sCD163 levels as a biomarker of disease severity in leprosy and visceral leishmaniasis

Submitted by Ana Maria Fiscina Sampaio ([email protected]) on 2018-04-05T13:29:54Z No. of bitstreams: 1 Silva LR sCD163 levels as a biomarker of disease....pdf: 1813439 bytes, checksum: 61b593e582b8cc501964a9d141747a1a (MD5)Approved for entry into archive by Ana Maria Fiscina Sampaio ([email protected]) on 2018-04-05T13:58:17Z (GMT) No. of bitstreams: 1 Silva LR sCD163 levels as a biomarker of disease....pdf: 1813439 bytes, checksum: 61b593e582b8cc501964a9d141747a1a (MD5)Made available in DSpace on 2018-04-05T13:58:17Z (GMT). No. of bitstreams: 1 Silva LR sCD163 levels as a biomarker of disease....pdf: 1813439 bytes, checksum: 61b593e582b8cc501964a9d141747a1a (MD5) Previous issue date: 2017Fundação de Apoio à Pesquisa e à Inovação Tecnológica do Estado de Sergipe (FAPITEC - http://www.fapitec.se.gov.br/)/SE/FUNTEC/Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Grants: CNPq n.12/2009, Processo n. 019.203.02712/2009-8 (ARJ).Universidade Federal de Sergipe. Hospital Universitário. Laboratório de Biologia Molecular. Aracaju, SE, Brasil / Universidade Federal de Sergipe. Departamento de Educação em Saúde de Lagarto. Lagarto, SE, BrasilUniversidade Federal de Sergipe. Hospital Universitário. Laboratório de Biologia Molecular. Aracaju, SE, Brasil / Universidade Federal de Sergipe. Departamento de Educação em Saúde de Lagarto. Lagarto, SE, BrasilUniversidade Federal de Sergipe. Hospital Universitário. Laboratório de Biologia Molecular. Aracaju, SE, Brasil / Universidade Federal de Sergipe. Departamento de Educação em Saúde de Lagarto. Lagarto, SE, BrasilUniversidade Federal de Sergipe. Hospital Universitário. Laboratório de Biologia Molecular. Aracaju, SE, BrasilUniversidade Federal de Sergipe. Hospital Universitário. Laboratório de Biologia Molecular. Aracaju, SE, BrasilUniversidade Federal de Sergipe. Hospital Universitário. Laboratório de Biologia Molecular. Aracaju, SE, BrasilUniversidade Federal de Sergipe. Hospital Universitário. Laboratório de Biologia Molecular. Aracaju, SE, Brasil / Universidade Federal de Sergipe. Departamento de Educação em Saúde de Lagarto. Lagarto, SE, BrasilFundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, BA, BrasilUniversidade de São Paulo. Faculdade de Medicina de Ribeirão Preto. Departamento de Bioquímica e Imunologia. Ribeirão Preto, SP, BrasilHoward University. Department of Biology. Washington, DC, USAInfectious Diseases Research Institute. Seattle, WA, USAInfectious Diseases Research Institute. Seattle, WA, USAUniversidade Federal de Sergipe. Hospital Universitário. Laboratório de Biologia Molecular. Aracaju, SE, BrasilUniversidade Federal de Sergipe. Hospital Universitário. Laboratório de Biologia Molecular. Aracaju, SE, BrasilCD163, receptor for the haptoglobin-hemoglobin complex, is expressed on monocytes/macrophages and neutrophils. A soluble form of CD163 (sCD163) has been associated with the M2 macrophage phenotype, and M2 macrophages have been shown to down-modulate inflammatory responses. In particular, previous studies have shown that M2 is closely associated with the most severe clinical presentation of leprosy (i.e. lepromatous leprosy (LL)), as well as tuberculosis. We hypothesized that sCD163 correlates with severity of diseases caused by intracellular pathogens

sCD163 is elevated in the serum of lepromatous leprosy patients.

<p>(A) Sera of leprosy patients with various clinical presentations were collected and sCD163 concentrations measured by ELISA. Household contacts without symptoms or signs of leprosy (Contacts) were used as a control group. The mean ± SD sCD163 levels in patients with indeterminate leprosy (IL) (n = 9; 114 ± 49,57 ng/mL), true tuberculoid leprosy (TT) (n = 14; 90,29 ± 44,06 ng/mL), borderline leprosy (BL) (n = 14; 97,71 ± 47,97 ng/mL) and lepromatous leprosy (LL) (n = 10; 177,6 ± 62,18 ng/mL), as well as contacts (n = 23; 90,78 ± 31,55 ng/mL) were compared by Mann-Whitney test. ROC curves comparing sCD163 concentrations from TT versus LL (B) and Contacts versus LL patients (C) were constructed and are shown.</p

Susceptibility of Balb/c and C57BL/6 mice to ROCV infection.

<p>Mice were infected with different doses of ROCV (n = 10 mice per group). Survival rate and body weight loss for C57BL/6 (A and B) and for Balb/c (C and D). Comparison of mortality rate between C57BL/6 and Balb/c (Fig 1E). Survival rate and body weight loss were measured for up to 21 days post-infection. Statistically significant differences in B and D: ****p<0.0001 was determined by one-way ANOVA with Dunnett's multiple comparisons test, in E: *p<0.01 by Log-rank (Mantel-Cox) test.</p

CCR2 plays a protective role in Rocio virus–induced encephalitis by promoting macrophage infiltration into the brain

Rocio virus (ROCV) is a highly neuropathogenic mosquito-transmitted flavivirus responsible for an unprecedented outbreak of human encephalitis during 1975-1976 in Sao Paulo State, Brazil. Previous studies have shown an increased number of inflammatory macrophages into the central nervous system (CNS) of ROCV-infected mice, implying a role for macrophages in the pathogenesis of ROCV. Here, we showed that ROCV infection results in increased expression of C-C chemokine ligand 2 (CCL2) in the blood and in infiltration of macrophages into the brain. Moreover, we showed using C-C chemokine receptor 2 (CCR2) knockout mice that CCR2 expression was essential for macrophage infiltration in the brains during ROCV infection and that the lack of CCR2 resulted in increased disease severity and mortality. Thus, our findings show the protective role of CCR2-mediated infiltration of macrophages in the brain during ROCV infection

CD163 expression is induced by <i>Leishmania</i> infection of neutrophils.

<p>(A) Gating strategy for the analysis of neutrophil phenotypes. Neutrophils were purified from healthy donors and infected with GFP-expressing <i>L</i>. <i>amazonensis</i> (5 parasites: 1 neutrophil) in RPMI 1640 plus 10% FBS. (B) 3h after infection, neutrophils were characterized by flow cytometry and data analyzed by FlowJo software (in duplicate, n = 5 experiments). Green and black bars represent, respectively, the GFP positive and negative cells. The white bars represent non-exposed group (unstimulated). The mean ± SD of parental percentage of GFP+, GFP- and non-exposed cells were compared by Friedman paired test with Dunn’s post test.</p

Evaluation of cross-protective immunity against ROCV after prior infections with different flaviviruses.

<p>Immunization and challenge regime (A). Mice (n = 40 per group) were infected twice with the different flaviviruses known to circulate in Brazil, and were then challenged (n = 20 per group) with 2.76x10⁷ PFU of ROCV and survival rates (B and E), clinical scores (C and F) and body weight loss (D and G) were determined up to 21 days post-infection. Statistically significant differences in B and E: *p<0.01, **p<0.001 and ****p<0.0001 was determined by Log-rank (Mantel-Cox) test. In F: ****p<0.0001 was analyzed by a student t-test. In D-G *p<0.01, **p<0.001 and ****p<0.0001 was determined by One-way ANOVA with Dunnett's multiple comparisons test. All statistically significant differences were with group control (Mock).</p

Variable cytokine production by CD163+ and CD163- monocyte/macrophages.

<p>(A) Gating strategy for analysis of the cytokine profiles of CD163+ and CD163- monocyte/macrophages. PBMC were collected from healthy donors and the adherent cells cultured for 5 days in RPMI 1640 plus 20% FBS. The cells were incubated with <i>L</i>. <i>amazonensis</i> strain (10 parasites: 1 macrophage) and <i>L</i>. <i>infantum-</i>isolate 1 (5:1), and incubated with antibodies specific for intracellular cytokines, prior to analysis by flow cytometry (n = 6 experiments, in duplicate). (B) Frequency of IL-4+ cells, (C) MFI of IL-4-PerCPCy5.5, (D) iMFI of IL-4 analysis, (E) Frequency of IL-10+ cells, (F) MFI of IL-10-APC, (G) iMFI of IL-10 analysis, (H) Frequency of IL-12+ cells, (I) MFI of IL-12-APC, (J) iMFI of IL-12 analysis, (K) Frequency of TNF-α+ cells, (L) MFI of TNF-α-PerCPCy5.5, (M) iMFI of TNF-α analysis.</p

sCD163 levels correlate with severity of VL.

<p>(A) sCD163 levels were measured in sera of VL patients of different clinical status. The mean ± SD sCD163 levels of patients with classical VL at D0 (D0-Classic, n = 33) (152,1 ± 67,86 ng/mL), D30 (n = 19) (98,79 ± 58,58 ng/mL) and of patients of severe VL at D0 (D0-SVL) (n = 13) (241,5 ± 76,88 ng/mL) were compared by Mann-Whitney test. Sera from <i>Leishmania</i>-infected individuals without symptoms or signs of VL (DTH+, n = 11, 72,55 ± 25,68 ng/mL) and healthy individuals from non-endemic regions (HC, n = 8, 49,0 ± 23,71 ng/mL) were included as control groups. Spearman correlation analyses between sCD163 concentrations were performed versus (B) spleen size, (C) liver size and (D) neutrophil count. (E) Paired analysis of sCD163 levels of VL patients before and after treatment (n = 15, p = 0.0455, paired t test). ROC curves of sCD163 concentration comparing HC versus (F) D0-Classic, (G) D0-Classic versus D30 and (H) D0 versus D0-SVL group.</p