Comparison of immunoreactive score, HER2/neu score and H score for the immunohistochemical evaluation of somatostatin receptors in bronchopulmonary neuroendocrine neoplasms.

AIMS: Due to the growing number of somatostatin receptor (SSTR) targeting analogues and radiopeptides used for the diagnosis and therapy of neuroendocrine neoplasms (NEN), the assessment of SSTR subtype status has increasingly gained predictive value. In pathology, the SSTR protein levels are detected routinely by immunohistochemistry (IHC); however, a lack of a standardized evaluation system persists. Thus, in the present investigation, three well-established semi-quantitative scoring systems [immunoreactive score (IRS), human epidermal growth factor receptor 2 (HER2)/neu score, H score] used commonly for SSTR-IHC evaluation in NEN were compared. METHODS AND RESULTS: A total of 240 formalin-fixed, paraffin-embedded tumour samples from 90 patients with bronchopulmonary NEN were examined by IHC and quantitative reverse transcription-polymerase chain reaction (qRT-PCR) for SSTR1, 2A, 3, 4 and 5 expression. Using both methods, SSTR1, 2A and 5 were the most frequently expressed subtypes. For all SSTR subtypes, all three scores correlated well with each other and with qRT-PCR data. However, the IRS was the most meaningful score with the best correlation to mRNA levels. CONCLUSIONS: Because a unified IHC scoring system for SSTR analysis is needed urgently to optimize the theranostics of NEN, among the scores tested, the IRS seems to be the most suitable according to our results. It provides sufficient accuracy combined with high practicability.

The evolution of vertebrate somatostatin receptors and their gene regions involves extensive chromosomal rearrangements.

BACKGROUND: Somatostatin and its related neuroendocrine peptides have a wide variety of physiological functions that are mediated by five somatostatin receptors with gene names SSTR1-5 in mammals. To resolve their evolution in vertebrates we have investigated the SSTR genes and a large number of adjacent gene families by phylogeny and conserved synteny analyses in a broad range of vertebrate species. RESULTS: We find that the SSTRs form two families that belong to distinct paralogons. We observe not only chromosomal similarities reflecting the paralogy relationships between the SSTR-bearing chromosome regions, but also extensive rearrangements between these regions in teleost fish genomes, including fusions and translocations followed by reshuffling through intrachromosomal rearrangements. These events obscure the paralogy relationships but are still tractable thanks to the many genomes now available. We have identified a previously unrecognized SSTR subtype, SSTR6, previously misidentified as either SSTR1 or SSTR4. CONCLUSIONS: Two ancestral SSTR-bearing chromosome regions were duplicated in the two basal vertebrate tetraploidizations (2R). One of these ancestral SSTR genes generated SSTR2, -3 and -5, the other gave rise to SSTR1, -4 and -6. Subsequently SSTR6 was lost in tetrapods and SSTR4 in teleosts. Our study shows that extensive chromosomal rearrangements have taken place between related chromosome regions in teleosts, but that these events can be resolved by investigating several distantly related species.

Immunohistochemical distribution of somatostatin and somatostatin receptor subtypes (SSTR1-5) in hypothalamus of ApoD knockout mice brain.

In the present study, the expression of somatostatin (SST) and somatostatin receptor subtypes (SSTR1-5) was determined in the hypothalamus of wild-type (wt) and apolipoprotein D knockout (ApoD(-/-)) mice brain. SST-like immunoreactivity, while comparable in most regions of hypothalamus, diminished significantly in arcuate nucleus of ApoD(-/-) mice. SSTR1 strongly localized in all major hypothalamic nuclei as well as in the median eminence and ependyma of the third ventricle of wt mice brain. SSTR1-like immunoreactivity increases in hypothalamus except in paraventricular nucleus of ApoD(-/-) mice. SSTR2 was well expressed in most of the hypothalamic regions whereas it decreases significantly in ventromedial and arcuate nucleus of ApoD(-/-) mice. SSTR3 and SSTR4-like immunoreactivity increases in ApoD(-/-) mice in all major nuclei of hypothalamus, median eminence, and ependymal cells of third ventricle. SSTR5 is well expressed in ventromedial and arcuate nucleus whereas weakly expressed in paraventricular nucleus. In comparison to wt, ApoD(-/-) mice exhibit increased SSTR5-like immunoreactivity in paraventricular nuclei and decreased receptor expression in ventromedial hypothalamus and arcuate nucleus. In conclusion, the changes in hypothalamus of ApoD(-/-) mice may indicate potential role of ApoD in regulation of endocrine functions of somatostatin in a receptor-dependent manner.

Pancreatic neuroendocrine tumors: clinical features, diagnosis and medical treatment: advances.

Pancreatic neuroendocrine tumors (pNETs) comprise with gastrointestinal carcinoids, the main groups of gastrointestinal neuroendocrine tumors (GI-NETs). Although these two groups of GI-NETs share many features including histological aspects; over-/ectopic expression of somatostatin receptors; the ability to ectopically secrete hormones/peptides/amines which can result in distinct functional syndromes; similar approaches used for tumor localization and some aspects of treatment, it is now generally agreed they should be considered separate. They differ in their pathogenesis, hormonal syndromes produced, many aspects of biological behaviour and most important, in their response to certain anti-tumour treatment (chemotherapy, molecular targeted therapies). In this chapter the clinical features of the different types of pNETs will be considered as well as aspects of their diagnosis and medical treatment of the hormone-excess state. Emphasis will be on controversial areas or recent advances. The other aspects of the management of these tumors (surgery, treatment of advanced disease, tumor localization) are not dealt with here, because they are covered in other chapters in this volume.

Colocalization of somatostatin receptors with DARPP-32 in cortex and striatum of rat brain.

Somatostatin (SST)-positive medium-sized aspiny interneurons are selectively spared in excitotoxicity. The biological effects of SST are mediated via five different receptors, namely somatostatin receptor (SSTR)1-5; however, SSTR subtype spared in excitotoxicity and involved in neuroprotection is not known. Dopamine- and cAMP-regulated phosphoprotein (DARPP-32) is predominantly expressed in medium-sized projection neurons that are most vulnerable in excitotoxicity. In the present study, we determined the colocalization of SST and SSTRs with DARPP-32 in rat brain cortical and striatal regions using immunofluorescence immunohistochemistry. We also determined the expression of DARPP-32 in SSTR1-5 immunoprecipitate prepared from cortex and striatum. SST-positive neurons in cortex and striatum are devoid of colocalization with DARPP-32. However, in cortical and striatal brain regions, three different neuronal populations either expressing SSTRs and DARPP-32 alone or displaying colocalization were identified. Quantitative analysis reveals that in cortex and striatum, SSTR1 and 5 are most predominant receptor subtypes colocalized with DARPP-32 followed by SSTR4, 2, and 3 in cortex whereas SSTR2, 4, and 3 in striatum. Importantly, DARPP-32 is expressed in SSTR1-5 immunoprecipitate prepared from cortex and striatum. Taken together, these results provide the first evidence that the SSTR-positive neurons lacking colocalization with DARPP-32 might be spared in excitotoxicity.

(99m)TC-octreotide scintigraphy and somatostatin receptor subtype expression in juvenile nasopharyngeal angiofibromas.

BACKGROUND: The main goal of the study was the analysis of somatostatin receptor (SSTR) expression on juvenile nasopharyngeal angiofibroma (JNA) cells and a subsequent analysis of the utility of SST analog-based scintigraphy in JNA diagnostics. METHODS: Nine JNA cases were analyzed. All tissue samples were analyzed for the expression of SSTRs. In 2 cases, scintigraphy was performed after the intravenous (IV) administration of an SST analog. MRI of the craniofacial region was subsequently performed. RESULTS: The SST analogues were accumulated in areas matching pathologic tissue in the nasopharynx. Immunohistochemical evaluation of the tissue samples proved the overexpression of SSTRs. CONCLUSIONS: SSTRs are overexpressed on JNA cells. The SST analog (99m)TC-octreotide is effectively bound to JNA cells. SST analogues might be used in the diagnostics and treatment of primary, recurrent, or residual JNA.

Differential expression of somatostatin receptor subtype-related genes and proteins in non-functioning and functioning adrenal cortex adenomas.

Adrenocortical adenomas display highly variable expressions of somatostatin receptor (SSTR) subtypes, whose expression is mandatory (although not always sufficient) to achieve the positive effects of somatostatin (SST) analog therapy. Immunohistochemistry (IHC) is the main method used to investigate receptor protein expression. The molecular biology method - polymerase chain reaction (PCR) - is also often used to investigate receptor expression. Nevertheless, the expression of receptor mRNA and the respective receptor protein is not always synchronized. The aim of this study was to investigate SSTR expression by IHC in adrenal adenomas, to compare the results to data obtained by real-time PCR and to determine whether hormonally functioning and non-functioning adenomas differ in this respect. Adrenocortical adenomas were removed surgically from 13 females and 2 males. The tissues were obtained from 9 non-functioning and 6 functioning adenomas. The intensity of IHC reaction was scored semiquantitatively by two independent observers. Real-time PCR was performed using pairs of primers in a reaction amplified along a gradient of temperatures. Amplified DNA was measured by monitoring SYBR-Green fluorescence. In non-functioning tumors, compatibility between IHC and PCR results was observed for SSTR 1 and 2 in 62.5% of the samples. Fifty percent of patients demonstrated compatibility for SSTR 4 and 5 and 37.5% for SSTR 3. In hormonally active adenomas, total compatibility of both methods was noted for SSTR 2 (100%). The compatibility obtained for SSTR 5 was 66.6%. We conclude that receptor gene and respective receptor protein expression are not always synchronized. Messenger RNA detection alone is not sufficient to predict the presence of the receptor protein acting as a target for SST and its analogs.

Immunohistochemical expression and localization of somatostatin receptor subtypes in androgen ablated prostate cancer.

OBJECTIVE: The aim was to examine the expression and localization of the five somatostatin receptors (termed SSTR1 to 5) in radical prostatectomies (RPs) from patients with prostatic adenocarcinoma (PCa) under complete androgen ablation (CAA) before operation. MATERIAL: The five SSTRs were evaluated in the epithelial, smooth muscle and endothelial cells of normal-looking epithelium (Nep), high-grade prostatic intraepithelial neoplasia (HGPIN) and PCa in 20 RPs with clinically detected PCa from patients under CAA. Twenty RPs with clinically detected PCa from hormonally untreated patients were used as control group. RESULTS: Concerning the secretory cells (i) Membrane staining was seen for SSTR3 and SSTR4; the mean percentages of positive cells, higher in SSTR3 than in SSTR4, decreased sharply in HGPIN and PCa compared with Nep; the mean percentages in the androgen ablated group were 30% to 90% lower than in the untreated; (ii) Cytoplasmic staining was seen for all five SSTRs; the mean percentages of positive cells in Nep, HGPIN and PCa of the untreated group were similar, and in general as high as 80% or more; in the treated group, the Nep values were similar to those in the untreated, whereas the values in HGPIN and PCa were lower for SSTR1, three and five, with a decrease of 30% for SSTR1; (iii) Nuclear staining was seen with SSTR4 and SSTR5, the mean percentages for the former being much lower than for the latter; treatment affected both HGPIN and PCa, whose proportions of stained cells were 30% to 55% lower than in the untreated group. Cytoplasmic staining in the basal cells was seen for all five SSTRs, both in Nep and HGPIN. The values in the treated group were lower than in the other, the difference between the two group being in general comprised between 10% and 40%. Treatment did not affect SSTR staining in the smooth muscle and endothelial cells. CONCLUSIONS: The present study expands our knowledge on the expression and localization of the five SSTRs in the prostate following CAA.

Gene expression of somatostatin receptor subtypes SSTR2a, SSTR3 and SSTR5 in peripheral blood of neuroendocrine lung cancer affected patients.

BACKGROUND: Somatostatin (SS) acts as a universal endocrine off-switch, and also inhibits the growth of neuroendocrine tumours through its specific receptors (SSTRs). Somatostatin receptors are G-protein-coupled receptors, which are encoded by five separate genes (SSTR1-5). Short peptide analogues demonstrate specific binding only for the subgroup consisting of SSTR2a, SSTR3 and SSTR5. Moreover, previous studies reported that expression of mRNA for SSTR2a correlated with therapeutic outcome in patients with carcinoid tumours treated with somatostatin analogs. PURPOSE: To develop and apply a Real Time Quantitative PCR technique (RT-qPCR) to compare and contrast the mRNA levels of SSTR2a, SSTR3 and SSTR5 in Neuroendocrine Lung Cancer affected patients. METHODS: Peripheral blood samples from 21 neuroendocrine lung cancer affected patients (14 SCLC, 6 LC and 1 LCNEC) subjected to scintigraphy with (111)In-DTPA-D-Phe(1)-octreotide (OctreoScan) and 24 healthy blood donors were investigated by RT-qPCR. mRNA levels for SSTR2a, SSTR3 and SSTR5 were measured in peripheral blood samples with a relative quantification method using plasmid dilutions as calibration curves and GAPDH as reference gene. RESULTS: A statistically significant increase in target genes/GAPDH copy number ratio was found for SSTR2a (median 38; IQR 22-141) and SSTR5 (median 51; IQR 19-499) in neuroendocrine lung cancer affected patients as compared with samples from healthy blood donors (P ≤ 0.0003 and P ≤ 0.0005). Since low levels of expression were detected in the control group for all three genes, optimal cut-off values were assessed using ROC curve analyses and were equal to 9.05 for SSTR2a and 16.97 for SSTR5. These cut off values resulted in a sensitivity of 86% (95%IC 65-95) for both markers and a specificity of 83% (95%IC 64-93%) and 79% (95%IC 60-91%) for SSTR2a and SSTR5 respectively. Comparison between OctreoScan results and RT-qPCR analysis demonstrated agreement in 76% of the cases. CONCLUSIONS: Our results suggest that SSTR2a and SSTR5 mRNAs are detectable in peripheral blood of neuroendocrine lung cancer affected patients using real-time quantitative PCR, with a good agreement with OctreoScan. The high sensitivity of this non-invasive molecular technique suggests that this method could represent a useful tool in the clinical management of neuroendocrine lung cancers.

Co-expression of somatostatin receptor subtypes and estrogen receptor-α mRNAs by non-functioning pituitary adenomas in young patients.

The expression by non-functioning adenomas (NFoma) of somatostatin receptor (SSTR) subtypes and estrogen receptor (ERα) is poorly understood. Consequently, the mRNAs of SSTR subtypes (SSTR) 1, 2, 3, and 5, dopamine receptor (D2R), and ERα were measured by real-time quantitative RT-PCR in 59 NFomas and 50 functioning adenomas; the latter included 30 GH-secreting adenomas (GHomas) and 20 prolactinomas (PRLomas). NFomas expressed higher levels of SSTR3 than functioning adenomas but had lower levels of SSTR2, SSTR5 and D2R mRNAs than GHomas. Their ERα levels were higher than those of GHomas. The SSTR subtype mRNA levels in NFomas correlated significantly with each other; there was also a good correlation between the SSTR subtypes and ERα in NFomas. These correlations were largely only observed in younger patients (<50 years). The present study describes the differential expression of SSTR subtypes in the largest number of NFoma patients studied thus far, and further proposes possible involvements of SSTR3 and estrogen in the pathophysiology of NFomas.

Somatostatin receptor subtypes in hormone-refractory (castration-resistant) prostatic carcinoma.

The aim of this study was to examine the tissue expression and localisation of the somatostatin receptors (SSTRs) in hormone-refractory (HR) prostate cancer (PCa). Five SSTRs were evaluated immunohistochemically in 20 radical prostatectomies (RPs) with Gleason score (GS) 3+3=6 PCa, in 20 RPs with GS 4+4=8 and 4+5=9 PCa, and 20 transurethral resection of the prostate specimens with HR PCa. The mean values in the cytoplasm (all five SSTRs were expressed), membrane (only SSTR3 and SSTR4 were expressed) and nuclei (only SSTR4 and SSTR5 were expressed) of the glands in HR PCa were 20-70% lower than in the other two groups, the differences being statistically significant. All five SSTRs were expressed in the smooth muscle and endothelial cells of HR PCa, the mean values being lower than in the other two groups. In conclusion, this study expands our knowledge on the expression and localisation of five SSTRs in the various tissue components in the HR PCa compared with hormone-sensitive PCa.

Immunohistochemical expression and localization of somatostatin receptor subtypes in prostate cancer with neuroendocrine differentiation.

The aim of the study is to examine the tissue expression and localization of the somatostatin receptors (SSTRs) in prostate cancer (PCa) with neuroendocrine (NE) differentiation. The five SSTR subtypes (SSTR1 to 5) were evaluated immunohistochemically in the secretory cells of normal-looking epithelium (Nep), high-grade prostatic intraepithelial neoplasia (HGPIN) and PCa in 20 radical prostatectomies (RPs) with Gleason score 3+3=6 acinar PCa; 20 RPs with GS 4+4=8 and 4+5=9 PCa; and 20 RPs with PCa with NE differentiation. The basal cells were evaluated in Nep and HGPIN. In all groups the stromal smooth muscle and endothelial cells were also analyzed. Concerning the secretory cells, (i) the greatest mean proportions of cells with strong cytoplasmic staining in PCa were seen for SSTR2, mainly in the group of RP with NE differentiation, and for SSTR4 in all three groups; the mean values in HGPIN were intermediate between Nep and PCa; (ii) Membrane staining was seen for SSTR3 and SSTR4; the mean percentages of positive cells, higher in SSTR3 than in SSTR4, decreased from Nep to HGPIN and PCa in all three RP groups; in the latter two, the mean percentages were similar; and (iii) Nuclear staining was seen with SSTR4 and SSTR5; for SSTR4, the mean percentages in the PCa of the three groups were higher than in HGPIN and Nep, the highest proportion being with PCa with NE differentiation. Concerning the basal cells, in Nep the mean proportions of cells with strong staining intensity were greater for SSTR1 and SSTR3 than for the other subtypes, the lowest being with SSTR2; in HGPIN the highest mean propositions of positive cells was with SSTR3, the proportions in the three RP groups being similar. Concerning the stromal smooth muscle and endothelial cells, the highest mean values being in SSTR1 and the lowest in SSTR5; for the former subtype the highest proportion of endothelial cells with strong intensity was seen in the RP NE group. In conclusion, this immunohistochemical study expands our knowledge on the expression and localization of five SSTRs in the various tissue components in the prostate with PCa with NE differentiation, compared with conventional PCa. Typing somatostatin receptor expression in NE tumours could be of relevance to target somatostatin analogue-based diagnostic approach and treatment.

Comparative analyses of sequence structure, evolution, and expression of four somatostatin receptors in orange-spotted grouper (Epinephelus coioides).

Somatostatins (SSs) and somatostatin receptors (SSTRs) play important roles in the growth, development and metabolism of vertebrates. In the present study, four SSTRs were isolated from orange-spotted grouper (Epinephelus coioides), a coral fish of high commercial value cultivated in Southeast Asia. Phylogenetic tree analysis grouped the four SSTRs as two distinct groups of SSTR1 and SSTR2/3/5. Four SSTRs exhibited high homology across the vertebrates. The expression of four grouper SSTR mRNAs was studied in 11 tissues. The highest level of SSTR1 mRNA was found in forebrain. The mRNAs of SSTR2 and SSTR3 were highly expressed in pituitary, forebrain and liver. The levels of SSTR5 mRNA were low in most tissues except for pituitary and intestine. The expression of four grouper SSTR mRNAs was investigated in seven embryonic stages and five early larval development stages. The highest levels of SSTR1 and 2 mRNAs appeared during hatching, while the highest levels of SSTR3 and 5 mRNAs were found in brain vesicle stage. Intraperitoneal injection of SS14 significantly increased the levels of all four SSTR mRNAs in pituitary and SSTR1, 3 mRNAs in liver in a dose-dependent manner, but no effect on SSTR2 and 5 in liver. These observations contribute to the understanding of the evolution of SSTR family and offer information on structure, distribution and function of fish SSTRs.

Somatostatin enhances the chemosensitivity of GBC-SD cell line to doxorubicin through arresting the cell cycle to S phase rather than through the P53/Bax-depended apoptosis way in vitro.

BACKGROUND/AIMS: As one of the mostly aggressive and fatal malignancy, gallbladder carcinoma has been known to be resistant to many anticancer drugs. Although it is under active investigation, it is still difficult to achieve satisfactory effect for most chemo-drugs on this tumor. It has previously reported that somatostatin could increase the chemosensitivity of gallbladder carcinoma cells (GBC-SD) and reduce the therapeutic dose of Doxorubicin in killing GBC-SD cells. SST could enhance the chemosensitivity of gallbladder carcinoma to Doxorubincin (DOX) by transient arresting cell cycle to S phase. We tried to clarify the mechanism by which SST utilized to enhance the chemosensitivity of GBC-SD cells to DOX. We further investigated whether the enhanced chemosensitivity of GBC-SD cells to DOX in the presence of SST is via apoptosis or cell cycle regulation. In addition, we also looked into related factors involved in cell cycle regulation and apoptosis. METHODOLOGY: Twenty-four hours after somatostatin treatment, doxorubicin was gradually added and the growth curve of GBC-SD cells was determined according to MTT test. Cell apoptosis was measured by flow cytometry (FCM) using Annexin V/ Propidium Iodide Binding. Cell cycle was also examined by FCM. The somatostatin receptor (SSTR) subtypes in GBC-SD cells were identified using immunocytochemistry and RT-PCR assay. The expressions of p53, Bax and phosphorylated RB (pRB) protein were examined using western blotting assay. RESULTS: When GBC-SD cells were treated with SST alone, no significant cell growth inhibition and cell apoptosis were observed. SST could induce a transient S phase arrest in GBC-SD cells. The mRNA expression of SSTR1, 2, 3, 4, 5 were all detected in GBC-SD cells, whereas only SSTR1, 2, 3 were detected in GBC-SD cells using immunocytochemistry assay. After GBC-SD cells were treated with SST for 24h, the expression level of p53 and Bax protein in GBC-SD cells was similar to that of the control group, however up-regulated pRB protein expression was observed (p < 0.05). CONCLUSIONS: Our results suggested that the synergistic inhibitory effect of somatostatin and doxorubicin co-treatment on GBC-SD cells was not due to SST induced apoptosis concerning the expression of p53 and Bax protein. Our data clearly showed all 5 SST receptor subtypes expressed in GBC-SD cells by RT-PCR and 3 SST receptors by immunocytochemistry. Accumulated evidence has been proved the relationship between cell cycle regulation and RB protein phosphorylation. In the chemosensitized GBC-SD cell line treated with SST, phosphorylated RB and cell cycle arrest were simultaneously manifested. We reasoned that somatostatin might enhance the chemosensitivity of GBC-SD cells to doxorubin through arresting the cell cycle at S phase, but not P53 and Bax protein induced cell apoptosis.

Differential ligand-mediated pituitary somatostatin receptor subtype signaling: implications for corticotroph tumor therapy.

OBJECTIVE: Pituitary targeted pharmacotherapy for Cushing's disease is challenging and ineffective. Unlike octreotide and lanreotide, the multisomatostatin receptor (SST) analog pasireotide that exhibits SST5 greater than SST2 binding affinity offers potential for treating Cushing's disease. Because corticotroph cells express SST5 more abundantly than SST2, pasireotide likely exerts superior corticotroph action mainly through SST5. However, there is no direct evidence for this assumption, and moreover, the ligand effect on corticotroph SST2 is not known. RESULTS: We used AtT20 mouse pituitary corticotroph tumor cells stably overexpressing SST2 or SST5 and TtT/GF mouse pituitary folliculostellate cells stably or transiently expressing SST receptors to examine ligand-receptor activation by SST2- and SST5-selective agonists. We show that pasireotide was more potent than either octreotide or somatostatin-14 in mouse corticotroph cells. Pasireotide potency is not affected by SST2 abundance, SST2 antagonist treatment, or octreotide cotreatment in SST2-overexpressing cells. Pasireotide also does not induce SST2 internalization and attenuates octreotide or SRIF14-induced SST2 internalization only at superphysiological dose ranges. In contrast, octreotide attenuates pasireotide potency in SST5-overexpressing cells. Moreover, short exposure to pasireotide causes prolonged inhibition of forskolin or CRH-induced cAMP accumulation, in contrast to somatostatin-14- and SST2-selective agonists that induced postwithdrawal cAMP rebound. Long-term pasireotide signaling effects are enhanced by SST5 overexpression. CONCLUSION: The results indicate that SST5 determines short- and long-term enhanced pasireotide action in corticotroph cells, whereas the ligand action on SST2 is negligible.

Expression of somatostatin receptor subtypes in human pituitary adenomas -- immunohistochemical studies.

BACKGROUND: The highly variable expression of SSTR subtypes in pituitary adenomas (PA) may partially explain why the subgroup of somatotropinomas or other adenomas do not respond to the therapeutic action of currently used long-acting somatostatin analogues like octreotide or lanreotide. MATERIAL AND METHODS: Our study summarizes the data on expression of all somatostatin receptor subtypes (SSTR 1-5), extended for 2A and 2B SSTR isoforms, revealed by means of immunohistochemistry in dependence to different hormonal phenotype of the tumour. RESULTS: The pattern of SSTR immunostaining (estimated according to the percentage frequency of appearance) was in acromegaly: SSTR 5 > SSTR 1 > SSTR 2A = SSTR 3 > SSTR 2B, in prolactinomas: SSTR 2B = SSTR 3 = SSTR 5 > SSTR 1 = SSTR 2A, in gonadotropinomas: SSTR 3 > SSTR 2B > SSTR 1 = SSTR 2A > SSTR 5, in corticotropinomas: SSTR 2A > SSTR 1 = SSTR 3 > SSTR 5 > SSTR 2B. In PA immunonegative for pituitary hormones, we noticed only a weak staining of all receptor subtypes including SSTR 4. In plurihormonal adenomas with positive GH phenotype the staining pattern was: SSTR 5 > SSTR 1 = SSTR 2B and in plurihormonal PA with negative GH phenotype: SSTR 1 = SSTR 5 > SSTR 2A = SSTR 2B = SSTR 3. In plurihormonal adenoma with ACTH immunopositivity, the staining pattern was: SSTR = SSTR 2A = SSTR 3 = SSTR 5. SSTR 1 and SSTR 5 were the most frequent subtypes of somatostatin receptor in plurihormonal adenomas without ACTH expression. CONCLUSIONS: Human PA represents a group of tumours with a much more differentiated appearance of somatostatin receptor subtypes. It is very important to determine the SSTR profile individually for each tumour to make an appropriate decision as to therapeutic strategy choice. Apart from applying SSTR 2 and SSTR 5-preferring octreotide and lanreotide - newly synthesized multiligand analogues, such as SOM 230, KE 108, or other SST selective analogues, may represent a further useful approach for the treatment, especially in cases other than somatotropinoma or thyrotropinoma.

Somatostatin receptor subtype 4 couples to the M-current to regulate seizures.

The K(+) M-current (I(M), Kv7) is an important regulator of cortical excitability, and mutations in these channels cause a seizure disorder in humans. The neuropeptide somatostatin (SST), which has antiepileptic properties, augments I(M) in hippocampal CA1 pyramidal neurons. We used SST receptor knock-out mice and subtype-selective ligands to investigate the receptor subtype that couples to I(M) and mediates the antiepileptic effects of SST. Using pentylenetetrazole as a chemoconvulsant, SST(2), SST(3), and SST(4) receptor knock-out mice all had shorter latencies to different seizure stages and increased seizure severity when compared with wild-type mice. However, the most robust differences were observed in the SST(4) knock-outs. When seizures were induced by systemic injection of kainate, only SST(4) knock-outs showed an increase in seizure sensitivity. We next examined the action of SST and subtype-selective SST agonists on electrophysiological parameters in hippocampal slices of wild-type and receptor knock-out mice. SST(2) and SST(4) appear to mediate the majority of SST inhibition of epileptiform activity in CA1. SST lacked presynaptic effects in mouse CA1, in contrast to our previous findings in rat. SST increased I(M) in CA1 pyramidal neurons of wild-type and SST(2) knock-out mice, but not SST(4) knock-out mice. Using M-channel blockers, we found that SST(4) coupling to M-channels is critical to its inhibition of epileptiform activity. This is the first demonstration of an endogenous enhancer of I(M) that is important in controlling seizure activity. SST(4) receptors could therefore be an important novel target for developing new antiepileptic and antiepileptogenic drugs.

Immunohistochemical localization of somatostatin receptor subtypes in benign and malignant adrenal tumours.

BACKGROUND: Somatostatin mediates its action through five receptor subtypes (sst1-5) that are widely distributed in various endocrine tissues and tumours. Because of the inhibitory effects of somatostatin, long-acting analogues have been synthesized. In contrast to their well-established use in neuroendocrine and pituitary tumours, little is known about their potential use in adrenal tumours. OBJECTIVE: We examined somatostatin receptor protein expression in adrenal tumours of various aetiologies. Immunostaining was performed with specific polyclonal antibodies for sst1-5. DESIGN: Seven benign and eight malignant pheochromocytomas (PHEOs), eight aldosterone-secreting adenomas (APAs), nine cortisol-secreting adenomas (CPAs), seven nonfunctioning adrenal tumours (NFAs) and 25 adrenal carcinomas (CAs) as well as eight normal adrenal glands were investigated. MEASUREMENTS: Staining pattern, distribution and subcellular localization of the somatostatin receptor subtypes were evaluated. RESULTS: In the majority of normal cortices the expression of sst1-5 was limited to the reticular zone. The medulla was predominantly positive for sst3. Most cortical adenomas were positive for all five subtypes. However, in the majority of these tumours, less than 30% of cells were positively stained. A high expression of sst4 was found in CPAs but only very few cortical carcinomas exhibited sst immunostaining. All benign PHEOs were positive for sst3. The majority presented with more than 60% of tumour cells stained. By contrast, only six out of eight malignant PHEOs were positive for sst3. CONCLUSIONS: Somatostatin receptor subtypes are expressed in PHEOs as well as in tumours of the adrenal cortex with tumour-specific distribution patterns. This may offer new diagnostic and therapeutic possibilities.

Vasculoprotective effects of somatostatin receptor subtypes.

We have shown that somatostatin agonist peptide CH275, selective to somatostatin receptor (sst) subtypes 1,4, was more effective in preventing intimal hyperplasia than the sst2,3,5-selective octreotide, raising the question what are the separate roles of the sst1- and 4-subtypes. Here, we dissect this observation further with highly subtype-selective peptidomimetics and demonstrate that, after rat carotid denudation, both the sst1- and 4-selective analogs (300 microg/kg/day, s.c.) increased lumen size, while only the sst4-selective analog significantly reduced intimal nuclei number, intimal area, and intima/media ratio. The 2,3,5-selective compounds had no effect on these parameters. The observed in vivo effects were further investigated ex vivo with explant outgrowth from pieces of vascular wall. The sst4-selective analog was more effective than the sst1-selective one in inhibiting the percent of outgrowth and the migration of cells from the explants while neither compound affected proliferation. Thus, selective targeting to sst4 should be considered when developing orally active vasculoprotective therapies.