Critical phosphate salt concentrations leading to altered micellar casein structures and functional intermediates.

We investigated the effect of different phosphate salts on the structural integrity of micellar casein (MC) at pH 7.0. With the increase of salt concentration, a reduction in turbidity was observed for the MC solutions, and it was modeled using an exponential decay function. The inflection point of the model was defined as the first critical salt concentration (C*), and it is suggested that the salt concentration initiates the disintegration of MC. For linear polyphosphates, C* decreased with the number of phosphate groups. Apparent viscosity (ηapp) of MC solutions increased with the increase of salt concentration, and they recorded a peak while the turbidity decreased to a minimum. The salt concentration that resulted in the highest ηapp was identified as the second critical salt concentration (C**). It is hypothesized that the interactions among protein species present in the mixtures are at an optimum state at C**. Both C* and C** were found to be dependent on the MC concentration. The work presented herein supports an understanding of the concentration effect of phosphate salts on MC for structuring dairy products.

The effect of emulsifying salts on the turbidity of a diluted milk system with varying pH and protein concentration.

Solutions of 10 commonly used emulsifying salts (ES) listed in the Code of Federal Regulations (21CFR133.179) for pasteurized process cheese were tested for their effect on the turbidity of a diluted milk system at different pH and protein concentrations to characterize the conditions that affect micellar structure. Emulsifying salt solutions were made by mixing the ES in a 1-in-20 dilution of water in skim milk ultrafiltrate (3 kDa molecular weight cut-off) to obtain ES concentrations from 0 to 248 mM. Skim milk was added to solutions containing nanopure water, skim milk ultrafiltrate, and a specific ES ranging in concentration from 0 to 248 mM and pH 5, 5.8, 6.8, 7.8, and 8.8. The turbidity of the samples was measured as the optical density at 400 nm immediately after mixing (time, t = 0), after 30 s (t = 30s), and after 30 min (t = 30min). Emulsifying salts were found to cause a decrease in the turbidity of the system, which was modeled using an exponential decay model, where C* represents a threshold salt concentration at which rapid dissociation occurs. At pH values 5.8 and 6.8, the ES caused the greatest decrease in turbidity of the diluted milk system. At pH 5, the ES had the least effect on the turbidity of the system. Sodium hexametaphosphate was found to have the strongest dissociative effect, with a C* value of 0.33 mM for t = 0 at pH 6.8. In contrast, the largest C* value calculated at pH 6.8 was monosodium phosphate at 278.22 mM. Increased time resulted in lower C* values. The model established for this study can be used to predict the dissociation of casein micelles in the presence of various types of ES.

Characterization and sub-cellular localization of SS1R, SS2R, and SS5R in human late-stage prostate cancer cells: effect of mono- and bi-specific somatostatin analogs on cell growth.

Somatostatin (SST) and SST receptors (SS1R, SS2R, SS3R, SS4R and SS5R) appear to play a significant role in the progression of human prostate cancer (PCa), which is associated with heterogeneity of SSRs expression and specific cell localization as we already demonstrated in the LNCaP cell line, an in vitro model of human androgen-dependent PCa. In this study, PC-3 and DU-145 human castration-resistant PCa cells were found to express all SSRs, while LNCaP expressed all but SS4R. A 48-h treatment with BIM-23244 (SS2R/SS5R) or BIM-23926 (SS1R) SST analogs was more effective in inhibiting cell proliferation, compared to BIM-23120 (SS2R), BIM-23206 (SS5R) and BIM-23704 (SS1R/SS2R). BIM-23926 (SS1R) treatment increased the amount of p21 and decreased phosphorylated (p) ERK1/2. BIM-23244 (SS2R/SS5R) led to p21 increment only in PC-3 cells, and to pERK1/2 reduction in both cell lines. SS1R/SS2R and SS2R/SS5R receptor dimers were natively present on cell membrane and their amount was increased by BIM-23704 (SS1R/SS2R) or BIM-23244 (SS2R/SS5R) treatment, respectively. SS1R, SS2R and SS5R were differently distributed among nuclear, lysosomal and microsomal compartment, according to their different recycling dynamics. These results show that, in PC-3, DU-145 and LNCaP cells, activation of SS1R and SS2R/SS5R leads to relevant antiproliferative effects.

Somatostatin dramatically stimulates growth hormone release from primate somatotrophs acting at low doses via somatostatin receptor 5 and cyclic AMP.

Somatostatin and cortistatin have been shown to act directly on pituitary somatotrophs to inhibit growth hormone (GH) release. However, previous results from nonprimate species indicate that these peptides can also directly stimulate GH secretion, at low concentrations. The relevance of this phenomenon in a nonhuman primate model was investigated in the present study by testing the impact of somatostatin/cortistatin on GH release in primary pituitary cell cultures from baboons. High doses (> 10(-10) m) of somatostatin/cortistatin did not alter basal GH secretion but blocked GH-releasing hormone (GHRH)- and ghrelin-induced GH release. However, at low concentrations (10(-17)-10(-13) m), somatostatin/cortistatin dramatically stimulated GH release to levels comparable to those evoked by GHRH or ghrelin. Use of somatostatin receptor (sst) specific agonists/antagonists, and signal transduction blockers indicated that sst2 and sst1 activation via intact adenylate cylcase and mitogen-activated protein kinase systems mediated the inhibitory actions of high-concentration somatostatin. By contrast, the stimulatory actions of low-dose somatostatin on GH release were mediated by sst5 signalling through adenylate cylcase/cAMP/protein kinase A and intracellular Ca(2+) pathways, and were additive with ghrelin (not GHRH). Notably, low-concentrations of somatostatin, similar to sst5-agonists, inhibited prolactin release. These results clearly demonstrate that the ultimate impact of somatostatin/cortistatin on hormone release is dose-dependent, cell type-selective and receptor-specific, where the stimulatory effects of low-concentration somatostatin/cortistatin on GH release extend to primates, thereby supporting the notion that this action is relevant in regulating GH secretion in humans.

Somatostatin-dopamine chimeras: a novel approach to treatment of neuroendocrine tumors.

A combination of basic research observations concerning the interaction of somatostatin (SST) and dopamine (DA) receptors, and clinical reports of enhanced efficacy of combined SST and DA analogue treatment in suppressing GH hypersecretion, lead to the concept of creating chimeric molecules combining structural features of both compound classes. The resulting SST/DA chimeras retain the ability to interact with receptors of both families and display greatly enhanced potency and efficacy, as compared with that of individual SST or DA receptor agonists. In vitro studies with pituitary adenoma cells from acromegalic patients have demonstrated that the chimeric molecules have exceptional activity with regard to suppression of GH and prolactin secretion. Similarly, potent suppression of ACTH secretion from Cushing's-causing corticotroph tumors, and suppression of nonfunctioning pituitary adenoma proliferation has been observed. The chimeric SST/DA compounds are also quite potent and efficacious in suppressing both GH and IGF1 in vivo when tested in nonhuman primates, with no effect on either insulin secretion or glycemic control. Initial clinical studies examining acute, subcutaneous administration of the chimeric SST/DA compound, BIM-23A760, revealed both prolonged circulating half-life and extended duration of biological effect. With chronic administration, however, BIM-23A760 was found to produce a metabolite with dopaminergic activity that gradually accumulates and interferes with the activity of the parent compound. Consequently, efforts are currently underway to produce a second-generation chimera for treatment of neuroendocrine disease.

Somatostatin and dopamine receptor interaction in prostate and lung cancer cell lines.

Somatostatin analogues inhibit in vitro cell proliferation via specific membrane receptors (SSTRs). Recent studies on transfected cell lines have shown a ligand-induced formation of receptor dimers. The aim of this study is 1) to evaluate the role of specific ligands in modulating receptor interactions in the androgen-dependent prostate cancer cell line, LNCaP, and in the non-small cell lung cancer line, Calu-6, by co-immunoprecipitation and immunoblot; and 2) to correlate the antiproliferative effect of these compounds with their ability in modulating receptor interactions. In LNCaP, we have demonstrated the constitutive presence of sstr1/sstr2, sstr2/sstr5, sstr5/dopamine (DA) type 2 receptor (D2R), and sstr2/D2R dimers. BIM-23704 (sstr1- and sstr2-preferential compound) increased the co-immunoprecipitation of sstr1/sstr2 and significantly inhibited proliferation (-30.98%). BIM-23244 (sstr2-sstr5 selective agonist) significantly increased the co-immunoprecipitation of sstr2/sstr5, and induced a -41.36% inhibition of proliferation. BIM-23A760, a new somatostatin/DA chimeric agonist with a high affinity for sstr2 and D2R and a moderate affinity for sstr5, significantly increased the sstr5/D2R and sstr2/D2R complexes and was the most powerful in inhibiting proliferation (-42.30%). The chimeric compound was also the most efficient in modulating receptor interaction in Calu-6, increasing the co-immunoprecipitation of D2R/sstr5 and inhibiting cell proliferation (-30.54%). However, behind BIM-23A760, BIM-53097 (D2R-preferential compound) also significantly inhibited Calu-6 proliferation (-17.71%), suggesting a key role for D2R in receptor cross talk and in controlling cell growth. Indeed, activation of monomeric receptors did not affect receptor co-immunoprecipitation, whereas cell proliferation was significantly inhibited when the receptors were synergistically activated. In conclusion, our data show a dynamic ligand-induced somatostatin and DA receptor interaction, which may be crucial for the antiproliferative effects of the new analogues.

Ghrelin in the regulation of body weight and metabolism.

Ghrelin, a peptide hormone predominantly produced by the stomach, was isolated as the endogenous ligand for the growth hormone secretagogue receptor. Ghrelin is a potent stimulator of growth hormone (GH) secretion and is the only circulatory hormone known to potently enhance feeding and weight gain and to regulate energy homeostasis following central and systemic administration. Therapeutic intervention with ghrelin in catabolic situations may induce a combination of enhanced food intake, increased gastric emptying and nutrient storage, coupled with an increase in GH thereby linking nutrient partitioning with growth and repair processes. These qualities have fostered the idea that ghrelin-based compounds may have therapeutic utility in treating malnutrition and wasting induced by various sub-acute and chronic disorders. Conversely, compounds that inhibit ghrelin action may be useful for the prevention or treatment of metabolic syndrome components such as obesity, impaired lipid metabolism or insulin resistance. In recent years, the effects of ghrelin on glucose homeostasis, memory function and gastrointestinal motility have attracted considerable amount of attention and revealed novel therapeutic targets in treating a wide range of pathologic conditions. Furthermore, discovery of ghrelin O-acyltransferase has also opened new research opportunities that could lead to major understanding of ghrelin physiology. This review summarizes the current knowledge on ghrelin synthesis, secretion, mechanism of action and biological functions with an additional focus on potential for ghrelin-based pharmacotherapies.

Effects of selective somatostatin analogs and cortistatin on cell viability in cultured human non-functioning pituitary adenomas.

Clinically "non-functioning" human pituitary adenomas (NFPA) constitute about 35% of pituitary adenomas. Somatostatin receptors (SSTR) expression in these adenomas has previously been described both in vitro and in vivo, without evidence for a correlation with tumor volume or the therapeutic efficacy of somatostatin analogs. This study was performed on 13 surgically removed pituitary macroadenomas, diagnosed before surgery as "non-functioning". In addition, 3 growth hormone (GH)-secreting adenomas served as controls. A specimen from each tumor was dispersed and digested to isolate and culture the tumor cells, and the in vitro effects of SSTR2 and SSTR5 selective analogs and Cortistatin (CST) (100nM) on cell viability were studied. The quantity of viable cells was estimated using the XTT method. RNA purification of tumor samples and subsequent RT-PCR studies for SSTR2 and SSTR5 expression were performed. Somatostatin analog with high affinity for SSTR2 reduced cell viability by 20-80% in 8 of 13 NFPAs studied, all expressing the SSTR2. The inhibitory effect on cell viability of SSTR5-selective analog was 15-80% in 10 of 13 NFPAs studied, all but three expressing the SSTR5. CST, however, effectively reduced cell viability in only 6 NFPAs. Cell viability was inhibited by all peptides studied in 2 out of 3 GH-secreting adenomas, expressing both receptors. The third adenoma responded to SSTR2 analog and expressed only SSTR2. These results suggest the involvement of SSTR2 and SSTR5 in the anti-proliferative effects of somatostatin; however, CST is less potent in reducing cell viability in these tumors.

Role of the corticotropin-releasing factor receptor type 2 in the control of food intake in mice: a meal pattern analysis.

The actions of corticotropin-releasing factor (CRF) and related peptides are mediated by two receptors (CRF(1) and CRF(2)). The respective role of each subtype in the control of food intake remains poorly known. In the present study, we examined the quantity and microstructure of ingestive behavior of knockout (KO) mice lacking CRF(2) receptors and their wild-type (WT) littermates. Under basal conditions, CRF(2) KO mice showed increased nocturnal food intake, evident as an increased zenith in circadian cosinor analysis of food intake. Microstructure analysis revealed that this greater food intake reflected increased meal size, rather than meal frequency, suggesting a decreased satiating value of food. Following acute restraint stress, CRF(2) KO mice showed an intact immediate anorectic response with increased latency to eat and decreased meal size. However, CRF(2) deletion abolished the prolonged phase of restraint-induced anorexia. CRF(2) KO mice did not differ from WT controls in feeding responses to food deprivation or injection of ghrelin receptor agonists. Independent of genotype, food deprivation increased food intake, with dramatic changes in meal size, meal frequency, water : food ratio and eating rate. Acyl-ghrelin or BIM-28131, a potent ghrelin analog, dose-dependently stimulated food intake by increasing meal size (ghrelin, BIM-28131) and meal number (BIM-28131), while slowing the average eating rate (BIM-28131) similarly in WT and KO mice. These results suggest that the CRF(2) receptor is involved in the control of meal size during the active phase of eating and following acute exposure to stress.

In vitro antiangiogenic activity of selective somatostatin subtype-1 receptor agonists.

BACKGROUND: Endothelial cells of human blood vessels (arteries and veins) show high levels of somatostatin subtype-1 receptor (sst(1)). The aim of the present study is to investigate the inhibitory effects of novel somatostatin analogs, highly selective for human sst(1), on in vitro angiogenesis and their modulation of vascular endothelial growth factor (VEGF) and vascular endothelial growth factor receptor-2 (VEGFR-2) expression. MATERIALS AND METHODS: Somatostatin analogs BIM-23745 and BIM-23926 were tested for their ability to prevent proliferation and migration of human endothelial HMEC-1 cells, to modulate VEGF and VEGFR-2 expression and to inhibit sprouting of microvessels from cultured human placental vessel explants in fibrin matrix for 28 days. RESULTS: The somatostatin sst(1 )receptor-selective agonists, BIM-23745 and BIM-23926 showed a suppression of endothelial proliferation (e.g. 10(-6) M BIM-23475, 40.0 +/- 2.1% vs. 100% of controls; 10(-7) M BIM-23926, 55.3 +/- 3.3% vs. 100% of controls), migration (e.g. 10(-7) M BIM-23475, 35.0 +/- 1.56% vs. 100% of controls; 10(-7) M BIM-23926, 53.7 +/- 1.77% vs. 100% of controls) and microvessel sprouting (e.g. 10(-8) M BIM-23475, 42.8 +/- 5.6% vs. 100% of controls; 10(-7) M BIM-23926, 17.2 +/- 11.8% vs. 100% of controls). A small but significant percentage of cells exposed to BIM-23745 and BIM-23926 for 24 h and for 72 h presented typical apoptotic morphology. Moreover, both the analogs significantly inhibit VEGF and VEGFR-2 gene expression in endothelial cells grown for 144 h in a fibrin matrix and the VEGF secretion in conditioned media. CONCLUSIONS: The inhibition of endothelial activities suggests potential therapeutic utility for administration of somatostatin sst(1 )receptor-selective agonists in the proliferative diseases involving angiogenesis.

A comparison of the effects of gastrin, somatostatin and dopamine receptor ligands on rat gastric enterochromaffin-like cell secretion and proliferation.

Gastrin regulates ECL cell histamine release and is a critical determinant of acid secretion. ECL cell secretion and proliferation is inhibited by gastrin antagonists and somatostatin but little is known about the role of dopamine agonists in this process. Since the ECL cell exhibits all three classes of receptor we evaluated and compared the effects of the gastrin receptor antagonist, (YF476), lanreotide (SST agonist) and novel dopaminergic agents (BIM53061 and BIM27A760) on ECL cell histamine secretion and proliferation. Highly enriched (>98%) ECL cell preparations prepared from rat gastric mucosa using a FACS approach were studied. Real-time PCR confirmed presence of the CCK2, SS2 and SS5 and D1 receptors on ECL cells. YF476 inhibited histamine secretion and proliferation with IC(50)s of 1.25 nM and 1.3 x 10(-11) M respectively, values 10-1000x more potent than L365,260. Lanreotide inhibited secretion and proliferation (2.2 nM, 1.9 x 10(-10) M) and increased YF476-inhibited proliferation a further 5-fold. The dopamine agonist, BIM53061, inhibited gastrin-mediated ECL cell secretion and proliferation (17 nM, 6 x 10(-10) M) as did the novel dopamine/somatostatin chimera BIM23A760 (22 nM, 4.9 x 10(-10) M). Our studies demonstrate that the gastrin receptor antagonist, YF476, is the most potent inhibitor of ECL cell histamine secretion and proliferation. Lanreotide, a dopamine agonist and a dopamine/somatostatin chimera inhibited ECL cell function but were 10-1000x less potent than YF476. Agents that selectively target the CCK2 receptor may provide alternative therapeutic strategies for gastrin-mediated gastrointestinal cell secretion and proliferation such as evident in the hypergastrinemic gastric carcinoids associated with low acid states.

Effects of somatostatin-14 and the receptor-specific somatostatin analogs on chromogranin A and alpha-subunit (alpha-SU) release from "clinically nonfunctioning" pituitary adenoma cells incubated in vitro.

The aim of the study was to examine the effect of somatostatin (SST) and its analogs on the release of chromogranin A (CgA) and alpha-subunit (alpha-SU) from clinically non-functioning pituitary adenomas incubated in vitro. Seven pituitary macroadenomas surgically removed were investigated. All of the tumors were diagnosed before surgery as non-functioning, but they expressed either gonadotropins or their subunits as detected by immunohistochemistry. Two tumors additionally expressed prolactin and growth hormone. All adenomas also expressed chromogranin A (CgA) and at least 3 of 5 subtypes of somatostatin receptors. The cells isolated from the examined tumors were exposed in vitro to either native SST-14 or the following receptor-specific SST analogs: BIM-23926 (agonist of sst1 receptor), BIM-23120 (agonist of sst2 receptor), BIM-23206 (agonist of sst5 receptor) and BIM23A387 (somatostatin/dopamine chimera). The concentration of CgA was measured by means of ELISA method and of alpha-SU was measured by an immunoradiometric method. It was found that the exposure on SST-14 resulted in the decrease of CgA and alpha-SU release from tumor cells in majority of samples, and the effect on CgA was positively correlated with the expression of sst3 and also with the sst2A/sst2B expressions ratio. The inhibitory effect of SST-14 on CgA and alpha-SU seems also to correlate negatively with the expression of sst2B. CgA inhibition also correlates positively with sst5 expression. Among the other compounds studied, only the sst2 agonist decreased the release in all the investigated samples. The remaining substances (agonists of sst1 and sst5 and SST/DA chimera) produced the divergent changes (increased or decreased release, depending on the sample). The data suggest that the inhibition of CgA (and possibly of alpha-SU) release by SST is mediated via subtypes sst2A, sst3 and sst5, whereas sst2B subtype may induce the opposite effect.

Regulation of insulin and glucagon secretion from rat pancreatic islets in vitro by somatostatin analogues.

Somatostatin is an inhibitor of hormone secretion through specific receptors (sst1-5). The aim of this study was to investigate the putative regulatory role of somatostatin analogues on the secretion of insulin and glucagon by rat pancreatic islets. After 48 h exposure only the non-selective agonists (somatostatin, octreotide and SOM-230) inhibited insulin accumulation. The inhibition of insulin secretion was accompanied by increased islet insulin contents. None of the analogues showed a consistent effect on the glucagon accumulation in the medium after 48 h. Since we observed a difference in the regulatory effect between the non-selective and selective analogues, combinations of selective analogues were studied. Combination of sst2+sst5 agonists inhibited the medium insulin accumulation, while combination of sst1+sst2 analogues caused a decrease in glucagon accumulation. After removal of somatostatin a rebound effect with increased insulin secretion were observed. This effect was reversed after 6 h. For SOM-230 insulin secretion continued to be suppressed even after the analogue was removed and returned to control values after 3 h. As for glucagon secretion there was an initial decline after culture with octreotide, while the other substances failed to induce any changes. In summary, non-selective somatostatin analogues or combinations of receptor selective analogues may cause inhibition of hormone secretion from rat pancreatic islets. For insulin and glucagon, combinations of sst2+sst5 and sst1+sst2, respectively may exert this effects. Thus, our data suggest that more than one sst must be involved to down-regulate islet glucagon and insulin secretion.

Novel synergistic mechanism for sst2 somatostatin and TNFalpha receptors to induce apoptosis: crosstalk between NF-kappaB and JNK pathways.

Somatostatin is a multifunctional hormone that modulates cell proliferation, differentiation and apoptosis. Mechanisms for somatostatin-induced apoptosis are at present mostly unsolved. Therefore, we investigated whether somatostatin receptor subtype 2 (sst2) induces apoptosis in the nontransformed murine fibroblastic NIH3T3 cells. Somatostatin receptor subtype 2 expression induced an executioner caspase-mediated apoptosis through a tyrosine phosphatase SHP-1 (Src homology domain phosphatase-1)-dependent stimulation of nuclear factor kappa B (NF-kappaB) activity and subsequent inhibition of the mitogen-activated protein kinase JNK. Tumor necrosis factor alpha (TNFalpha) stimulated both NF-kappaB and c-Jun NH2-terminal kinase (JNK) activities, which had opposite action on cell survival. Importantly, sst2 sensitized NIH3T3 cells to TNFalpha-induced apoptosis by (1) upregulating TNFalpha receptor protein expression, and sensitizing to TNFalpha-induced caspase-8 activation; (2) enhancing TNFalpha-mediated activation of NF-kappaB, resulting in JNK inhibition and subsequent executioner caspase activation and cell death. We have here unraveled a novel signaling mechanism for a G protein-coupled receptor, which directly triggers apoptosis and crosstalks with a death receptor to enhance death ligand-induced apoptosis.

Effect of Extended Irrigation and Host Resistance on Deoxynivalenol Accumulation in Fusarium-Infected Wheat.

Deoxynivalenol (DON) levels are not easily predicted from visual disease assessment, and it is thought likely that environmental conditions such as temperature and moisture influence DON accumulation. This field study examined the influence of environmental moisture on DON accumulation in Fusarium-infected wheat (Triticum aestivum). The effect of extended mist irrigation applied from inoculation (at anthesis) until harvest was compared with mist irrigation applied from inoculation until disease assessment (at early dough), as is generally applied in screening nurseries used for germplasm selection and cultivar improvement. DON concentrations were quantified in kernels at early dough, hard dough, kernel hard, and maturity. Kernels from plots with extended mist irrigation generally had lower DON concentrations than those from plots where mist irrigation was not applied following disease assessment. DON concentrations tended to decrease from disease assessment until harvest, regardless of the irrigation treatment. DON concentrations in the cultivars moderately resistant to Fusarium head blight were lower than those in the susceptible cultivar. Environmental moisture is an important factor determining the DON content of Fusarium-infected wheat.

Effects of chimeric somatostatin-dopamine molecules on human peripheral blood lymphocytes activation.

BIM 23A761, selective for somatostatin receptors subtypes 2, 5 and the dopamine receptor subtype 2, and BIM 23A757 with affinity for SSTR2 and DAR2 were studied on human PBL proliferation and activation. BIM 23A761 was significantly more potent than specific SSTR and DAR2 agonists in suppressing lymphocyte proliferation induced by mitogen or alloantigen, while BIM 23A757 was more potent than specific SSTR2 and DAR2 agonists in suppressing antigen induced proliferation only. Both molecules displayed enhanced potency in suppressing IFNgamma and IL-6 secretion compared with the SSTR and DAR2 analogs, while only BIM 23A761 was able to inhibit IL-2 secretion and its effect is more potent than the control analogs. Furthermore BIM 23A761 inhibit cell progression into the S phase and then into the G2/M, while BIM 23A757 inhibited bromodeoxyuridine incorporation only during the S phase. Both chimeric molecules resulted significantly more effective than the respective controls.

The effect of selective sst1, sst2, sst5 somatostatin receptors agonists, a somatostatin/dopamine (SST/DA) chimera and bromocriptine on the "clinically non-functioning" pituitary adenomas in vitro.

The aim of the work was to investigate the effects of somatostatin analogs acting selectively on sst1 (BIM-23926), sst2 (BIM-23120) and sst5 (BIM-23206) receptor subtypes on the viability of "clinically non-functioning" pituitary adenomas in vitro. The effects of native SST (SST-14), a SST/DA chimera (BIM-23A387) and a D(2)-dopamine receptor agonist bromocriptine (BC) were also examined. The study was performed on 10 surgically removed pituitary macroadenomas, diagnosed before surgery as "non-functioning". A part of each tumor was mechanically dispersed and digested with collagenase to isolate the tumoral cells. Another part of each tumor was fixed, embedded in paraffin and immunostained to reveal the pituitary hormones and SST receptor subtypes (sst1, sst2A, sst2B, sst3, sst4, sst5). The tumoral cell suspensions were incubated for 24 h with the substances mentioned above. The quantity of viable cells was estimated using the EZ4U system. The results were compared with the immunohistochemical evaluation of the hormonal profile of adenoma and the sst receptor subtype immunoreactivities present. The findings indicate that selective sst1, sst2 and sst5 receptors agonists, SST/DA chimera and D(2)-dopamine receptor agonist bromocriptine affect the viability of some, but not all, "clinically non-functioning" pituitary adenomas in vitro. The most effective was bromocriptine. The investigated somatostatin analogs including SST/DA chimera exerted roughly similar inhibitory effects. Further studies are needed to fully evaluate the potential usefulness of these compounds in the pharmacological treatment of "non-functioning" pituitary tumors.

PYY3-36 as an anti-obesity drug target.

The neuropeptide Y (NPY)/peptide YY (PYY) system has been implicated in the physiology of obesity for several decades. More recently ignited enormous interest in PYY3-36, an endogenous Y2-receptor agonist, as a promising anti-obesity compound. Despite this interest, there have been remarkably few subsequent reports reproducing or extending the initial findings, while at the same time studies finding no anti-obesity effects have surfaced. Out of 41 different rodent studies conducted (in 16 independent labs worldwide), 33 (83%) were unable to reproduce the reported effects and obtained no change or sometimes increased food intake, despite use of the same experimental conditions (i.e. adaptation protocols, routes of drug administration and doses, rodent strains, diets, drug vendors, light cycles, room temperatures). Among studies by authors in the original study, procedural caveats are reported under which positive effects may be obtained. Currently, data speak against a sustained decrease in food intake, body fat, or body weight gain following PYY3-36 administration and make the previously suggested role of the hypothalamic melanocortin system unlikely as is the existence of PYY deficiency in human obesity. We review the studies that are in the public domain which support or challenge PYY3-36 as a potential anti-obesity target.

Dopamine receptor subtype 2 and somatostatin receptor subtype 5 expression influences somatostatin analogs effects on human somatotroph pituitary adenomas in vitro.

Dopamine (DA) and somatostatin (SRIF) receptor agonists inhibit growth hormone (GH) secretion by pituitary adenomas. We investigated DA subtype 2 receptor (DR2) and SRIF receptor (sst) subtypes 2 and 5 expression in 25 GH-secreting pituitary adenomas and tested in primary culture the effects on GH and prolactin (PRL) secretion of sst agonists selectively interacting with sst2 (BIM-23120), sst5 (BIM-23206), and sst2 and sst5 (BIM-23244). All adenomas expressed sst2; eight adenomas expressed both sst5 and DR2, eight sst5 but not DR2, and eight DR2 but not sst5. One tissue lacked expression of DR2 and sst5. GH secretion was inhibited by BIM-23120 in all samples, while it was reduced by BIM-23206 only in adenomas not expressing DR2. BIM-23120's inhibitory effects correlated with sst2 and DR2 expression, whereas DR2 expression correlated inversely with BIM-23206 inhibitory effects on GH secretion. In seven mixed GH-/PRL-secreting pituitary adenomas, PRL secretion was inhibited in sst5-expressing tumors by BIM-23206, but not by BIM-23120. BIM-23244 reduced PRL secretion only in adenomas expressing sst2, sst5 and DR2. sst5 and DR2 expression correlated directly with BIM23206 inhibitory effects on PRL secretion. Our results suggest that adenomas expressing DR2 are less likely to respond to clinically available SRIF analogs in terms of GH secretion inhibition. Therefore, drugs interacting also with DR2 might better control secretion of pituitary adenomas.