Pathogen-based cancer therapies have been widely studied. Parasites, such as Toxoplasma gondii have elicited great interest in cancer therapy. Considering safety in clinical applications, we tried to develop an exosome-based immunomodulator instead of a live parasite for tumor treatment. The exosomes, called DC-Me49-exo were isolated from culture supernatants of dendritic cells (DCs) infected with the Me49 strain of T. gondii and identified. We assessed the antitumoral effect of these exosomes in a mouse model of colorectal cancer (CRC). Results showed that the tumor growth was significantly inhibited after treatment with DC-Me49-exo. Proportion of polymorphonuclear granulocytic bone marrow-derived suppressor cells (G-MDSCs, CD11b⁺Ly6G⁺) and monocytic myeloid-derived suppressor cells (M-MDSCs, CD11b⁺Ly6C⁺) were decreased in the DC-Me49-exo group compared with the control groups in vitro and in vivo. The proportion of DCs (CD45⁺CD11c⁺) increased significantly in the DC-Me49-exo group. Levels of interleukin-6 (IL-6) and granulocyte-macrophage colony-stimulating factor (GM-CSF) significantly decreased after treatment with DC-Me49-exo. Furthermore, we found that DC-Me49-exo regulated the lever of MDSC mainly by inhibiting the signal transducer and activator of transcription (STAT3) signaling pathway. These results indicated that exosomes derived from DCs infected with T. gondii could be used as part of a novel cancer therapeutic strategy by reducing the proportion of MDSCs.

Ticks, as blood-feeding arthropod vectors, have evolved their own unique mechanism to suppress host immune responses and evade immune defenses in order to complete blood-feeding. The immunoregulatory effect of tick bioactive molecules on hosts has been widely reported, and the cystatin family has been identified as one of the major immunomodulators. In previous studies, we obtained a novel tick salivary bioactive protein named RHcyst-1, which belongs to the type 1 cystatin family. Here, we demonstrated the effects of RHcyst-1 on the host immune response mainly on dendritic cell (DC) function. Understanding the function of tick-derived bioactive molecule may help to clarify the mechanisms of how ticks escape the host immune response and help to control ticks and tick-borne disease transmission. Bone marrow-derived DCs (BMDCs) were generated and induced by GM-CSF and IL-4 with or without RHcyst-1 addition. Flow cytometry was used to analyze the differentiation and maturation of BMDCs and T cell cytokine production. Quantitative real-time PCR (qRT-PCR) and western blot were used to measure changes in expression within STAT and p38 MAPK signaling pathways. Flow cytometry analysis revealed that RHcyst-1 inhibited the differentiation of BMDCs, but had no effect on the maturation of BMDCs. T cells co-cultured with DCs treated with RHcyst-1 produced significantly less TNF-α, IFN-γ and IL-2 than the control group. Further analysis showed that the mRNA level and phosphorylation of p38, ERK and STAT were significantly changed after RHcyst-1 added to bone marrow monocytes during the differentiation stage. Our results suggest that RHcyst-1 is one of the major immunosuppressive proteins of BMDC function from blood-feeding ticks.

Rhipicephalus haemaphysaloides is a widespread tick species in China and other South East Asian countries, where it is the vector of many pathogens. The objective of this study was to study the role of serpin (serine protease inhibitor) during the tick-host interaction. The differentiation of bone marrow-derived dendritic cells (BMDC) was induced in vitro, and the effect of RHS2 on the maturation of DCs was evaluated. The effects of RHS2 on T cell activation and cytotoxic T lymphocytes’ (CTLs) activity were analyzed by flow cytometry. Antibody subtypes after immunization of mice with RHS2 and OVA were determined. RHS2 can inhibit the differentiation of bone marrow-derived cells into DCs and promote their differentiation into macrophages. RHS2 can inhibit the maturation of DCs and the expression of CD80, CD86 and MHCII. The number of CD3+CD4+ and CD3+CD8+ T cells secreting IFN-γ, IL-2 and TNF-α was decreased, and the number of CD3+CD4+ T cells secreting IL-4 was increased, indicating that RHS2 can inhibit the activation of CD4 T cells and CD8 T cells, leading to inhibition of Th1 immune response. RHS2 inhibits the elimination of target cells by cytotoxic T lymphocytes. After immunization of mice with RHS2 and OVA, serum IgG2b was significantly reduced and IgM was increased. The results show that RHS2 has an inhibitory effect on the host immune response. Ticks have evolved various ways to circumvent adaptive immunity. Their serpin inhibits BMDC differentiation to reduce immune responses.

The study of tick olfaction is relatively new compared to that of insects, and the molecular mechanisms involved remain poorly understood. Despite several potential chemosensory genes identified in multiple tick species, these are yet to be validated through independent functional experiments. In this research, we cloned and analyzed a microplusin-like gene, HlonML-1, and investigated its role in the chemosensory activities of H. longicornis. The results showed that this gene’s amino acid sequences lack histidine residues essential for antimicrobial activity, and it is evolutionarily linked to putative chemosensory microplusins in ticks. Gene expression analyses indicated that HlonML-1 was significantly more abundant in ticks exposed to potential attractants and in the forelegs of H. longicornis than in non-exposed ticks and the hindlegs, respectively. Tick forelegs support the Haller’s organ, which is a sensory structure mostly specialized for chemosensation. Furthermore, Y-tube olfactometer assays indicated that silencing HlonML-1 significantly impaired adult ticks’ ability to detect selected odors, while their gustatory-related behavior remained unaffected compared to the control groups. Given its unique sequences, relative abundance in chemosensory tissues, and impact on odor detection, HlonML-1 is likely involved in the olfactory chemosensation of H. longicornis. Future research validating putative chemosensory microplusins in the genomes of various tick species may enhance our understanding of their olfactory functions in tick and lead to the identification of new molecular targets for developing tick repellents.