Fig. 2: Low Ser plasma levels induce muscle wasting in in vivo CRC models.

A Workflow for the in vivo assessment of the wasting effect of Ser dietary modulation. CRC cells were injected subcutaneously into the flank of Balb/c mice or Foxn1nu/nu Athymic-Nude mice. Two weeks later, tumor size was assessed by caliper measurement, and mice were randomly divided into experimental groups: control diet, Ser-Gly lacking diet (-S-G diet). Mice were fed with a diet containing Ser and Gly (+S + G) or lacking these two amino acids (-S-G) until the end of the experiment. Ser was added (+Ser) or not in drinking water (20 g/L) 5 days after diet change (n = 3). B Plasma Ser mice abundance in mice bearing HT29-PHGDH low-derived tumors. Relative Ser levels were quantified by GS-MS analysis in plasma from Foxn1nu/nu Athymic-Nude mice bearing HT29-PHGDH low-derived tumors and fed with +S + G or -S-G diet. Each dot represents a plasma sample derived from a single mouse. Data are represented as mean ± SEM, Student’s t test, **p < 0.01. C Evolution of mice weight over the time. Foxn1nu/nu Athymic-Nude mice bearing HT29-low-derived tumors were fed with a +S + G or -S-G two weeks after tumor cells injection (diet change). Mice’s weight was assessed every two days until experiment endpoint. Weight values are normalized to the average weight at the day of diet change. Data are represented as mean ± SEM of at least three mice. D, E Gastrocnemius muscle mass. D Representative high-resolution ultrasound images of gastrocnemius muscle of Athymic-Nude mice bearing HT29-low-derived tumors fed with +S + G or -S-G diet. E Quantification of gastrocnemius muscle volume of mice as in (D) measured by the dedicated in vivo imaging system (Vevo LAZRX photoacoustic imaging). Data are represented as mean ± SEM of three independent measurements, Student’s t test, *p < 0.05. F Cross-sectional histological staining of gastrocnemius muscle fiber. Representative images of Hematoxylin and Eosin (H&E) staining of gastrocnemius skeletal muscle collected from Athymic-Nude mice bearing HT29-low-derived tumors fed with +S + G or -S-G diet (magnification ×10). G Quantification of gastrocnemius myofiber cross-sectional area. Gastrocnemius skeletal muscles were collected from at least three Athymic-Nude mice bearing HT29-low-derived tumors fed with +S + G or -S-G diet, tissues were stained with H&E, and cross-sectional fibers diameter was qualified by ImageJ. Values are represented as mean ± SEM, Unpaired t-test, **p < 0.01. H Ser plasma levels following Ser supplementation in drinking water of mice. Ser levels were quantified by GS-MS analysis in plasma from Foxn1nu/nu Athymic-Nude mice bearing HT29-PHGDH low-derived tumors and fed with +S + G or -S-G diet. Ser was added (+Serine) or not in drinking water (20 g/L) 5 days after diet change. Each dot represents a plasma sample derived from a single mouse. Data are represented as mean ± SEM, One-way ANOVA with Sidak’s post hoc test, **p < 0.01. I Evolution of weight over the time. Foxn1nu/nu Athymic-Nude mice bearing HT29-PHGDH-low derived tumors were treated as in (F) and mice weight was monitored each two days until experiment endpoint. Data are reported as relative to the average weight at the day of diet change and represented as mean ± SEM of at least three mice.