Expression of Vascular Endothelial Growth Factor and Presence of Angiovascular Cells in Tissues from Different Thyroid Disorders

Background

Vascular endothelial growth factor (VEGF) is involved in tumor angiogenesis and other pathophysiological processes.

Materials and methods

We studied the localization of VEGF in human thyroid tissues to clarify its involvement in proliferative processes in a variety of thyroid disorders. Immunohistochemical analysis using purified rabbit polyclonal anti-human VEGF or anti-human CD34 antibody and a streptavidin–biotin peroxidase complex detection system was performed on 58 tissue specimens from 53 patients with different thyroid disorders and 5 normal thyroid glands.

Results

Vascular endothelial growth factor was not detected in normal thyroid follicular cells. However, some thyroid tumor cells expressed VEGF in the cytoplasm (papillary carcinoma, 10/18; follicular carcinoma, 1/3; medullary carcinoma, 2/2; follicular adenoma, 3/11; adenomatous goiter, 2/4). In benign follicular adenoma and adenomatous goiter, weak expression of VEGF was found in small areas of the tumor, whereas in malignant thyroid tumors, it was strongly expressed in many cells. However, VEGF was not expressed in anaplastic carcinoma, malignant lymphoma, or Graves’ disease. Angiovascular cells stained with CD34 antibody in tissues from different thyroid disorders reflected statistically significant differences in papillary carcinoma, follicular adenoma, and Graves’ disease compared with normal thyroids, and such cells showed a trend toward increases in medullary carcinoma and adenomatous goiter. In contrast, low vascularity was observed in anaplastic carcinoma, malignant lymphoma, and follicular carcinoma.

Conclusions

Because VEGF probably functions as a hypoxia-inducible angiogenic factor, overexpression of this mediator, concomitant with hypervascularity, may be induced more strongly in malignant thyroid tumors, which need more oxygen to proliferate, than in benign follicular tumors. However, neither VEGF nor CD34 was expressed in anaplastic thyroid carcinoma, which is an extremely poorly differentiated malignant tumor. CD34 but not VEGF was expressed in the hyperplastic thyroid tissues of Graves’ disease composed of nontransformed cells. Thus, the expression of VEGF concomitant with CD34 is suggested to reflect both the transformation and differentiation state of malignant tumors.     

生长抑素对人结肠癌裸鼠移植瘤的作用及机理

建立人结肠癌细胞株ＳＷ４８０裸鼠移植瘤模型,观察８肽生长抑素（ＳＭＳ２０１－９９５,ＳＭＳ）对移植瘤的及机理。结果：ＳＭＳ组及ＳＭＳ＋ＰＧ（５肽胃泌素）组移植瘤的体积、重量、瘤细胞内ＣＡＭＰ、ＤＮＡ、蛋白质含量Ｓ期和Ｇ２Ｍ期ＰＧ组显著低于对照组。本实验结果提示：凶素具有直接换制人结肠癌株ＳＷ４８０移植瘤生长的作用,又可抑制胃泌素对移植瘤的促增殖作用,这种作用机理是通过抑制瘤细胞内ＣＡＭＰ、ＤＮＡ和     

Classification, staging, prognostic factors and risk factors in thyroid carcinoma

Thyroid carcinomas arise from follicular cells (papillary, follicular, Hurthle, anaplastic), parafollicular cells (medullary) and stroma (lymphoma, sarcoma). Gradation and prognostic factors are different for every one of histological type. Most patients with papillary and follicular thyroid cancer have an excellent prognosis. At the other extreme is anaplastic thyroid cancer whose usual mean survival can be measured in months. Exposure to external radiation and living in endemic goiter area increase the frequency of thyroid cancer. Medullary thyroid carcinoma is often familial and may occur in associations with the multiple endocrine neoplasia syndromes.     

甲状旁腺素对人甲状腺髓样癌细胞增殖和凋亡的影响

目的探讨甲状旁腺素对人甲状腺髓样癌细胞体外增殖抑制及凋亡作用。方法体外培养甲状腺髓样癌细胞株,经甲状旁腺素和甲状旁腺素受体单抗干预处理后,倒置相差显微镜下观察细胞生长状况,流式细胞仪检测细胞凋亡。结果倒置相差显微镜下细胞变化明显,各浓度的甲状旁腺素和甲状旁腺素受体单抗均分别能有效地抑制甲状腺髓样癌细胞增殖、诱导细胞凋亡,凋亡作用呈时间和浓度依赖。当甲状旁腺素浓度为2．0μmol／L、甲状旁腺素受体单抗浓度为1．0μmol／L时,对细胞凋亡作用显著（P〈0．05）,凋亡率分别为13．24％及20．78％。结论甲状旁腺素对人甲状腺髓样癌细胞增殖有抑制作用并能诱导其凋亡。     

The helix-loop-helix protein,Id-1, is overexpressed and regulates growth in papillary thyroid cancer

BACKGROUND: Members of the Id helix-loop-helix proteins are key regulators of cell growth and differentiation in a variety of cell types. They are also required for cell cycle progression and regulate tumor angiogenesis. Furthermore, deregulated expression of Id proteins has been observed in some human malignancies. Therefore we hypothesized that the Id-1 gene may play a role in papillary thyroid carcinogenesis. METHODS: Id-1 gene expression was characterized by Northern blot analysis and Id-1 immunohistochemistry in human thyroid tissue. Id-1 gene expression and regulation was evaluated in a human papillary thyroid cancer cell line, TPC-1, by Northern blot analysis. RESULTS: The Id-1 gene was significantly overexpressed in papillary thyroid cancer compared with normal thyroid tissue from the same patients (n = 18) by both Northern blot analysis and semiquantitative Id-1 immunohistochemistry (P <.001). Id-1 immunoreactivity was primarily localized to the cytoplasm of the thyroid follicular cells. In the TPC-1 cell line, stimulation by TSH and serum up-regulated Id-1 mRNA expression 1.5- and 4.0-fold, respectively. Activation of the mitogen intracellular protein kinase A and protein kinase C signaling pathways also up-regulated Id-1 mRNA expression. Inhibition of Id-1 mRNA expression with Id-1 antisense oligonucleotide inhibited growth compared with control Id-1 sense and random oligonucleotides (P <.05). CONCLUSIONS: The Id-1 gene is overexpressed in papillary thyroid cancer. Id-1 may play a role in the regulation of growth in papillary thyroid cancer.     

Valproic acid inhibits growth, induces apoptosis, and modulates apoptosis-regulatory and differentiation gene expression in human thyroid cancer cells

BACKGROUND: Among the most promising new therapies for thyroid cancer are the histone deacetylase inhibitors. Valproic acid (VA) is an anticonvulsant that inhibits histone deacetylase activity at nontoxic concentrations. We hypothesized that VA would have antineoplastic effects on human thyroid cancer cells. METHODS: We treated 1 papillary and 3 follicular thyroid cancer cell lines with VA (0.5-2 mmol/L) for 24 to 72 hours. Cell proliferation was measured with a cell proliferation assay kit. Annexin V-fluorescein isothiocyanate was used to quantitate cells that were undergoing apoptosis. Quantitative polymerase chain reaction was used to measure expression of apoptosis-regulatory and differentiation genes. RESULTS: VA inhibited growth in all cell lines by 26% to 59% at 48 hours and up to 77% at 72 hours. Nineteen percent to 30% of VA-treated cells underwent apoptosis, compared with 4% to 8% of the control cells. Expression of pro survival genes bcl-2 and bcl-xl was down-regulated by 10% to 60%; expression of the proapoptosis gene bax was up-regulated by 23% to 85%. Sodium-iodide symporter and thyroglobulin messenger RNA expression were up-regulated by 93% to 370% in follicular cell lines but remained unchanged in the papillary cell line. CONCLUSION: VA inhibits growth, induces apoptosis, and modulates apoptosis-regulatory and differentiation gene expression in thyroid cancer cells. These findings suggest that VA may be useful clinically for patients with thyroid cancers of follicular cell origin.     

Medullary carcinoma of the thyroid gland. Clinical, pathological, and immunohistochemical features with review of the literature

Twenty medullary carcinomas of the thyroid gland were examined for the presence of immunoreactive calcitonin, thyroglobulin, glucagon, keratin, gastrin/CCK, carcinoembryonic antibody (CEA), insulin, serotonin, adreno-corticotropic hormone (ACTH), prostatic acid phosphatase, and somatostatin using the immunoperoxidase peroxidase-antiperoxidase technique. In addition, they were stained with mucicarmine, alcian blue/periodic acid-Schiff (PAS), Grimelius, Congo red, crystal violet, and Fontana-Masson stains. Calcitonin-immunoreactive cells were absent in one tumor and present in 19 tumors (95%). Thyroglobulin was present in seven tumors (35%). Twenty tumors contained CEA-immunoreactive cells (100%). Fourteen cases were immunoreactive to serotonin (70%) and 12 were positive for somatostatin (60%). Glucagon- and gastrin/CCK-immunoreactive cells were found in two cases each (10%). Four tumors (20%) contained ACTH-immunoreactive cells and three cases (15%) were positive for prostatic acid phosphatase. Five cases (25%) contained keratin-immunoreactive cells. One case was immunoreactive to insulin (5%). Grimelius-positive cells were present in 19 of the cases (95%). Mucin-containing cells were present in 65% of the cases. The validity of the immunocytochemical localizations was tested by specific absorption of each antibody with the corresponding antigen. The demonstration of immunoreactivity for multiple antigens in each of the 20 cases suggests that the origin of medullary thyroid carcinomas is from a neuroendocrine cell potentially capable of producing numerous hormone substances. In addition, as the neoplastic cells in 35% of the tumors contained hormonal substances as well as thyroglobulin, it is suggested that papillary or follicular tumors mixed with a neuroendocrine component exist more commonly than previously suspected. Finally, psammoma bodies might be present in pure medullary carcinoma of the thyroid gland.     

A population-based analysis of survival factors in differentiated and medullary thyroid carcinoma.

OBJECTIVE: The study purpose was to determine survival and prognostic factors for differentiated thyroid carcinoma (DTC). METHODS: Cases of DTC were extracted from the Surveillance, Epidemiology and End Results database from 1988 through 1998. Kaplan-Meier survival analysis was conducted for papillary, follicular, and medullary histologies. Cox proportional hazard analysis was used to examine the influence of age, gender, tumor size, local extension, and cervical node involvement on overall survival. RESULTS: A total of 18,118 cases were identified, including 15,820 (87.3%) papillary carcinomas, 1799 (9.9%) follicular carcinomas, and 499 (2.8%) medullary carcinomas. Mean survival (10-year survival) was 122 (87.7%), 117 (80.2%), and 108 (73.7%) months for papillary, follicular, and medullary tumors, respectively. For each histology, increasing age, male gender, and degree of local extension substantially reduced survival. Cervical metastasis did not influence survival for papillary or follicular carcinomas but approached significance for medullary carcinoma (P = 0.065). CONCLUSIONS: Degree of local extension in thyroid carcinoma should be subclassified to more accurately determine prognosis. Treatment of the neck should be considered for medullary thyroid carcinoma.     

Simultaneous medullary and papillary thyroid carcinoma with lymph node metastasis in the same patient: case report and review of the literature

We report a rare case of simultaneous medullary thyroid carcinoma on the left thyroid lobe with lymph node metastasis and papillary thyroid cancer on the right thyroid lobe. The 55-year-old woman was diagnosed with medullary thyroid carcinoma after left hemithyroidectomy for goitre. Completion thyroidectomy, central neck dissection and left modified neck dissection revealed the presence of papillary carcinoma on the right side. The extreme rarity and interesting pathological features are discussed and we raise the question of whether the finding of another thyroid cancer on the opposite side was coincidental or from possible activation of a common tumorigenic pathway for both follicular and parafollicular thyroid cells.     

Measurement of cellular DNA content in thyroid tumors by the flow cytometer

Cellular DNA contents measured by flow cytometer were analysed in relation to histopathological classification and clinicopathological findings in 94 patients with thyroid tumors. The DNA determination was carried out on both tumor tissues and surrounding thyroid tissues. As an indicator of tumor growth, proliferative index (PI) and DNA index were calculated from DNA histograms. The PI value (mean +/- SD) was 32.5 in medullary carcinoma, 31.3 +/- 10.2 in follicular carcinoma, 28.2 +/- 6.2 in papillary carcinoma, 21.6 +/- 4.4 in follicular adenoma, and 20.6 +/- 4.4 in adenomatous goiter, respectively, whereas the value in normal thyroid tissues was 4.1 +/- 2.2. PI values in the surrounding thyroid tissues in cases of follicular and papillary carcinomas were significantly (p less than 0.01) lower than those of the corresponding cancer tissues, but they were higher than that of the normal tissues. The DNA index and frequencies of aneuploidy were 1.15 and 50.0% in medullary carcinoma; 1.25 +/- 0.27 and 66.7% in follicular carcinoma; 1.19 +/- 0.25 and 64.2% in papillary carcinoma; 1.01 +/- 0.04 and 9.3% in follicular adenoma; and 1.00 +/- 0.00% in adenomatous goiter. The result implies that PI value and DNA index are relatively correlated with clinicopathological criteria of malignancy of individual thyroid tumors, and they may become a putative tool for determination of the biological malignancy.     

A twenty-year experience with thyroid carcinoma in children

During the past 20 years, 23 patients (7 males, 16 females) were operated on for thyroid carcinoma in our institution. The average age was 13.6 years (range, 22 months to 27 years). Our series includes papillary carcinoma in 11, follicular carcinoma in four, and medullary thyroid carcinoma in eight patients. Follow-up ranged from 8 months to 20.3 years, with an average of 7.5 years for well-differentiated carcinomas and 4.3 years for medullary thyroid carcinomas. All patients are presently alive with no evidence of progressive disease. Patients with papillary and follicular carcinomas underwent partial thyroidectomy; those with medullary carcinoma underwent total thyroidectomy. Serious complications included three permanent hypoparathyroidism and two tracheostomies, all after secondary neck explorations. The overall results observed in our series of patients seem to support the current conservative approach to well-differentiated thyroid carcinoma, reserving total thyroidectomy for medullary cancer of the thyroid. A more aggressive search for familial medullary carcinoma through use of pentagastrin stimulation leads to early detection and more effective therapy.     

A case with poorly differentiated follicular carcinoma of the thyroid produced and secreted CEA

Detection of thyroglobulin, CEA, and calcitonin in serum and tissue is very useful to make differential diagnosis among papillary, follicular, and medullary carcinoma of the thyroid. The case was a 70-year-old woman with a nodule of the thyroid, presenting elevated serum level of thyroglobulin and CEA (22.0 ng/ml). Both serum level of thyroglobulin and CEA decreased after surgery. One year and nine months after surgery, she died from systemic metastases of thyroid carcinoma. Histological examination of the tumor presented to be poorly differentiated type of follicular carcinoma, presenting positive of both thyroglobulin and CEA, and negative of calcitonin in the cytoimmunochemistry. This case was identified to be a rare case that tumor cells of poorly differentiated follicular carcinoma produced and secreted both thyroglobulin and CEA.     

Thyroid disorders. Part III: neoplastic thyroid disease.

This paper is part III of the series on thyroid disorders. Thyroid tumors are the most common endocrine neoplasms. Most of these tumors are benign hyperplastic or colloid nodules or benign follicular adenomas. However, 5% to 10% of the lesions that come to medical attention are carcinomas. A major clinical challenge is establishing which nodules are hyperplastic, benign, or malignant. History, clinical findings, ultrasonography, and fine-needle aspiration biopsy are the mainstays for diagnosis. There are 3 main histologic types of thyroid cancer: differentiated, medullary, and anaplastic. Differentiated lesions are subdivided into papillary, follicular, and Hurthle cell carcinomas. In addition, primary lymphoma may occur in the thyroid gland and other cancers may metastasize to the thyroid. An important neoplastic syndrome, multiple endocrine neoplasia type 2 (MEN2), involves medullary carcinoma of the thyroid gland. In 2002 there were 10 cases of thyroid cancer per 100 000 population. During the past 10 years the rate of thyroid cancer has been increasing 5% per year. The overall 10-year survival for papillary carcinoma is 80% to 90%, follicular carcinoma 65% to 75%, and medullary carcinoma 60% to 70%. The prognosis for anaplastic carcinoma is very poor and 5-year survival is rare. The dentist by inspection and palpation of the neck in the area of the thyroid gland may detect single or multiple lesions that may be benign or malignant. Patients with identified nodules or enlarged thyroid glands should be referred for diagnosis and treatment. Patients with thyroid cancer will benefit from the early detection and treatment of their lesions as early detection can lead to a cure or prolongation of their life.     

Cancer gene therapy using a replication-competent herpes simplex virus type 1 vector.

OBJECTIVE: The authors investigate the efficacy of hrR3, a viral vector derived from herpes simplex virus type 1 (HSV 1), in destroying colon carcinoma cells in vitro and in vivo. The effect of adding the prodrug ganciclovir in combination with hrR3 infection also is assessed. SUMMARY BACKGROUND DATA: Most cancer gene therapy strategies use viral vectors that are incapable of replication. The HSV 1 vector hrR3 is capable of replication, and its replication is cytotoxic to cells. hrR3 also possesses the HSV-thymidine kinase gene, which converts ganciclovir into a toxic metabolite. Thus, the addition of ganciclovir to hrR3-infected cells may enhance the ability of hrR3 to destroy tumor cells. To increase specificity for tumor cells, hrR3 has a mutated ribonucleotide reductase gene and replicates selectively in cells with high levels of endogenous rbonucleotide reductase. Actively dividing cells such as tumor cells have high levels of endogenous ribonucleotide reductase for synthesis of DNA precursors. The authors are interested in the use of HSV 1 vectors to treat liver metastases from colorectal cancer. METHODS: Ribonucleotide reductase expression in several colon carcinoma cell lines and in primary cultures of human hepatocytes was determined by Western blot analysis. hrR3-mediated cytotoxicity in the colon carcinoma cell lines was determined using an in vitro assay. The human colon carcinoma cell line HT29 was injected into the flanks of nude mice followed by intratumoral injection of hrR3. Tumor growth rate was assessed with and without the addition of intraperitoneal ganciclovir. RESULTS: Ribonucleotide reductase levels in colon carcinoma cell lines are much higher than in primary cultures of human hepatocytes. hrR3 efficiently destroys colon carcinoma cell lines in vitro. A single intratumoral injection of hrR3 into HT29 flank tumors significantly reduces tumor growth rate, and the administration of ganciclovir has no additive effect. CONCLUSIONS: The inherent cytotoxicity of hrR3 replication effectively destroys colon carcinoma cells in vitro and in vivo. This cytotoxicity is not enhanced in vivo by the addition of ganciclovir. In the future, more efficacious and selective HSV 1 vectors may be useful in the treatment of cancer.     

Flow cytometric DNA analysis of thyroid carcinoma

Abnormal DNA content has been considered as an additional criterion for determining the biological behavior of a tumor. Flowcytometric DNA analysis was done on 121 patients with thyroid carcinoma encountered during the period between 1975 and 1987. Tumor tissues were sampled from paraffin-embedded blocks and the histology of thyroid carcinoma found to consist of 91 papillary, 23 follicular, 2 medullary, 1 squamous cell and 4 anaplastic carcinomas. The incidence of aneuploidy in thyroid carcinoma was 7.4 per cent (9 patients) while that of diploidy was 92.6 per cent (112 patients). The aneuploid specimens consisted of 6 papillary, 1 follicular, 1 medullary and 1 anaplastic carcinomas and, of 4 anaplastic carcinoma patients with subsequent death within 6 months, only 1 was aneuploid. As an indicator of proliferative potential, S-phase fraction (SPF) was also determined by flow cytometry, but this could not be used as an independent prognostic factor. The aneuploid patients showed a significantly decreased survival rate (p<0.01). Thus, although DNA measurement proved useful for predicting the survival of aneuploid patients, there is some discrepancy between DNA content and the biological behavior of the tumor.     

Phenylacetate enhances the antiproliferative effect of retinoic acid in follicular thyroid cancer.

BACKGROUND: Retinoic acid (RA) has antiproliferative as well as redifferentiating effects in thyroid cancers. Similar effects have been seen with phenylacetate (PA) therapy. These observations prompted us to evaluate the potential antiproliferative effects of the combination of RA and PA in follicular thyroid cancer. METHODS: Three follicular cell lines were treated in vitro with varying concentrations of all-trans RA or PA alone or in combination. Growth was measured by dimethyl-thiazol-diphenyltetrazolium bromide assays. RESULTS: RA (1-2.5 micromol/L) and PA (1-10 mmol/L) alone inhibited cell growth in a time- and dose-dependent manner, with maximum effect at 5 days. The combination of RA and PA had synergistic antiproliferative effects. In the FTC-133 cell line, RA alone (2.5 micromol/L) inhibited growth 16% and PA alone (10 mmol/L) inhibited growth 35% versus controls, whereas the combination of the 2 inhibited growth by 60% at 5 days (P < .005). Similar results were seen with FTC-236 and FTC-238 cell lines. CONCLUSIONS: Our results support that RA and PA have antiproliferative effects in follicular thyroid cancer and are synergistic. The combination of RA and PA may be beneficial in the treatment of advanced thyroid cancers for which conventional therapy fails or as an adjuvant to radioactive iodine therapy in aggressive tumors.     

Surgical treatment of thyroid cancer

Surgery is the treatment of choice for thyroid cancer, often followed by I131 and thyroid hormone to control the local residual tumor and distant metastasis. Hundred and sixty-two patients with thyroid disease underwent surgery at the Division of Surgical Oncology of the Cancer Institute in Genoa. Thirty cases presented no malignant hot thyroid nodules, 37 of the other 132 cases were cancers (28%). In 23 cases (62%), the pathological diagnosis was papillary carcinoma, in ten cases (27%) follicular carcinoma, in four (11%) medullary carcinoma. In 13 cases (35%) (ten papillary carcinoma, two follicular carcinoma, one medullary carcinoma) the lesion was multicentric. Our data suggest that total thyroidectomy, performed in two steps, in most cases does not carry important post-operative morbidity but offers the greatest potential for cure. In our cases only one patient died (of the cancer). After radical surgery it is possible to detect and treat metastases by I131.     

Flow cytometric DNA analysis of thyroid carcinoma

Abnormal DNA content has been considered as an additional criterion for determining the biological behavior of a tumor. Flowcytometric DNA analysis was done on 121 patients with thyroid carcinoma encountered during the period between 1975 and 1987. Tumor tissues were sampled from paraffin-embedded blocks and the histology of thyroid carcinoma found to consist of 91 papillary, 23 follicular, 2 medullary, 1 squamous cell and 4 anaplastic carcinomas. The incidence of aneuploidy in thyroid carcinoma was 7.4 per cent (9 patients) while that of diploidy was 92.6 per cent (112 patients). The aneuploid specimens consisted of 6 papillary, 1 follicular, 1 medullary and 1 anaplastic carcinomas and, of 4 anaplastic carcinoma patients with subsequent death within 6 months, only 1 was aneuploid. As an indicator of proliferative potential, S-phase fraction (SPF) was also determined by flow cytometry, but this could not be used as an independent prognostic factor. The aneuploid patients showed a significantly decreased survival rate (p less than 0.01). Thus, although DNA measurement proved useful for predicting the survival of aneuploid patients, there is some discrepancy between DNA content and the biological behavior of the tumor.     

Fine needle aspiration biopsy of thyroid nodules

The clinical value of the fine needle aspiration of thyroid nodules was evaluated by comparing preoperative cytology to subsequent pathology in 109 patients undergoing thyroidectomy. Preoperative cytology was reported as insufficient cellular material (31 patients), benign goiter (27 patients), follicular neoplasm (22 patients), thyroiditis (12 patients), suspicious for papillary carcinoma (nine patients), Hurthle cell neoplasm (five patients), medullary carcinoma (one patient), lymphoma (one patient), and metastatic adenocarcinoma (one patient). Operative findings demonstrated that the overall sensitivity of fine needle aspiration in diagnosing thyroid neoplasia (carcinoma or adenoma) was 88% and its specificity was 80%. Operation verified the cytologic diagnosis of medullary carcinoma, lymphoma, metastatic adenocarcinoma, and seven of nine papillary carcinomas. Of the five patients with an aspiration biopsy diagnosis of Hurthle cell neoplasm, three patients had carcinoma and one had an adenoma. Four carcinomas and 12 follicular adenomas were found in patients with a cytologic diagnosis of follicular neoplasm. Thyroiditis was confirmed at operation in all 12 patients with this diagnosis on fine needle aspiration. One carcinoma was found in the 27 patients with benign goiter diagnosed on cytology. Fine needle aspiration is a valuable tool that can lead to earlier diagnosis and treatment of thyroid cancer. However, a negative aspiration does not supplant good clinical judgement in determining the need for thyroidectomy.