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A gene coding for ornithine decarboxylase (odcA) is differentially expressed during the Mucor circinelloides yeast-to-hypha transition.

The differential display technique was used to identify genes from Mucor circinelloides involved in the yeast-to-hypha transition. Using a limited set of primer combinations, cDNA fragments corresponding to mRNAs differentially expressed during the dimorphic transition were isolated. Northern analyses showed that the accumulation of the transcript detected by hybridisation with one of the cDNA fragments increased during the transition and was undetectable at the mycelial stage. Sequence analysis and database searches of this fragment revealed high similarity to ornithine decarboxylase (ODC) encoding genes. The odcA gene of M. circinelloides was isolated from genomic and cDNA libraries and characterised. Electrophoretic karyotyping and hybridisations showed that the odcA gene is single-copy and linked to the leuA and rDNA genes. The single transcript detected (2.1 kb), was considerably longer than the deduced ORF. Through non-radioactive primer extension analysis four transcription initiation sites were mapped to positions -61, -167, -239 and -436 from the start codon. The ODC mRNA levels increased during the yeast-to-hypha transition, reaching a maximum at 120 min, which was accompanied by a rise in ODC enzymatic activity. The expression pattern of the odcA gene showed that in M. circinelloides the ODC levels are transcriptionally regulated, in contrast with other dimorphic fungi in which a post-transcriptional regulation has been proposed.

5' Untranslated Regions↗

Identification of differentially expressed genes associated with androgen-independent growth of prostate cancer.

BACKGROUND: The human prostate cancer xenograft, CWR22, similar to most human prostate cancers, regresses after castration and recurs several months after the removal of androgen. Genes uniquely associated with proliferation were identified by comparison of tumors that exist in androgen absence but differ in proliferative capacity. METHODS: cDNA libraries from CWR22 tumors from 20-day castrate mice (proliferation undetectable) and recurrent CWR22 tumors (proliferation rate similar to androgen-dependent CWR22) were compared to evaluate the possibility that proliferation is triggered by either gain of function or loss of suppression. Differentially expressed genes were evaluated further for their temporal association with the onset of cellular proliferation using northern and western analysis and immunohistochemistry of a series of CWR22 tumors that spanned the transition from androgen-dependent to recurrent growth. RESULTS: Subtractive hybridization identified 11 candidate genes from among 1,057 clones examined. Northern analysis confirmed differential expression of 8 genes. Western analysis revealed an association between tomoregulin, translation elongation factor-1 alpha (EF-1 alpha), Mxi-1, and thioredoxin-binding protein 2/vitamin D up-regulated protein, and the onset of recurrent growth. Immunohistochemistry revealed expression of tomoregulin, EF-1 alpha, Mxi-1, and thioredoxin reductase-1 coincidental with the onset of cellular proliferation on day 120 after castration. CONCLUSIONS: One or more of these genes may represent an appropriate target to prevent, delay or treat recurrent prostate cancer.

Androgens↗

Downregulation of genes involved in DNA repair and differential expression of transcription regulators and phosphatases precede IgM-induced apoptosis in the Burkitt's lymphoma cell line BL60-2.

Apoptosis of lymphocytes recognizing self-antigens is an essential mechanism to protect the organism against autoimmune diseases. Programmed cell death of susceptible B cells occurs in response to surface IgM cross-linking mediated by self-antigens. This effect can be mimicked in the Burkitt's lymphoma line BL60-2 by addition of anti-IgM antibodies. In order to identify genes with differential expression in response to the apoptotic stimulus, total RNA prepared from BL60-2 cells before and at different points in time after IgM cross-linking was used for Atlas arrays, high-density oligonucleotide microarrays (GeneChip arrays, Affymetrix) and in RNase protection assays (RPA). One of our major observations was the downregulation of six genes involved in the ligation of DNA strand breaks, like DNA ligases and DNA-PK, indicating a shutdown of DNA repair mechanisms in apoptotic cells. In addition, we found changes on mRNA level for several transcription regulators, including early growth response genes 1 and 2, TAFII30 and topoisomerase I. Furthermore, we show accumulation of mRNA for the phosphatases CD45 and DUSP5 in anti-IgM stimulated BL60-2 cells. Our data provide a basis for further analysis of the differentially expressed genes and their roles in IgM-induced B cell death as well as in apoptosis in other cellular systems.

Apoptosis↗

Effects of imidacloprid metabolites on habituation in honeybees suggest the existence of two subtypes of nicotinic receptors differentially expressed during adult development.

Habituation of the proboscis extension reflex (PER) in honeybees (Apis mellifera) is age-dependent. Very young bees (< or =7 days old) require significantly less trials to abolish the response to multiple sucrose stimulations than older bees (> or =8 days old). A nicotinic agonist, imidacloprid, modifies this behaviour by increasing the number of trials in < or =7-day-old bees and by decreasing it in older bees [Neurobiol. Learn. Mem. 76 (2001) 183.]. Here we tested our hypothesis that this effect is associated with a differential expression of two subtypes of nicotinic acetylcholine receptors (nAChRs). By testing the effects of six metabolites of imidacloprid, we show that two of them, olefin and 5-hydroxy-imidacloprid, modify the number of trials needed to habituate the PER in a contrasting manner. Olefin increases the number of trials in both age groups, whereas 5-hydroxy-imidacloprid decreases the number of trials, but only in 8-day-old individuals. We conclude that olefin and 5-hydroxy-imidacloprid are specific agonists of two subtypes of an nAChR that are differentially expressed during adult maturation of young honeybees. Olefin is the agonist of an nAChR expressed in both age groups, whereas 5-hydroxy-imidacloprid is the agonist of a late-onset nAChR that is activated in 8-day-old bees. The implications of this finding for the honeybee biology are discussed.

Aging↗

Revealing differential expression patterns of piRNA in FACS blood cells of SARS-CoV-2 infected patients.

Non-coding RNA expression has shown to have cell type-specificity. The regulatory characteristics of these molecules are impacted by changes in their expression levels. We performed next-generation sequencing and examined small RNA-seq data obtained from 6 different types of blood cells separated by fluorescence-activated cell sorting of severe COVID-19 patients and healthy control donors. In addition to examining the behavior of piRNA in the blood cells of severe SARS-CoV-2 infected patients, our aim was to present a distinct piRNA differential expression portrait for each separate cell type. We observed that depending on the type of cell, different sorted control cells (erythrocytes, monocytes, lymphocytes, eosinophils, basophils, and neutrophils) have altering piRNA expression patterns. After analyzing the expression of piRNAs in each set of sorted cells from patients with severe COVID-19, we observed 3 significantly elevated piRNAs - piR-33,123, piR-34,765, piR-43,768 and 9 downregulated piRNAs in erythrocytes. In lymphocytes, all 19 piRNAs were upregulated. Monocytes were presented with a larger amount of statistically significant piRNA, 5 upregulated (piR-49039 piR-31623, piR-37213, piR-44721, piR-44720) and 35 downregulated. It has been previously shown that piR-31,623 has been associated with respiratory syncytial virus infection, and taking in account the major role of piRNA in transposon silencing, we presume that the differential expression patterns which we observed could be a signal of indirect antiviral activity or a specific antiviral cell state. Additionally, in lymphocytes, all 19 piRNAs were upregulated.

Humans↗

ORP-3, a human oxysterol-binding protein gene differentially expressed in hematopoietic cells.

Using differential display polymerase chain reaction, a gene was identified in CD34(+)-enriched populations that had with low or absent expression in CD34(-) populations. The full coding sequence of this transcript was obtained, and the predicted protein has a high degree of homology to oxysterol-binding protein. This gene has been designated OSBP-related protein 3 (ORP-3). Expression of ORP-3 was found to be 3- to 4-fold higher in CD34(+) cells than in CD34(-) cells. Additionally, expression of this gene was 2-fold higher in the more primitive subfraction of hematopoietic cells defined by the CD34(+)38(-) phenotype and was down-regulated with the proliferation and differentiation of CD34(+) cells. The ORP-3 predicted protein contains an oxysterol-binding domain. Well-characterized proteins expressing this domain bind oxysterols in a dose-dependent fashion. Biologic activities of oxysterols include inhibition of cholesterol biosynthesis and cell proliferation in a variety of cell types, among them hematopoietic cells. Characterization and differential expression of ORP-3 implicates a possible role in the mediation of oxysterol effects on hematopoiesis.

Antigens, CD34↗

Cloning and identification of genes differentially expressed in metastatic and non-metastatic rat rhabdomyosarcoma cell lines.

The objective of this study was to identify genes involved in invasion and metastasis using a rat rhabdomyosarcoma model (SMF-A and RMS-B cell lines). The SMF-A cell line was established from a metastatic nodule of an induced rhabdomyosarcoma in syngeneic F344 rats. Two cell lines with defined metastatic potentials, SMF-Ai and SMF-Da, were cloned from the SMF-A line. The cell line SMF-Ai is tumorigenic, highly invasive and highly metastatic. On the other hand, the revertant line SMF-Da is less tumorigenic, non-invasive and non-metastatic. We have isolated from a SMF-Ai cDNA library eight cDNA clones which are differentially expressed by the metastatic SMF-Ai and the non-metastatic SMF-Da cell line using Northern blot analysis. Five of these clones, smf-4, smf-6, smf-41, smf-42 and smf-44, are overexpressed in the SMF-Da cell line and have homology with beta-2-microglobulin, lactate dehydrogenase, ribosomal protein L38, ribosomal protein S4 and acidic ribosomal phosphoprotein P1, respectively. The three other clones, smf-7, smf-40 and smf-61, are overexpressed in SMF-Ai. Clones smf-40 and smf-61 show significant homology with the human TB3-1 gene and the human fus gene respectively. The clone smf-7 has no significant homology with known sequences. We also analyzed the expression of these clones in other rat rhabdomyosarcoma cell lines (RMS-B and their clones) and in tumors obtained by injection of these cell lines into rats or nude mice. Smf-61 and smf-7 were the only clones with a differential expression pattern associated with the invasive or metastatic potential of all cell lines examined. A preliminary study of the expression of smf-7 and smf-61 in other cancer cell lines also showed mRNA expression in two human rhabdomyosarcomas and a human epidermoid carcinoma suggesting the existence of genes homologous to smf-7 and smf-61 clones in human cancers. Our findings suggest an association between the expression of smf-7 and smf-61 and invasive or metastatic potential of rhabdomyosarcoma cells.

Animals↗

Identification of rat Gas7 isoforms differentially expressed in brain and regulated following kainate-induced neuronal injury.

The growth arrest-specific gene 7 (Gas7) is expressed primarily in the brain and is necessary for the formation of neurite in cultured cerebellar preneurons. The endogenous rat Gas7 (rGas7) is transiently elevated before nerve growth factor-promoted neurite outgrowths emerge in cultured PC12 cells. We report three Gas7 isoforms (a, b, and c) in rat tissues. Peptide microsequencing identified two Gas7 forms, rGas7-a (38 kDa) and rGas7-b (47 kDa). rGas7-c can be predicted from a transcription variant by alternative splicing. Although two open reading frames were predicted, a cloned rGas7 cDNA encoded mostly rGas7-a in mammalian cells. The overexpression of the rGas7 cDNA in PC12 cells sufficed to promote small lamellipodia- and filopodia-like cell processes that resemble the initial stages of neurite formation. Three rGas7 isoforms were differentially expressed in all of the brain subregions. Only rGas7-a was detected in rat cerebellum, as in mouse cerebellum. Kainate injury did not affect the level of rGas7-b, but the level of isoform c was substantially suppressed in the hippocampus. Immunohistochemistry reveals that Gas7 was expressed primarily in the pyramidal neurons of the hippocampus and was quickly attenuated before recovery in the CA3 area after kainate was administered. These results suggest that differential expression and unique regulation of Gas7 isoforms in brain subregions may be important in specialized brain functions. Conservation of Gas7 isoforms by alternative splicing in mammals is also considered.

Amino Acid Sequence↗

Localization and differential expression of FMRFamide-like immunoreactivity in the nematode Ascaris suum.

By immunocytochemical and immunohistochemical methods, FMRFamide-like immunoreactivity (FLI) was localized to many neurons and processes in the Ascaris nervous system, including the head, tail, and lateral lines. Some of these cells were identified; they included sensory neurons, interneurons, and motor neurons. FLI was also present in the pharyngeal neurons and in their varicosities near the surface of the pharynx. By HPLC analysis of extracts, only a subset of the FMRFamide-like peptides (FLPs) expressed in Ascaris heads, and heads from which the pharynx had been removed, were expressed in the pharynx. Furthermore, FLPs appeared to be differentially expressed in female heads and tails and male heads and tails. Acetone and acid methanol differentially extracted subforms of FLI from Ascaris heads and from C. elegans.

Animals↗

Differential expression of hepatic sterol carrier proteins in the streptozotocin-treated diabetic rat.

Sterol carrier protein-2 (SCP2) is a 13.2-kilodalton protein that has been implicated in intracellular cholesterol transport, whereas a related sterol carrier protein, sterol carrier protein-X (SCPx; 58 kilodaltons) has been suggested to function also in the beta-oxidation of fatty acids. Although diabetes-related hyperlipidemia and altered cholesterol metabolism have been extensively studied, the intracellular cholesterol transport capacity during hyperglycemic states has not been examined. The fact that beta-oxidation is increased in diabetes whereas hepatic cholesterol metabolism is reduced suggests that differential expression of these sterol carrier proteins may accompany diabetic dyslipidemia. In this study, SCP2 protein levels were reduced by 60% in mildly hypercholesterolemic (cholesterol, > 130 and < 150 mg/dl; P < 0.01) diabetic rats and by 90% in severely hypercholesterolemic (cholesterol, > 150 mg/dl; P < 0.002) diabetic animals. In contrast, hepatic SCPx protein expression increased (3.5-fold) after diabetes induction with streptozotocin (STZ). The decline in SCP2 was inversely related to serum cholesterol levels. Hepatic SCP messenger RNA levels examined by ribonuclease protection assay demonstrated that hepatic SCP messenger RNA was increased 2-fold in diabetic animals. Northern blot analysis indicated that both the 0.8-kilobase SCP2-specific and the 2.1-kilobase SCPx-specific transcripts increased after STZ injection. SCPx protein induction preceded the decline in SCP2 by 4-5 days. Insulin treatment reversed the increase in SCPx and prevented the decline in SCP2. We conclude that SCP2 and SCPx are differentially expressed in the STZ-diabetic rat and suggest that this change in SCP expression should be considered a potential contributing mechanism through which cholesterol metabolism may be altered in diabetes.

Acetyl-CoA C-Acetyltransferase↗

Differential expression of genes involved in skin homing, proliferation, and apoptosis in CD4+ T cells of patients with atopic dermatitis.

CD4+ T cells play a critical role in allergic diseases, both in the affected tissue as well as systemically. Our objective was to investigate the in vivo activation state of peripheral blood CD4+ T cells of atopic dermatitis (AD) patients by analyzing gene expression profiles of unstimulated CD4+ T cells. mRNA samples from blood CD4+ T cells, isolated from five AD patients and seven healthy controls (HC), were analyzed using oligonucleotide arrays. Differentially regulated genes were validated by quantitative PCR (Q-PCR) in a larger group of patients with AD, in a group of patients with allergic asthma (AA), and HC subjects. In addition, "typical" T helper type 1 (Th1)- and Th2-related genes were analyzed by Q-PCR. Microarray analysis revealed differential expression of 52 genes in AD patients. Q-PCR confirmed several differentially regulated genes in AD, including CCR10, CRTH2, C-JUN, and NR4A2. Two groups of genes with highly correlating gene expression levels involved in tissue homing and proliferation or apoptosis, respectively, were identified. No marked differences were found in gene expression levels of typical Th1 or Th2 genes in AD or in AA patients. This study demonstrates that peripheral blood, unstimulated CD4+ T cells in AD patients show differentially expressed genes involved in tissue homing, proliferation, and apoptosis. No marked expression differences of "typical" atopy genes were found.

Adult↗

The Isolation of differentially expressed mRNA sequences by selective amplification via biotin and restriction-mediated enrichment.

Molecular analysis of development frequently implies the isolation and characterization of genes with specific spatial and temporal expression patterns. Several methods have been developed to identify such DNA sequences. The most comprehensive technique involves the genomewide probing of DNA sequence microarrays with mRNA sequences. However, at present this technology is limited to the few organisms for which the entire genome has been sequenced. Here, we describe a subtractive hybridization technique, called selective amplification via biotin and restriction-mediated enrichment (SABRE), which allows the selective amplification of cDNA fragments representing differentially expressed mRNA species. The method involves the competitive hybridization of an excess of driver cDNA fragments (D) to a trace of tester cDNA fragments (T), and the subsequent purification of tester homohybrids (in which both strands are contributed by the tester cDNA). After competitive hybridization, cDNA fragments that are more abundant in the tester than in the driver are enriched in the tester homohybrids. However, as the fraction of tester homohybrids is very small [T(2)/(D + T)(2)], their purification requires highly efficient procedures. In SABRE, the isolation of tester homohybrids is afforded by a combination of three successive steps: removal of biotinylated terminal sequences from most of the heterohybrids by S1 nuclease digestion, capture of biotinylated hybrids with streptavidin-coated paramagnetic beads, and specific release of homohybrids from the beads by restriction nuclease digestion. If several rounds of SABRE selection are performed in series, even relatively rare differentially expressed mRNA sequences may result in the production of predominant cDNA fragments in the final tester homohybrid population.

Animals↗

Identification of differentially expressed genes in rat aortic allograft vasculopathy.

Graft vasculopathy is an important complication of long-surviving organ transplants, but its pathogenesis has remained elusive. We investigated rat aortic transplants with vasculopathy, aortic transplants without vasculopathy, and normal aortas for differentially expressed mRNA transcripts to gain further insight into the molecular mechanisms involved. Aortic transplants were performed in allogeneic or syngeneic recipients followed by removal after 1 or 5 months, RNA isolation, and differential display to identify mRNA transcripts the expression of which was modulated in conjunction with the transplant procedure and the development of vasculopathy. Using 80 random primers, 57 differentially displayed polymerase chain reaction products were identified, 18 of which were found in allografts but not in syngeneic grafts or normal vessels, whereas 15 were expressed in normal vessels and syngeneic grafts but not in allografts. Of the differentially displayed amplicons, 13 were successfully reamplified and used as probes for Northern analysis; differential expression was confirmed in 6 instances. DNA sequence analysis of these PCR products revealed identity with the immunoglobulin J chain in 2 instances, the ferritin heavy chain, a sequence related but not identical with Ras, and an established sequence tag recently isolated from a human fetal heart library; 1 sequence was not related to any known gene. To assess whether differential mRNA expression of the J-chain gene, a gene expressed in cells of B lymphocyte lineage, was associated with infiltration of the graft by B lymphocytes, tissue sections were stained with an antibody against the B cell marker CD45RA. Although the number of CD45RA-positive cells was low, there was a significant increase in the number of CD45RA-positive cells in the adventitia and intima of grafts with vasculopathy. Furthermore, immunostaining with anti-ferritin antiserum confirmed the presence of ferritin-positive cells within the inner layer of the graft vessel wall and dispersed in the intima, media, and adventitia. The question remains as to which of these genes are critically relevant in the pathogenesis of graft vasculopathy and whether they serve as targets for therapeutic interventions.

Animals↗

Cloning of the genes for human stromelysin and stromelysin 2: differential expression in rheumatoid synovial fibroblasts.

Stromelysin is a member of a gene family of metalloproteinases involved in extracellular matrix remodeling in normal and diseased processes. Primary cultures of rheumatoid synovial cells produce large amounts of metalloproteinase mRNA and proteins. We cloned a cDNA for human stromelysin from a rheumatoid synovial cell cDNA library, and we used the cDNA to isolate the gene for human stromelysin and a related gene, stromelysin 2. We sequenced parts of the genes and found that both are contained on approximately 14 kilobase pairs of DNA. Using an exon-containing fragment of the stromelysin 2 genomic clone as a specific probe in Northern blot analysis, we demonstrate the differential expression of stromelysin and stromelysin 2 in rheumatoid synovial cells, human foreskin fibroblasts, and rabbit synovial fibroblasts. In addition, using chimeric constructs of the stromelysin promoter linked to the bacterial gene chloramphenicol acetyltransferase (CAT), we show that the elements required for the tumor promoter phorbol myristate acetate (PMA), epidermal growth factor (EGF), and interleukin 1 beta (IL-1 beta) induction are contained on a 307 base pair fragment which includes approximately 270 base pairs (bp) of 5'-flanking DNA. The cloning of the human stromelysin and stromelysin 2 genes, the documentation of their differential expression, and the identification of transcriptional regulatory regions in the stromelysin gene will facilitate the study of metalloproteinase gene expression in normal processes and in diseases such as rheumatoid arthritis.

Animals↗

Differential expression of the melatonin receptor in human monocytes.

Earlier studies have shown that the pineal hormone melatonin activates human monocytes. It is reported here that melatonin induces the secretion of IL-1, IL-6, and TNF in fresh and 1-day in vitro cultured monocytes that also express the melatonin receptor (Kd = 270 +/- 60 pM; 42,000-48,000 receptors/cell). However, when monocytes were cultured in vitro for 2 days, the number of receptors decreased to 11,000 receptors/cell, with the same Kd. LPS activation of fresh or 1-day cultured monocytes did not result in any increase in melatonin receptor number. LPS activation of 2-day cultured monocytes led to an increase in the number of melatonin receptors, from 11,000 receptors/cell to the plateau of 42,000 to 48,000 receptors/cell. The loss of receptors by 2-day cultured monocytes was not irreversible. Melatonin did not induce the release of IL-1, TNF, or IL-6 in monocytes cultured in vitro for 3 days and for up to 15 days, and these long time cultured monocytes did not express the melatonin receptors even after activation by LPS. The loss of melatonin receptors by monocytes cultured in vitro for 3 days and for up to 15 days was irreversible. Therefore, it is shown for the first time that human monocytes express melatonin receptors. Furthermore, human monocytes express melatonin receptors differentially depending on their state of maturation, and it appears that in vitro monocyte differentiation and maturation negatively affect human monocyte melatonin receptor expression.

Cell Differentiation↗

Differential expression of low Mr GTP-binding proteins in human megakaryoblastic leukemia cell line, MEG-01, and their possible involvement in the differentiation process.

The expression of various low Mr GTP-binding proteins at various states of differentiation of a human megakaryoblastic leukemia cell line, MEG-01, was analyzed using thermocycle amplification of mRNA and immunoblotting. MEG-01 cells were found to express mRNAs of rap1A, rap1B, rap2B, ralA, rhoA, rac1, rac2, CDC42Hs, rab1, rab3B, rab6, ram and ran, but not rab4, and the proteins of Rap1, Rap2, RhoA, Rac1, Rac2, Rab3B, Rab4, Rab6 and Rab8 were expressed. Differentiation of MEG-01 cells induced by 100 nM 12-O-tetradecanoylphorbol-13-acetate revealed the considerable increases in mRNA expression of rap1B, rab3B, rab4, ram and ran whereas the levels of rap2B, rhoA and rac1 decreased. During the differentiation process, significant changes in protein levels of Rap1, RhoA, Rac1, Rac2, Rab3B, Rab4 and Rab6 were observed among three subcellular (cytosol, Triton X-100-soluble membrane and -insoluble cytoskeleton) fractions. The present investigation may be useful for the study of the megakaryocyte differentiation.

Blood Platelets↗

Effect of rosiglitazone on the differential expression of obesity and insulin resistance associated proteins in lep/lep mice.

Peroxisome proliferator-activated receptors (PPARs) participate in the molecular mechanism of pathologies with altered lipid homeostasis such as type 2 diabetes or obesity. The insulin sensitizer drug, rosiglitazone, has been shown to bind and activate PPAR-gamma1 in adipocytes and PPAR-gamma2 in hepatocytes. The identification of new molecular targets associated with fatty acid oxidation and PPAR-gamma nuclear receptor regulation in insulin resistance tissues is a key research goal. In the present study, we have used a proteomic approach to identify such targets. Lean and obese C57 Bl/6J lep/lep mice were given BRL49653, rosiglitazone, 10 mg/kg diet, by dietary admixture for 7 days. Rosiglitazone normalized the impaired glucose tolerance and dyslipidemia in lep/lep mice but had no significant effect in the lean mice. Samples of liver, white and brown adipose tissue, and muscle proteins were obtained and 100 microg of proteins was arrayed by two-dimensional gel electrophoresis. Thirty-four polypeptides were differentially expressed (p < 0.05) between lep/lep and lean mice and eleven were significantly (p < 0.05) modulated by rosiglitazone treatment of the obese mice. None of the proteins was modulated by rosiglitazone treatment of the lean mice. The identity of these differentially expressed proteins was made using tandem mass spectrometric analysis and revealed components of fatty acid and carbohydrate metabolism as well as proteins with unknown function.

Animals↗

[Preparation of the cDNA microarray on the differential expressed cDNA of senescence-accelerated mouse's hippocampus].

Alzheimer' s disease (AD) is the most common form of dementia in the elderly. AD is an invariably fatal neurodegenerative disorder with no effective treatment. Senescence-accelerated mouse prone 8 (SAMP8) is a model for studying age-related cognitive impairments and also is a good model to study brain aging and one of mouse model of AD. The technique of cDNA microarray can monitor the expression levels of thousands of genes simultaneously and can be used to study AD with the character of multi-mechanism, multi-targets and multi-pathway. In order to disclose the mechanism of AD and find the drug targets of AD, cDNA microarray containing 3136 cDNAs amplified from the suppression subtracted cDNA library of hippocampus of SAMP8 and SAMR1 was prepared with 16 blocks and 14 x 14 pins, the housekeeping gene beta-actin and G3PDH as inner conference. The background of this microarray was low and unanimous, and dots divided evenly. The conditions of hybridization and washing were optimized during the hybridization of probe and target molecule. After the data of hybridization analysis, the differential expressed cDNAs were sequenced and analyzed by the bioinformatics, and some of genes were quantified by the real time RT-PCR and the reliability of this cDNA microarray were validated. This cDNA microarray may be the good means to select the differential expressed genes and disclose the molecular mechanism of SAMP8's brain aging and AD.

Aging↗