Characterization of a renal tubular epithelial cell line which secretes the autologous target antigen of autoimmune experimental interstitial nephritis.

Abstract

Abstract. Proximal tubular epithelial cells from mice which develop autoimmune interstitial nephritis were found to express the nephritogenic target antigen, 3M-1. Anti-3M-1 mAbs (ct3M-1-Ab) were used to positively select for 3M-l-secreting tubular epithelium and, after stabilization in culture, this new cell line (MCT) was examined for the production of several moieties important to either immune interactions or to the development of extracellular matrix. Alkaline phosphatase-staining MCT cells also express epithelial growth factor receptors with a Kd of 0.87 nM and an epithelial growth factor receptor constant (R,,) of 2.1 x 104 receptors/ceil. MCT culture supernatants contain greater amounts of laminin, and types IV and V procollagens compared to types I and III procollagens, and growing MCT cells on type I collagen matrix causes them to preferentially secrete even more type IV and V procollagen.The 30,000-Mr 3M-1 antigen could be immunoprecipitated from biosynthetically labeled MCT cell supernatants with a3M-1-Ab. An identical-sized moiety was isolated by immunoaffinity chromatography from collagenase-solubilized mouse kidney tubular basement membranes. The 3M-I antigen can be found on the MCT cell surface by radioimmunoassay, or deposited in a linear array in the extracellular matrix surrounding the MCT cells in culture by immunofluorescence. Mature messenger RNA species for both class I and class II major histocompatibility complex (MHC) molecules were detected by Northern hybridization, and their corresponding cell surface gene products were detected by cytofluorography of MCT cells stained with haplotype-specific antibodies. Both the cell surface 3M-1 and the small amounts of detected class II MHC molecules appear to be biologically functional, as MCT cells can support the proliferation of 3M-Ispecific, class II MHC-restricted helper T cells in culture. These findings suggest that MCT cells provide all the necessary biological parameters for interfacing both as the target of a nephritogenic immune response, and as a potential source for new extracellular matrix which develops as a fibrogenic response to interstitial nephritis.

Prostaglandin E2 increases proximal tubule fluid reabsorption, and modulates cultured proximal tubule cell responses via EP1 and EP4 receptors

“…These cells were characterized and shown to possess many of the characteristics of the in vivo PT. 21 MCT cells were grown in DMEM: F-12 (1:1) media supplemented with 10% FBS, 1% Penicillin-Streptomycin, and 1% L-glutamine and were maintained at 5% CO 2 were carried out as previously described. 22 Briefly, EP siRNA was prepared according to the manufacturer's instructions to a final concentration of 5 nM in 100 μl DMEM-F12 and HiPerfect Transfection reagent (Qiagen), and cells were maintained for a total of 48 h. A universal scrambled siRNA (Qiagen) was used as a negative control for these experiments.…”

“…The kidneys were quickly removed, and 1-2 mm coronal slices were placed in chilled dissection dishes for freehand dissection of PTs, distinguished from other segments based on various properties: diameter difference, cell heterogeneity, and transluscency. 21 The microdissected PTs were then transferred to a chamber for in vitro perfusions and measurement of net fluid reabsorption (Jv). To confirm the involvement of EP 1 receptors in fluid reabsorption, PTs were also isolated from mice lacking the EP 1 receptor (FVB-EP 1 null mice), which our group generated and thoroughly characterized.…”

T cells reactive to an inducible heat shock protein induce disease in toxin-induced interstitial nephritis.

“…MCT cells, a proximal tubular epithelial cell line derived from SJL/J mice (15), were grown as previously described.…”

“…MCT ceils are a proximal tubular epithelial ceil line derived from SJL/J mice (15). We next evaluated whether this cell line also demonstrated inducible HSP70 expression.…”

Role of Atrophic Tubules in Development of Interstitial Fibrosis in Microembolism-Induced Renal Failure in Rat

“…11 Other previous studies have stressed the importance of damaged tubules in promoting tubulointerstitial injury, since tubular cells in a damaged kidney can express or secrete various cytokines including growth factors 12,13 and matrix proteins. 14,15 In addition, a transdifferentiation of tubular epithelial cells into myofibroblasts 16 and high proliferation index among atrophic tubular cells have been noted in end-stage human kidneys with interstitial fibrosis. 17 Since pathological changes in tubules occur at the same time as interstitial fibrosis in most animal models of renal disease, causal relationships between tubular changes and interstitial fibrosis have been difficult to delineate.…”