Investigating the role of CFTR in human and mouse myometrium

Abstract

Background: Abnormal cystic fibrosis transmembrane conductance regulator (CFTR) function in cystic fibrosis (CF) has been linked to airway smooth muscle abnormalities including bronchial hyperresponsiveness. However, a role for CFTR in other types of smooth muscle, including myometrium, remains largely unexplored. As CF life expectancy and the number of pregnancies increases, there is a need for an understanding of the potential role of CFTR in myometrial function.

Methods: We investigated the role of CFTR in human and mouse myometrium. We used immunofluorescence to identify CFTR expression, and carried out contractility studies on spontaneously contracting term pregnant and non-pregnant mouse myometrium and term pregnant human myometrial biopsies from caesarean sections.

Results: CFTR was found to be expressed in term pregnant mouse myometrium. Inhibition of CFTR, with the selective inhibitor CFTR_inh-172, significantly reduced contractility in pregnant mouse and human myometrium in a concentration-dependent manner (44.89 ± 11.02 term pregnant mouse, 9.23 ± 4.75 term-pregnant human; maximal effect at 60 μM expressed as a percentage of the pre-treatment control period). However, there was no effect of CFTR_inh-172 in non-pregnant myometrium.

Conclusion: These results demonstrate decreased myometrial function when CFTR is inhibited, which may have implications on pregnancy and labour outcome and therapeutic decisions for labour in CF patients.

Keywords: CFTR; Contractility; Cystic fibrosis; Human; Mouse; Myometrium; Pregnancy.

Fig. 1

The effect of different concentrations of CFTR_inh-172 (1–100 μM) on contractility of term pregnant and non-pregnant mouse myometrium in vitro.

Fig. 2

Effect of 10 μM CFTRinh172 in the absence and presence of 0.1 nM oxytocin. Scatter plot demonstrating the significant inhibitory effect of 10 μM CFTR_inh-172 on AUC, in the absence and presence of 0.1 nM oxytocin (Mann Whitney Test; p < 0.05).

Fig. 3

The effect of increasing concentrations of CFTR_inh-172 (1–60 μM) on contractility of human myometrium in vitro. (A), shows control period of contraction, followed by increased concentrations of CFTR_inh-172 (B–E). (F), shows the concentration-response curve of the AUC, with an pEC₅₀ value of 5.35 ± 0.10 compared to pregnant mouse pEC₅₀ value of 5.12 ± 0.14.

Fig. 4

The effect of CFTR_inh-172 on the response to high K⁺ solution in term pregnant mouse. Control high K+ responses are shown in (A) (protocol 1) and (E) (protocol 2); High K+ responses in the presence of CFTR_inh-172 are shown in (B) (protocol 1) and (F) (protocol 2). Dotted lines in A and B indicate maximum High K+ amplitude. Significant decrease in peak amplitude in the presence of CFTR_inh-172 (difference before and in presence of CFTR inhibitor; paired Student's t-test) and area under the curve (AUC) are shown as scatterplots in (C) and (D). Significant decrease of the steady-state plateau amplitude in the presence of CFTR_inh-172 compared to control (Mann Whitney test) is shown in (G).

Fig. 5

Immunofluorescence showing expression of CFTR in (A), actin in (B), CFTR and actin images (A and B) merged to show relative expression patterns shown in (C). DAPI (4,6-diamidino-2-phenylindole), staining of myometrial nuclei is shown in (D) and control no primary antibody shown in (E).