Specific pupylation as IDEntity reporter (SPIDER) for the identification of protein-biomolecule interactions.

Protein-biomolecule interactions play pivotal roles in almost all biological processes. For a biomolecule of interest, the identification of the interacting protein(s) is essential. For this need, although many assays are available, highly robust and reliable methods are always desired. By combining a substrate-based proximity labeling activity from the pupylation pathway of Mycobacterium tuberculosis and the streptavidin (SA)-biotin system, we developed the Specific Pupylation as IDEntity Reporter (SPIDER) method for identifying protein-biomolecule interactions. Using SPIDER, we validated the interactions between the known binding proteins of protein, DNA, RNA, and small molecule. We successfully applied SPIDER to construct the global protein interactome for m6A and mRNA, identified a variety of uncharacterized m6A binding proteins, and validated SRSF7 as a potential m6A reader. We globally identified the binding proteins for lenalidomide and CobB. Moreover, we identified SARS-CoV-2-specific receptors on the cell membrane. Overall, SPIDER is powerful and highly accessible for the study of protein-biomolecule interactions.

Modulation of innate immune response to viruses including SARS-CoV-2 by progesterone.

Whether and how innate antiviral response is regulated by humoral metabolism remains enigmatic. We show that viral infection induces progesterone via the hypothalamic-pituitary-adrenal axis in mice. Progesterone induces downstream antiviral genes and promotes innate antiviral response in cells and mice, whereas knockout of the progesterone receptor PGR has opposite effects. Mechanistically, stimulation of PGR by progesterone activates the tyrosine kinase SRC, which phosphorylates the transcriptional factor IRF3 at Y107, leading to its activation and induction of antiviral genes. SARS-CoV-2-infected patients have increased progesterone levels, and which are co-related with decreased severity of COVID-19. Our findings reveal how progesterone modulates host innate antiviral response, and point to progesterone as a potential immunomodulatory reagent for infectious and inflammatory diseases.

Retrospective quantitative detection of SARS-CoV-2 by digital PCR showing high accuracy for low viral load specimens.

INTRODUCTION: Accurate detection of severe acute respiratory syndrome coronavirus 2 is critical for diagnosis and disease status evaluation of Coronavirus disease 2019. We retrospectively evaluated the infection status and viral load of severe acute respiratory syndrome coronavirus 2 in Nantong city, China, using a quantitative digital polymerase chain reaction and reverse-transcription PCR. METHODOLOGY: A total of 103 clinical specimens from 31 patients were collected and tested by digital PCR and reverse-transcription PCR. RESULTS: The overall accuracy of digital PCR was 96.8%, which was higher than the overall accuracy of 87.1% for reverse-transcription PCR. 4 (3.88%) specimens for ORF1ab and 22 (21.36%) specimens for N gene were negative by reverse-transcription PCR but positive by digital PCR. 3 (2.91%, 3/103) specimens of ORF1ab were positive by reverse-transcription PCR but negative by digital PCR. The digital PCR assay exhibited higher sensitivity to measure the N gene than the ORF1ab gene (p < 0.01). CONCLUSIONS: Our results showed that digital PCR assay provides more reliable detection of Coronavirus disease 2019 than reverse-transcription PCR, especially for low viral load specimens.

Pathogenesis and Treatment of Cytokine Storm Induced by Infectious Diseases.

Cytokine storm is a phenomenon characterized by strong elevated circulating cytokines that most often occur after an overreactive immune system is activated by an acute systemic infection. A variety of cells participate in cytokine storm induction and progression, with profiles of cytokines released during cytokine storm varying from disease to disease. This review focuses on pathophysiological mechanisms underlying cytokine storm induction and progression induced by pathogenic invasive infectious diseases. Strategies for targeted treatment of various types of infection-induced cytokine storms are described from both host and pathogen perspectives. In summary, current studies indicate that cytokine storm-targeted therapies can effectively alleviate tissue damage while promoting the clearance of invading pathogens. Based on this premise, "multi-omics" immune system profiling should facilitate the development of more effective therapeutic strategies to alleviate cytokine storms caused by various diseases.

Tubular epithelial cell-to-macrophage communication forms a negative feedback loop via extracellular vesicle transfer to promote renal inflammation and apoptosis in diabetic nephropathy.

Background: Macrophage infiltration around lipotoxic tubular epithelial cells (TECs) is a hallmark of diabetic nephropathy (DN). However, how these two types of cells communicate remains obscure. We previously demonstrated that LRG1 was elevated in the process of kidney injury. Here, we demonstrated that macrophage-derived, LRG1-enriched extracellular vesicles (EVs) exacerbated DN. Methods: We induced an experimental T2DM mouse model with a HFD diet for four months. Renal primary epithelial cells and macrophage-derived EVs were isolated from T2D mice by differential ultracentrifugation. To investigate whether lipotoxic TEC-derived EV (EVe) activate macrophages, mouse bone marrow-derived macrophages (BMDMs) were incubated with EVe. To investigate whether activated macrophage-derived EVs (EVm) induce lipotoxic TEC apoptosis, EVm were cocultured with primary renal tubular epithelial cells. Subsequently, we evaluated the effect of LRG1 in EVe by investigating the apoptosis mechanism. Results: We demonstrated that incubation of primary TECs of DN or HK-2 mTECs with lysophosphatidyl choline (LPC) increased the release of EVe. Interestingly, TEC-derived EVe activated an inflammatory phenotype in macrophages and induced the release of macrophage-derived EVm. Furthermore, EVm could induce apoptosis in TECs injured by LPC. Importantly, we found that leucine-rich α-2-glycoprotein 1 (LRG1)-enriched EVe activated macrophages via a TGFßR1-dependent process and that tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-enriched EVm induced apoptosis in injured TECs via a death receptor 5 (DR5)-dependent process. Conclusion: Our findings indicated a novel cell communication mechanism between tubular epithelial cells and macrophages in DN, which could be a potential therapeutic target.

Management of Endovascular Treatment for Acute Ischemic Stroke During the COVID-19 Pandemic at a Single Institution in Beijing, China: A Brief Report.

BACKGROUND: The coronavirus disease (COVID-19) pandemic is currently a major challenge for health care systems around the world. For a time-sensitive emergency such as acute ischemic stroke (AIS), streamlined workflow times are essential to ensure good clinical outcomes. METHODS: The aim of this single-center, retrospective, observational study was to describe changes in stroke workflow patterns and clinical care during the COVID-19 pandemic. Data from AIS patients undergoing emergent endovascular treatment (EVT) between 23 January and 8 April 2020 were retrospectively collected and compared with data from patients admitted during a similar period in 2019. The primary outcome was difference in time from symptom onset to recanalization. Secondary outcomes included workflow times, clinical management, discharge outcomes, and health-economic data. RESULTS: In all, 21 AIS patients were admitted for emergent EVT during the 77-day study period, compared with 42 cases in 2019. Median time from symptom onset to recanalization was 132 minutes longer during the pandemic compared with the previous year (672 vs. 540 min, P=0.049). Patients admitted during the pandemic had a higher likelihood of endotracheal intubation (84.6% vs. 42.4%, P<0.05) and a higher incidence of delayed extubation after EVT (69.2% vs. 45.5%, P<0.05). National Institutes of Health Stroke Scale at hospital discharge was similar in the 2 cohorts, whereas neurointensive care unit stay was longer in patients admitted during the pandemic (10 vs. 7 days, P=0.013) and hospitalization costs were higher (123.9 vs. 95.2 thousand Chinese Yuan, P=0.052). CONCLUSION: Disruptions to medical services during the COVID-19 pandemic has particularly impacted AIS patients undergoing emergent EVT, resulting in increased workflow times. A structured and multidisciplinary protocol should be implemented to minimize treatment delays and maximize patient outcomes.

Temporal lung changes in high-resolution chest computed tomography for coronavirus disease 2019.

OBJECTIVE: To evaluate temporal lung changes in coronavirus disease 2019 (COVID-19) in high-resolution computed tomography (HRCT) and to determine the appropriate computed tomographic (CT) follow-up time. METHODS: Eighty-six patients with two or more HRCT scans who were diagnosed with COVID-19 were included. The CT score and major CT findings were evaluated. RESULTS: Eighty-two (95.3%) patients had lesions on the initial HRCT scans. Most scans showed bilateral, multifocal lung lesions, with multiple lobes involved and diffuse distribution. For fifty-seven patients with type I (progress compared with the initial CT score), the CT score reached a peak at 12 days and the nadir at 36 days. For twenty-nine patients with type II (no progress compared with the initial CT score), the lowest CT score was reached at 23 days. On the final HRCT scans (>21 days), patients with a reticular pattern were older than those without a reticular pattern. CONCLUSION: The appropriate follow-up time of CT scans is during the second week (approximately 12 days) and the fourth to fifth weeks (approximately 23-36 days) from the onset of illness. These times could help reduce the CT radiation dose and show timely changes in the course of the disease by CT.

The Perioperative Management of Subarachnoid Hemorrhage During the Coronavirus Disease 2019 Pandemic in China.

BACKGROUND: For most of the international community outside the epicenter, coronavirus disease 2019 (COVID-19) containment is normalizing, and daily medical practice runs parallel to preventing and treating COVID-19. This experience of simultaneously conducting emergent surgery and infection control for COVID-19 disease is useful outside the epicenter during the pandemic. CASE DESCRIPTION: In this single-center retrospective observational study, we enrolled patients with subarachnoid hemorrhage (SAH) who were emergently admitted from January 23 to April 8, 2020. Based on the COVID-19 triage, patients with SAH were divided into 3 categories: positive, negative, and under investigation. During 77 days, 90 patients with SAH were admitted at the center. The median age was 55 years (range, 18-80 years) and 40 patients (44.4%) were male. None was positive, 42 patients were negative, and 48 patients were under investigation for COVID-19 before surgery. During the same period, 9 patients were diagnosed with COVID-19 without nosocomial infection. CONCLUSIONS: Rescuing patients with SAH and containment of COVID-19 benefit from joint prevention and control, a centralized system of equipment distribution and personnel assignment, and quick workflow establishment.