摘要
in studying a relatively well-differentiated human hepatoma cell line (Hep G2),we found that the cells expressed mRNA for PTH-related peptide (PTHrP) and secreted biologically active PTHrP. In the present study, using RT/PCR and Northern analysis,we found that the Hep G2 liver cells also express mRNA for the PTH/PTHrP receptor and exhibit specific binding for radiolabeled N-terminal PTHrP. Therefore, we hypothesized:the cells would respond to endogenously produced peptide. Since PTHrP has been implicated as a potential regulator of cell growth, we asked whether PTHrP might act in an autocrine/paracrine fashion to affect growth of Hep G2 cells. Endogenous PTHrP production by the cells in culture was neutralized by adding aliquots of polyclonal antiserum to either synthetic PTHrP(1-34)or recombinant PTHrP(-5 to 139) to the cultured cells. Both antisera were shown to be capable of inhibiting the ability of conditioned growth medium to stimulate cAMP accumulation in ROS cells in a manner similar to the inhibition produced by the PTHrP antagonist,[Asn10 Leu11,d-Trp12]PTHrP (7-34).When subconfluent Hep G2 cells were exposed to increasing amounts of these two rabbit antisera (1:800￣1:100 dilution in growth medium)for 3 days,a dose-dependent 40%￣ 50% increase in cell growth was observed (vs treatment with nonimmune rabbit serum).The increased cell growth produced by the antisera could be inhibited by concurrent addition of a large concentration (10 μmol/L)of synthetic PTHrP(1-36).The results show that addition of antisera which can neutralize the N-terminal biologic activity of PTHrP caused an enhanced growth of HeP G2 cells in culture which could be inhibited by addition of synthetic N-terminal PTHrP.The findings suggest a possible local regulatory role for PTHrP in liver growth.
in studying a relatively well-differentiated human hepatoma cell line (Hep G2),we found that the cells expressed mRNA for PTH-related peptide (PTHrP) and secreted biologically active PTHrP. In the present study, using RT/PCR and Northern analysis,we found that the Hep G2 liver cells also express mRNA for the PTH/PTHrP receptor and exhibit specific binding for radiolabeled N-terminal PTHrP. Therefore, we hypothesized:the cells would respond to endogenously produced peptide. Since PTHrP has been implicated as a potential regulator of cell growth, we asked whether PTHrP might act in an autocrine/paracrine fashion to affect growth of Hep G2 cells. Endogenous PTHrP production by the cells in culture was neutralized by adding aliquots of polyclonal antiserum to either synthetic PTHrP(1-34)or recombinant PTHrP(-5 to 139) to the cultured cells. Both antisera were shown to be capable of inhibiting the ability of conditioned growth medium to stimulate cAMP accumulation in ROS cells in a manner similar to the inhibition produced by the PTHrP antagonist,[Asn10 Leu11,d-Trp12]PTHrP (7-34).When subconfluent Hep G2 cells were exposed to increasing amounts of these two rabbit antisera (1:800￣1:100 dilution in growth medium)for 3 days,a dose-dependent 40%￣ 50% increase in cell growth was observed (vs treatment with nonimmune rabbit serum).The increased cell growth produced by the antisera could be inhibited by concurrent addition of a large concentration (10 μmol/L)of synthetic PTHrP(1-36).The results show that addition of antisera which can neutralize the N-terminal biologic activity of PTHrP caused an enhanced growth of HeP G2 cells in culture which could be inhibited by addition of synthetic N-terminal PTHrP.The findings suggest a possible local regulatory role for PTHrP in liver growth.
