Angpt2/Tie2 autostimulatory loop controls tumorigenesis.

Invasive nonfunctioning (NF) pituitary neuroendocrine tumors (PitNETs) are non-resectable neoplasms associated with frequent relapses and significant comorbidities. As the current therapies of NF-PitNETs often fail, new therapeutic targets are needed. The observation that circulating angiopoietin-2 (ANGPT2) is elevated in patients with NF-PitNET and correlates with tumor aggressiveness prompted us to investigate the ANGPT2/TIE2 axis in NF-PitNETs in the GH3 PitNET cell line, primary human NF-PitNET cells, xenografts in zebrafish and mice, and in MENX rats, the only autochthonous NF-PitNET model. We show that PitNET cells express a functional TIE2 receptor and secrete bioactive ANGPT2, which promotes, besides angiogenesis, tumor cell growth in an autocrine and paracrine fashion. ANGPT2 stimulation of TIE2 in tumor cells activates downstream cell proliferation signals, as previously demonstrated in endothelial cells (ECs). Tie2 gene deletion blunts PitNETs growth in xenograft models, and pharmacological inhibition of Angpt2/Tie2 signaling antagonizes PitNETs in primary cell cultures, tumor xenografts in mice, and in MENX rats. Thus, the ANGPT2/TIE2 axis provides an exploitable therapeutic target in NF-PitNETs and possibly in other tumors expressing ANGPT2/TIE2. The ability of tumor cells to coopt angiogenic signals classically viewed as EC-specific expands our view on the microenvironmental cues that are essential for tumor progression.

The Diagnosis and Management of Endocrine Side Effects of Immune Checkpoint Inhibitors.

BACKGROUND: The immunologically mediated side effects of immune checkpoint inhibitors (CPI) often involve the endocrine system as well, and they can even be fatal, as in the case of unrecognized hypophysitis. Distinguishing such side effects from tumor-related changes is often difficult, because their clinical features can be nonspecific. METHODS: This review is based on publications retrieved by a selective search in PubMed, with special attention to international recommendations. RESULTS: Depending on their target molecules, the CPI now in use differ from one another in the incidence of side effects such as autoimmune thyroid disease (4-16%), hypophysitis (0.1- 18%), adrenalitis (0.7-8%), and autoimmune diabetes mellitus (0.5-2%). The typical clinical warning signs and laboratory constellations of hypophysitis include exhaustion, hyponatremia, and headache. Hypo- and hyperthyroidism and primary adrenocortical insufficiency likewise have nonspecific manifestations. Autoimmune diabetes mellitus often takes a fulminant course. Patients being treated with CPI should be monitored at close intervals, at least as frequently as the administration of the drug, so that endocrine side effects can be recognized in time. In case of doubt, glucocorticoid supplementation should be given whenever hypocortisolism is suspected, even before endocrine evaluation is completed and the results are available. Interrupting or discontinuing CPI treatment is rarely indicated. CONCLUSION: With the increasing number of patients being treated with CPI, more and more physicians from a wide variety of specialties, not necessarily working in specialized centers, now have to consider immunologically mediated endocrine side effects in the differential diagnosis, and treat them properly when they arise. These things should be done in collaboration with endocrinologists. The ongoing study of such side effects of the CPI now in use, and of those that will be introduced in the future, is important and will lead to improved understanding.

Human Cerebrospinal Fluid Induces Neuronal Excitability Changes in Resected Human Neocortical and Hippocampal Brain Slices.

Human cerebrospinal fluid (hCSF) has proven advantageous over conventional medium for culturing both rodent and human brain tissue. In addition, increased activity and synchrony, closer to the dynamic states exclusively recorded in vivo, were reported in rodent slices and cell cultures switching from artificial cerebrospinal fluid (aCSF) to hCSF. This indicates that hCSF possesses properties that are not matched by the aCSF, which is generally used for most electrophysiological recordings. To evaluate the possible significance of using hCSF as an electrophysiological recording medium, also for human brain tissue, we compared the network and single-cell firing properties of human brain slice cultures during perfusion with hCSF and aCSF. For measuring the overall activity from a majority of neurons within neocortical and hippocampal human slices, we used a microelectrode array (MEA) recording technique with 252 electrodes covering an area of 3.2 × 3.2 mm2. A second CMOS-based MEA with 4225 sensors on a 2 × 2 mm2 area was used for detailed mapping of action potential waveforms and cell identification. We found that hCSF increased the number of active electrodes and neurons and the firing rate of the neurons in the slices and induced an increase in the numbers of single channel and population bursts. Interestingly, not only an increase in the overall activity in the slices was observed, but a reconfiguration of the network could also be detected with specific activation and inactivation of subpopulations of neuronal ensembles. In conclusion, hCSF is an important component to consider for future human brain slice studies, especially for experiments designed to mimic parts of physiology and disease observed in vivo.

Long-term adult human brain slice cultures as a model system to study human CNS circuitry and disease.

Most of our knowledge on human CNS circuitry and related disorders originates from model organisms. How well such data translate to the human CNS remains largely to be determined. Human brain slice cultures derived from neurosurgical resections may offer novel avenues to approach this translational gap. We now demonstrate robust preservation of the complex neuronal cytoarchitecture and electrophysiological properties of human pyramidal neurons in long-term brain slice cultures. Further experiments delineate the optimal conditions for efficient viral transduction of cultures, enabling 'high throughput' fluorescence-mediated 3D reconstruction of genetically targeted neurons at comparable quality to state-of-the-art biocytin fillings, and demonstrate feasibility of long term live cell imaging of human cells in vitro. This model system has implications toward a broad spectrum of translational studies, regarding the validation of data obtained in non-human model systems, for therapeutic screening and genetic dissection of human CNS circuitry.

Consensus on Postoperative Recommendations After Transsphenoidal Surgery.

BACKGROUND: Guidelines for patient behavior following transsphenoidal surgery do not exist. To gain generally recommendations, the German pituitary working group conducted a study among pituitary surgeons to elucidate their opinions and customs of patients' counselling. METHODS: Questions concerning daily activities, exertion of sports and work life were addressed. It was asked to provide the postoperative time interval after which specific activities can be resumed both after a routine or an extended approach. RESULTS: Fourteen pituitary surgeons returned the completed questionnaire. Following routine operations, washing the hair was allowed within one week, blowing the nose after 3, flying on an airplane and driving a car after one, lifting heavy weights after 4, playing wind instruments after 6, use of CPAP (continuous positive airway pressure) device after 3, permit leisure sports after 2 to 4 weeks (except for scuba diving). Competitive sports can be resumed after 6 weeks. Occupation with mental demands was considered feasible after 2 weeks, with physical labor after 4 weeks. After extended transsphenoidal surgery, the recommended time interval was roughly twice as long compared to the routine approach. Driving a car was allowed within the first 4 weeks after surgery by some pituitary surgeons, while others allow driving only after 3 months analogous to the regulations after craniotomy. The risk of scuba diving was considered high. CONCLUSIONS: The data of our study and the literature, and expert opinions from related scientific fields resulted in a consensus on recommendations for patients' conduct to minimize risks after transsphenoidal surgery.

Human Cerebrospinal fluid promotes long-term neuronal viability and network function in human neocortical organotypic brain slice cultures.

Pathophysiological investigation of CNS-related diseases, such as epilepsy or neurodegenerative disorders, largely relies on histological studies on human post mortem tissue, tissue obtained by biopsy or resective surgery and on studies using disease models including animal models, heterologous expression systems or cell culture based approaches. However, in general it remains elusive to what extent results obtained in model systems can be directly translated to the human brain, calling for strategies allowing validation or even primary investigation in live human CNS tissue. In the work reported here, we prepared human organotypic slice cultures from access tissue of resective epilepsy surgery. Employing different culture conditions, we systematically compared artificial culturing media versus human cerbrospinal fluid (hCSF) obtained from patients with normal pressure hydrocephalus (NPH). Presented data demonstrates sustained cortical neuronal survival including not only maintenance of typical cellular electrophysiological properties and activity, such as robust action potential generation and synaptic connectivity, but also preservation of tonic and phasic network activity up to several weeks in vitro. As clearly delineated by immunocytochemistry, single cell patch clamp and extracellular recordings, we find that in contrast to artificial culturing media, hCSF significantly enhances neuron viability and maintenance of network activity.

Targeting hyperactivation of the AKT survival pathway to overcome therapy resistance of melanoma brain metastases.

Brain metastases are the most common cause of death in patients with metastatic melanoma, and the RAF-MEK-ERK and PI3K-AKT signaling pathways are key players in melanoma progression and drug resistance. The BRAF inhibitor vemurafenib significantly improved overall survival. However, brain metastases still limit the effectiveness of this therapy. In a series of patients, we observed that treatment with vemurafenib resulted in substantial regression of extracerebral metastases, but brain metastases developed. This study aimed to identify factors that contribute to treatment resistance in brain metastases. Matched brain and extracerebral metastases from melanoma patients had identical ERK, p-ERK, and AKT immunohistochemistry staining patterns, but there was hyperactivation of AKT (p-AKT) and loss of PTEN expression in the brain metastases. Mutation analysis revealed no differences in BRAF, NRAS, or KIT mutation status in matched brain and extracerebral metastases. In contrast, AKT, p-AKT, and PTEN expression was identical in monolayer cultures derived from melanoma brain and extracerebral metastases. Furthermore, melanoma cells stimulated by astrocyte-conditioned medium showed higher AKT activation and invasiveness than melanoma cells stimulated by fibroblast-conditioned medium. Inhibition of PI3K-AKT signaling resensitized melanoma cells isolated from a vemurafenib-resistant brain metastasis to vemurafenib. Brain-derived factors appear to induce hyperactivation of the AKT survival pathway and to promote the survival and drug resistance of melanoma cells in the brain. Thus, inhibition of PI3K-AKT signaling shows potential for enhancing and/or prolonging the antitumor effect of BRAF inhibitors or other anticancer agents in melanoma brain metastases.

MR imaging findings in colloid cysts of the sellar region: comparison with colloid cysts of the third ventricle and Rathke's cleft cysts.

RATIONALE AND OBJECTIVES: To identify magnetic resonance (MR) imaging characteristics allowing specific preoperative discrimination between colloid cysts (CCs) of the sellar region and third ventricle (CC3rdv) versus Rathke's cleft cysts (RCCs). MATERIALS AND METHODS: MR imaging data of 38 patients with histologically proven CCs/CC3rdv and RCC underwent retrospective analysis with respect to signal intensity and heterogeneity on T1- and T2-weighted images, presence of the dot sign, enhancement, size, location, and accompanying infundibular stalk abnormalities. RESULTS: Thirteen patients had CCs, 12 had CC3rdv, and 13 had RCCs. Signal intensity on T1-weighted images was partly or entirely hyperintense (n = 8), iso- or mixed iso/hypointense (n = 5) in CCs; hyperintense (n = 8), isointense, or mixed hypo/isointense (n = 3) in CC3rdv and hyperintense (n = 9); or mixed (n = 4) in RCCs. On T2-weighted images, signal intensity was hypointense (n = 12) or hyperintense (n = 1) in CCs, hypointense (n = 9) or hyperintense (n = 2) in CC3rdv, and hypointense (n = 5) or iso/hyperintense (n = 8) in RCCs. T2-weighted images were unavailable in two patients. Only one questionable enhancement was found in CCs, whereas an enhancing rim was consistently seen in RCCs. The dot sign was present in 7 CCs, 8 CC3rdv, and 4 RCCs. Mean cyst diameters were 12.6 mm for CCs and 14.5 mm for RCCs. RCCs showed more frequent and even solely suprasellar extent contrary to CCs. CONCLUSION: Cyst wall enhancement was found in all RCCs but in none of the CCs, making this feature a reliable discriminator between the two. Complementary, suprasellar extension was more frequent in RCCs, whereas signal hypointensity on T2w was more common in colloid cysts.

Transcriptome analysis of MENX-associated rat pituitary adenomas identifies novel molecular mechanisms involved in the pathogenesis of human pituitary gonadotroph adenomas.

Gonadotroph adenomas comprise 15-40% of all pituitary tumors, are usually non-functioning and are often large and invasive at presentation. Surgery is the first-choice treatment, but complete resection is not always achieved, leading to high recurrence rates. As gonadotroph adenomas poorly respond to conventional pharmacological therapies, novel treatment strategies are needed. Their identification has been hampered by our incomplete understanding of the molecular pathogenesis of these tumors. Recently, we demonstrated that MENX-affected rats develop gonadotroph adenomas closely resembling their human counterparts. To discover new genes/pathways involved in gonadotroph cells tumorigenesis, we performed transcriptome profiling of rat tumors versus normal pituitary. Adenomas showed overrepresentation of genes involved in cell cycle, development, cell differentiation/proliferation, and lipid metabolism. Bioinformatic analysis identified downstream targets of the transcription factor SF-1 as being up-regulated in rat (and human) adenomas. Meta-analyses demonstrated remarkable similarities between gonadotroph adenomas in rats and humans, and highlighted common dysregulated genes, several of which were not previously implicated in pituitary tumorigenesis. Two such genes, CYP11A1 and NUSAP1, were analyzed in 39 human gonadotroph adenomas by qRT-PCR and found to be up-regulated in 77 and 95% of cases, respectively. Immunohistochemistry detected high P450scc (encoded by CYP11A1) and NuSAP expression in 18 human gonadotroph tumors. In vitro studies demonstrated for the first time that Cyp11a1 is a target of SF-1 in gonadotroph cells and promotes proliferation/survival of rat pituitary adenoma primary cells and cell lines. Our studies reveal clues about the molecular mechanisms driving rat and human gonadotroph adenomas development, and may help identify previously unexplored biomarkers for clinical use.

Visualization of subdural strip and grid electrodes using curvilinear reformatting of 3D MR imaging data sets.

Curvilinear reformatting of 3D MR imaging data sets was used to visualize the position of subdural strip and grid electrodes relative to the underlying cerebral cortex in patients with epilepsy who were undergoing invasive electroencephalographic recordings. The contour of the cortical surface was delineated interactively, and topographical relationships among surface gyration, cortical lesions, and subdural electrodes were investigated by using serial convex planes parallel to the cortical surface. Electrode contacts could be marked and their positions projected to underlying areas at different depths. This method is apt for routine purposes and allows electrode positions to be displayed with respect to cortical and subcortical regions of interest.