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Abstract Recent advancements in computer vision within the field of artificial intelligence (AI) have made significant inroads into the medical domain. However, the application of AI for classifying renal pathology remains challenging due to the subtle variations in multiple renal pathological classifications. Vision Transformers (ViT), an adaptation of the Transformer model for image recognition, have demonstrated superior capabilities in capturing global features and providing greater explainability. In our study, we developed a ViT model using a diverse set of stained renal histopathology images to evaluate its effectiveness in classifying renal pathology. A total of 1861 whole slide images (WSI) stained with HE, MASSON, PAS, and PASM were collected from 635 patients. Renal tissue images were then extracted, tiled, and categorized into 14 classes on the basis of renal pathology. We employed the classic ViT model from the Timm library, utilizing images sized 384 × 384 pixels with 16 × 16 pixel patches, to train the classification model. A comparative analysis was conducted to evaluate the performance of the ViT model against traditional convolutional neural network (CNN) models. The results indicated that the ViT model demonstrated superior recognition ability (accuracy: 0.96–0.99). Furthermore, we visualized the identification process of the ViT models to investigate potentially significant pathological ultrastructures. Our study demonstrated that ViT models outperformed CNN models in accurately classifying renal pathology. Additionally, ViT models are able to focus on specific, significant structures within renal histopathology, which could be crucial for identifying novel and meaningful pathological features in the diagnosis and treatment of renal disease.

Abstract Background Right aortic arch (RAA) is a common congenital aortic arch abnormality. Fetuses with RAA frequently have good outcomes after birth. However, chromosomal abnormalities and genetic syndromes suggest poor prognosis for these patients. So far the underlying genetic etiology is still not identified in most RAA patients based on traditional genetic techniques and a problem is still debated whether fetuses with isolated RAA should be referred for CMA. Our study aims to investigate the genetic etiology of fetuses with right aortic arch (RAA) by chromosomal microarray analysis (CMA) and whole exome sequencing (WES) and evaluate the efficacy of CMA in fetal isolated RAA. Results Among these 153 fetuses, 99 (64.7%) with isolated RAA and 54 (35.3%) with non-isolated RAA; 25.5% (39/153) with additional intracardiac anomalies (ICA), and 19.0% (29/153) with extracardiac anomalies (ECA). Tetralogy of Fallot (n = 10) and persistent left superior vena cava (n = 11) are the most common ICA and ECA, respectively. CMA detected 15 clinically significant copy number variations (CNVs) in 14 cases (9.2%); microdeletion of 22q11.21 was the most common pathogenic CNVs (7.8%). The chromosomal abnormalities rate was higher in non-isolated RAA and RAA with ICA groups than in isolated RAA group (16.7% vs. 5.1%; 20% vs. 5.1%, both p < 0.05). From five cases further undergoing WES, a diagnostic variant in MTOR gene (c.7255G > A, de novo) was first reported in prenatal, extending the prenatal manifestation of Smith–Kingsmore syndrome (OMIM: 616638); a clinically relevant variant c.3407A > T in STAG2 was identified, being inherited from the healthy mother. Moreover, the premature birth and termination rates were higher in non-isolated RAA group than in isolated RAA group (11.1% vs. 1.0%; 37.0% vs. 2.0%, both p < 0.01). Conclusions We demonstrate that CMA and WES are useful diagnostic tools for fetal RAA, particularly non-isolated RAA, and all fetuses with RAA should be referred for CMA. The data probably aids in prenatal diagnosis and prenatal counseling of fetal RAA.

Abstract Backgroud A systematic analysis was conducted to investigate the molecular etiology of fetal cleft lip and/or palate (CL/P) and the association between various types of CL/P and copy number variations (CNVs), as well as their impact on birth outcomes. Methods In this retrospective study conducted between January 2016 and July 2022, a cohort of pregnancies diagnosed with fetal CL/P was enrolled and comprehensive clinical data for all cases were extracted from our medical record database, including demographic data about the pregnancies, ultrasound findings, results of Chromosomal microarray (CMA), as well as relevant pregnant and perinatal outcomes. Results Among the 358 cases, 32 clinically significant variants in 29 (8.1%) fetuses with CL/P were detected by CMA. In 338 singleton pregnancies, the diagnostic yield of CMA in the context of CL/P fetuses was determined to be 7.7% (26/338). CP cases exhibited a relatively higher prevalence of pathogenic/likely pathogenic CNVs at a rate of 25% (3/12), followed by CLP cases at 8.0% (23/288). Notably, the CL group did not demonstrate any pathogenic/likely pathogenic CNV findings among the examined cases (0/38). The diagnostic rate of clinically significant variants was notably higher in the non-isolated CL/P group than in the isolated CL/P group (11/33, 33.3% vs. 15/305, 4.9%, p

Abstract Background Neuroblastoma is a common pediatric cancer, where preclinical studies suggest that a mesenchymal-like gene expression program contributes to chemotherapy resistance. However, clinical outcomes remain poor, implying we need a better understanding of the relationship between patient tumor heterogeneity and preclinical models. Results Here, we generate single-cell RNA-seq maps of neuroblastoma cell lines, patient-derived xenograft models (PDX), and a genetically engineered mouse model (GEMM). We develop an unsupervised machine learning approach (“automatic consensus nonnegative matrix factorization” (acNMF)) to compare the gene expression programs found in preclinical models to a large cohort of patient tumors. We confirm a weakly expressed, mesenchymal-like program in otherwise adrenergic cancer cells in some pre-treated high-risk patient tumors, but this appears distinct from the presumptive drug-resistance mesenchymal programs evident in cell lines. Surprisingly, however, this weak-mesenchymal-like program is maintained in PDX and could be chemotherapy-induced in our GEMM after only 24 h, suggesting an uncharacterized therapy-escape mechanism. Conclusions Collectively, our findings improve the understanding of how neuroblastoma patient tumor heterogeneity is reflected in preclinical models, provides a comprehensive integrated resource, and a generalizable set of computational methodologies for the joint analysis of clinical and pre-clinical single-cell RNA-seq datasets.

Abstract Background Ciprofol is a promising sedative. This study aims to explore the median effective dose (ED50) of ciprofol in inhibiting responses to fiberoptic bronchoscopy in patients with pulmonary tuberculosis (PTB) of different genders and ages when combined with 0.15 μg/kg sufentanil, and to evaluate its efficacy and safety, providing a reference for the rational use of ciprofol in clinical practice. Methods PTB patients who underwent bronchoscopy examination and treatment at The Third People’s Hospital of Changzhou between May 2023 and June 2023 were selected and divided into four groups using a stratified random method. All patients received intravenous injection of 0.15 μg/kg sufentanil followed by injection of the test dose of ciprofol according to Dixon’s up-and-down method. The initial dose of ciprofol in all four groups was 0.4 mg/kg, with an adjacent ratio of 1:1.1. The next patient received a 10% increase in the dose of ciprofol if the previous patient in the same group experienced positive reactions such as choking cough, frowning, and body movements during the endoscopy. Otherwise, it was judged as a negative reaction, and the next patient received a 10% decrease in the dose of ciprofol. The transition from a positive reaction to a negative reaction was defined as a turning point, and the study of the group was terminated when seven turning points occurred. Hemodynamic parameters, oxygen saturation and adverse reactions were recorded at different time points in all groups. The Probit regression analysis method was used to calculate the ED50 of ciprofol in the four groups and compare between the groups. Results The ED50 of ciprofol combined with 0.15 μg/kg sufentanil for bronchoscopy in the four groups were 0.465 mg/kg, 0.433 mg/kg, 0.420 mg/kg and 0.396 mg/kg, respectively. Conclusion The ED50 of ciprofol used for fiberoptic bronchoscopy varied among PTB patients of different genders and ages. Trial registration The Chinese Clinical Trial Registry, ChiCTR2300071508, Registered on 17 May 2023.

Abstract Background The molecular mechanism of fetal cystic hygroma (CH) is still unclear, and no study has previously reported the transcriptome changes of single cells in CH. In this study, single-cell transcriptome sequencing (scRNA-seq) was used to investigate the characteristics of cell subsets in the lesion tissues of CH patients. Methods Lymphoid tissue collected from CH patients and control donors for scRNA-seq analysis. Differentially expressed gene enrichment in major cell subpopulations as well as cell-cell communication were analyzed. At the same time, the expression and interactions of important VEGF signaling pathway molecules were analyzed, and potential transcription factors that could bind to KDR (VEGFR2) were predicted. Results The results of scRNA-seq showed that fibroblasts accounted for the largest proportion in the lymphatic lesions of CH patients. There was a significant increase in the proportion of lymphatic endothelial cell subsets between the cases and controls. The VEGF signaling pathway is enriched in lymphatic endothelial cells and participates in the regulation of cell-cell communication between lymphatic endothelial cells and other cells. The key regulatory gene KDR in the VEGF signaling pathway is highly expressed in CH patients and interacts with other differentially expressed EDN1, TAGLN, and CLDN5 Finally, we found that STAT1 could bind to the KDR promoter region, which may play an important role in promoting KDR up-regulation. Conclusion Our comprehensive delineation of the cellular composition in tumor tissues of CH patients using single-cell RNA-sequencing identified the enrichment of lymphatic endothelial cells in CH and highlighted the activation of the VEGF signaling pathway in lymphoid endothelial cells as a potential modulator. Simple summary The molecular and cellular pathogenesis of fetal cystic hygroma (CH) remains largely unknown. This study examined the distribution and gene expression signature of each cell subpopulation and the possible role of VEGF signaling in lymphatic endothelial cells in regulating the progression of CH by single-cell transcriptome sequencing. The enrichment of lymphatic endothelial cells in CH and the activation of the VEGF signaling pathway in lymphatic endothelial cells provide some clues to the pathogenesis of CH from the perspective of cell subpopulations.

Abstract Background Patients with hypopharyngeal carcinoma (HPC) have a poor prognosis mainly because of lymphatic metastasis. This research aimed to determine the PKM2 role in lymphatic metastasis in HPC and the underlying molecular mechanism contributing to this phenomenon. Methods PKM2 in HPC was studied for its expression and its likelihood of overall survival using TCGA dataset. Western blotting, qRT-PCR, and IHC were employed to confirm PKM2 expression. Methods including gain- and loss-of-function were used to examine the PKM2 role in HPC metastasis in vitro and in vivo. In vitro and in vivo studies also confirmed lymphatic metastasis’s mechanism. Results Prominent PKM2 overexpression was seen in patients with lymphatic metastasis of HPC, and there was an inherent relationship between a high PKM2 level and poor prognosis. In vitro research showed that knocking down PKM2 decreased tumor cell invasion, migration, and proliferation while promoting apoptosis and inhibiting epithelial-mesenchymal transition, but overexpressing PKM2 had the reverse effect. Animal studies suggested that PKM2 may facilitate tumor development and lymphatic metastasis. Conclusions Our findings suggest that PKM2 may be a tumor’s promoter gene of lymphatic metastasis, which may promote lymphatic metastasis of HPC by regulating epithelial-mesenchymal transition. PKM2 may be a biomarker of metastatic potential, ultimately providing a basis for exploring new therapeutic targets.

Abstract Objective To determine the prevalence of sensitivity to common inhaled and food allergens among children with allergic rhinitis (AR) in Changzhou in eastern China and provide a basis for epidemiological research of pediatric allergic rhinitis and allergen avoidance in this region. Methods This was a retrospective observational study, a total of 1248 children with AR were enrolled at the Third People’s Hospital of Changzhou between January 2018 and December 2019. The serum-specific immunoglobulin E (sIgE) to 19 kinds of inhaled and food allergens and serum total IgE were detected with the AllergyScreen test (Mediwiss Analytic GmbH, Moers, Germany). All participants had a positive reaction to at least one allergen in the test (the sIgE concentration ≥ 0.35 IU/ml). Results Among the patients, 818 (65.54%) were male and 430 (34.46%) were female, with 81 (6.50%) aged 1–3 year, 501 (40.14%) aged 4–7 year, and 666 (53.36%) aged 8–14 year. The positivity rate of inhaled allergens was 80.05% (n = 999), while the positivity rate of food allergens was 66.19% (n = 826). 828 patients (66.35%) were sensitized to multiple allergens. The most common inhaled allergens were Dermatophagoides pteronyssinus (65.38%), mold mix (25.56%), house dust (20.67%), and dog hair dander (13.94%), and the most common food allergens were cow’s milk (30.31%), cashew nut (27.9%), egg (22.68%), and beef (12.98%). With an increase in age, the inhaled allergen positivity rate showed a significant increase (P

Abstract Objective To evaluate the prenatal and perinatal outcome of fetuses with extremely large nuchal translucency (eNT) thickness (≥ 6.5 mm). Methods 193 (0.61%) singleton fetuses with eNT were retrospectively included. Anomaly scan, echocardiography, and chromosomal and genetic test were included in our antenatal investigation. Postnatal follow-up was offered to all newborns. Results Major congenital anomalies included congenital heart defect (32.6%, 63/193), hydrops fetalis (13.5%, 26/193), omphalocele (9.3%, 18/193), and skeletal dysplasia (7.8%, 15/193) et al. Abnormal karyotype was identified in 81/115 (70.4%) cases including Turner syndrome (n = 47), Trisomy 18 (n = 17), Trisomy 21 (n = 9), and Trisomy 13 (n = 3). Chromosomal microarray analysis provided informative results with 3.6% (1/28) incremental diagnostic yield over conventional karyotyping. The diagnostic yield of exome sequencing is 10.0% (2/20). There was no significant increase [Odds Ratio (OR) = 1.974; 95% confidence interval 0.863–4.516; P = 0.104] in the incidence of chromosomal defects despite the presence of other structural anomalies. Only 13 fetuses were successfully followed up and survived at term, no one was found with developmental delay or mental retardation. Conclusions Extremely large NT has a high risk of chromosomal abnormality. CMA and ES improve chromosomal genomic and genetic diagnosis of fetal increased NT. When cytogenetic analysis and morphology assessment are both normal, the outcome is good.

Abstract Background Head and neck squamous cell carcinoma (HNSCC) is highly aggressive with a significant tropism of lymph nodes, which restricts treatment options and negatively impacts patient outcomes. Although progress has been made in understanding the molecular mechanisms underlying lymphatic metastasis (LM), these mechanisms remain elusive. ANXA6 is a scaffold protein that participates in tumor pathogenesis and autophagy regulation; however, how ANXA6 affects autophagy and LM in HNSCC cells remains unknown. Methods RNA sequencing was performed on HNSCC clinical specimens with or without metastasis as well as on The Cancer Genome Atlas dataset to investigate ANXA6 expression and survival. Both in vitro and in vivo studies were performed to investigate the role of ANXA6 in the regulation of LM in HNSCC. The molecular mechanism by which ANXA6 interacts with TRPV2 was examined at the molecular level. Results ANXA6 expression was significantly upregulated in HNSCC patients with LM and higher expression was associated with poor prognosis. ANXA6 overexpression promoted the proliferation and mobility of FaDu and SCC15 cells in vitro; however, ANXA6 knockdown retarded LM in HNSCC in vivo. ANXA6 induced autophagy by inhibiting the AKT/mTOR signaling pathway in HNSCC, thereby regulating the metastatic capability of the disease. Furthermore, ANXA6 expression positively correlated with TRPV2 expression both in vitro and in vivo. Lastly, TRPV2 inhibition reversed ANXA6-induced autophagy and LM. Conclusions These results indicate that the ANXA6/TRPV2 axis facilitates LM in HNSCC by stimulating autophagy. This study provides a theoretical basis for investigating the ANXA6/TRPV2 axis as a potential target for the treatment of HNSCC, as well as a biomarker for predicting LM.

Abstract Background Bone marrow mesenchymal stem cells (BMSCs) are commonly used in regenerative medicine. However, it is not clear whether transplantation of BMSCs can improve cardiac function of the X-Linked Muscular Dystrophy Mice (mdx) and how to detect it. We aimed to investigate the role of speckle tracking echocardiography (STE) in detecting cardiac function of the BMSCs-transplanted mdx in comparison with the untreated mdx. Methods The experimental mice were divided into the BMSCs-transplanted mdx, untreated mdx, and control mice groups (n = 6 per group). The BMSCs were transplanted via tail vein injections into a subset of mdx at 20 weeks of age. After four weeks, the cardiac functional parameters of all the mice in the 3 groups were analyzed by echocardiography. Then, all the mice were sacrificed, and the cardiac tissues were harvested and analyzed by immunofluorescence. The serum biochemical parameters were also analyzed to determine the beneficial effects of BMSCs transplantation. Results Traditional echocardiography parameters did not show statistically significant differences after BMSCs transplantation for the three groups of mice. In comparison with the control group, mdx showed significantly lower left ventricular (LV) STE parameters in both the long-axis and short-axis LV images (P

Abstract Background Exome sequencing (ES) is becoming more widely available in prenatal diagnosis. However, data on its clinical utility and integration into clinical management remain limited in practice. Herein, we report our experience implementing prenatal ES (pES) in a large cohort of fetuses with anomalies detected by ultrasonography using a hospital-based in-house multidisciplinary team (MDT) facilitated by a three-step genotype-driven followed by phenotype-driven analysis framework. Methods We performed pES in 1618 fetal cases with positive ultrasound findings but negative for karyotyping and chromosome microarray analysis between January 2014 and October 2021, including both retrospective (n=565) and prospective (n=1053) cohorts. The diagnostic efficiency and its correlation to organ systems involved, phenotypic spectrum, and the clinical impacts of pES results on pregnancy outcomes were analyzed. Results A genotype-driven followed by phenotype-driven three-step approach was carried out in all trio pES. Step 1, a genotype-driven analysis resulted in a diagnostic rate of 11.6% (187/1618). Step 2, a phenotype-driven comprehensive analysis yielded additional diagnostic findings for another 28 cases (1.7%; 28/1618). In the final step 3, data reanalyses based on new phenotypes and/or clinical requests found molecular diagnosis in 14 additional cases (0.9%; 14/1618). Altogether, 229 fetal cases (14.2%) received a molecular diagnosis, with a higher positive rate in the retrospective than the prospective cohort (17.3% vs. 12.4%, p

Abstract Background Epidermal growth factor receptor (EGFR) is both a driver oncogene and a therapeutic target in advanced head and neck squamous cell carcinoma (HNSCC). However, response to EGFR treatment is inconsistent and lacks markers for treatment prediction. This study investigated EGFR-induced epithelial-to-mesenchymal transition (EMT) as a central parameter in tumor progression and identified novel prognostic and therapeutic targets, and a candidate predictive marker for EGFR therapy response. Methods Transcriptomic profiles were analyzed by RNA sequencing (RNA-seq) following EGFR-mediated EMT in responsive human HNSCC cell lines. Exclusive genes were extracted via differentially expressed genes (DEGs) and a risk score was determined through forward feature selection and Cox regression models in HNSCC cohorts. Functional characterization of selected prognostic genes was conducted in 2D and 3D cellular models, and findings were validated by immunohistochemistry in primary HNSCC. Results An EGFR-mediated EMT gene signature composed of n = 171 genes was identified in responsive cell lines and transferred to the TCGA-HNSCC cohort. A 5-gene risk score comprising DDIT4, FADD, ITGB4, NCEH1, and TIMP1 prognosticated overall survival (OS) in TCGA and was confirmed in independent HNSCC cohorts. The EGFR-mediated EMT signature was distinct from EMT hallmark and partial EMT (pEMT) meta-programs with a differing enrichment pattern in single malignant cells. Molecular characterization showed that ITGB4 was upregulated in primary tumors and metastases compared to normal mucosa and correlated with EGFR/MAPK activity in tumor bulk and single malignant cells. Preferential localization of ITGB4 together with its ligand laminin 5 at tumor-stroma interfaces correlated with increased tumor budding in primary HNSCC tissue sections. In vitro, ITGB4 knock-down reduced EGFR-mediated migration and invasion and ITGB4-antagonizing antibody ASC8 impaired 2D and 3D invasion. Furthermore, a logistic regression model defined ITGB4 as a predictive marker of progression-free survival in response to Cetuximab in recurrent metastatic HNSCC patients. Conclusions EGFR-mediated EMT conveyed through MAPK activation contributes to HNSCC progression upon induction of migration and invasion. A 5-gene risk score based on a novel EGFR-mediated EMT signature prognosticated survival of HNSCC patients and determined ITGB4 as potential therapeutic and predictive target in patients with strong EGFR-mediated EMT.

Abstract Background Lymphatic metastasis was an independent prognostic risk factor for hypopharyngeal carcinoma and was the main cause of treatment failure. The purpose of this study was to screen the differential genes and investigate the mechanism of lymphatic metastasis in hypopharyngeal carcinoma. Methods Transcriptome sequencing was performed on primary tumors of patients, and differential genes were screened by bioinformatics analysis. The expression of differential genes was verified by qRT-PCR, western-blotting and immunohistochemical, and prognostic value was analyzed by Kaplan–Meier and log-rank test and Cox’s test. Next, FADU and SCC15 cell lines were used to demonstrate the function of differential genes both in vitro by EdU, Flow cytometry, Wound Healing and Transwell assays and in vivo by a foot-pad xenograft mice model. Proteomic sequencing was performed to screen relevant targets. In addition, in vitro and in vivo experiments were conducted to verify the mechanism of lymphatic metastasis. Results Results of transcriptome sequencing showed that RAF1 was a significantly differential gene in lymphatic metastasis and was an independent prognostic risk factor. In vitro experiments suggested that decreased expression of RAF1 could inhibit proliferation, migration and invasion of tumor cells and promote apoptosis. In vivo experiments indicated that RAF1 could promote tumor growth and lymphatic metastasis. Proteomic sequencing and subsequent experiments suggested that LAGE1 could promote development of tumor and lymphatic metastasis, and was regulated by RAF1. Conclusions It suggests that RAF1 can promote lymphatic metastasis of hypopharyngeal carcinoma by regulating LAGE1, and provide a basis for the exploring of novel therapeutic target and ultimately provide new guidance for the establishment of intelligent diagnosis and precise treatment of hypopharyngeal carcinoma.

Abstract Background Lymph node metastasis is the main cause of poor prognosis of head and neck squamous carcinoma (HNSCC) patients. N6-methyladenosine (m6A) RNA modification is an emerging epigenetic regulatory mechanism for gene expression, and as a novel m6A reader protein, IGF2BP2 has been implicated in tumor progression and metastasis. However, not much is currently known about the functional roles of IGF2BP2 in HNSCC, and whether IGF2BP2 regulates lymphatic metastasis through m6A modification in HNSCC remains to be determined. Methods The expression and overall survival (OS) probability of m6A-related regulators in HNSCC were analyzed with The Cancer Genome Atlas (TCGA) dataset and GEPIA website tool, respectively. The expression levels of IGF2BP2 were measured in HNSCC tissues and normal adjacent tissues. To study the effects of IGF2BP2 on HNSCC cell metastasis in vitro and in vivo, gain- and loss- of function methods were employed. RIP, MeRIP, luciferase reporter and mRNA stability assays were performed to explore the epigenetic mechanism of IGF2BP2 in HNSCC. Results We investigated 20 m6A-related regulators in HNSCC and discovered that only the overexpression of IGF2BP2 was associated with a poor OS probability and an independent prognostic factor for HNSCC patients. Additionally, we demonstrated that IGF2BP2 was overexpressed in HNSCC tissues, and significantly correlated to lymphatic metastasis and poor prognosis. Functional studies have shown that IGF2BP2 promotes both HNSCC cell migration as well as invasion via the epithelial-mesenchymal transition (EMT) process in vitro, and IGF2BP2 knockdown significantly inhibited lymphatic metastasis and lymphangiogenesis in vivo. Mechanistic investigations revealed that Slug, a key EMT-related transcriptional factor, is the direct target of IGF2BP2, and essential for IGF2BP2-regulated EMT and metastasis in HNSCC. Furthermore, we demonstrated that IGF2BP2 recognizes and binds the m6A site in the coding sequence (CDS) region of Slug and promotes its mRNA stability. Conclusions Collectively, our study uncovers the oncogenic role and potential mechanism of IGF2BP2, which serves as a m6A reader, in controlling lymphatic metastasis and EMT in HNSCC, suggesting that IGF2BP2 may act as a therapeutic target and prognostic biomarker for HNSCC patients with metastasis.

Abstract Background Single-cell RNA sequencing (scRNA-seq) provides new insights to address biological and medical questions, and it will benefit more from the ultralow input RNA or subcellular sequencing. Results Here, we present a highly sensitive library construction protocol for ultralow input RNA sequencing (ulRNA-seq). We systematically evaluate experimental conditions of this protocol, such as reverse transcriptase, template-switching oligos (TSO), and template RNA structure. It was found that Maxima H Minus reverse transcriptase and rN modified TSO, as well as all RNA templates capped with m7G improved the sequencing sensitivity and low abundance gene detection ability. RNA-seq libraries were successfully prepared from total RNA samples as low as 0.5 pg, and more than 2000 genes have been identified. Conclusions The ability of low abundance gene detection and sensitivity were largely enhanced with this optimized protocol. It was also confirmed in single-cell sequencing, that more genes and cell markers were identified compared to conventional sequencing method. We expect that ulRNA-seq will sequence and transcriptome characterization for the subcellular of disease tissue, to find the corresponding treatment plan.

Abstract Background The non-invasive quantitative evaluation of left ventricle (LV) function plays a critical role in clinical cardiology. This study proposes a novel ultrasonic biomechanics method by integrating both LV vortex and wall motion to fully assess and understand the LV structure and function. The purpose of this study was to validate the ultrasonic biomechanics method as a quantifiable approach to evaluate LV function. Methods Firstly, B-mode ultrasound images were acquired and processed, which were utilized to implement parameters for quantifying the LV vortex and wall motion respectively. Next, the parameters were compared in polyvinyl alcohol cryogen (PVA) phantoms with different degree of stiffness corresponding to different freezing and thawing cycles in vitro. Finally, the parameters were computed in vivo during one cardiac cycle to assess the LV function in normal and abnormal subjects in vivo. Results In vitro study, the velocity field of PVA phantom differed with stiffness (varied elasticity modulus). The peak of strain for wall motion decreases with the increase of elasticity modulus, and periodically changed values. Statistical analysis for parameters of vortex dynamics (energy dissipation index, DI; kinetic energy fluctuations, KEF; relative strength, RS; and vorticity, W) based on different elasticity (E) of phantom depicted the good viability of this algorithm. In vivo study, the results confirmed that subjects with LV dysfunction had lower vorticity and strain (S) compared to the normal group. Conclusion Ultrasonic biomechanics method can obtain the vortex and wall motion of left ventricle. The method may have potential clinical value in evaluation of LV dysfunction.

Abstract Background Pathology education provides information on pathology and guides students to become pathologists. Recently, the Ministry of Education of the People’s Republic of China required the establishment of the system of ‘High-quality Online and Offline Courses’, which indicates that online courses will play an important role in higher education. Furthermore, the number of pathologists currently cannot satisfy clinical needs. To solve this health issue and implement the policy from the Ministry of Education, it is necessary to improve the current state of pathology education. First, we need to know students’ opinions of the current courses and their professional choices. Methods Online questionnaires covering the quality of traditional courses, attitudes towards online courses, and suggestions for optimizing courses were designed and applied. Whether students want to become pathologists and the underlying reasons for this interest are also included in this survey. Participants are medical students from certain colleges in Nanjing. The collected data were assessed by statistical analyses, and p-values less than 0.05 were considered significant. Results Of the 342 valid responses, 60.94% of undergraduate students showed their interest in pathology courses, and among them, 48.72% expressed that they may become pathologists. However, the corresponding percentage is only 29.59% in the group without interest. To optimize curricula, the top two suggestions are introducing more clinical cases (undergraduate students, 64.45%; graduate students, 79.09%) and making the classes lively and interesting (undergraduate students, 59.77%; graduate students, 62.79%). Approximately 80.00% of students consider online courses to be good supplementary materials to traditional courses, and approximately half prefer an online-offline mixed learning model. Salary, interest, and employment status are the main factors influencing students’ professional choices. Conclusions Students are generally satisfied with traditional pathology courses, and online courses are good supplementary materials in their opinions. It has been suggested that clinical cases be introduced in classes. It is more likely that students who have an interest in pathology will become pathologists. The data from this survey also show that the main causes of the shortage of pathologists are a lack of engaging work and an unsatisfactory salary.

Abstract Background In order to better help doctors make decision in the clinical setting, research is necessary to connect electronic health record (EHR) with the biomedical literature. Pseudo Relevance Feedback (PRF) is a kind of classical query modification technique that has shown to be effective in many retrieval models and thus suitable for handling terse language and clinical jargons in EHR. Previous work has introduced a set of constraints (axioms) of traditional PRF model. However, in the feedback document, the importance degree of candidate term and the co-occurrence relationship between a candidate term and a query term. Most methods do not consider both of these factors. Intuitively, terms that have higher co-occurrence degree with a query term are more likely to be related to the query topic. Methods In this paper, we incorporate original HAL model into the Rocchio’s model, and propose a new concept of term proximity feedback weight. A HAL-based Rocchio’s model in the query expansion, called HRoc, is proposed. Meanwhile, we design three normalization methods to better incorporate proximity information to query expansion. Finally, we introduce an adaptive parameter to replace the length of sliding window of HAL model, and it can select window size according to document length. Results Based on 2016 TREC Clinical Support medicine dataset, experimental results demonstrate that the proposed HRoc and HRoc_AP models superior to other advanced models, such as PRoc2 and TF-PRF methods on various evaluation metrics. Among them, compared with the Proc2 and TF-PRF models, the MAP of our model is increased by 8.5% and 12.24% respectively, while the F1 score of our model is increased by 7.86% and 9.88% respectively. Conclusions The proposed HRoc model can effectively enhance the precision and the recall rate of Information Retrieval and gets a more precise result than other models. Furthermore, after introducing self-adaptive parameter, the advanced HRoc_AP model uses less hyper-parameters than other models while enjoys an equivalent performance, which greatly improves the efficiency and applicability of the model and thus helps clinicians to retrieve clinical support document effectively.

Abstract Introduction Pregnancy management in women with Wilson disease (WD) remains an important clinical problem. This research was conducted to investigate how to avoid worsening of WD symptoms during pregnancy and increase pregnancy success in women with WD by identifying the best pregnancy management approaches in these patients. Patients and methods The clinical data of 117 pregnancies among 75 women with WD were retrospectively analyzed. Related information of the fetus was also recorded and analyzed. At the same time, regression analysis was performed for data of 22 pregnant women without WD, as normal controls. Results Of a total of 117 pregnancies among the 75 women with WD and 31 pregnancies among the 22 control womenincluded in this study, there were 108 successful pregnancies and 9 spontaneous abortions. Among the 108 successful pregnancies, 97 women a history of copper chelation therapy before pregnancy; all 97 women stopped anti-copper therapy during pregnancy. The nine women with spontaneous abortion had no pre-pregnancy history of copper displacement therapy. The incidence of lower limb edema was higher in the WD group than in normal controls (P = 0.036). Compared with the control group, there was a higher proportion in the WD group of male infants (P = 0.022) and lower average infant birth weight (t = 3.514, P = 0.001). Conclusion It is relatively safe for women with WD patients to become pregnant. The best management method for pregnancy in women with WD may be intensive pre-pregnancy copper chelation therapy and no anti-copper treatment during pregnancy.

Abstract Background Many studies have demonstrated that the geometry of the carotid bifurcation enables prediction of blood flow variation associated with atherosclerotic plaque formation. The phase angle between the arterial wall circumferential strain and its instantaneous wall shear stress is known as stress phase angle (SPA). This parameter is used to evaluate hemodynamic factors of atherogenesis. Note that SPA can be numerically computed for the purpose of locating atherosclerosis in different artery geometries. However, there is no experimental data to verify its role in the location of atherosclerosis in different artery geometries. In this study, we use an ultrasonic biomechanical method to experimentally evaluate the role of SPA for locating atherosclerosis in carotid bifurcation. Results For carotid anthropomorphic vascular phantom experiments, the SPAs of common carotid arteries (CCAs), external carotid arteries (ECAs) and internal carotid arteries (ICAs) are − 148.53 ± 6.92°, − 153.95 ± 5.11°, and − 238.69 ± 1.72°, respectively. The corresponding SPAs are − 173.47 ± 0.065°, − 115.57 ± 4.83° and − 233.9 ± 8.12° for the polyvinyl alcohol (PVA-c) phantoms. In vivo mouse experiments indicated that the wall shear stress and circumferential strain were out of phase in the ICAs (− 280.08 ± 13.12°) to a greater extent as compared to CCAs (− 141.97 ± 8.03°) and ECAs (− 170.07 ± 9.24°). Conclusions The results suggested that SPA may be a useful indicator to locate the atherosclerosis position in carotid bifurcation.

Abstract Background An association between serum complement levels and poor renal prognosis in patients with immunoglobulin A nephropathy (IgAN) remains controversial. Methods We conducted a retrospective study examining the relationship between serum complement levels and prognosis in patients with IgAN. Between 2009 and 2013, patients with biopsy-confirmed IgAN were identified from the Second Affiliated Hospital of Wenzhou Medical College, China, and various parameters were documented during follow-up until 2015. The definition of the primary endpoint was a decrease of estimated glomerular filtration rate (eGFR) more than 30% from their baseline levels. Results A total of 403 patients (55.3% female, average 33.7 months of follow-up) were identified and enrolled, with the primary endpoint occurring in 39 (9.8%) patients. Among the patients selected, 202 (50.1%) received corticosteroid treatment alone or in combination with another immunosuppressant (GS group), while others did not receive immunosuppressive treatment (non-GS group). The incidence of the primary endpoint was slightly lower in the GS group compared to the non-GS group (7.0% versus 12.6%, p = 0.06). Multivariate Cox proportional-hazard regression analyses, adjusting for age, systolic and diastolic blood pressure, 24-h urine protein, and immunosuppressive therapy, showed that serum complement 4 (C4) levels (hazard ratio [HR] 2.4, 95% confidence interval [CI] 1.6-3.8, p

Abstract Background Our study was aimed to explore the clinical implication of chromosome microarray analysis (CMA) in genetically etiological diagnosis of children with congenital heart disease (CHD). Methods A total of 104 children with CHD with or without multiple congenital anomalies (MCA) or intellectual disabilities/developmental delay (ID/DD) but normal karyotype were investigated using Affymetrix CytoScan HD array. Result Pathogenic copy number variations (PCNVs) were identified in 29 children (27.9%). The detection rates in children with simple CHD and complex CHD were 31.1% (19/61) and 23.2% (10/43), respectively. The detection rates of PCNVs were 17.9% (7/39), 20% (5/25), 63.2% (12/19) and 23.8% (5/21) in isolated CHD, CHD plus MCA, CHD plus ID/DD, CHD plus MCA and ID/DD, respectively. The PCNVs rate of CHD plus ID/DD was significantly higher than that of isolated CHD. Two genomic loci including 15q11.2 deletion and 1q43-q44 deletion were considered as CHD locus. The DVL1, SKI, STIM1, CTNNA3 and PLN were identified as candidate genes associated with CHD phenotypes. Conclusion CMA can increase the diagnostic rate and improve the etiological diagnosis in children with CHD. We suggest CMA as a first-tier test in children with CHD, especially in children with CHD plus ID/DD.

Abstract Background Both copy number variant-sequencing (CNV-seq) and karyotype analysis have been used as powerful tools in the genetic aetiology of fetuses with congenital heart diseases (CHD). However, CNV-seq brings clinicians more confusions to interpret the detection results related to CHD with or without extracardiac abnormalities. Hence, we conducted this study to investigate the clinical value of CNV-seq in fetuses with CHD. Results A total of 167 patients with fetal CHD including 36 single CHD (sCHD), 41 compound CHD (cCHD) and 90 non-isolated CHD (niCHD) were recruited into the study. 28 cases (16.77%, 28/167) were revealed with chromosomal abnormalities at the level of karyotype. The pathogenic detection rate (DR) of CNV-seq (23.17%, 19/82) was higher than that of karyotyping (15.85%, 13/82) in 82 cases by CNV-seq and karyotyping simultaneously. The DR of pathogenic copy number variations (PCNVs) (31.43%) was higher in niCHD subgroup than that in sCHD and cCHD (9.52% and 23.08%). Conotruncal defect (CTD) was one of the most common heart malformations with the highest DR of PCNVs (50%) in 7 categories of CHD. In terms of all the pregnancy outcomes, 67 (40.12%) cases were terminated and 100 (59.88%) cases were live neonates. Only two among 34 cases with a pathogenic genetic result chose to continue the pregnancy. Conclusions CNV-seq combined with karyotyping is a reliable and accurate prenatal technique for identifying pathogenic chromosomal abnormalities associated with fetal CHD with or without extracardiac abnormalities, which can assist clinicians to perform detailed genetic counselling with regard to the etiology and related outcomes of CHD.

Abstract Background Ankyrin repeat domain 49 (ANKRD49) has been found to be highly expressed in multiple cancer including lung adenocarcinoma (LUAD) and lung squamous carcinoma (LUSC). However, the function of ANKRD49 in the pathogenesis of NSCLC still remains elusive. Previously, ANKRD49 has been demonstrated to promote the invasion and metastasis of A549 cells, a LUAD cell line, via activating the p38-ATF-2-MMP2/MMP9 pathways. Considering the heterogeneity of tumor cells, the function and mechanism of ANKRD49 in NSCLC need more NSCLC-originated cells to clarify. Methods Real-time qPCR was employed to test ANKRD49 expression levels in nine pairs of fresh NSCLC tissues and the corresponding adjacent normal tissues. The function of ANKRD49 was investigated using overexpression and RNA interference assays in lung adenocarcinoma cell line (NCI-H1299) and lung squamous carcinoma cell line (NCI-H1703) through gelatin zymography, cell counting kit-8, colony formation, wound healing, migration and invasion assays mmunoprecipitation was performed to in vitro. Immunoprecipitation was performed to test the interaction of c-Jun and ATF2. Chromatin immunoprecipitation was conducted to assess the transcriptional regulation of ATF2/c-Jun on MMP-2/9. Moreover, the tumorigenicity of ANKRD49 was evaluated in nude mice models and the involved signal molecular was also measured by immunohistochemical method. Results We found that the levels of ANKRD49 in cancerous tissues were higher than those in adjacent normal tissues. in vitro assay showed that ANKRD49 promoted the migration and invasion of NCI-H1299 and NCI-H1703 cells via enhancing the levels of MMP-2 and MMP-9. Furthermore, ANKRD49 elevated phosphorylation of JNK and then activated c-Jun and ATF2 which interact in nucleus to promote the binding of ATF2:c-Jun with the promoter MMP-2 or MMP-9. In vivo assay showed that ANKRD49 promoted lung metastasis of injected-NSCLC cells and the high metastatic rate was positively correlated with the high expression of ANKRD49, MMP-2, MMP-9, p-JNK, p-c-Jun and p-ATF2. Conclusion The present study indicated that ANKRD49 accelerated the invasion and metastasis of NSCLC cells via JNK-mediated transcription activation of c-Jun and ATF2 which regulated the expression of MMP-2/MMP-9. The molecular mechanisms of ANKRD49’s function is different from those found in A549 cells. The current study is a supplement and improvement to the previous research.

Abstract Background Atherosclerosis is now the main cause of cardiac-cerebral vascular diseases around the world. Disturbances in lipid metabolism have an essential role in the development and progression of atherosclerosis. Thus, we aimed to investigate lipid metabolism-related molecular clusters and develop a diagnostic model for atherosclerosis. Methods First, we used the GSE100927 and GSE43292 datasets to screen differentially expressed lipid metabolism-related genes (LMRGs). Subsequent enrichment analysis of these key genes was performed using the Metascape database. Using 101 atherosclerosis samples, we investigated the LMRG-based molecular clusters and the corresponding immune cell infiltration. After that, a diagnostic model for atherosclerosis was constructed using the least absolute shrinkage and selection operator (LASSO) and multivariate logistic regression. Finally, a series of bioinformatics techniques, including CIBERSORT, gene set variation analysis, and single-cell data analysis, were used to analyze the potential mechanisms of the model genes in atherosclerosis. Results A total of 29 LMRGs were found to be differentially expressed between atherosclerosis and normal samples. Functional and DisGeNET enrichment analyses indicated that 29 LMRGs are primarily engaged in cholesterol and lipid metabolism, the PPAR signaling pathway, and regulation of the inflammatory response and are also closely associated with atherosclerotic lesions. Two LMRG-related molecular clusters with significant biological functional differences are defined in atherosclerosis. A three-gene diagnostic model containing ADCY7, SCD, and CD36 was subsequently constructed. Receiver operating characteristic curves, decision curves, and an external validation dataset showed that our model exhibits good predictive performance. In addition, three model genes were found to be closely associated with immune cell infiltration, especially macrophage infiltration. Conclusions Our study comprehensively highlighted the intricate association between lipid metabolism and atherosclerosis and created a three-gene model for future clinical diagnosis.

Abstract Background Whole-cell tumor vaccines have shown much promise; however, only limited success has been achieved for the goal of eliciting robust tumor-specific T-cell responses. Methods Hepatocellular carcinoma (HCC) cells, H22 and Hepa1–6, were modified by blocking the STAT3 signaling pathway with a STAT3 decoy oligodeoxynucleotide, and the immunogenicity and possibility of using these cell lysates as a vaccine were evaluated. Results STAT3-blocked whole HCC cell lysates inhibited tumor growth and tumorigenesis, and prolonged the survival of tumor-bearing mice. In addition, STAT3-blocked whole HCC cell lysates stimulated the activation of T cells and natural killer (NK) cells, and enhanced the infiltration of cytotoxic CD8+ T cells in the tumor tissues. In addition, the maturation of dendritic cells (DCs) was enhanced, which promoted the generation of immunological memory against HCC. Furthermore, secondary immune responses could be primed as soon as these immunized mice were challenged with HCC cells, accompanied by T cell and NK cell activation and infiltration. Additionally, immunization with this vaccine decreased the generation of Tregs and the production of TGF-β and IL-10. Importantly, STAT3-blocked whole HCC cell lysates prevented HCC-mediated exhaustion of T cells and NK cells, showing low expression of checkpoint molecules such as PD-1 and TIGIT on T cells and NK cells in the immunized mice. Conclusions The newly generated STAT3-blocked whole-cell HCC vaccine has potential for cancer cell vaccination.

Abstract Objectives To compare the diagnostic performance of the Contrast-Enhanced Ultrasound (CEUS) Liver Imaging Report and Data System (LI-RADS) v2016 and v2017 in identifying the origin of tumor in vein (TIV). Methods From April 2014 to December 2018, focal liver lesions (FLLs) accompanied by TIV formation in patients at high risk for hepatocellular carcinoma (HCC) were enrolled. Histologic evaluation or composite imaging reference standard were served as the reference standard. Each case was categorized according to the CEUS LI-RADS v2016 and v2017, respectively. Diagnostic performance of CEUS LI-RADS v2016 and v2017 in identifying the originated tumor of TIV was validated via sensitivity, specificity, accuracy, positive predictive value (PPV) and negative predictive value. Results A total of 273 FLLs with TIV were analyzed finally, including 266 HCCs and 7 non-HCCs. In v2016, when adopting all TIV as LR-5V, the accuracy and PPV in identifying the originated tumor were both 97.4%. In v2017, when assigning TIV according to contiguous FLLs CEUS LI-RADS category, the accuracy and PPV were 61.9% and 99.4% in subclass of LR-5 as the diagnostic criteria of HCC, and 64.1% and 99.4% in subclass of LR-4/5 as the criteria of HCC diagnosis. There were significant differences in diagnostic accuracy between CEUS LI-RADS v2016 and v2017 in identifying the originated tumor of TIV (p

Abstract The development of optical organic nanoparticles (NPs) is desirable and widely studied. However, most organic dyes are water-insoluble such that the derivatization and modification of these dyes are difficult. Herein, we demonstrated a simple platform for the fabrication of organic NPs designed with emissive properties by loading ten different organic dyes (molar masses of 479.1–1081.7 g/mol) into water-soluble polymer nanosponges composed of poly(styrene-alt-maleic acid) (PSMA). The result showed a substantial improvement over the loading of commercial dyes (3.7–50% loading) while preventing their spontaneous aggregation in aqueous solutions. This packaging strategy includes our newly synthesized organic dyes (> 85% loading) designed for OPVs (242), DSSCs (YI-1, YI-3, YI-8), and OLEDs (ADF-1–3, and DTDPTID) applications. These low-cytotoxicity organic NPs exhibited tunable fluorescence from visible to near-infrared (NIR) emission for cellular imaging and biological tracking in vivo. Moreover, PSMA NPs loaded with designed NIR-dyes were fabricated, and photodynamic therapy with these dye-loaded PSMA NPs for the photolysis of cancer cells was achieved when coupled with 808 nm laser excitation. Indeed, our work demonstrates a facile approach for increasing the biocompatibility and stability of organic dyes by loading them into water-soluble polymer-based carriers, providing a new perspective of organic optoelectronic materials in biomedical theranostic applications.