Disseminated toxoplasmosis with atypical symptoms which developed with exacerbation of systemic lupus erythematosus.

Toxoplasma is a common parasite worldwide that mainly affects the brain, lungs and eyes. Although toxoplasmic encephalitis is a lethal disease without treatment, past case reports show most patients with systemic lupus erythematosus who developed toxoplasmic encephalitis were misdiagnosed and treated as neuropsychiatric systemic lupus erythematosus, which led to unfavorable outcomes. We herein describe a case of disseminated toxoplasmosis affecting all the above organs with atypical symptoms, which developed with exacerbation of systemic lupus erythematosus. She had initially manifested with retinochoroiditis without vitritis, mild cognitive impairment and an isolated lung mass. These are completely different from the classic symptoms of toxoplasmosis that have been reported in patients with HIV infection and/or those after hematopoietic transplantation. Our case, together with previously reported cases, suggests the manifestation of toxoplasmosis that develops in systemic lupus erythematosus patients can be different from that seen in conventional cases and varies between individual patients. Our case highlights both the difficulty in and the importance of diagnosing toxoplasmosis in patients with systemic lupus erythematosus and provides helpful information to identify this rare, devastating, yet treatable disease.

Successful treatment of toxoplasmic encephalitis diagnosed early by polymerase chain reaction after allogeneic hematopoietic stem cell transplantation: two case reports and review of the literature.

Toxoplasmic encephalitis represents a rare, but often fatal infection after allogeneic hematopoietic stem cell transplantation. Polymerase chain reaction (PCR)-based preemptive therapy is considered promising for this disease, but is not routinely applied, especially in low seroprevalence countries including Japan. We encountered 2 cases of toxoplasmic encephalitis after transplantation that were successfully treated. The diagnosis of toxoplasmic encephalitis in these cases was confirmed by PCR testing when neurological symptoms were observed. Both patients received pyrimethamine and sulfadiazine treatments within 2 weeks of the development of neurological symptoms, and remained free of recurrence for 32 and 12 months. These results emphasized the importance of the PCR test and immediate treatment after diagnosis for the management of toxoplasmic encephalitis.

Overexpression of IL-8 in the cornea induces ulcer formation in the SCID mouse.

AIMS: Although interleukin 8 (IL-8) is not produced in the normal cornea, it has been detected there in several pathological conditions. In this study, the direct effects of IL-8 overexpression on the cornea was examined. METHODS: The corneal surface of severe combined immunodeficiency mice was infected by the adenovirus vector encoding human IL-8 (IL-8/Ad5) and clinical and pathological changes were observed at various time points. RESULTS: Clinically, marked angiogenesis and ulcer formation in the cornea were observed by 12 hours and 24 hours, respectively. Histologically, prominent angiogenesis was observed in the corneal stroma at 12 hours. Cleft formation between the corneal epithelium and stroma, and neutrophil infiltration into the corneal stroma were seen at 16 hours. By 24 hours after the infection with IL-8/Ad5, a shallow ulcer was formed in the cornea. In contrast, infection with the control adenovirus carrying the beta galactosidase gene (LacZ) showed neither corneal ulceration nor neutrophil infiltration. Immunohistochemical analysis showed that infection with IL-8/Ad5 resulted in the production of IL-8 by corneal and conjunctival stromal cells. CONCLUSION: Our results indicate that IL-8 overexpression in corneal tissue causes ulcer formation in the cornea through chemoattraction of neutrophils, suggesting the aetiological role of IL-8 in some types of corneal ulcers.

Heat stress-induced modulation of host defense against Toxoplasma gondii infection in mice.

This study examined the effects of burn injury on murine immune response against Toxoplasma gondii infection. Male C57BL/6 mice were divided into 3 groups: T. gondii infection (group T), burn injury (group B), and burn injury followed by T. gondii infection (group BT). The survival of group BT was significantly lower than those of group B and group T. Parasite abundance in the tissues was determined by quantitative competitive-polymerase chain reaction. Group BT exhibited significantly higher numbers of T. gondii than group T. Antibody production against T.g.HSP30 in group BT was significantly lower than that in group T, whereas no significant difference was observed in SAG1-specific antibody production. Delayed-type hypersensitivity (DTH) specific for 2,4-dinitrofluorobenzene (DNFB) of both group B and group BT was significantly lower than that of group T. One week after infection, serum interferon-gamma (IFN-gamma) and interleukin (IL)-10 levels in group BT were significantly lower, whereas serum IL-6 levels were significantly higher than in group T Serum TNF-alpha levels in both group T and group BT were elevated at 1 wk after infection, although there was no significant difference between them. Serum IFN-gamma, IL-10, and TNF-alpha levels in group B were not elevated during the experimental term. In conclusion, the impaired antigen-specific antibody production and DTH response, together with the modulated patterns of cytokine responses, seemed to be strongly involved in the development of burn-induced immunosuppression and the consequent increased susceptibility to T. gondii infection in mice.

Involvement of MyD88 in host defense and the down-regulation of anti-heat shock protein 70 autoantibody formation by MyD88 in Toxoplasma gondii-infected mice.

This study investigated the influence of TLR (toll-like receptor)4, TLR2, and MyD88 in Toxoplasma gondii-infected wild-type (WT) mice and TLR4-, TLR2-, and MyD88-deficient mice. Ninety-five percent of MyD88-deficient mice died 10-16 days after intraperitoneal infection with 100 cysts of T. gondii Fukaya strain, whereas 95-100% of TLR4- and TLR2-deficient mice and WT C57BL/6 (B6) mice survived for more than 7 wk after T. gondii infection. The distribution of T. gondii in various organs of TLR4-, TLR2-, and MyD88-deficient mice and WT B6 mice was assessed 2 wk after T. gondii intraperitoneal infection using quantitative competitive polymerase chain reaction. In MyD88-deficient mice, high levels of T. gondii load were observed in the brain, tongue, heart, lungs, spleen, liver, mesenteric lymph node, and kidneys after infection. The T. gondii load was significantly increased in the lungs in both TLR4- and TLR2-deficient mice compared with WT B6 mice. High levels of anti-mouse heat shock protein (mHSP)70 autoantibody and anti-T. gondii HSP70 antibody production were detected in the sera from MyD88-deficient mice.

Organ infectivity of Toxoplasma gondii in interferon-gamma knockout mice.

To determine the influence of interferon (IFN)-gamma on the organ infectivity and on the genetic susceptibility of susceptible (C57BL/6) and resistant (BALB/c) strains after peroral infection with cysts of Toxoplasma gondii. IFN-gamma knockout (KO) mice in C57BL/6 and BALB/c backgrounds were utilized. The kinetics of the changes in T. gondii abundance were evaluated with a quantitative competitive polymerase chain reaction assay in various organs at different times after peroral infection. In IFN-gamma KO mice, a T. gondii-specific gene, SAG1, was detected in all organs examined, and the protozoan proliferated much more actively than in wild-type mice. The abundance of T. gondii was much higher in mesenteric lymph nodes and the heart than in other organs. In contrast, in the nervous system organs and kidneys, only a weakly detectable reaction was observed. Toxoplasma gondii grew at a more rapid rate in the organs of IFN-gamma KO C57BL/6 mice than in the organs of IFN-gamma KO BALB/c mice during the course of infection. Destruction of the IFN-gamma gene showed remarkable effects on the infectivity in both susceptible and resistant mice.

Establishment of gene-vaccinated skin grafting against Toxoplasma gondii infection in mice.

Vaccine effects of in vivo gene-vaccinated skin graft were evaluated against Toxoplasma gondii (T. gondii) infection. By using a gene gun, cDNA coding T. gondii SAG1 molecule was intracutaneously vaccinated into C57BL/6 (B6; a susceptible strain), BALB/c (a resistant strain) and (C57BL/6 x BALB/c) F1 (CBF1) mice, and the gene-vaccinated skin of these strains was transplanted to CBF1 mice. Regarding the antibody production against SAG1, CBF1-recipient mice transplanted with the SAG1 gene-vaccinated B6 skin were high responders, whereas CBF1 mice skin grafted with vaccinated skin of both BALB/c and CBF1 mice were low responders. The donor-derived LC/DC migrated to the draining lymph nodes of the recipients from the skin graft within 3 days. The vaccine effect against T. gondii challenge infection was obtained in CBF1 mice which received the skin graft of the SAG1 gene-vaccinated BALB/c mice.

Toxoplasma gondii Hsp70 as a danger signal in toxoplasma gondii-infected mice.

Toxoplasma gondii Hsp70, T gondii Hsp30/bag1, and surface antigen 1 messenger RNAs were shown to be useful in analyzing stage conversion of T gondii between bradyzoites and tachyzoites. The high-level expression of T gondii Hsp70 was correlated with mortality in interferon-gamma knockout mice infected with T gondii. Tgondii Hsp70 inhibited the induction of nitric oxide release by peritoneal macrophages of T gondii-infected mice. These findings identify T gondii Hsp70 as a danger signal during lethal, acute T gondii infection.

Lenses of SPARC-null mice exhibit an abnormal cell surface-basement membrane interface.

SPARC (secreted protein acidic and rich in cysteine) is a matricellular protein involved in cell-matrix interactions. We have shown previously that mice deficient in SPARC develop posterior cortical cataract early in life that progresses to a mature opacity and capsule rupture. To evaluate the primary effects of SPARC deficiency in the lens, we examined the lenses of SPARC-null and wild-type mice by electron microscopy and immunohistochemistry to investigate whether ultrastructural abnormalities occur at the basement membrane (capsule)-lens cell interface in SPARC-null mice. The most notable feature in the lenses of SPARC-null mice, relative to wild-type animals, was the modification of the basal surface of the lens epithelial and fiber cells at the basement membrane (capsule) interface. Electron microscopy revealed numerous filopodial projections of the basal surface of the lens epithelial and fiber cells into the extracellular matrix of the anterior, posterior, and equatorial regions of the lens capsule. In 1 week old precataractous lenses, basal invasive filopodia projecting into the capsule were small and infrequent. Both the size and frequency of these filopodia increased in precataractous 3-4 week old lenses and were prominent in the cataractous 5-6 week old lenses. By rhodamine-phalloidin labeling, we confirmed the presence of basal invasive filopodia projecting into the lens capsule and demonstrated that the projections contained actin filaments. In contrast to the obvious abnormal projections at the interface between the basal surface of the lens epithelial and fiber cells and the lens capsule, the apical and lateral plasma membranes of lens epithelial cells and lens fibers in SPARC-null mice were as smooth as those of wild-type mice. We conclude that the absence of SPARC in the murine lens is associated with a filopodial protrusion of the basal surface of the lens epithelium and differentiating fiber cells into the lens capsule. The altered structures appear prior to the opacification of the lens in the SPARC-null model. These observations are consistent with one or more functions previously proposed for SPARC as a modulator of cell shape and cell-matrix interactions.

Anti-HSP70 autoantibody formation by B-1 cells in Toxoplasma gondii-infected mice.

Formation of anti-Toxoplasma gondii HSP70 (TgHSP70) antibody cross-reactive to mouse HSP70 (mHSP70) was observed in the sera of BALB/c (a resistant strain) and C57BL/6 (B6; a susceptible strain) mice after peroral infection with T. gondii cysts of the Fukaya strain. The levels of anti-mHSP70 immunoglobulin G (IgG) autoantibody production in B6 mice were higher than those in BALB/c mice. The isotype and subclass of IgG of anti-TgHSP70 monoclonal antibodies cross-reactive to mHSP70 were mu and gamma3. Anti-mHSP70 autoantibody in T. gondii-infected BALB/c and B6 mice was shown to be produced by the CD5(+) subset of B cells (B-1a cells) but not by conventional B cells (B-2 cells). The epitopes recognized by anti-mHSP70 autoantibody were located primarily in the C-terminal fragment of mHSP70.

A role of carboxy-terminal region of Toxoplasma gondii-heat shock protein 70 in enhancement of T. gondii infection in mice.

We investigated the role of recombinant Toxoplasma gondii heat shock protein (rT.g.HSP) 70-full length, rT.g.HSP70-NH2-terminal region, or rT.g.HSP70-carboxy-terminal region in prophylactic immunity in C57BL/6 mice perorally infected with Fukaya cysts of T. gondii. At 3, 4, 5, and 6 weeks after infection, the number of T. gondii in the brain tissue of each mouse was measured by quantitative competitive-polymerase chain reaction (QC-PCR) targeting the surface antigen (SAG) 1 gene. Immunization with rT.g.HSP70-full length or r.T.g.HSP70-carboxy-terminal region increased the number of T. gondii in the brain tissue after T. gondii infection, whereas immunization with rT.g.HSP70-NH2-terminal region did not. These results suggest that T.g.HSP70-carboxy-terminal region as well as T.g.HSP70-full length may induce deleterious effects on the protective immunity of mice infected with a cyst-forming T. gondii strain, Fukaya.

Protective immunity induced by vaccination with SAG1 gene-transfected cells against Toxoplasma gondii-infection in mice.

To develop a vaccine by augmenting the protective cellular immunity against Toxoplasma gondii (T. gondii)-infection, T gondii SAG1 gene-transfectants were established by using RMA.S (H-2b), a murine transporter associated with the antigen processing (TAP) molecule-deficient lymphoma line, as a host antigen-presenting cell (APC). Immunization of C57BL/6 mice with the SAG1-transfected RMA.S induced CD8+ cytotoxic T lymphocytes (CTL) specific for not only SAG1-transfected RMA.S but also T gondii-infected RMA.S, and elicited protective responses to infection with a virulent T. gondii strain, RH.

SPARC deficiency leads to early-onset cataractogenesis.

PURPOSE: To determine the role of SPARC (secreted protein, acidic, and rich in cysteine) in cataractogenesis by examining mice deficient in a matricellular protein SPARC. METHODS: Mice were rendered SPARC-deficient by a targeted disruption of the gene. Slit-lamp microscopy and histology were used to examine the eyes of SPARC-null and wild-type mice from birth to 14 months of age. RESULTS: SPARC-null mice developed opacities in the posterior cortex of the eye as early as 1.5 months after birth. The diffuse cataracts appeared to progress toward the anterior cortex and reached maturity in many animals by 3.5 months of age. Early stages of cataractogenesis in SPARC-null mice included inhibition of normal lens fiber cell differentiation, degeneration of fiber cells, vacuole formation at the equator, and liquefaction of the cortex. No cataracts were detected in wild-type mice up to the age of 8 months. CONCLUSIONS: The early onset of cataracts in SPARC-null mice establishes that the gene is essential to the maintenance of lens transparency.

Abnormal naive and memory T lymphocyte subsets in the peripheral blood of patients with uveitis.

PURPOSE: To better define the role of lymphocytes in the pathogenesis of uveitis, we studied the expression of memory and naive cell markers on T lymphocytes from peripheral blood. METHODS: Surface antigens on T lymphocytes obtained from peripheral blood of 27 patients with uveitis, including 12 patients with Behçet's disease (BD), 7 patients with Vogt-Koyanagi-Harada disease (VKH), and 8 patients with idiopathic uveitis (IU), were detected by three-color flow cytometric analysis. Lymphocytes from 14 age-matched healthy control subjects were similarly evaluated. RESULTS: The percentage of T lymphocytes that were CD4+CD29+ lymphocytes (memory cells) was high in all patients with uveitis, while that of CD4+CD45RA+ lymphocytes (naive cells) was lower in patients with BD and VKH, although the difference was not statistically significant. The percentage of CD29+ cells within CD3+CD4+ cell population was significantly higher in patients with BD and VKH than in the controls (p < 0.01), and the percentage of CD45RA+ cells was significantly lower in BD patients than in controls (p < 0.01). The T lymphocyte subsets in patients with IU were similar to the controls. CONCLUSIONS: These results show an abnormal distribution of T lymphocytes in patients with uveitis associated with an underlying systemic disease.

Correlation between direct binding ability of synthetic T. gondii SAG1 peptides to HLA-A2 measured by a sensor for surface plasmon resonance and antigenicity of the peptides for T. gondii-infected cell-specific CTL.

Toxoplasma gondii (T. gondii) -infected B lymphoma cells present T. gondii antigens in the context of major histocompatibility complex molecules to T. gondii-specific CD8+ cytotoxic T cells (CTL). HLA-A2 molecules of T. gondii-infected human cells have been shown to be used in presenting T. gondii antigens to CD8+ CTL. SAG1, one of the major antigenic molecules of T. gondii, is an antigen for T. gondii-specific CTL, and represents a possible basis for vaccines. The direct binding of nonamer SAG1 peptides to HLA-A2 was assayed here using an automated biosensor system with a sensor for surface plasmon resonance detection. The antigenicity of synthetic SAG1 peptides to T. gondii-specific CD8+ CTL also was assayed. The present study found a high correlation between the binding ability of synthetic SAG1 peptides to HLA-A2 and the antigenicity of peptides to T. gondii-infected cell-specific CD8+ CTL.

Heat shock cognate protein 71-associated peptides function as an epitope for Toxoplasma gondii-specific CD4+ CTL.

HLA-DR-restricted CD4+ cytotoxic T-lymphocyte (CTL) lines specific for Toxoplasma gondii (T. gondii)-infected melanoma cells have been established from peripheral blood lymphocytes (PBLs) of a patient with chronic toxoplasmosis. The role of heat shock cognate protein (HSC) 71 in antigen (Ag) processing and presentation of T. gondii-infected melanoma cells to these CD4+ CTL lines was investigated. A human melanoma cell line (P36) pulsed with T. gondii-infected P36 cell-derived HSC71 was lysed by a T. gondii-specific CD4+ CTL line (Tx-HSC-1). The Tx-HSC-1 also killed T. gondii-infected P36 cells. The lytic activity of Tx-HSC-1 against P36 cells pulsed with T. gondii-infected P36 cell-derived HSC71 was inhibited by monoclonal antibodies (mAbs) against HSC71. Anti-human leukocyte antigen (HLA)-DR mAb also partially blocked the lytic activity, whereas anti-HLA-A,B,C mAb did not block the lytic activity. In addition, a flow cytometric analysis with these specific mAbs against HSC71 showed HSC71 to be expressed on the cell surface of T. gondii-infected P36 cells as well as uninfected P36 cells. These data indicate that HSC71 molecules are expressed on human melanoma cell line P36, and that HSC71 may play a potential role in Ag presentation and processing of T. gondii-infected P36 cells to CD4+ CTL.

Melanoma specific Th1 cytotoxic T lymphocyte lines in Vogt-Koyanagi-Harada disease.

AIMS/BACKGROUND: To determine the functional properties and cytokine production profiles of melanoma specific cytotoxic T lymphocytes (CTLs) induced from peripheral blood leucocytes of two patients with Vogt-Koyanagi-Harada disease (VKH). METHODS: Melanoma specific CTL lines were established by long term coculture with a human melanoma cell line (P-36). Cytotoxic activity against P-36 was measured by 51Cr release. The involvement of human leucocyte antigen (HLA) class I or class II molecules in the cytotoxicity of the CTL lines against P-36 was analysed using anti-HLA class I or anti-HLA class II monoclonal antibody (MAb). Surface molecules of CTL lines were analysed by flow cytometry using MAbs specific for CD4, CD8, CD16, CD25, CD56, HLA-DR, T cell antigen receptor (TCR) alpha beta and TCR gamma delta. Cytokine production and soluble interleukin 2 receptor (sIL-2R) secretion were determined by enzyme linked immunosorbent assays. mRNAs of cytokines were analysed using reverse transcription polymerase chain reaction (RT-PCR). RESULTS: CTLs showed strong cytotoxic activity against P-36. The CTL activity of the cell lines against P-36 was inhibited by the anti-HLA-DR MAb, whereas the MAb specific for monomorphic determinants of HLA-A, B, and C failed to block lytic activity. Flow cytometry identified the following surface molecules: CD4+, CD8-, CD16-, CD25+, CD56-, HLA-DR+, TCR alpha beta +, and TCR gamma delta-. CTLs constitutively produced a high level of IL-6. IL-6 production and sIL-2R secretion of CTLs were enhanced when CTLs were stimulated with P-36. CTLs also produced high levels of interferon gamma (IFN-gamma) and IL-2, but not IL-4. mRNAs of IL-2 and IFN-gamma were detected by RT-PCR in the CTLs. CONCLUSIONS: Melanoma specific HLADR restricted T helper 1 (Th1) CTLs may play a role in the immunopathogenesis of VKH.

Differential expression of tenascin in the skin during hapten-induced dermatitis.

Tenascin is a large extracellular matrix glycoprotein which is found in limited regions of normal adult tissues including the skin. We investigated the induction of tenascin expression in mouse skin during hapten-induced dermatitis. In the dorsal skin, hapten application first induced a transient expression of tenascin in deeper regions of the skin. Its distribution then spread over the whole dermis corresponding to the infiltration of Mac-2-positive macrophages. In the ear, tenascin was consistently found in the subcutaneous tissue on the inner side, but very little was seen on the outer side. Tenascin did appear transiently, however, on both sides under hapten treatment. In the early phase of allergic contact dermatitis, no apparent induction of tenascin expression was observed in the swollen ear. However, there was an abundant tenascin expression on both sides during healing. Tenascin expressed under normal conditions was mostly the 180-kDa isoform, while the 230-kDa isoform was markedly induced during healing of the dermatitis. These results suggest that tenascin, particularly the larger 230-kDa isoform, may play important roles in the pathogenesis and healing of hapten-induced dermatitis.