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Abstract Rationale The mechanisms that underlie the pathogenesis of chronic rhinosinusitis without nasal polyps (CRSsNP), chronic rhinosinusitis with nasal polyps (CRSwNP), and aspirin-exacerbated respiratory disease (AERD) are not clear. Objectives To first evaluate the inflammatory profiles of CRSsNP and CRSwNP tissues and then to investigate whether clinical differences observed between CRSwNP and AERD are in part secondary to differences in inflammatory mediator expression within nasal polyp (NP) tissues. Methods Expression levels of numerous inflammatory mediators were determined by quantitative real-time polymerase chain reaction, ELISA, and multiplex immunoassay. Measurements and Main Results CRSwNP NP had increased levels of type 2 mediators, including IL-5 (P < 0.001), IL-13 (P < 0.001), eotaxin-2 (P  < 0.001), and monocyte chemoattractant protein (MCP)-4 (P < 0.01), compared with sinonasal tissue from subjects with CRSsNP and control subjects. Expression of IFN-γ messenger RNA or protein was low and not different among the chronic rhinosinusitis subtypes examined. Compared with CRSwNP, AERD NP had elevated protein levels of eosinophil cationic protein (ECP) (P < 0.001), granulocyte–macrophage colony–stimulating factor (GM-CSF) (P < 0.01), and MCP-1 (P = 0.01), as well as decreased gene expression of tissue plasminogen activator (tPA) (P = 0.02). Despite the higher eosinophilia in AERD, there was no associated increase in type 2 mediator protein levels observed. Conclusions CRSwNP was characterized by a predominant type 2 inflammatory environment, whereas CRSsNP did not reflect a classic type 1 milieu, as has been suggested previously. AERD can be distinguished from CRSwNP by elevated ECP levels, but this enhanced eosinophilia is not associated with elevations in traditional type 2 inflammatory mediators associated with eosinophil proliferation and recruitment. However, other factors, including GM-CSF, MCP-1, and tPA, may be important contributors to AERD pathogenesis.

Animal model systems are invaluable for examining human diseases. Our laboratory recently established a mouse model of nasal polyps (NPs) and investigated similarities and differences between this mouse model and human NPs. We especially focus on the hypothesis that B cell activation occurs during NP generation in the murine model. After induction of ovalbumin-induced allergic rhinosinusitis, 6% ovalbumin and Staphylococcus aureus enterotoxin B (10 ng) were instilled into the nasal cavity of mice three times per week for 8 weeks. The development of structures that somewhat resemble NPs (which we will refer to as NPs) was confirmed by hematoxylin and eosin staining. The mRNA and protein levels of various inflammatory cell markers and mediators were measured by real-time PCR in nasal tissue and by ELISA in nasal lavage fluid (NLF), respectively. Total Ig isotype levels in NLF were also quantitated using the Mouse Ig Isotyping Multiplex kit (EMD Millipore, Billerica, MA) on a Luminex 200 instrument (Life Technologies, Grand Island, NY). Similar to human NPs, there were significant increases in gene expression of inflammatory cell markers, such as CD19, CD138, CD11c, and mast cell protease-6 in nasal tissue samples of the NP group compared with those of the control group. In further investigations of B cell activation, mRNA expressions of B cell activating factor and a proliferation-inducing ligand were found to be significantly increased in mouse NP tissue. B cell–activating factor protein concentration and IgA and IgG1 levels in NLF were significantly higher in the NP group compared with the control group. In this study, the NP mouse model demonstrated enhanced B cell responses, which are reminiscent of B cell responses in human NPs.

Chronic rhinosinusitis (CRS) is a heterogeneous chronic inflammatory disease of the nose and paranasal sinuses that presents without or with nasal polyps (CRSwNP). Notable features of CRSwNP are the frequent presence of type 2 allergic inflammation and high prevalence of Staphylococcus aureus (SA) colonization. As inflammation persists, sinus tissue undergoes epithelial damage and repair along with polyp growth, despite active medical management. Because one feature of damaged tissue is enhancement of growth factor signaling, we evaluated the presence of epidermal growth factor receptor (EGFR) ligands and matrix metalloproteinases (MMPs) in CRS. The objectives of this study were to analyze the expression of EGFR ligands and MMPs in patients with CRS and to investigate the possible role of SA on epithelial activation. Sinonasal tissues were collected during surgery from control subjects and patients with CRS. Tissues were processed as described previously for analysis of mRNA (RT-PCR) and proteins (ELISA) for the majority of EGFR ligands within the tissue extracts. CRS tissue was used for evaluation of the distribution of epiregulin (EREG), an EGFR ligand, and MMP-1 by immunohistochemistry. In parallel studies, expression of these genes and proteins was analyzed in cultured primary airway epithelial cells. Elevated expression of EREG and MMP-1 mRNA and protein was observed in uncinate and polyp tissue from patients with CRSwNP. Immunohistochemistry study of clinical samples revealed that airway epithelial cells expressed both of these proteins. Cultured primary human airway epithelial cells expressed MMP-1, and MMP-1 was further induced by stimulation with EREG or heat-killed SA (HKSA). The induction of MMP-1 by HKSA was blocked by an antibody against EREG, suggesting that endogenous EREG induces MMP-1 after stimulation with HKSA. EREG and MMP-1 were found to be elevated in nasal polyp and uncinate tissues in patients with CRSwNP. Elevated expression of EREG and MMP-1 may be related to polyp formation in CRS, and colonization of SA might further enhance this process.

Staphylococcus aureus (SA) colonization and infection is common, and may promote allergic or inflammatory airway diseases, such as asthma, cystic fibrosis, and chronic rhinosinusitis by interacting with airway epithelial cells. Airway epithelial cells not only comprise a physical barrier, but also play key roles in immune, inflammatory, repair, and remodeling responses upon encounters with pathogens. To elucidate the impact of SA on epithelial-mediated remodeling of allergic airways, we tested the hypothesis that SA can enhance the remodeling process. Normal human bronchial epithelial (NHBE) cells were stimulated with heat-killed SA (HKSA) or transforming growth factor (TGF) α. Cell extracts were collected to measure mRNA (real-time RT-PCR) and signaling molecules (Western blot); supernatants were collected to measure protein (ELISA) after 24 hours of stimulation. Epidermal growth factor receptor (EGFR) signaling inhibition experiments were performed using a specific EGFR kinase inhibitor (AG1478) and TGF-α was blocked with an anti–TGF-α antibody. HKSA induced both mRNA and protein for TGF-α and matrix metalloproteinase (MMP) 1 from NHBE cells by a Toll-like receptor 2–dependent mechanism. Recombinant human TGF-α also induced mRNA and protein for MMP-1 from NHBE cells; anti–TGF-α antibody inhibited HKSA-induced MMP-1, suggesting that endogenous TGF-α mediates the MMP-1 induction by HKSA. HKSA-induced MMP-1 expression was suppressed when a specific EGFR kinase inhibitor was added, suggesting that EGFR signaling was mediating the HKSA-induced MMP-1 release. Exposure or colonization by SA in the airway may enhance the remodeling of tissue through a TGF-α–dependent induction of MMP-1 expression, and may thereby promote remodeling in airway diseases in which SA is implicated, such as asthma and chronic rhinosinusitis.

Aspergillus fumigatus (AF) infection and sensitization are common and promote Th2 disease in individuals with asthma. Innate immune responses of bronchial epithelial cells are now known to play a key role in determination of T cell responses upon encounter with inhaled pathogens. We have recently shown that extracts of AF suppress JAK-STAT signaling in epithelial cells and thus may promote Th2 bias. To elucidate the impact of AF on human bronchial epithelial cells, we tested the hypothesis that AF can modulate the response of airway epithelial cells to favor a Th2 response and explored the molecular mechanism of the effect. Primary normal human bronchial epithelial (NHBE) cells were treated with AF extract or fractionated AF extract before stimulation with poly I:C or infection with human rhinovirus serotype 16 (HRV16). Expression of CXCL10 mRNA (real-time RT-PCR) and protein (ELISA) were measured as markers of IFN-mediated epithelial Th1–biased responses. Western blot was performed to evaluate expression of IFN regulatory factor-3 (IRF-3), NF-κB, and tyrosine-protein phosphatase nonreceptor type 11 (PTPN11), which are other markers of Th1 skewing. Knockdown experiments for protease-activated receptor-2 (PAR-2) and PTPN11 were performed to analyze the role of PAR-2 in the mechanism of suppression by AF. AF and a high-molecular-weight fraction of AF extract (HMW-AF; > 50 kD) profoundly suppressed poly I:C– and HRV16-induced expression of both CXCL10 mRNA and protein from NHBE cells via a mechanism that relied upon PAR-2 activation. Both AF extract and a specific PAR-2 activator (AC-55541) suppressed the poly I:C activation of phospho–IRF-3 without affecting activation of NF-κB. Furthermore, HMW-AF extract enhanced the expression of PTPN11, a phosphatase known to inhibit IFN signaling, and concurrently suppressed poly I:C–induced expression of both CXCL10 mRNA and protein from NHBE cells. These results show that exposure of bronchial epithelial cells to AF extract suppressed poly I:C and HRV16 signaling via a mechanism shown to involve activation of PAR-2 and PTPN11. This action of AF may promote viral disease exacerbations and may skew epithelial cells to promote Th2 inflammation in allergic airway disorders mediated or exacerbated by AF, such as asthma and chronic rhinosinusitis.

Up to 50% of patients with chronic rhinosinusitis (CRS) have comorbid asthma, and we have reported that a subset of CRS patients who have nasal polyps (CRSwNP) have elevated autoantigen‐specific antibodies within their nasal polyps (NP). While increases in the prevalence and/or severity of both asthma and autoimmunity in women are well characterized, it is not known whether CRSwNP is more severe or frequent in women than men. We sought to determine whether CRSwNP demonstrated sex‐specific differences in frequency and/or severity. Using a retrospectively collected database of tertiary care patients (n = 1393), we evaluated the distribution of sex in patients with CRSwNP with or without comorbid asthma or aspirin hypersensitivity. We further compared the severity of sinus disease between men and women with CRSwNP. Although women comprised 55% of CRS patients without NP (CRSsNP), a significantly smaller proportion of CRSwNP patients were female (38%, P < 0.001). Interestingly, women with CRSwNP were significantly more likely than men to have comorbid asthma (P < 0.001), and 61% of patients with the most severe form of disease (aspirin‐exacerbated respiratory disease (CRSwNP plus asthma plus aspirin sensitivity)) were women (P < 0.05). Women with CRSwNP were significantly more likely to have taken oral steroids, and were more likely to have a history of revision surgeries (P < 0.05) compared to men. These data suggest that women with CRSwNP have more severe disease than men in a tertiary care setting. Future studies are needed to elucidate the mechanisms that drive disease severity in men and women, paving the way for the development of personalized treatment strategies for CRSwNP based on sex.

SARS-CoV-2 continues to transmit and evolve in humans and animals. White-tailed deer ( Odocoileus virginianus ) have been previously identified as a zoonotic reservoir for SARS-CoV-2 with high rates of infection and probable spillback into humans. Here we report sampling 1,127 white-tailed deer (WTD) in Pennsylvania, and a genomic analysis of viral dynamics spanning 1,017 days between April 2021 and January 2024. To assess viral load and genotypes, RNA was isolated from retropharyngeal lymph nodes and analyzed using RT-qPCR and viral whole genome sequencing. Samples showed a 14.64% positivity rate by RT-qPCR. Analysis showed no association of SARS-CoV-2 prevalence with age, sex, or diagnosis with Chronic Wasting Disease. From the 165 SARS-CoV-2 positive WTD, we recovered 25 whole genome sequences and an additional 17 spike-targeted amplicon sequences. The viral variants identified included 17 Alpha, 11 Delta, and 14 Omicron. Alpha largely stopped circulating in humans around September 2021, but persisted in WTD as recently as March of 2023. Phylodynamic analysis of pooled genomic data from Pennsylvania documents at least 12 SARS-CoV-2 spillovers from humans into WTD, including a recent series of Omicron spillovers. Prevalence was higher in WTD in regions with crop coverage rather than forest, suggesting an association with proximity to humans. Analysis of seasonality showed increased prevalence in winter and spring. Multiple examples of recurrent mutations were identified associated with transmissions, suggesting WTD-specific evolutionary pressures. These data document ongoing infections in white-tailed deer, probable onward transmission in deer, and a remarkable rate of new spillovers from humans.

Little is known about how the observed fat-specific pattern of 3D-spatial genome organisation is established. Here we report that adipocyte-specific knockout of the gene encoding nuclear envelope transmembrane protein Tmem120a disrupts fat genome organisation, thus causing a lipodystrophy syndrome. Tmem120a deficiency broadly suppresses lipid metabolism pathway gene expression and induces myogenic gene expression by repositioning genes, enhancers and miRNA-encoding loci between the nuclear periphery and interior. Tmem120a −/− mice, particularly females, exhibit a lipodystrophy syndrome similar to human familial partial lipodystrophy FPLD2, with profound insulin resistance and metabolic defects that manifest upon exposure to an obesogenic diet. Interestingly, similar genome organisation defects occurred in cells from FPLD2 patients that harbour nuclear envelope protein encoding LMNA mutations. Our data indicate TMEM120A genome organisation functions affect many adipose functions and its loss may yield adiposity spectrum disorders, including a miRNA-based mechanism that could explain muscle hypertrophy in human lipodystrophy. Little is known how spatial genome organization is directed in fat; however, key proadipogenic genes reposition from the nuclear envelope (NE) to the interior during adipogenesis. Here the authors show that deletion of NE protein Tmem120a in adipocytes disrupts fat genome organization, which results in suppression of lipid metabolism pathways and induces myogenic gene expression.

Glycolysis and the tricarboxylic acid cycle (TCA) cycle are reprogrammed in cancer cells to meet bioenergetic and biosynthetic demands, including by engagement with the extracellular matrix (ECM). However, the mechanisms by which the ECM engagement reprograms core energy metabolism is still unknown. We showed that the canonical cell−ECM adhesion protein focal adhesion kinase (FAK, also known as PTK2) and, specifically, its kinase activity, is driving cellular energetics. Using a mouse stem cell model of glioblastoma, we showed that deletion of the FAK gene simultaneously inhibits glycolysis and glutamine oxidation, increases mitochondrial fragmentation, elevates phosphorylation of the mitochondrial protein MTFR1L at serine residue 235 (S235) and triggers a mesenchymal-to-epithelial transition. These metabolic and structural changes arise through altered contractility of actomyosin, as shown by myosin light chain type II (MYL2, also known as MLC2) phosphorylated (p) at S19. This process can be reversed by Rho-kinase (ROCK) inhibitors revealing mechanotransduction pathway control of both mitochondrial dynamics and glutamine oxidation. FAK-dependent metabolic programming is associated with regulation of cell migration, invasive capacity and tumour growth in vivo. Our work describes a previously unrecognised FAK–ROCK axis that couples mechanical cues to the rewiring of energy metabolism, linking cell shape, mitochondrial function and malignant behaviour.

The multidrug resistance-1 (MDR1) gene encodes an ATP-dependent efflux transporter that is highly expressed in the colon. In mice, loss of MDR1 function results in colitis with similarities to human inflammatory bowel diseases (IBD). Here, we show that MDR1 has an unexpected protective role for the mitochondria where MDR1 deficiency results in mitochondrial dysfunction with increased mitochondrial reactive oxygen species (mROS) driving the development of colitis. Exogenous induction of mROS accelerates, while inhibition attenuates colitis in vivo; these effects are amplified in MDR1 deficiency. In human IBD, MDR1 is negatively correlated to SOD2 gene expression required for mROS detoxification. To provide direct evidential support, we deleted intestinal SOD2 gene in mice and showed an increased susceptibility to colitis. We exploited the genome-wide association data sets and found many (∼5%) of IBD susceptibility genes with direct roles in regulating mitochondria homeostasis. As MDR1 primarily protects against xenotoxins via its efflux function, our findings implicate a distinct mitochondrial toxin+genetic susceptibility interaction leading to mitochondrial dysfunction, a novel pathogenic mechanism that could offer many new therapeutic opportunities for IBD.