Biomaterial Database

Semi-quantitative analysis of cytokine gene expression in blood and cerebrospinal fluid cells by reverse transcriptase polymerase chain reaction. 1995

P Rieckmann, and M Albrecht, and H Ehrenreich, and T Weber, and U Michel

Department of Psychiatry, University of Göttingen, Germany.

An easy, reproducible and semi-quantitative, non-radioactive method for the analysis of mRNA expression for various cytokines, (i.e., Interleukin (IL)-1 beta, IL-4, IL-6, tumor necrosis factor (TNF)-alpha, lymphotoxin (LT), transforming growth factor (TGF)-beta, interferon (IFN)-gamma and endothelin-1 (ET-1)) in cells from cerebrospinal fluid (CSF) and peripheral blood mononuclear cells (PBMC) has been established. By means of polymerase chain reaction primers that cover a splice junction, amplification of contaminating DNA was omitted. Densitometric scanning of ethidium bromide-stained agarose gels proved to be very sensitive for semiquantitative analysis of PCR products. Serial tenfold dilutions of cDNA revealed a log-linear regression from 10(6) to 10(2) cells under optimal cycle conditions. The intra- and inter-assay variability of the method was below 10%. With this assay, the cytokine expression pattern of as few as 10(4) mononuclear cells from blood or CSF was determined. This method made it possible to detect differences in the cytokine gene expression pattern of mononuclear cells from patients with different neurological diseases. CSF cells from 43 patients with various neurological diseases were analyzed. TNF-alpha, LT, and IL-1 mRNA were prominent in the CSF cells of most patients with bacterial meningitis. TNF-alpha, LT, IFN-gamma and IL-6 mRNAs were detected in patients with active multiple sclerosis, whereas TNF-alpha, IL-6, and endothelin-1 mRNA expression was found frequently in patients with HIV encephalitis. Pro-inflammatory cytokines were rarely detected in CSF cells from patients with non-inflammatory diseases of the central nervous system. In blood mononuclear cells from patients with multiple sclerosis, TNF-alpha mRNA expression was associated with disease activity. The sensitivity, specificity, velocity and reliability of this assay considerably facilitates the analysis of cytokine production in mononuclear cells even in conditions where only a limited number of cells is available for analysis.

UI	MeSH Term	Description	Entries
D008969	Molecular Sequence Data	Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories.	Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D011237	Predictive Value of Tests	In screening and diagnostic tests, the probability that a person with a positive test is a true positive (i.e., has the disease), is referred to as the predictive value of a positive test; whereas, the predictive value of a negative test is the probability that the person with a negative test does not have the disease. Predictive value is related to the sensitivity and specificity of the test.	Negative Predictive Value,Positive Predictive Value,Predictive Value Of Test,Predictive Values Of Tests,Negative Predictive Values,Positive Predictive Values,Predictive Value, Negative,Predictive Value, Positive
D002555	Cerebrospinal Fluid	A watery fluid that is continuously produced in the CHOROID PLEXUS and circulates around the surface of the BRAIN; SPINAL CORD; and in the CEREBRAL VENTRICLES.	Cerebro Spinal Fluid,Cerebro Spinal Fluids,Cerebrospinal Fluids,Fluid, Cerebro Spinal,Fluid, Cerebrospinal,Fluids, Cerebro Spinal,Fluids, Cerebrospinal,Spinal Fluid, Cerebro,Spinal Fluids, Cerebro
D006801	Humans	Members of the species Homo sapiens.	Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D001483	Base Sequence	The sequence of PURINES and PYRIMIDINES in nucleic acids and polynucleotides. It is also called nucleotide sequence.	DNA Sequence,Nucleotide Sequence,RNA Sequence,DNA Sequences,Base Sequences,Nucleotide Sequences,RNA Sequences,Sequence, Base,Sequence, DNA,Sequence, Nucleotide,Sequence, RNA,Sequences, Base,Sequences, DNA,Sequences, Nucleotide,Sequences, RNA
D012333	RNA, Messenger	RNA sequences that serve as templates for protein synthesis. Bacterial mRNAs are generally primary transcripts in that they do not require post-transcriptional processing. Eukaryotic mRNA is synthesized in the nucleus and must be exported to the cytoplasm for translation. Most eukaryotic mRNAs have a sequence of polyadenylic acid at the 3' end, referred to as the poly(A) tail. The function of this tail is not known for certain, but it may play a role in the export of mature mRNA from the nucleus as well as in helping stabilize some mRNA molecules by retarding their degradation in the cytoplasm.	Messenger RNA,Messenger RNA, Polyadenylated,Poly(A) Tail,Poly(A)+ RNA,Poly(A)+ mRNA,RNA, Messenger, Polyadenylated,RNA, Polyadenylated,mRNA,mRNA, Non-Polyadenylated,mRNA, Polyadenylated,Non-Polyadenylated mRNA,Poly(A) RNA,Polyadenylated mRNA,Non Polyadenylated mRNA,Polyadenylated Messenger RNA,Polyadenylated RNA,RNA, Polyadenylated Messenger,mRNA, Non Polyadenylated
D014409	Tumor Necrosis Factor-alpha	Serum glycoprotein produced by activated MACROPHAGES and other mammalian MONONUCLEAR LEUKOCYTES. It has necrotizing activity against tumor cell lines and increases ability to reject tumor transplants. Also known as TNF-alpha, it is only 30% homologous to TNF-beta (LYMPHOTOXIN), but they share TNF RECEPTORS.	Cachectin,TNF-alpha,Tumor Necrosis Factor Ligand Superfamily Member 2,Cachectin-Tumor Necrosis Factor,TNF Superfamily, Member 2,TNFalpha,Tumor Necrosis Factor,Cachectin Tumor Necrosis Factor,Tumor Necrosis Factor alpha
D015870	Gene Expression	The phenotypic manifestation of a gene or genes by the processes of GENETIC TRANSCRIPTION and GENETIC TRANSLATION.	Expression, Gene,Expressions, Gene,Gene Expressions
D016133	Polymerase Chain Reaction	In vitro method for producing large amounts of specific DNA or RNA fragments of defined length and sequence from small amounts of short oligonucleotide flanking sequences (primers). The essential steps include thermal denaturation of the double-stranded target molecules, annealing of the primers to their complementary sequences, and extension of the annealed primers by enzymatic synthesis with DNA polymerase. The reaction is efficient, specific, and extremely sensitive. Uses for the reaction include disease diagnosis, detection of difficult-to-isolate pathogens, mutation analysis, genetic testing, DNA sequencing, and analyzing evolutionary relationships.	Anchored PCR,Inverse PCR,Nested PCR,PCR,Anchored Polymerase Chain Reaction,Inverse Polymerase Chain Reaction,Nested Polymerase Chain Reaction,PCR, Anchored,PCR, Inverse,PCR, Nested,Polymerase Chain Reactions,Reaction, Polymerase Chain,Reactions, Polymerase Chain
D016207	Cytokines	Non-antibody proteins secreted by inflammatory leukocytes and some non-leukocytic cells, that act as intercellular mediators. They differ from classical hormones in that they are produced by a number of tissue or cell types rather than by specialized glands. They generally act locally in a paracrine or autocrine rather than endocrine manner.	Cytokine