Use of Rhino-Probe™ sampling for analyzing cross-shift variation in the expression of IL8, IL6 and TNF alpha in nasal epithelial cells from Colorado dairy workers

Introduction: Dairy work with its inherent exposure to bioaerosols, has long been associated with adverse inflammatory conditions in workers. In previous studies, dairy worker response to pro-inflammatory bioaerosols has been assessed by analysis of proteins (MPO, TNFα and IL8 etc) in nasal lavage fluid. Recent clinical studies have used Rhino-Probe Nasal Curettes to study inflammatory responses from exposure tobacco smoke and diesel exhaust. The object of this study was to utilize Rhino-probes™ in conjunction with RT-PCR to study the inflammatory response of Colorado dairy workers exposed to bioaerosols. Methods: Samples of nasal epithelial cells were collected from16 dairy workers pre and post workshift. Samples were stored in RNAlater (Qiagen) at -74°C. RNAlater was removed by adding phosphate buffer and pelletizing the cells by centrifuging. The supernatant was transferred to RNA lysis (RLT) buffer (Qiagen, RNeasy Micro Kit) and extracted with Qiagen QIAshredder and RNeasy Micro Kits. Total RNA content was quantified by Nano-Drop. mRNA transcripts for candidate genes GAPDH (housekeeping), IL8, IL6 and TNFalpha mRNA transcripts were quantified by RT-PCR (Bio-Rad iScript™ One-Step RT-PCR Kit with SYBR® Green; Instrument Bio-Rad CFX model). Results: A total of 32 paired (Pre and Post) samples were collected from the volunteers, 15 Hispanic males and 1 female. Of these, on 12 paired samples underwent RT-PCR. The remainder were rejected due to poor RNA yield and/or quality scores (260:280) of