| It is an important defense mechanism of nasal cavities and paranasal sinuses for goblet cells and submucosal gland cells to secret mucus. The mucus forms a layer of blanket on the superfacial surface of the epithelia of rhinosinus mucosa, which can entrap materials, prevent mechanical injury and maintain fluidity. However, in circumstances of chronic sinusitis and nasal polyps, the mucus may be produced excessively and its rheological properties changed. This makes it unsuitable to be transported by mucus-cilia system and results in mucus stasis, and, in turn, becomes a source of bacterial infection.The mucin concentration of the mucus has been shown to be the most important determinant of the viscosity and elasticity. Kim et al found MUC8 mRNA level increased clearly in the cultured epithelial cells of nasal polyps, compared with that in the epithelial cells of inferior turbinates. Ishinaga et al demonstrated dexamethasone suppressed MUC8 mRNA expression, which could be upregulated by lipopolysaccharide (LPS) in cultured human epithelial cells of nasal polyps. However, still not available is the data of investigation as to effects of corticosteroids on mucin gene expression of human nasal polyp tissue in vivo. The purpose of the present study was to elucidate the effects of corticosteroids on MUC8 gene expression of human nasal polyp tissue.Materials and methods1. Materials35 patients were included in the present study. They were some patients with chronic sinusitis and nasal polyps who received surgery in Department of Otorhinolaryngology of Second Affiliated Hospital of Zhejiang University between June 2002 and January 2003. Seventeen patients received corticosteroids therapy before surgery, aging from 16 to 60 ( average age 38.9 14.5 ), among them, twelve cases were male and 5 cases were female. We applied the staging system of chronic sinusitis with or without polyps and criteria of evaluation for therapeutic response to endoscopic sinus surgery, which was drawn up during National Conference of Rhinology at Haikou (1997), organized by Otorhinolaryngologic Branch Committee of Chinese Medical Association and Editorial Board of Chinese Journal of Otorhinolaryngology. Accordingly, eight cases were classified as type II stage 2, 8 as type II stage 3, and 1 as type III. These 17 patients were administered dexamethasone systemically for 4~7 days (average 4.7 1.4 days) before surgery. Ten of them were additionally received intranasal spray of corticosteroids. We made sure the other 18 patients, as control, didn't receive any kinds of corticosteroids therapy for 30 days before surgery, neither systemically nor topically. Among controls, 14 cases were male and 4 cases were female, aging from 14 to 58 (average age 36.3 15.2 ). They were composed of 4 cases of type II stage 1, 9 cases of type II stage 2, 4 cases of type II stage 3, and 1 case of type III. The samples of nasal polyp tissue were obtained at the time of surgery and immediately stored at 802. MethodsRNA IsolationFrozen tissue was homogenized in 1 ml of Trizol reagent. 0.2 ml of chloroform was then added to each sample. After vigorous vibration and centrifugation, RNA was present in the aqueous phase. After mixing with isopropyl alcohol, the RNA precipitate was washed with 75% ethanol. Then, the pellets were dissolved in DEPC water. The isolated RNA concentration and purity were identified through measurement of OD260 and OD280.Reverse Transcription of RNA8 g RNA pellets of each sample was taken and denatured at 70 for 5 minutes. Afterchilled on ice, the sample was added with 5 15 reverse transcriptase buffer, 1.25 1 10mM dNTPs, 1 1 50pmol Oligo dT, 1 1 200U/ 1 moloney murine leukemiavirus (M-MLV), 0.65 1 40U/ 1 RNasin, and supplemented with DEPC water to an end volume of 25 1. The mixture was kept at 42 for 60 minutes and inactivated at 95for 5 minutes. The generated cDNA samples were stored at - 20 until use. Polymerase chain reaction, PCRAmplification of MUC8 cDNA: The produced cDNA as above was amplified... |
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