Galectin-receptor interaction: a key player in liver fibrosis induced by Schistosoma japonicum infection.

BACKGROUND: Schistosoma japonicum eggs lodge in the liver and induce a fibrotic granulomatous immune response in the liver of host. Galectin 3 (Gal-3) is a protein implicated in fibrosis in multiple organs. However, the pathology and molecular mechanisms promoting hepatic granuloma formation remain poorly understood. METHODS: To investigate the effect of blocking galectin-receptor interactions by α-lactose on liver immunopathology in mice with S. japonicum infection, C57BL/6 mice were infected with S. japonicum and alpha (α)-lactose was intraperitoneally injected to block the interactions of galectins and their receptors. RESULTS: Compared with S. japonicum-infected mice, there were significantly decreased Gal-3 mRNA and protein expression levels, decreased intensity of Gal-3 fluorescence in the liver, decreased serum ALT and AST levels, decreased egg numbers of S. japonicum in the liver section, attenuated hepatic and spleen pathology, and alleviated liver fibrosis accompanied with decreased protein expression levels of fibrosis markers [α-smooth muscle actin (α-SMA), collagen I, and collagen IV] in the liver of S. japonicum-infected mice blocked galectin-receptor interactions with hematoxylin-eosin staining, Masson's trichrome staining, immunohistochemistry, or Western blot analysis. Compared with S. japonicum-infected mice, blocking galectin-receptor interactions led to increased eosinophil infiltration and higher eosinophil cationic protein (ECP) expression in the liver, accompanied by increased mRNA levels of eosinophil granule proteins [ECP and eosinophil peroxidase (EPO)], IL-5, CCL11, and CCR3 in the liver and decreased mRNA levels of Gal-3 and M2 macrophage cytokines (TGF-ß, IL-10, and IL-4) in the liver and spleen by using quantitative real-time reverse transcription-polymerase chain reaction. In addition, there were increased Beclin1 protein expression and protein expression ratio of LC3B-II/LC3B-I and decreased p62 protein expression and protein expression ratios of phospho-mTOR/mTOR and phospho-AKT/AKT by Western blot; increased double-labeled F4/80+/LC3B+ cells by immunofluorescence staining; increased M1 macrophage polarization in the liver of S. japonicum-infected mice blocked galectin-receptor interactions by flow cytometric analysis and immunofluorescence staining. CONCLUSIONS: Our data found that blockage of galectin-receptor interactions downregulated Gal-3, which in turn led to reduced liver functional damage, elevated liver eosinophil recruitment, promoted macrophage autophagy through the Akt/mTOR signaling pathway, and alleviated liver pathology and fibrosis. Therefore, Gal-3 plays a pivotal role during S. japonicum infection and could be a target of pharmacologic potential for liver fibrosis induced by S. japonicum infection.

Impact of Galectin-Receptor Interactions on Liver Pathology During the Erythrocytic Stage of Plasmodium berghei Malaria.

Hepatopathy is frequently observed in patients with severe malaria but its pathogenesis remains unclear. Galectins are evolutionarily conserved glycan-binding proteins with pleiotropic roles in innate and adaptive immune responses, and exhibit pivotal roles during Plasmodium spp. infection. Here, we analyzed the impact of blockage of galectin-receptor interactions by treatment with alpha (α)-lactose on liver immunopathology during the erythrocytic stage of malaria in mice infected with Plasmodium berghei ANKA (PbANKA). Our results found that compared with PbANKA-infected mice (malarial mice), blockage of galectin-receptor interactions led to decreased host survival rate and increased peripheral blood parasitemia; exacerbated liver pathology, increased numbers of CD68+ macrophages and apoptotic cells, and increased parasite burden in the livers on days 5 and 7 post infection (p.i.) as well as increased mRNA expression levels of galectin-9 (Gal-9) and its receptor, the T cell immunoglobulin domain and mucin domain protein 3 (Tim-3), interferon (IFN)α, IFNγ, and the triggering receptor expressed on myeloid cells (TREM)-1 in the livers or spleens of PbANKA-infected mice on day 7 p.i. Observed by transmission electron microscopy, the peritoneal macrophages isolated from malarial mice with α-lactose treatment had more pseudopodia than those from malarial mice. Measured by using quantitative real-time reverse transcription-polymerase chain reaction assay, the mRNA expression levels of Gal-9, IFNα, IFNß, IFNγ, and TREM-1 were increased in the peritoneal macrophages isolated from malarial mice with α-lactose treatment in comparison of those from malarial mice. Furthermore, significant positive correlations existed between the mRNA levels of Gal-9 and Tim-3/IFNγ/TREM-1 in both the livers and the peritoneal macrophages, and between Gal-9 and Tim-3/TREM-1 in the spleens of malarial mice; significant positive correlations existed between the mRNA levels of Gal-9 and IFNγ in the livers and between Gal-9 and IFNα in the peritoneal macrophages from malarial mice treated with α-lactose. Our data suggest a potential role of galectin-receptor interactions in limiting liver inflammatory response and parasite proliferation by down-regulating the expressions of IFNα, IFNγ, and TREM-1 during PbANKA infection.

Galectin-Receptor Interactions Regulates Cardiac Pathology Caused by Trichinella spiralis Infection.

The parasitic nematode Trichinella spiralis causes trichinellosis, a serious food-borne parasitic zoonosis worldwide. Infection with T. spiralis may also cause myocarditis. In the present study, we used mouse models to assess the impact of blockage of galectin-receptor interactions by α-lactose on cardiac immunopathology during acute T. spiralis experimental infection. Our data demonstrated that, after T. spiralis infection, blockage of galectin-receptor interactions resulted in cardiac dysfunction detected by transthoracic conventional echocardiography, and increased serum Gal-3 level, a biomarker of myocardial damage. In addition, there were increased eosinophil number in peripheral blood, and increased eosinophil infiltration in the heart and spleen tissues accompanied with increased mRNA levels of eosinophil granule proteins (including eosinophil cationic protein (ECP) and eosinophil peroxidase (EPO)) and IL-5 in these organs; increased cardiac fibrosis accompanied with increased Gal-3 and collagen 1 expressions in the hearts of mice with blockage of galectin-receptor interactions after T. spiralis infection. Correlation analysis showed that significant positive correlations existed between the mRNA levels of Gal-3 and ECP/EPO/eosinophil major basic protein/IL-5/CCL11/CCR3/α-SMA/collagen 1 in the hearts of both T. spiralis-infected mice and T. spiralis-infected mice with blockage of galectin-receptor interactions. Our data suggest that galectin-receptor interactions play a pivotal role during acute T. spiralis infection, and lack of galectin-receptor interactions upregulates Gal-3 which, in turn, leads to elevated heart eosinophil recruitment, exacerbated heart pathology and fibrosis, and heart functional damage.

Trichinella spiralis co-infection exacerbates Plasmodium berghei malaria-induced hepatopathy.

BACKGROUND: Although Plasmodium parasites and intestinal helminths share common endemic areas, the mechanisms of these co-infections on the host immune response remain not fully understood. Liver involvement in severe Plasmodium falciparum infections is a significant cause of morbidity and mortality. However, the effect of pre-existing Trichinella spiralis infection on the immune response and liver immune-pathogenesis in P. berghei ANKA (PbANKA)-infected mice needs to be elucidated. METHODS: Outbred Kunming mice were infected with T. spiralis and 9 days later were challenged with P. berghei ANKA (PbANKA), and the investigation occurred at 13 days after co-infection. RESULTS: Compared with PbANKA-mono-infected mice, T. spiralis + PbANKA-co-infected mice had similar survival rate but lower PbANKA parasitaemia; however, there were more severe hepatosplenomegaly, increased liver and spleen indexes, and increased liver pathology observed by hematoxylin and eosin staining; higher expression levels of galectin (Gal)-1, Gal-3, CD68+ macrophages, and elastase-positive neutrophils measured by immunohistochemical staining; upregulated mRNA expression levels of Gal-1, Gal-3, cytokines (interferon-gamma (IFNγ) and interleukin (IL)-6), and M1 macrophage polarization marker (inducible nitric oxide synthase (iNOS)) in the liver, and increased expression levels of Gal-1, IFNγ, IL-6, eosinophil cationic protein, eosinophil protein X, and M1 (IL-1ß and iNOS) and M2 (Ym1) macrophage polarization markers in the spleen of co-infected mice detected by using quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). In vitro study showed that compared with PbANKA-mono-infected mice, there were significantly increased expression levels of Gal-1, Gal-3, IL-6, IL-1ß, and iNOS in the peritoneal macrophage isolated from co-infected mice detected by using qRT-PCR. Correlation analysis revealed significant positive correlations between Gal-3 and IL-1ß in the peritoneal macrophages isolated from PbANKA-mono-infected mice, between Gal-3 and IFNγ in the spleen of co-infected mice, and between Gal-1 and Ym1 in the peritoneal macrophages isolated from co-infected mice. CONCLUSIONS: Our data indicate that pre-existing infection of T. spiralis may suppress P. berghei parasitaemia and aggravate malaria-induced liver pathology through stimulating Gal-1 and Gal-3 expression, activating macrophages, neutrophils, and eosinophils, and promoting mediator release and cytokine production.

Galectins, Eosinophiles, and Macrophages May Contribute to Schistosoma japonicum Egg-Induced Immunopathology in a Mouse Model.

Schistosomiasis is a severe public health problem, which can cause tissue fibrosis and can even be fatal. Previous studies have proven that galectins and different kinds of cells involve in the regulation of tissue fibrosis process. In this study, outbred Kunming mice were infected with Schistosoma japonicum (S. japonicum). Our results showed that compared with uninfected mice, there were severe egg granulomatous inflammation and tissue fibrosis in the livers, spleens, and large intestines of S. japonicum-infected mice at 8 weeks post-infection (p.i.), and the number of eosinophils by hematoxylin and eosin staining and CD68 macrophage-positive area by immunohistochemical staining were significantly increased. Detected by using quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR), at 8 weeks after S. japonicum infection, the mRNA expression levels of galectin (Gal)-1, Gal-3, CD69, eosinophil protein X (EPX), and chitinase 3-like protein 3 (Ym1) were significantly increased in liver, spleen, and large intestine; eotaxin-1 (CCL11) and eosinophil cationic protein were significantly increased in both liver and spleen; eotaxin-2 (CCL24) and Arginase1 (Arg1) were significantly increased in both spleen and large intestine; and CD200R was significantly increased in both liver and large intestine. However, interleukin (IL)-1ß and inducible nitric oxide synthase (iNOS) were only significantly increased in liver. The M2/M1 ratio of CD200R/CD86 genes was significantly increased in liver, and ratios of Ym1/IL-1ß and Ym1/iNOS were significantly increased in liver, spleen, and large intestine of S. japonicum-infected mice. Ex vivo study further confirmed that the levels of Gal-1, Gal-3, CD200R, Arg1, and Ym1 were significantly increased, and the ratios of CD200R/CD86 and Ym1/IL-1ß were significantly increased in peritoneal macrophages isolated from S. japonicum-infected mice at 8 weeks p.i. In addition, correlation analysis showed that significant positive correlations existed between mRNA levels of Gal-1/Gal-3 and EPX in liver, between Gal-3 and Ym1 in both liver and large intestine, and between Gal-3 and CD200R in peritoneal macrophages of S. japonicum-infected mice. Our data suggested that Gal-1, Gal-3, eosinophils, and macrophages are likely involved in the development of egg granulomatous response and fibrosis induced by S. japonicum infection.

Association of TREM-1, IL-1ß, IL-33/ST2, and TLR Expressions With the Pathogenesis of Ocular Toxoplasmosis in Mouse Models on Different Genetic Backgrounds.

Ocular toxoplasmosis (OT) is one of the most common causes of posterior uveitis. The signaling of triggering receptor expressed on myeloid cells (TREM)-1 amplifies inflammation, whereas TREM-2 signaling is anti-inflammatory. IL-1ß is a major driver of inflammation during infection. Toll-like receptors (TLRs) play important roles in protective immune response during Toxoplasma gondii infection, and interleukin (IL)-33 receptor (T1/ST2) signaling prevents toxoplasmic encephalitis in mice. However, the pathogenic mechanisms of OT are not yet well elucidated. To investigate the role of TREM-1, TREM-2, IL-1ß, IL-33/ST2, and TLRs in OT of susceptible C57BL/6 (B6) and resistant BALB/c mice, both strains of mice were intravitreally infected with 500 tachyzoites of the RH strain of T. gondii. Histopathological analysis showed that T. gondii-infected B6 mice had more severe ocular damage observed by light microscopy, higher number of neutrophil elastase-positive cells in the eyes detected by immunohistochemical staining, more T. gondii tachyzoites in the eyes observed by transmission electron microscopy, and higher mRNA expression levels of tachyzoite-specific surface antigen 1 detected by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR) in comparison of T. gondii-infected BALB/c mice. Detected by using qRT-PCR, the mRNA expression levels of TREM-1, IL-1ß, IL-33, ST2, TLR11, TLR12, and TLR13 were significantly higher in the eyes of T. gondii-infected B6 mice than those of T. gondii-infected BALB/c mice, whereas the mRNA expression levels of TLR3 and TLR9 were significantly higher in the eyes of T. gondii-infected BALB/c mice than those of T. gondii-infected B6 mice. Correlation analysis showed that significant positive correlations existed between TREM-1 and IL-1ß/IL-33/ST2/TLR9/TLR11 in the eyes of B6 mice and existed between TREM-1 and IL-33/ST2/TLR3/TLR9/TLR13 in the eyes of BALB/c mice after ocular T. gondii infection. Our data revealed that, compared with T. gondii-resistant BALB/c mice, ocular T. gondii infection can stimulate higher production of TREM-1, IL-33, ST2, TLR11, TLR12, and TLR13 in the eyes of T. gondii-susceptible B6 mice, however, whether those lead to more severe ocular pathology in the susceptible B6 mice remain to be further studied.

Activation of Mast Cells Promote Plasmodium berghei ANKA Infection in Murine Model.

Malaria, a mosquito-borne infectious disease, is a severe health problem worldwide. As reported, some anti-malarial drugs with anti-parasitic properties also block mast cells (MCs) activities. It is hypothesized that MCs activity may be correlated with the pathogenesis of malaria. Thus, the role of MCs on malarial pathogenesis and the involved physiological action and pathways need to be further investigated. This study aimed to investigate the effect of MCs activation on malaria disease severity using KunMing mice with Plasmodium berghei ANKA (PbANKA) infection treated with MCs degranulator (compound 48/80, C48/80) or MCs stabilizer (disodium cromoglycate, DSCG). PbANKA infection caused a dramatic increase in MCs density and level of MCs degranulation in cervical lymph node (CLN) and skin. Compared with infected control, C48/80 treatment had shortened survival time, increased parasitemia, exacerbated liver inflammation and CLN hyperplasia, accompanied with increase in vascular leakage and leukocyte number. The infected mice with C48/80 treatment also elevated the release of CCL2, CXCL1, and MMP-9 from MCs in CLN and skin, and TNF-α, IFN-γ, CCR2, and CXCR2 mRNA expression in CLN and liver. In contrast, the infected mice treated with DSCG showed longer survival time, lower parasitemia, improved liver inflammation and CLN hyperplasia, followed by a decline of vascular leakage and leukocyte number. Decreased MCs-derived CCL2, CXCL1, and MMP-9 from CLN and skin, mRNA expression in CLN and liver (TNF-α, IFN-γ, CCR2, and CXCR2) were also observed in infected mice with DSCG treatment. Our data indicated that MCs activation may facilitate the pathogenesis of PbANKA infection.

Galectin-3 and Galectin-9 May Differently Regulate the Expressions of Microglial M1/M2 Markers and T Helper 1/Th2 Cytokines in the Brains of Genetically Susceptible C57BL/6 and Resistant BALB/c Mice Following Peroral Infection With Toxoplasma gondii.

Toxoplasmic encephalitis (TE), an opportunistic infection, is a severe health problem in immunocompromised patients. Previous studies have revealed that C57BL/6 mice are susceptible and BALB/c mice are resistant to TE. To investigate the mechanisms involved in the immunopathogenesis of TE in susceptible C57BL/6 and resistant BALB/c mice, both strains of mice were perorally infected with the Prugniuad (Pru) strain of Toxoplasma gondii. Our results showed that compared with BALB/c mice, C57BL/6 mice infected with T. gondii Pru strain had more severe brain histopathological damage, and higher mRNA expression levels of tachyzoite-specific surface antigen 1, bradyzoite-specific antigen 1, interferon gamma (IFNγ), interleukin (IL)-10, arginase1 (Arg1) (M2 marker), galectin (Gal)-3, Gal-9, T. gondii microneme protein 1 (TgMIC1), TgMIC4, and TgMIC6 during the course of infection by using quantitative real-time reverse transcription-polymerase chain reaction. Further analysis displayed that BALB/c mice showed higher numbers of microglial cells and higher levels of IL-1ß, inducible nitric oxide synthase (iNOS) (M1 marker), and chitinase-3-like protein 3 (Ym1) (M2 marker) in the early infective stage [at day 14 or 35 post infection (p.i.)] compared with C57BL/6 mice, whereas C57BL/6 mice showed higher numbers of microglial cells and higher levels of IL-10, iNOS (M1 marker), and Ym1 (M2 marker) at days 35, 50, or 70 p.i. compared with BALB/c mice. Correlation analysis showed that significant positive correlations existed between Gal-3 and IL-4/IL-10/iNOS/Ym1 and between Gal-9 and IL-4/Ym1 in C57BL/6 mice; between Gal-3 and IFNγ/Arg1 and between Gal-9 and IFNγ/Arg1 in BALB/c mice. Together, our data demonstrated that different Gal-3 and Gal-9 expressions as well as different positive correlations were found between Gal-3 and T helper 1 (Th1)/Th2/M1/M2 cytokines or between Gal-9 and Th1/Th2/M2 cytokines in the brains of T. gondii Pru strain-infected C57BL/6 and BALB/c mice.

Large-scale Artemisinin-Piperaquine Mass Drug Administration With or Without Primaquine Dramatically Reduces Malaria in a Highly Endemic Region of Africa.

Background: Mass drug administration (MDA), with or without low-dose primaquine (PMQLD), is being considered for malaria elimination programs. The potential of PMQLD to block malaria transmission by mosquitoes must be balanced against liabilities of its use. Methods: Artemisinin-piperaquine (AP), with or without PMQLD, was administered in 3 monthly rounds across Anjouan Island, Union of Comoros. Plasmodium falciparum malaria rates, mortality, parasitemias, adverse events, and PfK13 Kelch-propeller gene polymorphisms were evaluated. Results: Coverage of 85 to 93% of the Anjouan population was achieved with AP plus PMQLD (AP+PMQLD) in 2 districts (population 97164) and with AP alone in 5 districts (224471). Between the months of April-September in both 2012 and 2013, average monthly malaria hospital rates per 100000 people fell from 310.8 to 2.06 in the AP+PMQLD population (ratio 2.06/310.8 = 0.66%; 95% CI: 0.02%, 3.62%; P = .00007) and from 412.1 to 2.60 in the AP population (ratio 0.63%; 95% CI: 0.11%, 1.93%; P < .00001). Effectiveness of AP+PMQLD was 0.9908 (95% CI: 0.9053, 0.9991), while effectiveness of AP alone was 0.9913 (95% CI: 0.9657, 0.9978). Both regimens were well tolerated, without severe adverse events. Analysis of 52 malaria samples after MDA showed no evidence for selection of PfK13 Kelch-propeller mutations. Conclusions: Steep reductions of malaria cases were achieved by 3 monthly rounds of either AP+PMQLD or AP alone, suggesting potential for highly successful MDA without PMQLD in epidemiological settings such as those on Anjouan. A major challenge is to sustain and expand the public health benefits of malaria reductions by MDA.

Increased IL-27/IL-27R expression in association with the immunopathology of murine ocular toxoplasmosis.

Interleukin 27 (IL-27) is a member of the IL-6/IL-12 family, and IL-27 receptor (IL-27R) consists of WSX-1 (the IL-27Rα subunit) and the signal-transducing subunit gp130. Human and mouse mast cells (MCs) express the IL-27R. To explore the expressions of IL-27/IL-27R subunits (WSX-1 and gp130) during acute ocular toxoplasmosis (OT), we established mouse model by intraocular injection of 500 Toxoplasma gondii RH strain tachyzoites. Histopathological changes were analyzed, MCs were counted by toluidine blue staining, and tryptase+/IL-27+ MCs were examined by immunofluorescence double-staining in the eyes and cervical lymph nodes (CLNs) of T. gondii-infected mice. The mRNA expressions of IL-27p28, WSX-1, gp130, and tachyzoite specific surface antigen 1 (SAG1) in the eyes and CLNs of T. gondii-infected mice, and the expressions of WSX-1 and gp130 in the murine mastocytoma cell line P815 infected with T. gondii tachyzoites in vitro were examined by using quantitative real-time reverse transcription-polymerase chain reaction. Our results showed that, after T. gondii infection, severe histopathological changes, increased numbers of total MCs and degranulated MCs, elevated expressions of IL-27p28, WSX-1, and gp130 were found in the eyes and CLNs, and significant correlations between the levels of IL-27 and SAG1 existed in the eyes and CLNs of T. gondii-infected mice. In addition, increased levels of WSX-1 and gp130 were examined in T. gondii-infected P815 cells. Our data suggested that IL-27/IL-27R expression induced by T. gondii infection may regulate MC-mediated immune response during acute OT in mouse model.

Expression of Toll­like receptor 4 on mast cells in gingival tissues of human chronic periodontitis.

The purpose of the present study was to examine the expression of Toll­like receptor 4 (TLR4) on mast cells in gingival tissues of human chronic periodontitis. A total of 68 donors, including 23 with mild chronic periodontitis, 25 with advanced chronic periodontitis and 20 healthy controls, were included in the present study. Gingival specimens from the donors were fixed in 4% neutral formalin, stained with hematoxylin and eosin for histologic observation, stained for immunohistochemical identification of TLR4 in gingival tissues, and stained with double immunofluorescence for the identification of TLR4 on mast cells in gingival tissues. The results revealed that the expression of TLR4 in the gingival tissues and on mast cells in the gingival tissues of patients with chronic periodontitis were significantly higher, compared with those of the normal control group (P<0.05). The expression levels of TLR4 in the gingival tissues and on mast cells in patients with advanced chronic periodontitis were significantly higher, compared with those in patients with mild chronic periodontitis (P<0.05). In conclusion, the expression of TLR4 in gingival tissues and on mast cells increased with the severity of chronic periodontitis, suggesting that TLR4, particularly mast cell TLR4, may be important in the disease process of human chronic periodontitis.

Disodium cromoglycate may act as a novel adjuvant for UV-attenuated Toxoplasma gondii vaccine in mouse model.

We have proven the beneficial effects during acute Toxoplasma gondii infection when mast cells were inhibited by disodium cromoglycate (DSCG). Here we investigated the adjuvant effect of DSCG on the protective efficacy of UV-attenuated T. gondii (UV-Tg) vaccine. Mice were infected with 102Tg alone or infected with 102Tg plus DSCG (Tg + DSCG), immunized with 105 UV-Tg and challenged with 102Tg (UV-Tg + Tg) or immunized with 105 UV-Tg plus DSCG and challenged with 102Tg (UV-Tg + DSCG + Tg). Compared to Tg group, Tg + DSCG, UV-Tg + Tg, and UV-Tg + DSCG + Tg showed significantly prolonged survival times, decreased parasite burdens, reduced liver histopathologies, and increased levels of Th1 and Th2 cytokines and IL-17 in the livers and spleens by using quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). Compared to UV-Tg + Tg, UV-Tg + DSCG + Tg had significantly longer survival time, lower tissue parasite burden and histopathological score, and higher levels of Th1 and Th2 cytokines and IL-17 in the livers or spleens. Our data suggest that DSCG may play an adjuvant role in the immunization induced by UV-attenuated T. gondii in mice, by promoting cellular immune response against T. gondii challenge.

Temporal changes in genetic diversity of msp-1, msp-2, and msp-3 in Plasmodium falciparum isolates from Grande Comore Island after introduction of ACT.

BACKGROUND: Malaria is still one of the serious public health problems in Grande Comore Island, although the number of annual cases has been greatly reduced in recent years. A better understanding of malaria parasite population diversity and transmission dynamics is critical for assessing the effectiveness of malaria control measures. The objective of this study is to investigate temporal changes in genetic diversity of Plasmodium falciparum populations and multiplicity of infection (MOI) in Grande Comore 10 years after introduction of ACT. METHODS: A total of 232 P. falciparum clinical isolates were collected from the Grande Comore Island during two sampling periods (118 for 2006‒2007 group, and 114 for 2013‒2016 group). Parasite isolates were characterized for genetic diversity and complexity of infection by genotyping polymorphic regions in merozoite surface protein gene 1 (msp-1), msp-2, and msp-3 using nested PCR and DNA sequencing. RESULTS: Three msp-1 alleles (K1, MAD20, and RO33), two msp-2 alleles (FC27 and 3D7), and two msp-3 alleles (K1 and 3D7) were detected in parasites of both sampling periods. The RO33 allele of msp-1 (84.8%), 3D7 allele of msp-2 (90.8%), and K1 allele of msp-3 (66.7%) were the predominant allelic types in isolates from 2006-2007 group. In contrast, the RO33 allele of msp-1 (63.4%), FC27 allele of msp-2 (91.1%), and 3D7 allele of msp-3 (53.5%) were the most prevalent among isolates from the 2013-2016 group. Compared with the 2006‒2007 group, polyclonal infection rates of msp-1 (from 76.7 to 29.1%, P < 0.01) and msp-2 (from 62.4 to 28.3%, P < 0.01) allelic types were significantly decreased in those from 2013‒2016 group. Similarly, the MOIs for both msp-1 and msp-2 were higher in P. falciparum isolates in the 2006-2007 group than those in 2013-2016 group (MOI = 3.11 vs 1.63 for msp-1; MOI = 2.75 vs 1.35 for msp-2). DNA sequencing analyses also revealed reduced numbers of distinct sequence variants in the three genes from 2006‒2007 to 2013‒2016: msp-1, from 32 to 23 (about 28% decline); msp-2 from 29 to 21 (about 28% decline), and msp-3 from 11 to 3 (about 72% decline). CONCLUSIONS: The present data showed dramatic reduction in genetic diversity and MOI among Grande Comore P. falciparum populations over the course of the study, suggesting a trend of decreasing malaria transmission intensity and genetic diversity in Grande Comore Island. These data provide valuable information for surveillance of P. falciparum infection and for assessing the appropriateness of the current malarial control strategies in the endemic area.

Tryptase and TIM-1 double-positive mast cells in different stages of human chronic periodontitis.

BACKGROUND/PURPOSE: Mast cells (MCs) play a critical role in the pathogenesis of allergic reactions and inflammatory conditions through the release of inflammatory mediators. T cell immunoglobulin mucin domain (TIM-1) has been reported to express in MCs. The aim of the present study was to examine the expression and analyze the quantification of TIM-1 on tryptase-positive MCs in different stages of human chronic periodontitis using double-immunofluorescence staining. MATERIALS AND METHODS: Individuals who participated in this study were divided into three groups: healthy control gingivae (n = 27), chronic slight periodontitis (n = 34), and chronic severe periodontitis (n = 31). Their gingival specimens were taken and fixed in 10% buffered formalin, stained with hematoxylin and eosin (HE) for histopathology, and stained with double-immunofluorescence (DIF) for identification of tryptase-TIM-1 double-positive MCs in gingival tissues. RESULTS: Compared with healthy controls, the densities (cells/mm2) of tryptase-TIM-1 double-positive MCs were significantly increased in both the chronic slight periodontitis (P < 0.05) and severe periodontitis groups (P < 0.01). However, compared with the chronic slight periodontitis group, both the score of gingival tissue inflammation and the density of tryptase-TIM-1 double-positive MCs in gingival tissue were significantly increased in the severe periodontitis groups (P < 0.05). Conclusion: By incorporating HE with double-immunofluorescence staining in human chronic periodontitis, the significantly increased number of tryptase-TIM-1 double-positive MCs had the similar tendency as the severity of periodontitis inflammation. Based on our results, we suggest that tryptase-TIM-1 double-positive MCs may play an important role in human chronic periodontitis.

Expression of matrix metalloproteinase-8 and matrix metalloproteinase-13 in mast cells of human periapical lesions.

OBJECTIVE: This study aimed to detect the expression of matrix metalloproteinase-8 (MMP-8) and MMP-13 in mast cells (MCs) of human periapical lesions and to discuss the pathogenic role of MCs in periapical lesions. METHODS: Ninety samples were divided into three groups: (1) periapical granuloma group (n=30); (2) periapical cyst group (n=30); (3) normal periodontal membrane group (n=30). The samples were fixed in 10% neutral formalin for over 48 h and made into serial sections. After H&E staining, histological changes were observed under the optical microscope. Moreover, double immunofluorescence (DIF) staining was performed to detect expression of MMP-8 and MMP-13 in MCs of periapical lesions under the fluorescence microscope. RESULTS: Compared with the normal control group, the number of MMP-8 and MMP-13 double positive MCs in the periapical lesions increased significantly (P<0.01). There was no significant difference in the density of MMP-8 and MMP-13 double positive MCs in the periapical cyst group and periapical granuloma group (P>0.05). CONCLUSION: The number of MCs increased significantly in periapical lesions and there was a considerable increase in the density of MMP-8 and MMP-13 double positive MCs. These results indicate that MCs positive for MMP-8 and MMP-13 might contribute to the pathogenesis of chronic apical periodontitis.

Mast cell activator compound 48/40 is not an effective adjuvant for UV-attenuated Toxoplasma gondii vaccine.

Toxoplasma gondii (T. gondii, Tg) is a globally distributed parasitic protozoan causing different forms of toxoplasmosis in humans. Mast cells (MCs) play a role during T. gondii infection. Several studies suggest that MC activator compound 48/80 (C48/80) may be an effective vaccine adjuvant resulting in a potent and protective antigen-specific immune response against bacteria or virus infections. The present study was performed to determine whether C48/80 had adjuvant activity for ultraviolet (UV)-attenuated T. gondii vaccine to induce protective immune responses against T. gondii in mouse model. Kunming mice were divided into the following groups: naive mice, naive mice administrated with C48/80 intraperitoneal (i.p.) injection, mice infected by i.p. injection of 104 T. gondii RH strain alone (Tg group), mice infected with 104 RH tachyzoites plus C48/80 administration (Tg + C48/80), mice immunized with UV-Tg alone, and mice immunized with UV-Tg plus C48/80 administration (UV-Tg + C48/80). All the vaccinated mice were challenged with 104 tachyzoites of T. gondii RH strain at the same time as the primary infection. The survival rates, liver histopathologies, liver parasite burdens, and mRNA expression levels of Th1 and Th2 cytokines in the livers and spleens detected by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR) were compared among the aforementioned groups after primary infection or challenge infection. The results showed that, compared to the Tg group or Tg + C48/80 group, the UV-Tg + Tg group and UV-Tg + C48/80 + Tg group had significantly prolonged survival time, lower liver histopathological scores, decreased liver parasite burdens, and increased levels of Th1 and Th2 cytokines in the livers and spleens. There was no significant difference of survival time between the UV-Tg + Tg group and the UV-Tg + C48/80 + Tg group; however, the UV-Tg + C48/80 + Tg group showed higher parasite burden, more severe liver histopathology, and decreased IL-4 level compared to the UV-Tg + Tg group. These results indicate that C48/80 had no adjuvant activity for the immunization induced by UV-attenuated T. gondii vaccine.

The Roles of Mast Cells in Parasitic Protozoan Infections.

Protozoan parasites such as Plasmodium spp., Leishmania spp., Trypanosoma spp., and Toxoplasma gondii are major causes of parasitic diseases in both humans and animals. The immune system plays a critical role against protozoa, but their immune mechanism remains poorly understood. This highlights the need to investigate the function of immune cells involved in the process of parasite infections and the responses of host immune system to parasite infections. Mast cells (MCs) are known to be central players in allergy and anaphylaxis, and it has been demonstrated that MCs have crucial roles in host defense against a number of different pathogens, including parasites. To date, there are many studies that have examined the interaction of helminth-derived antigens and MCs. As one of the major effector cells, MCs also play an important role in the immune response against some parasitic protozoa, but their role in protozoan infections is, however, less well characterized. Herein, we review the current knowledge about the roles of MCs and their mediators during infections involving highly pathogenic protozoa including Plasmodium spp., Leishmania spp., Trypanosoma spp., and T. gondii. We offer a general review of the data from patients and experimental animal models infected with the aforementioned protozoa, which correlate MCs and MC-derived mediators with exacerbated inflammation and disease progression as well as protection against the parasitic infections in different circumstances. This review updates our current understanding of the roles of MCs during parasitic protozoan infections, and the participation of MCs in parasitic protozoan infections could be of a potential therapeutic target.

Immunomodulatory activity of interleukin-27 in human chronic periapical diseases.

This study aims to observe expression of IL-27 on different cells in periapical tissues of different types of human chronic periapical diseases. Periapical tissue specimens of 60 donors, including healthy control (n=20), periapical granuloma group (n=20) and radicular cysts group (n=20), were fixed in 10% buffered formalin, stained with hematoxylin and eosin for histopathology. Then specimens were stained with double- immuno-fluorescence assay for identification of IL-27-tryptase (mast cells, MCs), IL-27-CD14 (mononuclear phagocyte cells, MPs) and IL-27-CD31 (endothelial cells, ECs) double-positive cells in periapical tissues. The results indicated that compared with healthy control, the densities (cells/mm2) of IL-27-tryptase, IL-27-CD14 and IL-27-CD31 double-positive cells were significantly increased in human chronic periapical diseases (periapical granuloma group and radicular cysts group) (P<0.001). The density of IL-27-tryptase double positive cells in radicular cysts group was significantly higher than those in periapical granuloma group (P<0.001). Densities of IL-27-CD14 and IL-27-CD31 double-positive cells in periapical granuloma group had no significant difference with those in radicular cysts group (P=0.170 and 0.138, respectively). IL-27-CD14 double positive cells density achieved to peak among three cell groups in radicular cysts groups. In conclusion, IL-27 expressed in MCs, MPs and ECs of human chronic periapical diseases with different degrees. IL-27-tryptase double-positive cells may participate in pathogenic mechanism of chronic periapical diseases, especially for formation of fibrous in periapical cysts. IL-27-CD14 and IL-27-CD31 double-positive cells may participate in immunologic response to resist periapical infection, and they may play an dual role in pathogenesis and localization of periapical diseases.

Expression of transforming growth factor-ß in mast cells in human chronic periapical diseases.

This study was undertaken to investigate the distribution of mast cells (MCs) and the expression of transforming growth factor-ß (TGF-ß) on tryptase positive MCs in different types of human periapical diseases. Periapical tissues of 78 participates were used in this study, including healthy control (n=28), periapical cyst (n=25), and periapical granuloma (n=25). The tissue samples were fixed in 10% formalin for at least 48 h, followed by staining with hematoxylin and eosin (HE) for histopathological examination. Then, they were stained with toluidine blue for MCs and MCs degranulation examination, or stained with double immunofluorescence for identification of tryptase-TGF-ß double positive MCs. The results showed that the density of tryptase-TGF-ß double positive MCs in periapical lesions was significantly higher than that of healthy control (P<0.01). The number of TGF-ß positive MCs in the periapical cyst was potently higher than that in the periapical granuloma (P<0.01). In addition, compared with toluidine blue staining, the number of MCs with double immunofluorescence staining was significantly increased (P<0.01). The TGF-ß positive MCs may play an important role in the pathogenesis of human chronic periapical diseases, particularly in the formation of fibrous tissues in periapical cyst. Double immunofluorescence staining is more sensitive than the traditional toluidine blue staining for identifying MCs.

Up-regulated TLR2 and TLR4 expressions in liver and spleen during acute murine T. gondii infection.

Toll-like receptors (TLRs) play a central role in the pathogen clearance and pathological processes. The liver is an important innate immune organ, in which Kupffer cells and hepatocytes are important innate immune cells. However, the role of TLR2 and TLR4 in the liver caused by Toxoplasma gondii infection remains less clear. In this study, mice were infected with T. gondii RH strain and the grades of liver and spleen injuries were histopathologically evaluated. TLR2+ and TLR4+ cells in the livers and spleens were detected by immunohistochemistry, and their messenger RNA (mRNA) expressions were detected using quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). The pathological severities in the livers and spleens were increased with time in T. gondii-infected mice. Compared with uninfected controls, obvious TLR2+ and TLR4+ cells were observed in the livers and spleens infected with T. gondii at 8 days post-infection, accompanied with significantly over-expressed mRNA levels of TLR2 and TLR4 in the livers and spleens after infection. Our data indicated that increased levels of TLR2 and TLR4 in the liver and spleen may play an important role during acute T. gondii infection.