Construction of an ER stress-related prognostic signature for predicting prognosis and screening the effective anti-tumor drug in osteosarcoma.

BACKGROUND: Osteosarcoma is the most common malignant primary bone tumor in infants and adolescents. The lack of understanding of the molecular mechanisms underlying osteosarcoma progression and metastasis has contributed to a plateau in the development of current therapies. Endoplasmic reticulum (ER) stress has emerged as a significant contributor to the malignant progression of tumors, but its potential regulatory mechanisms in osteosarcoma progression remain unknown. METHODS: In this study, we collected RNA sequencing and clinical data of osteosarcoma from The TCGA, GSE21257, and GSE33382 cohorts. Differentially expressed analysis and the least absolute shrinkage and selection operator regression analysis were conducted to identify prognostic genes and construct an ER stress-related prognostic signature (ERSRPS). Survival analysis and time dependent ROC analysis were performed to evaluate the predictive performance of the constructed prognostic signature. The "ESTIMATE" package and ssGSEA algorithm were utilized to evaluate the differences in immune cells infiltration between the groups. Cell-based assays, including CCK-8, colony formation, and transwell assays and co-culture system were performed to assess the effects of the target gene and small molecular drug in osteosarcoma. Animal models were employed to assess the anti-osteosarcoma effects of small molecular drug. RESULTS: Five genes (BLC2, MAGEA3, MAP3K5, STC2, TXNDC12) were identified to construct an ERSRPS. The ER stress-related gene Stanniocalcin 2 (STC2) was identified as a risk gene in this signature. Additionally, STC2 knockdown significantly inhibited osteosarcoma cell proliferation, migration, and invasion. Furthermore, the ER stress-related gene STC2 was found to downregulate the expression of MHC-I molecules in osteosarcoma cells, and mediate immune responses through influencing the infiltration and modulating the function of CD8+ T cells. Patients categorized by risk scores showed distinct immune status, and immunotherapy response. ISOX was subsequently identified and validated as an effective anti-osteosarcoma drug through a combination of CMap database screening and in vitro and in vivo experiments. CONCLUSION: The ERSRPS may guide personalized treatment decisions for osteosarcoma, and ISOX holds promise for repurposing in osteosarcoma treatment.

Impact of non-alcoholic fatty liver disease and fibrosis on mortality and kidney outcomes in patients with type 2 diabetes and chronic kidney disease: A multi-cohort longitudinal study.

AIM: To evaluate the impact of non-alcoholic fatty liver disease (NAFLD) presence and fibrosis risk on adverse outcomes in patients with type 2 diabetes and chronic kidney disease. METHODS: Data were sourced from two longitudinal cohorts: 1172 patients from the National Health and Nutrition Examination Survey (NHANES) and 326 patients from the kidney biopsy cohort at the West China Hospital of Sichuan University. Cox regression estimated hazard ratios (HRs) for NAFLD and liver fibrosis concerning adverse clinical outcomes. Subsequently, a two-sample Mendelian randomization study using genome-wide association study statistics explored NAFLD's potential causal link to cardio-cerebrovascular events. RESULTS: In the NHANES cohort, NAFLD stood as an independent risk factor for various outcomes: overall mortality [HR 1.53 (95% confidence interval, CI 1.21-1.95)], mortality because of cardio-cerebrovascular diseases [HR 1.63 (95% CI 1.12-2.37)], heart disease [HR 1.58 (95% CI 1.00-2.49)], and cerebrovascular disease [HR 3.95 (95% CI 1.48-10.55)]. Notably, advanced liver fibrosis, identified by a fibrosis-4 (FIB-4) score >2.67, exhibited associations with overall mortality, cardio-cerebrovascular disease mortality and heart disease mortality. Within the kidney biopsy cohort, NAFLD correlated with future end-stage kidney disease [ESKD; HR 2.17 (95% CI 1.41-3.34)], while elevated FIB-4 or NAFLD Fibrosis Scores predicted future ESKD, following full adjustment. Liver fibrosis was positively correlated with renal interstitial fibrosis and tubular atrophy in biopsies. Further Mendelian randomization analysis supported a causal relationship between NAFLD and cardio-cerebrovascular events. CONCLUSIONS: In patients with type 2 diabetes and chronic kidney disease, the NAFLD presence and elevated FIB-4 scores link to heightened mortality risk and ESKD susceptibility. Moreover, NAFLD shows a causal relationship with cardio-cerebrovascular events.

Estimated small dense low-density lipoprotein-cholesterol and the risk of kidney and cardiovascular outcomes in diabetic kidney disease.

AIMS: This study aimed to investigate the correlations between estimated small dense low-density lipoprotein-cholesterol (esd-LDL-c) and the development of end-stage kidney disease (ESKD), cardiovascular mortality, and all-cause mortality in individuals with diabetic kidney disease (DKD) or diabetes mellitus (DM) concomitant chronic kidney disease (CKD). METHODS: We analyzed the data from a biopsy-proven DKD cohort conducted at West China Hospital of Sichuan University between 2009 and 2021 (the DKD cohort) and participants with DM and CKD in the National Health and Nutrition Examination Survey (NHANES) 2011-2014 (the NHANES DM-CKD cohort). Cox regression analysis was also used to estimate associations between esd-LDL-c and the incidence of ESKD, cardiovascular mortality, and all-cause mortality. RESULTS: There were 175 ESKD events among 338 participants in the DKD cohort. Patients were divided into three groups based on esd-LDL-c tertiles (T1 < 33.7 mg/dL, T2 ≥ 33.7 mg/dL to <45.9 mg/dL, T3 ≥ 45.9 mg/dL). The highest tertile of esd-LDL-c was associated with ESKD (adjusted HR 2.016, 95% CI 1.144-3.554, p = .015). Furthermore, there were 99 deaths (39 cardiovascular) among 293 participants in the NHANES DM-CKD cohort. Participants were classified into three groups in line with the tertile values of esd-LDL-c in the DKD cohort. The highest tertile of esd-LDL-c was associated with cardiovascular mortality (adjusted HR 3.95, 95% CI 1.3-12, p = .016) and all-cause mortality (adjusted HR 2.37, 95% CI 1.06-5.32, p = .036). CONCLUSIONS: Higher esd-LDL-c was associated with increased risk of ESKD in people with biopsy-proven DKD, and higher cardiovascular and all-cause mortality risk among those with DM-CKD.

Novel Magnetic-Bead-Assisted Sequential Extraction Method for Liquid Chromatography-Mass Spectrometry (MS)/MS of Components with Diverse Properties: Gastrin Determination as a Case Study.

Sample preparation is the rate-limiting step in liquid chromatography-mass spectrometry (LC-MS)/MS-based clinical analysis when target analytes possess significantly different properties. Repeated solid-phase extraction (SPE) processes are typically required, resulting in low throughput and excessive consumption of labor, materials, and samples. In this study, we developed and validated a feasible and productive method to enrich target analytes with different properties during a single operation, while sufficiently removing matrix interferences to meet LC-MS/MS requirements. Gastrin determination was selected as the subject of this study. An automated magnetic-bead-assisted sequential extraction (MBASE) workflow was developed to simultaneously isolate nonsulfated gastrin-17 (G17ns), sulfated gastrin-17 (G17s), nonsulfated gastrin-34 (G34ns), and sulfated gastrin-34 (G34s) from human serum. It performs two different ion-exchange-based magnetic-bead extraction steps on one sample aliquot to produce one combined extract for LC-MS/MS analysis. When compared with the traditional SPE process, the MBASE workflow saves over 75% time and labor expenses as well as over 90% material cost, while providing even higher extraction efficiency. The MBASE LC-MS/MS method was validated as accurate and robust. Clinical sample test results demonstrated that the conventional chemiluminescence immunoassay method significantly under-estimated total gastrins in human serum, and the MBASE LC-MS/MS method could serve as an ideal tool to provide a comprehensive and accurate gastrin profile.

A robust LC-MS/MS method to measure 8-oxoGuo, 8-oxodG, and NMN in human serum and urine.

OBJECTIVE: To establish and validate a robust LC-MS/MS method for simultaneously measuring 8-oxoGuo, 8-oxodG, and NMN in serum and urine to evaluate the oxidative stress status. METHODS: A Waters TQ-XS triple quadrupole mass spectrometer system coupled with an Acquity UPLC Primer HSS T3 column was chosen. The clinical performance was verified according to the CLSI C62-A and EP-15 guidelines. Furthermore, matched serum and urine samples from 22 apparently healthy check-ups, 20 patients with atherosclerosis, and 18 individuals with dementia were evaluated. RESULTS: The recovery for serum 8-oxoGuo, urine 8-oxoGuo, serum 8-oxodG, urine 8-oxodG, serum NMN, and urine NMN was 88.8-112.4%, 102.4-114.1%, 88.5-107.7%, 94.9-102.6%, 98.4-108.9%, and 88.5-108.6%, respectively. Based on the inter-assay results, total coefficient of variation, matrix effect, and carryover, the LC-MS/MS method was deemed robust. The limit of quantification was 0.017, 0.018, and 0.150 nmol/L for 8-oxoGuo, 8-oxodG, and NMN, respectively, which are suitable for accurate measurements in human serum and urine samples. Higher 8-oxoGuo and 8-oxodG levels and lower NMN levels, indicative of significantly higher oxidative stress status, were found in patients with dementia compared to healthy subjects. CONCLUSION: We established and validated a robust LC-MS/MS method to simultaneously measure 8-oxoGuo, 8-oxodG, and NMN in serum and urine.

Total serum vitamin B12 (cobalamin) LC-MS/MS assay as an arbiter of clinically discordant immunoassay results.

OBJECTIVES: Measurement of the serum levels of vitamin B12 (VB12) is key for evaluating VB12 deficiency-dependent anemia. Immunoassay, the major method for determining VB12, tends to give false-normal results because of the presence of anti-intrinsic factor (IF-Ab) or other factors such as heterophilic antibodies et al. This study aimed to develop a liquid chromatography tandem mass spectrometry (LC-MS/MS) method that is helpful for distinguish false normal VB12 results measured by the immunoassay. METHODS: Different forms of VB12 were derivatized into CN-B12, which was collected through solid-phase extraction and analyzed via LC-MS/MS. 236 serum samples were measured both by LC-MS/MS and immunoassay, results were compared, and the IF-Ab effect was evaluated. RESULTS: The LC-MS/MS assay afforded a linear slope from 20 to 4,000 pmol/L for CN-B12. OH-VB12, methyl-VB12, and CoA-VB12 showed recovery within 89.3-109.5%. The intra-assay CV of VB12 was 2.6-4.1%, whereas the total CV was 9.3-9.8%. Passing-Bablok regression between LC-MS/MS and immunoassay results showed that the slope was 1.085 and the intercept was -15.691. The Bland-Altman plot showed that the mean difference and difference% were -34.6 pmol/L and 0.3%, respectively. Inter-rater agreement analysis showed that the linear weighted kappa value was 0.885, implying good agreement between the two methods. However, two samples were falsely elevated and one sample was falsely normal in the immunoassay compared with LC-MS/MS. The LC-MS/MS method helped in the distinction of false-normal VB12 results shown by the immunoassay. CONCLUSIONS: The VB12 LC-MS/MS method can be used as an arbiter of clinically discordant immunoassay results.

Prediction of potential hard sodium carbaboride compounds assuming sp3-bonded covalent clathrates.

Boron-carbon clathrates have attracted great attention due to their unique sp3-bonded structure and excellent electronic properties. Here, by performing first-principles calculations, we predicted six stoichiometric Na-B-C clathrates (NaBC11, Na2B2C10, NaB2C10, Na2B4C8, NaB4C8, and Na2B6C6) based on Na-doped boron-carbon clathrates. As a result, NaBC11, Na2B2C10, and NaB2C10 were found to become energetically favorable. Under ambient conditions, the electronic structure calculations show that NaBC11 and Na2B2C10 are indirect band gap semiconductors, and NaB2C10, Na2B4C8, and NaB4C8 exhibit metallic features. Na2B2C10 and Na2B4C8 are found to be synthesized at 22.7 and 14.2 GPa, respectively. Interestingly, the formation enthalpies of NaxB2C10 and NaxB4C8 (x = 0, 1, and 2) clathrates decrease in turn with the increased number of Na atoms in the same synthetic paths. Moreover, the ideal indentation strengths of NaBC11, Na2B2C10, and NaB2C10 approach 40 GPa, indicating that they are hard materials with superior hardness. These findings offer valuable insights for advancing the synthesis of boron-carbon clathrates.

High Thyrotropin Levels and Risk of Mortality in the Elderly With Subclinical Hypothyroidism: A Systematic Review and Meta-analysis.

OBJECTIVE: This study aims to determine whether elevated endogenous thyrotropin levels contribute to an increased risk of adverse outcomes, such as all-cause mortality in older adults with subclinical hypothyroidism. METHODS: Eight electronic databases were searched for relevant articles from inception until March 23, 2022. Cohort studies assessing the association between thyrotropin levels and the risk of mortality among older adults aged ≥60 years with subclinical hypothyroidism were eligible. The outcomes of interest were either all-cause or cardiovascular-related mortality. Two independent researchers assessed the eligibility of the studies and collected data through a previously defined data extraction form. The Newcastle-Ottawa Scale was used to evaluate the quality of evidence, and multivariate-adjusted hazard ratios (HRs) (95% Cl) were collected as the necessary risk estimate for synthesis. Random-effects models were applied for meta-analysis. RESULTS: Overall, 13 studies involving 44 514 participants were included in this meta-analysis. There were no significant differences in the risk of all-cause mortality (pooled HR: 1.18 [95% Cl: 0.95, 1.45], I2 = 94%) and cardiovascular-related mortality (pooled HR: 1.08 [95% Cl: 0.94, 1.23], I2 = 0%) between euthyroid older adults and older adults with subclinical hypothyroidism. The results remained the same when only older adults with thyrotropin ≥10 mIU/L were assessed (pooled HR for all-cause mortality and cardiovascular-related mortality, respectively: 1.53 [95% Cl: 0.81, 2.88], I2 = 22%, 1.35 [95% Cl: 0.63, 2.86], I2 = 43%). CONCLUSION: High thyrotropin levels are not associated with increased risk for all-cause mortality as well as cardiovascular-related mortality in older adults aged ≥60 years with subclinical hypothyroidism, suggesting an unnecessity in initialing treatment.

Effect of Probiotic Supplementation on the Gut Microbiota Composition of Infants Delivered by Cesarean Section: An Exploratory, Randomized, Open-label, Parallel-controlled Trial.

BACKGROUND: Infants born via cesarean section (CS) are at an increased risk of immune-related diseases later in life, potentially due to altered gut microbiota. Recent research has focused on the administration of probiotics in the prevention of gut microbiota dysbiosis in neonates delivered by CS. This study was performed to investigate the effects of probiotic supplementation on the gut microbiota of CS-delivered infants. METHODS: Thirty full-term neonates delivered by CS were randomized into the intervention (supplemented orally with a probiotic containing Bifidobacterium longum, Lactobacillus acidophilus, and Enterococcus faecalis for 2 weeks) and control groups. Stool samples were collected at birth and 2 weeks and 42 days after birth. The composition of the gut microbiota was analyzed using 16S rRNA sequencing technology. RESULTS: The applied bacterial strains were abundant in the CS-delivered infants supplemented with probiotics. Probiotics increased the abundance of some beneficial bacteria, such as Bacteroides, Acinetobacter, Veillonella, and Faecalibacterium. Low colonization of Klebsiella, a potentially pathogenic bacterium, was observed in the intervention group. CONCLUSIONS: Our results showed that probiotics supplemented immediately after CS enriched the gut microbiota composition and altered the pattern of early gut colonization. TRIAL REGISTRATION: registration number NCT05086458.

Nightshift work can induce oxidative DNA damage: a pilot study.

BACKGROUND: Regular sleep is very important for human health; however, the short-term and long-term effects of nightshift with sleep deprivation and disturbance on human metabolism, such as oxidative stress, have not been effectively evaluated based on a realistic cohort. We conducted the first long-term follow-up cohort study to evaluate the effect of nightshift work on DNA damage. METHODS: We recruited 16 healthy volunteers (aged 33 ± 5 years) working night shifts at the Department of Laboratory Medicine at a local hospital. Their matched serum and urine samples were collected at four time points: before, during (twice), and after the nightshift period. The levels of 8-oxo-7,8-dihydroguanosine (8-oxoG) and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), two important nucleic-acid damage markers, were accurately determined based on a robust self-established LC‒MS/MS method. The Mann-Whitney U or Kruskal-Wallis test was used for comparisons, and Pearson's or Spearman's correlation analysis was used to calculate the correlation coefficients. RESULTS: The levels of serum 8-oxodG, estimated glomerular filtration rate-corrected serum 8-oxodG, and the serum-to-urine 8-oxodG ratio significantly increased during the nightshift period. These levels were significantly higher than pre-nightshift work level even after 1 month of discontinuation, but no such significant change was found for 8-oxoG. Moreover, 8-oxoG and 8-oxodG levels were significantly positively associated with many routine biomarkers, such as total bilirubin and urea levels, and significantly negatively associated with serum lipids, such as total cholesterol levels. CONCLUSION: The results of our cohort study suggested that working night shifts may increase oxidative DNA damage even after a month of discontinuing nightshift work. Further studies with large-scale cohorts, different nightshift modes, and longer follow-up times are needed to clarify the short- and long-term effects of night shifts on DNA damage and find effective solutions to combat the negative effects.

The association of plasma NT-proBNP level and progression of diabetic kidney disease.

AIMS: Diabetic kidney disease (DKD) is the most common cause of end-stage kidney disease (ESKD). The identification of risk factors involved in the progression of DKD to ESKD is expected to result in early detection and appropriate intervention and improve prognosis. This study aimed to explore whether plasma N-terminal pro-B-type natriuretic peptide (NT-proBNP) was associated with kidney outcomes in patients with type 2 diabetes mellitus (T2DM) and biopsy-proven DKD. METHODS: Patients with biopsy-proven DKD who were followed up at West China Hospital over 12 months were enrolled. The kidney outcome was defined as progression to ESKD. The cutoff value of plasma NT-proBNP concentration was calculated by using receiver operating characteristic (ROC) curve analysis. The influence of NT-proBNP levels on kidney outcome in patients with DKD was assessed using Cox regression analysis. RESULTS: A total of 30 (24.5%) patients reached ESKD during a median follow-up of 24.1 months. The baseline serum NT-proBNP level had a significant correlation with baseline proteinuria, kidney function, glomerular lesions, interstitial fibrosis tubular atrophy (IFTA), and arteriolar hyalinosis. Multivariate Cox regression analysis indicated that increased NT-proBNP level was significantly associated with a higher risk of progression to ESKD (HR 6.43; 95% CI (1.65-25.10, p = 0.007), and each 1 SD increase in LG (NT-proBNP) was also associated with a higher risk (HR 2.43; 95% CI 1.94-5.29, p = 0.047) of an adverse kidney outcome after adjusting for confounding factors. CONCLUSIONS: A higher level of plasma NT-proBNP predicts kidney prognosis in patients with biopsy-proven DKD. This warrants further investigation into the potential mechanisms.

An innovative approach based on real-world big data mining for calculating the sample size of the reference interval established using transformed parametric and non-parametric methods.

BACKGROUND: Currently, the direct method is the main approach for establishment of reference interval (RI). However, only a handful of studies have described the effects of sample size on establishment of RI and estimation of sample size. We describe a novel approach for estimation of the sample size when establishing RIs using the transformed parametric and non-parametric methods. METHODS: A total of 3,697 healthy participants were enrolled in this study. We adopted a two-layer nested loop sample size estimation method to determine the effects of sample size on RI, using thyroid-related hormone as an example. The sample size was selected as the calculation result when the width of the confidence interval (CI) of the upper and lower limit of the RI were both stably < 0.2 times the width of RI. Then, we calculated the sample size for establishing RIs via transformed parametric and non-parametric methods for thyroid-related hormones. RESULTS: Sample sizes for thyroid stimulating hormone (TSH), as required by parametric and non-parametric methods to establish RIs were 239 and 850, respectively. Sample sizes required by the transformed parametric method for free triiodothyronine (FT3), free thyroxine (FT4), total triiodothyronine (TT3) and total thyroxine (TT4) were all less than 120, while those required by the non-parametric method were more than 120. CONCLUSION: We describe a novel approach for estimating sample sizes for establishment of RI. A corresponding open-source code has been developed and is available for applications. The established method is suitable for most analytes, with evidence based on thyroid-related hormones indicating that different sample sizes are required to establish RIs using different methods for analytes with different variations.

An automated magnetic bead extraction method for measuring plasma metanephrines and 3-methoxytyramine using liquid chromatography tandem mass spectrometry.

Liquid chromatography tandem mass spectrometry (LC-MS/MS) is used routinely in clinical diagnostics; however, automating the sample pretreatment is challenging. We established and evaluated an automated method based on the magnetic bead extraction principle (MBE) to measure normetanephrine (NMN), metanephrine (MN), and 3-methoxytyramine (3-MT). The target analytes were extracted, purified, and concentrated using different solvents and chemical bond-modified magnetic beads transferred via a magnetic bar. The linearity, recovery, matrix effect, and precision of MBE were evaluated thoroughly, and compared with traditional solid-phase extraction (SPE) using 131 plasma samples. The chromatography peaks of metanephrines and 3-MT, extracted via MBE, are symmetrical, without interfering peaks. The linearity was excellent with correlation coefficient (r) > 0.99. The MBE exhibited good reproducibility with within-run coefficient variations (CVs) of 1.96-2.00%, 4.06-5.75%, and 3.89-4.90% for MN, NMN, and 3-MT, respectively. The total CVs for MN, NMN, and 3-MT were 1.96-2.80%, 5.12-5.75%, and 5.44-6.27%, respectively. The relative recoveries for MN, NMN, and 3-MT varied between 93.5 and 107.4%, whereas their biases were all within 10%. The results for MN, NMN, and 3-MT extracted via MBE compared with SPE exhibited excellent correlation, with r > 0.99; the mean bias% for MN, NMN, and 3-MT were small (-2.9%, -3.2%, and -3.2%, respectively). In conclusion, the automated MBE method for measuring plasma metanephrines and 3-MT can be applied in future routine clinical diagnostics, and the MBE principle may indicate a new era for LC-MS/MS in clinical application.

Is pre-heat necessary for the measurement of 8-oxo-7,8-dihydroguanosine and 8-oxo-7,8-dihydro-2'-deoxyguanosine in urine samples.

BACKGROUND: It is currently unclear for the necessary of pre-heating urine samples for the accurate determination of 8-oxo-7,8-dihydroguanosine (8-oxoG) and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG). Thus, we conducted this study to evaluate the effect of pre-heat (i.e., to 37°C) on the accurate measurement of 8-oxoG and 8-oxodG in frozen urine samples. METHODS: Random urine samples from six healthy volunteers, six patients with renal dysfunction, and six patients with systematic diseases such as diabetes were collected, split, and stored at -80°C for up to 1 month. The frozen samples were thawed at room temperature (RT) or 37°C for different time, 10-fold diluted with ddH2O containing 1% formic acid, and determined by self-established LC-MS/MS method coupled with an ACQUITY™ Primer HSS T3 column. RESULTS: Thawing the samples at RT for 30 or 120 min, or at 37°C for 15 or 90 min did not affect the determination of 8-oxoG and 8-oxodG in urine samples. Moreover, no significant difference between thawing the urine samples at RT and 37°C was found after storing at -80°C for 1-3 months. CONCLUSION: It is not always necessary to pre-heat the frozen urine samples to release 8-oxoG and 8-oxodG from precipitates, which is associated with different pre-treatment and determination methods.

Development and internal validation of machine learning algorithms for end-stage renal disease risk prediction model of people with type 2 diabetes mellitus and diabetic kidney disease.

AIMS: Diabetic kidney disease (DKD) is the most common cause of end-stage renal disease (ESRD) and is associated with increased morbidity and mortality in patients with diabetes. Identification of risk factors involved in the progression of DKD to ESRD is expected to result in early detection and appropriate intervention and improve prognosis. Therefore, this study aimed to establish a risk prediction model for ESRD resulting from DKD in patients with type 2 diabetes mellitus (T2DM). METHODS: Between January 2008 and July 2019, a total of 390 Chinese patients with T2DM and DKD confirmed by percutaneous renal biopsy were enrolled and followed up for at least 1 year. Four machine learning algorithms (gradient boosting machine, support vector machine, logistic regression, and random forest (RF)) were used to identify the critical clinical and pathological features and to build a risk prediction model for ESRD. RESULTS: There were 158 renal outcome events (ESRD) (40.51%) during the 3-year median follow up. The RF algorithm showed the best performance at predicting progression to ESRD, showing the highest AUC (0.90) and ACC (82.65%). The RF algorithm identified five major factors: Cystatin-C, serum albumin (sAlb), hemoglobin (Hb), 24-hour urine urinary total protein, and estimated glomerular filtration rate. A nomogram according to the aforementioned five predictive factors was constructed to predict the incidence of ESRD. CONCLUSION: Machine learning algorithms can efficiently predict the incident ESRD in DKD participants. Compared with the previous models, the importance of sAlb and Hb were highlighted in the current model.HighlightsWhat is already known? Identification of risk factors for the progression of DKD to ESRD is expected to improve the prognosis by early detection and appropriate intervention.What this study has found? Machine learning algorithms were used to construct a risk prediction model of ESRD in patients with T2DM and DKD. The major predictive factors were found to be CysC, sAlb, Hb, eGFR, and UTP.What are the implications of the study? In contrast with the treatment of participants with early-phase T2DM with or without mild kidney damage, major emphasis should be placed on indicators of kidney function, nutrition, anemia, and proteinuria for participants with T2DM and advanced DKD to delay ESRD, rather than age, sex, and control of hypertension and glycemia.

Addition of glomerular lesion severity improves the value of anemia status for the prediction of renal outcomes in Chinese patients with type 2 diabetes.

We aimed to determine the utility of biopsy data and anemia for the prediction of renal outcomes in Chinese patients with type 2 diabetes. In total, 441 Chinese patients with type 2 diabetes and biopsy-confirmed diabetic nephropathy (DN) were enrolled in a retrospective study. Their renal pathology was assessed using the Renal Pathology Society system. Cox proportional hazards models were used to estimate hazard ratios (HRs) for end-stage renal disease (ESRD), and immunofluorescence staining was used to assess the expression of hypoxia-inducible factor (HIF)-α in patients' kidneys. We found that glomerular pathology classification was an independent pathological predictor of low hemoglobin concentration, according to linear and logistic regression analyses. Each 1 g/dL decrease in baseline hemoglobin concentration was associated with a 42% higher risk of an adverse renal outcome, after adjustment for clinical and pathologic covariates. In patients with severe glomerular lesions, the risk of progression to ESRD was significantly higher if mild or moderate/severe anemia was present, but in patients with mild glomerular lesions, the risk was only significantly higher in those with moderate or severe anemia than in the absence of anemia. Harrell's C Concordance was improved, but the Akaike information criterion was worsened by adding the glomerular pathology classification to the use of anemia status and clinical data. Immunofluorescence staining revealed that renal HIF-1α and HIF-2α expression was significantly higher in classes II-IV than class I. Thus, the addition of glomerular pathology classification increases the value of anemia status for the prediction of the progression to ESRD.

Analytical and Clinical Performance of a Liquid Chromatography-Tandem Mass Spectrometry Method for Measuring Gastrin Subtypes G34 and G17 in Serum.

BACKGROUND: Two major forms of gastrin, gastrin-17 (G17) and gastrin-34 (G34), exist in blood. However, conventional immunoassay methods can only quantify total gastrin or G17 alone. Here, we aimed to establish a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to quantify G17 and G34 simultaneously. METHODS: Serum samples were prepared by anion-exchange solid-phase extraction. The analytical performance of the LC-MS/MS method was validated and the method was compared to chemiluminescence immunoassay (CLIA) and radioimmunoassay (RIA). The G17 and G34 concentrations in 245 serum samples from healthy controls, individuals with gastrinoma, and individuals with other diseases were analyzed. RESULTS: The total runtime of the LC-MS/MS method was 6 min. No substantial matrix effect was observed with internal standard correction. The intraassay coefficients of variation (CVs) for G17 and G34 were 4.0%-14.2% and 4.4%-10.4%, respectively, and total CVs were 5.2%-14.1% and 4.6%-12.4%, respectively. The correlation coefficient between LC-MS/MS and CLIA was 0.87, and between LC-MS/MS and RIA was 0.84. The G17+G34 concentrations for 87.5% of individuals with gastrinoma were higher than the 95th percentile of healthy controls (18.1 pg/mL), whereas the concentrations for individuals with other diseases and gastrinoma overlapped. Based on the Youden indices calculated for G17+G34, G34, and G17, the most specific biomarker was G17 (96.9% clinical specificity at 209.8 pg/mL) for gastrinoma. CONCLUSIONS: This method should aid in the diagnosis of diseases associated with increased gastrin concentrations.

Establishment of Early Pregnancy Related Thyroid Hormone Models and Reference Intervals for Pregnant Women in China Based on Real World Data.

Thyroid hormone reference intervals are crucial for diagnosing and monitoring thyroid dysfunction during early pregnancy, and the dynamic change trend of thyroid hormones during pregnancy can assist clinicians to assess the thyroid function of pregnant women. This study aims to establish early pregnancy related thyroid hormones models and reference intervals for pregnant women. We established two derived databases: derived database* and derived database#. Reference individuals in database* were used to establish gestational age-specific reference intervals for thyroid hormones and early pregnancy related thyroid hormones models for pregnant women. Individuals in database# were apparently healthy non-pregnant women. The thyroid hormones levels of individuals in database# were compared with that of individuals in database* using nonparametric methods and the comparative confidence interval method. The differences in thyroid stimulating hormone and free thyroxine between early pregnant and non-pregnant women were statistically significant (p<0.0001). The reference intervals of thyroid stimulating hormone, free thyroxine and free triiodothyronine for early pregnant women were 0.052-3.393 µIU/ml, 1.01-1.54 ng/dl, and 2.51-3.66 pg/ml, respectively. Results concerning thyroid stimulating hormone and free thyroxine reference intervals of early pregnancy are comparable with those from other studies using the same detection platform. Early pregnancy related thyroid hormones models showed various change patterns with gestational age for thyroid hormones. Early pregnancy related thyroid hormones models and reference intervals for pregnant women were established, so as to provide accurate and reliable reference basis for the diagnosing and monitoring of maternal thyroid disfunction in early pregnancy.

Data mining: traditional spring festival associated with hypercholesterolemia.

BACKGROUND: Serum lipid concentrations are affected by long-term high-fat diets; thus, we hypothesize that lipid levels increase after the Spring Festival in China. METHOD: In total, 20,192 individuals (male: n=10,108, female: n=10,084) were enrolled in this retrospective cross-sectional study based on clinical data from the Laboratory Information System (LIS) and Hospital Information System (HIS) in Peking Union Medical College Hospital from 2014 to 2018. Total cholesterol (TC), triglycerides (TGs), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) were analyzed. RESULTS: The serum TC [male vs. female: (4.71 ± 0.90 vs. 4.56 ± 0.85) mmol/L], TG [male vs. female: (1.71 ± 1.56 vs. 1.02 ± 0.68) mmol/L], and LDL-C [male vs. female: (3.01 ± 0.77 vs. 2.73 ± 0.74) mmol/L] levels were significantly higher in males than in females (P < 0.001); serum HDL-C [male vs. female: (1.18 ± 0.28 vs. 1.50 ± 0.34) mmol/L] was significantly lower in males (P < 0.001). In February, the TC, TG, and LDL-C levels were 8.4%, 16.3%, and 9.3% higher than the lowest levels recorded, respectively. The prevalence of dyslipidemia of the two weeks before the Spring festival was significantly lower than that of the first week after the Spring festival (43.6% (168/385) vs. 54.1% (126/233), P=0.007). Additionally, the prevalence of dyslipidemia was statistically higher in the first week after the Spring Festival than in May-January. CONCLUSION: Higher TC, TG, and LDL-C in winter could be associated with high-fat diets during the Spring Festival. The Spring Festival was immediately followed by a higher lipid concentrations. Thus, we don't recommend lipid assessment or physical examination immediately after the holiday especially Spring festival.

Evaluation of bone metabolism-associated biomarkers in Tibet, China.

AIM: To measure and evaluate the distribution and possible contributing factors of seven bone metabolism-associated biomarkers in Tibet, a plateau province of China. METHODS: A total of 1615 individuals were recruited from Tibet at three different altitudes. The levels and possible contributing factors of serum calcium, serum phosphorus, ALP, 25OHD, PINP, CTX, and PTH were evaluated. RESULTS: In total, 1246 Tibetan adults (males: n = 543) were eventually enrolled in this study. Multiple linear regression recognized age, sex, altitude, and BMI as the major effect factors. The levels of ALP, PINP, and CTX in males continuously decreased with age; however, those in females increased after approximately 39 years of age. Males had higher 25OHD levels (23.9 vs. 15.4 ng/ml) but lower levels of serum phosphorus (1.12 vs. 1.19 mmol/L) and PTH (41.3 vs. 47.4 pg/ml) than females. Before the age of 50, males had higher levels of calcium, ALP, PINP, and CTX than females, and the opposite trend was observed after the age of 50. The highest levels of serum calcium and phosphorus and the lowest levels of PINP and CTX were found in the Shigatse/Lhasa region, suggesting a better bone metabolism status. Compared with reports from plain areas of China, significantly higher levels of PINP (65.3 vs. 49.36 ng/ml) and CTX (0.46 vs. 0.37 ng/ml) were recorded in Tibetan adults. CONCLUSION: A more active bone turnover status was found in Tibetan adults than in individuals from the plain areas of China.