery. Most studies on patients after resuscitated cardiac arrest (CA) have focused on survival or the extent of brain dysfunction ; the preet al. [Recently, Roberts et al.  reportedet al. , were inet al. [Several studies have also highlighted the high prevalence of acute kidney injury (AKI) in CA patients and how this may influence patient outcome in this setting. Domanovits et al.  showed tet al. . In addiet al. . Other set al. -11. Neveet al. ,12. Moreet al. ,14.Finally, AKI may have an impact on neurological recovery. In an experimental model of renal injury, AKI was found to contribute to the inflammatory injury in the hippocampus, altered blood–brain barrier permeability and potentiated oxidative stress in the cerebral tissue ,16. PatiComité d’Ethique Hospitalo-Facultaire Erasme-ULB) approved the study, but waived the need for informed consent because the study was purely observational. The study was performed in accordance with the ethical standards of the 1964 Declaration of Helsinki and its later amendments. All comatose patients  score <9) admitted after in-hospital CA (IHCA) or out-of-hospital CA (OHCA) (January 2008 to October 2012) were included in an institutional database, provided that they survived for at least 48 h after CA, so that at least three serum creatinine (sCr) measurements were obtained. Exclusion criteria were missing data on creatinine or urine output, AKI (for IHCA) before the occurrence of CA and chronic renal failure requiring hemodialysis.This retrospective study was performed in the Department of Intensive Care at Erasme Hospital, Brussels. The local Ethical Committee  and of a water-circulating blanket device . Midazolam (continuous infusion 0.03 to 0.1 mg/kg/h) and morphine (0.1 to 0.3 mg/kg/h) were administered during the hypothermic phase, while neuromuscular blocking agents  were given in the induction phase and, if needed, as a continuous infusion thereafter (1 to 3 mcg/kg/min). Re-warming was performed passively at a maximum rate of 0.5°C/h; sedation, analgesia and NMBAs were discontinued when the body temperature exceeded 37°C.2) at 35–45 mmHg and peripheral capillary oxygen saturation (SpO2) >94%. Invasive hemodynamic monitoring  was placed whenever needed and transesophageal echocardiography was performed within the first 8 to 12 h after ICU admission in all patients. Mean arterial pressure (MAP) was maintained at >65 to 70 mmHg using fluids, dobutamine and/or norepinephrine, whenever needed. Higher levels of MAP were targeted in patients with a history of hypertension and in patients with low (<60%) cerebral oximetry values . Intra-aortic balloon counterpulsation (IABP) or extracorporeal membrane oxygenation (ECMO) was initiated in cases of severe cardiogenic shock. A local insulin protocol was applied to keep blood glucose levels between 110 and 150 mg/dL in all patients.Patients were kept in a 30° semi-recumbent position; ventilation was set to maintain arterial partial pressure of carbon dioxide  were performed. Withdrawal of life-support was an interdisciplinary decision based on bilateral absence of the N20 response to somatosensory evoked potentials (SSEPs), persisting deep coma, or presence of status myoclonus or refractory status epilepticus.We collected data on demographics, pre-existing chronic diseases and CPR  in all patients. Serum creatinine levels on ICU admission were recorded, as was the CrCl on the first day of therapy, calculated from the 24-h urine collection, according to the following formula:sCr and urine output were collected daily during the entire ICU stay. Daily fluid balance and the administration of hydroxyethyl starch (HES) solutions were also noted. We also recorded use of any potentially nephrotoxic agents, including intravenous contrast media, angiotensin-converting enzyme inhibitors (ACEIs), angiotensin II receptor blockers (ARBs), non-steroidal anti-inflammatory drugs (NSAIDs), aminoglycosides, amphotericin B, colistin and immunosuppressant agents . Treatments with vasoactive drugs, mechanical ventilation, and continuous renal replacement therapy (CRRT) were recorded, as were length of ICU stay and ICU and hospital outcome. Neurological evaluation at 3 months after CA was assessed using the cerebral performance category score . The CPC evaluation was assessed during follow-up visits or by telephone interview with the general practitioner.AKI was defined as a daily urine output <0.5 mL/kg/h and/or an increase in sCr level by at least 0.3 mg/dL or >1.5 time increase from baseline values . In partt-test, or Mann–Whitney U-test, as appropriate. The significance of differences in sCr between groups was analyzed using two-way (time and group) analysis of variance for repeated measures (ANOVA), followed by Bonferroni post-hoc analysis. Multivariable logistic regression analysis with AKI development as the dependent variable was performed in all patients. Colinearity between variables was excluded prior to modelling; only variables associated with a higher risk of AKI (P <0.2) on a univariate basis were introduced in the multivariate model. Odds ratios (OR) with 95% confidence intervals (CI) were computed. The same analysis was then performed to identify independent predictors of favorable neurological outcome. A P-value <0.05 was considered as statistically significant.Statistical analyses were performed using the SPSS 18.0 for Windows NT software package . Descriptive statistics were computed for all study variables. The Kolmogorov-Smirnov test was used, and histograms and normal-quantile plots were examined to verify the normality of distribution of continuous variables. Discrete variables were expressed as counts (percentage) and continuous variables as means ± SD or median (25th to 75th percentiles). Demographics and clinical differences between groups  were assessed using the chi-square test, Fisher’s exact test, Student’s Of the 253 CA patients eligible over the study period, 54 were excluded  and favorable neurological outcome 19/51 (37%)) and 199 were included in the final cohort . Patients who developed AKI were significantly older than those who did not, were more likely to have chronic heart failure, pre-existing neurological or renal disease, and to be already receiving steroid or diuretic therapy. Patients who developed AKI also more frequently had a witnessed and IHCA than other patients, related to the larger number of witnessed CA among patients with IHCA than OHCA patients . Patients who developed AKI had a longer time to ROSC and received more epinephrine during CPR than other patients. These patients were also more likely to have shock, to be treated with vasopressors and dobutamine, and to have a longer duration of mechanical ventilation during the ICU stay than those who did not develop AKI. Finally, patients who developed AKI received more nephrotoxic agents, in particular aminoglycosides, had a greater fluid balance over the first 2 days after ICU admission and were more likely to receive diuretics during the ICU stay. Patients who developed AKI had greater hospital mortality  than those who did not and there was a trend towards fewer patients with a favorable neurological outcome at 3 months . Thirty-eight patients (19%) had ARC on the first day after ICU admission, and six of them eventually developed AKI, after a median of 3  developed AKI, after a median of 2  days; CKD was found in 21 patients (25% of the AKI patients) and CRRT was used in 24 patients (29% of the AKI patients) . Similar results were found for a 3-month favorable neurological outcome . The development of ARC, AKI or need for CRRT had no significant impact on neurological outcome  [et al., in a large cohort of 311 patients with OHCA, found that AKI had no impact on hospital survival [et al. reported that extra-cerebral organ dysfunction was common after CA but only cardiovascular dysfunction and altered gas exchange were associated with outcome [Our data provide no evidence that development of AKI contributes to poor neurological outcome in the setting of CA. By contrast, Hasper = 0.013) . All othsurvival . Robertset al. showed that creatinine levels on discharge were similar to baseline values in such patients, and that none of those being treated with CRRT during their ICU stay needed long-term hemodialysis thereafter [The study has some limitations. First, as a retrospective study, we may have missed some confounders that could influence the development of AKI. However, we collected a large amount of clinical data from the medical files, as well as data related to CA characteristics and the administration of potential nephrotoxic agents to minimize this bias. Of course, the generalizability of these findings may be limited because of different therapeutic strategies in the management of CA patients within other centers, which may influence both AKI occurrence and overall outcome. Second, during the four years of study enrolment, management of CA treatment may have changed somewhat , althougereafter .In this cohort, AKI occurred in almost half the patients who survived CA; however, it was not an independent prognostic factor for poor outcome. Age, epinephrine dose, cumulative fluid balance and presence of shock were independent predictors of the development of AKI in this population. ARC occurred in almost 20% of patients and was associated with reduced incidence of AKI. Renal failure had no clear correlation with poor neurological outcome.AKI is common in patients admitted after CA.Major risk-factors for AKI are older age, chronic renal disease, higher dose of epinephrine, in-hospital CA, presence of shock during the ICU stay, low creatinine clearance (CrCl) on admission and a high cumulative fluid balance at 48 h.AKI was not an independent predictor of poor 3-month neurological outcome.