ABSTRACT
c-Jun, a component of the activating protein-1 transcription factor family, has been known to play an important role in the control of cell proliferation. It is also suspected to be a critical mediator of tumor promotion. However, investigations of c-Jun activation patterns in inflammatory and inflammatory transforming skin diseases have not been described so far. In this work, we show the c-Jun activation pattern in skin samples of patients with cutaneous lichen planus (LP), squamous cell carcinoma (SCC), psoriasis and normal skin using an immunohistochemical approach and Western blot analysis. In addition, we studied the c-Jun activation pattern in histological samples of three patients in whom LP transformed to SCC. We show that c-Jun is rarely activated in normal skin and psoriasis in contrast to LP and SCC. Our results suggest that c-Jun activation in human skin is involved in (1) proliferation and (2) could potentially participate in the transformation of LP from an inflammatory to a carcinogenic state. Nevertheless, JNK1/2, an important c-Jun activating kinase, was not found to be differentially regulated in LP and SCC compared with normal skin.
Subject(s)
JNK Mitogen-Activated Protein Kinases/metabolism , Lichen Planus/enzymology , Neoplasms, Squamous Cell/enzymology , Psoriasis/enzymology , Skin Neoplasms/enzymology , Blotting, Western , Case-Control Studies , Enzyme Activation , Humans , Immunohistochemistry , Mitosis , Signal TransductionABSTRACT
A capsid protein of porcine circovirus 2 (PCV 2) serves as a diagnostic antigen for the detection of PCV 2-associated disease known as a postweaning multisystemic wasting syndrome (PMWS). In this report, a bacterial expression system was developed for the expression and purification of the full-length PCV 2 capsid (Cap) protein from a codon-optimized cap gene. Replacement of rare arginine codons located at the 5' end of the cap reading frame with codons optimal for E. coli was found to overcome the poor expression of the viral protein in the prokaryotic system. The Cap protein was purified to greater than 95% homogeneity by using a single cation-exchange chromatography at a yield of 10 mg per litre of bacterial culture. Despite the failure of the E. coli-expressed Cap protein to self-assemble into virus-like particles (VLPs), the immunization of mice with recombinant Cap yielded antibodies with the same specificity as those raised against native PCV 2 virions. In addition, the antigenic properties of the purified Cap protein were employed in a subunit-based indirect ELISA to monitor the levels of PCV 2 specific antibodies in piglets originating from a herd which was experiencing PCV 2 infection. These results pave the way for a straightforward large-scale production of the recombinant PCV 2 capsid protein and its use as a diagnostic antigen or a PCV 2 subunit vaccine.
Subject(s)
Antibodies, Viral/blood , Capsid Proteins/immunology , Circovirus/immunology , Escherichia coli/metabolism , Porcine Postweaning Multisystemic Wasting Syndrome/diagnosis , Swine Diseases/diagnosis , Swine/virology , Amino Acid Sequence , Animals , Capsid Proteins/genetics , Capsid Proteins/metabolism , Circovirus/genetics , Circovirus/metabolism , Escherichia coli/genetics , Immunization , Molecular Sequence Data , Porcine Postweaning Multisystemic Wasting Syndrome/prevention & control , Porcine Postweaning Multisystemic Wasting Syndrome/virology , Recombinant Proteins/genetics , Recombinant Proteins/immunology , Recombinant Proteins/metabolism , Swine Diseases/prevention & control , Swine Diseases/virology , Viral Vaccines/administration & dosage , Viral Vaccines/immunology , Virion/metabolismABSTRACT
The genetically detoxified Bordetella pertussis adenylate cyclase is a promising delivery system for immunodominant tuberculosis antigens in gamma interferon release assays. This system has not been evaluated in human immunodeficiency virus (HIV)-infected persons in high tuberculosis prevalence areas. A whole-blood gamma interferon release assay with Mycobacterium tuberculosis antigens (early-secreted antigenic target 6, culture filtrate protein 10, alpha-crystallin 2, and TB10.3) delivered by adenylate cyclase in addition to native tuberculosis antigens (without adenylate cyclase delivery) was evaluated in 119 adults in Khayelitsha Township, Cape Town, South Africa. Results were compared to tuberculin skin test results of 41 HIV-positive and 42 HIV-negative asymptomatic persons, in addition to 36 HIV-positive persons with recently diagnosed smear- or culture-positive pulmonary tuberculosis. Delivery of tuberculosis antigens by adenylate cyclase decreased by 10-fold the amount of antigen required to restimulate T cells. Furthermore, the responses of HIV-positive persons with a low response to native tuberculosis antigens were enhanced when these antigens were delivered by adenylate cyclase. When gamma interferon responses to the tuberculosis antigens (with or without delivery by adenylate cyclase) were combined, a significantly higher number of patients were scored positive than by tuberculin skin testing. Ex vivo responses to tuberculosis antigens delivered by adenylate cyclase are maintained in the context of HIV infection. Our findings suggest that the majority of those in this population are infected with tuberculosis, which is of significant public health importance.
