Effect Of Salivary Gland Extract Research Articles

In Vitro Assessment of the Anti-Proliferative and Anti-Viability Effects of Salivary Gland Extracts from Hyalomma ticks (Acari: Ixodidae) on Human Colorectal Cancer Cells

Background: The saliva and salivary glands of ticks possess a wide range of immuno-pharmacologically active mole­cules that effectively modulate the activity of enzymes, antibodies, and amines that have a role in different biological processes. Derived components from saliva and salivary glands of hard ticks Ixodidae have been characterized as poten­tial natural sources for discovering promising anti-cancer drug candidates.
 Methods: The anti-cancer activity of salivary gland extracts (SGEs) from Hyalomma anatolicum, Hyalomma drome­darii, Hyalomma marginatum, and Hyalomma schulzei was assessed. MTT assays and flow cytometry were done on the HT-29 colorectal cancer cell line to evaluate the anti-viability and proliferative inhibition.
 Results: Based on the MTT assay results, the SGEs from Hy. dromedarii had the highest and lowest substantial anti-viability effects on the HT-29 cancer cell and human foreskin fibroblast (HFF) normal cell, respectively. The cytometric assessment revealed a significant increase in the apoptosis and necrosis ratio of the HT-29 cancer cells after treatment with Hy. dromedarii SGEs.
 Conclusion: The results demonstrated that Hy. dromedarii SGEs have significant anti-proliferative, anti-viability, and apoptotic potential. The result of this study suggests that Hy. dromedarii SGEs is an appropriate candidate for further investigations to identify and purify the mechanisms and molecules involved in the anti-cancer activity of the SGEs. 

Dissociating effect of salivary gland extract from Ixodes ricinus on human fibroblasts: Potential impact on Borrelia transmission

Understanding the mechanism of pathogen transmission is essential for the development of strategies to reduce arthropod-borne diseases. The pharmaco- and immunomodulatory properties of insect and acarine saliva play an essential role in the efficiency of pathogen transmission. The skin as the site where arthropod saliva and pathogens are inoculated - represents the key interface in vector-borne diseases. We identified tick molecules potentially involved in pathogen transmission, using micro-HPLC and mass spectrometry, followed by in vitro assays on human skin cells. Histone H4 isolated from Ixodes ricinus salivary gland extract was identified as a molecule with a dissociating effect on human primary fibroblasts. This histone might be involved in the formation of the feeding pool formed around the tick mouthparts and responsible of tissue necrosis in the vertebrate host. Thanks to its selective antimicrobial activity, it may also sterilize the feeding pool and facilitate transmission of pathogens such as Borrelia burgdorferi sensu lato.

Immunomodulatory and morphophysiological effects of Rhipicephalus sanguineus s. l. (Acari: Ixodidae) salivary gland extracts

Rhipicephalus sanguineus s. l. is popularly known as the “brown dog tick” since dogs are its preferential hosts, but the species has been reported to parasitize other mammals, including humans, with significant medical-veterinary importance since it transmits several important pathogenic agents during the feeding period. The tick saliva is a complex mixture that has several functions, including the capability to modulate the hemostatic, inflammatory and immunologic systems of the host, allowing pathogens to settle. Despite knowledge about the immunosuppressive action of tick saliva, little is known about the mechanisms involved in this process and the morphophysiological effects caused by exposure to the salivary gland extract, taking into consideration the different periods of the glandular cycle. Thus, the objective of this study was to analyze the in vitro effects of salivary gland extracts obtained from R. sanguineus s. l. females fed on host rabbits for two (SGE2 – Salivary Gland Extracts of 2 days) and four days (SGE4 – Salivary Gland Extracts of 4 days) on J774 cells (monocyte macrophage cell line) and verify the occurrence of morphological and immunomodulatory alterations in these cells when exposed to different concentrations of these extracts. The results showed that: (i) SGE2 and SGE4 at the concentration of 4 μg/mL presented cytotoxicity to the J774 cells exposed for 24 and 48 hours; (ii) SGE2 at the concentrations of 2 μg/mL(48-hour exposure) and 1 μg/mL (24-hour exposure) and SGE4 at the concentrations of 2 and 1 μg/mL (48-hour exposure) showed proinflammatory activity, confirmed by the increased secretion of NO and proinflammatory cytokine (IL-2), and the presence of morphological characteristics detected by microscopy; and (iii) SGE2 and SGE4 at the concentrations of 0.5 and 0.1 μg/mL had immunomodulatory activity, demonstrated by decreases in the secretion of NO and proinflammatory cytokines (IL2, IL-6 and TNF-α) and increase in the synthesis of IL-10, confirmed by the morphophysiological analysis. These unprecedented data are extremely relevant for future research to identify the processes involved in the ectoparasite-host relationship, as well to develop more efficient tick control strategies.

Ixodid tick salivary gland extracts suppress human transforming growth factor-β1 triggered signalling pathways in cervical carcinoma cells

Ticks, very successful vectors and reservoirs of diverse viruses and other infectious agents, are able to counteract host defence mechanisms and wound healing processes to facilitate blood-feeding. Cutting the epidermis by the tick chelicerae and penetration of the hypostome into the dermis followed by feeding elicit a wound healing response, involving coagulation, inflammation, angiogenesis, synthesis of extracellular matrix and ground substances, as well as tissue remodelling. These repair processes of injured skin are coordinated by a wide array of cytokines, chemokines and growth factors. Tick salivary gland compounds, in addition to their vital anticoagulant and immunomodulatory functions, also possess significant anti-tumour properties and represent very promising sources of novel pharmaceuticals. Among life-threatening cancers, cervical carcinoma is still a high risk. Human transforming growth factor-beta 1 (huTGF-β1) inhibitors are significant in anti-cancer therapy. In this study, we investigated effects of salivary gland extracts (SGE) of females of two ixodid ticks, Dermacentor reticulatus (DR SGE) and Hyalomma anatolicum excavatum (HAE SGE), on signalling pathways triggered by huTGF-β1 in two cervical carcinoma cell lines, SiHa and HeLa, varying in their response to huTGF-β1. By dual luciferase reporter assay we monitored activation/inhibition of canonical SMAD or non-canonical signalling pathways in cells treated with huTGF-β1 and DR SGE/HAE SGE. Using specific monoclonal antibodies we tested effects of SGE on phosphorylation of ERK1/2 and/or AKT-1 molecules. We were the first to detect significant inhibition of huTGF-β1 induced signalling pathways, canonical as well as non-canonical, by DR and HAE SGE. Although the overall effects of both SGE on signalling pathways were similar, we revealed different patterns of effects of the two SGE on phosphorylation of the monitored signal transducers. Further investigations in this field are required.

Effect of O. porcinus Tick Salivary Gland Extract on the African Swine Fever Virus Infection in Domestic Pig

African swine fever is a haemorrhagic disease in pig production that can have disastrous financial consequences for farming. No vaccines are currently available and animal slaughtering or area zoning to restrict risk-related movements are the only effective measures to prevent the spread of the disease. Ornithodoros soft ticks are known to transmit the African swine fever virus (ASFV) to pigs in farms, following the natural epidemiologic cycle of the virus. Tick saliva has been shown to modulate the host physiological and immunological responses during feeding on skin, thus affecting viral infection. To better understand the interaction between soft tick, ASFV and pig at the bite location and the possible influence of tick saliva on pig infection by ASFV, salivary gland extract (SGE) of Ornithodoros porcinus, co-inoculated or not with ASFV, was used for intradermal auricular inoculation. Our results showed that, after the virus triggered the disease, pigs inoculated with virus and SGE presented greater hyperthermia than pigs inoculated with virus alone. The density of Langerhans cells was modulated at the tick bite or inoculation site, either through recruitment by ASFV or inhibition by SGE. Additionally, SGE and virus induced macrophage recruitment each. This effect was enhanced when they were co-inoculated. Finally, the co-inoculation of SGE and virus delayed the early local spread of virus to the first lymph node on the inoculation side. This study has shown that the effect of SGE was powerful enough to be quantified in pig both on the systemic and local immune response. We believe this model should be developed with infected tick and could improve knowledge of both tick vector competence and tick saliva immunomodulation.

The Effect of Salivary Gland Extract of <i>Lucillia Sericata</i> Maggots on the Human Dermal Fibroblasts Proliferation

Problem statement: The aim of study was to investigate the effect of Salivary Gland Extract (SGE) of Lucillia sericata maggots on Human Neonatal Fibroblasts (B-HNF-1) proliferation and migration within collagen/hyaluronan membrane. Chronic, non-healing wounds are major health care problem worldwide. Nowadays, Maggot Debridement Therapy (MDT) has attracted much attention due to its successfull application in a treatment of chronic wounds where conventional treatment has failed. Maggot Excretion/Secretion (ES) contains specific proteinases which are positivelly involved in the remodeling of Extracelallar Matrix (ECM) components. Approach: Immunohistochemical methods using primary monoclonal antibody against marker vimentin was employed for the visualization of fibroblasts cultured within collagen hyaluronan membrane. Results: We found that maggot SGE at concentration of 2.4 and 4.8 gL mL-1 significantly induced the migration of fibroblasts into the membrane after 5 and 10 days of cultivation. On the other hand, SGE at concentration of 7.2 gL mL-1 reduced the migration of fibroblasts. Conclusion: The obtained results suggest that medicinal maggots secretion may speed up the wound healing process through the stimulation of fibroblast migration. However, further detailed research is needed to fully elucidate the substance (s) responsible for fibrloblast stimulation and migration.

Salivary gland extract from engorged Ixodes ricinus (Acari: Ixodidae) stimulates in vitro growth of Borrelia burgdorferi sensu lato

In vitro effect of salivary gland extract from fed Ixodes ricinus, the competent vector of Lyme borreliosis in Europe, on the growth of Borrelia burgdorferi sensu lato (B. garinii, B. afzelii and B. burgdorferi sensu stricto) was examined in BSK-H medium. Motility rate, concentration of motile spirochetes and their morphology were estimated at intervals of 0, 2, 4, 6 and 8 days using darkfield microscopy. Salivary gland extract derived from I. ricinus stimulated markedly the growth of three genomic species of borreliae. The results confirm a substantial role of salivary glands in the mechanism of pathogen transmission to vertebrate host.

The Effects of Salivary Gland Extracts from Boophilus microplus Ticks on Mitogen-stimulated Bovine Lymphocytes

The effect of salivary gland extract (SGE) from the tick Boophilus microplus was examined in mitogen-stimulated lymphocytes in vitro. SGE was added to lymphocytes of seven cattle together with the mitogens concanavalin A (ConA), phytohaemagglutinin (PHA) and pokeweed mitogen (PWM). Semi-purified B cells from another seven cattle were stimulated with the mitogen lipopolysaccharide (LPS). PHA and ConA stimulated proliferation of lymphocytes to the same extent, but the inhibition due to SGE of Boophilus microplus on the proliferative response stimulated by PHA (39.0% +/- 9.3%) was less than the inhibition of proliferative response stimulated by ConA (75.4% +/- 6.9%). In contrast, SGE of B. microplus stimulated the proliferation of B cells in the presence of LPS in a dose-dependent manner. Enhanced stimulation of B cells by SGE at >4 microg in culture was greater than twice that observed when B cells were stimulated by LPS alone. SGE does not have a direct suppressive effect on bovine B cell proliferation; however, in vivo the effectiveness of B cell responses might be influenced by other immune factors, such as cytokine profiles.

Effect of Tick Saliva on Mechanisms of Innate Immune Response Against <I>Borrelia afzelii</I>

The effect of salivary gland extract (SGE) and saliva from the tick Ixodes ricinus (L.) on the interaction of Borrelia afzelii spirochetes with mouse macrophages as well as on the borreli-acidal effect of calf serum was studied. SGE reduced both the number of phagocytosing cells and phagocytosed bacteria. An inhibitory effect of SGE on the killing of spirochetes by the alternative pathway of complement activation also was observed. Both SGE and saliva down-regulated production of proinflammatory cytokine tumor necrosis factor-alpha by macrophages stimulated with interferon-gamma and live B. afzelii spirochetes. The production of another macrophage cytokine, interleukin-6, remained unchanged. SGE and saliva exerted a different effect on the production of nitric oxide by stimulated macrophages. Whereas SGE up-regulated NO production, saliva decreased it. The significance of immunosuppressive effects of tick saliva for the transmission of Borrelia spirochetes is discussed.

Tick modulation of the in-vitro expression of adhesion molecules by skin-derived endothelial cells

As a tick feeds, its saliva induces innate and acquired immune responses in the host, including leucocyte infiltration into the bite site. Tick salivary glands produce molecules, however, that counteract many host defences against blood feeding. The effects of salivary-gland extracts (SGE) of Dermacentor andersoni and Ixodes scapularis on the expression of various adhesion molecules [E-selectin, P-selectin, intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1)] by the sEND.1 cell line (which is based on cells from the subcutaneous tissue of mice) have now been investigated in vitro. The effects were found to differ with the tick species. The SGE of D. andersoni significantly down-regulated the expression of ICAM-1, whereas a similar extract prepared from I. scapularis significantly reduced the expression of P-selectin and VCAM-1. Tick salivary proteins therefore appear to have direct effects on adhesion-molecule expression, in addition to their previously established roles in down-regulating the pro-inflammatory cytokines that activate endothelial cells. It remains unclear exactly how the reduction of adhesion-molecule expression in the host's endothelial cells benefits the feeding tick but it may alter leucocyte migration to the bite site and/or reduce antigen presentation by the endothelial cells. It may also modulate the interactions between the host's leucocytes and any tick-borne pathogens, during initial infection of the endothelium.

Effect of salivary gland extract ofLeishmania vector,Lutzomyia longipalpis, on leukocyte migration in OVA-induced immune peritonitis

Salivary gland extracts (SGE) from Lutzomyia longipalpis potentate L. major infection by inducing a Th2 immune response. However, the effect of SGE on the effector phase of immune response is not known. Herein, we demonstrate that SGE inhibited neutrophil migration in ovalbumin (OVA)-induced peritonitis in immunized mice. SGE pretreatment of mice inhibited OVA-induced CD4+ and CD8+ T lymphocyte migration. The OVA-induced production of TNF-alpha, IL-1beta and leukotriene B4 (LTB4), neutrophil chemotactic mediators in this model, were inhibited by SGE. On the other hand, SGE enhanced production of IL-10 and IL-4. In naive mice, SGE also blocked LTB4-induced neutrophil migration, but not that induced fMLP. Moreover, co-incubation of LTB4 (but not fMLP, TNF-alpha and MIP-1alpha) with SGE inhibited the ability of LTB4 to induce neutrophil migration in vivo and in vitro. Altogether, the results suggest that SGE has anti-inflammatory properties that are associated with inhibition of TNF-alpha and LTB4 production and/or with the neutrophil chemotactic activity of LTB4. The effectiveness of SGE in inhibiting neutrophil migration and inflammatory mediators release in a Th1 immune inflammatory response model reinforces the need for isolation of the compounds responsible for these activities, which could be used as prototypes for the development new anti-inflammatory drugs.

Effect of tick salivary gland extract on the cytokine production by mouse epidermal cells

Previous studies have demonstrated that both tick saliva and Borrelia burgdorferi sensu lato antigens modulate the cytokine response of the host. In this paper, the effect of salivary gland extract (SGE) from Ixodes ricinus (L., 1758) ticks on cytokine production by primary cultures of mouse epidermal cells stimulated with Borrelia afzelii Canica, Nato, du Merle, Mazie, Baranton et Postic, 1993 spirochetes was analysed. Epidermal cells were derived from C3H/HeN mice, susceptible to Lyme disease, and BALB/c mice, which are resistant. In cultures from C3H/HeN mice, SGE down regulated production of tumour necrosis factor alpha (TNF-alpha) and up regulated Th2 cytokine, interleukin 4 (IL-4). Cultures from BALB/c mice produced higher basal levels of monitored cytokines, but their production was affected by SGE a different way. While Th2 cytokines IL-6 and IL-10 were down regulated, the effect on TNF-alpha and IL-4 was ambiguous. These results indicate that the effect of tick saliva on the epidermal cells of Lyme disease-susceptible C3H/HeN mice mirrors its effect on other cells of the immune system.

Manipulation of host cytokine network by ticks: a potential gateway for pathogen transmission

Ticks are obligatory blood-feeding arthropods that secrete various immunomodulatory molecules to antagonize host inflammatory and immune responses. Cytokines play an important role in regulating these responses. We investigated the extent to which ticks interact with the sophisticated cytokine network by comparing the effect of salivary gland extracts (SGE) of 3 ixodid tick species, Dermacentor reticulatus, Amblyomma variegatum and Ixodes ricinus, all of which are important vectors of tick-borne pathogens. Using specific ELISAs, anti-cytokine activity was demonstrated with 7 cytokines: IL-8, MCP-1, MIP-1alpha, RANTES, eotaxin, IL-2 and IL-4. The results varied between species, and between adult males and females of the same species. Relatively high activity levels were detected in saliva of female D. reticulatus, confirming that the observed anti-cytokine activities are an integral part of tick saliva secreted into the host. Results with fractionated SGE indicated that from 2 to 6 putative cytokine binding molecules are produced, depending on species and sex. Binding ability of SGE molecules was verified by cross-linking with radio-isotope labelled MIP-1alpha. By targeting different cytokines, ixodid ticks can manipulate the cytokine network, which will greatly facilitate blood-feeding and provide a gateway for tick-borne pathogens that helps explain why ticks are such efficient and effective disease vectors.

A comparison of the immunosuppressive effects of salivary gland extracts from two laboratory strains of Boophilus microplus

A comparison of the immunosuppressive effects of salivary gland extracts from two laboratory strains of Boophilus microplus

A comparison of the immunosuppressive effects of salivary gland extracts from two laboratory strains of Boophilus microplus

This study addresses three questions related to the immune response of cattle to tick salivary gland extracts. Firstly, is there a difference in the inhibition of proliferation of Concanavalin A (ConA) stimulated bovine lymphocytes induced by salivary gland extracts of the N and Y strains of Boophilus microplus? Second, is there a difference in the development rate of the Y and N tick strains? Third, does the host affect the inhibitory effect of salivary gland extract on the proliferation of ConA stimulated lymphocytes from the two tick strains? Salivary gland extract of the Y strain inhibited in vitro proliferation of lymphocytes stimulated by ConA significantly more than that of the N strain, when each strain was raised on different animals. A difference in the development rate was observed between the tick strains when raised on the same animal, with female ticks of the Y strain developing faster and reaching a greater fully engorged weight than ticks of the N strain. The difference in their rate of development did not appear to contribute to a difference in inhibitory effects of the salivary gland extracts and there was no difference between the inhibitory effects of salivary gland extracts from both strains. However, when Y strain ticks were raised on different animals, there was a significant difference in the inhibition of lymphocyte proliferation between the two salivary gland extracts. Therefore, it was concluded that there is no difference between the inhibitory effects of the two tick strains and that the host has an influence on salivary gland extract composition of B. microplus and its inhibitive properties.

Modulation of human lymphocyte proliferation by salivary gland extracts of ixodid ticks (Acari: Ixodidae): effect of feeding stage and sex

Ixodid ticks remain attached to their hosts for several days to weeks. During this extended feeding process new proteins involved in the modulation of host immune responses are expressed in tick salivary glands. In our study a stimulatory or inhibitory effect of salivary gland extracts (SGE) of unfed and partially fed female Ixodes ricinus (Linnaeus, 1758), female and male Amblyomma variegatum (Fabricius, 1794) and Rhipicephalus appendiculatus Neumann, 1901 ticks on human lymphocyte proliferation induced by Concanavalin A (ConA) and phytohaemagglutinin (PHA), respectively, was investigated. SGE of all female ticks examined suppressed proliferation of ConA-induced lymphocytes; highly significant suppression was observed in the presence of unfed I. ricinus and 9-day fed A. variegatum SGE. SGE of partially fed A. variegatum and I. ricinus females suppressed PHA responses of lymphocytes. Lymphocytes showed reduced PHA and ConA responses in the presence of SGE of unfed and 2-day fed R. appendiculatus females, while SGE of 6-day fed females enhanced PHA responses, but reduced their ConA responses; generally SGE of 2-day fed females displayed the strongest inhibition. Amblyomma variegatum male SGE slightly enhanced PHA, but significantly reduced ConA responses of lymphocytes and their inhibitory effect increased during feeding. SGE of unfed and 2-day fed R. appendiculatus males enhanced PHA and ConA responses and those of 6-day fed males suppressed lymphocyte proliferation. The results suggest that (i) species- and sex-specific differences exist in the effects of tick salivary gland antigens on human lymphocyte proliferation and (ii) effect of SGE on human lymphocyte responses to mitogens varies depending on the tick feeding status.

Modulation of bovine lymphocyte response by salivary gland extracts of the stable fly, Stomoxys calcitrans (Diptera: Muscidae).

The effect of salivary gland extract of the stable fly, Stomoxys calcitrans (L), on bovine lymphocyte proliferation was determined, and antibody reactivity to salivary gland proteins was characterized in cattle exposed to stable flies. Salivary glands were dissected from male and female flies (4-8 d after eclosion), and protein extracts were made by freeze-thaw cycles. Salivary gland extract (SGE, 1 and 5 microg) significantly inhibited mitogen-driven proliferation of bovine lymphocytes, compared with 1 and 5 microg of identically prepared midgut extract (ANOVA, P < 0.05). Phytohemagglutinin A (PHA) stimulated lymphocyte responses were suppressed by 61.7 and 79.5% (mean values) with 1 and 5 microg of SCE, whereas concanvalin A (Con A) stimulated responses were suppressed by 62.9 and 77.1% (1 and 5 microg). In contrast, midgut extract (1 and 5 microg) minimally suppressed PHA (12.7% +/- 12.6 and 18.7% +/- 15.5) and Con A-driven responses (13.8% +/- 20.5 and 24.6% +/- 14.9), respectively. Viability studies using propidium iodide and flow cytometry demonstrated that SGE was not cytotoxic. Two-color immunofluorescence studies identified T and B lymphocytes as the nonviable cells in the cultures. Western blot analysis of serum collected from five dairy cows during periods of low and high fly exposure identified an immunodominant 27 kDa protein among the salivary gland proteins. These results indicate that exposure of cattle to stable fly saliva during blood feeding results in an antibody response to salivary proteins and that the saliva has a potential to modulate T lymphocyte function.

Effect of salivary gland extracts from the tick, Boophilus microplus, on leucocytes from Brahman and Hereford cattle.

The effect of salivary gland extract (SGE) from Boophilus microplus on peripheral blood lymphocytes, neutrophils and monocytes from Brahman (Bos indicus) and Hereford (Bos taurus) cattle was investigated. SGE (8 micro g) significantly inhibited the proliferation response of lymphocytes to concanavalin A from both Brahman and Hereford cattle by 89% and 41%, respectively. The difference in inhibition between the two breeds was highly significant (P < 0.01), whilst at 1 micro g of SGE, significant inhibition of lymphocytes occurred only in Hereford cattle (34%). Flow cytometric analysis of monocytes and neutrophils showed that SGE (40 micro g) significantly reduced both the proportion of cells actively phagocytosing Escherichia coli labelled with fluorescein isothiocyanate (E. coli-FITC) and the uptake of E. coli-FITC in Brahman cattle. However, in Hereford cattle, a significant depression in uptake was only observed in neutrophils. The proportion of monocytes and neutrophils with oxidative activity was significantly suppressed in the presence of SGE in both breeds of cattle. These results indicate that peripheral blood leucocytes from different breeds of cattle respond differently to SGE.

Salivary gland extract from Ixodes ricinus ticks inhibits production of interferon-gamma by the upregulation of interleukin-10.

Tick saliva has been shown to modulate host immunity by a so far unknown mechanism. We have demonstrated an inverted effect of salivary gland extract (SGE), derived from partially fed Ixodes ricinus females, on the production of two cytokines, interferon (IFN)-gamma and interleukin (IL)-10, in vitro. While SGE markedly suppressed the elaboration of IFN-gamma by mouse splenocytes stimulated with lipopolysaccharide (LPS), the production of IL-10 was increased in comparison with SGE-untreated cultures. The suppressive effect of SGE could be abolished by the addition of an IL-10 neutralizing monoclonal antibody to splenocyte cultures. Similar results were obtained when live Borrelia afzelii spirochetes, which are transmitted in Europe by I. ricinus ticks, were used for the cytokine induction. These results suggest that tick saliva can upregulate the IL-10 production at the tick feeding site, which consecutively inhibits the elaboration of pro-inflammatory cytokines, for example IFN-gamma. This immunosuppression may facilitate the establishment of tick-transmitted pathogens in the host.

Effects of infestation by Rhipicephalus sanguineus on lymphocyte blastogenic responses to mitogens in dogs.

Mitogen blastogenic responses of lymphocytes from dogs infested with adult Rhipicephalus sanguineus and the effects of salivary gland extracts (SGE) of the tick on the blastogenic responses of lymphocytes from normal dogs were studied. Infestation by R. sanguineus significantly suppressed concanavalin A, phyto-hemagglutinin and pokeweed mitogen responses of lymphocytes from dogs. The inhibition of lymphocyte responses of dogs in the first infestation was greater than that in the second infestation. SGE from R. sanguineus also suppressed all mitogen blastogenic responses of lymphocytes from healthy dogs in vitro. These suppressive effects of SGE on the blastogenic responses of PBL to mitogens were significantly inhibited by trypsin digestion. It is suggested that some proteins in SGE contribute to the suppressive effects of SGE on the blastogenic responses of peripheral blood lymphocytes from dogs.