Demonstration of Smooth Muscle Contractile Protein Antigens in Liver and Epithelial Cells

Abstract

The presence of antismooth muscle autoantibodies in the sera of patients with active chronic hepatitis, and also in some other conditions not necessarily associated with liver disease, was discovered by use of cryostat sections from composite tissue blocks for routine immunofluorescent autoantibody tests. On sections that included rat stomach, liver, and kidney, these sera by the indirect test exhibited well-defined staining of smooth muscle fibers in the stomach mucosa and submucosa and of the smooth muscle coat of blood vessels in all three organs. It was noted2 that many such positive sera also displayed definite staining in areas that lack smooth muscle, for example, renal glomeruli and liver parenchymal cells; the latter cells were outlined t o yield a "polygonal" staining pattern. Both the smooth muscle and nonsmooth muscle staining patterns with positive sera were removed by adsorption with crude extracts of human uterine smooth muscle t i~s u e .~ Positive sera often also stain microfilamentous structures in the cytoplasm and near the surface of various different cell types in tissue culture, and this staining is also prevented by prior absorption with smooth muscle e~t r a c t .~In addition, we have noted the ability of some positive sera t o stain bile ductule cells in liver sections and also the apical regions of epithelial cells in gastric mucosa and kidney tubules, and we have suggested that different contractile proteins present in various cell types are the target antigens.T o elucidate the appearance of smooth muscle antibody (SMA) in a variety of human diseases that d o not primarily affect smooth muscle tissue itself, we have studied the distribution of the different contractile smooth muscle-type proteins in tissues other than smooth muscle by means of antisera raised against the different individual proteins. We have used immunofluorescence and immunoperoxidase labeling for this purpose and have obtained additional information about the intracellular localization of contractile proteins in epithelial and liver cells. MATERIALS AND METHODS Preparation of Contractile ProteinsPregnant and nonpregntnt uteri obtained from the operating room were immediately frozen at -70 C. When required, the tissue was rapidly thawed, cut into pieces, homogenized, and treated with high-ionic-strength (0.3 M) KC1 to extract the myosin and then with acetone to produce a dried tissue powder. Actin was prepared from the powder as previously d e~c r i b e d .~ Briefly, the 489

Motility of bile canaliculi in the living animal: implications for bile flow.

“…It has been generally considered that these are rigid channels that lack motility and that function as mere conduits conveying bile to the portal bile ducts. However, the bile canaliculi are surrounded by a rich investment of actin filaments (20,21,28,29,33,34,39,40,50,53). Using isolated hepatocyte couplets and time lapse microscopy, it has been shown that bile canaliculi repeatedly open and close and that this motion is accompanied by the expelling of a bolus of bile (52,55).…”

Smooth muscle associated antigen in astrocytes and astrocytomata

“…H[ctorologouis a-itisera raised against contractile proteins including actin, myosinl, meromyosin and tropomyosin give p)atterns of stainiing similar to that obtained with SMA serum with the reactionls abolislhed by absorption with the corresponding pLre antigen (Trenchev, Snieyd anid Holborow, 1974). Antiserato actini and meromyosin have also been demonstrated to react with microfilaments ultrastructurally (Holborow et al, 1975). These observations suggest that SMA serum contains a group of antibodies to antigenically related contractile proteins associated with microfilaments and validate its use as a marker for these proteins in tissues.…”

What is Actin Doing in the Liver Cell?†

“…Actin filaments are present throughout the cytoplasm of the hepatocyte but are especially numerous beneath the cell membranes in the region of bile canaliculi. This has been shown by electron microscopy (17,18), heavy meromyosin binding (18,19), by the use of anti-smooth muscle (20) and specific anti-actin antibodies (21), and by biochemical analysis of liver cell fractions (2,22). It must be remembered that bile canaliculi have no smooth muscle investment, elastica, or any structural support aside from that provided by membranes of contiguous liver cells.…”