Coexpression of type 2 immune targets in sputum-derived epithelial and dendritic cells from asthmatic subjects.

BACKGROUND: Noninvasive sputum sampling has enabled the identification of biomarkers in asthmatic patients. Studies of discrete cell populations in sputum can enhance measurements compared with whole sputum in which changes in rare cells and cell-cell interactions can be masked. OBJECTIVE: We sought to enrich for sputum-derived human bronchial epithelial cells (sHBECs) and sputum-derived myeloid type 1 dendritic cells (sDCs) to describe transcriptional coexpression of targets associated with a type 2 immune response. METHODS: A case-control study was conducted with patients with mild asthma (asthmatic cases) and healthy control subjects. Induced sputum was obtained for simultaneous enrichment of sHBECs and sDCs by using flow cytometry. Quantitative PCR was used to measure mRNA for sHBEC thymic stromal lymphopoietin (TSLP), IL33, POSTN, and IL25 and downstream targets in sDCs (OX40 ligand [OX40L], CCL17, PPP1R14A, CD1E, CD1b, CD80, and CD86). RESULTS: Final analyses for the study sample were based on 11 control subjects and 13 asthmatic cases. Expression of TSLP, IL33, and POSTN mRNA was increased in sHBECs in asthmatic cases (P = .001, P = .05, and P = .04, respectively). Expression of sDC OX40L and CCL17 mRNA was increased in asthmatic cases (P = .003 and P = .0001, respectively). sHBEC TSLP mRNA expression was strongly associated with sDC OX40L mRNA expression (R = 0.65, P = .001) and less strongly with sDC CCL17 mRNA expression. sHBEC IL33 mRNA expression was associated with sDC OX40L mRNA expression (R = 0.42, P = .04) but not sDC CCL17 mRNA expression. CONCLUSIONS: Noninvasive sampling and enrichment of select cell populations from sputum can further our understanding of cell-cell interactions in asthmatic patients with the potential to enhance endotyping of asthmatic patients.

Therapeutic hypothermia after cardiac arrest in a patient with systemic sclerosis and Raynaud phenomenon.

Therapeutic hypothermia favorably impacts neurologic outcomes in patients after cardiopulmonary arrest, although the appropriate target temperature is less clear. Its safety profile in patients with systemic sclerosis (SSc) and Raynaud phenomenon (RP), who may be at increased risk for ischemic complications, has not been addressed in the literature, to our knowledge. Digital lesions are commonly seen in patients with SSc, and cold-induced myocardial ischemia has also been reported. We describe a case of a man with SSc, RP, and digital ulcers who underwent therapeutic hypothermia after cardiopulmonary arrest. He regained full neurologic function, and except for digital necrosis, no hypothermia-associated adverse events were observed. Other risk factors for ischemia, such as cocaine use, may have contributed to the development of the digital necrosis. However, clinicians should be aware of the risk for ischemic complications in patients with SSc and RP when considering the appropriate target temperature after cardiopulmonary arrest.

Leptin signaling targets the thyrotropin-releasing hormone gene promoter in vivo.

The regulation of TRH gene expression in the paraventricular nucleus of the hypothalamus (PVH) by leptin is critical for normal function of the thyroid axis in rodents and humans. The TRH neuron in the PVH expresses both leptin and melanocortin-4 receptors, suggesting that both signaling systems may regulate TRH gene expression in vivo. Indeed, the TRH promoter responds to both of these signaling pathways in cell culture through identified cis-acting elements, which include signal transducer and activator of transcription (STAT) 3 and cAMP-response element binding protein binding sites that mediate leptin and melanocortin responses, respectively. To determine whether leptin signaling can directly target the TRH promoter in vivo, we developed a chromatin immunoprecipitation assay to use on leptin-treated animals. After a single injection of leptin in fasting animals, we could detect a significant increase in TRH gene expression in the PVH that correlated well with the induction of phosphorylated-STAT3 in the hypothalamus. Furthermore, using a STAT3 antibody, we could immunoprecipitate the STAT-binding site containing regions of both the TRH promoter and the promoter of the suppressor of cytokine signaling-3 gene, another well-defined target of leptin action. In contrast, upstream regions of these promoters that lack STAT sites were not precipitated. Taken together these experiments demonstrate that STAT3 mediates transcriptional effects of leptin in vivo and that the TRH promoter is a likely direct site of leptin action. In addition, these experiments demonstrate that chromatin immunoprecipitation can be used to characterize leptin-signaling in vivo.