The human carotid body transcriptome with focus on oxygen sensing and inflammation--a comparative analysis

Abstract

The carotid body (CB) is the key oxygen sensing organ. While the expression of CB specific genes is relatively well studied in animals, corresponding data for the human CB are missing. In this study we used five surgically removed human CBs to characterize the CB transcriptome with microarray and PCR analyses, and compared the results with mice data. In silico approaches demonstrated a unique gene expression profile of the human and mouse CB transcriptomes and an unexpected upregulation of both human and mouse CB genes involved in the inflammatory response compared to brain and adrenal gland data. Human CBs express most of the genes previously proposed to be involved in oxygen sensing and signalling based on animal studies, including NOX2, AMPK, CSE and oxygen sensitive K+ channels. In the TASK subfamily of K+ channels, TASK-1 is expressed in human CBs, while TASK-3 and TASK-5 are absent, although we demonstrated both TASK-1 and TASK-3 in one of the mouse reference strains. Maxi-K was expressed exclusively as the spliced variant ZERO in the human CB. In summary, the human CB transcriptome shares important features with the mouse CB, but also differs significantly in the expression of a number of CB chemosensory genes. This study provides key information for future functional investigations on the human carotid body.

Figure 1. Haematoxylin–eosin staining of the human carotid body

A representative section of the human carotid body stained with hematoxylin–eosin.

Figure 2. Expression of selected chemosensory genes in the human carotid body

Relative expression of selected genes was determined by real time PCR. Data were analysed according to the common ΔC_t method (see Methods), normalized to the reference gene TBP and related to the expression of tyrosine hydroxylase (TH). The resulting values are expressed as means, n = 5, with the exception of Maxi-K_Strex, which was expressed only in the CBs of patients B and E. *Combined data using previously published results from patients A–C (Fagerlund et al. 2010) and data from two additional patients D and E. #Data from patients B and C. The full gene names are shown in Supplement S13.

Figure 3. Expression of TASK-like channels and Maxi-K in mouse and human carotid bodies analysed with PCR

A, expression of TASK-1 and TASK-3 K⁺ channels in the DBA/2J mouse CB. cDNA from three DBA/2J mice (DBA/2J-1-3) was used as a template to generate TASK-1 (515 bp) and TASK-3 (413 bp) PCR fragments. Primer sequences and PCR conditions are described by Ortega-Saenz et al. (2010). Amplification of actin was used as a loading control. B, expression of splice variants of Maxi-K in the human CB. A set of primers generating 615 and/or 777 bp long amplicons indicative of the expression of ZERO (without the Strex exon) and/or Strex isoforms of Maxi-K, respectively, was used for PCR analysis of cDNA from two human CBs (patients B and E). Note the absence of the 777 bp fragment in both CBs under stringent (1.0 m MgCl₂) and less stringent (1.5 mm MgCl₂) PCR conditions. Even increased number of cycles (40) and lower annealing temperature (55°C) failed to produce the 777 bp amplicon (results not shown). An additional, more Strex-specific set of primers (see Methods) was similarly ineffective (results not shown). M, DNA ladder.