Unusual Tubular Inclusions in the Mitochondrial Matrix from Human Livers with Cholelithiasis

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1 Arch. histol. jap., Vol. 44, No. 3 (1981) p Unusual Tubular Inclusions in the Mitochondrial Matrix from Human Livers Cholelithiasis Torao YAMAMOTO and Takeharu HISATSUGU Department of Anatomy (Prof. T. YAMAMOTO), Faculty of Medicine, Kyushu University, Fukuoka and Department of Surgery (Prof. T. HISATSUGU), Saga Medical College, Saga, Japan Received November 10, 1980 Summary. Unusual tubular in the mitochondrial matrix of human hepatocytes were found in 7 out of 15 cases cholelithiasis. These were composed of tubular structures which were about 50nm in diameter and varying in number. The wall of the individual tubules appeared to consist of about 10 tubular subunits a diameter of about 5nm. Although some relationship between occurrence of these tubular and production of gallstones might be suggested, their origin and functional significance are unknown. It has been well known that the mitochondria of human hepatocytes often contain inclusion bodies, the most frequent of which are crystalline of filamentous. These have been observed in the mitochondrial matrix of hepatocytes not only under pathological (JEZEQUEL, 1959; EKHOLM and EDLUND, 1960; SCHAFFNER et al., 1963; BHAGWAT and Ross, 1971; and others), but also normal conditions (MUGNAINI, 1964; WILLS, 1965; MA and BIEMPICA, 1971). During the course of ultrastructural examination of the liver biopsies from patients cholelithiasis, we have found unusual tubular in the mitochondrial matrix which have probably not been reported so far. The present paper deals some ultrastructural aspects of these interesting intramitochondrial. MATERIALS AND METHODS Small pieces of the liver were removed by means of surgical biopsy from fifteen patients cholelithiasis. Out of these fifteen cases, seven were confirmed during operation to possess mixed stones consisting of cholesterol and calcium bilirubinate in the gall bladder, four both in the gall bladder and the biliary ducts. In four cases pure bilirubinate stones were found in the common ducts. Liver biopsies were also carried out for two patients out cholelithiasis as controls. Immediately after biopsy, small pieces of the liver were cut into small bits in a drop of fixative, then transferred into and fixed in 3% glutaraldehyde buffered 0.1 M cacodylate at ph 7.4 for 2hr, and postfixed in 1% osmic acid adjusted to ph

2 258 T. YAMAMOTO the same embedded toluidin in blue, Fig. 1. buffer and resin. examined in Tubular the dehydration sections in an ordinary filamentous cholelithiasis. matrix cholelithiasis. After Thick Intramitochondrial seen 2. T. HISATSUGU: for 2hr. epoxy patient Fig. and light in the light mitochondrial in and ethanol, microscopy microscope. Bundles of mitochondria. 18,000 in graded for the lattice Thin biopsied pattern specimens were were stained sections for elec- hepatocyte from of filaments can a be 30,000 matrix from a human hepatocyte

3 Tubular Iron microscopy were stained 100 B electron microscope. Fig. 3. Tubular arrows) Fig. 4. profiles of both from A cross-sectional structures Inclusions lead tartrate in longitudinal a human tubules can be Mitochondrial solution (arrow) hepatocyte view of tubular individual in the clearly and examined and cross-sectional cholelithiasis. at higher seen. Matrix in a JEM (double 45,000 magnification. 120, Subunit

4 260 T. YAMAMOTO and T. HISATSUGU: OBSERVATIONS The mitochondria of human livers cholelithiasis did not show any marked differences in appearance from those of normal human livers. Most common were the wide and a few microns long. Occasionally giant mitochondria in irregular profile could be seen in some hepatocytes. In all the livers cholelithiasis examined, some hepatocytes contained mitochondria possessing inclusion bodies in their matrix. These inclusion bodies were of two types: one was of filamentous in crystal array and the other was of tubular ones arranged in parallel. Filamentous showed the same structural patterns as those described in many previous reports and were observed so frequently in almost all the hepatocytes examined here (Fig. 1). Giant mitochondria tended to contain more frequently these than did other, normal mitochondria. On the other hand, tubular were observed in 7 cases out of 15, but not in the livers out any cholelithiasis. These consisted of tubular elements varying in number (Fig. 2). In longitudinal section, individual tubules showed two parallel thin lines and sometimes an additional central line between them (Fig. 2, 3). Such longitudinal profiles of these tubules tended to be slightly tapered toward their ends, suggesting their slow twisting along the length (Fig. 3). In cross sections, individual tubules showed as a whole a ring profile a diameter of 45-60nm, mostly 50nm (Fig. 3, 4). At higher magnification a wall of tubules was found to be formed by a circular array of about 10 fine dots of subunit structures, and often contain one or two central dots (Fig. 4). Further careful observation revealed that such fine dots of subunits appeared to be of ring form in profile and about 5nm in diameter. There was no consistent relationship between the occurrence of tubular and the localization or the chemical nature of gallstones. The values of serum glutamic oxaloacetic transaminase (SGOT), serum glutamic pyruvic transaminase (SGPT), or alkaline phosphatase examined in clinical laboratory tests also did not show any significant correlation the occurrence of these. DISCUSSION There have been many reports dealing the filamentous in the matrix of mitochondria from human hepatocytes. These are known to occur in the hepatocytes not only under pathological but also under normal conditions. Besides these filamentous, the present observation has demonstrated tubular in the mitochondrial matrix of hepatocytes cholelithiasis. Similar have been described in the epithelial cells of the uriniferous tubule of snakes but not in the hepatocytes (KUROSUMI, MATSUZAWA and WATARI, 1966; YAMAMOTO, EBE and KOBAYASHI, 1969). No report has been published on the occurrence of this kind of tubular in human cells. The tubules in the human liver are twice as large in diameter (about 50nm) as those (about 25nm) in the snake kidney. The present study has revealed

5 Tubular Inclusions in the Mitochondrial Matrix 261 a more detailed structural organization of these, in which approximately 10 tubular subunits a diameter of about 5nm seem to be arrayed circularly, forming individual tubules of 50nm in dimeter as a whole. Though the origin and functional significance of these are not known at present, they are presumably proteineous in nature. These tubules seem somewhat similar in appearance to the cytoplasmic microtubules reported elsewhere (LEDBETTER and PORTER, 1963; MCNABB and SANDBORN, 1963; SLAUTTERBACK, 1963; SANDBORN et al., 1964; and others). However, judging from their cross sectional views, they are in reality different from the latter in structural organization. Since the subunit structure of these tubules resembles each component of filamentous in terms of dimension and appearance, there may be the possibility that the tubular are a type of filamentous inclusion, in which filamentous components are arranged not in crystalline but in circular form. An alternative possibility might be that the tubular are composed of actin filaments involved in the movement of mitochondria, because of the structural resemblance of their subunits to the microvillous filaments of the intestinal absorptive cells which are known to be actin (MOOSEKER and TILNEY, 1975). Of interest may be the fact that the tubular were observed in relatively high frequency (about 46%) in the liver biopsies of cholelithiasis, but not in those of the 2 cases out cholelithiasis. Thus, one might be tempted to presume that this fact reflects such metabolic disorders in the liver as results in the production of gallstones. However, the present study does not provide any concrete evidence to support or deny this assumption. Further study must be required in this connection.

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