Investigation of the immunomodulatory effects and molecular mechanisms of cichoric acid on cyclophosphamide-induced immunosuppression in mice

Immunosuppressive diseases (ISD) are relatively common in veterinary clinical practice. The lack of therapeutic drugs for ISD severely impacts the healthy breeding of livestock and poultry. Cichoric Acid (CA), a natural active ingredient derived from traditional medicinal plants, has demonstrated a notable immunoenhancing effect. Nevertheless, the underlying mechanisms by which CA attenuates cyclophosphamide (CTX)-mediated immunosuppressive effects in mice have not yet been fully elucidated. In vivo , CA demonstrated a significant increase in body weight of CTX-induced immunosuppressed mice, elevated thymus and spleen indices, boosted levels of white blood cells, red blood cells, lymphocytes, neutrophils, and hemoglobin in the blood, while reducing average platelet volume and platelet distribution width. Histopathological findings indicated CA notably ameliorated severe thymic atrophy and cortical disappearance, as well as mitigated splenic atrophy and white pulp structural damage. CA demonstrated significant immunomodulatory effects on multiple lymphocyte populations. It substantially enhanced B-cell function by promoting proliferation and activation, as evidenced by increased production of IgM and IgG antibodies and elevated expression of CD19 protein in spleen. Concurrently, CA influenced T-cell immunity by modulating the differentiation of CD4+ and CD8+ T-cell subsets and stimulating the proliferation of CD3+ T cells. Furthermore, CA treatment markedly altered cytokine secretion profiles. It significantly suppressed the levels of pro-inflammatory cytokines, including tumor necrosis factor-α (TNF-α) and interleukin-12 (IL-12), while promoting the secretion of anti-inflammatory mediators such as interleukin-10 (IL-10) and transforming growth factor-β (TGF-β). These cytokines shift effectively inhibited macrophage polarization towards the M1 phenotype, thereby contributing to an overall enhancement of immune regulatory function. In vitro , CA markedly boosted cell viability and phagocytosis in immunosuppressive RAW264.7 macrophages stimulated by phosphoramide mustard (PM). It enhanced macrophage-derived exosomes (Exos) secretion, modulated mRNA, lncRNA and microRNA expression within Exos, influenced Ras, PI3K/Akt, MAPK, and B cell receptor signaling pathways, and triggered the upregulation of PI3K/Akt signaling pathway-associated proteins in B cells, thereby eliciting immunoregulatory responses. In conclusion, CA demonstrated a significant immunopotentiation on mice with CTX-induced immunosuppression via regulating the crosstalk of macrophages and B cells by Exos. These findings established a robust foundation for the potential treatment of ISD in veterinary clinical settings. • CA reversed CTX-induced immunosuppression in mice by restoring body weight, thymus/spleen indices, and organ atrophy. It improved hematological parameters and lymphocyte subsets. • CA enhanced B-cell proliferation, antibody production, and CD19 expression in spleen. It balanced CD4+/CD8+ T-cell ratios and stimulated CD3+ T-cell proliferation while regulating inflammatory cytokines. • CA improved RAW264.7 macrophage viability and phagocytosis, and shifted M1 polarization to M2. • CA-enhanced macrophage-derived exosomes modulated ncRNAs profiles, upregulated PI3K/Akt proteins in B cells, and facilitated macrophage-B-cell communication to restore immune homeostasis.