Interleukin-10(IL-10) inhibits production of a wide range of cytokines in various cell types and transcriptionally inhibits lipopolysaccharide(LPS)-induced expression of proinflammatory mediators. Cytokine expression by macrophages is an important aspect to ochestrate inflammatory responses. As an approach to identify mechanistic targets of IL-10, it was examined the time course for expression of KC(murine homologue of Gro) gene in murine peritoneal macrophages stimulated with LPS with or without IL-10. The effect of IL-10 on LPS induced KC mRNA expression was delayed and only seen after 1 hour treatment. Pretreatment with IL-10 did not eliminate the delayed inhibitory response nor increase the magnitude of suppression. These effects did not depend upon time of IL-10 treatment but the time of LPS treatment. LPS-induced KC mRNA expression by inhibitoy action of IL-10 was not controlled at the level of transcription. The result indicates that IL-10 acts late in the process of KC gene expression and that the prominant site of action may be mRNA stability or translation.
Animals ; Cytokines ; Gene Expression* ; Interleukin-10* ; Macrophages ; Macrophages, Peritoneal* ; Mice* ; RNA Stability ; RNA, Messenger

Previously, we reported that DJ-1, encoded by a Parkinson's disease (PD)-associated gene, inhibits expression of proinflammatory mediators in interferon-gamma (IFN-gamma)-treated astrocytes and microglia through inhibition of STAT1 activation. Here, using microglia and astrocytes cultured from wild-type (WT) and DJ-1-knockout (KO) mouse brains, we examined how DJ-1 regulates suppressor of cytokine signaling 1 (SOCS1), a negative feedback regulator of STAT1 (signal transducer and activator of transcription) that is also induced by STAT1. We found that IFN-gamma significantly increased SOCS1 mRNA expression in WT microglia and astrocytes, but not in KO cells, although STAT1 was highly activated in these latter cells. We further found that SOCS mRNA stability was decreased in DJ-1-KO cells, an effect that appeared to be mediated by the microRNA, miR-155. IFN-gamma increased the levels of miR-155 in DJ-1-KO cells but not in WT cells. In addition, an miR-155 inhibitor rescued SOCS1 expression and decreased STAT1 activation in DJ-1-KO cells. Taken together, these results suggest that DJ-1 efficiently regulates inflammation by maintaining SOCS1 expression through regulation of miR-155 levels, even under conditions in which STAT1 activation is decreased.
Animals ; Astrocytes* ; Brain ; Inflammation ; Interferon-gamma ; Mice ; Microglia* ; MicroRNAs ; Parkinson Disease ; RNA Stability ; RNA, Messenger ; Transducers

To investigate the method of RNA isolation from human embryonic tissues and the factors influencing the quality of RNA, the RNA from human embryonic tissues obtained with drug-induced labor or non-drug induced labor were isolated by using grind with liquid nitrogen or homogenizer without liquid nitrogen. The results showed that the positive rates of RNA integrity in grind with liquid nitrogen group and in homogenizer without liquid nitrogen group were 68.42% and 29.79% respectively, and there was significant difference between these two groups; however, there was no statistic difference in positive rate of RNA integrity, OD(260)/OD(280) ratio and beta-actin gene expression level between the drug-induced labor group and non-drug induced labor group. It is concluded that pulverize of tissue in liquid nitrogen remains the integrity of RNA isolated and may be applied for RNA isolation from human embryonic tissues. The quality of RNA is not affected by different methods of induction of maternal labor.
Embryo, Mammalian ; metabolism ; Freezing ; Humans ; Nitrogen ; pharmacology ; RNA ; isolation & purification ; RNA Stability ; drug effects

Ribonucleic acid (RNA) has potential use in forensic science for the determination of postmortem interval. We report the first study on serial sampling of messenger RNA (mRNA) from surgical specimens to determine if there is a correlation between mRNA quantity and elapsed time. Skin tissues were collected from modified radical mastectomy specimens. After a defined period of time, bisected skin sections were cut and frozen in liquid nitrogen. Serial collection of the specimens was conducted, and frozen sections were obtained from all samples. Quantitative real-time reverse transcription-polymerase chain reaction was performed using the extracted RNA to measure the transcriptional activity of 2 selected housekeeping genes. The selected loci were mRNA sequences that exhibited time-dependent quantitative changes in a previous study. We collected 44 samples from 9 different patients, with 3-10 samples collected per patient. The amount of mRNA transcripts present in the serial samples showed a weak time-dependent correlation trend only in some cases. Further studies to evaluate different target mRNA sequences are necessary, as is exploration of additional methods to evaluate mRNA transcript degradation.
Forensic Sciences ; Frozen Sections ; Genes, Essential ; Humans ; Mastectomy, Modified Radical ; Nitrogen ; RNA ; RNA Stability ; RNA, Messenger ; Skin

The effects of bear bile on the expression of the p53 protein in cancer cell lines were investigated by Western blot and RT-PCR methods. The p53 protein expression was increased after addition of bear bile in HaCaT, KUMA3, KUMA4 and KUMA6 cell lines that carried mutations n the p53 gene, but it is not ncreased n HCT116 and KUMA5 cell lines with the normal p53 gene as determined by Western blot method. The level of p53 mRNA was not altered in all cell lines after the treatment of bear bile as determined RT-PCR. These results indicate that the increment of the p53 protein after addition of bear bile was caused by increase in the protein stability but not by increase in protein synthesis. It can be concluded that bear bile affects positively only in the cell lines which have genetic variation of the p53 gene, and does not affect in the cell lines which have wild-type p53 gene, on the other hand.
Bile* ; Blotting, Western ; Cell Line* ; Genes, p53 ; Genetic Variation ; Hand ; Protein Stability ; RNA, Messenger

Skin atrophy is one of the most common side effects of topical steroid use and results from decreased skin proalphaI collagen mRNA production due to specific alterations in collagen gene transcription or from reduced collagen mRNA stability. This report describes a case of a 8 year-old-girl who visited our hospital with a complaint of ambiguous genitalia. She was observed to have atrophic labia majora and atrophic striae resembling scrotal rugae. However, her serum levels of FSH, LH, 17-KS, 17-OHCS, 5-DHT, DHT and DHEA-S were all in normal range and her karyotype was 46,XX. Abdominal and pelvic ultrasonogram showed a normal uterus and ovaries. Careful history taking revealed frequent use of steroid ointment due to vulvar pruritus for 2 years and she was finally diagnosed to have atrophy of external genitalia induced by long-term use of topical steroid.
Atrophy* ; Collagen ; Disorders of Sex Development ; Female ; Genitalia* ; Karyotype ; Ovary ; Pruritus ; Reference Values ; RNA Stability ; RNA, Messenger ; Skin ; Ultrasonography ; Uterus

Mutation and reduction of mitochondrial DNA (mtDNA) have been suggested as factors in the pathogenesis of several metabolic diseases. Recently, we demonstrated that C1qTNF-related protein-6 (CTRP6) is involved in fatty acid metabolism in muscle cells. In this study, we showed that expression of CTRP6 was up-regulated in mtDNA-depleted C2C12 cells, which displayed a marked decrease in cellular mtDNA and ATP content. Replacement of mtDNA normalized the expression level of CTRP6 similar to that in normal C2C12 cells, indicating that CTRP6 expression was up-regulated by mtDNA depletion. However, CTRP6 promoter activity remained unchanged in mtDNA-depleted cells. We also found that mtDNA depletion inhibited decay of CTRP6 mRNA. Taken together, mtDNA depletion induces an increase in CTRP6 expression by increasing mRNA stability.
Adiponectin/*genetics/metabolism ; Animals ; Cell Line ; DNA, Mitochondrial/*metabolism ; Mice ; Promoter Regions, Genetic ; RNA Stability ; RNA, Messenger/*metabolism ; Up-Regulation

Circular RNAs (circ RNAs) are a novel type of RNA that, unlike linear RNAs, form a covalently closed continuous loop and are highly represented in the eukaryotic transcriptome. They share a stable structure, high expression and often exhibit tissue/developmental-stage-specific expression. Emerging evidence indicates that circRNAs might play important roles in human disease, such as cancer, neurological disorders and atherosclerotic vascular disease risk. The huge potentials of circRNAs are recently being discovered from the laboratory to the clinic. CircRNAs might be developed as a potential novel and stable biomarker and potential drugs used in disease diagnosis and treatment. Here, we review the current understanding of the roles of circRNAs in human disease and their potential clinic significance in disease.
Biomarkers ; analysis ; Disease ; genetics ; Gene Expression Regulation ; Humans ; Neoplasms ; diagnosis ; genetics ; therapy ; RNA ; genetics ; RNA Stability ; Signal Transduction ; genetics

The let-7 miRNA was one of the first miRNAs discovered in the nematode, Caenorhabditis elegans, and its biological functions show a high level of evolutionary conservation from the nematode to the human. Unlike in C. elegans, higher animals have multiple isoforms of let-7 miRNAs; these isoforms share a consensus sequence called the 'seed sequence' and these isoforms are categorized into let-7 miRNA family. The expression of let-7 family is required for developmental timing and tumor suppressor function, but must be suppressed for the self-renewal of stem cells. Therefore, let-7 miRNA biogenesis must be carefully controlled. To generate a let-7 miRNA, a primary transcript is produced by RNA polymerase II and then subsequently processed by Drosha/DGCR8, TUTase, and Dicer. Because dysregulation of let-7 processing is deleterious, biogenesis of let-7 is tightly regulated by cellular factors, such as the RNA binding proteins, LIN28A/B and DIS3L2. In this review, we discuss the biological functions and biogenesis of let-7 miRNAs, focusing on the molecular mechanisms of regulation of let-7 biogenesis in vertebrates, such as the mouse and the human.
Animals ; Base Sequence ; Gene Expression Regulation ; Humans ; MicroRNAs ; biosynthesis ; chemistry ; genetics ; metabolism ; RNA Processing, Post-Transcriptional ; RNA Stability ; Transcription, Genetic

OBJECTIVE: To explore the regulatory function of RNA binding motif protein 38 (RBM38) in human acute myeloid leukemia cells HL-60 and its mechanism. METHODS: The lentivirus carriers of overexpressed and knockdown RBM38 were constructed. After HL-60 cells were transfected, Western blot was used to analyze the expression level of RBM38 in HL-60 cells. The cell proliferation and cycle of HL-60 were detected by CCK-8 assay and flow cytometry assay, respectively. RNA immunoprecipitation coupled real-time PCR (RIP-qPCR) was used to detect the combination of RBM38 with mRNAs. Actinomycin D treatment followed by real-time PCR (AcD-qPCR) was used to detect the effect of RBM38 on the stability of target mRNAs. RESULTS: RBM38 in HL-60 cells was overexpressed or inhibited by lentivirus transduction. Overexpressed RBM38 promoted the cell cycle and proliferation of HL-60, while RBM38 knockdown repressed the two processes. RBM38 showed an interaction with FZD1 mRNA and enhancement of its stability. CONCLUSION RBM38 can regulate cell proliferation of HL-60 by improving the stability of FZD1 mRNA.
Cell Proliferation ; Frizzled Receptors ; HL-60 Cells ; Humans ; Leukemia, Myeloid, Acute ; RNA Stability ; RNA-Binding Proteins/metabolism*