These findings may help to explain why IC occurs primarily in women, why IC and other associated symptoms tend to be worst at the midcycle, and may also suggest that prognostic usefulness of staining bladder biopsies for OR. These results are particularly interesting because oestradiol has been shown to augment MC histamine secretion in response to the neuropeptide, substance P, which may be released in certain neuroinflammatory disorders which have a high prevalence in women, such as IC and irritable bowel syndrome.
The use of in vitro hepatic microsomal models as a method of studying the biotransformation of xenobiotics can be complicated by the insolubility of a lipophilic substrate. Addition of solvent to an in vitro system is an approach used to increase solubility of such substrates, but solvents may interact with the microsomal enzymes and cause changes in the kinetic parameters. This study focused on the solvents polyethylene glycol-400 (PEG) and acetone and their influence on the human microsomal in vitro biotransformation of the antineoplastic agent tamoxifen (TAM). The antianxiety agent alprazolam (ALP), a verified P450 3A substrate, was also studied. TAM is a nonpolar drug that is metabolized to two oxidative metabolites, N-desmethyltamoxifen (DMT) and 4-hydroxytamoxifen (4-OH-TAM), by cytochrome P450 isoforms. DMT is formed primarily by 3A isoforms, and the pathway(s) responsible for 4-OH-TAM formation are unknown. Biotransformation of ALP is mediated by P450 3A isoforms, which form alpha-hydroxyalprazolam (alpha-OH-ALP) and 4-hydroxyalprazolam (4-OH-ALP). Both PEG and acetone at 5% of the total microsomal incubation volume were found to solubilize TAM in the in vitro system. However, both solvents had an effect on the P450 mediated metabolism of ALP and TAM. For ALP, PEG was a noncompetitive inhibitor of alpha-OH-ALP (mean Ki = 2.06%) and 4-OH-ALP (mean Ki = 2.37%) formation. Acetone stimulated the production of alpha-OH-ALP, but had no apparent influence on 4-OH-ALP formation. The solvent's influence on TAM metabolism varied. PEG decreased the amount of DMT formed by the microsomes as compared to the system containing no solvent (control); however, 4-OH-TAM formation in the presence of PEG was 146-226% of controls. Samples containing acetone produced smaller quantities of both DMT (39-63%) and 4-OH-TAM (45-69%) as compared to controls. Since PEG and acetone increased the solubility of TAM in the incubation buffer but inhibited or accelerated enzymatic reactions compared to buffer alone, actual solubility in buffer was not a determinant of the rate of TAM metabolism. TAM appeared to be taken up into the microsomal fraction, making it available for biotransformation by the P450 isoforms. Although solvents may increase the solubility of nonpolar agents in aqueous systems, careful evaluation of the effects of solvent on metabolite formation, as compared to buffer controls, is needed in order to properly evaluate an in vitro metabolic model.
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