TFF3 as a Diagnostic Biomarker in Kidney Transplant Patients.

Intestinal trefoil factor 3 (TFF3) is a protein secreted by many cell types, and its serum and urine levels vary in patients with kidney disease. Therefore, the present study aimed to determine the diagnostic value of TFF3 in allogeneic kidney transplant patients included in the one-year follow-up. To analyze the influence of the diagnostic method used, we studied the type of biological material and the time elapsed since renal transplantation on the parameter's value. The study also aimed to investigate the relationship between TFF3 levels and creatinine and estimated glomerular filtration rate (eGFR) values in the serum and urine of the patients studied. The study used blood and urine samples from adult patients (n = 19) 24-48 h, 6 months, and 12 months after kidney transplantation. We collected one-time blood and urine from healthy subjects (n = 5) without renal disease. We applied immunoenzymatic ELISA and xMap Luminex flow fluorimetry to determine TFF3 in serum and urine. There was a significant difference in TFF3 levels in the serum of patients collected on the first one or two days after kidney transplantation compared to the control group (determined by ELISA and Luminex) and six months and one year after kidney transplantation (ELISA). We observed a correlation between creatinine concentration and urinary TFF3 concentration (ELISA and Luminex) and a negative association between eGFR and urinary (ELISA) and serum (Luminex) TFF3 concentration in patients on the first and second days after kidney transplantation. We noted significant correlations between eGFR and TFF3 levels in the serum and urine of patients determined by the two methods six months and one year after transplantation. In women, we observed that urinary TFF3 concentration increased significantly with increasing creatinine and that with increasing eGFR, urinary TFF3 concentration determined by two methods decreased significantly. In the present study, the choice of diagnostic method for the determination of TFF3 in serum and urine significantly affected the concentration of this biomarker. The values of this parameter determined by ELISA were higher than those assessed using the Luminex assay. Based on the presented results, we can conclude that TFF3 has great potential to monitor renal transplant patients. Determination of this protein in parallel with creatinine and eGFR levels in serum and urine may provide helpful diagnostic information.

TFF3 mediates the NF-κB/COX2 pathway to regulate PMN-MDSCs activation and protect against necrotizing enterocolitis.

Intestinal trefoil factor 3 (TFF3) plays an important role in repairing the intestinal mucosa. However, the detailed mechanism regarding immune regulation by TFF3 is not well defined. Here, we reported that treatment of mouse BM cells and human peripheral blood mononuclear cells from healthy volunteers with TFF3 activated polymorphnuclear myeloid-derived suppressor cells (PMN-MDSCs) in vitro. We also found that prostaglandin E2 is a major TFF3-mediated MDSC target, and that NF-κB/COX2 signaling was involved in this process. Moreover, TFF3 treatment or transfer of TFF3-derived PMN-MDSCs (TFF3-MDSCs) to experimental necrotizing enterocolitis (NEC) mice caused PMN-MDSC accumulation in the lamina propria (LP), which was associated with decreased intestinal inflammation, permeability, bacterial loading, and prolonged survival. Interestingly, no NEC severity remission was observed in Rag1 KO mice that were given TFF3-MDSCs, but coinjection with CD4+ T cells significantly relieved NEC inflammation. Overall, TFF3 mediates the NF-κB/COX2 pathway to regulate PMN-MDSC activation and attenuates NEC in a T-cell-dependent manner, which suggests a novel mechanism in preventing NEC occurrence.

Pretreatment with intestinal trefoil factor alleviates stress-induced gastric mucosal damage via Akt signaling.

BACKGROUND: Stress-related gastric mucosal damage or ulcer remains an unsolved issue for critically ill patients. Stress ulcer prophylaxis has been part of routine intensive care, but uncertainty and controversy still exist. Co-secreted with mucins, intestinal trefoil factor (ITF) is reported to promote restitution and regeneration of intestinal mucosal epithelium, although the mechanism remains unknown. AIM: To elucidate the protective effects of ITF on gastric mucosa and explore the possible mechanisms. METHODS: We used a rat model of gastric mucosal damage induced by water immersion restraint stress and lipopolysaccharide-treated human gastric epithelial cell line to investigate the potential effects of ITF on damaged gastric mucosa both in vivo and in vitro. RESULTS: ITF promoted the proliferation and migration and inhibited necrosis of gastric mucosal epithelia in vitro. It also preserved the integrity of gastric mucosa by upregulating expressions of occludin and zonula occludens-1. In the rat model, pretreatment with ITF ameliorated the gastric mucosal epithelial damage and facilitated mucosal repair. The protective effects of ITF were confirmed to be exerted via activation of Akt signaling, and the specific inhibitor of Akt signaling LY249002 reversed the protective effects. CONCLUSION: ITF might be a promising candidate for prevention and treatment of stress-induced gastric mucosal damage, and further studies should be undertaken to verify its clinical feasibility.

Intestinal Trefoil Factor 3 Alleviates the Intestinal Barrier Function Through Reducing the Expression of TLR4 in Rats with Nonalcoholic Steatohepatitis.

BACKGROUND: Previous studies have reported that nonalcoholic steatohepatitis (NASH) is relevant to intestinal mucosal barrier dysfunction. AIM OF THE STUDY: To investigate the effects of intestinal trefoil factor 3 (TFF3) on intestinal barrier function and endotoxin/toll-like receptor 4(TLR4) expression in NASH rats. METHODS: Sixty NASH rats were divided into control, NASH and NASH-TFF3 treated group. Intestinal permeability, serum levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), endotoxin (ET), diamine oxidase (DAO) and liver index were examined. HE and PAS staining were performed to observe the histopathology of liver and terminal ileum. Expression of TFF3 and occludin were detected by immunohistochemical staining. mRNA and protein expression of TLR4, nuclear factor-κB (NF-κB), Mucin-2(Muc2) were detected by RT-qPCR and Western Blot. Interleukin (IL) -1ß and IL-10 levels in the ileum were measured by ELISA. RESULTS: In NASH group, levels of AST, ALT, ET, DAO, NAS, liver index and intestinal permeability were higher while occludin expressions were lower than control and NASH-TFF3 treated groups (p <0.05). Histopathology examination showed pathological damages of liver and ileum were alleviated in NASH-TFF3 treated group. NASH-TFF3 treated group had decreased expression levels of TLR4 and NF-κB and increased expression levels of Muc2 than NASH group. Besides, NASH group showed increased IL-1ß and IL-10 levels compared with control group. NASH-TFF3 treated group showed decreased IL-1ß level however increased IL-10 level compared with NASH group. CONCLUSION: Recombinant human TFF3 (rhTFF3) can reduce the expression of TLR4, reduce intestinal permeability, alleviate liver damage and thus may play a therapeutic role in the treatment of NASH rats.

miR￣7 regulates intestinal trefoil factor and affects intestinal epithelial cell proliferation and migration / 中国小儿急救医学

Objective To investigate the regulation of miR-7 on intestinal trefoil factor (TFF3) and its effect on the proliferation and migration of intestinal epithelial cells. Methods miR-7 mimics and miR-7 inhibitor were transfected into LS174T cells respectively. The experiment was divided into 5 groups,including blank control group,miR-7 mimic negative control group,miR-7 mimic group,miR-7 inhibitor negative con-trol group,miR-7 inhibitor group. MTT assay was used to detect cell proliferation. Cell scratch assay was used to detect the effect of miR-7 on cell migration. Western blot was used to detect the change of TFF3 protein in each group. Results Compared with the blank control group,the miR-7 mimic negative control group and the miR-7 inhibitor negative control group,the OD value of the miR-7 mimic group decreased significantly, the difference was statistically significant(P<0. 05); the cell scratch interval increased,the cell migration rate decreased,the difference was statistically significant( P<0. 05); and the TFF3 protein expression was accompanied(P<0. 05). The OD value of the miR-7 inhibitor group significantly increased,the difference was statistically significant(P<0. 05); the cell scratch gap decreased,the cell migration rate was enhanced, the difference was statistically significant( P <0. 05); and the expression of TFF3 protein increased ( P <0. 05). Conclusion miR-7 can regulate the expression of TFF3 and further inhibit the proliferation and migration of LS174T cells.

Intestinal trefoil factor increased the Bcl-2 level in a necrotizingenterocolitis neonate rat model.

BACKGROUND/AIM: The aim of this study was to investigate the therapeutic effect of intestinal trefoil factor (ITF) on necrotizing enterocolitis (NEC) by observing the pathological changes and detecting the protein level differences in Caspase-3, Bax, and Bcl-2 in an NEC neonate rat model. MATERIALS AND METHODS: A Wistar rat model of NEC was established and 30 one-day-old neonate Wistar rats were randomly divided into three groups including a normal control (group A), NEC rats treated with 0.2 ml physiological saline through intraperitoneal (i.p.) injection (group B), and NEC rats treated with 0.2 mg ITF by i.p injection (group C). RESULTS: Compared with group B, there were statistically significant differences in Caspase-3, Bax, and Bcl-2 levels in groups A and C(P < 0.05). Moreover, there was a significant difference in the Bcl-2 level between groups A and B (P < 0.05). CONCLUSION: ITF alleviated injury of the intestinal tract in neonate rats with NEC and this mechanism was possibly related to a reduction in the expression of Caspase-3 and Bax and the increase in Bcl-2 expression.

Value of citrulline, intestinal fatty acid binding protein, and intestinal trefoil factor in the diagnosis of acute gastroin-testinal injury in critically ill children / 临床儿科杂志

ObjectiveTo compare the values of serum citrulline, intestinal fatty acid binding protein (IFABP) and intes-tinal trefoil factor (ITF) in diagnosis of acute gastrointestinal injury in critically ill children.MethodsA total of 84 critically ill children were enrolled. The serum citrullin, IFABP, and ITF were measured by high performance liquid chromatography and enzyme-linked immunosorbent assay. The testing results and clinical data were analyzed.ResultsCompared with non gastroin-testinal injury group, the serum citrulline level was signiifcantly lower and IFABP and ITF levels were signiifcantly higher in gas-trointestinal injury group (allP<0.05). In critically ill children, the serum citrulline level was negatively correlated with C-reactive protein, procalcitonin and hospitalization time (r=-0.36 to -0.31,P<0.01).ConclusionsThe levels of citrulline, IFABP and ITF have diagnostic values for acute gastrointestinal injury in critically ill children. The level of citrulline may relfect the degree of acute gastrointestinal injury in critically ill children.

Association between trefoil factor 3 gene variants and idiopathic recurrent spontaneous abortion.

Trefoil factor 3 (TFF3) gene is an inflammatory mediator expressed in human endometrium during the window of implantation. The aim of this study was to evaluate the possible genetic association of TFF3 variants in recurrent spontaneous abortion. Women with a history of recurrent spontaneous abortion (n = 164) and healthy pregnant women (n = 143) were genotyped for five TFF3 polymorphisms (rs225439 G/A, rs533093 C/T, rs225361 A/G, rs11701143 T/C and rs77436142 G/C). In addition, haplotypes formed within the gene were analysed. Within the recurrent spontaneous abortion group, women who at some point had given birth and childless women had 4.19 ± 1.75 and 5.34 ± 3.42 consecutive spontaneous abortions, respectively. Women who had experience recurrent spontaneous abortions had a lower allele frequency of the rs11701143 promoter region minor C allele compared with fertile women (0.02 versus 0.05, P = 0.015). Patients with rs225361 AG genotype had significantly more successful pregnancies before spontaneous abortion than those with homozygous AA and GG genotypes (P = 0.014). No significant differences in haplotype frequencies between patients and controls were detected. Possible genetic risk factors identified that might contribute to the pathogenesis of idiopathic recurrent spontaneous abortion were TFF3 gene variants.

ITF increases the transcriptional activity of ITF promoter via the JAK-STAT3 signal transduction pathway / 中南大学学报(医学版)

Objective: To investigate the effect ofintestinal trefoil factor (ITF) on the transcriptional activity of ITF promoter and to explore the regulatory mechanismofJanus kinase/signal transducersand activators of transcription(JAK/STAT) on ITF promoter. Methods: The 5' flanking sequence of the ITF gene was cloned from human whole blood genomic DNA by PCR. ITF promoter fragment was cloned and inserted into the pGL3-Basic vector to construct recombinant vector. ITF promoter vector was stimulated with ITF at various concentrations and the luciferase activity was measured. The JAK-STAT3 signal transductionpathway was then blocked by a speciifc inhibitor AG490 to determine the signal pathway involved in ITF promoter activity. Results: Restriction endonuclease analysis and DNA sequencing confirmed that the recombinant plasmid, containing ITF promoter, was constructed successfully. After transient transfection, the activity of ITF promoter was increased signiifcantly in the presence of ITF (P<0.05). Blockage of the JAK-STAT3 signal transduction pathway with AG490 signiifcantly reduced the ITF promoter activity (P<0.05). Conclusion: ITF increases the transcriptional activity of ITF promoter via the JAK-STAT3 signal transduction pathway.

Research Progress of protective effect of intestinal trefoil factor on the gastrointestinal tract / 医学研究生学报

Intestinal trefoil factor (ITF or TFF3) is a member of trefoil factor family which is most studied , mainly in the in-testinal tract .TFF3 plays an important role in gastrointestinal mucosal protection and epithelial repair .Action mechanism includes in-teraction with mucin, migration, anti-apoptosis, angiogenesis, immune regulation, interaction with its receptor, etc.Recombinant TFF3 has been approved as a national drug and its therapeutic indications for repair mucosa , prevention and treatment of gastrointesti-nal ulcers and inflammation .Furthermore, the classic TFF3 receptor and signaling pathway is highly valuable to make TFF 3 become an effective treatment for gastrointestinal injury disease .

Experimental study of chitosan as carrier of intestinal trefoil factor gene / 中国药学杂志

OBJECTIVE: To construct an optimal human intestinal trefoil factor (hITF) gene delivery system.

Study on the effects of intestinal trefoil factor on gastric mucosal epithelial cell proliferation and its signal transduction mechanism / 医学研究生学报

Objective To explore the effects of intestinal trefoil factor ( ITF) on gastric mucosal epithelial cell proliferation and its possible molecular mechanism . Methods The cultured GES-1 cells were treated with ITF in the concentration of 100 ng/mL and 500 ng/mL in vitro, and then were observed using microscope for the morphological analysis .The Cell Counting Kit-8 ( CCK-8) was used to detect the proliferation activity of GES-1.The cultured GES-1 cells were treated with 100 ng/mL ITF and the specific inhibitor of PI3K/Akt signaling pathway LY294002 (15μmol/L) in vitro, and then were observed using microscope for the morphological analysis . The proliferation activity of treated GES-1cells was detected using CCK-8 and the expressions of p-Akt and Akt of PI3K/Akt signaling pathway were determined by Western blot . Results Compared with the control group , the proliferation activity of GES-1 cells in-creased after being treated with ITF and the higher concentration of ITF induced the higher proliferation activity .LY294002 inhibited the increased proliferation activity of GES-1induced by ITF.The data of Western blot indicated that ITF induced the expression of p -Akt and activated the P3IK/Akt signaling pathway to modulate the proliferation activity of GES -1 cells.However, LY294002 inhibited the PI3K/Akt signaling pathway and then decreased the proliferation activity of GES -1 cells. Conclusion ITF could promote the proliferation ac-tivity of GES-1 cells by activating PI3K/Akt signaling pathway .

Trefoil factor 3 peptide regulates migration via a Twist-dependent pathway in gastric cell.

Trefoil factor 3 (TFF3) is a member of the TFF-domain peptide family and essential in regulating cell migration and maintaining mucosal integrity in gastrointestinal tract. However, the role of TFF3 and its downstream regulating mechanisms in cancer cell migration remain unclear. We previously reported that TFF3 prolonged the up-regulation of Twist protein to modulate IL-8 secretion in intestinal epithelial cells. In this study, we investigated the role of Twist protein in TFF3-induced migration of SGC7901 cells. While Twist was activated by TFF3, siRNA-mediated knockdown of Twist abolished TFF3-induced cell migration. Furthermore, the migration related marker CK-8 as well as ZO-1 and MMP-9 was also regulated by TFF3 via a Twist-dependent mechanism. Our study suggests that Twist, as an important potential downstream effector, plays a key role in TFF3-modulated metastasis in gastric cancer and can be a promising therapeutic target against intestinal-type gastric cancer.

Repair of damaged intestinal mucosa in a mouse model of sepsis.

BACKGROUND: The intestine is not only the main target attacked by sepsis but also the vital organ which mediated sepsis. The recovery of the damaged intestinal barrier structure and function is related to the occurrence and outcome of multiple organ dysfunction syndrome (MODS). How to protect and reduce the damage of the intestinal mucosa and how to promote the reconstruction of the intestinal mucosa have been the important topics in sepsis for many years. This study aimed to investigate the influential factors of intestinal mucosal reconstruction after intestinal epithelial injury in vivo in a mouse model of sepsis. METHODS: Mice were subjected to cecal ligation and puncture (CLP) for induction of sepsis to assess intestinal mucosal damage, epithelial cell apoptosis, and transformed number of goblet cells, and to detect the concentration of TNF-α, IL-1 and TGF-ß1 and TFF3 (trefoil factor 3) expression in the small intestinal mucosa. All above were performed by HE staining, western blot, ELISA and immunohistochemistry respectively. The experimental animals were divided into a sepsis group and a sham-operation group. The animals with sepsis were separately killed at 6 (7 animals), 24 (7 animals) and 48 hours (7 animals) after CLP. RESULTS: Injured intestinal mucosa was observed in the 3 groups under a light microscope, in which damage scores in the 24-hour and 48-hour groups were higher than in the 6-hour group and no difference was found between the two groups. Moreover, less of goblet cells or other epithelial cells adjacent to the injured surface migrated into the wound to cover the denuded area. The number of goblet cells was substantially decreased in the three CLP groups compared with the sham-operation group. Protein levels of IL-1 and TNF-α were significantly increased by 3-4 fold at all time points when compared with the sham-operation group, and cleaved caspase-3 by 4 fold. Although TFF3 expression was modestly increased for 6 hours after the onset of CLP, it appeared to decline at 24 hours and 48 hours as shown by Western blot. A similar tendency was observed upon TGF-ß1, i.e. the protein level was not elevated at 24 hours and 48 hours, but increased modestly at 6 hours. CONCLUSIONS: Sepsis from CLP shows less restitution on the surface of injured intestinal mucosa. There is evidence that both constant inflammatory reaction and epithelial cell apoptosis may affect mucosal reestablishment of the intestine at the onset of sepsis. Mucosa after severe sepsis showed the state of high inflammation, and declined goblet cell function and mucosal reconstruction, which affected the repair of damaged intestinal barrier. Constant inflammatory reaction, and declined goblet cell function and mucosal reconstruction ability may affect the reestablishment of intestinal mucosa at the onset of sepsis.

The expression of intestinal trefoil factor (ITF) in colon of cytomegalovirus infected mouse and the involvement of ganciclovir / 重庆医学

Objective To observe the expression of ITF in colon of mice infected by mouse cytomegalovirus (MCMV) and the in-volvement of ganciclovir(GCV) .Methods Forty-eight BALB/c young mice were randomly divided into blank group ,virus group and GCV group ,each with 8 mice .Mice in virus group and GCV group received injection of 100 μL MCMV virus suspension (TCID50105 .31 /mL) ,and GCV group was given intraperitoneal injection of GCV once a day at the dose of 50 mg/kg from day 0 (24 hours after vaccination of virus ) ,for 14 days .At the same time the virus group and blank group were given the same dose of normal saline as controls .Murine cytomegalovirus loads in livers and colons were measured by PCR .The expression levels of ITF in mRNA in colon were detected by RT-PCR .Results After MCMV injection ,mice in virus group manifested aggressively apparent poor appetite ,less activity ,fur laxly ,unresponsiveness to stimuli ,growth retardation ,body weight not increased .All liver and colon tissue MCMV-DNA PCR electrophoresis of virus group had positive strip ,while the blank group did not .GCV group also showed less bright positive strip when compared with the virus group .Expression level of ITF mRNA was significant higher in GCV group than virus group ,there was statistically significant difference(P<0 .05) .Expression of ITF mRNA in virus group were higher than that in blank group ,there was statistical difference(P<0 .05) .Conclusion ITF is regarded as a fast reaction peptide in the course of mucosa impairments ,so ITF plays a protective role in delayed healing process after acute MCMV infection .

Application of molecular biomarkers in differential diagnosis of benign and malignant follicular thyroid tumors / 中华内分泌代谢杂志

Twenty-nine thyroid tissue samples were collected from patients with thyroid nodules.The total RNA were extracted,the gene expressions of TFF3,HMGA2,C1orf24,and DDIT3 were detected by RT-PCR.In comparison with normal thyroid tissues,the expression of C1orf24 mRNA was decreased in the follicular thyroid adenoma (FTA) group,but increased in the follicular thyroid carcinomas (FTC) group.The expressions of DDIT3 and HMGA2 mRNA were increased in both FTA and FTC groups,and were even higher in the latter.The expressions of TFF3 mRNA level were decreased in FTA and FTC.The data suggested that molecular markers C1ort24,DDIT3,HMGA2,and TFF3 in thyroid tissue seem to be helpful in the differential diagnosis between follicular adenomas and carcinomas.

Repair of damaged intestinal mucosa in a mouse model of sepsis / 世界急诊医学杂志（英文）

BACKGROUND:The intestine is not only the main target attacked by sepsis but also the vital organ which mediated sepsis. The recovery of the damaged intestinal barrier structure and function is related to the occurrence and outcome of multiple organ dysfunction syndrome (MODS). How to protect and reduce the damage of the intestinal mucosa and how to promote the reconstruction of the intestinal mucosa have been the important topics in sepsis for many years. This study aimed to investigate the influential factors of intestinal mucosal reconstruction after intestinal epithelial injuryin vivo in a mouse model of sepsis.METHODS:Mice were subjected to cecal ligation and puncture (CLP) for induction of sepsis to assess intestinal mucosal damage, epithelial cell apoptosis, and transformed number of goblet cells, and to detect the concentration of TNF-α, IL-1 and TGF-β1 and TFF3 (trefoil factor 3) expression in the small intestinal mucosa. All above were performed by HE staining, western blot, ELISA and immunohistochemistry respectively. The experimental animals were divided into a sepsis group and a sham-operation group. The animals with sepsis were separately killed at 6 (7 animals), 24 (7 animals) and 48 hours (7 animals) after CLP.RESULTS:Injured intestinal mucosa was observed in the 3 groups under a light microscope, in which damage scores in the 24-hour and 48-hour groups were higher than in the 6-hour group and no difference was found between the two groups. Moreover, less of goblet cells or other epithelial cells adjacent to the injured surface migrated into the wound to cover the denuded area. The number of goblet cells was substantially decreased in the three CLP groups compared with the sham-operation group. Protein levels of IL-1 and TNF-α were significantly increased by 3-4 fold at all time points when compared with the sham-operation group, and cleaved caspase-3 by 4 fold. Although TFF3 expression was modestly increased for 6 hours after the onset of CLP, it appeared to decline at 24 hours and 48 hours as shown by Western blot. A similar tendency was observed upon TGF-β1, i.e. the protein level was not elevated at 24 hours and 48 hours, but increased modestly at 6 hours.CONCLUSIONS:Sepsis from CLP shows less restitution on the surface of injured intestinal mucosa. There is evidence that both constant inflammatory reaction and epithelial cell apoptosis may affect mucosal reestablishment of the intestine at the onset of sepsis. Mucosa after severe sepsis showed the state of high inflammation, and declined goblet cell function and mucosal reconstruction, which affected the repair of damaged intestinal barrier. Constant inflammatory reaction, and declined goblet cell function and mucosal reconstruction ability may affect the reestablishment of intestinal mucosa at the onset of sepsis.

Relationship of intestinal trefoil factor for regulating of nuclear factor-κB and tumor necrosis factor-α and the protective effect of intestinal injury / 中华实用儿科临床杂志

Objective To investigate the relationship of intestinal trefoil factor(HF) for regulating of nuclear factor(NF)-κB and tumor necrosis factor(TNF)-α and the protective effect of intestinal injury.Methods Twenty-four 10-day Wistar rats were divided into 3 groups:the control group(NS group,n =8),given 9 g/L sodium injection intraperitoneally,1 mL/kg; the lipopolysaccharide (LPS) group (n =8),given LPS (5 g/L),5 mg/kg intraperitoreally; ITF group(n=8),given 1TF(5 g/L) 0.1 mL/each plus LPS 5 mg/kg intraperitoreally.Rats were sacrificed 3 hours after injection.A segment of distal ileum was dissected.The pathologic changes of small intestine were observed under the optical microscope(hematoxylin-eosin staining).The expressions of NF-κB mRNA and protein were detected by reverse transcription polymerase chain reaction(RT-PCR),immunohistochemistry,respectively.TNF-α level in intestinal tissues was measured by enzyme linked immunosorbent assay.Results The structure of small intestine in the control group remained normal.The inflammatory cells infiltration and the edema of interstitial substance and epithelium were observed in LPS group and ITF group,while the change in the ITF group was significantly lower than that in LPS group.The expressions of NF-κB mRNA and protein in ITF group were significantly lower than those in LPS group(all P ＜ 0.01).The secretion of TNF-α in the rTF group was significantly lower than that in LPS group(P ＜ 0.01).Conclusion Protective effects of ITF on intestinal injury are related to the down regulation of NF-κB mRNA and protein expressions and the reduction of the secrete of mediators of inflammation TNF-α.

Relationship between intestinal permeability and expression of intestinal trefoil factor mRNA in mice after total body irradiation / 中华放射医学与防护杂志

Objective To investigate the change of the intestinal permeability,the expression level of intestinal trefoil factor (ITF) mRNA and the relationship between them after total body irradiation (TBI),and explore the effect of TBI on the development of intestinal permeability and the expression level of ITF mRNA.Methods Twenty two BALB/c mice were randomly divided into 4 equal groups: 3 groups at 4,8 and 12 d after TBI with the total dose of 8.0 Gy and the dose rate of 1.0 Gy/min respectively,and a control group.Lactulose (L) and mannitol (M) were perfused into the esophagus before the experiment and urine samples were collected.Liquid chromatography was used to measure the L/M excretion ratio in the urine samples collected 4,8,and 12 days after the TBI.And then the mice were killed with their intestine were taken out.The expression of ITF mRNA in the jejunum tissue was detected by real-time fluorescence quantitative PCR.Results The urine L/M ratio levels of the groups 4,8 and 12 days after TBI were (0.5092 ± 0.0352),(0.7174 ± 0.0116),and (0.7295 ± 0.0533) respectively,all significantly higher than that of the control group [(0.2908 ± 0.0533),F = 321.47,P ＜ 0.05].The ITF mRNA expression levels of groups 4,8 and 12 days after TBI were (0.78612 ±0.1428),(0.2521 ±0.1223),and (0.2306 + 0.0221 ) respectively,all significantly lower than that of the control group [( 1.3498 + 0.0476),F = 235.71 ,P ＜ 0.05].The urine L/M ratio was significantly negatively correlated with the expression of ITF mRNA in all TBI groups (r = - 0.985,P ＜ 0.01 ).Conclusions The intestinal permeability increases and the expression level of ITF mRNA decreases after TBI.The urine L/M ratio is negatively correlated with the expression level of ITF mRNA after TBI.ITF is involved in protection against intestinal permeability induced by TBI.

Protective effect of intestinal trefoil factor on methotrexate-induced injury in intestinal mucosa / 中华消化杂志

Objective To investigate the potential mechanism of intestinal trefoil factor(ITF)against methotrexate (MTX)- induced injury in intestinal mucosa. Methods Cultured IEC-6 cells were divided into groups as follows: blank group, MTX treated group, ITF treated group and experimental group treated with gradient concentrations of ITF plus MTX. Expression of E-cadherin mRNA was determined by Real-Time polymerase chain reaciton (RT- PCR). The activity of matrix metalloproteinase(MMP)-2 and MMP-9 was measured by gelatin zymogramphy. Caspases-3 activity was measured by colorimetric assay. Cell proliferation was assessed by cell counting kit-8 (CCK-8)assay. Migration of IEC-6 in vitro was observed using modified Boyden chamber assay. Results The expression of E-cadherin mRNA in experimental group (treated with 0.1 mg/ml or 1 mg/ml of ITF) was significantly down-regulated (0. 538±0. 109 or 0. 528±0. 132, respectively) in comparison with MTX treated group (0. 763±0. 139) with significant difference (P=0. 021 or P=0. 025, respectively). There was no significant difference in activity of MMP-2 and MMP-9 among groups (P＞0. 05). When compared with MTX treated group (0. 090 ±0. 011 ), the activity of Caspase3 in experimental group (treated with 0. 1 mg/ml or 1 mg/ml of ITF) was significantly decreased (0. 077±0. 009, P=0. 032 or 0. 044±0. 009,P=0. 005, respectively). There was no statistical difference in cell proliferation between experimental group (treated with 1 μg/ml, 0.01 mg/ml, 0. 1 mg/ml or 1.0 mg/ml of ITF) and MTX treated group (P=0. 132,0. 150,0. 114 or 0. 367, respectivley). More migratory cells attached to the bottom surface of the membrane in experiment group (treated with 0. 1 mg/ml or 1 mg/ml of ITF) in comparison with MTX treated group (P ＜0. 001 ). Moreover, more migratory cells were found in experimental group treated with 1.0 mg/ml of ITF than those in group treated with 0. 1 mg/ml of ITF (P＜0. 001). Conclusions Without cell proliferation, the protective effect of ITF is related to its functions of promoting cell migration and inhibiting cell apoptosis, which may down-regulate expression of E-cadherin mRNA.