Prognosis of pediatric BCP-ALL with IKZF1 deletions and impact of intensive chemotherapy: Results of SCCLG-2016 study.

BACKGROUND: IKZF1 deletion (IKZF1del) is associated with poor prognosis in B-cell precursor acute lymphoblastic leukemia (BCP-ALL). But the prognosis of IKZF1del combined with other prognostic stratification factors remains unclear. Whether intensified treatment improves BCP-ALL prognosis has not been determined. METHODS: A retrospective analysis was performed on 1291 pediatric patients diagnosed with BCP-ALL and treated with the South China Children's Leukemia 2016 protocol. Patients were stratified based on IKZF1 status for comparison of characteristics and outcome. Additionally, IKZF1del patients were further divided based on chemotherapy intensity for outcome assessments. RESULTS: The BCP-ALL pediatric patients with IKZF1del in south China showed poorer early response. Notably, the DFS and OS for IKZF1del patients were markedly lower than IKZF1wt group (3-year DFS: 88.7% [95% CI: 83.4%-94.0%] vs. 93.5% [95% CI: 92.0%-94.9%], P = .021; 3-year OS: 90.7% [95% CI: 85.8% to 95.6%] vs. 96.1% [95% CI: 95% to 97.2%, P = .003]), with a concurrent increase in 3-year TRM (6.4% [95% CI: 2.3%-10.5%] vs. 2.9% [95% CI: 1.9%-3.8%], P = .025). However, the 3-year CIR was comparable between the two groups (5.7% [95% CI: 1.8%-9.5%] vs. 3.7% [95% CI: 2.6%-4.7%], P = .138). Subgroup analyses reveal no factor significantly influenced the prognosis of the IKZF1del cohort. Noteworthy, intensive chemotherapy improved DFS from 85.7% ± 4.1% to 94.1% ± 0.7% in IKZF1del group (P = .084). Particularly in BCR::ABL positive subgroup, the 3-year DFS was remarkably improved from 53.6% ± 20.1% with non-intensive chemotherapy to 100% with intensive chemotherapy (P = .026). CONCLUSIONS: Pediatric BCP-ALL patients with IKZF1del in South China manifest poor outcomes without independent prognostic significance. While no factor substantially alters the prognosis in the IKZF1del group. Intensified chemotherapy may reduce relapse rates and improve DFS in patients with IKZF1del subset, particularly in IKZFdel patients with BCR::ABL positive.

From the Public Health Perspective: a Scalable Model for Improving Epidemiological Testing Efficacy in Low- and Middle-Income Areas.

The globe is an organically linked whole, and in the pandemic era, COVID-19 has brought heavy public safety threats and economic costs to humanity as almost all countries began to pay more attention to taking steps to minimize the risk of harm to society from sudden-onset diseases. It is worth noting that in some low- and middle-income areas, where the environment for epidemic detection is complex, the causative and comorbid factors are numerous, and where public health resources are scarce. It is often more difficult than in other areas to obtain timely and effective detection and control in the event of widespread virus transmission, which, in turn, is a constant threat to local and global public health security. Pandemics are preventable through effective disease surveillance systems, with nonpharmacological interventions (NPIs) as the mainstay of the control system, effectively controlling the spread of epidemics and preventing larger outbreaks. However, current state-of-the-art NPIs are not applicable in low- and middle-income areas and tend to be decentralized and costly. Based on a 3-year case study of SARS-CoV-2 preventive detection in low-income areas in south-central China, we explored a strategic model for enhancing disease detection efficacy in low- and middle-income areas. For the first time, we propose an integrated and comprehensive approach that covers structural, social, and personal strategies to optimize the epidemic surveillance system in low- and middle-income areas. This model can improve the local epidemic detection efficiency, ensure the health care needs of more people, reduce the public health costs in low- and middle-income areas in a coordinated manner, and ensure and strengthen local public health security sustainably.

The RAS-signaling-pathway-mutation-related prognosis in B-cell acute lymphoblastic leukemia: A report from South China children's leukemia group.

The next-generation sequencing technologies application discovers novel genetic alterations frequently in pediatric acute lymphoblastic leukemia (ALL). RAS signaling pathway mutations at the time of relapse ALL frequently appear as small subclones at the time of onset, which are considered as the drivers in ALL relapse. Whether subclones alterations in the RAS signaling pathway should be considered for risk group stratification of ALL treatment is not decided yet. In this work, we investigate the RAS signaling pathway mutation spectrum and the related prognosis in pediatric ALL. We employed an NGS panel comprising 220 genes. NGS results were collected from 202 pediatric ALL patients. 155 patients (76.7%) harbored at least one mutation. The incidences of RAS signaling pathway mutations are different significantly between T-ALL and B-ALL. In B-ALL, the RAS pathway is mostly involved, and NRAS (17.6%), KRAS (22.7%), and PTPN11 (7.7%) were the three most frequently mutated genes. Co-occurring mutations of CREBBP and NRAS, FLT3, or PTPN11 (p = 0.002, p = 0.009, and p = 0.003, respectively) were found in this cohort. The 3-year RFS rates for the RAS signaling pathway mutation-positive and negative cases was 76.5 % versus 89.7 % (p = 0.012). Four cases relapsed in the lately 3 years were RAS signaling pathway mutation-positive. RAS signaling pathway mutation is an important biomarker for poorer relapse-free survival in pediatric B-ALL patients despite good early MRD levels.

DeKinomics pulse-chases kinase functions in living cells.

Cellular context is crucial for understanding the complex and dynamic kinase functions in health and disease. Systematic dissection of kinase-mediated cellular processes requires rapid and precise stimulation ('pulse') of a kinase of interest, as well as global and in-depth characterization ('chase') of the perturbed proteome under living conditions. Here we developed an optogenetic 'pulse-chase' strategy, termed decaging kinase coupled proteomics (DeKinomics), for proteome-wide profiling of kinase-driven phosphorylation at second-timescale in living cells. We took advantage of the 'gain-of-function' feature of DeKinomics to identify direct kinase substrates and further portrayed the global phosphorylation of understudied receptor tyrosine kinases under native cellular settings. DeKinomics offered a general activation-based strategy to study kinase functions with high specificity and temporal resolution under living conditions.

Current landscape and future prospects of RET and ROS1 targets.

RET and ROS1 are becoming key targets for targeted therapy. To show current landscape of ROS1 and RET targets, a patent analysis was performed. The present results indicated that inhibitor structures of ROS1 target demonstrated unique elements compared with inhibitor structures of RET or BRAF targets. Our study was the first time to uncover that a number of inhibitor structures of ROS1 target contained sulfur and boron elements. The inhibitors of RET target could be developed for treatment of various cancers, including lung cancer, thyroid cancer, and other solid tumor, while the inhibitors of ROS1 target are virtually developed for treatment of lung cancer. Our findings provide a new insight for drug discovery of ROS1 and RET target.

Patent landscape of platelet growth factor receptor and c-KIT targets.

Type III receptor tyrosine kinase, e.g., PDGFR, are associated with various autoimmune diseases. To show the status of PDGFR and c-KIT targets, we performed the US patent analysis. The present study showed that the R&D of c-KIT target was much earlier than the R&D of PDGFR targets. Currently, the PDGFR-based target demonstrates more applications in the development of biological therapy. Our findings indicated that some inhibitors of c-KIT target contained sulfur elements or 1,3-diazine rings. The c-KIT target has more competitive edges for chemical drug discovery than the PDGFR target. c-KIT and PDGFR targets are currently preferable for drug discovery in autoimmune diseases. This study was the first to show R&D differentiation between PDGFR and c-KIT targets in drug development.

TIPE2 ameliorates neuroinflammation and cognitive impairment in sepsis-associated encephalopathy through regulating RhoA/ROCK2-NF-κB signaling pathway.

Sepsis-associated encephalopathy (SAE) is an acute brain dysfunction induced by systemic inflammation caused by sepsis and is one of the most common types of encephalopathy in intensive care units. Deteriorative neuroinflammation is closely related to the development of brain injury, which often transforms into common pathological manifestations in patients with severe sepsis. Therefore, taking necessary preventive and protective measures for potential brain injury and promptly reducing neuroinflammatory injury is necessary to improve the long-term prognoses of patients. Tumor necrosis factor-α-induced protein 8-like 2 (TIPE2) can play a significant protective role in septic lung injury, but studies on its expression and role in neurological diseases are rare. In the present study, we found that TIPE2 can expressed in microglia and ameliorate brain injury caused by SAE by suppressing neuroinflammation. The RhoA/ROCK2 pathway is the central coordinator of tissue injury response, and the activation of RhoA participates in the lipopolysaccharide-induced activation of the nuclear factor kappa B (NF-κB) signaling pathway. The activation of RhoA and phosphorylation of NF-κB was enhanced after TIPE2 deficiency. Importantly, TIPE2 negatively regulates inflammatory responses in vivo and in vitro and plays a protective role in SAE by inhibiting the activation of RhoA/ROCK2-NF-κB signaling pathways. The ultimate aim of our proposed project is to provide a theoretical basis for the development of a novel strategy for the early prevention and therapy of SAE.

Exploration for cobalt/nitrogen-doped catalyst to creatinine degradation via peroxymonosulfate activation: toxicity evaluation, statistical modeling, and mechanisms study.

Developing multifunctional catalysts applied in diversiform modes via advanced oxidation processes (AOPs) is a promising and attractive approach for organic pollution degradation. Herein, a novel hollow bamboo-like structural cobalt/nitrogen-doped carbonized material (CoC/N) was employed as a catalyst for AOPs, in which CoC/N was prepared in situ through calcining a Co-based coordination polymer. When CoC/N was utilized as a peroxymonosulfate (PMS) activator, the catalyst stood out prominent activities for effective CA oxidation. Furthermore, a five-level central composite rotatable design (CCRD) model describing CA decay as a function of PMS concentration, CoC/N dosage, and solution pH value was successfully constructed and engaged to explore the optimal operating conditions. Finally, the possible degradation mechanism of CA in CoC/N-PMS system was proposed by quantum chemistry calculation and LC/MS analysis. This work shed light on the structural morphology of the catalyst and its PMS synergy degradation pathway, which promotes its applications in miscellaneous pollutant degradation. A new Co/N-doped material was used to degrade unconventionality organic pollutant creatinine (CA) for the first time, in which the scientific approaches of five-level central composite rotatable design (CCRD) model, response surface methodology (RSM) and density function theory (DFT) were employed to evaluate the material performance and CA degradation pathway. The toxicity evaluation, statistical modeling and mechanisms study have been investigated meticulously.

Age, pelvic incidence, facet joint angle and pedicle-facet angle as correlative factors for isthmic spondylolisthesis: a retrospective case control study.

BACKGROUND: Isthmic spondylolisthesis (IS) is a common clinical disease with a high incidence rate. However, most current researches explain the clear pathogenesis from a single perspective. The aim of our study was to explore the relationships between multiple parameters in patients and find the potential risk factors of this disease. METHODS: Our study retrospectively included 115 patients who were diagnosed with isthmic spondylolisthesis and the same number of individuals without spondylolisthesis. The following parameters were measured or collected: age, pelvic incidence (PI), facet joint angle (FJA) and pedicle-facet angle (P-F angle). The radiographic files were imported into Mimics Medical 20.0 and all collected data were analyzed using SPSS, version 26.0, statistical software. RESULTS: The age was higher in IS group than in control group. PI was also higher in the IS group (50.99 ± 7.67) than in the control group (43.77 ± 9.30) significantly (P = 0.009). There was significant difference in cranial and average FJA tropism in L3-L4 level (P = 0.002, P = 0.006, respectively) and in L4-L5 level (P < 0.001). P-F angle of L4-L5 level showed significantly larger in IS group than in control group (P = 0.007).The logistic regression analysis showed a larger age, a greater L3-L4 cranial FJA tropism, and a greater L4-L5 cranial FJA tropism were potential predictors of IS, with an OR of 1.07, 1.28, and 1.39 respectively. The thresholds of the predictors were 60 years, 5.67°, and 8.97° according to the ROC curve. The linear regression equation was established: degree of slippage (%) = 0.220*age - 0.327* L3-4 cranial FJA tropism - 0.346* L4-5 average FJA tropism (F = 3.460, P = 0.011, r = 0.659). CONCLUSIONS: Our study revealed that isthmic spondylolisthesis may be related to multiple factors rather than a single factor. Age, PI, PJA and P-F angle are potentially associated with spondylolisthesis.

Optimal Shortcuts to Adiabatic Control by Lagrange Mechanics.

We combined an inverse engineering technique based on Lagrange mechanics and optimal control theory to design an optimal trajectory that can transport a cartpole in a fast and stable way. For classical control, we used the relative displacement between the ball and the trolley as the controller to study the anharmonic effect of the cartpole. Under this constraint, we used the time minimization principle in optimal control theory to find the optimal trajectory, and the solution of time minimization is the bang-bang form, which ensures that the pendulum is in a vertical upward position at the initial and the final moments and oscillates in a small angle range.

Relationship between methylenetetrahydrofolate reductase gene polymorphisms and methotrexate drug metabolism and toxicity.

Background: Acute lymphoblastic leukemia (ALL) is the most common malignancy in children, and methotrexate (MTX) is the key drug for ALL. Studies on the relationship between High-Dose methotrexate (HD-MTX) toxicity and methylenetetrahydrofolate reductase (MTHFR) C677T and A1298C genes have drawn different conclusions. This study aimed to investigate the relationship between the polymorphism of MTHFR C677T and A1298C genes and the toxicity responses of MTX. Methods: The MTHFR C677T and A1298C genotypes of 271 children with ALL who received HD-MTX chemotherapy in southern China from September 2017 to June 2021 were analyzed, and the toxicity of HD-MTX was evaluated and analyzed according to Common Terminology Criteria for Adverse Events (CTCAE) 5.0. Results: The MTHFR C677T and A1298C gene polymorphisms were not correlated with the 48-hour MTX blood concentrations (P>0.05). Unconditional logistic regression model analysis also revealed that the risk of liver function impairment [odds ratio (OR) =1.656, 95% confidence interval (CI): 1.179-2.324, P<0.05] and mucosal damage (OR =1.508, 95% CI: 1.042-2.183, P<0.05) were 1.656 and 1.508 times higher for the heterozygous mutant (CT), and homozygous mutant (TT) mutant type than for the wild-type (CC), wild-type, respectively. The risk of neutropenia and liver function impairment were 0.498 (OR =0.498, 95% CI: 0.251-0.989, P<0.05) and 6.067 (OR =6.067, 95% CI: 1.183-31.102, P<0.05) times higher in low-risk children with CT+TT mutant genotypes than in those with CC wild genotypes, respectively. Furthermore, the risk of mucosal damage was 1.906 times higher in high-risk children with the CT+TT genotype than in those with the CC genotype (OR =1.906, 95% CI: 1.033-3.518, P<0.05). The MTHFR A1298C genotypes differed in the incidence of liver function damage and gastrointestinal toxic reactions in children with ALL. Nonetheless, no increased risk of liver function impairment nor gastrointestinal reactions in children with the heterozygous mutant (AC)+CC mutation was observed. Conclusions: Advancements in MTHFR genotype testing in children with ALL and the introduction of personalised treatments based on genotype results during HD-MTX chemotherapy will help to predict, prevent, and reduce the occurrence of adverse MTX-related toxic reactions.

Effect of total knee arthroplasty for valgus knee correction on clinical outcome and patellar position.

PURPOSE: The purpose was to investigate the effect of different degrees of valgus deformity correction on patellar position and clinical outcome in patients with valgus knees after total knee arthroplasty (TKA). METHODS: We retrospectively analyzed and followed 118 patients with valgus knees. Based on the post-operative hip-knee-ankle (HKA), patients were divided into three groups: neutral (±3°), mild (3-6°), and severe (> 6°). Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC), range of motion (ROM), and Knee Society Score (KSS) were used to evaluate post-operative clinical efficacy. Also, the patellar tilt angle (ε-angle), congruence angle (θ-angle), and Insall-Salvati index (ISI) were used to represent the patellar position. Post-operative observation indicators included HKA, angle of the femur (α-angle), tibial angle (ß-angle), femoral component flexion angle (γ-angle), and tibial component posterior slope angle (δ-angle). RESULTS: All patients showed significant improvements in HKA, ROM, WOMAC, and KSS after operation (P < 0.001). Regarding patellar position, the ISI values decreased to varying degrees (P < 0.05). The patellar tilt angle was significantly increased in the severe valgus group compared to that in the mild valgus and neutral groups (P < 0.001). Univariate analysis showed that the degree of post-operative residual valgus was significantly affected by WOMAC, KSS, α-, ε-, and θ-angles. CONCLUSION: Minor valgus undercorrection did not affect the short-term outcome after TKA; however, when the residual valgus angle was > 6°, the post-operative scores were significantly reduced. Inadequate valgus correction does not result in significant changes in patellar height but may increase the risk of poor patellar tracking.

Tongguanteng injection reverses paclitaxel resistance via upregulation of TAB1 expression in ovarian cancer in vitro and in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: Tongguanteng injection (TGT), the water extract from the stem of the Traditional Chinese hebal medicine of Marsdenia tenacissima (Roxb.) Wight et Arn. has been used as anticancer remedy for decades. TGT was not only used in the treatment of many malignant cancers extensively, but also an adjuvant anticancer drug with chemotherapeutics clinically. AIM OF THE STUDY: To evaluate the effects of TGT on reversing paclitaxel (PTX) resistance and investigate the potential mechanism related to TAB1 in ovarian cancer (OC) in vitro and in vivo. MATERIALS AND METHODS: The synergistic effect and reversal ratio were determined by CCK8 assay and median-effect principle after the combination of TGT and PTX in OC A2780 and its PTX-resistant (A2780/T) cells. The biological functions in cell apoptosis, migration and invasion of A2780/T cells treated by PTX 4 µM with TGT 20, 40, 80 mg⋅mL-1 for 24 h were evaluated by colony formation, flow cytometry, wound healing and transwell assays. Proteomics technique and bioinformatic analysis were used to indentify the change of TAB1 expression in A2780/T cells induced by TGT. The association between TAB1 expression and human OC was analyzed by gene expression databases. In A2780/T cells, western blotting and colony formation assays were used to investigate the relationship between TAB1 expression and PTX resistance after TAB1 overexpression by TAB1 plasmids. The mechanism of TGT and PTX regulating TAB1 and its related proteins were explored by western blotting and flow cytometry assays after TAB1 knock-down using siTAB1. Moreover, TUNEL staining, immunohistochemistry (IHC) and histopathology were used to observe the antitumor effects, TAB1 and p-p38 expression and the tissues impairments in nude mice xenograft model established by A2780/T cells after the co-treatment with TGT and PTX by in vivo. RESULTS: TGT combined with PTX showed the synergistic effect (CI<1), which could reverse the IC50 values of PTX in OC A2780 and A2780/T cells about 23.50 and 6.44 times, respectively. Besides, TGT combined with PTX could significantly inhibit the migration, invasion and promote apoptosis of A2780/T cells. We identified that TGT could induce TAB1 expression in A2780/T cells by proteomics analysis. TAB1 downregulation was significantly associated with tumorigenesis and poor prognosis in OC patients and PTX resistance in A2780/T cells. Furthermore, TGT could activate TAB1/TAK1/p38 MAPK signaling pathway targeting TAB1 and regulate the expression of Bax, Bcl-2 proteins to improve the sensitivity of A2780/T cells to PTX. TGT combined with PTX also showed a greater inhibition in tumor growth than PTX monotherapy in vivo. These promising results show the efficacy of TGT in reversing PTX resistance and provide a potential strategy that targeting TAB1/TAK1/p38 MAPK signaling pathway may improve the chemotherapy sensitivity in OC. CONCLUSIONS: Our results revealed that Tongguanteng injection could reverse paclitaxel resistance and the potential mechanism might be associated with the activation of TAB1/TAK1/p38 MAPK signaling pathway in OC in vitro and in vivo. TAB1 might be a pivotal target for reversing PTX resistance. This study will provide a theoretical basis for the combination of Tongguanteng injection and paclitaxel in clinic.

Metabolic profiling reveals genotype-associated alterations in carotenoid content during banana postharvest ripening.

Banana fruits have attracted considerable attention for health-promoting effects attributed to ubiquitous functional metabolites. However, genotype-dependent accumulation patterns of carotenoids in banana remain largely unclear. Here, we performed a systematic metabolomic investigation of 18 banana cultivars of the AAA, AAB, or ABB genome groups. Our results indicate that the levels of soluble sugars increase during postharvest ripening regardless of genotype, whereas amino acids (AAs) and tricarboxylic acid (TCA) cycle-derived organic acids display genotype-dependent patterns. The levels of AAs derived from the glycolytic pathway increased, whereas those derived from the TCA cycle significantly decreased during ripening. The carotenoid composition in banana pulp was genotype-specific, and the contents of α-carotene were the highest in AAA-genome bananas. Moreover, high α-carotene and ß-carotene contents in banana were correlated with elevated levels of TCA cycle-derived AAs and decreased levels of glycolysis-derived AAs. Taken together, these findings provide a comprehensive understanding of genotype-associated carotenoid accumulation, thereby facilitating the breeding of future high carotenoid banana cultivars.

Proteomic Difference Analysis of Whole Blood and Bloodstains.

OBJECTIVES: To study the changes of protein levels in peripheral blood after it dried. METHODS: The proteins from whole blood and bloodstains were detected by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and normalized by the label-free quantification (LFQ) method. The differential proteins were analyzed by using R 4.2.1 software, limma and edgeR package. The analysis of biological function, signaling pathway and subcellular localization for the differential proteins was then performed. RESULTS: A total of 623 and 596 proteins were detected in whole blood and bloodstains, respectively, of which 31 were statistically significant in the quantitative results, including 10 up-regulated and 21 down-regulated proteins in bloodstains. CONCLUSIONS: The protein abundances in whole blood and bloodstains are highly correlated, and the variation of protein abundances may be related to the changes of endogenous and structural proteins in cells. The application of proteomics technology can assist the screening and identification of protein biomarkers, thereby introducing new biomarkers for forensic research.

Linolenic Acid-Derived Oxylipins Inhibit Aflatoxin Biosynthesis in Aspergillus flavus through Activation of Imizoquin Biosynthesis.

Oxylipins play important signaling roles in aflatoxin (AF) biosynthesis in Aspergillus flavus. We previously showed that exogenous supply of autoxidated linolenic acid (AL) inhibited AF biosynthesis in A. flavus via oxylipins, but the molecular mechanism is still unknown. Here, we performed multiomics analyses of A. flavus grown in media with or without AL. Targeted metabolite analyses and quantitative reverse transcription (qRT)-polymerase chain reaction (PCR) showed that the imizoquin (IMQ) biosynthetic pathway was distinctly upregulated in the presence of AL. 13C-glucose labeling confirmed in parallel that the tricarboxylic acid cycle was also enhanced by AL, consistent with observed increases in mycelial growth. Moreover, we integrated thermal proteome profiling and molecular dynamics simulations to identify a potential receptor of AL; AL was found to interact with a transporter (ImqJ) located in the IMQ gene cluster, primarily through hydrophobic interactions. Further analyses of strains with an IMQ pathway transcription factor overexpressed or knocked out confirmed that this pathway was critical for AL-mediated inhibition of AF biosynthesis. Comparison of 22 assembled A. flavus and Aspergillus oryzae genomes showed that genes involved in the IMQ pathway were positively selected in A. oryzae. Taken together, the results of our study provide novel insights into oxylipin-mediated regulation of AF biosynthesis and suggest potential methods for preventing AF contamination of crops.

Dynamic Phosphotyrosine-Dependent Signaling Profiling in Living Cells by Two-Dimensional Proximity Proteomics.

Tyrosine phosphorylation (pTyr)-dependent signaling pathways play a vital role in various biological processes, which are spatiotemporally assembled and dynamically regulated on a minute scale by pTyr in living cells. Studying these pTyr-mediated signaling complexes is therefore challenging due to the highly dynamic nature of the protein complexes and the low abundance of pTyr. In this study, we adopted minute-resolution APEX2-based proximity labeling (PL) in living cells and Src SH2 superbinder-based pTyr peptide enrichment for simultaneously profiling these protein complexes and associated pTyr sites from the same affinity-purified sample. Upon different time courses of EGF stimulation of the living cells stably expressing APEX2-FLAG-GRB2, we constructed two-dimensional time-course curves for both interactome and tyrosine phosphoproteome. Well-annotated pTyr signaling complexes in EGFR signaling and located at the endosome were quantified with tightly correlated time-course curves for both interacting proteins and pTyr sites. Importantly, the correlated time-course curves for EGFR and endosomal HGS were well validated by targeted-parallel reaction monitoring (PRM)-MS analysis. Taking advantage of the high sensitivity of the PRM assay, the low-abundant pTyr peptide EGFR pY1110, which cannot be identified in the data-dependent acquisition (DDA) analysis, could be well quantified. Collectively, this two-dimensional proximity proteomic strategy is promising for comprehensively characterizing pTyr-mediated protein complexes with high sensitivity in living cells.

Efficiency Analysis of Uncle-Nephew Relationship Identification by Increasing STR Markers and Adding Reference Samples.

OBJECTIVES: To estimate the system efficiency of uncle-nephew relationship identification by increasing STR markers and adding reference samples based on the test results of simulated data and real samples, so as to provide references for selecting the appropriate number of STRs and reference samples for uncle-nephew relationship identification. METHODS: Five common models of uncle-nephew relationship identification were constructed by adding different reference samples. In each model, the likelihood ratio (LR) for 10 000 pairs of uncle-nephew relationships and 10 000 pairs of unrelated individuals were simulated by detecting 19, 39 or 55 STRs, and the system efficiency at different thresholds was simulated. The samples of the Han population in Zhejiang were collected, and 55 autosomal STRs were obtained by using SiFaSTRTM 23plex kit, Goldeneye® DNA ID 22NC kit and AGCU 21+1 PCR amplification kit. When 19, 39 and 55 STRs were detected, the LR of each model and system efficiency under different thresholds were calculated and compared with the simulation results. RESULTS: Under the same detection system, the calculated results of simulated data and corresponding true samples were basically consistent. In the same model, there was a positive correlation between the system efficiency of uncle-nephew relationship identification and the number of STRs detected. Moreover, the system efficiency of introducing relatives was higher than identifying only two individuals. The order of preference for the introduction of relatives was the full sibling (or mother) of the uncle and the full sibling (or mother) of the nephew. CONCLUSIONS: The system efficiency of uncle-nephew relationship identification could be improved by increasing the number of STRs and introducing known relatives, which would provide the basis for selecting the most appropriate detection system and reference individuals in actual cases.

Integrated and High-Throughput Approach for Sensitive Analysis of Tyrosine Phosphoproteome.

Tyrosine phosphorylation (pTyr) regulates various signaling pathways under normal and cancerous states. Due to their low abundance and transient and dynamic natures, systematic profiling of pTyr sites is challenging. Antibody and engineered binding domain-based approaches have been well applied to pTyr peptide enrichment. However, traditional methods have the disadvantage of a long sample preparation process, which makes them unsuitable for processing limited amount of samples, especially in a high-throughput manner. In this study we developed a 96-well microplate-based approach to integrate all the sample preparation steps starting from cell culture to MS-compatible pTyr peptide enrichment in three consecutive 96-well microplates. By assembling an engineered SH2 domain onto a microplate, nonspecific adsorption of phosphopeptides is greatly reduced, which allows us to remove the Ti-IMAC purification and three C18 desalting steps (after digestion, pTyr enrichment, and Ti-IMAC purification) and, therefore, greatly simplifies the entire pTyr peptide enrichment workflow, especially when processing a large number of samples. Starting with 96-well microplate-cultured, pervanadate-stimulated cells, our approach could enrich 21% more pTyr sites than the traditional serial pTyr enrichment approach and showed good sensitivity and reproducibility in the range of 200 ng to 200 µg peptides. Importantly, we applied this approach to profile tyrosine kinase inhibitor-mediated EGFR signaling pathway and could well differentiate the distinct response of different pTyr sites. Collectively, the integrated 96-well microplate-based approach is valuable for profiling pTyr sites from limited biological samples and in a high-throughput manner.

Integrated Analysis of Gene Co-Expression Network and Prediction Model Indicates Immune-Related Roles of the Identified Biomarkers in Sepsis and Sepsis-Induced Acute Respiratory Distress Syndrome.

Sepsis is a series of clinical syndromes caused by immunological response to severe infection. As the most important and common complication of sepsis, acute respiratory distress syndrome (ARDS) is associated with poor outcomes and high medical expenses. However, well-described studies of analysis-based researches, especially related bioinformatics analysis on revealing specific targets and underlying molecular mechanisms of sepsis and sepsis-induced ARDS (sepsis/se-ARDS), still remain limited and delayed despite the era of data-driven medicine. In this report, weight gene co-expression network based on data from a public database was constructed to identify the key modules and screen the hub genes. Functional annotation by enrichment analysis of the modular genes also demonstrated the key biological processes and signaling pathway; among which, extensive immune-involved enrichment was remarkably associated with sepsis/se-ARDS. Based on the differential expression analysis, least absolute shrink and selection operator, and multivariable logistic regression analysis of the screened hub genes, SIGLEC9, TSPO, CKS1B and PTTG3P were identified as the candidate biomarkers for the further analysis. Accordingly, a four-gene-based model for diagnostic prediction assessment was established and then developed by sepsis/se-ARDS risk nomogram, whose efficiency was verified by calibration curves and decision curve analyses. In addition, various machine learning algorithms were also applied to develop extra models based on the four genes. Receiver operating characteristic curve analysis proved the great diagnostic and predictive performance of these models, and the multivariable logistic regression of the model was still found to be the best as further verified again by the internal test, training, and external validation cohorts. During the development of sepsis/se-ARDS, the expressions of the identified biomarkers including SIGLEC9, TSPO, CKS1B and PTTG3P were all regulated remarkably and generally exhibited notable correlations with the stages of sepsis/se-ARDS. Moreover, the expression levels of these four genes were substantially correlated during sepsis/se-ARDS. Analysis of immune infiltration showed that multiple immune cells, neutrophils and monocytes in particular, might be closely involved in the process of sepsis/se-ARDS. Besides, SIGLEC9, TSPO, CKS1B and PTTG3P were considerably correlated with the infiltration of various immune cells including neutrophils and monocytes during sepsis/se-ARDS. The discovery of relevant gene co-expression network and immune signatures might provide novel insights into the pathophysiology of sepsis/se-ARDS.