AIP) gene are associated with pituitary adenoma, acromegaly and gigantism. Identical alleles in unrelated pedigrees could be inherited from a common ancestor or result from recurrent mutation events.Mutations in the aryl hydrocarbon receptor-interacting protein  haplotypes; coalescent-based approximate Bayesian estimation of the time to most recent common ancestor (tMRCA) of the derived allele; forward population simulations to estimate current number of allele carriers; proposal of mutation mechanism; protein structure predictions; co-immunoprecipitation and cycloheximide chase experiments.Observational, inferential and experimental study, including: AIP with additional haploblocks privately shared between subsets of the pedigrees, indicating the existence of an evolutionarily recent common ancestor, the ‘English founder’, with an estimated median tMRCA of 47 generations (corresponding to 1175 years) with a confidence interval . The mutation occurred in a small tandem repeat region predisposed to slipped strand mispairing. The resulting seven amino-acid duplication disrupts interaction with HSP90 and leads to a marked reduction in protein stability.Nine European-origin, unrelated c.805_825dup-positive pedigrees  included 16 affected  and nine unaffected carriers. All pedigrees shared a 2.79 Mbp haploblock around The c.805_825dup allele, originating from a common ancestor, associates with a severe clinical phenotype and a high frequency of gigantism. The mutation is likely to be the result of slipped strand mispairing and affects protein–protein interactions and AIP protein stability. AIP, MIM: 60555)  allele, with loss of heterozygosity identified in a number of cases  . AIP is : 60555)  and a C-: 60555) , 6. Diseescribed , 7. AIP of cases , 9.Mutations leading to a truncated protein are dispersed over the entire gene, while missense mutations are more common in the C-terminal end of the molecule . SeveralAIP exon 6 in-frame duplication (rs267606578/EF643650) in three affected members of a family from the United Kingdom  was performed as previously published  around AIP  in the iAIP . HaplotyAIP , incorpoAIP . We usedAIP , using Lipofectamine 2000 (Life Technologies) following the manufacturer’s instructions. Cells were harvested 48 h later by trypsinization and resuspended in 1.5 mL of lysis buffer  supplemented with Complete Protease Inhibitor Cocktail (Roche)). After being cleared by centrifugation, lysates were cleaned up by incubation with 50 µL of Protein G Sepharose 4 Fast Flow  and divided in thirds for incubation with 5 µg of anti-Myc  or anti-HA (Sigma-Aldrich H3663) mouse monoclonal antibodies, or mouse IgG (Sigma-Aldrich I5381), as appropriate. The co-immunoprecipitation was performed as previously described , but two clinically non-functioning pituitary adenomas (NFPA) were also observed . Two FIPThe overall average age of onset of symptoms was 19.1 years .AIP shared among c.805_825dup allele-carrying chromosomes in all nine pedigrees, as well as additional haploblocks shared among subsets of chromosomes, strongly supporting the existence of a recent common ancestor; the ‘English founder’  haploblock around founder’ . We obseWe obtained tMRCA estimates of 47 (9–113) generations ), based on HapMap genetic distances  and 48  and B. AThe estimated current number of carriers per generation – obtained through forward simulations – was 32 (9–299) , based on HapMap genetic distances, and 32 (9–250), based on Rutgers map distances  and D. TWe studied the structure of the DNA sequence around the 21 base-pair tandem duplication allele. We have identified a region in the normal sequence with two short repeated sequences separated by an intervening sequence  and B, win silico prediction. As compared with WT AIP, the steady state levels of the p.F269_H275dup protein were markedly reduced, and this led us to hypothesize an effect of the mutation on AIP protein stability as well, consistent with what has already been shown for several other AIP mutations (P < 0.001) and even that of the truncating mutant p.R304*  (The mutation results in the duplication of seven amino-acids in the third TPR domain of AIP (TPR3). These can adopt a helical conformation, thus adding a 1.9 extra helical turn to the TPR3 alpha helix and increasing its length by 10.5 Å. The TPR3 domain is important for the interaction of AIP with other proteins, in particular, HSP90 and TOMM7 peptides , D and Eutations . Cyclohe < 0.05) , suggestAIP mutations are reported in about 20% of FIPA pedigrees    or the rout 20%) . This is unknown .de novo mutations. These patients were tested for AIP mutations because of their clinical characteristics , confirming previous reports of relatively high prevalence of AIP mutations in apparently sporadic GH-secreting macroadenomas of the youth  countries and show that all the affected subjects, the majority suffering from young-onset somatotropinomas, share a common ancestral haplotype. The mutation results in the duplication of seven amino-acids in third TPR domain of AIP, leading to the disruption of protein–protein interactions and markedly reduced protein stability.In summary, we report the same R S and M K serve on Pfizer Ltd’s advisory board. The other authors declare no conflict of interest.S R was supported by the EC-REA Marie Sklodowska-Curie IEF postdoctoral fellowship (grant 303006/2011). A D was supported by an MRC postdoctoral fellowship (grant MR/K021613/1). D I is supported by a Diabetes UK George Alberti Clinical Research Training Fellowship. M K’s work on familial pituitary adenomas is supported by the Medical Research Council, Barts and the London Charity and Pfizer Ltd.


Rapid eye movement sleep behavior disorder (RBD) and Parkinson’s disease (PD) are two distinct clinical diseases but they share some common pathological and anatomical characteristics. This study aims to confirm the clinical features of RBD in Chinese PD patients.One hundred fifty PD patients were enrolled from the Parkinson`s disease and Movement Disorders Center in  Department of Neurology, Shanghai General Hospital from January 2013 to August 2014. This study examined PD patients with or without RBD as determined by the REM Sleep Behavior Disorder Screening Questionnaire (RBDSQ), assessed motor subtype by Unified PD Rating Scale (UPDRS) III at “on” state, and compared the sub-scale scores representing tremor, rigidity, appendicular and axial. Investigators also assessed the Hamilton Anxiety Scale (HAMA), Hamilton Depression Scale (HAMD), Mini-Mental State Examination (MMSE), Clinical Dementia Rating (CDR), and Parkinson’s disease Sleep Scale (PDSS).+ and pRBD− groups. pRBD+ group had lower MMSE scores, higher PDSS scores, and pRBD+ PD patients had more prominent proportion in anxiety, depression, constipation, hallucination and a greater prevalence of orthostatic hypotension.One hundred fourty one PD patients entered the final study. 30 (21.28%) PD patients had probable RBD (pRBD) diagnosed with a RBDSQ score of 6 or above. There were no significant differences for age, including age of PD onset and PD duration, gender, smoking status, alcohol or coffee use, presence of anosmia or freezing, UPDRS III, and H-Y stages between the pRBD+ PD patients exhibited greater changes in non-motor symptoms. However, there was no increase in motor deficits.pRBD Rapid eye movement sleep (REM) is characterized by decreased or absent muscle tone (atonia), desynchronization of the electroencephalogram, with the presence of saw tooth waves, and autonomic instability. Rapid eye movement sleep behavior disorder (RBD) is a form of parasomnia during which patients develop limb or body movements, which correlate with dream enactment behavior. The abnormal physiology of RBD is loss of muscle atonia  during otherwise intact REM sleep .The standard RBD diagnostic criteria are based on the 2nd edition of the International Classification of Sleep Disorders (ICSD) . ChahineRBD has a close relationship with neurodegenerative diseases, especially those with α-synucleinopathy pathology such as PD, dementia with Lewy bodies (DLB) and multiple system atrophy (MSA) , 6. ReceOne hundred fifty PD patients were enrolled from the Parkinson’s disease and Movement Disorder Center in the Department of Neurology, Shanghai General Hospital, Shanghai Jiao Tong University, Shanghai, China from January 2013 to August 2014. The diagnosis of PD was made by two movement disorder specialists according to the UK Parkinson’s Disease Society (UKPDS) Brain Bank Criteria. Patients with severe dementia (CDR ≥ 2), or other central nervous system disorders were excluded from this study. The study was approved by the Institution’s Ethics Committee and all recruited patients consented to participate in the study.Of 150 patients, 5 patients were excluded because of probable DLB, 3 patients had probable MSA, and 1 patient had vascular Parkinsonism. A total of 141 patients were enrolled in the study. Patient evaluation was performed by movement disorder specialists. Parkinsonism staging was evaluated according to the Hoehn & Yahr staging scale. Part III of the Unified PD Rating Scale (UPDRS III) was performed during the “on” state. Motor subtype was analyzed by predominance of tremor or rigidity (UPDRS sub-scores) and predominance of limb or axial features (UPDRS sub-scores) , Hamilto2 tests were used where appropriate. A P-P plot was used to test normal distribution. A p value <0.05 was considered to be significant.The data were analyzed using the Statistical Package for Social Sciences (SPSS) 19. The data is presented as mean, counts and percentages, and the adjusted difference in means. Analysis of descriptive variables was performed using two-tailed t tests. Mann-Whitney U tests and X+ group, and 56 male (50.45%) in pRBD− group (p = 0.655). Thirty patients (21.28%) were diagnosed with probable RBD (pRBD) based on a RBD screening questionnaire score ≥ 6. If the cut-off score was set at 7 or 5, the incidence of RBD was 17.02% and 26.24% respectively, with little difference in the clinical features (Table + group was 68.33 ± 8.76 versus 69.32 ± 9.75 years in the pRBD− group (p = 0.618). Mean PD duration years is 4.13 ± 4.216 in pRBD+ and 4.65 ± 3.570 in pRBD− group (p = 0.5). Smoking, alcohol and coffee consumption were infrequent in both groups (NS). There was no difference between the numbers of patients who took levodopa or dopamine agonists, and the levodopa equivalent dosage was similar in both groups. However, the pRBD+ group had greater antidepressant and anti-hypnotic use : 18 male (60%) in pRBDes Table . The mea+ PD patients had significantly higher rates of anxiety  and depression   (Table p = 0.538)  (Table + group and 6.3% (7) in pRBD− group (p = 0.032) . Mean H-Y stage was also similar between the two groups . When limb scores in UPDRS III was compared with axial scores, the ratio showed no difference between pRBD+ and pRBD− groups . When rigidity scores in UPDRS III were compared with resting tremor score, the ratio was also not significantly different . Overall there was no difference between pRBD+ group and pRBD− group for motor severity and motor subtype (Table p = 0.75).The UPDRS III score was similar in the pRBDedunculopontine nucleus (PPN) and locus cerulean (LC)/ dorsal subcoeruleus (subCD) are compromised in both PD and RBD [Probable RBD is common in early PD and predicts future cognitive decline, particularly in attention and memory domains . The ped and RBD –15. Auto and RBD , and con and RBD , 17. Dys and RBD . PD pati and RBD . The bas and RBD . The app and RBD .Our study had compared motor and non-motor symptoms in PD patients with and without RBD. The results were similar if we used a RBDSQ cut off point at 6 or 7 Table . OverallIn summary, this study systematically investigated the clinical features of PD patients with RBD. There are several potential weaknesses. We used the RBDSQ to detect RBD in PD patients which is easier and more readily available than PSG. The sample size is relatively small and it might have given false negatives and false positives for diagnosing RBD without PSG. We diagnosed the patient with anosmia and constipation only based on self-report and not using objective examination. Neurological image and electrophysiology will be valuable for further study.+ PD patients, while the non-motor symptoms are prominent, such as mood, sleep, constipation, cognition and orthostatic hypotension. However, further studies and laboratory tests are needed to improve the understanding of RBD in PD.The present study demonstrated that there were no significant differences in motor deficits in pRBD


The aim of the present study was to construct a cost-effective noninvasive diagnostic index for prediction of hepatic steatosis in patients with hepatitis B virus(HBV) infection. From January 2011 to January 2015, a total of 364 consecutive subjects who underwent liver biopsies were enrolled. The Receiver-operating characteristic(ROC) curves and Obuchowski measure were constructed to evaluate the diagnostic accuracy of the new index. The AUROCs of steatosis index of patients with HBV infection (SIHBV) in predicting of steatosis were 0.929  in the model group and 0.855  in the validation group respectively. Comparisons of AUROCs demonstrated that SIHBV was significantly superior to Korean Score, fatty liver index (FLI), hepatic steatosis index (HSI), lipid accumulation product(LAP), and fatty liver disease (FLD) index for prediction of hepatic steatosis in model group and validation group. Especially for patients with hepatic steatosis percentage of 5.0–9.9% and 10.0–19.9%, SIHBV had a sensitivity of 63.6% and 79.2%, whereas it were 29.1% and 45.8% for Ultrasonography . In conclusion, as a cost-effective, simple, noninvasive, and readily available method, SIHBV may act as a massive screening tool before further examinations such as MRI, CT, transient elastography, or liver biopsy, especially for developing countries. Which is the major cause of elevated ALT level? CHB or NAFLD? For newly diagnosed CHB patients with elevated ALT level caused by NAFLD, it is not necessary to receive anti-viral therapy for the moment. For patients with abnormal liver function and low HBV DNA load in the course of anti-viral therapy, there is no need to adjust the anti-viral treatment if elevated ALT level was caused by NAFLD. Determination that whether fatty liver exists in CHB patients is very important for the final clinical decision. Therefore, CHB patients with elevated ALT level need a high cost-effective, noninvasive, sensitive, and convenient method for detection of NAFLD.Liver biopsy is still the gold standard for assessing hepatic steatosis but is limited by invasiveness and sampling error9et al. has reported that US could only detect 55% and 72% of patients with hepatic steatosis percentage of 10–19% and 20–29%, respectively;meanwhile, US could not detect hepatic steatosis while hepatic steatosis presented in less than 10% of hepatocytesMost primary hospitals select Ultrasonography(US) as basic screening tool for detection of NAFLD in consideration of clinical maneuverability and cost-effectiveness. From the point of view of cost-effectiveness, US is a high cost-effective noninvasive method for detection of fatty liver. However, US is limited by low sensitivity for mild steatosis and inability to differentiate mild fibrosis from steatosiset al. has conducted a noninvasive diagnostic index named steatoTest, showing an AUROC of 0.790 (SE 0.03) for hepatic steatosis ≥5% by biopsy in the training groupet al. has established Fatty Liver Index (FLI) in a population of 496 subjects with hepatic steatosis diagnosed by US in 2006, with an AUROC of 0.840 (95% CI 0.810–0.870)et al. has constructed another diagnostic index named lipid accumulation product (LAP), with an AUROC of 0.80 (95% CI 0.760–0.840)15. However, these studies were performed in general population, which may not be suitable for patients with HBV infection.Several noninvasive diagnostic indexes have been established to predict hepatic steatosis in general population. In 2005, Poynard Therefore, we performed this retrospective study to construct a high cost-effective and noninvasive diagnostic index for prediction of hepatic steatosis in patients with HBV infection. To assess the diagnostic accuracy of this new index, we compared it with US and five other noninvasive indexes for detection of liver steatosis in CHB patients.3 copies/ml. The exclusion criteria were as follows: liver cancer(n = 5);co-infection with hepatitis C virus, hepatitis D virus or human immunodeficiency virus(n = 8); autoimmune liver diseases suah as autoimmune hepatitis, primary biliary cirrhosis, and primary sclerosing cholangitis(n = 3); alcohol ingestion >20 g/day (n = 15); hereditary and metabolic liver diseases such as Wilson’s disease, hemochromatosis, and α-1-antitrypsin deficiency(n = 2). The patients with missing data in terms of BMI, TG, LDL, UA, and HGB were ruled out(n = 34).This study included 431consecutive patients who had been diagnosed with HBV infection and had undergone liver biopsies in Department of Infectious Diseases of The Shunde First People’s Hospital between January 2011 and January 2015. The Patients were enrolled based on the following criteria: chronic hepatitis B(CHB) defined as hepatitis B surface antigen (HBsAg) positivity for more than 6 months; HBV-DNA >10Therefore, a total of 364 patients  were recruited into the present study finally. There were no significant differences in terms of demographic and clinical parameters between patients included and excluded (data not shown). The written informed consent were obtained from patients before inclusion. This study was approved by the ethics committee of The First People’s Hospital of Shunde. All clinical investigations were conducted according to the principles of Declaration of Helsinki.All patients systematically underwent complete biochemical workups, ultrasonography, and liver biopsy within 2 days. Blood samples of the subjects were obtained before liver biopsy. Biochemical tests were performed by commercial assays in our hospital laboratory. The serum HBV-DNA load was detected using a Real-Time polymerase chain reaction (PCR) System .2.Waist circumference was measured at the midway point between the costal margin and the iliac crest. Body mass index (BMI) was calculated as weight (kg)/height (m)The Korean Score, fatty liver index (FLI), hepatic steatosis index (HSI), lipid accumulation product(LAP), and fatty liver disease (FLD) index were calculated according to the formulas reported in the original articles131415162 ≤ BMI < 25 kg/m2, 1 point; 25 kg/m2 ≤ BMI, 2 points.Korean Score was calculate according to the following parameters:ALT/AST ratio > 1.5, 1 point; γ-GTP (IU/L) > 50, 1 point; Triglyceride (mg/dL) > 150, 1 point; 23 kg/mWe performed this study according to the STARD recommendation for the optimal quality in reporting diagnostic accuracy.We used Menghini’s method of one second rapid vacuum aspiration to get liver tissue. A minimum length of 2.0 cm with at least 11 portal tracts was required for qualified liver specimens. The specimens were fixed, paraffin-embedded, and stained with haematoxylin and eosin (HE). Histological grading of necro-inflammation (G0-G4) and staging of the liver fibrosis (S0-S4) were carried out according to Scheuer’s classificationP < 0.05 was considered statistically significant.Continuous data were expressed as mean ± SD or median (quartile range) depending on the normality of the data. Categorical variables were expressed as proportions. Continuous variables were compared by one-way ANOVA analysis of variance or Kruskal-Wallis H test, depending on the normality of the data; Nominal or ordinal variables were compared by Kruskal-Wallis H test. All variables that significantly associated with hepatic steatosis in univariate logistic regression analyses were included in forward stepwise multivariate logistic regression analysis to conduct a predictive index for hepatic steatosis. The area under the receiver operator characteristic curve(AUROC) were calculated to evaluate the diagnostic accuracy of new index. The AUROC values of these indexes were compared by DeLong’s test21222122A total of 364 patients were included in the study, 118 (32.4%) of whom were diagnosed with hepatic steatosis. All patients included in the present study were randomly divided into two groups: model group (n = 182) and validation group (n = 182). The baseline characteristics of subjects in model group and validation group were summarized in All variables that significantly associated with hepatic steatosis in univariate logistic regression analyses were included in forward stepwise multivariate logistic regression analysis to conduct a predictive index for hepatic steatosis. The results of logistic analyses were summarized in According to the results of multivariate logistic regression analysis, we constructed a new diagnostic indexl named steatosis index of patients with HBV infection (SIHBV):P < 0.05), which was higher than that of Korean, LAP, HSI, FLI, and FLD. Comparisons of AUROCs using method suggested by DelongThe AUROC of SIHBV was calculated to assess the diagnostic accuracy for prediction of hepatic steatosis . The AURP < 0.01).The AUROCs of SIHBV, FLI, LAP, HSI, FLD, and Korean Score  in prediThe spectrum effect reflects the inherent variation of performance of diagnostic models in different population. Considering of spectrum effect caused by different distributions of hepatic steatosis stage, the results of comparisons of AUROCs by Delong test need further confirmation.The Obuchowski method can fully take into account of the spectrum effect and the ordinal scale system2122The overall diagnostic accuracy of SIHBV (Obuchowski measure = 0.924)for hepatic steatosis was significantly higher than that of FLD (Obuchowski measure = 0.879), Korean (Obuchowski measure = 0.867), FLI (Obuchowski measure = 0.860), LAP (Obuchowski measure = 0.821), and HSI (Obuchowski measure = 0.811).P < 0.05).We further explored the clinical utility of SIHBV according to different thresholds of hepatic steatosis. For a threshold of 33% of hepatic steatosis, 27 and 337 patients were defined as with or without hepatic steatosis. The AUROCs of SIHBV, FLI, LAP, HSI, FLD, and Korean Score  in prediP < 0.01).For a threshold of 20%, 39 and 325 patients were defined as with or without hepatic steatosis. The AUROCs of SIHBV, FLI, LAP, HSI, FLD, and Korean Score  in prediP = 0.001 and P = 0.018). For hepatic steatosis percentage of 20.0–29.9%, sensitivity of SIHBV was higher than that of US, although the difference was not statistically significant(P > 0.05). For hepatic steatosis percentage of 33.0%–100.0%, SIHBV had the same sensitivity as US (P > 0.05). The comparison results of Chi square test also indicated that the total correct rate of SIHBV was significantly superior to US(P = 0.027) In all 118 patients with steatosis diagnosed by liver biopsy, US only detected 29.1, 45.8, 58.3, and 81.5% of patients with hepatic steatosis percentage of 5.0–9.9%, 10.0–19.9%, 20.0–32.9, and 33.0–100.0%, respectively . The posTo explore the clinical utility of SIHBV for prediction of hepatic steatosis, the optimal Cut-off values were determined according to positive likelihood ratio (PLR) ≈ 10.0 for confirming diagnosis of hepatic steatosis and negative likelihood ratio (NLR) ≈0.1 for excluding diagnosis of hepatic steatosisAmong the 364 patients included in the study, 175 (48.1%) patients had a value of SIHBV less than or equal to 0.18 and 105 (28.8%) patients had a value of SIHBV higher than or equal to 0.48 . The cutP < 0.01). Interestingly, the further comparison results of Chi square test indicated that SIHBV was significantly superior to US for hepatic steatosis percentage of 5.0–9.9% and 10.0–19.9%, demonstrating that SIHBV was valuable for detection of mild hepatic steatosis and could help clinicians to find mild hepatic steatosis in patients with HBV infection. With a lower value of 0.18 and a higher value of 0.48, the patients were divided into three groups with different prevalence risk of hepatic steatosis, facilitating clinicians assess the risk of NAFLD and distinct that whether elevated ALT level was caused by NAFLD. Comparisons of Obuchowski measure confirmed that SIHBV was superior to other five noninvasive indexes for hepatic steatosis.We constructed a new diagnostic index for prediction of hepatic steatosis in patients with HBV infection, consisting of BMI, hemoglobin, age, TG and SUA. The AUROC of SIHBV in predicting hepatic steatosis were 0.929 (0.889–0.970) in the model group and 0.855 (0.794–0.917) in the validation group. Comparisons of AUROCs indicated that SIHBV was significantly superior to US, FLI, LAP, HSI, FLD, and Korean Score for prediction of hepatic steatosis in model group and validation group  of 364 patients can be free from liver biopsy with a overall correct rate of 90.0% for diagnosis of hepatic steatosis. Second, patients with CHB can save medical expenditures in the course of treatment by replacing US with SIHBV, which is actually a free method for BMI, hemoglobin, age, TG, and SUA can be freely obtained from routine examinations. Most importantly, SIHBV can help doctors determine the real cause of elevated ALT in the course of treatment or selection of suitable patients for antiviral therapy, especially hepatic steatosis percentage is less than 20%.et al. has found that patients with HBV and/or HCV infections had higher liver enzyme levels in comparison with virus-negative patients (P < 0.0001), whereas the prevalence of the metabolic syndrome was similar in this 2 groupsIt has been reported that the prevalence of NAFLD and non-alcoholic steato-hepatitis (NASH) were 76 and 56% in patients with Type 2 Diabetes Mellitus(T2DM) and normal plasma aminotransferase levels, indicating that NAFLD and NASH were common in patients with T2DMThe present study has several strengths: First, SIHBV consists of readily-available laboratory parameters which can be freely obtained from routine examinations, so SIHBV is easy to be implement in the primary hospital. Second, the prevalence and stages of hepatic steatosis were different in various studies, which should be taken into account for assessment of diagnostic accuracy of noninvasive indexes. To avoid the spectrum effect and the risk of multiple testing, we performed comparisons of diagnostic accuracy of six noninvasive indexes by Obuchowski method. The results of Obuchowski measure were in agreement with that of Delong test. Third, hepatic steatosis in the present study was diagnosed by liver biopsy, which is considered the gold standard for assessing hepatic steatosis. Fourth, the present study was performed in patients with CHB, so SIHBV is more suitable for detection of hepatic steatosis in patients with HBV infection.There were two limitations in the present study. The first limitation was that the sample size of the present study was relatively small, which might affect reliability of the results. The second limitation of this study was that all subjects were patients in our hospital, which might reduce the representative of the study population. To further validate the diagnostic accuracy and clinical utility of SIHBV for hepatic steatosis in patients with HBV infection, further studies should base on a large scale and multi-center population.In summary, for prediction of hepatic steatosis in patients with HBV infection, SIHBV has a better diagnostic accuracy, which is significantly superior to US, FLI, LAP, HSI, FLD, and Korean Score. SIHBV need further external validation in large population before it was used for prediction of hepatic steatosis in patients with HBV infection.How to cite this article: Zhang, Z. et al. Diagnostic accuracy and clinical utility of a new noninvasive index for hepatic steatosis in patients with hepatitis B virus infection. Sci. Rep.6, 32875; doi: 10.1038/srep32875 (2016).


Acute gastroenteritis (AGE) causing dehydration with or without dysnatremias is a common childhood health challenge. While it is accepted that oral rehydration therapy is preferred, clinical factors or parent and healthcare provider preferences may lead to intravenous rehydration (IVR). Isotonic solutions are increasingly recommended in most scenarios requiring IVR. Nevertheless, children with AGE, having ongoing losses of water and electrolytes, represent a unique population.To evaluate the association between acquired dysnatremias and IVR in children with AGE.A systematic search of MEDLINE database was conducted through September 14, 2016. Observational studies and clinical trials conducted in high-income countries were included. The Grades of Recommendation, Assessment, Development, and Evaluation approach was used to evaluate the overall quality of evidence for each outcome.603 papers were identified of which 6 were included . Pooling of patient data was not possible due to significantly different interventions or exposures. Single studies results demonstrated that within 24 h, administration of isotonic saline was not associated with a significant decline in serum sodium while hypotonic solutions  were associated, in one study, with mean serum sodium declines from 1.3 mEq/L  in 133 young infants (aged 1–28 months), to 5.7 (SD 3.1) mEq/L in a subgroup of 18 older children . Both isotonic and hypotonic saline were shown to be associated with improvement of baseline hyponatremia in different studies. Baseline hypernatremia was corrected within 4–24 h in 81/83 (99.6%) children using hypotonic saline IVR.There is a paucity of publications assessing the risk for acquired dysnatremias associated with IVR in children with AGE. Current high-quality evidence suggests that, short-term use of isotonic solutions is safe and effective in most children with AGE; hypotonic solutions may also be appropriate in some subpopulations, however, the quality of available evidence is low to very low. Further research investigating outcomes associated with IVR use beyond 24 h focusing on specific age groups is required. Dehydration, complicating acute gastroenteritis (AGE), continues to be a leading cause of morbidity and mortality around the globe . NumerouBoth oral and IVR can cause or exacerbate electrolyte disturbances related to sodium homeostasis. The risk for hypernatremia associated with oral rehydration solutions (ORSs) containing excessive amounts of salt and carbohydrates was dramatically reduced by the introduction of reduced total osmolality ORS containing equimolar concentrations of sodium and glucose –7. WhileTo form an evidence-based opinion on this topic, we conducted a systematic review of clinical trials and observational studies that investigated the risk of dysnatremias in relation to IVR solution composition and rate of administration. We focused on children in high-income countries given the significant differences in patients, pathogens, and management practices that exist, driven by economic factors.This review was planned, conducted, and reported in adherence with Preferred Reporting Items for Systematic Reviews and Meta-Analyses standards of quality for reporting systematic review .We sought to identify studies to address the following PICO question : in chilhttp://data.worldbank.org/income-level/high-income); and (6) with any outcome measure of serum sodium after IVR treatment. All eligible studies, regardless of report focus, were included. Review articles, and those containing duplicate data were excluded. No language, age, or study design restrictions were employed.Eligible studies contained the following elements: (1) children with AGE; (2) all patients received IVF therapy; (3) compared different types of IVF, or different infusion rates or different IVF volumes, or investigated risk factors in patient characteristics ; (4) clinical or biochemical documentation of hydration status, fluids administered, and serum sodium level before and after fluid therapy; (5) conducted in a high-income country (We searched MEDLINE (1946 to September 2016) on September 14, 2016. The search used key terms including Gastroenteritis, Diarrh*ea, Vomiting, Volume expansion, Fluid management, IVF, Hydration, Dehydration, Intravascular volume expansion, Fluid, Saline, Rehydration, Hyponatr*emia, and Hypernatr*emia. The MEDLINE search strategy is appended . Experts in the field were also invited to suggest relevant studies. All published studies with sufficient information, irrespective of language of publication, publication year, publication type, and publication status, were eligible for inclusion. An updated search performed on May 5, 2017 identified no additional eligible studies.Potentially relevant publications were identified through independent screening of search result titles and abstracts by two trained study reviewers that included a pediatric emergency medicine physician (Jianling Xie) and a pediatric nephrologist (Silviu Grisaru). The reviewers had an overall agreement of 97.6%. Duplicate articles were manually reconciled.The full text of potentially relevant citations was obtained and then reviewed by two independent reviewers (Silviu Grisaru and Jianling Xie) using standard, predefined eligibility criteria. A consensus approach with input from the senior investigator (Stephen B. Freedman) was employed to resolve disagreements. Reasons for inclusion and exclusion were documented.Data were extracted by two independent reviewers (Jianling Xie and Silviu Grisaru) and any disagreement was resolved by discussion to reach consensus. We collected information on the study design, research topic, nature of participant characteristics, IVF type, infusion rate and volume, follow-up interval, and outcomes type and outcome data. We only collected data published, unpublished data were not sought. We did not perform meta-analysis as all included studies investigated different interventions or exposures that precluded data synthesis, thus only a narrative summary of results was generated.The Newcastle–Ottawa scale (NOS)  was usedSix hundred and two abstracts were identified by the initial search and one additional study was suggested by an expert in the field; at the end of the screening process and application of exclusion/inclusion criteria, six eligible studies were included in this review Figure . The eliThe assessment of risk of bias is summarized in Table n = 102) comparing the effect of two IVR solution compositions  on sodium balance . Sixty-three percent (30/48) of children who were hyponatremic at baseline and were treated with large-volume bolus were isonatremic after 4 h of IVR, compared with 44% (15/34) in the standard volume group (P = 0.10). Among all participants, irrespective of the bolus size, hyponatremia at baseline was associated with a mean serum sodium increase of 2.6 (SD 2.1) mEq/L compared with only 0.4 (SD 2.1) mEq/L among children who were not hyponatremic (P < 0.001). Baseline isonatremic was associated with a plasma sodium decrease of ≥2.0 mEq/L in 17% (23/135) of children, compared with only 2% (2/82) among those who were hyponatremic (P = 0.001). Fifty-nine children (53%) in the large-volume bolus group and 39 (37%) in the standard volume group experienced a serum sodium increase ≥2.0 mEq/L, however, no participants developed hypernatremia after 4 h. The conclusion based on these results was that both large and standard volume boluses of 0.9% saline are not associated with development or worsening of hyponatremia after short-term IVR. Moreover, large-volume bolus fluid therapy was associated with a more rapid correction of hyponatremia  in 2011 . Patientn = 77, including 38 in the 0.9% saline group). Inclusion criteria consisted of AGE in children aged 0.5–11 years with a Gorelick dehydration score ≥4  for IVR of children with dehydration caused by AGE . The priscore ≥4 . Prior tSánchez-Bayle et al. published in 2014 results from a single center prospective observational study that included 205 children aged 1–28 months  with AGE who received IVR . Most stKahn et al. reported on 40 infants less than 6 months of age who were admitted to the intensive care unit for management of AGE associated hypernatremic dehydration (serum sodium ≥155 mEq/L) . ParticiP < 0.0005), more likely to be female  and received a larger volume initial saline bolus . Among the 19 children that had mild hyponatremia at baseline, 5 remained hyponatremic at follow-up. The overall change in mean serum sodium among all 121 participants was −1.7 (SD 4.3) mEq/L. The main limitation of this study stems from its retrospective design forcing grouping together children that received different IVR hypotonic solutions some of whom received an initial saline bolus of variable volume  of the studied populations. Only three short-term clinical trials that focused on different research questions or primary outcomes were identified. Three short-term observational studies analyzed outcome time points that varied from 4 to 24 h. Four included studies were subject to a moderate to high risk of bias Tables  and 6. WSG, JX, SS, and SF conceptualized and designed the study, and designed the data collection instruments. SG and JX performed the data extraction. SS and SF supervised and coordinated data collection and analyses. SG drafted the initial manuscript. SG, JX, SS, and SF reviewed and critically revised the drafts of the manuscript and approved the final manuscript as submitted.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.


Despite regulatory issues surrounding the use of animal-derived cell culture supplements, most clinical cardiac cell therapy trials using mesenchymal stromal cells (MSCs) still rely on fetal bovine serum (FBS) for cell expansion before transplantation. We sought to investigate the effect of human serum from heart failure patients (HFS) on cord blood MSCs (CB-MSCs) during short-term culture under regular conditions and during simulated acute and chronic stress. Cell survival, proliferation, metabolic activity, and apoptosis were quantified, and gene expression profiles of selected apoptosis and cell cycle regulators were determined. Compared to FBS, HFS and serum from healthy donors (CS) showed similar effects by substantially increasing cell survival during chronic and acute stress and by increasing cell yields 5 days after acute stress. Shortly after the termination of acute stress, both HFS and CS resulted in a marked decrease in apoptotic cells. Transcriptome analysis suggested a decrease in TNF-mediated induction of caspases and decreased activation of mitochondrial apoptosis. Our data confirm that human serum from both healthy donors and heart failure patients results in increased cell yields and increased resistance to cellular stress signals. Therefore, we consider autologous serum a valid alternative to FBS in cell-based therapies addressing severe heart disease. Based on the findings of Kawada et al.  and Chen, includi culture , 20. Oth culture . In fact culture .In the current study, we sought to investigate whether the short-term incubation of a “virginal” model cell product with human serum of CHF has an immediate effect on cell proliferation and metabolism. We chose cord blood-derived MSCs (CB-MSCs) as a model cell type, since CB-MSCs are more proliferative, not senescent, and have not been subjected to exogenous noxae . CB-MSCsA literature search on MEDLINE and clinicaltrials.gov was conducted to identify clinical trials testing MSCs for cardiovascular regeneration in the past five years. The search was limited to study protocols or results published between May 1, 2012, and May 31, 2017. A combination of terms for mesenchymal stem cells  and disease-related keywords  was used to identify relevant trials. Vocabulary and syntax were adjusted across databases. Each study protocol was screened for cell type used and, if applicable, for cell culture medium formulation used for ex vivo expansion of cells .n = 12) and volunteers (n = 12) (p < 0.05 versus CHF patients) without a history of cardiovascular disease.In accordance with the Declaration of Helsinki, this study was approved by the Ethics Committee of Charité-Universitätsmedizin Berlin, Berlin, with informed consents signed by all patients ((n = 12) . A thorog for 15 minutes at 4°C to remove the clot and all remaining cellular particles. Serum supernatants were then sterile filtered, aliquoted, flash frozen in liquid nitrogen, and then stored at −80°C for later use. Serum from each donor (n = 12) was tested individually on CB-MSCs to account for possible patient-specific confounding factors. CHF is associated with interstitial and intravascular volume retention, and patients usually show relative hyperproteinemia which could influence the quantitative bioactivity of heart failure serum (HFS) as compared to control serum (CS). Accordingly, the total protein concentration in CS was significantly higher than that in HFS . We noted that the lowest serum protein concentration in one of our heart failure patients (4.6 g/dL) was within the range of protein concentration found in our FBS sample (4.5 g/dL). Since we hypothesized that differences in serum impact on cells arise from its bioactive contents, that is, cytokines, exosomes, short nucleotides, or other trophic factors, we decided to eliminate the variance in total protein concentration. In order to minimize this systematic error, serum concentrations were normalized concentration within our samples to 4.5 g/dL, by dilution with Dulbecco's phosphate-buffered saline (DPBS) containing Ca2+ and Mg2+ . Total protein concentration of each serum sample was quantified using the Pierce BCA Protein Assay Kit .Venous blood was collected in S-Monovettes®  by using the BD Vacutainer Safety-Lok blood collection set . Whole blood samples were left undisturbed at room temperature for 30 minutes and then centrifuged at 35002 in T175 flasks and cultured under 21% O2 and 5% CO2 at 37°C. Partial media changes were performed every 3 days. All cultured cells were screened for the presence of mycoplasma . All experiments were performed on cells between passages 4 and 6. CB-MSC phenotype and their ability to differentiate into nonhematopoietic cell types have been repeatedly confirmed in previous experiments by our group [Human CB-MSCs were provided by courtesy of Dr. K. Bieback, who isolated them from umbilical cord blood with the mothers' consent and approval of the local ethics committee and expanded the cells based on a previously published protocol . Prior t3 cells/cm2 into 96-well plates . After 24 hours of incubation under standard cell culture conditions to allow for attachment (day 0), cells were deprived of glucose and provided with either 10% human serum or FBS in their culture media and cultivated for 5 days at 1% O2. In parallel, as a control condition, cells were kept under regular culture conditions with the exception of supplementing the media with FBS, CS, or HFS. Culture media was changed every other day.Glucose deprivation in a setting of low oxygen tension triggers oxidative stress in MSCs , 30. To in vitro model of “simulated ischemia reperfusion injury” where the cells are deprived of glucose, serum, and oxygen (0.2% O2) for 4 hours and then transferred back into regular culture conditions  as well as serum and glucose deprivation for 4 hours, followed by 4 hours of simulated reperfusion in normal culture settings. The proliferative capacity of CB-MSCs (or recovery) during the first 5 days after the acute stress trigger was tested in culture with the different sera .To test how CB-MSCs cultured with different sera behave in response to an acute stress trigger (AcS), we chose an nditions . This seOne part of 3--5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) and phenazine methosulfate (PMS) solution was added to 5 parts of culture media, and the cells were incubated for 4 hours at 37°C. Absorbance (OD) was measured at 490 nm and 650 nm as a reference wavelength. Subsequently, nuclei were stained with Hoechst 33342 (Life Technologies) in the dark for 20 minutes at room temperature and washed with DPBS. Cell numbers were counted in the Operetta High-Content Imaging System  at 380 nm excitation and 445 nm emission. Cell survival rate after AcS is depicted as the percentage of initially plated cells. Cell survival rate is depicted as the percentage of cells counted after AcS, as compared to the percentage of cells plated. BrdU incorporation was used  to quantify cell proliferation. Cells cultured in 96-well plates were incubated with BrdU labeling solution at 37°C for 4 hours, and the following steps were performed without interruptions according to the manufacturer's instructions. Absorbance was measured in the ELISA reader  at 370 nm with the reference wavelength set to 492 nm.μM H2O2 were used as Annexin V-positive control, while cells incubated on ice after 10 minutes of 65°C warm water bath were used as ethidium homodimer III-positive control. FITC- and/or EthD-III-positive cells were then quantified using the MACSQuant VYB  and analyzed with FlowJo 10 .Two different assays were performed to quantify apoptosis. Fluorochrome-labeled inhibitors of caspases (FLICA) assays were used to detect caspase activity in CB-MSCs. The polycaspase probe (SR-VAD-FMK), provided by ImmunoChemistry Technologies , recognizes all the different types of activated polycaspases. Cells were incubated with the polycaspase probe for 45 minutes at 37°C, under gentle agitation every 10 minutes. Then, nuclear counterstaining with Hoechst 33342 was performed. Cells were then scanned with the Operetta High-Content Imaging System at 570 nm and 380 nm excitation and 615 nm and 445 nm emission wavelengths. The data were analyzed using the Columbus software (PerkinElmer). Additionally, the differential detection of late and early stages of apoptosis was performed by FITC-Annexin V and ethidium homodimer III (EthD-III) staining, using a kit provided by Promokine . CB-MSCs were harvested after the AcS and stained according to the manufacturer's protocol. Cells in normal culture were used as negative control. CB-MSCs treated for 24 hours with 200 http://realtimeprimers.com , using their recommended amplification protocol. All the CT values were normalized to mRNA expression of HPRT1. We collected total RNA after 1 day of cultivation with FBS, CS, and HFS and then 4 hours after AcS, as well as 1, 3, and 5 days after AcS under continued cultivation with either FBS, CS, and HFS. The fold change of the mRNA expression before AcS of 119 genes was included in a heatmap, using average-linkage hierarchical clustering. The grouping of the genes in the heatmap was determined by calculating the distance among the groups by the Spearman rank correlation method.Total RNA was isolated using the NucleoSpin RNA isolation kit . The purity and integrity of isolated RNA were determined by spectrophotometry and gel electrophoresis. The RNA samples from biological replicates were pooled before cDNA synthesis to account for biological variation . cDNA waU test was applied to test for differences between groups. In cases where Levene's test showed heterogeneity, Welch ANOVA and Games-Howell post hoc tests were performed. The differences were considered statistically significant at p < 0.05. Repeated measures ANOVA was performed along with the experiments that had three or more time points. Statistical analyses were performed using the IBM SPSS Statistics software Version 22 .All results are shown as mean ± SEM. Unless stated otherwise, experiments were performed with group sizes of at least four control samples (FBS) and 12 human serum samples. Normal distribution and homogeneity of variances (Levene's test) were tested. With a normal distribution provided, ANOVA with Tukey's post hoc analysis was done; otherwise, the Kruskal-Wallis test or Mann–Whitney NCT00810238; CHART-1, NCT01768702) to expand autologous BM-MSCs prior to implantation. In one trial , human serum from healthy donors with the blood group AB was used to expand human umbilical cord MSCs. In one-third of the published trials, cell products that did not require any cell culture after isolation and prior to transplantation were used. This includes trials utilizing adipose-derived stromal vascular cells and bone marrow-derived mononuclear cell preparations.As shown in p = 0.03)  , indicat2. Combined with glucose starvation, this model significantly inhibited CB-MSC proliferation regardless of the source of the supplemented serum  and CS (72.9% ± 5.6%) was significantly higher than the cells treated with FBS (48.7% ± 2.1%)  (p = 0.04)  . Moreove = 0.04) . During  = 0.04) . As expe = 0.04) . After t = 0.04) .In our previous work, we showed that IL-6 titers are significantly higher in sera from patients with CHF . IL-6 faIn order to better understand the pathways regulating apoptosis and cell cycle that are affected during acute stress under the influence of the different sera, we performed an RT-PCR array with panels of mRNAs coding for the proteins involved in apoptosis and cell cycle pathways . Our anaAs mentioned in the previous section, the increased yield of CB-MSCs cultured with human serum could only in part be explained by the increased proliferation. After AcS, we observed significantly lower cell loss and decrease in apoptosis in the cells treated with human serum. The array analysis confirmed the activation of expression of multiple antiapoptotic mRNAs in HFS- and CS-treated cells, especially at the early time points after AcS. The most prominent was an increase in the expression of TRAF1 (TNF receptor-associated factor 1) and TRAF2 as well as mRNA expression of the BIRC-family. By binding the TRAF1/2 complex to the intracellular domain of the TNF receptor, TRADD (tumor necrosis factor receptor type 1-associated DEATH domain protein) inhibits the activation of caspases . BIRC2 (in vitro hypoxia/reoxygenation model mirrors the situation in myocardial ischemia incompletely, but cell behavior cannot be directly observed in respective in vivo experimental models. The culture period of 5 days is short; in cell products requiring longer MSC expansion, the impact of the different sera may differ. We chose this period because we were interested in primary CB-MSC proliferative behavior and stress response. Because cells usually reached confluence at day 5, passaging/subcultivation would be required for longer periods, impeding direct readouts regarding cell number and so on. The average serum donor age differed by roughly 10 years between the CS and HFS groups. This may have impacted the biologic activity of the sera. However, given that both groups were more or less middle-aged while the cells were neonatal, this confounding factor should be of little relevance. Finally, it may be argued that FBS is a pooled product, while the human sera were used individually. We chose not to pool the human sera because we were interested in the heterogeneity of serum activity and because in the clinical setting, autologous human serum would obviously not be pooled. Taken together, we feel that our data support the concept of using autologous serum for cardiac cell therapy, but other experimental designs would be required to understand biology and therapeutic relevance in greater detail.Clearly, our At odds with current clinical practice, human serum for MSC expansion in 2D culture is not only equivalent but also may even be superior to FBS in terms of proliferative capacity and resilience to acute and chronic “ischemic” stress. This is also the case when potentially autologous serum from patients with advanced heart failure is used. Using autologous serum simplifies GMP grade translational cell expansion, ought to reduce costs, avoids potential problems with xenogenic biomolecules, and may even have positive preconditioning effects on therapeutic cells.


MK levels were measured by enzyme-linked immunosorbent assay in 151 recruited DTC patients after exercising strict inclusion and exclusion criteria. There were 28 TgAb positive DTC patients with metastases and 123 DTC patients without metastases. The value of pre-131I-ablative MK to predict metastasis was assessed by receiver operating characteristic (ROC) curves in these two groups of patients. MK levels in the TgAb positive DTC patients were significantly higher than the DTC patients without metastases. ROC showed good predictability of MK, with an area under the curve of 0.856 (P < 0.001), and a diagnostic accuracy of 83% at the optimal cut-off value of 550 pg/ml. In conclusion, we show that MK can potentially be used as a surrogate biomarker for predicting DTC metastases when Tg is not suitable due to TgAb positivity.Serum thyroglobulin (Tg) is the main post-operative tumor biomarker for patients with differentiated thyroid cancer (DTC). However, the presence of thyroglobulin antibodies (TgAb) can interfere with Tg level and invalidate the test. In this study, we aimed to investigate the predicative value of midkine (MK) as a cancer biomarker for DTC patients with positive TgAb before the first  Serum thyroglobulin (Tg) is generally regarded as the mainstay post-operative tumor biomarker for patients with differentiated thyroid cancer (DTC)4739et al.10et al.et al.et al.18F-FDG PET/CT could be an effective alternative in the detection of recurrence or metastasis, and to resolve the TgAb controversy for such patients. The authors argued that TgAb level should be regarded as related with the immune system rather than the tumor load141117The false Tg result can have serious consequences for the follow-up of DTC patients, because it masks the disease status131I ablative therapyMidkine (MK) is a pleiotropic growth factor prominently expressed during embryogenesis yet down-regulated to a low level in healthy adults. In various pathologies, most notably in cancer, strikingly enhanced MK over-expression has been reported2021131I ablative therapy.In this study, we aimed to evaluate the predictive value of serum MK for DTC patients with positive TgAb before the first In our analysis cohort, we had 28 TgAb positive DTC patients with metastases (group 1), and 123 without metastases (group 2). All 151 patients were diagnosed with PTC. During the recruitment period, we had a total number of 849 DTC patients in the MK clinical investigation, among them 211 patients had positive TgAb. The prevalence of DTC patients with positive TgAb in our cohort was 24.85% (211/849). However, after implementing exclusion criteria, 60 cases were not included in the current analysis. Specifically, there were 5 cases with other malignancies , 22 cases of ischemic diseases , 1 autoimmune diseases (1 multiple sclerosis), 5 kidney diseases , 1 neural diseases (1 Alzheimer’s disease), 5 inflammation (5 rheumatoid arthritis), 10 hypertension (including 1 hypertensive nephropathy), 16 diabetes (including 4 diabetic nephropathy).131I therapy. MK was significantly higher in group 1 than in group 2, while age, TgAb, Tg, free triiodothyronine (FT3), free thyroxine (FT4) and thyroid stimulating hormone (TSH) did not display any significant differences between groups  showed a significantly worse 131I-avid metastasis-free survival .The current research focused on MK and thyroid cancer, which is the subject of several previous investigations. An immunohistochemistry study described strongly expressed MK protein in PTC, yet very faint or no expression of MK in normal follicular epithelial cellsIn this investigation, we aimed to assess whether MK could be a surrogate biomarker when Tg was not suitable to monitor the disease due to TgAb positivity in DTC patients. We displayed that MK was a good marker for predicting DTC metastases with an area under the curve value of 0.856 (P < 0.001), and a diagnostic accuracy of 83% with an MK cut-off value of 550 pg/ml. These results further confirmed our previous findings: a diagnostic accuracy of 89% for DTC metastatic prediction with an MK threshold level of 505 pg/ml14et al.et al.In addition to DTC, studies indicate that MK can outperform several currently used blood tumor specific biomarkers, such as alpha fetoprotein (AFP) for hepatocellular carcinoma (HC)32Although all the above evidence proves that MK can be a sensitive malignant biomarker for DTC, its shortcomings are also obvious, which are also the limitations of the current study. As pointed out by JonesWe demonstrate that MK can potentially be used as a surrogate biomarker for the prediction of DTC metastases when Tg is not suitable to monitor the disease due to TgAb positivity. Further longitudinal studies need to be conducted to validate the current finding.131I treatment were asked to consider entering the MK clinical investigation. If the patients consented, their serum MK levels were measured consequently along with their Tg and TgAb tests during follow-up. For the recruitment of the current investigation, MK database archive retrieval (from January 2011 till December 2015) was performed to identify DTC patients with positive TgAb. Patients with a confirmed post-surgical diagnosis of DTC in pathology and a positive TgAb prior 131I therapy were included in the study. Patients with MK influencing co-morbidities such as other malignancies, ischemic diseases, autoimmune diseases, kidney diseases, neural diseases, inflammation, hypertension and diabetes were excluded from the study. Patients with a negative TgAb prior 131I therapy were also excluded.Patients’ recruitment protocol was described previously131I for thyroid ablation, after a preparation of thyroid hormone withdrawal. Blood tests were conducted less than 2 days before 131I therapy. Routine examinations, including body weight, blood pressure, serum thyroid hormones, blood routine, biochemical indices , etc., are all measured. Detailed medical histories are collected. Four to 5 days after 131I administration, whole body scan (as well as tomography imaging if necessary) was performed by using a dual-detector SPECT/CT machine equipped with high-energy collimators. Before September 2014, Discovery VH SPECT/CT machine  was used for imaging. In September 2014, a new SPECT/CT machine, Discovery NM/CT 670 , was installed and used in our institution. Neck ultrasonography was also performed with frequency range from 3 MHZ to 12 MHZ using PHILIPS HD11 XE  to check cervical lymph nodes. Patients were positioned supinely with the neck hyper-extended and imaged by ultrasound. Approximately 6 months after the initial 131I therapy, a second 131I treatment protocol was implemented to those with confirmed metastases, or a diagnostic 131I scan to those without evidence of metastases.Management protocol for the DTC patients was conducted generally according to the 2009 guideline from the American Thyroid Association131I scan was performed, evaluation of each DTC patient was done. For this study, follow-up data were available for at least 6 months after the first ablation for all recruited DTC patients. Metastases were defined as positive hot spots in post-therapeutic 131I SPECT/CT scanFinal diagnosis was made by our panel of nuclear medicine physicians in consensus, based on a comprehensive consideration of imaging, serological and other clinical materials. All DTC patients were closely followed. After each The Institutional Review Board of Tianjin Medical University General Hospital approved the ethical and methodological aspects of this investigation. All participants provided their written informed consents to participate in this study. We confirm that all methods were performed in accordance with the relevant guidelines and regulations.FT3, FT4 and TSH assays were conducted on a fully automated ADVIA Centaur analyzer . Tg and TgAb were assessed on a fully automated IMMULITE 2000 analyzer . These assays were based on chemiluminescent reaction principle.The calibration references for the above indices were: FT3, 3.50–6.50 pmol/L; FT4, 11.50–23.50 pmol/L; TSH, 0.30–5.00 μIU/mL; Tg, 0–55.00 ng/mL; TgAb, 0-40.00 IU/mL. In the current study, TgAb > 40 IU/mL was defined as positive, otherwise negative.2O2 and tetramethylbenzidine) were added to the wells (100 μl per well) for a 20-minute reaction. Step 5, 1 mol/L H2SO4 (stop solution) was added (50 μl per well), and the optical densities of the wells were measured at 450 nm with a Multiskan MS Plate Reader . Step 6, after creating a standard curve by a four parameter logistic curve-fit method, concentrations of samples were determined.Fasting blood samples were obtained and centrifuged to collect serums, which were aliquoted, and stored at −80 °C until use. MK concentrations (reported as pg/mL) were measured in the following steps by using a commercial kit  in enzyme-linked immunosorbent assay methodology. Step 1, 100 μl of each serum sample or standard was incubated in 96-well microplate (pre-coated with goat anti-human MK antibody) for 2 hours at room temperature. Step 2, after washing three times, biotinylated goat anti-human MK antibody was added and incubated with captured MK for 2 hours at room temperature. Step 3, after another thrice washing, 100 μl aliquots of streptavidin-conjugated horseradish-peroxidase were added and allowed to react for 30 minutes in dark place. Step 4, after plate washing, substrate solutions T test. ROC curves were drawn and diagnostic efficacies were then determined. After the optimal cut-off MK value was selected, sensitivity, specificity, diagnostic accuracy, PPV  and NPV  for differential diagnosis were assessed. A P value not exceeding 0.05 was considered as statistically significant.All data were presented as mean ± standard deviation, which was statistically analyzed by SPSS 17.0  software. Differences of indices between two groups of patients were measured by independent samples How to cite this article: Jia, Q. et al. Serum midkine as a surrogate biomarker for metastatic prediction in differentiated thyroid cancer patients with positive thyroglobulin antibody. Sci. Rep.7, 43516; doi: 10.1038/srep43516 (2017).Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.


Adipose tissue is not only the main organ for energy storage, but it also has endocrine properties, producing “adipokines” responsible for energy homeostasis, insulin sensitivity, and inflammation. Leptin, produced by adipocytes, is the key hormone in appetite regulation and suppression of orexigenic, hypothalamic neuropeptide Y (NPY). We wanted to establish and compare levels of leptin and NPY in different obesity types in childhood, and to investigate their correlations with auxological parameters. Twenty-one obese children (seven girls and 14 boys), divided into two groups, were compared with 14 controls. The mean age of the study group was 10.81 ± 3.69 years and the mean puberty stage was 2.21. The mean body mass index (BMI) was 32.80 kg/m (range 23.30– 47.02) and the mean overweight 30.73 kg (range 8.00–74.00). The mean leptin level was higher in boys and in the group with central obesity, but was not significant. Leptin/NPY ratio and leptin/BMI ratio was also higher in the central obesity group and there was a more significant difference compared with controls. We found significant correlation of the leptin level with body mass (BM), body mass excess (BME), and BMI (p < 0.05). The mean leptin level in obese children was very high (36.39 ng/ml). Leptin and NPY levels showed inverse values in two different obesity types. Results are suggestive for leptin resistance rather than leptin deficiency in our group of obese children. Orexitropic signaling proteins correlated significantly with auxological parameters. Determination of the leptin and NPY concentrations provided evidence that obesity represents disease with neuroendocrine dysfunction and high leptin/NPY ratio, which could be a useful marker for central obesity.


Lymnaea stagnalis) embryos collected over 16 years from water bodies contaminated by Chernobyl fallout, and from control locations. The fraction of embryo cells with aberrations increased dramatically (>10-fold) and non-linearly over a dose rate range of 0.03–420 μGy/h (0.00026–3.7 Gy/year). NTE were very important for describing the non-linearity of this radiation response: the TE-only model (without NTE) performed dramatically worse than the TE+NTE model. NTE were predicted to reach ½ of maximal intensity at 2.5 μGy/h (0.022 Gy/year) and to contribute >90% to the radiation response slope at dose rates <11 μGy/h (0.1 Gy/year). Internally-incorporated 90Sr was possibly more effective per unit dose than other radionuclides. The radiation response shape for chromosomal aberrations in snail embryos was consistent with data for a different endpoint: the fraction of young amoebocytes in adult snail haemolymph. Therefore, radiation may affect different snail life stages by similar mechanisms. The importance of NTE in our model-based analysis suggests that the search for modulators of NTE-related signaling pathways could be a promising strategy for mitigating the deleterious effects of chronic irradiation.The biological effects of chronic ionizing radiation exposure can be difficult to study, but important to understand in order to protect the health of occupationally-exposed persons and victims of radiological accidents or malicious events. They include targeted effects (TE) caused by ionizations within/close to nuclear DNA, and non-targeted effects (NTE) caused by damage to other cell structures and/or activation of stress-signaling pathways in distant cells. Data on radiation damage in animal populations exposed over multiple generations to wide ranges of dose rates after the Chernobyl nuclear-power-plant accident are very useful for enhancing our understanding of these processes. We used a mechanistically-motivated mathematical model which includes TE and NTE to analyze a large published data set on chromosomal aberrations in pond snail ( Chronic ionizing radiation exposure can affect human health and ecosystem functioning –3. For eIn general, reduction of the radiation dose rate reduces the deleterious effects per unit dose , 10. In However, if the exposure is protracted over very long times such as multiple cell cycles or multiple organism generations, additional phenomena come into play: metabolism, reproduction, developmental pathways and cell proliferation/differentiation programs must operate under continuously stressful conditions, and susceptibility to radiation effects often changes with the organism’s age/stage. In other words, the biological responses to acute and chronic irradiations can be qualitatively different because radiation acts as a transient stressor in the first scenario and as a continuous one in the second , 12.The ability of a given organism to resist one type of radiation stress does not necessarily correlate with its ability to resist the other. Examples of poor correlations between resistance to acute and chronic irradiation have been found in prokaryotes –15 and iIonizing radiation acts on cells through multiple mechanisms. Ionizations in or very close to DNA (e.g. in the first hydration layer) can produce DSBs and other forms of DNA damage which can kill the cell or potentially transform it into a pre-malignant state , 19. TheThe signaling pathways involved in NTE are complex and incompletely understood. However, their consequences can be quantitatively modeled in a simple and tractable manner by using the following assumptions: (1) Irradiated cells “activate” nearby cells in an “on-off” (binary) manner. (2) Activated cells accumulate DNA damage  at an elevated rate. Eventually they can revert to the background state, but this process may be very slow in some cases (e.g. require years-decades). (3) If the radiation dose is protracted, cells remain activated longer. The result is an increased yield of damaged cells–an inverse dose-rate effect, where chronic irradiation becomes more effective per unit dose than acute irradiation. Such effects have been observed after exposure to densely ionizing radiation, e.g. for lung cancer in radon-exposed miners –29. (4) in vitro and in vivo data  represents a decrease in radiation-induced effects over time: T0 is the year when observations began , T is tarameter . This exdM(R)/dR, is:The “slope” of the radiation response could be estimated at any selected dose rate by differentiating Gudkov et al.  did not 137Cs, 90Sr, 238Pu, 239+240Pu, 241Am; (b) the dose rate from 137Cs and 90Sr in the water; and (c) the dose rate from external γ-rays . The solution for LL is:In other words, eference , and F(RN–K)!] + ln[N!]–ln[K!] do not contain F(R) and, therefore, have no role in determining the best-fit values of parameters bac, a, b, q and c. They can be omitted for convenience.The terms ln, applies directly to the dose rate, rather than to the biological effects:In addition to the TE+NTE and TE-only models, we used the IT metrics ΔAICc and W to compare the performances of various other model versions. Specifically, an NTE-only model was generated by setting  0 in Eq . A time-WF(i) for specific radionuclides, which were described earlier, into each of these model structures . For each version, Poisson and NB error distributions were analyzed separately.Multiple other model versions were produced by substituting dose rete estimates produced using weighting factors The data on chromosomal aberrations in snail embryos exposed to different radiation dose rates  are show−4 and 1.8–2.6×10−3 Gy/year), but the steepest rate of increase occurred between lightly and highly contaminated locations (dose rates between 1.8–2.6×10−3 and 0.31–0.51 Gy/year). The increase became less steep at high contamination levels (dose rates between 0.31–0.51 and 3.07–3.68 Gy/year) [The radiation response shape was clearly non-linear. Specifically, the response showed some increase between uncontaminated (background) locations and lightly contaminated locations  was approximately 0.022 Gy/year, i.e. 2.5 μGy/h (b) was approximately 0.24 (a) was about 0.008 year/Gy, i.e. 7.0×10−5 h/μGy  with time after the Chernobyl accident in all water bodies was consistent with the data  was about 0.05 year-1  to the total dose rate experienced by snails differed substantially among different water bodies [90Sr was weighted by a factor of 25   . This fo90Sr contribution decreased the differences in radiation levels between the highly contaminated water bodies . This occurred because the fraction of the total dose rate which was due to internal radionuclide incorporation differed strongly among locations: e.g. internal exposure contributed only 5% to the total dose rate in Globokoye lake, but >50% in Azbuchin lake and Yanosvky crawl [a overlapped zero in the internal Sr model , whereas at higher dose rates their predicted contribution to the radiation response declined , Fig 5. s), which scaled the dose response from the embryo data to the adult data, but did not alter its shape. The best-fit value of s was 12.65, and the fitted curve was visually consistent with the data ..Fig 6). Lymnaea stagnalis. The analyzed data sets on chromosomal aberrations in snail embryo cells and on the composition of adult snail haemolymph were presented graphically in a publication by Gudkov et al. [We conducted a mechanistically-motivated quantitative analysis of the effects of chronic irradiation from Chernobyl nuclear-power plant accident fallout on natural populations of a common aquatic invertebrate: the pond snail v et al. . We recov et al. , 31. CerThese data showed a non-linear radiation response . The robL. stagnalis is reported by Golubev et al. [The cell signaling pathways responsible for radiation-induced NTE are actively studied but the complex mechanisms involved remain incompletely understood , 43–45. v et al. : snails 90Sr was 25 . 90Sr and its decay product 90Y are almost pure β emitters. Although the DNA damaging and cytotoxic potencies of β-particles are not as high as those of α-particles, the high importance of 90Sr (rather than some α-emitting radionuclides) in L. stagnalis is plausible because molluscs accumulate 90Sr in their shells [90Sr may be highly relevant for molluscs not only because of its radioactivity, but because of the combination of its radioactivity with its chemical properties, which result in its long-term incorporation into mollusk tissues.Our analysis provided some evidence that different radionuclides may have different effectiveness in inducing NTE. For example, the best-fit weighting factor for internally accumulated r shells . In otheInteractions of radiation with other environmental factors , and with chemical pollutants and/or pathogens, could in principle affect chromosomal aberration yields in the studied snails. Variations in snail population structure (e.g. age/stage distributions) at different times or locations also could confound the effects of radiation. Unfortunately, such factors could not be investigated using the analyzed data from Gudkov et al. , 31. HowL. stagnalis on a physiological level. Most of the effects at low dose rates (<1 Gy/year) were likely to be caused by NTE Click here for additional data file.


While six herbal constituents of HJG, Rehmannia root, Dioscorea rhizome, Rhizoma Alismatis, Poria sclerotium, Moutan bark, and Cinnamon bark, could induce neurite outgrowth effects, the effect was strongest with HJG (500 μg/ml). Second, we demonstrated that HJG-induced neurite outgrowth was blocked by an inhibitor of cAMP response element binding protein (CREB), KG-501 . Moreover, HJG was observed to induce CREB phosphorylation 20–90 min after treatment  and CRE-mediated transcription in cultured PC12 cells . These results suggest a CREB-dependent mechanism underlies the neurotrophic effects of HJG. Finally, we examined improvements of memory impairment following HJG treatment using a Morris water maze in AD model animals (CI + Aβ rats). Repeated oral administration of HJG improved memory impairment  and induced CREB phosphorylation within the hippocampus . Together, our results suggest that HJG possesses neurotrophic effects similar to those of NGF, and can ameliorate cognitive dysfunction in a rat dementia model via CREB activation. Thus, HJG could potentially be a substitute for neurotrophic factors as a treatment for dementia.Hachimijiogan (HJG) is a traditional herbal medicine that improves anxiety disorders in patients with dementia. In this study, we tested the hypothesis that HJG exerts neurotrophic factor-like effects to ameliorate memory impairment in Alzheimer disease (AD) model rats. First, we describe that HJG acts to induce neurite outgrowth in PC12 cells (a rat pheochromocytoma cell line) like nerve growth factor (NGF) in a concentration-dependent manner (3 μg/ml HJG,  Recently, there have been global increases in the number of patients with dementia disorders, such as AD. However, as the pathogenesis of dementia has not yet been fully elucidated, radical curative therapies have not been established. Novel dementia drugs that do not adversely affect ADL are required. In the study of neurodegenerative diseases characterized by neuronal cell death (such as AD), the role of neurotrophic factors has recently drawn attention. Neurotrophic factors are substances that promote neuronal survival, differentiation, and regeneration. Although various trophic factors have been identified, research has indicated particularly important roles for NGF and BDNF .in vivo half-life that is too short to enable delivery to a target tissue or cell. This makes it difficult to directly apply neurotrophic factors as therapeutic agents for AD  is an herbal medicine used in China and Japan comprising eight herbs: RR, Cornus fruit, DR, AR, PS, MB, CB, and Aconite root. HJG has traditionally been used to treat diabetes mellitus, hypertension, and nephrotic syndrome. It has also been used to treat dysuria, lumbago, lack of energy, and poor eyesight in older individuals. Furthermore, it has been reported that HJG treatment improved cognitive dysfunction in patients with dementia . AdditioRehmannia glutinosa Libosch var. purpurea Makino, Scrophulariaceae), Cornus fruit , DR , AR , PS , MB , CB , and Aconite root .Dry powdered extracts of HJG and its constituents (excluding Cornus fruit and Aconite root) supplied by Tsumura & Co.  were dissolved in distilled water. HJG is a dried extract of the following eight herbs: RR . Dimethyl sulfoxide and KG-501 (a CREB inhibitor) were purchased from Sigma–Aldrich . Anti-CREB, anti-phospho-CREB, and anti-rabbit horseradish peroxidase-conjugated secondary antibodies were purchased from Cell Signaling Technology . Anti-rabbit β-actin antibodies were obtained from Abcam .Donepezil hydrochloride was purchased from Tokyo Chemical Industry . For animal treatments, HJG and DPZ were dissolved in distilled water. Three weeks after CI, HJG, DPZ, and vehicle (distilled water) were orally administered daily for 7 days.2. The medium was changed two or three times per week. Cells treated with NGF (50 ng/ml) were used as positive controls.PC12 (rat pheochromocytoma) cells were cultured in RPMI-1640 with 5% (v/v) fetal bovine serum, 10% (v/v) horse serum, 100 units/ml penicillin, and 100 μg/ml streptomycin. Cells were grown to confluence at 37°C in 5% CO5 cells/ml) were seeded on collagen-coated six-well plates and cultured for 24 h. Cells were treated with different dilutions of HJG (1–500 μg/ml), or each herb at the concentration based on the original mixing ratio of constituents, and cultured for 72 h. PC12 cells were photographed using an inverted microscope  and phase-contrast objectives. Images of two fields per well were taken with 40–50 cells per field. The length and number of neurites were measured for 20 independent cells per field using Image J 1.44 software (NIH Image Software), and the neurite lengths for each cell were summed. Each experiment was conducted in triplicate.PC12 cells (1 × 10Proteins were extracted from cells at appropriate time points after drug treatment using RIPA buffer  containing a protease inhibitor cocktail . Protein concentrations were measured using a BCA Protein Assay Reagent Kit . Proteins were separated on SDS polyacrylamide gels, transferred to polyvinylidene fluoride membranes, and probed with specific antibodies. Immunoreactive polypeptides were detected with chemiluminescence using ImmunoStar LD .5 cells/well) were seeded in collagen-coated 12-well plates for 24 h. Cells were co-transfected with the pGL4.29 luciferase reporter vector containing a CRE and pGL4.74 Renilla luciferase vector (Promega) using Lipofectamine 3000 (Life Technologies). Forty-eight hours after transfection, cells were treated with vehicle (0.1% DMSO), HJG, or its constituents for 8 h and harvested using Passive Lysis Buffer (Promega).PC12 cells  according to the manufacturer’s instructions. We normalized the intensity of luciferase reactions measured in the lysates of transfectants to their Renilla luciferase activity, which was used as an internal control.ad libitum except during the restricted feeding period. We restricted food intake for rats (8–10 g each day), and their body weight was maintained at ∼80% of free-feeding levels during the eight-arm radial maze tasks. We conducted all procedures relating to animal care and use according to regulations dictated by the Experimental Animal Care and Use Committee of Fukuoka University (No. 1405744).Male Wistar rats weighing 250–300 g were obtained from Kyudo Co., Ltd. . Rats were housed in groups of 4–5 per cage at 23 ± 2°C with a relative humidity of 60 ± 10%, and maintained under a 12-h light–dark cycle. Food and water were available Aggregated Aβ was prepared as previously described . Aβ42 peL = ±1.3 mm, and H = 3.3 mm from the bregma. Three days after cannulation, the common carotid arteries were compressed by clips and cerebral circulation was interrupted for 10 min to create CI.Cerebral ischemia + Aβ rats were prepared by combining an intracerebroventricular Aβ injection with transient CI as previously described . Rats weThe first injection of Aβ was performed 1 h after CI. Aβ was injected daily for 7 days. Aβ (600 pmol/20 μl/day) was injected bilaterally using injection cannulas  connected by polyethylene tubing to a perfusion pump  and driven at a rate of 1 μl/min.Apparatus and procedures for the water maze  were modified from those originally developed by A trial consisted of placing the rat by hand into the water facing the wall of the pool at one of three starting positions located in a quadrant that did not contain the platform. The platform (12 cm in diameter) was located in a constant position in the middle of one quadrant with its top surface located 2 cm below the surface of the water. During each block of three trials, the rat started once at each of three starting positions. Rats that demonstrated a swimming time on the 5th day within 20 s of the average were deemed to have memorized the maze. In the probe test, the platform was removed from the tank and rats were allowed to swim freely for 120 s. Their performance in the probe test was assessed by swimming time to first reach the platform position.t-test. Differences between the control group and each treatment group were subjected to one-way analysis of variance (ANOVA) followed by Dunnett’s test. Multiple comparisons were evaluated with Tukey’s test after one-way ANOVA. Values of p < 0.05 were considered statistically significant.Results are indicated as mean ± SEM for three or more experiments. Experiments with two groups were analyzed using Student’s in vitro neuronal model characterized by the extension of protrusions and differentiation into sympathetic cells following stimulation with NGF . PC12 cells treated with HJG for 3 days exhibited clearly different morphologies compared with the control group, including protrusion elongation . By measuring the length and number of neurites, we observed a significant increase in total neurite length as the concentration of HJG increased from 3 to 500 μg/ml compared with the control group . Neurite outgrowth was represented by neurite extension , neurite numbers , and length per individual neurite . Using these methods, we searched for effective herbal constituents of HJG capable of exerting an elongation effect. The total length of neurites was increased significantly compared with the control group . Examination of CREB phosphorylation with Western blots showed increased phosphorylation of CREB 20–90 min after treatment with HJG, suggesting CREB activation by HJG . Subsequently, we investigated neurite outgrowth induced by HJG administered with KG-501 (10 μM), a CREB inhibitor. Co-treatment with KG-501 significantly inhibited the neurite outgrowth effect of HJG .We investigated the effect of HJG on CREB, which exists downstream of neurotrophic factor intracellular signaling pathways. CREB plays an important role in neurogenesis and memory formation .Cyclic AMP response element-binding protein, a transcription factor, is known to increase the expression of target genes in a CRE-dependent manner. Therefore, we examined the effect of HJG on transcriptional activity of CREB using a CRE reporter vector. As a result, we confirmed a significant and concentration-dependent increase in CRE-dependent transcriptional activity with HJG treatment , a CREB inhibitor. Neurite outgrowth was observed following treatment with both HJG and its six constituents, and was suppressed by KG-501 treatment . However, while PS showed a particularly strong effect, CREB-dependent neurite outgrowth was also observed following treatment with the other five herbal constituents . Furthermore, as there exist various pathways by which neurite outgrowth occurs, we acknowledge that factors and pathways other than CREB likely contribute to the effect of HJG on neurite outgrowth.Next, we examined the effect of each constituent on CREB transcriptional activity using a CRE reporter vector. Among the HJG constituents, we observed particularly significant increases of CRE-dependent transcription activity following treatment with PS . In animals with combined CI and intraventricular administration of Aβ (CI + Aβ rats), treatment with HJG showed a concentration-dependent amelioration  of previously significantly longer swim times to first reach the platform position . DPZ, an inhibitor of acetylcholinesterase, also reduced swim time . There were no significant differences in swimming speed between any of the groups (data not shown). Therefore, both HJG and DPZ (a treatment for AD) showed similar amelioration of memory disorder in dementia model rats.After confirming that HJG has neurotrophic factor-like effects and activates CREB Figure 5C). When dementia model rats underwent 7 days of continuous oral HJG administration, there was a significant increase in phosphorylated CREB . Therefore, we suggest that HJG improved memory impairment by restoring phosphorylated CREB in the hippocampus of CI + Aβ rats.Given the confirmation that HJG improved cognitive function, we subsequently examined activation of CREB by HJG in the brain of CI + Aβ rats. Phosphorylated CREB protein in the hippocampus was quantified with Western blot . In the current study, we showed that the CREB-mediated pathway is mechanistically related to the neurotropic effects of HJG. Considering that HJG is a multicomponent drug, other mechanisms may also be associated with these effects, which is an important point for understanding the complete neuropharmacological actions of HJG.First, we examined neurite outgrowth following HJG treatment by examining neurite elongation effects in PC12 cells. Our results showed that HJG induced neurite outgrowth in a similar manner to neurotrophic factors. In addition, our results suggested that the constituents of HJG did not significantly influence the total number of neurites, but did elicit neurite elongation activity. We next attempted to identify the candidate constituent herb(s). As shown in We previously showed that memory impairment in CI + Aβ rats was ameliorated by HJG administration for 7 days after ischemia treatment . In our In this study, HJG elicited CREB phosphorylation in cultured PC12 cells and AD model rats. However, it is still not clear how HJG induces CREB activation in detail. Some candidate pathways are involved in activation of CREB. Many neurotrophic factors and neurotransmitters act as membrane receptors to trigger signaling pathways that regulate CREB phosphorylation . PhosphoCyclic AMP response element-binding protein has been reported to regulate the transcription of various genes, as well as neuronal differentiation, cell survival, protrusion elongation, and memory/learning ability . The actWe have shown that HJG exerts neurotrophic effects via CREB activation, similar to that of neurotrophic factors. Furthermore, HJG ameliorates cognitive dysfunction in dementia model rats via CREB activation. Our results suggest that HJG could be a potential substitute for neurotrophic factors as a treatment for dementia. As CREB is not only related to memory formation but also depression and anxiety , HJG mayKK, TW, and KI designed the study. KK, HF, HS, KH, AF, and HM conducted the research and analyzed the data. KK, TW, and MK wrote the paper. SK, MK, and KI supervised the experiments. All authors read and approved the final manuscript.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.


Shedding of the endothelial glycocalyx layer (EGL) is known to occur during major surgery, but its degradation associated with minimally invasive video-assisted thoracoscopy (VATS) remains unclear. We investigated if serum biomarkers of EGL disruption were elevated during VATS lobectomy, and whether the urinary trypsin inhibitor (UTI) ulinastatin exerted a protective effect during this procedure. Sixty ASA II-III lung cancer patients undergoing elective VATS lobectomy were divided equally into UTI and control groups. UTI group patients received intravenous UTI during surgery. Serum levels of syndecan-1 and heparan sulfate were examined before (T0) and at the end of surgery (T1). Serum albumin and hemoglobin were measured before surgery (BOD) and on the first postoperative day (POD1).P = 0.022⁎) and increased even more significantly in patients whose surgery lasted >3 h . Serum albumin levels on POD1 were significantly lower in control group compared with UTI group . In control group, syndecan-1 levels were significantly elevated at T1 compared with T0 . This trial is registered with  Lung resection is the preferred therapeutic choice for suitable lung cancer cases. However, acute lung injury (ALI) and induced acute respiratory distress syndrome (ARDS) are the leading cause of death after thoracic surgery, leading to significantly reduced 1-year survival . MinimalThe endothelial glycocalyx layer (EGL) is a dynamic layer mainly consisting of glycoprotein and proteoglycan layers at the luminal surface of the vascular endothelium. Although the precise mechanism is not well understood, this endothelial surface layer is believed to act as a barrier to circulating cells and large molecules involved in fluid homeostasis and regulation, and it thus plays as key role in vascular permeability and edema formation , 9–13. IChiCTR-IOC-17010416). Sixty ASA II-III lung cancer patients undergoing elective VATS lobectomy were selected to participate in the study. Informed consent was obtained from all patients. Exclusion criteria were body mass index >35 kg/m2, cardiac failure (New York Heart Association class >II), pulmonary hypertension , clinically relevant obstructive or restrictive lung diseases, obvious coagulation disorders, severe pulmonary or systemic infections, treatment with immunodepressant drugs in the 8 weeks before surgery, and tobacco abuse.Ethical approval for this clinical cohort study (number 2016-sci-122) was provided by the Ethics Committee of Beijing Chaoyang Hospital Affiliated to Capital Medical University, Beijing, China, and the study was registered at clinicaltrialecrf.org (n = 30) and UTI group (n = 30) based on a list of random numbers generated by the RAND function in MS Excel. General anesthesia was induced with midazolam 0.02 mg/kg, propofol 1.5–2.5 mg/kg, and sufentanil 0.5 μg/kg. Tracheal intubation using a double-lumen endotracheal tube was facilitated by administration of rocuronium (0.8 mg/kg). The correct position of the tube was confirmed by fiberoptic bronchoscopy. Anesthesia was maintained by continuous infusion of propofol (3–5 mg/kg/h) and remifentanil (0.1–0.4 μg/kg/min), and administration of rocuronium. Supplemental sevoflurane 0.5–1 v/v% was used if needed. The patients were placed in the lateral decubitus position for VATS lobectomy with OLV. The tidal volume was set to 6 ml/kg, the fraction of inspired oxygen (FiO2) was adjusted to maintain oxyhemoglobin saturation >95%, and the respiratory rate was set to maintain PaCO2 between 35 and 45 mmHg. When the main procedure was finished, double-lung ventilation was restored for reconstruction. Patients in the UTI group received intravenous ulinastatin for injection  with a loading dose of 5000 IU/kg at the beginning of surgery, followed by infusion of 1000 IU/kg/h during surgery. The control group was given the same volume of normal saline solution. Lactated Ringer's solution and 6% 130/0.4 hydroxyethyl starch were infused during the procedure, but no exogenous albumin solution was applied.The patients were divided randomly into a control group  levels, fibrinogen degradation products (FDP), fibrinogen (Fbg), D-dimer, and international normalized ratio (INR) were measured before surgery (BOD) and on the first postoperative day (POD1).Blood samples were collected from the radial artery before surgery (T0) and at the end of surgery (T1). The blood samples were centrifuged after placed stable for 1 hour and the serum was then stored at −80°C for future use. The levels of the glycocalyx components syndecan-1  and HS  were measured by enzyme-linked immunosorbent assay (ELISA). Arterial blood gases were analyzed and the PaOP < 0.05. Mean values were compared between groups by independent sample t-tests, and mean values within groups were compared by paired t-tests. Numerical data were compared by Pearson's χ2 tests. SPSS 15 and GraphPad Prism software (version 6) were used for statistical analysis and for creating the graphs, respectively.Normally distributed data are presented as mean ± standard error of the mean. The results of statistical tests were considered significant if P = 0.022), whereas the increase in syndecan-1 levels was not significant in the UTI group . The procedure lasted >3 h in 15 control and 19 UTI cases (long-duration groups). Further analysis of these long-duration cases showed that syndecan-1 levels were highly significantly elevated at T1 compared with T0 in the control group , whereas the increase was still not significant in the UTI group  (P = 0.031). There was no significant difference in HGB levels between the two groups over the same time periods  . There w= 0.138) . Serum a= 0.138) , and POD periods , and theer group . Fbg, FDer group , but theer group .In this study, we treated patients undergoing VATS lobectomy with the UTI ulinastatin at the routine dose permitted in China, as used in other studies , 19. SerDeterioration of the EGL is known to be caused by major surgery, and serum syndecan-1 and HS are important biomarkers of this process –24. Diff2 may also contribute to EGL disruption [2/FiO2 and lactic acid, though this may have been because the sample size was not powerful enough to investigate the relationships between biomarkers of EGL shedding and clinical outcomes such as poor tissue oxygenation. It is also possible that our results were affected by not using a unified stepwise method to analyze lung recruitment [The function of the EGL in the setting of VATS lobectomy has not previously been explored. The thickness of the endothelial surface layer in pulmonary vessels is considered to exceed that in other vascular beds, indicating an important barrier function in lung vessels . It is tsruption , 30. Glysruption . All thesruption . Furthersruption , 34. Sersruption , 36. Neusruption , while nsruption , 38. In sruption , and furruitment , and thiALI after lung resection remains a poorly understood condition with high mortality and limited therapeutic options. EGL dysfunction has demonstrated a close relationship with ALI and ARDS. This study demonstrated EGL shedding following VATS lobectomy and confirmed that the applied dose of UTI could alleviate this degradation and decrease albumin extravasation, thus preserving vascular permeability. These results suggest that the glycocalyx may represent a therapeutic target for improving ALI following lung resection. Further studies are required to investigate EGL function in the perioperative setting and to explore the effects of different UTI doses on the EGL and clinical outcomes.