Figure 5

Expression of the CaR decreases PTH mRNA levels and confers responsivity to a low [Ca ²⁺ ] _o and the calcimimetic R568. A. Representative Northern blot analysis for PTH mRNA in cells transiently transfected in triplicate with hPTH and either the CaR plasmid or empty vector. After transfection cells were grown in 1.2 or 0.2 mM Ca²⁺ medium for an additional 48 h. Ethidium bromide staining of the membrane is shown as loading control. B. qRT PCR for PTH corrected for co-transfected GH mRNAs for cells treated as in A. C. qPCR for PTH and HPRT mRNAs from cells expressing the hPTH gene and the CaR (CaR+) or empty vector (CaR-) in cells grown in 1.2 mM Ca²⁺ supplemented with either R568 or vehicle. The results in A-C represent one of three repeat experiments performed in triplicate with similar results. D. Representative Northern blot for PTH and GFP mRNA levels in cells transfected with PTH and GFP and either CaR (CaR+) or empty vector (CaR-). The cells were grown in 0.2 mM calcium supplemented with R568 or vehicle. Quantification of the results is shown below the gel. The results are presented as mean ± SE of two repeat experiments performed in triplicate. *, P < 0.05, CaR+:CaR-; **, P < 0.05, CaR+, treated:CaR+, untreated.