CRISPR Screening and Comparative LC-MS Analysis Identify Genes Mediating Efficacy of Delamanid and Pretomanid against Mycobacterium tuberculosis.

Tuberculosis (TB), the leading cause of death from bacterial infections worldwide, results from infection with Mycobacterium tuberculosis (Mtb). The antitubercular agents delamanid (DLM) and pretomanid (PMD) are nitroimidazole prodrugs that require activation by an enzyme intrinsic to Mtb; however, the mechanism(s) of action and the associated metabolic pathways are largely unclear. Profiling of the chemical-genetic interactions of PMD and DLM in Mtb using combined CRISPR screening reveals that the mutation of rv2073c increases susceptibility of Mtb to these nitroimidazole drugs both in vitro and in infected mice, whereas mutation of rv0078 increases drug resistance. Further assays show that Rv2073c might confer intrinsic resistance to DLM/PMD by interfering with inhibition of the drug target, decaprenylphophoryl-2-keto-b-D-erythro-pentose reductase (DprE2), by active nicotinamide adenine dinucleotide (NAD) adducts. Characterization of the metabolic pathways of DLM/PMD in Mtb using a combination of chemical genetics and comparative liquid chromatography-mass spectrometry (LC-MS) analysis of DLM/PMD metabolites reveals that Rv0077c, which is negatively regulated by Rv0078, mediates drug resistance by metabolizing activated DLM/PMD. These results might guide development of new nitroimidazole prodrugs and new regimens for TB treatment.

Five-Year Outcomes of Bioresorbable Stent Therapy for Coronary Heart Disease: A Systematic Review and Meta-Analysis of Randomized Controlled Trials.

Background: The efficacy of bioresorbable vascular scaffolds (BVS) compared to metallic stents for the treatment of coronary heart disease remains controversial. The analysis of clinical outcomes at five years following the initial treatment has yet to be reviewed. This study sought to assess the five-year outcomes in randomized controlled trials of BVS in the treatment of coronary heart disease using a systematic review and meta-analysis. Methods: A systematic database search was conducted from their inception to June 30th, 2023 using various Medical Subject Headings (MeSH) terms including: "Coronary Disease", "Bioresorbable stent", "Randomized controlled trials". Results: After a rigorous selection process, a total of five high-quality articles were finally included in this study. Each trial demonstrated a low risk of bias. After 5 years, bioresorbable stents showed outcomes similar to conventional metal stents in terms of cardiac mortality. However, they were inferior in terms of lesion revascularization rates, in-stent thrombosis rates, target lesion failure, target vessel failure, and myocardial infarction. Conclusions: While bioresorbable stents are comparable to metallic stents in terms of cardiac mortality rates, they exhibit significant drawbacks that warrant clinical consideration.

Absence of PD-L1 signaling hinders macrophage defense against Mycobacterium tuberculosis via upregulating STAT3/IL-6 pathway.

The blockade of programmed death-ligand 1 (PD-L1) pathway has been clinically used in cancer immunotherapy, while its effects on infectious diseases remain elusive. Roles of PD-L1 signaling in the macrophage-mediated innate immune defense against M.tb is unclear. In this study, the outcomes of tuberculosis (TB) in wild-type (WT) mice treated with anti-PD-1/PD-L1 therapy and macrophage-specific Pdl1-knockout (Pdl1ΔΜΦ) mice were compared. Treatment with anti-PD-L1 or anti-PD-1 benefited protection against M.tb infection in WT mice, while Pdl1ΔΜΦ mice exhibited the increased susceptibility to M.tb infection. Mechanistically, the absence of PD-L1 signaling impaired M.tb killing by macrophages. Furthermore, elevated STAT3 activation was found in PD-L1-deficient macrophages, leading to increased interleukin (IL)-6 production and reduced inducible nitric oxide synthase (iNOS) expression. Inhibiting STAT3 phosphorylation partially impeded the increase in IL-6 production and restored iNOS expression in these PD-L1-deficient cells. These findings provide valuable insights into the complexity and mechanisms underlying anti-PD-L1 therapy in the context of tuberculosis.

Flu-CED: A comparative transcriptomics database of influenza virus-infected human and animal models.

BACKGROUND: The continuing emergence of influenza virus has highlighted the value of public databases and related bioinformatic analysis tools in investigating transcriptomic change caused by different influenza virus infections in human and animal models. METHODS: We collected a large amount of transcriptome research data related to influenza virus-infected human and animal models in public databases (GEO and ArrayExpress), and extracted and integrated array and metadata. The gene expression matrix was generated through strictly quality control, balance, standardization, batch correction, and gene annotation. We then analyzed gene expression in different species, virus, cells/tissues or after antibody/vaccine treatment and imported sample metadata and gene expression datasets into the database. RESULTS: Overall, maintaining careful processing and quality control, we collected 8064 samples from 103 independent datasets, and constructed a comparative transcriptomics database of influenza virus named the Flu-CED database (Influenza comparative expression database, https://flu.com-med.org.cn/). Using integrated and processed transcriptomic data, we established a user-friendly website for realizing the integration, online retrieval, visualization, and exploration of gene expression of influenza virus infection in different species and the biological functions involved in differential genes. Flu-CED can quickly query single and multi-gene expression profiles, combining different experimental conditions for comparative transcriptome analysis, identifying differentially expressed genes (DEGs) between comparison groups, and conveniently finding DEGs. CONCLUSION: Flu-CED provides data resources and tools for analyzing gene expression in human and animal models infected with influenza virus that can deepen our understanding of the mechanisms underlying disease occurrence and development, and enable prediction of key genes or therapeutic targets that can be used for medical research.

Application of combined CRISPR screening for genetic and chemical-genetic interaction profiling in Mycobacterium tuberculosis.

CRISPR screening, including CRISPR interference (CRISPRi) and CRISPR-knockout (CRISPR-KO) screening, has become a powerful technology in the genetic screening of eukaryotes. In contrast with eukaryotes, CRISPR-KO screening has not yet been applied to functional genomics studies in bacteria. Here, we constructed genome-scale CRISPR-KO and also CRISPRi libraries in Mycobacterium tuberculosis (Mtb). We first examined these libraries to identify genes essential for Mtb viability. Subsequent screening identified dozens of genes associated with resistance/susceptibility to the antitubercular drug bedaquiline (BDQ). Genetic and chemical validation of the screening results suggested that it provided a valuable resource to investigate mechanisms of action underlying the effects of BDQ and to identify chemical-genetic synergies that can be used to optimize tuberculosis therapy. In summary, our results demonstrate the potential for efficient genome-wide CRISPR-KO screening in bacteria and establish a combined CRISPR screening approach for high-throughput investigation of genetic and chemical-genetic interactions in Mtb.

The Effects of ATIR Blocker on the Severity of COVID-19 in Hypertensive Inpatients and Virulence of SARS-CoV-2 in Hypertensive hACE2 Transgenic Mice.

Angiotensin-converting enzyme 2 (ACE2) is required for the cellular entry of the severe acute respiratory syndrome coronavirus 2. ACE2, via the Ang-(1-7)-Mas-R axis, is part of the antihypertensive and cardioprotective effects of the renin-angiotensin system. We studied hospitalized COVID-19 patients with hypertension and hypertensive human(h) ACE2 transgenic mice to determine the outcome of COVID-19 with or without AT1 receptor (AT1R) blocker treatment. The severity of the illness and the levels of serum cardiac biomarkers (CK, CK-BM, cTnI), as well as the inflammation markers (IL-1, IL-6, CRP), were lesser in hypertensive COVID-19 patients treated with AT1R blockers than those treated with other antihypertensive drugs. Hypertensive hACE2 transgenic mice, pretreated with AT1R blocker, had increased ACE2 expression and SARS-CoV-2 in the kidney and heart, 1 day post-infection. We conclude that those hypertensive patients treated with AT1R blocker may be at higher risk for SARS-CoV-2 infection. However, AT1R blockers had no effect on the severity of the illness but instead may have protected COVID-19 patients from heart injury, via the ACE2-angiotensin1-7-Mas receptor axis.

Preclinical and clinical progress for HDAC as a putative target for epigenetic remodeling and functionality of immune cells.

Genetic changes are difficult to reverse; thus, epigenetic aberrations, including changes in DNA methylation, histone modifications, and noncoding RNAs, with potential reversibility, have attracted attention as pharmaceutical targets. The current paradigm is that histone deacetylases (HDACs) regulate gene expression via deacetylation of histone and nonhistone proteins or by forming corepressor complexes with transcription factors. The emergence of epigenetic tools related to HDACs can be used as diagnostic and therapeutic markers. HDAC inhibitors that block specific or a series of HDACs have proven to be a powerful therapeutic treatment for immune-related diseases. Here, we summarize the various roles of HDACs and HDAC inhibitors in the development and function of innate and adaptive immune cells and their implications for various diseases and therapies.

[Establishment of an Iron-overloaded Mouse Model with Tuberculosis and Analysis of the Iron Metabolism Index].

Objective To establish a mouse model of exogenous iron overload combined with tuberculosis(TB). Methods C57BL/6N mice were divided into negative control, low-, medium-, and high-dose iron groups and received intraperitoneal injection of iron dextran at 0, 3.75, 7.50, and 15.00 mg/dose(3 times/week for 4 weeks), respectively.After 4 weeks, the organ morphology and body weight of the mice were evaluated.The content of serum iron, ferritin, transferrin, and transferrin receptor was determined by ELISA.Heart, liver, spleen, lung, kidney, and small intestine were analyzed for tissue iron content and iron deposition pathology.Mycobacterium tuberculosis(Mtb)standard strain H37Rv was injected via tail vein to infect the mice receiving moderate-dose iron to establish an iron-overloaded mouse model of active TB.HE staining and Mtb culture were employed to analyze tuberculous lesions and bacterial loads of lung, spleen and liver tissues. Results The weight gain percentages of mice in the negative control, low-, medium-, and high-dose iron groups were 25.47%, 25.22%, 24.74%, and 21.36%, respectively, which was significantly lower in the high-dose group than in the negative control(F=17.235, P=0.027), low-dose(F=15.206, P=0.031), and medium-dose(F=11.061, P=0.036)groups.Liver had the highest iron content, followed by spleen, kidney, and small intestine.The iron content in heart and lung tissues of the low-dose group had no significant difference compared with those of the negative control group(F=19.023, P=0.715;F=23.193, P=0.902).Serum iron and ferritin in the iron-overloaded mice increased in a dose-dependent manner, while transferrin and transferrin receptor had no significant changes.HE and Prussian blue staining showed that the iron-overloaded mice had different degrees of iron deposition in tissues and high-dose iron caused liver and kidney damage.The lung(F=23.227, P=0.017), spleen(F=19.023, P=0.021), and liver(F=17.392, P=0.009)of the iron-overloaded mice with TB had a significantly shorter time of bacterial culture than those of the TB-infected mice without iron overload.The lung(F=21.012, P=0.007), spleen(F=20.173, P=0.002), and liver(F=19.091, P=0.005)of the iron-overloaded mice with TB had significantly higher bacterial loads than those of the TB-infected mice without iron overload. Conclusions The exogenous iron-overloaded mouse model with similar symptoms to patients with clinical iron overload can be established by intraperitoneal injection of medium-dose(7.50 mg/dose, 3 times/week for 4 weeks)iron dextran.Mtb injection through the tail vein can help construct a mouse model of iron overload combined with active TB.

The double-sided effects of Mycobacterium Bovis bacillus Calmette-Guérin vaccine.

Bacillus Calmette-Guérin (BCG), the only vaccine proven to be effective against tuberculosis (TB), is the most commonly used vaccine globally. In addition to its effects on mycobacterial diseases, an increasing amount of epidemiological and experimental evidence accumulated since its introduction in 1921 has shown that BCG also exerts non-specific effects against a number of diseases, such as non-mycobacterial infections, allergies and certain malignancies. Recent Corona Virus Disease 2019 (COVID-19) outbreak has put BCG, a classic vaccine with significant non-specific protection, into the spotlight again. This literature review briefly covers the diverse facets of BCG vaccine, providing new perspectives in terms of specific and non-specific protection mechanisms of this old, multifaceted, and controversial vaccine.