Short communication: optimization of semi-quantitative RT PCR analysis for CPT I gene expression in rainbow trout (Oncorhynchus mykiss)

A key enzyme in mitochondrial β¬oxidation, carnitine palmitoyltransferase (CPT) I, is transcriptionally regulated in mammals, but this enzyme also experiences allosteric modulations (Harano et al., 1985; Murthy and Pande, 1987; Bezaire et al., 2004). CPT I is located on the inner side of the outer mitochondrial membrane and catalyses the conversion of acyl-CoA to fatty acylcarnitine (Kerner and Hoppel, 2000; Price et al., 2000). Quantitative RT-PCR is a reliable technique for measuring transcripts in small amounts of tissue (Spriewald et al., 2000). With this technique, multiple mRNAs can be assayed simultaneously in a relatively short period of time. Here we describe the standard procedure, optimized in our laboratory, to assess CPT I levels with β¬actin as an internal control in rainbow trout, and all the necessary controls to ensure a quantitative analysis. RNA Extraction and Reverse Transcription Total cellular RNA was isolated from liver of rainbow trout using RNX reagent (Cinnagen-Iran). To obtain cDNA, 1 µg of total RNA was subjected to reverse transcription polymerase chain reaction (RT-PCR) with MuLV reverse transcriptase using the RevertAidTM M¬MuLV Reverse Transcriptase Kit (Fermentase Life Science, Germany) and random hexamer primer. Reaction conditions in the reverse transcription step are mostly dependent on the enzyme and the primers of choice. Whereas other protocols to require the use of specific primers, we prefer to reverse transcribe the total RNA population with random hexamers so that different PCR analyses could be performed on the same cDNA sample

Incidental sinus abnormalities in 256 patients referred for brain MRI

Background: Imaging abnormalities in the paranasal sinuses are regularly noted as incidental findings on MRI, however, little is known about their prevalence in the Iranian population. The purpose of this study was to classify these findings in the paranasal sinuses as seen on MRI and to investigate the prevalence, according to site and type of paranasal abnormality. Methods: In this cross-sectional study, the T2-weighted axial MRI of 256 patients with diseases unrelated to their paranasal sinuses were reviewed between May 2002 and June 2003. The findings were categorized according to the anatomic location and the imaging characteristics of the abnormality. The abnormalities recorded included total sinus opacification, mucoperiosteal thickening >5mm, air fluid levels and retention cysts or polyps. Unilateral or bilateral involvement and septal deviation were also noted. A sinus was considered normal if it was fully aerated and no soft-tissue density was apparent within the cavity. Results: Among our cases, 111 (43.5%) were male and 145 (56.5%) were female. Of these patients, abnormalities in one or more of the sinus groups were found in 110 subjects (42.9%), 55.5% of which were male and 44.5% were female (P=0.001). Maxillary sinus abnormalities were observed in 66.4% of the patients, while ethmoid sinus abnormalities were found in 63.6%. Of the ethmoid abnormalities, 21% were found in the anterior section, 9% in the middle ethmoid, and 8% in the posterior ethmoid. The most common abnormality found was mucosal thickening. Among our cases, 23.4% had septal deviation, which was significantly higher among those with sinusitis (29% versus 19.1%; P<0.01). Of those patients with sinus involvement, 16% were involved in the sphenoid sinus and 5% in the frontal sinus. The results obtained from the patients with sinus abnormality revealed that 85% suffered from cough, nasal obstruction, runny nose, facial pain and post nasal discharge and 24% had been diagnosed with chronic sinusitis by physician. Conclusion: Our results showed that there was a high rate of incidental abnormalities in the paranasal sinuses that are unrelated to the patient's presenting problems