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• W1573598806 abstract "OBJECTIVE Both in vivo and in vitro evidence indicates that primary hyperparathyroidism is characterized by a reduced sensitivity to extracellular calcium ([Ca 2+ ] o ). The existence of alterations in the expression and signalling of calcium sensing receptor (CaSR) in parathyroid neoplasia is still uncertain. In order to clarify the role of CaSR in the reduced [Ca 2+ ] o sensing of parathyroid neoplasia we investigated PTH secretion and intracellular effectors triggered by CaSR activation as well as the levels of expression of CaSR and CaSR coupled G proteins (Gq/G 11 ) in parathyroid adenomas and primary hyperplasia. MATERIALS AND METHODS The study included 27 parathyroid adenomas, 4 cases of primary hyperplasia and pools of normal parathyroid biopsies. Tissues were either snap frozen in liquid nitrogen or placed in sterile medium for cell dispersion. The effects of increasing [Ca 2+ ] o on in vitro PTH release, intracellular cAMP levels and intracellular calcium ([Ca 2+ ] i ) in cells loaded with the Ca 2 + indicator fura‐2 were evaluated. CaSR mRNA levels were assessed by semiquantitative RT‐PCR analysis, using GAPDH as internal standard, while CaSR protein was detected by western blot analysis using a specific polyclonal antibody. Purified antisera selective for G 11 α and Gqα were used to detect this class of proteins. RESULTS In basal conditions (at 0.5 m m [Ca 2+ ] o ) in vitro PTH released ranged from 29.4 to 1186 pg/well/60 minutes. Increasing [Ca 2+ ] o from 0.5 to 1, 2.5 and 5 m m caused a variable effect. One group ( n = 7) showed a significant but partial reduction of PTH release (of 17 to 60% of basal levels) that occurred at physiological [Ca 2+ ] o concentrations (1 m m ) while the remainder showed either inhibition detectable only at 2.5 m m ( n = 15) or total ( n = 9) resistance to [Ca 2+ ] o . In the responsive cells, [Ca 2+ ] o (1–5 m m ) caused a pertussis toxin‐insensitive [Ca 2+ ] i rise (ranging from 10% to 260%), due to Ca 2+ release from intracellular stores, and an inhibition of forskolin‐stimulated cAMP levels. By RT‐PCR almost all tumours tested showed a substantial reduction in CaSR mRNA levels when compared to the normal tissue (CaSR/GAPDH ratio: 3.1 ± 0.5 vs . 15.5 ± 3.1; P < 0.001), which was confirmed by immunoblotting analysis demonstrating low levels of CaSR protein in tumour tissues. Moreover, low amounts of G 11 α and Gqα, the G proteins involved in CaSR coupling, were observed in the majority of pathological tissues. CONCLUSIONS The study shows that the activation of the calcium sensing receptors expressed in adenomatous parathyroid glands modulates intracellular effectors in a similar way to those operating in the normal parathyroid. Although a reduction of calcium sensing receptor expression is probably involved in the poor inhibition of PTH release induced by [Ca 2+ ] o, this is not the only factor altering [Ca 2+ ] o sensing in parathyroid adenomas, since tumours characterized by different in vitro sensitivity to [Ca 2+ ] o showed similar CaSR levels. The low content of G proteins of the Gq subfamily might represent an additional alteration leading to a defective [Ca 2+ ] o sensing." @default.
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• W1573598806 date "2000-03-01" @default.
• W1573598806 modified "2023-10-17" @default.
• W1573598806 title "Calcium-sensing receptor expression and signalling in human parathyroid adenomas and primary hyperplasia" @default.
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• W1573598806 doi "https://doi.org/10.1046/j.1365-2265.2000.00933.x" @default.
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