Approach to PTHrP gene therapy for oral cancer

在以往的临床免疫组化研究中，作者发现甲状旁腺相关蛋白（PTHrP）在口腔鳞状细胞癌中表达，其定位对应于高度角化和组织学恶性程度较低的区域。本研究旨在探讨PTHrP对口腔鳞状细胞增殖和分化的影响。我们使用的鳞状细胞癌细胞系，SCC-25，来自舌癌，和永生化角质细胞系，NDUSD-1，来自人牙龈组织。将PTHrP（1-34）、（34-53）和（107-139）片段加入培养基中。据报道，每个PTHrP片段具有不同的生理功能，例如调节平滑肌张力、跨上皮钙转运以及组织和器官发育、分化和增殖。为了评估增殖，我们计数了细胞的数量。通过使用外皮蛋白、细胞角蛋白（CK）10、CK 14、CK 5、CK 6和CK 18进行免疫荧光染色，评价片段影响分化的能力。我们还评估了细胞内cAMP浓度。在用片段刺激后，在任一细胞系中细胞数量或免疫荧光染色没有明显变化。在加入片段后，两种细胞系中cAMP的浓度均未发生变化。PTHrP片段（1-34）对NDUSD-1细胞CK表达无明显影响。虽然已经报道PTHrP在反义RNA研究中影响角质形成细胞的分化，但是我们没有观察到外源性添加的PTHrP对所检查的细胞系中的细胞分化和增殖的任何影响。这些片段在口腔鳞状细胞中的内源性功能需要进一步研究。此外，我们研究了PTHrP的体外作用，通过使用cDNA阵列研究。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.