Abstract-We previously showed that hydrogen peroxide (H 2 O 2 ) contributes to flow-induced dilation in human coronary resistance arteries (HCRAs); however, the source of this H 2 O 2 is not known. We hypothesized that the H 2 O 2 is derived from superoxide (O 2 •Ϫ ) generated by mitochondrial respiration. HCRAs were dissected from right atrial appendages obtained from patients during cardiac surgery and cannulated with micropipettes. H 2 O 2 -derived radicals and O 2•Ϫ were detected by electron spin resonance (ESR) using BMPO as the spin trap and by histofluorescence using hydroethidine (HE, 5 mol/L) and dichlorodihydrofluorescein (DCFH, 5 mol/L). Diameter changes to increases in pressure gradients (20 and 100 cm H 2 O) were examined in the absence and the presence of rotenone (1 mol/L), myxothiazol (100 nmol/L), cyanide (1 mol/L), mitochondrial complex I, III, and IV inhibitors, respectively, and apocynin (3 mmol/L), a NADPH oxidase inhibitor. At a pressure gradient of 100 cm H 2 O, ubisemiquinone and hydroxyl radicals were detected from effluents of vessels. Including superoxide dismutase and catalase in the perfusate reduced the ESR signals. Relative ethidium and DCFH fluorescence intensities in HCRAs exposed to flow were enhanced (1.45Ϯ0.15 and 1.57Ϯ0.12, respectively compared with no-flow) and were inhibited by rotenone (0.87Ϯ0.17 and 0.95Ϯ0.07). Videomicroscopic studies showed that rotenone and myxothiazol blocked flow-induced dilation (% max. dilation at 100 cm H 2 O: rotenone, 74Ϯ3% versus 3Ϯ13%; myxothiazol, 67Ϯ3% versus 28Ϯ4%; PϽ0.05). Neither cyanide nor apocynin altered flow-induced dilation. These results suggest that shear stress induced H 2 O 2 formation, and flow-induced dilation is derived from O 2•Ϫ originating from mitochondrial respiration.

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Forensic DNA analysis of sexual assault evidence requires unambiguous differentiation of DNA profiles in mixed samples. To investigate the feasibility of magnetic bead-based separation of sperm from cell mixtures using a monoclonal antibody against MOSPD3 (motile sperm domain-containing protein 3), 30 cell samples were prepared by mixing 10(4) female buccal epithelial cells with sperm cells of varying densities (10(3), 10(4), or 10(5) cells/mL). Western blot and immunofluorescence assays showed that MOSPD3 was detectable on the membrane of sperm cells, but not in buccal epithelial cells. After biotinylated MOSPD3 antibody was incubated successively with the prepared cell mixtures and avidin-coated magnetic beads, microscopic observation revealed that each sperm cell was bound by two or more magnetic beads, in the head, neck, mid-piece, or flagellum. A full single-source short tandem repeat profile could be obtained in 80% of mixed samples containing 10(3) sperm cells/mL and in all samples containing ≥10(4) sperm cells/mL. For dried vaginal swab specimens, the rate of successful detection was 100% in both flocked and cotton swabs preserved for 1 day, 87.5% in flocked swabs and 40% in cotton swabs preserved for 3 days, and 40% in flocked swabs and 16.67% in cotton swabs preserved for 10 days. Our findings suggest that immunomagnetic bead-based separation is potentially a promising alternative to conventional methods for isolating sperm cells from mixed forensic samples.