Combined tetrahydrocannabinol and cannabidiol to treat pain in epidermolysis bullosa: a report of three cases.

Epidermolysis bullosa (EB) is a genetic blistering disorder characterized by intense pain related to disease pathology and care-based interventions. Opioid-based therapies underpin pain care in EB; however, they are unable to provide adequate analgesia in a significant proportion of patients. Cannabinoid-based medicines (CBMs) have been studied increasingly for pain conditions of various aetiologies and pose as a novel dimension for pain care in EB. We present three patients with EB who were prescribed pharmaceutical-grade sublingually administered CBMs comprising tetrahydrocannabinol and cannabidiol. All three patients reported improved pain scores, reduced pruritus and reduction in overall analgesic drug intake.

Effect of dust exposure and nitrogen oxides on lung function parameters of German coalminers: a longitudinal study applying GEE regression 1974-1998.

PURPOSE: Workplace limits for dust and nitrogen oxides are under review in Germany and the EU. We conducted a study on German coal miners to determine the effects of exposure on lung function. METHODS: Longitudinal inception cohort study (1974-1998) on miners who began working underground at two coal mines between 1974 and 1979. We determined the number of shifts worked underground, the exposure to coal mine dust, quartz dust, nitrogen oxides (NO, NO(2)), smoking behavior, and three lung function parameters (FVC, FEV(1), FEV(1)/FVC). General estimation equation (GEE) models were fitted. RESULTS: 1,369 miners worked an average 3,017 shifts (S) underground. The mean respirable coal mine dust concentration was 1.89 mg/m(3) (quartz: 0.067 mg/m(3)), and the nitrogen oxide concentrations were 0.58 ppm (NO) and 0.007 ppm (NO(2)). On average, 9 measurements of lung function were available per miner. Compared to reference values, the findings were unexceptionable (103, 101, and 99%) on average. GEE-regression models did not reveal detrimental dust exposure effects. Nitrogen oxides (NO (x) = NO + NO(2)) showed small but clearly insignificant effects on lung function: delta FVC = -0.0008 ml/(220 ppmS), P = 0.86, delta FEV(1) = -0.003 ml/(220 ppmS), P = 0.50 and delta FEV(1)%FVC = -0.07%/(220 ppmS), P = 0.22. CONCLUSIONS: The effect of dust exposure on lung function described in older British and American coal miner studies was not confirmed. This can be explained partly by differences in methods (here: longitudinal studies, no prior exposure), but also by lower dust levels. NO (x) exposures showed no relevant influence on lung function-a result confirming findings from British coal mining.

Crystal structures of human gephyrin and plant Cnx1 G domains: comparative analysis and functional implications.

The molybdenum cofactor (Moco) consists of a unique and conserved pterin derivative, usually referred to as molybdopterin (MPT), which coordinates the essential transition metal molybdenum (Mo). Moco is required for the enzymatic activities of all Mo-enzymes, with the exception of nitrogenase and is synthesized by an evolutionary old multi-step pathway that is dependent on the activities of at least six gene products. In eukaryotes, the final step of Moco biosynthesis, i.e. transfer and insertion of Mo into MPT, is catalyzed by the two-domain proteins Cnx1 in plants and gephyrin in mammals. Gephyrin is ubiquitously expressed, and was initially found in the central nervous system, where it is essential for clustering of inhibitory neuroreceptors in the postsynaptic membrane. Gephyrin and Cnx1 contain at least two functional domains (E and G) that are homologous to the Escherichia coli proteins MoeA and MogA, the atomic structures of which have been solved recently. Here, we present the crystal structures of the N-terminal human gephyrin G domain (Geph-G) and the C-terminal Arabidopsis thaliana Cnx1 G domain (Cnx1-G) at 1.7 and 2.6 A resolution, respectively. These structures are highly similar and compared to MogA reveal four major differences in their three-dimensional structures: (1) In Geph-G and Cnx1-G an additional alpha-helix is present between the first beta-strand and alpha-helix of MogA. (2) The loop between alpha 2 and beta 2 undergoes conformational changes in all three structures. (3) A beta-hairpin loop found in MogA is absent from Geph-G and Cnx1-G. (4) The C terminus of Geph-G follows a different path from that in MogA. Based on the structures of the eukaryotic proteins and their comparisons with E. coli MogA, the predicted binding site for MPT has been further refined. In addition, the characterized alternative splice variants of gephyrin are analyzed in the context of the three-dimensional structure of Geph-G.

Mapping of brain stem neuronal circuitry active during swallowing.

A poorly understood neural circuit in the brain stem controls swallowing. This experiment studied the swallowing circuit in the rat brain stem by means of fos immunocytochemistry. The fos protein is a marker of activated neurons, and under experimental conditions, repetition of a behavior causes the fos protein to be produced in the neurons involved in that behavior. The fos technique has been successfully used to delineate neural circuits involved in reflex glottic closure, cough, and vocalization; however, the technique has not been used to map the swallowing circuit. Nine rats were used in this study. Swallows were evoked in anesthetized rats for 1 hour, then, after a 4-hour delay to allow maximum fos production, the rats were painlessly sacrificed by perfusion. The brain stems were removed and sectioned in the frontal plane, and every fourth section was immunoreacted for fos protein. All sections were examined by light microscopy, and cells positive for fos were marked on drawings of brain stem structures for different levels throughout the brain stem. Control animals underwent sham experiments. After subtraction of the areas of fos labeling seen in controls, all experimental rats showed fos-labeled neurons in very discrete and localized areas, including practically all regions implicated by prior neurophysiology studies of swallowing. The distribution of labeled neurons was more dispersed through the brain stem than current theories of swallowing would suggest. Specifically, recent studies of swallowing control have focused on the nucleus of the solitary tract (NST) and the region surrounding the nucleus ambiguus (periambigual area) just rostral to the obex. These areas contained fos-labeled neurons, but unexpectedly, heavy labeling was found in the same areas caudal to the obex. Areas containing the heaviest labeling were specific subnuclei of the NST and surrounding reticular formation; the periambigual area; and the intermediate reticular zone in the pons and caudal medulla. Interestingly, none of these anatomic structures had uniform fos labeling; this finding suggests that the unlabeled areas are involved in other oromotor behaviors, or that the specific protocol did not activate the full population of swallowing-related neurons. A notable finding of this study is a candidate for the central pattern generator (CPG) of swallowing. Careful lesioning studies in cats strongly suggest that a region in the rostral-medial medulla contains the CPG for swallowing, although the exact location of the CPG was never pinpointed. In the homologous region of the rat brain stem, fos labeling was only found in a small group of neurons within the gigantocellular reticular formation that may be a candidate for the CPG. In summary, correlation with prior physiology experiments suggests that this experiment appears to have delineated many, if not all, of the components of the swallowing circuit for the first time in any mammal. In addition, other areas were found that might also be swallowing-related. One notable example is a small group of fos-labeled cells that may be the CPG for swallowing. Further studies are required to clarify the specific roles of the fos-labeled neurons seen in this study.

Soluble human interleukin-4 receptor is produced by activated T cells under the control of metalloproteinases.

BACKGROUND: Soluble interleukin 4 receptors (sIL-4R) are present in biological fluids. In contrast to mice, in man no distinct mRNA coding for sIL-4R has been described, suggesting that human sIL-4R is exclusively produced by proteolytic cleavage of the cell surface receptor. It is not known whether human sIL-4R is actively produced during an immune response. METHODS: Human purified T cells, CD4+, CD8+, CD45RA+ and CD45R0+ T cell subpopulations were activated in vitro. sIL-4R was determined in the supernatants, cell surface IL-4R was measured by flow cytometry and RT-PCR. RESULTS: Recombinant sIL-4R inhibited IL-4-mediated proliferation and IL-5 upregulation by T cells. sIL-4R could be detected at low levels in supernatants of nonactivated T cells, but at high levels following TCR engagement. This response was paralleled by enhanced transcription and de novo synthesis of the human cell surface IL-4R. Both, activated naive CD45RA+ and memory CD45R0+ T cells, produced sIL-4R with long-lasting kinetics. IL-4 increased sIL-4R production by activated CD45RA+, but there was less of an increase by CD45R0+ T cells. In addition, interferon-gamma enhanced sIL-4R production. Cycloheximide and dexamethasone inhibited sIL-4R production by activated T cells, but did not abolish constitutive release of sIL-4R. Phosphoramidon and 1,10-phenanthroline dose-dependently inhibited shedding of the IL-4R, even in nonactivated T cells. CONCLUSION: The production of human sIL-4R by T cells is regulated by TCR stimuli, IL-4 and IFN-gamma and needs the activity of metalloproteinases. Thus, sIL-4R should be regarded as inducible and due to its IL-4-antagonizing activity an immunoregulatory molecule.

[Hemodynamic effects of a single intravenous administration of prostaglandin E1 in a patient sample with chronic NYHA-stage II/III heart failure]. / Hämodynamische Effekte einer einmaligen intravenösen Applikation von Prostaglandin E1 bei einem Patientenkollektiv mit chronischer Herzinsuffizienz im NYHA-Stadium II/III.

We investigated the hemodynamic effects of a single infusion of PGE1 (60 micrograms infused over a period of 2 h--this is the single dose used in courses of treatment for peripheral occlusive arterial disease) in patients with chronic heart failure NYHA class II-III. The ejection fraction of these patients was < 55%, their average age was 58.4 years (standard deviation 10 years), and their condition was stable. Nineteen of the patients had coronary heart disease and one patient had myocarditis. The hemodynamic data were obtained invasively by catheterization of the right and left heart. Blood pressure and pulse rate were measured manually. Intravenous infusion of 60 micrograms PGE1 over a period of 2 hours did not significantly alter contractility or hemodynamics. Dp/dtmax, dp/dtmax/p, and dp/dt DP40, which are parameters of left ventricular contractility, determined with the aid of a catheter-tip manometer, did not differ significantly over time from those in the placebo control group. Similarly, the other data furnished no evidence that administration of PGE1 had any hemodynamic or myocardial effects. Hence, it is reasonable to state that it is safe to administer PGE1 to patients with peripheral occlusive arterial disease.

Hemodynamic effects of a single intravenous infusion of prostaglandin E1 in patients with clinically moderate to severe chronic heart failure.

In a placebo-controlled, double-blind study, we investigated the hemodynamic effects of a single infusion of prostaglandin E ( 1 ) (PGE ( 1 ); 60 microg infused over a period of 2 hours, the unit dosage used in courses of treatment for peripheral occlusive arterial disease) in 20 patients with moderate to severe chronic heart failure (New York Heart Association functional class II or III). Ejection fraction before therapy was less than 55%, and average age was 58.4 +/- 10 years in these clinically stable patients. Nineteen patients had coronary heart disease and one patient had had myocarditis underlying heart failure. Hemodynamic data were obtained by right- and left-heart catheterization and by Doppler echocardiography. Blood pressure and pulse rate were measured manually. Intravenous infusion of 60 microg PGE ( 1 ) over a period of 2 hours did not significantly alter contractility or hemodynamics. Dp/dt max, dp/dt max/p and dp/dt DP40, measures of left ventricular contractility determined with a catheter-tip manometer, did not differ significantly over time in PGE ( 1 ) -treated patients and those who received placebo. Other measures also failed to reveal PGE ( 1 ) -induced myocardial effects. We conclude that it is safe to administer PGE ( 1 ) to patients with peripheral occlusive arterial disease irrespective of heart failure.

Effects of chronic hypoxia on capillary flow and hematocrit in rat skeletal muscle.

The cremasteric microcirculation was studied in rats exposed to chronic hypoxia. Control male weanling Sprague-Dawley rats (n = 8) were raised for 42-49 days at 752 mmHg. Hypoxic rats (n = 9) were reared for 3 days at 551 mmHg, 4 days at 461 mmHg, 3 days at 371 mmHg, and 31-38 days at 311 mmHg (6,000 m). Red blood cells labeled with fluorescein isothiocyanate were injected. The exposed cremaster was observed using fluorescence microscopy. Rats acutely breathed 10, 21, and 30% O2 spontaneously in random order. Hypoxia-adapted animals had greater (P less than 0.01) red cell flux (10.6 +/- 1.0 vs. 5.7 +/- 0.4/s), capillary hematocrits, capillary-to-systemic hematocrit ratios (0.42 +/- 0.02 vs. 0.33 +/- 0.02), and reduced red cell spacing (11.4 +/- 1.3 vs. 22.3 +/- 2.0 microns) than controls under 21% O2. Chronically hypoxic rats also demonstrated significantly (P less than 0.05) larger capillary diameters (6.52 +/- 0.04 vs. 6.15 +/- 0.06 microns) and greater perfused (135 +/- 5 vs. 94 +/- 3 mm/mm3) and anatomic (182 +/- 5 vs. 151 +/- 8 mm/mm3) microvessel length-densities at 21% O2. Results were generally similar for 10 and 30% O2. Bulk capillary blood flow was significantly (P less than 0.01) greater in controls (2.75 +/- 0.32 vs. 1.87 +/- 0.12 pl/s) only under 30% O2. Our experiments demonstrate that numerous physiological, in addition to anatomic, alterations can occur in the cremasteric microcirculation in response to chronic hypoxia.

Intraperitoneal injection of zymosan in mice induces pain, inflammation and the synthesis of peptidoleukotrienes and prostaglandin E2.

Intraperitoneal injection of zymosan in mice induced rapid extravasation and accumulation of plasma protein in the peritoneal cavity. Neutrophils began to appear in the peritoneal cavity after a lag period of approximately 3 hours. The injected mice exhibited a pain response (writhing) during the first 30 minutes after injection, but writhing ceased before protein or cell accumulation had reached maximum levels. The injection of zymosan induced synthesis of PGE2 (measured by RIA) which reached maximum levels at 30 minutes, then declined slowly. Peptido-leukotriene levels (detected by bioassay, RIA and HPLC) increased rapidly after injection, reached a peak within an hour of injection and declined to undetectable levels within 4 hours. The early peptido-LT was predominantly LTC4, while later, LTE4 was the major component. LTD4 levels remained low throughout and no LTB4 was detected at any time. Indomethacin treatment elevated levels of peptido-LTs, reduced PGE2 levels and inhibited writhing. Phenidone reduced peptido-LT levels. In vitro studies demonstrated that zymosan stimulates LTC4 synthesis by peritoneal cells whereas LTE4, LTD4, LTB4 or monoHETES were not detectable (using HPLC methods). The source of enzymes responsible for the in vivo metabolism of LTC4 to LTD4 and LTE4 could not be identified.

Evaluation of methods of measuring glomerular and nutrient blood flow in rat kidneys.

In normal rats, glomerular plasma flow rates (GPF) were estimated from the uptake of microspheres, and single-nephron filtration rates were estimated by Hanssen's technique in order to calculate single-nephron filtration fractions (SNFF) for outer (C1), middle, (C2), and deep (C3) nephrons. With large microspheres (15 micron), SNFF averaged 0.19, 0.41, and 0.63, and with small microspheres (9 micron), SNFF averaged 0.25, 0.48, and 0.42 for areas C1, C2, and C3, respectively. Kidney filtration fractions (FF) averaged 0.36. Because microsphere experiments in normal rats have generally suggested that SNFF-C1 and FF are similar, we conclude that both types of microspheres overestimated outer cortical plasma flow rates, and probably underestimated inner cortical plasma flow rates. In addition, not all of the smaller microspheres were trapped in the glomeruli. Nutrient blood flow rates were estimated from the uptake of 86Rb. Values ranged from 7.3 ml/g per min in the outer cortex to 4.7 ml/g per min in the inner cortex. Because these values are very similar to measurements made by several other techniques, we conclude that 86Rb uptake adequately estimates cortical blood flow. Medullary blood flow estimates, however, increased with time and were probably too high.

Intrarenal mechanisms of salt retention after bile duct ligation in rats.

In order to study renal salt-retaining mechanisms during the early stages of ascites formation, rats were subjected to bile duct ligation. After this procedure, plasma volumes were found to be reduced and hematocrits slightly increased. The whole-kidney glomberular filtration rate and plasma flows were reduced to 59 and 57% of control values, but the filtration fraction was unchanged. Absolute sodium excretion, as well as the fraction of the filtered sodium load excreted, was also significantly reduced. When micropuncture techniques were used to examine the function of single superficial nephrons, the glomerular filtration rate in these nephrons was found to be reduced to 70% of controlled values, and fractional reabsorption was found to be increased at all accessible sites along the nephron. Filtration by intermediate and juxtamedullary nephrons, determined by Hanssen's technique, was reduced to 55 and 48% of control values. By the use of radioactive microspheres, it was demonstrated that blood flow to superficial, intermediate, and juxtamedullary nephrons was reduced to 49, 59, and 73% of control values. Filtration by superficial nephrons decreased much more than plasma flow--a finding which suggests that the measured increase in fractional reabsorption was associated with an increase in the superficial nephron filtration fraction. From this study, it appears that two factors play an important part in the sodium retention observed in the initial stages of ascites formation following bile duct ligation in rats: (a) a decrease in the filtered sodium load and (b) increased fractional reabsorption by the superficial nephrons--the nephrons which show the least decrease in filtration.