Marco Volante

Squamous cell carcinoma of the lung compared with other histotypes shows higher messenger RNA and protein levels for thymidylate synthase

In patients with cancer, one of the main mechanism of resistance to antimetabolite drugs is related to higher levels of thymidylate synthase (TS) activity.

Poorly differentiated thyroid carcinoma: the Turin proposal for the use of uniform diagnostic criteria and an algorithmic diagnostic approach.

Poorly differentiated (PD) thyroid carcinomas lie both morphologically and behaviorally between well-differentiated and undifferentiated (anaplastic) carcinomas. Following the original description of this entity, different diagnostic criteria have been employed, resulting in wide discrepancies and confusion among pathologists and clinicians worldwide. To compare lesions occurring in different geographic areas and the diagnostic criteria applied in those countries, we designed a study with a panel of internationally recognized thyroid pathologists to develop consensus diagnostic criteria for PD carcinomas. Eighty-three cases were collected from Europe, Japan, and the United States, and circulated among 12 thyroid pathologists. Diagnoses were made without any knowledge of the clinical parameters, which were subsequently used for survival analysis. A consensus meeting was then held in Turin, Italy, where an agreement was reached concerning the diagnostic criteria for PD carcinoma. These include (1) presence of a solid/trabecular/insular pattern of growth, (2) absence of the conventional nuclear features of papillary carcinoma, and (3) presence of at least one of the following features: convoluted nuclei; mitotic activity ≥3×10 HPF; and tumor necrosis. An algorithmic approach was devised for practical use in the diagnosis of this tumor.

Expression of somatostatin receptor types 1-5 in 81 cases of gastrointestinal and pancreatic endocrine tumors A correlative immunohistochemical and reverse-transcriptase polymerase chain reaction analysis

Abstract. Somatostatin receptors (SSTRs) have been extensively mapped in human tumors by means of autoradiography, reverse-transcriptase polymerase chain reaction (RT-PCR), in situ hybridization (ISH) and immunohistochemistry (IHC). We analyzed the SSTR type 1–5 expression by means of RT-PCR and/or IHC in a series of 81 functioning and non-functioning gastroenteropancreatic (GEP) endocrine tumors and related normal tissues. Moreover, we compared the results with clinical, pathological and hormonal features. Forty-six cases (13 intestinal and 33 pancreatic) were studied for SSTR 1–5 expression using RT-PCR, IHC with antibodies to SSTR types 2, 3, 5 and ISH for SSTR2 mRNA. The vast majority of tumors expressed SSTR types 1, 2, 3 and 5, while SSTR4 was detected in a small minority. Due to the good correlation between RT-PCR and IHC data on SSTR types 2, 3, and 5, thirty-five additional GEP endocrine tumors were studied with IHC alone. Pancreatic insulinomas had an heterogeneous SSTR expression, while 100% of somatostatinomas expressed SSTR5 and 100% gastrinomas and glucagonomas expressed SSTR2. Pre-operative biopsy material showed an overlapping immunoreactivity with that of surgical specimens, suggesting that the SSTR status can be detected in the diagnostic work-up. It is concluded that SSTRs 1–5 are heterogeneously expressed in GEP endocrine tumors and that IHC is a reliable tool to detect SSTR types 2, 3 and 5 in surgical and biopsy specimens.

OBESTATIN PROMOTES SURVIVAL OF PANCREATIC β-CELLS AND HUMAN ISLETS AND INDUCES EXPRESSION OF GENES INVOLVED IN THE REGULATION OF β-CELL MASS AND FUNCTION

OBJECTIVE—Obestatin is a newly discovered peptide encoded by the ghrelin gene whose biological functions are poorly understood. We investigated obestatin effect on survival of β-cells and human pancreatic islets and the underlying signaling pathways. RESEARCH DESIGN AND METHODS—β-Cells and human islets were used to assess obestatin effect on cell proliferation, survival, apoptosis, intracellular signaling, and gene expression. RESULTS—Obestatin showed specific binding on HIT-T15 and INS-1E β-cells, bound to glucagon-like peptide-1 receptor (GLP-1R), and recognized ghrelin binding sites. Obestatin exerted proliferative, survival, and antiapoptotic effects under serum-deprived conditions and interferon-γ/tumor necrosis factor-α/interleukin-1β treatment, particularly at pharmacological concentrations. Ghrelin receptor antagonist [D-Lys3]-growth hormone releasing peptide-6 and anti-ghrelin antibody prevented obestatin-induced survival in β-cells and human islets. β-Cells and islet cells released obestatin, and addition of anti-obestatin antibody reduced their viability. Obestatin increased β-cell cAMP and activated extracellular signal–related kinase 1/2 (ERK1/2) and phosphatidylinositol 3-kinase (PI 3-kinase)/Akt; its antiapoptotic effect was blocked by inhibition of adenylyl cyclase/cAMP/protein kinase A (PKA), PI 3-kinase/Akt, and ERK1/2 signaling. Moreover, obestatin upregulated GLP-1R mRNA and insulin receptor substrate-2 (IRS-2) expression and phosphorylation. The GLP-1R antagonist exendin-(9-39) reduced obestatin effect on β-cell survival. In human islets, obestatin, whose immunoreactivity colocalized with that of ghrelin, promoted cell survival and blocked cytokine-induced apoptosis through cAMP increase and involvement of adenylyl cyclase/cAMP/PKA signaling. Moreover, obestatin 1) induced PI 3-kinase/Akt, ERK1/2, and also cAMP response element–binding protein phosphorylation; 2) stimulated insulin secretion and gene expression; and 3) upregulated GLP-1R, IRS-2, pancreatic and duodenal homeobox-1, and glucokinase mRNA. CONCLUSIONS—These results indicate that obestatin promotes β-cell and human islet cell survival and stimulates the expression of main regulatory β-cell genes, identifying a new role for this peptide within the endocrine pancreas.

Somatostatin receptor type 2A immunohistochemistry in neuroendocrine tumors: a proposal of scoring system correlated with somatostatin receptor scintigraphy

Typing somatostatin receptor expression in neuroendocrine tumors is of relevance to target somatostatin analogue-based diagnostic approach and treatment. The expanding use of immunohistochemistry to detect somatostatin receptors is to date not paralleled by an accurate methodological setting and standardized interpretation of the results. A multicentric study was designed to compare somatostatin receptor immunohistochemical expression with in vivo scintigraphic data and verify its usefulness in the clinical management of neuroendocrine tumors. After methodological setting by testing different somatostatin receptor antibodies, 107 cases of neuroendocrine tumors with available somatostatin receptor scintigraphy data and pathological material were retrospectively analyzed for somatostatin receptor types 2A, 3 and 5 immunohistochemical expression, and compared with scintigraphic images and, whenever available, with the clinical response to somatostatin analogue treatment. Restricting ‘positive cases’ to the presence of a membrane pattern of staining, an overall somatostatin receptor type 2A immunohistochemistry/somatostatin receptor scintigraphy agreement of 77% (χ2 test P<0.0001) was reached. Lower concordance ratios were detected in preoperative and metastatic tumor samples, possibly as a consequence of somatostatin receptor expression heterogeneity. Pure somatostatin receptor type 2A cytoplasmic staining showed poor correlation with somatostatin receptor scintigraphy (54% concordance rate). The immunohistochemical detection of somatostatin receptor types 3 and 5, which showed almost exclusively a cytoplasmic pattern, did not improve the concordance with scintigraphic data. In a pilot series, somatostatin receptor type 2A immunohistochemistry correlated with clinical response in 75% of cases. In conclusion, we propose a scoring system for somatostatin receptor type 2A immunohistochemistry in neuroendocrine tumors correlated with in vivo data, based on the evidence that only membrane (rather that cytoplasmic) staining should be considered for a reliable, standardized and clinically relevant report.

Poorly differentiated carcinomas of the thyroid with trabecular, insular, and solid patterns: a clinicopathologic study of 183 patients.

The term poorly differentiated (PD) carcinoma was proposed 20 years ago to define aggressive, follicular‐derived thyroid carcinomas with behavior intermediate between follicular/papillary and anaplastic carcinomas. Among the variable histologic patterns recognized in such tumors, trabecular‐insular‐solid (TIS) areas usually are predominant. Conversely, some authors pointed out that PD carcinomas are characterized by unequivocal, high‐grade histology with atypias, high mitotic counts, and necrosis rather than by a specific growth pattern.

Ghrelin Expression in Fetal, Infant, and Adult Human Lung:

Ghrelin is a recently identified hormone with potent growth hormone (GH)-releasing activity. It is produced by rat and human gastric endocrine cells and by the pituitary, hypothalamus, placenta, and by gastroenteropancreatic tumors. No evidence of ghrelin production by foregut-derived organs other than stomach has been provided to date. The aim of the present study was to investigate ghrelin expression by human fetal (20 cases), infant (13 cases), and adult (seven cases) lungs by immunohistochemistry, in situ hybridization, and RT-PCR. Expression of the GH secretagogue receptor, the endogenous receptor for ghrelin, was also investigated by RT-PCR. Ghrelin protein was found in the endocrine cells of the fetal lung in decreasing amounts from embryonic to late fetal periods. Its expression was maintained in newborns and children under 2 years but was virtually absent in older individuals. Scattered positive cells were also found in the trachea and the esophagus. Ghrelin mRNA was detected in adult lung by the more sensitive RT-PCR technique. GHS receptor mRNA was detected in nine cases of infant and adult lungs, possibly indicating the existence of local autocrine circuits. We conclude that the fetal lung is an additional source of circulating ghrelin, whose functions at the respiratory tract level remain to be clarified.

RET/PTC activation in hyalinizing trabecular tumors of the thyroid.

Hyalinizing trabecular tumor (HTT) of the thyroid is a neoplasm of follicular derivation with a histogenesis that is still the subject of debate. Morphologic affinities between HTT and papillary carcinoma, including nuclear pseudoinclusions and grooves, suggest that these tumors may be of similar origin. The authors investigated the relationship between these two types of tumors by assessing HTT for the presence of rearrangements of the proto-oncogene rearranged during transfection (RET) that, in thyroid tumors, are specific for papillary carcinoma. A series of 14 HTTs, including two cases associated with classic papillary carcinoma, was studied by means of immunohistochemistry and reverse transcription–polymerase chain reaction. Seven follicular adenomas with focal hyalinized trabecular areas served as control cases. Three of the 14 HTT cases under consideration displayed rearrangements of RET generating the RET/papillary thyroid carcinoma type 1 (PTC1) oncogene. In another case, RET expression was detected focally by immunohistochemistry alone. Finally, in one mixed HTT–papillary carcinoma sample, RET/PTC1 expression was detected, but only in the papillary component. None of the control follicular adenomas contained rearrangements of RET/PTC. These findings demonstrate that a comparable percentage (28.6%) of HTTs and papillary carcinomas exhibit the same RET proto-oncogene alterations. Thus, HTT may represent the “hyalinizing trabecular” variant of papillary carcinoma rather than a separate entity.

Ghrelin in Fetal Thyroid and Follicular Tumors and Cell Lines : Expression and Effects on Tumor Growth

Ghrelin, a growth hormone-releasing hormone produced by gastroenteropancreatic endocrine cells, hypothalamus, and pituitary, was recently identified in medullary thyroid carcinomas and derived cell lines. However, no data exist on its expression in either normal or neoplastic thyroid follicular cells. We analyzed ghrelin expression by immunohistochemistry, in situ hybridization, and reverse transcriptase-polymerase chain reaction in 15 fetal, 4 infant, and 10 adult thyroids, and in 54 tumors of follicular origin. We also analyzed the effects of ghrelin on cell proliferation in N-PAP and ARO thyroid carcinoma cell lines. Ghrelin-binding sites were investigated using reverse transcriptase-polymerase chain reaction to detect its growth hormone secretagogue receptor (GHS-R) mRNA and an in situ-binding localization procedure. Strong ghrelin immunoreactivity was found in fetal but not in infant or adult thyroids. Ghrelin protein and mRNA were present, in variable amounts, in benign and malignant tumors. Normal thyroids, thyroid tumors, and cell lines showed ghrelin binding sites by binding localization, in the absence of the specific GHS receptor mRNA (with the exception of one normal thyroid). Moreover, ghrelin induced dose-dependent inhibition of growth in cell lines. In conclusion, ghrelin is expressed in fetal but not in adult thyroid, and is re-expressed in tumors; the presence of ghrelin receptors other than GHS-R in normal and neoplastic adult thyroid is suggested; ghrelin inhibits cell proliferation of thyroid carcinoma cell lines in vitro.

Retrieved endogenous biotin : a novel marker and a potential pitfall in diagnostic immunohistochemistry

Antigen retrieval (AR) procedures are based on the effect of heating (by either microwave or pressure cooking treatments) on routinely fixed and paraffin embedded tissues. We observed that AR procedures restore the reactivity of endogenous biotin (EB) and report on the distribution of EB following AR in a series of routinely fixed and embedded tissues.