Characterization of site-specific N-glycosylation signatures of isolated uromodulin from human urine.

Uromodulin (Umod, Tamm-Horsfall protein) is the most abundant urinary N-glycoprotein produced exclusively by the kidney. It can form filaments to antagonize the adhesion of uropathogens. However, the site-specific N-glycosylation signatures of Umod in healthy individuals and patients with IgA nephropathy (IgAN) remain poorly understood due to the lack of suitable isolation and analytical methods. In this study, we first presented a simple and fast method based on diatomaceous earth adsorption to isolate Umod. These isolated glycoproteins were digested by trypsin and/or Glu-C. Intact N-glycopeptides with or without HILIC enrichment were analyzed using our developed EThcD-sceHCD-MS/MS. Based on the optimized workflow, we identified a total of 780 unique intact N-glycopeptides (7 N-glycosites and 152 N-glycan compositions) from healthy individuals. As anticipated, these glycosites exhibited glycoform heterogeneity. Almost all N-glycosites were modified completely by the complex type, except for one N-glycosite (N275), which was nearly entirely occupied by the high-mannose type for mediating Umod's antiadhesive activity. Then, we compared the N-glycosylation of Umod between healthy controls (n = 9) and IgAN patients (n = 9). The N-glycosylation of Umod in IgAN patients will drastically decrease and be lost. Finally, we profiled the most comprehensive site-specific N-glycosylation map of Umod and revealed its alterations in IgAN patients. Our method provides a high-throughput workflow for characterizing the N-glycosylation of Umod, which can aid in understanding its roles in physiology and pathology, as well as serving as a potential diagnostic tool for evolution of renal tubular function.

Unexpected Thymine Oxidation and Collision-Induced Thymine-Pt-guanine Cross-Linking on 5'-TpG and 5'-GpT by a Photoactivatable Diazido Pt(IV) Anticancer Complex.

The photochemical products of dinucleotides 5'-TpG/5'-GpT with a photoactivatable anticancer Pt(IV) complex (trans,trans,trans-[Pt(N3)2(OH)2(py)2], py = pyridine; 1) were characterized by electrospray ionization mass spectrometry. The primary MS showed the main products were monoplatinated and diplatinated adducts for both the dinucleotides accompanied by the formation of minor triplatinated dinucleotides, indicating that T-N3 and G-N1 may be platination sites additional to the well-known G-N7 site. Surprisingly, a series of minor platinated adducts with oxidation of guanine and/or thymine were observed. Although guanine is more sensitive to oxidation than thymine, thymine can compete with guanine for complex 1-induced oxidation, of which the oxidation adducts were identified as cis- and trans-diastereomers of 5,6-dihydroxy-5,6-dihydrothymidine (cis,trans-ThdGly), 5-formyl-2'-deoxyuridine (5-FormdUrd), and 5-(hydroxymethyl)-2'-deoxyuridine (5-HMdUrd), respectively. While for guanine, apart from 8-hydroxyguanine (8-OH-G) and N-formylamidoiminohydantoin (RedSp), other guanine oxidized adducts such as spiroiminodihydantoin (Sp), dehydroguanidinohydantoin (DGh), and 2,6-diamino-4-hydroxy-5-formamidopyrimidine (FapyG) were also identified. MS/MS analysis showed that unique fragments with a Pt moiety [Pt(N3)(py)] cross-linking the G and T bases were formed during the fragmentation of monoplatinated dinucleotides. Such binding mode to and oxidative damages on DNA bases imposed by the diazido Pt(IV) complex are apparently distinct from those of cisplatin, perhaps accounting for its unique mechanism of action.

Quantification of myocardial strain in patients with isolated left ventricular non-compaction and healthy subjects using deformable registration algorithm: comparison with feature tracking.

BACKGROUND: Systolic dysfunction of the left ventricle is frequently associated with isolated left ventricular non-compaction (iLVNC). Clinically, the ejection fraction (EF) is the primary index of cardiac function. However, changes of EF usually occur later in the disease course. Feature tracking (FT) and deformable registration algorithm (DRA) have become appealing techniques for myocardial strain assessment. METHODS: Thirty patients with iLVNC (36.7 ± 13.3 years old) and fifty healthy volunteers (42.3 ± 13.6 years old) underwent cardiovascular magnetic resonance (CMR) examination on a 1.5 T MR scanner. Strain values in the radial, circumferential, longitudinal directions were analyzed based on the short-axis and long-axis cine images using FT and DRA methods. The iLVNC patients were further divided based on the ejection fraction, into EF ≥ 50% group (n = 11) and EF < 50% group (n = 19). Receiver-operating-characteristic (ROC) analysis was performed to assess the diagnostic performance of the global strain values. Intraclass correlation coefficient (ICC) analysis was used to evaluate the intra- and inter-observer agreement. RESULTS: Global radial strain (GRS) was statistically lower in EF ≥ 50% group compared with control group [GRS (DRA)/% vs. controls: 34.6 ± 7.0 vs. 37.6 ± 7.2, P < 0.001; GRS (FT)/% vs. controls: 37.4 ± 13.2 vs. 56.9 ± 16.4, P < 0.01]. ROC analysis of global strain values derived from DRA and FT demonstrated high area under curve (range, 0.743-0.854). DRA showed excellent intra- and inter-observer agreement of global strain in both iLVNC patients (ICC: 0.995-0.999) and normal controls (ICC: 0.934-0.996). While for FT analysis, global radial strain of normal controls showed moderate intra-observer (ICC: 0.509) and poor inter-observer agreement (ICC: 0.394). CONCLUSIONS: In patients with iLVNC, DRA can be used to quantitatively analyze the strain of left ventricle, with global radial strain being an earlier marker of LV systolic dysfunction. DRA has better reproducibility in evaluating both the global and segmental strain.

Proteomic Strategy for Identification of Proteins Responding to Cisplatin-Damaged DNA.

A new proteomic strategy combining functionalized magnetic nanoparticle affinity probes with mass spectrometry was developed to capture and identify proteins specifically responding to 1,2-d(GpG) intrastrand cisplatin-cross-linked DNA, the major DNA lesion caused by cisplatin and thought to induce apoptosis. A 16-mer oligodeoxynucleotide (ODN) duplex and its cisplatin-cross-linked adduct were immobilized on magnetic nanoparticles via click reaction, respectively, to fabricate negative and positive affinity probes which were very stable in cellular protein extracts due to the excellent bio-orthogonality of click chemistry and the inertness of covalent triazole linker. Quantitative mass spectrometry results unambiguously revealed the predominant binding of HMGB1 and HMGB2, the well-established specific binders of 1,2-cisplatin-cross-linked DNA, to the cisplatin-cross-linked ODN, thus validating the accuracy and reliability of our strategy. Furthermore, 5 RNA or single-stranded DNA binding proteins, namely, hnRNP A/B, RRP44, RL30, RL13, and NCL, were demonstrated to recognize specifically the cisplatinated ODN, indicating the significantly unwound ODN duplex by cisplatin cross-linking. In contrast, the binding of a transcription factor TFIIFa to DNA was retarded due to cisplatin damage, implying that the cisplatin lesion stalls DNA transcription. These findings promote understanding in the cellular responses to cisplatin-damaged DNA and inspire further precise elucidation of the action mechanism of cisplatin.

In Situ Liquid Secondary Ion Mass Spectrometry: A Surprisingly Soft Ionization Process for Investigation of Halide Ion Hydration.

The understanding of ion solvation phenomena is of significance due to their influences on many important chemical, biological, and environmental processes. Mass spectrometry (MS)-based methods have been used to investigate this topic with molecular insights. As ion-solvent interactions are weak, ionization processes should be as soft as possible in order to retain solvation structures. An in situ liquid secondary ion MS (SIMS) approach developed in our group has been recently utilized in investigations of Li ion solvation in nonaqueous solution, and it detected a series of solvated Li ions. As traditionally SIMS has long been recognized as a hard ionization process with strong damage occurring at the sputtering interface, it is very interesting to study further how soft in situ liquid SIMS can be. In this work, we used halide ion hydration as an example to compare the ionization performance of the in situ liquid SIMS approach with regular electrospray ionization MS (ESI-MS). Results show that, although ESI has been recognized as a soft ionization method, nearly no solvated halide ions were detected by ESI-MS analysis, which acquired only strong signals of salt ion clusters. In contrast, in liquid SIMS spectra, a series of obvious hydrated halide ion compositions could be observed. We further evaluated the hydration numbers of halide ions and revealed the effects of the ion size, charge density, and polarizability on the hydration phenomenon. Our findings demonstrated that the in situ liquid SIMS approach is surprisingly soft, and it is expected to have very broad applications on investigation of various ion-solvent interactions and many other interesting chemical processes (e.g., the initial nucleation of nanoparticle formation) in liquid environment.

Discovery of Cisplatin Binding to Thymine and Cytosine on a Single-Stranded Oligodeoxynucleotide by High Resolution FT-ICR Mass Spectrometry.

The clinically widely-used anticancer drug, cisplatin, binds strongly to DNA as a DNA-damaging agent. Herein, we investigated the interaction of cisplatin with a 15-mer single-stranded C,T-rich oligodeoxynucleotide, 5'-CCTT4CTT7G8C9T10TCTCC-3' (ODN15), using ultra-high resolution Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) in conjunction with tandem mass spectrometry (top-down MS). Top-down MS analysis with collision-induced dissociation (CID) fragmentation of the mono-platinated and di-platinated ODN15 provided abundant and informative Pt-containing or Pt-free a/[a - B], w and internal fragments, allowing the unambiguous identification of T4, T7, C9, and T10 as the platination sites on the cisplatin-ODN15 adducts. These results revealed that, in addition to the well-established guanine site, the unexpected thermodynamic binding of cisplatin to cytosine and thymine bases was also evident at the oligonucleotide level. Furthermore, the binding models of cisplatin with cytosine and thymine bases were built as the Pt coordinated to cytosine-N(3) and thymine-N(3) with displacement of the proton or tautomerization of thymine. These findings contribute to a better understanding of the mechanism of action of cisplatin and its preference for gene loci when the drug binds to cellular DNA, and also demonstrate the great potential and superiority of FT-ICR MS in studying the interactions of metallodrugs with large biomolecules.

Facile synthesis of graphene oxide/palygorskite composites for Pb(II) rapid removal from aqueous solutions.

As a kind of earth-abundant and cheap natural clay mineral, palygorskite (Pal) was facilely modified by grafting with graphene oxide (GO) to fabricate GO/Pal composites for rapid removal of Pb(II) from aqueous solutions. The results of characterization confirmed that the GO/Pal composites were successfully grafted between GO sheets and Pal nanorods. The effects of pH, adsorbent dosage, adsorption time, initial Pb(II) concentration and temperature on the adsorption of Pb(II) onto the GO/Pal composites as adsorbents were systematically investigated. The maximum adsorption capacity over 106.6 mg/g was obtained within a short adsorption time of less than 1 h even at 298.15 K. The adsorption of Pb(II) was a fast process that more accurately followed the pseudo-second-order kinetic equation. This process also could be described better with the Langmuir equation model than the Freundlich model. The negative values of ΔG° and the positive values of ΔH° and ΔS° indicated that it was a spontaneous, endothermic and entropy-increasing adsorption process. Compared with pristine Pal and GO powders, such the GO/Pal composites as a cost-efficient and eco-friendly adsorbents could significantly improve the adsorption properties of Pb(II) and would have potential application in the industrial wastewater treatment for rapid removal of Pb(II).

Increased myocardial extracellular volume assessed by cardiovascular magnetic resonance T1 mapping and its determinants in type 2 diabetes mellitus patients with normal myocardial systolic strain.

BACKGROUND: Cardiac magnetic resonance (CMR) T1 mapping and tissue-tracking strain analysis are useful quantitative techniques that can characterize myocardial tissue and mechanical alterations, respectively, in patients with early diabetic cardiomyopathy. The purpose of this study was to assess the left ventricular myocardial T1 value, extracellular volume fraction (ECV), and systolic strain in asymptomatic patients with type 2 diabetes mellitus (T2DM) and their underlying relationships with clinical parameters. METHODS: We recruited 50 T2DM patients (mean age: 55 ± 7 years; 28 males) and 32 sex-, age-and BMI-matched healthy volunteers to undergo contrast-enhanced CMR examinations. The myocardial native T1, post-contrast T1 and ECV values of the left ventricle were measured from T1 and ECV maps acquired using the modified Look-Locker inversion recovery technique. The left ventricular global systolic strain and the strain rates were evaluated using routine cine images and tissue-tracking analysis software. The baseline clinical and biochemical indices were collected before the CMR examination. RESULTS: The myocardial ECV and native T1 values were significantly higher in the diabetic patients than in the controls. (ECV: 27.4 ± 2.5% vs. 24.6 ± 2.2%, p < 0.001; native T1: 1026.9 ± 30.0 ms vs. 1011.8 ± 26.0 ms, p = 0.022). However, the left ventricular global systolic strain, strain rate, volume, myocardial mass, ejection fraction, and left atrial volume were similar between the diabetic patients and the healthy controls. In the diabetic patients, the native T1 values were independently correlated with the hemoglobin A1c levels (standardized ß = 0.368, p = 0.008). The ECVs were independently associated with the hemoglobin A1c levels (standardized ß = 0.389, p = 0.002), angiotensin-converting enzyme inhibitor (ACEI) treatment (standardized ß = - 0.271, p = 0.025) and HCT values (standardized ß = - 0.397, p = 0.001). CONCLUSIONS: Type 2 diabetes mellitus patients with normal myocardial systolic strain exhibit increased native T1 values and ECVs indicative of myocardial extracellular interstitial expansion, which might be related to poor glycemic control. The amelioration of myocardial interstitial matrix expansion might be associated with ACEI treatment. A valid assessment of the association of glucose control and ACEI treatment with myocardial fibrosis requires notably larger trials.

Synthesis of Triarylmethanes via Palladium-Catalyzed Suzuki Coupling of Trimethylammonium Salts and Arylboronic Acids.

An efficient palladium-catalyzed Suzuki coupling of 1,1-diarylmethyl-trimethylammonium triflates with arylboronic acids is reported. This reaction offers a novel approach to triarylmethane derivatives in good to excellent yields with the palladium-catalyzed C-N bond cleavage as the key feature. Broad substrate scope regarding both reaction partners are observed. Moreover, reactive functional groups such as vinyl and formyl groups are conserved in this transformation.

Selective binding of an organoruthenium complex to G-rich human telomeric sequence by tandem mass spectrometry.

RATIONALE: Human telomeric DNA is reported to be a potential target for anticancer organometallic ruthenium(II) complexes, however, the interaction sites were not clearly discriminated and identified. METHODS: In the current study, tandem mass spectrometry (MS/MS) using collision-induced dissociation (CID) was firstly introduced to identify the interaction sites of an organometallic ruthenium(II) complex [(η6 -biphenyl)Ru(en)Cl][PF6 ] (1; en = ethylenediamine) with 5'-T1 T2 A3 G4 G5 G6 -3' (I), the repeating unit of human telomeric DNA, in both positive- and negative-ion mode at a low reaction molar ratio (1/I = 0.2) which was applied to preserve the site selectivity. RESULTS: Mass spectrometric results showed that mono-ruthenated I was the main product under the conditions. In positive-ion mode, MS/MS results indicated that ruthenium complex 1 binds to T2 or G6 in strand I. However, in negative-ion mode, no efficient information was obtained for exact identification of ruthenation sites which may be attributed to losses of fragment ions due to charge neutralization by the coordination of the positively charged ruthenium complex to the short MS/MS fragments. CONCLUSIONS: This is the first report of using top-down MS to characterize the interactions of organometallic ruthenium(II) complexes and human telomeric DNA. Thymine can be thermodynamically competitive with guanine for binding to ruthenium complexes even at low reaction molar ratio, which inspired us to explore in greater depth the significance of thymine binding.

Binding of Organometallic Ruthenium Anticancer Complexes to DNA: Thermodynamic Base and Sequence Selectivity.

Organometallic ruthenium(II) complexes [(η6-arene)Ru(en)Cl][PF6] (arene = benzene (1), p-cymene (2), indane (3), and biphenyl (4); en = ethylenediamine) are promising anticancer drug candidates both in vitro and in vivo. In this paper, the interactions between ruthenium(II) complexes and 15-mer single- and double-stranded oligodeoxynucleotides (ODNs) were thermodynamically investigated using high performance liquid chromatography (HPLC) and electrospray ionization mass spectroscopy (ESI-MS). All of the complexes bind preferentially to G8 on the single strand 5'-CTCTCTT7G8T9CTTCTC-3' (I), with complex 4 containing the most hydrophobic ligand as the most reactive one. To the analogs of I (changing T7 and/or T9 to A and/or C), complex 4 shows a decreasing affinity to the G8 site in the following order: -AG8T- (K: 5.74 × 104 M-1) > -CG8C- > -TG8A- > -AG8A- > -AG8C- > -TG8T- (I) ≈ -CG8A- (K: 2.81 × 104 M-1). In the complementary strand of I, the G bases in the middle region are favored for ruthenation over guanine (G) bases in the end of oligodeoxynucleotides (ODNs). These results indicate that both the flanking bases (or base sequences) and the arene ligands play important roles in determining the binding preference, and the base- and sequence-selectivity, of ruthenium complex in binding to the ODNs.

Prognostic Significance of Neutrophil-to-Lymphocyte Ratio in Ovarian Cancer: A Systematic Review and Meta-Analysis of Observational Studies.

BACKGROUND AND AIMS: Published data on the prognostic role of neutrophil-to-lymphocyte ratio (NLR) in ovarian cancer are controversial. We conducted this meta-analysis to obtain a more accurate assessment of prognostic significance of NLR in ovarian cancer. MATERIALS AND METHODS: We conducted a systematic literature search using the electronic databases PubMed, Web of Science, and Embase up to May 2016. Hazard ratio (HR) and odd ratio (OR) with 95% confidence interval (95% CI) were calculated. Subgroup analyses were carried out to explore the source of heterogeneity. Statistical analysis was performed using Stata 10.0. RESULTS: A total of 12 studies, consisting of 3,854 patients, which met our criterion were selected in this meta-analysis. Our pooled results showed that high pre-treatment NLR level was significantly associated with poorer overall survival (OS) (HR: 1.69, 95% CI 1.29-2.22) and shorter progression free survival (PFS) (HR 1.63, 95% CI 1.27-2.09). Additionally, increased NLR was also significantly correlated with advanced FIGO stage (OR 2.32, 95% CI1.79-3.00), higher serum level of CA-125 (OR 3.33, 95% CI 2.43-4.58), more extensive ascites (OR 3.54, 95% CI 2.31-5.42) as well as less chemotheraputic response (OR 0.53, 95% CI 0.40-0.70). The findings from most of subgroup meta-analyses were consistent with those from the overall meta-analyses. CONCLUSIONS: Elevated pre-treatment NLR could served as a predicative factor of poor prognosis for ovarian cancer patients.

[Leptin gene C2549A polymorphism in minority Hui and Uygur children with obesity].

OBJECTIVE: To investigate the relationship of leptin gene polymorphism with obesity in ethnic minority Hui and Uygur children in China. METHODS: Sixty-eight ethnic minority (35 Hui and 33 Uygur) children with obesity and 69 age-matched minority (36 Hui and 33 Uygur) children without obesity were recruited from six primary schools in the sub-urban areas of Urumqi. Venous blood was sampled from all subjects after fasting for 12 hours. Leptin gene C2549A polymorphism was determined by polymerase chain reaction (PCR) and restriction fragment length polymorphism methods. Blood concentrations of lipids, leptin and insulin were measured with biochemical methods and radioimmunoassys, respectively. RESULTS: In the 137 children tested, the prevalence of AA, AC and CC genotype was 9.5%, 33.6% and 56.9%, respectively. A allele frequency was significantly different between the two ethnic (i.e. Hui and Uygur) groups (P<0.05). A allele frequency and AA+ AC genotype frequency were not significantly different between obese and non-obese children in both ethnic groups (P>0.05). Blood leptin levels were not significantly different between obese and non-obese children with an AA+AC or CC genotype in both ethnic groups (P>0.05). CONCLUSIONS: Leptin gene polymorphisms exist in Hui and Uygur children. The C2549A polymorphism is not significantly associated with the prevalence of obesity in both Hui and Uygur children.

A validated model to predict spread through air space in lung adenocarcinoma.

Background: Spread through air space (STAS) is currently considered to be a significant predictor of a poor outcome of pulmonary adenocarcinoma. Preoperative prediction of STAS is of great importance for treatment planning. The aim of the present study was to establish a nomogram based on computed tomography (CT) features for predicting STAS in lung adenocarcinoma and to assess the prognosis of the patients with STAS. Methods: A retrospective cohort study was performed in Wuhan Union Hospital from December 2015 to March 2021. The sample was divided into training and testing cohorts. Clinicopathologic and radiologic variables were recorded. The independent risk factors for STAS were determined by stepwise regression and then incorporated into the nomogram. Receiver operating characteristic (ROC) curves and calibration curves analysed by the Hosmer-Lemeshow test were used to evaluate the performance of the model. Decision curve analysis (DCA) was conducted to determine the clinical value of the nomogram. The Kaplan-Meier method was used for survival analysis and the multivariable Cox proportional hazards regression model was used to identify independent predictors for recurrence-free survival (RFS) and overall survival (OS). Results: The sample included 244 patients who underwent surgical resection for primary lung adenocarcinoma. The training cohort included 199 patients (68 STAS-positive and 131 STAS-negative patients), and the testing cohort included 45 patients (15 STAS-positive and 30 STAS-negative patients). The preoperative CT features associated with STAS were shape, ground-glass opacity (GGO) ratio and spicules. The nomogram including these three factors had good discriminative power, and the areas under the ROC curve were 0.875 and 0.922 for the training and testing data sets, respectively, with well-fitted calibration curves. DCA showed that the nomogram was clinically useful. STAS-positive patients had significantly worse OS and RFS than STAS-negative patients (both P<0.01). OS and RFS at 5-year for STAS-positive patients were 63.1% and 59.5%, respectively. Multivariate analysis showed that age [hazard ratio (HR), 1.1; 95% confidence interval (CI): 1.035-1.169; P=0.002], diameter (HR, 1.06; 95% CI: 1.04-1.11; P=0.03) and surgical margin (HR, 32.8; 95% CI: 6.8-158.3; P<0.001) were independent risk factors for OS. Adjuvant therapy (HR, 7.345; 95% CI: 2.52-21.41; P<0.001), N stage (N2) (HR, 0.239; 95% CI: 0.069-0.828; P=0.02) and surgical margin (HR, 15.6; 95% CI: 5.9-41.1; P<0.001) were found to be independent risk factors for RFS. Conclusions: The outcome of STAS-positive patients was worse. The nomogram incorporating the identified CT features could be applied to facilitate individualized preoperative prediction of STAS and selection of rational therapy.

Integrated metabolomics revealed the photothermal therapy of melanoma by Mo2C nanosheets: toward rehabilitated homeostasis in metabolome combined lipidome.

Melanoma, the most aggressive and life-threatening form of skin cancer, lacks innovative therapeutic approaches and deeper bioinformation. In this study, we developed a photothermal therapy (PTT) based on Mo2C nanosheets to eliminate melanoma while utilizing integrated metabolomics to investigate the metabolic shift of metabolome combined lipidome during PTT at the molecular level. Our results demonstrated that 1 mg ml-1 Mo2C nanosheets could efficiently convert laser energy into heat with a strong and stable photothermal effect (74 ± 0.9 °C within 7 cycles). Furthermore, Mo2C-based PTT led to a rapid decrease in melanoma volume (from 3.299 to 0 cm2) on the sixth day, indicating the effective elimination of melanoma. Subsequent integrated metabolomics analysis revealed significant changes in aqueous metabolites (including organic acids, amino acids, fatty acids, and amines) and lipid classes (including phospholipids, lysophospholipids, and sphingolipids), suggesting that melanoma caused substantial fluctuations in both metabolome and lipidome, while Mo2C-based PTT helped improve amino acid metabolism-related biological events (such as tryptophan metabolism) impaired by melanoma. These findings suggest that Mo2C nanosheets hold significant potential as an effective therapeutic agent for skin tumors, such as melanoma. Moreover, through exploring multidimensional bioinformation, integrated metabolomics technology provides novel insights for studying the metabolic effects of tumors, monitoring the correction of metabolic abnormalities by Mo2C nanosheet therapy, and evaluating the therapeutic effect on tumors.

Simultaneous generation of gradients with gradually changed slope in a microfluidic device for quantifying axon response.

Over the past decades, various microfluidic devices have been developed to investigate the role of the molecular gradient in axonal development; however, there are very few devices providing quantitative information about the response of axons to molecular gradients with different slopes. Here, we propose a novel laminar-based microfluidic device enabling simultaneous generation of multiple gradients with gradually changed slope on a single chip. This device, with two asymmetrically designed peripheral channels and opposite flow direction, could generate gradients with gradually changed slope in the center channel, enabling us to investigate simultaneously the response of axons to multiple slope gradients with the same batch of neurons. We quantitatively investigated the response of axon growth rate and growth direction to substrate-bound laminin gradients with different slopes using this single-layer chip. Furthermore, we compartmented this gradient generation chip and a cell culture chip by a porous membrane to investigate quantitatively the response of axon growth rate to the gradient of soluble factor netrin-1. The results suggested that contacting with a molecular gradient would effectively accelerate neurites growth and enhance axonal formation, and the axon guidance ratio obviously increased with the increase of gradient slope in a proper range. The capability of generating a molecular gradient with continuously variable slopes on a single chip would open up opportunities for obtaining quantitative information about the sensitivity of axons and other types of cells in response to gradients of various proteins.

The inflammatory cytokine profiles and ocular biometric characteristics of primary angle-closure glaucoma.

OBJECTIVE: To investigate the pathogenesis of primary angle-closure disease (PACG) by measuring the anatomical structures of the anterior and posterior segments of the eye and inflammatory markers in the peripheral blood. METHODS: This case-control study enrolled patients diagnosed with acute PACG (APACG) and chronic PACG (CPACG). It also enrolled control subjects without PACG. The anterior and posterior anatomical features were measured in all study participants. The levels of interleukin (IL)-6, tumour necrosis factor-α and the neutrophil-to-lymphocyte ratio (NLR) in the peripheral blood were measured. RESULTS: This study analysed a total of 99 eyes: 34 eyes from 34 patients with APACG, 28 eyes from 28 patients with CPACG and 37 eyes from 37 control patients with senile cataract. The axis length, corneal diameter, anterior chamber depth and anterior chamber volume were significantly smaller in the APACG and CPACG groups compared with the controls. The level of IL-6 in the peripheral blood of patients with PACG was significantly lower than that of the controls. The NLR in the peripheral blood of patients with PACG was significantly greater than that of the controls. CONCLUSIONS: Changes in the ocular anatomy and some inflammatory markers might be involved in the pathogenesis of PACG.

PIWI-Interacting RNA Pathway Genes: Potential Biomarkers for Clear Cell Renal Cell Carcinoma.

Background: Clear cell renal cell carcinoma (ccRCC) is one of the most lethal malignancies in the urinary system, yet effective diagnostic and prognostic markers are lacking. Recently, several of piRNA pathway genes have been reported to be associated with cancer diagnosis and prognosis, but their role in ccRCC is still unclear. Methods: We analysed the expression of 27 piRNA pathway genes in 539 kidney renal clear cell carcinoma (KIRC) and 72 nontumor tissue samples (data from TCGA), and 12 mRNAs were significantly different. The aim was to sift the piRNA pathway genes that are correlated with ccRCC patient survival and to construct a piRNA pathway gene risk prognostic model using Kaplan-Meier survival curve and ROC curve, respectively. Results: 5 piRNA pathway genes (TDRD7, GPAT2, PLD6, SUV39H1, and DOM3Z) were picked out and used to construct the piRNA pathway gene risk model. Kaplan-Meier survival curve analysis showed that compared with that of the low-risk group of ccRCC patients, the OS of the high-risk group of ccRCC patients was significantly reduced. The predictive performance of the prognostic risk model was measured using a ROC curve, which individually showed AUC values for 1 year of 0.707, for 3 years of 0.713, and for 5 years of 0.701. Moreover, the mRNA and protein expression levels of TDRD7 were overexpressed in the ccRCC datasets (data from our cohort, TCGA, GEO, and CPTAC) and ccRCC cell lines, and the expression levels correlated with the clinicopathological characteristics in ccRCC. The Tumor Immune Estimation Resource (TIMER) showed that the mRNA expression level of TDRD7 was positively related to tumor immune infiltrating cells (TICs) in ccRCC. Mechanistically, gene set enrichment analysis (GSEA) was performed to uncover the mechanism of TDRD7 in ccRCC. In summary, the piRNA pathway genes,especially TDRD7, may be potential cancer diagnostic and prognostic biomarkers of ccRCC.

Site-specific N-glycosylation characterization of micro monoclonal immunoglobulins based on EThcD-sceHCD-MS/MS.

Monoclonal immunoglobulin produced by clonal plasma cells is the main cause in multiple myeloma and monoclonal gammopathy of renal significance. Because of the complicated purification method and the low stoichiometry of purified protein and glycans, site-specific N-glycosylation characterization for monoclonal immunoglobulin is still challenging. To profile the site-specific N-glycosylation of monoclonal immunoglobulins is of great interest. Therefore, in this study, we presented an integrated workflow for micro monoclonal IgA and IgG purification from patients with multiple myeloma in the HYDRASYS system, in-agarose-gel digestion, LC-MS/MS analysis without intact N-glycopeptide enrichment, and compared the identification performance of different mass spectrometry dissociation methods (EThcD-sceHCD, sceHCD, EThcD and sceHCD-pd-ETD). The results showed that EThcD-sceHCD was a better choice for site-specific N-glycosylation characterization of micro in-agarose-gel immunoglobulins (~2 µg) because it can cover more unique intact N-glycopeptides (37 and 50 intact N-glycopeptides from IgA1 and IgG2, respectively) and provide more high-quality spectra than sceHCD, EThcD and sceHCD-pd-ETD. We demonstrated the benefits of the alternative strategy in site-specific N-glycosylation characterizing micro monoclonal immunoglobulins obtained from bands separated by electrophoresis. This work could promote the development of clinical N-glycoproteomics and related immunology.

Deep diagnostic agent forest (DDAF): A deep learning pathogen recognition system for pneumonia based on CT.

BACKGROUND: Even though antibiotics agents are widely used, pneumonia is still one of the most common causes of death around the world. Some severe, fast-spreading pneumonia can even cause huge influence on global economy and life security. In order to give optimal medication regimens and prevent infectious pneumonia's spreading, recognition of pathogens is important. METHOD: In this single-institution retrospective study, 2,353 patients with their CT volumes are included, each of whom was infected by one of 12 known kinds of pathogens. We propose Deep Diagnostic Agent Forest (DDAF) to recognize the pathogen of a patient based on ones' CT volume, which is a challenging multiclass classification problem, with large intraclass variations and small interclass variations and very imbalanced data. RESULTS: The model achieves 0.899 ± 0.004 multi-way area under curves of receiver (AUC) for level-I pathogen recognition, which are five rough groups of pathogens, and 0.851 ± 0.003 AUC for level-II recognition, which are 12 fine-level pathogens. The model also outperforms the average result of seven human readers in level-I recognition and outperforms all readers in level-II recognition, who can only reach an average result of 7.71 ± 4.10% accuracy. CONCLUSION: Deep learning model can help in recognition pathogens using CTs only, which might help accelerate the process of etiological diagnosis.