Whole blood transcriptomics reveals the enrichment of neutrophil activation pathways during erythema nodosum leprosum reaction.

Introduction: Patients with the multibacillary form of leprosy can develop reactional episodes of acute inflammation, known as erythema nodosum leprosum (ENL), which are characterized by the appearance of painful cutaneous nodules and systemic symptoms. Neutrophils have been recognized to play a role in the pathogenesis of ENL, and recent global transcriptomic analysis revealed neutrophil-related processes as a signature of ENL skin lesions. Methods: In this study, we expanded this analysis to the blood compartment, comparing whole blood transcriptomics of patients with non-reactional lepromatous leprosy at diagnosis (LL, n=7) and patients with ENL before administration of anti-reactional treatment (ENL, n=15). Furthermore, a follow-up study was performed with patients experiencing an ENL episode at the time of diagnosis and after 7 days of thalidomide treatment (THAL, n=10). Validation in an independent cohort (ENL=8; LL=7) was performed by RT-qPCR. Results: An enrichment of neutrophil activation and degranulation-related genes was observed in the ENL group, with the gene for the neutrophil activation marker CD177 being the most enriched gene of ENL episode when compared to its expression in the LL group. A more pro-inflammatory transcriptome was also observed, with increased expression of genes related to innate immunity. Validation in an independent cohort indicated that S100A8 expression could discriminate ENL from LL. Supernatants of blood cells stimulated in vitro with Mycobacterium leprae sonicate showed higher levels of CD177 compared to the level of untreated cells, indicating that the leprosy bacillus can activate neutrophils expressing CD177. Of note, suggestive higher CD177 protein levels were found in the sera of patients with severe/moderate ENL episodes when compared with patients with mild episodes and LL patients, highlighting CD177 as a potential systemic marker of ENL severity that deserves future confirmation. Furthermore, a follow-up study was performed with patients at the time of ENL diagnosis and after 7 days of thalidomide treatment (THAL, n=10). Enrichment of neutrophil pathways was sustained in the transcriptomic profile of patients undergoing treatment; however, important immune targets that might be relevant to the effect of thalidomide at a systemic level, particularly NLRP6 and IL5RA, were revealed. Discussion: In conclusion, our study reinforces the key role played by neutrophils in ENL pathogenesis and shed lights on potential diagnostic candidates and novel therapeutic targets that could benefit patients with leprosy.

Genome-Wide Screening of mRNA Expression in Leprosy Patients.

Leprosy, an infectious disease caused by Mycobacterium leprae, affects millions of people worldwide. However, little is known regarding its molecular pathophysiological mechanisms. In this study, a comprehensive assessment of human mRNA was performed on leprosy skin lesions by using DNA chip microarrays, which included the entire spectrum of the disease along with its reactional states. Sixty-six samples from leprotic lesions (10TT, 10BT, 10BB, 10BL, 4LL, 14R1, and 10R2) and nine skin biopsies from healthy individuals were used as controls (CC) (ages ranged from 06 to 83 years, 48 were male and 29 female). The evaluation identified 1580 differentially expressed mRNAs [Fold Change (FC) ≥ 2.0, p ≤ 0.05] in diseased lesions vs. healthy controls. Some of these genes were observed in all forms of the disease (CD2, CD27, chit1, FA2H, FAM26F, GZMB, MMP9, SLAMF7, UBD) and others were exclusive to reactional forms (Type "1" reaction: GPNMB, IL1B, MICAL2, FOXQ1; Type "2" reaction: AKR1B10, FAM180B, FOXQ1, NNMT, NR1D1, PTX3, TNFRSF25). In literature, these mRNAs have been associated with numerous pathophysiological processes and signaling pathways and are present in a large number of diseases. The role of these mRNAs maybe studied in the context of developing new diagnostic markers and therapeutic targets for leprosy.

A doença de Jorge Lobo e a coloraçäo pele prata / Jorge Lobo's disease and methenamine silver staining

FUNDAMENTOS - A análise dos cortes histológicos de lesöes cutâneas corados pela prata metenamina (PM) e hematoxilina-eosina (HE) revelam diferenças na coloraçäo do P.loboi, agente causador da doença de Jorge Lobo. Nos cortes corados pela PM observa-se também depósito de uma substância granulosa, de tonalidade marrom-escura, semelhante àquela exibida pelo parasita. OBJETIVOS - Interpretar as alteraçöes da coloraçäo do P.loboi corado pela PM em cortes histológicos e suspensöes fúngicas e comparar os resultados com os obtidos por meio da coloraçäo vital pelo diacetato de fluoresceína-brometo de etídio (DF-BE). Interpretar, também, a natureza do material granuloso e argentófilo observado em algumas localizaçöes das lesöes cutâneas. PACIENTE E MÉTODOS - Biópsias da pele de cinco pacientes foram processadas em soluçäo salina 0,85 porcento e incluídas em parafina para obtençäo de cortes histológicos. A suspensäo de fungos foi corada pelo DF-BE e PM, e os cortes histológicos, pela HE e PM com a finalidade de avaliar a viabilidade dos fungos. RESULTADOS - Os percentuais de viabilidade do P.loboi em suspensäo variaram de 37 a 51 porcento e de 39 a 59 porcento com os corantes DF-BE e PM, respectivamente. Nos cortes histológicos corados pela PM, os percentuais variaram de 36 a 57 porcento. Observou-se material granuloso argentófilo em todos os cortes analisados, tanto no interior de células em que havia grande número de cápsulas vazias quanto em áreas com número de fungos reduzido. CONCLUSÖES - Os resultados obtidos sugerem que, por meio da coloraçäo dos cortes histológicos pela PM, é possível estimar o índice de viabilidade do P.loboi, embora a coloraçäo dos fungos em suspensäo pelo DF-BE seja método mais preciso. As observaçöes feitas também sugerem que a presença do material argentófilo está relacionada com a destruiçäo do P.loboi no interior dos macrófagos