Lanternfish (Benthosema pterotum) Hydrolysates: Dual Action on Immune Regulation

Background: Lanternfish ( Benthosema pterotum ), an abundant deep-sea resource with a unique protein profile, holds significant potential as a source of bioactive peptides. This study was designed to investigate and characterize the immunomodulatory properties of protein hydrolysates derived from lanternfish, prepared using Alcalase (BPHA) and Flavourzyme (BPHF), on the RAW 264.7 murine macrophage cell line. Methods: Lanternfish protein was enzymatically hydrolyzed, and the resulting peptides were fractionated to isolate those with a molecular weight below 3 kDa. RAW 264.7 macrophages were treated with B. pterotum hydrolyzed using Alcalase enzymes (BPHA) and B. pterotum hydrolyzed using Flavourzyme enzymes (BPHF) at concentrations ranging from 100 to 500 µg/mL. The cytotoxicity of the hydrolysates was evaluated using an 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) assay. To assess immunomodulatory activity, the gene expression levels of pro-inflammatory mediators, including Inducible nitric oxide synthase (iNOS), Tumor necrosis factor alpha (TNF-α), Interleukin-1 beta (IL-1β), and Interleukin-6 (IL-6), were quantified by quantitative real-time PCR in both resting macrophages and in cells stimulated with lipopolysaccharide (LPS) to induce an inflammatory state. Results: The MTT assay confirmed that both BPHA and BPHF were noncytotoxic across all tested concentrations. In resting (nonstimulated) macrophages, both hydrolysates significantly upregulated the expression of iNOS, TNF-α, IL-1β, and IL-6 in a dose-dependent manner, demonstrating a clear immunostimulatory effect. In contrast, when macrophages were exposed to an inflammatory challenge with LPS, the hydrolysates exhibited potent anti-inflammatory activity by significantly downregulating the expression of these same pro-inflammatory genes. Conclusions: Protein hydrolysates derived from lanternfish display a notable dual immunomodulatory capability. They function as immunostimulants in a normal physiological state and act as anti-inflammatory agents during an inflammatory response. These findings underscore the potential of lanternfish hydrolysates as a valuable functional ingredient for applications in nutraceuticals and functional foods aimed at balancing and modulating the immune system.