Biomedical subjects

R J Ruch

Publications and source records attributed to R J Ruch.

70 records · Page 4Linked to original sources

PubMed1987

Strain and species effects on the inhibition of hepatocyte intercellular communication by liver tumor promoters.

The effect of the liver tumor promoters phenobarbital (PB), 1,1-bis(4-chlorophenyl)-2,2,2-trichlorethane (DDT), and dieldrin on gap junction-mediated intercellular communication between primary cultured hepatocytes from male mice (B6C3F1), C3H, C57BL, and Balb/c strains) and male F344 rats was determined. Intercellular communication was detected autoradiographically as the passage and incorporation of [5-3H]uridine nucleotides from prelabelled donor hepatocytes to donor-contacting recipient hepatocytes. At non-toxic concentrations, PB (20-500 micrograms/ml) inhibited intercellular communication between B6C3F1, C3H, and Balb/c mouse hepatocytes and F344 rat hepatocytes, but not between C57BL mouse hepatocytes. DDT (1-10 micrograms/ml) inhibited intercellular communication between hepatocytes from all 4 strains of mice and the F344 rat. Dieldrin (1-10 micrograms/ml) inhibited intercellular communication between hepatocytes from the 4 strains of mice but not between rat hepatocytes. These findings showed a good correlation with the in vivo liver tumor promoting/hepatocarcinogenic actions of PB, DDT and dieldrin in the 4 mouse strains and the F344 rat strain.

Animals↗

PubMed1986

Effects of tumor promoters, genotoxic carcinogens and hepatocytotoxins on mouse hepatocyte intercellular communication.

Intercellular communication via gap junctions may be an important mechanism of cellular growth control. Tumor promoters can inhibit intercellular communication between cultured cells, while genotoxic carcinogens apparently lack this capability. The inhibition of intercellular communication by tumor promoters may be an essential mechanism by which tumor promotion occurs in vivo. In this study, the liver tumor promoters phenobarbital, lindane (1,2,3,4,5,6-hexachlorocyclohexane, gamma-isomer), DDT (1,1-Bis[4-chlorophenyl]-2,2,2-trichloroethane), Aroclor 1254 (a polychlorinated biphenyl mixture) and dieldrin inhibited intercellular communication between male B6C3F1 mouse hepatocytes in primary culture. Intercellular communication was detected as the passage of [5-3H]uridine nucleotides from pre-labelled donor hepatocytes to non-labelled recipient hepatocytes. Mouse hepatocyte intercellular communication was also inhibited by the skin tumor promoter TPA (12-0-tetradecanoyl-phorbol-13-acetate), but not by the bladder tumor promoter saccharin. The genotoxic hepatocarcinogens dimethylnitrosamine, diethylnitrosamine, benzo[a]pyrene and 2-acetylaminofluorene, and the hepatocytotoxins bromobenzene, acetaminophen, carbon tetrachloride, chloroform and methotrexate had no effect on mouse hepatocyte intercellular communication at non-cytotoxic levels. These results suggest that the ability to inhibit mouse hepatocyte intercellular communication is an effect specific to tumor promoters.

Animals↗

PubMed1986

Antioxidant prevention of tumor promoter induced inhibition of mouse hepatocyte intercellular communication.

The liver tumor promoters, phenobarbital (20-500 micrograms/ml), lindane (1,2,3,4,5,6-hexachlorocyclohexane, gamma-isomer; 0.1-5.0 micrograms/ml), and DDT (1,1-bis[4-chlorophenyl]-2,2,2-trichloroethane; 0.5-10.0 micrograms/ml), and the hydrogen peroxide-generating enzyme, glucose oxidase (0.01-0.10 units/ml) inhibited gap junctional intercellular communication between B6C3F1 mouse hepatocytes in primary culture. Addition of the antioxidants, superoxide dismutase (100 units/ml), DPPD (N,N'-diphenyl-1,4-phenylenediamine; 25 microM), and vitamin E (DL-alpha-tocopherol acetate; 100 microM), to tumor promoter-treated cultures prevented the inhibition of hepatocyte intercellular communication. DPPD and vitamin E, prevented the inhibition of hepatocyte intercellular communication by glucose oxidase. Superoxide dismutase had no effect on the inhibition of intercellular communication caused by glucose oxidase. These results suggest that activated oxygen species are produced during liver tumor promoter treatment of cultured mouse hepatocytes and are responsible for the inhibition of mouse hepatocyte intercellular communication by the promoters.

Animals↗

PubMed1986

Mechanisms of chloroform and carbon tetrachloride toxicity in primary cultured mouse hepatocytes.

Mechanisms of chloroform (CHCl3) and carbon tetrachloride (CCl4) toxicity to primary cultured male B6C3F1 mouse hepatocytes were investigated. The cytotoxicity of both CHCl3 and CCl4 was dose- and duration-dependent. Maximal hepatocyte toxicity, as determined by lactate dehydrogenase leakage into the culture medium, occurred with the highest concentrations of CHCl3 (5 mM) and CCl4 (2.5 mM) used and with the longest duration of treatment (20 hr). CCl4 was approximately 16 times more toxic than CHCl3 to the hepatocytes. The toxicity of these compounds was decreased by adding the mixed function oxidase system (MFOS) inhibitor, SKF-525A (25 microM) to the cultures. The addition of diethyl maleate (0.25 mM), which depletes intracellular glutathione (GSH)-potentiated CHCl3 and CCl4 toxicity. The toxicity of CHCl3 and CCl4 could also be decreased by adding the antioxidants N,N'-diphenyl-p-phenylenediamine (DPPD) (25 microM), alpha-tocopherol acetate (Vitamin E) (0.1 mM), or superoxide dismutase (SOD) (100 U/mL) to the cultures. These results suggest that: in mouse hepatocytes, both CHCl3 and CCl4 are metabolized to toxic components by the MFOS; GSH plays a role in detoxifying those metabolites; free radicals are produced during the metabolism of CHCl3 and CCl4; and free radicals may be important mediators of the toxicity of these two halomethanes.

Animals↗

PubMed1986

Carcinogenicity of chlorinated methane and ethane compounds administered in drinking water to mice.

The chlorinated hydrocarbons chloroform (CHCl3), 1,1-dichlorethane (1,1-DCE) and 1,2-dichloroethane (1,2-DCE) have been detected in finished drinking water. When administered to B6C3F1 mice by gavage in corn oil, these compounds have been shown to induce hepatic tumors. The present study examines the effect on liver tumor incidence of continuous treatment of CHCl3 (600 mg/L and 1800 mg/L), 1,1-DCE (835 mg/L and 2500 mg/L), and 1,2-DCE (835 mg/L and 2500 mg/L) administered in drinking water to male B6C3F1 mice using a two-stage (initiation/promotion) treatment protocol. Seventy 4-week-old male B6C3F1 mice constituted each treatment group. Of these mice, 35 were initiated by treatment with diethylnitrosamine (DENA) (10 mg/L) in the drinking water for 4 weeks. The remaining 35 received deionized drinking water. Each group was subsequently treated with one of two concentrations of CHCl3, 1,1-DCE, or 1,2-DCE in drinking water for 52 weeks. An additional group received phenobarbital (PB) (500 mg/L) and served as the positive control for liver tumor promotion. Mice were sampled after 24 weeks (10 mice) and 52 weeks (25 mice). At sampling, liver and lung tumors were detected. None of the compounds increased the number or incidence of lung or liver tumors by themselves. PB promoted liver tumor formation (but not lung tumors) in the DENA-initiated mice. 1,1-DCE and 1,2-DCE did not affect the incidence or number of liver or lung tumors in the DENA-initiated animals. CHCl3, however, inhibited liver and lung tumorigenesis in the DENA-initiated mice.

Animals↗

PubMed1986

A nonviral, virus strain-specific antigen expressed on rat cells transformed by avian sarcoma virus.

Five of six rat sarcomas, induced by the Schmidt-Ruppin (SR) strain of avian tumor virus, expressed a Mr 60,000 tumor cell surface antigen (TSA), immunoprecipitable from non-ionic detergent extracts. Expression of the antigen was exclusive to rat cells transformed by the SR virus strain. Moreover, expression of TSA appeared restricted by cell type. The five TSA-positive SR-transformed rat cell lines tested were apparently of fibroblastic origin, but cultured rat cerebral endothelial cells (RCE-T1), transformed by SR virus, showed no expression of TSA. However, the antigen emerged on cultured tumors obtained after histoincompatible transplantation of these cells into newborn rats of another strain (tumor digest cells). Investigation of TSA for immunological relationship to viral structural antigens and the src gene product indicated that the TSA is distinct from any of these and more probably derives from a virus-directed alteration in a host molecule.

Animals↗

PubMed1986

Effect of phenobarbital on the development of liver tumors in juvenile and adult mice.

The effect of long-term exposure to phenobarbital (CAS: 50-06-6) subsequent to tumor initiation on the development of liver tumors in BALB/c and (C57BL/6 X C3H/Anf)F1 (B6C3F1) mice was determined. In male B6C3F1 mice that received either 15 or 45 ppm diethylnitrosamine [(DENA) CAS: 55-18-5] between 6 and 10 weeks of age, subsequent treatment with 500 ppm sodium phenobarbital in the drinking water resulted in the promotion of liver tumors. However, in male B6C3F1 mice initiated on day 15 of age with 25 mg DENA/kg, beginning long-term treatment of 500 ppm sodium phenobarbital at 4 weeks of age inhibited the development of liver tumors, whereas in male BALB/c mice initiated with 25 mg DENA/kg on day 15 of age, beginning the long-term treatment with 500 ppm sodium phenobarbital at 4 weeks of age promoted the development of liver tumors. Hence phenobarbital can either enhance or inhibit the formation of liver tumors, depending both on the mouse strain used and the animal's age at the start of exposure.

Adenoma↗

PubMed1985

Trout hepatocyte culture: isolation and primary culture.

Rainbow trout (Salmo gairdneri) hepatocytes were isolated using a two-step perfusion through the portal vein. A typical perfusion yielded 2.92 X 10(6) liver cells with a mean viability of 96.3%. Hepatocytes comprised 93.4% of the total cell isolate. Survival of hepatocytes in suspension culture was dependent on fetal bovine serum concentration and temperature of incubation. Serum concentrations of 5, 10, and 20% produced the highest survival during primary culture. Hepatocyte survival was in inverse proportion to the incubation temperature. Trout hepatocyte DNA synthesis and mitosis decreased during the culture period. Cytochrome p450 activity decreased rapidly during the first 2 d of culture and then remained low but measurable during the remaining 8 d of culture. Culture temperature also influenced the p450 activity with lower temperatures producing greater activity. Morphologic changes occurred in the cells during culture. Isolated hepatocytes self-aggregated, forming strands and clumps that increased in size with time in culture. Junctional complexes between cells were evident within the aggregates. Nuclear atypia, increases in size and number of autophagic vacuoles, and the appearance of bundles of intermediate filaments also were observed with increased time in culture.

Animals↗

PubMed1985

Selective resistance to cytotoxic agents in hepatocytes isolated from partially hepatectomized and neoplastic mouse liver.

Hepatocytes were isolated from B6C3F1 male mouse neoplastic livers (containing hepatocellular adenomas and carcinomas) or two-thirds partially hepatectomized livers and tested in primary culture for their cytotoxic response to hepatotoxins. Partially hepatectomized mouse hepatocytes were less sensitive to lindane, methotrexate, diethylnitrosamine and adriamycin, and more sensitive to cycloheximide compared to normal mouse hepatocytes. Neoplastic hepatocytes were less sensitive to lindane and methotrexate, did not differ in cytotoxic response to diethylnitrosamine and adriamycin and were more sensitive to cycloheximide compared to normal mouse hepatocytes.

Animals↗

PubMedDate not supplied

Inhibition of connexin43 gap junctional intercellular communication by TPA requires ERK activation.

The phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA), is a potent inhibitor of gap junctional intercellular communication (GJIC). This inhibition requires activation of protein kinase C (PKC), but the events downstream of this kinase are not known. Since PKC can activate extracellular signal regulated kinases (ERKs) and these also downregulate GJIC, we hypothesized that the inhibition of GJIC by TPA involved ERKs. TPA treatment (10 ng/ml for 30 min) of WB-F344 rat liver epithelial cells strongly activated p42 and p44 ERK-1 and -2, blocked gap junction-mediated fluorescent dye-coupling, and induced connexin43 hyperphosphorylation and gap junction internalization. These effects were completely prevented by inhibitors of PKC (bis-indolylmaleimide I; 2 microM) and ERK activation (U-0126; 10 microM). These data suggest that ERKs are activated by PKC in response to TPA treatment and are downstream mediators of the gap junction effects of the phorbol ester.

Animals↗

PubMedDate not supplied

Defective gap junctional intercellular communication in lung cancer: loss of an important mediator of tissue homeostasis and phenotypic regulation.

Gap junctions provide direct pathways for the exchange of molecules and ions between neighboring cells, a process known as gap junctional intercellular communication (GJIC). This GJIC is important for homeostasis and regulation of mitosis, differentiation, and apoptosis. Gap junctions are present in lung airway and alveolar epithelial cells and, in addition to the above roles, might coordinate ciliary beating and surfactant secretion. GJIC is decreased in human and mouse lung carcinoma cells because of reduced expression of the gap junction protein, connexin43 (Cx43), and defects in signal transduction pathways that mediate Cx43 function. This reduced GJIC is important in the behavior of lung carcinoma cells because forced expression of Cx43 in lung carcinoma cells inhibits their growth and tumorigenicity. In this report, we summarize our studies on the role of GJIC in lung neoplasia.

Animals↗

PubMedDate not supplied

Role of gap junctions in lung neoplasia.

Reduced gap junctional intercellular communication (GJIC) has been noted in many types of neoplastic cells and may contribute to the neoplastic phenotype. This study assessed GJIC (by fluorescent dye-coupling) and gap junction protein (connexin) expression in mouse and human lung carcinoma cell lines and investigated whether reduced GJIC was involved in their neoplastic phenotype. Dye-coupling and connexin43 (Cx43) expression were much lower in most of the carcinoma lines (16 of 22) compared to nontransformed lung epithelial cells. Other connexins were not detected. A poorly communicating mouse lung carcinoma cell line (E9) was transfected with Cx43 or transduced with Cx32 and several stable clones were isolated that had 2- to 4-fold increased dye coupling. When evaluated for growth in vitro, the population doubling times were increased and the saturation densities were decreased in the clones. When assessed for tumorigenicity, the parental E9 cells formed tumors with a 100% incidence (6/6 mice), whereas the clones varied in tumorigenic response (0-88% incidence). The best communicating clone (E9-2) was not tumorigenic. The highly communicating Cx32 clone, E9/32-9, gave a tumor incidence of 88%. These results suggest that restoration of GJIC by forced connexin expression can reduce the growth and tumorigenicity of lung carcinoma cells in a connexin-specific manner.

Adenocarcinoma↗

PubMedDate not supplied

The role of oval cells and gap junctional intercellular communication in hepatocarcinogenesis.

The role of oval cells, and Gap Junctional Intercellular Communication (GJIC) in hepatic differentiation and neoplasia is controversial. Oval cells accumulate in great number when hepatocyte regeneration is blocked following massive hepatotoxicity or after treatment with some hepatocarcinogens. This suggests oval cells are facultative stem cells or close progeny of liver stem cells that are activated only under specific conditions. Studies with oval cell lines clearly indicate that they can differentiate into hepatocytes and that neoplastic derivatives of oval cells can produce hepatocellular and biliary neoplasms. Because hepatocytes express Cx32 and biliary cells express Cx43, the differentiation of oval cells into hepatocytes or In addition, because Cx32 hemichannels and Cx43 hemichannels cannot form heterotypic patent channels, the type of connexin expressed by the differentiating oval cell will determine whether it communicates with hepatocytes or biliary epithelial cells, respectively. This communication may be necessary for the further differentiation and regulated growth of the differentiating oval cells and impairment of this GJIC may contribute to the formation of hepatocellular and cholangiocellular neoplasms. The type of connexin expressed may also determine the susceptibility of the differentiating oval cells to the various types of rodent liver tumor promoters. Thus, three major points have been developed here. First, Cx32 or Cx43 expression and GJIC with hepatocytes or biliary epithelial cells, respectively, may determine the final differentiated fate of oval cells. Secondly, blocked GJIC may determine whether oval cells progress to hepatocellular or cholangiocellular carcinoma. Lastly, the ability of tumor promoters to block Cx32 or Cx43-mediated GJIC in differentiating oval cells may determine whether these agents promote the formation of hepatocellular or cholangiocellular carcinomas. Thus, GJIC may be the key factor in the differentiation of oval cells and blocked GJIC may promote their neoplastic transformation in a lineage-specific manner. In this chapter, we have outlined several new hypotheses on the role of oval cells and GJIC in hepatocarcinogenesis. We hope that other investigators will consider our ideas, but realize these views will be contentious to many. Our intent, however, was to stimulate discussion and debate, even argument, because truth often arises amidst controversy and may be found in the most peculiar places.

Animals↗

PubMedDate not supplied

The role of gap junctional intercellular communication in neoplasia.

Gap junctions are comprised of proteinaceous, plasma membrane channels that link the interiors of adjacent cells and permit cells to directly exchange small (< 1,000 Daltons) molecules and ions. This exchange, termed gap junctional intercellular communication (GJIC), appears to be involved in growth regulation. Growth controlling factors may pass between cells through the junctions. The loss of gap junctions or impairment of their permeability has been observed in many neoplastic cells and cells treated with growth promoting carcinogens and other agents. The loss of GJIC appears to be an important event in the conversion of a normal cell into a neoplastic one. On the other hand, the restoration of GJIC in neoplastic cells by transfection with gap junction protein (connexin) cyclic deoxyribonucleic acids (cDNAs) or by stimulating endogenous connexin gene expression has led to the reversal of the neoplastic phenotype. The biology of gap junctions and their role in growth regulation and neoplasia are reviewed.

Animals↗

PubMedDate not supplied

Enhancement of gap junctional intercellular communication by dibutyryl cyclic AMP in lung epithelial cells.

Reduced gap junctional intercellular communication (GJIC) occurs in neoplastic cells and contributes to their phenotype. Cyclic AMP agonists inhibit lung cancer cell growth and enhance GIIC in other cell types, but little is known about their effects on lung epithelial cell gap junctions. We have examined whether N6, 2'-O-dibutyryladenosine 3':5'-cyclic mono-phosphate (DBcAMP) affected GJIC, gap junction protein (connexin43) expression, and the growth of non-transformed and neoplastic mouse lung epithelial cells. DBcAMP (0.01.1 mM) stimulated GJIC (assayed by fluorescent dye microinjection) and connexin43 expression (assessed by Northern and Western blotting) and reduced their proliferation. These results suggest an association between cAMP growth inhibition and enhanced GJIC in lung epithelial cells.

Animals↗

PubMedDate not supplied

Role of protein kinase C in the deficient gap junctional intercellular communication of K-ras-transformed murine lung epithelial cells.

BACKGROUND: Protein kinase C (PKC) activity is increased and gap junctional intercellular communication (GJIC) is decreased frequently in Ras-transformed cells. We investigated the roles of Ras and PKC in the deficient gap junctional intercellular communication (GJIC) of K-ras-transformed E9 mouse lung carcinoma cells. METHODS: GJIC was measured by fluorescent dye microinjection. Ras activity was blocked with lovastatin or a K-ras antisense oligonucleotide. PKC activity was inhibited with GF 109203X or apigenin or was downregulated by overnight treatment with 12-O-tetradecanoylphorbol-13-acetate. The content and phosphorylation of the gap junction protein, connexin43 (Cx43), was assessed by Western blot. RESULTS: E9 cell GJIC was increased two-three fold by lovastatin, the K-ras antisense oligonucleotide, and PKC inhibition/downregulation. Cx43 content and phosphorylation were unchanged, however. CONCLUSIONS: Oncogenic Ras blocks GJIC in E9 cells through a PKC-dependent mechanism, but this does not directly involve Cx43 expression or phosphorylation.

Animals↗