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Results for “MESSENGER-RNA”

Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 127 records · Page 7
All sourcesNASA NTRSDOE OSTI
DOE OSTI2018

Sterol regulatory element binding protein 1 trans-activates 25-hydroxy vitamin D3 24-hydroxylase gene expression in renal proximal tubular cells

Highlights: • SREBP1 activates CYP24A1 transcription in a kidney proximal tubule cell line. • SREBP1 binds to a putative SRE in mouse Cyp24a1 to activate transcription. • Mice injected with thyroid hormone T{sub 3} had reduced renal Srebp1c and Cyp24a1 levels. • SREBP1 trans-activation of CYP24A1 expression could regulate vitamin D{sub 3} levels. The physiological activity of the steroid derived hormone vitamin D is regulated by several enzymatic steps. Both 25-hydroxy vitamin D{sub 3} 1α-hydroxylase (CYP27B1) and 25-hydroxyvitamin D{sub 3} 24-hydroxylase (CYP24A1) modulate blood levels of 1,25-dihydroxyvitamin D{sub 3}, an activated form of vitamin D. We previously demonstrated that CYP27B1 expression was trans-activated by sterol regulatory element binding protein 1 (SREBP1), although whether SREBP1 also regulates CYP24A1 transcription was unclear. Here we investigated the ability of SREBP1 to affect CYP24A1 transcription. In a luciferase reporter assay, mouse and human CYP24A1 promoter activity was strongly activated by SREBP1 in opossum kidney proximal tubular cells (OK-P). Three putative SREs (pSREs) were found in the mouse Cyp24a1 gene promoter and the SREBP1 protein showed specific binding to the pSRE1 element in EMSAs. Site-directed mutagenesis of the pSRE1 element strongly decreased SREBP1-mediated Cyp24a1 gene transcription. Moreover, siRNA-mediated SREBP1 knock-down repressed CYP24A1 expression in human renal proximal tubular epithelial cells (HKC-8). In animal studies, mice given various doses of thyroid hormone (T{sub 3}) showed dose-dependent reductions in renal Srebp1c and Cyp24a1 mRNA levels. Taken together, our results suggest that SREBP1 trans-activates CYP24A1 expression through SREBP binding elements present in the promoter.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Role of acid responsive genes in the susceptibility of Escherichia coli to ciclopirox

Highlights: • Ciclopirox antibacterial activity was analysed by genome-wide expression profiling. • Ciclopirox affects expression of glutamate-dependent acid resistance pathway genes. • Down-regulation of hns transcription likely increases ciclopirox activity. • Susceptibility to ciclopirox accompanies increased cell size and decreased motility. • The findings identify probable pathways for ciclopirox action against E. coli. Antibiotic resistance poses a huge threat to the effective treatment of bacterial infections. To circumvent the limitations in developing new antibiotics, researchers are attempting to repurpose pre-developed drugs that are known to be safe. Ciclopirox, an off-patent antifungal agent, inhibits the growth of Gram-negative bacteria, and genes involved in galactose metabolism and lipopolysaccharide (LPS) biosynthesis are plausible antibacterial targets for ciclopirox, since their expression levels partially increase susceptibility at restrictive concentrations. In the present study, to identify new target genes involved in the susceptibility of Escherichia coli to ciclopirox, genome-wide mRNA profiling was performed following ciclopirox addition at sublethal concentrations, and glutamate-dependent acid resistance (GDAR) genes were differentially regulated. Additional susceptibility testing, growth analyses and viability assays of GDAR regulatory genes revealed that down-regulation of evgS or hns strongly enhanced susceptibility to ciclopirox. Further microscopy and phenotypic analyses revealed that down-regulation of these genes increased cell size and decreased motility. Our findings could help to maximise the efficacy of ciclopirox against hard-to-treat Gram-negative pathogens.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Evaluation of synthetic promoters in Physcomitrella patens

Highlights: • Cis-elements can be randomly assembled to construct short plant promoters. • Synthetic promoters have higher mRNA expression level than the endogenous PpAct7. • Synthetic promoters have similar protein expression levels to AtUBQ10 promoter. Securing a molecular toolbox including diverse promoters is essential for genome engineering. However, native promoters have limitations such as the available number or the length of the promoter. In this work, three short synthetic promoters were characterized by using the yellow fluorescent protein Venus. All of the tested promoters were active and showed higher mRNA expression than housekeeping gene PpAct7, and similar protein expression level to the AtUBQ10 promoter. This study shows that few cis-regulatory elements are enough to establish a strong promoter for continuous expression of genes in plants. Along with this, the study enhance the number of available promotors to be used in P. patens. It also demonstrates the potential to construct multiple non-native promoters on demand, which would aid to resolve one bottleneck in multiple pathway expression in P. patens and other plants.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Transcription of blunt snout bream (Megalobrama amblycephala) HIF3α and its localization in the nucleus under both normoxic and hypoxic conditions

Highlights: • HIF3α cDNA was characterized the in hypoxia-sensitive blunt snout bream. • HIF3α mRNA was stably expressed during embryogenesis and different adult tissues. • HIF3α mRNA was upregulated in the liver, brain, and kidney under hypoxia. • HIF3α was distributed in the nucleus under normoxic and hypoxic conditions. • Hypoxia simulation caused a significant increase in HIF3α protein levels. • There was a nuclear localization signal (NLS) sequence at the C-terminus of HIF3α. Although hypoxia-inducible factor (HIF) 1α and 2α function as master regulators of the transcriptional response to hypoxia, the function of HIF3α and its responses to hypoxic stress remain unclear in teleost fish. Here, we characterized the HIF3α cDNA in hypoxia-sensitive blunt snout bream (Megalobrama amblycephala), with 3059 bp length, consisting of an open reading frame (ORF) encoding 643 amino acid residues. Blunt snout bream HIF3α mRNA was stably expressed during stages of embryonic development and in adult tissues. After a 4 h hypoxia stress, HIF3α mRNA of the juvenile fish was significantly upregulated in the liver, brain, and kidney, and restored to the pretreatment levels after a 24 h recovery. When tagged with enhanced green fluorescent protein (EGFP) and transfected into cultured HeLa cells, blunt snout bream HIF3α was mainly distributed in the nucleus under normoxia. Treatment of the cells with CoCl{sub 2} to mimic hypoxic conditions showed that there was no effect about the nuclear localization of HIF3α but a statistically significant increase in HIF3α protein levels. A nuclear localization signal (NLS) sequence at the C-terminus of HIF3α may exert positive effects in the process of nuclear localization. These results suggest that blunt snout bream HIF3α could be involved in different physiological functions under normoxia and hypoxia conditions.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Overexpression of NRK1 ameliorates diet- and age-induced hepatic steatosis and insulin resistance

NAD{sup +} is a co-enzyme in redox reactions and a substrate required for activity of various enzyme families, including sirtuins and poly(ADP-ribose) polymerases. Dietary supplementation of NAD{sup +} precursors nicotinamide mononucleotide (NMN) or nicotinamide riboside (NR) protects against metabolic disease, neurodegenerative disorders and age-related physiological decline in mammals. Here we sought to identify the roles of nicotinamide riboside kinase 1 (NRK1) plays in regulating hepatic NAD{sup +} biosynthesis and lipid metabolism. Using adenovirus mediated gene transduction to overexpress or knockdown NRK1 in mouse liver, we have demonstrated that NRK1 is critical for maintaining hepatic NAD{sup +} levels and triglyceride content. We have further shown that the hepatic expression of Nmrk1 mRNA is significantly decreased either in mice treated with high-fat diet or in aged mice. However, adenoviral delivery of NRK1 in these diet- and age-induced mice elevates hepatic NAD{sup +} levels, reduces hepatic steatosis, and improves glucose tolerance and insulin sensitivity. Our results provide important insights in targeting NRK1 for treating hepatic steatosis.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Frequencies and expression levels of programmed death ligand 1 (PD-L1) in circulating tumor RNA (ctRNA) in various cancer types

Highlights: • Blood were collected from gastric, colorectal, lung, breast, prostate cancer patients. • PD-L1 expression was detected in the plasma at varying frequencies. • No PD-L1 mRNA was detected in cancer-free individuals. • PD-L1 mRNA can be detected and quantitated in ctRNA of cancer patients. Precision medicine and prediction of therapeutic response requires monitoring potential biomarkers before and after treatment. Liquid biopsies provide noninvasive prognostic markers such as circulating tumor DNA and RNA. Circulating tumor RNA (ctRNA) in blood is also used to identify mutations in genes of interest, but additionally, provides information about relative expression levels of important genes. In this study, we analyzed PD-L1 expression in ctRNA isolated from various cancer types. Tumors inhibit antitumor response by modulating the immune checkpoint proteins programmed death ligand 1 (PD-L1) and its cognate receptor PD1. The expression of these genes has been implicated in evasion of immune response and resistance to targeted therapies.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Genome-wide profiles of metastasis-associated mRNAs and microRNAs in salivary adenoid cystic carcinoma

Highlights: • Different expressed miRNAs and mRNAs were profiled in lowly and highly metastatic SACC cell lines. • Multiple regulatory interactions between miRNAs and mRNAs may contribute to SACC metastasis. • miR-338–5p/3p and their target LAMC2 function on metastasis of multiple types of cancer. Salivary adenoid cystic carcinoma (SACC) is often accompanied with poor prognosis due to local recurrence, distant metastasis, and perineural invasion. The mechanism involved in SACC metastasis is not yet fully understood. In this study, we profiled the expression of messenger RNA (mRNA) and microRNA (miRNA) in a SACC cell line, ACC-2, and a highly metastatic SACC cell line, ACC-M, using high-throughput sequencing. We discovered that: (1) differentially expressed (DE) mRNAs and DE miRNAs are potentially involved in SACC metastasis; (2) multiple regulatory interactions between DE miRNAs and DE mRNAs exist; and (3) miR-338–5p/3p target LAMC2 to impair motility and invasion of ACC-M and MDA-MB-231 cells. In conclusion, our study integrated the regulatory effects of miRNAs and mRNAs on SACC metastasis and provided a potential application for miRNAs in future therapeutic intervention.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Long non-protein coding RNA DANCR functions as a competing endogenous RNA to regulate osteoarthritis progression via miR-577/SphK2 axis

Highlights: • DANCR was increased in OA. • DANCR inhibition suppressed OA chondrocytes proliferation and induced cell apoptosis. • DANCR-miR-577/SphK2 axis play important roles in OA. Long noncoding RNAs (lncRNAs) have been known to be involved in multiple diverse diseases, including osteoarthritis (OA). This study aimed to explore the role of differentiation antagonizing non-protein coding RNA (DANCR) in OA and identify the potential molecular mechanisms. The expression of DANCR in cartilage samples from patients with OA was detected using quantitative reverse transcription–polymerase chain reaction. The effects of DANCR on the viability of OA chondrocytes and apoptosis were explored using cell counting kit 8 assay and flow cytometry assay, respectively. Additionally, the interaction among DANCR, miR-577, and SphK2 was explored using dual-luciferase reporter and RIP assays. The present study found that DANCR was significantly upregulated in patients with OA. Functional assays demonstrated that DANCR inhibition suppressed the proliferation of OA chondrocytes and induced cell apoptosis. The study also showed that DANCR acted as a competitive endogenous RNA to sponge miR-577, which targeted the mRNA of SphK2 to regulate the survival of OA chondrocytes. In conclusion, the study revealed that lncRNA DANCR might promote the proliferation of OA chondrocytes and reduce apoptosis through the miR-577/SphK2 axis. Thus, lncRNA DANCR might be considered as a potential therapeutic target for OA treatment.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Matrix stiffness regulate apoptotic cell death in HIV-HCV co-infected hepatocytes: Importance for liver fibrosis progression

Highlights: • HIV-HCV co-infection increases HIV/HCV RNAs levels and apoptosis in hepatocytes. • Fibrotic matrix stiffness enhances HIV/HCV RNAs and apoptosis in hepatocytes. • Apoptotic infected hepatocytes induce pro-fibrotic changes in hepatic stellate cells. HIV-HCV co-infection causes rapid progression of liver fibrosis. These outcomes to liver cirrhosis can be improved, but not stopped by specific antiviral therapies. Due to high significance of HIV-HCV interactions for morbidity and mortality in co-infected patients, our attention was attracted to the multi-component pathogenesis of fibrosis progression as the transition to end-stage liver disease development. In this study, we hypothesize that increased matrix stiffness enhances apoptosis in HCV-HIV-co-infected hepatocytes and that capturing of apoptotic bodies (AB) derived from these infected hepatocytes by hepatic stellate cells (HSC) drives the fibrosis progression. As the source of viruses, JFH1 (HCV genotype 2a) and HIV-1{sub ADA} (either purified or containing in infected macrophage supernatants) were chosen. Using Huh7.5-CYP (RLW) cells and primary human hepatocytes mono-infected with HCV and HIV or co-infected, we have shown that both HCV and HIV RNA levels were increased in co-infected cells, which was accompanied by hepatocyte apoptosis. This apoptosis was attenuated by azidothymidine treatment. The levels of both infections and apoptosis were more prominent in primary hepatocytes cultured on substrates mimicking fibrotic stiffness (24 kPa-stiff) compared to substrates mimicking healthy liver (2.4 kPa-soft). The engulfment of AB from pathogen-exposed hepatocytes activated pro-fibrotic mRNAs in HSC. Overall, the increased matrix stiffness is not only a consequence of liver inflammation/fibrosis, but the condition that further accelerates liver fibrosis development. This is attributed to the switching of HSC to pro-fibrotic phenotype by capturing of excessive amounts of apoptotic HCV- and HIV-infected hepatocytes.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Bitter tastant quinine modulates glucagon-like peptide-1 exocytosis from clonal GLUTag enteroendocrine L cells via actin reorganization

Highlights: • Various bitter taste receptors were expressed in GLUTag cells. • Quinine increased the intracellular Ca{sup 2+} levels but did not promote GLP-1 secretion. • Quinine caused actin reorganization and suppressed exocytosis of GLP-1 vesicles. • Application of forskolin in quinine-treated GLUTag cells enhanced GLP-1 exocytosis. Enteroendocrine L cells in the gastrointestinal tract secrete glucagon-like peptide-1 (GLP-1), which plays an important role in glucose homeostasis. Here we investigated the effect of bitter tastant quinine on GLP-1 secretion using clonal GLUTag mouse enteroendocrine L cells. We found that GLUTag cells expressed putative quinine receptors at mRNA levels. Although application of quinine resulted in an increase of intracellular Ca{sup 2+} levels, which was mediated by Ca{sup 2+} release from the endoplasmic reticulum and Ca{sup 2+} influx through voltage-sensitive Ca{sup 2+} channels, quinine had little effect on GLP-1 secretion. Total internal reflection fluorescence microscopy and immunocytochemistry revealed that GLP-1-containing vesicles remained unfused with the plasma membrane and facilitated actin polymerization beneath the plasma membrane after application of quinine, respectively. Interestingly, application of forskolin together with quinine induced GLP-1 exocytosis from the cells. These results suggest that quinine does not induce GLP-1 secretion because it facilitates Ca{sup 2+} increase and actin reorganization but not cAMP increase, and both Ca{sup 2+} and cAMP are essential for GLP-1 secretion.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

DDX1 regulates alternative splicing and insulin secretion in pancreatic β cells

DEAD-box helicase 1 (DDX1) is a multifunction protein involved in diverse cellular processes including transcription, viral replication, mRNA/miRNA processing, and tRNA splicing. Here, we report a novel function of DDX1 in mRNA alternative splicing in pancreatic β cells. By performing integrated data analysis of high-throughput RNA sequencing (RNA-Seq), and cross-linking and immunoprecipitation coupled with deep sequencing (CLIP-Seq), we identify hundreds of alternative splicing genes that are targeted by DDX1. These DDX1-targeted alternative splicing genes are mainly associated with calcium ion binding, high voltage-gated calcium channel, and transmembrane transporter. Functionally, silencing DDX1 impairs calcium influx and insulin secretion in the pancreatic β cells. These results reveal an important role for DDX1 in the regulation of gene alternative splicing and insulin secretion in pancreatic β cells.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

HDAC-mediated deacetylation of KLF5 associates with its proteasomal degradation

Highlights: • Highlights: (for review). • Silencing HDAC1/2 increased KLF5 protein by preventing proteasomal degradation. • Deacetylase activity was required for HDAC1/2-mediated KLF5 degradation. • The acetylation-mimicking form of KLF5 was prevented from degradation. • The binding of KLF5 with HDAC1/2 was not affected by its acetylation status. Krüppel-like factor 5 (KLF5) is a basic transcription factor that regulates diverse cellular processes during tumor development. Acetylation of KLF5 at lysine 369 (K369) reverses its function from promoting to suppressing cell proliferation and tumor growth. In this study, we examined the regulation of KLF5 by histone deacetylases in the prostate cancer cell line DU 145. While confirming the functions of HDAC1/2 in KLF5 deacetylation and the promotion of cell proliferation, we found that the knockdown of HDAC1/2 upregulated KLF5 protein but not KLF5 mRNA, and the increase in KLF5 protein level by silencing HDAC1/2 was at least in part due to decreased proteasomal degradation. Deacetylase activity was required for HDAC1/2-mediated KLF5 degradation, and mutation of KLF5 to an acetylation-mimicking form prevented its degradation, even though the mutation did not affect the binding of KLF5 with HDAC1/2. Mutation of K369 to arginine, which prevents acetylation, did not affect the binding of KLF5 to HDAC1 or the response of KLF5 to HDAC1/2-promoted degradation. These findings provide a novel mechanistic association between the acetylation status of KLF5 and its protein stability. They also suggest that maintaining KLF5 in a deacetylated form may be an important mechanism by which KLF5 and HDACs promote cell proliferation and tumor growth.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Down-regulation of TRAF4 targeting RSK4 inhibits proliferation, invasion and metastasis in breast cancer xenografts

Highlights: • Knockdown of TRAF4 inhibited the growth of xenograft tumors in nude mice. • Knockdown of TRAF4 significantly decreased the lung and liver metastases. • There is a negative correlation between TRAF4 and RSK4 mRNA and proteins. TRAF4 is interacting with RSK4 possibly by the induction of AKT/NF-κB signal axis. Ribosomal S6 protein kinase 4 (RSK4) was known as a novel tumor suppressor gene, and the tumor necrosis factor receptor-associated factor 4 (TRAF4) was linked to carcinogenesis. The purpose of this study is to further investigate the effect of the TRAF4 gene on cell proliferation, invasion and metastasis in vivo and explore whether there is an interaction between TRAF4 and RSK4 in breast cancer. MDA-MB-231 cells were transfected with lentivirus TRAF4-shRNA to specifically block the expression of TRAF4, or transfected with lentivirus negative-shRNA as a negative control. Four-six weeks female BALB/c nude mice were randomly assigned to three groups (n = 14): TRAF4-shRNA, negative and control, and then inoculated subcutaneously with the corresponding cells. In-vivo metastasis model was constructed by injecting above cells into tail vein. Tumor proliferation was assessed in terms of the tumor growth curve, tumor size and weight. Invasion and metastasis were evaluated by the histopathologic examination in lung or/and liver tissues. Measurement of TRAF4 and RSK4 expression and their correlation factors (AKT, NF-κB, TGF-β1, TNF-α, MMP2 and MMP9) were performed by immunohistochemistry, western blot or fluorescence quantitative RT-PCR. We found that the size and weight of tumors in TRAF4-shRNA group was significantly smaller than the negative and blank group, and the number of the lung and liver metastases lesions was also fewer (P < 0.05). And TRAF4 and its correlation factors (P-AKT, P-NF-κB, TGF-β1, TNF-α, MMP2 and MMP9) in the TRAF4-shRNA group were significantly decreased compared with the negative and blank group. However, the expression of RSK4 mRNA and protein in TRAF4-shRNA group were significantly increased. Collectively, TRAF4 knockdown significantly inhibited proliferation, invasion and metastasis in the xenograft nude mouse model, possibly involving in the interaction with RSK4 through down-regulation of AKT signaling pathway and then inactivating NF-κB pathway.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Respective roles of the mitogen-activated protein kinase (MAPK) family members in pancreatic stellate cell activation induced by transforming growth factor-β1 (TGF-β1)

Activated pancreatic stellate cells (PSCs) play a crucial role in the progression of pancreatic fibrosis. Transforming growth factor-β (TGF-β) is one of the strongest stimulator inducing fibrosis. The mitogen-activated protein kinase (MAPK) proteins (including ERK, JNK and p38 MAPK) are known to contribute to PSC activation and pancreatic fibrosis. Previous studies have identified PSC activation induced by TGF-β1 is related to MAPK pathway, but the respective role of MAPK family members in PSC activation still unclear, and which family member may be the key mediator in mice PSC activation still controversial. In this study, we investigated the influence of different MAPK family member (JNK, ERK, and p38 MAPK) on mice PSC activation using an in vivo and in vitro model. The results showed p-JNK, p-ERK and p-p38 MAPK were all over-expressed in CP group, and p-JNK, p-ERK, and p-p38 MAPK were co-expressed with activated PSC. In vitro, TGF-β1 induced JNK and ERK over-expression in PSCs. In contrast, p38 MAPK expression in PSC showed only a very weak increase. JNK- and ERK-specific inhibitors inhibited FN and α-SMA mRNA expression in PSCs, and a p38 MAPK inhibitor had no effect on PSC activation. These findings indicate that JNK and ERK were directly involved in the PSCs activation induced by TGF-β1 and the development of pancreatic fibrosis. p38 MAPK participate in the progression of CP, but it does not respond to TGF-β1 directly and may not be regarded as the target of TGF-β1 induced PSC activation.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Protective role of AgRP neuron's PDK1 against salt-induced hypertension

Highlights: • AgRP neuron specific PDK1 KO mice responded to high salt diet with hypertension. • The KO mice showed elevated urinary noradrenalin excretion under high salt diet. • The KO mice showed elevated NUCB2 mRNA expression in paraventricular nucleus (PVN). • PVN NUCB2 silencing counteracted elevated noradrenalin excretion and hypertension. • AgRP neuron's PDK1 serves to regulate PVN nesfatin-1 neurons and blood pressure. In the hypothalamic arcuate nucleus (ARC), orexigenic agouti-related peptide (AgRP) neurons regulate feeding behavior and energy homeostasis. The 3-phosphoinositide-dependent protein kinase-1 (PDK1) in AgRP neurons serves as a major signaling molecule for leptin and insulin, the hormones regulating feeding behavior, energy homeostasis and circulation. However, it is unclear whether PDK1 in AGRP neurons is also involved in regulation of blood pressure. This study explored it by generating and analyzing AgRP neuron-specific PDK1 knockout (Agrp-Pdk1{sup flox/flox}) mice and effect of high salt diet on blood pressure in KO and WT mice was analyzed. Under high salt diet feeding, systolic blood pressure (SBP) of Agrp-Pdk1{sup flox/flox} mice was significantly elevated compared to Agrp-Cre mice. When the high salt diet was switched to control low salt diet, SBP of Agrp-Pdk1{sup flox/flox} mice returned to the basal level observed in Agrp-Cre mice within 1 week. In Agrp-Pdk1{sup flox/flox} mice, urinary noradrenalin excretion and NUCB2 mRNA expression in hypothalamic paraventricular nucleus (PVN) were markedly upregulated. Moreover, silencing of NUCB2 in the PVN counteracted the rises in urinary noradrenalin excretions and SBP. These results demonstrate a novel role of PDK1 in AgRP neurons to counteract the high salt diet-induced hypertension by preventing hyperactivation of PVN nesfatin-1 neurons.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Transplantation of periaortic adipose tissue inhibits atherosclerosis in apoE −/− mice by evoking TGF-β1-mediated anti-inflammatory response in transplanted graft

Highlights: • Transplantation of tPAT reduces the plaque size in apoE{sup -/-} mice, concomitant with increased serum TGF-β1levels. • TGF-β1 levels show significant inverse correlation with plaque size in tPAT-transplanted mice. • Neutralizing TGF-β antibody abrogates anti-atherogenic effect of tPAT transplantation. • TGF-β-positive staining is colocalized with Mac-2-positive cells in grafted tPAT. • mRNA expression levels of alternatively activated macrophages genes and IL-4 are significantly increased in grafted tPAT. Perivascular adipose tissue (PAT) is associated with vascular homeostasis; however, its causal effect on atherosclerosis currently remains undefined. Here, we investigated the effect of experimental PAT transplantation on atherosclerosis. The thoracic periaortic adipose tissue (tPAT) was dissected from 16-week-old wild-type mice and transplanted over the infrarenal aorta of 20-week-old apoE deficient (apoE{sup −/−}) mice fed high-cholesterol diet for 3 months. Oil-red O staining after 4 weeks showed a significant 20% decrease in the atherosclerotic lesion of suprarenal aorta compared with that of sham control mice, while that of infrarenal aorta showed no difference between the two groups. TGF-β1 mRNA expression was significantly higher in grafted tPAT than donor tPAT, accompanied by a significant increase in serum TGF-β1 concentration, which was inversely correlated with the suprarenal lesion area (r = −0.63, P = 0.012). Treatment with neutralizing TGF-β antibody abrogated the anti-atherogenic effect of tPAT transplantation. Immunofluorescent analysis of grafted tPAT showed that TGF-β-positive cells were co-localized with Mac-2-positive cells and this number was significantly increased compared with donor tPAT. There was also marked increase in mRNA expression of alternatively activated macrophages-related genes. Furthermore, the percentage of eosinophils in stromal vascular fraction of donor tPAT was much higher than that in epididymal white adipose tissue, concomitant with the significantly higher protein level of IL-4. IL-4 mRNA expression levels in grafted tPAT were increased in a time-dependent manner after tPAT transplantation. Our findings show that tPAT transplantation inhibits atherosclerosis development by exerting TGF-β1-mediated anti-inflammatory response, which may involve alternatively activated macrophages.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

FOXM1 promotes proliferation in human hepatocellular carcinoma cells by transcriptional activation of CCNB1

Highlights: • FOXM1 and CCNB1 are prognostic indicators in human hepatocellular carcinoma. • FOXM1 transcriptionally upregulates CCNB1 expression levels in HCC cells. • CCNB1 is essential for proliferation in HCC cells. • FOXM1 modulates HCC cell proliferation by targeting CCNB1. The transcription factor Forkhead box protein M1 (FOXM1) plays critical roles in cancer development and progression, including human hepatocellular carcinoma (HCC). However, the regulatory role and underlying mechanisms of FOXM1 is still limited. Here, we found that the high level expression of FOXM1 and CCNB1 is closely associated with poor prognosis in HCC patients. And FOXM1 and CCNB1 were overexpressed concomitantly in liver tumor tissues. Knockdown of FOXM1 significantly inhibited the expression levels of CCNB1 in HCC cell lines at both the mRNA and protein levels. Mechanistic studies revealed that FOXM1 binds directly to the promoter region of CCNB1 and regulates the expression levels of the CCNB1 gene in the transcriptional level. Furthermore, the loss of functional and rescue experiments showed that CCNB1 is essential for FOXM1-driven proliferation in HCC cells. In the present study, our results partially explained the dysregulated expression of FOXM1 play an important role in proliferation of human hepatocellular carcinoma cells via transcriptional activation of CCNB1 expression. And it also highlights a FOXM1/CCNB1 axis could be a potential target for the treatment of HCCs.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

The orphan G protein-coupled receptor 25 (GPR25) is activated by Apelin and Apela in non-mammalian vertebrates

Highlights: • The orphan receptor GPR25 was cloned from zebrafish, pigeons and spotted gars. • GPR25 is activated by Apelin and Apela in non-mammalian vertebrates. • Like Apelin receptor, GPR25 is coupled to Gi-cAMP signaling pathway. • GPR25 mRNA is widely expressed in zebrafish/spotted gar/human tissues. G protein-coupled receptor 25 (GPR25) is an orphan G protein-coupled receptor in vertebrates, that has been implicated to be associated with autoimmune diseases and regulate blood pressure in humans. However, the endogenous ligand of GPR25 remains unknown in vertebrates. Here, we reported that in non-mammalian vertebrates (zebrafish, spotted gars, and pigeons), GPR25 could be activated by Apelin and Apela peptides, which are also the two endogenous ligands of vertebrate Apelin receptor (APLNR). Using the pGL3-CRE-luciferase reporter assay and confocal microscopy, we first demonstrated that like APLNR, zebrafish GPR25 expressing in HEK293 cells could be effectively activated by zebrafish Apelin and Apela peptides, leading to the inhibition of forskolin-stimulated cAMP production and receptor internalization. Like zebrafish GPR25, pigeon and spotted gar GPR25 could also be activated by Apelin and Apela, and their activation could inhibit forskolin-induced cAMP accumulation. Interestingly, unlike zebrafish (/spotted gar/pigeon) GPR25, human GPR25 could not be activated by Apelin and Apela under the same experimental conditions. RNA-seq analysis further revealed that GPR25 is expressed in a variety of tissues, including the testes and intestine of zebrafish/spotted gars/humans, implying the potential roles of GPR25 signaling in many physiological processes in vertebrates. Taken together, our data not only provides the first proof that the orphan receptor GPR25 possesses two potential ligands ‘Apelin and Apela’ and its activation decreases intracellular cAMP levels in non-mammalian vertebrates, but also facilitates to unravel the physiological roles of GPR25 signaling in vertebrates.

60 APPLIED LIFE SCIENCES↗