Preparation of highly purified porcine theca cells

Kataoka, Nobuhiko; Taii, Shunzo; Kita, Magdalena; Mori, T.

doi:10.1530/jrf.0.1020073

Cited by 5 publications

(5 citation statements)

References 28 publications

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“…Morphologically, the cultured granulosa cells maintained a characteristic round (or polygonal) shape throughout our culture conditions [ 25 , 31 ]. Purified granulosa cells from large and medium follicles displayed steroidogenic criteria [ 20 ]: (i) the granulosa cells did not synthesize estradiol unless aromatized androgens (i.e., androstenedione and testosterone) were added, and (ii) FSH significantly stimulated progesterone production in granulosa cells. When conducting the cell culture and reagent incubation experiments, we performed at least four independent experiments as in previous literature [ 22 , 25 , 31 ].…”

Section: Methodsmentioning

confidence: 99%

“…Female sex hormone production is complex and regulated by interactions between granulosa cells and theca cells. Progesterone is the main secretory product of granulosa cells and diffuses into theca cells to serve as a substrate for androgen biosynthesis [ 20 , 21 , 22 ]. Thereafter, theca cells subsequently release androgens for granulosa cells to convert androgens into estrogens.…”

Section: Introductionmentioning

confidence: 99%

“…FSH-induced progesterone release is dually regulated through two distinct intracellular signaling systems, including adenyl cyclase/cAMP- and L-type calcium channel-mediated pathways. FSH increases progesterone [ 20 , 21 , 23 , 24 , 25 ] and estradiol production [ 20 , 24 , 26 ], which is regulated via the cAMP-related signaling pathway [ 23 , 24 , 26 ]. It further activates P450scc (cytochrome P450 side-chain cleavage), 3β-HSD or P450arom in granulosa cells [ 25 , 27 , 28 , 29 ].…”

Section: Introductionmentioning

confidence: 99%

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Amphetamine-Decreased Progesterone and Estradiol Release in Rat Granulosa Cells: The Regulatory Role of cAMP- and Ca2+-Mediated Signaling Pathways

Chen

Wang

et al. 2021

Biomedicines

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The purpose of this study is to evaluate the amphetamine effects on progesterone and estradiol production in rat granulosa cells and the underlying cellular regulatory mechanisms. Freshly dispersed rat granulosa cells were cultured with various test drugs in the presence of amphetamine, and the estradiol/progesterone production and the cytosolic cAMP level were measured. Additionally, the cytosolic-free Ca2+ concentrations ([Ca2+]i) were measured to examine the role of Ca2+ influx in the presence of amphetamine. Amphetamine in vitro inhibited both basal and porcine follicle-stimulating hormone-stimulated estradiol/progesterone release, and amphetamine significantly decreased steroidogenic enzyme activities. Adding 8-Bromo-cAMP did not recover the inhibitory effects of amphetamine on progesterone and estradiol release. H89 significantly decreased progesterone and estradiol basal release but failed to enhance a further amphetamine inhibitory effect. Amphetamine was capable of further suppressing the release of estradiol release under the presence of nifedipine. Pretreatment with the amphetamine for 2 h decreased the basal [Ca2+]i and prostaglandin F2α-stimulated increase of [Ca2+]i. Amphetamine inhibits progesterone and estradiol secretion in rat granulosa cells through a mechanism involving decreased PKA-downstream steroidogenic enzyme activity and L-type Ca2+ channels. Our current findings show that it is necessary to study the possibility of amphetamine perturbing reproduction in females.

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Amphetamine-Decreased Progesterone and Estradiol Release in Rat Granulosa Cells: The Regulatory Role of cAMP- and Ca2+-Mediated Signaling Pathways

Chen

Wang

et al. 2021

Biomedicines

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“…GC were collected as thecal layers were deposited into a Petri dish containing medium 199 supplemented with 25 mM Hepes, 100 U/ml penicillin, 100 g/ml streptomycin, 10 g/ml gentamicin, and 0.5 g/ml fungizone (M199). A modification of the method described by Kataoka et al [24] was used to obtain the final theca cell preparations. Thecal layers were washed with M199 over a 177-m mesh to remove loosely adhering GC.…”

Section: Theca and Granulosa Cell Isolation And Culturementioning

confidence: 99%

“…The 1-day attachment period for the TC eliminates GC contamination, since 2 days are required for GC to attach [23]. In addition, this method of TC isolation results in less than 3% contamination with GC [24].…”

Section: Theca and Granulosa Cell Isolation And Culturementioning

confidence: 99%

Relaxin Secretion and Gene Expression in Porcine Granulosa and Theca Cells Are Stimulated during In Vitro Luteinization1

Ohleth

Bagnell

1999

Biology of Reproduction

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During formation of the corpus luteum, the primary source of relaxin switches from theca cells (TC) to granulosa-derived, large luteal cells. What controls this shift in relaxin production is poorly understood. The objective of this study was to observe the effect of luteinization on relaxin gene expression and secretion by porcine granulosa (GC) and TC using an in vitro model. TC and GC from medium-sized porcine follicles (4-6 mm) were treated for up to 8 days with LH (250 ng/ml) and/or insulin-like growth factor-I (IGF-I; 10 ng/ml). Media were assayed for relaxin and progesterone by RIA, changes in cell morphology were recorded, and total RNA was subjected to reverse transciption-polymerase chain reaction to monitor relaxin gene expression. In vitro luteinization, induced with LH + IGF-I treatment, was confirmed in both GC and TC by a change in morphology and a sustained, significant rise in progesterone secretion. In luteinizing GC, relaxin secretion was first detected after 5 treatment days, and steadily rose until it became significantly higher (p < 0.001) by treatment Days 7-8. In contrast, relaxin release from luteinizing TC was significant after only 2 days of treatment (p < 0.05) and increased consistently over the 8-day culture period (p < 0.001). In GC, relaxin mRNA was not detected until treatment Day 4 and became significantly higher (p < 0.001) by Day 8, the final treatment day. Relaxin transcript in luteinizing TC was low on treatment Days 2-4 and significantly higher (p < 0.01) by treatment Days 6 and 8. In summary, the present study demonstrates that hormones important in the control of luteinization are essential for regulating relaxin gene expression and secretion by GC and TC in the porcine follicle.

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