Hepatitis B Virus Infection Among Leprosy Patients: A Case for Polymorphisms Compromising Activation of the Lectin Pathway and Complement Receptors.

Thousands of leprosy patients not only suffer from physical deformities, but also either have or have had hepatitis B virus (HBV) coinfection. Polymorphisms of the complement system modulate susceptibility to leprosy, but genetic susceptibility to past or present HBV infection is unknown. We used sequencing and multiplex sequence-specific PCR to genotype 72 polymorphisms of seven genes (MBL2, FCN1, FCN2, FCN3, MASP1, MASP2, C3) encoding components of the lectin pathway, and two genes encoding complement receptors (CR1, VSIG4) in 190 patients, of which 74 were positive for HBsAg and/or anti-HBc (HBV+, 93.2% with a resolved infection) and 116 lepromatous patients, and 408 HBV-blood donors. In addition, we tested for levels of proteins of the lectin pathway. We found no difference between serum concentrations of mannan-binding lectin (MBL), MBL-associated serine proteins (MASP-1, MASP-2, MASP-3, MAp44), ficolin-3 (FCN-3), soluble complement receptor 1 (sCR1) and MBL mediated C4 activation, measured by ELISA or TRIFMA in up to 167 HBV+ and HBV- patients. Haplotypes lowering protein levels or encoding dysfunctional proteins increased susceptibility to HBV infection: MBL2*LYQC (OR = 3.4, p = 0.02), MASP1*AC_CC (OR = 4.0, p = 0.015) and MASP2*1C2-l (OR = 5.4, p = 0.03). Conversely, FCN1*3C2 haplotype, associated with higher gene expression, was protective (OR = 0.56, P = 0.033). Other haplotypes associated with HBV susceptibility were: MASP2*2B1-i (OR = 19.25, P = 0.003), CR1*3A (OR = 2.65, P = 0.011) and VSIG4*TGGRCG (OR = 12.55, P = 0.014). Some polymorphisms in ficolin genes associated with lower protein levels increased susceptibility to leprosy/HBV infection: FCN*1 (OR = 1.66, P = 0.029), FCN2*GGGCAC (OR = 6.73, P = 0.008), and FCN3*del_del_C (OR = 12.54, P = 0.037), and to lepromatous disease/HBV infection: FCN2*TA (OR = 2.5, P = 0.009), whereas FCN2*MAG was associated with increased FCN-2 expression and resistance against coinfection (OR = 0.29, P = 0.026). These associations were independent of demographic factors and did not increase susceptibility to leprosy per se, except MASP2*1C2-l. Associations for FCN2, FCN3, MASP1, MASP2, and VSIG4 variants were also independent of each other. In conclusion, polymorphisms compromising activation of the lectin pathway of complement increase susceptibility to HBV infection, with ficolin polymorphisms playing a major role in modulating the susceptibility among leprosy patients.

Complement receptor 1 (CR1, CD35) association with susceptibility to leprosy.

BACKGROUND: Pathophysiological mechanisms are still incompletely understood for leprosy, an urgent public health issue in Brazil. Complement receptor 1 (CR1) binds complement fragments C3b/C4b deposited on mycobacteria, mediating its entrance in macrophages. We investigated CR1 polymorphisms, gene expression and soluble CR1 levels in a case-control study with Brazilian leprosy patients, aiming to understand the role of this receptor in differential susceptibility to the disease. METHODOLOGY: Nine polymorphisms were haplotyped by multiplex PCR-SSP in 213 leprosy patients (47% multibacillary) and 297 controls. mRNA levels were measured by qPCR and sCR1 by ELISA, in up to 80 samples. PRINCIPAL FINDINGS: Individuals with the most common recombinant haplotype harboring rs3849266*T in intron 21 and rs3737002*T in exon 26 (encoding p.1408Met of the York Yka+ antigen), presented twice higher susceptibility to leprosy (OR = 2.43, p = 0.017). Paucibacillary patients with these variants presented lower sCR1 levels, thus reducing the anti-inflammatory response (p = 0.040 and p = 0.046, respectively). Furthermore, the most ancient haplotype increased susceptibility to the multibacillary clinical form (OR = 3.04, p = 0.01) and presented the intronic rs12034383*G allele, which was associated with higher gene expression (p = 0.043), probably increasing internalization of the parasite. Furthermore, there was an inverse correlation between the levels of sCR1 and mannose-binding lectin (initiator molecule of the lectin pathway of complement, recognized by CR1) (R = -0.52, p = 0.007). CONCLUSIONS: The results lead us to suggest a regulatory role for CR1 polymorphisms on mRNA and sCR1 levels, with haplotype-specific effects increasing susceptibility to leprosy, probably by enhancing parasite phagocytosis and inflammation.

Evaluation of rheumatoid factor and anti-citrullinated peptide antibodies in relation to rheumatological manifestations in patients with leprosy from Southern Brazil.

BACKGROUND: Leprosy patients may present several osteoarticular complaints, which require further evaluation of inflammatory diseases, such as rheumatoid arthritis (RA). Therefore, an adequate clinical assessment in addition to testing for rheumatoid factors (RF) and anticyclic citrullinated peptide antibodies (anti-CCP), can be useful in order to establish the correct diagnosis. METHOD: In this study, the relation of RF and anti-CCP with rheumatological manifestations was evaluated in 97 leprosy patients from Southern Brazil. The results were compared to RA patients and healthy controls from the same geographical area and ethnic background. RESULTS: Neuropathy was observed in 71.1% and arthritis in 35.1% of the leprosy patients. A high frequency of RF positivity was observed among the leprosy patients (41.2%, 40/97), with RF immunoglobulin A (IgA) significantly associated with arthritis (OR = 7.9, 95% CI = 1.5-40.6 P = 0.008). Anti-CCP was observed in 9.3% (9/97) of the patients, with anti-CCP2 being the most frequent subtype. Only 4.1% (4/97) of the patients were RF and anti-CCP concomitantly positive. RF IgM showed a significant association with leprosy when compared to healthy controls (P < 0.0001) whereas for anti-CCP2 no significant results were observed (P = 0.0585). However, both biomarkers showed a strong association with RA when compared to leprosy in patients from the same geographical area and ethnic background (anti-CCP2 OR = 38.6; 95% CI = 16.49-90.26; P < 0.0001 and RF IgM OR = 4.51; 95% CI = 2.62-7.77; P < 0.0001). CONCLUSION: Due to the similarity of some rheumatological manifestations in leprosy with other inflammatory diseases, such as RA, clinical and laboratorial evaluation of affected patients must be carefully assessed in order to achieve proper diagnosis and treatment.

Placental alkaline phosphatase in pediatric adrenocortical cancer.

The germline R337H mutation in the TP53 gene is considered to be responsible for the increased incidence of adrenocortical tumors (ACTs) in children from Brazil. High level production of hormones in ACTs (>95%) cause virilization alone (60%), Cushing syndrome (<5%), the mixed type (30%), or other rarer manifestations. ACT probably develops owing to events occurring during the final stages of intrauterine life based on the very common early onset of signs and symptoms shortly after birth. In this study, we determined by immunohistochemistry and enzyme assays whether placental alkaline phosphatase (PLAP) is expressed in pediatric ACTs. Immunohistochemical analysis revealed positive p53 expression in 88% of the tested ACTs (29 of 33). PLAP was detected at a slightly lower frequency based on immunohistochemical (17 of 33, 51%) and enzyme activity analyses (9 of 16, 56%). In conclusion, probably at a certain time point during adrenocortical development (end of gestation to early postnatal period), some fetal zone cells survive owing to defective apoptosis and develop into childhood ACT, maintaining some characteristics of the embryonal period, such as PLAP expression. Further studies of PLAP should investigate the functional role, if any, of PLAP in such tumors.

Mitotane associated with cisplatin, etoposide, and doxorubicin in advanced childhood adrenocortical carcinoma: mitotane monitoring and tumor regression.

PURPOSE: To define a mitotane dose for pediatric patients with adrenocortical cancer (ACC) that maintains therapeutic plasma levels (TL) between 14 and 20 microg/mL and to verify its antitumor efficacy in association with 8 cycles of cisplatin, etoposide, and doxorubicin (CED). METHODS: Powdered mitotane was dissolved in a medium chain triglyceride oil and administered to 11 children with ACC (2.4 to 15.4 y of age); an initial low dose was increased to 4 g/m2/d. Ten of the 11 children had a germline TP53 R337H mutation. Mitotane plasma levels were determined using high-performance liquid chromatography. RESULTS: The mitotane dose to maintain TL in 7 patients ranged from 1.0 to 5.3 g/m2/d. Six children reached mitotane levels of 10 microg/mL in 3.6 months (1.5 to 5.0 mo), whereas 5 children took 8 months (6.5 to 12.5 mo). Minor to partial tumor remission was found in 5 patients (<1 y) and complete remission was found in 2 patients. Of the 3 patients who are alive at the time of report, 1 patient has been without disease for 16 months, and 2 patients have progressive disease. All patients had recurrent metastatic disease (2 to 9 times). Mitotane toxic effects were nausea, diarrhea, vomiting, neurologic alterations, gynecomastia, a rare case of hypertensive encephalopathy, and CED-related hematologic toxic effects. CONCLUSIONS: Mitotane daily dose to maintain TL is variable and monitoring should start 1.5 months after the beginning of treatment. CED combined with mitotane is the best available pharmacologic treatment for ACC, but further studies are required to characterize different profiles of therapeutic response.

Amostra para controle de qualidade do eritrograma estavel durante 100 dias / Sample for quality control of red blood cells measurements with stability for up to 100 days

Eritrocitos de sangue humano de doadores saudaveis foram suspensos em meio CE de preservaçao, incubados durante 1 hora em glutaraldeidoa 0,5g/l, sob constante homogeneizaçao, e ressuspensos em meio CE contendo vitamina E (concentraçao final de 21 mg/l). Os resultados obtidos indicam que o tratamento e conservaçao de eritrocitos nas condiçoes especificadas podem levar a obtençao de amostras adequadas ao controle de qualidade do eritrograma, validas durante aproximadamente tres meses.