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Objective@#To investigate the effects of sodium butyrate on intestinal barrier of the severe scald mice and the related mechanism.@*Methods@#Eighteen C57BL/6 female mice, aged eight to twelve weeks, were divided into sham scald group, pure scald group, and scald+ sodium butyrate group according to random number table, with 6 mice in each group. Back of each mouse in pure scald group and scald+ sodium butyrate group were immersed into 90 ℃ water for 9 s, causing full-thickness scald of 30% total body surface area, while back of each mouse in sham scald group were immersed into 37 ℃ water for 9 s, causing sham injury. All of the mice in 3 groups were intraperitoneally injected with 1 mL sterile lactated Ringer′s solution immediately after injury. Besides, mice in scald+ sodium butyrate group were intraperitoneally injected with 300 mg/kg sodium butyrate at 30 min before injury and immediately after injury, while mice in sham scald group and pure scald group were intraperitoneally injected with the same volume of sterile phosphate buffer solution. At post injury hour (PIH) 24, portal vein of mice in 3 groups was harvested, intestinal permeability was measured by fluorescin isothiocyanate-dextran fluorescence probe tracing method, then lileal tissue of mice in 3 groups was harvested, protein expressions of zonula occludens l (ZO-1), occludin, claudin-1, claudin-2, nucleotide-binding oligomerization domain-containing protein-like receptor family pyrin domain containing 3 (NLRP3), interleukin-1β (IL-1β), and IL-18 were detected by Western blotting, and distribution of ZO-1 in intestinal mucosa was observed by indirect immunofluorescence. Data were processed with one-way analysis of variance, least-significant difference test, and Bonferroni correction.@*Results@#(1) At PIH 24, the intestinal permeability of mice in sham scald group, pure scald group, and scald+ sodium butyrate group was 0.88±0.19, 2.62±0.48, 1.23±0.16, respectively. Compared with that in sham scald group, the intestinal permeability of mice in pure scald group was significantly elevated (P<0.01), while the intestinal permeability of mice in scald+ sodium butyrate group showed no obvious change (P>0.05). Compared with that in pure scald group, the intestinal permeability of mice in scald+ sodium butyrate group was significantly decreased (P<0.01). (2) At PIH 24, compared with those in sham scald group, the protein expressions of ZO-1, occludin, and claudin-1 of mice in pure scald group and scald+ sodium butyrate group were significantly decreased (P<0.05), while the protein expression of claudin-2 was significantly increased (P<0.05). At PIH 24, compared with those of pure scald group, the protein expressions of ZO-1 and occludin of mice in scald+ sodium butyrate group were significantly elevated (P<0.05), while the protein expression of claudin-2 was significantly decreased (P<0.05), the protein expression of claudin-1 showed no significant difference (P>0.05). (3) At PIH 24, compared with those in sham scald group, the protein expressions of NLRP3, IL-1β, and IL-18 of mice in pure scald group and scald+ sodium butyrate group were significantly increased (P<0.05). Compared with those of pure scald group, the protein expressions of NLRP3, IL-1β, and IL-18 of mice in scald+ sodium butyrate group were significantly decreased (P<0.05). (4) At PIH 24, ZO-1 in intestinal mucosa of mice in sham scald group was distributed smoothly, continuously and homogeneously along the membrane. ZO-1 in intestinal mucosa of mice in pure scald group was distributed unsmoothly with breaks. The distribution of ZO-1 in intestinal mucosa of mice in scald+ sodium butyrate group was ameliorated compared with that in pure scald group.@*Conclusions@#Sodium butyrate can inhibit the activation of NLRP3 inflammasome and decrease the production of IL-1β and IL-18 in intestinal mucosa of severe scald mice, which protects the intestinal barrier function by alleviating the alteration of tight junction protein expression and localization.
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Objective@#To investigate the significance of intestinal fatty acid binding protein (IFABP) in the evaluation of intestinal barrier dysfunction of mice at the early stage of severe burn injury.@*Methods@#Thirty-six 8-week-old C57BL/6 male mice were collected and divided into normal control group (n=6) and scald group (n=30) according to random number table. Back of each mouse in scald group was placed into hot water of 90 ℃ for 10 s, causing full-thickness scald (hereinafter refer to as burn) of 30% total body surface area, while mice in normal control group were not inflicted with burns. Six mice in normal control group were taken, and 6 mice in scald group at 1, 2, 6, 12, and 24 h post injury were taken respectively. The portal vein blood of each mouse was extracted and the plasma was separated to measure intestinal permeability with fluorescin isothiocyanate-dextran fluorescence probe tracing method and plasma IFABP content by enzyme-linked immunosorbent assay. The distal ileum tissue of mice in normal control group and scald group at each time point post injury was collected to observe the morphology of the intestinal mucosa tissue by hematoxylin-eosin staining. Data were processed with one-way analysis of variance and Student-Newman-Keuls test, and pearson correlation test was used to analyze the correlation between intestinal permeability and plasma IFABP content of burned mice.@*Results@#(1) At 1, 2, 6, 12, and 24 h post injury, the intestinal permeability of mice in scald group was 2.7±0.8, 5.4±2.5, 7.3±4.2, 12.4±6.1, 1.4±0.7, respectively, obviously higher than 1.0±0.4 of normal control group (P<0.05 or P<0.01). The intestinal permeability of mice in scald group showed an increasing trend post injury, reaching the peak at 12 h post injury, and rapidly falling back at 24 h post injury. (2) At 1, 2, 6, 12, and 24 h post injury, the plasma IFABP content of mice in scald group was (64±11), (59±12), (76±18), (111±22), and (66±10) ng/mL, obviously higher than (35±8) ng/mL in normal control group (P<0.05 or P<0.01). The plasma IFABP content of mice in scald group showed an increasing trend post injury, reaching the peak at 12 h post injury, and rapidly decreasing at 24 h post injury. (3) Uniform thickness of mucosa, intact epithelia, regularly arranged villi, and no inflammatory cell infiltration were observed in ileum of mice in normal control group. In ileum of mice in scald group, shortened villi of mucosa with different degrees, edema of lamina propria, and infiltration of neutrophils were observed at 1 and 2 h post injury; obviously damaged and partially exfoliated ileal mucosa, disorderly arranged and broken villi, degenerated and necrotic epithelial cells, dilated central lacteal, and infiltration of lymphocytes and neutrophils were observed at 6 and 12 h post injury; the damage of ileal mucosa was alleviated, and basically intact epithelia, dilated central lacteal, and infiltration of inflammatory cells were observed at 24 h post injury. (4) There was a significantly positive correlation between the intestinal permeability and the plasma IFABP content of burned mice (r=0.841, P<0.05).@*Conclusions@#The plasma IFABP can be used as a good biological indicator for the evaluation of intestinal barrier dysfunction of mice at the early stage of severe burn injury.
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Objective@#To investigate the effects of short chain fatty acid (SCFA) on barrier disruption of human intestinal epithelial cell induced by endotoxin/lipopolysaccharide (LPS) and the related mechanism.@*Methods@#The human intestinal epithelial cell line Caco-2 was used to reproduce monolayer-cells. Cells were divided into control group, LPS group, and SCFA+ LPS group according to the random number table. Cells in control group were only routinely cultured with DMEM medium. Cells in LPS group were cultured with DMEM medium and LPS with final mass concentration of 10 μg/mL. Cells in SCFA+ LPS group were cultured with DMEM medium, LPS and SCFA (consisting of 0.5 mmol/L acetate, 0.01 mmol/L propionate, and 0.01 mmol/L butyrate) with final mass concentration of 10 μg/mL. At post culture hour (PCH) 0, 1, 2, 6, 12, and 24, transepithelial electrical resistance (TER) of cells was determined with an ohmmeter, with sample number of 72. Another portion of cells were divided and treated as above, and then Western blotting was employed to detect the protein expressions of zonula occludens 1 (ZO-1), occludin, and claudin-1 at PCH 24, with sample number of 6. Another portion of cells were divided and treated as above and then immunofluorescence was used to observe cellular morphology and distribution of ZO-1. Data were processed with analysis of variance of factorial design, one-way analysis of variance, least-significant difference test, and Bonferroni correction.@*Results@#(1) Compared with that in control group, TER of cells in LPS group was significantly reduced from PCH 1 to 24 (P<0.01), while TER of cells in SCFA+ LPS group showed no obvious change (P>0.05). TER of cells in SCFA+ LPS group was significantly higher than that in LPS group from PCH 1 to 24 (P<0.01). (2) Compared with the protein expressions of ZO-1, occludin, and claudin-1 of cells in control group (1.25±0.10, 1.17±0.04, and 1.24±0.20), those of cells in LPS group (0.74±0.23, 0.76±0.11, and 0.77±0.11) at PCH 24 were significantly decreased (P<0.05), while those of cells in SCFA+ LPS group (1.23±0.46, 1.05±0.09, and 1.01±0.13) showed no significant differences (P>0.05). Protein expressions of occludin and claudin-1 of cells in SCFA+ LPS group were significantly higher than those in LPS group at PCH 24 (P<0.05). Protein expression of ZO-1 of cells in SCFA+ LPS group was higher than that in LPS group at PCH 24 with no significant difference (P>0.05). (3) At PCH 24, cells in control group were compact in arrangement with pebble-like appearance, and ZO-1 was distributed smoothly and continuously along the cell membrane. In LPS group, cells were sparse in arrangement with change in appearance, and ZO-1 was distributed uncontinuously along the cell membrane with curls and breaks. In SCFA+ LPS group, the appearance of cells and distribution of ZO-1 were remarkably ameliorated compared with those in LPS group.@*Conclusions@#SCFA can alleviate the barrier disruption of human intestinal epithelial cell induced by LPS through interdicting the abnormal distribution of ZO-1 and decrease of TER and tight junction proteins′ expressions.