Epidermal Fatty Acid-binding Protein Expression Research Articles

Concomitant decrease of E- and A-FABP expression predicts worse survival in urothelial bladder cancer patients

Non-muscle invasive bladder cancers (NMIBC) pTa-pT1 are depicted by a high risk of recurrence and/or progression with an unpredictable clinical evolution. Our aim was to identify, from the original resection specimen, tumors that will progress to better manage patients. We previously showed that A-FABP (Adipocyte- Fatty Acid Binding Protein) loss predicted NMIBC progression. Here we determined by immunohistochemistry the prognostic value of E-FABP (Epidermal-Fatty Acid Binding Protein) expression in 210 tumors (80 pTa, 75 pT1, 55 pT2-T4). Thus, E-FABP low expression was correlated with a high grade/stage, the presence of metastatic lymph nodes, and visceral metastases (p < 0.001). Unlike A-FABP in NMIBC, E-FABP low expression was not associated with RFS or PFS in Kaplan–Meier analysis. But patients of the overall cohort with a high E-FABP expression had a longer mOS (53.8 months vs. 29.3 months, p = 0.029). The immunohistochemical analysis on the same NMIBC tissue sections revealed that when A-FABP is absent, a high E-FABP expression is detected. E-FABP could compensate A-FABP loss. Interestingly, patients, whose original tumor presents both low E-FABP and negative A-FABP, had the worse survival, those maintaining the expression of both markers had better survival. To conclude, the combined evaluation of A- and E-FABP expression allowed to stratify patients with urothelial carcinoma for optimizing treatment and follow-up.

Salivary and Lacrimal Gland Alterations of the Epidermal Fatty Acid-Binding Protein (E-FABP) in Non-Obese Diabetic Mice.

The purpose of this study was to investigate the changes in E-FABP in the salivary and lacrimal glands of the Sjögren syndrome (SS) model non-obese diabetic mice (NOD). Cotton thread and ocular vital staining tests were performed on 10-week NOD male mice (n = 24) and age- and sex-matched wild-type (WT) mice (n = 25). Tear and saliva samples were collected at sacrifice for E-FABP ELISA assays. Salivary and lacrimal gland specimens underwent immunohistochemistry stainings for E-FABP. Real-time RT-PCR was also performed for the quantification of mRNA expression levels in the salivary and lacrimal glands. Corneal vital staining scores in the NOD mice were significantly higher compared with those for the wild-type mice (p = 0.0001). The mean tear E-FABP level showed a significantly lower concentration in the NOD mice (p = 0.001). The mean saliva E-FABP level also showed a significantly lower concentration in the NOD mice (p = 0.04). Immunohistochemistry revealed intense E-FABP staining in the LG acinar epithelium and less intense staining in the acinar epitheliae of the SGs in the NOD mice compared to the WT mice. Real-time RT-PCR for the mRNA expression of E-FABP showed a significantly decreased expression in the SG and a significant increase in the LG of the NOD mice compared to the WT mice. In conclusion, the E-FABP showed marked alterations in the tear film, saliva, lacrimal, and salivary glands of the NOD mouse, which may help explain the ocular surface changes in relation to the dry eye disease in this SS model mouse and keratoconjunctivitis sicca in SS patients.

Abstract PS8-27: Consumption of a high-fat diet rich in linoleic acid promotes mammary tumor growth

Abstract Consumption of high-fat diets rich in linoleic acid promotes mammary tumor growth Rong Jin1, Jiaqing Hao1, Edward Sauter2, Bing Li1 1Department of Microbiology and Immunology, University of Louisville, Louisville, KY2Hartford Healthcare Cancer Institute, Hartford, CT *To whom correspondence should be addressed: Bing Li, Department of Microbiology and Immunology, University of Louisville, 505 South Hancock Street, Louisville, KY, 40202. Telephone: 502-852-2678. Email: b.li@louisville.edu. AbstractBackground: The most recent Dietary Guidelines in 2015-2020 for Americans from the U.S. Department of Health and Human Services (DHH) and U.S. Department of Agriculture (USDA) recommends a healthy eating pattern with more oils but less saturated fats. Oils predominantly consist of unsaturated fats with either monounsaturated or polyunsaturated fatty acids (FAs). As high fat diets (HFDs) contribute to obesity, which increases the risk of breast cancer, in this report we evaluate whether the consumption of HFDs rich in different unsaturated fatty acids induces similar degrees of obesity and further determines the impact of different oils on mammary tumor growth using our unique murine mouse models. Hypothesis: Consumption of different oils impacts obesity-associated mammary tumor development. Methods: Weaned C57BL/6 wild type (WT) mice and epidermal fatty acid binding protein (E-FABP)-deficient (genetically depleted E-FABP) female mice were randomly grouped and fed either a safflower oil HFD (45% fat, rich in 18:2 polyunsaturated FAs) or an olive oil HFD (45% fat, rich in 18:1 monounsaturated FAs), or a control low fat diet (LFD). After 5 months on the diets, E0771 mammary tumor cells (0.5 × 106/mouse) were injected into the mammary fat pad and tumor volume was measured at 3-day intervals for tumor growth. Immune cell phenotype and functions were evaluated before and after tumor injection. The student’s t-test and/or ANOVA were used to compare immune cell function, obesity and tumor sizes in WT and E-FABP-/- mice, respectively. Results: Compared to the LFD diet, consumption of either safflower oil HFD or olive oil HFD was able to induce a similar degree of mouse obesity in WT mice. E-FABP deficiency has no obvious impact on HFD-induced mouse weight increase. Interestingly, while safflower oil-induced obesity significantly increased mammary tumor growth, olive oil-induced obesity did not promote mammary tumor growth when compared to LFD-fed lean mice. Immunophenotypic and functional analyses showed that mice on the safflower oil diet exhibited lower numbers of CD8+ T cells with reduced production of TNFa than mice on the olive oil diet or LFD. Moreover, safflower oil-associated tumor growth was compromised in mice genetically lacking of E-FABP expression. Mechanistically, E-FABP expression in T cells, especially in CD8+ T cells, mediated 18:2 linoleic acid-induced ROS production and T cell death, thus leading to impaired anti-tumor immunity in safflower oil-fed obese mice. Impact: Although oils are healthy dietary components as recommended by DHH and USDA, different oils exhibit diverse immunoregulatory effects in vivo. Overconsumption of oils with 18:2 FAs may cause obesity-associated breast cancer development. Citation Format: Rong Jin, Jiaqing Jiaqing, Di Yin, Yanmei Yi, Hua Yuan, Satuer Edward, Bing Li. Consumption of a high-fat diet rich in linoleic acid promotes mammary tumor growth [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS8-27.

Mycosis fungoides and Sézary syndrome tumor cells express epidermal fatty acid-binding protein, whose expression decreases with loss of epidermotropism.

Fatty acid binding protein (FABP) is a family of transport proteins for fatty acid (FA). Epidermal FABP (E-FABP) is highly expressed by resident memory T cells (TRM ) in the skin. It supports the uptake of exogenous FA for long-term survival of skin TRM . Mycosis fungoides (MF) is regarded as malignancy of skin TRM . In this study, we investigated E-FABP expression in psoriasis vulgaris (PV), atopic dermatitis (AD), MF, and Sézary syndrome (SS). E-FABP mRNA levels in PV were much higher than those in healthy controls. E-FABP mRNA levels in AD and MF/SS lesional skin were also significantly higher than those of normal skin. By immunohistochemical staining, E-FABP was positive in MF/SS lesional skin. Interestingly, E-FABP was stained positive in epidermotropic lymphoid cells in patch, plaque, and erythrodermic lesions of MF/SS, suggesting that a part of tumor cells expressed E-FABP. In tumorous lesions, however, most dermal tumor cells were negative for E-FABP. Immunohistochemical staining using patch/plaque lesions and tumorous lesions from the same patients also revealed that E-FABP expression decreased in tumorous lesions. Our study has suggested that MF/SS tumor cells express E-FABP, whose expression decreases with loss of epidermotropism.

Effect of methotrexate treatment on the expression of epidermal-fatty acid-binding protein (E-FABP) and apolipoproteins in patients with psoriasis.

IntroductionEpidermal-fatty acid-binding protein (E-FABP) is a marker of transiently amplifying cells which are formed from stem cells in epidermis. Their role is an uptake of fatty acids and metabolism. Psoriatic keratinocytes overexpress E-FABPs, which leads to acanthosis and may explain the lipid’s disturbances in psoriasis.AimAssessment of FABP and apolipoprotein expression in patients treated with methotrexate (MTX).Material and methodsFABP expression in the lesional and perilesional psoriatic skin from 11 male patients compared to 5 healthy skin samples were evaluated by immunohistochemistry. FABP, apolipoprotein A1 (ApoA1) and B (ApoB) serum levels were assessed by ELISA. These parameters were evaluated before and after treatment with subcutaneous MTX (15 mg/wk for 12 weeks).ResultsExpression of E-FABP was lower in the control group than in the lesional and perilesional psoriatic skin, before and after treatment. After treatment the expression decreased in the lesional and perilesional skin. Serum E-FABP was higher in the control group (482.855 ±240.550 pg/ml) compared to patients, but not statistically significantly. After MTX treatment, a statistically significant reduction was observed in psoriatic patients. ApoA1 levels did not differ in the control and patients groups, both before and after treatment. In contrast, ApoB levels did not differ statistically between the control group (1447.126 ±311.11 ng/ml) and patients before treatment, while they were the lowest after treatment (1081.67 ±117.83 ng/ml vs. 808.306 ±103.72 ng/ml; p < 0.01).ConclusionsOur study confirms the beneficial effect of MTX, not only as an anti-proliferative effect, but also reducing the cardiovascular risk by decreasing atherogenic ApoB.

Altered epidermal fatty acid-binding protein expression in hepatocellular carcinoma predicts unfavorable outcomes.

ObjectiveHepatocellular carcinoma (HCC) is a rapidly proliferating malignancy that requires large amounts of fatty acids to synthesize cellular membranes and provide energy. Epidermal fatty acid-binding protein (EFABP) is uniquely expressed in epidermal cells, but its role and expression in HCC are not clear.Subjects and methodsA total of 804 HCC specimens were collected to construct a tissue microarray (TMA) and for immunohistochemistry (IHC) analysis. The relationship between EFABP expression and clinical features of patients with HCC was analyzed.ResultsThe EFABP IHC score for HCC tissue was 0.76±0.69, being significantly higher than that for matched nontumorous tissue (0.48±0.55; P<0.001). Using the median IHC score (ie, 0.8) in the tumorous tissue, a high level of EFABP expression was found in 57.3% (461/804) of the cases. Patients with HCC displaying high EFABP expression had poorer tumor differentiation (P=0.029), more vascular invasion (P=0.006), and a higher proportion of late TNM stage disease (P=0.042). Kaplan–Meier analysis revealed that the patients with high EFABP expression had significantly worse outcomes in terms of overall survival (P=0.003), worse disease-free survival (P=0.021), and a higher probability of recurrence (P=0.014). Multivariate analysis indicated that EFABP expression was an independent prognostic variable for overall survival (P=0.021) and disease-free survival (P=0.044). For HCC recurrence, only vascular invasion (P=0.020) and EFABP expression (P=0.026) were independent risk factors.ConclusionOur data revealed that EFABP expression was increased in HCC samples. High EFABP expression was correlated with shorter survival times in patients with HCC and served as an independent factor for worse outcomes. Our study therefore provides a promising bio-marker for the prognostic prediction of HCC and a potential therapeutic target for the disease.

Inhibition of tumor growth by a newly-identified activator for epidermal fatty acid binding protein.

Our previous studies have demonstrated that expression of epidermal fatty acid binding protein (E-FABP) in tumor associated macrophages (TAMs) promotes macrophage anti-tumor activity by enhancing IFNβ responses in tumor models. Thus, E-FABP represents a new protective factor in enhancing tumor immune surveillance against tumor development. Herein, we report the compound 5-(benzylamino)-2-(3-methylphenyl)-1,3-oxazole-4-carbonitrile (designated EI-05) as a novel E-FABP activator for inhibition of mammary tumor growth. EI-05 was selected from the ZINC compound library using molecular docking analysis based on the crystal structure of E-FABP. Although EI-05 is unable to bind E-FABP directly, it significantly increases E-FABP expression in macrophages during inflammation. Stimulation of macrophages with EI-05 remarkably enhances lipid droplet formation and IFNβ production, which further promotes the anti-tumor activity of macrophages. Importantly, administering EI-05 in vivo significantly inhibits mammary tumor growth in a syngeneic mouse model. Altogether, these results suggest that EI-05 may represent a promising drug candidate for anti-tumor treatment through enhancing E-FABP activity and IFNβ responses in macrophages.

Expressions and significances of CRABPII and E-FABP in non-small cell lung cancer

背景与目的细胞视黄酸结合蛋白（cellular retinoic acid-binding protein Ⅱ, CRABPⅡ）和表皮型脂肪酸结合蛋白（epidermal fatty acid-binding protein, E-FABP）作为维甲酸（retinoic acid, RA）的转运蛋白，通过RA信号传导通路，从正反两方面影响细胞的增殖和凋亡。本研究旨在探讨CRABPII和E-FABP在非小细胞肺癌（non-small cell lung cancer, NSCLC）中的表达及意义。方法利用组织芯片技术和免疫组织化学SP法检测54例正常肺组织、287例NSCLC原发癌组织以及103例淋巴结转移癌组织中CRABPII和E-FABP的表达。结果CRABPII在NSCLC原发癌组织中的表达与患者的性别、肿瘤的有无转移、TNM分期有关（P < 0.05）。E-FABP在NSCLC原发癌组织中的阳性表达率分别高于正常肺组织和淋巴结转移癌组织（P < 0.05）。在NSCLC原发癌组织中，E-FABP的表达与肿瘤的病理分级、有无转移有关（P < 0.05）。在NSCLC中，E-FABP的阳性表达较CRABPII占优势（P < 0.05），两种蛋白的差异性表达与肿瘤的大小、病理分级、有无转移、TNM分期有关（P < 0.05），瘤体愈大，肿瘤发生转移，临床分期愈晚，E-FABP的表达愈占优势。Kaplan-Meier单因素生存分析显示：CRABPII的表达、CRABPII与E-FABP的差异性表达与NSCLC患者的预后有关（P < 0.05）。结论E-FABP在NSCLC中高表达，其表达的增强可能与NSCLC的发生和演进有关；CRABPII可能在NSCLC的发展过程中起负向调节作用，CRABPII阴性表达患者术后生存率更高，对NSCLC患者预后的评估有一定价值。

Increased expression of epidermal fatty acid-binding protein by alveolar macrophages during acute rejection of rat lungs

In the lung, epidermal fatty acid-binding protein (E-FABP) is expressed by alveolar macrophages (AM) and alveolar epithelial cells type II (AEII). E-FABP may regulate macrophage activation and is involved in the metabolism of surfactant phospholipids. As macrophage activation and surfactant dysfunction are associated with rejection, we hypothesize that E-FABP expression is changed during acute rejection of pulmonary grafts. Orthotopic left lung transplantations were performed in the Dark Agouti to Lewis and in the isogeneic Lewis to Lewis rat strain combinations. E-FABP expression was analyzed in the lung by immunohistochemistry, immunoblotting and quantitative reverse transcription-polymerase chain reaction (RT-PCR). Alveolar leukocytes obtained by bronchoalveolar lavage were analyzed by RT-PCR. Immunohistochemistry of isografts revealed strong E-FABP immunoreactivity in AEII and a moderate immunoreactivity in AM. In allografts undergoing acute rejection, AM exhibiting increased E-FABP immunoreactivity accumulated. Immunoblots revealed a single band at 15 kDa, which corresponds to the expected molecular mass of E-FABP. The levels of E-FABP mRNA were higher in allografts than in isografts and control lungs. Furthermore, alveolar leukocytes isolated by bronchoalveolar lavage from allografts displayed higher E-FABP mRNA expression levels than leukocytes from isografts and controls. In conclusion, we demonstrate for the first time upregulation of E-FABP expression in AM during severe inflammation.

Expression of E‐FABP in PC12 cells increases neurite extension during differentiation: involvement of n‐3 and n‐6 fatty acids

Epidermal fatty acid-binding protein (E-FABP), a member of the family of FABPs, exhibits a robust expression in neurons during axonal growth in development and in nerve regeneration following nerve injury. This study examines the impact of E-FABP expression in normal neurite extension in differentiating pheochromocytoma cell (PC12) cultures supplemented with selected long chain free fatty acids (LCFFA). We found that E-FABP binds to a broad range of saturated and unsaturated LCFFAs, including those with potential interest for neuronal differentiation and axonal growth such as C22:6n-3 docosahexaenoic acid (DHA), C20:5n-3 eicosapentaenoic acid (EPA), and C20:4n-6 arachidonic acid (ARA). PC12 cells exposed to nerve growth factor (NGFDPC12) exhibit high E-FABP expression that is blocked by mitogen-activated protein kinase kinase (MEK) inhibitor U0126. Nerve growth factor-differentiated pheochromocytoma cells (NGFDPC12) antisense clones (NGFDPC12-AS) which exhibit low E-FABP expression have fewer/shorter neurites than cells transfected with vector only or NGFDPC12 sense cells (NGFDPC12-S). Replenishing NGFDPC12-AS cells with biotinylated recombinant E-FABP (biotin-E-FABP) protein restores normal neurite outgrowth. Cellular localization of biotin-E-FABP in NGFDPC12 was detected mostly in the cytoplasm and in the nuclear region. Treatment of NGFDPC12 with DHA, EPA, or ARA further enhances neurite length but it does not trigger further induction of TrkA or MEK phosphorylation or E-FABP mRNA observed in differentiating PC12 cells without LCFFA supplementation. Significantly, DHA and EPA neurite stimulating effects are higher in NGFDPC12-S than in NGFDPC12-AS cells. These findings are consistent with the scenario that neurite extension of differentiating PC12 cells, including further stimulation by DHA and EPA, requires sufficient cellular levels of E-FABP.

Analysis on the phenotype of E-FABP-gene knockout mice.

The fatty acids are shown to be critical in the maintenance of the water permeability barrier that is ascribed to the lipids in the intracellular milieu of the cornified cell layer in the epidermis. In view of this importance in the skin, we examined the phenotype of epidermal fatty acid binding protein (E-FABP)-deficient mice. In spite of total lack of E-FABP expression in the various tissues of E-FABP deficient mice, these animals appeared normal in gross and histological examination. In Northern blot analysis for other FABPs, the gene expression of heart (H-)-type FABP is specifically elevated in the liver of neonatal heterozygous and homozygous mice, suggesting the functional compensation of H-FABP for E-FABP deficiency during their development. In functional analyses of the skin, the basal transepidermal water loss (TEWL) of the adult homozygous mice showed lower levels compared with the wild-type mice, and the impairment of recovery in TEWL was observed in the homozygous mice when the lipid barrier of the skin was disrupted by acetone. These results demonstrate that E-FABP is responsible for the water permeability barrier of the skin, although the molecular mechanism remains to be further elucidated.