Club cell protein (CC16) in plasma,bronchial brushes,BAL and urine following an inhaled allergen challenge in allergic asthmatics

Background: Club cell protein (CC16) is a pneumoprotein secreted by epithelial club cells. CC16 possesses anti-inflammatory properties and is a potential biomarker for airway epithelial damage. We studied the effect of inhaled allergen on pulmonary and systemic CC16 levels.

Methods: Thirty-four subjects with allergic asthma underwent an inhaled allergen challenge. Bronchoscopy with bronchoalveolar lavage (BAL) and brushings was performed before and 24?h after the challenge. CC16 was quantified in BAL and CC16 positive cells and CC16 mRNA in bronchial brushings. CC16 was measured in plasma and urine before and repeatedly after the challenge. Thirty subjects performed a mannitol inhalation challenge prior to the allergen challenge.

Results: Compared to baseline, CC16 in plasma was significantly increased in all subjects 0–1?h after the allergen challenge, while CC16 in BAL was only increased in subjects without a late allergic response. Levels of CC16 in plasma and in the alveolar fraction of BAL correlated significantly after the challenge. There was no increase in urinary levels of CC16 post-challenge. Mannitol responsiveness was greater in subjects with lower baseline levels of CC16 in plasma.

Conclusions: The increase in plasma CC16 following inhaled allergen supports the notion of CC16 as a biomarker of epithelial dysfunction.     

Migrating action potential complexes--a feature of normal jejunal myoelectric activity in the rat

To determine if migrating action potential complexes (MAPCs) are a feature of normal motility, Hooded-Lister rats (100-150 g) were surgically prepared with three pairs of bipolar jejunal electrodes spaced 2.5 cm apart and with a jejunostomy tube for motility recording. Animals were studied conscious and unrestrained on postoperative day 14 after an 18-h fast. Intestinal myoelectric and motor activity was recorded for a 1-h interval in 24 animals that continued to fast and in 12 animals that were allowed to feed for 10 min. Fasting rats had a jejunal slow wave frequency of 32 +/- 2 contractions/min which did not differ significantly after feeding. Migrating myoelectric complexes (MMC) were clearly identified in all fasting animals and had a cycle period of 10.0 +/- 3.6 min. MAPCs were seen during phase II in 83% of MMCs and had an average distribution of 4.2 +/- 3.9/MMC. Feeding abolished the MMC and initiated a continuous irregular pattern of electrical spiking and associated contractile activity. Migrating action potential complexes were seen after feeding with a frequency of 1.8 +/- 0.4/min. It is concluded that MAPCs are a common feature of normal interdigestive phase II and also of postprandial jejunal motility in the rat. This supports the hypothesis that the MAPC is a basic propulsive motor pattern intrinsic to normal intestinal function.     

Relationship of postprandial motilin, gastrin, and pancreatic polypeptide release to intestinal motility during vagal interruption.

Experiments were performed to determine how postprandial motilin, gastrin, and pancreatic polypeptide plasma concentrations measured during vagal blockade relate to coincident small intestinal motility patterns. Feeding produced a postprandial pattern of intestinal motility coincident with a sustained increase in gastrin and pancreatic polypeptide and a decline in motilin plasma concentrations. Vagal blockade replaced the fed pattern with one similar to migrating motor complex (MMC) activity. Highest motilin plasma concentrations were observed during phase III of this MMC-like activity, as occurs in the fasted state. Vagal blockade reduced but did not abolish the postprandial increase in plasma gastrin and pancreatic polypeptide concentrations. Termination of vagal cooling produced a decline in motilin and an elevation in gastrin and pancreatic polypeptide concentrations, coincident with the return of the fed pattern. In conclusion, during vagal blockade in the fed state (i) motilin, but not gastrin or pancreatic polypeptide plasma concentrations, fluctuate with the MMC-like activity, and any measurement of motilin concentration under these circumstances must be interpreted on the basis of gut motility patterns, and (ii) gastrin and pancreatic polypeptide concentrations are marginally elevated, but these changes are not enough to disrupt the MMC or have any motor effect. Lastly, the fed pattern and the postprandial changes in motilin, gastrin, and pancreatic polypeptide concentrations are in part dependent upon intact vagal pathways.     

The effects of cholecystokinin-octapeptide and pentagastrin on electrical and motor activities of canine colonic circular muscle

The effects of cholecystokinin-octapeptide (CCK-OP) and pentagastrin on electrical and motor activities of circular muscle of the canine colon were studied with the sucrose gap technique. Additional organ bath experiments were performed to further characterize the motor response to the peptides and to elucidate their site of action. The electrical activity consisted of slow waves having an initial potential followed by a plateau potential, at a regular frequency of 4.5 cycles/min. Both peptides prolonged the duration and increased the amplitude of the plateau phase of the slow waves. Concomitantly, the slow wave frequency was reduced. In addition, CCK-OP increased spiking activity. Both spiking activity and the prolonged plateau potential generated contractile activity, prolonged phasic contraction occurring with slow waves with a prolonged plateau. In organ bath experiments, both CCK-OP and pentagastrin increased the basal tone of the muscle strips and prolonged the duration of the phasic contractions. The prolongation of the duration of the contractions was not antagonized by tetrodotoxin (TTX) and atropine. CCK-OP but not pentagastrin increased the force of contractions, this action was not affected by atropine but was reduced in the presence of TTX, suggesting that the increase in force may be partially mediated by noncholinergic excitatory nerves. The increase in basal tension by the peptides was enhanced in the presence of TTX indicating that myenteric inhibitory neurones were tonically active under our experimental conditions. The results provide the electrophysiological basis for CCK-OP and pentagastrin induced changes in colonic motility.     

Interaction of cortisol with the neural retina of the chick embryo in culture.

The uptake of cortisol and the kinetics of hormone-receptor interaction in the cytosol and cell nucleus were investigated in the intact tissue in organ culture. Cortisol is concentrated by the neural retina. The accumulation of the free steroid is temperature dependent but the effect of temperature decreases with the increase of cortisol in medium. Cortisol binding to specific receptors in the cytosol shows a sigmoidal type of kinetics which correlates well with the kinetics of glutamine-synthetase induction by cortisol. The temperature dependent translocation of the receptor-hormone complexes to the nuclei and the effect of detergents on the binding to nuclei are presented.     

Feline lower esophageal sphincter sling and circular muscles have different functional inhibitory neuronal responses

The lower esophageal sphincter (LES) has a circular muscle component exhibiting spontaneous tone that is relaxed by nitric oxide (NO) and a low-tone sling muscle that contracts vigorously to cholinergic stimulation but with little or no evidence of NO responsiveness. This study dissected the responses of the sling muscle to nitrergic innervation in relationship to its cholinergic innervation and circular muscle responses. Motor responses were induced by electrical field stimulation (EFS; 1-30 Hz) of muscle strips from sling and circular regions of the feline LES in the presence of cholinergic receptor inhibition (atropine) or NO synthase inhibition [NG-nitro-L-arginine (L-NNA)+/-atropine]. This study showed the following. First, sling muscle developed less intrinsic resting tone compared with circular muscle. Second, with EFS, sling muscle contracted (most at 50% by 5 Hz. Third, on neural blockade with atropine or L-NNA+/-atropine, 1) sling muscle, although predominantly influenced by excitatory cholinergic stimulation, had a small neural NO-mediated inhibition, with no significant non-NO-mediated inhibition and 2) circular muscle, although little affected by cholinergic influence, underwent relaxation predominantly by neural release of NO and some non-NO inhibitory influence (at higher EFS frequency). Fourth, the sling, precontracted with bethanecol, could relax with NO and some non-NO inhibition. Finally, the tension range of both muscles is similar. In conclusion, sling muscle has limited NO-mediated inhibition to potentially augment or replace sling relaxation effected by switching off its cholinergic excitation. Differences within the LES sling and circular muscles could provide new directions for therapy of LES disorders.     

Dasyatispora levantinae gen. et sp. nov., a new microsporidian parasite from the common stingray Dasyatis pastinaca in the eastern Mediterranean

A new microsporidian infecting the Mediterranean common stingray Dasyatis pastinaca (Linnaeus, 1758) is described from Iskenderun Bay, Turkey. The parasite invades the disc muscles, producing slender, spindle-shaped subcutaneous swellings that develop into massive, elongated, tumor-like protuberances measuring up to 11 x 4 cm. Severity of the infection may vary from light (1 or 2 small lesions) to intense, with large parts of the dorsal surface covered with lumps and protrusions. These masses contained a yellowish-white caseous substance consisting of degraded host tissue and microsporidian sporophorous vesicles, which in turn contained developing sporonts, sporoblasts and spores. The ripe spore contained a uni-nucleate sporoplasm and large posterior vacuole, and measured 3.8-4.3 x 2.6-2.8 microm. Infection prevalence was 21% in a sample of 143 host individuals examined. All the infected stingray individuals were within the weight class of 300 to 800 g (200 to 305 mm disc width). Phylogenetic analyses of rDNA sequences indicate that this microsporidian belongs to the Pleistophoridae and clusters with species of the genera Ovipleistophora Pekkarinen, Lom & Nilsen, 2002 and Heterosporis Schubert, 1969. However, the morphology, development and host differ distinctly from all reported species, including those belonging to these 2 genera, and it is thus assigned to a newly erected genus and named Dasyatispora levantinae gen. et sp. nov. This is the first record of a microsporidian infection in a batoid. It is also the first microsporidian species to be formally described from an elasmobranch.     

Native folding of aggregation-prone recombinant proteins in Escherichia coli by osmolytes, plasmid- or benzyl alcohol-overexpressed molecular chaperones

When massively expressed in bacteria, recombinant proteins often tend to misfold and accumulate as soluble and insoluble nonfunctional aggregates. A general strategy to improve the native folding of recombinant proteins is to increase the cellular concentration of viscous organic compounds, termed osmolytes, or of molecular chaperones that can prevent aggregation and can actively scavenge and convert aggregates into natively refoldable species. In this study, metal affinity purification (immobilized metal ion affinity chromatography [IMAC]), confirmed by resistance to trypsin digestion, was used to distinguish soluble aggregates from soluble nativelike proteins. Salt-induced accumulation of osmolytes during induced protein synthesis significantly improved IMAC yields of folding-recalcitrant proteins. Yet, the highest yields were obtained with cells coexpressing plasmid-encoded molecular chaperones DnaK-DnaJ-GrpE, ClpB, GroEL-GroES, and IbpA/B. Addition of the membrane fluidizer heat shock-inducer benzyl alcohol (BA) to the bacterial medium resulted in similar high yields as with plasmid-mediated chaperone coexpression. Our results suggest that simple BA-mediated induction of endogenous chaperones can substitute for the more demanding approach of chaperone coexpression. Combined strategies of osmolyte-induced native folding with heat-, BA-, or plasmid-induced chaperone coexpression can be thought to optimize yields of natively folded recombinant proteins in bacteria, for research and biotechnological purposes.