Optical Coherence Tomography Findings in Patients Presenting With In-Stent Restenosis: A Prospective Observational Study of Patterns of Neointimal Hyperplasia and Associated Risk Factors.

Introduction Morphological features of neointimal tissue play a pivotal role in the pathophysiology of in-stent restenosis (ISR) after percutaneous coronary intervention (PCI). This study was designed to qualitatively and quantitatively assess neointimal characteristics of lesions using optical coherence tomography (OCT) in patients presenting with ISR.  Methods This was a single-center, prospective, observational study performed at a tertiary-care center in India. Patients diagnosed with stable angina and acute coronary syndrome with post-procedural angiographically documented restenosis (>50%) were included. Results A total of 34 patients with ISR were studied. Neointimal hyperplasia was classified as (i) homogenous group (n = 18) and (ii) non-homogenous group (n = 16). Fourteen (77.8%) diabetics belonged to the homogenous group. Predominant plaque characteristics such as neoatherosclerosis, cholesterol crystals, and calcium were documented in 14 (77.8%), 12 (66.7%), and 11 (61.1%) patients in the homogenous group and 10 (62.5%), 10 (62.5%), and 9 (56.2%) patients in the non-homogenous group, respectively. Unexpanded stent struts were identified in 11 (61.1%) and 11 (68.8%) patients in the homogenous and non-homogenous groups, respectively. Mean strut thickness was 93.73 ± 31.03 µm and 83.54 ± 18.0 µm, ISR was 72.50 ± 15.93% and 65.37 ± 21.69%, the neointimal thickness was 588.06 ± 167.82 µm and 666.25 ± 218.05 µm, and neointimal hyperplasia was 54.54 ± 11.23% and 59.26 ± 8.86% in the homogenous and non-homogenous groups, respectively. Conclusion Neoatherosclerosis and stent underexpansion were predominantly observed in our study and only diabetes was found to be significantly associated with homogenous neointimal hyperplasia.

CRISPR/Cas9-Mediated CtBP1 Gene Editing Enhances Chemosensitivity and Inhibits Metastatic Potential in Esophageal Squamous Cell Carcinoma Cells.

Innovative therapeutic strategies for esophageal squamous cell carcinoma (ESCC) are urgently required due to the limited effectiveness of standard chemotherapies. C-Terminal Binding Protein 1 (CtBP1) has been implicated in various cancers, including ESCC. However, the precise expression patterns and functional roles of CtBP1 in ESCC remain inadequately characterized. In this study, we aimed to investigate CtBP1 expression and its role in the resistance of ESCC to paclitaxel, an effective chemotherapeutic agent. Western blotting and immunofluorescence were applied to assess CtBP1 expression in the TE-1 and KYSE-50 cell lines. We observed the marked expression of CtBP1, which was associated with enhanced proliferation, invasion, and metastasis in these cell lines. Further, we successfully generated paclitaxel resistant ESCC cell lines and conducted cell viability assays. We employed the CRISPR/Cas9 genome editing system to disable the CtBP1 gene in ESCC cell lines. Through the analysis of the drug dose-response curve, we assessed the sensitivity of these cell lines in different treatment groups. Remarkably, CtBP1-disabled cell lines displayed not only improved sensitivity but also a remarkable inhibition of proliferation, invasion, and metastasis. This demonstrates that CtBP1 may promote ESCC cell malignancy and confer paclitaxel resistance. In summary, our study opens a promising avenue for targeted therapies, revealing the potential of CtBP1 inhibition to enhance the effectiveness of paclitaxel treatment for the personalized management of ESCC.

Tissue proteome analysis for profiling proteins associated with lymph node metastasis in gallbladder cancer.

Lymph node (LN) metastasis is the earliest sign of metastatic spread and an established predictor of poor outcome in gallbladder cancer (GBC). Patients with LN positive GBC have a significantly worse survival (median survival- 7 months) than patients with LN negative disease (median survival- ~ 23 months) in spite of standard treatment which includes extended surgery followed by chemotherapy, radiotherapy and targeted therapy. This study aims at understanding the underlying molecular processes associated with LN metastasis in GBC. Here, we used iTRAQ-based quantitative proteomic analysis using tissue cohort comprising of primary tumor of LN negative GBC (n = 3), LN positive GBC (n = 4) and non-tumor controls (Gallstone disease, n = 4), to identify proteins associated with LN metastasis. A total of 58 differentially expressed proteins (DEPs) were found to be specifically associated with LN positive GBC based on the criteria of p value ≤ 0.05, fold change ≥ 2 and unique peptides ≥ 2. These include the cytoskeleton and associated proteins such as keratin, type II cytoskeletal 7 (KRT7), keratin type I cytoskeletal 19 (KRT19), vimentin (VIM), sorcin (SRI) and nuclear proteins such as nucleophosmin Isoform 1 (NPM1), heterogeneous nuclear ribonucleoproteins A2/B1 isoform X1 (HNRNPA2B1). Some of them are reported to be involved in promoting cell invasion and metastasis. Bioinformatic analysis of the deregulated proteins in LN positive GBC using STRING database identified 'neutrophil degranulation' and 'HIF1 activation' to be among the top deregulated pathways. Western blot and IHC analysis showed a significant overexpression of KRT7 and SRI in LN positive GBC in comparison to LN negative GBC. KRT7, SRI and other proteins may be further explored for their diagnostics and therapeutic applications in LN positive GBC.

In silico analysis to identify novel ceRNA regulatory axes associated with gallbladder cancer.

Competitive endogenous RNA (ceRNA) networks are reported to play a crucial role in regulating cancer-associated genes. Identification of novel ceRNA networks in gallbladder cancer (GBC) may improve the understanding of its pathogenesis and might yield useful leads on potential therapeutic targets for GBC. For this, a literature survey was done to identify differentially expressed lncRNAs (DELs), miRNAs (DEMs), mRNAs (DEGs) and proteins (DEPs) in GBC. Ingenuity pathway analysis (IPA) using DEMs, DEGs and DEPs in GBC identified 242 experimentally observed miRNA-mRNA interactions with 183 miRNA targets, of these 9 (CDX2, MTDH, TAGLN, TOP2A, TSPAN8, EZH2, TAGLN2, LMNB1, and PTMA) were reported at both mRNA and protein levels. Pathway analysis of 183 targets revealed p53 signaling among the top pathway. Protein-protein interaction (PPI) analysis of 183 targets using the STRING database and cytoHubba plug-in of Cytoscape software revealed 5 hub molecules, of which 3 of them (TP53, CCND1 and CTNNB1) were associated with the p53 signaling pathway. Further, using Diana tools and Cytoscape software, novel lncRNA-miRNA-mRNA networks regulating the expression of TP53, CCND1, CTNNB1, CDX2, MTDH, TOP2A, TSPAN8, EZH2, TAGLN2, LMNB1, and PTMA were constructed. These regulatory networks may be experimentally validated in GBC and explored for therapeutic applications.

Early warning and diagnosis of liver cancer based on dynamic network biomarker and deep learning.

Background: Early detection of complex diseases like hepatocellular carcinoma remains challenging due to their network-driven pathology. Dynamic network biomarkers (DNB) based on monitoring changes in molecular correlations may enable earlier predictions. However, DNB analysis often overlooks disease heterogeneity. Methods: We integrated DNB analysis with graph convolutional neural networks (GCN) to identify critical transitions during hepatocellular carcinoma development in a mouse model. A DNB-GCN model was constructed using transcriptomic data and gene expression levels as node features. Results: DNB analysis identified a critical transition point at 7 weeks of age despite histological examinations being unable to detect cancerous changes at that time point. The DNB-GCN model achieved 100% accuracy in classifying healthy and cancerous mice, and was able to accurately predict the health status of newly introduced mice. Conclusion: The integration of DNB analysis and GCN demonstrates potential for the early detection of complex diseases by capturing network structures and molecular features that conventional biomarker discovery methods overlook. The approach warrants further development and validation.

Bistable insulin response: The win-win solution for glycemic control.

To satisfy both the safety and rapidity of glycemic control, muscles' insulin response must be bistable, as theoretically predicted. Here, we test the bistability hypothesis by combining cellular experiments (to measure the threshold values in vitro) with mathematical modeling (to test the relevance of bistability in vivo). We examine bistability in C2C12 myotubes by both single-cell analysis (FÓ§rster resonance energy transfer) and cultured cells analysis (immunoblot). These technologies demonstrate bistable insulin response, with typical switch-on and switch-off thresholds of approximately 300 and 100 pM, respectively. Our mathematical model demonstrates the indispensability of bistability in interpreting experimental data, reveals fine details of plasma glucose-insulin dynamics, and explains unclear phenomena. These results suggest that the body's ability to simultaneously avoid both hypoglycemia and hyperglycemia is mediated by bistability. The switch-on threshold is a promising biomarker for metabolic complications due to its deep quantitative connection with body composition, which is easy to measure.

Proteomic profiling of cell line-derived extracellular vesicles to identify candidate circulatory markers for detection of gallbladder cancer.

Gallbladder cancer (GBC) is the sixth most common gastrointestinal tract cancer with a very low overall survival and poor prognosis. Profiling of cancer-derived extracellular vesicles (EVs) is an emerging strategy for identification of candidate biomarkers for the detection and prognosis of the disease. The aim of the study was to analyse the protein content from GBC cell line- derived EVs with emphasis on proteins which could be used as candidate biomarkers for the detection of GBC. NOZ and OCUG-1 cell lines were cultured and EVs were isolated from conditioned media. LC-MS/MS analysis of total EV proteins led to the identification of a total of 268 proteins in both the cell lines. Of these, 110 proteins were identified with ≥2 unique peptides with ≥2 PSMs in at least two experimental and technical replicate runs. STRING (Search Tool for the Retrieval of Interacting Genes/Proteins) database was used to perform bioinformatics analysis of 110 proteins which showed 'cell adhesion molecule binding', 'integrin binding', 'cadherin binding' among the top molecular functions and 'focal adhesion' to be among the top pathways associated with the EV proteins. A total of 42 proteins including haptoglobin (HP), pyruvate kinase (PKM), annexin A2 (ANXA2), thrombospondin 1 (THBS1), were reported to be differentially abundant in GBC tissue. Of these, 16 proteins were reported to be differentially abundant in plasma and plasma-derived EVs. We infer these proteins to be highly important to be considered as potential circulatory biomarkers for the detection of GBC. To check the validity of this hypothesis, one of the proteins, haptoglobin (HP) as a representative case, was analysed in plasma by quantitative Enzyme- linked immunosorbent assay (ELISA) and we observed its increased levels in GBC in comparison to controls (p value= 0.0063). Receiver operating characteristic (ROC) curve analysis for GBC vs controls showed an Area under the ROC Curve (AUC) of 0.8264 for HP with 22% sensitivity against 100% specificity. We propose that HP along with other candidate proteins may be further explored for their clinical application.

A prospective observational study to evaluate Glutathione S-transferase gene polymorphism and its association with Antitubercular drugs induced liver injury in tertiary hospital.

BACKGROUND: Anti-TB drugs are most common cause of idiosyncratic hepatotoxicity worldwide. Reactive metabolite formed during drug metabolism has been involved in a clinical toxicity are described as 'idiosyncratic' drug induce liver injury (DILI). We have observed the distribution of glutathione S -transferase (GST) gene polymorphism & its association with drug-induced liver injury in patients taking anti-tubercular treatment. METHODS: A prospective observational study including 96 patients receiving anti-tubercular treatment. Blood sample was collected for LFT and gene extraction after ruling out other cause of liver injury. DNA extraction for GST gene was done follow by polymerase chain reaction to identify homozygous null mutation at GSTM1 and GSTT1 loci. Association of GSTM1 and GSTT1 gene with DILI was seen. RESULTS: Out of 96 tubercular patients under treatment, drug induced liver injury was found in 21 (21.9%) patients and 75 does not develop DILI, GST M1 gene null mutation was observed in 14 (66.7%), GST T1 gene null mutation was observed in 9 (42.9%), Both GST gene null mutation was observed in 8 (38.1%) in DILI group. CONCLUSION: The GSTM1 gene null mutation and both GSTM1 and T1 gene null mutation were a risk factor for the development of DILI. But there is no significant association between GSTT1 gene null mutation and DILI in TB patients.

Quantitative proteomic analysis of GnRH agonist treated GBM cell line LN229 revealed regulatory proteins inhibiting cancer cell proliferation.

BACKGROUND: Gonadotropin-releasing hormone (GnRH) receptor, a rhodopsin-like G-protein coupled receptor (GPCR) family member involved in GnRH signaling, is reported to be expressed in several tumors including glioblastoma multiforme (GBM), one of the most malignant and aggressive forms of primary brain tumors. However, the molecular targets associated with GnRH receptor are not well studied in GBM or in other cancers. The present study aims at investigating the effect of GnRH agonist (Gosarelin acetate) on cell proliferation and associated signaling pathways in GBM cell line, LN229. METHODS: LN229 cells were treated with different concentrations of GnRH agonist (10-10 M to 10-5 M) and the effect on cell proliferation was analyzed by cell count method. Further, total protein was extracted from control and GnRH agonist treated cells (with maximum reduction in cell proliferation) followed by trypsin digestion, labeling with iTRAQ reagents and LC-MS/MS analysis to identify differentially expressed proteins. Bioinformatic analysis was performed for annotation of proteins for the associated molecular function, altered pathways and network analysis using STRING database. RESULTS: The treatment with different concentrations of GnRH agonist showed a reduction in cell proliferation with a maximum reduction of 48.2% observed at 10-6 M. Quantitative proteomic analysis after GnRH agonist treatment (10-6 M) led to the identification of a total of 29 differentially expressed proteins with 1.3-fold change (23 upregulated, such as, kininogen-1 (KNG1), alpha-2-HS-glycoprotein (AHSG), alpha-fetoprotein (AFP), and 6 downregulated, such as integrator complex subunit 11 (CPSF3L), protein FRG1 (FRG1). Some of them are known [KNG1, AHSG, AFP] while others such as inter-alpha-trypsin inhibitor heavy chain H2 (ITIH2), ITIH4, and LIM domain-containing protein 1 (LIMD1) are novel to GnRH signaling pathway. Protein-protein interaction analysis showed a direct interaction of KNG1, a hub molecule, with GnRH, GnRH receptor, EGFR and other interactors including ITIH2, ITIH4 and AHSG. Overexpression of KNG1 after GnRH agonist treatment was validated using Western blot analysis, while a significant inhibition of EGFR was observed after GnRH agonist treatment. CONCLUSIONS: The study suggests a possible link of GnRH signaling with EGFR signaling pathways likely via KNG1. KNG1 inhibitors may be investigated independently or in combination with GnRH agonist for therapeutic applications.

Evaluating the groundwater potential of Wadi Al-Jizi, Sultanate of Oman, by integrating remote sensing and GIS techniques.

Groundwater resources are highly stressed due to their overuse, especially in the arid region. This study is aimed at discovering potential groundwater resource zones using currently available data and state-of-the-art methods. This will lead to effective management of scarcely available and rapidly depleting groundwater resources in the Wadi Al-Jizi catchment, located in the Al-Batinah region. Data on terrain characteristics, geology, and geomorphology was integrated using remote sensing techniques and geographical information system (GIS). The result from this exercise was used for the identification of areas with a high potential for groundwater availability. These areas were classified into five types, namely, excellent, good, medium, low, and very low representing 11%, 59.5%, 26%, and 3.5% of the total area, respectively. The present study shows that the integration of all the weighted parameters shows promising results in the zonation of groundwater. This study shall be useful to the decision-makers in highlighting potential drilling as well as recharge sites in the area.

Quantitative tissue proteome profile reveals neutrophil degranulation and remodeling of extracellular matrix proteins in early stage gallbladder cancer.

Gallbladder cancer (GBC) is an aggressive malignancy of the gastrointestinal tract with a poor prognosis. It is important to understand the molecular processes associated with the pathogenesis of early stage GBC and identify proteins useful for diagnostic and therapeutic strategies. Here, we have carried out an iTRAQ-based quantitative proteomic analysis of tumor tissues from early stage GBC cases (stage I, n=7 and stage II, n=5) and non-tumor controls (n=6) from gallstone disease (GSD). We identified 357 differentially expressed proteins (DEPs) based on ≥ 2 unique peptides and ≥ 2 fold change with p value < 0.05. Pathway analysis using the STRING database showed, 'neutrophil degranulation' to be the major upregulated pathway that includes proteins such as MPO, PRTN3, S100A8, MMP9, DEFA1, AZU, and 'ECM organization' to be the major downregulated pathway that includes proteins such as COL14A1, COL1A2, COL6A1, COL6A2, COL6A3, BGN, DCN. Western blot and/or IHC analysis confirmed the elevated expression of MPO, PRTN3 and S100A8 in early stage of the disease. Based on the above results, we hypothesize that there is an increased neutrophil infiltration in tumor tissue and neutrophil degranulation leading to degradation of extracellular matrix (ECM) proteins promoting cancer cell invasion in the early stage GBC. Some of the proteins (MPO, MMP9, DEFA1) associated with 'neutrophil degranulation' showed the presence of 'signal sequence' suggesting their potential as circulatory markers for early detection of GBC. Overall, the study presents a protein dataset associated with early stage GBC.

Plasma-derived candidate biomarkers for detection of gallbladder carcinoma.

Gallbladder carcinoma (GBC) is a major cancer of the gastrointestinal tract with poor prognosis. Reliable and affordable biomarker-based assays with high sensitivity and specificity for the detection of this cancer are a clinical need. With the aim of studying the potential of the plasma-derived extracellular vesicles (EVs), we carried out quantitative proteomic analysis of the EV proteins, using three types of controls and various stages of the disease, which led to the identification of 86 proteins with altered abundance. These include 29 proteins unique to early stage, 44 unique to the advanced stage and 13 proteins being common to both the stages. Many proteins are functionally relevant to the tumor condition or have been also known to be differentially expressed in GBC tissues. Several of them are also present in the plasma in free state. Clinical verification of three tumor-associated proteins with elevated levels in comparison to all the three control types-5'-nucleotidase isoform 2 (NT5E), aminopeptidase N (ANPEP) and neprilysin (MME) was carried out using individual plasma samples from early or advanced stage GBC. Sensitivity and specificity assessment based on receiver operating characteristic (ROC) analysis indicated a significant association of NT5E and ANPEP with advanced stage GBC and MME with early stage GBC. These and other proteins identified in the study may be potentially useful for developing new diagnostics for GBC.

Comprehensive strategies of machine-learning-based quantitative structure-activity relationship models.

Early quantitative structure-activity relationship (QSAR) technologies have unsatisfactory versatility and accuracy in fields such as drug discovery because they are based on traditional machine learning and interpretive expert features. The development of Big Data and deep learning technologies significantly improve the processing of unstructured data and unleash the great potential of QSAR. Here we discuss the integration of wet experiments (which provide experimental data and reliable verification), molecular dynamics simulation (which provides mechanistic interpretation at the atomic/molecular levels), and machine learning (including deep learning) techniques to improve QSAR models. We first review the history of traditional QSAR and point out its problems. We then propose a better QSAR model characterized by a new iterative framework to integrate machine learning with disparate data input. Finally, we discuss the application of QSAR and machine learning to many practical research fields, including drug development and clinical trials.

Determination of split renal function in voluntary renal donors by multidetector computed tomography and nuclear renography: How well do they correlate?

BACKGROUND: The use of computed tomography (CT) for estimation of split renal function (SRF) has been reported previously. However, most of these studies have small samples, and many do not account for the renal attenuation at CT. OBJECTIVE: The aim of this study was to compare multidetector computed tomography (MDCT) volumetry-attenuation-based SRF with that obtained via Tc99m-diethylenetriaminepentaacetic acid (DTPA) renal scintigraphy in voluntary renal donors. METHODS: Between January 2017 and January 2020, 526 voluntary renal donors were enrolled prospectively. All donors underwent contrast CT and DTPA scan before surgery. The semiautomatic region of interest (ROI) tool was applied slice by slice on axial CT images acquired in the arterial phase. The renal contour was drawn semiautomatically with mouse clicks around the renal parenchyma, and the renal volume was ascertained. Using renal volume and attenuation, SRF was determined and compared with results obtained at DTPA imaging. RESULTS: The mean age was 44.91 ± 10.97 years (mean ± s.d.). There was no significant difference in SRF based on DTPA and MDCT volumetry for the left kidney (49.18% ± 3.40% vs. 49.15% ± 3.38%, p = 0.540) and for the right kidney (50.82% ± 3.40% vs. 50.86% ± 3.39%, p = 0.358). A very good correlation was observed between the two methods for the left kidney (r = 0.953, p = 0.000) and the right kidney (r = 0.955, p = 0.000). On simple linear regression analysis, 90.8% of DTPA SRF values for the left kidney and 91.3% of DTPA SRF values for the right kidney could be predicted correctly using the corresponding MDCT SRF values. CONCLUSION: MDCT volumetry-attenuation-derived estimation of SRF for living renal donors could be an alternative to renal scintigraphy-based SRF estimation.

Computational and Experimental Studies Reveal That Thymoquinone Blocks the Entry of Coronaviruses Into In Vitro Cells.

INTRODUCTION: Since December 2019, severe acute respiratory syndrome-related coronavirus-2 (SARS-CoV-2) has caused the coronavirus disease 2019 (COVID-19) pandemic in China and worldwide. New drugs for the treatment of COVID-19 are in urgent need. Considering the long development time for new drugs, the identification of promising inhibitors from FDA-approved drugs is an imperative and valuable strategy. Recent studies have shown that the S1 and S2 subunits of the spike protein of SARS-CoV-2 utilize human angiotensin-converting enzyme 2 (hACE2) as the receptor to infect human cells. METHODS: We combined molecular docking and surface plasmon resonance (SPR) to identify potential inhibitors for ACE2 from available commercial medicines. We also designed coronavirus pseudoparticles that contain the spike protein assembled onto green fluorescent protein or luciferase reporter gene-carrying vesicular stomatitis virus core particles. RESULTS: We found that thymoquinone, a phytochemical compound obtained from the plant Nigella sativa, is a potential drug candidate. SPR analysis confirmed the binding of thymoquinone to ACE2. We found that thymoquinone can inhibit SARS-CoV-2, SARS-CoV, and NL63 pseudoparticles infecting HEK293-ACE2 cells, with half-maximal inhibitory concentrations of 4.999, 7.598, and 6.019 µM, respectively. The SARS-CoV-2 pseudoparticle inhibition had half-maximal cytotoxic concentration of 35.100 µM and selection index = 7.020. CONCLUSION: Thymoquinone is a potential broad-spectrum inhibitor for the treatment of coronavirus infections.

Role of Computed Tomography in Estimation of Residual Renal Function at One Year after Donor Nephrectomy: Comparison with Nuclear Scintigraphy.

Computed tomography (CT) is routinely used for preoperative anatomical evaluation of renal donors, but recent studies show that it may be useful as a predictor of renal function too. This study aimed to compare CT volumetry-attenuation-based glomerular filtration rate (GFR) with Tc99m-diethylene-triamine-pentaacetic acid (DTPA) renal scintigraphy GFR and predict post-donor nephrectomy renal function. This is a prospective study involving 39 renal donors at a tertiary care hospital in northern India from 2014 to 2017. Renal volume and attenuation were determined by CT using a semiautomatic tool, and split renal function (SRF) was calculated for each side. CT-GFR was obtained using CT-SRF and global GFR from modification of diet in renal disease equation. At 12 months after nephrectomy, GFR was estimated from serum creatinine (eGFR). Predonation CT-GFR of preserved kidney was used to predict post-nephrectomy renal function which was compared with post-donation renal function calculated using serum creatinine as well as with DTPA-GFR. Pearson correlation coefficient and scatter diagram were used to assess and test the linear relationship between two continuous variables. The Statistical Package for the Social Sciences, version 23, was used for data analysis. There was a highly significant change in mean DTPA-GFR and mean CT-GFR compared to eGFR values (50.79-64.25, 26.50%, and 50.77-64.25, 26.55%, respectively). Similarly, there was a very good correlation between DTPA-GFR and eGFR (r = 0.968, P <0.001) as well as CT-GFR and eGFR (r = 0.968, P <0.001). CT volumetry-attenuation-based GFR accurately predicts post-donor nephrectomy renal function at 1 year similar to Tc99m-DTPA renal scintigraphy.

Can face masks offer protection from airborne sneeze and cough droplets in close-up, face-to-face human interactions?-A quantitative study.

Day-to-day observations reveal numerous medical and social situations where maintaining physical distancing is either not feasible or not practiced during the time of a viral pandemic, such as, the coronavirus disease 2019 (COVID-19). During these close-up, face-to-face interactions, a common belief is that a susceptible person wearing a face mask is safe, at least to a large extent, from foreign airborne sneeze and cough droplets. This study, for the first time, quantitatively verifies this notion. Droplet flow visualization experiments of a simulated face-to-face interaction with a mask in place were conducted using the particle image velocimetry setup. Five masks were tested in a snug-fit configuration (i.e., with no leakage around the edges): N-95, surgical, cloth PM 2.5, cloth, and wetted cloth PM 2.5. Except for the N-95 mask, the findings showed leakage of airborne droplets through all the face masks in both the configurations of (1) a susceptible person wearing a mask for protection and (2) a virus carrier wearing a mask to prevent the spreading of the virus. When the leakage percentages of these airborne droplets were expressed in terms of the number of virus particles, it was found that masks would not offer complete protection to a susceptible person from a viral infection in close (e.g., <6 ft) face-to-face or frontal human interactions. Therefore, consideration must be given to minimize or avoid such interactions, if possible. This study lends quantitative support to the social distancing and mask-wearing guidelines proposed by the medical research community.

Immunoproteomics approach revealed elevated autoantibody levels against ANXA1 in early stage gallbladder carcinoma.

BACKGROUND: Early diagnosis is important for the timely treatment of gallbladder carcinoma (GBC) patients and may lead to increased survival outcomes. Here, we have applied serological proteome analysis (SERPA), an immunoproteomics approach, for the detection of 'tumor-associated antigens (TAAs) that elicit humoral response' in early stage GBC patients. METHODS: Total protein from pooled tumor tissue of GBC patients (n = 7) was resolved by two-dimensional gel electrophoresis (2-DE) followed by immunoblotting using pooled blood plasma from healthy volunteers (n = 11) or gallstone disease (GSD) cases (n = 11) or early stage GBC (Stage I and II) (n = 5) or GBC stage IIIA (n = 9). 2-D gel and immunoblot images were acquired and analyzed using PDQuest software to identify immunoreactive spots in GBC cases in comparison to controls. Proteins from immunoreactive spots were identified by liquid chromatography- tandem mass spectrometric analysis (LC-MS/MS). Autoantibody levels for two of the functionally relevant proteins were investigated in individual plasma samples (52 cases and 89 controls) by dot blot assay using recombinant proteins. RESULTS: Image analysis using PDQuest software identified 25 protein spots with significantly high or specific immunoreactivity in GBC cases. Mass spectrometric analysis of 8 corresponding protein spots showing intense immunoreactivity (based on densitometric analysis) in early stage GBC or GBC stage IIIA cases led to the identification of 27 proteins. Some of the identified proteins include ANXA1, HSPD1, CA1, CA2, ALDOA and CTSD. Among the two proteins, namely ANXA1 and HSPD1 verified using a cohort of samples, significantly elevated autoantibody levels against ANXA1 were observed in early stage GBC cases in comparison to healthy volunteers or GSD cases (unpaired t-test, p < 0.05). Receiver operating characteristic (ROC) curve analysis for ANXA1 showed an Area under the Curve (AUC) of 0.69, with 41.7% sensitivity against a specificity of 89.9% for early stage GBC. IHC analysis for ANXA1 protein showed 'high' expression levels in 72% of GBC cases whereas all the controls showed 'low' expression levels. CONCLUSIONS: The study suggests that the ANXA1 autoantibody levels against ANXA1 may be potentially employed for early stage detection of GBC patients. Other proteins could also be explored and verified in a large cohort of clinical samples.

Physical mapping of introgressed chromosome fragment carrying the fertility restoring (Rfo) gene for Ogura CMS in Brassica juncea L. Czern & Coss.

KEY MESSAGE: Rfo is located on a radish chromosome fragment (~ 108 Kb), which is seated in the middle of a pretty large C genome translocation at the distal region of chromosome A09 of B. juncea. Ogura cytoplasmic male sterility (CMS) is used to produce hybrids in Indian mustard (Brassica juncea L.). Fertility restorers for this CMS were developed by cross-hybridizing B. juncea (AABB; 2n = 36) with B. napus (AACC; 2n = 38) carrying radish Rfo gene. This hybrid production system is normally stable, but many commercial mustard hybrids show male sterile contaminants. We aimed to identify linkage drag associated with Rfo by comparing hybridity levels of 295 handmade CMS x Rfo crosses. Although Rfo was stably inherited, hybridity was < 85 percent in several combinations. Genome re-sequencing of five fertility restorers, mapping sequencing reads to B. juncea reference and synteny analysis with Raphanus sativus D81Rfo genomic region (AJ550021.2) helped to detect ~ 108 Kb of radish chromosome (R) fragment substitution in all fertility restorers. This radish segment substitution was itself located amidst a large C genome translocation on the terminal region of chromosome A09 of B. juncea. The size of alien segment substitution varied from 11.3 (NTCN-R9) to 22.0 Mb (NAJR-102B-R). We also developed an in silico SSR map for chromosome A09 and identified many homoeologous A to the C genome exchanges in the introgressed region. A to the R genome exchanges were rare. Annotation of the substituted fragment showed the gain of many novel genes from R and C genomes and the loss of B. juncea genes from the corresponding region. We have developed a KASPar marker for marker-aided transfer of Rfo and testing hybridity levels in seed production lots.

A novel pathogenic missense variant in CNNM4 underlying Jalili syndrome: Insights from molecular dynamics simulations.

BACKGROUND: Jalili syndrome (JS) is a rare cone-rod dystrophy (CRD) associated with amelogenesis imperfecta (AI). The first clinical presentation of JS patients was published in 1988 by Jalili and Smith. Pathogenic mutations in the Cyclin and CBS Domain Divalent Metal Cation Transport Mediator 4 (CNNM4) magnesium transporter protein have been reported as the leading cause of this anomaly. METHODS: In the present study, a clinical and genetic investigation was performed in a consanguineous family of Pakistani origin, showing characteristic features of JS. Sanger sequencing was successfully used to identify the causative variant in CNNM4. Molecular dynamics (MD) simulations were performed to study the effect of amino acid change over CNNM4 protein. RESULTS: Sequence analysis of CNNM4 revealed a novel missense variant (c.1220G>T, p.Arg407Leu) in exon-1 encoding cystathionine-ß-synthase (CBS) domain. To comprehend the mutational consequences in the structure, the mutant p.Arg407Leu was modeled together with a previously reported variant (c.1484C>T, p.Thr495Ile) in the same domain. Additionally, docking analysis deciphered the binding mode of the adenosine triphosphate (ATP) cofactor. Furthermore, 60ns MD simulations were carried out on wild type (p.Arg407/p.Thr495) and mutants (p.Arg407Leu/p.Thr495Ile) to understand the structural and energetic changes in protein structure and its dynamic behavior. An evident conformational shift of ATP in the binding site was observed in simulated mutants disrupting the native ATP-binding mode. CONCLUSION: The novel identified variant in CNNM4 is the first report from the Pakistani population. Overall, the study is valuable and may give a novel insight into metal transport in visual function and biomineralization.