Roles of Reactive Oxygen Species in CXCL8 and CCL2 Expression in Response to the 30-kDa Antigen of <Emphasis Type="Italic">Mycobacterium tuberculosis</Emphasis>

Background  The 30-kDa antigen (Ag) of Mycobacterium tuberculosis (M. tuberculosis) is a strong inducer of innate and adaptive immune responses in human tuberculosis. The generation of reactive oxygen species (ROS) plays an important role in inflammatory signaling as well as antimicrobial defense. Materials and Methods  In this study, we investigated the role of ROS in the activation of mitogen-activated protein kinases (MAPKs) and secretion of the CXC chemokine ligand 8 (CXCL8) and CC chemokine ligand 2 (CCL2) by human monocytes stimulated with the 30-kDa Ag of M. tuberculosis H37Rv. Results  Treatment of human monocytes with the 30-kDa Ag activated rapid superoxide generation. In addition, the 30-kDa Ag activated mRNA and protein expression of CXCL8 and CCL2 in human primary monocytes through nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-dependent ROS generation. Analysis of MAPK activation (extracellular signal-regulated kinase (ERK) 1/2 and p38) showed rapid phosphorylation of both subfamilies in response to the 30-kDa Ag. In addition, 30-kDa-induced MAPK activation was inhibited in a dose-dependent manner by pretreatment with ROS scavengers. Toll-like receptor (TLR) 2 was required for ROS generation, chemokine production, and MAPK activation following stimulation with the 30-kDa Ag. Using highly specific signaling pathway inhibitors, we found that both p38 and ERK1/2 activation are essential for 30-kDa Ag-induced CCL2 but not CXCL8 production in human monocytes. Conclusion  These results indicate that TLR2–ROS signaling plays a crucial role in the 30-kDa Ag-mediated expression of CXCL8 and CCL2 in human monocytes. In addition, both p38 and ERK1/2 activation are essential for 30-kDa Ag-stimulated CCL2 production by monocytes.     

Identification of an immunostimulating protein from Mycobacterium leprae.

Despite the recent identification of a number of Mycobacterium leprae proteins, the major immunogenic determinants of this organism remain obscure. We isolated from M. leprae a potent immunostimulatory preparation, designated the MLP fraction, which contains a major protein of 35 kilodaltons (kDa). This protein was precipitated by monoclonal antibody ML03-A1, which recognizes a 35-kDa protein of M. leprae, and by sera obtained from patients with lepromatous leprosy. Neither sera from healthy controls nor sera from patients with pulmonary tuberculosis recognized the 35-kDa protein, and only one of four serum samples from patients with borderline tuberculoid leprosy reacted with this protein. The MLP fraction stimulated T-cell proliferation in patients with leprosy whose T cells proliferate in response to whole M. leprae cells. Apparently, the T-cell epitope associated with MLP is also expressed on M. tuberculosis and M. bovis BCG, since patients with pulmonary tuberculosis and BCG-vaccinated individuals demonstrated significant responses to the MLP fraction. The 35-kDa M. leprae protein, purified to homogeneity in the laboratory of P. J. Brennan, stimulated T-cell proliferative responses in all MLP-responsive subjects. These findings suggest that the 35-kDa protein present in MLP is an immunostimulatory component of M. leprae. In addition to serving as a useful probe for study of the T-cell anergy associated with lepromatous disease, this protein may ultimately be useful as a component of a vaccine designed to provide protection against infection with M. leprae.     

Urine protein electrophoresis and immunofixation electrophoresis supplement one another in characterizing proteinuria

Urine protein electrophoresis (UPE) is often considered to have limited usefulness in evaluating proteinuria that is not associated with gammopathies. Unusual protein bands that are detected by UPE are commonly characterized by immunofixation electrophoresis (IFE). In this paper, electrophoretic gel patterns are shown to illustrate the greater sensitivity of IFE, compared to UPE. However, UPE remains useful for three applications: (1) UPE provides distinctive patterns that can indicate the source of proteinuria and is useful in assessing renal diseases that are independent of gammopathy; (2) combined use of UPE and IFE can avoid misinterpretations and repeated analyses of urine proteins, and (3) UPE can be used in conjunction with IFE to improve the quantitation of Bence-Jones proteinuria (BJP).     

Clinical profiles of four patients with Rett syndrome carrying a novel exon 1 mutation or genomic rearrangement in the MECP2 gene

Rett syndrome (RTT) is a neurodevelopmental disorder caused by mutations in the X-linked MECP2 gene encoding methyl CpG binding protein 2 (MeCP2). Recently, a new isoform of MeCP2 including exon 1 was identified. This new isoform is more abundantly expressed in brain than the isoform including exons 2-4. Very little is known about the phenotypes associated with mutations in exon 1 of MECP2 since only a limited number of RTT patients carrying such mutations have been identified so far. In this study, we screened a cohort of 20 girls with RTT for exon 1 mutations by sequencing and multiplex ligation-dependent probe amplification (MLPA). We identified one girl with a novel exon 1 mutation (c.30delCinsGA) by sequencing and three with genomic rearrangements by MLPA. Comparison of the phenotypes showed that the girls carrying a mutation or rearrangement encompassing exon 1 were more severely affected than the girls with rearrangements not affecting exon 1.     

Expression of podoplanin in thymoma: its correlation with tumor invasion, nodal metastasis, and poor clinical outcome

Recent studies have shown that podoplanin overexpression is associated with lymph node metastasis and poor clinical outcome in several malignant tumors. To investigate the role of podoplanin in thymoma, we examined 111 thymomas by immunohistochemistry using monoclonal antibody D2-40, which recognizes podoplanin. The tumors consisted of 8 type A, 40 type AB, 15 type B1, 23 type B2, 15 type B3, and 10 combined thymomas according to the World Health Organization histological classification system and of 41 stage I, 28 stage II, 16 stage III, 20 stage IVa, and 6 stage IVb thymomas according to the Masaoka staging system. We have found podoplanin expression in 0 (0%) type A, 4 (10%) type AB, 4 (27%) type B1, 16 (70%) type B2, 10 (67%) type B3, and 7 (70%) combined thymomas and in 5 (12%) cases of stage I, 7 (25%) of stage II, 11 (69%) of stage III, 12 (60%) of stage IVa, and all (100%) of stage IVb thymomas. Podoplanin was significantly expressed in B2/B3/combined thymomas and advanced stage thymomas (P < .0001). On survival analysis, podoplanin expression was significantly associated with an increased risk of death for the whole group of thymomas (P = .0002), although it was not identified as an independent prognostic factor in multivariate analysis. The significant survival curve differences of podoplanin expression were also seen for stage III/IVa/IVb thymomas (P = .0409) and B2/B3/combined thymomas (P = .0478). In conclusion, D2-40 immunostaining seems to be valuable for predicting the aggressive and metastatic potential of thymomas and the prognosis of the patients.