Activation of EP4 receptors prevents endotoxin-induced neutrophil infiltration into the airways and enhances microvascular barrier function.

BACKGROUND AND PURPOSE: Pulmonary vascular dysfunction is a key event in acute lung injury. We recently demonstrated that PGE2 , via activation of E-prostanoid (EP)4 receptors, strongly enhances microvascular barrier function in vitro. The aim of this study was to investigate the beneficial effects of concomitant EP4 receptor activation in murine models of acute pulmonary inflammation. EXPERIMENTAL APPROACH: Pulmonary inflammation in male BALB/c mice was induced by LPS (20 µg per mouse intranasally) or oleic acid (0.15 µL·g-1 , i.v. ). In-vitro, endothelial barrier function was determined by measuring electrical impedance. KEY RESULTS: PGE2 activation of EP4 receptors reduced neutrophil infiltration, pulmonary vascular leakage and TNF-α concentration in bronchoalveolar lavage fluid from LPS-induced pulmonary inflammation. Similarly, pulmonary vascular hyperpermeability induced by oleic acid was counteracted by EP4 receptor activation. In lung function assays, the EP4 agonist ONO AE1-329 restored the increased resistance and reduced compliance upon methacholine challenge in mice treated with LPS or oleic acid. In agreement with these findings, EP4 receptor activation increased the in vitro vascular barrier function of human and mouse pulmonary microvascular endothelial cells and diminished the barrier disruption induced by LPS. The EP2 agonist ONO AE1-259 likewise reversed LPS-induced lung dysfunction without enhancing vascular barrier function. CONCLUSION AND IMPLICATIONS: Our results show that activation of the EP4 receptor strengthens the microvascular barrier function and thereby ameliorates the pathology of acute lung inflammation, including neutrophil infiltration, vascular oedema formation and airway dysfunction. This suggests a potential benefit for EP4 agonists in acute pulmonary inflammation.

Heteromerization of GPR55 and cannabinoid CB2 receptors modulates signalling.

BACKGROUND AND PURPOSE: Heteromerization of GPCRs is key to the integration of extracellular signals and the subsequent cell response via several mechanisms including heteromer-selective ligand binding, trafficking and/or downstream signalling. As the lysophosphatidylinositol GPCR 55 (GPR55) has been shown to affect the function of the cannabinoid receptor subtype 2 (CB2 receptor) in human neutrophils, we investigated the possible heteromerization of CB2 receptors with GPR55. EXPERIMENTAL APPROACH: The direct interaction of human GPR55 and CB2 receptors heterologously expressed in HEK293 cells was assessed by co-immunoprecipitation and bioluminescence resonance energy transfer assays. The effect of cross-talk on signalling was investigated at downstream levels by label-free real-time methods (Epic dynamic mass redistribution and CellKey impedance assays), ERK1/2-MAPK activation and gene reporter assays. KEY RESULTS: GPR55 and CB2 receptors co-localized on the surface of HEK293 cells, co-precipitated in membrane extracts and formed heteromers in living HEK293 cells. Whereas heteromerization led to a reduction in GPR55-mediated activation of transcription factors (nuclear factor of activated T-cells, NF-κB and cAMP response element), ERK1/2-MAPK activation was potentiated in the presence of CB2 receptors. CB2 receptor-mediated signalling was also affected by co-expression with GPR55. Label-free assays confirmed cross-talk between the two receptors. CONCLUSIONS AND IMPLICATIONS: Heteromers, unique signalling units, form in HEK293 cells expressing GPR55 and CB2 receptors. The signalling by agonists of either receptor was governed (i) by the presence or absence of the partner receptors (with the consequent formation of heteromers) and (ii) by the activation state of the partner receptor.

The GPCR-associated sorting protein 1 regulates ligand-induced down-regulation of GPR55.

BACKGROUND AND PURPOSE: Many GPCRs, including the CB(1) cannabinoid receptor, are down-regulated following prolonged agonist exposure by interacting with the GPCR-associated sorting protein-1 (GASP-1). The CB(1) receptor antagonist rimonabant has also recently been described to be an agonist at GPR55, a cannabinoid-related receptor. Here we investigated the post-endocytic properties of GPR55 after agonist exposure and tested whether GASP-1 is involved in this process. EXPERIMENTAL APPROACH: We evaluated the direct protein-protein interaction of GPR55 with GASP-1 using (i) GST-binding assays and (ii) co-immunoprecipitation assays in GPR55-HEK293 cells with endogenous GASP-1 expression. We further tested the internalization, recycling and degradation of GPR55 using confocal fluorescence microscopy and biotinylation assays in the presence and absence of GASP-1 (lentiviral small hairpin RNA knockdown of GASP-1) under prolonged agonist [rimonabant (RIM), lysophosphatidylinositol (LPI)] stimulation. KEY RESULTS: We showed that the prolonged activation of GPR55 with rimonabant or LPI down-regulates GPR55 via GASP-1. GASP-1 binds to GPR55 in vitro, and this interaction was required for targeting GPR55 for degradation. Disrupting the GPR55-GASP-1 interaction prevented post-endocytic receptor degradation, and thereby allowed receptor recycling. CONCLUSION AND IMPLICATIONS: These data implicate GASP-1 as an important regulator of ligand-mediated down-regulation of GPR55. By identifying GASP-1 as a key regulator of the trafficking and, by extension, functional expression of GPR55, we may be one step closer to gaining a better understanding of this receptor in response to cannabinoid drugs. LINKED ARTICLES: This article is part of a themed section on Cannabinoids in Biology and Medicine. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2012.165.issue-8. To view Part I of Cannabinoids in Biology and Medicine visit http://dx.doi.org/10.1111/bph.2011.163.issue-7.

The trapezoid form of the trochlea tali.

The wedge-shaped superior surface of the trochlea tali may be seen in a proximal view as a trapezium. The angle of this trapezium was calculated by measuring the length, and the smallest and largest widths of the trochlea, and was found to be 16.04 degrees for the left talus and 12.48 degrees for the right talus, respectively. The difference between these angles is highly significant. Due to biomechanical features of the talocrural joint an increase in the angle of the trochlea tali also intensifies the incongruency of this joint in plantar-flexion. Thus, this incongruency is larger on the left side. With this diminution of the joint-surface contact, increased internal rotation or "wobbling" movements are possible. An explanation for these facts might be found in footedness, similar to handedness. When constructing a prosthesis one has to consider that the trochlea tali cannot be exactly mirrored from one side to the other but may have to be calculated separately for each side. The fact that footedness seems to correlate with the angle of the trochlea tali can be also helpful.

An HLA class-II allele frequent in Eskimos and Amerindians is found in the Tyrolean Ice Man.

DNA was extracted from specimens derived from the calcaneus of the Tyrolean Ice Man under sterile conditions in a laboratory, where no DNA extractions and PCR experiments had been performed before. Agarose gel electrophoresis and ethidium bromide staining did not reveal any evidence of genomic DNA in the preparation obtained, indicating a high degree of DNA degradation. Nevertheless, we performed PCR amplifications with this sample using primer pairs specific for HLA class II alleles. HLA-DRB and DQB1 alleles were amplified in a nested PCR approach. In one of the reactions, we observed a distinct amplification product, which we directly sequenced. By comparing the obtained nucleotide sequence with a database of HLA alleles we assigned the HLA-DRB1*1402 type to the amplified sample. None of the investigators involved possesses this allele, indicating that no contamination with modern DNA had occurred. The HLA-DRB1*1402 allele is extremely rare in Europe, but is common in Inuits and South American Indians and has previously only once been identified in the laboratory.

Otorhinolaryngologic computer-assisted biopsies of the Iceman.

BACKGROUND: The Iceman is a prehistoric, completely preserved, 5300-year-old male human mummy. OBJECTIVE: To obtain the first biopsy specimens from inside the Iceman while meeting an extended standard of hygiene and following precise intraoperative guidance to the site of biopsy and keeping tissue damage to a minimum. DESIGN: Biopsy specimens from the nose, the maxillary sinus, and the larynx of the Iceman were obtained. Special caution had to be taken while performing the biopsies to not contaminate the Iceman with heavy metals or remnants of microorganisms. SUBJECT: The Iceman, a cadaver kept frozen in a glacier for 5300 years. The Iceman is in an excellent state of preservation and will allow fundamental histological, morphological, and molecular genetic insights into early man. INTERVENTION: The biopsies were planned and executed with the aid of Interventional Video Tomography, a system that guides the surgeon to the target area by combining live video with existing imaging modalities. The system does not need mechanical fixation of the subject (the Iceman) and is barely in physical contact with the subject; thus, it was the ideal tool for guiding the surgeon to the site of the biopsy samplings through a tiny canal into the nose, the maxillary sinus, and the larynx of the Iceman. RESULTS: We have obtained a number of tissue samples by precisely guided 3-dimensional navigation. Unnecessary tissue damage was avoided. CONCLUSIONS: Visual inspection of the extracted mucosa showed typical human cadaver tissue, despite its age, without clinical abnormalities. Currently, the samples are being investigated by various international scientific groups.

[3D computer-assisted ENT biopsies of the Iceman]. / 3D-computerunterstützte HNO-Biopsien des Eismannes.

The University of Innsbruck possesses a unique prehistoric, completely conserved 5300-year-old human cadaver. We report our experiences during which ENT specialists collected samples from various cavities inside the Iceman. Guidance of biopsy instruments was accomplished with computer-assisted navigation based on Interventional Video Tomography. This technology allows surgical guidance by interlinking currently available imaging modalities with live endoscopic video. The system operates without patient fixation and is practically free of external contact. Apart from sterility, special precautionary measures were necessary to avoid contamination with heavy metals or microorganisms. Visual inspection of the samples of mucosa from the nose, maxillary sinus and larynx revealed the typical patterns of a human cadaver without overt pathology.

Skull of a 5,300-year-old mummy: reproduction and investigation with CT-guided stereolithography.

PURPOSE: In September 1991, a mummified corpse was discovered soon after it was released by the receding ice of the Similaun glacier in the Tyrolean Alps. This body proved to be an astonishingly well-preserved man from the late Neolithic Age or early Bronze Age. To preserve the fragile mummy, scientific investigations were required to be as noninvasive and nondestructive as possible. MATERIALS AND METHODS: A radiologic investigation that included conventional radiography, digital radiography, and whole-body computed tomography (CT) was performed from the CT data, the skull was duplicated by means of stereolithography. RESULTS: The copy of the prehistoric skull was validated by means of comparison of measurements obtained from the original CT images and from external physical measurements of the intact head of the mummy. CONCLUSION: The CT images, radiologic findings, and duplicated skull are expected to provide anthropologists and other interested scientists relatively accurate information without the need to handle the mummy, to expose it to the considerable risks of repeated partial thawing, or to perform invasive exploratory studies such as autopsy.

[Value of the neurovascular guide plate for pelvic surgery]. / Die Bedeutung der Gefäss-Nerven-Leitplatte für die Beckenchirurgie.

In the past few years great effort has been made to reduce the morbidity after oncologic operations in the small pelvis of the male. In this respect it is essential for urologic surgeons to orient themselves with the anatomic structures that mark the course of the important vessels and nerves. For this reason an anatomic study of the connective tissue in the male pelvis was performed in which dissections and serial sections were analyzed. The results show that the connective tissue in the male pelvis does not represent a monomorphic block but consists of several portions of varying density. The looser portions are also known as the "connective tissue spaces" of the pelvis. The comparatively dense portions--the so-called "vessel-nerve guiding plate" and its three "connective tissue pillars"--envelop the vessels and nerves supplying the corresponding pelvic organs. Therefore, they provide ideal landmarks for urologic surgery.

[A new model fo the evaluation of measurements of the neurocranium]. / Ein neuer Ansatz zur metrischen Erfassung des Neurocraniums.

A simple and user-friendly model for trigonometric description of the neurocranium based on newly defined points of measurement is presented. This model not only provides individual description, but also allows for an evaluation of developmental and phylogenetic aspects.

Some anthropological aspects of the prehistoric Tyrolean ice man.

The corpse of a Late Neolithic individual found in a glacier in Oetztal is unusual because of the intact nature of all body parts that resulted from the characteristics of its mummification process and its protected geographical position with regard to glacier flow. Anthropological data indicate that the man was 25 to 40 years old, was between 156 and 160 centimeters in stature, had a cranial capacity of between 1500 and 1560 cubic centimeters, and likely died of exhaustion.