Enhanced metanephric specification to functional proximal tubule enables toxicity screening and infectious disease modelling in kidney organoids (preprint)

While pluripotent stem cell-derived kidney organoids represent a promising approach for the study of renal disease, renal physiology and drug screening, the proximal nephron remains immature with limited evidence for key functional solute channels. This may reflect early mispatterning of the nephrogenic mesenchyme or insufficient maturation. In this study, prolonged differentiation and modification of media conditions to enhance metanephric nephron progenitor specification resulted in the induction of nephrons containing elongated and aligned proximal nephron segments together with SLC12A1+ loops of Henle. Nephron proximal segments showed superior HNF4A gene and protein expression, as well as upregulation of key functional transporters, including SLC3A1/2, SLC47A1, and SLC22A2. The striking proximo-distal orientation of nephrons was shown to result from localised WNT antagonism originating from the centre of the organoid. Functionality of such transporters was evidenced by albumin and organic cation uptake, as well as appropriate KIM-1 upregulation in response to the nephrotoxicant, cisplatin. PT-enhanced organoids also possessed improved expression of receptors associated with SARS-CoV2 entry, rendering these organoids susceptible to infection and able to support viral replication without co-location of ACE2 and TMPRSS2. These PT-enhanced organoids provide an accurate model with which to study human proximal tubule maturation, inherited and acquired proximal tubular disease, and drug and viral responses.

Integrated immune networks in SARS-CoV-2 infected pregnant women reveal differential NK cell and unconventional T cell activation (preprint)

Although pregnancy poses a greater risk for severe COVID-19, the underlying immunological changes associated with SARS-CoV-2 infection during pregnancy are poorly understood. We defined immune responses to SARS-CoV-2 in pregnant and non-pregnant women during acute and convalescent COVID-19 up to 258 days post symptom onset, quantifying 217 immunological parameters. Additionally, matched maternal and cord blood were collected from COVID-19 convalescent pregnancies. Although serological responses to SARS-CoV-2 were similar in pregnant and non-pregnant women, cellular immune analyses revealed marked differences in key NK cell and unconventional T cell responses during COVID-19 in pregnant women. While NK, γδ T cells and MAIT cells displayed pre-activated phenotypes in healthy pregnant women when compared to non-pregnant age-matched women, activation profiles of these pre-activated NK and unconventional T cells remained unchanged at acute and convalescent COVID-19 in pregnancy. Conversely, activation dynamics of NK and unconventional T cells were prototypical in non-pregnant women in COVID-19. In contrast, activation of αβ CD4 + and CD8 + T cells, T follicular helper cells and antibody-secreting cells was similar in pregnant and non-pregnant women with COVID-19. Elevated levels of IL-1β, IFN-γ, IL-8, IL-18 and IL-33 were also found in pregnant women in their healthy state, and these cytokine levels remained elevated during acute and convalescent COVID-19. Collectively, our study provides the first comprehensive map of longitudinal immunological responses to SARS-CoV-2 infection in pregnant women, providing insights into patient management and education during COVID-19 pregnancy.

Immune responses in COVID-19 respiratory tract and blood reveal mechanisms of disease severity (preprint)

Although the respiratory tract is the primary site of SARS-CoV-2 infection and the ensuing immunopathology, respiratory immune responses are understudied and urgently needed to understand mechanisms underlying COVID-19 disease pathogenesis. We collected paired longitudinal blood and respiratory tract samples (endotracheal aspirate, sputum or pleural fluid) from hospitalized COVID-19 patients and non-COVID-19 controls. Cellular, humoral and cytokine responses were analysed and correlated with clinical data. SARS-CoV-2-specific IgM, IgG and IgA antibodies were detected using ELISA and multiplex assay in both the respiratory tract and blood of COVID-19 patients, although a higher receptor binding domain (RBD)-specific IgM and IgG seroconversion level was found in respiratory specimens. SARS-CoV-2 neutralization activity in respiratory samples was detected only when high levels of RBD-specific antibodies were present. Strikingly, cytokine/chemokine levels and profiles greatly differed between respiratory samples and plasma, indicating that inflammation needs to be assessed in respiratory specimens for the accurate assessment of SARS-CoV-2 immunopathology. Diverse immune cell subsets were detected in respiratory samples, albeit dominated by neutrophils. Importantly, we also showed that dexamethasone and/or remdesivir treatment did not affect humoral responses in blood of COVID-19 patients. Overall, our study unveils stark differences in innate and adaptive immune responses between respiratory samples and blood and provides important insights into effect of drug therapy on immune responses in COVID-19 patients.

Immune responses in COVID-19 respiratory tract and blood reveal mechanisms of disease severity (preprint)

Although the respiratory tract is the primary site of SARS-CoV-2 infection and the ensuing immunopathology, respiratory immune responses are understudied and urgently needed to understand mechanisms underlying COVID-19 disease pathogenesis. We collected paired longitudinal blood and respiratory tract samples (endotracheal aspirate, sputum or pleural fluid) from hospitalized COVID-19 patients and non-COVID-19 controls. Cellular, humoral and cytokine responses were analysed and correlated with clinical data. SARS-CoV-2-specific IgM, IgG and IgA antibodies were detected using ELISA and multiplex assay in both the respiratory tract and blood of COVID-19 patients, although a higher receptor binding domain (RBD)-specific IgM and IgG seroconversion level was found in respiratory specimens. SARS-CoV-2 neutralization activity in respiratory samples was detected only when high levels of RBD-specific antibodies were present. Strikingly, cytokine/chemokine levels and profiles greatly differed between respiratory samples and plasma, indicating that inflammation needs to be assessed in respiratory specimens for the accurate assessment of SARS-CoV-2 immunopathology. Diverse immune cell subsets were detected in respiratory samples, albeit dominated by neutrophils. Importantly, we also showed that dexamethasone and/or remdesivir treatment did not affect humoral responses in blood of COVID-19 patients. Overall, our study unveils stark differences in innate and adaptive immune responses between respiratory samples and blood and provides important insights into effect of drug therapy on immune responses in COVID-19 patients.

Integrated immune networks in SARS-CoV-2 infected pregnant women reveal differential NK cell and unconventional T cell activation (preprint)

Although pregnancy poses a greater risk for severe COVID-19, the underlying immunological changes associated with SARS-CoV-2 infection during pregnancy are poorly understood. We defined immune responses to SARS-CoV-2 in pregnant and non-pregnant women during acute and convalescent COVID-19 up to 258 days post symptom onset, quantifying 217 immunological parameters. Additionally, matched maternal and cord blood were collected from COVID-19 convalescent pregnancies. Although serological responses to SARS-CoV-2 were similar in pregnant and non-pregnant women, cellular immune analyses revealed marked differences in key NK cell and unconventional T cell responses during COVID-19 in pregnant women. While NK cells, {gamma}{delta} T cells and MAIT cells displayed pre-activated phenotypes in healthy pregnant women when compared to non-pregnant age-matched women, activation profiles of these pre-activated NK and unconventional T cells remained unchanged at acute and convalescent COVID-19 in pregnancy. Conversely, activation dynamics of NK and unconventional T cells were prototypical in non-pregnant women in COVID-19. In contrast, activation of {beta} CD4+ and CD8+ T cells, T follicular helper cells and antibody-secreting cells was similar in pregnant and non-pregnant women with COVID-19. Elevated levels of IL-1{beta}, IFN-{gamma}, IL-8, IL-18 and IL-33 were also found in pregnant women in their healthy state, and these cytokine levels remained elevated during acute and convalescent COVID-19. Collectively, our study provides the first comprehensive map of longitudinal immunological responses to SARS-CoV-2 infection in pregnant women, providing insights into patient management and education during COVID-19 pregnancy.