Dapsone-induced hepatic complications: it's time to think beyond methemoglobinemia.

Drug-induced liver injury is an important cause of hepatotoxicity and poses a challenging clinical problem with respect to both diagnosis and management. Patients susceptible to hepatotoxicity on exposure to dapsone is constantly on the rise. Dapsone (4,4'-diaminodiphenylsulfone) is clinically used alone or in combination with rifampicin for the treatment of a variety of dermatological disorders such as acne, dermatitis herpetiformis, psoriasis, Toxoplasma gondii infections, leprosy and pneumocystis carinii pneumonia in AIDS patients. However, the clinical use of dapsone is limited because of dose-dependent adverse hematological reactions. The cholestatic injury caused by dapsone and its N- hydroxylated metabolites hinders bile flow and causes oxidative stress and hepatic necrosis, further, leading to hemolysis responsible for hepatitis due to iron overload in the liver. Hence, clinicians' awareness of the hepatotoxic potential of dapsone is highly warranted.

Hematological alterations in lepromatous leprosy: A cross-sectional observational study.

BACKGROUND: Dapsone has been the mainstay for the treatment of leprosy since its discovery in the 1940s. However, hematological disturbances are not uncommon in leprosy patients on daily dapsone therapy. Hence, the present study was conducted to document the hematologic alterations observed in lepromatous leprosy patients treated with Dapsone 100 mg daily. METHODOLOGY: A cross-sectional observational study was conducted amongst 32 lepromatous leprosy patients treated with Dapsone 100 mg daily. A complete hemogram was conducted for all the study recruits. The test results were compared against the standard average values for adults for the given variables. The one sample t-test was employed to compare the difference between the study values and the standard normal values for adults. The statistical significance was considered at p < 0.05. RESULTS: The study reveals a marked decrease in hemoglobin concentration in patients on dapsone, 100 mg daily. Other hematological alterations found were reduced platelet count, reduced mean platelet volume, reduced Hematocrit, reduced Mean Corpuscular hemoglobin, reduced Mean Corpuscular hemoglobin concentration. (p < 0.05). CONCLUSION: Treatment of lepromatous leprosy with 100 mg daily Dapsone therapy may lead to hematological alterations. These findings are suggestive of dapsone-induced hemolysis.

Antarctic rocks from continental Antarctica as source of potential human opportunistic fungi.

We assessed the diversity of culturable fungi associated with rocks of continental Antarctica to evaluate their physiological opportunistic virulence potential in vitro. The seventy fungal isolates obtained were identified as nine species of Acremonium, Byssochlamys, Cladosporium, Debaryomyces, Penicillium, and Rhodotorula. Acremonium sp., D. hansenii, P. chrysogenum, P. citrinum, P. tardochrysogenum, and R. mucilaginosa were able to grow at 37 °C; in addition, B. spectabilis displayed a high level of growth at 37 and 45 °C. Thirty-one isolates of P. chrysogenum, P. citrinum, and P. tardochrysogenum were able to produce partial haemolysis on blood agar at 37 °C. Acremonium sp., P. citrinum, and P. tardochrysogenum showed spore sizes ranging from 2.81 to 5.13 µm diameters at 37 °C. Of these, P. chrysogenum and P. tardochrysogenum displayed macro- and micro morphological polymorphism. Our results suggest that rocks of the ultra-extreme cold and dry environment of Antarctica harbour cryptic fungi phylogenetically close to opportunistic pathogenic and mycotoxigenic taxa with physiologic virulence characteristics in vitro.

Hemolytic anemia in patients receiving daily dapsone for the treatment of leprosy.

INTRODUCTION: Multidrug therapy for leprosy is currently done with dapsone, clofazimine and rifampicin. Dapsone is known to cause hemolytic anemia (HA) and this adverse event during MDT seems to be more frequent than reported. The aim of this report is to discuss and grade HA due to dapsone during MDT treatment for leprosy. METHODS: This is a retrospective study of 194 leprosy patients from a Leprosy Control Programme Unit in Vit6ria-ES, Brazil. RESULTS: HA was observed in 48 (24.7%) patients and occurred within the first 3 months in 51% of these. Mean hematocrit levels fell from 38.5 to 31.5 and hemoglobin from 12.8 to 10.3. CONCLUSION: Dapsone used in the MDT regime for leprosy decreases the hematocrit and hemoglobin levels due to a low grade hemolysis, which can result in significant anemia.

The functional state of the complement system in leprosy.

Ninety-one patients with different clinical forms of leprosy, 36 lepromatous (LL), 33 tuberculoid (TL), and 22 dimorphic (DL), and 31 healthy volunteer donors were included in this study. Total complement system (CS) activity was assessed by hemolytic methods, whereas individual components were quantified by the enzyme-linked immunosorbent assay. Under conditions allowing initiation of cascade by the classic pathway (CP) but not alternative pathway (AP) activation, significant CS consumption was detected only in sera from patients with LL. In this group of patients, C4 but not factor B (fB) or C3 was significantly reduced, whereas mannose-binding lectin (MBL) serum levels were significantly higher. These results indicate that the CP is involved in CS activation in patients infected with Mycobacterium leprae manifesting LL clinical form of leprosy. An association is likely between circulating immune complexes and MBL high serum levels for initiation of CS activation in patients with LL form of leprosy.

A case of post-partum borderline tuberculoid leprosy complicated by a median nerve abscess, peptic ulceration and rifampicin-induced haemolytic renal failure.

We report a case of borderline tuberculoid leprosy complicated by a median nerve abscess, acute renal failure secondary to rifampicin-induced haemolysis and duodenal ulceration secondary to steroid use. Rifampicin induced hameolysis is a rare and probably under-reported complication of leprosy multi-drug therapy. It should be considered when patients complain of flu-like symptoms after taking their monthly rifampicin.

Effect of listeriolysin O-loaded erythrocytes on Mycobacterium avium replication within macrophages.

OBJECTIVE: To evaluate the efficacy of erythrocytes loaded with the haemolytic toxin listeriolysin O against Mycobacterium avium replication within human macrophages. METHODS: Recombinant listeriolysin O was loaded in human erythrocytes by a procedure of hypotonic dialysis and isotonic resealing. Loaded erythrocytes were modified to allow them to be recognized and taken up by human macrophages infected with M. avium. The antimycobacterial activity of the erythrocytes loaded with listeriolysin O was evaluated by supernatant and intracellular cfu counts on days 4 and 7 post-erythrocyte administration. RESULTS: Recombinant listeriolysin O was encapsulated in human erythrocytes to reach final concentrations ranging from 1 to 4 ng/mL of erythrocytes. Erythrocytes loaded with increasing quantities of recombinant protein were able to reduce (at most by 50%) M. avium replication in a dose-dependent fashion when administered to infected macrophages. CONCLUSIONS: Erythrocytes loaded with listeriolysin O are effective against M. avium replication within macrophages. We are confident that the strategy presented could be useful against mycobacteria other than M. avium (such as Mycobacterium tuberculosis and Mycobacterium leprae) by itself or as part of an antimycobacterial treatment.

Identification and mutagenesis by allelic exchange of choE, encoding a cholesterol oxidase from the intracellular pathogen Rhodococcus equi.

The virulence mechanisms of the facultative intracellular parasite Rhodococcus equi remain largely unknown. Among the candidate virulence factors of this pathogenic actinomycete is a secreted cholesterol oxidase, a putative membrane-damaging toxin. We identified and characterized the gene encoding this enzyme, the choE monocistron. Its protein product, ChoE, is homologous to other secreted cholesterol oxidases identified in Brevibacterium sterolicum and Streptomyces spp. ChoE also exhibits significant similarities to putative cholesterol oxidases encoded by Mycobacterium tuberculosis and Mycobacterium leprae. Genetic tools for use with R. equi are poorly developed. Here we describe the first targeted mutagenesis system available for this bacterium. It is based on a suicide plasmid, a selectable marker (the aacC4 apramycin resistance gene from Salmonella), and homologous recombination. The choE allele was disrupted by insertion of the aacC4 gene, cloned in pUC19 and introduced by electroporation in R. equi. choE recombinants were isolated at frequencies between 10(-2) and 10(-3). Twelve percent of the recombinants were double-crossover choE mutants. The choE mutation was associated with loss of cooperative (CAMP-like) hemolysis with sphingomyelinase-producing bacteria (Listeria ivanovii). Functional complementation was achieved by expression of choE from pVK173-T, a pAL5000 derivative conferring hygromycin resistance. Our data demonstrate that ChoE is an important cytolytic factor for R. equi. The highly efficient targeted mutagenesis procedure that we used to generate choE isogenic mutants will be a valuable tool for the molecular analysis of R. equi virulence.

Serious side effects of rifampin on the course of WHO/MDT: a case report.

A male born in 1935 was diagnosed as having lepromatous leprosy when he was 17 years old. In addition to dapsone (DDS) monotherapy, he had been treated with rifampin (RMP) for 2 terms: first with 450 mg a day for 2 years when he was 39 years old; second with 150 mg a day for 2 months after a 1-year interval from the first regimen. During these entire courses with RMP, no complication was noted. When he was 64 years old in 1999, a diagnosis of relapsed borderline tuberculoid (BT) leprosy was made, and he was started on the multibacillary (MB) regimen of the World Health Organization multidrug therapy (WHO/MDT). After the third dose of monthly RMP, he developed a flu-like syndrome and went into shock. A few hours later, intravascular hemolysis occurred followed by acute renal failure. He was placed on hemodialysis for 7 series and recovered almost completely about 2 months later. The immune complexes with anti-RMP antibody followed by complement binding may have accounted for these symptoms. Twenty-four reported cases of leprosy who had developed side effects of RMP under an intermittent regimen were analyzed; 9 of the cases had had prior treatment with RMP but 15 had not. Adverse effects were more likely to occur in MB cases and were more frequent during the first 6 doses of intermittent regimens. The cases with prior treatment with RMP had had a higher incidence of serious complications such as marked hypotension, hemolysis and acute renal failure. However, many exceptions were also found, and we could not verify any fully dependable factor(s) to predict the side effects of RMP. More field investigation is desirable, and monthly administration of RMP must be conducted under direct observation through the course of WHO/MDT.

Avaliacao de provas laboratoriais utilizadas para detectar deficiencias da glicose-6-fosfato desidrogenase (G6PD) no sangue total / Evaluation of laboratory tests used to detect glucose-6-phosphate dehydrogenase (G6PD) deficiencies in whole blood

A deficiência do G6PD é frequente entre brasileiros. A exposição de seus portadores a determinados medicamentos pode desencadear hemolise. Sendo assim, conveniente uma pré-determinação da G6PD antes do tratamento com medicamentos oxidantes. Avaliamos duas metodologias, para detectar pacientes de G6PD no sangue. A técnica Qualitativa, apesar de mais pratica e econômica não apresenta resultados precisos e exatos, servindo apenas como triagem. Enquanto a técnica Quantitativa, mesmo sendo de custo elevado, dificultando a sua aquisição, apresenta vantagens para determinar a atividade enzimática com maior exatidão.

Hemolytic drugs aniline and dapsone induce iron release in erythrocytes and increase the free iron pool in spleen and liver.

Incubation of rat erythrocytes with the hydroxylated metabolites of aniline and dapsone (4-4'-diaminodiphenylsulfone), phenylhydroxylamine and dapsone hydroxylamine, respectively, induced marked release of iron and methemoglobin formation. On the contrary, no release of iron nor methemoglobin formation was seen when the erythrocytes were incubated with the parent compounds (aniline and dapsone). The acute intoxication of rats with aniline or dapsone induced a marked increase in the erythrocyte content of free iron and methemoglobin, indicating that the xenobiotics are effective only after biotransformation to toxic metabolites in vivo. Prolonged administration of aniline or dapsone to rats produced continuous release of iron from erythrocytes. Marked iron overload was seen in the spleen and in the liver Kupffer cells, as detected histochemically. The spleen weight in these subchronically treated animals was significantly increased. The free iron pool was markedly increased in the spleen and to a lower extent in the liver. The possible relationships between iron release in erythrocytes, oxidative damage seen in senescent cells, hemolysis, overwhelmed capacity of spleen and liver to keep iron in storage forms and subsequent increase in low molecular weight, catalitically active iron is discussed.

Characterization of a haemolysin from Mycobacterium tuberculosis with homology to a virulence factor of Serpulina hyodysenteriae.

Scrutiny of sequence data from the Mycobacterium leprae genome sequencing project identified the presence of a gene encoding a 268-amino-acid polypeptide which is highly similar to a pore-forming haemolysin/cytotoxin virulence determinant, TlyA, from the swine pathogen Serpulina hyodysenteriae. Using degenerate oligonucleotide primers based on the TlyA sequences, the Mycobacterium tuberculosis homologue was amplified and this product was used to obtain the clone and sequence a 2.5 kb fragment containing the whole M. tuberculosis tlyA gene. tlyA encodes a 267-amino-acid protein with a predicted molecular mass of 28 kDa. TlyA homologues were identified by PCR in M. leprae, Mycobacterium avium and Mycobacterium bovis BCG, but appeared absent in Mycobacterium smegmatis, Mycobacterium vaccae, Mycobacterium kansasii, Mycobacterium chelonae and Mycobacterium phlei. The M. tuberculosis gene appeared to be the first gene in an operon containing at least two other genes. Introduction of the M. tuberculosis tlyA gene into M. smegmatis using a mycobacterial shuttle expression plasmid converted non-haemolytic cells into those exhibiting significant haemolytic activity. Similarly, inducible haemolytic activity was observed in sonicated bacteria when tlyA was expressed as a His6-tagged fusion protein in Escherichia coli. tlyA mRNA was detected in both M. tuberculosis and M. bovis BCG using RT-PCR, confirming that this gene is expressed in organisms cultured in vitro.

[Vitamin E as protective agent against hemolysis in leprosy patients under dapsone treatment]. / Vitamina E como agente protector de hemólisis en pacientes de lepra tratados con dapsona.

Dapsone (4,4'diaminodiphenyl-sulphone) commonly used in the treatment of patients who suffer from leprosy, is a strongly oxidative drug, producing damage to the red cell membrane. This study investigated whether Vitamin E would have a protective effect on the red cell membrane from oxidant damage caused by Dapsone in patients with leprosy. We have studied 16 patients for 4 months, divided into two groups. Group 1 (n = 7) dapsone (DDS): 100 mg/day; Group 2 (n = 9) dapsone: 100 mg/day in addition with Vitamin E: 800 U/day. We did not include patients with low levels of Glucose-6-Phosphate Dehydrogenase (G-6-PD) because of their sensibility to this drug. At the beginning of the treatment we determined the level of G-6-PD. All patients showed a normocytic normochromic anemia with a decrease in Haptoglobine levels (below 5 mg/dl). Statistical analyses showed that reticulocyte counts did not present significant differences between groups all through evolution. As for methemoglobin (Hi) we observed in Group 1 an increase between the first and the fourth month, which was not seen in group 2. Statistical analyses of the results suggest that oral Vitamin E confers partial protective effect and does not correct the hemolysis parameters produced by Dapsone treatment except for Hi levels which were more sensitive to the oxidant damage.

Vitamina E como agente protector de hemolisis en pacientes de lepra tratados con dapsona / Vitamin E as protective agent agains hemolysis in leprosy patients under dapsone treatment

La dapsona (4,4-diamino difenilsulfona) (DDS) utilizada en el tratamiento de pacientes con lepra, produce como efecto secundario una hemólisis intravascular debida a una comprobada lesión oxidativa a nivel de la membrana del eritrocito. Con el objeto de evaluar el efecto protector de la vitamina E sobre la hemólisis producida por la droga se estudiaron 16 pacientes durante cuatro meses, divididos en dos grupos: Grupo 1(n = 7) inicia tratamiento con dapsona por administración oral: 100 mg/dia y Grupo 2 (n = 9) dapsona mßs vitamina E: 800 U/dYa. Al inicio del tratamiento se determinó el nível enzimático de Glucosa-6-fosfato dehidrogenasa (G-6PD), siendo excluidos aquéllos que presentaban bajos niveles de dicha enzima dada su sensibilidad al fármaco. Todos los pacientes presentaban una anemia normocítica normocrómica, con un marcado descenso de Haptoglobina (menos de 5 mg/dl). El análisis estadístico demuestra que los valores de reticulocitos no presentan diferencias significativas entre los grupos y a lo largo del tiempo de estudio. En el caso de la metahemoglobina (Hi) se observó en el grupo 1 un aumento significativo entre el mes de inicio y al final del tratamiento, no obteniéndose los mismos resultados para el grupo 2, no obstante ser el parámetro más sensible a la agresión oxidativa. En la formación de cuerpos de Heinz se obtuvo los mismos resultados que en el caso de la Hi. Se concluye que si bien la vitamina E no corrige todos los parámetros de laboratorio valorados, evitaría la formación de metahemoglobina producida como resultado del tratamiento con dapsona. (AU)

Vitamina E como agente protector de hemolisis en pacientes de lepra tratados con dapsona / Vitamin E as protective agent agains hemolysis in leprosy patients under dapsone treatment

La dapsona (4,4'-diamino difenilsulfona) (DDS) utilizada en el tratamiento de pacientes con lepra, produce como efecto secundario una hemólisis intravascular debida a una comprobada lesión oxidativa a nivel de la membrana del eritrocito. Con el objeto de evaluar el efecto protector de la vitamina E sobre la hemólisis producida por la droga se estudiaron 16 pacientes durante cuatro meses, divididos en dos grupos: Grupo 1(n = 7) inicia tratamiento con dapsona por administración oral: 100 mg/dia y Grupo 2 (n = 9) dapsona mßs vitamina E: 800 U/dÝa. Al inicio del tratamiento se determinó el nível enzimático de Glucosa-6-fosfato dehidrogenasa (G-6PD), siendo excluidos aquéllos que presentaban bajos niveles de dicha enzima dada su sensibilidad al fármaco. Todos los pacientes presentaban una anemia normocítica normocrómica, con un marcado descenso de Haptoglobina (menos de 5 mg/dl). El análisis estadístico demuestra que los valores de reticulocitos no presentan diferencias significativas entre los grupos y a lo largo del tiempo de estudio. En el caso de la metahemoglobina (Hi) se observó en el grupo 1 un aumento significativo entre el mes de inicio y al final del tratamiento, no obteniéndose los mismos resultados para el grupo 2, no obstante ser el parámetro más sensible a la agresión oxidativa. En la formación de cuerpos de Heinz se obtuvo los mismos resultados que en el caso de la Hi. Se concluye que si bien la vitamina E no corrige todos los parámetros de laboratorio valorados, evitaría la formación de metahemoglobina producida como resultado del tratamiento con dapsona.

Reconstruction of anti-leprosy drug depleted complement haemolytic activity by addition of zymosan-treated sera (a source of C142) and CratEDTA (a source of C3-C9).

This paper describes the mechanism of in vitro interaction of human serum complement system with anti-leprosy drugs (dapsone and clofazimine) and anti-lepra reaction drugs such as chloroquine. These drugs could inhibit the complement-mediated lysis of erythrocytes both via direct and alternative pathways, but only at hypertherapeutic doses. Attempts were made to restore the drug depleted complement-mediated lysis of erythrocytes by adding zymosan-treated guinea-pig sera (a source of C142) and also by adding Crat-EDTA sera (a source of C3-C9). Destroyed complement-mediated haemolytic activity by dapsone could be restored by early complement (C142) components, while complement-mediated haemolytic activity blocked by clofazimine could be regenerated by adding both late (C3-C9) and early (C142) complement component. However, chloroquine-mediated inhibition of the complement-mediated haemolysis activity could not be appreciably restored by adding both early and late complement reagents.

Intravascular hemolysis and acute renal failure following intermittent rifampin therapy.

Renal failure is a rare complication associated with the use of rifampin. Intravascular hemolysis leading to acute renal failure following rifampin therapy is extremely rare. Two patients with leprosy who developed hemolysis and acute renal failure following rifampin are reported.