A DNA Microarray Analysis of Rat Intestine, Spleen and Liver Transcriptome after Oral Administration of Lavender Oil
ABSTRACT: Lavender oil (LO) – a commonly used oil in aromatherapy with well defined volatile components linalool and linalyl acetate – use in non-traditional medicine is increasing globally. To understand and provide evidence for the potential positive effects of LO on the body, we have established an animal model for investigating in this current study, orally administered LO effects genome-wide in rat intestine, spleen and liver. The rats were administrated LO at 5 mg/kg (usual threaupeutic dose in humans) followed by screeing of differentially expressed genes in the tissues utlilizing a 4x44K whole genome rat chip (Agilent microarray platform) in conjunction with a dye-swap approach, one of the novelties of this study. Fourteen days after treatment (LO) and in comparison with a control group (sham), a total of 156 and 154 up (>1.5 fold)- and down (<0.75 fold)-regulated genes, 174 and 66 up- (>1.5 fold)- and down (<0.75 fold)-regulated genes, and 222 and 322 up- (>1.5 fold)- and down (<0.75 fold)-regulated genes showed differential expression at the mRNA level, in the intestine, spleen and liver, respectively. Reverse transcription-polymerase chain reaction (RT-PCR) validation of highly up- and down-expressed genes revealed the regulation of Papd4, Lrp1b, Alb, Cyr61, Cyp2c, and Cxcl1 genes, by LO, as examples in these tissues. Using bioinformatics functionally categorization of differentially expressed genes was done by their Gene Ontology (GO) revealing their diverse functions and potential roles in LO-mediated effects in rat. Present results are a first such inventory of genes affected by the essential oil of Lavender (LO) in an animal model. Rats were administrated LO at 5 mg/kg (the usual threaupeutic dose in humans) followed by the screeing of differentially expressed genes in the intestine, spleen, and liver utlilizing a 4x44K whole genome rat chip (Agilent) and the dye-swap approach.
Project description:Lavender oil (LO) is commonly used oil in aromatherapy, with well-defined volatile components linalool and linalyl acetate, in non-traditional medicine. To understand and demonstrate the potential positive effects of LO on the body, we have established an animal model for investigating the orally administered LO effects genome-wide in the rat tissues. In this data submission, we investigate the effect of LO ingestion in the blood. These results are the first such inventory of genes that are affected by lavender essential oil (LO) in the blood of an animal model, forming the basis for further functional analysis and investigation. Briefly, rats were administered LO at 5 mg/kg (usual therapeutic dose in humans) followed by screening of differentially expressed genes in the tissues, using a 4×44-K whole-genome rat chip (Agilent microarray platform; Agilent Technologies, Palo Alto, CA, USA) in conjunction with a dye-swap approach.
Project description:Human epidemiological evidence has led scientists to theorize that undernutrition during gestation is an important early origin of adult diseases. Animal models have successfully demonstrated that maternal diet could contribute to some of adult diseases. Undernutrition is perceived harmful in pregnant women whereas calorie restriction is a strategy proven to extend healthy and maximum life span in adult. This diagrammatically opposite effect of nutritional condition might provide us hints to search for genes underlying health conditions. Here, we have initiated a study examining the effect of undernutrition on maternal and fetal livers, utilizing high-throughput DNA microarray analysis for screening genome-wide changes in their transcriptomes. Briefly, pregnant mice were exposed to food deprivation (FD) on gestation day (GD) 17, and caesarean section was performed on GD18. Control mice were supplied with chow ad libitum till sacrifice. Total RNA extracted from mother and fetal livers for each control and treatment (FD) was analyzed with an Agilent mouse whole genome DNA chip. Total of 3058 and 3126 up (>1.5 fold)- and down (<0.75 fold)-regulated genes, and 1475 and 1225 up- (>1.5 fold)- and down (<0.75 fold)-regulated genes showed differential expression at the mRNA level, in the maternal and fetal livers, respectively. Interestingly, 103 genes up-regulated in mother were down-regulated in fetus, whereas 108 down-regulated maternal genes were up-regulated in fetus; these 211 genes are potential candidates related to longevity or health. The role of some of these genes, in context of the proposed mechanisms for developmental origins of health and disease is discussed. Comparison between mouse control and FD livers in fetus and mother was performed. Five to ten biological replicates were used, and pooled total RNA from each condition (control and FD) was dye-swaped.
Project description:A diet rich in nucleic acids and protamin protein, termed as nucleoprotein was used for the study. Mice were fed with NP diets for 4 weeks followed by removal of the liver and spleen. Total RNA extracted from livers and spleens was pooled in each group (low NP or LNP-control, and 1.2% NP-treatment, diets), prior to DNA microarray analysis (Agilent mouse whole genome 4 x 44K). Results revealed 1373 & 3386 up (>1.5 fold)- and down (<0.75 fold)-regulated genes in the liver, and 252 & 1838 up- and down-regulated genes in the spleen, respectively following 1.2%NP diet. Analysis of genes related to NP diets will be discussed. To investigate the effect of NP, we added S-nuclegen® at a concentration of 1.2% into CLEA basic purified diet (CLEA JAPAN, Inc., Tokyo, Japan) known to include nucleic acids (NAs) at much lower amounts than standard diet. By HPLC analysis CLEA basic purified diet (low NP), and 1.2% NP diet included 0.03%, and 0.5% NAs, respectively. Male C57BL/6J mice (13 weeks) were fed with NP diet for 4 weeks. The liver and spleen were removed and deep frozen in liquid nitrogen. Whole blood was also sampled from these mice, and frozen in liquid nitrogen. Total RNA extracted from livers and spleens was pooled in each group (control, lowNP or LNP; and treatment, 1.2%NP), prior to DNA microarray analysis (Agilent mouse whole genome 4 x 44K).
Project description:A diet rich in nucleic acids and protamin protein, termed as nucleoprotein (NP), has been attracting a great deal of attention in food science for its beneficial effects. In the present study, we performed a global gene expression profiling in mice fed with NP diet rich in nucleoproteins from the salmon testis. Since our recent research has revealed anti-inflammatory effect of the NP diet, we induced inflammation by lipopolysaccharide (LPS) injection in the mice for this analysis study. Mice were fed with NP diet for 4 weeks followed by a single injection of LPS. The liver and spleen were removed 6 h post-LPS injection. Total RNA extracted from livers (L) and spleens (S) was pooled in each group (control and NP diet), prior to DNA microarray analysis (Agilent mouse whole genome 4 x 44K). Results revealed 322 & 702 up (>1.5 fold)- and down (<0.75 fold)-regulated genes in the liver, and 325 & 501 up- and down-regulated genes in the spleen, respectively following NP diet. Analysis of genes related to inflammation suggests increased activity of immune function during acute period of LPS injection, which may contribute to early demise of inflammation. NP diet can be expected to be useful for inhibition of inflammatory reactions whose over-accumulation is thought to be related to the acceleration of aging process. To investigate the effect of NP, we added S-nuclegen® at a concentration of 1.2% into CLEA basic purified diet (CLEA JAPAN, Inc., Tokyo, Japan) known to include nucleic acids (NAs) at much lower amounts than standard diet. By HPLC analysis CLEA basic purified diet (low NP), and 1.2% NP diet included 0.03%, and 0.5% NAs, respectively. Male C57BL/6J mice (7 weeks) were fed with NP diet for 4 weeks followed by a single injection of LPS at a dose of 10 mg/kg. The liver and spleen were removed 6 h post-LPS injection. Total RNA extracted from livers and spleens was pooled in each group (control and NP diet), prior to DNA microarray analysis (Agilent mouse whole genome 4 x 44K).
Project description:7,12-Dimethylbenz[a]anthracene (DMBA)-induced rat mammary carcinoma is a well-recognized model, however, the genetic alterations during its carcinogenesis have yet to be determined. We used laser capture microdissection to specifically isolate cells from terminal end buds (TEBs), the origin of carcinoma, at 2 weeks after sesame oil- treatment (control) or DMBA-treatment (DMBA-TEBs), ductal carcinoma in situ (DCIS) and invasive mammary carcinoma (MC). Using an oligonucleotide microarray representing 20600 rat probe sequences, we analyzed gene expression profiles and validated mRNA and protein levels of genes of interest by real-time quantitative PCR, immunohistochemistry and immunofluorescence. The number of differentially expressed genes was smallest in DMBA-TEBs (63), followed by DCIS (798) and MC (981). Only the expression of PEP-19, an anti-apoptic gene, showed significant increases in DMBA-TEBs (4-fold), DCIS (10-fold) and MC (16-fold). MMP-13 expression was increased markedly in DCIS (19-fold) and MC (61-fold) while OPN expression was increased 6-fold in DCIS and 8-fold in MC. MMP-7 expression was increased 4-fold in MC. Nidogen-1; a participant in the assembly of basement membranes, TSP-2; an inhibitor of angiogenesis and COUP-TFI; a transcription repressor showed significant decreases in DCIS (4-, 9-, and 17-fold, respectively) and MC (10-, 37-, and 100-fold). Network analyses with IPA software revealed that the most significant network was centered on Akt groups in DCIS and ERK groups in MC. The present findings provide insights into the molecular changes and suggest the importance of PEP-19 overexpression very early on during mammary carcinogenesis. Keywords: Comparative experiments of sesame oil treatment（control）and 7,12-Dimethylbenz[a]anthracene（DMBA）treatment group. Or comparative experiments of tissue type Experiment：7,12-Dimethylbenz[a]anthracene (DMBA)-induced rat mammary carcinoma is a well-recognized model, however, the genetic alterations during carcinogenesis have yet to be determined. We used laser capture microdissection to specifically isolate the cells from terminal end buds (TEBs), the origin of MC, at 2 weeks after sesame oil-(control) or DMBA-treatment (DMBA-TEBs), ductal carcinoma in situ (DCIS) and invasive mammary carcinoma (MC). At 50 days after birth, inbred Sprague-Dawley female rats were given 1ml sesame oil (control) or 20 mg DMBA (Nacalai tesque, Inc. Kyoto Japan) dissolved in 1ml sesame oil, by gastric intubations. Animals in control group were euthanized after 2 weeks (W) and DMBA-treated group were euthanized after 2 weeks (W), 6W and 8W, respectively. From for 4 sample(control, DMBA-TEBs, DCIS, MC), we performed a hybridization repeatedly, respectively.
Project description:To investigate the effects of corn oil (CO), common drug vehicle, on the gene expression profiles in rat thymus with microarray technique. Female Wistar Rats were administered daily with normal saline (NS), CO 2, 5, 10 ml/kg for 14 days, respectively. Then, the thymus samples of rats were collected for microarray test and histopathology examination. The microarray data showed that 0, 40, 458 differentially expressed genes (DEGs) in 2, 5, 10 ml/kg CO group compared to NS group, respectively. The altered genes were associated with immune response, cellular response to organic cyclic substance, regulation of fatty acid beta-oxidation, et al. However, no obvious histopathologic change was observed in the three CO dosage groups. These data show that 10 ml/kg CO , that dosage has been determined as the vehicle in drug safety assessment , can cause obvious influence on gene expression in rat thymus. Our study suggest that the dosage of CO gavage as the vehicle for water-in-soluble agents in drug development should be no more than 5 ml/kg if agents’ molecular effects in thymus want to be assessed. Gene expression in thymus from female Wistar rats daily administered with 2, 5, 10 ml/kg of corn oil or 10 ml/kg of saline by gavage for 14 consecutive days were measured using Agilent Rat Whole Genome 8×*60K array.
Project description:Microbiota from rats fed with wheat aleurone and plant omega fatty acids In this study we investigated how an AX-rich WA and ALA from linseed oil (LO) modulate the gut microbiota of rats. Wistar rats were fed a standard diet and received either an iso-energetic control oil (PO), control oil + aleurone (A+PO), linseed oil (LO) or linseed oil + aleurone (A+LO) during 12 weeks. Feacal samples were recovered after the 12 week treatments. DNA extractions were performed using using the Qiagen's DNA Stool Kit (Qiagen, West Sussex, UK). 10ng of DNA template were amplified by PCR (16S gene) and purified using Qiagen's Qiaquick PCR purification kit (Qiagen, West Sussex, UK). 1ug of purified PCR product were labelled with either Cy3 or Cy5 using Genomic DNA ULS Labelling kit (Agilent Technologies, Palo Alto, CA). 250ng of labelled DNA were hybridized on the microarray for 24h at 65°C. Washings were performed as recommended by the manufacturer. Microarray scanning was performed on a Surescan Microarray scanner (Agilent Technologies, Palo Alto, CA). Data were extracted using the Feature extraction software (Agilent Technologies, Palo Alto, CA). The retained intensity value for each probe was the ratio between the spot’s median intensity signals and the median of background signals. A 13 chip study was realized to analyze the feacal microbiota of rats treated with either an iso-energetic control oil (PO), control oil + aleurone (A+PO), linseed oil (LO) or linseed oil + aleurone (A+LO) during 12 weeks. Each microarray corresponding to hybridization with 250ng of labelled 16S rRNA gene amplicons from 2 rat DNA faecal samples. Microbiota structure and diversity were assessed using the HuGChip (Tottey et al., 2013). Each probe (4441) was synthetized in three replicates. On the same array, 2 different samples were hybridized. One labelled with the Cy3 dye and one with the Cy5 dye. The results were processed as single channel (13 raw data files available on Series records for 25 samples).
Project description:We have demonstrated that fish oil/pectin (FO/P) diets protect against colon cancer compared to corn oil/cellulose (CO/C) by upregulating apoptosis and suppressing proliferation. To elucidate the mechanisms whereby FO/P diets induce apoptosis and suppress proliferation during the tumorigenic process, we analyzed the temporal gene expression profiles from exfoliated rat colonocytes. KEYWORDS: Fish oil/pectin, pectin, exfoliated colonocytes, gene expression, apoptosis, colon cancer, chemoprevention Rats consumed diets containing FO/P or CO/C and were injected with azoxymethane (AOM, 2x, 15 mg/kg BW, s.c.). Feces collected at the initiation, aberrant crypt foci (ACF), and tumor stages of colon cancer (24 h (n=20), 7 wk (n=37), and 28 wk (n=28) after AOM injection, respectively) was used for poly A(+) RNA extraction. Gene expression signatures were determined using Codelink arrays.
Project description:Our group has been systematically investigating effects of the neuropeptide pituitary adenylate-cyclase activating polypeptide (PACAP) on the ischemic brain. To do so, we have established and utilizing the permanent middle cerebral artery occlusion (PMCAO) mouse model, where PACAP38 (1 pmol) injection is given intracerebroventrically in comparison to a control saline (0.9% NaCl) injection, in conjunction with high-throughput DNA microarray analysis. In previous studies, we have accumulated a large volume of data (gene inventory) from the whole brain (ipsilateral and contralateral hemispheres) by this approach. In our latest research, we have targeted specifically infract/ischemic core (hereafter IC) and the penumbra (hereafter P) post-PACA38 injection, for re-examining the transcriptome at 6 and 24 h post-treatment. The aim of the current study is simple – to delineate the specificity of expression and localization of differentially expressed molecular factors influenced by PACAP38. Prior to this highly expensive and time-consuming omic analysis, we checked the validity of our hypothesis and experimental strategy wherein, ischemic core and penumbra were carefully sampled and compared to the corresponding contralateral (healthy) IC and P regions at 6 and 24 h, post PACAP38 or saline injections to reveal expected differences in gene and protein expression by traditional reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blot analyses, respectively (Hori et al., 2014). Our present results using the mouse 4x44K whole genome DNA chip reveal numerous changes (≧/≦ 1.5/0.75-fold) at both 6 h (654 and 456, and 522 and 449 up- and down-regulated genes for IC and P, respectively; and 24 h (2568 and 2684, and 1947 and 1592 up- and down-regulated genes for IC and P, respectively) after PACAP38 treatment. Using bioinformatics analysis by pathway- or specific-disease-state focused gene classifications and Ingenuity Pathway Analysis (IPA), these genes were functionally classified and discussed. Taken together, the DNA microarray analysis provides not only a great resource for further study, but also reinforces the importance of region-specific analyses in genome-wide identification of target molecular factors that might play a role in the neuroprotective function of PACAP38. In the present study, we used three mice each in PMCAO groups for PACAP38 and saline injections, respectively, that exhibited neurological grades G1 and G2 for the subsequent downstream analysis. Six or 24 h post-injection of PACAP38 or saline, the mice were removed from their cages, decapitated, and their brains carefully removed on ice. The right (ipsilateral; ischemic) and left (contralateral) hemispheres were dissected, and from each hemisphere the ischemic core and penumbra regions and corresponding healthy core and penumbra were carefully removed with a sterile scalpel, and placed in 2 ml Eppendorf tubes. The samples were then quickly immersed in liquid nitrogen and stored in -80 ºC prior to further analysis. Effect of the intracerebroventricular PACAP38 administration into ischemic mouse brain was evaluated at the molecular level in the ischemic core and penumbra of ipsilateral (right) hemisphere over the saline injection by whole genome DNA microarray analysis (4x44K, Agilent).
Project description:In order to gain insight into the effects of aging on susceptibility to environmental toxins, we characterized the expression of xenobiotic metabolizing enzymes (XMEs) from the livers of male Brown Norway and F344 rats across the adult lifespan. To examine metabolic processes across lifespan after challenge with a xenobiotic compound, Brown Norway rats were exposed to 1.0 g/kg body weight toluene by oral gavage in corn oil (4ml/kg body weight) or corn oil alone. Experiment Overall Design: Brown Norway rats: Analysis of gene expression profiles for XMEs in male Brown Norway rats 4, 12, and 24 months old. Rats were exposed to 1g/kg toluene by oral gavage, and sacrificed after 4 hours. Total RNA was isolated from liver samples and gene expression analyzed using Affymetrix Rat 230 2.0 full-genome GeneChips. Data from 18 samples, with three rats in each of the control age groups and dosed age groups, were analyzed. Experiment Overall Design: F344 rats: Analysis of gene expression profiles for XMEs in male F344 6, 11, 18, and 24 months old. Total RNA was isolated from liver samples and gene expression analyzed using Affymetrix Rat 230 2.0 full-genome GeneChips. Data from 16 samples, with four rats in each of the 4 age groups, were analyzed.