Lucille London

Respiratory Reovirus 1/L Induction of Intraluminal Fibrosis, a Model of Bronchiolitis Obliterans Organizing Pneumonia, Is Dependent on T Lymphocytes

Bronchiolitis obliterans organizing pneumonia (BOOP) is a clinical syndrome characterized by perivascular/peribronchiolar leukocyte infiltration leading to the development of intraalveolar fibrosis. We have developed an animal model of BOOP where CBA/J mice infected with 1 x 10(6) plaque-forming units (PFU) reovirus 1/L develop follicular bronchiolitis and intraalveolar fibrosis similar to human BOOP. In this report, we demonstrate a role for T cells in the development of intraluminal fibrosis associated with BOOP. Corticosteroid treatment of reovirus 1/L-infected mice both inhibited the development of fibrotic lesions when administered early in the time-course and promoted the resolution of fibrotic lesions when corticosteroid administration was delayed. Further, the depletion of either CD4(+) or CD8(+) T cells before reovirus 1/L infection also inhibited fibrotic lesion development. Both corticosteroid treatment and depletion of CD4(+) or CD8(+) T cells also resulted in decreased expression of the proinflammatory and profibrotic cytokines, interferon (IFN)-gamma and monocyte chemoattractant protein-1 (MCP-1). Further, treatment of mice with a neutralizing monoclonal antibody to IFN-gamma also significantly inhibited the development of fibrosis. Taken together, these results suggest a significant role for T cells in the development of reovirus 1/L-induced BOOP fibrotic lesions in CBA/J mice and suggests that T(H)1-derived cytokines, especially IFN-gamma, may play a key role in fibrotic lesion development.

Polychlorinated biphenyl mixtures (Aroclors) inhibit LPS-induced murine splenocyte proliferation in vitro

The immune system is believed to be a sensitive indicator for adverse polychlorinated biphenyl (PCB)-induced health effects. Four commercial PCB mixtures (Aroclors) or six individual PCB congeners were evaluated for their effect on splenocyte viability and lipopolysaccharide (LPS)-induced splenocyte proliferation in vitro in two strains of mice, C57B1/6 (high affinity aromatic hydrocarbon receptor (AhR) complex) and DBA/J (low affinity AhR complex). All four Aroclors, the selected individual noncoplanar congeners, or two tertiary mixtures containing one congener from each class significantly decreased the in vitro LPS-induced proliferation of murine splenocytes in either strain of mice without inducing a significant decrease in viability. In contrast, selected individual coplanar or mono-ortho-coplanar congeners did not inhibit splenocyte proliferation or viability at any concentration. These results suggest that mixtures of PCBs and/or congener class (specifically, noncoplanar congeners) may be more highly immunotoxic than individual planar and mono-ortho-coplanar congeners alone. Thus, this in vitro assay has revealed a more complex pattern of immunotoxicity of Aroclors versus individual congeners than has previously been reported or anticipated based on both in vivo derived immunotoxic data and standard comparisons to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). These results have important practical significance since mixtures of PCB congeners were used industrially and now contaminate the environment.

Inhibition of LPS-induced splenocyte proliferation by Ortho-substituted polychlorinated biphenyl congeners

Polychlorinated biphenyls (PCBs) are persistent environmental contaminants, and their ubiquitous nature has prompted studies of their potential health hazards. As a result of their lipophilic nature, PCBs accumulate in breast milk and subsequently affect the health of offspring of exposed individuals. Biological effects of PCBs in animals have mostly been attributed to coplanar congeners, although effects of ortho congeners also have been demonstrated. To investigate the relationship of immunotoxicity and chlorine substitution pattern, the effects of PCB congeners and mixtures of ortho and non-ortho-substituted constituents of Aroclor 1242 on splenocytes from C57B1/6 mice were examined. The immunotoxic endpoints investigated included splenocyte viability, lipopolysaccharide (LPS)-induced splenocyte proliferation, and LPS-induced antibody secretion. Congeners with multiple ortho chlorines preferentially inhibited splenocyte proliferation as compared with non- or mono-ortho-substituted congeners. However, mixtures of non- and mono-ortho-substituted congeners and multi-ortho-substituted congeners inhibited LPS-induced splenocyte proliferation and antibody secretion at similar concentrations. Exposure of splenocytes to these mixtures did not activate the aryl hydrocarbon receptor (AhR) signal transduction pathway. These results suggest individual multi-ortho-substituted congeners preferentially inhibit LPS-induced splenocyte proliferation, while congeners not exhibiting an effect individually may have additive effects in a mixture to produce an immunotoxic response through an AhR-independent pathway.

Optimal induction of upper respiratory tract immunity to reovirus 1/L by combined upper and lower respiratory tract inoculation

There has been an increasing interest in developing vaccines which are both easy to administer and which elicit functionally protective immune responses at mucosal and/or systemic sites. Intranasally administered vaccines meet the criteria of ease of administration and are thought to stimulate respiratory-mucosal immunity via interaction with nasal associated lymphoid tissues (NALT). The aim of this study was to gain a better understanding of how best to stimulate respiratory-mucosal immunity using a murine model of respiratory reovirus infection. Either a predominantly upper respiratory tract infection or a combination upper and lower respiratory tract infection was established by administering the same virus dose in either a small or large inoculum volume. These studies demonstrate that stimulation of NALT alone by an upper respiratory tract infection does not induce an optimal primary antibody response even in the nasal cavity. Effective immunity of both the upper and lower respiratory tract was obtained when a combination upper and lower respiratory tract infection was established. These results have important clinical implications since they suggest that effective respiratory mucosal immunity will be best achieved by the combined stimulation of both the upper and lower respiratory tract and will likely require both intranasal as well as inhaled aerosol delivery of antigen to the lower respiratory tract in humans.

Reovirus Serotype 1/Strain Lang-Stimulated Activation of Antigen-Specific T Lymphocytes in Peyer’s Patches and Distal Gut-Mucosal Sites: Activation Status and Cytotoxic Mechanisms

Intraduodenal priming of mice with reovirus serotype 1/strain Lang (reovirus 1/L) stimulates gut lymphocytes and generates precursor and effector CTLs. Our earlier studies demonstrated that germinal center and T cell Ag (GCT) is a marker which identifies reovirus 1/L-specific precursor CTL and effector CTL in Peyer’s patches (PP) of reovirus 1/L-inoculated mice. In this study, we characterized the expression of the activation markers, GCT and CD11c, on reovirus 1/L-stimulated gut lymphocytes and the effector mechanisms involved in reovirus 1/L-specific cytotoxicity. We found that intraduodenal reovirus 1/L inoculation of mice induced the expression of both GCT and CD11c on PP lymphocytes (PPL), intraepithelial lymphocytes (IEL), and lamina propria lymphocytes (LPL), and these activated cells expressed Fas ligand (FasL). The majority of the GCT+CD11c+ IEL and LPL expressed a phenotype, TCRαβ+Thy-1+CD8+ similar to that expressed on reovirus 1/L-stimulated PPL. However, splenic lymphocytes expressed GCT but not CD11c after stimulation with reovirus 1/L. Perforin, Fas-FasL, and TRAIL pathways were found to be involved in PPL, IEL, and LPL cytotoxic activity against reovirus 1/L-infected targets. In PPL, perforin and Fas-FasL pathways were more effective than TRAIL. In IEL, all three cytotoxic mechanisms were equally as effective. However, LPL prefer Fas-FasL and TRAIL over perforin. Further, we demonstrated the preferential migration of GCT+ PPL to the intraepithelial compartment and the lamina propria. These results suggest that GCT and CD11c can be used as activation markers for gut lymphocytes and CD11c can also be used to differentiate between activated gut and systemic lymphocytes.

Erratum: Correction: Curcumin Modulates the Inflammatory Response and Inhibits Subsequent Fibrosis in a Mouse Model of Viral-Induced Acute Respiratory Distress Syndrome (PLoS ONE (2015) 10:8 (e0134982))

In Fig 4A, as the result of an error in preparation of the Western blot images, incorrect bands were shown for ß-actin, saline-treated α-SMA and saline-treated Tenascin. Bands labelled ßactin were taken from the α-SMA blot by mistake; the band labelled saline-treated α-SMA was taken from a different lane showing curcumin-treated lysate in the same blot; the band labelled saline-treated Tenascin was from a different blot. Additionally, α-SMA, E-cadherin and ß-actin expression were measured on a single blot that was stripped and re-probed multiple times, while Tenascin (TN-C) was probed on a different blot in an independent experiment with ß-actin antibodies used concurrently. The ß-actin bands from the independent Tenascin blot were not shown in the original Fig 4A. Here we provide a revised Fig 4 showing the correct ß-actin and α-SMA bands, and a revised Fig 4 legend. The original uncropped blot images are provided as a supplementary file, with arrows indicating the correct sized bands. Multiple exposures are provided for the E-cadherin blot. The revised figure has been prepared using lanes 1, 6, and 7 of the original E-cadherin blot to match the lanes used from the ß-actin and α-SMA blots; the original published figure used lanes 1, 4, and 5 (lanes 4 and 6 both used lysates from untreated mice on day 14; lanes 5 and 7 both used lysates from curcumin-treated mice on day 14). The errors in the original figure do not affect the results demonstrated in Fig 4A, which are further confirmed by the additional methods shown in Fig 4B and 4C.