Ophthalmological findings in patients suspected with COVID-19 at a tertiary hospital in Pernambuco, Brazil / Achados oftalmológicos em pacientes com suspeita de COVID-19 atendidos na triagem do HC-UFPE

ABSTRACT Purpose: This study evaluated the frequency of the most common ophthalmological, neurological, and systemic findings in symptomatic patients seen at a COVID-19 screening service at Hospital das Clínicas - Universidade Federal de Pernambuco. Methods: A total of 104 patients under clinical suspicion of SARS-CoV-2 infection underwent medical evaluation through an ophthalmological and systemic symptoms survey. All participants selected for the study underwent COVID-19 RT-PCR testing. Results: The mean age was 38.8 years, with 44.23% between 31 and 40 years old, 68.27% female, and 31.73% male. The most common symptoms in patients with a positive RT-PCR test were cough (69.23%), fever (42.3%), hyposmia (38.46%), myalgia (38.46%), and ageusia (30.77%). In the positive group, 34.61% presented with ophthalmological symptoms: burning (19.23%), eye pain (11.54%), foreign body sensation (7.7%), hyperemia (7.7%), and tearing (3.84%). Conclusions: Systemic clinical features were characteristic of upper respiratory infection, but neurological findings of hyposmia and anosmia proved to be important markers for suspicion of SARS-CoV-2 infection. Ophthalmic symptoms in patients with COVID-19 were similar to those observed in other viral conditions and may precede systemic conditions. A high rate of self-medication was observed for general symptoms compared with ophthalmological conditions.

RESUMO Objetivo: Avaliar a frequência dos sintomas oftalmológicos, neurológicos e sistêmicos mais comuns em pacientes sintomáticos atendidos no serviço de triagem de COVID-19 do Hospital das Clínicas da UFPE. Métodos: Cento e quatro pacientes com suspeita clínica de infecção por Sars-Cov-2 foram submetidos a avaliação médica e aplicação de questionário sobre a sintomatologia oftalmológica, neurológica e sistêmica. Todos os participantes do estudo tiveram exame de RT-PCR para COVID-19 solicitado. Resultados: A média de idade foi de 38,8 anos, com 44,23% entre 31 e 40 anos. Mulheres corresponderam a 68,27% dos atendimentos e homens a 31,73%. Os sintomas mais frequentes nos pacientes com RT-PCR positivo foram: tosse (69,23%), febre (42,3%), mialgia (38,46%), hiposmia (38,46%), e ageusia (30,77%). Neste grupo, os sintomas oftalmológicos estiveram presentes em 34,61%, sendo: ardor (19,23%), dor ocular (11,54%), sensação de corpo estranho (7,7%), hiperemia (7,7%) e lacrimejamento (3,84%) os mais encontrados. Conclusões: O quadro clínico sistêmico foi característico de infecção respiratória alta, porém os achados neurológicos de hiposmia e anosmia mostraram-se importantes marcadores para a suspeição dos casos de infecção por COVID-19. Os sintomas oftalmológicos dos pacientes com COVID-19 foram semelhantes aos presentes em outros quadros virais, podendo preceder o quadro sistêmico. Houve uma alta taxa de automedicação para os sintomas gerais quando comparado ao quadro oftalmológico.

Ophthalmological findings in patients suspected with COVID-19 at a tertiary hospital in Pernambuco, Brazil.

PURPOSE: This study evaluated the frequency of the most common ophthalmological, neurological, and systemic findings in symptomatic patients seen at a COVID-19 screening service at Hospital das Clínicas - Universidade Federal de Pernambuco. METHODS: A total of 104 patients under clinical suspicion of SARS-CoV-2 infection underwent medical evaluation through an ophthalmological and systemic symptoms survey. All participants selected for the study underwent COVID-19 RT-PCR testing. RESULTS: The mean age was 38.8 years, with 44.23% between 31 and 40 years old, 68.27% female, and 31.73% male. The most common symptoms in patients with a positive RT-PCR test were cough (69.23%), fever (42.3%), hyposmia (38.46%), myalgia (38.46%), and ageusia (30.77%). In the positive group, 34.61% presented with ophthalmological symptoms: burning (19.23%), eye pain (11.54%), foreign body sensation (7.7%), hyperemia (7.7%), and tearing (3.84%). CONCLUSIONS: Systemic clinical features were characteristic of upper respiratory infection, but neurological findings of hyposmia and anosmia proved to be important markers for suspicion of SARS-CoV-2 infection. Ophthalmic symptoms in patients with COVID-19 were similar to those observed in other viral conditions and may precede systemic conditions. A high rate of self-medication was observed for general symptoms compared with ophthalmological conditions.

Heterologous Chimeric Construct Comprising a Modified Bacterial Superantigen and a Cruzipain Domain Confers Protection Against Trypanosoma cruzi Infection.

Chagas disease is an endemic chronic parasitosis in Latin America affecting more than 7 million people. Around 100 million people are currently at risk of acquiring the infection; however, no effective vaccine has been developed yet. Trypanosoma cruzi is the etiological agent of this parasitosis and as an intracellular protozoan it can reside within different tissues, mainly muscle cells, evading host immunity and allowing progression towards the chronic stage of the disease. Considering this intracellular parasitism triggers strong cellular immunity that, besides being necessary to limit infection, is not sufficient to eradicate the parasite from tissues, a differential immune response is required and new strategies for vaccines against Chagas disease need to be explored. In this work, we designed, cloned and expressed a chimeric molecule, named NCz-SEGN24A, comprising a parasite antigen, the N-terminal domain of the major cysteine protease of T. cruzi, cruzipain (Nt-Cz), and a non-toxic form of the staphylococcal superantigen (SAg) G, SEG, with the residue Asn24 mutated to Ala (N24A). The mutant SAg SEGN24A, retains its ability to trigger classical activation of macrophages without inducing T cell apoptosis. To evaluate, as a proof of concept, the immunogenicity and efficacy of the chimeric immunogen vs. its individual antigens, C3H mice were immunized intramuscularly with NCz-SEGN24A co-adjuvanted with CpG-ODN, or the recombinant proteins Nt-Cz plus SEGN24A with the same adjuvant. Vaccinated mice significantly produced Nt-Cz-specific IgG titers after immunization and developed higher IgG2a than IgG1 titers. Specific cell-mediated immunity was assessed by in-vivo DTH and significant responses were obtained. To assess protection, mice were challenged with trypomastigotes of T. cruzi. Both schemes reduced the parasite load throughout the acute phase, but only mice immunized with NCz-SEGN24A showed significant differences against control; moreover, these mice maintained 100% survival. These results encourage testing mutated superantigens fused to specific antigens as immune modulators against pathogens.

Cambio de actitud de estudiantes de Enfermería hacia la enfermedad mental mediante aprendizaje experiencial / Change of attitude by Nursing students towards mental illnesses through experiential learning

Introducción: La reforma psiquiátrica y promulgación del Plan Nacional de Salud Mental y Psiquiatría, plantea el desafío de un cambio de actitud hacia las personas con enfermedad mental. Es debido a la estigmatización y discriminación que los pacientes sufren violaciones de derechos humanos, negándoseles la posibilidad de gozar del grado más alto de salud. Para Enfermería de la Universidad Del Desarrollo es un desafío y deber formar profesionales respetuosos de la diversidad y capaces de generar cuidados de enfermería especializados, por lo que se diseñó un curso que incorporó Aprendizaje Experiencial. Objetivo: Promover un cambio de actitud hacia la enfermedad mental en estudiantes de Enfermería de tercer año mediante la incorporación del Aprendizaje Experiencial. Métodos: Estudio de cohorte. Se analizó la existencia de diferencias estadísticamente significativas (test t, 95 por ciento confianza y 0,05 significancia) en los puntajes obtenidos por 52 estudiantes de Enfermería, luego de aplicar la Escala de medición de actitudes hacia los enfermos mentales en futuros técnicos de salud, pre y post realización del curso, previa firma de consentimiento informado. Resultados: El puntaje promedio aumentó de 59,3 (de: 10,5) a 64,5 puntos (de: 8,8). La diferencia promedio en el puntaje total del test antes y después del curso es de 5,2 puntos. Al distinguir entre ítems favorables y desfavorables los puntajes promedios aumentan post realización del curso. Conclusión: Es posible promover un cambio de actitud hacia la enfermedad mental en estudiantes de Enfermería mediante el Aprendizaje Experiencial(AU)

Introduction: The psychiatric reform and the promulgation of the National Plan of Mental Health and Psychiatry posed the challenge of a change of attitude towards people with mental illnesses. Due to stigmatization and discrimination, patients suffer human rights violations, being denied the possibility of enjoying the highest degree of health. For Nursing at Universidad del Desarrollo it is a challenge and a duty to train professionals who are respectful of diversity and capable of generating specialized nursing care, so a course was designed that incorporated experiential learning. Objective: To promote a change of attitude towards mental illnesses in third-year Nursing students through the incorporation of experiential learning. Methods: Cohort study. We analyzed the existence of statistically significant differences (t-test, 95 percent confidence interval and 0.05 significance) in the scores obtained by 52 Nursing students, after applying the Attitudes towards the Mentally Ill Measuring Scale in future health technicians, before and after the completion of the course, after signing informed consent. Results: The average score increased from 59.3 (from: 10.5) to 64.5 points (from: 8.8). The average difference in the total score of the test before and after the course is 5.2 points. Upon distinguishing between favorable and unfavorable items, the average scores increase after the completion of the course. Conclusion: It is possible to promote a change of attitude towards mental illnesses in Nursing students by means of experiential learning(AU)

egc Superantigens Impair Monocytes/Macrophages Inducing Cell Death and Inefficient Activation.

Bacterial superantigens (SAgs) are enterotoxins that bind to MHC-II and TCR molecules, activating as much as 20% of the T cell population and promoting a cytokine storm which enhances susceptibility to endotoxic shock, causing immunosuppression, and hindering the immune response against bacterial infection. Since monocytes/macrophages are one of the first cells SAgs find in infected host and considering the effect these cells have on directing the immune response, here, we investigated the effect of four non-classical SAgs of the staphylococcal egc operon, namely, SEG, SEI, SEO, and SEM on monocytic-macrophagic cells, in the absence of T cells. We also analyzed the molecular targets on APCs which could mediate SAg effects. We found that egc SAgs depleted the pool of innate immune effector cells and induced an inefficient activation of monocytic-macrophagic cells, driving the immune response to an impaired proinflammatory profile, which could be mediated directly or indirectly by interactions with MHC class II. In addition, performing surface plasmon resonance assays, we demonstrated that non-classical SAgs bind the gp130 molecule, which is also present in the monocytic cell surface, among other cells.

Spatially offset Raman spectroscopy for in vivo bone strength prediction.

Bone strength is a worldwide health concern. Although multiple techniques have been developed to evaluate bone quality, there are still gaps to be filled. Here we report a non-invasive approach for the prediction of bone strength in vivo using spatially offset Raman spectroscopy. Raman spectra were acquired transcutaneously from the tibiae of mice from 4 to 23 weeks old and subsequently on the exposed bones. Partial least squares regression was applied to generate predictions of the areal bone mineral density (aBMD), volumetric bone mineralization density (vBMD), and maximum torque (MT) of each tibia as quantified by dual-energy X-ray absorptiometry, microCT imaging, and biomechanical tests, respectively. Significant correlations were observed between Raman spectral predictions and the reference values in all three categories. To our knowledge, this is the first demonstration of Raman spectroscopy predicting a biomechanical bone parameter (MT) in vivo with an uncertainty much smaller than the spread in the reference values.

Kinetic and thermodynamic studies of the interaction between activating and inhibitory Ly49 natural killer receptors and MHC class I molecules.

Natural killer (NK) cells are lymphocytes of the innate immune system that eliminate virally infected or malignantly transformed cells. NK cell function is regulated by diverse surface receptors that are both activating and inhibitory. Among them, the homodimeric Ly49 receptors control NK cell cytotoxicity by sensing major histocompatibility complex class I molecules (MHC-I) on target cells. Although crystal structures have been reported for Ly49/MHC-I complexes, the underlying binding mechanism has not been elucidated. Accordingly, we carried out thermodynamic and kinetic experiments on the interaction of four NK Ly49 receptors (Ly49G, Ly49H, Ly49I and Ly49P) with two MHC-I ligands (H-2Dd and H-2Dk). These Ly49s embrace the structural and functional diversity of the highly polymorphic Ly49 family. Combining surface plasmon resonance, fluorescence anisotropy and far-UV circular dichroism (CD), we determined that the best model to describe both inhibitory and activating Ly49/MHC-I interactions is one in which the two MHC-I binding sites of the Ly49 homodimer present similar binding constants for the two sites (∼106 M-1) with a slightly positive co-operativity in some cases, and without far-UV CD observable conformational changes. Furthermore, Ly49/MHC-I interactions are diffusion-controlled and enthalpy-driven. These features stand in marked contrast with the activation-controlled and entropy-driven interaction of Ly49s with the viral immunoevasin m157, which is characterized by strong positive co-operativity and conformational selection. These differences are explained by the distinct structures of Ly49/MHC-I and Ly49/m157 complexes. Moreover, they reflect the opposing roles of NK cells to rapidly scan for virally infected cells and of viruses to escape detection using immunoevasins such as m157.

Uptake and intracellular trafficking of superantigens in dendritic cells.

Bacterial superantigens (SAgs) are exotoxins produced mainly by Staphylococcus aureus and Streptococcus pyogenes that can cause toxic shock syndrome (TSS). According to current paradigm, SAgs interact directly and simultaneously with T cell receptor (TCR) on the T cell and MHC class II (MHC-II) on the antigen-presenting cell (APC), thereby circumventing intracellular processing to trigger T cell activation. Dendritic cells (DCs) are professional APCs that coat nearly all body surfaces and are the most probable candidate to interact with SAgs. We demonstrate that SAgs are taken up by mouse DCs without triggering DC maturation. SAgs were found in intracellular acidic compartment of DCs as biologically active molecules. Moreover, SAgs co-localized with EEA1, RAB-7 and LAMP-2, at different times, and were then recycled to the cell membrane. DCs loaded with SAgs are capable of triggering in vitro lymphocyte proliferation and, injected into mice, stimulate T cells bearing the proper TCR in draining lymph nodes. Transportation and trafficking of SAgs in DCs might increase the local concentration of these exotoxins where they will produce the highest effect by promoting their encounter with both MHC-II and TCR in lymph nodes, and may explain how just a few SAg molecules can induce the severe pathology associated with TSS.

Ocular toxoplasmosis: evaluation of lacrimal-specific secretory IgA levels in both patients with active and inactive phases of the disease.

Ocular toxoplasmosis can result in recurrent uveitis. Studies have shown that a correlation between active ocular toxoplasmosis and the presence of anti-Toxoplasma gondii secretory IgA (SIgA) in tears. This study compares anti-T. gondii SIgA levels in patients' tears during the acute and inactive phases of toxoplasmic uveitis. Twenty-nine positive tear specific SIgA for T. gondii patients with acute toxoplasmic uveitis were selected and were followed-up for at least two years, when the anti-T. gondii SIgA tears levels were determined. Specific SIgA for T. gondii was negative in 22 patients (75.86%) and positive in seven patients (24.13%) of whom six (85.7%) were followed over three years. Average SIgA levels during the acute phase are 1.54 and decrease significantly to 0.72 (p = 0.0001) during the inactive phase of disease. Because anti-T. gondii SIgA in the tear is negative in 75.86% of patients after the acute phase of infection, T. gondii SIgA levels may be used as a complementary diagnostic marker for active ocular toxoplasmosis.

Ocular toxoplasmosis: evaluation of lacrimal - specific secretory IgA levels in both patients with active and inactive phases of the disease

Ocular toxoplasmosis can result in recurrent uveitis. Studies have shown that a correlation between active ocular toxoplasmosis and the presence of anti-Toxoplasma gondii secretory IgA (SIgA) in tears. This study compares anti-T. gondii SIgA levels in patients' tears during the acute and inactive phases of toxoplasmic uveitis. Twenty-nine positive tear specific SIgA for T. gondii patients with acute toxoplasmic uveitis were selected and were followed-up for at least two years, when the anti-T. gondii SIgA tears levels were determined. Specific SIgA for T. gondii was negative in 22 patients (75.86 percent) and positive in seven patients (24.13 percent) of whom six (85.7 percent) were followed over three years. Average SIgA levels during the acute phase are 1.54 and decrease significantly to 0.72 (p = 0.0001) during the inactive phase of disease. Because anti-T. gondii SIgA in the tear is negative in 75.86 percent of patients after the acute phase of infection, T. gondii SIgA levels may be used as a complementary diagnostic marker for active ocular toxoplasmosis.

Anti-Toxoplasma gondii secretory IgA in tears of patients with ocular toxoplasmosis: immunodiagnostic validation by ELISA.

Toxoplasma gondii causes posterior uveitis and the specific diagnosis is based on clinical criteria. The presence of anti-T. gondii secretory IgA (sIgA) antibodies in patients' tears has been reported and an association was found between ocular toxoplasmosis and the anti-T. gondii sIgA isotype in Brazilian patients. The purpose of this study was to provide an objective validation of the published ELISA test for determining the presence of anti-T. gondii sIgA in the tears of individuals with ocular toxoplasmosis. Tears from 156 patients with active posterior uveitis were analysed; 82 of them presented characteristics of ocular toxoplasmosis (standard lesion) and 74 patients presented uveitis due to other aetiologies. Cases of active posterior uveitis were considered standard when a new inflammatory focus satellite to old retinochoroidal scars was observed. The determination of anti-T. gondii sIgA was made using an ELISA test with crude tachyzoite antigenic extracts. Tears were collected without previous stimulation. Detection of sIgA showed 65.9% sensitivity (95% CI = 54.5-74.4), 71.6% specificity (95% CI = 59.8-81.2), a positive predictive value of 72% (95% CI = 60.3-81.5) and a negative predictive value of 65.4% (95% CI = 54.0-75.4). sIgA reactivity was higher in the tears of patients with active posterior uveitis due to T. gondii (p < 0.05). The test is useful for differentiating active posterior uveitis due to toxoplasmosis from uveitis caused by other diseases.

Anti-Toxoplasma gondii secretory IgA in tears of patients with ocular toxoplasmosis: immunodiagnostic validation by ELISA

Toxoplasma gondii causes posterior uveitis and the specific diagnosis is based on clinical criteria. The presence of anti-T. gondii secretory IgA (sIgA) antibodies in patients' tears has been reported and an association was found between ocular toxoplasmosis and the anti-T. gondii sIgA isotype in Brazilian patients. The purpose of this study was to provide an objective validation of the published ELISA test for determining the presence of anti-T. gondii sIgA in the tears of individuals with ocular toxoplasmosis. Tears from 156 patients with active posterior uveitis were analysed; 82 of them presented characteristics of ocular toxoplasmosis (standard lesion) and 74 patients presented uveitis due to other aetiologies. Cases of active posterior uveitis were considered standard when a new inflammatory focus satellite to old retinochoroidal scars was observed. The determination of anti-T. gondii sIgA was made using an ELISA test with crude tachyzoite antigenic extracts. Tears were collected without previous stimulation. Detection of sIgA showed 65.9 percent sensitivity (95 percent CI = 54.5-74.4), 71.6 percent specificity (95 percent CI = 59.8-81.2), a positive predictive value of 72 percent (95 percent CI = 60.3-81.5) and a negative predictive value of 65.4 percent (95 percent CI = 54.0-75.4). sIgA reactivity was higher in the tears of patients with active posterior uveitis due to T. gondii (p < 0.05). The test is useful for differentiating active posterior uveitis due to toxoplasmosis from uveitis caused by other diseases.

[Clinical characteristics of 64 individuals carrying active posterior presumptively toxoplasmic uveitis, in Pernambuco]. / Características clínicas de 64 indivíduos portadores de uveítis posterior activa presumiblemente toxoplásmica en Pernambuco.

PURPOSE: To describe clinical characteristics of posterior active uveitis presumptively by Toxoplasma gondii (PAUPT) in patients with typical lesion. Tranversal study. METHODS: Sixty-four patients with retinochoroiditis scatter and active satellite lesions examined in Pernambuco, Brazil. All were older than 10 years and immunocompetent. Gender, age, skin color, and residence were recorded. Previous uveitis, visual accuracy, intraocular pressure (IOP), and ocular examination were analyzed. RESULTS: 52% were males, most of them with white skin (68.8%). Mean age 29 years (+/-10.87). Eighty-four percent of the patients lived in the metropolitan area. 56.2% were having the first episode of uveitis. In the damaged eye, visual accuracy mean was 20/200, IOP mean 14.5 mmHg (+/-64). Hyperemia of the conjunctiva was observed in 29.7% of the patients and alterations of the cornea in 51.6%. There were cells in the aqueous humor in 62.7%. 6.2% had posterior synechiae. All had vitreous damage and 45.3% retinal vasculitis. In 42.2% of the patients, lesions were located in zone I of Holland and 90.6% had the size of one discus diameter or greater. Neuritis was observed in 28.2%. Uveitis was more frequent in the right eye (54.7%). CONCLUSION: PAUPT affects young people and the main symptom was reduction of visual acuity. IOP mean was normal. Alterations of the vitreous were observed in all cases. Injuries were equal to one discus diameter or greater and located in zone I of Holland.

Características clínicas de 64 indivíduos portadores de uveítis posterior activa presumiblemente toxoplásmica en Pernambuco / Clinical characteristics of 64 individuals carrying active posterior presumptively toxoplasmic uveitis, in Pernambuco

OBJETIVO: Describir características clínicas de uveítis posterior activa, presumiblemente por Toxoplasma gondii (UPAPT) en portadores de lesión típica. Estudio tranversal. MÉTODOS: 64 portadores de UPAPT con retinocorroiditis cicatrizada y lesión satélite activa, mayores de 10 años, inmunocompetentes, examinados en Permambuco, Brazil. Se analizó: sexo, edad, color de la piel, procedencia, uveítis anteriores, agudeza visual, presión ocular y exámen ocular. RESULTADOS: Masculino en 52 por ciento. Edad media 29 años (±10,87). Piel blanca en 68,8 por ciento. Domicilio en la área metropolitana en 80,4 por ciento. Primer episodio de uveítis en 56,2 por ciento. Media de visión en ojo afectado 20/200. Presión ocular media 14,5 mmHg (±7,64) en ojo afectado. Conjuntiva hiperemiada en 29,7 por ciento. Alteraciones corneales en 51,6 por ciento. Células en el humor acuoso en 67,2 por ciento. Sinéquias posteriores en 6,2 por ciento. Compromiso vítreo en 100 por ciento. Vasculitis retiniana en 45,3 por ciento. Lesiones localizads en la zona I de Holland en 42,2 por ciento, siendo de tamaño igual o mayor de un diámetro de disco en 90,6 por ciento. Neuritis en 28,2 por ciento. CONCLUSIÓN: UPAPT afecta adultos jóvenes, siendo el síntoma principal la disminución de la visión. Presión ocular media normal. Compromiso vítreo en todos los casos. Com mayor frequencia las lesiones fueron mayores de un diámetro de disco localizadas en la zona I de Holland.

PURPOSE: To describe clinical characteristics of posterior active uveitis presumptively by Toxoplasma gondii (PAUPT) in patients with typical lesion. Tranversal study. METHODS: Sixty-four patients with retinochoroiditis scatter and active satellite lesions examined in Pernambuco, Brazil. All were older than 10 years and immunocompetent. Gender, age, skin color, and residence were recorded. Previous uveitis, visual accuracy, intraocular pressure (IOP), and ocular examination were analyzed. RESULTS: 52 percent were males, most of them with white skin (68.8 percent). Mean age 29 years (±10.87). Eighty-four percent of the patients lived in the metropolitan area. 56.2 percent were having the first episode of uveitis. In the damaged eye, visual accuracy mean was 20/200, IOP mean 14.5 mmHg (±64). Hyperemia of the conjunctiva was observed in 29.7 percent of the patients and alterations of the cornea in 51.6 percent. There were cells in the aqueous humor in 62.7 percent. 6.2 percent had posterior synechiae. All had vitreous damage and 45.3 percent retinal vasculitis. In 42.2 percent of the patients, lesions were located in zone I of Holland and 90.6 percent had the size of one discus diameter or greater. Neuritis was observed in 28.2 percent. Uveitis was more frequent in the right eye (54.7 percent). CONCLUSION: PAUPT affects young people and the main symptom was reduction of visual acuity. IOP mean was normal. Alterations of the vitreous were observed in all cases. Injuries were equal to one discus diameter or greater and located in zone I of Holland.

Lacrimal secretory IgA in active posterior uveitis induced by Toxoplasma gondii

It is quite difficult to diagnose active toxoplasmosis in patients with ocular toxoplasmosis. Active posterior uveitis presumably due to Toxoplasma gondii infection (APUPT) is seldom produced during a prime-infection; hence most patients do not show high IgM antibodies. High levels of IgA have been described in active toxoplasmosis. The purpose of this study was to investigate possible association between APUPT and the specific anti-parasite sIgA in tears. The study was carried out as case-control. Tears of 25 clinically confirmed APUPT patients and 50 healthy control subjects were analyzed. All were IgG seropositive. Specific sIgA was determined by ELISA assay using T. gondii RH strain crude extract. Anti-T. gondii sIgA was found in 84 percent of the cases and in 22 percent of the control subjects. The intensity of the reaction was higher in APUPT cases (P = 0.007). There was strong association between APUPT patients and lacrimal sIgA (odds-ratio 18.61, P = 0.0001). ELISA test sensitivity was 84 percent and specificity 78 percent . Our data suggest that anti-T.gondii secretory IgA found in tears may become an important marker for active ocular toxoplasmosis.

The roles of thymidylate synthase and p53 in regulating Fas-mediated apoptosis in response to antimetabolites.

Fas (CD95/Apo-1) is a member of the tumor necrosis factor receptor family. Receptor binding results in activation of caspase 8, leading to activation of proapoptotic downstream molecules. We found that expression of Fas was up-regulated >10-fold in MCF-7 breast and HCT116 and RKO colon cancer cell lines after treatment with IC(60) doses of 5-fluorouracil (5-FU) and raltitrexed (RTX). Combined treatment with the agonistic Fas antibody CH-11 and either 5-FU or RTX resulted in a highly synergistic induction of apoptosis in these cell lines. Similar results were obtained for another antifolate, Alimta. Induction of thymidylate synthase expression inhibited Fas induction in response to RTX and Alimta, but not in response to 5-FU. Furthermore, thymidylate synthase induction abrogated the synergy between CH-11 and both antifolates but had no effect on the synergistic interaction between 5-FU and CH-11. Inactivation of p53 in MCF-7 and HCT116 cell lines blocked 5-FU- and antifolate-mediated up-regulation of Fas. Furthermore, Fas was not up-regulated in response to 5-FU or antifolates in the p53-mutant H630 colon cancer cell line. Lack of Fas up-regulation in the p53-null and -mutant lines abolished the synergistic interaction between 5-FU and CH-11. Interestingly, synergy was still observed between the antifolates and CH-11 in the p53-null HCT116 and p53-mutant H630 cell lines, although this was significantly reduced compared with the p53 wild-type cell lines. Our results indicate that Fas is an important mediator of apoptosis in response to both 5-FU and antifolates.

Lacrimal secretory IgA in active posterior uveitis induced by Toxoplasma gondii.

It is quite difficult to diagnose active toxoplasmosis in patients with ocular toxoplasmosis. Active posterior uveitis presumably due to Toxoplasma gondii infection (APUPT) is seldom produced during a prime-infection; hence most patients do not show high IgM antibodies. High levels of IgA have been described in active toxoplasmosis. The purpose of this study was to investigate possible association between APUPT and the specific anti-parasite sIgA in tears. The study was carried out as case-control. Tears of 25 clinically confirmed APUPT patients and 50 healthy control subjects were analyzed. All were IgG seropositive. Specific sIgA was determined by ELISA assay using T. gondii RH strain crude extract. Anti-T. gondii sIgA was found in 84% of the cases and in 22% of the control subjects. The intensity of the reaction was higher in APUPT cases (P = 0.007). There was strong association between APUPT patients and lacrimal sIgA (odds-ratio 18.61, P = 0.0001). ELISA test sensitivity was 84% and specificity 78%. Our data suggest that anti-T.gondii secretory IgA found in tears may become an important marker for active ocular toxoplasmosis.

Identification of 5-fluorouracil-inducible target genes using cDNA microarray profiling.

The fluoropyrimidine 5-Fluorouracil (5-FU) is widely used in the treatment of cancer. To identify novel downstream mediators of tumor cell response to 5-FU, we used DNA microarray technology to identify genes that are transcriptionally activated by 5-FU treatment in the MCF-7 breast cancer cell line. Of 2400 genes analyzed, 619 were up-regulated by >3-fold. Highly up-regulated genes (>6-fold) with signal intensities of >3000 were analyzed by Northern blot. Genes that were consistently found to be up-regulated were spermine/spermidine acetyl transferase (SSAT), annexin II, thymosin-beta-10, chaperonin-10, and MAT-8. Treatment of MCF-7 cells with the antifolate tomudex and DNA-damaging agent oxaliplatin also resulted in up-regulation of each of these targets. The 5-FU-induced activation of MAT-8, thymosin-beta-10, and chaperonin-10 was abrogated by inactivation of p53 in MCF-7 cells, whereas induction of SSAT and annexin II was significantly reduced in the absence of p53. Moreover, each of these genes contained more than one potential p53-binding site, suggesting that p53 may play an important regulatory role in 5-FU-induced expression of these genes. In addition, we found that basal expression levels of SSAT, annexin II, thymosin beta-10, and chaperonin-10 were increased (by approximately 2-3-fold), and MAT-8 expression dramatically increased (by approximately 10-fold) in a 5-FU-resistant colorectal cancer cell line (H630-R10) compared with the parental H630 cell line, suggesting these genes may be useful biomarkers of resistance. These results demonstrate the potential of DNA microarrays to identify novel genes involved in mediating the response of tumor cells to chemotherapy.