Expression of sodium iodide symporter in benign and malignant human thyroid tissues

The extent of human sodium iodide symporter (hNIS) expression in different kinds of human thyroid cancer tissues and cell lines remains controversial. In this study, polyclonal antibodies to hNIS were used to analyze the expression of symporter protein in benign and malignant human thyroid tissues. Formalin-fixed, paraffin wax-embedded tissue sections were used. Staining was performed using primary polyclonal antibody of rabbit anti-human hNIS diluted in PBS (1:500). Results showed that 2 of 3 normal tissue, 3 of nodular hyperplasia, one follicular adenoma, 3 of 11 papillary thyroid carcinoma, 1 of 5 follicular carcinoma and none of 3 metastatic thyroid epithelial tissue specimens stained positively for hNIS. A higher percentage of positive staining for symporter protein was found in benign thyroid tissues including normal thyroid tissue, nodular hyperplasia, and adenoma (60%). In contrast, papillary and follicular thyroid carcinomas demonstrated lower symporter protein expression (20%). In conclusion, although the number of tissue samples examined in this study was small, hNIS staining found a higher ratio of symporter protein expression in normal and benign thyroid tissues compared with malignant tissues. Determination of the reason for discrepancies in the expression of hNIS in in vivo and in vitro studies will require further investigation.     

Expression of the sodium iodide symporter and thyroglobulin genes are reduced in papillary thyroid cancer.

Altered expression of the gene encoding the sodium iodine symporter (NIS) may be an important factor that leads to the reduced iodine accumulation characteristic of most benign and malignant thyroid nodules. Both up- and down-regulation of NIS gene expression have been reported in thyroid cancer using several different methods. The goal of the present study was to accurately identify alterations in NIS gene expression in benign and malignant thyroid nodules using an accurate real-time quantitative RT-PCR assay system. Total RNA was prepared from 18 benign thyroid nodules, 20 papillary thyroid cancers, and 23 normal thyroid samples from 38 subjects. Quantitative RT-PCR was used to measure NIS and thyroglobulin (TG) mRNA expression in normal thyroid tissue and in each nodular tissue sample. Papillary thyroid cancer samples had significantly lower NIS mRNA expression (72 +/- 41 picogram equivalents [pg Eq]), than did benign nodules (829 +/- 385 pg Eq), or normal tissues (1907 +/- 868 pg Eq, P = 0.04). Most important, in the paired samples, NIS gene expression was decreased in each papillary thyroid cancer compared with normal tissue (69% median decrease; range, 40-96%; P = .013). Eleven of the 12 benign nodules also demonstrated lower NIS gene expression than the normal tissue (49% decrease; range, 2-96%; P = .04). Analysis of the paired samples demonstrated that Tg mRNA expression was significantly lower in each of the thyroid cancer samples than in corresponding normal tissue (759 +/- 245 pg Eq vs. 1854 +/- 542 pg Eq, P = .03). We have demonstrated a significant decrement in NIS gene expression in all papillary thyroid cancers and in over 90% of benign nodules examined compared with adjacent normal thyroid tissue, using a highly accurate quantitative RT-PCR technique. Similarly, thyroid cancers demonstrated significantly lower TG mRNA expression than corresponding normal thyroid. Reduced NIS expression may be an important factor in the impairment of iodine-concentrating ability of neoplastic thyroid tissues.     

HLA class II antigen expression and the autoimmune thyroid response in patients with benign and malignant thyroid tumors

To further understand the relationship between the immune system and the neoplastic human thyroid cell we investigated the degree of intrathyroidal lymphocytic infiltration and thyroid HLA class II expression in 17 patients with thyroid tumors. In another 60 thyroid tumor patients the association of thyroidal lymphocytic infiltration with thyroid autoantibody production was analyzed. In total 117 thyroid tissues were examined including tissue obtained at autopsy (n = 28), fetal thyroid tissue (n = 4), thyroid samples obtained from areas distant from benign follicular adenomas (n = 5), and 80 abnormal thyroids including patients with benign (n = 53) or malignant (n = 24) thyroid tumors and Hashimoto's thyroiditis (n = 3). Normal adult and fetal thyroid tissue had no significant lymphocytic infiltration and no detectable HLA-DR, -DP, or -DQ antigens on their thyroid follicular epithelial cells. The degree of lymphocytic infiltration in the nonneoplastic thyroid tissue of thyroid glands with benign and malignant thyroid tumors varied considerably and correlated with the presence and titer of serum thyroid autoantibodies measured by sensitive ELISA techniques. However, all but one of the benign follicular adenomas had thyroid cells negative for HLA class II determinants despite the presence of infiltrating lymphocytes, while 7 of 10 thyroid carcinomas expressed class II antigen (principally HLA-DR) even when only minor degrees of lymphocytic infiltration were present. These data indicate a correlation between lymphocytic infiltrates and serum thyroid autoantibody titers but the relationship with HLA class II expression is more complex. Since we have previously shown that HLA class II antigen expression can be induced by local interferon-gamma secretion, presumably from activated T cells, we conclude that estimates of simple thyroid lymphocytic infiltration and serum autoantibody secretion do not correlate with the degree of intrathyroidal T-cell activation. Furthermore, our observation of increased expression of HLA class II antigens in thyroid cancer suggests considerable cellular heterogeneity in susceptibility to HLA class II antigen induction in human thyroid disease.     

Differential nuclear and cytoplasmic expression of PTEN in normal thyroid tissue, and benign and malignant epithelial thyroid tumors

Germline mutations in PTEN (MMAC1/TEP1) are found in patients with Cowden syndrome, a familial cancer syndrome which is characterized by a high risk of breast and thyroid neoplasia. Although somatic intragenic PTEN mutations have rarely been found in benign and malignant sporadic thyroid tumors, loss of heterozygosity (LOH) has been reported in up to one fourth of follicular thyroid adenomas (FAs) and carcinomas. In this study, we examined PTEN expression in 139 sporadic nonmedullary thyroid tumors (55 FA, 27 follicular thyroid carcinomas, 35 papillary thyroid carcinomas, and 22 undifferentiated thyroid carcinomas) using immunohistochemistry and correlated this to the results of LOH studies. Normal follicular thyroid cells showed a strong to moderate nuclear or nuclear membrane signal although the cytoplasmic staining was less strong. In FAs the neoplastic nuclei had less intense PTEN staining, although the cytoplasmic PTEN-staining intensity did not differ significantly from that observed in normal follicular cells. In thyroid carcinomas as a group, nuclear PTEN immunostaining was mostly weak in comparison with normal thyroid follicular cells and FAs. The cytoplasmic staining was more intense than the nuclear staining in 35 to 49% of carcinomas, depending on the histological type. Among 81 informative tumors assessed for LOH, there seemed to be an associative trend between decreased nuclear and cytoplasmic staining and 10q23 LOH (P = 0.003, P = 0.008, respectively). These data support a role for PTEN in the pathogenesis of follicular thyroid tumors.     

The correlation of sodium iodide symporter and BRAFV600E mutation in classical variant papillary thyroid carcinoma

BRAF^V600E mutation was analyzed by real-time polymerase chain reaction in 96 consecutive cases with classical variant papillary thyroid cancer, and immunohistochemical staining of Na +/I − symporter (NIS) protein was evaluated. Localization (intracellular or membranous), density, and the intensity of cytoplasmic staining were characterized semiquantitatively. Extrathyroidal invasion, surgical margin positivity, and lymph node metastasis were compared with BRAF^V600E mutation and NIS expression. Eighty-eight patients who had at least 24-month follow-up were also included in survival analysis. BRAF^V600E mutation was determined in 78.1% (75/96) and functional NIS activity in 74% (71/96) of the cases. There were statistically significant differences in mean ages between BRAF^V600E mutation–positive (48.6) and BRAF^V600E mutation–negative cases (37.3; Levene test, P = .419; Student t test, P = .001). The surgical margin positivity (46.7%) and extrathyroidal extension percentage (54.7%) in the BRAF^V600E mutation–positive group were higher than the negative (28.6% and 33.3%, respectively) group, without statistical significance (P = .138 and P = .084, respectively). Functional NIS activity was higher in BRAF^V600E mutation–positive cases (78.1%) than mutation-negative ones (57.1%; P = .047). The possibility of moderate and intense cytoplasmic staining in BRAF^V600E mutation–positive cases (72%) was 6.3 times higher than the possibility of weak staining (28%) in the mutation-positive cases (95% confidence interval, 2.2-18.8; P = .001). Functional NIS expression is higher in patients with classical variant papillary thyroid cancer with BRAF^V600E mutation. However, the clinical features were not found to be associated with NIS expression. There may be different mechanisms determining the outcome of therapy.     

Galectin-3、CK19和TPO表达在甲状腺良恶性肿瘤病理诊断中的价值

目的比较甲状腺病变中galectin-3、CK19和甲状腺过氧化物酶（thyroid peroxidase,TPO）蛋白表达的差异,寻找有助于诊断良恶性肿瘤的标志物。方法采用免疫组化EnVision二步法检测134例甲状腺癌（123例乳头状癌、11例滤泡癌）、34例甲状腺腺瘤、20例结节性甲状腺肿和10例桥本甲状腺炎中galectin-3、CK19和TPO蛋白的表达。结果Galectin-3、CK19和TPO在甲状腺乳头状癌（包括主要的3个亚型）与良性病变中的表达差异有显著性（P〈0．01）,在乳头状癌各亚型间和各良性病变之间的表达差异均无显著性（P〉0．05）。滤泡癌和良性病变中galectin-3表达差异有显著性（P〈0．01）。滤泡癌和腺瘤中CK19、TPO的表达差异有显著性（P〈0．05,P〈0．01）。结论甲状腺恶性肿瘤特别是乳头状癌中galectin-3和CK19的表达有增高,而TPO表达缺失。3种蛋白分别是诊断甲状腺肿瘤特别是乳头状癌的有用标志物,联合使用结果更可靠。     

Immunohistochemical expression of galectin-3 in benign and malignant thyroid lesions

CONTEXT: The expression of galectin-3, a human lectin, has been shown to be highly associated with malignant behavior of thyroid lesions. DESIGN: We studied the immunohistochemical expression pattern of galectin-3 in a variety of follicular-derived thyroid lesions (13 benign and 62 malignant), including Hürthle cell and follicular carcinoma, papillary carcinomas and variants, and anaplastic and poorly differentiated carcinomas. RESULTS: Immunoreactivity was strongest in papillary thyroid carcinomas, whereas staining was less intense in Hürthle cell and anaplastic carcinomas, and even weaker in the follicular variant of papillary thyroid carcinoma. Staining was absent or weak in the 3 follicular thyroid carcinomas and was negative in both insular carcinomas. In several tumors, staining was stronger at the advancing invasive edge of the lesion than in the central portion of the tumor. Galectin-3 was also expressed focally and weakly in reactive follicular epithelium and entrapped follicles in chronic lymphocytic thyroiditis. A variety of thyroid lesions showed prominent endogenous, biotin-like activity, which could cause flaws in interpretation if a biotin-detection system were used. CONCLUSION: We conclude that galectin-3 immunostaining, when used in biotin-free detection systems, may be useful as an adjunct to distinguish benign from malignant thyroid lesions.     

Sodium/iodide symporter expression in primary lung cancer and comparison with glucose transporter 1 expression

The aim of the present study was to evaluate the expression of sodium/iodide symporter (NIS) and glucose transporter 1 (Glut1) in 139 primary lung cancers on immunohistochemistry, and to determine the diagnostic utility of NIS as an imaging reporter. Immunoreactivity for NIS and Glut1 was noted in 75 (54.0%) and 72 (51.8%) of the 139 cases, respectively. Analysis of NIS expression on Western blot confirmed the immunohistochemistry. NIS expression was significantly higher in the adenocarcinomas than in the other carcinomas, and Glut1 expression was significantly higher in the squamous cell carcinomas than in the other carcinomas (each P  < 0.0001). The frequency of NIS expression in those carcinomas lacking Glut1 expression was significantly higher than in those with Glut1 expression ( P  = 0.012). Among 64 adenocarcinomas, the frequency of the NIS(+)/Glut1(−) phenotype was 61.0%, which was the most frequent expression pattern. By studying the expression pattern of NIS in lung cancer, the present paper provides a helpful foundation for examining the potential utility of NIS-mediated radioiodide as an alternative diagnostic modality, especially for the management of patients with lung adenocarcinoma lacking Glut1 expression.     

hTPO和hNIS共转染提高人胶质瘤、肺癌及甲状腺癌细胞摄放射性碘能力

目的 研究共转染人甲状腺过氧化物酶基因（hTPO）及人钠碘转运体基因（hNIS）后胶质瘤、肺癌及甲状腺癌细胞的摄碘能力变化。方法 克隆,重组,包装并扩增纯化得到重组腺病毒（AdTPO）,测定病毒滴度。使用脂质体转染法将hNIS基因转染入人胶质瘤、肺癌及甲状腺癌细胞系中,经过G418抗生素筛选获得稳定表达hNIS的细胞系,只转染hNIS基因的细胞系为对照组;使用重组腺病毒将hTPO基因转导入对照组中,共转染hNIS及hTPO基因的细胞系为实验组;未转入hTPO和hNIS基因的原始肿瘤细胞系为空白对照组。然后进行3组细胞系的体外摄125I 率及各细胞系的125I外流实验。结果 在各肿瘤细胞系中,实验组细胞的摄碘率和有效半衰期,较对照组细胞和空白对照组细胞均有所提高,实验组细胞的摄碘率：H460组为59628 1281,U251组为7968 1261,ARO组为52971 2162,FRO组为49638 1281,三组间总体具有统计学差异（P<0.01）。结论 将hTPO和hNIS基因共转染至肿瘤后,能有效提高肿瘤的摄碘能力并延长细胞内碘滞留时间。     

Evaluation of coxsackievirus and adenovirus receptor expression in human benign and malignant thyroid lesions

Giaginis C, Zarros A, Alexandrou P, Klijanienko J, Delladetsima I, Theocharis S. Evaluation of coxsackievirus and adenovirus receptor expression in human benign and malignant thyroid lesions. APMIS 2010; 118: 210–21. Coxsackievirus and adenovirus receptor (CAR) expression on tumor cells is associated with sensitivity to adenoviral infection, being considered as a surrogate marker for monitoring and/or predicting adenovirus‐mediated gene therapy. The aim of this study was to evaluate the clinical significance of CAR expression in human benign and malignant thyroid lesions. CAR protein expression was assessed immunohistochemically on paraffin‐embedded thyroid tissues from 107 patients with benign and malignant lesions and was statistically analyzed in relation to histopathologic type; tumor size; lymph node metastasis; capsular, lymphatic and vessel invasion; as well as follicular cells’ proliferative capacity. CAR immunoreactivity was characterized as negative/weak in 53 (49.53%), moderate in 31 (28.97%) and strong in 23 (21.50%) of 107 thyroid cases. CAR immunoreactivity was significantly increased in malignant compared with that in benign thyroid lesions (p = 0.00002). Both malignant and benign thyroid lesions with enhanced follicular cells’ proliferative capacity showed significantly increased CAR immunoreactivity (p = 0.00027). In malignant thyroid lesions, enhanced CAR immunoreactivity was significantly associated with larger tumor size (p = 0.0067). The current data revealed that CAR immunoreactivity could be considered of diagnostic utility in thyroid neoplasia. Further research effort is warranted to delineate whether CAR could be considered clinically important for both diagnosis and future (gene) therapeutic applications in thyroid neoplasia.     

Comparison of metabolic ratios of urinary estrogens between benign and malignant thyroid tumors in postmenopausal women

Background

Estrogen metabolism may be associated with the pathophysiological development of papillary thyroid carcinoma (PTC).

Methods

To evaluate the differential estrogen metabolism between benign and malignant PTCs, estrogen profiling by gas chromatography–mass spectrometry was applied to urine samples from postmenopausal patients with 9 benign tumors and 18 malignant stage I and III/IV PTCs.

Results

The urinary concentration of 2-methoxyestradiol was significantly lower in the stage I malignant patients (3.5-fold; P?<?0.025) than in the benign group. The metabolic ratios of 16α-OH-estrone/estrone and estriol/estradiol, which are responsible for 16α-hydroxylase activity, were increased more than 2.5-fold in the advanced-stage malignant PTC (P?<?0.02 each). The more than 6.2-fold decrease in the urinary 2-/16α-hydroxylase ratio in stage III/IV malignant PTC was consistent with the ratio in postmenopausal patients with endocrine gland cancers. In addition, reductive 17β-hydroxysteroid dehydrogenase (17β-HSD; estradiol/estrone or estriol/16α-OH-estrone) was present at significantly higher levels in subjects with stage III/IV malignant PTCs than in benign subjects (>3.5-fold difference; P?<?0.002). In particular, the estriol/16α-OH-estrone ratio differentiated between the benign and early-stage malignant patients (P?<?0.01).

Conclusions

Increased 16α-hydroxylation and/or a decreased 2-/16α-ratio, as well increased reductive 17β-HSD, with regard to estrogen metabolism could provide potential biomarkers. The devised profiles could be useful for differentiating malignant thyroid carcinomas from benign adenomas in postmenopausal women.

     

Genetic and epigenetic alterations of 9p21 gene products in benign and malignant tumors of the head and neck

The multistep process of tumorigenesis has not been decoded to date, although numerous investigations into probable molecular changes have meanwhile been conducted. However, not only DNA changes or loss of alleles cause deregulation of gene function, but also epigenetic alterations (e.g. methylation) result in functional loss. The INK4a-ARF (CDKN2A) locus, located on chromosome 9p21, encodes two functionally distinct tumor suppressor genes, p14ARF and p16INK4a, which play active roles in the p53 and Rb tumor suppressive pathways. We therefore examined not only p16 and p14 proteins, but also alterations of the INK4a-ARF locus, including methylation and loss of heterozygosity in benign and malignant tumors of the head and neck (squamous cell carcinomas and pleomorphic adenomas). In benign pleomorphic adenomas, methylation of p14ARF was found in 1 out of 42 (2%) cases, whereas alterations of p16INK4a occurred in 12/42 (29%) pleomorphic adenomas. In HNSCC, methylation of p16INK4a occurred in 16 out of 50 (32%) carcinomas. P14ARF was found to be methylated in 8 out of 50 cases (16%). Our results demonstrate that alterations of the INK4a-ARF locus are frequent and important events not only in the carcinogenesis of malignant, but also in benign tumors.     

Androgen metabolism in malignant and benign bone tumors

The comparison of activities of the key enzymes of androgen metabolism in morphological variants of bone sarcomas and benign tumors suggests that malignant tumors of different histogenesis not only metabolize the main androgen testosterone, but also inactivate 5α-dihydrotestosterone, the main regulatory androgen in bones. It is possible that androgen metabolism in bone tumors is aimed at the formation of other androgens, in particular, 3α-and 5β-diols, which can be involved in regulatory processes in bone tissue. Further studies will disclose clinical significance of androgen metabolism and individual androgens in human bone tumors. Translated fromByulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 129, No. 1, pp. 90–92, January, 2000     

A transporter gene (sodium iodide symporter) for dual purposes in gene therapy: imaging and therapy

Radioiodide uptake (RAIU) in thyroid follicular epithelial cells, mediated by a plasma membrane transporter, sodium iodide symporter (NIS), provides a first step mechanism for thyroid cancer detection by radioiodide injection and effective radioiodide treatment for patients with invasive, recurrent, and/or metastatic thyroid cancers after total thyroidectomy. NIS gene transfer to tumor cells may significantly and specifically enhance internal radioactive accumulation of tumors following radioiodide administration, and result in better tumor control. NIS gene transfers have been successfully performed in a variety of tumor animal models by either plasmid-mediated transfection or virus (adenovirus or retrovirus)-mediated gene delivery. These animal models include nude mice xenografted with human melanoma, glioma, breast cancer or prostate cancer, rats with subcutaneous thyroid tumor implantation, as well as the rat intracranial glioma model. In these animal models, non-invasive imaging of in vivo tumors by gamma camera scintigraphy after radioiodide or technetium injection has been performed successfully, suggesting that the NIS can serve as an imaging reporter gene for gene therapy trials. In addition, the tumor killing effects of 131I after NIS gene transfer have been demonstrated in in vitro clonogenic assays and in vivo radioiodide therapy studies, suggesting that NIS gene can also serve as a therapeutic agent when combined with radioiodide injection. Better NIS-mediated tumor treatment by radioiodide requires a more efficient and specific system of gene delivery with better retention of radioiodide in tumor. Results thus far are, however, promising, and suggest that NIS gene transfer followed by radioiodide treatment will allow non-invasive in vivo imaging to assess the outcome of gene therapy and provide a therapeutic strategy for a variety of human cancers.     

Sodium/iodide‐symporter: distribution in different mammals and role in entero‐thyroid circulation of iodide

The sodium (Na⁺)/iodide (I^–)‐symporter (NIS) is abundantly expressed and accumulates iodide in thyroid follicular cells. The NIS is also found in extrathyroidal tissues, particularly gastric mucosa. Controversies exist on the localization of extrathyroidal NIS. We have studied the presence of both NIS peptide and NIS messenger RNA (mRNA) in the digestive tract and thyroid from different mammals. The role of gastric NIS is enigmatic and we aimed to unravel its possible involvement in iodide transport. Methods: Distribution and expression of NIS were studied using immunocytochemistry and in situ hybridization. Iodide transport in the gastrointestinal tract was measured after oral or intravenous (i.v.) administration of ¹²⁵I to rats with or without ligation of the pylorus. Results: All thyroid follicular cells in rat and mouse expressed NIS, whereas a patchy staining was noted in man, pig and guinea‐pig. Gastric mucosa surface epithelium in all species and ductal cells of parotid gland in guinea‐pig, rat and mouse expressed NIS. In parietal cells and in endocrine cells of intestines and pancreas NIS immunoreactivity but no NIS mRNA was found. Studies of ¹²⁵I uptake showed marked iodide transport from the circulation into the gastric lumen. Conclusions: The localization of NIS varies slightly among mammals. To establish expression of NIS in a particular cell type the need to correlate the presence of both NIS protein by immunocytochemistry and NIS mRNA by in situ hybridization is emphasized. An entero‐thyroidal circulation of iodide mediated principally by gastric NIS, but possibly also by NIS in salivary glands is suggested.     

乳腺良恶性肿瘤新生血管超微结构及血管生成相关分子的表达

目的 探讨乳腺良恶性不同肿瘤在新生血管超微结构及其血管生成相关分子表达方面的差异性.方法 应用透射电镜观察乳腺良恶性肿瘤新生血管超微结构改变,免疫组化技术检测CD34、VEGF及其受体Flk-1/KDR在两组肿瘤中的表达特性.结果 恶性组新生血管内皮细胞紧密连接开放,基膜不连续,缺乏平滑肌成分.内皮细胞胞体大,细胞核大,畸形,核仁增大、边集,核质比例增大,胞质内吞饮泡多.较多的单个内皮细胞呈裂隙状,血管腔闭塞或明显狭窄.恶性组MVD高于良性组(P<0.05),微血管丰富区位于痛巢边缘.VEGF在乳腺癌性上皮细胞及癌周血管内皮细胞呈强阳性表达,Flk-1/KDR在乳腺恶性肿瘤m管内皮细胞旱强阳性表达,VEGF及Flk一1/KDR尤其在癌灶边缘呈强阳性表达,良性组几乎不表达(P<0.05).结论 乳腺癌新生血管内皮细胞在超微结构及分子表达上具有异质性,VEGF或受体Flk-1/KDR可能是乳腺癌早期诊断及治疗的分子靶标,癌灶边缘可能是下一步进行乳腺癌分子影像观察的重点靶区.