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.

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.

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.