ABSTRACT
Background: Although respiratory failure is the hallmark of severe disease, it is increasingly clear that Coronavirus Disease 2019 (COVID-19) is a multi-system disorder. The presence of gastrointestinal (GI) involvement by Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has been suggested by epidemiological, clinical, non-human primate, in-vitro (enteroid) and ex-vivo (human biopsy) studies. Having recently documented persistence of SARS-CoV-2 within the intestinal epithelium 7 months after infection, here we aimed to study mucosal immune cell abnormalities in individuals with prior history of COVID-19. Methods: Individuals with previous COVID-19 diagnosis (by either RT–PCR or seroconversion) and controls (without RT-PCR or serological evidence of prior COVID-19 infection) undergoing endoscopic evaluation were recruited into the study (Table 1). Colonic and small intestinal (duodenal and ileal) biopsies were analyzed by multiparameter flow cytometry for mucosal immune cell populations including myeloid cells (classical and non-classical monocytes, dendritic cell subsets), T cells (subsets and activation state), B cells (including plasma cells) and NK cells. Persistence of viral antigens was determined by immunofluorescence microscopy (n=30) using a previously published anti-nucleocapsid (NP) antibody. Results: Thirty subjects with a previous history of COVID-19 (post-COVID), median of 4 months from diagnosis (range 1-10 months), were recruited and compared with 40 normal volunteer (NV) controls. Relative to controls, post-COVID subjects displayed higher frequencies of classical (CD14+) monocytes in both, the colon and the small bowel, while significantly higher frequencies of conventional dendritic cells (cDC)1 (lin-HLA-DRhiCD14- CD11c+CD141+) and cDC2 (lin-HLA-DRhiCD14-CD11c+CD1c+) were noted in the colon. Among NK subsets, CD56bright CD16- NK cells were significantly higher in the colon of post-COVID subjects. Among T cell subsets, CD8+ tissue resident memory T cells (CD8+CD69+CD103+) were significantly increased in colon of post-COVID subjects compared to NV. Among B cell subsets, plasma cells (CD3-CD27+CD38hi) trended higher (p= 0.06), while mucosal B cells (CD3-CD19+) were significantly lower in the terminal ileum of post-COVID subjects compared to NV. Finally, with IF, we detected SARS-CoV-2 NP in 10 out of 30 (33%) of post-COVID subjects (Figure 1). Conclusion: Innate and adaptive immune cell abnormalities persist in the intestinal mucosa of post-COVID subjects for up to 10 months and may reflect viral persistence or immune cell dysregulation in the intestines. These findings have major implications for understanding the pathogenesis of long-term sequelae of COVID-19, including long-haul COVID.(Table Presented)(Figure Presented)
ABSTRACT
Background: Although respiratory failure is the hallmark of severe disease, it is increasingly clear that Coronavirus Disease 2019 (COVID-19) is a multi-system disorder. The presence of gastrointestinal (Gl) involvement by Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has been suggested by epidemiological, clinical, non-human primate, invitro (enteroid) and ex-vivo (human biopsy) studies. Having recently documented persistence of SAR-CoV-2 within the intestinal epithelium 7 months after infection, here we aimed to study mucosal immune cell abnormalities in individuals with prior history of COVID-19. Methods: Individuals with previous COVID-19 diagnosis (by either RT- PCR or seroconversion) and controls (without RT-PCR or serological evidence of prior COVID-19 infection) undergoing endoscopic evaluation were recruited into the study (Table 1,2). Colonic and small intestinal (duodenal and ileal) biopsies were analyzed by multiparameter flow cytometry for mucosal immune cell populations including myeloid cells (classical and non-classical monocytes, dendritic cell subsets), T cells (subsets and activation state), B cells (including plasma cells). Persistence of viral antigens was determined by immunofluorescence microscopy (n=30) using a previously published anti-nucleocapsid (NP) antibody. Results: Thirty subjects with a previous history of COVID-19 (post- COVID), median of 4 months from diagnosis (range 1-10 months), were recruited and compared with 40 normal volunteer (NV) controls. Relative to controls, post-COVID subjects displayed higher frequencies of classical (CD14+) monocytes in both, the colon and the small bowel, while significantly higher frequencies of conventional dendritic cells (cDC) 1 (lin-HLA-DRhiCD14-CD11c+CD141+) and cDC2 (lin-HLA-DRhiCD14-- CD11c+CD1c+) were noted in the colon only. Among T cell subsets, CD8+ tissue resident memory T cells (CD8+CD69+CD103+) were significantly increased in colon of post-COVID subjects compared to NV. Among B cell subsets, plasma cells (CD3-CD27+CD38hi) trended higher (p=0.06), while mucosal B cells (CD3-CD19+) were significantly lower in the terminal ileum of post-COVID subjects compared to NV. Finally, with IF, we detected SARS-CoV-2 NP in 10 out of 30 (33%) of post-COVID subjects (Figure 1). There were no significant correlations of these cell populations with either time after the infection or IF positivity. Conclusion: Innate and adaptive immune cell abnormalities persist in the intestinal mucosa of post-COVID subjects for up to 10 months and may reflect viral persistence or immune cell dysregulation in the intestines. These findings have major implications for understanding the pathogenesis of long term sequela of COVID-19, including long-haul COVID.
ABSTRACT
Introduction: SARS-CoV-2, the causative pathogen for COVID-19, engages host ACE2 receptor for cellular entry. The brush border of the small intestines express high levels of ACE2. Gastrointestinal (GI) manifestations are common among COVID-19 patients. However, to date, there is limited information regarding intestinal response to SARS-CoV-2 infection. Methods: Intestinal biopsies were obtained from 17 COVID-19 patients (17.3 ± positive nasal swab) for cellular and transcriptomic analyses using mass cytometry and RNA-sequencing, respectively. Ten uninfected individuals served as compartment (EC) and lamina propria (LP) were analyzed separately. Results: The cellular profiles from LP of COVID-19 patients showed reduced frequencies of CD206+ conventional dendritic cells (CDC2s) and plasmacytoid (CD123+) dendritic cells Effector T cell (PD1+CD38+) frequency was increased in the LP and Intraepithelial lymphocytes (IEL) were increased in the EC of COVID-19 patients, with a concomitant decrease in CD206+ CDC2s. RNA sequencing active downregulation of genes involved in inflammatory pathways including IBD-associated pathways, while an upregulation of intestinal barrier function Gene expression of Neuropilin-1 (NRP-1), a putative SARS-CoV-2 receptor as well as key inflammatory cytokines (IL-1b, IFN-g, CCL24 and CXCL8) were significantly reduced in controls. A low intensity antiviral host response signature was observed predominantly in EC as opposed to LP suggesting viral localization to epithelium. Conclusions: Epithelial, myeloid and lymphoid cell alterations characterize intestinal response to SARS-CoV-2 infection with an unanticipated downregulation of key inflammatory pathways that have been implicated in adverse outcomes associated with These data stand in contrast to the inflammatory response reported in the systemic compartments and identify a potential mitigating role of the GI
ABSTRACT
Background: Host proteins ACE-2 and TMPRSS2 facilitate SARS-CoV-2 infection and are expressed in the lungs as well as the intestinal tract, particularly in the small bowel. Gastrointestinal symptoms represent the most common extrapulmonary manifestation of COVID-19. Viral RNA has been isolated from fecal samples from COVID-19 patients, where it can persist longer than detection in nasopharyngeal swabs. While SARS-CoV-2 infection of enterocytes has been demonstrated in vitro, in vivo studies are lacking. Methods: Small intestinal biopsies from patients who underwent clinically indicated endoscopic procedures after a positive SARS-CoV-2 nasopharyngeal swab (n=27) or were found to have positive serology (n=2) were analyzed by immunofluorescence (IF) (n=25) and electron microscopy (EM) (n=14) for the presence of virus. Clinical details were also collected. Results: Sixteen of 29 patients had detectable SARS-CoV-2 antigen by either IF or EM (Figure 1). Virus was restricted to the epithelium and patchy in distribution. Virus was detected as soon as 15 days after symptom onset and persisted up to 6 months after symptom resolution. Five patients were nasopharyngeal swab positive at the time of procedure and, of these, 4 had detectable antigen on biopsy. Despite the presence of virus, only 9/16 patients had any signs of inflammation on histology, and when present, this was mild. In two patients where virus was present at 3 months and 4 months, additional biopsies were obtained at 7 months and 6 months, respectively. Viral antigen was persistently detected in both patients and both patients were nasopharyngeal swab negative for all procedures. Interestingly, only 37.5% (6 of 16) of patients with virus detected in the small bowel had GI symptoms (diarrhea, nausea or vomiting) during their acute COVID-19 illness as compared to 46.1% (6/13) of patients where no virus could be detected in the intestines. Conclusion: SARS-CoV-2 infects enterocytes in humans in vivo and can persist in the intestines up to 7 months following symptoms resolution. This persistence is not associated with an overt inflammatory infiltrate and does not appear to correlate with presence of GI symptoms in the acute COVID-19 setting.
ABSTRACT
Background: SARS-CoV-2, the etiopathological agent for COVID-19, engages host ACE2 receptor for cellular entry. The brush border of the small intestines express high levels of ACE2 in physiological conditions. Gastrointestinal (GI) manifestations are common among COVID-19 patients. However, to date, there is limited information regarding intestinal response to SARS-CoV-2 infection. Methods: Intestinal biopsies were obtained from 17 COVID-19 patients (17.3-17.5 days from the last positive nasal swab) for cellular and transcriptomic analyses using mass cytometry and RNA-sequencing. Ten COVID-uninfected individuals served as controls. The epithelial compartment (EC) and lamina propria (LP) were analyzed separately. Results: The cellular profiles of intestinal tissues from COVID-19 patients showed reduced frequencies of CD206+ CDC2s and plasmacytoid dendritic cells in the LP of COVID-19 patients by mass cytometry. Effector T cell (PD1+CD38+) frequency was increased in the LP and blood of COVID-19 patients. Intraepithelial lymphocytes (IEL) were increased in the EC of COVID-19 patients, with a concomitant decrease in CD206+ CDC2s. RNA sequencing revealed an active downregulation of genes involved in inflammatory pathways including Th17 and IBD-associated pathways, while an upregulation of intestinal barrier function (mucin biosynthesis), amino acid metabolism and mineral absorption pathways was noted. Gene expression of Neuropilin-1 (NRP-1), a putative SARSCoV-2 receptor as well as key inflammatory cytokines (IL-1β, IFNγ, CCL24 and CXCL8) were significantly reduced in COVID-19 patients compared to controls. A low intensity antiviral host response signature was observed predominantly in EC reflecting the cellular localization of the virus. Conclusion: Epithelial, myeloid and lymphoid cell alterations characterize intestinal response to SARS-CoV-2 infection with an unanticipated downregulation of key inflammatory pathways that have been implicated in adverse outcomes associated with COVID-19. These data stand in contrast to reports from the pulmonary and systemic compartments and identify a potential mitigating role of the GI tract in COVID-19-associated immunopathology.