ABSTRACT
BACKGROUND: The inflammation in acne vulgaris is widely thought to be induced by an immunological reaction, but the role of Propionibacterium acnes is unclear. OBJECTIVES: To examine the local host response mechanism of a keratinocyte cell line 3 h and 6 h after stimulation with viable and heat-killed P. acnes. METHODS: The quantitative expression of cytokines was measured at the mRNA level by real-time reverse transcription-polymerase chain reaction. RESULTS: The coincubation of a keratinocyte cell line with viable, but not heat-killed, P. acnes modulated an adequate cytokine response for interleukin (IL)-1beta, granulocyte/macrophage colony-stimulating factor and IL-8. High-performance liquid chromatographic analysis of the in vivo porphyrin pattern secreted by P. acnes revealed a predominance of coproporphyrin III in acne lesions. This same porphyrin fraction also modestly induced IL-8 expression by keratinocytes. CONCLUSIONS: This cytokine pattern may favour a chemotactic response and implicates P. acnes and coproporphyrin III in the recruitment of inflammatory cells to the site of infection and in the development of acne lesions.
Subject(s)
Acne Vulgaris/immunology , Coproporphyrins/immunology , Cytokines/biosynthesis , Keratinocytes/immunology , Propionibacterium acnes/immunology , Acne Vulgaris/metabolism , Acne Vulgaris/microbiology , Adolescent , Adult , Antigens, Bacterial/immunology , Cell Line , Chemotactic Factors/immunology , Coproporphyrins/biosynthesis , Cytokines/genetics , Cytokines/immunology , Humans , Inflammation Mediators/metabolism , RNA, Messenger/genetics , Reproducibility of Results , Reverse Transcriptase Polymerase Chain Reaction/methods , Skin/metabolismABSTRACT
Monoclonal antibodies against Pd-coproporphyrin I have been obtained. The antibody specificity for free as well as for conjugated Pd-coproporphyrin I is characterized. Affinity constants are estimated for 3 monoclonal antibodies effectively interacting with free Pd-coproporphyrin I. A comparative study on the binding of monoclonal antibodies with analogues and derivatives of Pd-coproporphyrin I has revealed that the antigen is mainly located inside the antibody paratope. The protein adjoins complementary to the metalloporphyrin in such a manner that antibodies obtained discern only isomer I, and to some degree, isomer III of coproporphyrin.
Subject(s)
Coproporphyrins/immunology , Metalloporphyrins/immunology , Animals , Antibodies, Monoclonal , Antibody Affinity , Antibody Specificity , Hybridomas , Immunoenzyme Techniques , Immunoglobulin G/immunology , Immunoglobulin M/immunology , Mice , Mice, Inbred BALB CABSTRACT
The synthesis of protein conjugates with the new high-efficient fluorescent labile coproporphyrin-I was optimized. A number of conjugates of monoclonal antibodies with different coproporphyrin-I content were synthesized, and their spectral properties were studied in water and micellar solutions, i.e. adsorption, excitation and emission spectra, fluorescence quantum yields, fluorescence pH-dependences. The binding constants of coproporphyrin-I and its protein conjugates with serum albumin were determined. The antibodies labelled with coproporphyrin-I retain the functional activity and photochemically stable in water solutions. The sensitivity of fluorometric detection of coproporphyrin-I and its conjugates with proteins is more than 10 times greater than in case of FITC.
