De Lisle RC, Roach E, Jansson K. Effects of laxative and N-acetylcysteine on mucus accumulation, bacterial load, transit, and inflammation in the cystic fibrosis mouse small intestine. Am J Physiol Gastrointest Liver Physiol 293: G577-G584, 2007. First published July 5, 2007; doi:10.1152/ajpgi.00195.2007.-The accumulation of mucus in affected organs is characteristic of cystic fibrosis (CF). The CF mouse small intestine has dramatic mucus accumulation and exhibits slower interdigestive intestinal transit. These factors are proposed to play cooperative roles that foster small intestinal bacterial overgrowth (SIBO) and contribute to the innate immune response of the CF intestine. It was hypothesized that decreasing the mucus accumulation would reduce SIBO and might improve other aspects of the CF intestinal phenotype. To test this, solid chow-fed CF mice were treated with an osmotic laxative to improve gut hydration or liquid-fed mice were treated orally with N-acetylcysteine (NAC) to break mucin disulfide bonds. Treatment with laxative or NAC reduced mucus accumulation by 43% and 50%, respectively, as measured histologically as dilation of the intestinal crypts. Laxative and NAC also reduced bacterial overgrowth in the CF intestine by 92% and 63%, respectively. Treatment with laxative normalized small intestinal transit in CF mice, whereas NAC did not. The expression of innate immune response-related genes was significantly reduced in laxative-treated CF mice, whereas there was no significant effect in NACtreated CF mice. In summary, laxative and NAC treatments of CF mice reduced mucus accumulation to a similar extent, but laxative was more effective than NAC at reducing bacterial load. Eradication of bacterial overgrowth by laxative treatment was associated with normalized intestinal transit and a reduction in the innate immune response. These results suggest that both mucus accumulation and slowed interdigestive small intestinal transit contribute to SIBO in the CF intestine.small intestinal bacterial overgrowth MICE that lack the expression of the cystic fibrosis (CF) transmembrane conductance regulator (Cftr) gene have a severe intestinal phenotype with many of the problems that occur in human CF patients. These include the accumulation of mucus, intestinal obstruction, slowed small intestinal transit, small intestinal bacterial overgrowth (SIBO), inflammation, and failure to thrive (4 -6, 30). Of these changes, bacterial overgrowth is of interest because it can be responsible for the majority of the other aspects of the intestinal phenotype (22, 34).Secreted gel-forming mucins, mainly Muc2 in the intestine, bind bacteria and help carry them aborally for efficient clearance from the small intestine. When mucus clearance is impaired, bacteria can proliferate in the mucus and may use it as an energy source (36). Mucus accumulation in the CF intestine is in part due to the dehydrated, acidic environment as well as the altered glycosylation of mucins (38, 40), which may increase the viscosity of mucus. Our labora...

Alport disease is caused by mutations in genes encoding the ␣3, ␣4, or ␣5 chains of type IV collagen, which form the collagenous network of mature glomerular basement membrane (GBM). In the absence of ␣3, ␣4, ␣5 (IV) collagen, ␣1, ␣2 (IV) collagen persists, which ordinarily is found only in GBM of developing kidney. In addition to dysregulation of collagen IV, Alport GBM contains aberrant laminins, which may contribute to the progressive GBM thickening and splitting, proteinuria, and renal failure seen in this disorder. This study sought to characterize further the laminin dysregulation in collagen ␣3(IV) knockout mice, a model of Alport disease. With the use of confocal microscopy, laminin ␣1 and ␣5 abundance was quantified, and it was found that they co-distributed in significantly large amounts in areas of GBM thickening. In addition, labeling of entire glomeruli for laminin ␣5 was significantly greater in Alport mice than in wild-type siblings. Reverse transcriptase-PCR from isolated glomeruli demonstrated significantly more laminin ␣5 mRNA in Alport mice than in wild-type controls, indicating upregulated transcription of Lama5. For testing glomerular barrier function, ferritin was injected into 2-wk-old Alport and control mice, and GBM was examined by electron microscopy. Highest ferritin levels were seen in Alport GBM thickenings beneath effaced podocyte foot processes, but morphologically normal GBM was significantly permeable as well. We concluded that (1) ultrastructurally normal Alport GBM residing beneath differentiated podocyte foot processes is inherently and abnormally permeable, and (2) upregulation of Lama5 transcription and concentration of laminin ␣1 and ␣5 within Alport GBM thickenings contribute to abnormal permeabilities.

II.

Eradication of bacterial overgrowth in CF mice significantly decreased mucus secretion and accumulation in intestinal crypts without an effect on mucin gene expression. It is proposed that bacterial overgrowth stimulates mucus secretion, which contributes to its accumulation in the small intestine. Control of bacterial overgrowth is expected to reduce mucus accumulation and may improve intestinal function and overall health in CF.

This VTEP protocol was determined to afford a good risk-to-benefit ratio for a wide variety of neurosurgical procedures.

BackgroundCystic fibrosis (CF) is caused by mutations in the CFTR gene that impair the function of CFTR, a cAMP-regulated anion channel. In the small intestine loss of CFTR function creates a dehydrated, acidic luminal environment which is believed to cause an accumulation of mucus, a phenotype characteristic of CF. CF mice have small intestinal bacterial overgrowth, an altered innate immune response, and impaired intestinal transit. We investigated whether lubiprostone, which can activate the CLC2 Cl- channel, would improve the intestinal phenotype in CF mice.MethodsCftrtm1UNC (CF) and wildtype (WT) littermate mice on the C57BL/6J background were used. Lubiprostone (10 μg/kg-day) was administered by gavage for two weeks. Mucus accumulation was estimated from crypt lumen widths in periodic acid-Schiff base, Alcian blue stained sections. Luminal bacterial load was measured by qPCR for the bacterial 16S gene. Gastric emptying and small intestinal transit in fasted mice were assessed using gavaged rhodamine dextran. Gene expression was evaluated by Affymetrix Mouse430 2.0 microarray and qRT-PCR.ResultsCrypt width in control CF mice was 700% that of WT mice (P < 0.001). Lubiprostone did not affect WT crypt width but, unexpectedly, increased CF crypt width 22% (P = 0.001). Lubiprostone increased bacterial load in WT mice to 490% of WT control levels (P = 0.008). Conversely, lubiprostone decreased bacterial overgrowth in CF mice by 60% (P = 0.005). Lubiprostone increased gastric emptying at 20 min postgavage in both WT (P < 0.001) and CF mice (P < 0.001). Lubiprostone enhanced small intestinal transit in WT mice (P = 0.024) but not in CF mice (P = 0.377). Among other innate immune markers, expression of mast cell genes was elevated 4-to 40-fold in the CF intestine as compared to WT, and lubiprostone treatment of CF mice decreased expression to WT control levels.ConclusionsThese results indicate that lubiprostone has some benefits for the CF intestinal phenotype, especially on bacterial overgrowth and the innate immune response. The unexpected observation of increased mucus accumulation in the crypts of lubiprostone-treated CF mice suggests the possibility that lubiprostone increases mucus secretion.

To localize binding sites for peptide YY (PYY) in the pancreas we utilized a slide-mount autoradiographic technique on frozen sections of rat pancreas incubated with 125I-Tyr36-PPY. Saturable autoradiographic labeling was located over pancreatic blood vessels, whereas islets, acinar cells, ducts, and neural elements did not appear to be specifically labeled. Isolated vascular fragments were prepared by collagenase digestion of rat pancreas. Binding experiments with 125I-Tyr36-PYY showed saturable binding to the fraction enriched in blood vessels but not to acini. Inhibition of 125I-Tyr36-PYY binding by nonradioactive neuropeptide Y (NPY) and PYY were similar, with half-maximal inhibition at 31.2 +/- 5 pM (n = 6); the potency of pancreatic polypeptide (PP) was 10,000 times lower. The binding site was classified as belonging to a Y1 type of NPY and/or PYY receptors, since [Leu31,Pro34]NPY, a specific Y1-receptor agonist, inhibited binding similar to NPY. To further localize the bound [125I-Tyr36]PYY within the blood vessels, light- and electron-microscopic autoradiographs were prepared and quantitated. Autoradiographic grains were located predominantly over vascular smooth muscle cells, although saturable localization was also seen over endothelial cells. It is concluded that in the pancreas Y1 receptors are predominantly located on vascular smooth muscle cells.

The uptake of 1251-insulin by rat pancreas was studied in vivo. Following fixation and light microscope autoradiography, saturable uptake of 1251-insulin was quantitatively demonstrated on acinar and duct cells but not on blood vessels and islets of Langerhans. Electron microscopy revealed the localization of '251-insulin to the basolateral cell membranes of acinar and duct cells.497