Immunostimulatory Activity without Pathological Effects of Fish Protein Hydrolysate from Clarias Catfish

Introduction
Fish protein hydrolysate (FPH), in particular, contains small, crucial peptides that offer beneficial health effects. However, research on its immunological effects remains limited. This study investigates the impact of fish hydrolysate on immune-related tissues (blood and spleen) and visceral organs (liver and kidneys) in rats.

Methods
FPH was produced from the hydrolysis of Clarias catfish-minced meat using Alcalase. In vivo evaluation was conducted using freeze-dried FPH. Thirty male Wistar rats (150–200 g) were divided into five groups: placebo, three FPH doses (0.125, 0.250, and 0.500 mg/g BW), and one commercial product (0.125 mg/g BW). Treatments were given orally by gavage, diluted in 1 mL of distilled water, once daily for 14 days. On day 15, rats were euthanized, and blood, spleen, liver, and kidneys were collected. Blood parameters were analyzed using an automatic hematology analyzer. Organ weight and size were measured morphometrically. White pulp area, glomerular width, and liver sinusoid size were measured using a microruler in ImageJ. Splenic IgM expression was assessed via immunohistochemical (IHC) staining and quantified using the ImmunoRatio plugin in ImageJ. A docking study was conducted between two peptides with the Mas receptor and Toll-like receptor 4 (TLR4).

Results
Studies confirmed that catfish-derived FPH does not cause pathological changes in the spleen, liver, or kidneys, aside from liver sinusoid dilation. Additionally, it significantly increases (p < 0.05) circulating leukocytes, differential leukocyte counts, and IgM expression in the spleen. Two peptides from catfish-derived FPH showed good affinity for the Mas receptor and TLR4.

Conclusion
These findings indicate that catfish-derived FPH can improve immune function without causing damage to organs.