Single-cell transcriptome reveals highly complement activated microglia cells in association with pediatric tuberculous meningitis.

Background: Tuberculous meningitis (TBM) is a devastating form of tuberculosis (TB) causing high mortality and disability. TBM arises due to immune dysregulation, but the underlying immune mechanisms are unclear. Methods: We performed single-cell RNA sequencing on peripheral blood mononuclear cells (PBMCs) and cerebrospinal fluid (CSF) cells isolated from children (n=6) with TBM using 10 xGenomics platform. We used unsupervised clustering of cells and cluster visualization based on the gene expression profiles, and validated the protein and cytokines by ELISA analysis. Results: We revealed for the first time 33 monocyte populations across the CSF cells and PBMCs of children with TBM. Within these populations, we saw that CD4_C04 cells with Th17 and Th1 phenotypes and Macro_C01 cells with a microglia phenotype, were enriched in the CSF. Lineage tracking analysis of monocyte populations revealed myeloid cell populations, as well as subsets of CD4 and CD8 T-cell populations with distinct effector functions. Importantly, we discovered that complement-activated microglial Macro_C01 cells are associated with a neuroinflammatory response that leads to persistent meningitis. Consistently, we saw an increase in complement protein (C1Q), inflammatory markers (CRP) and inflammatory factor (TNF-α and IL-6) in CSF cells but not blood. Finally, we inferred that Macro_C01 cells recruit CD4_C04 cells through CXCL16/CXCR6. Discussion: We proposed that the microglial Macro_C01 subset activates complement and interacts with the CD4_C04 cell subset to amplify inflammatory signals, which could potentially contribute to augment inflammatory signals, resulting in hyperinflammation and an immune response elicited by Mtb-infected tissues.

Discovering novel biomarkers for diagnosis and treatment monitoring of active pulmonary tuberculosis by ion metabolism analysis.

Tuberculosis (TB) is a highly lethal infectious disease that poses a global threat. Timely and accurate biomarker for TB diagnosis and treatment monitoring remains a pressing need. Ions, the crucial trace element for humans, may be potential targets for TB diagnosis and the forecasting of TB development. To explore the potential of ions as biomarkers, we measured and compared the levels of various ions in whole blood and plasma samples from healthy control (HC), pulmonary TB patients (TB), cured pulmonary TB patients (RxTB), and other non-TB pneumonia patients (PN) by using ultra-high performance liquid chromatography-tandem mass spectrometry. Our study demonstrated that Cu (AUC = 0.670), Pb (AUC = 0.660), and Zn (AUC = 0.701) in whole blood exhibited promising diagnostic performance for TB. Then we used a neural network (NNET) for TB prediction, the AUC values used to differentiate definite TB from HC or PN in plasma were 0.867 and 0.864, respectively. The AUC values used to differentiate definite TB from HC or PN in whole blood were 0.818 and 0.660, respectively. Our correlation analysis showed that Zn (r= 0.356, p=0.001) and Cu (r= 0.361, p=0.0004) in plasma are most closely related to disease severity. Additionally, six ions (Cu, Sb, V, Mn, Fe, Sr) in plasma and whole blood were altered following anti-TB therapy. These results showed that ions could be diagnostic biomarkers for TB. Furthermore, the level of particular ions can forecast the degree of lung damage and the success of the TB treatment. In conclusion, this study highlights the possibility of using ions from blood samples to enable rapid tuberculosis diagnosis.

Oxygen uptake during the last bouts of exercise incorporated into high-intensity intermittent cross-exercise exceeds the V˙ O2max of the same exercise mode.

Oxygen uptake (V˙ O2) was measured during a non-exhaustive high-intensity intermittent cross-exercise (HIICE) protocol consisting of four alternating bouts of 20 â€‹s running (R) and three bouts of bicycle exercise (BE) at ∼160% and ∼170% maximal oxygen uptake (V˙ O2max), respectively, with 10 â€‹s between-bout rests (sequence R-BE-R-BE-R-BE-R). The V˙ O2 during the last BE ([52.2 â€‹± â€‹5.0] mL·kg-1·min-1) was significantly higher than the V˙ O2max of the BE ([48.0 â€‹± â€‹5.4] mL·kg-1·min-1, n â€‹= â€‹30) and similar to that of running. For clarifying the underlying mechanisms, a corresponding HIICE-protocol with BE and arm cranking ergometer exercise (AC) was used (sequence AC-AC-BE-AC-BE-AC-AC-BE). In some experiments, thigh blood flow was occluded by a cuff around the upper thigh. Without occlusion, the V˙ O2 during the AC ([39.2 â€‹± â€‹7.1] mL·kg-1·min-1 [6th bout]) was significantly higher than the V˙ O2max of AC ([30.2 â€‹± â€‹4.4] mL·kg-1·min-1, n â€‹= â€‹7). With occlusion, the corresponding V˙ O2 ([29.8 â€‹± â€‹3.9] mL·kg-1·min-1) was reduced to that of the V˙ O2max of AC and significantly less than the V˙ O2 without occlusion. These findings suggest that during the last bouts of HIICE may exceed the of the specific exercise, probably because it is a summation of the V˙ O2 for the ongoing exercise plus excess post-oxygen consumption (EPOC) produced by the previous exercise with a higher V˙ O2max.

Terminal modifications independent cell-free RNA sequencing enables sensitive early cancer detection and classification.

Cell-free RNAs (cfRNAs) offer an opportunity to detect diseases from a transcriptomic perspective, however, existing techniques have fallen short in generating a comprehensive cell-free transcriptome profile. We develop a sensitive library preparation method that is robust down to 100 µl input plasma to analyze cfRNAs independent of their 5'-end modifications. We show that it outperforms adapter ligation-based method in detecting a greater number of cfRNA species. We perform transcriptome-wide characterizations in 165 lung cancer, 30 breast cancer, 37 colorectal cancer, 55 gastric cancer, 15 liver cancer, and 133 cancer-free participants and demonstrate its ability to identify transcriptomic changes occurring in early-stage tumors. We also leverage machine learning analyses on the differentially expressed cfRNA signatures and reveal their robust performance in cancer detection and classification. Our work sets the stage for in-depth study of the cfRNA repertoire and highlights the value of cfRNAs as cancer biomarkers in clinical applications.

The gut and lung microbiota in pulmonary tuberculosis: susceptibility, function, and new insights into treatment.

INTRODUCTION: Tuberculosis (TB) is a chronic infectious disease caused by mycobacterium tuberculosis (Mtb) that poses a major threat to human health. AREAS COVERED: Herein, we aim to review the alteration of the microbiota in gut and respiratory during TB development, the potential function and mechanisms of microbiota in the pathogenesis of Mtb infection, and the impact of antibiotic treatment on the microbiota. In addition, we discuss the potential new paradigm for the use of microbiota-based treatments such as probiotics and prebiotics in the treatment of TB. EXPERT OPINION: Studies have shown that trillions of micro-organisms live in the human gut and respiratory tract, acting as gatekeepers in maintaining immune homeostasis and respiratory physiology and playing a beneficial or hostile role in the development of TB. Anti-TB antibiotics may cause microecological imbalances in the gut and respiratory tract, and microbiome-based therapeutics may be a promising strategy for TB treatment. Appropriate probiotics and prebiotics supplementation, along with antimycobacterial treatment, will improve the therapeutic effect of TB treatment and protect the gut and respiratory microbiota from dysbiosis.

Three E2F target-related genes signature for predicting prognosis, immune features, and drug sensitivity in hepatocellular carcinoma.

Background: Hepatocellular carcinoma (HCC) is extremely malignant and difficult to treat. The adenoviral early region 2 binding factors (E2Fs) target pathway is thought to have a major role in tumor growth. This study aimed to identify a predictive E2F target signature and facilitate individualized treatment for HCC patients. Methods: We constructed an E2F target-related gene profile using univariate COX and LASSO regression models and proved its predictive efficacy in external cohorts. Furthermore, we characterized the role of the E2F target pathway in pathway enrichment, immune cell infiltration, and drug sensitivity of HCC. Results: Lasso Cox regression created an E2F target-related gene signature of GHR, TRIP13, and CDCA8. HCC patients with high risk were correlated with shorter survival time, immune evasion, tumor stem cell characteristics and high sensitivity to Tipifarnib and Camptothecin drugs. Conclusion: Hepatocellular carcinoma prognosis was predicted by an E2F target signature. This finding establishes the theoretical usefulness of the E2F target route in customized identification and treatment for future research.

Piezodynamic Eradication of Both Gram-Positive and Gram-Negative Bacteria by Using a Nanoparticle Embedded Polymeric Membrane.

Nowadays, bacterial infection is regarded as a serious threat to humankind, which needs to be taken care of. The emergence of antibiotic resistance and multidrug resistance (MDR) is rendering this situation more troublesome. However, several alternative treatment regimens have aided such diseases quite well in the recent past, among which dynamic antibacterial therapies combat this situation quite well. Among various dynamic therapies, piezodynamic therapy is a very recent avenue, in which mechanical stimuli have been exploited to treat bacterial infections. Herein, piezo-active bismuth ferrite-loaded poly(vinylidene fluoride-co-hexafluoropropylene) polymer has been utilized to eradicate gram-positive bacteria (E. faecalis) and gram-negative bacteria (E. coli). The sample has been designed in a free-standing membrane form, which, under soft ultrasound (~10 kHz), generates reactive radicals to ablate bacteria. Initially, the structure and morphology of the membrane have been substantiated by using X-ray diffraction and scanning electron microscopy methods; besides, Fourier transform infrared spectrum of the sample depicts a tremendously high value of polarizability and further confirms the piezo-activity of the membrane. More than 99% of E. coli and E. faecalis have been successfully eradicated within 30 min of ultrasound. Moreover, the solid-state structure and hydrophobic nature of the membrane help us to reuse it in a cyclic manner, which is possibly reported herein for the very first time. This novel membrane could be deployed in healthcare systems and pigment industries and could be exploited as a self-cleaning material.

Berbamine promotes macrophage autophagy to clear Mycobacterium tuberculosis by regulating the ROS/Ca2+ axis.

Drug-resistant tuberculosis (TB) poses a major threat to global TB control; consequently, there is an urgent need to develop novel anti-TB drugs or strategies. Host-directed therapy (HDT) is emerging as an effective treatment strategy, especially for drug-resistant TB. This study evaluated the effects of berbamine (BBM), a bisbenzylisoquinoline alkaloid, on mycobacterial growth in macrophages. BBM inhibited intracellular Mycobacterium tuberculosis (Mtb) growth by promoting autophagy and silencing ATG5, partially abolishing the inhibitory effect. In addition, BBM increased intracellular reactive oxygen species (ROS), while the antioxidant N-acetyl-L-cysteine (NAC) abolished BBM-induced autophagy and the ability to inhibit Mtb survival. Furthermore, the increased intracellular Ca2+ concentration induced by BBM was regulated by ROS, and BAPTA-AM, an intracellular Ca2+-chelating agent, could block ROS-mediated autophagy and Mtb clearance. Finally, BBM could inhibit the survival of drug-resistant Mtb. Collectively, these findings provide evidence that BBM, a Food and Drug Administration (FDA)-approved drug, could effectively clear drug-sensitive and -resistant Mtb through regulating ROS/Ca2+ axis-mediated autophagy and has potential as an HDT candidate for TB therapy. IMPORTANCE It is urgent to develop novel treatment strategies against drug-resistant TB, and HDT provides a promising approach to fight drug-resistant TB by repurposing old drugs. Our studies demonstrate, for the first time, that BBM, an FDA-approved drug, not only potently inhibits intracellular drug-sensitive Mtb growth but also restricts drug-resistant Mtb by promoting macrophage autophagy. Mechanistically, BBM activates macrophage autophagy by regulating the ROS/Ca2+ axis. In conclusion, BBM could be considered as an HDT candidate and may contribute to improving the outcomes or shortening the treatment course of drug-resistant TB.

Application and Development of Minimally Invasive Techniques in the Treatment of Spinal Metastases.

With the improvement of medical technology, the quality of life and prognosis of patients with malignant tumors have been greatly improved, and surgical treatment strategies for patients with spinal metastatic tumors have received extensive attention. Traditional open surgery for spinal metastases has problems such as large trauma, slow recovery, and influence on subsequent systemic treatment. Minimally invasive spine surgery has similar clinical outcomes to traditional open surgery, but minimally invasive spine surgery is less invasive and has a shorter recovery time. Minimally invasive spine surgery was initially applied to non-neoplastic diseases such as spinal degeneration and trauma, and was gradually applied to the treatment of spinal metastatic tumors and spinal deformities. For patients with spinal metastases, a shorter recovery time is helpful for early postoperative radiotherapy, thereby achieving a more satisfactory tumor control effect. This review discusses the application of minimally invasive spine surgery in the treatment of spinal metastatic tumors from the concept, surgical purpose, indications, and surgical selection, so as to provide reference for clinical practice.

Mycobacterium tuberculosis Utilizes Host Histamine Receptor H1 to Modulate Reactive Oxygen Species Production and Phagosome Maturation via the p38MAPK-NOX2 Axis.

Tuberculosis (TB), which is caused by the single pathogenic bacterium, Mycobacterium tuberculosis, is among the top 10 lethal diseases worldwide. This situation has been exacerbated by the increasing number of cases of multidrug-resistant TB (MDR-TB) and extensively drug-resistant TB (XDR-TB). Histamine is an organic nitrogenous compound that mediates a plethora of cell processes via different receptors. The expression of histamine receptor H1 (HRH1), one of the four histamine receptors identified to date was previously reported to be augmented by M. tuberculosis infection, although the underlying mechanism is unclear. In the present study, we applied confocal microscopy, flow cytometry, and Western blotting to show that HRH1 expression was enhanced in macrophages following mycobacterial infection. Furthermore, by combining techniques of gene knockdown, immunoprecipitation, intracellular bacterial burden analysis, fluorescence labeling, and imaging, we found that M. tuberculosis targeted the host HRH1 to suppress NOX2-mediated cROS production and inhibit phagosome maturation and acidification via the GRK2-p38MAPK signaling pathway. Our findings clarified the underlying mechanism of the M. tuberculosis and host HRH1 interaction and may provide useful information for the development of novel antituberculosis treatments. IMPORTANCE Once engulfed in macrophage phagosomes, M. tuberculosis adopts various strategies to take advantage of the host environment for its intracellular survival. Histamine is an organic nitrogen-containing compound that mediates a plethora of cellular processes via different receptors, but the crosstalk mechanism between M. tuberculosis and HRH1 in macrophages is not clear. Our results revealed that M. tuberculosis infection enhanced HRH1 expression, which in turn restrained macrophage bactericidal activity by modulating the GRK2-p38MAPK signaling pathway, inhibiting NOX2-mediated cROS production and phagosome maturation. Clarification of the underlying mechanism by which M. tuberculosis utilizes host HRH1 to favor its intracellular survival may provide useful information for the development of novel antituberculosis treatments.

Comprehensive identification of immuno-related transcriptional signature for active pulmonary tuberculosis by integrated analysis of array and single cell RNA-seq.

BACKGROUND: Tuberculosis (TB) continues to be a major cause of morbidity and mortality worldwide. However, the molecular mechanism underlying immune response to human infection with Mycobacterium tuberculosis (Mtb) remains unclear. Assessing changes in transcript abundance in blood between health and disease on a genome-wide scale affords a comprehensive view of the impact of Mtb infection on the host defense and a reliable way to identify novel TB biomarkers. METHODS: We combined expression profiling by array and single cell RNA-sequencing (scRNA-seq) via 10X Genomics platform to better illustrate the immuno-related transcriptional signature of TB and explore potential diagnostic markers for differentiating TB from latent tuberculosis infection (LTBI) and healthy control (HC). FINDINGS: Pathway analysis based on differential expressed genes (DEGs) revealed that immune transcriptional profiling could effectively differ TB with LTBI and HC. Following WGCNA and PPI network analysis based on DEGs, we screened out three key immuno-related hub genes (ADM, IFIT3 and SERPING1) highly associated with TB. Further validation found only ADM expression significantly increased in TB patients in both adult and children's datasets. By comparing the scRNA-seq datasets from TB, LTBI and HC, we observed a remarkable elevated expression level and proportion of ADM in TB Myeloid cells, further supporting that ADM expression changes could distinguish patients with TB from LTBI and HC. Besides, the hsa-miR-24-3p-NEAT1-ADM-CEBPB regulation pathway might be one of the critical networks regulating the pathogenesis of TB. Although further investigation in a larger cohort is warranted, we provide useful and novel insight to explore the potential candidate genes for TB diagnosis and intervention. INTERPRETATION: We propose that the expression of ADM in peripheral blood could be used as a novel biomarker for differentiating TB with LTBI and HC.

Long Non-coding RNAs in Tuberculosis: From Immunity to Biomarkers.

Tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb) is the leading lethal infectious disease with 1.3 million deaths in 2020. Despite significant advances have been made in detection techniques and therapeutic approaches for tuberculosis, no suitable diagnostic tools are available for early and precise screening. Many studies have reported that Long non-coding RNAs (lncRNAs) play a regulatory role in gene expression in the host immune response against Mtb. Dysregulation of lncRNAs expression patterns associated with immunoregulatory pathways arose in mycobacterial infection. Meanwhile, host-induced lncRNAs regulate antibacterial processes such as apoptosis and autophagy to limit bacterial proliferation. In this review, we try to summarize the latest reports on how dysregulated expressed lncRNAs influence host immune response in tuberculosis infection. We also discuss their potential clinical prospects for tuberculosis diagnosis and development as molecular biomarkers.

Single-cell immune profiling reveals functional diversity of T cells in tuberculous pleural effusion.

Orchestration of an effective T lymphocyte response at infection sites is critical for protection against Mycobacterium tuberculosis (Mtb) infection. However, the local T cell immunity landscape in human tuberculosis is poorly defined. Tuberculous pleural effusion (TPE), caused by Mtb, is characterized by an influx of leukocytes to the pleural space, providing a platform suitable for delineating complex tissue responses to Mtb infection. Using single-cell transcriptomics and T cell receptor sequencing, we analyzed mononuclear cell populations in paired pleural fluid and peripheral blood of TPE patients. While all major cell clusters were present in both tissues, their relative proportions varied significantly by anatomic location. Lineage tracking analysis revealed subsets of CD8 and CD4 T cell populations with distinct effector functions specifically expanded at pleural sites. Granzyme K-expressing CD8 T cells were preferentially enriched and clonally expanded in pleural fluid from TPE, suggesting that they are involved in the pathogenesis of the disease. The findings collectively reveal the landscape of local T cell immunity in tuberculosis.

The emerging role of non-coding RNAs from extracellular vesicles in Alzheimer's disease.

Alzheimer's disease is an age-dependent neurodegenerative disease. Recently, different non-coding RNAs (ncRNAs), including microRNAs, long non-coding RNAs, and circular RNAs, have been found to contribute to Alzheimer's disease's pathogenesis. Extracellular vehicles could be enriched in ncRNAs and in their role in mediating intercellular communication. Signatures of extracellular vesicular ncRNAs have shown them to be a potential biomarker in Alzheimer's disease. This perspective discusses the potential role of extracellular vehicle ncRNAs in Alzheimer's disease, providing a theoretical basis for extracellular vesicular ncRNAs in Alzheimer's disease, from pathogenesis to diagnosis and treatment.

Current approaches in laboratory testing for SARS-CoV-2.

The outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which began in Wuhan, Hubei Province, China, has rapidly spread to produce a global pandemic. It is now clear that person-to-person transmission of SARS-CoV-2 has been occurring and that the virus has been dramatically growing in recent months. Early, rapid and accurate diagnosis is of great significance for curtailing the spread of SARS-CoV-2. There are currently several diagnostic techniques (e.g. viral culture and nucleic acid amplification test) being used to detect the virus. However, the sensitivity and specificity of these methods are quite different, with the sample source and detection limit varying greatly. This study reviewed all types and characteristics of the currently available laboratory diagnostic assays for detecting SARS-CoV-2 infection and summarized the selection strategies of testing and sampling sites at different disease stages to improve the diagnostic accuracy of Coronavirus Disease 2019 (COVID-19).

Evaluations of the serological test in the diagnosis of 2019 novel coronavirus (SARS-CoV-2) infections during the COVID-19 outbreak.

We developed a chemiluminescence immunoassay method based on the recombinant nucleocapsid antigen and assessed its performance for the clinical diagnosis of severe acute respiratory syndrome coronavirus (SARS-CoV)-2 infections by detecting SARS-CoV-2-specific IgM and IgG antibodies in patients. Full-length recombinant nucleocapsid antigen and tosyl magnetic beads were used to develop the chemiluminescence immunoassay approach. Plasmas from 29 healthy cohorts, 51 tuberculosis patients, and 79 confirmed SARS-CoV-2 patients were employed to evaluate the chemiluminescence immunoassay method performance for the clinical diagnosis of SARS-CoV-2 infections. A commercial ELISA kit (Darui Biotech, China) using the same nucleocapsid antigen was used for the in-parallel comparison with our chemiluminescence immunoassay method. The IgM and IgG manner of testing in the chemiluminescence immunoassay method showed a sensitivity and specificity of 60.76% (95% CI 49.1 to 71.6) and 92.25% (95% CI 83.4 to 97.2) and 82.28% (95% CI 72.1 to 90.0) and 97.5% (95% CI 91.3 to 99.7), respectively. Higher sensitivity and specificity were observed in the chemiluminescence immunoassay method compared with the Darui Biotech ELISA kit. The developed high sensitivity and specificity chemiluminescence immunoassay IgG testing method combined with the RT-PCR approach can improve the clinical diagnosis for SARS-CoV-2 infections and thus contribute to the control of COVID-19 expansion.

Ibrutinib suppresses intracellular mycobacterium tuberculosis growth by inducing macrophage autophagy.

Tuberculosis (TB) is a major cause of morbidity and mortality worldwide. The host-directed therapy is a promising strategy for TB treatment that synergize with anti-TB treatment drugs. In this study, we found that the anti-chronic lymphocytic leukemia drug, ibrutinib, inhibited the growth of intracellular Mtb in human macrophages. Mechanisms studies showed that ibrutinib treatment significantly decreased p62 and increased LC3b proteins in Mtb infected macrophages. In addition, ibrutinib increased LC3b colocalization with intracellular Mtb and auto-lysosome fusion. Furthermore, inhibition of autophagy by using siRNA targeting ATG7 abolished the effect of ibrutinib-mediated suppression of intracellular Mtb. Next, we found that ibrutinib induced autophagy was through inhibition of BTK/Akt/mTOR pathway. Finally, we confirmed that ibrutinib treatment significantly reduced Mtb load in mediastinal node and spleen of Mtb infected mice. In conclusion, our data suggest that ibrutinib is a potential host-directed therapy candidate against TB.