Role of antigen-presenting cells in non-ulcerated skin lesions caused by Leishmania (Leishmania) infantum chagasi.

In Central America, infection by Leishmania (Leishmania) infantum chagasi causes visceral leishmaniasis and non-ulcerated cutaneous leishmaniasis (NUCL). This work aimed to evaluate the participation of subpopulations of antigen-presenting cells in skin lesions of patients affected by NUCL through double-staining immunohistochemistry using cellular and intracellular markers. Twenty-three skin biopsies from patients affected by NUCL were used. Histological sections stained by HE were used for histopathological study. Immunohistochemical studies were performed using primary antibodies against Langerhans cells, dermal dendritic cells, T lymphocytes, and the cytokines IL-12, IFN-γ, TNF-α, iNOS, and IL-10. The histopathological lesions were characterized by an inflammatory infiltrate, predominantly lymphohistiocytic, of variable intensity, with a diffuse arrangement associated with epithelioid granulomas and discreet parasitism. Double-staining immunohistochemistry showed higher participation of dendritic cells producing the proinflammatory cytokine IL-12 in relation to the other evaluated cytokines. Activation of the cellular immune response was marked by a higher density of CD8 Tc1-lymphocytes followed by CD4 Th1-lymphocytes producing mainly IFN-γ. The data obtained in the present study suggest that antigen-presenting cells play an important role in the in situ immune response through the production of proinflammatory cytokines, directing the cellular immune response preferentially to the Th1 and Tc1 types in NUCL caused by L. (L.) infantum chagasi.

Macrophage Polarization in the Skin Lesion Caused by Neotropical Species of Leishmania sp.

Macrophages play important roles in the innate and acquired immune responses against Leishmania parasites. Depending on the subset and activation status, macrophages may eliminate intracellular parasites; however, these host cells also can offer a safe environment for Leishmania replication. In this sense, the fate of the parasite may be influenced by the phenotype of the infected macrophage, linked to the subtype of classically activated (M1) or alternatively activated (M2) macrophages. In the present study, M1 and M2 macrophage subsets were analyzed by double-staining immunohistochemistry in skin biopsies from patients with American cutaneous leishmaniasis (ACL) caused by L. (L.) amazonensis, L. (V.) braziliensis, L. (V.) panamensis ,and L. (L.) infantum chagasi. High number of M1 macrophages was detected in nonulcerated cutaneous leishmaniasis (NUCL) caused by L. (L.) infantum chagasi (M1 = 112 ± 12, M2 = 43 ± 12 cells/mm2). On the other side, high density of M2 macrophages was observed in the skin lesions of patients with anergic diffuse cutaneous leishmaniasis (ADCL) (M1 = 195 ± 25, M2 = 616 ± 114), followed by cases of localized cutaneous leishmaniasis (LCL) caused by L. (L.) amazonensis (M1 = 97 ± 24, M2 = 219 ± 29), L. (V.) panamensis (M1 = 71 ± 14, M2 = 164 ± 14), and L. (V.) braziliensis (M1 = 50 ± 13, M2 = 53 ± 10); however, low density of M2 macrophages was observed in NUCL. The data presented herein show the polarization of macrophages in skin lesions caused by different Leishmania species that may be related with the outcome of the disease.

Leishmania amazonensis downregulates macrophage iNOS expression via Histone Deacetylase 1 (HDAC1): a novel parasite evasion mechanism.

The induced expression of nitric oxide synthase (iNOS) controls the intracellular growth of Leishmania in infected macrophages. Histones deacetylases (HDACs) negatively regulate gene expression through the formation of complexes containing transcription factors such as NF-κB p50/50. Herein, we demonstrated the occupancy of p50/p50_HDAC1 to iNOS promoter associated with reduced levels of H3K9Ac. Remarkably, we found increased levels of HDAC1 in L. amazonensis-infected macrophages. HDAC1 upregulation was not found in L. major-infected macrophages. The parasite intracellular load was reduced in HDAC1 knocked-down macrophages, which presented increased nitric oxide levels. HDAC1 silencing led to the occupancy of CBP/p300 to iNOS promoter and the rise of H3K9Ac modification. Importantly, the immunostaining of skin samples from hiporeactive cutaneous leishmaniasis patients infected with L. amazonensis, revealed high levels of HDAC1. In brief, L. amazonensis induces HDAC1 in infected macrophages, which contribute to parasite survival and is associated to hiporeactive stage found in L. amazonensis infected patients.

The dentist as a child abuse informer: a case report

Introducition: Domestic violence against children interferes in their psychological development, leading to sequels that manifest and persist up to adulthood. Physical evidence of domestic violence is easily observed in the orofacial complex and eventually becomes detected by dentists. Case Report: We report the case of a 9-year-old victim of maltreatment who was diagnosed during dental treatment. The existence of physical injuries (a hematoma in the left orbit and burns on the left hand and in the lips) aroused the attention of the pediatric dentistry, whose brought the case to the responsible authorities. Custody of the child was granted to the grandmother by a court decision, which enabled the recovery of health and quality of life. Conclusion: Professionals must properly conduct cases through complaints in order to protect children from future occurrences.

Introdução: A violência doméstica contra as crianças interfere no seu desenvolvimento psicológico, levando a sequelas que se manifestam e persistem até a idade adulta. A evidência física da violência doméstica é facilmente observada no complexo orofacial e, eventualmente, é detectada pelos dentistas. Relato do Caso: Relatamos o caso de uma vítima de maus-tratos, de 9 anos de idade, que foi diagnosticada durante o tratamento odontológico. Um odontopediatra, durante as consultas de rotina, após identificar injúrias físicas (hematoma na órbita esquerda e queimaduras na mão esquerda e lábios), suspeitou tratar-se de maus tratos, levando o caso às autoridades responsáveis. A custódia da criança foi concedida à avó por uma decisão judicial, o que permitiu a recuperação da saúde e qualidade de vida. Conclusão: Os profissionais devem conduzir adequadamente os casos de abuso, a fim de proteger as crianças de ocorrências futuras.

A cross-sectional study on canine Leishmania (L.) infantum chagasi infection in Amazonian Brazil ratifies a higher prevalence of specific IgG-antibody response than delayed-type hypersensitivity in symptomatic and asymptomatic dogs.

This was a cross-sectional study which analyzed the prevalence and the clinical and immunological spectrum of canine Leishmania (L.) infantum chagasi infection in a cohort of 320 mongrel dogs living in an endemic area of American visceral leishmaniasis in the Amazonian Brazil by using, mainly, the indirect fluorescence antibody test (IFAT-IgG) and the delayed-type hypersensitivity (DTH), and the parasite research by the popliteal lymph node aspiration. The IFAT and DTH reactivity recognized three different immune response profiles: (1) IFAT((+))/DTH((-)) (107 dogs), (2) IFAT((-))/DTH((+)) (18 dogs), and (3) IFAT((+))/DTH((+)) (13 dogs), providing an overall prevalence of infection of 43% (138/320). Thus, the specific prevalence of IFAT( (+) )/DTH( (-) ) 33.4% (107/320) was higher than those of IFAT( (-) )/DTH( (+) ) 5.6% (18/320) and IFAT( (+) )/DTH( (+) ) 4.0% (13/320). Moreover, the frequency of these profiles among 138 infected dogs showed that the IFAT( (+) )/DTH( (-) ) rate of 77.5% (107/138) was also higher than those of 13.0% (18/138) of IFAT( (-) )/DTH( (+) ) and 9.5% (13/138) of IFAT( (+) )/DTH( (+) ) rates. The frequency of asymptomatic dogs (76%-105) was higher than those of symptomatic (16.6%-23) and oligosymptomatic ones (7.4%-10). A total of 16 (11.6%) L. (L.) i. chagasi isolates were obtained from infected dogs, all from the IFAT( (+) ) /DTH( (-) ) profile: 41% (9/22) from symptomatic, 33.3% (3/9) from oligosymptomatic, and 5.2% (4/76) from asymptomatic dogs. These findings strongly suggested that despite the higher frequency of asymptomatic dogs (76%-105), the majority (72.4%-76) was characterized by the IFAT( (+) ) /DTH( (-) ) profile with a doubtful immunogenetic resistance against infection.

Leishmania sp. identification by PCR associated with sequencing of target SSU rDNA in paraffin-embedded skin samples stored for more than 30 years.

Paraffin-embedded samples commonly stored at educational and research institutions constitute tissues banks for follow-up or epidemiological studies; however, the paraffin inclusion process involves the use of substances that can cause DNA degradation. In this study, a PCR protocol was applied to identify Leishmania strains in 33 paraffin-embedded skin samples of patients with American cutaneous leishmaniasis. DNA was obtained by the phenol-chloroform protocol following paraffin removal and then used in PCR or nested PCR based on the nucleotide sequence of the small subunit ribosomal RNA (SSU rDNA). The amplicons obtained were cloned and sequenced to determine the single nucleotide polymorphism that distinguishes between different Leishmania species or groups. This assay allowed to distinguish organisms belonging to the subgenus Viannia and identify L. (Leishmania) amazonensis and L. (L.) chagasi of the Leishmania subgenus. Of the 33 samples, PCR and nested PCR identified 91% of samples. After sequencing the PCR product of 26 samples, 16 were identified as L. (L.) amazonensis, the other 10 contain organisms belonging to the L. (Viannia) sub-genus. These results open a huge opportunity to study stored samples and promote relevant contributions to epidemiological studies.

A prospective study on the dynamics of the clinical and immunological evolution of human Leishmania (L.) infantum chagasi infection in the Brazilian Amazon region.

This prospective study was carried out from October 2003 to December 2005 and involved a cohort of 946 individuals of both genders, aged 1-89 years, from an endemic area for American visceral leishmaniasis (AVL), in Pará State, Brazil. The aim of the study was to analyze the dynamics of the clinical and immunological evolution of human Leishmania (L.) infantum chagasi infection represented by the following clinical-immunological profiles: asymptomatic infection (AI); symptomatic infection (SI=AVL); subclinical oligosymptomatic infection (SOI); subclinical resistant infection (SRI); and indeterminate initial infection (III). Infection diagnosis was determined by the indirect fluorescent antibody test and leishmanin skin test. In total, 231 cases of infection were diagnosed: the AI profile was the most frequent (73.2%), followed by SRI (12.1%), III (9.9%), SI (2.6%) and SOI (2.2%). The major conclusion regarding evolution dynamics was that the III profile plays a pivotal role from which the cases evolve to either the resistant, SRI and AI, or susceptible, SOI and SI, profiles; only one of the 23 III cases evolved to SI, while most evolved to either SRI (nine cases) or SOI (five cases) and eight cases remained as III.

A cross-sectional study on the clinical and immunological spectrum of human Leishmania (L.) infantum chagasi infection in the Brazilian Amazon region.

The objectives of this study were to identify individuals with symptomatic and/or asymptomatic infection due to Leishmania (L.) infantum chagasi; to study the two types of infection, both clinically and immunologically, and to determine the prevalence rate of infection at the beginning of the study. This was a cross-sectional study with a cohort of 946 individuals, of both genders, from the age of 1 year, living in the municipality of Barcarena, PA, Brazil, an area endemic for American visceral leishmaniasis (AVL). The leishmanin skin test (LST) and the indirect fluorescent test (IFAT), were used for the diagnosis of infection. One hundred and twenty cases of infection were diagnosed, with a prevalence rate of 12.6%; eight cases showed high seroreactivity (1280-10240, IgG) in IFAT and no LST reaction; four of these cases were typical AVL and four had subclinical oligosymptomatic infection. Using two immunological methods with a clinical examination of the infected individuals enabled the identification of five clinical-immunological profiles which may promote a better understanding of the interaction between L. (L.) i. chagasi and the human immune response: asymptomatic infection (AI) 73.4%; subclinical resistant infection (SRI) 15%; subclinical oligosymptomatic infection (SOI) 3%; symptomatic infection (AVL) 3% and indeterminate initial infection (III) 5%.

A longitudinal study on the transmission dynamics of human Leishmania (Leishmania) infantum chagasi infection in Amazonian Brazil, with special reference to its prevalence and incidence.

This was a longitudinal study carried out during a period over 2 years with a cohort of 946 individuals of both sexes, aged 1 year and older, from an endemic area of American visceral leishmaniasis (AVL) in Pará State, Brazil. The object was to analyze the transmission dynamics of human Leishmania (Leishmania) infantum chagasi infection based principally on the prevalence and incidence. For diagnosis of the infection, the indirect fluorescent antibody test (IFAT) and leishmanin skin test (LST) were performed with amastigote and promastigote antigens of the parasite, respectively. The prevalence by LST (11.2%) was higher (p < 0.0001) than that (3.4%) by IFAT, and the combined prevalence by both tests was 12.6%. The incidences by LST were also higher (p < 0.05) than those by IFAT at 6 (4.7% x 0.6%), 12 (4.7% x 2.7%), and 24 months (2.9% x 0.3%). Moreover, there were no differences (p > 0.05) between the combined incidences by both tests on the same point surveys, 5.2%, 6.3%, and 3.6%. During the study, 12 infected persons showed high IFAT IgG titers with no LST reactions: five children and two adults developed AVL (2,560-10,120), and two children and three adults developed subclinical oligosymptomatic infection (1280-2560). The combined tests diagnosed a total of 231 cases of infection leading to an accumulated prevalence of 24.4%.

Reviewing the role of the dendritic Langerhans cells in the immunopathogenesis of American cutaneous leishmaniasis.

The role of dendritic Langerhans cells (LCs) in the immunopathogenesis of American cutaneous leishmaniasis (ACL) was reviewed in the light of more recent clinical and immunological features of ACL, caused by the principal human pathogenic leishmanial parasites found in Brazil: Leishmania (Viannia) braziliensis and L. (L.) amazonensis. The report shows a species-specific correlation between the LC density and the CD4+ and CD8+ T-cell profiles in the cellular infiltrate of skin lesions of ACL patients, providing the conclusion that LCs might be influencing the dichotomy of interaction between L. (V.) braziliensis and L. (L.) amazonensis with the human T-cell immune response. While L. (V.) braziliensis shows a clear tendency to direct infection to the hypersensitivity pole of the ACL clinical-immunological spectrum marked by a strong Th1-type immune response, L. (L.) amazonensis shows the opposite, directing infection to the hyposensitivity pole associated with a marked Th2-type immune response. These are probably the main immunological mechanisms of LCs regarding the immune response dichotomy that modulates infection outcome by these Leishmania parasites.

Revisão sobre a patogenia da leishmaniose tegumentar americana na Amazônia, com ênfase à doença causada por Leishmania (V. ) braziliensis e Leishmania (L. ) amazonensis / Revisiting the pathogenesis of the american tegumentary leishmaniasis in Amazonian, with emphasis to the disease due to Leishmania (V. ) braziliensis and Leishmania (L.) amazonensis

A patogenia da leishmaniose tegumentar americana (LTA) na Amazônia foi revisada à luz dos mais recentes aspectos associados ao espectro clínico, histopatológico e imunopatológico da doença causada por Leishmania (V.) braziliensis e Leishmania (L.) amazonensis. Esta revisão mostrou a existência de uma dicotomia entre as duas espécies de Leishmania e a resposta imune celular; enquanto a L. (V.) braziliensis mostra forte tendência em dirigir a infecção, a partir da forma central do espectro clínico-imunológico, a leishmaniose cutânea localizada (LCL), para o pólo imunológico hiperreativo, representado pela leishmaniose cutâneo-mucosa (LCM), com exacerbação da hipersensibilidade e perfil da resposta CD4 tipo-Thl, a L. (L.) amazonensis mostra o oposto, dirige a infecção para o pólo imunológico hiporreativo, representado pela leishmaniose cutânea anérgica difusa (LCAD), com forte inibição da hipersensibilidade e perfil da resposta CD4 tipo- Th2. Entre a forma central LCL e as formas polares LCM e LCAD a infecção passa por uma fase intermediária, a leishmaniose cutânea disseminada borderline (LCDB), com inibição parcial da hipersensibilidade e peifil da resposta CD4 Thl + Th2. Estes são, provavelmente, os principais mecanismos imunológicos que modulam a patogenia da LTA causada por L. (V.) braziliensis e L. (L.) amazonensis.

The pathogenesis of American tegumentary leishmaniasis (ATL) was reviewed ifl the light of more recent features of clinical, histopathological and immunopathological spectrum of disease caused by Leishmania (V.) braziliensis and Leishmania (L.) amazonensis. This review has shown a dichotomy in the interaction between these two species of Leishmania with the human .cellular immune response; while L. (V.) braziliensis shows a clear tendency to direct infection, from the localized cutaneous leishmaniasis (LCL) in the center of the clinical-immunological spectrum of disease, to the hyperactive immunologic pole represented by mucocutaneous leishmaniasis (MCL), which shows exacerbated hypersensitivity reaction and CD4 Thl-type immune response, L. (L.) amazonensis shows the opposite,. directing infection to the hypoactive immunologic pole consisted by anergic diffuse cutaneous leishmaniasis (ADCL), associated with a marked inhibition of hypersensitivity reaction and CD4 Th2type immune response. Between the central LCL and thetwo polar MCL and ADCL forms the infection may present an intermediary phase, borderline disseminated cutaneous leishmaniasis (BDCL), which shows partial inhibition of hypersensitivity reaction and a mixed CD4 Thl plus Th2 immune response. These are probably the main immunological mechanisms regarding the immune response dichotomy that modulates the pathogenesis of ATL caused by these Leishmania parasites.

Correlation between the components of the insulin-like growth factor I system, nutritional status and visceral leishmaniasis.

The role of the insulin-like growth factor I (IGF-I) system and nutritional status was studied in 241 children from a Brazilian area endemic for visceral leishmaniasis (VL). Thirty-nine children had the active form, 20 were oligosymptomatic, 38 were asymptomatic and 144 were not infected. Serum concentrations of growth hormone (GH), total and free IGF-I and IGF binding-protein 3 (IGFBP3) were measured by radioimmunoassay. Nutritional status was evaluated by anthropometric indicators and biochemical measurements. Total and free IGF-I and IGFBP3 were significantly reduced in the active form. Z scores for total and free IGF-I and for IGFBP3 were found to be significantly lower for active VL and oligosymptomatic individuals than for asymptomatic individuals, but never reached values

Experimental model of chronic osteomyelitis caused by Leishmania (L) amazonensis.

Experimental animal models have been used for the study of the physiopathogenesis of leishmaniasis, on some occasions with success, while in other situations such as bone alterations that accompany tegumentary leishmaniasis, especially in diffuse cutaneous form (DCL), the mechanisms are still unknown. In the present study, we determined these alterations in an animal model susceptible to Leishmania (L) amazonensis. Amastigotes of L. (L) amazonensis isolated from patients with diffuse cutaneous leishmaniasis (DCL) were inoculated into the hind paws of eight BALB/c mice, macroscopic and histopathological aspects were analyzed. After 90 and 120 days of evolution, histopathological analysis demonstrated a mononuclear cell infiltrate rich in plasma cells and intense parasitism of intra- and extra-medullary macrophages, with areas of bone necrosis and discrete involvement of cartilaginous tissue. The results show that the inflammatory process developed during L. (L) amazonensis infection might cause bone tissue destruction and secondarily affect the joints.