Dermal application of nitric oxide releasing acidified nitrite-containing liniments significantly reduces blood pressure in humans

Vascular ischemic diseases, hypertension, and other systemic hemodynamic and vascular disorders may be the result of impaired bioavailability of nitric oxide (NO). NO but also its active derivates like nitrite or nitroso compounds are important effector and signal molecules with vasodilating properties. Our previous findings point to a therapeutical potential of cutaneous administration of NO in the treatment of systemic hemodynamic disorders. Unfortunately, no reliable data are available on the mechanisms, kinetics and biological responses of dermal application of nitric oxide in humans in vivo. The aim of the study was to close this gap and to explore the therapeutical potential of dermal nitric oxide application. We characterized with human skin in vitro and in vivo the capacity of NO, applied in a NO-releasing acidified form of nitrite-containing liniments, to penetrate the epidermis and to influence local as well as systemic hemodynamic parameters. We found that dermal application of NO led to a very rapid and significant transepidermal translocation of NO into the underlying tissue. Depending on the size of treated skin area, this translocation manifests itself through a significant systemic increase of the NO derivates nitrite and nitroso compounds, respectively. In parallel, this translocation was accompanied by an increased systemic vasodilatation and blood flow as well as reduced blood pressure. We here give evidence that in humans dermal application of NO has a therapeutic potential for systemic hemodynamic disorders that might arise from local or systemic insufficient availability of NO or its bio-active NO derivates, respectively.     

Mnl2, a novel component of the ER associated protein degradation pathway

In eukaryotes, membrane and soluble proteins of the secretory pathway enter the endoplasmic reticulum (ER) after synthesis in an unfolded state. Directly after entry, most proteins are modified with glycans at suitable glycosylation sites and start to fold. A protein that cannot fold properly will be degraded in a process called ER associated degradation (ERAD). Failures in ERAD, either by loss of function or by premature degradation of proteins, are a cause of severe diseases. Therefore, the search for novel ERAD components to gain better insight in this process is of high importance. Carbohydrate trimming is a relevant process in ER quality control. In this work a novel putative yeast mannosidase encoded by the open reading frame YLR057W was identified and named Mnl2. Deletion of MNL2 diminished the degradation efficiency of misfolded CPY^* in the absence of the cognate mannosidase Mnl1, indicating a specific role in ERAD.     

Markers of enhanced cholesterol absorption are a strong predictor for cardiovascular diseases in patients without diabetes mellitus

Objective

Hypercholesterolemia is a major risk factor for cardiovascular disease (CVD), and diabetes mellitus and statin treatment affect cholesterol metabolism. The aim of the present study was to evaluate markers of cholesterol metabolism and determine their relationship with CVD in patients without diabetes mellitus who were not receiving statin treatment.

Methods

In addition to conventional CVD risk factors, plasma levels of campesterol and sitosterol (indicators of cholesterol absorption) and lathosterol (an indicator of cholesterol synthesis) were determined in 835 consecutive patients referred for coronary angiography. Coronary artery disease was evaluated by coronary angiograms, carotid atherosclerosis and peripheral vascular disease were assessed by Doppler ultrasound, and cerebrovascular accidents and transient ischemic attacks were identified by medical history.

Results

After excluding patients with known diabetes mellitus and those receiving statin treatment, 177 patients were included in the analysis. Compared to patients without CVDs (n = 111), patients with concomitant CVDs (n = 66) had a reduced lathosterol-to-cholesterol ratio (1.25 ± 0.61 vs. 1.38 ± 0.63, P < 0.05) and an increased campesterol-to-cholesterol ratio (1.81 ± 1.04 vs. 1.50 ± 0.69, P < 0.05), indicating that enhanced absorption and reduced synthesis of cholesterol is associated with CVD development. Logistic regression analysis including all established cardiovascular risk factors (age, sex, total cholesterol, arterial hypertension, body mass index and smoking) revealed that campesterol and the campesterol-to-cholesterol ratio were significant predictors of concomitant CVD in this patient population.

Conclusion

In patients without diabetes mellitus, markers of enhanced cholesterol absorption were a strong predictor for concomitant CVD.     

Different capacity of monocyte subsets to phagocytose iron-oxide nanoparticles

Objective

To explore the capacity of human CD14⁺CD16⁺⁺ and CD14⁺⁺CD16^- monocytes to phagocyte iron-oxide nanoparticles in vitro.

Methods

Human monocytes were labeled with four different magnetic nanoparticle preparations (Ferumoxides, SHU 555C, CLIO-680, MION-48) exhibiting distinct properties and cellular uptake was quantitatively assessed by flow cytometry, fluorescence microscopy, atomic absorption spectrometry and Magnetic Resonance Imaging (MRI). Additionally we determined whether cellular uptake of the nanoparticles resulted in phenotypic changes of cell surface markers.

Results

Cellular uptake differed between the four nanoparticle preparations. However for each nanoparticle tested, CD14⁺⁺CD16^- monocytes displayed a significantly higher uptake compared to CD14⁺CD16⁺⁺ monocytes, this resulted in significantly lower T1 and T2 relaxation times of these cells. The uptake of iron-oxide nanoparticles further resulted in a remarkable shift of expression of cell surface proteins indicating that the labeling procedure affects the phenotype of CD14⁺CD16⁺⁺ and CD14⁺⁺CD16^- monocytes differently.

Conclusion

Human monocyte subsets internalize different magnetic nanoparticle preparations differently, resulting in variable loading capacities, imaging phenotypes and likely biological properties.     

Molecular cloning and functional characterization of the bovine (Bos taurus) organic anion transporting polypeptide Oatp1a2 (Slco1a2)

We describe the cloning, functional characterization and tissue localization of a novel membrane transporter of the OATP/Oatp-gene family obtained from liver and kidney of cattle (Bos taurus). The carrier protein exhibits highest sequence identity to the human OATP1A2 (previously called OATP-A) and is, therefore, named bovine Oatp1a2. Bovine Oatp1a2 received the gene symbol Slco1a2 that is identical to the SLC classification of human OATP1A2 (SLCO1A2, previously called SLC21A3) and is likely an orthologue of the human gene. Two different full-length bOatp1a2 cDNAs of 2316-bp and 3504-bp were obtained and encoded for a 666 amino acid membrane protein, which contains twelve putative transmembrane spanning domains. Bovine Oatp1a2 expression was detected in liver, kidney, brain and adrenal gland. Uptake studies in cRNA-injected oocytes demonstrated that bOatp1a2 transports estrone-3-sulfate and taurocholate, with K(m) values of 9.6 microM and 51 microM, respectively, and estradiol-17beta-glucuronide. However, the structurally-related heart glycosides ouabain (1 microM) and digoxin (1 microM) are neither transported by bovine Oatp1a2 nor by human OATP1A2. We conclude that based on the tested substrates bovine Oatp1a2 shows functional homology to human OATP1A2.     

Short/branched-chain acyl-CoA dehydrogenase deficiency due to an IVS3+3A>G mutation that causes exon skipping

Short/branched-chain acyl-CoA dehydrogenase deficiency (SBCADD) is an autosomal recessive disorder of l-isoleucine catabolism. Little is known about the clinical presentation associated with this enzyme defect, as it has been reported in only a limited number of patients. Because the presence of C5-carnitine in blood may indicate SBCADD, the disorder may be detected by MS/MS-based routine newborn screening. It is, therefore, important to gain more knowledge about the clinical presentation and the mutational spectrum of SBCADD. In the present study, we have studied two unrelated families with SBCADD, both with seizures and psychomotor delay as the main clinical features. One family illustrates the fact that affected individuals may also remain asymptomatic. In addition, the normal level of newborn blood spot C5-acylcarnitine in one patient underscores the fact that newborn screening by MS/MS currently lacks sensitivity in detecting SBCADD. Until now, seven mutations in the SBCAD gene have been reported, but only three have been tested experimentally. Here, we identify and characterize an IVS3+3A>G mutation (c.303+3A>G) in the SBCAD gene, and provide evidence that this mutation is disease-causing in both families. Using a minigene approach, we show that the IVS3+3A>G mutation causes exon 3 skipping, despite the fact that it does not appear to disrupt the consensus sequence of the 5′ splice site. Based on these results and numerous literature examples, we suggest that this type of mutation (IVS+3A>G) induces missplicing only when in the context of non-consensus (weak) 5′ splice sites. Statistical analysis of the sequences shows that the wild-type versions of 5′ splice sites in which +3A>G mutations cause exon skipping and disease are weaker on average than a random set of 5′ splice sites. This finding is relevant to the interpretation of the functional consequences of this type of mutation in other disease genes.     

Regulatory effects of synthetic liver X receptor- and peroxisome-proliferator activated receptor agonists on sterol transport pathways in polarized cerebrovascular endothelial cells

The blood-brain barrier contributes to maintain brain cholesterol metabolism and protects this uniquely balanced system from exchange with plasma lipoprotein cholesterol. Brain capillary endothelial cells, representing a physiological barrier to the central nervous system, express apolipoprotein A-I (apoA-I, the major high-density lipoprotein (HDL)-associated apolipoprotein), ATP-binding cassette transporter A1 (ABCA1), and scavenger receptor, class B, type I (SR-BI), proteins that promote cellular cholesterol mobilization. Liver X receptors (LXRs) and peroxisome-proliferator activated receptors (PPARs) are regulators of cholesterol transport, and activation of LXRs and PPARs has potential therapeutic implications for lipid-related neurodegenerative diseases. To clarify the functional impact of LXR/PPAR activation, sterol transport along the: (i) ABCA1/apoA-I and (ii) SR-BI/HDL pathway was investigated in primary, polarized brain capillary endothelial cells, an in vitro model of the blood-brain barrier. Activation of LXR (24(S)OH-cholesterol, TO901317), PPARalpha (bezafibrate, fenofibrate), and PPARgamma (troglitazone, pioglitazone) modulated expression of apoA-I, ABCA1, and SR-BI on mRNA and/or protein levels without compromising transendothelial electrical resistance or tight junction protein expression. LXR-agonists and troglitazone enhanced basolateral-to-apical cholesterol mobilization in the absence of exogenous sterol acceptors. Along with the induction of cell surface-located ABCA1, several agonists enhanced cholesterol mobilization in the presence of exogenous apoA-I, while efflux of 24(S)OH-cholesterol (the major brain cholesterol metabolite) in the presence of exogenous HDL remained unaffected. Summarizing, in cerebrovascular endothelial cells apoA-I, ABCA1, and SR-BI represent drug targets for LXR and PPAR-agonists to interfere with cholesterol homeostasis at the periphery of the central nervous system.     

Der Einfluß der Umweltfaktoren auf das Wachstum und den Häutungsrhythmus der StrandkrabbeCarcinides maenas

Zusammenfassung 1. Junge Strandkrabben von 4–16 mm Carapaxbreite wurden bis zur Geschlechtsreife unter konstanten Umweltbedingungen aufgezogen.2. Die Dauer ihrer Häutungsintervalle nimmt bei konstanter Temperatur mit der Körpergröße stetig zu.3. Die Dauer der Häutungsintervalle hängt von der Temperatur und der Ernährung ab. Von der Tageslänge scheint sie weitgehend unabhängig zu sein.4. Der relative Grösßenzuwachs bei jeder Häutung ist im gesamten untersuchten Größenbereich und bei den verschiedenen Temperaturen bei allen Häutungen gleich: Bei den Häutungen verdoppelt sich jeweils das Körpervolumen.5. Augenstielamputationen und Verlust von Extremitäten wirken auf den Häutungsrhythmus in gleicher Weise: Die Schwankungsbreite in der Dauer der Häutungsintervalle ist vermindert. Die Häutungsintervalle sind in 20° C deutlich, in 10° C nur geringfügig verkürzt.6. Durch die Anwesenheit größerer Artgenossen werden die Häutungen verzögert. Die optische Wahrnehmung spielt dabei keine Rolle.7. Aus diesen Ergebnissen wird folgendes geschlossen: Der ausschlaggebende Faktor für die Auslösung von Häutungen ist ein bestimmter Größenzuwachs. Temperatur und Ernährung beeinflussen den Häutungsrhythmus dadurch, daß sie das Tempo des Wachstums bestimmen. Die winterliche Häutungsruhe in Freilandpopulationen wird nicht durch den Kurztag bedingt, sondern durch die Kälte. Diese hemmt lediglich das Wachstum, sie verhindert nicht die Häutungen über das häutungshemmende Hormon. Dieses vermindert vielmehr die Temperaturabhängigkeit des Häutungsrhythmus, indem es die Häutungen im Warmen stärker verzögert als im Kalten. Es gestattet die Anpassung des Häutungstermins an die individuelle Lage der Tiere. Es hemmt in Anwesenheit größerer Artgenossen die Häutung. Beim Verlust mehrerer Gliedmaßen wird seine Sekretion eingestellt, so daß die nächste Häutung vorzeitig erfolgt. Das häutungshemmende Hormon bedingt dementsprechend die große individuelle Variation in der Dauer der Häutungsintervalle.

---

The effect of environmental factors on growth and moulting rhythm in the shore crab,Carcinides maenas

Young crabs (carapace width 4 to 16 mm) were raised under controlled conditions in the laboratory. The time intervals between subsequent moults increase at all test temperatures with increasing body size. The length of intermoult periods varies with temperature and feeding. It is not affected by day length. Moulting takes place as soon as a certain increase in size is attained. In comparable size groups, the amount of this increase is identical in all test temperatures. Moreover, the relation of increase to initial size is constant over the whole size range investigated. The body volume doubles at each moult. Eyestalk amputations and loss of extremities have similar effects: They shorten the intermoult periods at 20° C considerably, but at 10° C they do so only slightly; furthermore, the amplitude of fluctuations is narrowed. The presence of large specimens tends to retard moulting in smaller ones; this response is independent of visual stimuli. The following assumptions are made: Low temperatures retard the moulting rhythm directly by slowing down growth. They are not acting via the moult inhibiting hormone. Loss of several extremities causes a stop of hormone delivery resulting in shortened intermoult periods. Recognition by touch of a larger specimen causes increased hormone delivery and thus retardation of the subsequent moulting process.