ISOLATION AND CHARACTERIZATION OF THE LOWER PORTION OF THE THIN LIMB OF HENLE IN PRIMARY CULTURE

To further characterize cells of the lower portion of the thin limb of Henle (TLH1p) under defined conditions in vitro, we developed a techn ique to enrich this cell population in suspension. TLH1p cells were is olated by enzymatic digestion of rat inner medulla, elimination of col lecting ducts by lectin-coated beads, and differential centrifugation. Immunohistochemical staining of primary cultures of TLH1p cells with various markers revealed the preparations to be >90% pure. The hormona l stimulation pattern of PGE(2) and cAMP production by arginine vasopr essin, angiotensin II, and dopamine in the isolated cells also argued against significant contamination by other cell types. Staining with a n antibody against the aquaporin-1 water channel showed the distributi on of cells from the ascending and descending limbs to be approximatel y equal in the isolated population. This technique allows the enrichme nt of cells from the lower portion of the thin limb of Henle in suspen sion to a very high degree of purity with the option to start primary cultures. Because these segments of the tubular system in particular a re relatively inaccessible for microdissection, the presented method r enders the possibility of addressing new questions regarding these tub ular segments under defined conditions in vitro.