Inhibition of growth of PC-82 human prostate cancer line xenografts in nude mice by bombesin antagonist RC-3095 or combination of agonist [D-Trp6]-luteinizing hormone-releasing hormone and somatostatin analog RC-160.

The effects of treatment with a bombesin receptor antagonist [D-Tpi6, Leu13 psi (CH2NH) Leu14]BN(6-14)(RC-3095) and the combination of an agonist of luteinizing hormone-releasing hormone [D-Trp6]-LH-RH and somatostatin analog D-Phe-Cys-Tyr-D-Trp-Lys-Val- Cys-Trp-NH2 (RC-160) were studied in nude mice bearing xenografts of the hormone-dependent human prostate tumor PC-82. During the 5 weeks of treatment, tumor growth was decreased in all treated groups compared with controls. Bombesin antagonist RC-3095 and the combination of [D-Trp6]-LH-RH and RC-160 caused a greater inhibition of tumor growth than [D-Trp6]-LH-RH or RC-160 alone as based on measurement of tumor volume and percentage change in tumor volume. The largest decrease in tumor weight was also seen in the groups treated with the bombesin antagonist and with the combination of RC-160 and [D-Trp6]-LH-RH. Serum prostatic-specific antigen levels were greatly decreased, and insulin-like growth factor I (IGF-I) as well as growth hormone levels were reduced in all treated groups. Specific binding sites for [D-Trp6]-LH-RH, epidermal growth factor (EGF), IGF-I, and somatostatin (SS-14) were found in the tumor membranes. Receptors for EGF were significantly down-regulated by treatment with the bombesin antagonist or RC-160. Combination of LH-RH agonists with somatostatin analog RC-160 might be considered for improvement of hormonal therapy for prostate cancer. The finding that bombesin antagonist RC-3095 inhibits the growth of PC-82 prostate cancer suggests the merit of further studies to evaluate the possible usefulness of antagonists of bombesin in the management of prostatic carcinoma.     

Rickettsia-macrophage interactions: host cell responses to Rickettsia akari and Rickettsia typhi

The existence of intracellular rickettsiae requires entry, survival, and replication in the eukaryotic host cells and exit to initiate new infection. While endothelial cells are the preferred target cells for most pathogenic rickettsiae, infection of monocytes/macrophages may also contribute to the establishment of rickettsial infection and resulting pathogenesis. We initiated studies to characterize macrophage-Rickettsia akari and -Rickettsia typhi interactions and to determine how rickettsiae survive within phagocytic cells. Flow cytometry, microscopic analysis, and LDH release demonstrated that R. akari and R. typhi caused negligible cytotoxicity in mouse peritoneal macrophages as well as in macrophage-like cell line, P388D1. Host cells responded to rickettsial infection with increased secretion of proinflammatory cytokines such as interleukin-1beta (IL-1beta) and IL-6. Furthermore, macrophage infection with R. akari and R. typhi resulted in differential synthesis and expression of IL-beta and IL-6, which may correlate with the existence of biological differences among these two closely related bacteria. In contrast, levels of gamma interferon (IFN-gamma), IL-10, and IL-12 in supernatants of infected P388D1 cells and mouse peritoneal macrophages did not change significantly during the course of infection and remained below the enzyme-linked immunosorbent assay cytokine detection limits. In addition, differential expression of cytokines was observed between R. akari- and R. typhi-infected macrophages, which may correlate with the biological differences among these closely related bacteria.     

Strain and substrain differences in context- and tone-dependent fear conditioning of inbred mice

The performance of C57BL/6J (6J), C57BL/6N (6N), DBA/2J (2J) and DBA/2N (2N) mice in context- and tone-dependent fear conditioning was determined 24 h after fear conditioning to evaluate and compare different behavioral measures as indices of emotional learning. Freezing, the change in activity and the size of the explored area were evaluated as behavioral parameters indicating fear. Additionally, the heart rate (HR) increase elicited by tone presentation was evaluated as an autonomic indicator of fear. During the context-dependent memory test, freezing was high only in 6J and 6N mice, whereas a drop of activity and a reduced exploratory area was measured in all strains. During the tone-dependent memory test, high freezing, low activity, reduced exploratory area and a strong HR increase were demonstrated only in 6N and 6J mice, whereas behavioral and HR changes of 2J and 2N mice were always low. In extinction tests, context- and tone-dependent freezing of 6J mice decayed significantly faster than the freezing of 6N mice, whereas in both substrains the conditioned tachycardia to tone extinguished similarly in the home cage. The data demonstrate that monitoring of additional behavioral measures besides freezing and autonomic measures is necessary to interpret differences in associative learning performance of mouse strains that could be related to a differential expression of fear.     

Regulatory mechanisms of fear extinction and depression-like behavior.

Human anxiety is frequently accompanied by depression, and when they co-occur both conditions exhibit greater severity and resistance to treatment. Little is known, however, about the molecular processes linking these emotional and mood disorders. Based on previously reported phosphorylation patterns of extracellular signal-regulated kinase (ERK) in the brain, we hypothesized that ERK's upstream activators intertwine fear and mood regulation through their hippocampal actions. We tested this hypothesis by studying the upstream regulation of ERK signaling in behavioral models of fear and depression. Wild-type and ERK1-deficient mice were used to study the dorsohippocampal actions of the putative ERK activators: mitogen-activated and extracellular signal-regulated kinase (MEK), protein kinase C (PKC), and cAMP-dependent protein kinase (PKA). Mice lacking ERK1 exhibited enhanced fear extinction and reduced depression caused by overactivation of ERK2. Both behaviors were reversed by inhibition of MEK, however the extinction phenotype depended on hippocampal, whereas the depression phenotype predominantly involved extrahippocampal MEK. Unexpectedly, inhibition of PKC accelerated extinction and decreased depression by ERK-independent mechanisms, whereas inhibition of PKA did not produce detectable molecular or behavioral effects in the employed paradigm. These results indicate that, contrary to fear conditioning but similar to mood stabilization, extinction of fear required upregulation of MEK/ERK and downregulation of ERK-independent PKC signaling. The dissociation of these pathways may thus represent a common mechanism for fear and mood regulation, and a potential therapeutic option for comorbid anxiety and depression.     

Inhibition of human epithelial ovarian cancer cell growth in vitro by somatostatin analog RC-160

In this study, we investigated the effects of somatostatin analog RC-160 on the growth of the OV-1063 human epithelial ovarian cancer cell line in vitro. RC-160 inhibited cell proliferation, as measured by cell number, and [(3)H]thymidine incorporation into DNA at 10(-9)-10(-5) M. In OV-1063 cells, (125)I-labeled RC-160 was bound to one class of specific, saturable binding sites with high affinity (K(d) = 0.2 +/- 0.03 nM) and low capacity (5,500 binding sites per cell). (125)I-labeled RC-160 could be displaced by unlabeled RC-160. Ligand binding was dependent on time and temperature. Receptor internalization assay showed that the ligand-receptor complex was internalized at 37 degrees C, which indicates the presence of biologically active somatostatin receptors on OV-1063 cells. These results suggest that somatostatin analog RC-160 can suppress the growth of OV-1063 human epithelial ovarian cancer cells by a direct action and that the inhibitory effect of somatostatin analog is mediated through the high-affinity somatostatin receptors.     

Partial angiotensin-converting enzyme inhibition during acute orthostatic stress in persons with tetraplegia

INTRODUCTION: Individuals with tetraplegia rely on the renin-angiotensin system for orthostatic blood pressure control. OBJECTIVES: To determine the effect of partial angiotensin-converting enzyme (ACE) inhibition on heart rate (HR), active plasma renin (PR), and mean arterial blood pressure (MAP) during acute orthostasis in subjects with tetraplegia (n = 7) and nondisabled persons (n = 8). METHODS: Subjects were instructed to avoid caffeine and alcohol for 24 hours before testing and to report to the laboratory between 10 AM and 1 PM. Progressive head-up tilt (15 degrees, 25 degrees, 35 degrees, and 45 degrees) was performed on 2 separate days; Day 1: without ACE inhibition; Day 2: after intravenous (IV) infusion of enalaprilat (0.625 mg). RESULTS: HR was reduced during orthostasis in the tetraplegia compared with the nondisabled group (P < 0.0001), and was unaffected by ACE inhibition in either group. PR was not increased with orthostasis in either group, but was increased after ACE inhibition in both groups (P < 0.001). MAP was not affected by orthostasis in either group, but was reduced with ACE inhibition in both groups (P < 0.01). In the tetraplegia group, MAP was initially reduced after ACE inhibition, but was maintained thereafter with increasing angles of tilt, and no subject complained of symptomatic orthostatic hypotension. CONCLUSION: Subjects with tetraplegia were tolerant of an acute bout of orthostatic stress after partial ACE inhibition. This may have clinical relevance because of the increased prevalence of type 2 diabetes mellitus in this population and the use of ACE inhibitors for the treatment of progressive renal and cardiovascular disease.