TY - JOUR
T1 - Expression of parathyroid hormone-related peptide (PTH-rp) and its receptor in the porcine ovary
T2 - Regulation by transforming growth factor-β and possible paracrine effects of granulosa cell PTH-rp secretion on theca cells
AU - Garmey, J. C.
AU - Schnorr, J. A.
AU - Bruns, M. E.
AU - Bruns, D. E.
AU - Seaner, R. M.
AU - Ferguson, J. E.
AU - Jayes, F. C.L.
AU - Aguirre, C.
AU - Veldhuis, J. D.
N1 - Copyright:
Copyright 2018 Elsevier B.V., All rights reserved.
PY - 2000
Y1 - 2000
N2 - Parathyroid hormone-related peptide (PTH-rp) and the PTH-rp receptor are expressed in certain cancers as well as in many normal tissues. To evaluate the expression of this Ca2+-regulating hormone and its receptor in porcine ovary, we isolated partial cDNAs encoding homologous PTH-rp and PTH-rp receptor using reverse transcription-polymerase chain reaction (RT-PCR). The cDNA encoding PTH-rp (419 base pairs [bp]) was 92% and 87% homologous to human and rat sequences, respectively, while the PTH-rp receptor clone (167 bp) was 94% and 91% identical to the human and rat genes. Qualitative estimates of PTH-rp mRNA by RT-PCR indicated that the PTH-rp gene is expressed at high levels in the corpus luteum but is undetectable in granulosa and theca cells isolated from small (1-5 mm) and medium-sized (5-8 mm) antral follicles. In contrast, PTH-rp receptor transcripts were most abundant in corpora lutea and theca cells, and least abundant (albeit detectable) in granulosa cells. Regulation of PTH-rp protein production was assessed in serum-free monolayer cultures of porcine granulosa cells. Transforming growth factor (TGF)-β1 (100 ng/ml) increased PTH-rp concentrations (assayed by two-site immunoradiometric assay of culture media) as well as corresponding PTH-rp mRNA accumulation (assessed by RT-PCR) in a time-dependent manner, with maximal responses of 3- to 5-fold at 96 h. TGF- β1 dose-response studies revealed an ED50 of 0.24-0.38 ng/ml with a maximal effect at 30 ng/ml. Other growth factors and hormones, including insulin, insulin-like growth factor (type I), epidermal growth factor, FSH, estradiol, and interleukin-1, failed to alter PTH-rp secretion. Biological effects of PTH-rp were evident in purified porcine theca cells. Using the Ca2+-sensitive fluorescent indicator dye, fura-2, and digital imaging videomicroscopy, we found that PTHrp (1 μM) stimulated intracellular free calcium ion concentrations ([Ca2+](i)) in single porcine theca cells. The [Ca2+](i) elevation was characterized by a slow and prolonged rise. After PTH-rp stimulation, theca cells maintained responsiveness to hormone stimulation by LH, which elicited a typical theca cell [Ca2+](i) response. Our results allow a hypothesis of a paracrine intrafollicular signaling system involving interaction between theca cell-derived TGF-β and granulosa cell-derived PTH-rp, with feedback by PTH-rp on theca cells. Alternatively, expression of mRNAs encoding PTH-rp and its receptor in corpora lutea suggests that this peptide may play a role in luteal cell function. The precise role of this intraovarian PTH-rp system will require further study.
AB - Parathyroid hormone-related peptide (PTH-rp) and the PTH-rp receptor are expressed in certain cancers as well as in many normal tissues. To evaluate the expression of this Ca2+-regulating hormone and its receptor in porcine ovary, we isolated partial cDNAs encoding homologous PTH-rp and PTH-rp receptor using reverse transcription-polymerase chain reaction (RT-PCR). The cDNA encoding PTH-rp (419 base pairs [bp]) was 92% and 87% homologous to human and rat sequences, respectively, while the PTH-rp receptor clone (167 bp) was 94% and 91% identical to the human and rat genes. Qualitative estimates of PTH-rp mRNA by RT-PCR indicated that the PTH-rp gene is expressed at high levels in the corpus luteum but is undetectable in granulosa and theca cells isolated from small (1-5 mm) and medium-sized (5-8 mm) antral follicles. In contrast, PTH-rp receptor transcripts were most abundant in corpora lutea and theca cells, and least abundant (albeit detectable) in granulosa cells. Regulation of PTH-rp protein production was assessed in serum-free monolayer cultures of porcine granulosa cells. Transforming growth factor (TGF)-β1 (100 ng/ml) increased PTH-rp concentrations (assayed by two-site immunoradiometric assay of culture media) as well as corresponding PTH-rp mRNA accumulation (assessed by RT-PCR) in a time-dependent manner, with maximal responses of 3- to 5-fold at 96 h. TGF- β1 dose-response studies revealed an ED50 of 0.24-0.38 ng/ml with a maximal effect at 30 ng/ml. Other growth factors and hormones, including insulin, insulin-like growth factor (type I), epidermal growth factor, FSH, estradiol, and interleukin-1, failed to alter PTH-rp secretion. Biological effects of PTH-rp were evident in purified porcine theca cells. Using the Ca2+-sensitive fluorescent indicator dye, fura-2, and digital imaging videomicroscopy, we found that PTHrp (1 μM) stimulated intracellular free calcium ion concentrations ([Ca2+](i)) in single porcine theca cells. The [Ca2+](i) elevation was characterized by a slow and prolonged rise. After PTH-rp stimulation, theca cells maintained responsiveness to hormone stimulation by LH, which elicited a typical theca cell [Ca2+](i) response. Our results allow a hypothesis of a paracrine intrafollicular signaling system involving interaction between theca cell-derived TGF-β and granulosa cell-derived PTH-rp, with feedback by PTH-rp on theca cells. Alternatively, expression of mRNAs encoding PTH-rp and its receptor in corpora lutea suggests that this peptide may play a role in luteal cell function. The precise role of this intraovarian PTH-rp system will require further study.
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U2 - 10.1095/biolreprod62.2.334
DO - 10.1095/biolreprod62.2.334
M3 - Article
C2 - 10642570
AN - SCOPUS:0033968580
SN - 0006-3363
VL - 62
SP - 334
EP - 339
JO - Biology of Reproduction
JF - Biology of Reproduction
IS - 2
ER -