Correction: Promoter-regulated in vivo asymmetric self-assembly strategy to synthesize heterogeneous nanoparticles for signal amplification.

Correction for 'Promoter-regulated in vivo asymmetric self-assembly strategy to synthesize heterogeneous nanoparticles for signal amplification' by Chen Chen et al., Nanoscale, 2022, 14, 16180-16184, https://doi.org/10.1039/D2NR04661J.

Bifunctional Protein TC1 Mediated One-Pot Strategy for Robust Immobilization of DNA with High Accessibility.

Immobilizing DNA with high accessibility at the interface is attractive but challenging. Current methods often involve multiple chemical reactions and derivatives. In this study, an endonuclease, TC1, is introduced to develop a robust strategy for immobilizing DNA with enhanced accessibility. TC1 enables direct immobilization of DNA onto a solid support through self-catalytic DNA covalent coupling and robust solid adsorption capabilities. This method demonstrates high accessibility to target molecules, supported by the improved sensitivity of DNA hybridization and aptamer-target recognition assays. TC1-mediated DNA immobilization is a one-pot reaction that does not require chemical derivatives, making it promising for the development of high-performance DNA materials and technologies.

A bacteria-derived tetramerized protein ameliorates nonalcoholic steatohepatitis in mice via binding and relocating acetyl-coA carboxylase.

Increased de novo lipogenesis (DNL) is a major feature of nonalcoholic steatohepatitis (NASH). None of the drugs targeting the catalytic activity of acetyl-CoA carboxylase (ACC), the rate-limiting enzyme in the DNL process, have been approved by the FDA. Whether cytosolic ACC1 can be regulated spatially remains to be explored. Herein, we find that streptavidin (SA), which is a bacterium-derived tetrameric protein, forms cytosolic condensates and efficiently induces a spatial re-localization of ACC1 in liver cells, concomitant with inhibited lipid accumulation. Both SA tetrameric structure and multivalent protein interaction are required for condensate formation. Interestingly, the condensates are further characterized as gel-like membraneless organelle (SAGMO) and significantly restrict the cytosolic dispersion of ACC1 and fatty acid synthase. Notably, AAV-mediated delivery of SA partially blocks mouse liver DNL and ameliorates NASH without eliciting hypertriglyceridemia. In summary, our study shows that insulating lipogenesis-related proteins by SAGMO might be effective for NASH treatment.

Phosphorothioated DNA Engineered Liposomes as a General Platform for Stimuli-Responsive Cell-Specific Intracellular Delivery and Genome Editing.

Intracellular protein delivery is highly desirable for protein drug-based cell therapy. Established technologies suffer from poor cell-specific cytosolic protein delivery, which hampers the targeting therapy of specific cell populations. A fusogenic liposome system enables cytosolic delivery, but its ability of cell-specific and controllable delivery is quite limited. Inspired by the kinetics of viral fusion, we designed a phosphorothioated DNA coatings-modified fusogenic liposome to mimic the function of viral hemagglutinin. The macromolecular fusion machine docks cargo-loaded liposomes at the membrane of target cells, triggers membrane fusion upon pH or UV light stimuli, and facilitates cytosolic protein delivery. Our results showed efficient cell-targeted delivery of proteins of various sizes and charges, indicating the phosphorothioated DNA plug-in unit on liposomes could be a general strategy for spatial-temporally controllable protein delivery both in vitro and in vivo.

Rational design of a genome-based insulated system in Escherichia coli facilitates heterologous uricase expression for hyperuricemia treatment.

Hyperuricemia is a prevalent disease worldwide that is characterized by elevated urate levels in the blood owing to purine metabolic disorders, which can result in gout and comorbidities. To facilitate the treatment of hyperuricemia through the uricolysis, we engineered a probiotic Escherichia coli Nissle 1917 (EcN) named EcN C6 by inserting an FtsP-uricase cassette into an "insulated site" located between the uspG and ahpF genes. Expression of FtsP-uricase in this insulated region did not influence the probiotic properties or global gene transcription of EcN but strongly increased the enzymatic activity for urate degeneration, suggesting that the genome-based insulated system is an ideal strategy for EcN modification. Oral administration of EcN C6 successfully alleviated hyperuricemia, related symptoms and gut microbiota in a purine-rich food-induced hyperuricemia rat model and a uox-knockout mouse model. Together, our study provides an insulated site for heterologous gene expression in EcN strain and a recombinant EcN C6 strain as a safe and effective therapeutic candidate for hyperuricemia treatment.

Single-cell transcriptome landscape and antigen receptor dynamic during SARS-CoV-2 vaccination.

Vaccination by inactivated vaccine is an effective strategy to prevent the COVID-19 pandemic. However, the detailed molecular immune response at single-cell level is poorly understood. In this study, we systematically delineated the landscape of the pre- and post-vaccination single-cell transcriptome, TCR (T cell antigen receptor) and BCR (B cell antigen receptor) expression profile of vaccinated candidates. The bulk TCR sequencing analysis of COVID-19 patients was also performed. Enrichment of a clonal CD8+ T cell cluster expressing specific TCR was identified in both vaccination candidates and COVID-19 patients. These clonal CD8+ T cells showed high expression of cytotoxicity, phagosome and antigen presentation related genes. The cell-cell interaction analysis revealed that monocytes and dendritic cells could interact with these cells and initiate phagocytosis via ICAM1-ITGAM and ITGB2 signaling. Together, our study systematically deciphered the detailed immunological response during SARS-CoV-2 vaccination and infection. It may facilitate understanding the immune response and the T-cell therapy against COVID-19.

Promoter-regulated in vivo asymmetric self-assembly strategy to synthesize heterogeneous nanoparticles for signal amplification.

Signal amplification is commonly used to enhance the sensitivity of biological analysis. Here, we present a strategy involving in vivo asymmetric self-assembly combined with promoter strength regulation to synthesize heterogeneous nanoparticles for signal amplification. Two expression vectors were constructed by genetically inserting, respectively, signal and binding molecules into the hepatitis B core antigen protein (HBcAg) structure. Because of differential expression of the two recombinant proteins in the presence of a strong promoter (T7) and a weak promoter (Tac-1) and spontaneous asymmetric self-assembly in vivo, heterogeneous HBcAg nanoparticles (NPs) with a high ratio of signal-bearing to target-binding molecules were obtained. These nanoparticles contained a large number of green fluorescent proteins as signal molecules and a small number of B1 immunoglobulin-binding domains from protein G for antibody binding, thus enabling sensitive immunoassays. As a proof of concept, improved sensitivity for antibody detection was achieved using the heterogeneous nanoparticle conjugated with a secondary antibody molecule.

Intracellular Delivery of Micron-Sized Magnetic Particles through a Virus Infection Pathway.

Micron-sized magnetic particles (M-MPs) have low toxicity, strong magnetic signals, and long-term retention capability, which are significant advantages for their application in biomedical imaging. Unfortunately, M-MPs are only internalized by few cell types, such as macrophages and phagocytes, and because of this lack of active intracellular delivery, their applications are restricted. The emergence of self-assembled virus-like particles (VLPs) offers a viable approach to drive M-MPs into cells, although the specific mechanism has not been revealed. In this study, we investigated in detail the intracellular pathway of M-MPs mediated by VLPs using a fluorescence co-localization method. The results indicated that the intracellular movement of M-MPs was consistent with the virus infection pathway, specifically caveolae-dependent endocytosis, transportation through microtubules, and accumulation in the endoplasmic reticulum. This study provides experimental support for the active transport of M-MPs into other cell types, thereby further extending their applications.

Dysregulation of Innate and Adaptive Immune Responses in Asymptomatic SARS-CoV-2 Infection with Delayed Viral Clearance.

Asymptomatic infection with SARS-CoV-2 is a major concern in the control of the COVID-19 pandemic. Many questions concerning asymptomatic infection remain to be answered, for example, what are the differences in infectivity and the immune response between asymptomatic and symptomatic infections? In this study, based on a cohort established by the Wuchang District Health Bureau of Wuhan in the early stage of the COVID-19 pandemic in Wuhan in 2019, we conducted a comprehensive analysis of the clinical, virological, immunological, and epidemiological data of asymptomatic infections. The major findings of this study included: 1) the asymptomatic cohort enrolled this study exhibited low-grade but recurrent activity of viral replication; 2) despite a lack of overt clinical symptoms, asymptomatic infections exhibited ongoing innate and adaptive immune responses; 3) however, the immune response from asymptomatic infections was not activated adequately, which may lead to delayed viral clearance. Given the fragile equilibrium between viral infection and host immunity, and the delayed viral clearance in asymptomatic individuals, close viral monitoring should be scheduled, and therapeutic intervention may be needed.

Distinct gut microbiota and health outcomes in asymptomatic infection, viral nucleic acid test re-positive, and convalescent COVID-19 cases.

Gut microbiota composition is suggested to associate with coronavirus disease 2019 (COVID-19) severity, but the impact of gut microbiota on health outcomes is largely unclear. We recruited 81 individuals from Wuhan, China, including 13 asymptomatic infection cases (Group A), 24 COVID-19 convalescents with adverse outcomes (Group C), 31 severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) re-positive cases (Group D), and 13 non-COVID-19 healthy controls (Group H). The microbial features of Groups A and D were similar and exhibited higher gut microbial diversity and more abundant short-chain fatty acid (SCFA)-producing species than Group C. Group C was enriched with opportunistic pathogens and virulence factors related to adhesion and toxin production. The abundance of SCFA-producing species was negatively correlated, while Escherichia coli was positively correlated with adverse outcomes. All three groups (A, C, and D) were enriched with the mucus-degrading species Akkermansia muciniphila, but decreased with Bacteroides-encoded carbohydrate-active enzymes. The pathways of vitamin B6 metabolic and folate biosynthesis were decreased, while selenocompound metabolism was increased in the three groups. Specifically, the secondary bile acid (BA) metabolic pathway was enriched in Group A. Antibiotic resistance genes were common among the three groups. Conclusively, the gut microbiota was related to the health outcomes of COVID-19. Dietary supplementations (SCFAs, BA, selenium, folate, vitamin B6) may be beneficial to COVID-19 patients.

Case Report: A Re-Positive Case of SARS-CoV-2 Associated With Glaucoma.

The current pandemic of coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has already become a global threat to the human population. Infection with SARS-CoV-2 leads to a wide spectrum of clinical manifestations. Ocular abnormalities have been reported in association with COVID-19, but the nature of the impairments was not specified. Here, we report a case of a female patient diagnosed with glaucoma on re-hospitalization for ocular complications two months after being discharged from the hospital upon recovery from COVID-19. Meanwhile, the patient was found re-positive for SARS-CoV-2 in the upper respiratory tract. The infection was also diagnosed in the aqueous humor through immunostaining with antibodies against the N protein and S protein of SARS-CoV-2. Considering the eye is an immune-privileged site, we speculate that SARS-CoV-2 survived in the eye and resulted in the patient testing re-positive for SARS-CoV-2.

Serological investigation of asymptomatic cases of SARS-CoV-2 infection reveals weak and declining antibody responses.

Without an effective vaccine against SARS-CoV-2, the build-up of herd immunity through natural infection has been suggested as a means to control COVID-19. Although population immunity is typically estimated by the serological investigation of recovered patients, humoral immunity in asymptomatic subjects has not been well studied, although they represent a large proportion of all SARS-CoV-2 infection cases. In this study, we conducted a serosurvey of asymptomatic infections among food workers and performed serological and cellular response analyses of asymptomatic subjects in Wuhan, the original epicenter of the COVID-19 outbreak. Our data showed that up to 5.91% of the food workers carried SARS-CoV-2 IgG antibodies asymptomatically; however, in 90.4% of them, the antibody level declined over a 2-week period. IgM and IgG antibodies, including neutralizing antibodies, were significantly lower in asymptomatic subjects than in recovered symptomatic patients with similar disease courses. Furthermore, the asymptomatic subjects showed lymphopenia and a prominent decrease in the B-cell population, as well as a low frequency of antibody-secreting cells and a low cytokine response. These factors probably contributed to the low and unsustained antibody levels in asymptomatic subjects. Our results show that asymptomatic subjects are likely to be vulnerable to SARS-CoV-2 reinfection, and neither the proportion of population immunity nor the breadth of immune responses is sufficient for herd immunity.

Auxiliary Module Promotes the Synthesis of Carboxysomes in E. coli to Achieve High-Efficiency CO2 Assimilation.

Carboxysomes (CBs) are protein organelles in cyanobacteria, and they play a central role in assimilation of CO2. Heterologous synthesis of CBs in E. coli provides an opportunity for CO2-organic compound conversion under controlled conditions but remains challenging; specifically, the CO2 assimilation efficiency is insufficient. In this study, an auxiliary module was designed to assist self-assembly of CBs derived from a model species cyanobacteria Prochlorococcus marinus (P. marinus) MED4 for synthesizing in E. coli. The results indicated that the structural integrity of synthetic CBs is improved through the transmission electron microscope images and that the CBs have highly efficient CO2-concentrating ability as revealed by enzyme kinetic analysis. Furthermore, the bacterial growth curve and 13C-metabolic flux analysis not only consolidated the fact of CO2 assimilation by synthetic CBs in E. coli but also proved that the engineered strain could efficiently convert external CO2 to some metabolic intermediates (acetyl-CoA, malate, fumarate, tyrosine, etc.) of the central metabolic pathway. The synthesis of CBs of P. marinus MED4 in E. coli provides prospects for understanding their CO2 assimilation mechanism and realizing their modular application in synthetic biology.

Myoglobin and C-reactive protein are efficient and reliable early predictors of COVID-19 associated mortality.

Since the emergence of SARS-CoV-2, numerous studies have been attempting to determine biomarkers, which could rapidly and efficiently predict COVID-19 severity, however there is lack of consensus on a specific one. This retrospective cohort study is a comprehensive analysis of the initial symptoms, comorbidities and laboratory evaluation of patients, diagnosed with COVID-19 in Huoshenshan Hospital, Wuhan, from 4th February to 12th March, 2020. Based on the data collected from 63 severely ill patients from the onset of symptoms till the full recovery or demise, we found not only age (average 70) but also blood indicators as significant risk factors associated with multiple organ failure. The blood indices of all patients showed hepatic, renal, cardiac and hematopoietic dysfunction with imbalanced coagulatory biomarkers. We noticed that the levels of LDH (85%, P < .001), HBDH (76%, P < .001) and CRP (65%, P < .001) were significantly elevated in deceased patients, indicating hepatic impairment. Similarly, increased CK (15%, P = .002), Cre (37%, P = 0.102) and CysC (74%, P = 0.384) indicated renal damage. Cardiac injury was obvious from the significantly elevated level of Myoglobin (52%, P < .01), Troponin-I (65%, P = 0.273) and BNP (50%, P = .787). SARS-CoV-2 disturbs the hemolymphatic system as WBC# (73%, P = .002) and NEUT# (78%, P < .001) were significantly elevated in deceased patients. Likewise, the level of D-dimer (80%, P < .171), PT (87%, P = .031) and TT (57%, P = .053) was elevated, indicating coagulatory imbalances. We identified myoglobin and CRP as specific risk factors related to mortality and highly correlated to organ failure in COVID-19 disease.

Sustainability of SARS-CoV-2 Induced Humoral Immune Responses in COVID-19 Patients from Hospitalization to Convalescence Over Six Months.

Understanding the persistence of antibody in convalescent COVID-19 patients may help to answer the current major concerns such as the risk of reinfection, the protection period of vaccination and the possibility of building an active herd immunity. This retrospective cohort study included 172 COVID-19 patients who were hospitalized in Wuhan. A total of 404 serum samples were obtained over six months from hospitalization to convalescence. Antibodies in the specimens were quantitatively analyzed by the capture chemiluminescence immunoassays (CLIA). All patients were positive for the anti-SARS-CoV-2 IgM/IgG at the onset of COVID-19 symptoms, and the IgG antibody persisted in all the patients during the convalescence. However, only approximately 25% of patients can detect the IgM antibodies, IgM against N protein (N-IgM) and receptor binding domain of S protein (RBD-IgM) at the 27th week. The titers of IgM, N-IgM and RBD-IgM reduced to 16.7%, 17.6% and 15.2% of their peak values respectively. In contrast, the titers of IgG, N-IgG and RBD-IgG peaked at 4-5th week and reduced to 85.9%, 62.6% and 87.2% of their peak values respectively at the end of observation. Dynamic behavior of antibodies and their correlation in age, gender and severity groups were investigated. In general, the COVID-19 antibody was sustained at high levels for over six months in most of the convalescent patients. Only a few patients with antibody reducing to an undetectable level which needs further attention. The humoral immune response against SARS-CoV-2 infection in COVID-19 patients exhibits a typical dynamic of acquired immunity.

Rhomboidal Pt(II) Metallacycle-Based Hybrid Viral Nanoparticles for Cell Imaging.

Supramolecular coordination complexes (SCCs) have emerged as anticancer agents. Tracking the movement of these metallic anticancer agents plays an important role in the field of biomedicines. Herein, we describe a method for tracking the movement of a rhomboidal Pt(II) metallacycle agent using the quantum dots encapsidation in vitro self-assembly system of viral proteins. When incubated with living Vero cells, self-assembly of hybrid viral nanoparticles were employed for simultaneous cell imaging and visual transmission of the Pt(II) metallacycle agent. Considering these results, we believe that the multifunctional biomaterials consisting of a supramolecular coordination complex and quantum dots provide a new alternative for probing of the delivery of Pt(II) metallacycle drugs.

SARS-CoV-2 proteome microarray for global profiling of COVID-19 specific IgG and IgM responses.

We still know very little about how the human immune system responds to SARS-CoV-2. Here we construct a SARS-CoV-2 proteome microarray containing 18 out of the 28 predicted proteins and apply it to the characterization of the IgG and IgM antibodies responses in the sera from 29 convalescent patients. We find that all these patients had IgG and IgM antibodies that specifically bind SARS-CoV-2 proteins, particularly the N protein and S1 protein. Besides these proteins, significant antibody responses to ORF9b and NSP5 are also identified. We show that the S1 specific IgG signal positively correlates with age and the level of lactate dehydrogenase (LDH) and negatively correlates with lymphocyte percentage. Overall, this study presents a systemic view of the SARS-CoV-2 specific IgG and IgM responses and provides insights to aid the development of effective diagnostic, therapeutic and vaccination strategies.