RESUMEN
Herein, we performed microarray experiments in Schwann cells infected with live M. leprae and identified novel differentially expressed genes (DEG) in M. leprae infected cells. Also, we selected candidate genes associated or implicated with leprosy in genetic studies and biological experiments. Forty-seven genes were selected for validation in two independent types of samples by multiplex qPCR. First, an in vitro model using THP-1 cells was infected with live Mycobacterium leprae and M. bovis bacillus Calmette-Guérin (BCG). In a second situation, mRNA obtained from nerve biopsies from patients with leprosy or other peripheral neuropathies was tested. We detected DEGs that discriminate M. bovis BCG from M. leprae infection. Specific signatures of susceptible responses after M. leprae infection when compared to BCG lead to repression of genes, including CCL2, CCL3, IL8 and SOD2. The same 47-gene set was screened in nerve biopsies, which corroborated the down-regulation of CCL2 and CCL3 in leprosy, but also evidenced the down-regulation of genes involved in mitochondrial metabolism, and the up-regulation of genes involved in lipid metabolism and ubiquitination. Finally, a gene expression signature from DEG was identified in patients confirmed of having leprosy. A classification tree was able to ascertain 80% of the cases as leprosy or non-leprous peripheral neuropathy based on the expression of only LDLR and CCL4. A general immune and mitochondrial hypo-responsive state occurs in response to M. leprae infection. Also, the most important genes and pathways have been highlighted providing new tools for early diagnosis and treatment of leprosy.
Asunto(s)
Quimiocinas/metabolismo , Lepra/metabolismo , Metabolismo de los Lípidos , Mitocondrias/metabolismo , Transcriptoma , Células Cultivadas , Quimiocinas/genética , Análisis por Conglomerados , Femenino , Perfilación de la Expresión Génica , Regulación de la Expresión Génica/inmunología , Interacciones Huésped-Patógeno , Humanos , Lepra/inmunología , Lepra/microbiología , Masculino , Mitocondrias/microbiología , Mycobacterium bovis/inmunología , Mycobacterium leprae/inmunología , Análisis de Secuencia por Matrices de Oligonucleótidos , Nervios Periféricos/metabolismo , Células de Schwann/inmunología , Células de Schwann/metabolismo , Células de Schwann/microbiologíaRESUMEN
Thalidomide (Thal) can suppress the growth of established, as well as explanted tumors in mice. We wanted to determine if it could suppress the ability of tumor cells to assemble and establish a primary tumor at the injection site. Using the mouse 4T1 mammary tumor model, we fed Thal to mice for 4 days, then injected 10(5) 4T1 cells into the interscapular region of Balb/c mice. After 20 days on treatment with Thal, all seven control mice, fed with meal had tumors ranging from 3 to 93 mm(3) (median 20). Two of the eight mice fed with meal + Thal had no tumors, and the remaining mice had tumors ranging from 2 to 22 mm(3) (median 5). The median volume of the tumors in the control group was significantly more than that of mice treated with Thal (p = 0.03, Mann-Whitney test). In vitro treatment of the 4T1cells with Thal did not inhibit their ability to proliferate, to adhere to plastic, or to bind to Concanavalin-A. Thal caused a marked reduction in the ability of the 4T1 cells to assemble into palpable tumors.
Asunto(s)
Inhibidores de la Angiogénesis/farmacología , Neoplasias Mamarias Experimentales/tratamiento farmacológico , Neoplasias Mamarias Experimentales/metabolismo , Neoplasias Mamarias Experimentales/patología , Talidomida/farmacología , Animales , Línea Celular Tumoral , Femenino , Ratones , Ratones Endogámicos BALB C , Factores de TiempoRESUMEN
Erythema nodosum leprosum (ENL) is an inflammatory reaction that may occur in multibacillary leprosy patients, and thalidomide is the treatment of choice. Its cause and the mechanism by which thalidomide suppresses ENL are not known. In the skin lesions, im- mune complexes and split products of complement are found. The activation of complement could precipitate ENL, and thalidomide could suppress the inflammation by inhibiting the activation of complement. To determine if thalidomide could suppress the activation of complement, we first incubated normal serum with thalidomide and with M. leprae or zymosan. The amount of residual functional complement was then assessed by determining the dilution of serum required to lyses sheep erythrocytes sensitized by rabbit antibodies (CH50 Assay). M. leprae and zymosan activated complement. The residual complement activity in the serum incubated with M. leprae or with zymosan was equivalent to that incubated with M. leprae or zymosan in the presence of thalidomide, hydrolyzed thalidomide and metabolites of thalidomide. Thalidomide did not inhibit the activation of complement by zymosan, a known initiator of complement activation by the alternative pathway, or by M. leprae.
Asunto(s)
Activación de Complemento/efectos de los fármacos , Eritema Nudoso/tratamiento farmacológico , Leprostáticos/farmacología , Lepra Lepromatosa/tratamiento farmacológico , Mycobacterium leprae/efectos de los fármacos , Mycobacterium leprae/inmunología , Talidomida/farmacología , Animales , Proteínas del Sistema Complemento/análisis , Eritema Nudoso/inmunología , Eritema Nudoso/fisiopatología , Humanos , Lepra Lepromatosa/inmunología , Lepra Lepromatosa/fisiopatología , Hígado/enzimología , Ratones , Mycobacterium leprae/metabolismo , Conejos , OvinosRESUMEN
Thalidomide and analogues are a class of immunomodulatory drugs or IMiDS. Thalidomide was initially approved by the U.S. Food and Drug Administation for treatment of erythema nodosum in leprosy and is now approved for multiple myeloma as well. A second generation IMiD, lenalidomide, is also approved for multiple myeloma and refractory myelodysplastic syndrome. Discovery of this class of drugs has been serendipitous and empirical, as the drug targets have been unknown. In this review, the authors integrate recent identification of drug targets of IMiDS, which include the inducible form of nitric oxide synthase (iNOS), Rho GTPase and caspase-1, with the developments in the understanding of the molecular biology of human inflammatory, infectious and neoplastic skin disorders. Because thalidomide reemerged through leprosy, the original disease classified by the T cell, the authors have also emphasized advances in the understanding of T-cell subsets in human skin disorders.
Asunto(s)
Dermatitis/tratamiento farmacológico , Inmunomodulación , Neoplasias Cutáneas/tratamiento farmacológico , Talidomida/análogos & derivados , Talidomida/uso terapéutico , Síndrome de Behçet/tratamiento farmacológico , Dermatitis/inmunología , Eritema Nudoso/tratamiento farmacológico , Eritema Nudoso/inmunología , Humanos , Lupus Eritematoso Cutáneo/tratamiento farmacológico , Lupus Eritematoso Cutáneo/inmunología , Linfoma Cutáneo de Células T/tratamiento farmacológico , Linfoma Cutáneo de Células T/inmunología , Sarcoidosis/tratamiento farmacológico , Sarcoidosis/inmunología , Neoplasias Cutáneas/inmunología , Talidomida/farmacologíaRESUMEN
OBJECTIVE: The immune-mediated events that precipitate erythema nodosum leprosum (ENL) are not well understood. One component may be the complexing of antibody with antigens released from infected macrophages, the activation of complement and the subsequent local inflammation. We assess here the ability of highly-purified, disrupted M. leprae, to activate complement. RESULTS: Intact and sonically-disrupted crude and alkali-purified nu/nu mouse-derived M. leprae suspensions were incubated with normal serum and a hemolytic titer (CH50) was determined as a measure of complement fixation. Crude M. leprae consumed complement, and disrupted preparations more than the intact. Purified M. leprae preparations did not consume complement unless disrupted. CONCLUSION: M. leprae, if disrupted, can activate complement. This supports a hypothesis that links released antigens with ENL, and may explain the increased probability of an occurrence of ENL following chemotherapy.
Asunto(s)
Activación de Complemento , Eritema Nudoso/inmunología , Lepra Lepromatosa/inmunología , Mycobacterium leprae/inmunología , Animales , Eritema Nudoso/fisiopatología , Humanos , Ratones , Ratones Desnudos , Mycobacterium leprae/fisiología , SonicaciónRESUMEN
Thalidomide is a drug that can enhance mitogen- and antigen-stimulated cells' ability to synthesize IL-2. To assess if thalidomide could concomitantly enhance the synthesis of IFN-gamma and incorporation of [H3]-thymidine, peripheral blood mononuclear cells (PBMC) were incubated in the presence or absence of thalidomide and staphylococcal enterotoxin A (SEA), anti-CD3, Con-A or PHA. After 18 h, the cultures were sampled for IL-2. At the termination of the 3-day cultures, they were assayed for IFN-gamma and incorporation of [H3]-thymidine. Regardless of the mitogen used to stimulate the PBMC, the thalidomide-treated PBMC produced more IL-2 than controls. Thalidomide enhanced IFN-gamma synthesis in the Con-A and anti-CD3-stimulated PBMC. It suppressed the ability of SEA and PHA-stimulated PBMC to incorporate [H3]-thymidine, whereas it enhanced incorporation of [H3]-thymidine in PBMCs stimulated with anti-CD3. When the PBMC were enriched for CD4+ or CD8+ cells, the SEA- and anti-CD3-stimulated CD4+ cells responded far better than the CD8+ cells in the synthesis of IL-2 and incorporation of [H3]-thymidine. In the thalidomide-treated SEA-stimulated CD4+ and CD8+ cells, thalidomide acted as a costimulant to enhance the synthesis of IL-2. In the anti-CD3-stimulated thalidomide-treated cultures of PBMC enriched for CD4+ cells, thalidomide acted as a costimulant to enhance the incorporation [H3]-thymidine. Thalidomide cooperated with all of the mitogens to enhance T-cell synthesis of IL-2. However, depending on the stimulant, thalidomide could suppress or enhance PBMC incorporation of [H3]-thymidine. The SEA-stimulated cells targeted by thalidomide to suppress incorporation of [H3]-thymidine were CD4+. CD4+ cells stimulated with anti-CD3 were enhanced by thalidomide in their ability to synthesize IL-2 and to incorporate [H3]-thymidine. Increased production of IL-2 by activated T cells may be a mechanism through which thalidomide exerts its immunomodulatory effects.