Murine Epidermal Ceramide Synthase 4 Is a Key Regulator of Skin Barrier Homeostasis.

Epidermal barrier dysfunction is associated with a wide range of highly prevalent inflammatory skin diseases. However, the molecular processes that drive epidermal barrier maintenance are still largely unknown. Here, using quantitative proteomics, lipidomics, and mouse genetics, we characterize epidermal barrier maintenance versus a newly established barrier and functionally identify differential ceramide synthase 4 protein expression as one key difference. We show that epidermal loss of ceramide synthase 4 first disturbs epidermal lipid metabolism and adult epidermal barrier function, ultimately resulting in chronic skin barrier disease characterized by acanthosis, hyperkeratosis, and immune cell accumulation. Importantly, prolonged barrier dysfunction induced by loss of ceramide synthase 4 induced a barrier repair response that largely recapitulates molecular programs of barrier establishment. Collectively, this study provides an unbiased temporal proteomic characterization of barrier maintenance and disturbed homeostasis and shows that lipid homeostasis is essential to maintain adult skin barrier function to prevent disease.

Recent Zika Virus Isolates Induce Premature Differentiation of Neural Progenitors in Human Brain Organoids.

The recent Zika virus (ZIKV) epidemic is associated with microcephaly in newborns. Although the connection between ZIKV and neurodevelopmental defects is widely recognized, the underlying mechanisms are poorly understood. Here we show that two recently isolated strains of ZIKV, an American strain from an infected fetal brain (FB-GWUH-2016) and a closely-related Asian strain (H/PF/2013), productively infect human iPSC-derived brain organoids. Both of these strains readily target to and replicate in proliferating ventricular zone (VZ) apical progenitors. The main phenotypic effect was premature differentiation of neural progenitors associated with centrosome perturbation, even during early stages of infection, leading to progenitor depletion, disruption of the VZ, impaired neurogenesis, and cortical thinning. The infection pattern and cellular outcome differ from those seen with the extensively passaged ZIKV strain MR766. The structural changes we see after infection with these more recently isolated viral strains closely resemble those seen in ZIKV-associated microcephaly.

Endogenous Il10 alleviates the systemic antiviral cellular immune response and T cell-mediated immunopathology in select organs of acutely LCMV-infected mice.

The immunoregulatory cytokine IL-10 suppresses T-cell immunity. The complementary question, whether IL-10 is also involved in limiting the collateral damage of vigorous T cell responses, has not been addressed in detail. Here, we report that the particularly strong virus-specific immune response during acute primary infection with the lymphocytic choriomeningitis virus (LCMV) in mice is significantly further increased in Il10-deficient mice, particularly regarding frequencies and cytotoxic activity of CD8(+) T cells. This increase results in exacerbating immunopathology in select organs, ranging from transient local swelling to an increased risk for mortality. Remarkably, LCMV-induced, T cell-mediated hepatitis is not affected by endogenous Il10. The alleviating effect of Il10 on LCMV-induced immunopathology was found to be operative in delayed-type hypersensitivity footpad-swelling reaction and in debilitating meningitis in mice of both the C57BL/6 and BALB/c strains. These strains are prototypic counterpoles for genetically imprinted type 1-biased versus type 2-biased T cell-mediated immune responses against various infectious pathogens. However, during acute LCMV infection, neither systemic cytokine patterns nor the impact of Il10 on LCMV-induced immunopathology differed conspicuously between these two strains of mice. This study documents a physiological role of Il10 in the regulation of a balanced T-cell response limiting immunopathological damage.

Acid sphingomyelinase is required for protection of effector memory T cells against glucocorticoid-induced cell death.

The activity of acid sphingomyelinase (aSMase) was previously reported to be involved in glucocorticoid-induced cell death (GICD) of T lymphocytes. This mechanism in turn is believed to contribute to the therapeutic efficacy of glucocorticoids (GCs) in the treatment of inflammatory diseases. In this study, we reassessed the role of aSMase in GICD by using aSMase knockout mice. The absence of aSMase largely abolished the partial protection that effector memory CD4(+) T cells in wild-type mice possess against GICD. Reduced IL-2 secretion by aSMase-deficient CD4(+) T cells suggested that a lack of this important survival factor might be the cause of these cells' enhanced susceptibility to GICD. Indeed, addition of IL-2 restored the protection against GICD, whereas neutralization of IL-2 abrogated the otherwise protective effect seen in wild-type effector memory CD4(+) T cells. The therapeutic implications of the altered sensitivity of aSMase-deficient T cells to GICD were assessed in models of inflammatory disorders; namely, experimental autoimmune encephalomyelitis and acute graft-versus-host disease. Surprisingly, GC treatment was equally efficient in both models in terms of ameliorating the diseases, regardless of the genotype of the T cells. Thus, our data reveal a hitherto unrecognized contribution of aSMase to the sensitivity of effector memory CD4(+) T cells to GICD and call into question the traditionally attributed importance of GICD of T cells to the treatment of inflammatory diseases by GCs.

Interleukin-4 impairs granzyme-mediated cytotoxicity of Simian virus 40 large tumor antigen-specific CTL in BALB/c mice.

In this report we analyzed the impact of interleukin-4 (IL-4) on tumor-associated simian virus 40 (SV40) large T-antigen (TAg)-specific CD8+ cytotoxic T cells during rejection of syngeneic SV40 transformed mKSA tumor cells in BALB/c mice. Strikingly, challenge of naïve mice with low doses of mKSA tumor cells revealed a CD8+ T cell-dependent prolonged survival time of naïve IL-4-/- mice. In mice immunized with SV40 TAg we observed in IL-4-/- mice, or in wild type mice treated with neutralizing anti-IL-4 monoclonal antibody, a strongly enhanced TAg-specific cytotoxicity of tumor associated CD8+ T cells. The enhanced cytotoxicity in IL-4-/- mice was accompanied by a significant increase in the fraction of CD8+ tumor associated T-cells expressing the cytotoxic effector molecules granzyme A and B and in granzyme B-specific enzymatic activity. The data suggest that endogenous IL-4 can suppress the generation of CD8+ CTL expressing cytotoxic effector molecules especially when the antigen induces only a very weak CTL response.

Severe impairment in early host defense against Listeria monocytogenes in mice deficient in acid sphingomyelinase.

The phagolysosomal compartment is crucial for the defense against infection with intracellular pathogens. Within this compartment, the TNF- and IFN-gamma-responsive acid sphingomyelinase (ASMase) generates the signaling molecule ceramide, resulting in the activation of proteases like cathepsin D. To investigate the possible role of ASMase as a mediator of the antibacterial effects of TNF and IFN-gamma, ASMase(-/-) mice were infected with Listeria monocytogenes. ASMase(-/-) mice showed a dramatically increased susceptibility to L. monocytogenes (LD(50) approximately 100 CFU) when compared with syngeneic wild-type mice (LD(50) approximately 10,000 CFU). In L. monocytogenes-challenged ASMase(-/-) mice, IFN-gamma serum levels as well as IL-1 beta and IL-6 secretion by macrophages were similar to those observed in wild-type C57BL/6 mice. Although macrophages and granulocytes from ASMase(-/-) mice showed intact production of reactive nitrogen intermediates and oxidative burst, ASMase(-/-) macrophages proved completely incapable of restricting the growth of L. monocytogenes in vitro. The results of this study suggest that ASMase is crucially required for the intracellular control of L. monocytogenes in macrophages and granulocytes by nonoxidative mechanisms.