Longitudinal Magnetic Resonance Imaging Tracking of Transplanted Neural Progenitor Cells in the Spinal Cord Utilizing the Bright-Ferritin Mechanism.

Human neural progenitor cells (hNPCs) hold promise for treating spinal cord injury. Studies to date have focused on improving their regenerative potential and therapeutic effect. Equally important is ensuring successful delivery and engraftment of hNPCs at the injury site. Unfortunately, no current imaging solution for cell tracking is compatible with long-term monitoring in vivo. The objective of this study was to apply a novel bright-ferritin magnetic resonance imaging (MRI) mechanism to track hNPC transplants longitudinally and on demand in the rat spinal cord. We genetically modified hNPCs to stably overexpress human ferritin. Ferritin-overexpressing (FT) hNPCs labeled with 0.2 mM manganese provided significant T1-induced bright contrast on in vitro MRI, with no adverse effect on cell viability, morphology, proliferation, and differentiation. In vivo, 2 M cells were injected into the cervical spinal cord of Rowett nude rats. MRI employed T1-weighted acquisitions and T1 mapping on a 3 T scanner. Conventional short-term cell tracking was performed using exogenous Mn labeling prior to cell transplantation, which displayed transient bright contrast on MRI 1 day after cell transplantation and disappeared after 1 week. In contrast, long-term cell tracking using bright-ferritin allowed on-demand signal recall upon Mn supplementation and precise visualization of the surviving hNPC graft. In fact, this new cell tracking technology identified 7 weeks post-transplantation as the timepoint by which substantial hNPC integration occurred. Spatial distribution of hNPCs on MRI matched that on histology. In summary, bright-ferritin provides the first demonstration of long-term, on-demand, high-resolution, and specific tracking of hNPCs in the rat spinal cord.

Hypothalamic astrocyte NAD+ salvage pathway mediates the coupling of dietary fat overconsumption in a mouse model of obesity.

Nicotinamide adenine dinucleotide (NAD)+ serves as a crucial coenzyme in numerous essential biological reactions, and its cellular availability relies on the activity of the nicotinamide phosphoribosyltransferase (NAMPT)-catalyzed salvage pathway. Here we show that treatment with saturated fatty acids activates the NAD+ salvage pathway in hypothalamic astrocytes. Furthermore, inhibition of this pathway mitigates hypothalamic inflammation and attenuates the development of obesity in male mice fed a high-fat diet (HFD). Mechanistically, CD38 functions downstream of the NAD+ salvage pathway in hypothalamic astrocytes burdened with excess fat. The activation of the astrocytic NAMPT-NAD+-CD38 axis in response to fat overload induces proinflammatory responses in the hypothalamus. It also leads to aberrantly activated basal Ca2+ signals and compromised Ca2+ responses to metabolic hormones such as insulin, leptin, and glucagon-like peptide 1, ultimately resulting in dysfunctional hypothalamic astrocytes. Our findings highlight the significant contribution of the hypothalamic astrocytic NAD+ salvage pathway, along with its downstream CD38, to HFD-induced obesity.

Realizing Minimally Perturbed, Nonlocal Chiral Metasurfaces for Direct Stokes Parameter Detection.

Recent development in nonlocal resonance based chiral metasurfaces draws great attention due to their abilities to strongly interact with circularly polarized light at a relatively narrow spectral bandwidth. However, there still remain challenges in realizing effective nonlocal chiral metasurfaces in optical frequency due to demanding fabrications such as 3D-multilayered or nanoscaled chiral geometry, which, in particular, limit their applications to polarimetric detection with high-Q spectra. Here, we study the underlying working principles and reveal the important role of the interaction between high-Q nonlocal resonance and low-Q localized Mie resonance in realizing effective nonlocal chiral metasurfaces. Based on the working principles, we demonstrate one of the simplest types of nonlocal chiral metasurfaces which directly detects a set of Stokes parameters without the numerical combination of transmitted values presented from typical Stokes metasurfaces. This is achieved by minimally altering the geometry and filling ratio of every constituent nanostructure in a unit cell, facilitating consistent-sized nanolithography for all samples experimentally at a targeted wavelength with relatively high-Q spectra. This work provides an alternative design rule to realizing effective polarimetric metasurfaces and the potential applications of nonlocal Stokes parameters detection.

Comparison of Particulate Steroid Injection vs Nonparticulate Steroid Injection for Lumbar Radicular Pain: A Systematic Review and Meta-analysis.

OBJECTIVE: The objective of this study was to identify the difference on pain intensity and disability between particulate and nonparticulate steroid injections in patients with lumbar radicular pain. Subgroup analysis by study design, type of particulate steroid, and follow-up duration were performed. DATA SOURCES: We performed the literature search in the PubMed, Embase, and Cochrane Library up March, 2023. STUDY SELECTION: Studies, including randomized controlled trials (RCTs) and nonrandomized studies, that compared particulate steroid injection and nonparticulate steroid injection in patients with lumbar radicular pain were independently reviewed by 2 reviewers for eligibility for inclusion. DATA EXTRACTION: Outcomes of interest were pain intensity and disability. Two reviewers independently assessed the quality of included studies using the revised Cochrane Risk of Bias (RoB2.0) tool for RCTs and the Risk of Bias in Nonrandomized Studies of Interventions Tool (ROBINS-I) for nonrandomized studies. Effect sizes were estimated using mean difference (MD) and standardized mean difference (SMD). DATA SYNTHESIS: A total of 10 studies were included in this meta-analysis. The results showed no significant difference in visual analog scale, disability score and the numbers of patients with 50% pain reduction between particulate and nonparticulate steroid injection groups (P>.05). Particulate steroid injections showed significant better effect in pain scale in RCTs (MD=0.62; 95% CI 0.08-1.16, P=.02). In subgroup analysis with steroid types, methylprednisolone showed better effect compared with dexamethasone, while dexamethasone showed better effect compared with betamethasone. CONCLUSIONS: This meta-analysis suggested no significant differences between the particulate and nonparticulate steroid groups in pain or disability score. Therefore, considering the safety profile of nonparticulate steroids, nonparticulate steroid injection may be helpful in patients with lumbar radicular pain.

Injection of a PMMA-doped MSC spheroid gel for the treatment of painful osteoporotic vertebral compression fractures.

We aimed to develop a biocompatible treatment to overcome the limitations of polymethyl methacrylate (PMMA) vertebroplasty for osteoporotic compression fracture patients. We synthesized an injectable hydrogel containing PMMA. Mesenchymal stem cell (MSC) spheroids were included in the injectable PMMA-doped gel (= PMMA-doped spheroid gel). In vitro, the osteogenic/anti-inflammatory effects of the embedded spheroids were investigated by the quantitative real-time polymerase chain reaction method. In vivo, we used ovariectomy (OVX)-induced osteoporotic rats with injured femurs to investigate the pain-relief effects. The OVX rats were divided into four groups according to the materials injected (non, PMMA, PMMA gel, and PMMA-spheroid gel) into the lesion. The immunofluorescence (IF) intensity levels of painful markers in dorsal root ganglia (DRG) were measured. In vitro, a volumetric ratio of the gel of 8 (gel):2 (PMMA) was non-cytotoxic for MSCs and promoted the expression of osteogenic/anti-inflammatory markers. In vivo, the values of several bone parameters in the PMMA-doped spheroid gel group showed remarkable increases compared to those in the PMMA group. In addition, the IF intensity levels of the painful markers were noticeably decreased in the PMMA-spheroid gel group. We, therefore, suggest that this treatment can be useful for osteoporotic vertebral compression fracture patients.

Axon guidance gene-targeted siRNA delivery system improves neural stem cell transplantation therapy after spinal cord injury.

BACKGROUND: Neural stem cells (NSCs) derived from the embryonic spinal cord are excellent candidates for the cellular regeneration of lost neural cells after spinal cord injury (SCI). Semaphorin 3 A (Sema3A) is well known as being implicated in the major axon guidance of the growth cone as a repulsive function during the development of the central nervous system, yet its function in NSC transplantation therapy for SCI has not been investigated. Here, we report for the first time that embryonic spinal cord-derived NSCs significantly express Sema3A in the SCI environment, potentially facilitating inhibition of cell proliferation after transplantation. METHODS: siRNA-Sema3A was conjugated with poly-l-lysin-coated gold nanoparticles (AuNPs) through a charge interaction process. NSCs were isolated from embryonic spinal cords of rats. Then, the cells were embedded into a dual-degradable hydrogel with the siRNA- Sema3A loaded-AuNPs and transplanted after complete SCI in rats. RESULTS: The knockdown of Sema3A by delivering siRNA nanoparticles via dual-degradable hydrogels led to a significant increase in cell survival and neuronal differentiation of the transplanted NSCs after SCI. Of note, the knockdown of Sema3A increased the synaptic connectivity of transplanted NSC in the injured spinal cord. Moreover, extracellular matrix molecule and functional recovery were significantly improved in Sema3A-inhibited rats compared to those in rats with only NSCs transplanted. CONCLUSIONS: These findings demonstrate the important role of Sema3A in NSC transplantation therapy, which may be considered as a future cell transplantation therapy for SCI cases.

Anti-Inflammatory Effects of Tegoprazan in Lipopolysaccharide-Stimulated Bone-Marrow-Derived Macrophages.

The purpose of this study was to investigate the anti-inflammatory effect of tegoprazan (TEGO) in lipopolysaccharide (LPS)-stimulated bone-marrow-derived macrophages (BMMs). To this end, compared to methylprednisolone (MP; positive control), we evaluated whether TEGO effectively differentiates LPS-stimulated BMMs into M2-phenotype macrophages. Moreover, the expression of pro- and anti-inflammatory cytokines genes influenced by TEGO was measured using quantitative real-time polymerase chain reaction (qRT-PCR) analysis. TEGO was found to reduce nitric oxide (NO) production in BMMs significantly. In addition, TEGO significantly decreased and increased the gene expression levels of pro-inflammatory and anti-inflammatory cytokines, respectively. In addition, we evaluated the phosphorylated values of the extracellular signal-regulatory kinase (ERK) and p38 in the mitogen-activated protein (MAP) kinase signaling pathway through Western blotting. TEGO significantly reduced the phosphorylated values of the ERK and p38. In other words, TEGO suppressed the various pro-inflammatory responses in LPS-induced BMMs. These results show that TEGO has the potential to be used as an anti-inflammatory agent.

High- versus Low-dose Steroid Injection for Adhesive Capsulitis (Frozen Shoulder): A Systematic Review and Meta-analysis.

BACKGROUND: Intraarticular steroid injections are a commonly used and proven treatment for frozen shoulder; however, there is no scientific basis for a certified dose. OBJECTIVES: This study aimed to identify the difference between high- and low-dose steroid injections treatments and suggest an appropriate dose. STUDY DESIGN: Systematic review and meta-analysis. METHODS: The MEDLINE, EMBASE, and Cochrane electronic databases were searched through February 15, 2023 for eligible randomized controlled trials. The effects of high- and low-dose steroid injections were calculated as standardized mean differences (SMD) in pain, shoulder range of motion (ROM), and functional improvement. The Grading of Recommendations Assessment, Development and Evaluation (GRADE) approach was used to evaluate evidence quality. RESULTS: Four studies with 274 patients were included in the final analysis. The meta-analysis showed that improvement in pain (SMD, 0.10; 95% CI, -0.12 to 0.32), ROM (SMD, 0.07; 95% CI, -0.05 to 0.19), and functional improvement (SMD, 0.08; 95% CI, -0.10 to 0.26) did not differ significantly between the high- and low-dose steroid injections. Subgroup follow-up analyses also showed no clinically significant differences in SMD for pain, ROM, and functional scale measurement in any subgroups (after 3 weeks, 6 weeks, and one year). One article described that, although there was no significant difference in adverse events frequency between the high- and low-dose groups, flushing tended to occur more frequently in the high-dose group. LIMITATIONS: Limitations are the small number of studies included in the meta-analysis, no disease stage considered, and a short follow-up period. CONCLUSIONS: This meta-analysis suggests there are no significant differences between the high- and low-dose steroid groups in pain, ROM, or functional improvement. Therefore, considering the side effects of high-dose steroids, starting with low-dose steroids is recommended. However, further studies are needed to establish exact protocols according to disease severity. KEY WORDS: Frozen shoulder, adhesive capsulitis, steroids, triamcinolone acetonide, injections, intraarticular, optimal dose, meta-analysis, randomized controlled trial.

Current Status of Epidemiology, Diagnosis, Therapeutics, and Vaccines for the Re-Emerging Human Monkeypox Virus.

Monkeypox (Mpox) virus, a member of the Poxviridae family, causes a severe illness similar to smallpox, which is characterized by symptoms such as high fever, rash, and pustules. Human-to-human transmission cases have been reported but remained low since the first recorded case of human infection occurred in the Congo in 1970. Recently, Mpox has re-emerged, leading to an alarming surge in infections worldwide since 2022, originating in the United Kingdom. Consequently, the World Health Organization (WHO) officially declared the '2022-23 Mpox outbreak'. Currently, no specific therapy or vaccine is available for Mpox. Therefore, patients infected with Mpox are treated using conventional therapies developed for smallpox. However, the vaccines developed for smallpox have demonstrated only partial efficacy against Mpox, allowing viral transmission among humans. In this review, we discuss the current epidemiology of the ongoing Mpox outbreak and provide an update on the progress made in diagnosis, treatment, and development of vaccines for Mpox.

Circulating blood eNAMPT drives the circadian rhythms in locomotor activity and energy expenditure.

Nicotinamide adenine dinucleotide (NAD+) is an essential cofactor of critical enzymes including protein deacetylase sirtuins/SIRTs and its levels in mammalian cells rely on the nicotinamide phosphoribosyltransferase (NAMPT)-mediated salvage pathway. Intracellular NAMPT (iNAMPT) is secreted and found in the blood as extracellular NAMPT (eNAMPT). In the liver, the iNAMPT-NAD+ axis oscillates in a circadian manner and regulates the cellular clockwork. Here we show that the hypothalamic NAD+ levels show a distinct circadian fluctuation with a nocturnal rise in lean mice. This rhythm is in phase with that of plasma eNAMPT levels but not with that of hypothalamic iNAMPT levels. Chemical and genetic blockade of eNAMPT profoundly inhibit the nighttime elevations in hypothalamic NAD+ levels as well as those in locomotor activity (LMA) and energy expenditure (EE). Conversely, elevation of plasma eNAMPT by NAMPT administration increases hypothalamic NAD+ levels and stimulates LMA and EE via the hypothalamic NAD+-SIRT-FOXO1-melanocortin pathway. Notably, obese animals display a markedly blunted circadian oscillation in blood eNAMPT-hypothalamic NAD+-FOXO1 axis as well as LMA and EE. Our findings indicate that the eNAMPT regulation of hypothalamic NAD+ biosynthesis underlies circadian physiology and that this system can be significantly disrupted by obesity.

Near-flat top bandpass filter based on non-local resonance in a dielectric metasurface.

Localized light matter interaction at a resonant nanostructure facilitates spectrally selective transmission of light, which has led to demonstrations of ultrathin metasurface based optical filters. Unfortunately, due to the nature of Lorentzian spectral line shape in such resonances, it is inevitable to suffer significant spectral crosstalk. In this work, we demonstrate a conceptually new type of spectral filter which exhibits near flat-top bandpass with minimized spectral overlaps. To realize this, we leverage the recent development of non-local resonance in dielectric nanostructure to design a double-layered optical filter with performance comparable to the ideal spectral filters. The designed metasurface shows averaged transmission of more than 90% across the target spectral band and suppressed transmission of less than 10% out of the spectral band.

Real-Time Visualizing Nucleation and Growth of Electrodes for Post-Lithium-Ion Batteries.

ConspectusUntil recently, most studies on nucleation and growth mechanisms have employed electrochemical transient measurements, and numerous models have been established on various metal electrode elements. Contrary to the conventional tip-induced nucleation and growth model, a base-induced nucleation and growth mode was discovered not so long ago, which highlighted the importance of direct real-time observations such as visualization. As analysis techniques developed, diverse in situ/operando imaging methods have spurred the fundamental understanding of complex and dynamic battery electrochemistry. Experimental observations of alkali Li and Na metals are limited and difficult because their high reactivity makes not only the fabrication but also the analysis itself challenging. Na metal has high reactivity to electrolytes. Accordingly, it is difficult to visualize the Na deposition in real-time due to gas evolution and resolution limitation. Only a few studies have examined the Na deposition and dissolution reactions in operando. It is generally believed that the Mg anode is free from the dendrite growth of Mg metal, and Mg deposition preferentially occurs along the surface direction. However, whether the Mg anode always follows the dendrite-free growth has currently become a controversial topic and is being discussed and redefined based on real-time imaging analyses. In addition, a variety of morphological evolutions in the metal anodes are required to be systematically distinguished by key parameters. Real-time imaging analysis can directly confirm the solid-liquid-solid multiphase conversion reactions of S and Se cathodes. S and Se elements belong to the same chalcogen group, but their crystal structures and morphological changes significantly differ in each electrode during deposition and dissolution reactions. Therefore, it is necessitated to discuss the nucleation and growth behaviors by examining intrinsic properties of each element in chalcogen cathodes. Considering that a mechanistic understanding of the Se cathode is in its infancy, its nucleation and growth behaviors must be further explored through fundamental studies. In this Account, we aim to discuss the nucleation and growth behaviors of metal (Li, Na, and Mg) anodes and chalcogen (S and Se) cathodes. To elucidate their nucleation and growth mechanisms, we overview the morphological evolutions on the electrode surface and interface by in situ/operando visualizations. Our recent studies covering Li, Na, Mg, S, and Se electrodes verified by operando X-ray imaging are used as critical resources in understanding their nucleation and growth behaviors. Overall, with validation of the complex and dynamic nucleation and growth behaviors of metal and chalcogen electrodes by in situ/operando visualization methods, we hope that this Account can contribute to supporting the fundamental knowledge for the development of high-energy-density metal and chalcogen electrodes.

Ocular tropism of SARS-CoV-2 in animal models with retinal inflammation via neuronal invasion following intranasal inoculation.

Although ocular manifestations are reported in patients with COVID-19, consensus on ocular tropism of SARS-CoV-2 is lacking. Here, we infect K18-hACE2 transgenic mice with SARS-CoV-2 using various routes. We observe ocular manifestation and retinal inflammation with production of pro-inflammatory cytokines in the eyes of intranasally (IN)-infected mice. Intratracheal (IT) infection results in dissemination of the virus from the lungs to the brain and eyes via trigeminal and optic nerves. Ocular and neuronal invasions are confirmed using intracerebral (IC) infection. Notably, the eye-dropped (ED) virus does not cause lung infection and becomes undetectable with time. Ocular and neurotropic distribution of the virus in vivo is evident in fluorescence imaging with an infectious clone of SARS-CoV-2-mCherry. The ocular tropic and neuroinvasive characteristics of SARS-CoV-2 are confirmed in wild-type Syrian hamsters. Our data can improve the understanding regarding viral transmission and clinical characteristics of SARS-CoV-2 and help in improving COVID-19 control procedures.

PLK1-ELAVL1/HuR-miR-122 signaling facilitates hepatitis C virus proliferation.

The liver-specific microRNA, miR-122, plays an essential role in the propagation of hepatitis C virus (HCV) by binding directly to the 5'-end of its genomic RNA. Despite its significance for HCV proliferation, the host factors responsible for regulating miR-122 remain largely unknown. In this study, we identified the cellular RNA-binding protein, ELAVL1/HuR (embryonic lethal-abnormal vision-like 1/human antigen R), as critically contributing to miR-122 biogenesis by strong binding to the 3'-end of miR-122. The availability of ELAVL1/HuR was highly correlated with HCV proliferation in replicon, infectious, and chronically infected patient conditions. Furthermore, by screening a kinase inhibitor library, we identified rigosertib, an anticancer agent under clinical trials, as having both miR-122-modulating and anti-HCV activities that were mediated by its ability to target polo-like kinase 1 (PLK1) and subsequently modulate ELAVL1/HuR-miR-122 signaling. The expression of PLK1 was also highly correlated with HCV proliferation and the HCV positivity of HCC patients. ELAVL1/HuR-miR-122 signaling and its mediation of PLK1-dependent HCV proliferation were demonstrated by performing various rescue experiments and utilizing an HCV mutant with low dependency on miR-122. In addition, the HCV-inhibitory effectiveness of rigosertib was validated in various HCV-relevant conditions, including replicons, infected cells, and replicon-harboring mice. Rigosertib was highly effective in inhibiting the proliferation of not only wild-type HCVs, but also sofosbuvir resistance-associated substitution-bearing HCVs. Our study identifies PLK1-ELAVL1/HuR-miR-122 signaling as a regulatory axis that is critical for HCV proliferation, and suggests that a therapeutic approach targeting this host cell signaling pathway could be useful for treating HCV and HCV-associated diseases.

Influence of Zika virus 3'-end sequence and nonstructural protein evolution on the viral replication competence and virulence.

Zika virus (ZIKV) has been circulating in human networks over 70 years since its first appearance in Africa, yet little is known about whether the viral 3'-terminal sequence and nonstructural (NS) protein diverged genetically from ancient ZIKV have different effects on viral replication and virulence in currently prevailing Asian lineage ZIKV. Here we show, by a reverse genetics approach using an infectious cDNA clone for a consensus sequence (Con1) of ZIKV, which represents Asian ZIKV strains, and another clone derived from the MR766 strain isolated in Uganda, Africa in 1947, that the 3'-end sequence -UUUCU-3' homogeneously present in MR766 genome and the -GUCU-3' sequence strictly conserved in Asian ZIKV isolates are functionally equivalent in viral replication and gene expression. By gene swapping experiments using the two infectious cDNA clones, we show that the NS1-5 proteins of MR766 enhance replication competence of ZIKV Con1. The Con1, which was less virulent than MR766, acquired severe bilateral hindlimb paralysis when its NS1-5 genes were replaced by the counterparts of MR766 in type I interferon receptor (IFNAR1)-deficient A129 mice. Moreover, MR766 NS5 RNA-dependent RNA polymerase (RdRp) alone also rendered the Con1 virulent, despite there being no difference in RdRp activity between MR766 and Con1 NS5 proteins. By contrast, the Con1 derivatives expressing MR766 Nsps, like Con1, did not develop severe disease in wild-type mice treated with an IFNAR1 blocking antibody. Together, our findings uncover an unprecedented role for ZIKV NS proteins in determining viral pathogenicity in immunocompromised hosts.

Transplantation of neuron-inducing grafts embedding positively charged gold nanoparticles for the treatment of spinal cord injury.

In this study, we aimed to investigate the recovery after traumatic spinal cord injury (SCI) by inducing cellular differentiation of transplanted neural stem cells (NSCs) into neurons. We dissociated NSCs from the spinal cords of Fisher 344 rat embryos. An injectable gel crosslinked with glycol chitosan and oxidized hyaluronate was used as a vehicle for NSC transplantation. The gel graft containing the NSC and positively charged gold nanoparticles (pGNP) was implanted into spinal cord lesions in Sprague-Dawley rats (NSC-pGNP gel group). Cellular differentiation of grafted NSCs into neurons (stained with ß-tubulin III [also called Tuj1]) was significantly increased in the NSC-pGNP gel group (***p < 0.001) compared to those of two control groups (NSC and NSC gel groups) in the SCI conditions. The NSC-pGNP gel group showed the lowest differentiation into astrocytes (stained with glial fibrillary acidic protein). Regeneration of damaged axons (stained with biotinylated dextran amines) within the lesion was two-fold higher in the NSC-pGNP gel group than that in the NSC gel group. The highest locomotor scores were also found in the NSC-pGNP gel group. These outcomes suggest that neuron-inducing pGNP gel graft embedding embryonic spinal cord-derived NSCs can be a useful type of stem cell therapy after SCI.

Immunological and Pathological Peculiarity of Severe Acute Respiratory Syndrome Coronavirus 2 Beta Variant.

Diverse severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants have emerged since the beginning of the COVID-19 pandemic. We investigated the immunological and pathological peculiarity of the SARS-CoV-2 beta variant of concern (VoC) compared to the ancestral strain. Comparative analysis of phenotype and pathology revealed that the beta VoC induces slower disease progression and a prolonged presymptomatic period in the early stages of SARS-CoV-2 infection but ultimately causes sudden death in the late stages of infection in the K18-hACE2 mouse model. The beta VoC induced enhanced activation of CXCL1/2-CXCR2-NLRP3-IL-1ß signal cascade accelerating neutrophil recruitment and lung pathology in beta variant-infected mice, as evidenced by multiple analyses of SARS-CoV-2-induced inflammatory cytokines and transcriptomes. CCL2 was one of the most highly secreted cytokines in the early stages of infection. Its blockade reduced virus-induced weight loss and delayed mortality. Our study provides a better understanding of the variant characteristics and need for treatment. IMPORTANCE Since the outbreak of COVID-19, diverse SARS-CoV-2 variants have been identified. These variants have different infectivity and transmissibility from the ancestral strains. However, underlying molecular mechanisms have not yet been fully elucidated. In our study, the beta variant showed distinct pathological conditions and cytokine release kinetics from an ancestral strain in a mouse model. It was associated with higher neutrophil recruitment by increased levels of CXCL1/2, CXCR2, and interleukin 1ß (IL-1ß) at a later stage of viral infection. Our study will provide a better understanding of SARS-CoV-2 pathogenesis.

Current Updates on COVID-19 Vaccines and Therapeutics: As of June 2022.

Since COVID-19, caused by SARS-CoV-2 infection, has become a global issue, many vaccines and therapeutic candidates have been developed or are being developed against the COVID-19 endemic and the next wave. However, it is difficult to overcome the spread and mutation rate of SARS-CoV-2 in the COVID-19 pandemic because development of vaccines and therapeutics involves considerable social cost and time, as well as research capabilities. Thus, assessing the development status of these agents is important for advancing efficient research strategies. In this review, we summarize the status of 141 vaccines and 345 therapeutic candidates under development worldwide, according to their development stage and characteristics. As of June 2022, 32 vaccines and 12 therapeutics have been approved for emergency use. Although the development of four of these therapeutics was terminated owing to their low efficacy against various variants of SARS-CoV-2, many new candidates that have completed phase 3 clinical trials have been awaiting phase 4 clinical trials or full approval by the Food and Drug Administration (FDA). These efforts are expected to contribute to establishing an efficient research strategy to overcome the COVID-19 pandemic and facilitate its transition toward an endemic phase. Electronic Supplementary Material ESM: The online version of this article (doi: 10.1007/s12257-022-0188-4) contains supplementary material, which is available to authorized users.

Human Alphacoronavirus Universal Primers for Genome Amplification and Sequencing.

Rapid and accurate sequencing covering the entire genome is essential to identify genetic variations of viral pathogens. However, due to the low viral titers in clinical samples, certain amplification steps are required for viral genome sequencing. At present, there are no universal primers available for alphacoronaviruses and that, since these viruses have diverse strains, new primers specific to the target strain must be continuously developed for sequencing. Thus, in this study, we aimed to develop a universal primer set valid for all human alphacoronaviruses and applicable to samples containing trace amounts of the virus. To this aim, we designed overlapping primer pairs capable of amplifying the entire genome of all known human alphacoronaviruses. The selected primers, named the AC primer set, were composed of 10 primer pairs stretching over the entire genome of alphacoronaviruses, and produced PCR products of the expected size (3-5 kb) from both the HCoV-229E and HCoV-NL63 strains. After genome amplification, an evaluation using various sequencing platforms was carried out. The amplicon library sequencing data were assembled into complete genome sequences in all sequencing strategies examined in this study. The sequencing accuracy varied depending on the sequencing technology, but all sequencing methods showed a sequencing error of less than 0.01%. In the mock clinical specimen, the detection limit was 10-3 PFU/ml (102 copies/ml). The AC primer set and experimental procedure optimized in this study may enable the fast diagnosis of mutant alphacoronaviruses in future epidemics.

Escape and Over-Activation of Innate Immune Responses by SARS-CoV-2: Two Faces of a Coin.

In the past 20 years, coronaviruses (CoVs), including SARS-CoV-1, MERS-CoV, and SARS-CoV-2, have rapidly evolved and emerged in the human population. The innate immune system is the first line of defense against invading pathogens. Multiple host cellular receptors can trigger the innate immune system to eliminate invading pathogens. However, these CoVs have acquired strategies to evade innate immune responses by avoiding recognition by host sensors, leading to impaired interferon (IFN) production and antagonizing of the IFN signaling pathways. In contrast, the dysregulated induction of inflammasomes, leading to uncontrolled production of IL-1 family cytokines (IL-1ß and IL-18) and pyroptosis, has been associated with COVID-19 pathogenesis. This review summarizes innate immune evasion strategies employed by SARS-CoV-1 and MERS-CoV in brief and SARS-CoV-2 in more detail. In addition, we outline potential mechanisms of inflammasome activation and evasion and their impact on disease prognosis.