Prognostic importance of tumour-infiltrating memory T cells in oesophageal squamous cell carcinoma.

Memory T cells survive for many months and years and are critically important for host defence in humans. In tumour immunity, they have been also suggested to play a significant role in tumour progression and metastasis. However, the role of memory T cells in actual human cancer remains largely unknown. In this study, the clinical importance of tumour-infiltrating CD45RO(+) memory T cells was investigated in human oesophageal squamous cell carcinoma (OSCC). CD45RO(+) T cells were evaluated by immunohistochemistry in primary OSCC tumours from 105 patients. Patients were classified into two groups as CD45RO(+hi) or CD45RO(+lo) based on the number of cells stained positively for CD45RO. No significant difference was observed between CD45RO status and several clinicopathological prognostic factors. However, the postoperative overall and disease-free survival for CD45RO(+hi) patients was significantly better than for CD45RO(+lo) patients. Furthermore, there were significant correlations of CD45RO status in the primary tumour with postoperative lymph node and pulmonary recurrence, suggesting that memory T cells may control postoperative metastatic recurrence. Most importantly, CD45RO(+) memory T cell status has a significant prognostic value for OSCC independently of conventional tumour-node-metastasis (TNM) classification. Our study may provide a rationale for developing a novel immunotherapy in intentional induction of memory T cells for the treatment of oesophageal cancer.

A comparison of surgery and radiation therapy for cT1 esophageal squamous cell carcinoma.

Surgery and radiation therapy have been used to treat esophageal squamous cell carcinoma. However, treatment outcomes have not yet been extensively investigated. The aim of this study was to compare surgery and radiation therapy for clinical T1 esophageal squamous cell carcinoma. A total of 67 clinical T1 esophageal squamous cell carcinoma patients were treated between January 1997 and December 2005; 29 had undergone radical esophagectomy (surgery group) and 38 were treated with definitive radiation therapy (radiation group). The mean patient age was lower in the surgery group than in the radiation group. In surgery group, respiratory complications, anastomotic leaks, recurrent nerve palsies, and anastomotic stenosis occurred in 7, 8, 6, and 5 patients, respectively. In radiation group, leucopenia, esophagitis, pericarditis were observed in 15, 3, and 3 patients, respectively. The 5-year overall survival rate for the surgery group was 68.9%, and 74.3% for the radiation group. There were no significant difference between groups (P= 0.3780). The 5-year relapse-free survival rate in the surgery group was 61.8% and 38.8% in the radiation group. The relapse-free survival rate was significantly higher in the surgery group than in the radiation group (P= 0.0051). The 5-year overall and relapse-free survival rates for tumors invaded into but not through the muscularis mucosa were 83.3% and 75.0%, respectively, in the surgery group and 78.8% and 33.3%, respectively, in the radiation group. There were no significant differences. The 5-year overall survival rates for patients with tumors that invaded the submucosal layer was 64.9% in the surgery group and 66.5% in the radiation group. This difference was not significant (P= 0.8712). The 5-year relapse-free survival rate in the surgery group (56.0%) was significantly higher than that in the radiation group (41.8%; P= 0.0219). In conclusion, surgery may become a standard treatment for cT1 esophageal cancer that can offer longer relapse-free survival, particularly for patients with tumors that invade the submucosa.

Practical evaluation of Mung bean seed pasteurization method in Japan.

The majority of the seed sprout-related outbreaks have been associated with Escherichia coli O157:H7 and Salmonella. Therefore, an effective method for inactivating these organisms on the seeds before sprouting is needed. The current pasteurization method for mung beans in Japan (hot water treatment at 85 degrees C for 10 s) was more effective for disinfecting inoculated E. coli O157:H7, Salmonella, and nonpathogenic E. coli on mung bean seeds than was the calcium hypochlorite treatment (20,000 ppm for 20 min) recommended by the U.S. Food and Drug Administration. Hot water treatment at 85 degrees C for 40 s followed by dipping in cold water for 30 s and soaking in chlorine water (2,000 ppm) for 2 h reduced the pathogens to undetectable levels, and no viable pathogens were found in a 25-g enrichment culture and during the sprouting process. Practical tests using a working pasteurization machine with nonpathogenic E. coli as a surrogate produced similar results. The harvest yield of the treated seed was within the acceptable range. These treatments could be a viable alternative to the presently recommended 20,000-ppm chlorine treatment for mung bean seeds.

Monoclonal antibody 7H6 reacts with a novel tight junction-associated protein distinct from ZO-1, cingulin and ZO-2.

The tight junction is an essential element of the intercellular junctional complex; yet its protein composition is not fully understood. At present, only three proteins, ZO-1 (Stevenson, B. R., J. D. Siliciano, M. S. Mooseker, and D. A. Goodenough. 1986. J. Cell Biol. 103:755-766), cingulin (Citi, S., H. Sabanay, R. Jakes, B. Geiger, and J. Kendrick-Jones. 1988. Nature (Lond.). 333:272-275) and ZO-2 (Gumbiner, B., T. Lowenkopf, and D. Apatira. 1991. Proc. Natl. Acad. Sci. USA. 88:3460-3464) are known to be associated with the tight junction. We have generated a monoclonal antibody (7H6) against a bile canaliculus-rich membrane fraction prepared from rat liver. This 7H6 antigen was preferentially localized by immunofluorescence at the junctional complex regions of hepatocytes and other epithelia, and 7H6-affiliated gold particles were shown electron microscopically to localize at the periphery of tight junctions. Immunoblot analysis of a bile canaliculus-rich fraction of rat liver using 7H6, anti-ZO-1 antibody (R26.4C), and anti-cingulin antibody revealed that 7H6 reacted selectively with a 155-kD protein, whereas R26.4C reacted only with a 225-kD protein. Anti-cingulin antibody reacted solely with 140 and 108-kD proteins, indicating that the protein recognized by 7H6 is immunologically different from ZO-1 and cingulin. Immunoprecipitation of detergent extracts obtained from metabolically labeled MDCK cells with R26.4C coprecipitated a 160-kD protein, which corresponds to ZO-2, with ZO-1. However, 7H6 did not react with the 160-kD protein. These results strongly suggest that the 7H6 antibody recognizes a novel tight junction-associated protein different from ZO-1, cingulin and ZO-2.

Evaluation of the efficacy and safety of olopatadine and fexofenadine compared with placebo in Japanese cedar pollinosis using an environmental exposure unit.

BACKGROUND: Second-generation oral H1-antihistamines have become a mainstay of treatment for the symptoms of seasonal allergic rhinitis; however, the effect of olopatadine has not been widely reported to date. OBJECTIVES: To evaluate the efficacy of 2 oral H1-antihistamines, olopatadine and fexofenadine, in the treatment of the nasal symptoms of Japanese cedar pollinosis and their possible side effects. METHODS: This was a randomized, double-blind, placebo-controlled, crossover study conducted in an environmental exposure unit (EEU). Twenty volunteers suffering from Japanese cedar pollinosis were randomly divided into 3 groups and exposed to cedar pollen in the EEU with oral administration of olopatadine hydrochloride (5 mg), fexofenadine hydrochloride (60 mg), or placebo 1 hour prior to pollen exposure. Nasal symptoms, activity impairment, and subjective sleepiness were self-assessed during the study period. Attention was measured using the digit cancellation test. The trial was repeated after 4 and 7 weeks. RESULTS: Compared with placebo, olopatadine significantly improved nasal symptoms and activity impairment during pollen exposure (P < .05). There was no significant relief of nasal discharge or nasal congestion with fexofenadine throughout the 5-hour exposure to cedar pollen. Furthermore, olopatadine significantly reduced nasal congestion during the first 2 hours, as well as sneezing and nasal discharge 4 hours after admission to the EEU compared with fexofenadine (P < .05). There was no significant difference in the effect on subjective sleepiness among the 3 groups, and all 3 agents had little effect on attention. CONCLUSIONS: These findings suggest that olopatadine is more effective than placebo and fexofenadine in improving nasal symptoms of Japanese cedar pollinosis.

Combination treatments for killing Escherichia coli O157:H7 on alfalfa, radish, broccoli, and mung bean seeds.

In this study, the effectiveness of prolonged dry-heat treatment (50 degrees C) alone or in combination with chemical treatments (1% oxalic acid, 0.03% phytic acid, 50% ethanol, electrolyzed acidic water, and electrolyzed alkaline water) in eliminating Escherichia coli O157:H7 on laboratory-inoculated alfalfa, radish, broccoli, and mung bean seeds was compared with that of dry-heat treatment in combination with irradiation treatment. Dry-heat treatment for 17 or 24 h alone could reduce E. coli O157:H7 numbers to below detectable levels in radish, broccoli, and alfalfa seeds, but was unable to reduce the pathogen numbers to below the detectable level in mung bean seeds. In addition, dry-heat treatment for 17 h plus sanitizer treatments were effective in greatly reducing pathogen populations on radish, broccoli, and alfalfa seeds, without compromising the quality of the sprouts, but these treatments did not eliminate the pathogen from radish and alfalfa seeds. Seventeen hours of dry heat followed by a 1.0-kGy dose of irradiation completely eliminated E. coli O157:H7 from radish and mung bean seeds, whereas only a minimum radiation dose of 0.25 kGy was required to completely eliminate the pathogen from broccoli and alfalfa seeds. Dry heat in combination with radiation doses of up to 1.0 kGy did not negatively impact the seed germination rate or length of alfalfa, broccoli, and radish seeds or the length of alfalfa, broccoli, and radish sprouts, but did decrease the length of mung bean sprouts.

A novel red-emitting K2Ca(PO4)F:Eu2+ phosphor with a large Stokes shift.

We report a K2CaPO4F:Eu2+ phosphor with a new crystal structure. This phosphor has a large Stokes shift and converts near-ultraviolet light to red luminescence without absorption of other visible light. The mechanism was elucidated by applying a constrained density functional theory to the solved crystal structure.

Effect of lysed Enterococcus faecalis FK-23 on allergen-induced immune responses and intestinal microflora in antibiotic-treated weaning mice.

BACKGROUND: Recent epidemiological studies have indicated that early life receipt of antibiotics may be associated with an increased risk of developing atopic disorder. Lysed Enterococcus faecalis FK-23 (LFK), a probiotic product of E faecalis, has been shown to have inhibitory effects on allergen-induced immune responses in mice. OBJECTIVE: The purpose of this study was to evaluate the effects of LFK on immune responses and intestinal microflora in antibiotic-treated, and allergen-sensitized weaning mice. METHODS: Three-week-old BALB/c mice were sensitized with cedar pollen allergen to establish the experimental model. The allergen-induced peritoneal accumulation of eosinophils, serum levels of total and allergen-specific immunoglobulin (Ig) E and IgG2a, and the intestinal bacterial flora were determined in the control, antibiotic, LFK and antibiotic-LFK groups (n = 7 in all groups). Orally administered erythromycin, one kind of macrolide antibiotic, was used for the experiments. RESULTS: There was no significant difference in the allergen-induced peritoneal accumulation of eosinophils and serum specific IgE and IgG2a levels in erythromycin-treated mice compared to a control group. However, the ratio of serum total IgE to IgG2a levels was significantly increased in erythromycin-treated mice relative to that found either in LFK-treated mice or in erythromycin-treated mice with LFK supplementation. The total aerobes, total anaerobes and Enterococcus species of intestinal microflora were not significantly different among all groups. Lactobacillus species were distinctly eliminated in the mice exposed to erythromycin on day 7 and totally recovered in erythromycin-treated mice with LFK intervention on day 28, but could not be recovered in the erythromycin-treated mice without LFK intervention. CONCLUSIONS: Our results suggest that LFK may improve the intestinal ecosystem disturbed by antibiotic use, and thereby prevent subsequent development of atopy. However, whether different antibiotics have different effects on immune responses needs to be addressed further.

Effect of probiotic Bifidobacterium longum BB536 [corrected] in relieving clinical symptoms and modulating plasma cytokine levels of Japanese cedar pollinosis during the pollen season. A randomized double-blind, placebo-controlled trial.

Probiotic microorganisms have been shown to be effective in the treatment of allergic inflammation and food allergy, but their efficacy remains controversial. This study tested the effect of a yogurt supplemented with a probiotic strain Bifidobacterium longum BB536 in the treatment of Japanese cedar pollinosis (JCPsis). Forty subjects with a clinical history of JCPsis were given yoghurt either containing BB536 (BB536 yoghurt) or without BB536 (placebo yoghurt) at 2 X 100 g per day for 14 weeks, in a randomized, double-blind, placebo-controlled trial. Subjective symptoms and self-care measures were recorded daily and blood samples were taken before and during the intervention (at weeks 4, 9, and 14) to measure the blood parameter levels related to JCPsis. Yoghurt supplemented with BB536 significantly alleviated eye symptoms compared with placebo yoghurt (odds ratio 0.31; 95% confidence interval 0.10-0.97; p = 0.044). Although no statistically significant differences were detected, nasal symptoms such as itching, rhinorrhea, and blockage, as well as throat symptoms tended to be relieved with the BB536 yoghurt. BB536 tended to suppress the decreasing blood levels of interferon-gamma (IFN-y) and the increasing blood eosinophil rates; a significantly higher IFN-gamma level was observed for the difference from baseline at week 4. A decreased trend in the difference from baseline levels of JCP-specific IgE levels was also observed at week 4 in the BB536 group compared with the placebo group. In conclusion, these results suggest that intake of BB536-supplemented yoghurt may relieve JCPsis symptoms, probably through a modulating effect on Th balance.

Induction of a novel Ca2+-dependent chymotrypsin-like serine protease by tumor promoters in rat livers.

Induction of a microsomal Ca2+-dependent serine protease by hepatic tumor promoters was studied. Male F344 rats were fed a diet containing one of the following promoting agents: phenobarbital (CAS: 50-06-6), dichlorophenyltrichloroethane (CAS: 50-29-3), butylated hydroxytoluene, ethyl-alpha-chlorophenoxyisobutyrate (CAS: 128-95-0), or 17-alpha-ethynylestradiol (CAS: 57-63-6) or a nonpromoting agent, diphenylhydantoin (CAS: 57-41-0), for 1 week. By treatment with promoters, the protease activity in the microsomal fraction was increased to threefold to fivefold that of control, whereas only a slight increase of activity was found after diphenylhydantoin treatment. The Ca2+-dependent protease activity was determined with the use of N-benzoyl-L-tyrosine ethyl ester as the substrate in a medium containing 50 mM CaCl2 for its maximal activity. This protease was preferentially localized in the smooth microsomal membrane and strongly inhibited by diisopropyl phosphorofluoridate (CAS: 55-91-4), and the optimum pH of the activity was 7.8. It appears that the Ca2+-dependent serine protease measured by using a chymotrypsin substrate is a novel protease, and induction of its activity by hepatic tumor-promoting agents is a common and specific phenomenon.

A microsomal butyrylesterase appearing in rat livers during development, regeneration, and carcinogenesis, and after phenobarbital treatment.

A microsomal butyrylesterase (L-I) was purified from the livers of male W rats treated with phenobarbital, and an antiserum against this purified L-I was raised in a rabbit. By the Ouchteriony double-diffusion test, a precipitin line was observed between the anti-L-I antiserum and each Triton X-100 extract of livers during development, regeneration after partial hepatectomy, and carcinogenesis and of hyperplastic nodules and hepatomas, all of which revealed L-I in their esterase isoenzyme patterns. These precipitin lines exhibited esterase activity. The fusion of the lines of these tissue extracts and that of the purified L-I indicated the presence of an antigen site common to their esterases. The extracts of adult and fetal livers and also of hepatomas resembling fetal liver in the esterase isoenzyme pattern did not produce a precipitin line with anti-L-I antiserum.

Kinetics of phenotypic maturation of remodeling of hyperplastic nodules during liver carcinogenesis.

Hyperplastic nodules appearing during the preneoplastic phase of liver carcinogenesis were divided into two types, persistent and remodeling, according to the pattern of staining for gamma-glutamyltransferase. In the resistant cell model of liver carcinogenesis used in this study, hyperplastic nodules, uniformly staining for gamma-glutamyltransferase, rapidly emerge by 4 weeks after a single injection of diethylnitrosamine and brief selection by dietary 2-acetylaminofluorene plus partial hepatectomy. By 6 weeks, a majority of nodules (about 75%) show an obvious irregularity and loss of uniformity in staining for gamma-glutamyltransferase while the remaining nodules continue to be uniformly stained. The number of irregularly stained nodules increases over the next 18 weeks until over 95% of nodules show the nonuniform loss of enzyme activity. The progressive loss of enzyme activity is accompanied by architectural remodeling to normal-appearing liver. This is associated with the increasing disappearance of many obvious nodules from the liver as the remodeling ones blend imperceptibly with the surrounding liver. The uniformly stained nodules show the persistence of hepatocyte arrangements in plates two or more cells thick and in acini and of cytoplasmic hypertrophy characteristic of persistent hyperplastic nodules. Labeling indices are much higher in hepatocytes of the persistent uniformly stained nodules than in the remodeling ones. The possibility of exploiting this phase of the model further for in-depth analysis of the nodule-to-carcinoma sequence is discussed.

Demonstration of embryonic-type hemoglobin in extramedullary hematopoietic cells in the liver during experimental liver carcinogenesis by 3'-methyl-4-dimethylaminoazobenzene.

Using an immunofluorescent technique, we demonstrated the production of rat embryonic-type hemoglobin in extra-medullary hematopoietic cells, which appeared in the liver of adult Wistar rats fed a diet containing 0.06% 3'-methyl-4=dimethylaminoazobenzene. Specific anti-rat embryonic hemoglobin serum was prepared by immunizing a rabbit with hemoglobin from a 15- or 16-day-old Wistar rat fetus and absorbing with adult rat liver hemogenate and red blood cells. The appearance of hematopoietic foci was observed at both precancerous and cancerous stages during 3'-methyl-4-dimethylaminoazobenzene-induced liver carcinogenesis. At the cancerous stage, almost all these foci were observed in poorly differentiated hepatocellular carcinomas. The hematopoietic cells seen at both stages were stained positively with antiembryonic hemoglobin serum, but red blood cells and other types of cells in the liver did not show this specific fluorescence. These results together with earlier observations on the appearance of fetal characteristics strongly suggest that a fetal environment prevails in the adult rat liver during hepatic carcinogenesis.

Effects of delays in the cell cycle on the induction of preneoplastic and neoplastic lesions in rat liver by 1,2-dimethylhydrazine.

This study was designed to explore further the relationship between cell proliferation and the induction of early putative preneoplastic lesions by carcinogens. Rats were given a non-necrogenic dose of 1,2-dimethylhydrazine 24 hr before being subjected to partial hepatectomy. Beginning 4 hr later, hydrocortisone was injected 10 times at 4-hr intervals to delay progression through the cell cycle, including inhibition of DNA synthesis by at least 85% for about 40 hr. At the appropriate time thereafter, the putative preneoplastic hepatocytes were selectively stimulated to grow in vivo into gamma-glutamyltransferase-positive focal lesions. Animals given hydrocortisone showed a large decrease (71%) in the number of gamma-glutamyltransferase-positive foci. In contrast, when hydrocortisone was given at 6 days after partial hepatectomy, no inhibition in the induction of hepatic lesions was observed. In the next experiments, rats were treated with 1,2-dimethylhydrazine and were subjected to partial hepatectomy at 12, 24, or 48 hr or 1 week thereafter. A significant number of gamma-glutamyltransferase-positive foci was found when partial hepatectomy was performed at 12 or 24 hr but far fewer were found when the operative partial hepatectomy was delayed to 48 hr or 1 week later. Similarly, in long-term experiments, six of 14 animals developed primary hepatocellular carcinoma 13 months after the time of injection of 1,2-dimethylhydrazine when partial hepatectomy was performed at 12 hr, while none of the animals developed liver cancer when the operation was performed at 48 hr. These results imply that the majority of biochemical lesions induced by 1,2-dimethylhydrazine that are relevant to the induction of liver preneoplasia and neoplasia are short-lived and that their persistence is associated with some cellular activity closely related to the cell cycle.

Consistent disruption of the AML1 gene occurs within a single intron in the t(8;21) chromosomal translocation.

The AML1 gene on chromosome 21 was rearranged by the t(8;21) chromosomal translocation in acute myeloid leukemia (AML). Southern blot analysis of 21 AML patients with t(8;21), including three with complex translocations, t(8;V;21), demonstrated that all the breakpoints occurred at random within a single intron between two coding exons of AML1. Clustering of the breakpoints in the restricted intron suggests the formation of a unique fusion gene between the AML1 gene and a presumable counterpart gene on chromosome 8. Nucleotide sequencing of the breakpoint region revealed that the translocation event was accompanied by deletion of a short stretch of nucleotides.

Promotion of liver cancer development by brief exposure to dietary 2-acetylaminofluorene plus partial hepatectomy or carbon tetrachloride.

Adult male Fischer rats were exposed to a necrogenic dose (200 mg/kg) of diethylnitrosamine or to nonnecrogenic doses of N-methyl-N-nitrosourea, 1,2-dimethylhydrazine, or benzo(a)pyrene following partial hepatectomy or sham hepatectomy. This treatment by itself led to no hepatocellular carcinomas by 8 to 18 months, except in animals given N-methyl-N-nitrosourea, which showed a 30% incidence by 12 months. With each treatment regimen, exposure to dietary 2-acetylaminofluorene for 2 weeks coupled with partial hepatectomy or the administration of a necrogenic dose of CCl4, was associated with an incidence of 68 to 94% of cancer at 8, 12, or 18 months, depending upon the initiating carcinogen used. Appropriate controls showed either no hepatocellular carcinoma or a much lower incidence. It is concluded that the 2-week exposure to dietary 2-acetylaminofluorene plus partial hepatectomy or the administration of CCl4 has a strong promoting effect on liver carcinogenesis with four different chemical carcinogens.

Randomly controlled study of chemotherapy versus chemoimmunotherapy in postoperative gastric cancer patients.

From September 1979 through March 1981, a total of 302 patients with gastric cancer and undergoing gastrectomy at the Department of Surgery at Chiba University Hospital and its 14 affiliated hospitals was studied for clinical effectiveness of immunotherapy with Nocardia rubra cell wall skeleton. The patients were stratified by gross stage of cancer and degree of operative curability. They were then assigned randomly to either chemotherapy group or chemotherapy plus immunotherapy group. Immunotherapy used was intradermal injection of 400 micrograms of N. rubra cell wall skeleton which was given weekly for the first month and monthly thereafter. After the specimen was examined microscopically, the patients were classified by histological stage of cancer and radicality of surgical intervention into curative or noncurative groups. The patients were surveyed for survival period in December 1981. The postoperative survival rate was compared in patients of histologically curative or noncurative resection cases between the two treatment groups. No statistical difference was detected between the groups in age, sex, or operative procedures that might influence the patient's survival. As a result, statistical intergroup difference in survival rates was not seen in patients of the curative group, probably due to a short observation period. However, the intergroup difference in survival rates was statistically significant in patients of the noncurative group (p less than 0.01). These results indicate the adjunctive effect of N. rubra cell wall skeleton as an immunotherapeutic agent in patients undergoing gastrectomy for gastric cancer.

ADP-ribosylation of myelin basic protein by cholera toxin.

Cholera toxin ADP-ribosylates four types of myelin basic proteins (MBPs) of Mr 14,000, 17,500, 19,000 and 22,000 in rat brain myelin. On an analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, MBP underwent mono- and multi-(ADP-ribosyl)ation by cholera toxin and thus modified MBP migrated on the gel as several discrete protein bands, the molecular masses of which were apparently larger by 500-2000 daltons than that of the corresponding untreated MBP. On average, 1.1 mol of ADP-ribosyl residue was incorporated into 1 mol of MBP. Four types of purified MBPs were also ADP-ribosylated by cholera toxin dependent on GTP and the protein factor for the ADP-ribosylation. The results show evidence that MBP is one of major and specific substrates of cholera toxin in brain membranes.

Asymmetric binding of cytochrome b5 to the membrane of human erythrocyte ghosts.

The intact, amphipatic form of cytochrome b5 could bind to unsealed ghosts, but not to resealed ghosts, suggesting that the cytochrome could bind only to the inner (cytoplasmic) surface of the ghost membrane. This was further confirmed by the finding that the cytochrome could bind to closed, inside-out vesicles prepared from the ghosts. This asymmetric binding was not due to the exclusive localization of sialic acid and sugar chains on the outer surface of the ghosts membrane, because the cytochrome could not bind to ghosts even after enzymatic removal of these components. Although liposomes consisting of phosphatidylcholine or both phosphatidylcholine and sphingomyelin could effectively bind the cytochrome, this binding capacity was progressively decreased as increasing amount of cholesterol was included in the composition of phosphatidylcholine liposomes. Removal of cholesterol from resealed ghosts by incubation with egg phosphatidylcholine liposomes resulted in the binding of cytochrome b5 to the outer surface of the treated ghosts. The possibility is discussed that the asymmetric binding is due to preferential localization of cholesterol in the outer leaflet of the lipid bilayer that constitutes the ghost membrane.

Induction of a microsomal butyrylesterase in rat liver by phenobarbital treatment.

A carboxylesterase (carboxylic-ester hydrolase, EC 3.1.1.1) was induced in the liver of rats after repeated administration of phenobarbital. This enzyme migrated most rapidly towards the anode among Triton X-100-solubilized liver esterases by electrophoresis on the cellulose acetate membrane and it was tentatively designated as L-I. L-I increased in the microsomal fraction and was mainly concentrated in the fraction of smooth endoplasmic reticulum that proliferated after phenobarbital treatment. L-I- was separated from other liver esterases and purified about 800-fold. The purified L-I was homogeneous as judged by polyacrylamide gel electrophoresis and exhibited a molecular weight of about 55 000 as determined by gel filtration. L-I split the 1-naphthyl ester of butyric acid faster than esters of acetic, propionic or valeric acids; it therefore seemed to be a butyrylesterase.