A method for simultaneous acquisition of information on cytochemical and microangiographic characteristics of tissue after perfusion with contrast medium was developed. The technique is based on the ...use of hydrosoluble resin as embedding material and is illustrated by the demonstration of gamma-glutamyltranspeptidase activity and of the microcirculation in rat liver during chemically induced hepatocarcinogenesis and experimental metastasis. The method is applicable to investigations aimed at correlating microangiographic and cytochemical characteristics of normal or altered tissues.
The use of sonicated phospholipid vesicles (liposomes) as carriers of 2-3'-(methoxycarbonylamino)-phenyl-3-phenyl-6-methoxycarbonylamino-4-(3H)- quinazolone (NSC-251635), a water-insoluble ...antimitotic compound, was investigated in mice. NSC-251635 was incorporated in egg yolk lecithin, cholesterol, and stearylamine (4:3:1) liposomes. In vitro, NSC-251635 in suspension or entrapped in liposomes was not toxic for L1210 cells. In vivo, after ip or iv injections to CDF1 mice bearing intraperitoneal or intravenous L1210 leukemia, NSC-251635 was active only when it was incorporated in the liposomes and not when it was given as a suspension in Klucel or in saline. The NSC-251635 liposome preparation induced significantly prolonged survival of the treated animals.
Neoplastic liver cell colonies were induced in the livers of isogeneic F344 rats by intraportal injection of a hepatic cell suspension from diethylnitrosamine-treated donor rats. Examination of the ...livers 12 days after cell implantation revealed well-demarcated groups of liver cells. The colonies showed alterations of the normal hepatocyte phenotype, which were clearly demonstrated by histologic, cytochemical, and electron microscope techniques. The hepatocytes were markedly deficient in glucose-6-phosphatase and bile canalicular ATPase activities, and they contained numerous mitotic figures. Scanning and transmission electron microscopy allowed characterization of hepatocyte interfaces and the shape of sinusoids and the biliary network. The nodular colonies displayed disorganized, thickened trabeculae separated by dilated sinusoids. In these colonies the hepatocytes proliferated intensely and formed, inside the host parenchyma, revascularized, integrated nodular structures. However, these hepatocytes showed ultrastructural anomalies: large nuclei with prominent nucleoli, many free polysomes, and areas of proliferated smooth endoplasmic reticulum in connection with unfolded cisternae of the rough endoplasmic reticulum. All of these features agreed with the hypothesis previously proposed that the colonies may be precursors of the hepatocarcinomas that ultimately develop in animals given injections of treated liver cells. Direct confirmation, however, still is needed.
The cytochemical localization of glucose-6-phosphatase (G6Pase) and its biochemical quantification were studied in isolated and cultured adult rat parenchymal cells. Appropriate technical conditions ...were chosen to assume adequate ultrastructural preservation and retention of enzyme activity. Isolated hepatocytes separated by collagenase perfusion were shortly fixed in glutaraldehyde and entrapped in a pellet of fibrin. Frozen sections, 50 microns in thickness were incubated for cytochemical demonstration of G6Pase, in a slightly modified Wachstein-Meisel medium. Hepatocytes in culture, fixed for 1 min in glutaraldehyde, were impregnated in a 10% cryoprotective glycerol solution and quickly frozen in liquid nitrogen at -170 degrees C in order to induce penetration of the substrate. In these conditions, a homogeneous distribution of the enzyme was observed in both isolated and cultured cells. The cytochemical reaction appears continuous in the smooth and rough endoplasmic cisternae and in the nuclear envelope. Lead phosphate deposits, although evenly distributed, are reduced in intensity after 48 h culture. Biochemical determinations reveal the presence of a high specific enzymatic activity in isolated cells (108 nmolP/min/mg proteins), which decreases in culture, respectively to 70 and 50% of the original value, after 24 and 48 h culture. G6Pase induction by glucagon was obtained after 48 and 72 h in culture.
Oral administration of diethylnitrosamine (DENA) at the final concentration of 70 mg/kg, 24h after partial hepatectomy induces the appearance of preneoplastic foci composed of structurally altered ...and enzymatic deficient liver parenchymal cells of large size. These lesions grow up and form hyperplastic nodules, 0.5 to 2mm in diameter, 9 to 10 months after DENA treatment. Adult rat hepatocytes were isolated by collagenase perfusion from treated rats, sacrificed 6 months after the carcinogen application. They were separated by elutriation in distinct subpopulations, according to cell size and degree of ploidy. Fractions elutriated at high flow rates are enriched in preneoplastic hepatocytes of large size, which display a heterogeneous distribution of microvilli. Hyperplastic nodules were examined in situ by scanning electron microscopy, in 1 to 2 mm thick tissue slices, 10 months after the carcinogen administration. The nodules, easily detected at this stage of transformation, are composed of hypertrophied hepatocytes which exhibit heterogeneous cell surfaces. The parenchymal cells reveal frequently reduced interhepatocytic faces provided with sinuous bile canaliculi and sinusoidal faces covered with closely apposed, short microvilli.
