Lundberg P, Lundgren I, Mukohyama H, Lehenkari P P, Horton M A, Lerner U H
Department of Odontology, Oral Cell Biology, Umeå University, Umeå, Sweden.
Endocrinology. 2001 Jan;142(1):339-47. doi: 10.1210/endo.142.1.7912.
Three distinct complementary DNAs for vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating peptide (PACAP) receptors have been cloned and designated VIP-1 receptor (VIP-1R), VIP-2 receptor (VIP-2R), and PACAP receptor (PACAP-R). In the present study, we have characterized the binding sites on primary mouse calvarial osteoblasts for VIP and related peptides. By analyzing the cAMP response, the rank order of response observed was PACAP 38 > PACAP 27 > helodermin > VIP > helospectin > glucagon > PHI >>> secretin. The VIP-2R/PACAP-R antagonist, PACAP 6-38, inhibited both VIP- and PACAP-stimulated cAMP formation. Binding studies using an atomic force microscopy (AFM) technique showed high affinity binding for VIP and PACAP 38, but not for secretin. Radioligand binding studies using (125)I-VIP and (125)I-PACAP 38 demonstrated a more specific and higher affinity binding for PACAP 38 than for VIP. Secretin failed to inhibit both (125)I-VIP and (125)I-PACAP 38 binding. RT-PCR demonstrated that undifferentiated mouse calvarial osteoblasts express messenger RNA for VIP-2R, but not for VIP-1R or PACAP-R. When the osteoblasts were cultured for 20 days to induce bone noduli formation, VIP-1R, in addition to VIP-2R, were expressed when the nodules started to mineralize at 12 days. Taken together, these data demonstrate that mouse calvarial osteoblasts express functional VIP-2R with higher affinity binding for PACAP than for VIP and that the VIP-1R expression is induced during osteoblastic differentiation.
已克隆出三种不同的血管活性肠肽（VIP）和垂体腺苷酸环化酶激活肽（PACAP）受体互补DNA，并分别命名为VIP-1受体（VIP-1R）、VIP-2受体（VIP-2R）和PACAP受体（PACAP-R）。在本研究中，我们已对原代小鼠颅骨成骨细胞上VIP及相关肽的结合位点进行了特性分析。通过分析环磷酸腺苷（cAMP）反应，观察到的反应强度顺序为：PACAP 38＞PACAP 27＞胃泌酸调节素＞VIP＞蛙皮肽＞胰高血糖素＞PHI（胰高血糖素样肽I）＞促胰液素。VIP-2R/PACAP-R拮抗剂PACAP 6-38可抑制VIP和PACAP刺激的cAMP生成。使用原子力显微镜（AFM）技术进行的结合研究显示，对VIP和PACAP 38具有高亲和力结合，但对促胰液素则不然。使用（125）I-VIP和（125）I-PACAP 38进行的放射性配体结合研究表明，与VIP相比，PACAP 38具有更特异且更高亲和力的结合。促胰液素不能抑制（125）I-VIP和（125）I-PACAP 38的结合。逆转录聚合酶链反应（RT-PCR）表明，未分化的小鼠颅骨成骨细胞表达VIP-2R的信使核糖核酸（mRNA），但不表达VIP-1R或PACAP-R的mRNA。当成骨细胞培养20天以诱导骨结节形成时，在第12天结节开始矿化时，除了VIP-2R外，还表达了VIP-1R。综上所述，这些数据表明，小鼠颅骨成骨细胞表达功能性VIP-2R，对PACAP的亲和力高于对VIP的亲和力，并且在成骨细胞分化过程中诱导VIP-1R表达。
