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Fig. 7 | Respiratory Research

Fig. 7

From: Functional changes in long-term incubated rat precision-cut lung slices

Fig. 7

Gene expression of long-term cultivated rat precision-cut lung slices (PCLS). PCLS were analysed for messenger ribonucleic acid (mRNA) expression of endothelin-1 (Edn1, A), endothelin-2 (Edn2, B), endothelin-3 (Edn3, C), endothelin receptor type A (Ednra, D), endothelin receptor type B (Ednrb, E), thromboxane A2 receptor (Tbxa2r, F), thromboxane A synthase 1 (Tbxas1, G), acetylcholinesterase (Ache, H), muscarinic acetylcholine receptor M1 (Chrm1, I), muscarinic acetylcholine receptor M2 (Chrm2, J) and muscarinic acetylcholine receptor M3 (Chrm3, K) in relation to a reference gene index (Ref. gene index) consisting of ribosomal protein lateral stalk subunit P0 (Rplp0) and beta-actin (Actb). For this purpose, PCLS were snap frozen in liquid nitrogen after denoted cultivation time, ribonucleic acid (RNA) isolation was performed by a combination of phenol–chloroform extraction and magnetic-bead technique. Afterwards, reverse transcription (RT) and quantitative polymerase chain reaction (qPCR) were performed. For each timepoint, measurements were performed in triplets, n = 5, per timepoint an average of 6–8 slices was snap frozen in liquid nitrogen, of which 100 mg of the slices were used for RNA isolation. RT-qPCR was performed with an input of 230 ng RNA. Data is expressed as mean + standard deviation (SD), dots represent individual data points. Statistics were carried out using the general mixed model analysis (PROC GLIMMIX, SAS 9.4, SAS Institute Inc., Cary, NC, USA), statistical differences to the control group (PCLS of day 1) are indicated by asterisks (*0.01 ≤ p < 0.05; **0.001 ≤ p < 0.01, ***p < 0.001)

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