title: Isolation and characterization of resident macrophages from the smooth muscle layers of murine small intestine.
abstract: Macrophages within the murine tunica muscularis were isolated and cultured for physiological studies. Following dispersion, macrophages were identified by phagocytotic activity of fluorescein isothiocyanate (FITC)-dextran. Immediately following isolation, macrophages were rounded and possessed fluorescent granula but developed a ramified shape after 3-4 days in culture. Resident and cultured macrophages were immunopositive for F4/80 and I-Ad/I-Ed. Greater than 90% of F4/80 positive cultured cells were FITC-dextran positive. Macrophages had resting membrane potentials (RMP) of -33.3 +/- 1.5 mV after 1 day in culture, which increased to -53.9 +/- 4.4 mV after 3-4 days. The change in RMP was associated with the development of an inward rectifying K+ current, and a decrease in a voltage-dependent, inactivating outward current. After 3-4 days in culture the inflammatory mediated substances adenosine triphosphate (ATP), platelet-activating factor and bacterial lipopolysaccharide induced increases in cytoplasmic Ca2+ ([Ca2+]i). Forskolin suppressed the ATP-induced increase in [Ca2+]i. Macrophages exhibited oxidative bursts, measured by oxidation of dihydrorhodamine-123 to rhodamine-123. Oxidative bursts coincided with a reduction in intracellular pH. Macrophages expressed a proton conductance that may participate in pH maintenance during reactive oxygen production. These results suggest that resident macrophages in the intestine may play a role in the immunological protection of the gut.
authors.affiliation: Department of Veterinary Pharmacology, Graduate School of Agriculture and Life Sciences, The University of Tokyo, Tokyo, Japan. aozaki@mail.ecc.u-tokyo.ac.jp
authors.forename: H
authors.initials: H
authors.lastname: Ozaki
authors.forename: T
authors.initials: T
authors.lastname: Kawai
authors.forename: C W
authors.initials: CW
authors.lastname: Shuttleworth
authors.forename: K-J
authors.initials: KJ
authors.lastname: Won
authors.forename: T
authors.initials: T
authors.lastname: Suzuki
authors.forename: K
authors.initials: K
authors.lastname: Sato
authors.forename: H
authors.initials: H
authors.lastname: Horiguchi
authors.forename: M
authors.initials: M
authors.lastname: Hori
authors.forename: H
authors.initials: H
authors.lastname: Karaki
authors.forename: S
authors.initials: S
authors.lastname: Torihashi
authors.forename: S M
authors.initials: SM
authors.lastname: Ward
authors.forename: K M
authors.initials: KM
authors.lastname: Sanders
mesh_terms: D000818:Animals; D002118:Calcium / Q000032:analysis; D002478:Cells, Cultured; D006863:Hydrogen-Ion Concentration; D007421:Intestine, Small / Q000166:cytology / Q000276:immunology*; D007424:Intracellular Fluid / Q000737:chemistry; D008264:Macrophages / Q000166:cytology / Q000502:physiology*; D008564:Membrane Potentials; D051379:Mice; D009130:Muscle, Smooth / Q000166:cytology / Q000276:immunology*; D018408:Patch-Clamp Techniques; D016897:Respiratory Burst / Q000502:physiology
chemical_list: D002118:Calcium