RESUMO
INTRODUCTION: Mycobacterium abscessus is an emerging pulmonary pathogen with limited treatment options. Nitric oxide (NO) demonstrates antibacterial activity against various bacterial species, including mycobacteria. In this study, we evaluated the effect of adjunctive inhaled NO therapy, using a novel NO generator, in a CF patient with pulmonary M. abscessus disease, and examined heterogeneity of response to NO in vitro. METHODS: In the compassionate-use treatment, a 24-year-old CF patient with pulmonary M. abscessus was treated with two courses of adjunctive intermittent NO, first at 160 p.p.m. for 21 days and subsequently by escalating the dose up to 240 p.p.m. for 8 days. Methemoglobin, pulmonary function, 6 min walk distance (6MWD), qualify of life and sputum microbiology were assessed. In vitro susceptibility tests were performed against patient's isolate and comparison clinical isolates and quantified by Hill's slopes calculated from time-kill curves. RESULTS: M. abscessus lung infection eradication was not achieved, but improvements in selected qualify of life domains, lung function and 6MWD were observed during the study. Inhaled NO was well tolerated at 160 p.p.m. Dosing at 240 p.p.m. was stopped due to adverse symptoms, although methemoglobin levels remained within safety thresholds. In vitro susceptibility tests showed a dose-dependent NO effect on M. abscessus susceptibility and significant heterogeneity in response between M. abscessus clinical isolates. The patient's isolate was found to be the least susceptible strain in vitro. CONCLUSION: These results demonstrate heterogeneity in M. abscessus susceptibility to NO and suggest that longer treatment regimens could be required to see the reduction or eradication of more resistant pulmonary strains.
RESUMO
CrO3 is cytotoxic for human epithelial 293 kidney cells over a narrow concentration range of approximately 2-8 microM (D50 approximately 3.0 microM); significantly greater toxicity is observed in clonogenic assays (D50 approximately 0.1-1.0 microM). Survival of a small fraction of cells (< or = 0.1%) at CrO3 concentrations between 10(-5) to 10(-3) M, and first-order kinetics of cytotoxicity, rationalized the derivation of a new panel of transformed human epithelial cell lines resistant to cytotoxic concentrations of CrO3 over the range of 5-100 microM. Wild-type and Cr-resistant 293 cell lines display similar morphology under phase microscopy, but wild-type cells grow faster and reach stationary phase sooner than Cr-resistant cells. The Cr-resistant phenotype is stable, and it is specific, since Cr-resistant cells are killed by NiSO4 or by CdCl2 at concentrations equivalent to those that kill wild-type cells. Toxicity analysis curves subjected to target theory suggest that the Cr-resistant cell lines have fewer Cr-sensitive "targets" and have undergone a "loss of function" compared to wild-type cells. This loss of function may be related to significantly lower rates of uptake of Na2(51)CrO4,which correlate inversely with CrO3 concentrations used for the selection and maintenance of the Cr-resistant lines, and to reduced levels of an approximately 96-kDa protein in comparison to wild-type cells. This new panel of Cr-resistant transformed human epithelial kidney cell lines will be useful in comparative studies of genetic resistance and sensitivity to human Cr(VI) toxicity, sulfate transport, and growth control differences between wild-type and Cr-resistant cells.
