Hematoxylin and eosin tissue stain in Mohs micrographic surgery: a review.

BACKGROUND: Several factors may affect Mohs micrographic surgery (MMS) tissue section quality. Although other factors may affect section integrity and ease of processing, tissue stains are the cornerstone of histologic diagnosis. When performed incorrectly, visualization and discrimination of microscopic details may be suboptimal and even impossible. Patient harm may result if an accurate diagnosis cannot be made. OBJECTIVE: To review hematoxylin and eosin (H&E) technique, the most commonly used staining technique in MMS tissue preparation. MATERIALS AND METHODS: Review of literature. Tissue sections were H&E stained using a linear stainer where one reagent container was omitted on each trial run to evaluate microscopic differences in staining quality. RESULTS: The chemical nature of H&E staining is discussed, along with commonly encountered problems, pitfalls, and troubleshooting tips. H&E stained sections are presented to demonstrate histologic appearance in the absence of certain reagents. CONCLUSIONS: The success of MMS depends on high-quality tissue sections. The staining process should be optimized to be reproducible and reliable. To readily identify and resolve poor staining quality, a firm understanding of the principles upon which tissue staining is based and its pitfalls is necessary.

Management of traumatic facial nerve injuries.

Management of facial nerve injuries requires knowledge and skills that should be in every facial plastic surgeon's armamentarium. This article will briefly review the anatomy of the facial nerve, discuss the assessment of facial nerve injury, and describe the management of facial nerve injury after soft tissue trauma.

Downregulation of Friend leukemia virus integration 1 as a feedback mechanism that restrains lipopolysaccharide induction of matrix metalloproteases and interleukin-10 in human macrophages.

The E26 transformation-specific (Ets) proteins are a family of transcription factors with important roles in a variety of cellular processes ranging from proliferation and differentiation to transformation and metastasis. Tissue-specific expression of Ets proteins and their ability to interact with other families of transcription factors contribute to their versatility. In this study, we investigated the regulation of Ets factors in primary human monocytes and macrophages, and their role in matrix metalloprotease (MMP) and cytokine production. The macrophage-activating Toll-like receptor ligand, lipopolysaccharide (LPS), induced the expression of Ets family members epithelium-specific Ets factor 3 (ESE-3) and TEL-2 but rapidly suppressed Friend leukemia virus integration 1 (FLI-1) expression. Modulation of FLI-1 expression using either RNA interference or forced expression identified a positive role for FLI-1 in contributing to LPS-induced expression of MMP-1, MMP-3, MMP-10, and interleukin-10 (IL-10). Thus, the rapid downregulation of FLI-1 expression after LPS stimulation attenuates the induction of various MMPs and IL-10 under inflammatory conditions. In contrast, the expression of IL-6 and TNFα and the effects of interferon (IFN)γ on LPS responses were not dependent on FLI-1. Our results define a novel FLI-1-mediated self-regulatory feedback loop that limits MMP expression and thus may attenuate extent of tissue destruction associated with inflammatory responses.

Treatment of benign lymphoid hyperplasia of the orbit with rituximab.

Benign lymphoid hyperplasia is a disorder characterized by polyclonal lymphocytic infiltration of orbital tissues, predominantly with B-cells. Rituximab is a monoclonal antibody directed against CD20, a B-cell marker. Two patients with recurrent orbital masses involving the lacrimal glands were treated with rituximab. The diagnosis of benign lymphoid hyperplasia with predominance of CD20 cells was confirmed in both cases based on a surgical biopsy. Both patients had been previously treated with standard therapies, including high-dose steroids, and one patient had failed external-beam radiation therapy. They both responded well to treatment with intravenous rituximab. Neither patient experienced any side effects associated with rituximab.

Lipopolysaccharide-induced expression of matrix metalloproteinases in human monocytes is suppressed by IFN-gamma via superinduction of ATF-3 and suppression of AP-1.

Matrix metalloproteinases (MMPs) are induced during inflammatory responses and are important for immune regulation, angiogenesis, wound healing, and tissue remodeling. Expression of MMPs needs to be tightly controlled to avoid excessive tissue damage. In this study, we investigated the regulation of MMP expression by inflammatory factors in primary human monocytes and macrophages. IFN-gamma, which augments inflammatory cytokine production in response to macrophage-activating factors such as TLR ligands, instead broadly suppressed TLR-induced MMP expression. Inhibition of MMP expression was dependent on STAT1 and required de novo protein synthesis. IFN-gamma strongly enhanced TLR-induced expression of the transcriptional repressor activating transcription factor (ATF-3) in a STAT1-dependent manner, which correlated with recruitment of ATF-3 to the endogenous MMP-1 promoter as detected by chromatin immunoprecipitation assays. RNA interference experiments further supported a role for ATF-3 in suppression of MMP-1 expression. In addition, IFN-gamma suppressed DNA binding by AP-1 transcription factors that are known to promote MMP expression and a combination of supershift, RNA interference and overexpression experiments implicated AP-1 family member Fra-1 in the regulation of MMP-1 expression. These results define an IFN-gamma-mediated homeostatic loop that limits the potential for tissue damage associated with inflammation, and identify transcriptional factors that regulate MMP expression in myeloid cells in inflammatory settings.

Role of STAT3 in type I interferon responses. Negative regulation of STAT1-dependent inflammatory gene activation.

Type I interferons (IFNalpha/beta) induce antiviral responses and have immunomodulatory effects that can either promote or suppress immunity and inflammation. In myeloid cells IFNalpha/beta activates signal transducers and activators of transcription STAT1, STAT2, and STAT3. STAT1 and STAT2 mediate the antiviral and inflammatory effects of IFNalpha/beta, but the function of IFNalpha/beta-activated STAT3 is not known. We investigated the role of STAT3 in type I IFN signaling in myeloid cells by modulating STAT3 expression and the intensity of STAT3 activation using overexpression and RNA interference and determining the effects on downstream signaling and gene expression. IFNalpha-activated STAT3 inhibited STAT1-dependent gene activation, thereby down-regulating IFNalpha-mediated induction of inflammatory mediators such as the chemokines CXCL9 (Mig) and CXCL10 (IP-10). At the same time, IFNalpha-activated STAT3 supported ISGF-3-dependent induction of antiviral genes. STAT3 did not suppress STAT1 tyrosine phosphorylation or nuclear translocation but instead sequestered STAT1 and suppressed the formation of DNA-binding STAT1 homodimers. These results identify a regulatory function for STAT3 in attenuating the inflammatory properties of type I IFNs and provide a mechanism of suppression of STAT1 function that differs from previously described suppression of tyrosine phosphorylation. The results suggest that changes in the relative expression and activation of STAT1 and STAT3 that occur during immune responses determine the nature of cellular responses to type I IFNs.

IFN-gamma-primed macrophages exhibit increased CCR2-dependent migration and altered IFN-gamma responses mediated by Stat1.

Priming of macrophages with IFN-gamma increases cellular responsiveness to inflammatory stimuli, including IFN-gamma itself. We described previously that priming with subactivating concentrations of IFN-gamma increased Stat1 expression and resulted in enhanced activation of Stat1 and of a subset of IFN-gamma-responsive genes when primed macrophages were restimulated with low doses of IFN-gamma. In this study, we determined the effects of IFN-gamma priming on the macrophage transcriptome and on transcriptional responses to high saturating concentrations of IFN-gamma. At baseline, primed macrophages expressed a small subset of IFN-gamma-inducible genes, including CCR2, and exhibited increased migration in response to CCL2. Activation of gene expression by high concentrations of IFN-gamma was altered in primed macrophages, such that activation of a subset of IFN-gamma-inducible genes was attenuated. A majority of genes in this "less induced" category corresponded to genes that are induced by IFN-gamma via Stat1-independent but Stat3-dependent pathways and have been implicated in inflammatory tissue destruction. One mechanism of attenuation of gene expression was down-regulation of Stat3 function by increased levels of Stat1. These results reveal that priming enhances migration to inflammatory chemokines and identify IFN-gamma-inducible genes whose expression is attenuated by high levels of Stat1. The increase in Stat1 expression during priming provides a mechanism by which physiological regulation of the relative abundance of Stat1 and Stat3 impacts on gene expression. Our results also suggest that, in addition to inducing hypersensitivity to inflammatory stimuli, IFN priming delivers a homeostatic signal by attenuating IFN-gamma induction of certain tissue-destructive genes.

Homeostatic role of interferons conferred by inhibition of IL-1-mediated inflammation and tissue destruction.

In addition to their well known immune and proinflammatory activities, IFNs possess homeostatic functions that limit inflammation and tissue destruction in a variety of conditions such as arthritis, osteolysis, and multiple sclerosis. The mechanisms underlying the homeostatic actions of IFNs are not well understood. We report here that both type I and type II IFNs (IFN-alpha, IFN-beta, and IFN-gamma, respectively) suppressed a broad range of proinflammatory and tissue-destructive activities of IL-1, including induction of inflammatory mediators, production of matrix metalloproteinases, macrophage tissue invasion, and cartilage degradation. IFN-alpha attenuated IL-1-mediated cell recruitment in vivo. IFNs completely suppressed the activation of IL-1 signal transduction pathways in macrophages. The mechanism of IFN-mediated inhibition of IL-1 action and signaling was modulation of IL-1R expression, which was also observed in vivo. IFN-gamma-mediated down-regulation of IL-1R type I expression was dependent on Stat1, a transcription factor typically considered to be a key mediator of macrophage activation by IFNs. These results identify cellular and molecular mechanisms that contribute to the homeostatic role of IFNs in limiting inflammation and associated tissue destruction.