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ABSTRACT
Background: Previous studies suggest that intestinal epithelial stem cells (IESC), critical drivers of homeostasis and regeneration, include two subpopulations: crypt-based columnar and “position +4” stem cells, identified by Lgr5 and Bmi1 biomarkers, respectively. Teduglutide is an enterotrophic counterpart of glucagon-like peptide 2. This study aimed to investigate the response of putative IESC to surgical injury and teduglutide administration on an animal model of intestinal resection and anastomosis. Methods: Wistar rats (n = 59) were distributed into four groups: “Ileal Resection” versus “Laparotomy”, subsequently subdivided into “Postoperative Teduglutide Administration” versus “No Treatment”; and sacrificed at third or seventh days, with ileal sample harvesting. Flow cytometry was used to analyze epithelial stem cells with monoclonal antibodies against Lgr5, Bmi1 and also CD44, CD24, CD166, and Grp78 surface markers. Results: Surgical trauma induced an increase of epithelial stem cells population at third day (9.0 ± 0.3 versus 5.7 ± 0.3%, p = 0.0001), which was more intense and involved all subpopulations after ileal resection. At seventh day, teduglutide was significantly associated with higher proportion of Lgr5+/Bmi1− cells (5.8 ± 0.1 versus 2.9 ± 0.3%, p = 0.005) and, on the contrary, lower percentage of Lgr5−/Bmi1+ cells (0.03 ± 0.01 versus 1.9 ± 0.1%, p = 0.049) after ileal resection; and higher proportion of Lgr5+/Bmi1+ cells (1.7 ± 0.1 versus 1.1 ± 0.2%, p = 0.028) after isolated laparotomy. After surgery, Lgr5+/Bmi1− and Lgr5−/Bmi1+ subpopulations demonstrated an inverse correlation and both correlated negatively with Grp78 labeling index. Lgr5−/Bmi1+ and CD44+/CD24low/CD166+/Grp78+ cells proportions exhibited a high grade positive correlation. Conclusion: Those observations support the existence of two epithelial stem cells subpopulations with distinct behavior after surgical injury and teduglutide treatment.
ABBREVIATIONS
| Bmi1 | = | B cell-specific Maloney murine leukemia virus integration region 1. |
| CK18 | = | Cytokeratin 18. |
| CD45 | = | Cluster of differentiation 45. |
| CD31 | = | Cluster of differentiation 31. |
| CD166 | = | Cluster of differentiation 166. |
| CD24 | = | Cluster of differentiation 24. |
| CD44 | = | Cluster of differentiation 44. |
| FACS | = | Fluorescence-activated cell sorting. |
| Grp78 | = | 78 KDa glucose-regulated protein. |
| Hopx | = | Homeodomain-only. |
| K3EDTA | = | K3 ethylenediaminetetraacetic acid. |
| Lgr5 | = | Leucine-rich repeat-containing G-protein coupled receptor. |
| Lrig1 | = | Leucine-rich repeats and immunoglobulin-like domains 1. |
| SEM | = | Standard error of the mean. |
| Tert | = | Telomerase reverse transcriptase. |
| USF | = | United States Pharmacopeia. |
| vs. | = | versus |
| α-Sma | = | α-smooth muscle actin. |
| σ | = | Spearman's correlation coefficient. |
ACKNOWLEDGMENTS
Not applicable.
DECLARATION OF INTEREST
The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper.
FUNDING
The authors declare no funding for the research.
ORIGINAL COMMUNICATION
This manuscript represents an original research that was not submitted elsewhere for publication and that will not be sent to another journal until a decision is made concerning publication.
This study was partially presented as an oral communication at the 37th European Society for Clinical Nutrition and Metabolism (ESPEN) Congress, Lisbon, in September 8th, 2015 and the abstract was published in Clinical Nutrition [Costa BP, Sarmento-Ribeiro AB, Gonçalves AC, Botelho F, Abrantes M, F. Castro Sousa: Response of putative intestinal epithelial stem cells to surgical injury and teduglutide administration on an animal model. Clin Nutr 2015; 34 (Supp1):S19].
ETHICS APPROVAL
Study was implemented in accordance with the national animals' welfare guidelines and was authorized by the Ethics Committee of Faculty of Medicine, University of Coimbra, Coimbra, Portugal.
STATEMENT OF AUTHORSHIP
All the authors have made substantial contributions to the preparation of the manuscript.
AUTHORS' CONTRIBUTIONS
Study conception and design: Costa B
Acquisition of data: Costa B, Gonçalves AC, Abrantes AM, Matafome P, Alves R
Analysis and interpretation of data: Costa B
Drafting of manuscript: Costa B
Critical review: Seiça R, Sarmento-Ribeiro AB, Botelho MF, Castro Sousa F