Effects of dietary high fat on prostate intraepithelial neoplasia in TRAMP mice.

Increased fat intake is known to be a major cause of prostate cancer. In this study, we investigated the effect of dietary high fat on prostate intraepithelial neoplasia using transgenic adenocarcinoma mouse prostate (TRAMP) mice. Six-week-old male TRAMP mice were fed AIN93G (control group, 4.0 kcal/kg, n=6) and AIN93G-HFD (experimental group, 4.8 kcal/kg, n=7) for 10 weeks. Prostate histopathology, urogenital tract (UGT) weight, epididymal white adipose tissue weight, argyrophilic nucleolar organizer regions (AgNORs) counts, and serum leptin levels were examined. AIN93G-HFD fed group showed progressed neoplastic lesions in the prostate (P<0.05) compared to AIN93G fed group. AIN93G-HFD intake resulted in a increase in the weight of UGT (P<0.05) and epididymal white adipose tissue. The number of Ag-NOR positive dots significantly increased in each prostate lobe and final serum leptin levels in AIN93G-HFD fed group were about twice those of AIN93G fed group (P<0.05). Dietary high fat was related to the prostate cancer progression in the early stage of TRAMP mice and increased serum leptin levels, suggesting that the regulation of dietary components could delay the progression of prostate cancer.

Prevalence of murine norovirus infection in Korean laboratory animal facilities.

Currently, murine noroviruses (MNV) are the most prevalent viral pathogens identified in laboratory animal facilities. While several reports exist concerning the prevalence of MNV in North American research facilities, very few reports are available for other parts of the world, including Korea. This study evaluated the prevalence of MNV infection in 745 murine sera collected from 15 animal facilities in Korea by enzyme linked immunosorbent assay (ELISA). Positive cases were subcategorized by murine strain/genetics, housing environments and animal sources. In summary, 6.6% of inbred/outbred mice purchased from commercial vendors were seropositive, 9.6% of in-house colonies were seropositive and 27.0% of genetically modified mice (GMM) were seropositive. Partial gene amplification of fecal isolates from infected animals showed that they were homologous (100%) with MNV-4.

Microbiological quality assessment of laboratory mice in Korea and recommendations for quality improvement.

Regular monitoring of commercial laboratory rodents and institutional research animal residents is essential for microbiological quality control programs. The objective of our study was to investigate the recent prevalence of infectious pathogens in laboratory mice from eight experimental animal vendors and 56 institutional animal facilities in Korea. Our investigation was conducted in 2006-2007. Specific Pathogen Free (SPF) mice from four commercial breeders were clean according to serological, bacteriological, parasitological, and histopathological examination results. However, mice from one intermediate vendor that distributed SPF animals from main commercial vendors to local districts had Syphacia obvelata and Mycoptes musculinus infections. Additionally, mice from conventional animal breeders were highly contaminated. Among the 56 institutional animal facilities, mouse hepatitis virus (MHV), Sendai virus and Mycoplasma pulmonis positive results were obtained in 23.2, 8.9, and 1.8% of animals tested, respectively. These results indicate that quarantine and eradication efforts of infectious pathogens in these facilities are sub-optimal and need to be improved. The use of commercial conventional mice for research should be eliminated and appropriate vendor selection as well as thorough quarantine before releasing animals into a facility are needed. Finally we suggest qualified veterinary experts are needed at each animal facility to ensure an adequate health surveillance program.

Highly expressed recombinant human follicle-stimulating hormone from Chinese hamster ovary cells grown in serum-free medium and its effect on induction of folliculogenesis and ovulation.

OBJECTIVE: To develop efficient Chinese hamster ovary (CHO) cells that express recombinant human FSH (rhFSH) in serum-free conditions and to investigate the effect of this newly synthesized rhFSH on folliculogenesis and ovulation. DESIGN: Experimental study. SETTING: Seoul National University, South Korea. ANIMALS: Forty immature hypophysectomized rats and 40 androgen-sterilized mice. INTERVENTION(S): A stable single CHO cell that expresses rhFSH at a high level was obtained by introducing the human chorionic gonadotropin (hCG) alpha-subunit and FSH beta-subunit genes. After purification processing, we investigated the effect of this newly synthesized rhFSH on folliculogenesis in hypophysectomized rats and ovulation in androgen-sterilized mice. MAIN OUTCOME MEASURE(S): The ovary weight, uterine weight, number of follicles, and ovarian morphology were evaluated in immature hypophysectomized rats. The number of ovulated oocytes and ovarian morphology were examined in androgen-sterilized mice. RESULT(S): After purification processing, we analyzed the new rhFSH using matrix-associated laser desorption ionization-time of flight and found that this new rhFSH increased both ovarian weight and uterine weight in hypophysectomized rats and induced ovulation in androgen-sterilized mice. CONCLUSION(S): This newly synthesized rhFSH might be safely used in anovulatory infertile woman as well as in ovulation induction protocols for subfertile women.

Acute and Subchronic Inhalation Toxicity of n-Octane in Rats.

OBJECTIVES: We have investigated the toxic effects of the inhalation of subchronic and acute levels of n-octane. METHODS: The rats were exposed to n-octane of 0, 2.34, 11.68 and 23.36 mg/L (n = 5 rats/group/gender) in an acute inhalation test (Organization for Economic Co-operation and Development (OECD) TG 403), or to 0, 0.93, 2.62 and 7.48 mg/L (n = 10 rats/group/gender) for a subchronic inhalation test (OECE TG 413), to establish a national chemical management system consistent with the Globally Harmonized Classification System (GHS). RESULTS: Acutely-exposed rats became lethargic but recovered following discontinuation of inhalation. Other clinical symptoms such as change of body weight and autopsy finds were absent. The LC50 for the acute inhalation toxicity of n-octane was determined to exceed 23.36 mg/L and the GHS category was 'not grouping'. Subchronically-treated rats displayed no significant clinical and histopathological differences from untreated controls; also, target organs were affected hematologically, biochemically and pathologically. Therefore, the no observable adverse effect level was indicated as exceeding 7.48 mg/L and the GHS category was 'not grouping' for the specific target organ toxicity upon repeated exposure. CONCLUSION: However, n-octane exposure should be controlled to be below the American Conference of Industrial Hygienists recommendation (300 ppm) to prevent inhalation-related adverse health effects of workers.

Estrogen-responsive transient expression assay using a brain aromatase-based reporter gene in zebrafish (Danio rerio).

Whereas endogenous estrogens play an important role in the development, maintenance, and function of female and male reproductive organs, xenoestrogens present in the environment disrupt normal endocrine function in humans and wildlife. Various in vivo and in vitro assays have been developed to screen these xenoestrogens. However, traditional in vivo assays are laborious and unsuitable for large-scale screening, and in vitro assays do not necessarily replicate in vivo functioning. To overcome these limitations, we developed a transient expression assay in zebrafish, into which a brain aromatase (cyp19a1b)-based estrogen-responsive reporter gene was introduced. In response to 17beta-estradiol (10(-6) M) and heptachlor (10(-6) M), zebrafish embryos carrying the reporter construct expressed enhanced green fluorescent protein in the olfactory bulb, telencephalon, preoptic area, and mediobasal hypothalamus. This system will serve to model the in vivo conversion and breakdown of estrogenic compounds and thus provide a rapid preliminary screening method to estimate their estrogenicity.

Developmental toxicity and brain aromatase induction by high genistein concentrations in zebrafish embryos.

Genistein is a phytoestrogen found at a high level in soybeans. In vitro and in vivo studies showed that high concentrations of genistein caused toxic effects. This study was designed to test the feasibility of zebrafish embryos for evaluating developmental toxicity and estrogenic potential of high genistein concentrations. The zebrafish embryos at 24 h post-fertilization were exposed to genistein (1 x 10(-4) M, 0.5 x 10(-4) M, 0.25 x 10(-4) M) or vehicle (ethanol, 0.1%) for 60 h. Genistein-treated embryos showed decreased heart rates, retarded hatching times, decreased body length, and increased mortality in a dose-dependent manner. After 0.25 x 10(-4) M genistein treatment, malformations of survived embryos such as pericardial edema, yolk sac edema, and spinal kyphosis were also observed. TUNEL assay results showed apoptotic DNA fragments in brain. This study also confirmed the estrogenic potential of genistein by EGFP expression in the brain of the mosaic reporter zebrafish embryos. This study first demonstrated that high concentrations of genistein caused a teratogenic effect on zebrafish embryos and confirmed the estrogenic potential of genistein in mosaic reporter zebrafish embryos.

Protective effects of vitamin E against 3,3',4,4',5-pentachlorobiphenyl (PCB126) induced toxicity in zebrafish embryos.

3,3',4,4',5-Pentachlorinated biphenyls 126 (PCB126) is a global environmental contaminant that can induce cellular oxidative stress. We investigated whether vitamin E can protect against toxicity from PCB126 during zebrafish (Danio rerio) development. Zebrafish embryos were exposed to 100nM PCB126 and compared with a second group that was co-exposed with 100muM vitamin E until 5 days post fertilization. PCB126 induced pericardial sac edema, yolk sac edema, and growth retardation in zebrafish embyos. In contrast, vitamin E co-exposure group did not show any gross changes. Real-time PCR results showed that vitamin E co-exposure group were restored to control group for the expression levels of heat shock protein 70 Cognate, aryl hydrocarbon receptor type-2, cytochrome P450 1A, and superoxide dismutase-1. These data give insights into the use of vitamin E to reduce PCB126-mediated toxicity and into the use of zebrafish embryos for exploring mechanisms underlying the oxidative potential of AHR agonists.

Benomyl induction of brain aromatase and toxic effects in the zebrafish embryo.

Benomyl is a benzimidazole fungicide that has been widely used on a variety of food crops and ornamental plants. It is known to cause adverse effects on reproductive systems, including decreased testicular and epididymal weights and reduced epididymal sperm counts and fertility. The brain aromatase gene is up-regulated by estrogens and estrogen mimics and considered a target gene to screen estrogen mimics. This study was designed to test the estrogenic potential and toxic effects of benomyl in the zebrafish system, and validated this system as a model that may correspond to the effect of benomyl in rodents. Concentrations of 20 x 10(-6), 40 x 10(-6) and 80 x 10(-6) M of benomyl-treated embryos showed decreased survival, hatching and heart rates, and increased incidence of malformations, such as pericardial edema, spinal lordosis, elongated heart, head edema, eye lens protrusion and caudal fin disappearance. Benomyl induced enhanced green fluorescent protein (EGFP) expression in the mediobasal hypothalamus (MBH) in transient zebrafish embryos with a brain aromatase-based reporter gene. In this study, we determined that benomyl has estrogenic potential based on zebrafish brain aromatase gene induction, and that benomyl is toxic at 20 x 10(-6) M concentration and higher. These results demonstrate the usefulness of zebrafish embryos as an in vivo system to examine the estrogenic and developmental toxic potential of unknown compounds.

Detection of antibodies against SARS-Coronavirus using recombinant truncated nucleocapsid proteins by ELISA.

Severe acute respiratory syndrome (SARS) is a lifethreatening emerging respiratory disease caused by the coronavirus, SARS-CoV. The nucleocapsid (N) protein of SARS-CoV is highly antigenic and may be a suitable candidate for diagnostic applications. We constructed truncated recombinant N proteins (N1 [1-422 aa], N2 [1- 109 aa], and N3 [110-422 aa]) and determined their antigenicity by Western blotting using convalescent SARS serum. The recombinants containing N1 and N3 reacted with convalescent SARS serum in Western blotting. However, the recombinant with N2 did not. In ELISA using N1 or N3 as the antigens, positive results were observed in 10 of 10 (100%) SARS-CoV-positive human sera. None of 50 healthy sera gave positive results in either assay. These data indicate that the ELISA using N1 or N3 has high sensitivity and specificity. These results suggest that the middle or C-terminal region of the SARS N protein is important for eliciting antibodies against SARS-CoV during the immune response, and ELISA reactions using N1 or N3 may be a valuable tool for SARS diagnosis.

Search for potential target site of nucleocapsid gene for the design of an epitope-based SARS DNA vaccine.

It is believed today that nucleocapsid protein (N) of severe acute respiratory syndrome (SARS)-CoV is one of the most promising antigen candidates for vaccine design. In this study, three fragments [N1 (residues: 1-422); N2 (residues: 1-109); N3 (residues: 110-422)] of N protein of SARS-CoV were expressed in Escherichia coli and analyzed by pooled sera of convalescence phase of SARS patients. Three gene fragments [N1 (1-1269 nt), N2 (1-327 nt) and N3 (328-1269 nt)-expressing the same proteins of N1, N2 and N3, respectively] of SARS-N were cloned into pVAX-1 and used to immunize BALB/c mice by electroporation. Humoral (by enzyme-linked immunosorbent assay, ELISA) and cellular (by cell proliferation and CD4(+):CD8(+) assay) immunity was detected by using recombinant N1 and N3 specific antigen. Results showed that N1 and N3 fragments of N protein expressed by E. coli were able to react with sera of SARS patients but N2 could not. Specific humoral and cellular immunity in mice could be induced significantly by inoculating SARS-CoV N1 and N3 DNA vaccine. In addition, the immune response levels in N3 were significantly higher for antibody responses (IgG and IgG1 but not IgG2a) and cell proliferation but not in CD4(+):CD8(+) assay compared to N1 vaccine. The identification of antigenic N protein fragments has implications to provide basic information for the design of DNA vaccine against SARS-CoV. The present results not only suggest that DNA immunization with pVax-N3 could be used as potential DNA vaccination approaches to induce antibody in BALB/c mice, but also illustrates that gene immunization with these SARS DNA vaccines can generate different immune responses.

Protective and therapeutic effects of an extract mixture of alder tree, labiate herb, milk thistle green bean-rice bran fermentation, and turnip against ethanol-induced toxicity in the rat.

An herbal extract mixture and yogurt added to the herbal extract mixture were tested for their protective and therapeutic effects on ethanol-induced liver injury. The herbal extract mixture, yogurt and commercial drugs were used for treatment for two weeks prior to administering a single oral dose of ethanol (3 g/kg body weight). The herbal extract mixture and yogurt added to the herbal extract mixture were found to provide protection against ethanol-induced toxicity comparable to the commercial drug treatment, according to the serum and histopathological analysis. It was also shown that co-treatment with herbal extract mixture and yogurt against a triple oral dose of ethanol (2 g/kg body weight, over one week) provided protection against ethanol toxicity. After the initial set of experiments, the herbal extract mixture and yogurt treatments were extended for three more weeks. When compared to the positive control, further treatment with both the herbal extract and yogurt significantly reduced liver injury and resulted in a lower grade of lipid deposition.

Quantitative GFP fluorescence as an indicator of arsenite developmental toxicity in mosaic heat shock protein 70 transgenic zebrafish.

In transgenic zebrafish (Danio rerio), green fluorescent protein (GFP) is a promising marker for environmental pollutants. In using GFP, one of the obstacles which we faced was how to compare toxicity among different toxicants or among a specific toxicant in different model species with the intensity of GFP expression. Using a fluorescence detection method, we first validated our method for estimating the amount of GFP fluorescence present in transgenic fish, which we used as an indicator of developmental toxicity caused by the well-known toxicant, arsenite. To this end, we developed mosaic transgenic zebrafish with the human heat shock response element (HSE) fused to the enhanced GFP (EGFP) reporter gene to indicate exposure to arsenite. We confirmed that EGFP expression sites correlate with gross morphological disruption caused by arsenite exposure. Arsenite (300.0 microM) caused stronger EGFP fluorescence intensity and quantity than 50.0 microM and 10.0 microM arsenite in our transgenic zebrafish. Furthermore, arsenite-induced apoptosis was demonstrated by TUNEL assay. Apoptosis was inhibited by the antioxidant, N-acetyl-cystein (NAC) in this transgenic zebrafish. The distribution of TUNEL-positive cells in embryonic tissues was correlated with the sites of arsenite toxicity and EGFP expression. The EGFP values quantified using the standard curve equation from the known GFP quantity were consistent with the arsenite-induced EGFP expression pattern and arsenite concentration, indicating that this technique can be a reliable and applicable measurement. In conclusion, we propose that fluorescence-based EGFP quantification in transgenic fish containing the hsp70 promoter-EGFP reporter-gene construct is a useful indicator of development toxicity caused by arsenite.

Evaluation of phoP and rpoS mutants of Salmonella enterica serovar Typhi as attenuated typhoid vaccine candidates: virulence and protective immune responses in intranasally immunized mice.

The attenuation and immunoenhancing effects of rpoS and phoP Salmonella enterica serovar strain Typhi (Salmonella typhi) mutants have not been compared. Here, three S. typhi deletion mutants (phoP, rpoS, and rpoS-phoP double mutant) are constructed and these mutants are characterized with respect to invasiveness, virulence, and protective immune response compared with wild-type Ty2. It was found that phoP and phoP-rpoS deletion mutants are less invasive to HT-29 cells than the wild-type Ty2 and the rpoS single-deleted strain. The LD(50) of immunized mice was higher for phoP than for rpoS mutants, and the highest for the phoP-rpoS double mutant. In addition, all S. typhi mutants showed an increase in the specific serum IgG levels and T-cell-mediated immunity, and showed equal protection abilities against a wild-type Ty2 challenge after two rounds of immunization in BALB/c mice. It is concluded that phoP genes appear to play a more important role than rpoS genes in both cellular invasion and virulence of S. typhi, but not in immunogenicity in mice. Furthermore, the data indicate that the phoP-rpoS double mutant may show promise as a candidate for an attenuated typhoid vaccine.

Arsenite-induced apoptosis is prevented by antioxidants in zebrafish liver cell line.

This study evaluated oxidative stress-induced apoptosis as a possible mechanism of arsenite toxicity in zebrafish liver cell line (ZFL cells). The heat shock protein 70 (HSP70), a chaperone protein, appears to provide protection against oxidative stress and apoptosis. Using the MTT assay, we demonstrated that survival of ZFL cells treated with arsenite for 24h decreased in a dose-dependent manner. The possible mechanisms that promote the cytotoxicity of arsenite were addressed. Cell viability assays revealed that arsenite caused a dose-dependent increase in cell death, and pretreatment of the ZFL cells with antioxidants blunted these effects. Antioxidants such as N-acetyl-cysteine (NAC, 5 mM) and dithiothreitol (DTT, 80 microM) significantly prevented ZFL cells from arsenite-induced death. Nuclear staining was performed using 1 microg/ml Hoechst, and cells were analyzed with a fluorescent microscope. Arsenite (30 microM) induced massive apoptosis that was identified by morphology and condensation and fragmentation of the nuclei of the ZFL cells. Pretreatment with NAC or DTT before arsenite insult effectively protected the cells against oxidative stress-induced apoptosis from the arsenite. Using a transfected human hsp 70 promoter-enhanced green fluorescent protein (EGFP) reporter, pHhsp70-EGFP, the induction of HSP70 against oxidative stress-induced apoptosis by arsenite was observed. The induction of HSP70 by arsenite increased in a dose-dependent manner, and pretreatment of transfected ZFL cells with NAC or DTT before arsenite insult reduced EGFP expression. Taken together, our results provide evidence that stimulation of the heat shock response is a sensitive biomarker of arsenic exposure and that arsenite causes oxidative stress-induced apoptosis in ZFL cells.

Microbiological monitoring of guinea pigs reared conventionally at two breeding facilities in Korea.

In this study, microbiological monitoring of guinea pigs reared conventionally in two facilities was performed twice in 2004, with a three-month-interval between surveys. This study was based on the recommendations of the FELASA Working Group, with some modifications. In serological tests in the first survey, some animals from facility A showed positive results for Encephalitozoon cuniculi, Sendai virus, pneumonia virus of mice (PVM), and Reovirus-3 (Reo-3); facility B showed a positive result only for E. cuniculi. The results of the second survey were similar to the first, except for the presence of Sendai virus; all animals from the two facilities were Sendai virus-negative in the second experiment. No pathogenic bacteria were cultured in the organs of any of the animals in the first survey. However, in the second survey, Bordetella bronchiseptica was cultured from the lung tissue of two 10-week-old animals from facility A. Chlamydial infection was examined by the Macchiavello method, but no animal showed positive results. Tests using fecal flotation or the KOH wet mount method showed no infection of endoparasites, protozoa, ectoparasites, or dermatophytes in any animal in both surveys. However, in the histopathological examination, an infection of protozoa-like organisms was observed in the cecum of some animals from facility A. The present study revealed that microbiological contamination was present in guinea pigs reared conventionally in two facilities in Korea, suggesting that there is a need to improve environmental conditions in order to eradicate microbial contamination.

Human originated bacteria, Lactobacillus rhamnosus PL60, produce conjugated linoleic acid and show anti-obesity effects in diet-induced obese mice.

Many previous studies have reported that conjugated linoleic acid could be produced by starter culture bacteria, but the effects of the bacteria were not investigated. Moreover, there was no evidence of the conjugated linoleic acid-producing bacteria having potential health or nutritional effects related to conjugated linoleic acid, including reducing body fat. Here, we investigated the anti-obesity effect of Lactobacillus rhamnosus PL60, a human originated bacterium that produces t10, c12-conjugated linoleic acid, on diet-induced obese mice. After 8 weeks of feeding, L. rhamnosus PL60 reduced body weight without reducing energy intake, and caused a significant, specific reduction of white adipose tissue (epididymal and perirenal). Although the size of epididymal adipocytes was not reduced by L. rhamnosus PL60, apoptotic signals and UCP-2 mRNA levels increased in adipose tissue. Liver steatosis, a well known side effect of CLA, was not observed by L. rhamnosus PL60 treatment; on the contrary it seemed to be normalized. Results showed that the amount of conjugated linoleic acid produced by Lactobacillus rhamnosus PL60 was enough to produce an anti-obesity effect.

Specific activation of the human HSP70 promoter by copper sulfate in mosaic transgenic zebrafish.

Heat shock proteins (HSPs) play a central role in cell protection and repair upon stresses, such as that caused by heat and heavy metals. Copper sulfate inducibility of a pHhsp70 construct expressing the enhanced green fluorescent protein (EGFP) gene under the control of the exogenous human hsp70 promoter was tested in transfected CHSE 214 cells and transgenic zebrafish (Danio rerio). We developed a transient expression system, using mosaically transgenic zebrafish, which allows rapid analysis of transgenic expression. Transfected CHSE 214 cells which had been exposed to 250 nM and 2.5 microM copper sulfate for up to 24h showed increased EGFP expression in a dose-dependent manner. The 1.5 microM copper sulfate caused stronger EGFP fluorescence than the 1.0 microM copper sulfate in transgenic zebrafish. Most of the expression was spotty and was detected in the gills, dorsal and ventral retina, myotubes of the trunk, and skin epithelium. Transgenic zebrafish exposed to copper sulfate exhibited gross dysmorphogenesis, edema and trunk abnormalities, such as spinal lordosis, in vertebral development 5 days after fertilization. This transgenic zebrafish system was sensitive enough to detect copper sulfate at doses below the median lethal concentration (the LC50 was calculated to be 1.2 microM (95% confidence interval of 0.6-1.9 microM)). These results indicate that zebrafish could be useful transgenic biosensor systems for the detection of xenobiotic toxicants in the environment.

Use of PCR-restriction fragment length polymorphism for the identification of zoonotic mycobacteriosis in zebrafish caused by Mycobacterium abscessus and Mycobacterium chelonae.

Skin ulcers, scoliosis, and dropsy-like scale edema were observed in laboratory-maintained zebrafish. Affected fish had multifocal granulomas not only in internal organs such as the liver, intestine, genital organs, kidney, muscle, and spleen but also in the fin, epithelium, gills, and sclera of the eyes. Large numbers of acid-fast-rod-shaped bacteria were observed within the necrotic centers of well-demarcated, multifocal granulomas with Gram's stain and Ziehl-Neelson's stain. The size of the Mycobacterium spp. was 1-2 microm x 2-3 microm with a double-layered cell wall, based upon electron-microscopical features. Definitive diagnosis of these outbreaks was obtained by culture on selective media followed by PCR-restriction fragment length polymorphism analysis (PRA) of the rpoB gene for species identification. The amplified 360-bp products of the rpoB gene of mycobacteria isolated from zebrafish were digested with MspI restriction enzyme, which revealed unique band patterns matching those of Mycobacterium abscessus and Mycobacterium chelonae which are responsible for skin and soft tissue infection caused by rapidly growing mycobacteria in humans. This is the first documentation of the precise identification of zoonotic non-tuberculous mycobacteria isolated from laboratory-maintained zebrafish by the PRA of the rpoB gene; this study thus provides a great deal of useful epidemiological information and reduces the likelihood that epizootics will occur.

Potential antimicrobial effects of human lactoferrin against oral infection with Listeria monocytogenes in mice.

Listeria monocytogenes is a food-borne pathogen that causes serious listeriosis in humans. Antimicrobial effects of human lactoferrin (hLF) against L. monocytogenes have been clearly demonstrated in in vitro studies. However, in vivo studies have not been reported yet. This study investigated whether the oral administration of hLF could inhibit oral infection of listeria in BALB/c mice. The MICs for several strains of L. monocytogenes were determined, and the most sensitive strain was used for the animal work. hLF was administered to BALB/c mice for 7 days, commencing 4 days before oral infection. The effect of hLF was determined by bacterial enumeration and histopathological analysis of the liver and spleen, which are well-known as the major targets of oral listeria infection in mice. In bacterial enumeration, hLF decreased the number of L. monocytogenes cells in the liver. Histopathologically, the size and frequency of necrotic foci in the liver samples decreased with hLF administration. However, these changes were not observed in the spleen samples. The mRNA levels of inflammatory cytokines, such as interleukin (IL)-1beta, tumour necrosis factor (TNF)-alpha and interferon (IFN)-gamma, decreased in the liver of mice receiving hLF. This study has shown that hLF decreases the hepatic colonization of L. monocytogenes, hepatic necrosis and expression of inflammatory cytokines. It revealed that perorally given hLF could mediate antimicrobial and anti-inflammatory activities remote from the gut (i.e. in the liver) of mice challenged with L. monocytogenes.