A Novel Method of Preparation of Small Intestinal Brush Border Membrane Vesicles by Polyethylene Glycol Precipitation

Ramamoorthy, Prabhu; Balasubramanian, K.A.

doi:10.1006/abio.2000.4908

Cited by 36 publications

(18 citation statements)

References 29 publications

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“…The distal part of the jejunum and the ileum of control and CCl 4 -treated rats was used for the isolation of brush border membranes (BBM) and surfactant as described earlier. 25,26 Purity of the isolated BBM was checked by enrichment of the marker enzyme alkaline phosphatase. Isolated BBM were assessed for their ability to transport glucose by uptake measurements carried out using the rapid filtration technique, at room temperature as described.…”

Section: Methodsmentioning

confidence: 99%

Intestinal mucosal alterations in rats with carbon tetrachloride‐induced cirrhosis

et al. 2006

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Spontaneous bacterial peritonitis is a major cause of mortality after liver cirrhosis. Altered permeability of the mucosa and deficiencies in host immune defenses through bacterial translocation from the intestine due to intestinal bacterial overgrowth have been implicated in the development of this complication. Molecular mechanisms underlying the process are not well known. In order to understand mechanisms involved in translocation of bacteria, this study explored the role of oxidative stress in mediating changes in intestinal mucosal glycosylation and luminal bacterial content during cirrhosis. CCl 4 -induced cirrhosis in rats led to prolonged oxidative stress in the intestine, accompanied by increased sugar content of both intestinal brush border and surfactant layers. This was accompanied by changes in bacterial flora in the gut, which showed increased hydrophobicity and adherence to the mucosa. Inhibition of xanthine oxidase using sodium tungstate or antioxidant supplementation using vitamin E reversed the oxidative stress, changes in brush border membrane sugar content, and bacterial adherence. In conclusion, oxidative stress in the intestine during cirrhosis alters mucosal glycosylation, accompanied by an increased hydrophobicity of luminal bacteria, enabling increased bacterial adherence onto epithelial cells. This might facilitate translocation across the mucosa, resulting in complications such as spontaneous bacterial peritonitis. (HEPATOLOGY 2006;43:837-846.)

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Section: Methodsmentioning

confidence: 99%

Intestinal mucosal alterations in rats with carbon tetrachloride‐induced cirrhosis

et al. 2006

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“…This material was used for the preparation of 3 % homogenates using Tris/ mannitol buffer [2 mM Tris/HCl (pH 7.1) containing 50 mM mannitol] as described previously [19].…”

Section: Preparation Of Mucosal Homogenate and Assay Of Enzymesmentioning

confidence: 99%

Oral glutamine attenuates indomethacin-induced small intestinal damage

2004

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The use of NSAIDs (non-steroidal anti-inflammatory drugs), although of great therapeutic value clinically, is limited by their tendency to cause mucosal damage in the gastrointestinal tract. In the small intestine, the effects these drugs have been shown to produce include inhibition of cyclo-oxygenase, mitochondrial dysfunction and free radical-induced oxidative changes, all of which contribute to the mucosal damage seen. Glutamine is a fuel preferentially used by enterocytes and is known to contribute to maintaining the integrity of these cells. In the present study, we investigated the effect of glutamine on indomethacin-induced changes in the small intestinal mucosa. Rats were given 2% glutamine or glutamic acid or isonitrogenous amino acids, glycine or alanine, in the diet for 7 days. Indomethacin was then administered orally at a dose of 40 mg/kg of body weight. After 1 h, the small intestine was removed and used for the measurement of parameters of oxidative stress and mitochondrial and BBM (brush border membrane) function. Evidence of oxidative stress was found in the mucosa of the small intestine of drug-treated rats, as indicated by significantly increased activity of xanthine oxidase (P < 0.001) and myeloperoxidase (P < 0.001), with corresponding decreases in the levels of several free radical scavenging enzymes and alpha-tocopherol (P < 0.001 in all cases). Levels of products of peroxidation were also significantly elevated (P < 0.001 for all the parameters measured). In addition, oxidative stress was evident in isolated intestinal mitochondria and BBMs (P < 0.001 for all the parameters measured), with associated alterations in function of these organelles (P < 0.001 for all the parameters measured). Supplementation of the diet with glutamine or glutamic acid prior to treatment with indomethacin produced significant amelioration in all the effects produced by the drug in the small intestine (P < 0.001 for all the parameters measured). Glycine and alanine were found to be much less effective in these respects.

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“…It is believed that these unique conditions for BBMV immobilization on the L1 sensor chip are principally governed by BBMV composition containing phospholipids with different charges and proteins. Although BBMV composition is a little bit different depending on preparation methods and animal species, a significant portion (> 40%) is known to be occupied by proteins (14,20). Because the hydrophobic interaction between phospholipids and lipophilic moieties of dextran matrix is the most important factor for a capture on the sensor chip, the stable immobilization of BBMVs might be limited within the specific condition, found in this study, primarily due to the lack of phospholipids in BBMVs.…”

Section: Formation Of the Reconstructed Bbmvsmentioning

confidence: 80%

“…Therefore, small intestinal brush border membrane vesicles (BBMVs), prepared from the small intestinal mucosa, are gaining recognition as an alternative tool to investigate the active transport because they contain phospholipids, hydrolytic enzymes, and carrier proteins that are responsible for many biologic binding events (11,12). In particular, BBMVs can be used to estimate the absorption of drugs and nutrients that may be orally administrated because of their structural and functional similarity to real intestinal membrane (13,14). Nevertheless, the extended use of BBMVs has often been hampered by the need of chromophoric or radiolabeled compounds.…”

Section: Introductionmentioning

confidence: 99%

Preparation and Characterization of Reconstructed Small Intestinal Brush Border Membranes for Surface Plasmon Resonance Analysis

Cho

Park

et al. 2004

Pharm Res

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Purpose. To prepare the surface generated by small intestinal brush border membrane vesicles (BBMVs) for the surface plasmon resonance (SPR) analysis, which allows the real-time measurement of binding events occurring on the intestinal membrane. Methods. BBMVs were isolated from Sprague-Dawley rats, suspended in HEPES-buffered saline, and flowed over the surface of a SPR sensor chip composed of dextran derivatives modified with lipophilic residues. The surface coverage was determined from binding of bovine serum albumin to BBMV-immobilized sensor chip. The performance of BBMVs immobilized was evaluated by their interaction with otilonium bromide and bile salts. Results. The stable BBMV surface was achieved when BBMV suspension was flowed over the sensor chip for 8 h at a rate of 2 l/min. The flow of otilonium bromide resulted in an increased SPR signal because of its binding to calcium channel, which is known to be distributed over the gastrointestinal tact. When bile salts were flowed over ileal and duodenal BBMV surfaces, respectively, a slightly higher SPR signal was observed in the ileal BBMV surface, indicating the specific interaction of bile salts with bile acid transporters. Conclusions. BBMV surfaces may be useful for the estimation of binding events on the intestinal membrane by SPR analysis, especially for the drugs that are orally administrated.KEY WORDS: surface plasmon resonance; intestinal brush border membrane; otilonium bromide; bile salts.

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A Novel Method of Preparation of Small Intestinal Brush Border Membrane Vesicles by Polyethylene Glycol Precipitation

Cited by 36 publications

References 29 publications

Intestinal mucosal alterations in rats with carbon tetrachloride‐induced cirrhosis

Intestinal mucosal alterations in rats with carbon tetrachloride‐induced cirrhosis

Oral glutamine attenuates indomethacin-induced small intestinal damage

Preparation and Characterization of Reconstructed Small Intestinal Brush Border Membranes for Surface Plasmon Resonance Analysis

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