Background and Objectives: The current study was performed to evaluate the prevalence of periodontitis and to examine the association between reduced pulmonary function and periodontitis using Sixth Korea National Health and Nutrition Examination Survey (KNHANES) in 2014. Materials and Methods: A cross-sectional evaluation was conducted to estimate the prevalence of periodontitis and to examine the association between periodontitis and reduced pulmonary function while adjusting for sociodemographic characteristics and current smoking status in survey participants between 40 and 79 years old. The presence of periodontitis was evaluated by community periodontal index defined by the World Health Organization, and the assessments of reduced pulmonary function data were made as “normal,” “restrictive impairment,” or “obstructive impairment.” Results: A total of 4004 survey participants representing 25.4 million Koreans were included in the study. Overall, 41.1% of the study population were determined to have periodontitis, and 22.1% had reduced pulmonary function; 7.9% and 14.2% had restrictive- and obstructive- pulmonary impairments, respectively. Age, male gender, and current smoking status were positive predictors for periodontitis. Insurance coverage by workplace and higher education were protective factors against periodontitis. The association between periodontitis and restrictive impairment (adjusted odds ratio (OR) = 1.059, 95% CI 0.729–1.540) or obstructive impairment  was not significant. Conclusions: For Koreans, 40–79 years old, age, smoking status, gender, education, and insurance coverage were significant predictors of periodontitis. The prevalence of periodontitis was not significantly associated with reduced pulmonary function. To better understand the relationship between periodontitis and reduced pulmonary function, well-designed and larger scale epidemiologic studies are needed. Chronic obstructive pulmonary disease (COPD) is a generic term used to describe chronic lung diseases, including emphysema, chronic bronchitis, small airways disease, and non-reversible asthma . It affePeriodontitis is a chronic inflammatory disease that can lead to tissue destruction as a result of the perturbation of the homeostasis between the subgingival microflora and the host defense in susceptible individuals ,5. ImmunRecently, more pieces of evidence have shown that a poor oral and periodontal condition is a potential risk factor for systemic diseases, strongly indicating the two-way interrelationship between oral and general systemic health ,10,11. FA link between periodontitis and COPD has been proposed. Several studies have suggested that periodontitis is a co-factor for COPD . It has The purpose of the current study was to evaluate the prevalence of periodontitis and to examine the association between lung function test results, as indicators for COPD, and periodontitis using a national-level population data.The study used a nationwide, population-based, cross-section survey known as the Korea National Health and Nutrition Examination Survey version 6 (KNHANES VI-1) data, specifically KNHANES 2014 data . The surOut of 7550 survey participants from the 2014 survey data, a total of 4004 participants, who were between 40 and 79 years old, were included in the analysis. This age group was determined as the study group because the pulmonary function test was collected only from these participants. Baseline characteristics, including sociodemographic characteristics , lifestyle factor on current smoking status, pulmonary function test assessments, and clinical condition of periodontitis, were collected. 1). Obstructive impairment was defined as FEV1/FVC < 0.70, and restrictive impairment was defined as FEV1/FVC ≥ 0.70 and FVC < 80%, while normal lung function was defined as FEV1/FVC ≥ 0.70 and FVC ≥ 80% [Pulmonary function test included measurements of forced vital capacity (FVC) and forced expiratory volume in one second  community periodontal index (CPI) was used to assess the periodontal status . Using aThe analysis of the study focused on estimating the prevalence of recorded periodontitis diagnosis and evaluating its predictive factors. Using the unweighted sample data and the assigned sampling weight variable adjusted for the survey participants on pulmonary function test and complex sampling design, national-level estimates were calculated when they were aggregated to generate weighted frequencies. Descriptive statistics were used to calculate the frequencies of demographic characteristics and the prevalence of disease status. Univariable and multivariable logistic regression analyses were performed using weighted data. To assess the predictive factors for periodontitis diagnosis as the dependent variable (yes/no), a multivariable logistic regression model was constructed with pulmonary function test assessment as the independent variable with “normal lung function” as the reference group and determined the adjusted ORs with 95% confidence intervals (CIs) while adjusting confounders available from the data source and relevant to healthcare access, such as age, gender, current smoking status, level of education, marital status, and health insurance type, reflecting the Korean health insurance structure. p < 0.05. The study used publicly available national-level data without patient identifiers or linkable information. The study was approved by the Institutional Review Board from the Chonnam National University Dental Hospital, Gwangju, Korea .All analyses in the study used weighted data to yield national-level frequencies using SPSS version 23.0 . The significance level was set at From the KNHANES VI 2014 survey including 7550 study samples, a total of 4004 survey participants whose ages were between 40 and 79 years old representing 25.4 million Koreans were included in the study population. The mean (SD) age of the survey participants was 58.48 (SD 11.09). Demographic characteristics are described in The prevalence of periodontitis was 41.1%. as determined by CPI. and 22.1% of the study population had reduced pulmonary function; 7.9% and 14.2% had restrictive- and obstructive- pulmonary impairments, respectively .Univariate and multivariate associations were evaluated by logistic regression analysis using weighted data. Age and current smoking status (adjusted odds ratio (OR) = 2.330, 95% CI 1.745–3.110) showed a statistically significant relationship with periodontitis . Male paFor the pulmonary function assessments, obstructive pulmonary impairment, but not restrictive pulmonary impairment, was a significant predictor of periodontitis from univariate analysis . However, the association between periodontitis and restrictive pulmonary impairment  or obstructive pulmonary impairment  did not reach a statistical significance.In the present study, it was found that the prevalence of periodontitis was determined to be 41.1% among Korean population aged 40–79, as compared to 25.3% that was observed among adults aged over 19 in our previous study using identical national survey data . This fiIn this study using a nationally representative population, we could not find any statistically significant association between reduced lung function and the prevalence of chronic periodontitis. Although several studies in the literature evaluated the association between pulmonary diseases and periodontitis ,23,30, i1/FVC <70%) was used as a criterion for COPD, as defined by the Global Initiative for Chronic Obstructive Lung Disease (GOLD) guidelines, while other epidemiological studies adopted the value less than 65% of FEV1/FVC for COPD [The strengths of our study include the source of the data from national representative samples and the quality of the data from a quantitative analysis of the impaired and/or reduced lung function. More specifically, “obstructive” lung function (FEVfor COPD . AlthougA link between periodontitis and respiratory diseases, including COPD, has been proposed, as they may share common pathological processes resulting from immunological and inflammatory activities. It was suggested that periodontal microbes might play a role in the pathogenesis of COPD , as dentThis study has several limitations, primarily due to the nature of the KNHANES IV as a secondary dataset. First, the periodontitis was assessed by the CPI, i.e., a simpler method of assessing the needs for periodontal treatment in a community setting. The CPI is reported to overestimate or underestimate the prevalence of periodontitis because the use of representative teeth does not truly reflect the whole periodontal condition . BesidesDespite several pieces of evidence suggesting a link between COPD and periodontitis, the exact mechanisms involving the association have not been fully elucidated. Therefore, well-designed future prospective observational studies or large scale big data analyses are required to clarify the causality, biological mechanisms, and clinical relationships in the future. Also, more research is required to improve and simplify technologies to assess the COPD condition reflecting the inflammatory and destructive process in the lung or to indicate responsiveness to treatment.The results of the present study revealed that age, smoking status, gender, education, and insurance coverage were significant predictors of periodontitis. The reduced pulmonary function, both obstructive- and restrictive-pulmonary impairments, was not a significant predictor for the prevalence of periodontitis. Well-designed epidemiologic studies are needed to better understand a potential link and further find a causal relationship between periodontitis and obstructive pulmonary impairment.


Ibuprofen is a commonly used medication in the United States and is used both by prescription and over the counter, while hypokalemia is a life-threatening condition caused by various etiologies, one of which is the side effect of medications. Ibuprofen is well-known for its various nephrotoxic side effects, including hyperkalemia as a common electrolyte abnormality, however, renal tubular acidosis leading to hypokalemia with the use of ibuprofen has been reported rarely. We present here two cases of life-threatening hypokalemia due to over-the-counter use of large doses of ibuprofen and describe its management. Ibuprofen-induced hypokalemia was first described by Gaul et al.  in 1999,Patient 1A 52-year-old African American male with a medical history of uncontrolled hypertension, chronic obstructive pulmonary disease (COPD), current smoker with a 20 pack-year smoking history, and arthritis was sent to the emergency department (ED) by his primary care provider, with severe hypokalemia on routine lab work. On a review of systems, he endorsed chronic right shoulder pain, back pain, loss of weight about 15 pounds in the last two months, loss of appetite, and difficulty with walking up and down the stairs for the past few days before presentation. On presentation to the ED, vitals were within normal limits. Examination showed normal strength in both the upper and lower extremities with no focal weakness. Initial workup showed significant hypokalemia of 1.9 mMol/L, bicarbonate 17 mMol/L, anion gap (AG) 16 mMol/L, and albumin 3.6 gm/dL  of KCL on day one. The urine potassium to creatinine ratio was 48.9 mEq/gm, indicating renal potassium loss. A paraneoplastic syndrome causing hypokalemia was suspected as a possible etiology but adrenocorticotropic hormone (ACTH) was found to be low (<5 pg/ml). Hyperaldosteronism was ruled out, as the serum aldosterone was < 1.0 ng/dL, and he remained normotensive through the hospital course. Urine anion gap was found to be negative, as detailed in Table The patient continued to have hypokalemia despite daily replacement, as shown in Figure After ruling out the other possible etiologies, a diagnosis of Ibuprofen-induced hypokalemia and proximal RTA was reached. The patient was aggressively treated with intravenous fluids (IVF) and he received a total of 1080 mEq of KCL replacement over 11 days of hospitalization Figure  along wiPatient 2A 63-year-old African American female with a history of chronic venous stasis ulcers, an unknown history of renal disease presented to the ED with complaints of epigastric pain, nausea, and vomiting for 10 days. On presentation, vitals were within normal limits and physical examination was unremarkable except for chronic ulcers on bilateral lower extremities. Her initial lab results were significant for a potassium of 2.0 mMol/L, bicarbonate of 17 mMol/L, AG of 16 mMol/L, blood urea nitrogen (BUN) of 85 mg/dL, and creatinine of 3.9 mg/dL. The rest of the labs are outlined in Table Potassium on day two of admission remained low at 2.3 mMol/L and even after 180 mEq of KCL was repleted, it continued to be low at 2.7 mMol/L on day four. Meanwhile, the urine potassium/creatinine ratio was obtained on day four, and it was 20.6 mEq/gm, indicating urine loss of potassium. The patient remained normotensive, and the serum aldosterone was 2 ng/dL, ruling out hyperaldosteronism. The urine anion gap was found to be positive 12.7 and urine pH was 6 pointing toward the distal renal tubular acidosis (dRTA).After ruling out the other possible etiologies, a diagnosis of ibuprofen-induced hypokalemia and distal RTA was reached. She was aggressively treated with fluid and electrolyte replacement, as shown in Figure Ibuprofen, a nonsteroidal anti-inflammatory drug (NSAID) is among the very commonly used medications with over 21.3 million prescriptions in 2016 , and theThe most prominent abnormalities seen in our patients are hypokalemia, metabolic acidosis likely secondary to proximal RTA, with a negative urinary anion gap in patient 1, and distal RTA, with a positive urinary anion gap in patient 2, and alkaline urine pH in the setting of ibuprofen overuse. After ruling out other possible etiologies for RTA, a diagnosis of ibuprofen-induced RTA, leading to hypokalemia, was made in both the patients.The cases described here highlight the importance of inquiring about OTC medication history and considering ibuprofen as one of the differentials in patients with a combination of refractory hypokalemia and RTA. They also stress the need to further educate patients regarding the multiple nephrotoxic effects of excessive NSAID use, including RTAs, and emphasize the importance of limiting NSAID use.


Growth Differentiation Factor 11 (GDF11), a member of the super family of the Transforming Growth Factor β, has gained more attention in the last few years due to numerous reports regarding its functions in other systems, which are different to those related to differentiation and embryonic development, such as age-related muscle dysfunction, skin biology, metabolism, and cancer. GDF11 is expressed in many tissues, including skeletal muscle, pancreas, kidney, nervous system, and retina, among others. GDF11 circulating levels and protein content in tissues are quite variable and are affected by pathological conditions or age. Although, GDF11 biology had a lot of controversies, must of them are only misunderstandings regarding the variability of its responses, which are independent of the tissue, grade of cellular differentiation or pathologies. A blunt fact regarding GDF11 biology is that its target cells have stemness feature, a property that could be found in certain adult cells in health and in disease, such as cancer cells. This review is focused to present and analyze the recent findings in the emerging research field of GDF11 function in cancer and metabolism, and discusses the controversies surrounding the biology of this atypical growth factor. On May 2013 the research groups, led by doctors Amy J. Wagers and Richard T. Lee, published outstanding work suggesting that the growth differentiation factor 11 (GDF11) could be a good candidate for the age-related heart hypertrophy reversion observed in the model of heterochronic parabiosis . At thatThe findings, beyond this unfortunate motto, revealed that GDF11 could exert functions in adult systems, in addition of those characterized in embryonic and fetal tissues. The works by the groups of doctors Wagers and Lee provided evidence that the main target cells are those with certain stemness phenotype, such as the satellite cells in the muscle, which are the progenitor ones for new functional muscle cells.If GDF11 targets cells with stemness capacity, then many cancer cells should be targeted by this growth factor.Many cancer cells gain stemness capacity and this correlates with aggressiveness and poor prognosis. The findings raised by the group of doctors, Wagers and Lee, position cancer cells as a target of GDF11 since they proved that stemness is a key condition for GDF11 effect. However, the results could be opposite depending of the cancer cell origin, metabolic status, or the stage of the cancer. We must wait for incoming works in the next few years, perhaps months, revealing a more precise mechanism regarding these apparent controversies in cancer and metabolism.This work is focused to review the general knowledge of GDF11, and its functions in cancer biology and metabolism, taking into consideration recent findings in the specialized literature and in the public databases and scientific on-line resources.GDF11 , is a member of the super family of the Transforming Growth Factor beta (TGF-β) and a subfamily of the BMP which is widely secreted in many species, including mouse, rat and human, and it is accepted as a key factor in embryo development, particularly in the anterior/posterior patterning –9.GDF11 was identified by McPherron et al. in 1999, who cloned the human and mouse GDF11 and characterized its function in pattering the axial skeleton .ENSG00000135414). Two splice variants products have been identified, according to Ensembl  is one of the main acting proteins on GDF11, activating the mature GDF11 by proteolytic process at basic sites of the pro-domain . The eliIn humans, GDF11 is expressed in practically all tissues, but is particularly relevant in the brain (hippocampus), the kidneys, the endometrium, and the heart muscle; while the liver is the organ with the lowest expression , 18.As a member of the BMP family, GDF11 uses the canonical receptors and the SMAD proteins for signaling. The GDF11 dimer  binds the activin receptors type II A or B , proteins with serine/threonine kinase activity; leading to the recruitment and transphosphorylation of two type I serine/threonine kinase receptors, also known as activin-like kinase receptors (ALK), particularly the 4, 5, or 7 , 20. TheGDF11 can also transduce by non-canonical pathways. Mitogen activated protein kinase (MAPK) is perhaps the main non-SMAD pathway controlled by the growth factor, activating routes such as p38, AKT, and JNK , 26, howNegative regulation of the GDF11-mediated signaling can also occur at different levels. Extracellularly, GDF11 can be negatively regulated by the interaction with many proteins such as follistatin , 34, GDFThe BMP and activin membrane-bound inhibitor (BAMBI), a co-receptor that is not functional due to it lacks cytosolic domain, has been suggested to be another negative regulator in plasma membrane, but that still remains to be confirmed .In the cytosol, GDF11 follows the canonical negative regulation of the family. It has been reported that GDF11 is regulated by SMAD7  and SMADSancta sanctorum” of the “miraculous” molecule, when the laboratory of Amy Wagers , decreasing proliferation, motogenesis, and invasion. These characteristics were associated with transcriptional repression of cyclin D1 and A, and the overexpression of p27 . GDF11 eRemarkably, the invasion experiments using the chick embryo chorioallantoic membrane (CAM) model , 59 reveSimilar results were obtained in the hepatoblastoma cell lines, HepG2 and SMMC-7721: the treatment with GDF11 up to 72 h reduced cell viability. Although SMMC-7721 cells are probably a HELA-derivative cell line, the effect was also present . This reEpcam, promo1 (CD133), cd24, and ck19, that was associated with the repression of Snail and N-cadherin, and the overexpression of occluding and E-cadherin, strongly indicating a mesenchymal to epithelial transition  represents one of the most lethal cancers worldwide . It has https://www.proteinatlas.org) provides evidence from 176 patients: those with high GDF11 expression (n = 61) exhibited better survival rates, compared with those with low expression   . These o< 0.001) .Interestingly, another member of the family GDF15 is directly correlated with poor survival in PC patients, and it is proposed as a better marker than CA-125 , again r−/− animals the pancreas size are 2-fold smaller than wild type.As observed in HCC, in PC, the targets of GDF11 are poorly differentiated cells. In the mouse embryo, GDF11 is expressed in the pancreatic epithelium, at embryonic day E12-E14 , as it h+ progenitor cells and GDF11 induces β-cell differentiation , the expression of GDF11 was significantly higher compared with normal tissue . The claIt is known that lymphangiogenesis is a fundamental phenomenon for colorectal cancer dissemination . RecentlSome reports have pointed to the pro-tumorigenic properties of GDF11, with major or minor confidence of rigorous scientific approach.n = 19) in comparison with non-metastatic oral cancer (n = 15). Authors also sustain that GDF11 induced epithelial to mesenchymal transition by downregulating epithelial markers such as E-cadherin, and the overexpression of vimentin or metalloproteinase 9.In oral squamous cell carcinoma, Qin and coauthors  showed tIn uveal melanoma, GDF11 expression was significantly upregulated compared with surrounding tissue, the expression was higher in stage IV and substantially greater in the deceased cases regarding living cases . The mulhttps://www.cbioportal.org) indicates that GDF11 is altered in 1% of database patients. Gdf11 gene in some cancers, and The study by Liu et al.  also shoIt is particularly relevant to consider the convertase PCSK5, a key regulator of GDF11 activity. Pcsk5 gene presents a high frequency of genomic alterations in 3% of the patients, according to cBioportal, being particularly relevant in melanoma, endometrial carcinoma, and stomach adenocarcinoma, among others . Missens−/− mouse, the maturation and number of β- and α-cells are normal, however, another group led by Harmon reported that the gdf11−/− mouse exhibited impairment maturation of β-cells and an increment in α-cells, which could produce glucagon in comparison with the wild type mouse (The impact of GDF11 in the development of pancreas implies that the growth factor could exert some metabolic regulation on this organ in the adult, particularly in the endocrine pancreas . Dichmanpe mouse . This coRecently, a work by Anon-Hidalgo et al.  reportedPparg, Cebpa, and the executer proteins Plp, Cd36, Plin1, Adipoq, among others, in a mechanism associated to the canonical signal transduction mediated by SMAD2/3 (Luo et al. published that GDF11 decreased lipid content in human mesenchymal stem cells and the mouse 3T3-L1 cell line. This was associated with the repression of adipogenic genes, such as the transcription factors  SMAD2/3 . The rep SMAD2/3 . The mRNThe evidence sustains the metabolic intervention by GDF11, at least in terms of lipid homeostasis, and again in cells with stemness features. This could be relevant in cancer, since lipid overload is one of the main characteristics required for a proper cancer cell proliferation , 79. In Although the evidence of GDF11 regulation of the energetic and lipid metabolism is limited, it clearly indicates an effect tending to maintain the cellular energetic homeostasis. More research is required to characterize the mechanism underlying metabolic regulation by the growth factor, particularly in cancer cells.GDF11 is an intriguing non-conventional growth factor, perhaps the most fascinating new member of the TGF-β superfamily. It transduces, as practically all members, by the canonical SMAD and non-canonical MAPK pathways, but its functions can be quite variable, even contradictory, depending of the cell lineage, tissue , or evenPerhaps it is time that GDF11, due to its diverse functionality, constitutes its own subfamily as an atypical and versatile member of the TGF-β family.We must be cautious to oversimplify its functions. The controversies found clearly indicate that GDF11 displays particular activities depending of cell type, grade of differentiation, and pathological or normal conditions. This remarkable atypical member of the TGF-β family must be carefully studied in clear and well-controlled biological systems. The knowledge, regarding GDF11, will surely be increased in the next few years. The mechanism of action in each particular cancer or cell type must be elucidated to clarify these controversies, and perhaps they will stop being such, thanks to the mechanistic enlightenments obtained in the incoming research in the field.AS-N, MG-R, GP-V, and LC-R conception and preparation of the manuscript. LB, VS, and RM-L reviewed and corrected the manuscript. MCG-R and LG-Q final review of the manuscript and financial support.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The causes of birth defects (BDs) are complex and include genetic and environmental factors and/or their interactions. More research is needed to describe the epidemiology of BDs within specific regions of China. This study focused on differences in the prevalence of BDs based on ethnicity in a large city in Guangxi Province, China.Surveillance data of infants born in 114 registered hospitals in Liuzhou between 2011 and 2015 were analyzed to determine the epidemiology of BDs across five major ethnic groups. We calculated the prevalence of BDs and relative risk of BDs by ethnicity.There were 260,722 perinatal infants of which 6581 had BDs, with the average prevalence of 25.24 per 1000 perinatal infants (PIs). Prevalence data showed an obvious uptrend over the past 5 years. Han had the highest prevalence of total BDs (28.98‰), followed by Zhuang (25.19‰), Yao (18.50‰), Miao (15.78‰) and Dong (14.24‰). Relative to the Han; Zhuang, Miao, Yao, and Dong had a lower risk of musculoskeletal and urogenital malformations; Miao and Yao had a lower risk of cardiovascular malformation; and Dong had a lower risk of cardiovascular and craniofacial malformation. Several maternal risk factors were found to be associated with BDs .This study provided a comprehensive description of ethnic differences in the risk of BDs in Liuzhou City, China. Observed ethnic differences in the risk of BDs may be related to genetic susceptibilities, environment, cultural customs, or to potential combinations of these factors. The World Health Organization estimates that approximately 260,000 deaths  globally were caused by birth defects (BD) s in 2004 . It is eThe causes of BDs are complex and multifactorial. Nearly 50% of BDs cannot be ascribed to a specific cause , 9. In 1Guangxi is one of the western provinces of China, and the Chinese minorities account for majority of the population in Guangxi province (37.2%) . LiuzhouThe diagnosis of BD was based on the “International Statistical Classification of Diseases and Related Health Problems, Tenth Edition” (ICD-10) and Chinese National Criteria of BDs . BDs werAll surveillance data of BDs were collected from the obstetrics departments or neonatal departments according to the “National Office for Maternal and Child Health Surveillance” formulated by the National Health and Family Planning Commission. The surveying population was perinatal infants  born in the 114 registered hospitals of Liuzhou from 2011 to 2015. In total, infants were followed six times in the first year of life. The schedule for follow-up monitoring was as follows: Within 7 days for the first time-point, and then at 6 weeks, 3 months, 6 months, 8 months and 12 months old. Infant BDs were reported to the monitoring system when diagnosed within 7 days of life.Each delivery that was associated with a BD was reported using a registration card for BDs submitted by physicians in obstetrics and gynecology, pediatric or neonatal medicine through an online hospital-based survey, required by the Chinese government. Each case report card recorded basic maternal information, , Hepatitis B Virus (HBV), and syphilis infections), birth information , diagnoses of specific BDs, symptoms, medication use during early pregnancy, and family history. In addition to the case report card, a quarterly table for each registered hospital was completed by professional physicians. Each quarterly table contained 3 months of data such as the number of perinatal births, maternal age, residence, ethnicity, occupation, pregnancy history, gestational age of birth, gestational age at first antenatal care attendance, number of antenatal visits, infant gender, number of BDs, and maternal illness.Both case report cards and quarterly tables were reviewed and audited by maternal and child health hospitals and health administrative departments. Periodic quality control measures were in place at the monitored hospitals and occurred quarterly at the county-level and bi-annually at the city-level or province-level to assure reporting accuracy.P < 0.05 in the bivariate analyses were included in the model. Adjusted odds ratios (ORs) and their 95% confidence intervals (CIs) were calculated. All statistical analyses were performed using the SAS 9.4 statistical software package . A significance level of 0.01 was adopted in bivariate comparisons and multivariate analysis due to the large sample size.The prevalence rates and 95% confidence intervals of overall BDs were calculated for the whole sample and stratified by the five major ethnic groups across all years (2011–2015) of the study. A line chart was then constructed to graphically display the longitudinal trends of the prevalence of BDs by the different ethnic groups. The prevalence rates of the top fifteen BDs were also calculated and compared among the six major ethnic groups using chi-square tests. Multiple logistic regression analysis was performed to further examine the association of ethnicity with BDs, controlling for potential confounders including maternal age, gestational age, number of antenatal care visits, gestational age at first antenatal care visit, number of previous abortion, and syphilis infection. The dependent variable of logistic regression analysis was whether the infant had a birth defect (no/yes). Only variables identified as significantly associated with the dependent variable at A total of 260,722 perinatal infants were monitored from 2011 to 2015. Of the perinatal infants, the top 5 ethnic groups were represented as 44.7% were Han, 36.6% were Zhuang, 7.5% were Miao, 6.9% were Dong, and 2.4% were Yao . The total prevalence rate increased by 1.49% annually from 2011 to 2015 on average.Of the 260,722 perinatal infants, 6581 had BDs, with an average prevalence of 25.2 (95% CI: 24.6–25.9) per 1000 PIs  , followed by Zhuang (25.2 per 1000), Yao (18.5 per 1000), Miao (15.8 per 1000) and Dong (14.2 per 1000) Table . CompareFrom 2011 to 2015, 89 classes (or combinations of classes) of BDs in total were found in our sample, and the top fifteen classes of BDs were musculoskeletal malformation, cardiovascular, craniofacial, urogenital system, other BDs, neurologic, gastrointestinal system, genetic syndrome, respiratory system, musculoskeletal with cardiovascular systems, cardiovascular with other BDs, cardiovascular with urogenital systems, cardiovascular with craniofacial systems, musculoskeletal with other BDs, and craniofacial with musculoskeletal systems. Relative to the Han, Zhuang had significantly lower risks in musculoskeletal and urogenital system malformation ; Miao and Yao had significantly lower risks in musculoskeletal system , cardiovascular system , urogenital malformation  and other BDs ; Dong had statistical significant lower risks in the musculoskeletal system , cardiovascular system , craniofacial system , urogenital malformation  and other BDs   had lower risk of overall BDs. Mother’s age (> 35 years), gestational age (< 36 weeks), a higher number of antenatal care visits (12 or more), and a higher number of reported abortions (8 or more) were significantly positively associated with overall BDs Table .Table 7OOur study performed a comprehensive analysis of ethnic differences in the perinatal infant prevalence of BDs in Liuzhou, Guangxi, China over a 5 year time period. The average prevalence rate of total perinatal infants’ BDs was 25.2 per 1000 PIs in the past 5 years, with an upward trend increasing by 1.49% on average each year. The ethnic distribution in our study sample was consistent with the ethnic distribution in Liuzhou, Guangxi as reported in the 2012 Chinese population census , 23.The average prevalence rate (25.2 per 1000 PIs) was higher than 20.2/1000, which was reported in a prior analysis of BDs in perinatal children in Guangxi from 2001 to 2010 . The upwP < 0.001) [Our average prevalence rate was the highest among similar studies in various regions of China, such as Hunan  from 2005 to 2014; Inner Mongolia  from 2005 to 2008; and overall China  in 2009 , 25, 29.< 0.001) . In summThe causes of BDs are complex, including a variety of risk factors such as: advanced maternal age, exposures during the pregnancy, geographical location exposures, and race and ethnicity , 8, 33. Most previous Chinese studies showed that prevalence rates of overall BDs were different between rural and urban environments. For example, in the Inner Mongolia study, in the analysis of BDs in the Xinjiang multi-ethnic region, and in the government BDs Prevention Report in 2012, all concluded that a higher incidence of BDs in rural areas than urban areas , 29, 37.The top six BD categories covered most BDs (86.8%) in our study. In a study of BDs in Guangxi from 2001 to 2010, the top six BDs were Schridde syndrome, polydactylism, congenital heart disease (CDH), total cleft lip, ear anomaly and hypospadias. This contrasted to the top six in our study which included musculoskeletal BDs as number one, and then cardiovascular, craniofacial, urogenital, other BDs, and neurologic BDs .Thalassemia gene carriers in Guangxi comprise more than 20% of the population , which wThe prevalence of CDH was increasing in our report, which is consistent with outcomes of other current studies from within China; and this would likely be due to the use of fetal ultrasound in antenatal care examinations and the improved surveying system online in China supported by the national finances in Maternal and Child Health Care System , 28, 40.Our study found ethnic differences in the overall risk of BDs and types of BDs. To our knowledge, this is the first report of ethnic differences in the epidemiology of BDs in a hospital-based surveying system with internal quality controls in Guangxi, China. Our study provided a comprehensive description of ethnic differences in the risk of BDs in the most representative city  in Guangxi.


R = 0.85, p < 0.05). The final SEM also validated our hypothesis that the AI explained 79.3% and 84.4% of the biomass variations in the alpine steppe and the meadow, respectively. Furthermore, we found that the soils with higher carbon to nitrogen ratio and soil total nitrogen had larger biomass, whereas soil organic carbon had a negative effect on biomass in alpine steppe; however, opposite effects of soil factors on biomass were observed in an alpine meadow. The findings demonstrated that the AI was the most critical factor affecting biomass in the alpine grasslands, and different reaction mechanisms of biomass response to the AI existed in the alpine steppe and alpine meadow.The vegetation dynamic  is an important index used to evaluate the ecosystem function of grassland ecosystem. However, the critical factors that affect vegetation biomass are disputed continuously, and most of the debates focus on mean annual precipitation (MAP) or temperature (MAT). This article integrated these two factors, used the aridity index (AI) to describe the dynamics of MAP and MAT, and tested the hypothesis that vegetation traits are influenced primarily by the AI. We sampled 275 plots at 55 sites  across an alpine grassland of the northern Tibet Plateau, used correlation analysis and redundancy analysis (RDA) to explore which key factors determine the biomass dynamic, and explained the mechanism by which they affect the vegetation biomass in different vegetation types via structural equation modelling (SEM). The results supported our hypothesis, in all of the environmental factors collected, the AI made the greatest contribution to biomass variations in RDA , and the correlation between the AI and biomass was the largest ( The typurpurea) . Alpine urpurea) .2 area). Plant BIO in these quadrats was harvested by a clipping method flush with the ground. All species information, including the vegetation coverage within each quadrat, was identified by a field guide  along a transect at spatial intervals of 50 km in August 2012 , Table 1ld guide , and theld guide . After tin situ into one composite sample. The soil organic carbon (SOC) and soil total nitrogen (STN) concentrations were determined using a vario MACRO cube elemental analyser (http://www.worldclim.org/), and the outliers of the meteorological data acquired has been modified or removed. The database mentioned includes information on current global climate based on data collected from 1950 to 2000 and interpolated to a resolution grid of 2.5 arc min  using information on latitude, longitude, and elevation \docThe data package required in this study includes the target variable of BIO (BIO), and biotic factors  and abiotic factors  were selected to explore the response of BIO to these environment factors. First, we conducted a Pearson’s correlation analysis (Corrplot package) using R software 2.11.1  to screeTo demonstrate the reliability of our conclusions, we only used the AI and remaining independent variables collected to explore the mechanism of biomass variation. The structural equation modelling (SEM) approach was adopted to explain the mechanism by which the key factors affect BIO, both directly and indirectly, in alpine steppe and meadow. SEM is a common multivariate technique that has been used in recent studies to evaluate explicitly the causal relations among multiple interacting variables . SEM difR =  − 0.76; p < 0.05), MAP , C: N , and the AI  in alpine grasslands. In addition, these environmental factors above were also closely related to each other, example for AI, it was significantly correlated with MAP  and MAT .According to The results showed that the interactions among MAP, MAT, C: N, and the AI were the main drivers of the response variable , explain−2 in steppe, with a mean of 32.4 gm−2, and 3.18 to 149.30 g m−2 in meadow, with a mean of 59.4 g m−2  was the primary limiting environmental factor for vegetation traits in alpine grassland. Additionally, the different AI govern the different response patterns of BIO in the alpine steppe and the alpine meadow, respectively. The findings imply the important role of AI regulating vegetation dynamics, which need to be explored deeply in future research. Nevertheless, due to the unevenly distribution of the sampling sites throughout the vast region of northern Tibet plateau, the spatial heterogeneity of vegetation across regional environmental gradients limited this research. Hence, we should pay more attention to reduce spatial heterogeneity  to verify the effect of AI on variations of biomass.10.7717/peerj.7272/supp-1Supplemental Information 1the MAT, MAP, AI, SOC, C: N, STN, SR, BIO and COV represent mean annual temperature, mean annual precipitation, aridity index, soil organic carbon, the soil ratio of carbon to nitrogen, soil total nitrogen, species richness, aboveground biomass and coverage, respectively.Click here for additional data file.


Aims: To determine the incidence, risk factors, etiology, and associations of hyponatremia in community-dwelling elderly with geriatric morbidity and mortality. Materials: Elderly participants of a single center home-based primary care program were included. Method: Retrospective chart review was conducted on demographic and clinical variables, comorbid diseases, frailty by Fried criteria and biochemical tests over a 1-year period. Primary outcome measure was a composite of falls, fractures due to falls, and hospitalization witnessed within the first year of enrollment into the program. Secondary outcome was all-cause mortality. Results: The study population (n = 608) had a mean age of 84.3 ± 9.3 years and was largely female (77.1%) and African-American (89.5%). Mean follow-up was 41.5 months. Frailty was seen in 44.4%. Incidence of all-cause mortality was 26.9%. Initial hyponatremia occurred in 8.71% (n = 53), and persistent hyponatremia (> 6 months of low serum sodium) in 4.1% (n = 25) of the study population. The major causes of hyponatremia included multiple potential causes, idiopathic syndrome of inappropriate anti-diuretic hormone (SIADH) and medications (thiazides and selective serotonin reuptake inhibitor (SSRI)). Primary outcome was independently associated with frailty (Odds ratio (OR) of 2.33) and persistent but not initial hyponatremia (OR 3.52). Secondary outcome was independently associated with age > 75 years (OR 2.88) and Afro-American race (OR 2.09) only but not to frailty or hyponatremia. Conclusions: Hyponatremia is common in home-bound elderly patients and its persistence independently contributes to falls, fractures, and hospitalization but not mortality. Our study highlights a new association of hyponatremia with frailty and underscores the need to study time-dependent association of hyponatremia with epidemiological outcomes.  Hyponatremia is the most common electrolyte disorder in clinical practice , with hiTo our knowledge, the incidence and risk factors for hyponatremia in the frail elderly population has not been studied. The implication of such research is profound as this population is unique not only because of the burden of comorbid illness but also the high prevalence of disability, limited functional Status and frailty independently contributing to falls, injuries, disability, institutionalization, mood disorders, and other adverse health outcomes . Given tst January 2000 to 31st December 2012. The HBPC program comprises an interdisciplinary team including physicians, nurse practitioners, social workers, and office staff. Patients are seen as medically necessary at home, and urgent visits are made within 24 hours. The program sees patients within a 5 mile radius of the sponsor hospital, WHC. Follow up visits vary from once a month to once every 4 months, depending on the acuity of care. The present study was approved by the institutional ethics committee and was in accordance with the declaration of Helsinki. All the provisions of Health Insurance Portability and Accountability Act (HIPAA) act were satisfied to meet patient confidentiality. A retrospective chart review was conducted using the electronic medical records (EMR) of patients enrolled in the house call program (HBPC) at Medstar Washington Hospital Center (WHC), a community hospital and referral center in Washington, DC from 1st 2012. Primary outcome was defined as a composite of falls, fractures , and hospitalization (related or unrelated to hyponatremia) recorded during the first year of enrollment into the house call program. Secondary outcome was all-cause mortality. We hypothesized that baseline or persistent hyponatremia was a predictor of both primary and secondary outcomes. Inclusion criteria included patients with age of more than 65 years enrolled in the house call program with complete medical records for at least 1 year. Exclusion criteria included patients whose follow-up data was less than 1 year. Two independent reviewers analyzed charts, and data was cross-checked to maintain internal consistency of results. We reviewed the records from the time of the first physician note in the EMR until 1 year after and included demographics, past medical problems, medications, hospitalizations, incidence of fall, fractures, or injuries related to fall, and terminal events. Race/ethnicity was as per documented in the EMR and was recorded in view of prior studies showing implications on the etiology of hyponatremia , 9. MortHyponatremia was defined as serum sodium < 136 mEq/L on 2 or more occasions (corrected for hyperglycemia using a correction factor of 1.6 mEq/L decrease in serum sodium concentration for every 100 mg/dL increase in plasma glucose concentration). Initial hyponatremia was defined on the basis of the first recorded Na after enrollment into the program, but not persisting for more than 6 months after the initial measurement. Persistent hyponatremia was defined as an Na < 136 mEq/L for more than 6 consecutive months during the study year. Mild hyponatremia was defined as serum sodium between 130 and 135 mEq/L, moderate hyponatremia as serum sodium between 125 and 129 mEq/L, and severe hyponatremia as serum sodium less than 125 mEq/L. Functional limitation was assessed by frailty using the Fried Index, which was present if the patient had at least 3 of the following : documen1 each: Myocardial infarct, congestive heart failure, peripheral vascular disease, dementia, cerebrovascular disease, chronic lung disease, connective tissue disease, ulcer, chronic liver disease, diabetes. 2 each: Hemiplegia, moderate or severe kidney disease, diabetes with end organ damage, tumor, leukemia, lymphoma. 3 each: Moderate or severe liver disease. 6 each: Malignant tumor, metastasis, AIDS. Chronic kidney disease was defined as persistent estimated glomerular filtration rate (GFR) of < 60 mL/min over a 3-month period as measured by the modification of diet in renal disease (MDRD) equation. 2-tests were used to investigate differences in categorical clinical variables between hyponatremic and normonatremic groups. Logistic regression was used to determine independent predictors for primary and secondary outcomes. p-value of < 0.05 was considered significant. Statistical Analysis System software version 9.2  was used to perform the analysis. Continuous clinical variables were analyzed using the nonparametric Wilcoxon rank sum tests since normality assumption was not satisfied. χOut of a total of 958 patient charts, 608 patients were found eligible for analysis. Demographic variables are shown in The etiology of hyponatremia was divided between three categories: euvolemic, hypervolemic, and hypovolemic on the basis of documentation of volume status. Of the 53 patients who had initial hyponatremia, 37 (69.8%) were euvolemic, 5 (9.4%) were hypervolemic, and 11 (20.7%) were hypovolemic A. Of theIn euvolemic initial hyponatremic patients (n = 37), the predominant etiology identified was multiple potential causes (n = 11), thiazides (n = 8) followed by selective serotonin reuptake inhibitor (SSRI) (n = 6), idiopathic SIADH (n = 5), unknown or unclear (n = 4) malignancy (n = 2), and psychogenic polydipsia (n = 1) B. In euvWhen clinical and demographic variables were analyzed on the basis of initial hyponatremia versus normonatremia  Figure A, signifThe composite primary outcome within 1 year of observation was seen in 56.08% of the total population and on bivariate analysis A; signifIncidence of all-cause mortality was 26.9%. This included patients opting for palliative and hospice care as well as those receiving acute care in the hospital. When a subset analysis was done for overall mortality B, factorHyponatremia is the most common electrolyte disturbance in the elderly population . Point pWhile symptomatic hyponatremia is easily detected and treated, prognosis, therapeutic challenges, and implications of mild chronic hyponatremia have recently been highlighted. Most notable amongst the poor outcomes associated with this condition is the propensity to falls in the setting of neurocognitive dysfunction causing gait ataxia and poor attention span . This haOur study explored the epidemiology of hyponatremia in a vulnerable elderly home-bound population. The average age of the participants in the population was 84.3 years, which is much higher than previous studies , 20, 28.One of the distinct findings of the study was the variation in the etiology of hyponatremia with respect to duration. While physician-determined euvolemic hyponatremia was the most common cause , hypovolemic hyponatremia tends to decrease with time , while the incidence of hypervolemic hyponatremia increases with time . This is partly due to the easy recognition of overt signs and etiology of hypovolemia and its treatment. Not surprisingly, studies in acute settings have reported dehydration (presumably hypovolemia) as the most common cause . We founAge-related increase in the incidence of SIADH is a distinct phenomenon that has been previously reported , 18, 37.One of the novel findings of the current study is the association of frailty with hyponatremia in the elderly and its emergence as an independent risk factor for primary outcomes in addition to persistent hyponatremia. This is not surprising given the known association of frailty with falls, disability, osteoporosis, admission to hospital, postoperative complications, length of stay in hospital, and even death , 42. FraThere is a paucity of literature on the mechanisms of hyponatremia in frailty. Studies on the hypothalamic/-pituitary adrenal axis in frail elderly patients have shown suboptimal response to ACTH stimulation, suggesting unrecognized adrenal insufficiency as a possible cause of functional limitation and hyponatremia . AnotherPredicting all-cause mortality in an outpatient elderly population is difficult, and a recent systematic-analysis has revealed that most prognostic indices are either biased or weakly generalizable . ContrarSome of the strengths of the current study include an attempt to use hyponatremia as a time-dependent and as a baseline variable to investigate its role in contributing to geriatric morbidity. Most studies showing associations have been largely case-control studies , 7, 35, Some weaknesses of the current study include a small sample size, short follow-up period, and the fact that data were derived from a single center. Also, the retrospective nature of the data has inherent risks of being subject to selection and information bias. We tried to limit information bias with a detailed protocol for data collection and simultaneous data collection by two independent workers to ensure reproducibility of data. The data on etiological assessment of hyponatremia are also limited by the lack of a uniform protocol for renal consults or laboratory assessment. This is especially significant as volume assessment in hyponatremia by physical examination has poor sensitivity and specificity . As a coDespite all its shortcomings, we feel that our study is significant in shedding light on the epidemiology of hyponatremia in a very elderly population where the burden of disabling comorbid conditions is substantial yet patients are not at risk of imminent death. Given the significant association between frailty and hyponatremia, we believe we have identified a potential confounder in epidemiological studies on hyponatremia which may also serve as the elusive etiologic link between hyponatremia and its poor prognostic implications in the geriatric population. The current study also underscores the need to use serum sodium as a time-dependent variable to further investigate the issue of causality. The authors wish to thank the biostatistical team from the Medstar Research Institute, Washington, DC for their help in data analysis. The authors also thank the Office of the Graduate Medical Education, Georgetown University/Washington Hospital Center for funding the bio statistical analysis of the study. Furthermore, the authors would like to state that they have no financial interests, affiliation, or relationship attached to the conduct, analysis, or publication of the current research work.


Angelica sinensis (Oliv.) Diels is a widely used medicinal plant mainly originated in Gansu, China. Angelica sinensis is greatly demanded in the clinical practice of Chinese medicine due to its broad pharmacological activities of hematopoietic and anti-inflammatory properties. But, the percentage of early flowering in Angelica sinensis arrives to 20%~30%, which severely affects its quality and quantity. Here, transcriptome profiling and digital gene expression analysis were applied to study the mechanism of early flowering in Angelica sinensis. A total of 49,183,534 clean reads were obtained and assembled into 68,262 unigenes, and 49,477 unigenes (72.5%) could be annotated to a minimum of one database in the Nr, Nt, Swiss-Pro, GO, COG and KEGG. Taking the above transcriptome data as a reference, digital gene expression result showed that 5,094 genes expression level were significant changed during early flowering. These annotated genes offered much information promoting that the biosynthesis of secondary metabolites pathway, the hormone signal transduction pathway, and the transcription regulation system may be closely related to the early flowering phenomenon of Angelica sinensis. Further expression patterns of key genes contribute to early flowering were analyzed using quantitative real-time PCR. The transcriptome result offered important gene expression information about early flowering in Angelica sinensis. Angelica sinensis (Oliv.) Diels (AS) is a world-renowned plant medicine originating in Gansu, China. The root of AS is traditional Chinese medicinal materials, which was firstly recorded in “Shen Nong Ben Cao Jing” in the Han Dynasty. AS is suggested as a tonic, hematopoietic, antitumor2, and anti-inflammatory4 properties for the treatment of menstrual disorders, amenorrhea, dysmenorrhea. It is also considered as a healthy food material in Asia, Europe and America5. Thus, AS is great required in the world due to its widely used.AS reaches to 20%~30%, once the early flowering occurs, the roots of AS will be lignified and cannot be used as medicine, which severely affects its quality and quantity. Previous researches were focused on the function of ecological and nutritional factors in affecting early flowering of AS. However, the molecular mechanism of early flowering of AS is still elusive. To date, there is no effective methods to control and prevent the early flowering of AS. The molecular mechanism of early flowering in AS needs to be further studied.However, the early flowering rate of 6. Digital gene expression (DGE) analysis was also considered as a valuable tool to quantitative comparison gene expression, which can efficiently screen and comment differentially expressed genes7. In non-model plants, combine these two methods can promote the identification of different expression genes. The combination of transcriptome sequencing and DGE can provide more sensitive and efficient analysis of gene expression changes, and can also promote gene expression comparisons for the sample without reference databases8.The transcriptome assembly provided a powerful tool for high-throughput research at the genetic levelAS. DGE libraries from apical meristem of vegetative growth AS and flower buds of early flowering AS were built. By comparing changes in gene expression between these different groups, we can understand more deeply in the molecular mechanism of early flowering in AS.Here, Illumina sequencing technology and DGE system were performed to study the gene expression changes in early flowering of AS is a triennial medicinal plant, which flowers in the third year. Early flowering is frequent in May of the second year. The flower buds of early flowering AS and apical meristem of vegetative growth AS were collected in June of the second year in the field of Minxian (located at 34°29′ North latitude and 103°57′ East longitude), Gansu Province, China. The samples were stored at −80 °C. The medicinal plant was identified by Hui Yan, associate professor of Pharmacognosy, Nanjing University of Chinese Medicine.Total RNA from the plant materials was extracted and identified through a TRIzol/chloroform  referenced to the manufacture’s protocols.AS and apical meristem of vegetative growth AS for transcriptome analysis. If the RIN ≥8 and a 260/280 nm absorption ratio ≥ 1.8, RNA was used to set up transcriptome library. After the RNA extraction, mRNA was purified from total RNA by binding the RNA to magnetic beads. Then, mRNA was broken into short fragments. The cleaved RNA fragments were used as templates to synthesize the first-strand cDNA, after that DNA polymerase I and RNase H were added to synthesize the second-strand cDNA.The RNA was extracted from the flower buds of early flowering Next, suitable fragments were used as templates for PCR amplification, which yielded as the cDNA library for sequencing.De novo assembling of the clean reads by using Trinity method10. In order to understand the function of the unigenes, they were searched against the public databases, including NCBI Nr and Nt, Swiss-Prot, GO, COG, and KEGG database, with E value ≤ 10−5.The clean reads were screened from the raw data by filtering out poly-N, the low-quality reads . Then, the unigenes were generated by AS Samples were performed in parallel using an Illumina Gene Expression Sample Preparation Kit  and ZT2: late stage flower bud of early flowering AS ). Each experimental group consists of three biological sample replicates .DGE library preparation of the three groups of AS and flower bud of early flowering AS were mapped with the transcriptome library above. Reads per kilobase of per million mapped reads (RPKM) was used to measure the gene expression level. If the genes are satisfied with two conditions, false discovery rate (FDR) ≤ 0.001 and an absolute value of log2Ratio ≥ 1, they were defined for significant expression differences. The different expression genes (DEGs) were then compared with the transcriptome library of AS above.The clean reads from the sample of apical meristem of vegetative growth −△△CT method. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as the endogenous control.In order to verify the reliability of the DGE results, qRT-PCR was applied using LightCycler 480 SYBR Green I Master Mix  and a LightCycler 480 II Real-Time PCR instrument . Briefly, 1 μL of cDNA template from different group was used for reaction. The result of each gene repeated at least 3 times. The candidate genes expression changes were analyzed using 2AS has not yet been carried out, it is necessary to complete the sequencing of transcriptome of AS to provide a reference for screening differential expression of genes during early flowering. After filtering the adaptors and low-quality sequences, there are 49,183,534 clean reads , followed by Ricinus communis (12.9%), Populus trichocarpa (12.2%)  Fig. .Table 3SUsing WEGO software program, 326,207 unigenes were categorized in three categories: biological process, cellular component, and molecular function  between samples were identified using an algorithm. The matching percentage of clean reads and reference genes ranged from 87.47% and 88.90% in three DGE libaary. Among all reads, 67.50–69.74% per library was uniquely mapped to the reference genome, and 78.62–80.20% of reads was a perfect match to the reference gene  in the early flowering AS, the expression of the two genes (PHYB and ELF4) decreased, and the expression of the five genes  remained unchanged. In vernalization pathway, we detected an increase in the expression of four genes  in the early flowering AS, the expression of the SOC1 decreased. In gibberellin pathway, we detected an increase in the expression of two genes  in the early flowering AS, and the expression of the gibberellin 20-oxidase remained unchanged. In autonomous pathway, all 4 key genes  were no significant difference.In the study of the flowering mechanism of plant, most of the key genes are involved in photoperiodic pathway, vernalization pathway, autonomous pathway, gibberellin pathway. In photoperiodic pathway, we detected an increase in the expression of four genes  in ZC, ZT1 and ZT2 were shown in Fig. −ΔΔCt method was applied to calculate the relative expression of the genes. The DGE sequencing data was measured by the log2 value of samples. DGE sequencing and qRT-PCR showed significantly positive correlation (R2 = 0.951) in linear regression analysis  unigenes were matched with public databases. Our results will contribute to future genomic studies on AS and other Umbelliferae species. However, there were still nearly one third of the unigenes cannot be matched in public databases. Similar phenomena existed in transcriptome assemble of other plant, such as Lycoris aurea11 and Tagetes erecta12. The reason may be that the gene expression information of Umbelliferae is too little and the uniqueness of the gene expression of Umbelliferae. DGE was often used in combination with RNA sequencing to screen for differences in gene expression in different tissue of plant or to study disease mechanisms. Thus, a DGE analysis of apical meristem of vegetative growth AS and flower buds of early flowering AS was carried out to preliminarily clarify the mechanism of early flowering. According to the DGE results, a total of 5,094 and 4,556 transcripts were differently expressed between ZT1 and ZC, as well as ZT2 and ZC.Now, lack of genomic and transcriptome data limited the research of the mechanism of early flowering of AS. By contrast, all key genes in autonomous pathway are not changed. There are similarities in the gene expression of early flowering in AS and normal flowering in model plant, but at the same time there are still some differences in gene expression. These different genes are the focus of our future research work.In Arabidopsis, there are four classic pathways which controlled the flower time. In our study, some key genes in photoperiodic pathway, vernalization pathway and gibberellin pathway are up-regulated in early flowering 15. Our results were agreed with previous findings. In cryptochromes genes, CRY1 and CRY2 were both reported can promote flowering, our results were much the same between other groups16. GI and CO are regulated circadian clock, CO was considered as a gene that accelerates flowering in response to long days. FT is the target gene of CO, which is restricted to a similar time of day as expression of CO. FT was considered as one of the three integrons which can promote flowering17. CO, FT and GI were all found high expressed in early flowering AS. Photoperiodic pathway should be involved in the early flowering phenomenon of AS.In photoperiodic pathway, there were 4 genes  expressed higher in ZT. By contrast, there were only two genes (PHYB and ELF4) expressed higher in ZC. There were 5 genes  no significant difference. In phytochromes genes, PHYA was reported can promote flowering. On contrast, PHY, PHYD, PHYE inhibit floweringAS. By contrast, there were only one genes (SOC1) expressed higher in ZC. SOC1 is a major floral pathway integrator, which encodes a MADS box transcription factor and is one of the key floral activators integrating multiple floral inductive pathways, namely, long-day, vernalization, autonomous, and gibberellin-dependent pathways18, but SOC1 expression is obviously decreased in our experiment. FLC, an upstream negative regulator of SOC1, is high expressed, although VRN1 and VIN3 which control vernalization-mediated FLC silencing are both high expressed19. This should the reason of SOC1 expression decreased.In vernalization pathway, there were 4 genes  expressed higher in early flowering 21. In the GA-biosynthetic pathway, GA 20-oxidases and gibberellin 3 beta-hydroxylase 2 are both key enzymes22. In our study, gibberellin 3 beta-hydroxylase 2 were expressed higher in early flowering AS and there were no significant difference in gibberellin 20-oxidase expression level. The LFY homologs play a major role in the initiation of flowering23. LFY was also considered as one of the three integrons which can promote flowering24, which was positive regulated by GA. In our study, LFY was higher expressed in early flowering AS. Gibberellin pathway should be involved in the early flowering phenomenon of AS.Gibberellins (GAs) are essential for the development of fertile flowers in many plants, and may also be required immediately after fertilization25, the MADS-box transcriptional regulator that inhibits the activity of genes required to switch the meristem from vegetative to floral development27. One of the many pathways that regulate FLC expression is the autonomous promotion pathway composed of FCA, FY, FLD, FPA, FVE, LD, and FLK28. In our experiment, all 4 key genes  were no significant difference. The proteins involved in autonomous pathway have no changes in early flowering in AS.A central player in the floral transition is the floral repressor FLC29, S-adenosylmethionine synthetase (SAMS), S-adenosylmethionine decarboxylase (SAMDC)30, Spermidine synthase (SPDS)31, polyamine oxidase (PAOs) are key enzymes in polyamine metabolism. ADC, SAMDC and SPDS expression are up-regulated in early flowering sample.In fact, in addition to the classic four pathways that regulate plant flowering, we have also discovered changes in the expression of other genes. Plant polyamines are also an important class of plant growth regulators. Arginine decarboxylase (ADC)AS was major effected by the genes involved in photoperiodic pathway and GA pathway. Vernalization pathway and autonomous pathway no significantly changes in early flowering. This also should be the difference between the early flowering and normal flowering. These results provide basic information for exploring the molecular mechanisms that influence the early flowering of AS.In conclusion, early flowering of AS is very limited. Here, we combined RNA-Seq and DGE to study the molecular mechanism of early flowering of AS. We got 49,183,534 clear reads and assembled into 68,262 unigenes, the average length of each unigene was 728 bp.Now, effective genetic information on AS. Based on DGE study, many important genes regulating early flowering of AS were discovered and further analyzed. In this paper, we proposed a putative network underlying an overview of known floral regulators present and differentially regulated during floral induction of AS (Fig. AS.The result of sequencing provided effective gene expression profile information for genomic research of  AS Fig. , which pDataset 1


Among the few supportive care measures available for the management of dysphagia, Photobiomodulation (PBM) therapy, defined as the therapeutic use of light, has shown significant promise. In this case report, effective management of chronic dysphagia post hormonal therapy in a breast cancer patient was made. Experts in the supportive care in cancer and PBM proposed and requested further investigations of the protocol used in this case report in the management of dysphagia. In this case report, the protocol of PBM was proposed by experts in supportive care in cancer. Functional outcome swallowing scale for staging oropharyngeal dysphagia was used to assess the effectiveness of the treatment in pre-operative, per and post-operative stage. This case reports that PBM is effective in the management of dysphagia, a side effect of hormonal therapy in a cancer patient. Swallowing is considered a complex biomechanical interaction of anatomy and physiology in which the precise coordination of over twenty-five pairs of muscles in the oral cavity, pharynx, larynx, and esophagus is required . This co2+ signaling, with ROS regulating Ca2+ signaling and Ca2+ signaling affecting mitochondrial activities and ROS production [Among the few supportive care measures available for the management of dysphagia, Photobiomodulation (PBM), that is the therapeutic use of light, has shown significant promise . Visibleoduction . Howeveroduction . Additiooduction . PBM impoduction . Currentoduction ,11,12 anoduction . Furtheroduction ,8,9. TheA 45-year-old female patient with a history of breast cancer, previously treated by left mastectomy 4.5 years ago, chemotherapy 2.5 years ago and radiotherapy 1.5 years ago, started third -line hormonal treatment with Everolimus (AFINITOR 10 mg) and Exemestane (AROMASINE 25 mg) in addition to Zoledronic acid (ZOMETA 4 mg/100 ml). After three months of hormonal therapy, the patient complained of chronic impairment in swallowing in the upper gastrointestinal tract, from the lips to the upper esophageal sphincter. The patient was diagnosed with chronic dysphagia by a certified speech-language therapist. For the assessment of the severity of dysphagia and the follow-up, a functional outcome swallowing scale (FOSS) for staging oropharyngeal dysphagia was used . The trei Assessment of DysphagiaThe functional outcome swallowing scale for staging oropharyngeal dysphagia, proposed by John. R Salassa in 1999, was used for the assessment of dysphagia before treatment (T0), at day two of treatment (T2), immediately after treatment (T5) and fifteen days after treatment (T15). The FOSS was at stage II before the treatment .ii Treatment Protocol of Dysphagia2 for 10 seconds at each point, in a continuous and non-contact mode, tip of 320 µm. Bilaterally four points on the soft palate, bilaterally four points on the oropharynx. Extra-oral application was made with an energy density of 3 J/cm2, 10 seconds of irradiation on each point, tip of 320 µm, in a continuous and non-contact mode. The irradiation was made on the lateral and ventral pharynx and larynx, midline neck and lateral neck anterior to sternocleidomastoid muscle , there was occasional cough, symptomatic, a little difficulty in swallowing and prolonged meal time with the sensation of food getting stuck in the throat or esophagus (between stage II and I). After the treatment (T5), the patient had complete physiological function and was asymptomatic. Therefore, at stage 0 of FOSS, which persisted after 15 days (T15). A significant reduction of dysphagia (from stage II to Stage 0) was noted after 5 sessions of PBM with the protocol used. Therefore, PBM therapy successfully treated the hormonal therapy induced dysphagia .Dysphagia may result in numerous important sequelae such as weight loss, aspiration, pneumonia and chronic bronchial inflammation . AspiratIt can be concluded that the therapeutic use of photobiomodulation, according to specific protocol, is considered a good approach for the treatment of dysphagia induced by hormonal therapy in a breast cancer patient.


ISL1 (ISL LIM homeobox 1) has emerged as the major susceptibility gene for classic bladder exstrophy (CBE), in a multifactorial disease model. Here, GWAS (Genome wide association studies) discovery and replication studies, as well as the re-sequencing of ISL1, identified sequence variants ) associated with CBE. Here, we aimed to determine the molecular and functional consequences of these sequence variants and estimate the dependence of ISL1 protein on other predicted candidates. We used: (i) computational analysis of conserved sequence motifs to perform an evolutionary conservation analysis, based on a Bayesian algorithm, and (ii) computational 3D structural modeling. Furthermore, we looked into long non-coding RNAs (lncRNAs) residing within the ISL1 region, aiming to predict their targets. Our analysis suggests that the ISL1 protein specific N-terminal LIM domain (which harbors the variant c.137C > G), limits its transcriptional ability, and might interfere with ISL1-estrogen receptor α interactions. In conclusion, our analysis provides further useful insights about the ISL1 gene, which is involved in the formation of the BEEC, and in the development of the urinary bladder.Genetic factors play a critical role in the development of human diseases. Recently, several molecular genetic studies have provided multiple lines of evidence for a critical role of genetic factors in the expression of human bladder exstrophy-epispadias complex (BEEC). At this point,  The bladder exstrophy–epispadias complex (BEEC) is the most severe of all human congenital anomalies of the kidney and urinary tract (CAKUT), and involves the abdominal wall, pelvis, all of the urinary tract, the genitalia, and occasionally the spine and anus . Within Despite advances in surgical techniques, and improved understanding of the underlying anatomical defects, in later life many male and female patients experience chronic upper and lower urinary tract infections, sexual dysfunction, and urinary, or in the case of cloacal exstrophies, both urinary and fecal incontinence .ISL1 (ISL LIM homeobox 1) gene, a master control gene expressed in pericloacal mesenchyme. Multiple markers in this region showed evidence for an association with CBE, including 84 markers with genome-wide significance , where ΔGref and ΔGalt are the MFEs of the reference and altered transcript, respectively. The detailed information regarding all IDs , tool versions and databases, has been provided in To assess the impacts of SNPs on lncRNA secondary structures, we first extracted lncRNA transcript sequences from the human reference genome (hg38 version), according to the lncRNA transcript BED file, as Ref-transcripts. The secondary structure of the lncRNA was predicted using the RNAfold program , which cWe analyzed 72 genes that, according to literature, are putatively associated with BEEC , for funISL1 associated markers, rs9291768 and rs6874700, were analyzed for their potential association with any regulatory function , we identified one protein-coding gene, PARP8 , in the vicinity of rs9291768. Since ISL1 and PARP8 are the only protein coding genes mapped closely to rs9291768, we considered the mRNA of these two genes for further analysis. Our analysis, using web-tool LncTar, confirmed that neither ISL1 nor PARP8 mRNA is a target of this lncRNA (NONCODE_v5_lncRNA: NONHSAT249106) and therefore, the potential functional consequence of SNP rs9291768 has been assumed to be neutral. However, we cannot exclude the possibility that lncRNA NONHSAT249106 may have a functional impact on the transcripts of distantly located genes. All rsIDs associated with non-coding regions were subjected to GWAVA analysis. We found that all rsIDs showed a GWAVA score less than 0.5, which indicates that the variants are non-functional and they are likely to be associated with the disease conditions . From thfunction a. While on (ISL1 . In addiNONHSAT249106), which could potentially alter its stability, expression, or function, we compared the minimal free energy (MFE) secondary structure and the centroid secondary structure generated from the transcripts of both reference and altered IncRNAs, by using the RNAfold program. We found that the lncRNA secondary structure of both the reference and altered (re9291768) transcript showed a similar secondary structure, with MFE = −11.40Kcal/mol, suggesting that rs9291768 entails no structural change in lncRNA structure (NONHSAT249106), one other notable lncRNA, (LonLOC642366), overlaps with first exon of the ISL1 gene, whose function is still unclear.Considering the effect of SNP rs9291768 on the secondary structure of lncRNA , was investigated to build the three-dimensional structure of ISL1 wild type, and the p.Ala46Gly variant protein. The structures were generated using the wild type amino acid sequences (UniProt ID: P61371) and the mutant amino acid sequence (with p.Ala46Gly), by the Iterative Threading ASSEmbly Refinement method (I-TASSER) available online. For generating both models, I-TASSER used the template of the 4JCJ crystal structure from the PDB database . Since aISL1 at position c.137 might have functional consequences. Being a transcription factor, ISL1 protein acts by binding to the promoter of the target genes, and thereby, modulates their gene expression. From a structural point of view, the p.Ala46Gly is present at a loop turn, close to the N-terminal region of the ISL1 protein, and is also close to the DNA binding homeobox domain (amino acids 181–240). Since the LIM domain of ISL1 is known to interact with estrogen receptor α, the p.Ala46Gly variant protein may influence ISL1-estrogen receptor α interactions.We assume that the C>G variant in ISL1 gene alone are not sufficient enough to create a severe phenotype. Functional PPI analysis via STRING resulted into a network of 50 proteins A. This wTo better understand the functionality of these selective 50 genes in a STRING network, we investigated their putative involvement in different developmental stages of mice B. We fouISL1 gene in the etiology of the BEEC, our analysis provides novel insights about the consequences of genetic variants embedded within the ISL1 genomic landscape. In the present study, we investigated four genomic variants, and one recently introduced missense variant, linked to the BEEC. By performing structural analysis, we showed that although rs9291768, which is a part of lncRNA (NONCODE_v5_lncRNA: NONHSAT249106), ISL1 mRNA does not seem to be the main target of this lncRNA. Our amino acid sequence conservation analysis using Consurf, demonstrates that the p.Ala46Gly position (in LIM domain of ISL1 gene) is evolutionary conserved. However, the homeobox domain (amino acids 181-240), at the C-terminal region of ISL1, plays a unique role in DNA binding and the amino acid change of p.Ala46Gly, which resides at a loop turn close to the N-terminal region. Thus, it might have functional effects possibly interfering with the binding of ISL1 to estrogen receptor α. In an attempt to strengthen our understanding of the involvement of the ISL1 protein has been very strongly conserved during evolution, perhaps due to the involvement of different protein interaction networks. The multiplicity of ISL1 function is mirrored by its role in different diseases, such as BEEC, diabetes type I/II, and congenital heart defects ,32,33. TISL1 gene alone is not sufficient enough to create a severe phenotype. In addition, pathway enrichment analysis further suggests an involvement of hedgehog, hippo, and Wnt signaling pathways, in the etiology of BEEC. This in turn suggests that regardless of the complexity of the BEEC, pathway crosstalk also plays a significant role in pathogenesis of the disease. Importantly, the molecular framework of pathway integration, with regard to the ISL1 protein, needs further investigation.To understand the functional role of the ISL1 protein and other mutually exclusive proteins, for their involvement in the formation of the BEEC in more depth, we selected 72 genes which have been putatively linked to the BEEC, and performed PPI analysis. We found an interaction network between 50 proteins previously associated with the BEEC. Our approach does not predict the chronological contribution of these proteins to the disease phenotype. As the ISL1 protein shows up as a major interaction partner in our network analysis, we speculate that the involvement of mutational events in the ISL1. Previously, GWAS has identified the ISL1 locus to be associated with BEEC. Here, we investigated whether the most significantly associated variants rs9291768 and rs6874700 might constitute potential targets of the lncRNA (NONHSAT249106), residing in their close vicinity. However, the results of our analysis do not support this hypothesis. Interestingly, our analysis suggests that the ISL1 variant c.137C > G, results in decreased protein stability. In addition, by integrating multiple parameters, we provide novel insights about the involvement of ISL1 in the etiology of BEEC.In conclusion, our study characterized three genetic variants within the coding and non-coding region of


The aim of this study was to develop and validate an improved, simple, and sensitive method for the simultaneous determination of seven types  of biogenic amines (BAs) in wine matrices. For this reason, a modified QuEChERS combined with ultra-performance liquid chromatography coupled to a triple quadrupole mass spectrometry (UHPLC-QqQ-MS/MS) method was investigated. The optimization of UHPLC-QqQ-MS/MS separation and QuEChERS procedure was performed. Under optimum conditions, the excellent chromatographic performance of the whole separation was accomplished within 6.3 min analyzing time. Meanwhile, the recoveries ranged from 77.2% to 101.7%, while relative standard deviation (RSD) remained between 0.0% and 9.4%. The limit of detection  and the limit of quantification  were lower than those permitted by legislation in food matrices, which demonstrated the high sensitivity and applicability of this efficient method. This validated method was also applied in a pilot study to analyze BAs in 81 wine samples from Hexi Corridor Region , CAD, HEX, HIS, PEA, PUT, and TYR were detected to varying degrees in the samples. However, when compared with the existing standards, the BAs in all 81 wine samples did not exceed the prescribed limit value or toxic dose (2–40 mg/L). Moreover, a statistical approach was also conducted using Pearson correlation analysis, and to evaluate their concentrations in terms of wine parameters . The results showed that, among the seven kinds of BAs, the concentration of HIS had a certain correlation with alcoholic degree and grape variety. In addition, the level of PEA had a certain correlation with the wine pH and wine storage time. It is worth noting that this seems to be the first report regarding the application of QuEChERS-UHPLC-QqQ-MS/MS in the analysis of BAs in wine in this region. Biogenic amines (BAs) are toxic nitrogenous organic compounds that are mainly produced by microbial decarboxylation of free amino acids. Previous research found that the BAs appeared in a wide variety of food, such as meat  and fishBAs serve as an important component in organisms and play a key role in maintaining the basic physiological functions of biological cells. For example, a small amount of BAs is able to control the activity of the nervous system, eliminate free radicals, regulate blood pressure, enhance metabolic activity, and intestinal immunological competence . NeverthGiven the seriousness and urgency of the above problems, higher requirements have been placed on the BA determination methods and techniques. However, the analytical determination of BAs, both individually and simultaneously, is not an easy task because of their structure and generally low concentrations present in complex matrices. Many techniques have been developed to quantify these compounds in food products, including gas chromatography-mass spectrometry (GC-MS) , ion chrAdditionally, in order to minimize the loss of target analytes and subsequent decrease of recoveries, a lot of effort has been made to develop and standardize techniques of sample preparation. Traditional liquid–liquid extraction (LLE) , dispersThe Hexi Corridor Region  is a historic and major wine production area in China. The dry and cold climate throughout the year guarantees a reduction in pests and diseases in the process of grape cultivation. Therefore, it has also been known as the major production area of organic grapes and wines in China. In order to improve the overall quality of local wine, it is necessary to ensure the basic safety of wine production. The aim of the study was to develop QuEChERS procedures and the rapid UHPLC-QqQ-MS/MS method to carry out the simultaneous determination of seven common BAs (hexylamine (HEX), tryptamine (TRY), HIS, TYR, PUT, CAD, and PEA) in wines. Meanwhile, an additional goal was to apply the optimized and validated method to wine samples from the local wineries. As far as we know, this study can provide the first report regarding the application of QuEChERS-UHPLC-QqQ-MS/MS in BA analysis of wine in this region.2CO3), and sodium chloride (NaCl) were from Dengfeng Chemical Co., Ltd. . Distilled water was obtained from Watsons . Octadecylsilane (C18) sorbent and primary secondary amine (PSA) were acquired from Agilent Technologies .The BA reference standards of CAD, HEX, HIS, PEA, PUT, TRY and TYR were purchased from Dr. Ehrenstorfer . 1,7-diaminoheptane  was obtained from Pu Tian Tong chuang Biotechnology Co., Ltd. . HPLC grade methanol, acetonitrile, and formic acid were from Merck KGaA , analytical grade trichloromethane, butanol, ethyl acetate, anhydrous sodium carbonate (Nav/v). The standard solutions were stored under refrigerator conditions (4 °C) and protected from light. Prior to use, the working mixed standard solution was further diluted with 0.1% formic acid-water solution (v/v) to obtain the specific concentration required for the experiment.The individual stock solutions of CAD, HEX, HIS, PEA, PUT, TRY, TYR, and DAH at 100 mg/L were prepared in distilled water. A working mixed standard solution at 1 mg/L and internal standard solution at 100 mg/L were prepared immediately before use by diluting the individual stock solution in 0.1% formic acid-water solution , and 10% methanol-acetonitrile (v/v) were compared. The peak intensities and recoveries of seven kinds of BAs were used as indexes to determine the extraction solvent by a series of experiments.Under the condition of adjusting the pH value of sample solution to 12, the extraction effects of acetonitrile, ethyl acetate, trichloromethane-butanol , and the combined additive amount of NaCl and Na2CO3  were optimized. The recoveries of seven kinds of BAs were used as indexes to determine the optimal added salt amount and salt types by a series of experiments.Under the optimal pH and extraction solvent determined above, the sole additive amount of NaCl and NaIn order to improve recoveries provided by the aforementioned procedure, various dSPE sorbents  and sorbent mixtures (C18 + PSA) were tested during the clean-up step, respectively.v/v), 1 g NaCl and 2 g Na2CO3 were added, and the tubes were shaken for 5 min at room temperature. After extraction, the tubes were centrifuged for 5 min  with a 3K30 centrifuge ). The upper organic phase was transferred into a 15 mL vial and this extraction step was repeated. The extracted solution was diluted by 10 mL trichloromethane-butanol . Aliquots of 1 mL above solution containing 100 μL formic acid was evaporated under a gentle stream of nitrogen and reconstituted to 1 mL with 0.1% formic acid-water solution (v/v) for analysis. The extracts were stored at 4 °C until analysis in order to preserve their stability. Prior to injection, the sample was filtered through a 0.22 μm polyvinylidene fluoride (PVDF) filter ). Lastly, the optimized QuEChERS extraction procedure was employed to extract BAs from the wine samples. Briefly, 2.0 mL of sample was added to a 50 mL centrifuge tube with 30 μL DAH and 5 mL distilled water. The pH of the solution was adjusted to 12 by adding NaOH and homogenized on vortex mixer  for 2 min. Then, 5 mL of trichloromethane-butanol  mass spectrometer with an electrospray ionisation (ESI) interface, all from Agilent Technologies , operating in positive ion mode. Chromatographic separation was performed using an Eclipse Plus C18  from Agilent Technologies . In addition, 0.1% formic acid-water solution (eluent A) and 0.1% formic acid-acetonitrile solution (eluent B) were used as the mobile phase at a constant flow of 0.2 mL/min. Column temperature was set at 30 °C and the injection volume was 2 µL.+. The following settings were used: nebulizer, 45 psi; drying gas temperature, 300 °C; drying gas flow, 10 L/min; capillary voltage, 4000 V. Fragmentor voltage and collision energies were optimized for each analyte during infusion of the pure standard, and the most abundant fragment ion was chosen for the selected reaction monitoring  conditions in ESInitoring .Performance characteristics of the optimized method were established by a validation procedure with samples, studying linearity, LODs and LOQs, precision, and recovery.n = 3), and DAH concentration was 300 μg/L. Calculation curves were performed based on the average peak areas by internal standard calibration. LODs and LOQs were calculated as produce chromatographic peak at signal-to-noise ratio (S/N) of 3 and 10, respectively.The linearity calibration standards were prepared by combining a series of mixed working standard solutions into the blank wine samples to yield the desired concentrations in the range of 0.5 µg/L to 800 µg/L for each analyte  and interday precision, and they were expressed as relative standard deviation (RSD). For intraday precision, results were obtained from the injection of six spiked samples at concentration levels of 50, 100 and 500 μg/L, while for interday precision, three spiked samples, at the same aforementioned concentration levels, were analysed in five different days.Recovery studies were performed by spiking blank wine samples with the corresponding volume of the multi-compound working standard solutions (with 300 μg/L of DAH). Three concentrations were evaluated  by spiking three blank wine samples at each concentration level, and the pretreatment and instrumental parameters of samples are consistent before and after spiking. Wine samples were collected in local wineries  and supermarkets. Eighty-one wine samples, which include 15 commercial brands, were selected during the years 2008–2017 from various wineries. At least 3 L/sample were collected, and all samples were stored in the dark at 4 °C before analysis. BA identification and quantitation analyses in wine samples were performed using Agilent’s Mass Hunter Quantitative Analysis Software (version B.07.00). Microsoft Excel 2010  was used to calculate the mean, standard deviation and RSD. Statistical significance of the data and Pearson correlation analysis were performed using SPSS 19.0 . Data was subjected to one-way analysis of variance (ANOVA) and multiple comparative analysis was performed on Duncan’s test.+ precursor ions under the ESI+ mode. Then, dissociation with nitrogen was induced, and different collision energies were tested to identify the most abundant product ion scanned in the DMRM mode  added to the mobile phase was tested. It was found that when a small amount of formic acid was added to the acetonitrile-water system, the separation between the target objects became more complete, and the peak response value increased significantly , and 10% methanol-acetonitrile (v/v) were compared in this experiment. By comparing the peak abundance of the seven kinds of BAs, it was found that acetonitrile and trichloromethane-butanol solutions demonstrated better overall extraction efficiency for seven BAs, which assumed significant difference from other extraction agents . Even though non-purified treatment increased the replacement of nitrogen blowing solvent, it reduced the use of adsorbent and saved the treatment time. Therefore, after overall consideration, non-purification treatment was finally chosen as the optimized QuEChERS scheme.In order to further reduce the possible matrix effect in the extraction and detection processes, and to improve the detection accuracy, the purifying effects of C18, PSA and C18+PSA as adsorbent were investigated. The results showed that the purifying effects of these substances were significantly different and would affect the recovery ratio of PUT and CAD to varying degrees e. In addR2) higher than 0.9900 over the examined concentration range as showed in In terms of method validation, linear range, LOD, LOQ, precision, and recovery were studied, respectively. Good linear relationships were achieved with linear regression coefficients  and grape variety (p < 0.05). In addition, the level of PEA had a certain correlation with the wine pH (p < 0.01) and wine storage time (p < 0.05). In addition, the concentration of PEA was found to be related to the amount of CAD and HEX (p < 0.01). The content of TYR in wine was correlated with the presence of HIS (p < 0.01) and CAD (p < 0.05). Meanwhile, the total concentration of BAs in wine was significantly positively correlated with that of HIS, PUT and TYR (p < 0.01), namely, the total amount of BAs in wine is mainly determined by the concentration level of HIS, PUT, and TYR. This is consistent with the rule previously discovered regarding the number of various types of BAs in wine, as well as the research results obtained by Marques et al. [Furthermore, the effects of the storage time, the grape variety, the wine types, and the basic physicochemical indexes  of the 81 wine samples on the content of BAs were analyzed using Pearson correlation analysis . The ress et al.  and Garcs et al. . FurtherAt present, various methods have been developed for the analysis of BAs in foods. Initially, spectrofluorimetric techniques were used to determine BAs. More recently, chromatographic methods are the techniques most commonly applied to analyze several BAs simultaneously . Owing tMeanwhile, no laws or regulations on the dose of BAs in alcoholic beverages have been enacted in China. However, researchers have still proposed the toxic dose of some BAs in alcoholic beverages. According to the standard , the BAs2 additive amount, pH, alcoholic degree, and dissolved oxygen content [There are three possible sources of BAs in wine: already existing in the grape musts; formed by yeasts during alcoholic fermentation; or produced by the decarboxylation action of lactic acid bacteria during malolactic fermentation  was added to the must, the highest HIS values could be obtained from 3.7 to 8.3 mg/L) . Previou mg/L was content ,36.Today’s lifestyle and global markets have led to a large consumption of wine, with the new production and protection systems and the development of complex food chains, which in many cases requires a deeper understanding of how these wines are handled and forces us to face the new issues and challenges in providing safe products. Therefore, the safety of wine can be ensured by selecting grapes of good quality and the certain of sanitary conditions at the production stage, thus avoiding the contamination of infectious microbe from producing BAs in the fermentation process, inoculating excellent yeast strains and lactic acid bacteria strains without the activity of amino acid decarboxylase, and adopting appropriate brewing technology and reasonable storage environment. Although it is currently difficult to solve some of these problems, they should be studied and incorporated into future regulations to ensure wine safety and consumer health.A reliable and fast method to determine and quantify seven kinds of BAs in wine matrices has been developed and validated in this study. The optimization of MS/MS parameter, UHPLC separation, and QuEChERS procedure, which includes matrix pH condition, solvent, salt type and amount, clean-up agent, were performed, and the whole separation of BAs was accomplished within 6.3 min of analyzing time. Under optimum conditions, linear range, LOD, LOQ, precision, and recovery are good performance and quality results in terms of trueness, reproducibility, and repeatability. This validated method was applied in a pilot study to analyze BAs in 81 wine samples from the Hexi Corridor Region, CAD, HEX, HIS, PEA, PUT, and TYR were detected to varying degrees in the samples. Pearson correlation analysis showed that, among the seven kinds of BAs, the concentration of HIS had a certain correlation with alcoholic degree and grape variety. In addition, the level of PEA had a certain correlation with the wine pH and wine storage time. It was also noticed that the levels of HIS, PUT, and TYR strongly influenced the amine contents of wine. However, when compared with the existing standard, all of the BAs in the 81 kinds of wines did not exceed the prescribed limit value or toxic dose. That is to say, the food safety risks of BAs to wines in the Hexi Corridor Region are relatively low. However, the routine monitoring to BAs is still necessary to guarantee the wine safety in this area ,38.