The Efficacy of the Hepatocyte Spheroids for Hepatocyte Transplantation

The safety and short-term efficacy of hepatocyte transplantation (HCTx) have been widely proven. However, issues such as reduced viability and/or function of hepatocytes, insufficient engraftment, and lack of a long-term effect have to be overcome for widespread application of HCTx. In this study, we evaluated hepatocyte spheroids (HSs), formed by self-aggregation of hepatocytes, as an alternative to hepatocytes in single-cell suspension. Hepatocytes were isolated from C57BL/6 J mice liver using a three-step collagenase perfusion technique and HSs were formed by the hanging drop method. After the spheroids formation, the HSs showed significantly higher mRNA expression of albumin, ornithine transcarbamylase, glucose-6-phosphate, alpha-1-antitrypsin, low density lipoprotein receptor, coagulation factors, and apolipoprotein E (ApoE) than 2 dimensional (2D)-cultured hepatocytes ( p < 0.05). Albumin production by HSs was significantly higher than that by 2D-cultured hepatocytes (9.5 ± 2.5 vs 3.5 ± 1.8 μg/dL, p < 0.05). The HSs, but not single hepatocytes, maintained viability and albumin mRNA expression in suspension (92.0 ± 2.8% and 1.03 ± 0.09 at 6 h). HSs (3.6 × 106 cells) or isolated hepatocytes (fSH, 3.6 × 106 cells) were transplanted into the liver of ApoE knockout (KO-/-) mice via the portal vein. Following transplantation, serum ApoE concentration (ng/mL) of HS-transplanted mice (1w: 63.1 ± 56.7, 4w: 17.0 ± 10.9) was higher than that of fSH-transplanted mice (1 w: 33.4 ± 13.0, 4w: 13.7 ± 9.6). In both groups, the mRNA levels of pro-inflammatory cytokines (IL-6, IL-1β, TNF-α, MCP-1, and MIP-1β) were upregulated in the liver following transplantation; however, no significant differences were observed. Pathologically, transplanted HSs were observed as flat cell clusters in contact with the portal vein wall on day 7. Additionally, ApoE positive cells were observed in the liver parenchyma distant from the portal vein on day 28. Our results indicate that HS is a promising alternative to single hepatocytes and can be applied for HCTx.

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Effects of alfalfa saponin extract on the performance and cholesterol metabolism of laying hens

10.7287/peerj.preprints.1461v1 ◽

2015 ◽

Author(s):

Senhao Zhang ◽

Yinghua Shi ◽

Minggen Liang ◽

Jia Li ◽

Chengzhang Wang

Keyword(s):

Bile Acid ◽

Mrna Expression ◽

Laying Hens ◽

Cholesterol Metabolism ◽

Low Density Lipoprotein ◽

Density Lipoprotein ◽

Low Density ◽

Mrna Levels ◽

Density Lipoprotein Receptor ◽

Yolk Cholesterol

The experiment was performed to determine the effects of alfalfa saponin extract (ASE) on the performance and cholesterol metabolism of laying hens. A total of 150 Hy-Line Brown hens with 28 weeks old, were randomly divided into five treatment groups (five replicates per treatment with six hens per replicate). Diets containing 0, 60, 120, 240, and 480 mg ASE/kg were fed to hens for 77 days. The shell thickness had a trend to increase. The yolk cholesterol and liver bile acid decreased significantly (ASE 60 and 480 mg/kg groups for yolk cholesterol, and ASE 60 and 240 mg/kg groups for liver bile acid). Fecal bile acid has an elevation trend as ASE increased. The expression of very low density apolipoprotein-Ⅱ (apoVLDL-Ⅱ) gene was not affected by adding ASE. However, the mRNA expression of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase gene and cholesterol 7α-hydroxylase (CYP7A1) gene were significantly up-regulated. The mRNA expression of very-low-density-lipoprotein receptor(VLDLR) gene was suppressed due to adding ASE supplementation in the diet. These findings indicated that dietary ASE could regulate cholesterol levels in hens by up-regulating the mRNA levels of HMG-CoA and CYP7A1 and suppressing the expression of VLDLR.

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Effects of alfalfa saponin extract on the performance and cholesterol metabolism of laying hens

10.7287/peerj.preprints.1461 ◽

2015 ◽

Author(s):

Senhao Zhang ◽

Yinghua Shi ◽

Minggen Liang ◽

Jia Li ◽

Chengzhang Wang

Keyword(s):

Bile Acid ◽

Mrna Expression ◽

Laying Hens ◽

Cholesterol Metabolism ◽

Low Density Lipoprotein ◽

Density Lipoprotein ◽

Low Density ◽

Mrna Levels ◽

Density Lipoprotein Receptor ◽

Yolk Cholesterol

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Oxidative Stress and Inflammation in Sows with Excess Backfat: Up-Regulated Cytokine Expression and Elevated Oxidative Stress Biomarkers in Placenta

Animals ◽

10.3390/ani9100796 ◽

2019 ◽

Vol 9 (10) ◽

pp. 796

Author(s):

Yuanfei Zhou ◽

Tao Xu ◽

Yinghui Wu ◽

Hongkui Wei ◽

Jian Peng

Keyword(s):

Oxidative Stress ◽

Mrna Expression ◽

Low Density Lipoprotein ◽

Density Lipoprotein ◽

Late Gestation ◽

Backfat Thickness ◽

Mrna Levels ◽

Oxidative Stress Biomarkers ◽

Total Antioxidant ◽

The Impact

In sows, excess backfat during late gestation is associated with increased farrowing difficulties and influences the fetus, but the impact of backfat thickness on placental inflammation, oxidative stress, and vascular development has not been defined. In this study, 120 sows were divided into six groups based on backfat thickness (≤16, 17–18, 19–20, 21–22, 23–24, and ≥25 mm) in late gestation. The placental lipids, reactive oxygen species (ROS), malondialdehyde (MDA), and total antioxidant capacity (TAC) levels, inflammatory-related cytokine and angiogenesis were determined. The concentrations of triglycerides, total cholesterol, low density lipoprotein cholesterol (LDL–C), and free fatty acid (FFA) linearly increased (p < 0.05) associated with increased late gestation backfat. ROS and MDA increased and TAC decreased (p < 0.05) as the backfat thickness increased. The mRNA expression of toll-like receptors (TLR) 2, TLR4, tumor necrosis factor (TNF) α, interleukin (IL)–1β, IL–6, and monocyte chemoattractant protein (MCP)–1 increased with increased backfat in late gestation. There were no differences in IL–8 and IL–10 mRNA expression among sows with different backfat thickness. Placental vessel density initially increased and then decreased with increasing backfat thickness of sows. Similarly, the mRNA levels of vascular endothelial growth factor (VEGF) were also increased and then decreased. Excessive backfat in late gestation was associated with greater oxidative stress, greater expression of proinflammatory cytokines, and decreased expression of placental angiogenic regulators.

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Tanshinone IIA Promotes Macrophage Cholesterol Efflux and Attenuates Atherosclerosis of apoE-/- Mice by Omentin-1/ABCA1 Pathway

Current Pharmaceutical Biotechnology ◽

10.2174/1389201020666190404125213 ◽

2019 ◽

Vol 20 (5) ◽

pp. 422-432 ◽

Cited By ~ 3

Author(s):

Yu-lin Tan ◽

Han-xiao Ou ◽

Min Zhang ◽

Duo Gong ◽

Zhen-wang Zhao ◽

...

Keyword(s):

Atherosclerotic Plaque ◽

Lipid Content ◽

Plasma Levels ◽

Cholesterol Efflux ◽

Density Lipoprotein ◽

Tanshinone Iia ◽

Lipoprotein Cholesterol ◽

Mrna Levels ◽

Cellular Lipid ◽

Oil Red O

Background: Tanshinone IIA (Tan IIA) and Omentin-1 have a protective role in the cardiovascular system. However, if and how Tan IIA and Omentin-1 regulate cholesterol metabolism in macrophages has not been fully elucidated. Objective: To investigate the possible mechanisms of Tan IIA and Omentin-1 on preventing macrophage cholesterol accumulation and atherosclerosis development. Methods: The effect of Tan IIA on the protein and mRNA levels of Omentin-1 and ATP-binding cassette transporter A1 (ABCA1) in macrophages was examined by Western blot and qRT-PCR assay, respectively. Cholesterol efflux was assessed by liquid scintillation counting (LSC). Cellular lipid droplet was measured by Oil Red O staining, and intracellular lipid content was detected by high performance liquid chromatography (HPLC). In addition, the serum lipid profile of apoE−/− mice was measured by enzymatic method. The size of atherosclerotic lesion areas and content of lipids and collagen in the aortic of apoE−/− mice were examined by Sudan IV, Oil-red O, and Masson staining, respectively. Results: Tan IIA up-regulated expression of Omentin-1 and ABCA1 in THP-1 macrophages, promoting ABCA1-mediated cholesterol efflux and consequently decreasing cellular lipid content. Consistently, Tan IIA increased reverse cholesterol transport in apoE−/− mice. Plasma levels of high-density lipoprotein cholesterol (HDL-C), ABCA1 expression and atherosclerotic plaque collagen content were increased while plasma levels of low-density lipoprotein cholesterol (LDL-C) and atherosclerotic plaque sizes were reduced in Tan IIA-treated apoE−/− mice. These beneficial effects were, however, essentially blocked by knockdown of Omentin-1. Conclusion: Our results revealed that Tan IIA promotes cholesterol efflux and ameliorates lipid accumulation in macrophages most likely via the Omentin-1/ABCA1 pathway, reducing the development of aortic atherosclerosis.

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Postnatal Smoke Exposure Further Increases the Hepatic Nicotine Metabolism in Prenatally Smoke Exposed Male Offspring and Is Linked with Aberrant Cyp2a5 Methylation

International Journal of Molecular Sciences ◽

10.3390/ijms22010164 ◽

2020 ◽

Vol 22 (1) ◽

pp. 164

Author(s):

Khosbayar Lkhagvadorj ◽

Zhijun Zeng ◽

Karolin F. Meyer ◽

Laura P. Verweij ◽

Wierd Kooistra ◽

...

Keyword(s):

Dna Methylation ◽

Mrna Expression ◽

Nicotine Dependence ◽

Susceptibility Gene ◽

Smoke Exposure ◽

Adverse Outcomes ◽

Male Offspring ◽

Adult Offspring ◽

Mrna Levels ◽

Nicotine Metabolism

Prenatal smoke exposure (PreSE) is a risk factor for nicotine dependence, which is further enhanced by postnatal smoke exposure (PostSE). One susceptibility gene to nicotine dependence is Cytochrome P450 (CYP) 2A6, an enzyme responsible for the conversion of nicotine to cotinine in the liver. Higher CYP2A6 activity is associated with nicotine dependence and could be regulated through DNA methylation. In this study we investigated whether PostSE further impaired PreSE-induced effects on nicotine metabolism, along with Cyp2a5, orthologue of CYP2A6, mRNA expression and DNA methylation. Using a mouse model where prenatally smoke-exposed adult offspring were exposed to cigarette smoke for 3 months, enzyme activity, mRNA levels, and promoter methylation of hepatic Cyp2a5 were evaluated. We found that in male offspring, PostSE increased PreSE-induced cotinine levels and Cyp2a5 mRNA expression. In addition, both PostSE and PreSE changed Cyp2a5 DNA methylation in male groups. PreSE however decreased cotinine levels whereas it had no effect on Cyp2a5 mRNA expression or methylation. These adverse outcomes of PreSE and PostSE were most prominent in males. When considered in the context of the human health aspects, the combined effect of prenatal and adolescent smoke exposure could lead to an accelerated risk for nicotine dependence later in life.

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Reprogramming mRNA Expression in Response to Defect in RNA Polymerase III Assembly in the Yeast Saccharomyces cerevisiae

International Journal of Molecular Sciences ◽

10.3390/ijms22147298 ◽

2021 ◽

Vol 22 (14) ◽

pp. 7298

Author(s):

Izabela Rudzińska ◽

Małgorzata Cieśla ◽

Tomasz W. Turowski ◽

Alicja Armatowska ◽

Ewa Leśniewska ◽

...

Keyword(s):

Mrna Expression ◽

Ribosome Biogenesis ◽

Rna Polymerase Iii ◽

Mrna Levels ◽

Rna Seq ◽

Yeast Saccharomyces Cerevisiae ◽

Protein Coding ◽

General Transcription Factor ◽

Pol I ◽

Pol Iii

The coordinated transcription of the genome is the fundamental mechanism in molecular biology. Transcription in eukaryotes is carried out by three main RNA polymerases: Pol I, II, and III. One basic problem is how a decrease in tRNA levels, by downregulating Pol III efficiency, influences the expression pattern of protein-coding genes. The purpose of this study was to determine the mRNA levels in the yeast mutant rpc128-1007 and its overdose suppressors, RBS1 and PRT1. The rpc128-1007 mutant prevents assembly of the Pol III complex and functionally mimics similar mutations in human Pol III, which cause hypomyelinating leukodystrophies. We applied RNAseq followed by the hierarchical clustering of our complete RNA-seq transcriptome and functional analysis of genes from the clusters. mRNA upregulation in rpc128-1007 cells was generally stronger than downregulation. The observed induction of mRNA expression was mostly indirect and resulted from the derepression of general transcription factor Gcn4, differently modulated by suppressor genes. rpc128-1007 mutation, regardless of the presence of suppressors, also resulted in a weak increase in the expression of ribosome biogenesis genes. mRNA genes that were downregulated by the reduction of Pol III assembly comprise the proteasome complex. In summary, our results provide the regulatory links affected by Pol III assembly that contribute differently to cellular fitness.

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Ex vivo mRNA expression of toll-like receptors during latent tuberculosis infection

BMC Immunology ◽

10.1186/s12865-021-00400-4 ◽

2021 ◽

Vol 22 (1) ◽

Author(s):

Birhan Alemnew ◽

Soren T. Hoff ◽

Tamrat Abebe ◽

Markos Abebe ◽

Abraham Aseffa ◽

...

Keyword(s):

Mrna Expression ◽

Expression Patterns ◽

Fold Change ◽

Mrna Levels ◽

Healthy Children ◽

Toll Like Receptors ◽

Relative Expression ◽

Latently Infected ◽

Latent Tb ◽

Latent Tb Infection

Abstract Background Understanding immune mechanisms, particularly the role of innate immune markers during latent TB infection remains elusive. The main objective of this study was to evaluate mRNA gene expression patterns of toll-like receptors (TLRs) as correlates of immunity during latent TB infection and further infer their roles as potential diagnostic biomarkers. Methods Messenger RNA (mRNA) levels were analysed in a total of 64 samples collected from apparently healthy children and adolescents latently infected with tuberculosis (n = 32) or non-infected (n = 32). Relative expression in peripheral blood of selected genes encoding TLRs (TLR-1, TLR-2, TLR-4, TLR-6 and TLR-9) was determined with a quantitative real-time polymerase chain reaction (qRT-PCR) using specific primers and florescent labelled probes and a comparative threshold cycle method to define fold change. Data were analysed using Graph-Pad Prism 7.01 for Windows and a p-value less than 0.05 was considered statistically significant. Results An increased mean fold change in the relative expression of TLR-2 and TLR-6 mRNA was observed in LTBI groups relative to non-LTBI groups (p < 0.05), whereas a slight fold decrease was observed for TLR-1 gene. Conclusions An increased mRNA expression of TLR-2 and TLR-6 was observed in latently infected individuals relative to those non-infected, possibly indicating the roles these biomarkers play in sustenance of the steady state interaction between the dormant TB bacilli and host immunity.

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The mechanism of increased intestinal palmitic acid absorption and its impact on hepatic stellate cell activation in nonalcoholic steatohepatitis

Scientific Reports ◽

10.1038/s41598-021-92790-z ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Masakazu Hanayama ◽

Yasunori Yamamoto ◽

Hiroki Utsunomiya ◽

Osamu Yoshida ◽

Shuang Liu ◽

...

Keyword(s):

Liver Fibrosis ◽

Portal Vein ◽

Palmitic Acid ◽

Intestinal Absorption ◽

Nonalcoholic Steatohepatitis ◽

Cell Activation ◽

Stellate Cell ◽

Mrna Levels ◽

Nonalcoholic Fatty Liver ◽

Chylomicron Formation

AbstractDietary palmitic acid (PA) promotes liver fibrosis in patients with nonalcoholic steatohepatitis (NASH). Herein, we clarified the intestinal absorption kinetics of dietary PA and effect of trans-portal PA on the activation of hepatic stellate cells (HSCs) involved in liver fibrosis in NASH. Blood PA levels after meals were significantly increased in patients with NASH compared to those in the control. Expression of genes associated with fat absorption and chylomicron formation, such as CD36 and MTP, was significantly increased in the intestine of NASH model rats compared with that in the controls. Plasma levels of glucagon-like peptide-2, involved in the upregulation of CD36 expression, were elevated in NASH rats compared with those in the controls. Furthermore, portal PA levels after meals in NASH rats were significantly higher than those in control and nonalcoholic fatty liver rats. Moreover, PA injection into the portal vein to the liver in control rats increased the mRNA levels associated with the activation of HSCs. Increased intestinal absorption of diet-derived PA was observed in NASH. Thus, the rapid increase in PA levels via the portal vein to the liver may activate HSCs and affect the development of liver fibrosis in NASH.

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Clinical relevance of kallikrein-related peptidase 6 (KLK6) and 8 (KLK8) mRNA expression in advanced serous ovarian cancer

Biological Chemistry ◽

10.1515/hsz-2016-0177 ◽

2016 ◽

Vol 397 (12) ◽

pp. 1265-1276 ◽

Cited By ~ 14

Author(s):

Nancy Ahmed ◽

Julia Dorn ◽

Rudolf Napieralski ◽

Enken Drecoll ◽

Matthias Kotzsch ◽

...

Keyword(s):

Ovarian Cancer ◽

Mrna Expression ◽

Cox Regression ◽

Multivariable Analysis ◽

Progression Free Survival ◽

Predictive Marker ◽

Residual Tumor ◽

Mrna Levels ◽

Serous Ovarian Cancer ◽

Cox Regression Analysis

Abstract Most members of the kallikrein-related peptidase family have been demonstrated to be dysregulated in ovarian cancer and modulate tumor growth, migration, invasion, and resistance to chemotherapy. In the present study, we assessed the mRNA expression levels of KLK6 and KLK8 by quantitative PCR in 100 patients with advanced serous ovarian cancer FIGO stage III/IV. A pronounced correlation between KLK6 and KLK8 mRNA expression (rs = 0.636, p < 0.001) was observed, indicating coordinate expression of both peptidases. No significant associations of clinical parameters with KLK6, KLK8, and a combined score KLK6+KLK8 were found. In univariate Cox regression analysis, elevated mRNA levels of KLK6 were significantly linked with shortened overall survival (OS) (hazard ratio [HR] = 2.07, p = 0.007). While KLK8 values were not associated with patients’ outcome, high KLK6+KLK8 values were significantly associated with shorter progression-free survival (HR = 1.82, p = 0.047) and showed a trend towards significance in the case of OS (HR = 1.82, p = 0.053). Strikingly, in multivariable analysis, elevated KLK6 mRNA values, apart from residual tumor mass, remained an independent predictive marker for poor OS (HR = 2.33, p = 0.005). As KLK6 mRNA and protein levels correlate, KLK6 may represent an attractive therapeutic target for potent and specific inhibitors of its enzymatic activity.

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Birdsong Decreases Protein Levels of FoxP2, a Molecule Required for Human Speech

Journal of Neurophysiology ◽

10.1152/jn.90415.2008 ◽

2008 ◽

Vol 100 (4) ◽

pp. 2015-2025 ◽

Cited By ~ 58

Author(s):

Julie E. Miller ◽

Elizabeth Spiteri ◽

Michael C. Condro ◽

Ryan T. Dosumu-Johnson ◽

Daniel H. Geschwind ◽

...

Keyword(s):

Mrna Expression ◽

Target Genes ◽

Expression Patterns ◽

Vocal Learning ◽

Brain Regions ◽

Motor Deficits ◽

Mrna Levels ◽

Adult Males ◽

Protein Levels ◽

Area X

Cognitive and motor deficits associated with language and speech are seen in humans harboring FOXP2 mutations. The neural bases for FOXP2 mutation-related deficits are thought to reside in structural abnormalities distributed across systems important for language and motor learning including the cerebral cortex, basal ganglia, and cerebellum. In these brain regions, our prior research showed that FoxP2 mRNA expression patterns are strikingly similar between developing humans and songbirds. Within the songbird brain, this pattern persists throughout life and includes the striatal subregion, Area X, that is dedicated to song development and maintenance. The persistent mRNA expression suggests a role for FoxP2 that extends beyond the formation of vocal learning circuits to their ongoing use. Because FoxP2 is a transcription factor, a role in shaping circuits likely depends on FoxP2 protein levels which might not always parallel mRNA levels. Indeed our current study shows that FoxP2 protein, like its mRNA, is acutely downregulated in mature Area X when adult males sing with some differences. Total corticosterone levels associated with the different behavioral contexts did not vary, indicating that differences in FoxP2 levels are not likely attributable to stress. Our data, together with recent reports on FoxP2's target genes, suggest that lowered FoxP2 levels may allow for expression of genes important for circuit modification and thus vocal variability.

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