An Active Fraction of Trillium tschonoskii Promotes the Regeneration of Intestinal Epithelial Cells After Irradiation

Despite significant scientific advances toward the development of safe and effective radiation countermeasures, no drug has been approved for use in the clinic for prevention or treatment of radiation-induced acute gastrointestinal syndrome (AGS). Thus, there is an urgent need to develop potential drugs to accelerate the repair of injured intestinal tissue. In this study, we investigated that whether some fractions of Traditional Chinese Medicine (TCM) have the ability to regulate intestinal crypt cell proliferation and promotes crypt regeneration after radiation. By screening the different supplements from a TCM library, we found that an active fraction of the rhizomes of Trillium tschonoskii Maxim (TT), TT-2, strongly increased the colony-forming ability of irradiated rat intestinal epithelial cell line 6 (IEC-6) cells. TT-2 significantly promoted the proliferation and inhibited the apoptosis of irradiated IEC-6 cells. Furthermore, in a small intestinal organoid radiation model, TT-2 promoted irradiated intestinal organoid growth and increased Lgr5+ intestinal stem cell (ICS) numbers. More importantly, the oral administration of TT-2 remarkably enhanced intestinal crypt cell proliferation and promoted the repair of the intestinal epithelium of mice after abdominal irradiation (ABI). Mechanistically, TT-2 remarkably activated the expression of ICS-associated and proliferation-promoting genes and inhibited apoptosis-related gene expression. Our data indicate that active fraction of TT can be developed into a potential oral drug for improving the regeneration and repair of intestinal epithelia that have intestinal radiation damage.

Activation of G protein coupled estrogen receptor prevents chemotherapy-induced intestinal mucositis by inhibiting the DNA damage in crypt cell in an extracellular signal-regulated kinase 1- and 2- dependent manner

Chemotherapy-induced intestinal mucositis (CIM) is a common adverse reaction to antineoplastic treatment with few appropriate, specific interventions. We aimed to identify the role of the G protein coupled estrogen receptor (GPER) in CIM and its mechanism. Adult male C57BL/6 mice were intraperitoneally injected with 5-fluorouracil to establish the CIM model. The selective GPER agonist G-1 significantly inhibited weight loss and histological damage in CIM mice and restored mucosal barrier dysfunction, including improving the expression of ZO-1, increasing the number of goblet cells, and decreasing mucosal permeability. Moreover, G-1 treatment did not alter the antitumor effect of 5-fluorouracil. In the CIM model, G-1 therapy reduced the expression of proapoptotic protein and cyclin D1 and cyclin B1, reversed the changes in the number of TUNEL+ cells, Ki67+ and bromodeoxyuridine+ cells in crypts. The selective GPER antagonist G15 eliminated all of the above effects caused by G-1 on CIM, and application of G15 alone increased the severity of CIM. GPER was predominantly expressed in ileal crypts, and G-1 inhibited the DNA damage induced by 5-fluorouracil in vivo and vitro, as confirmed by the decrease in the number of γH2AX+ cells in the crypts and the comet assay results. Referring to the data from GEO dataset we verified GPER activation restored ERK1/2 activity in CIM and 5-fluorouracil-treated IEC-6 cells. Once the effects of G-1 on ERK1/2 activity were abolished with the ERK1/2 inhibitor PD0325901, the effects of G-1 on DNA damage both in vivo and in vitro were eliminated. Correspondingly, all of the manifestations of G-1 protection against CIM were inhibited by PD0325901, such as body weight and histological changes, the mucosal barrier, the apoptosis and proliferation of crypt cells. In conclusion, GPER activation prevents CIM by inhibiting crypt cell DNA damage in an ERK1/2-dependent manner, suggesting GPER might be a target preventing CIM.

PSXVI-7 Supplemental effects of Lactobacillus extract postbiotics in prevention of post-weaning diarrhea caused by enterotoxigenic F18+ Escherichia coli in nursery pigs

This study determined the supplemental effects of Lactobacillus extract (LBE) postbiotics on intestinal health and prevention of postweaning diarrhea caused by F18+ Escherichia coli (ETEC) in nursery pigs. Sixty-four weaned pigs (6.6 ± 0.7 kg BW) were allotted in a RCBD to 4 dietary treatments (NC: no-challenge; PC: challenge/no-treat; BMD: challenge/bacitracin; LBE: challenge/LBE 0.2%) and fed diets for 28 d. At d 7, challenged groups were orally inoculated with ETEC (2.4 x 1010 CFU) and NC group received sterile solution. Growth performance was analyzed weekly and pigs were euthanized on d 28 to measure intestinal health. Data were analyzed using the SAS 9.4. During post-challenge period, PC tended to decrease (P = 0.067) ADG (373 to 284 g/d), whereas BMD increased (P < 0.05) ADG (284 to 408 g/d); LBE tended to increase (P = 0.081) ADG (284 to 370 g/d). PC increased fecal score (P < 0.05, 3.4 to 3.9) on d 14, whereas BMD decreased it (P < 0.05, 3.9 to 3.5) on d 21. PC increased (P < 0.05) protein carbonyl (0.76 to 1.12 nmol/mg protein), crypt cell proliferation (28 to 36%), and Helicobacter rodentium (0.4 to 3.7%). However, BMD decreased (P < 0.05) crypt cell proliferation (36 to 32%) and Helicobacter spp. (15.0 to 1.4%); and increased (P < 0.05) villus height (309 to 377 µm), Bifidobacterium boum (0.04 to 2.0%), Pelomonas spp. (1.5 to 8.5%), and Microbacterium ginsengisoli (0.5 to 3.0%). LBE reduced (P < 0.05) crypt cell proliferation (36 to 27%) and Helicobacter rodentium (3.7 to 0.04%); and increased (P < 0.05) Lactobacillus salivarius (0.3 to 4.1%) and Propionibacterium acnes (0.4 to 7.4%). Collectively, ETEC reduced growth performance by adversely affecting microbiota and intestinal health. BMD and LBE improved growth performance by enhancing intestinal health and increasing beneficial microbiota in ETEC challenged pigs.

Unveiling the Roles of Low-Density Lipoprotein Receptor-Related Protein 6 in Intestinal Homeostasis, Regeneration and Oncogenesis

Intestinal epithelial self-renewal is tightly regulated by signaling pathways controlling stem cell proliferation, determination and differentiation. In particular, Wnt/β-catenin signaling controls intestinal crypt cell division, survival and maintenance of the stem cell niche. Most colorectal cancers are initiated by mutations activating the Wnt/β-catenin pathway. Wnt signals are transduced through Frizzled receptors and LRP5/LRP6 coreceptors to downregulate GSK3β activity, resulting in increased nuclear β-catenin. Herein, we explored if LRP6 expression is required for maintenance of intestinal homeostasis, regeneration and oncogenesis. Mice with an intestinal epithelial cell-specific deletion of Lrp6 (Lrp6IEC-KO) were generated and their phenotype analyzed. No difference in intestinal architecture nor in proliferative and stem cell numbers was found in Lrp6IEC-KO mice in comparison to controls. Nevertheless, using ex vivo intestinal organoid cultures, we found that LRP6 expression was critical for crypt cell proliferation and stem cell maintenance. When exposed to dextran sodium sulfate, Lrp6IEC-KO mice developed more severe colitis than control mice. However, loss of LRP6 did not affect tumorigenesis in ApcMin/+ mice nor growth of human colorectal cancer cells. By contrast, Lrp6 silencing diminished anchorage-independent growth of BRafV600E-transformed intestinal epithelial cells (IEC). Thus, LRP6 controls intestinal stem cell functionality and is necessary for BRAF-induced IEC oncogenesis.

The murine intestinal stem cells are highly sensitive to the modulation of the T3/TRα1-dependent pathway

The thyroid hormone T3 and its nuclear receptor TRα1 control gut development and homeostasis through the modulation of intestinal crypt cell proliferation. Despite increasing data, in depth analysis on their specific action on intestinal stem cells is lacking.By using ex vivo 3D organoid cultures and molecular approaches we observed early responses to T3 involving the T3-metabolizing enzyme Dio1 and the transporter Mct10, accompanied by a complex response of stem cell- and progenitor-enriched genes. Interestingly, specific TRα1 loss-of-function (inducible or constitutive) was responsible for low ex vivo organoid development and impaired stem cell activity. T3-treatment of animals in vivo not only confirmed the positive action of this hormone on crypt cell proliferation but also demonstrated its key action in modulating i) the number of the stem cells, ii) the expression of their specific markers and iii) the commitment of progenitors into lineage-specific differentiation.In conclusion, T3 treatment or TRα1 modulation has a rapid and strong effect on intestinal stem cells, broadening our perspectives in the study of T3/TRα1-dependent signaling in these cells.

Unveiling the Roles of Low-Density Lipoprotein Receptor-Related Protein 6 in Intestinal Homeostasis, Regeneration and Oncogenesis

Intestinal epithelial self-renewal is tightly regulated by signaling pathways controlling stem cell proliferation, determination and differentiation. In particular, Wnt/β-catenin signaling controls crypt cell division and survival and is required for maintenance of the intestinal stem cell niche. Most colorectal cancers are also initiated by mutations activating the Wnt/β-catenin pathway. Wnt signals are transduced through Frizzled receptors and LRP5/LRP6 coreceptors to downregulate GSK3β activity, resulting in increased nuclear β-catenin. Herein, we explored if LRP6 expression is required for maintenance of intestinal homeostasis, regeneration and oncogenesis. Mice with an intestinal epithelial cell-specific deletion of Lrp6 (Lrp6IEC-KO) were generated and their phenotype analyzed. No difference in intestinal architecture or in proliferative and stem cell numbers was found in Lrp6IEC-KO mice in comparison to controls. Nevertheless, using ex vivo intestinal organoid cultures, we found that LRP6 expression was critical for crypt cell proliferation and stem cell maintenance. When exposed to dextran sodium sulfate, Lrp6IEC-KO mice developed more severe colitis than control mice. However, loss of LRP6 did not affect tumorigenesis in Apc Min/+ mice nor growth of human colorectal cancer cells. By contrast, Lrp6 silencing diminished anchorage-independent growth of BRafV600E-transformed IEC. Thus, LRP6 controls intestinal stem cell functionality and is necessary for BRAF-induced IEC oncogenesis.