*3.6. Immunomodulatory E*ff*ects*

Allergic disease is a prevalent aberrant immune responsive against innocuous environmental proteins (antigens) [137]. Allergen-specific CD4+ T cells involved in the initiation of allergic reactivity can develop into either type 1 or type 2 helper T cells [138]. CD4+ T cells stimulated in the presence of interleukin-12 and γ-interferon can develop into type 1 helper T cells [138], while interleukin-4 promotes the development of type 2 helper T cells and inhibits the generation of type 1 helper T cells [139]. Since type 1 and 2 helper T cells regulate each other, interleukin-12 functions not only to induce the type 1 helper T-cell response but also to regulate the type 2 helper T-cell response [140]. Interleukin-12 strongly suppresses the production of IgE by preventing type 2 helper T-cell development [141]. Allergen-specific IgE induces the pathogenesis of allergic disorder [142].

Hasegawa et al. [143] described the e ffects of a *Chlorella* (*C. vulgaris*) hot-water extract on antigen specific response in mice. A 2% ( *w*/*w*) *Chlorella* hot-water extract diet or control diet (without *Chlorella* extract) was given to mice for two weeks before intraperitoneal administration of casein/complete Freund's adjuvant (an immunostimulant). Mice that received the hot-water extract exhibited suppressed IgE production and mRNA expression of interleukin-6 involved in the type 2 helper T-cell response. They also exhibited increased levels of interleukin-12 and g-interferon mRNA, increasing the type 1 helper T-cell response and suppressing the type 2 helper T-cell response. These results sugges<sup>t</sup> that *Chlorella* hot-water extract supplementation might be useful for suppressing allergic responses with a predominant type 2 helper T-cell response. To clarify the mechanisms underlying the immunomodulatory activity of *Chlorella* hot-water extract, soluble polysaccharides were isolated from *C. pyrenoidosa* hot-water extract and characterized [144]. GC-MS analysis indicated that the major monosaccharide components of the soluble polysaccharides are rhamnose (31.8%), glucose (20.4%), galactose (10.3%), mannose (5.2%), and xylose (1.3%). These soluble polysaccharides were intraperitoneally administrated (100 mg/kg of body weight) to 6–8-week-old mice. After 24 h, lipopolysaccharide as an antigen was administrated to mice, and their serum was collected 1.5 h later [144]. The soluble polysaccharides induced interleukin-1b secretion in macrophages via the toll-like receptor protein kinase signaling pathway. Interleukin-1β is one of the most important mediators of inflammation and host responses to infection [145]. These results sugges<sup>t</sup> that*Chlorella* hot-water-soluble polysaccharides could be used as an agen<sup>t</sup> source to stimulate anti-microorganism activity.

Halperin et al. [146] evaluated the e ffect of *C. pyrenoidosa* supplementation (200 or 400 mg) on the immune response to influenza vaccination. After 28 days of *Chlorella* supplementation, the antibody response to the influenza vaccine was not elevated in the overall study population but was increased in participants aged 50–55 years.

Salivary secretory immunoglobulin A (SIgA) plays a crucial role in mucosal immune function and is the first line of defense against pathogenic microbial invasion in human [147]. To evaluate whether *Chlorella* supplementation increases salivary SIgA secretion in humans, a blind, randomized, crossover study was conducted in participants administered *Chlorella* (*C. pyrenoidosa*) (6 g/day) or placebo for four weeks [148]. Although no di fference was observed in salivary SIgA levels before and after placebo ingestion, salivary SIgA levels were significantly elevated after *Chlorella* ingestion than at baseline. The SIgA secretion rate increased significantly after *Chlorella* supplementation. These results sugges<sup>t</sup> that four-week *Chlorella* supplementation increases salivary SIgA secretion and improves mucosal immune function in humans.

Natural killer cells are the predominant innate lymphocyte subsets that mediate antitumor and antiviral responses [149]. To evaluate the e ffect of *Chlorella* supplementation on natural killer cell activity and early inflammatory response in humans, a randomized, double-blinded, placebo-controlled trial was conducted in healthy adults ingested with *Chlorella* (*C. vulgaris*) (5 g/day) or placebo [150]. After eight weeks of supplementation, serum interferon-γ and interleukin-1β levels were significantly elevated and that of interleukin-12 tended to increase in the *Chlorella* group. Natural killer cell activities were significantly elevated in the *Chlorella* group. These results sugges<sup>t</sup> a beneficial immunostimulatory effect of short-term *Chlorella* supplementation that increases natural killer cell activity and produces interferon-γ, interleukin-12, and interleukin-1β.
