Accumulating evidence has shown that diabetes has an impact on bone metabolism with conflicting results. Furthermore, little is known about the relationship of prediabetes with bone mineral density (BMD). Therefore, we explored the association between BMD and glucometabolic status in adults in the US.In this cross-sectional study, we extracted and analyzed data from the National Health and Nutrition Examination Survey (NHANES) from 2005 to 2018. A total of 14610 subjects aged 40 ≥ years diagnosed with diabetes, prediabetes, or normal glucose regulation (NGR) and had available data on BMD were eligible.The prevalence of prediabetes and diabetes in US adults aged 40 ≥ years were 39.2% and 26.4%, respectively. After multivariable adjustment, we found an increasing trend of BMD at the total hip, femoral neck, and lumbar spine with glucometabolic conditions from NGR and prediabetes to diabetes in adults aged ≥ 40 years in the US. This trend was more prominent in women than that in men. Fasting plasma glucose (FPG) and HbA1c levels were also positively correlated with BMD.Glucometabolic conditions were significantly associated with BMD values in US adults. Diabetes mellitus (DM) is a chronic metabolic disorder characterized by insufficient insulin production, endogenous insulin resistance, or both , 2, resuSubstantial evidence has linked diabetes with disorders of bone metabolism and skeletal fragility, which have been suggested to be novel complications of long-term exposure to an uncontrolled blood glucose environment , 7. The Prediabetes, defined as the presence of impaired fasting glucose (IFG), impaired glucose tolerance (IGT), and/or mildly elevated hemoglobin A1c (HbA1c), precedes the onset of diabetes. Emerging insights into macrovascular disorders such as cardiovascular disease, stroke, and peripheral vascular disease are relevant to prediabetes –12. CurrIn this study, we investigated BMD trends in US adults aged ≥ 40 years with diabetes, prediabetes, and normal glucose regulation (NGR) using nationally representative sample data from the National Health and Nutrition Examination Survey (NHANES) from 2005 to 2018.The data for this cross-sectional study were obtained from the NHANES, a survey conducted by the National Center for Health Statistics (NCHS) of the Centers for Disease Control and Prevention (CDC) to assess the health and nutritional status of a nationally representative sample of the US civilian population using a multistage, stratified, clustered probability design. Questionnaire interviews, laboratory tests, and physical examinations were included in the survey. The continuous datasets of the NHANES 2005–2006, 2007–2008, 2009–2010, 2013–2014 and 2017–2018 were combined in this study. NHANES 2011–2012, 2015–2016 were excluded because of the lack of data on the lumbar spine and femur BMD during these periods. The NHANES study protocol was approved by the NCHS Ethics Review Board, and written informed consent was obtained from all participants  scans were only administered to eligible survey participants aged ≥ 40 years in the 2013–2014 cycle. This study population was limited to subjects aged ≥40 years who were diagnosed with diabetes, prediabetes, or NGR with available data on BMD. Among the total 50463 participants for the 5-cycle survey, after the exclusion of 17445 subjects without glucose status (n=33018) and 10250 without valid BMD data (n=22768), 14610 subjects aged ≥40 years were enrolled in our study for final analysis.Diabetes status was confirmed when subjects had a positive response to the question, “Have you ever been told by a doctor that you have diabetes?”. Furthermore, the diagnosis of diabetes was also based on American Diabetes Association (ADA) criteria , which iBMD of the total femur and lumbar spine was assessed using DXA scans with Hologic QDR-4500A fan-beam densitometers  in the NHANES mobile examination center (MEC). The detailed examination protocol for DXA is described on the NHANES website. BMD values of the total hip, femoral neck, trochanter, intertrochanter, Ward’s triangle, and vertebrae L1–L4 were obtained from the DXA profiles. Based on the osteoporosis diagnostic criteria of the World Health Organization (WHO) , BMD val2). The systolic blood pressure (SBP) and diastolic blood pressure (DBP) were measured. Data on serum creatinine (sCr), uric acid (UA), total cholesterol (TC), triglyceride (TG), alkaline phosphatase (ALP), serum calcium, and serum phosphorus were also extracted from the database. The detailed process of the data collection was obtained from the NHANES website.Demographic information, including age, gender , race/ethnicity , educational level , marital status , ratio of family income to poverty , smoking at least 100 cigarettes in life (yes/no), and vigorous recreational activities (yes/no) were collected from the NHANES database. Body mass index (BMI) was calculated as the weight (kg) divided by the squared height  and 95% confidence interval (95% CI) were estimated. Model 1 was unadjusted. Model 2 was adjusted for age, sex, and race. Model 3 was adjusted for model 2 adjustments plus BMI, education level, marital status, ratio of family income to poverty, smoking at least 100 cigarettes in life, vigorous recreational activities, ALP, total calcium, uric acid, creatinine, and cholesterol. SPSS software , STATA 15.0 software  were used for statistical analyses. A two-sided P value < 0.05 is considered statistically significant.Of the 14610 participants, 5730 (39.2%) had prediabetes and 3864 (26.4%) had diabetes, accounting for 65.6% of the entire cohort. Participants with diabetes and prediabetes differed from those with NGR in several demographic and biochemical indices Table 1.P-trend <0.05) after the adjustment of three models . After stratification by sex, an interesting phenomenon was that men aged ≥ 40 years had no significant association between lumbar BMD and glucose status after multivariate adjustment (P=0.149), whereas the trend remained stable in all skeletal sites in the corresponding women , while further adjustment for BMI, other socio-demographics, and biochemical indices attenuated its significance (P=0.110).As shown in After adjustment for multivariate factors, there was enough statistical evidence to show a positive association between HbA1c and BMD in all skeletal sites for the whole cohort  and both sexes Table 4.In the current study, after adjusting for multiple potential confounders, we reported an increasing trend of BMD at the total hip, femoral neck, and lumbar spine with glucometabolic conditions from NGR and prediabetes to diabetes in US adults ≥40 years. This trend was more prominent in women than that in men. Fasting blood glucose and HbA1c levels were also positively related to BMD. To the best of our knowledge, this is the first study to evaluate BMD values in US adults with normal, mild, and overt elevated plasma glucose levels with nationally representative data.There is a paucity of research on the link between diabetes and BMD values. Some studies have reported that patients with diabetes have decreased BMDs compared to NGR subjects , 22, whiThe relationship between HbA1c, hyperglycemia, and BMD had conflicting results. A study from Japan  indicateOur study focused on middle-aged and elderly US individuals and found an increasing trend in BMD in patients with prediabetes and diabetes. However, these results were not in line with the findings in youth with diabetes and prediabetes. An NHANES study of 2005–2006  enrolledAccumulating evidence from cohort studies and meta-analyses has shown that diabetes is associated with an elevated risk of fragility fractures , 30, desIn the current study, we analyzed data from the NHANES, which is unique in collecting demographic, health, and nutritional information. The NHANES sample represents the national US civilian population. In addition, the interview and examination processes were under high-quality control to ensure the reliability of the data. This is the key strength of this study. Furthermore, diabetes and prediabetes diagnosis were based on self-reported diagnosis and on the results of OGTT and HbA1c, increasing the diagnostic accuracy of glucose metabolism. However, this study had several unavoidable limitations. First, because of the cross-sectional nature of the NHANES, it is difficult to conclude on the cause-effect relationship. Second, anti-diabetic medicine was not included in the diabetes group, which may negatively or positively influence BMD. Third, although we examined the changes in socio-demographics, BMI, and biochemical indexes, other potential residual confounders, such as vitamin D and calcium supplementation, use of glucocorticoids may still need to be considered. Therefore, more prospective studies are required to investigate bone metabolism at different glucose levels.In conclusion, based on nationally representative data extracted from the NHANENS database from 2005 to 2018, we found a positive association between glucometabolic conditions and BMD values in US adults. To better understand the mechanisms underlying bone metabolism in individuals with prediabetes and diabetes, more longitudinal research is needed to explain BMD changes from normoglycemia to prediabetes and diabetes.The original contributions presented in the study are included in the article/supplementary material. Further inquiries can be directed to the corresponding authors.The studies involving human participants were reviewed and approved by NHANES. The patients/participants provided their written informed consent to participate in this study.JY extracted data from NHANES database, analyzed the data, wrote the manuscript. PJ and J-BZ conceived the idea for the study and edited the manuscript. All authors read and approved the final manuscript. All authors contributed to the article and approved the submitted version.This work was supported by the National Natural Science Foundation of China , the Beijing Municipal Administration of Hospitals’ Youth Program (grant number QML20170204), and Excellent Talents in the Dongcheng District of Beijing.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.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Lung adenocarcinoma (LUAD) is the most common type of Non-small-cell lung cancer (NSCLC). Distant metastasis of lung adenocarcinoma reduces the survival rate. we aim to develop a nomogram in order to predict the survival of patients with metastatic lung adenocarcinoma.We retrospectively collected patients who were initially diagnosed as metastatic LUAD from 2010 to 2015 from SEER database. Based on the multivariate and univariate Cox regression analysis of the training cohorts, independent prognostic factors were assessed. The nomogram prediction model was then constructed based on these prognostic factors to predict the overall survival at 12, 24 and 36 months after surgery. Nomogram were identified and calibrated by c-index, time-dependent receiver operating characteristic curve (time-dependent AUC) and calibration curve. Decision curve analysis (DCA) was used to quantify the net benefit of the nomogram at different threshold probabilities, and to better compare with the TNM staging system, we calculated the c-index of this nomogram as well as the net reclassification improvement (NRI) and the integrated discrimination improvement (IDI).A total of 1102 patients with metastatic LUAD who met the requirements were included for analysis. They were randomly divided into 774 in the training cohorts and 328 in the validation cohorts. As can be seen from the calibration plots, the predicted nomogram and the actual observations in both of the training and validation cohorts were generally consistent. The time dependent AUC values of 12 months, 24 months and 36 months were 0.707, 0.674 and 0.686 in the training cohorts and 0.690, 0.680 and 0.688 in the verification cohorts, respectively. C-indexes for the training and validation cohorts were 0.653 (95%CI 0.626-0.68)and 0.663 (95%CI 0.626-1), respectively. NRI and IDI show that the model is more clinical applicable than the existing staging system. In addition, our risk scoring system based on Kaplan Meier (K-M) survival curve can accurately divide patients into three hierarchy risk groups.This has led to the development and validation of a prognostic nomogram to assist clinicians in determining the prognosis of patients with metastatic lung adenocarcinoma after primary site surgery. As one of the most common forms of cancer, lung cancer has a high mortality rate among the 13 regions of the world indicating that lung cancer is a serious threat to human health and life .It is estimated that the 5-year relative survival rate of patients with LUAD is only 5% due to the fact that about 57% of the patients have advanced stage and metastatic disease . A lung A tumor-node-metastases (TNM) staging system is one of the main indicators used in predicting survival and determining treatment options . AlthougIn clinical practice, nomograms are widely used in terms of prognosis for cancer as a simple statistical tool . By evalConsequently, the objective of this study is to identify prognostic factors that are relevant to LUAD after surgery by examining data in the Surveillance, Epidemiology, and End Results (SEER) database of LUAD patients and to develop a prediction model for survival after surgery. An evaluation of the prediction model was assessed using calibration curves, receiver operating characteristic (ROC) curves, and decision curve analyses (DCA) .The data for the patients were obtained through the US SEER database using SEER*Stat software.Participants should meet the following criteria to be considered for inclusion in the study:(1) diagnosed with pulmonary adenocarcinoma by histology from 2010 to 2015; (2) patients with distant metastasis at the initial diagnosis; (3)received primary site surgery (4) patients with important variable information being fully documented. This study excluded patients diagnosed by autopsy or death certificate. In addition, patients with survival time < 1 month were also excluded. A random sample of patients selected for the study was divided into two groups, a training group (70%) and a testing group (30%).A human subject or any personally identifiable information was not included in the data. Consequently, informed consent was not required for this portion of the study.In this study, 15 variables were included to identify independent prognostic factors in patients with postoperative patients with metastatic LUAD. The demographic variables include age, race, gender and marital status. The clinicopathological characteristics of the tumor include primary location, laterality, tumor grade, N stage, T stage,liver metastasis, brain metastasis, lung metastasis, bone metastasis and treatment information, including chemotherapy and radiotherapy. Among the primary outcomes of this study was overall survival (OS). It was defined as the interval between the date of diagnosis and the date of death from whatever cause. Using X-tile software, we determined the best age and survival cut-off values . Additionally, the T stage can be divided into the following stages: T0, T1, T2, T3, and T4. It was discussed that the N stage consisted of N0 (No), N1-N3 (Yes). The M stage was defined as M0 for no metastasis and M1 for positive metastasis. We used the eighth edition of the TNM staging system to define T, N, and M for patients in the clinical stage.The R software randomly divided all patients into training and verification cohorts according to a ratio of 7:3. For the purpose of comparing the variables between the training cohort and the validation cohort, we used Chi-square test. In order to determine the independent prognostic factors of postoperative patients with metastatic LUAD, we first used univariate Cox regression analysis, and the variables with P < 0.05 were included in subsequent multivariate Cox regression analysis. The variables with P < 0.05 were identified as independent prognostic factors. The nomogram prediction model was then constructed based on these prognostic factors to predict the overall survival at 12-, 24- and 36-months after surgery. We explored the differences in survival rates between patients with different subtypes of lung adenocarcinoma based on the analysis of KM survival curves of the invasive subtypes included in the study.To compare the predicted event and the actual event, calibration plots were drawn for the 12-, 24-, and 36-month OS probability. In order to assess the model’s ability to distinguish between events and nonevents, receiver operating characteristic curves (ROC) and area under curves (AUC) were used. In addition, ROC curves or time-varying ROC curves of all independent variables were generated and compared with the AUC values of the corresponding nomogram. We established a calibration curve with a decision curve analysis (DCA) curve for evaluating the nomogram. A comparison of the nomogram with the conventional TNM staging system was based on calculating the C-index. In addition, we also calculated the net reclassification improvement (NRI) and the integrated discrimination improvement (IDI) of the nomogram. Finally, according to the best cut-off value of risk score, patients in the training cohorts and verification cohorts were divided into high-risk group, medium risk group and low-risk group, and the logarithmic rank test of Kaplan Meier (K-M) survival curve was performed to verify the prognostic value of nomogram. According to the calculated total points of each patient from the nomogram, a risk classification system was created by using X-tile program for Postoperative patients with metastatic lung adenocarcinoma. All statistical analysis was conducted using SPSS 25.0 and R software (version 3.6.1) software. P <0.05  was regarded as statistically significant in this study.In this study, 1102 postoperative patients with metastatic LUAD were included based on the inclusion and exclusion criteria The overall patient screening and study process is shown in We have updated the histological typing of lung adenocarcinoma in the seer database according to the histological subtype classification system for LUAD proposed by the International Association for the Study of Lung Cancer, the American Thoracic Society and the European Respiratory Society in 2011 . The rea.In order to identify independent prognostic factors in LUAD patients, we performed univariate and multivariate Cox proportional hazards regression analysis. On the basis of a Univariate Cox regression analysis, the prognostic factors of patients with metastatic LUAD included age, gender, race, N stage, primary location, differentiation grade, tumor size, bone metastasis, liver metastasis, radiotherapy, and chemotherapy. In a multivariate Cox regression analysis, we found age, gender, primary location, N stage, bone metastasis, liver metastasis, radiotherapy, and chemotherapy significantly correlated with the postoperative prognosis of postoperative LUAD patients We constructed a nomogram for postoperative patients with metastatic LUAD according to the selected independent prognostic factors. In order to construct the model, we used these eight variables. Following this, a nomogram was constructed based on the training cohorts for predicting 12-, 24-, and 36-month OS Figure 1C-index values in the training cohorts was 0.653 (95%CI 0.626-0.68) and in the validation cohorts they was 0.663 (95%CI 0.626-1).The time dependent AUC values of 12 months, 24 months and 36 months were 0.707, 0.674 and 0.686 in the training cohorts and 0.690, 0.680 and 0.688 in the verification cohorts, respectively. Figure 2Compared to the FIGO criteria-based tumor staging system, the nomogram was found to offer clinical benefits for predicting tumor behavior. The discrimination ability of the nomogram and the eighth version of the TNM staging system was compared in the training and validation cohorts using the IDI and NRI indices see in Table 4 Using the nomogram we were able to calculate a risk stratification based on the total points. Based on the X-tile analysis, 303 and 372 scores were selected as the optimal cutoff points , but over the past decade, they have mainly focused on anti-programmed death protein-1 (PD-1) and anti-programmed death ligand 1 (PD-L1) therapies, as well as immune checkpoint blockade therapy (ICB) used to treat these patients . The treIndividualized treatment is clearly becoming a paradigm for the treatment of patients with metastatic lung adenocarcinoma. To choose the right treatment, you’ve got to analyze histological features, individual genetic characteristics (mutations), antitumor drug resistance, tumor microenvironment, etc. With this personalized information, treatment will be greatly facilitated, and the nomogram we have created is one of the tools to provide information on patient survival. Hence, it is of paramount importance to study the prognosis of postoperative patients with metastatic LUAD.In a survival analysis of different pathological subgroups of lung adenocarcinoma patients, it was found to be more likely that those patients who had predominantly lepidic and acinar forms of the disease would survive longer than those with predominantly solid and papillary forms. Other previous studies have confirmed these findings, with colloid-predominant or solid-predominant subtypes of lung adenocarcinoma often representing a poorer prognosis, suggesting that different subtypes of lung cancer can also predict survival in any given patient to some extent , 23. PD-There are limitations to the current TNM staging systems in predicting mortality in cancer patients. As a result, it is essential to identify patients at high risk level after surgical resection . The nomAccordingly, the best cutoff value for OS was determined using the X-tile software, in order to divide the patients into three subgroups . Results from the Kaplan-Meier and Cox hazard ratio models showed significant differences between the three groups. It has been proven that nomogram can identify patients in the high-risk group more accurately than other staging systems. High-risk groups suffer from poor prognoses compared to patients at lower risk levels. In fact, it is vital to pay more attention to patients with total points≥372 than to those without.All the patients we included have undergone surgery at the primary site, and there is no similar prognostic study for these patients. NAKAZAWA et al. results suggested that existing liver and bone metastasis adversely affected the outcome of the disease . The preTraditional staging of TNM has always included high N as having a poor prognosis, which is also reflected in our prognostic model. The results of a study on 167 patients with non-small cell lung cancer who underwent complete resection showed a 5-year survival rate of 20% for patients classified as N1 and 21% for patients classified as N2, with no significant differences in survival between the two groups of patients . It is nTaking these results into account, we suggest that baseline characteristics in patients with metastatic LUAD, such as gender and age, can be considered independent prognostic factors. In previous retrospective studies , 34, we All patients in this study underwent surgical treatment at the primary site, so the operation was not included in the nomogram, which does not mean that the surgical treatment has no effect on the survival rate of patients. For stage IV NSCLC patients with one or more synchronous metastases, a surgical strategy combined with systemic therapy, such as radiotherapy or chemotherapy can be effective , 37.Adjuvant chemotherapy and adjuvant radiotherapy are controversial factors. Patients with NSCLC are usually treated with chemotherapy that uses platinum-based drugs. Radiation therapy, if necessary, is often used in conjunction with chemotherapy. The need to improve the long-term survival of patients with locally advanced NSCLC indicates the importance of a combination therapy, in which chemotherapy is a crucial component of controlling distant metastases in order to increase their overall survival rates . Cancer An analysis of a previous meta-analysis reported that postoperative chest radiotherapy has an overall negative effect on survival, resulting in a 21.3% relative increase in odds of dying [hazard ratio (HR)1.21 (95% CI 1.08-1.34)]; meaning, an absolute reduction of 7% in survival from 55% to 48% at 2 years owing to the decrease in survival rates possible , 44. OneThere is enough sample size in SEER database to collect large sample data for research, which makes the results of this study very convincing. Even so, there will always be a certain amount of limitations associated with this study. In the first place, since this is a retrospective study, there will always be some effect of bias. Second, we cannot make a quantitative study on the specific chemotherapy and radiotherapy schemes adopted by patients and the relevant data of surgical margins state. Third, the seer database lacks information on specific tumor markers that could improve prognostic accuracy, as well as certain hematological indicators: neutrophils, platelets, and absolute lymphocyte values. Finally, there is a lack of clinical external validation data to evaluate our nomogram. Despite these limitations, our first online version of the nomogram and nomogram for surgically resected metastatic LUAD patients has high clinical applicability and provides individualized and accurate survival predictions for each patient.The survival rate of patients with metastatic LUAD after primary site surgery was predicted using a nomogram. Compared with other staging systems, the model has good prediction accuracy and clinical utility, and can provide a reference for clinicians to formulate treatment plans.https://seer.cancer.gov/.Publicly available datasets were analyzed in this study. This data can be found here: XT, CM, and SP contributed to conception and design of the study. BZ organized the database. CM performed the statistical analysis. CM wrote the first draft of the manuscript. YG, BZ, SL, and SP wrote sections of the manuscript. All authors contributed to manuscript revision, read, and approved the submitted version.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.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Bipolar disorder (BD) is easy to be misdiagnosed as major depressive disorder (MDD), which may contribute to a delay in treatment and affect prognosis. Circadian rhythm dysfunction is significantly associated with conversion from MDD to BD. So far, there has been no study that has revealed a relationship between circadian rhythm gene polymorphism and MDD-to-BD conversion. Furthermore, the prediction of MDD-to-BD conversion has not been made by integrating multidimensional data. The study combined clinical and genetic factors to establish a predictive model through machine learning (ML) for MDD-to-BD conversion.By following up for 5 years, 70 patients with MDD and 68 patients with BD were included in this study at last. Single nucleotide polymorphisms (SNPs) of the circadian rhythm genes were selected for detection. The R software was used to operate feature screening and establish a predictive model. The predictive model was established by logistic regression, which was performed by four evaluation methods.It was found that age of onset was a risk factor for MDD-to-BD conversion. The younger the age of onset, the higher the risk of BD. Furthermore, suicide attempts and the number of hospitalizations were associated with MDD-to-BD conversion. Eleven circadian rhythm gene polymorphisms were associated with MDD-to-BD conversion by feature screening. These factors were used to establish two models, and 4 evaluation methods proved that the model with clinical characteristics and SNPs had the better predictive ability.The risk factors for MDD-to-BD conversion have been found, and a predictive model has been established, with a specific guiding significance for clinical diagnosis. Bipolar disorder (BD), including depressive and manic phases, is a common mental disorder with high morbidity, disability, and suicide risk, which seriously impairs social functioning –3. The pSome reports have identified risk factors that are related to conversion to BD, most of which are clinical risk factors, such as the age of onset of the MDD , family Machine learning (ML) studies complex distributions and determines possible connections based on complex and conditional relationships between variables. In addition, it verifies the dependability of the test results through repeated cross-validation . In receThis study aimed to find clinical risk factors for MDD-to-BD conversion. Based on that, we tried to establish a predictive model through ML by combining clinical and genetic factors. We hypothesized that by combining clinical characteristics with circadian rhythm gene polymorphisms, the predictive model could aid in the accurate clinical identification of BD. In this study, we compared the clinical characteristics of patients from MDD and BD groups to identify clinical risk factors. Then, we selected clinical and genetic features by three methods. Finally, we established a predictive model and evaluated its predictive ability.This study was carried out among inpatients at Zhongda Hospital. We recruited 500 patients with an initial diagnosis of MDD, and their clinical variables were collected to establish a database. The inclusion criteria for MDD were shown as follows. All recruited subjects were between 15 and 65 years old, having presented with depressive symptoms and received an initial diagnosis of MDD from two senior psychiatrists according to the Diagnostic and Statistical Manual of the American Psychiatric Association (DSM-IV). All were initially inpatients and scored 18 or over on the 17-item Hamilton Depression Rating Scale (HDRS-17) .A cohort of 299 of the initially recruited subjects was followed up successfully after 5 years and the diagnosis was determined independently by two senior psychiatrists. Of these 299 subjects, those with a score of more than 6 on the Young Mania Rating Scale (YMRS)  met the t-test was carried out to check for differences between two independent means (two groups). The input parameters included: tails two; parent distribution normal; effect size d 0.5; α err prob 0.05; power (1– β err prob) 0.08; allocation ratio N2/N1 1. Each group should have more than 64 patients.Exclusion criteria for MDD included documented history of diagnoses on Axis 1  of DSM-IV, pregnancy, lactation, abnormalities of blood measures, heart, liver and kidney function after examination, or a history of brain organic diseases and endocrine diseases. The exclusion criteria of the BD group were the same as those of the MDD group, except for the diagnosis of BD. This study was approved by the Ethical Committee of Zhongda Hospital (2021ZDSYLL062-P01), in accordance with the Declaration of Helsinki. The sample size was calculated by G Power. A two-sample A previous similar cohort study showed the rate of conversion from MDD to BD in 3 years as 12.4% . AssuminIn the patients with a stable diagnosis of MDD, a subgroup of 70 subjects was selected by random sampling. There was no significant difference in general clinical characteristics between these 70 patients with MDD and the patients who were not selected. The clinical characteristic variables in both MDD and BD groups included gender, the age of the first onset of MDD, family history of affective disorder, suicide attempt, psychotic symptoms, and the number of hospitalizations.According to the circadian rhythm pathway, 6 candidate genes were selected: CLOCK, ARNTL, CSKN1E, PER1, PER2, and PER3 . In totaFasting venous blood was collected from all subjects at 8:00 in the morning into an EDTA anticoagulant tube and stored at −80°C until genotyping. The genomic deoxyribonucleic acid (DNA) was extracted from blood cells by that TIANamp Blood DNA Extraction Kit (Tiangen) and divided into pieces for testing.Genomic DNA was sheared and subjected to Illumina paired-end DNA library preparation. All the SNPs were genotyped using FastTaget. The captured libraries were loaded onto the Illumina HiSeq platforms. Sequencing was performed at Genesky Biotechnologies Inc., Shanghai, China. The obtained data was directly imported into GenomeStudio software for analysis to obtain the genotype data.P < 0.05 was considered statistically significant. Haploview 4.1 program was used to test for gene detection rate and minor allele frequency (MAF). The SNPs with a detection rate lower than 90% or MAF lower than 5% were eliminated   . The log < 0.05) .The result of characteristics selected by RFE included gender, age at the first onset, suicide attempt, the number of hospitalizations, rs4146388 in ARNTL, rs1056547, and rs9312661 in CLOCK, rs10462024 and rs2585408 in PER1, rs2304671 and rs10462023 in PER2. The AUC value was 0.852.A GBDT method was used to select the top ten variables according to the importance of the variables: gender, age of the first onset, suicide attempt, the number of hospitalizations, rs4146388 in ARNTL, rs1056547, rs9312661 and rs3736544 in CLOCK, rs228669 and rs2859387 in PER3. The AUC value was 0.802.For RF with 5 cross-validation results, according to the smallest Gini index, the top five variables in each classification were chosen. They were gender, age of the first onset, suicide attempt, the number of hospitalizations, rs4146388 in ARNTL, rs2585405 in PER1. The AUC value was 0.616.p-value = 0.805). The calibration curve of model2 (p = 0.027), indicating an improved predictive ability in model2 compared with model1. We chose model2 as the final predictive model based on the above results. Moreover, five cross-validations was used as internal validation for model2 (Two models were established: the first model (model1) contained the age of first onset, gender, suicide attempts, and number of hospitalizations; and the second model (model2) included the age of first onset, gender, suicide attempts, number of hospitalizations, rs4146388 in ARNTL, rs1056547, rs9312661, and rs3736544 in CLOCK, rs2585408, rs10462024 and rs2585405 in PER1, rs2304671 and rs10462023 in PER2, rs228669 and rs2859387 in PER3. Their evaluation was performed by AUC value , calibraf model2  was closf model2 , suggestr model2 . The mear model2  for its According to these results, the age of onset was the risk factor for MDD-to-BD conversion using logistic regression analysis, RFE, GBDT, and RF. Furthermore, suicide attempts and the number of hospitalizations were also associated with MDD-to-BD conversion. Eleven circadian rhythm gene polymorphisms were associated with MDD-to-BD conversion by feature screening. In this study, the characteristics mentioned above were used to establish two models by ML, and four evaluation methods proved that the model with clinical characteristics and SNPs had the better predictive ability.The conversion rate of our study was 12.7%, similar to a prospective cohort study which showed the rate of conversion from MDD to BD in 3 years was 12.4% . The conThe logistic regression analysis, RFE, and GBDT showed that age at first onset is a risk factor for the disease. We found that the younger the age of onset, the higher the risk of BD, which is consistent with previous studies. Baldessarini et al. found a different rate of switch-risk between adult patients with MDD and adolescent patients with MDD, with the adolescent rate of 5.62%/year (9.33%/1.66 years) and an adult rate of 1.26%/year (3.66%/2.90 years) , indicatWhile using GBDT and RF to screen the characteristics, it was found that suicide attempts were also closely related to MDD-to-BD conversion. Previous studies have clustered many risk factors and identified the four most important variables, including suicide attempts . It was This study provided preliminary evidence that several SNPs of multiple circadian rhythm genes were associated with MDD-to-BD conversion. Some polymorphisms, such as rs2859387 of PER3 and rs4146388 of ARNTL, were related to the occurrence of BD in previous studies. We found rs4146388 in ARNTL was associated with MDD-to-BD conversion. Significant epistasis was found in the ARNTL (rs4146388 and rs7107287) and PER3 (rs2172563), indicating that the sum effects influence the occurrence of BD .Mansour et al. found that the genotype distribution of PER3 rs2859387 was significantly different between BD and healthy control subjects . At the The purpose of the feature selection process is to use an adequate but minimal number of features to achieve optimal prediction results of the model . Two metIn this study, four methods were performed to evaluate the effectiveness of the model. Although the AUC value of the model2 with SNPs showed no difference from that of the model1 without SNPs, three other feature selection methods showed that the predictive ability of the model with both clinical characteristics and SNPs was improved. The AUC value of model2 was lower than model1. It may be due to the fact that the AUC is an insensitive metric when evaluating improvement in predictive performance . The AUCThere are some limitations. First of all, the sample size of this study is small; however, we did the power analysis, and this study achieved the requirement of the minimum number of patients. Second, the dropout rate of patients who followed up was relatively high, impacting the sample size. More studies with a larger sample size, more follow-up times, and more extended follow-up periods are needed to be carried out to get a lower dropout rate and validate the predictive model.We have found that the age of onset was a risk factor for MDD-to-BD conversion using logistic regression analysis, RFE, GBDT, and RF. Furthermore, suicide attempts and the number of hospitalizations were also associated with MDD-to-BD conversion. We showed that eleven circadian rhythm gene polymorphisms were associated with MDD-to-BD conversion by feature screening. A model with both clinical features and genetic polymorphisms had a better predictive ability. In future research, a follow-up study with a larger sample is needed to verify our predictive model and modify the accuracy of the predictive model.The original contributions presented in the study are included in the article/The studies involving human participants were reviewed and approved by the Ethical Committee of Zhongda Hospital. Written informed consent to participate in this study was provided by the participants' legal guardian/next of kin.LC: formal analysis, writing—original drafting, writing—reviewing and editing, and validation. YH and BC: formal analysis. TS, ZC, TT, and CG: data curation. SC: data curation and blood collection. WC: formal analysis and reviewing and editing. YY and ZZ: conceptualization and methodology. ZX: conceptualization, methodology, investigation, data curation, project administration, and writing—reviewing and editing. All authors contributed to the article and approved the submitted version.This work was funded by the Natural Science Foundation of Jiangsu Province (No. BK20181272), Jiangsu Provincial Medical Youth Talent (No. QNRC2016825), National Natural Science Foundation of China (Nos. 81301167 and 81971277), and Key Research and Development Program  Foundation of Jiangsu Province (No. BE2019714).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.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


The independent risk factors were screened by performing a univariate analysis and binary logistic regression analysis, and then, all cases were randomly divided into the training set and the test set in a 7:3 ratio for constructing and validating the model. We then constructed the prediction models by the traditional logical regression method and the XGBoost method and tested the prediction performance of the two models by resampling.P < 0.05). The area under the receiver operating characteristic curve (AUC) score and the F1 score with the training set of the XGBoost model  were significantly higher than those of the logistic model . In the test set, the AUC and F1 scores of the XGBoost model  were also higher than those of the logistic model .The results of the binary logistic regression analysis showed that several factors, including the age of men and women, abnormal ovarian structure, prolactin (PRL), anti-Müllerian hormone (AMH), activated partial thromboplastin time (APTT), anticardiolipin antibody (ACA), and thyroid peroxidase antibody (TPO-Ab), independently influenced missed abortion significantly (We established a prediction model based on the XGBoost algorithm, which can accurately predict the risk of missed abortion in patients with IVF-ET. This model performed better than the traditional logical regression model. Due to the complexity of the etiology, the pathogenesis of missed abortion is not known , and even threaten the life of the individual . For womPredictive models are widely used in the medical field, and the common modeling methods include traditional logical regression  and machMachine learning is the core of artificial intelligence, and its data processing, induction, and synthesis abilities are better than those of other statistical methods. A lifting algorithm based on the regression tree called extreme gradient boosting (XGBoost) is a machine learning hot spot. Because of its short training time and high precision, it is widely used in the medical field . The XGBThis retrospective study was approved by the Ethics Committee of the Affiliated Women and Children’s Hospital of Qingdao University (QFELL-YJ-2022–18). As this was a retrospective study, informed patient consent was waived by the Ethics Committee of the Affiliated Women and Children’s Hospital of Qingdao University.n  =  340), and those in the normal pregnancy population were placed in the control group (n  =  677).Data were collected from 1,017 infertile women who were treated in the Reproductive Center of Qingdao Women’s and Children’s Hospital from September 2019 to May 2022. The inclusion criteria were as follows: (1) Patients who received complete treatment of IVF-ET and completed follow-up in our center; (2) patients with intrauterine pregnancy diagnosed by ultrasound after treatment; (3) patients diagnosed with a singleton pregnancy. The exclusion criteria were as follows: (1) Couples receiving donated sperm or eggs; (2) patients with ectopic pregnancy; (3) patients with multiple pregnancies; (4) pregnancy failure due to special reasons such as medication or trauma; (5) patients who also had severe heart, liver, lung, kidney, and other organ disorders; (6) incomplete information in the patient database. Based on the pregnancy outcome, individuals who had missed abortions were placed in the observation group (The results of ultrasonic examinations showed that missed abortion was associated with one of the following criteria : (1) theNeisseria gonorrhoeae, TORCH test, etc.  genetic factors, including chromosomal abnormalities in men and women ; (2) femst, etc. ; (7) abnst, etc. ; (8) utest, etc. ; (9) malst, etc. ; (10) otst, etc. .T-test was conducted for normally distributed continuous variables, the Mann–Whitney U test was conducted for non-normally distributed continuous variables, and the Chi-squared test was conducted for classified variables. All differences between groups were considered to be statistically significant at P < 0.05. The independent risk factors were determined by performing a binary logistic regression analysis.All data were analyzed using the SPSS 20.0 statistical software . Continuous variables that followed a normal distribution were expressed as the mean  ± standard deviation. Continuous variables that did not follow a normal distribution were expressed as the median (25th–75th percentile), and classified variables were expressed as frequency (percentage). Based on the data, the https://www.python.org/download/releases/3.0/) to construct the prediction model based on XGBoost and traditional logical regression. The train_test_split function of the sklearn package was used to split the dataset into two; 70% of it formed the training dataset, and 30% formed the test dataset (All independent risk factors were processed using Python (version 3.0;  dataset . The datTrichomonas vaginitis, and the TORCH test were removed because of the large proportion gap between the classification groups. The results for the remaining factors are shown in In total, 1,017 cases were included following the strict standard, including 48 influencing factors. Among these factors, data on the Rh blood group, Chlamydia infection, gonorrhea, A binary logistic regression model was established with pregnancy outcomes as the dependent variable and the statistically significant variables mentioned above as independent variables. The results are shown in We constructed the prediction model based on the influencing factors presented in Assisted reproductive technology has been used in the clinic for many years. IVF-ET technology is also the last hope of many infertile couples. Although the clinical pregnancy rate of patients treated with IVF-ET can increase by up to 46.9%, the live birth rate is very low, around 38.1% only . Missed We found an abnormal coagulation function, such as the shortening of APTT and PT, in women with missed abortions, suggesting the hypercoagulable state of the blood. This might be because the hypercoagulable state of the blood can selectively affect blood circulation in the uterus and placenta, form microthrombus in the placenta, cause a local placental infarction, decrease placental blood supply, and cause ischemia and hypoxia in the embryo and fetus, thus arresting the growth of the embryo . TherefoThe hypothalamic-pituitary-ovarian axis regulates the complex endocrine system. Abnormalities in any part of the axis can lead to adverse pregnancy outcomes. In this study, we found that the level of PRL also increased significantly in the observation group, and hyperprolactinemia was common in pituitary dysfunction or space-occupying lesions in the pituitary. An increase in prolactin levels can inhibit the synthesis and release of gonadotropin, adversely affect the development of follicles and embryos, and cause ovulation disorders, resulting in infertility or missed abortion . Additioetc. Embryos formed by such gametes are at a greater risk of spontaneously stopping development (in vitro fertilization. Although China has implemented the three-child policy, the reproductive desire of young women has decreased, and the proportion of conception among relatively older women in China has increased. Therefore, improving the early pregnancy monitoring of elderly patients receiving ART is important.The age of the individual strongly affects the success of pregnancy. As the age of a woman increases, the quality of oocytes decreases, leading to errors during oocyte meiosis, the formation of aneuploidy, chromosome translocation, inversion, elopment . The reselopment . In our We also found that the level of FT4 in women with embryo termination was lower than that in normal people, and the positive rate of TPO-Ab was significantly higher, suggesting that thyroid dysfunction might affect embryos adversely. Hypothyroidism might cause adverse effects, such as embryo termination and fetal malformations . TPO-Ab The last influencing factor we discuss in this study is the immune factor. ACA and ANA are human autoimmune antibodies. We found that the level of these two antibodies was higher in women with missed abortions. ACA might act on the membrane phospholipids of placental vascular endothelial cells and platelets in the early stage of pregnancy, block prostacyclin synthesis, and lead to placental embolism. ANA might influence DNA replication and immune abnormalities . Only a Based on the above factors, we constructed an XGBoost-based missed abortion risk prediction model for patients treated with IVF-ET. The results showed that the prediction performance of the model was better than the prediction performance of the traditional logical regression model. Although many researchers have studied the risk factors for missed abortion, Along with our prediction model, when predicting the risk of missed abortion, clinicians should identify high-risk factors in advance, perform personalized prevention and treatment of these high-risk factors before IVF-ET treatment and during early pregnancy, and monitor patients after confirming clinical pregnancy to avoid a missed diagnosis, which might cause embryos to remain in the uterus for a long time.With the development of big data, similar prediction models based on machine learning might be constructed by more researchers. These studies might effectively deal with massive clinical databases and use an unbiased method to identify new information variables that are not easy for clinicians to find, to increase the convenience for personalized diagnosis and treatment platforms. However, its application still has some limitations. First, machine learning lacks human emotion and cannot completely replace the human brain in making decisions. Clinicians need to use it as a theoretical basis to make decisions according to the actual situation of the patients. Second, there is a problem of information leakage. Constructing a model requires patient information. Our health system needs to be improved and fortified to ensure the privacy and security of patients.One limitation of this study was that the inclusion index did not reach an ideal state. Embryonic chromosomes, bad living habits, a history of close contact with pets, and environmental factors were not included because this was a retrospective study. Some of the clinical data used had certain limitations, and standardizing the subjective indicators was challenging. A more accurate forecasting model needs to be constructed with all the indicators. Another limitation was that the study population included individuals from a small area and lacked geographical and ethnic variation. Future studies should consider including data from multiple medical centers for the external verification of the model.To summarize, we found that the independent influencing factors of missed abortion included female age, male age, abnormal ovarian structures, PRL, AMH, APTT, ACA, and TPO-Ab. We constructed a prediction model based on the XGBoost algorithm, which could be used to accurately predict the risk of missed abortion in patients with IVF-ET. This model performed better than the traditional logical regression model. The findings of this study might provide a theoretical basis for preventing abortion among women who are at a high risk of missed abortion.10.7717/peerj.14762/supp-1Data S1Click here for additional data file.


We conducted a systematic review of cohort studies comparing the risk of heart failure in people with differing metabolic health and obesity statuses. We searched three electronic databases , where the studies of the relationships of metabolic health and obesity statuses with heart failure were included. Fixed-effects or random-effects models were used to estimate the summary relative risks [RRs]. Ten cohort studies were selected. Compared with individuals with normal metabolic health and body mass, the pooled RRs  for heart failure were 1.23  for metabolic healthy overweight individuals, 1.52  for metabolic healthy individuals with obesity, 1.56  for metabolically unhealthy normal-weight individuals, 1.75  for metabolically unhealthy overweight individuals, and 2.28  for metabolic unhealthy individuals with obesity. A sensitivity analysis suggested that no single study had a substantial effect on the results. The Egger’s and Begg’s tests showed no evidence of publication bias. People with overweight or obesity were at a higher risk of heart failure, even if metabolically healthy. In addition, compared with metabolically healthy normal-weight individuals; metabolically unhealthy normal-weight individuals, and those with overweight or and obesity, were at higher risk of heart failure. Heart failure has become a global public health concern, with an estimated 64.34 million people being affected . Heart fThe relationships of obesity status and metabolic health status with the risk of heart failure have not been thoroughly evaluated. A cohort study by Voulgari et al. published in 2011 was the first on this topic. They reported that compared metabolically healthy normal-weight individuals, metabolically unhealthy normal-weight individuals are at a higher risk of heart failure, whereas MHO individuals are at a non-significantly lower risk . HoweverGiven the accumulating evidence and inconsistency of the results published to date, we aimed to conduct a systematic review and meta-analysis to compare the risks of heart failure among people with differing metabolic health and obesity statuses.We performed a meta-analysis that was designed, performed, and reported according to the Meta-analyses Of Observational Studies in Epidemiology (MOOSE) guidelines . We searA study was determined as eligible for inclusion in the meta-analysis when the following criteria were all met: (1) exposure defined using metabolic health status and obesity status, (2) outcomes including heart failure, (3) cohort design, and (4) risk estimates with 95% confidence intervals (CIs) for the relationship between the defined exposure and outcome were reported. The duplicate studies presented as the published articles titles, authors, and publication year were deleted, by using Endnote software, version 9, Clarivate, Philadelphia, US. In this process, three authors  worked independently to screen each record. After screening, the final study selection was completed by evaluating the full text of the selected ones, and any disagreements were resolved by discussion.X.W. and J.Y.D. reviewed the selected articles independently and discussed to generate a standard form to record the following data from each included study to describe their characteristics: the family name of the first author, year of publication, characteristics of the participants , sample size, study design, number of cases, duration of follow-up, assessments of metabolic health and obesity, assessments of heart failure, risk estimates with 95% CIs, and the results of the fully adjusted statistical model.Study quality was evaluated using the Newcastle-Ottawa Scale, which includes assessments of study sample selection, group comparability, and the assessment of exposures and outcomes. Each study could be awarded a maximum of 9 points, with ≥7 points indicating high quality and <7 points indicating low quality. Two authors (X.W. and J.Y.D.) independently conducted the quality assessment.2) and those in the single US study [2). The metabolically healthy normal-weight group was treated as the reference group for the analysis. Relative risks (RRs) and their 95% CIs were used as the common measures of the relationships, and hazard ratios or incidence rate ratios were converted to RRs. The heterogeneity of the RRs across the studies was tested using Cochran’s Q statistic and p < 0.10, and we also calculated the I2 statistic to estimate the degree of inconsistency across the studies [We defined six groups according to the metabolic health status and obesity status of the participants: a metabolically healthy normal-weight group, a metabolically healthy overweight group, a metabolically healthy obesity group, a metabolically unhealthy normal-weight group, a metabolically unhealthy overweight group, and a metabolically unhealthy obesity group. Three studies combined individuals with overweight or obesity; we regarded those in the two Asian studies ,20 as haUS study  as havin studies . Accordi studies . Subgrou studies . We used studies . StatistWe initially obtained 591 records from the three databases. The majority of the records were excluded by scanning the title and abstract, mainly because the research question was not relevant or because they were duplicates. A total of 13 studies were selected for full-text review. Of these, we excluded two studies ,29 that The characteristics of the included studies are shown in p for heterogeneity > 0.10), and a fixed-effects model was used to generate the results. In contrast, a random-effects model was used for the metabolically healthy obesity group (p for heterogeneity < 0.001). Compared with the metabolic healthy normal-weight group, the pooled RRs of heart failure were 1.23  for the metabolic healthy overweight group and 1.52  for the metabolically healthy obesity group.p for heterogeneity < 0.001), and a random-effects model was used for the pooled analysis. Compared with the metabolically healthy normal-weight group, the pooled RRs of heart failure were 1.56  for the metabolically unhealthy normal-weight group, 1.75  for the metabolically unhealthy overweight group, and 2.28  for the metabolic unhealthy obesity group.p for publication bias > 0.26).We next performed a sensitivity analysis by omitting one study in turn and combining the remaining data, and no single study was found to have a substantial effect on the overall results for any exposure . In addiIn the present meta-analysis of 8,092,751 individuals, we evaluated the relationships of metabolic health status and obesity status with the risk of heart failure. We found that people with overweight and obesity had approximately 23% and 52% higher risks of developing heart failure, even if they were metabolically healthy. Compared with the metabolically healthy normal-weight group, the metabolically unhealthy normal-weight group was associated with an approximately 56% higher risk of heart failure, and this risk increased to 75% and 128% for the metabolically unhealthy overweight and obesity groups.There is growing interest in the relationship between MHO and cardiovascular disease (CVD). Despite the absence of metabolic abnormalities in individuals with obesity, early changes may already be present that contribute to a higher risk of occurrence of CVD. MHO was found to be associated with abnormalities in several markers of subclinical CVD, including impaired vasoreactivity, high carotid artery intima-media thickness and aortic pulse wave velocity, high left ventricular mass and impaired function, and higher prevalences of coronary and aortic calcification ,33. PrevThe potential mechanisms involved in this increase in risk of heart failure include hemodynamic changes and changes in cardiac anatomy. Adipose tissue may affect the circulating blood volume, resulting in increases in cardiac output and ventricular stroke volume, thereby predisposing toward left ventricular enlargement, hypertrophy, and concentric remodeling, all of which are linked to the development of heart failure ,38. ThusWe found that metabolically unhealthy individuals with normal weight were still at a higher risk of heart failure, and this group had a similar estimated risk to the MHO group (approximately 50%) in the present study. Previous prospective cohort studies have shown that the risk of CVD in metabolically unhealthy normal-weight individuals are at even higher risk than those with MHO ,42,43. TMetabolic health status and obesity status may change during a study. In most studies, a single baseline measurement of exposure factors was made, and we found only two studies ,31 that Several limitations of this study should be acknowledged. First, heterogeneity is a common limitation of meta-analyses. In the present study, considerable heterogeneity was identified in the analyses of the metabolically unhealthy groups. Only the studies by Caleyachetty et al.  and FaucIn conclusion, the present meta-analysis of previous cohort studies showed that people with overweight or obesity are at a higher risk of heart failure, even if they are metabolically healthy. Specifically, compared with metabolically healthy normal-weight individuals, metabolically unhealthy normal-weight individuals and those with overweight or obesity were found to be at a higher risk of heart failure. Therefore, the promotion of both metabolic health and weight management may represent an important means of reducing the incidence of heart failure.


Ficus carica (FC) extract in rats of two-generation F0 (parents) and F1 (offspring) in either sex. The F. carica extract was initially tested for acute and sub-chronic toxicity. Extracts were also tested for fertility assessment and effects on reproductive hormones like follicle-stimulating hormone (FSH), luteinizing hormone (LH), gonadotropin-releasing hormone (GnRH), estradiol, testosterone, dihydrotestosterone (DHT), dehydroepiandrosterone-sulfate (DHEAS), insulin, progesterone, and prolactin. The antioxidant activity of the extract was also evaluated by testing mRNA expressions of SOD2, GPX1, CAT, and GR in male testes and female ovaries. The animals treated with 100 mg/kg FC extract produced a more pronounced fertility effect in both genders. Expression of CAT, SOD2, GPX1, and GR was found to be increased in the reproductive organs of both sexes. Histology of the testes reveals increased spermatogenesis and increased folliculogenesis in ovaries. The hormone profile showed an increase in FSH, DHT, estradiol, and DHEAS levels in males and FSH, LH, estrogen, and DHEAS in females. The results of the study establish the effectiveness of natural products in improving fertility issues in either sex.The basic purpose of pharmacology is to look into the benefits of natural remedies and make them available to the general populace. Herbal medicines are now considered to be the future of humanity. The current study explored the effects of  Ficus carica Linn. (figs) had been traditionally used as a laxative and for other medicinal benefits related to cardiovascular, respiratory, spasmolytic, and anti-inflammatory effects [F. carica (Moraceae) have been assessed for hepatoprotective effect in carbon tetrachloride-induced liver toxicity in rats [F. carica contains three hydroxycinnamic acids (3-O- and 5-O-caffeoylquinic acids and ferulic acid), two flavonoid glycosides (quercetin 3-O-glucoside and quercetin 3-O-rutinoside), and two furanocoumarins . Except for ferulic acid, all other phenolic compounds are present in the leaf, pulp, and peel [F. carica is rich in amino acids including five essential amino acids histidine, leucine, lysine, phenylalanine, and tryptophan, and eight nonessential amino acids like alanine, and asparagine, cysteine, glutamine, glycine, ornithine, serine, and tyrosine. Cysteine, tryptophan, and tyrosine are found in greater concentrations than other amino acids [F. carica also contains coumarin which possesses anticancer, antianemic, and antioxidant activities. It has an alkaloid (quinine) that possesses antimalarial activity, anthocyanin  with antioxidant and radical scavenging actions, as well as hydrocarbon which possesses antioxidant, hemoptysis, and antiseptic activities [F. carica on the liver, kidney, hematological parameters, and fertility in male and female rats. effects . The lea in rats . The the in rats . The use in rats . It seem in rats . F. cariand peel . F. carino acids , 6. F. cThe research was executed after the consent from the Board for Advanced Studies and Research, the University of Karachi, allocated number 04642/pharm, dated September 2019, and followed by the permission of the DRC, Department of Pharmacology to use the animals as per the guidelines of the NIH.F. carica (FC) fruits were procured from a fruit shop in Karachi; Prof. Huma Shareef at the Department of Pharmacognosy Institute of Pharmaceutical Sciences, JSMU, Karachi identified it. The 70% hydro ethanol extract of FC was prepared as per the method of Elghareeb [F. carica extract was 25% (w/w) [The albino Wistar rat used in the study were aged 11–13 weeks  and raised in the animal's house of ICCBS, University of Karachi. Animals weigh about 170 to 200 grams. The rats were housed in a room maintained at 25°C; with 45% humidity and exposed to a natural daily 12 hr light/dark cycle. The rats were retained in the standard cages in the animal house. Pelleted rodent food and filtered tap water was provided ad libitum. The rats were acclimatized for two weeks. Females were subjected to vaginal smear examination for two weeks before treatment. The vaginal samples were collected by separating the lips of the female genital organ and placing them in a sterile cotton swab into the vagina. The swab was turned two to three times against the vaginal wall to collect the smear and moved on a clean glass slide; the smears were air-dried and stained with Wright Giemsa stain and observed by the microscope (×100 magnification) . FemalesThe acute toxicity study was conducted following OECD Guideline-423 a healthy albino Wistar female rat with 170–200 gm weight was used in the present study. The experiment was conducted on three female rats. Under fasting conditions, the rats were examined following the administration of a single dose of 500 mg/kg FC extract; food was suspended for another 4 hours after administration of the extract. Toxicity markers were noted for the first 24 hours, every hour, and each day for 14 days from the start of the study. The rats were under surveillance for 14 days and observed body weight, posture, signs of seizure, limb paralysis, body tone, gross behavioral changes, and mortality .Wistar rats in three groups of ten animals  were given 70% FC hydroethanol extract by mouth in doses of 50 mg/kg and 100 mg/kg as a single daily dose for 90 days. The control animals were given the vehicle. All rats were kept in the same environments, with food and water available at all times, throughout the study. Toxicity was evaluated through gross behavior, blood chemistry, and enzyme analysis. The plasma was obtained by collecting blood in EDTA tubes which were then used for blood analysis chemistry and enzyme analysis .Virgin female albino Wistar rats aged 13 weeks and male albino Wistar rats aged 11 weeks were divided into 3 groups. Each group consists of six males and six females. Dosing was started 70 days before the mating and continued throughout the mating in males and continued in females throughout gestation and lactation. Three days before mating, the female rats were exposed to the soiled bedding of a mature male rat to synchronize their estrous cycles . Day-1 p1 generation offspring when weaned and throughout their growth and maturation to adulthood, as well as during the breeding. Females of the F1 generation who were pregnant continue to get the FC extract throughout their pregnancy until the F2 generation offspring were weaned. From each litter, two males and two females were chosen at random; for mating with another pup of a different litter at the same dose, and level to produce the next generation. One-week acclimatization period was given before the exposure to FC extract to the F1 generation. The groups were formed in a pattern similar to the F0 generation and mated after 12 weeks of maturity, males received dosing throughout mating, and females received dosing throughout the gestational and lactation period. Each litter was examined immediately on delivery for the number of pups, stillbirths, live births, and the presence of overall defects. The neonates were cautiously witnessed on postnatal days zero (birth), four, seven, fourteen, and 21 days. Dead pups were grossly scrutinized for probable defects and the cause of disease. Reproductive toxicity endpoints show the animal's retorts to the FC extract from conception to weaning. This analysis reflects the total effect of the FC extracts at all stages, which is a more profound indicator for reproduction studies. Animals of the F0 and F1 generations were divided into three groups as follows:The control group comprised six males and six females given once daily 1 ml of distilled water by mouth.Treated groups were given 50 mg/kg and 100 mg/kg of FC extract by gastric gavage tube.The exact doses of the FC extract were given to certain FBlood samples for hormonal measurement were obtained during the estrus phase. The cardiac puncture procedure was used to collect blood samples. The blood was allowed to stand for 10 minutes in centrifuge tubes to clot at area temperature and centrifuged at 3500 rpm for 10 minutes. The serum was transferred to a different container, and the sera were stored at−80°C. ELISA was used for the assessment of testosterone, LH, FSH, estrogen, progesterone, prolactin, DHT, GnRH, DHEAS, and Insulin.μm thickness, and the mounted sections on a glass slide were deparaffinized by xylene. Ethanol series  and distilled water were used for rehydration and stained using Hematoxylin and Eosin. Stained microscopic slides were evaluated and compared in groups. The numbers of preantral, antral, corpus luteum, corpus Albicans and cystic follicles, and the stromal reaction were determined [Ovaries of the control group were removed during the estrus phase. Excised ovaries were protected in 10% formalin after cleaning the adherent connective tissues, then processed by usual procedures and embedded in paraffin. The ovaries were longitudinally and serially sectioned at five termined .The testes of the selected rats from all the groups in the study were detached and weighed. The testes were fixed in Bouin's fluid at room temperature for 24 h, tan tissues were handled routinely. For a short, while the tissues were transferred to 70% alcohol and dried by moving through ascending grades of alcohol; the tissues were then cleared in xylene and lastly embedded in paraffin wax 5 m sections were cut using a rotary microtome and stained with hematoxylin and eosin. The stained slides were seen in the microscope and images were taken.μg of total RNA. The oxidative stress enzyme indicators, i.e., CAT, SOD2, GPX1, and GR expressions in ovaries and testes were evaluated by RT-qPCR. The total reaction mixture volume was 25 microliters with thermal cycling in three steps. The reaction mixture was first gone through denaturation for 10 min at 95°C then again denatured for 15sec at 95°C followed by annealing for 30 sec at 60°C and final extension for 30 sec at 72°C. Denaturation, annealing, and extension comprised 40 cycles. Thermo Scientific Maxima SYBR Green/ROX qPCR master mix (2X) kit was used and the parallel sense and antisense were shown in RNA from the ovaries and testes was extracted by the triazole method. Reverse Aid First-strand complementary DNA (cDNA) synthesis kit (Thermo Scientific) was used for the synthesis of first-strand cDNA from 1 0 and F1 generations hormone profiles in each studied group. These means were matched by one-way ANOVA, and post hoc analysis was done using Tukey's test for multiple comparisons. P < 0.05 was deliberated statistically significant, and P < 0.01 was highly significant. Data were also compared for male and female samples separately in F0 and F1 generation parameters.Data were stored and analyzed using IBM-SPSS version 23.0. Mean with standard error to the mean was reported for liver, cardiac, kidney markers, blood profiles, and FF. carica extract in the dose of 500 mg/kg survived during 14 days of observation. Oral administration of this extract at 500 mg/kg did not yield any substantial changes in behavior, breathing, skin effects, defecation, loss of hair, postural abnormalities, lacrimation, salivation, loss in food intake, reflex sensitivity, and body weight of female rats.All animals were given the F. carica extract at 50 and 100 mg/kg exerted no side effects during the 90 days of the treatment period. However, there was an increase in animal body weight as compared to the control after 90 days. The percentage weight change in the males treated with 50 mg/kg was 28–34% and those treated with 100 mg/kg was 31–42%, while in females the percentage weight change at the same doses was 20–30%, and 25–35%, respectively, as compared to control.The 0 and F1 generation of albino Wistar rats following the administration of the two doses against a control group. The F0 generation  in the liver [F. carica. Whereas, in the males of the F1 generation, a highly significant increase in the levels of FSH, LH, and DHEAS was observed in a dose-dependent manner. GnRH, estradiol, DHT, testosterone, progesterone, and insulin were comparable to control animals. F. carica increased the levels of FSH more than that of LH, DHT, and estrogen which is therefore responsible for the increase in fertility in parent males. The outcomes of the present study are in accordance with a previous study by Haredy et al. [F. carica and increased endogenous activity of antioxidant enzymes (CAT and SOD2) and semen quality, while there was a significant decrease in markers of oxidative stress in testes, i.e., NO, 8OHdG, MDA and GSSG. Thus, it may be concluded that increased gene expression tends to reduce oxidative stress and in turn increases overall reproductive activity.Both FSH and LH enhance testicular growth in males while estradiol, DHT, and DHEAS are essential for libido, erectile function, and spermatogenesis in males . The finhe liver . Hence wy et al. , who repF. carica increases testosterone production and has a positive effect on FSH. This indicates that F. carica has the potential to increase spermatogenesis in males which was also evident by histological changes in testes as well as fertility data in both generations showing conception and copulation more efficiently without any toxicity in the groups treated with F. carica in a dose-dependent manner.Animals treated with 100 mg/kg FC showed increased expression of antioxidants in the testes with an increase in the conversion of testosterone into estrogen and DHT, while there was also an increase in FSH levels, Abu Bakar et al.  showed tF. carica has significantly increased insulin in males of the F0 generation and may be responsible for increased fertility due to elevated levels of FSH and LH.The previous study shows that in insulin-dependent diabetes testosterone synthesis and function of Leydig cells are decreased due to the lack of insulin, thus reducing LH levels, sperm output, fertility, and FSH . However0 generation, there was a highly significant increase in the levels of FSH and estradiol, while LH was increased significantly in both groups treated with F. carica, whereas there was a significant increase in DHEAS level at 100 mg/kg and a highly significant increase in prolactin levels. In contrast, females of the F1 generation showed a highly significant increase in the levels of FSH at 100 mg/kg and a highly significant increase in estradiol and DHEAS in both the groups treated with F. carica in a dose-dependent manner.In females of the FF. carica increased the level of FSH, LH, and estrogen in parents and offspring which indicates that the treatment with F. carica increases the proliferation of follicles which is also evident by histology and fertility data; since copulation occurred in less time, these results were repeated in F1 generation in a dose-dependent manner. The expression of SOD2, GPX1, CAT, and GR was increased in ovaries in the group treated with F. carica 100 mg/kg except for the expression of GPX 1 in F0 generation; this shows the antioxidant potential of the extract. F. carica increased prolactin levels in the female which indicates that it has the potential to increase milk production in lactating mothers.FSH is crucial for the survival, propagation, and distinction of preovulatory mural granulosa and excites the expression of hundreds of genes, comprising those required for the production of estradiol and expression of the LH receptor . The higF. carica extract resulted in a highly significant increase in DHEAS levels and hence may be effective in the management of andropause in men. Andropause is a steady process nearly similar to menopause in women and occurs with a steady and persistent drop in testosterone and dehydroepiandrosterone production in men [F. carica extract may also be used in the management of menopausal women as it enhances the levels of estrogen and DHEAS. Both age and declining estradiol levels had significant detrimental effects on sexual functioning, libido, and sexual responsiveness in females. The drug is effective in dengue as it increases the levels of platelets. A hallmark of dengue infection is thrombocytopenia which is associated with abnormal platelet function [The n in men . The F. function .F. carica is effective in enhancing conception and fertility in males and females by promoting ovulation and spermatogenesis. It also showed increased gene expression of antioxidant enzymes in both F0 and F1 generation producing an overall supportive effect in the reproduction process of both genders without any harmful effect on hematic, renal, and hepatic parameters. Results show that it may also be effective in andropause men and menopausal women. F. carica may also be effective in the management of dengue fever since has increased platelet and RBC count. However, the study has some constraints such as a lack of human studies which are essential for the general use of any drug in humans. Nonetheless, human studies may be performed prospectively to endorse the safety of F. carica extract in humans.


In patients with COVID-19 sEng did not predict mortality or correlate with markers of organ dysfunction. In contrast, in septic shock the level of sEng was significantly higher in patients with early mortality . Moreover, sEng levels correlated with signs of circulatory failure . The predominant clinical problem in patients with COVID-19 was ARDS, and although they often showed signs of other organ dysfunction, circulatory failure was exceptional. This potentially explains the difference between sEng levels in COVID-19 and septic shock. In conclusion, we have confirmed that sEng may reflect the extent of the circulatory failure in septic shock patients and thus could be potentially used for the early identification of patients with the highest degree of endothelial dysfunction who would benefit from endothelium-targeted individualized therapy.Sepsis is a clinical syndrome characterized by a dysregulated response to infection. It represents a leading cause of mortality in ICU patients worldwide. Although sepsis is in the point of interest of research for several decades, its clinical management and patient survival are improving slowly. Monitoring of the biomarkers and their combinations could help in early diagnosis, estimation of prognosis and patient's stratification and response to the treatment. Circulating soluble endoglin (sEng) is the cleaved extracellular part of transmembrane glycoprotein endoglin. As a biomarker, sEng has been tested in several pathologic conditions where its elevation was associated with endothelial dysfunction. In this study we have tested the ability of sEng to predict mortality and its correlation with other clinical characteristics in the cohort of septic shock patients ( Under the current definition, sepsis is described as a life-threatening organ dysfunction that is caused by a deregulated immune response to infection . Sepsis COVID-19, caused by the SARS-CoV-2 virus, progresses in some patients to very severe respiratory failure due to ARDS (an incidence rate of ~5%) , 7. AlthEndoglin (Eng) is an adhesion molecule, a sub-unit of the receptor system for TGF-β. It is the most frequently localized on the surface of endothelial cells, but its expression has been demonstrated in many other cell types, such as the cells of innate and adaptive immunity , 13. TheRecently, we proved that sEng can be potentially used as a prognostic biomarker in patients suffering from septic shock . In COVIIn this trial, we sought to test the prognostic potential (in prediction of early (D3), D28 and/or D90 mortality) of sEng in a larger and more precisely defined cohort of patients suffering from septic shock and the cohort of patients with a severe COVID-19 and compare the result between these two diagnoses. Secondly, we wanted to determine whether sEng levels are correlated with other laboratory and clinical characteristics of patients including the source of sepsis, and thus bring new insights into sEng pathophysiology and function in critical infectious states.The trial included patients admitted to the intensive care unit (ICU) of the Department of Anesthesiology and Intensive Care at St. Anne's University Hospital in Brno with sepsis requiring administration of catecholamines (the septic shock cohort) and with a severe course of COVID-19 (the COVID-19 cohort). The trial was approved by the local Ethics Committee and patients were enrolled on the basis of informed consent. A total of 37 consecutive septic shock patients hospitalized in year 2019 were included in the septic shock cohort. All patients were treated according to the current recommendations for sepsis treatment . InclusiA total of 40 patients were included in the COVID-19 cohort, who were admitted to the ICU with a severe course of COVID-19 during the 2021 global pandemic. All enrolled patients required artificial pulmonary ventilation at the time of admission. Exclusion criteria were the same for both cohorts: ongoing cancer, chronic immunosuppressive therapy, age under 18 years, and current pregnancy.Venous blood samples were collected from patients during the first 24 h after ICU admission. Immediately after collection, the samples were transported to the laboratory and centrifuged. The plasma obtained in this manner was frozen and stored at −80°C until it was analyzed. The plasma sEng concentration was measured in triplicate using the Human Endoglin/CD105 Quantikine ELISA Kit . The absorbance of the samples was measured using a Multiscan GO Microplate Spectrophotometer (Thermo Scientific) at 450 nm (wavelength correction of 540/570 nm) and the average value for three samples was calculated. Levels of routinely used biomarkers (C-reactive protein (CRP), creatinine, lactate, bilirubin) were analyzed by standard techniques in a hospital laboratory as part of routine daily patient diagnostics.p < 0.05. Spearman's rank correlation coefficient was used to measure the strength of the relationship between the two variables , noradrenalin dose, leukocytes, lactate, CRP, creatinine, bilirubin and oxygenation index. The Mann–Whitney U-test was used to test the difference in continuous variables between the two groups; the Kruskal-Wallis test was used for 3 or more groups; and Fisher's exact test was used to test for statistical dependence.A statistical analysis was performed in the R programming language  in the R studio environment (version 1.4.1106). The data are presented as the median (1st quartile−3rd quartile). The statistical significance was set at A total of 37 patients were enrolled in the septic shock cohort and 40 in the COVID-19 cohort. Patient characteristics are described in 9/L] and elevated lactate levels [2.7 (2.0–4.8) vs. 1.6 (1.2–2.1) mmol/L] at admission. There were no other statistically significant differences between the groups. All patients were mechanically ventilated upon admission and the SOFA score at admission was 11 (10–14). The most common comorbidities were obesity, diabetes mellitus, and ischemic heart disease and peripheral ischemic disease. The most common source of sepsis was pneumonia .The deceased patients were significantly older p74.5 (72.3–79.5) vs. 68.0 (66.0–75.5)] and had more pronounced leukocytosis .p = 0.210; p = 0.514, respectively; p = 0.019]. The sEng level did not correlate with most clinical data ; urosepsis and soft tissue infections had a higher median sEng than pneumonia, abdominal and chest infections ; organ failure ; circulatory failure ; severity of condition ]. Similarly to sepsis, a severe course of COVID-19 represents a generalized response to infection. However, a shock condition, in the sense of circulatory failure with hypotension and the need for higher doses of catecholamines and severe endothelial dysfunction, is usually not manifested here. Our data show that in the septic cohort, sEng level correlated with the degree of circulatory failure. Therefore, we suggest that in septic shock, sEng primarily reflects the degree of circulatory failure based on endothelial dysfunction and is released mainly from endothelium. Moreover, in patients with a fulminant course of sepsis , refractory circulatory failure is usually the dominant clinical problem. This is consistent with our finding of significantly higher sEng levels in the early deceased patients. Conversely, in the COVID-19 cohort, sEng did not correlate with mortality, lactate or the catecholamine dose –C. In seNeither from our data nor from the existing evidence, we are able to say unequivocally that the elevation of sEng accurately reflects the degree of endothelial dysfunction. Undoubtedly, there are multiple mechanisms that can increase sEng levels in such a complex syndrome as sepsis . For exaThe fact that sEng is elevated in early deceased septic patients raises the important question whether it is released only as a consequence of the severe condition or it plays some specific role in the increased mortality and could be a potential target for new therapies. To date we are not able to answer this clearly. However, there is evidence that sEng could have a detrimental effect. For example, sEng reduces the activity of the endothelial nitric oxide synthetase (eNOS) and subsequently the natural production of nitric oxide (NO), reduces endothelial turnover, increases the capillary permeability in lungs, kidneys and liver and increases leucocyte adhesion , 35. MorChanges in sEng levels have been described in association with a variety of comorbidities, in particular, higher comorbidities associated with atherosclerosis, hypertension, diabetes mellitus (DM), obesity, and dyslipidemia . In contThe dependence of sEng levels on the source of sepsis in the septic cohort is the novel and also interesting result. Patients with a urinary tract infection had the highest sEng levels. The expression of Eng by the urothelium has not been found in any published study to date, but an increased expression by the endothelium has been found in cases of urothelial carcinoma, for example . In gene9/L; CRP 151.1 vs. 237.5 mg/L).One limitation of the trial is its monocentric nature and the relatively small numbers of patients included in each cohort. Although the primary objective was not to compare the cohorts with each other, the differences in patients in the septic vs. COVID-19 cohorts pose a limitation too. The COVID-19 patients were younger and had fewer comorbidities compared to the patients with bacterial sepsis. In the COVID-19 patients, severe respiratory failure with a worse oxygenation index was evident (87.5 vs. 143.5); however, the dysfunction of other organs was already expressed to a much lesser extent than in those patients with bacterial sepsis  mortality , 44, 45.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by Local Ethics Committee of the Saint Anne's University Hospital in Brno, Czech Republic. The patients/participants provided their written informed consent to participate in this study.VT, MH, and KH: study preparation and design. VT and MH: patient recruitment and sample collection. MS and MŠ: data curation. AM, MHK, and OM: lab analyses. VT and AM: writing—original draft preparation. MH, JF, VŠ, and JP: writing—review and editing. VŠ and JP: supervision. JF: funding acquisition. All authors have read and agreed to the published version of the manuscript.This work has been supported by the Ministry of Health of the Czech Republic (Grant No. NU21-06-00408) and DRO . JF was supported by the European Social Fund and European Regional Development Fund, Project MAGNET (No. CZ.02.1.01/0.0/0.0/15_003/0000492). The funders had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.