BACKGROUND Fibrosis in the peripheral nerve is the end stage of leprous neuropathy and the cause of the resulting permanent neural function impairments. Preventive measures to avoid this irreversible pathological state are a relief strategy for leprosy sufferers. OBJECTIVES The present study describes the frequency of fibrosis along with its characterisation and pathogenic development. METHODS Six-hundred-and-thirteen nerve samples were sorted from 278 neural leprosy (NL) and 335 non-leprosy neuropathy patients (ON). The total number of samples was histologically examined by routine staining methods (haematoxylin-eosin, Wade staining and Gomori’s trichrome) and fibrosis was evaluated via semi-quantitative estimation. FINDINGS Fibrosis was most frequent in the NL group (33% against 0.4% in ON) while fibrosis in association with endoneurial microfasciculation was found in 38 (41.3%) of the NL samples in the examination of semithin sections. Pericytic activation in the perivascular environment was confirmed to be the source of the fibroblasts and perineurial cells delimiting microfascicles. End-stage fibrosis in leprosy displays an arrangement of microfascicles devoid of neural components (i.e., Schwann cells and axons) lined by an intermediate phenotype of fibroblastic-perineurial cells filled with bundles of collagen fibres. MAIN CONCLUSIONS The present study underscores that fibrosis is frequently the severe end stage of neural leprosy NL pathogeny after analysing the notably distinct development of fibrosis within the neural environment.

The clinical course of leprosy is often interrupted by reactions, which are acute inflammatory episodes that can be classified as type I or type II. Type II reactions can present as cutaneous lesions that resemble erythema multiforme (EM). EM is classically associated with drug allergies or pre-existing viral infections. However, the differential diagnostic criteria of the diverse causative agents remain controversial. The aim of this study was to determine both the clinical relevance and the morphological and immunohistochemical characteristics of the EM-like lesions during the course of type II leprosy reactions. Twenty-seven skin biopsies were taken from typical EM-like lesions of multibacillary patients and reviewed; their histological features were correlated to their clinical aspects. Then, a computer-assisted morphometric analysis was performed to measure the extent of angiogenesis during these acute episodes. The histopathological and immunohistochemical analysis of the EM lesions revealed that they shared the same features that have been previously described for ENL, including immunopositivity in the identical cell-mediated immune markers. Our results point to leprosy as the cause of the EM-like lesions in our patients. Therefore, leprosy should be considered in the differential diagnosis of EM.

It is important to understand the mechanisms that enable peripheral neurons to regenerate after nerve injury in order to identify methods of improving this regeneration. Therefore, we studied nerve regeneration and sensory impairment recovery in the cutaneous lesions of leprosy patients (LPs) before and after treatment with multidrug therapy (MDT). The skin lesion sensory test results were compared to the histopathological and immunohistochemical protein gene product (PGP) 9.5 and the p75 nerve growth factor receptors (NGFr) findings. The cutaneous neural occupation ratio (CNOR) was evaluated for both neural markers. Thermal and pain sensations were the most frequently affected functions at the first visit and the most frequently recovered functions after MDT. The presence of a high cutaneous nerve damage index did not prevent the recovery of any type of sensory function. The CNOR was calculated for each biopsy, according to the presence of PGP and NGFr-immunostained fibres and it was not significantly different before or after the MDT. We observed a variable influence of MDT in the recovery from sensory impairment in the cutaneous lesions of LPs. Nociception and cold thermosensation were the most recovered sensations. The recovery of sensation in the skin lesions appeared to be associated with subsiding inflammation rather than with the regenerative activity of nerve fibres.

Nerve biopsy examination is an important auxiliary procedure for diagnosing pure neural leprosy (PNL). When acid-fast bacilli (AFB) are not detected in the nerve sample, the value of other nonspecific histological alterations should be considered along with pertinent clinical, electroneuromyographical and laboratory data (the detection of Mycobacterium leprae DNA with polymerase chain reaction and the detection of serum anti-phenolic glycolipid 1 antibodies) to support a possible or probable PNL diagnosis. Three hundred forty nerve samples [144 from PNL patients and 196 from patients with non-leprosy peripheral neuropathies (NLN)] were examined. Both AFB-negative and AFB-positive PNL samples had more frequent histopathological alterations (epithelioid granulomas, mononuclear infiltrates, fibrosis, perineurial and subperineurial oedema and decreased numbers of myelinated fibres) than the NLN group. Multivariate analysis revealed that independently, mononuclear infiltrate and perineurial fibrosis were more common in the PNL group and were able to correctly classify AFB-negative PNL samples. These results indicate that even in the absence of AFB, these histopathological nerve alterations may justify a PNL diagnosis when observed in conjunction with pertinent clinical, epidemiological and laboratory data.

O efeito das reações hansenianas sobre a inervação cutânea de pacientes hansenianos foi avaliado em seis pacientes (três com reação reversa e três com eritema nodoso leprosum). Cortes congelados de biópsias de lesões cutâneas reacionais colhidas na ocasião da reação e de biópsias colhidas após a remissão do quadro reacional na mesma região ocupada previamente pela lesão foram marcados pela técnica de imunofluorescência indireta utilizando os anticorpos anti-NGFr e anti-PGP 9.5. Não foi encontrada diferença significativa na quantificação de fibras positivas para NGFr e para PGP 9.5 entre as biópsias colhidas durante a reação e as biópsias colhidas no período de remissão. Entretanto, no grupo de biópsias da reação houve uma significativa diferença entre a quantidade de fibras NGFr-positivas e as fibras imunomarcadas para PGP 9.5. Essa diferença contudo foi atribuída aos diferentes aspectos que a mesma fibra pode assumir quando marcadas com NGFr ou com PGP 9..5 separadamente. O presente estudo também mostrou que a avaliação das condições das fibras nervosas de um órgão deve ser realizada com marcadores para o axônio e para células de Schwann.

The effects of reactional episodes on the cutaneous nerve fibers of leprosy patients was assessed in six patients (three with reversal reactions and three with erythema nodosum leprosum). Cryosections of cutaneous biopsy of reactional lesions taken during the episode and of another sample during the remission period were immunostained with anti-NGFr and anti-PGP 9.5 (indirect immunofluorescence). We found no significant statistical difference in the number of NGFr- and PGP 9.5-positive fibers between the reactional and post-reactional groups. A significant difference was detected between the number of NGFr and PGP 9.5-stained fibers inside of the reactional group of biopsy cryosections but this difference was ascribed to the distinct aspects of the nerve fibers displayed whether stained with anti-NGFr or with anti-PGP 9.5; NGFr-positive branches looked larger and so interpreted as containing more fibers. In addition, a substantial number NGFr-positive fibers were PGP 9.5-negative. No differences in the number of stained fibers among the distinct cutaneous regions examined (epidermis + upper dermis, mid and deep dermis) was detected. In conclusion, the number of PGP- and NGFr-positive fibers were not significantly different in the reactional and post-reactional biopsies in the present study. NGFr-staining of the nerve fibers is different from their PGP-imunoreactivity and the evaluation of the nerve fiber status on an innervated target organ should be carried out choosing markers for both components of nerve fibers (Schwann cells and axons).

A marcação imuno-histoquímica dos neuropeptídios (SP, alfa-MSH e gama-MSH, CGRP, VIP) e a imunomarcação de mastócitos e suas subpopulações triptase-positivas e triptase/quimase positivas foram realizadas em biópsias cutâneas de seis pacientes com reações hansenianas (três reações reversas e três eritema nodosos). A imunorreatividade nessas biópsias foi comparada com a obtida nas biópsias após a remissão das reações. Foi encontrado aumento relativo de mastócitos triptase-positivos no interior do infiltrado inflamatório das biópsias reacionais quando comparadas com as biópsias pós-reacionais. Além disso, o número total de mastócitos e o número relativo de mastócitos triptase-positivos foi significativamente maior no interior do infiltrado inflamatório do que na derme não ocupada pelo infiltrado. Em relação aos neuropeptídios não foram encontradas diferenças significativas na imunomarcação dos dois grupos de biópsias. O aumento relativo de mastócitos triptase-positivos sugere que eles possam estar implicados na deflagração do processo reacional. O papel dos neuropeptídios na gênese das reações ainda não pode ser descartado.

The immunohistochemical identification of neuropeptides (calcitonin gene-related peptide, vasoactive intestinal polypeptide, substance P, alpha-melanocyte stimulating hormone and gamma-melanocyte stimulating hormone) quantification of mast cells and their subsets (tryptase/chymase-immunoreactive mast cells = TCMC and tryptase-immunoreactive mast cells = TMC) were determined in biopsies of six patients with leprosy reactions (three patients with type I reaction and three with type II). Biopsies were compared with those taken from the same body site in the remission stage of the same patient. We found a relative increase of TMC in the inflammatory infiltrate of the reactional biopsies compared to the post-reactional biopsy. Also, the total number of mast cells and the TMC/TCMC ratio in the inflammatory infiltrate was significantly higher than in the intervening dermis of the biopsies of both periods. No significant difference was found regarding neuroptide expression in the reactional and post-reactional biopsies. The relative increase of TMC in the reactional infiltrates could implicate this mast cell subset in the reported increase of the immune response in leprosy reactions.

Quarenta e quatro pacientes com sinais de neuropatia periférica foram acompanhados no Ambulatório de Hanseníase da Fundação Oswaldo Cruz por período que variou de 4 meses até 4 anos, com o intuito de confirmar ou afastar o diagnóstico de neuropatia hanseniana. Os sintomas neurológicos apresentados foram hipoestesia (41), parestesia (28), espessamento neural (22), dor nos nervos (20), paresia (20), amiotrofia (8). Dez pacientes dos 44 tiveram o diagnóstico de hanseníase confirmado. A confirmação diagnóstica se deu através da biópsia de áreas hipoestésicas sem lesão dermatológica (3 pacientes), pelo aparecimento de estados reacionais (duas reações reversas e 4 neurites reacionais) e pelo aparecimento de lesão cutânea não reacional característica da forma "borderline" lepromatosa. O acompanhamento clínico regular dos pacientes sem diagnóstico no primeiro exame mostrou ser um método que permitiu o diagnóstico da forma neurítica pura da hanseníase, quando os métodos objetivos de diagnóstico não confirmarem de imediato a doença.

Forty-four patients with neuritic leprosy were individually followed for periods ranging from 4 months to almost 4 years for the purpose of ascertaining the presence and/ or absence of leprosy. The neural symptoms presented were sensory impairment (41), parasthesia (28), nerve enlargement (22), nerve tenderness (20), paresia (20), amyotrophy (8). Leprosy was diagnosed in ten out of the total number of patients studied. Leprosy was confirmed by the appearance of reactional neuritis (4), reversal reaction (2), biopsy of the hypoesthesic area (3) and the appearance of non-reactional cutaneous lesion. We reported an experience in the diagnosis of neuritic leprosy and its most frequent clinical presentation with which clinicians have to be acquainted. We can also state that the clinical follow-up was an effective strategy for the diagnosis of the disease when diagnostic facilities are not available or have not confirmed the diagnosis.