Estrogen induces epidermal growth factor (EGF) receptor and its ligands in human fallopian tube: involvement of EGF but not transforming growth factor-alpha in estrogen-induced tubal cell growth in vitro.

We studied the estrogen-dependent expression of epidermal growth factor (EGF), transforming growth factor (TGF) alpha, and EGF receptor gene transcripts in human fallopian tubes in vivo and in vitro. Competitive polymerase chain reaction (PCR) was performed on the fallopian tube RNA samples from the postmenopausal women with or without estrogen replacement. Amounts of EGF, TGF alpha, and EGF receptors gene transcripts in the estrogen-treated group (n = 3) were significantly (P < 0.01) more than those in the untreated group (n = 3). Competitive PCR also showed that EGF, TGF alpha, and EGF receptor gene transcripts level in tubal cells were increased by estrogen in vitro: messenger RNA levels of these factors were significantly (P < 0.01, n = 3) increased in cells incubated with 10(-8) M estrogen compared with those in cells without estrogen treatment. We studied whether EGF and/or TGF alpha is involved in the estrogen-induced tubal cell growth in vitro. Estrogen enhanced the [3H]-thymidine incorporation into the cell in dose- and time-dependent manners in culture: estrogen treatment for more than 12 h significantly (P < 0.05) enhanced the [3H]-thymidine incorporation into the cell at 10(-8) M. The estrogen-induced cell growth was observed in association with the increase in EGF, TGF alpha, and EGF receptor messenger RNA levels by estrogen. If the EGF and/or TGF alpha is involved in the cell growth, then the estrogen-induced cell growth should be suppressed by blocking the action of EGF and/or TGF alpha. Therefore, we examined the effects of neutralizing monoclonal antibodies against EGF, TGF alpha, and EGF receptors. Anti-EGF antibody significantly reduced the estrogen-induced increase in [3H]-thymidine incorporation, whereas anti-TGF alpha antibody failed to show the effect. Anti-EGF receptor antibody showed a significant suppressive effect on the estrogen-induced increase in [3H]-thymidine incorporation. Moreover, the growth inhibitory effect by 1 microgram/ml anti-EGF was restored by 10(-8) M EGF but not by TGF alpha even at 10(-6) M. All these data suggest that estrogen induces EGF and TGF alpha/EGF receptors in the human fallopian tube and that EGF but not TGF alpha may be involved in the estrogen-induced human tubal cell growth in vitro.

Menstrual cycle specific expression of epidermal growth factor receptors in human fallopian tube epithelium.

We studied the expression of epidermal growth factor (EGF) receptor protein and messenger RNA (mRNA) in human fallopian tubes at three stages of the menstrual cycle: early follicular (n = 3), late follicular (n = 3) and luteal (n = 3). Immunohistochemical studies in the ampullary portion of the tubes showed that specific staining was localized to the epithelium and the vascular endothelium. Staining of the epithelium was intense at the late follicular and luteal stages, while it was weak at the early follicular stage. 125I-EGF binding study in the tubal plasma membranes revealed a class of high-affinity EGF receptors. Although dissociation constants were similar between the stages, numbers of binding sites at the late follicular and luteal stages were significantly (P < 0.01) greater than those at the early follicular stage. Western blotting showed that tubal plasma membranes contain M(r) 170,000 EGF receptor protein. The amounts were significantly (P < 0.01, n = 3) greater at the late follicular and luteal stages than those at the early follicular stage. Reverse transcription and polymerase chain reaction (RT-PCR) revealed that EGF receptor mRNA was expressed in all the 9 RNA samples (n = 3 for each stage) from the tubal ampullary portion. The amounts were significantly (P < 0.01, n = 3) greater at the late follicular and luteal stages than those at the early follicular stage (by a competitive PCR). Increase in the amounts of EGF receptor protein and mRNA occurred in association with an increase in serum oestradiol but not progesterone levels. Next we examined whether EGF receptor and its ligands (EGF and transforming growth factor alpha) are directly induced by oestrogen. We found that specific staining for EGF receptor and its ligands in the tubal epithelium was detected (by immunohistochemistry) in postmenopausal women with oestrogen replacement (n = 3), but not in subjects without oestrogen replacement (n = 3). These results suggested that EGF receptors in the human tubal epithelium are expressed in relation to specific stages of the menstrual cycle and that the expression may be induced by oestrogen.

A novel sulfonylurea receptor forms with BIR (Kir6.2) a smooth muscle type ATP-sensitive K+ channel.

We have isolated a cDNA encoding a novel isoform of the sulfonylurea receptor from a mouse heart cDNA library. Coexpression of this isoform and BIR (Kir6.2) in a mammalian cell line elicited ATP-sensitive K+ (KATP) channel currents. The channel was effectively activated by both diazoxide and pinacidil, which is the feature of smooth muscle KATP channels. Sequence analysis indicated that this clone is a variant of cardiac type sulfonylurea receptor (SUR2). The 42 amino acid residues located in the carboxyl-terminal end of this novel sulfonylurea receptor is, however, divergent from that of SUR2 but highly homologous to that of the pancreatic one (SUR1). Therefore, this short part of the carboxyl terminus may be important for diazoxide activation of KATP channels. The reverse transcription-polymerase chain reaction analysis showed that mRNA of this clone was ubiquitously expressed in diverse tissues, including brain, heart, liver, urinary bladder, and skeletal muscle. These results suggest that this novel isoform of sulfonylurea receptor is a subunit reconstituting the smooth muscle KATP channel.