Journal of Histochemistry and Cytochemistry, Vol. 48, 613-622, May 2000, Copyright © 2000, The Histochemical Society, Inc.


ARTICLE

Fatty Acid Synthase Is Expressed Mainly in Adult Hormone-sensitive Cells or Cells with High Lipid Metabolism and in Proliferating Fetal Cells

Takashi Kusakabea, Masahiro Maedab, Nobuo Hoshia, Takashi Suginoa, Kazuo Watanabea, Takeaki Fukudaa, and Toshimitsu Suzukia
a Department of Pathology, Fukushima Medical University School of Medicine, Fukushima, Japan
b Immunobiological Laboratories, Fujioka, Gunma, Japan

Correspondence to: Toshimitsu Suzuki, Dept. of Pathology, Fukushima Medical U. School of Medicine, 1-Hikariga-oka, Fukushima, 960-1295, Japan. E-mail: tmt@fmu.ac.jp


  Summary
Top
Summary
Introduction
Materials and Methods
Results
Discussion
Literature Cited

Animal fatty acid synthase (FAS) is a homodimer protein which synthesizes long-chain fatty acids and is rich in liver, brain, breast, and lung. However, the precise cellular localization of FAS in human tissues has not been elucidated. Immunohistochemistry with a new antibody to human FAS revealed that in adult human tissues FAS is distributed mainly in cells with high lipid metabolism (adipocytes, corpus luteum, hepatocytes, sebaceous glands, and Type II alveolar cells), in hormone-sensitive cells (anterior pituitary, apocrine gland, breast, endometrium, prostate, seminal vesicle, and adrenal cortex), and in a subset of epithelial cells of duodenum and stomach, colon absorptive cells, cerebral neurons, basket cells of cerebellum, decidua, uroepithelium, and epidymis. In fetal cells at 20 weeks of gestation, FAS was mainly present in proliferative epithelial cells of the digestive and respiratory systems, proximal renal tubules, adrenocortical cells, and mesenchymal and hematolymphoid cells. Staining was significant in nonproliferating cells, as observed in adult, and in sympathetic ganglion cells, Leidig cells of testis, and Langhans cells of chorionic villi. FAS is maintained in hormone-sensitive cells and/or cells active in lipid metabolism in the adult and is expressed in proliferating cells in the fetus, suggesting active fatty acid synthesis for energy utilization or membrane lipids. (J Histochem Cytochem 48:613–622, 2000)

Key Words: fatty acid synthase, cellular localization, fetal tissues, adult tissues


  Introduction
Top
Summary
Introduction
Materials and Methods
Results
Discussion
Literature Cited

ANIMAL FATTY ACID SYNTHASE (FAS: EC 2.3.1.85) ( Poncari 1981 ) is a multifunctional enzyme that catalyzes the synthesis of long-chain fatty acids by using acetyl-CoA as a primer, malonyl-CoA as a two-carbon donor, and NADPH as a reductant ( Bressler and Wakil 1961 ; Wakil 1989 ). FAS synthesizes mainly palmitate, as shown in a human breast cancer cell line in which FAS synthesized fatty acids composed of 80% palmitate, 10% myristate, and 10% stearate (Kuhajda et al. 1994 ). The enzyme is a homodimer whose molecular weight has been reported to be 480 kD ( Thompson et al. 1981 ) or 550 kD ( Kuhajda et al. 1994 ) and the synthase subunit to be 260 kD ( Kuhajda et al. 1994 ) or 270 kD ( Amy et al. 1989 ; Jayakumar et al. 1995 ). Most information about FAS is derived from biochemical studies, and the FAS level has been described to be low in normal human tissues due to its downregulation by dietary lipids ( McCarthy and Hardie 1984 ; Weiss et al. 1986 ). It has also been reported that FAS expression is controlled by hormones (insulin, glucagon, glucocorticoids, and thyroid hormone) and nutrients (glucose and fatty acids) (Sul and Wang 1998 ). Antibodies, such as OA-519, against haptoglobin-related protein epitopes that recognize FAS (Kuhajda et al. 1994 ) have been used mainly in tumor biology research (Kuhajda et al. 1989a , Kuhajda et al. 1989b ; Shurbaji et al. 1991 , Shurbaji et al. 1992 ; Redston et al. 1992 ; Jensen et al. 1993 , Jensen et al. 1995 ; Epstein et al. 1995 ; Hardman et al. 1995 ; Pizer et al. 1995 ; Alo et al. 1996 ; Gansler et al. 1997 ; Rashid et al. 1997 ), which indicates that high expression of FAS in breast, ovarian, and prostate cancer is associated with a poor prognosis (Kuhajda et al. 1989b ; Shurbaji et al. 1991 , Shurbaji et al. 1992 ; Epstein et al. 1995 ; Hardman et al. 1995 ; Jensen et al. 1995 ; Alo et al. 1996 ). However, this is not the case in colorectal carcinoma (Rashid et al. 1997 ). FAS has also been detected in endometrial glands and stromal cells from the proliferative to the early secretory phase, and their FAS expression is closely associated with cell proliferation and expression of progesterone or estrogen receptors ( Escot et al. 1990 ; Pizer et al. 1997 ). After cessation of cell proliferation in the mid- to the late secretory phase, the endometrial tissues become FAS-negative. However, the precise cellular localization and/or distribution of FAS has not been elucidated in normal human tissues, including human fetal tissues that exhibit active cell proliferation. Because human FAS cDNA has been cloned and its deduced amino acid sequence determined (Jayakumar et al. 1995 ; Semenkovich et al. 1995 ), we recently developed a new FAS antibody using a FAS peptide.

Here we describe the cellular distribution of FAS in normal human tissues in fetuses, newborns, infants, and adults.


  Materials and Methods
Top
Summary
Introduction
Materials and Methods
Results
Discussion
Literature Cited

Tissues
Normal human tissues were sampled at autopsy from three newborns, one infant, and three adults. For the central nervous system, seven adult brains (four females and three males, 29–69 years old) were employed. All cases were free of hormone (insulin and/or corticosteroid) or transfusion treatment. Fetal tissues, placenta, and decidua from 6, 20, 25, and 36 weeks of gestation were obtained from spontaneous abortions or stillbirths. In addition, curetted endometrium, surgically removed gallbladder for treatment of cholecystolithiasis, and extirpated breast for treatment of mammary carcinoma were also employed. Tissues were fixed with 10% neutral formalin and paraffin-embedded. Fetal brains were not included because they had not been processed for routine histological examination.

Antibodies
A polyclonal antibody was produced by immunizing New Zealand White rabbits by SC injections of a synthetic peptide composed of 17 amino acids corresponding to residues 1283–1299 of human brain FAS, which was coupled with keyhole limpet hemocyanin (Difco; Detroit, MI). The 17 amino-acid peptide is the sequence of the 3' terminus of the predicted human brain FAS gene product ( Jayakumar et al. 1995 ), which does not contain the acyl carrier protein or the thioesterase active site, which is also in the 3' terminus (Semenkovich et al. 1995 ). Antisera were fractionated through protein A Avide Gel (Bio Probe International; Tustin, CA) to obtain IgG and were specifically purified by antigen-bound activated thiol Avide Gel F affinity column chromatography (Bio Probe International). Polyclonal antibodies against anterior pituitary hormones (adrenocorticotropin, growth hormone, thyroid-stimulating hormone, luteinizing hormone, and prolactin) were purchased from DAKO (Kyoto, Japan) and polyclonal anti-apoprotein antibody from IBL (Fujioka; Gunma, Japan). Mouse monoclonal antibody against Ki-67 antigen (MIB-1) was from Immunotech SA (Marseille, France).

Western Blotting
Human gastric carcinoma cell line TKGW from IBL was employed because immunostaining of FAS by our antibody was positive in this cell line. A gastric cancer cell line, MKN-1, maintained in our laboratory and negative for FAS immunostaining, was also used. Both were homogenized in lysis buffer containing CHAPS and protease inhibitors (pefabloc, leupeptin, pepstatin, and aprotinin) and centrifuged at 15,000 x g for 30 min. The supernatant was mixed with sample buffer (18.5 ml (1 mol)/liter Tris-HCI, pH 7.8, 15 ml 2-mercaptoethanol, 30 ml glycerol, and distilled water, to 100 ml) in a ratio of 2:1. Protein concentration was determined by the Lowry method, and 10 µg protein was subjected to SDS-PAGE and then blotted onto a nitrocellulose membrane. The membrane was then incubated with 2 µg/ml anti-FAS antibody IgG or FAS antibody absorbed by 5 µg antigen, followed by anti-rabbit goat IgG coupled to horseradish peroxidase (IBL). The peroxidase reaction product was visualized with Konica Immunostain HRP1S-50B (Konica; Tokyo, Japan).

Immunohistochemistry
Immunohistochemical procedures were described previously ( Hiraki et al. 1997 ). In short, 3-µm-thick sections were treated with methanol–hydrogen peroxide for 20 min to block endogenous peroxidase activity and then with 5% skimmed milk (Yukijirushi; Sapporo, Japan) in PBS for 30 min to avoid nonspecific adsorption of IgG. The concentration of the primary FAS antibody was 2 µg IgG/ml. The antibodies against anterior pituitary hormones were prediluted and the MIB-1 was diluted to 1:100. For MIB-1, the sections were microwaved for 15 min to retrieve the antigen. All slides were incubated with the antibodies overnight at 4C, after which they were reacted with biotin-labeled anti-rabbit IgG (Nichirei kit; Tokyo, Japan) for the polyclonal antibodies or anti-mouse Igs (Nichirei kit) for the mouse monoclonal antibody, and then with streptavidin-conjugated peroxidase (Nichirei kit) for 20 min at room temperature. Peroxidase reaction product was visualized with the Graham–Karnovsky solution ( Graham and Karnovsky 1966 ) containing 65 mg of NaN3 /dl to block endogenous peroxidase activity. Nuclei were counterstained with 1% methyl green.

In breast cancer tissues known to be FAS-positive ( Thompson et al. 1981 ; Kuhajda et al. 1989b ; Chalbos et al. 1990 ; Joyeaux et al. 1990 ; Shurbaji et al. 1991 ; Jensen et al. 1995 ; Alo et al. 1996 ; Milgraum et al. 1997 ), the specificity of FAS immunostaining was confirmed by omission of the primary antibody and use of preimmune rabbit IgG (2 µg/ml) or primary antibody absorbed with 5 µg antigen for the first step of the immunostaining procedure.


  Results
Top
Summary
Introduction
Materials and Methods
Results
Discussion
Literature Cited

Specificity of the FAS Antibody and Validity of Immunostaining
The antibody purified by affinity chromatography reacted with a 270-kD protein in the extracts of liver and TKGW cells but did not react with the extract of MKN-1 by Western blotting, as shown in Fig 1A. The 270-kD protein has the same molecular weight as the monomer of human brain FAS (Jayakumar et al. 1995 ), which is believed to be formed through disruption of disulfide bonds in the dimer (Stoops and Wakil 1982 ) by 2-mercaptoethanol in the sample buffer. Absorbed antibody or normal rabbit IgG did not reveal any bands in the TKGW cell extract by Western blotting (Fig 1A).



View larger version (136K):
[in this window]
[in a new window]
 
Figure 1. Western blotting (A) reveals a major single band of about 270 kD in the extract of TKGW gastric carcinoma cell line detected by anti-FAS peptide antibody. Antibody absorbed with the homologous antigen or normal rabbit IgG fails to react with the extract of TKGW, and the antibody is nonreactive with the extract of FAS-negative MKN1 cells. Intense labeling of breast carcinoma cells by the FAS antibody (B) and complete abrogation of the labeling in the cancer cells by immunostaining with the absorbed FAS antibody (C) are noted. Bar = 20 µm.

As shown below, the conditions of immunostaining with the FAS antibody were established by immunohistochemistry with human breast cancer tissue, which is known to be FAS-positive (Thompson et al. 1981 ; Kuhajda et al. 1989b ; Chalbos et al. 1990 ; Joyeaux et al. 1990 ; Shurbaji et al. 1991 ; Jensen et al. 1995 ; Alo et al. 1996 ; Milgraum et al. 1997 ). The antibody clearly labeled breast carcinoma cells (Fig 1B), but the normal rabbit IgG (not shown) and antibody absorbed with the homologous antigen were negative (Fig 1C). FAS was distributed homogeneously in the cytoplasm and had a granular appearance in the intensely stained cells.

FAS-positive Cells in Adults
The results with human adult tissues are summarized in Table 1. Strongly FAS-positive cells were mainly nonproliferating, and limited amounts of FAS were detected in areas of active cell turnover, which consisted of cells in the proliferative zone (Ki-67-labeled) of gastroduodenal mucosa, hematopoietic cells in bone marrow, proliferating gland cells, and stromal cells of endometrium. Nonproliferating FAS-positive cells in the CNS were cerebral cortical neurons of the fronto-occipital lobes, mainly in the third to fifth layers (Fig 2A), some neurons in the basal ganglia, hippocampus, paraventriculus, pons and medulla oblongata, basket cells of the cerebellum (Fig 2B), and a few astrocytes. Axons of basket cells that terminate on or attach to Purkinje cell bodies were also positive. Hippocampal basket cells were not labeled. There were no gender differences in the frequency of FAS-positive neurons in cerebral cortices. Cerebellar cortical basket cells showed similar positive staining in the seven brains examined, whereas cerebral cortical neurons exhibited mild fluctuation in the frequency of FAS staining from brain to brain. FAS-positive cells were also found in the digestive system [absorptive cells of the large intestine (Fig 2C ) and the appendix vermiformis and hepatocytes in Zone 1 to Zone 2 (Fig 2D)], some Type II alveolar cells of lung (Fig 3A) also positive for surfactant apoprotein, in the urogenital system [a subset of epithelial cells of the epididymis and seminal vesicle, main prostate glandular cells ( Fig 3B), mucosal and submucosal glands of the prostate, decidualized stromal cells of the endometrium, lutein cells of the ovary, and umbrella cells of the bladder], anterior pituitary cells (Fig 3C) (partly identical with GH and/or ACTH cells), subcutaneous and intra-abdominal fat cells, apocrine gland and sebaceous gland of skin (Fig 3D), tissue mast cells, adrenal zona fasciculata cells, and epithelial cells of ducts and acini of the breast.



View larger version (157K):
[in this window]
[in a new window]
 
Figure 2. FAS-labeled cells in adult tissues. Cortical neurons mainly in the third to fifth layer show FAS-positivity (frontal lobe). Polygonal larger neurons and a few neuron processes are labeled (A). Positive cells in the molecular layer of cerebellum are largely basket cells, and many axons, even around the Purkinje cells, are also FAS-immunoreactive (B ). Absorptive cells of colon mucosa (C) and hepatocytes (D ) are positive. Bars = 20 µm.



View larger version (143K):
[in this window]
[in a new window]
 
Figure 3. Some Type II alveolar cells of lung (A), main prostate glands ( B), a subset of anterior pituitary cells (C), and sebaceous gland cells of skin (D) are FAS-positive. Bars = 50 µm.


 
View this table:
[in this window]
[in a new window]
 
Table 1. FAS-positive cells in adult tissues

FAS-positive Cells in Fetuses
In the 6-week-old fetus, hepatocytes and hemopoietic cells, including erythroblasts, were positive. Because each visceral organ was easily identified, two 20-week-old fetuses were examined precisely except for their brains. The results are summarized in Table 2. The FAS-positive cells were predominantly those labeled with Ki-67 in areas of cell proliferation, although the Ki-67-labeled cells exceeded the number of FAS-positive cells. Positive cells in the digestive system were basal layer cells of squamous epithelium of tongue and esophagus, nonmucinous salivary gland cells (Fig 4A and Fig 4B), glandular cells of gastrointestinal mucosa, pancreatic acinar cells, and hepatocytes. In the cardiopulmonary system, positive cells were found in the endocardium and tracheobronchial glands, bronchopulmonary epithelium, chondrocytes of tracheobronchial cartilage, and vascular endothelial cells (Fig 4C and Fig 4D). Positive cells of the urogenital system were found in the S-shaped proximal tubules of kidney ( Fig 4E and Fig 4F ) and in the prostate gland. In the endocrine system, anterior pituitary cells, thyroid follicular cells, and subcapsular adrenocortical cells were positive (Fig 4G and Fig 4H). In addition, thymocytes and nodal lymphocytes in the periphery of the tissue were weakly positive. Schwann cells and mesenchymal cells of peripheral nerves in sympathetic ganglia and the Auerbach plexus of the gastrointestinal tract, a few neuroblasts in the adrenal medulla, fibroblasts (especially in the perivertebral region), striated muscle cells of tongue, and hematopoietic cells, including erythroblasts and megakaryocytes, showed cytoplasmic FAS, but the frequency of positive cells was low. However, cardiomyocytes and smooth muscle cells of visceral organs were hardly labeled despite being positive with Ki-67. Some Ki-67-negative cells were FAS-positive in the fetus, but this was limited and consisted of adipocytes, ganglion cells of sympathetic ganglia (proliferating Schwann cells were also positive) and the Auerbach plexus, Leidig cells of testis (proliferating seminiferous tubules were negative) (Fig 5A and Fig 5B), sebaceous gland, urinary tract uroepithelial cells (Ki-67-labeled cells were negative) (Fig 5C and Fig 5D), and the upper layer of adrenocortical cells (Fig 4G ). Tissue mast cells and osteoblasts were also positive. Placenta from the sixth week of gestation or later showed FAS-positive cytotrophoblasts. Syncytiotrophoblasts and other components of the placenta were negative. Tissues of later gestational ages (25 and 36 weeks), newborns at 1 month, infants at 6 months, 10 months, and infants at 3 years had much more limited distributions of FAS-positive cells, almost similar to those of adult tissues. For example, a 1-month-old infant was FAS-negative in proximal tubules of kidney, subcapsular adrenocortical cells, Schwann cells of sympathetic ganglion, and mesenchymal cells (fibroblasts, chondrocytes, endothelial cells), which had ceased to proliferate.



View larger version (137K):
[in this window]
[in a new window]
 
Figure 4. FAS-positive cells (A,C,E,G) and alternative Ki-67 labeling ( B,D,F, H) in a 20-week fetus: salivary gland (A,B), endothelial cells of a blood vessel (C,D, arrows), kidney (E,F ), and adrenal (G,H). In the adrenal there is intense positivity in the subcapsular cortical cells heavily labeled by Ki-67 and also in the upper layer of cortical cells. Bars = 50 µm.



View larger version (129K):
[in this window]
[in a new window]
 
Figure 5. FAS (A,C) and Ki-67 immunostaining (B,D) in a 20-week fetus. Generally, FAS-positive cells are not labeled by Ki-67: Leidig cells of testis in contrast to Ki-67-labeling in the seminiferous tubules (A,B) and bladder uroepithelium are Ki-67-negative (C,D). Bars = 50 µm.


 
View this table:
[in this window]
[in a new window]
 
Table 2. FAS-positive cells in fetus at 20 weeks of gestation


  Discussion
Top
Summary
Introduction
Materials and Methods
Results
Discussion
Literature Cited

Long-chain fatty acids, such as palmitate, are essential constituents of membrane lipids and are also important substrates for cellular energy metabolism. FAS synthesizes mainly palmitate, as stated above. Expression of FAS mRNA in nonfetal human tissues was high in brain, liver, lung, intro-abdominal adipose tissue (Shurbaji et al. 1992 ; Jayakumar et al. 1995 ), and in duct cells and acinar cells of breast (Chalbos et al. 1990 ). However, the precise expression of FAS protein at the cellular level has not been clarified in normal human tissues, because the FAS antibody used thus far failed to label fat cells in cryostat sections (Joyeaux et al. 1990 ) and the antibody OA-519 also failed to react with normal or hyperplastic prostate (Shurbaji et al. 1992 ) and normal breast ( Jensen et al. 1995 ). As shown in this study, our new antibody raised against human brain FAS peptide recognizes FAS protein in human cells and can be used with formalin-fixed, paraffin-embedded tissues. The reason(s) for difference between the antibody described and ours is not clear, but the antibody reported may have a low affinity for the antigen(s) or may recognize a different antigen epitope(s) in the FAS molecule. With immunohistochemistry, we showed that FAS is present in various cells of tissues from the human fetus, newborn/infant, and adult. There are two types of FAS expression, one of which is closely associated with cell proliferation and constitutive expression in resting cells. In this study, the proliferative activity was evaluated with anti-Ki-67. The suitability of monoclonal anti-Ki-67 antibody for detecting proliferative activity was established previously because Ki-67 is known to be expressed in nuclei of cells cycling from G1- to M-phase of the cell cycle (Gerdes et al. 1984a , Gerdes et al. 1984b ). The expression of FAS associated with cell proliferation was clearly observed in human fetal tissues at 20 weeks of gestation. FAS was detected not only in developing epithelial cells but also in growing mesenchymal cells. FAS-containing cells were not always Ki-67-positive, but the frequency of negative cells was consistently lower than that of positive ones. Because the half-life of FAS mRNA is 4 hr ( Xu and Rooney 1997 ) and Ki-67 expression lasts from G1 - to the M-phase of the cell cycle, this finding appears reasonable. However, cardiomyocytes, seminiferous tubules, and esophagogastrointestinal tract smooth muscle cells lacked FAS expression despite their apparent Ki-67 labeling. In the FAS-negative proliferating cells, another type of FAS that does not react with the antibody we used might be present, since there are minor differences in the amino acid sequences of brain FAS and FAS in the hepatocellular carcinoma cell line HepG2 (Jayakumar et al. 1995 ). Alternatively, fatty acid binding protein, which removes fatty acids from the circulation and is rich in fetal and adult heart (Watanabe et al. 1993 ), might function in the heart in place of FAS. Some resting cells of the 20-week fetus were also FAS-positive. These included adipocytes, ganglion cells of peripheral nerves or sympathetic ganglia, Leidig cells of testis, sebaceous gland, uroepithelium, and fetal adrenocortical cells. Cells of this type might utilize FAS for active lipid metabolism and, among these fat cells, sebaceous glands and umbrella cells of urinary bladder remained FAS-positive in adult tissues. Although a few replicating cells in stomach and duodenum, as well as bone marrow cells, were FAS-positive in the adult, positive cells were found mostly among resting cells that have active lipid metabolism and/or hormone sensitivity. Sebaceous gland cells, fat cells, hepatocytes, and Type II alveolar cells (surfactant or dipalmitate lecithin producers) are cells with active lipid metabolism. Apocrine gland (sex hormone-sensitive), prostate, endometrium, mammary glands, seminal vesicle (testosterone-sensitive), and anterior pituitary cells are hormone-sensitive cells. Adrenocortical cells show both characteristics. Several cells that express FAS do not fall into these two categories: ganglion cells of cerebral cortices, glia, basket cells of cerebellum, mast cells, decidua, and cytotrophoblasts. Basket cells of cerebellar cortex are well-known inhibitory neurons, and similar inhibitory neurons, also called basket cells, exist in the hippocampus but these lacked FAS, indicating that FAS expression is not a characteristic of all inhibitory neurons. The physiological role of FAS in basket cells and some neurons of the cortex or sympathetic ganglia might be elucidated by neurophysiological research.

It was reported that decidualized stroma cells in the late secretory phase and decidua of pregnancy were intensely positive for FAS not associated with cell proliferation (Pizer et al. 1997 ). In the present study, these phenomena were confirmed and cytotrophoblasts of chorionic villi also contained FAS, but its biological significance in pregnancy requires investigation. Hormone dependency of FAS expression has been described in human cycling endometrium ( Pizer et al. 1997 ), as stated earlier, and hormonal stimulation induces upregulation of FAS expression in hormone-sensitive cells such as normal breast (Joyeaux et al. 1990 ), mastopathy, breast carcinoma (Chalbos et al. 1990 ), lactating breast ( Smith and Ryan 1979 ), and a prostate cancer cell line ( Swinnen et al. 1997 ). Insulin administration has been reported to rapidly induce FAS mRNA in diabetic mouse liver ( Paulauskis and Sul 1989 ), and dexamethasone causes induction of FAS in cultured fetal rat lung (Pope et al. 1988 ). In these cases, the relationship between the induction of FAS and cell growth is not clear, whereas growth factors such as fibroblast growth factor-1 induce cell growth and upregulate FAS expression in fibroblasts ( Hsu et al. 1993 ). The precise biological significance of FAS expression in normal and cancer tissues is not as clear, but FAS might be expressed and synthesized in coordination with increased demand for fatty acid metabolism and/or membrane synthesis in response to regulated or unregulated cell proliferation, and intense expression of the FAS in surface uroepithelium of both fetus and adult may play a role in excluding water through the hydrophobic nature of fatty acids.

In conclusion, under normal conditions FAS is highly expressed not only in proliferating cells and those with high lipid metabolism but also in hormone-sensitive cells and those with permeability barriers.


  Acknowledgments

Supported in part by a Grant-in-Aid for Scientific Research (no. 10670168) from the Ministry of Education, Science, Sports and Culture of Japan.

We thank H. Hiraki for his outstanding help with the immunohistochemical studies.

Received for publication August 18, 1999; accepted January 5, 2000.


  Literature Cited
Top
Summary
Introduction
Materials and Methods
Results
Discussion
Literature Cited

Alo PL , Visca P , Marci A , Mangoni A , Botti C , Di Tondo U (1996 ) Expression of fatty acid synthase (FAS) as a predictor of recurrence in stage I breast carcinoma patients. Cancer 77:474-482 [Medline]

Amy CM , Witkowski A , Naggert J , Williams B , Randhawa Z , Smith S (1989 ) Molecular cloning and sequencing of cDNA encoding the entire rat fatty acid synthase. Proc Natl Acad Sci USA 86:3114-3118 [Abstract]

Bressler R , Wakil SJ (1961 ) Studies on the mechanism of fatty acid synthesis: IX. The conversion of malonyl coenzyme A to long chain fatty acids. J Biol Chem 236:1643-1651

Chalbos D , Escot C , Joyeux C , Tissot–Carayon M-J , Pages A , Rochefort H (1990 ) Expression of the progestin-induced fatty acid synthetase in benign mastopathies and breast cancer as measured by RNA in situ hybridization . J Natl Cancer Inst 82:602-606 [Abstract]

Epstein JI , Carmichael M , Partin AW (1995 ) OA-519 (fatty acid synthase) as an independent predictor of pathologic stage in adenocarcinoma of the prostate. Urology 45:81-86 [Medline]

Escot C , Joyeux C , Mathieu M , Maudelonde T , Pages A , Rochefort H , Chalbos D (1990 ) Regulation of fatty acid synthetase ribonucleic acid in the human endometrium during the menstrual cycle. J Clin Endocrinol Metab 70:1319-1324 [Abstract]

Gansler TS , Hardman W, III , Hunt DA , Schaffel S , Henninger RA (1997 ) Increased expression of fatty acid synthase (OA-519) in ovarian neoplasms predicts shorter survival. Hum Pathol 28:686-692 [Medline]

Gerdes J , Dallenbach F , Lennert K (1984 ) Growth fraction in malignant non-Hodgkin's lymphoma (NHL) as determined in situ with the monoclonal antibody Ki-67. Hematol Oncol 2:365-371, a [Medline]

Gerdes J , Lemke H , Baisch H , Wacker HH , Schwab U , Stein H (1984 ) Cell cycle analysis of a cell proliferation-associated human nuclear antigen defined by the monoclonal antibody Ki-67. J Immunol 133:1710-1715, b [Abstract/Free Full Text]

Graham RC, Jr , Karnovsky MH (1966 ) The early stage of absorption of injected horseradish peroxidase in proximal tubules of mouse kidney. Ultrastructural cytochemistry by a new technique. J Histochem Cytochem 14:291-302 [Medline]

Hardman W , Gansler T , Schaffel S , Hennigar R (1995 ) OA-519 immunostaining portends poor prognosis in ovarian cancer . Mod Pathol 8:90A

Hiraki H , Hoshi N , Hasegawa H , Tanigawa T , Emura I , Seito T , Yamaki T , Fukuda T , Watanabe K , Suzuki T (1997 ) Regular immunohistochemical localization of endothelin-1 and endothelin-B receptor in normal, hyperplastic and neoplastic human adrenocortical cells. Pathol Int 47:117-125 [Medline]

Hsu DKW , Donohue PJ , Alberts GF , Winkles JA (1993 ) Fibroblast growth factor-1 induces phosphofruktokinase, fatty acid synthase and Ca2+-ATPase mRNA expression in NIH3T3 cells . Biochem Biophys Res Commun 197:1483-1491 [Medline]

Jayakumar A , Tai M-H , Huang W-Y , Al-Feel W , Hsu M , Abu-Elheiga L , Chirala SS , Wakil SJ (1995 ) Human fatty acid synthase: properties and molecular cloning . Proc Natl Acad Sci USA 92:8695-8699 [Abstract]

Jensen V , Holm–Nielsen P , Melsen F (1993 ) OA-519 expression in node negative breast cancer. Breast Cancer Res Treat 27:1-2

Jensen V , Ladekari M , Holm–Nielsen P , Melsen F , Soerensen FB (1995 ) The prognostic value of oncogenic antigen 519 (OA-519) expression and proliferative activity detected by antibody MIB-1 in node negative breast cancer. J Pathol 176:343-352 [Medline]

Joyeaux C , Chalbos D , Rochefort H (1990 ) Effects of progestins and menstrual cycle on fatty acid synthetase and progesterone receptor in human mammary glands. J Clin Endocrin Metab 70:1319-1324 [Abstract]

Kuhajda FP , Jenner K , Wood FW , Henninger RA , Jacobs LB , Dick JD , Pasternack GR (1994 ) Fatty acid synthesis: a potential selective target for anti-neoplastic therapy. Proc Natl Acad Sci USA 91:6379-6383 [Abstract]

Kuhajda FP , Katumuluwa AI , Pasternack GR (1989 ) Expression of haptoglobin-related protein and its potential role as a tumor antigen. Proc Natl Acad Sci USA 86:1188-1192, a [Abstract]

Kuhajda FP , Piantadosis S , Pasternack GR (1989 ) Haptoglobin-related protein (Hpr) epitopes in breast cancer as a predictor of recurrence of the disease. N Engl J Med 321:636-641, b [Abstract]

McCarthy AD , Hardie DG (1984 ) Fatty acid synthase: an example of protein evolution by gene fusion. Trends Biochem Sci 9:60-63

Milgraum LZ , Witters LA , Pasternack GR , Kuhajda FP (1997 ) Enzymes of the fatty acid synthesis pathway are highly expressed in in situ breast carcinoma. Clin Cancer Res 3:2115-2120 [Abstract]

Paulauskis JD , Sul HS (1989 ) Hormonal regulation of mouse fatty acid synthase gene transcription in liver. J Biol Chem 264:574-577 [Abstract/Free Full Text]

Pizer ES , Kurman RJ , Pasternack GR , Kuhajda FP (1997 ) Expression of fatty acid synthase is closely linked to proliferation and stromal decidualization in cycling endometrium. Int J Gynecol 16:45-51

Pizer ES , Wood FD , Pasternack GR , Kuhajda FP (1995 ) Altered fatty acid biosynthesis in endometrial carcinoma. Mod Pathol 8:95A

Poncari DAK (1981 ) Fatty acid synthase from human liver. Methods in Enzyomol 71:73-79

Pope TS , Smart DA , Rooney SA (1988 ) Hormonal effects on fatty-acid synthase in cultured fetal rat lung; induction by dexamethasone and inhibition of activity by triiodothyronine. Biochim Biophys Acta 959:169-177 [Medline]

Rashid A , Pizer ES , Moga M , Milgraum LZ , Zahurak M , Pasternack GR , Kuhajda FP , Hamilton SR (1997 ) Elevated expression of fatty acid synthase and fatty acid synthetic activity in colorectal neoplasia. Am J Pathol 150:201-208 [Abstract]

Redston MS , Kern SE , Vogelstein B , Hamilton SR (1992 ) Expression of OA-519 (haptoglobin-related protein epitopes) in colorectal carcinomas: comparison with molecular genetic alterations and metastasis. Mod Pathol 5:47A

Semenkovich CF , Coleman T , Fiedorek FT, Jr (1995 ) Human fatty acid synthase mRNA: tissue distribution, genetic mapping, and kinetics of decay after glucose deprivation. J Lipid Res 36:1507-1521 [Abstract]

Shurbaji MS , Kuhajda FP , Pasternack GR , Thurmond TS (1992 ) Expression of oncogenic antigen 519 (OA-519) in prostate cancer is a potential progressive indicator. Am J Clin Pathol 97:686-691 [Medline]

Shurbaji MS , Pasternack GR , Kuhajda FP (1991 ) Expression of haptoglobin-related protein in primary and metastatic breast cancers. A longitudinal study of 48 fatal tumors. Am J Clin Pathol 96:238-242 [Medline]

Smith S , Ryan P (1979 ) Asynchronous appearance of two enzymes concerned with medium chain fatty acid synthesis in developing rat mammary gland. J Biol Chem 254:8932-8936 [Medline]

Stoops JK , Wakil SJ (1982 ) The reaction of chicken liver fatty acid synthase with 5, 5'-dithiobis (2-nitrobenzoic acid). Biochem Biophys Res Commun 104:1018-1024 [Medline]

Sul HS , Wang D (1998 ) Nutritional and hormonal regulation of enzymes in fat synthesis : studies of fatty acid synthase and mitochondrial glycerol-3 phosphate acyltransferase gene transcription. Annu Rev Nutr 18:331-351 [Medline]

Swinnen JV , Esquenet M , Goossens K , Heyns W , Verhoeven G (1997 ) Androgens stimulate fatty acid synthase in the human prostate cancer cell line LNCaP. Cancer Res 57:1086-1090 [Abstract]

Thompson BJ , Stern A , Smith A (1981 ) Purification and properties of fatty acid synthetase from a human breast cell line. Biochim Biophys Acta 662:125-130 [Medline]

Wakil SJ (1989 ) Fatty acid synthase, a proficient multifunctional enzyme. Biochemistry 28:4523-4530 [Medline]

Watanabe K , Suzuki T , Wakabayashi H , Ono T , Veerkamp E , Kolb JH (1993 ) Immunohistochemical distribution of heart type fatty acid-binding protein immunoreactivity in normal human tissue and in acute myocardial infarct. J Pathol 170:59-65 [Medline]

Weiss L , Hoffmann GE , Schreiber R , Andres H , Fuchs E , Körber E , Kolb HJ (1986 ) Fatty-acid biosynthesis in man, a pathway of minor importance. Purification, optimal assay conditions, and organ distribution of fatty-acid synthase. Biol Chem Hoppe Seyler 367:905-912 [Medline]

Xu Z-X , Rooney SA (1997 ) Glucocorticoids increase fatty-acid synthase mRNA stability in fetal rat lung. Am J Physiol 272:L860-864[Abstract/Free Full Text]