Ventilated Patients With COVID-19 Show Airflow Obstruction.

OBJECTIVE: Many patients with coronavirus disease 2019 (COVID-19) need mechanical ventilation secondary to acute respiratory distress syndrome. Information on the respiratory system mechanical characteristics of this disease is limited. The aim of this study is to describe the respiratory system mechanical properties of ventilated COVID-19 patients. DESIGN, SETTING, AND PATIENTS: Patients consecutively admitted to the medical intensive care unit at the University of Iowa Hospitals and Clinics in Iowa City, USA, from April 19 to May 1, 2020, were prospectively studied; final date of follow-up was May 1, 2020. MEASUREMENTS: At the time of first patient contact, ventilator information was collected including mode, settings, peak airway pressure, plateau pressure, and total positive end expiratory pressure. Indices of airflow resistance and respiratory system compliance were calculated and analyzed. MAIN RESULTS: The mean age of the patients was 58 years. 6 out of 12 (50%) patients were female. Of the 21 laboratory-confirmed COVID-19 patients on invasive mechanical ventilation, 9 patients who were actively breathing on the ventilator were excluded. All the patients included were on volume-control mode. Mean [±standard deviation] ventilator indices were: resistive pressure 19 [±4] cmH2O, airway resistance 20 [±4] cmH2O/L/s, and respiratory system static compliance 39 [±16] ml/cmH2O. These values are consistent with abnormally elevated resistance to airflow and reduced respiratory system compliance. Analysis of flow waveform graphics revealed a pattern consistent with airflow obstruction in all patients. CONCLUSIONS: Severe respiratory failure due to COVID-19 is regularly associated with airflow obstruction.

B cell depletion in murine lupus using cytotoxic T lymphocytes in vivo: Feasibility and benefit.

Given the promising results in human lupus with B cell depletion, we tested whether in vivo cytotoxic T lymphocyte (CTL) could eliminate autoreactive B cells in the setting of murine lupus. Using the parent-into-F1 (P â†’ F1) model to generate CTL that eliminate B cells, we found that transfer ofNZB parental splenocytes into lupus-prone female NZB/W F1 mice resulted in profound B cell reduction whereas NZW â†’ F1 mice exhibited defective B cell elimination. Using pre-disease or early disease B/W mice as hosts, NZB â†’ F1 mice exhibited B cell depletion and improved proteinuria but no improvement in survival whereas NZW â†’ F1 mice had significantly reduced proteinuria and prolonged survival. Thus, despite the defective IL-2 environment in B/W F1 mice, generation of CTL and B cell depletion is feasible in NZB â†’ F1 mice. The surprising increase in survival for NZW â†’ F1 mice despite defective B cell elimination suggests that NZW splenocytes may contain a beneficial down regulatory cell.

Both perforin and FasL are required for optimal CD8 T cell control of autoreactive B cells and autoantibody production in parent-into-F1 lupus mice.

To test the relative roles of perforin (pfp) vs. FasL in CTL control of autoreactive B cell expansion, we used the parent-into-F1 model of murine graft-vs.-host disease in which donor CD8 CTL prevent lupus like disease by eliminating activated autoreactive B cells. F1 mice receiving either pfp or FasL defective donor T cells exhibited an intermediate short-term phenotype. Pairing of purified normal CD4 T cells with either pfp or FasL defective CD8 T cell subsets resulted in impaired host B cell elimination and mild lupus like disease that was roughly equivalent in the two experimental groups. Thus, in addition to major roles in tumor and intracellular pathogen control, pfp mediated CD8 CTL killing plays a significant role in controlling autoreactive B cell expansion and lupus downregulation that is comparable to that mediated by FasL killing. Importantly, both pathways are required for optimal elimination of activated autoreactive B cells.

In vivo IL-4 prevents allo-antigen driven CD8+ CTL development.

IL-4 has been shown to suppress acute graft vs. host disease (GVHD) in irradiated hosts. Here we evaluated whether IL-4 suppresses acute GVHD in the un-irradiated parent-into-F1 GVHD model with relevance to renal allograft rejection. IL-4 completely suppressed CD8 CTL when administered with donor cells however this effect was lost if its administration was delayed 3days. IL-4 did not inhibit donor CD8+ T cell homing to the host spleen but rather prevented donor CD8+ T cell differentiation into CTLs. Studies with IL-4Rα-deficient donor cells or recipient mice demonstrated that IL-4 effects on the host, rather than, or in addition to IL-4 effects on donor cells, were critical for suppression of CTL. Because IL-4 decreased all splenic dendritic cell populations and increased neutrophil and CD8+ T cells, IL-4 may suppress donor CD8+ CTL by decreasing Ag presentation and/or increasing host myeloid and CD8+ T cell suppression of donor T cells.

Intrinsic Differences in Donor CD4 T Cell IL-2 Production Influence Severity of Parent-into-F1 Murine Lupus by Skewing the Immune Response Either toward Help for B Cells and a Sustained Autoantibody Response or toward Help for CD8 T Cells and a Downregulatory Th1 Response.

Using the parent-into-F1 model of induced lupus and (C57BL/6 × DBA2) F1 mice as hosts, we compared the inherent lupus-inducing properties of the two parental strain CD4 T cells. To control for donor CD4 recognition of alloantigen, we used H-2(d) identical DBA/2 and B10.D2 donor T cells. We demonstrate that these two normal, nonlupus-prone parental strains exhibit two different T cell activation pathways in vivo. B10.D2 CD4 T cells induce a strong Th1/CMI pathway that is characterized by IL-2/IFN-γ expression, help for CD8 CTLs, and skewing of dendritic cell (DC) subsets toward CD8a DCs, coupled with reduced CD4 T follicular helper cells and transient B cell help. In contrast, DBA/2 CD4 T cells exhibit a reciprocal, lupus-inducing pathway that is characterized by poor IL-2/IFN-γ expression, poor help for CD8 CTLs, and skewing of DC subsets toward plasmacytoid DCs, coupled with greater CD4 T follicular helper cells, prolonged B cell activation, autoantibody formation, and lupus-like renal disease. Additionally, two distinct in vivo splenic gene-expression signatures were induced. In vitro analysis of TCR signaling revealed defective DBA CD4 T cell induction of NF-κB, reduced degradation of IκBα, and increased expression of the NF-κB regulator A20. Thus, attenuated NF-κB signaling may lead to diminished IL-2 production by DBA CD4 T cells. These results indicate that intrinsic differences in donor CD4 IL-2 production and subsequent immune skewing could contribute to lupus susceptibility in humans. Therapeutic efforts to skew immune function away from excessive help for B cells and toward help for CTLs may be beneficial.

In vivo maturation of allo-specific CD8 CTL and prevention of lupus-like graft-versus-host disease is critically dependent on T cell signaling through the TNF p75 receptor but not the TNF p55 receptor.

A third signal is required for maturation of effector CD8 CTL in addition to TCR and CD28 engagement. Inflammatory cytokines can provide a third signal; however, in nonpathogen settings (i.e., antitumor responses), the identity of the third signal is not clear. A useful model for in vivo CD8 CTL in the absence of exogenous pathogens is the alloantigen-driven parent-into F1 model of acute graft-versus-host disease (GVHD) characterized by a strong TNF-dependent donor antihost CD8 CTL T cell response. To determine whether TNF acts directly on donor T cells in a signal 3 manner, F1 mice received TNFR 1 (p55) knockout (KO) and/or TNFR 2 (p75) KO donor T cells. Donor p75 KO but not p55KO donor T cells failed to induce acute GVHD phenotype and instead induced a lupus-like chronic GVHD both short and long term because of quantitative and qualitative donor T cell defects, that is, reduced perforin, IFN-γ, and TNF production. Transfer of mixed or matched purified CD4 and CD8 T cells from wild type or p75KO donors demonstrated that optimal CTL maturation required p75 signaling in both CD4 and CD8 T cells. Despite defective p75KO CD4 help for CD8 CTL, p75KO CD4 help for B cells and autoimmunity was intact. These results provide a mechanism by which impaired CD8 CTL could contribute to reduced antiviral and antitumor responses and autoimmunity reported in patients receiving TNF blockers. Our results support the idea that selective p55 blockade may be beneficial by reducing inflammation without compromising CD8 CTL.

The parent-into-F1 murine model in the study of lupus-like autoimmunity and CD8 cytotoxic T lymphocyte function.

The transfer of homozygous C57Bl/6 (B6) or DBA/2 (DBA) parental strain T cells into normal B6D2F1 mice in the parent-into-F1 (p → F1) model results in a graft-vs.-host disease (GVHD) that takes one of the following two forms: (a) acute GVHD seen with B6 → F1 mice and mediated by donor CD8 cytotoxic T cells that eliminate host lymphocytes and (b) a chronic lupus-like GVHD seen with DBA → F1 mice and mediated by donor CD4 T cell cognate help to autoreactive B cells resulting in autoantibody production and renal disease similar to human lupus. Importantly, these two phenotypes can be distinguished by flow cytometry as early as 2 weeks after donor cell transfer. The p → F1 model can be used to screen for agents that alter lupus development. Additionally, the model is useful for preclinical screening of biologic agents with immunomodulatory potential. Agents that selectively inhibit CD8 T cell function will convert acute GVHD to chronic GVHD in B6 → F1 mice. Conversely, agents that promote CD8 CTL function will convert chronic GVHD to acute GVHD in DBA → F1 mice. Agents that completely suppress T cell function will block both phenotypes. The model is also useful for examining the effects of T cell mutations by transferring mutant T cells into wild-type hosts and assessing the effects on disease phenotype. Differences observed from wild-type T cells → F1 can be directly ascribed to alterations in mutant T cell function. Because of the early 2-week phenotype development, the p → F1 model is well suited to screening of potential immunomodulatory therapeutic compounds and the assessment of T cell mutations on in vivo function.

Donor CD8 T cells and IFN-gamma are critical for sex-based differences in donor CD4 T cell engraftment and lupus-like phenotype in short-term chronic graft-versus-host disease mice.

The transfer of unfractionated DBA/2J (DBA) splenocytes into B6D2F(1) (DBA → F(1)) mice results in greater donor CD4 T cell engraftment in females at day 14 that persists long-term and mediates greater female lupus-like renal disease. Although donor CD8 T cells have no demonstrated role in lupus pathogenesis in this model, we recently observed that depletion of donor CD8 T cells prior to transfer eliminates sex-based differences in renal disease long-term. In this study, we demonstrate that greater day 14 female donor CD4 engraftment is also critically dependent on donor CD8 T cells. Male DBA → F(1) mice exhibit stronger CD8-dependent day 8-10 graft-versus-host (GVH) and counter-regulatory host-versus-graft (HVG) responses, followed by stronger homeostatic contraction (days 10-12). The weaker day 10-12 GVH and HVG in females are followed by persistent donor T cell activation and increasing proliferation, expansion, and cytokine production from days 12 to 14. Lastly, greater female day 14 donor T cell engraftment, activation, and cytokine production were lost with in vivo IFN-γ neutralization from days 6 to 14. We conclude the following: 1) donor CD8 T cells enhance day 10 proliferation of donor CD4 T cells in both sexes; and 2) a weaker GVH/HVG in females allows prolonged survival of donor CD4 and CD8 T cells, allowing persistent activation. These results support the novel conclusion that sex-based differences in suboptimal donor CD8 CTL activation are critical for shaping sex-based differences in donor CD4 T cell engraftment at 2 wk and lupus-like disease long-term.

Enhancement of suboptimal CD8 cytotoxic T cell effector function in vivo using antigen-specific CD80 defective T cells.

T cell upregulation of B7 molecules CD80 and CD86 limits T cell expansion in immunodeficient hosts; however, the relative roles of CD80 separate from CD86 on CD4 versus CD8 T cells in a normal immune system are not clear. To address this question, we used the parent-into-F1 (P→F1) murine model of graft-versus-host disease and transferred optimal and suboptimal doses of CD80 and/or CD86 knockout (KO) T cells into normal F1 hosts. Enhanced elimination of host B cells by KO T cells was observed only at suboptimal donor cell doses and was greatest for CD80 KO→F1 mice. Wild-type donor cells exhibited peak CD80 upregulation at day 10; CD80 KO donor cells exhibited greater peak (day 10) donor T cell proliferation and CD8 T cell effector CTL numbers versus wild-type→F1 mice. Fas or programmed cell death-1 upregulation was normal as was homeostatic contraction of CD80 KO donor cells from days 12-14. Mixing studies demonstrated that maximal host cell elimination was seen when both CD4 and CD8 T cells were CD80 deficient. These results indicate an important role for CD80 upregulation on Ag-activated CD4 and CD8 T cells in limiting expansion of CD8 CTL effectors as part of a normal immune response. Our results support further studies of therapeutic targeting of CD80 in conditions characterized by suboptimal CD8 effector responses.