The galectin-4-glycan axis in intestinal inflammation: friend or foe?

MASSARO, MORA; CAGNONI, ALEJANDRO J; MANSELLE COCCO, MONTANA N; CUTINE, ANABELA M; MERLO, JOAQUÍN P; VEIGAS, FLORENCIA; GATTO, SABRINA; MORALES, ROSA M; PEREZ-SAEZ, JUAN M; LENIS, BLADIMIRO; TOSCANO, MARTA A; RABINOVICH, GABRIEL A; MARIÑO, KARINA V

Mar del Plata

Congreso; 2022 Reunion Anual Sociedad Argentina de Inmunologia; 2022

Sociedad Argentina de Inmunología

Inflammatory Bowel Diseases (IBD) including ulcerative colitis (UC) and Crohn’s disease (CD) constitute a group of chronic disorders that affect the gastrointestinal tract. Limitations of the currently available treatments generate a need for new therapeutic targets. Galectin-4 (Gal4) is a tandem-repeat type lectin with two carbohydrate recognition (CRD) domains that is predominantly expressed in intestinal epithelial cells, but its role in IBD is still under debate. In this work, we analyzed single cell RNAseq transcriptomic data from UC patients and found that Gal4 is downregulated when compared to healthy controls (p-val=4.9e-68). We then evaluated the influence of Gal4 using transgenic mice with a specific depletion of Gal4 in the intestinal epithelium (CreVill+/-/FloxGal4+/+). Although development of dextran sulfate sodium (DSS) acute colitis in these mice did not show statistically significant differences in weight loss and colon weight/length ratio when compared to WT littermates, we found significant differences in the T cell compartment, as mesenteric lymph nodes showed increased proportion CD4+ CD62L+ CD44+ memory T cells (p-val=0.009). No differences in the myeloid or B cell compartments were found. Next, and in order to explore the functional role of this protein, we have optimized the recombinant expression and purification of human Gal4 in E. coli BL21 (DE3) cells. As the His-Gal4 construct did not show optimal results, vector pET-28a-SUMO-Gal4 was finally selected for transformation. For purification, the SUMO tag was cleaved, and two sequential affinity columns (Ni-NTA and lactose) were used, obtaining 2 mg/L of active hGal4. Activity was biochemically tested by solid phase assays and in vitro, as hGal4 was able to induce interleukin-6 secretion in activated (anti-CD3 and anti-CD28) splenocytes. In summary, with these preliminary results we set up the basis for further characterization of the role of the Gal4-glycan axis in intestinal inflammation.