Expression of the calcium-activated potassium channel in upper and lower segment human myometrium during pregnancy and parturition

Abstract

Background: Large conductance calcium-activated potassium channel (BKCa) plays an important role in the control of uterine contractility during pregnancy. The change from uterine quiescence to enhanced contractile activity may be associated with the spatial and temporal expression of BKCa within myometrium. The objectives of this study were to examine the expression of BKCa alpha- and beta-subunit in upper segment (US) and lower segment (LS) regions of uterus, and to investigate for the possibly differential expression of these proteins in US and LS myometrium obtained from three functional states: (1) non-pregnant (NP); (2) term pregnant not in labour (TNL) and (3) term pregnant in labour (TL).

Methods: Myometrial biopsies were collected from non-pregnant women at hysterectomy and pregnant women at either elective caesarean section or emergency caesarean section. Protein expression level and cellular localization of BKCa alpha- and beta-subunit in US and LS myometrium were determined by Western blot analysis and immunohistochemistry, respectively.

Results: BKCa alpha- and beta-subunit were predominantly localized to myometrial smooth muscle in both US and LS myometrium obtained from non-pregnant and pregnant patients. The level of BKCa alpha-subunit in US but not in LS was significantly higher in NP myometrium than those measured in myometrium obtained during pregnancy. Lower expression of BKCa alpha-subunit in both US and LS was found in TL than in TNL biopsies. Expression of beta-subunit in both US and LS myometrium was significantly reduced in TL group compared with those measured in TNL group. There was no significant difference in BKCa beta-subunit expression in either US or LS between NP and TNL group.

Conclusion: Our results suggest that expression of BKCa alpha- and beta-subunit in pregnant myometrium is reduced during labour, which is consistent with the myometrial activity at the onset of parturition.

Figure 1

Immunolocalization of BK_Ca channel α- and β-subunit in non-pregnant myometrium. Positive staining for the α-subunit of BK_Ca (arrow) in (A) US myometrium and (B) LS myometrium. Positive staining for the β-subunit (arrow) in (C) US myometrium and (D) LS myometrium. (E-H) Negative controls. The primary antibody was substituted with (E) normal rabbit serum or (G) PBS. Sections were stained with (F) α-subunit preabsorption antibody or (H) β-subunit preabsorption antibody. Original magnification ×400.

Figure 2

Immunolocalization of BK_Ca channel α- and β-subunit in myometrium from pregnant women at term in labour or not in labour. A-D shows representative sections for positive staining for α-subunit in (A) US myometrium not in labour, (B) LS myometrium not in labour, (C) US myometrium in labour and (D) LS myometrium in labour. E-H shows representative sections for positive staining for β-subunit in (E) US myometrium not in labour, (F) LS myometrium not in labour, (G) US myometrium in labour and (H) LS myometrium in labour. Arrow: positive staining. Original magnification ×400.

Figure 3

Western blot analysis of BK_Ca α- and β-subunit in pregnant US and LS myometrium. (A) Representative immunoblots showing the expression of the 110 kDa α-subunit and the 35 kDa β-subunit in human myometrium. (B, C) The expression values of α- and β-subunit from all the pregnant patients was combined to given an overall expression profile in the US and LS. Data were expressed as mean ± SEM.

Figure 4

Semiquantitation of Western blot signals of BK_Ca α-subunit in US and LS myometrium. Myometrial tissues were obtained from non-pregnant women (n = 8) and pregnant women at term before the onset of labour (n = 10) or during active labour (n = 10). (A) Levels of α-subunit in US samples. (B) α-subunit expression in LS samples. Representative protein bands were presented on the top of the histogram. Data were expressed as mean ± SEM. *P < 0.05, **P < 0.01 with NP; #P < 0.05, ##P < 0.01 with TNL.

Figure 5

Semiquantitation of Western blot signals of BK_Ca β-subunit in US and LS myometrium. Myometrial biopsies were obtained from nonpregnant women (n = 8) and pregnant women at term not in labour (n = 10) or during active labour (n = 10) (A) BK_Ca β-subunit expression in US, (B) BK_Ca β-subunit level in LS. Representative protein bands were presented on the top of the histogram. Data were expressed as mean ± SEM. **P < 0.01 with NP; #P < 0.05, ##P < 0.01 with TNL.