Assessment of cell line competence for studies of pharmacological GPR30 modulation

Sousa, Cátia; Ribeiro, M.; Rufino, Ana T.; Leitão, Alcino J.; Mendes, Alexandrina Ferreira

doi:10.1080/10799893.2016.1203943

Cited by 10 publications

(13 citation statements)

References 33 publications

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“…These data imply that GPR30 may play a crucial role in the anti-NASH effects of DHEA. Previous studies showed that GPR30 is not a direct target of E2 in vivo and in vitro [ [33] , [34] , [35] ], whereas E2 usually bind to and activates the classical ERs, including ERα and ERβ [ 31 ]. Thus, the conversion of DHEA into E2 may indirectly enhanced the expression level of GPR30 by binding another target, such as the classical ERs.…”

Section: Resultsmentioning

confidence: 99%

The sex steroid precursor dehydroepiandrosterone prevents nonalcoholic steatohepatitis by activating the AMPK pathway mediated by GPR30

Wang

Yao

et al. 2021

Redox Biology

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Section: Resultsmentioning

confidence: 99%

The sex steroid precursor dehydroepiandrosterone prevents nonalcoholic steatohepatitis by activating the AMPK pathway mediated by GPR30

Wang

Yao

et al. 2021

Redox Biology

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“…On SDS-PAGE, in carotid artery lysates, GPER was resolved in 2 bands, namely, a major band with a corresponding molecular weight of ≈ 50 kDa-as previously described in other tissues- 54 and a minor band of GPER (≈43 kDa) as reported by other investigators. 55,56 As we have seen in vitro, with VSMC primary culture, after carotid ligation or vascular injury, there was a detectable downregulation of GPER content both with regard to protein content (to 15±6% of control; n=3; Figure 3D) and mRNA content (to 28±3% of control; n=3; Figure 3E). …”

Section: Carotid Injury Leads To An Attenuation Of Gper Expressionmentioning

confidence: 69%

Extent of Vascular Remodeling Is Dependent on the Balance Between Estrogen Receptor α and G-Protein–Coupled Estrogen Receptor

et al. 2016

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Abstract-Estrogens are important regulators of cardiovascular function. Some of estrogen's cardiovascular effects are mediated by a G-protein-coupled receptor mechanism, namely, G-protein-coupled estrogen receptor (GPER). Estradiol-mediated regulation of vascular cell programmed cell death reflects the balance of the opposing actions of GPER versus estrogen receptor α (ERα). However, the significance of these opposing actions on the regulation of vascular smooth muscle cell proliferation or migration in vitro is unclear, and the significance in vivo is unknown.To determine the effects of GPER activation in vitro, we studied rat aortic vascular smooth muscle cells maintained in primary culture. GPER was reintroduced using adenoviral gene transfer. Both estradiol and G1, a GPER agonist, inhibited both proliferation and cell migration effects that were blocked by the GPER antagonist, G15. To determine the importance of the GPER-ERα balance in regulating vascular remodeling in a rat model of carotid ligation, we studied the effects of upregulation of GPER expression versus downregulation of ERα. Reintroduction of GPER significantly attenuated the extent of medial hypertrophy and attenuated the extent of CD45 labeling. Downregulation of ERα expression comparably attenuated the extent of medial hypertrophy and inflammation after carotid ligation. These studies demonstrate that the balance between GPER and ERα regulates vascular remodeling. Receptor-specific modulation of estrogen's effects may be an important new approach in modifying vascular remodeling in both acute settings like vascular injury and perhaps in longer term regulation like in hypertension. Our initial focus on ERα (versus ER β , which is also expressed in rat aortic vascular smooth muscle cells [VSMCs]) was based on our demonstration that in isolated rat VSMCs, estradiol's (E2) antiapoptotic effects were mediated via ERα. 30 In contrast, E2 acting via GPER was proapoptotic. However, the in vivo implications of these receptor-specific opposing effects of estradiol on growth regulation have yet to be established.In these studies, we examined the process of vascular remodeling to understand how E2 regulates vascular growth via GPER versus ERα. Remodeling occurs subsequent to several vascular stressors including hypertension, 31 atherosclerosis, 32 and vascular injury 33 and can be associated with both adaptive responses (as in vascular injury) and maladaptive responses (as in hypertension and in vascular restenosis). The VSMC hypertrophy and hyperplasia that occurs in remodeling processes has been linked to a shift in the balance between proliferative and apoptotic mechanisms and a dedifferentiation of VSMCs from a contractile phenotype, as seen under normal conditions in situ, to a so-called synthetic phenotype, 34 as seen after vascular injury. This latter phenotype is characterized as having higher proliferative rates and greater migratory behavior. This synthetic phenotype is more similar to the phenotype of VSMCs maintained in primary culture, 35 th...

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“…As observed in other cell types, including human breast carcinoma cell lines and bovine aortic endothelial cells (BAEC; Sousa et al, 2017), in human chondrocytes, the molecular weight of the bands detected by the carboxy-and amino terminal-specific anti-GPR30 antibodies were different (Figure 2a,b). Moreover, the pattern of bands observed in human chondrocytes is different from that F I G U R E 4 Evaluation of the functional state of GPR30 expressed in primary human chondrocytes.…”

Section: Discussionmentioning

confidence: 59%

Expression and function of the nonclassical estrogen receptor, GPR30, in human cartilage and chondrocytes

Ribeiro

Sousa

Rufino

et al. 2020

Journal Cellular Physiology

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Estrogen hormones are important for cartilage homeostasis, but nothing is known regarding the expression and role of the membrane G protein‐coupled estrogen receptor (GPER), G protein‐coupled receptor 30 (GPR30), in adult articular chondrocytes. Using immunohistochemistry of cartilage sections, quantitative real‐time polymerase chain reaction and Western blot of chondrocyte extracts, we found that these cells express GPR30. Nonetheless, the pattern of bands detected by two distinct antibodies does not overlap, suggesting that the proteins detected represent partially degraded forms of the receptor. Treatment with GPR30 agonists did not induce Akt or ERK1/2 phosphorylation, two known GPR30‐activated signaling pathways, suggesting that GPR30 is not functional in human chondrocytes. Therefore, the protective anti‐osteoarthritic role of estrogen hormones in cartilage homeostasis is likely independent of GPR30. This study was performed using human cartilage collected from the distal femoral condyles of multiorgan donors at the Bone and Tissue Bank of the University and Hospital Center of Coimbra.

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Assessment of cell line competence for studies of pharmacological GPR30 modulation

Abstract: Cells that do not express the 44 and 50-55 kDa species do not respond to GPR30 agonists. Thus, the presence or absence of these GPR30 species is a simple and rapid manner to determine whether a given cell line is suitable for pharmacological or molecular studies of GPR30 modulation.

Cited by 10 publications

References 33 publications

The sex steroid precursor dehydroepiandrosterone prevents nonalcoholic steatohepatitis by activating the AMPK pathway mediated by GPR30

The sex steroid precursor dehydroepiandrosterone prevents nonalcoholic steatohepatitis by activating the AMPK pathway mediated by GPR30

Extent of Vascular Remodeling Is Dependent on the Balance Between Estrogen Receptor α and G-Protein–Coupled Estrogen Receptor

Expression and function of the nonclassical estrogen receptor, GPR30, in human cartilage and chondrocytes

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