The Caco-2 cell model established as a tool for in vitro investigations of intestinal drug transport processes has been widely used because of its growth characteristics, i.e., it forms polarized monolayers in cultures and differentiates into cells with high homology to human intestinal epithelial absorptive cells. Caco-2 cell cultures have provided a major conceptual advance in our understanding of intestinal drug absorption, biotransformation and bioavailability at the cellular level. Caco-2 cells have received considerable attention from the pharmaceutical industry because they have been widely accepted as a potent in vitro model membrane to screen for potential absorption problems in drug discovery programs. However, the Caco-2 monolayers model is still not perfect. The tightness of the monolayers resembles more colonic than small intestinal tissue, resulting in poor permeabilities for hydrophilic compounds traversing the epithelium via the aqueous paracellular pathway. Caco-2 cells have no mucus layer that is a potential barrier to drug absorption and display low expression of cytochrome P450 which are drug metabolizing enzymes. Further refinements of the Caco-2 cell culture model are needed to better predict human intestinal drug transport. To optimize Caco-2 model, the following technics have been used: modifying the condition of the cell culture, using molecular cloning strategies and inducing the expression of relevant enzymes. They are described in this review.
Biological Availability ; Biological Transport ; Caco-2 Cells ; cytology ; Colon ; physiology ; Epithelial Cells ; cytology ; Humans ; Intestinal Absorption ; Models, Biological

Colonic duplication is a rare congenital anomaly of the alimentary tract. In most cases, symptomatic duplications of the colon are recognized and treated by childhood. It is uncommon for these lesions to be detected in the adulthood since they present with vague symptoms if at all. We experienced a case of asymptomatic tubular duplication of the transverse colon in a 40-year-old female. Barium enema revealed a tubular duplication of the transverse colon. The duplicated segment arose from the mid ascending colon and incorporated just proximal to the splenic flexure, running parallel to the transverse colon and communicating with it at both ends. Colonoscopy demonstrated a normal colonic mucosa in the duplicated segment. The diameter of its lumen gradually narrowed proximally and the colonoscope could not be passed through the proximal opening of the segment. The patient did not need any treatment. Duplications of the alimentary tract can be found at any age. The possibility of congenital lesions in the adult population should not be overlooked.
Adult ; Colon, Transverse/*abnormalities/radiography ; Colonoscopy ; Female ; Humans ; Intestinal Mucosa/cytology

OBJECTIVE: To explore the molecular mechanisms of colonic epithelial aging related proteins and aged colonic epithelial susceptibility to tumor. METHODS: The proteins of normal human colonic epithelial tissue from young and old people were separated by 2-dimensional gel electrophoresis (2DGE), respectively. Then gels were stained by silver, scanned by imagescanner and analyzed with PDQuest software. The differentially expressed protein spots of colonic epithelium between the old and the young groups were identified by peptide mass fingerprint based on matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) and database searching. RESULTS: Well-resolved and reproducible 2DGE maps of normal human colonic epithelium from the young and the old were acquired. Nineteen more than 2 fold differentially expressed protein spots were identified representing 17 different proteins by MALDI-TOF-MS. The functions of these proteins involve in metabolism, energy generation, transportation, antioxidation, translation and protein folding. CONCLUSION Seventeen aging related proteins of human colonic epithelium identified indicate that injury of mitochondrial function and decline of antioxidant capability are important reasons for the aging of human colonic epithelium. These data provided useful clues for elucidating the mechanisms of colonic epithelial aging and aged colonic epithelial susceptibility to cancer.
Aging ; metabolism ; Cells, Cultured ; Cellular Senescence ; genetics ; Chloride Channels ; biosynthesis ; genetics ; Colon ; cytology ; Electron-Transferring Flavoproteins ; biosynthesis ; genetics ; Epithelial Cells ; cytology ; Humans ; Intestinal Mucosa ; cytology ; Proteins ; metabolism

The effect of acupuncture in the treatment of young pigs with induced enteropathogenic Escherichia coli diarrhea was histopathologically evaluated by routine hematoxylin and eosin stain. Thirty two pigs weighed 4-5kg and aged 21days old were used in this study. The animals with diarrhea were treated with traditional acupuncture, or enrofloxacin. In the group treated with traditional acupuncture, acupoint GV1 (Jiaochao) was used and in the group treated with antibiotics, enrofloxacin was injected intramuscularly. Ten pigs were inoculated with E. coli, but were not treated and served as nontreated control group. At postinoculation day 6, all pigs of the acupuncture and antibiotic treated groups recovered from diarrhea. In the ascending and descending colons of the nontreated control group, severe infiltration of inflammatory cells in the lamina propria was observed and in the fundic stomach, destruction of the fundic gland architecture and necrotic lesions were observed, however, in the same sites of the acupuncture and antibiotics treated groups, the mucosae of the colon and stomach were relatively similar to those of the normal group. These results indicate that acupuncture treatment is effective in controlling induced E. coli diarrhea in pigs at its early stage.
Acupuncture ; Animals ; Colon/cytology/microbiology/pathology ; Diarrhea/therapy/*veterinary ; Escherichia coli Infections/therapy/*veterinary ; Gastric Mucosa/cytology/microbiology/pathology ; Intestinal Mucosa/cytology/microbiology/pathology ; Male ; Stomach/cytology/microbiology/pathology ; Swine ; Swine Diseases/*microbiology/therapy

To study the acute and long-term effects of local gut inflammation on the sensitivity of the spinal sensory neurons, the expressions of vanilloid receptor 1 (VR1) and calcitonin gene-related peptide (CGRP) in the colon-innervated primary sensory neurons in dorsal root ganglia (DRG) were examined in rats with trinitrobenzenesulfonic acid (TNBS)-induced experimental colitis. The neurons projecting to the distal colon were identified by DiI(3) retrograde labelling. Macroscopic examination, mean damage score and myeloperoxidase (MPO) activity were determined to assess the inflammatory status of the colon tissue. The number of CGRP and VR1 immunoreactive neurons at different stages of inflammation (on days 7, 21 and 42 after TNBS treatment) were compared. On day 7 after TNBS treatment, macroscopic damage of the mucosa could be easily detected and the percentage of colon-innervated DRG neurons expressing CGRP and VR1 increased nearly two folds respectively [(95.38±9.45)% vs (42.86±.02)% for CGRP, (89.23±8.21)% vs (32.54±4.58)% for VR1]. When the colon inflammatory reaction was resolved on days 21 and 42 after TNBS treatment, the percentage of colon-innervated DRG neurons expressing CGRP and VR1 were still higher than that in the control group [(86.25±8.21)%, (68.28±7.12)% vs (42.86±5.02)% for CGRP; (67.22±6.52)%, (56.25±4.86)% vs (32.54±4.58)% for VR1]. These results suggest that the local gut inflammation increases the expressions of CGRP and VR1 in gut-innervated DRG sensory neurons. More importantly, this abnormal status persists even after the gut inflammatory reaction has been resolved for certain time.
Animals ; Calcitonin Gene-Related Peptide ; metabolism ; Colitis ; physiopathology ; Colon ; innervation ; Ganglia, Spinal ; cytology ; Inflammation ; physiopathology ; Neurons, Afferent ; cytology ; Rats ; Rats, Sprague-Dawley ; Sensory Receptor Cells ; cytology ; Spinal Cord ; cytology ; TRPV Cation Channels ; metabolism

OBJECTIVETo investigate the changes in the morphology and cytoskeleton (CSL) content of the CSL in the colonic smooth muscle cells (SMCs) of rats in early postscald stage, so as to elucidate the mechanism of dysfunction of gastrointestinal motility.

METHODSSeventy Wistar rats were randomly divided into normal control (n = 10, without scald) , and scald ( n = 60, with 10 cm x 7 cm wound inflicted on the back) groups. The colonic smooth muscle tissue of 10 normal rats and scalded rats were harvested at 1, 3, 6, 12, 24 and 48 postscald hours( PSH) and divided into two parts: one for histologic examination, and the other for the detection of CSL changes in the colonic smooth muscle tissue by flowcytometry method.

RESULTSThe electron microscope examination showed that the arrangement of cytoskeleton of SMC of the scalded rats during 1 to 3 PSH was disordered, and sparse, and the condensed area was uneven, with fragmentation. But the morphology and distribution of CSL gradually restored to normal state during 6 to 12 PSH, and it approached that of normal group at 48 PSH. The CSL content in the colonic smooth muscle tissue of scalded rats was obviously increased at 1 PSH (610+/-23) , decreased thereafter, evidently lower than that in control group at 3 PSH (92+/-17) , and then it started to increase at 12 PSH, exceeding the normal value at 24 PSH, and continued to rise until 48 PSH. There was significant difference in CSL content in the colonic smooth muscle tissue of the rats between scald and control group ( P < 0.05 or 0. 01).

CONCLUSIONChanges in the morphology and CSL content in the colonic smooth muscle tissue can be observed at early stage after a scald, which imply the kinetic balance between damage and repair in the body. In addition, changes in CSL content in the colonic smooth muscle tissue may be important factors in producing colonic dysfunction, damage of intestinal wall structure, and dynamic abnormalities of the colonic smooth muscle.

Actins ; metabolism ; Animals ; Burns ; metabolism ; pathology ; Colon ; cytology ; Cytoskeleton ; ultrastructure ; Disease Models, Animal ; Microtubules ; Myocytes, Smooth Muscle ; cytology ; metabolism ; Rats ; Rats, Wistar

OBJECTIVETo study the kinetogenic effects of serum containing Semen Arecae, Dandelion, Semen raphani and Atractylodes macrocephala on the colonic smooth muscle cells of rats.

METHODSSerum containing Chinese materia medicas was made according to standard methods. Smooth muscle cells were isolated from the muscle layers of Wistar rat's colon, referred to modified Bitar's method. The contractile response of colonic smooth muscle cells to serum containing Chinese materia medicas (10%, 50%, 100% concentration) and other medicines (blank and 1 x 10(-3) mol/L acetylcholine) were separately observed. The contractility was presented by the decrease of the cell length between the drug groups and the control.

RESULTSSerum containing each Chinese materia medica can make dose-dependent contraction at different concentrations (P < 0.05), but the strongest effect of each serum had no significant difference (P > 0.05).

CONCLUSIONSerum containing Semen Arecae, Dandelion, Semen raphani and Atractylodes macrocephala can make notable contraction on colonic smooth muscle cells in rats.

Animals ; Cells, Cultured ; Colon ; cytology ; Drugs, Chinese Herbal ; pharmacology ; Medicine, Chinese Traditional ; Muscle Contraction ; drug effects ; Muscle, Smooth ; cytology ; Myocytes, Smooth Muscle ; drug effects ; Rats

OBJECTIVETo induce DNA oxidative damage in colorectal crypt cells by hydrogen peroxide in vitro.

METHODSHydrogen peroxide was diluted into 100, 50, 10, 5 and 1 micromol/L with RPMI 1640. Colorectal crypt cells were treated with peroxide for 10 min, 30 min, 1 h, 1.5 h, 12 h and 24 h respectively. The survival of colorectal crypt cell was measured by MTT method, and the DNA oxidative damage special product, 8-OhdG was detected with immunohistochemical staining. Liner regression was used to measure the time trend of survival rate with SPSS 10.0 software.

RESULTSurvival rate of colorectal crypt cell was 60% and 80% after 10 min of hydrogen peroxide treatment. The longer treatment of hydrogen peroxide, the lower survival rate; the survival rate was reduced to 30% in 24 h. After 10 or 30 min treatment of 100 or 50 micromol/L hydrogen peroxide, the survival rates of colorectal crypt cells were reduced by 20% compared with those of 10, 5 and 1 micromol/L hydrogen peroxide. However, while cells were treated with different concentrations of hydrogen peroxide for 1.0 h or above, there were no differences in cell survival rates. The time trend test results demonstrated that the survival rates of colorectal crypt cells treated with 10, 5 and 1 micromol/L hydrogen peroxide were significantly decreased with the time length of treatment. Colorectal crypt cells treated with different concentrations of hydrogen peroxide for 15 minutes were positively stained brown in cytoplasm and nuclear by immunohistochemistry with 8-OhdG monoclonal antibody.

CONCLUSIONHydrogen peroxide could induce DNA oxidative damage in colorectal crypt cells. And treated with 1 - 10 micromol/L hydrogen peroxide for 10 - 30 min, DNA oxidative damage is apt to be induced in colorectal crypt cell.

Carbazoles ; analysis ; Cells, Cultured ; Colon ; cytology ; drug effects ; metabolism ; Humans ; Hydrogen Peroxide ; Models, Biological ; Oxidative Stress ; drug effects ; Propanolamines ; analysis ; Stem Cells ; cytology ; drug effects

AIMTo examine the ability of calcium ionophore (CI) to induce tryptase and histamine release from human mast cells and its mechanisms.

METHODSEnzymatically dispersed cells from human colons were challenged with CI, and the cell supernatants after challenge were collected. Tryptase release was determined with a sandwich ELISA procedure and histamine release was measured using a glass fibre-based fluorometric assay.

RESULTSCI was able to induce a concentration dependent release of histamine and tryptase from human colon mast cells following 15 min incubation. The maximum of induced histamine and tryptase release were approximately 5.3 and 2.8 fold more than the levels of spontaneous release, respectively. CI at the concentrations higher than 1.0 micromol/L was able to induce significantly more histamine than tryptase release from mast cells. The time course revealed that the action of CI on mast cells started from 10 s, peaked at 6 min and lasted at least 15 min following incubation. Pertussis toxin and metabolic inhibitors were able to inhibit mast cell response to CI.

CONCLUSIONHuman colon mast cells were able to release tryptase and histamine in response to CI. The process seemed to be associated with the activation of a G-protein coupled receptor on the membrane of mast cells and requires cell energy supply.

Calcium Ionophores ; pharmacology ; Cells, Cultured ; Colon ; cytology ; Histamine ; metabolism ; Humans ; Mast Cells ; drug effects ; metabolism ; secretion ; Tryptases ; metabolism

OBJECTIVETo study the influence of serum from scalded rats on the cytoskeleton of colonic smooth muscle cells (CSMC) of rats cultured in vitro, and to probe the possible mechanism of gastrointestinal motility disorder after burn.

METHODSCSMC isolated from healthy adult Wistar rat were cultured and divided into scald serum group (SS) and normal serum group (NS) according to the random number talbi. Two normal Wistar rats were used, one of which was inflicted with deep partial-thickness scald. Serum was obtained from blood collected from these two rats respectively and diluted to 20% in concentration. Serum from scald and normal rats were respectively added to the culture of CSMC in SS and NS groups. The expression of actin and the relative content of β-tubulin in CSMC was respectively determined with flow cytometry and Western blot at post treatment hour (PTH) 1, 3, 6, and 12 (with 10 samples in each group at each time point). Data were processed with t test.

RESULTSFluorescence intensity of actin in SS group at PTH 1, 3, 6, and 12 was respectively 59 ± 4, 26 ± 6, 39 ± 6, and 42 ± 6, all significantly lower than those in NS group (95 ± 10, 91 ± 10, 102 ± 9, and 97 ± 9, with t value respectively 10.528, 18.069, 18.748, 16.647, P < 0.05 or P < 0.01). In SS group, the fluorescence intensity decreased to the nadir at PTH 3, and then increased persistently at PTH 6 and 12. (2) Relative content of β-tubulin in SS group at PTH 1, 3, 6, and 12 was respectively 14.44 ± 0.26, 8.61 ± 0.19, 11.76 ± 0.31, and 12.13 ± 0.29, all significantly less than those in NS group (22.37 ± 1.15, 21.87 ± 1.79, 23.24 ± 1.55, and 21.99 ± 2.02, with t value respectively 21.176, 23.365, 23.000, 15.273, P values all below 0.01). In SS group, the relative content of β-tubulin decreased to the nadir at PTH 3 and increased slowly at PTH 6 and 12.

CONCLUSIONSThe reduction of CMSC content which has the tendency of increasing later, can be attributed to the influence of scald serum in initial stage. This may be related to the tolerance and adaptation to scald serum and self-repair of CMSC.

Animals ; Burns ; metabolism ; Cells, Cultured ; Colon ; cytology ; Cytoskeleton ; metabolism ; Male ; Microtubules ; metabolism ; Myocytes, Smooth Muscle ; metabolism ; Rats ; Rats, Wistar ; Serum