Approach to PTHrP gene therapy for oral cancer

在先前的临床免疫组化研究中，作者发现口腔鳞状细胞癌中表达甲状旁腺激素相关蛋白（PTHrP），其定位与角化程度较高的区域和组织恶性肿瘤的塔级相对应。本研究旨在探讨PTHrP对口腔鳞状细胞增殖和分化的影响。我们使用了来自舌癌的鳞状癌细胞系SCC-25和来自人类牙龈组织的永生化角化细胞系NDUSD-1。在培养基中加入PTHrP（1-34）、（34-53）和（107-139）片段。据报道，每一个PTHrP片段都有不同的生理功能，如调节平滑肌张力、经上皮钙转运、组织器官发育、分化和增殖。为了评估增殖，我们计算了细胞的数量。利用免疫荧光染色法，用天花素、细胞角蛋白（CK）10、CK14、CK5、CK6和CK18对这些片段影响分化的能力进行评估。我们还评估了细胞内cAMP浓度。细胞数量和免疫荧光染色均未见明显变化。加入片段后，两种细胞系的cAMP浓度均未发生变化。在cDNA芯片研究中，PTHrP片段（1-34）没有增加ndsd -1中CK的表达。虽然在反义RNA研究中有报道称PTHrP影响角质形成细胞的分化，但我们未观察到外源添加PTHrP对所检测细胞系细胞分化和增殖的影响。这些片段在口腔鳞状细胞中的内源性功能有待进一步研究。此外，我们还利用cDNA阵列研究了PTHrP在体外的作用。PTHrP不能降低影响细胞增殖的基因表达。

In previous clinical immunohistochemical studies, the authors found that parathyroid hormone-related protein (PTHrP) was expressed in oral squamous cell carcinoma in which the localization corresponded to regions of higher keratinization and tower grade of histologic malignancy. The purpose of this study was to examine the effect of PTHrP on proliferation and differentiation of oral squamous cells. We used a squamous carcinoma cell line, SCC-25, derived from tongue cancer, and an immortalized keratino cyte cell line, NDUSD-1, derived from human gingival tissue. PTHrP (1-34), (34-53), and (107-139) fragments were added to the culture medium. It has been reported that each of PTHrP fragments has different physiologic functions, such as regulation of smooth muscle tone, transepithelial calcium transport, and tissue and organ development, differentiation, and proliferation. For assessment of proliferation, we counted the number of cells. The ability of the fragments to affect differentiation was evaluated by using immunofluorescent staining with involucrin, cytokeratin (CK)10, CK14, CK5, CK6, and CK18. We also assessed intracellular cAMP concentration. No changes in cell number or immunofluorescence staining were evident in either cell line following stimulation with the fragments. The concentration of cAMP did not change in either cell line after adding the fragments. PTHrP fragment (1-34) did not increase CK expressions in NDUSD-1 in a cDNA microarray study. Although it has been reported that PTHrP affects differentiation of keratinocytes on antisense RNA study, we did not observe any influence of exogenously added PTHrP on cell differentiation and proliferation in the examined cell lines. The endogenous function of the fragments in oral squamous cells requires further study. Furthermore we examined in vitro effects of PTHrP by using cDNA array study. PTHrP could not reduce gene expression affecting cell proliferation.