Differential expression of SM22 isoforms in myofibroblasts and smooth muscle cells from rabbit bladder

The E-11 and 1-B8 monoclonal antibodies raised to the smooth muscle (SM)-sp ecific SM22 protein from pig stomach were used to study the in vivo and in vitro phenotypic characteristics of myofibroblasts (MF) and SM cells (SMC) from the bladder detrusor muscle and serosal thickening of male rabbit. The 22-kDa SM22 band found in the SM extract appeared to be composed of distin ct isoforms when examined in non-equilibrium two-dimensional gel electropho resis (2D-EF): alpha (the most basic), beta, gamma, and delta (the most aci dic) in the ratio of 34(alpha):23(beta):36(gamma):8(delta). Western blots o f 2D-electrophoresed bladder extracts treated with E-11 and 1-B8 showed tha t alpha, beta, and delta, but not gamma isoforms were labeled with E-11, wh ereas alpha, beta, and gamma isoforms were stained with 1-B8. This differen tial immunoreactivity was not influenced by phosphorylation. The tissue dis tribution of SM22 immunostaining was heterogeneous in the bladder SM and se rosal thickening developed as a consequence of partial outflow obstruction of the urinary bladder. At the cellular level, the 1-B8 and E-11 antibodies stained the SMC in a "diffuse" (the whole cytoplasm) and "honeycomb" (the peripheral cytoplasm) manner, whereas MF immunostaining was quite homogeneo us. The two antibodies also reacted with cultured primary bladder SMC and M F grown in low serum conditions showing a heterogeneous SM22 cell distribut ion but an identical subcellular localization, i.e., the actin-containing f ilamentous network, distinguishable in part from that found in vivo. The im munocytochemical, Western blotting and 2D-EF patterns of MF from thickened serosa indicated that the gamma isoform alone is expressed in this tissue. This SM22 variant appeared before the completion of the cellular transition from MF to fully differentiated SMC. This pattern is reminiscent of bladde r ontogenesis where SM22 expression in the developing bladder wall precedes that of SM myosin. Taken together these data suggest that: (i) SM22 isofor ms are differently assorted in MF vs. SMC; (ii) SM22 is a SMC-lineage marke r inasmuch as its expression occurs in an experimental condition characteri zed by a time-related cell phenotypic transition from MF to SMC, and (iii) cell conversion ability of serosal cells in the adult might take place via the reactivation of a specific "foetal" gene programme.