RESUMEN
COVID-19 induces re-circulating long-lived memory B cells (MBC) that, upon re-encounter with the pathogen, are induced to mount immunoglobulin responses. During convalescence, antibodies are subjected to affinity maturation, which enhances the antibody binding strength and generates new specificities that neutralize virus variants. Here, we performed a single-cell RNA sequencing analysis of spike-specific B cells from a SARS-CoV-2 convalescent subject. After COVID-19 vaccination, matured infection-induced MBC underwent recall and differentiated into plasmablasts. Furthermore, the transcriptomic profiles of newly activated B cells transiently shifted toward the ones of atypical and CXCR3+ B cells and several B-cell clonotypes massively expanded. We expressed monoclonal antibodies (mAbs) from all B-cell clones from the largest clonotype that used the VH3-53 gene segment. The in vitro analysis revealed that some somatic hypermutations enhanced the neutralization breadth of mAbs in a putatively stochastic manner. Thus, somatic hypermutation of B-cell clonotypes generates an anticipatory memory that can neutralize new virus variants.
Asunto(s)
Anticuerpos Neutralizantes , Anticuerpos Antivirales , COVID-19 , SARS-CoV-2 , Hipermutación Somática de Inmunoglobulina , SARS-CoV-2/inmunología , Humanos , Hipermutación Somática de Inmunoglobulina/genética , COVID-19/inmunología , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Células B de Memoria/inmunología , Glicoproteína de la Espiga del Coronavirus/inmunología , Glicoproteína de la Espiga del Coronavirus/genética , Anticuerpos Monoclonales/inmunología , Linfocitos B/inmunología , Región Variable de Inmunoglobulina/genética , Región Variable de Inmunoglobulina/inmunología , Memoria Inmunológica/inmunología , Vacunas contra la COVID-19/inmunologíaRESUMEN
During the last decade, a wide variety of cellular RNA sensors and structural characteristics of their agonists have been identified. On the basis of this knowledge, RNA formulations were developed as innovative adjuvant candidates. In contrast to DNA, RNA does not have genotoxic potential and is rapidly degraded. In many aspects, RNA mimics viral infections and induces considerably strong immune responses. Additionally, RNA-based adjuvants can be designed so that distinct RNA sensors can be triggered according to requirements of individual vaccines. Furthermore, RNA can be synthesized in vitro in a cell-free system, and recent developments in formulation technology have led to reduced RNA degradation within the body. These features qualify RNA as a promising adjuvant candidate. Here, we discuss latest developments in the field of RNA-based adjuvants and highlight differences between human and mouse nucleic acid sensors, which constitute a challenge in the development of RNA-based adjuvants. Finally, we discuss how RNA-based adjuvants are currently handled with regard to regulatory requirements.
Asunto(s)
Adyuvantes Inmunológicos , ARN/inmunología , Vacunas Virales/inmunología , Virosis/inmunología , Animales , Regulación Gubernamental , Humanos , Inmunidad , Ratones , Receptores de Reconocimiento de Patrones/metabolismoRESUMEN
Among innovative adjuvants conferring a Th1-shift, RNAdjuvant is a promising candidate. This adjuvant consists of a 547-nt uncapped noncoding ssRNA containing polyU repeats that is stabilized by a cationic carrier peptide. Whereas vaccination of mice with an influenza subunit vaccine induced moderate virus-specific IgG1, vaccination together with RNAdjuvant significantly enhanced this IgG1 and additionally promoted the formation of IgG2b/c, which is indicative of Th1 responses. Furthermore, such sera neutralized influenza virus, whereas this effect was not detected upon vaccination with the subunit vaccine alone. Similarly, upon vaccination with virus-like particles displaying vesicular stomatitis virus G protein, RNAdjuvant promoted the formation of virus-specific IgG2b/c and enhanced neutralizing IgG responses to an extent that mice were protected against lethal virus infection. RNAdjuvant induced dendritic cells to upregulate activation markers and produce IFN-I. Although these effects were strictly TLR7 dependent, RNAdjuvant-mediated augmentation of vaccine responses needed concurrent TLR and RIG-I-like helicase signaling. This was indicated by the absence of the adjuvant effect in vaccinated MyD88-/-Cardif-/- mice, which are devoid of TLR (with the exception of TLR3) and RIG-I-like helicase signaling, whereas in vaccinated MyD88-/- mice the adjuvant effect was reduced. Notably, i.m. RNAdjuvant injection induced local IFN-I responses and did not induce systemic effects, implying good tolerability and a favorable safety profile for RNAdjuvant.
Asunto(s)
Adyuvantes Inmunológicos , Inmunoglobulina G/sangre , Vacunas contra la Influenza/inmunología , Glicoproteínas de Membrana/inmunología , ARN no Traducido/inmunología , Receptor Toll-Like 7/inmunología , Vacunas de Partículas Similares a Virus/inmunología , Adyuvantes Inmunológicos/efectos adversos , Animales , Anticuerpos Antivirales/sangre , Proteína 58 DEAD Box/inmunología , Proteína 58 DEAD Box/metabolismo , Inmunoglobulina G/inmunología , Vacunas contra la Influenza/administración & dosificación , Glicoproteínas de Membrana/administración & dosificación , Glicoproteínas de Membrana/metabolismo , Ratones , Ratones Endogámicos BALB C , Factor 88 de Diferenciación Mieloide/deficiencia , Factor 88 de Diferenciación Mieloide/metabolismo , Células TH1/inmunología , Receptor Toll-Like 7/metabolismo , Vacunación , Vacunas de Subunidad/inmunología , Vacunas de Partículas Similares a Virus/administración & dosificación , Proteínas del Envoltorio Viral/administración & dosificación , Proteínas del Envoltorio Viral/inmunologíaAsunto(s)
Betula , Citomegalovirus , Humanos , Monocitos , Alérgenos , Polen , Células Dendríticas , Extractos Vegetales/farmacología , ÁrbolesRESUMEN
OBJECTIVE: Systemic sclerosis (SSc) is a severe rheumatic disease, causing fibrotic tissue rearrangement. Aberrant Toll-like receptor 8 (TLR8) transcripts in plasmacytoid dendritic cells (pDC) were recently linked to SSc pathogenesis, which is at least partially mediated by increased type I interferon (IFN-I) responses. Here, we addressed the functional role of TLR8 signaling in different immune cell subsets of patients with SSc. METHODS: Monocytes, conventional dendritic cells (cDC), and pDC, from blood and skin of patients with SSc were analyzed for TLR8 protein expression. To assess TLR function, cytokine responses upon TLR7 and TLR8 stimulation were studied. To identify relevant alterations specific for SSc (n = 16), patients with primary Sjögren's syndrome (pSS) (n = 10) and healthy controls (HC) (n = 13) were included into the study. RESULTS: In all individuals, TLR8 was expressed in monocytes and cDC, but not in pDC. The TLR8 expression levels were overall similar in patients with SSc, pSS, and HC. Additionally, in all study participants, TLR8 stimulation of pDC did not induce IFN-I expression. In contrast, monocytes from patients with SSc revealed increased interleukin-10 (IL-10) responses upon TLR8 (SSc vs. HC: p = 0.0126) and TLR7/8 stimulation (SSc vs. HC: p = 0.0170). CONCLUSION: TLR8 protein is not expressed in pDC of patients with SSc. Accordingly, they do not respond to TLR8 stimulation. In contrast, monocytes of patients with SSc respond to TLR8 stimulation with increased IL-10 responses. Therefore, TLR8 signaling in monocytes participates in SSc pathogenesis by conferring aberrant IL-10 expression.