Jingru Guo

Progress in the biological function of alpha-enolase

Alpha-enolase (ENO1), also known as 2-phospho-D-glycerate hydrolase, is a metalloenzyme that catalyzes the conversion of 2-phosphoglyceric acid to phosphoenolpyruvic acid in the glycolytic pathway. It is a multifunctional glycolytic enzyme involved in cellular stress, bacterial and fungal infections, autoantigen activities, the occurrence and metastasis of cancer, parasitic infections, and the growth, development and reproduction of organisms. This article mainly reviews the basic characteristics and biological functions of ENO1.

Selection of Reliable Reference Genes for Real-time qRT-PCR Analysis of Zi Geese (Anser anser domestica) Gene Expression.

Zi geese (Anser anser domestica) belong to the white geese and are excellent layers with a superior feed-to-egg conversion ratio. Quantitative gene expression analysis, such as Real-time qRT-PCR, will provide a good understanding of ovarian function during egg-laying and consequently improve egg production. However, we still don’t know what reference genes in geese, which show stable expression, should be used for such quantitative analysis. In order to reveal such reference genes, the stability of seven genes were tested in five tissues of Zi geese. Methodology/Principal Findings: The relative transcription levels of genes encoding hypoxanthine guanine phosphoribosyl transferase 1 (HPRT1), β-actin (ACTB), β-tubulin (TUB), glyceraldehyde-3-phosphate-dehydrogenase (GADPH), succinate dehydrogenase flavoprotein (SDH), 28S rRNA (28S) and 18S rRNA (18S) have been quantified in heart, liver, kidney, muscle and ovary in Zi geese respectively at different developmental stages (1 d, 2, 4, 6 and 8 months). The expression stability of these genes was analyzed using geNorm, NormFinder and BestKeeper software. Conclusions: The expression of 28S in heart, GAPDH in liver and ovary, ACTB in kidney and HPRT1 in muscle are the most stable genes as identified by the three different analysis methods. Thus, these genes are recommended for use as candidate reference genes to compare mRNA transcription in various developmental stages of geese.

The Favored Mechanism for Coping with Acute Cold Stress: Upregulation of miR-210 in Rats

Background/Aims: The main aim of this study was to determine the mechanisms by which rno-miR-210-3p affects changes in gene expression, metabolism, apoptosis and proliferation of cells under acute cold stress (ACS) conditions. Methods: The treatment group (n=6, weight 340±20 g) was exposed to ACS (temperature 4±0.5°C, relative humidity 45±0.5%) and the control group (n=6, weight 340±20 g) to normal temperature (NT) (temperature 24±0.5°C, relative humidity 45±0.5%). Rat liver samples were collected for qRT-PCR and western blot analyses to detect relative expression of rno-miR-210-3p, ISCU, Rap1b, ATP1b1, GPD1, E2F3, RAD52, PSMB6 and GPD2. For cell experiments, 100 pmol/dish rno-miR-210-3p mimic and 150 pmol/dish rno-miR-210-3p inhibitor were used. Mitochondrial glucose flux and glycolysis were measured using the XFe24 Extracellular Flux Analyzer. Cells were collected for apoptosis analysis 24 h after transfection and proliferation was quantified using the WST-1 Cell Proliferation and Cytotoxicity Assay Kit (Beyotime, Shanghai, China), according to the manufacturerʹs instructions. Results: In the rat experiment, expression of rno-miR-210-3p under ACS was increased sharply while ISCU, E2F3, RAD52, and PSMB6 levels declined, along with protein expression of ISCU and PSMB6. In cell experiments, ISCU, Rap1b, ATP1b1, GPD1, E2F3, RAD52, PSMB6 and GPD2 genes were downregulated while ISCU and PSMB6 protein expression decreased with upregulation of rno-miR-210-3p. Conversely, in response to decreased rno-miR-210-3p expression, ISCU, E2F3, RAD52, PSMB6 and GPD2 genes were upregulated, in addition to ISCU and PSMB6 proteins. Upregulation of miR-210 inhibited cell proliferation and induced cell death whereas its downregulation promoted cell proliferation. Upregulation or downregulation of miR-210 promoted glycolysis and mitochondrial respiration of BRL cells. However, downregulation of miR-210 caused acid production in cells. Conclusion: Expression of rno-miR-210-3p is significantly increased under ACS. Upregulation of rno-miR-210-3p inhibits the expression of ISCU, Rap1b, ATP1b1, GPD1, E2F3, RAD52, PSMB6 and GPD2 genes, promotes glycolysis of liver and enhances the mitochondrial respiratory capacity of cells, but may also cause cell death. Our findings collectively indicate that regulation of rno-miR-210-3p is a preferential mechanism of choice used by the body to cope with ACS.

Impact of prenatal cold stress on placental physiology, inflammatory response, and apoptosis in rats

Prenatal cold stress is one of the earliest factors affecting mammalian health, and is associated with neonatal growth retardation and immune dysfunction, thus increasing disease susceptibility. The mechanisms underlying these observations remain unclear; hence, the objective of this study was to elucidate placental responses to cold stress. 60 maternal rats were randomly allocated to either stressed (n = 30) or non-stressed (control, n = 30) treatment conditions and 30 pubs (n=15) were used for the pups analysis. We found that maternal exposure to cold stress resulted in decreased body temperature, increased food intake without body weight gain, and a high level of plasma corticosterone (CORT) between gestational day (GD) 14 and GD21. In addition, gestation cold stress induced the placental expression of heat shock protein 70 (HSP70), IκBα, glucocorticoid receptor (GR), mineralocorticoid receptor (MR), 11β-hydroxysteroid dehydrogenase 2 (11β-HSD2), interferon (IFN) regulatory factor 3 (IRF3), Caspase-3 proteins and altered the ratio of B-cell lymphoma-extra large (Bcl-xL) to Bcl-associated x (Bax) proteins on gestational GD15, GD17, GD19, and GD21, also resulted in the production of interleukin (IL)-1β. Next, gestational cold stress provoked a decrease in plasma GH levels of 21-day-old offspring, and the body weights of offspring were have no differences from postnatal day (PD) 1–21. Taken together, our results indicate that gestational cold stress induces placental apoptosis and the activation of NF-kB via HSP70/TLR4/NF-κB signaling pathways in the placenta, these changes may affect placental function and fetus development.

Prenatal cold stress: Effect on maternal hippocampus and offspring behavior in rats

HighlightsCold stress inhibits the phosphorylation of ERK1/2 in the hippocampus.Cold stress induces RBM3 or BDNF protein expression.Cold stress induces phosphorylation of P65 on Ser536.Prenatal cold stress showed a reduction in male and female offspring anxiety‐like behavior. ABSTRACT In mammals, environmental factors including cold stress exert dramatic effects on adult health during late gestation, the cold stress response refers to an organism’s response to cold. Indeed, cells and organs, including the hippocampus, are coordinated to respond to prevent hypothermia. The hippocampus act as an important brain structure that regulates the activity of the hypothalamic–pituitary–adrenal (HPA) axis, and suppress the stress reaction through feedback regulation of the HPA axis. To evaluate the response of the hippocampus during prenatal cold stress, we established a prenatal cold stress rat model. The molecular and signaling pathways responsible for the hippocampus cold exposure response were investigated. We assessed the glucocorticoid receptor, mineralocorticoid receptor, brain‐derived neurotrophic factor (BDNF), RNA‐binding motif protein 3 (RBM3), heat shock protein 70, protein expression, and extracellular signal‐regulated protein kinases 1 and 2 (ERK1/2) and nuclear factor‐kappa B pathways. Male and female offspring behavior were evaluated. Cold stress reduced the BDNF level in the maternal hippocampus in contrast to the increase in RBM3. BDNF has been shown to induce and RBM3 inhibits ERK phosphorylation. We measured p‐ERK1/2 and showed low‐level phosphorylation in the hippocampus after cold stress. Furthermore, we demonstrated that cold stress enhanced phosphorylation of P65 on Ser536, and led to apoptosis of the hippocampus in a caspase 3‐independent manner. Behavioral tests were performed on pubescent male and female offspring, both of which showed evidence of reduced anxiety‐like behavior. In summary, a more thorough understanding of these mechanisms may lead to maternal intervention that can reverse the damage of prenatal stressors or prevent the damage altogether and improve the physical quality of neonatal rats.

Effect of adenovirus-mediated up-regulation of α-enolase gene products on follicle-stimulating hormone receptor mRNA and luteinizing hormone receptor mRNA of granular cells from goose F1 follicles.

Enolases are glycolytic enzymes in the glycolytic pathway. In order to evaluate the effect of ENO1 on follicle-stimulating hormone receptor (FSHR) mRNA and luteinizing hormone receptor (LHR) mRNA of primary granular cell from goose F1 follicles, the recombinant plasmid adenovirus carrying ENO1 were constructed and infected the primary culture granular cells. The granular cells were randomly divided into three groups: recombinant adenovirus infected (pAd-CMV-ENO1), empty vector infected (pAd-CMV-Null) and no virus (mock control). The expression levels of FSHR mRNA and LHR mRNA of granular cells were examined by qRT-PCR. The results showed the group pAd-CMV-ENO1 had significantly higher FSHR mRNA expression levels than the other two groups (P < 0.05), but had significantly lower LHR mRNA expression levels than the other two groups (P < 0.05). The results suggested that ENO1 could improve the combination rate between FSH and FSHR to accelerate the proliferation and differentiation and steroidogenesis in poultry gonadal tissues.

GABAB receptor mediate hippocampal neuroinflammation in adolescent male and female mice after cold expose

Stress induces many non-specific inflammatory responses in the mouse brain, especially during adolescence. Although the impact of stress on the brain has long been reported, the effects of cold stress on hippocampal neuroinflammation in adolescent mice are not well understood; furthermore, whether these effects are gender specific are also not well established. Adolescent male and female C57BL/6 mice were exposed to 4 °C temperatures for 12 h, after which behavior was assessed using the open field test. Using western blotting and immunohistochemistry we also assessed glial cell numbers and microglial activation, as well as inflammatory cytokine levels and related protein expression levels. We found that in mice subjected to cold stress: 1) There were significant behavioral changes; 2) neuronal nuclei densities were smaller and total cell numbers were significantly decreased; 3) nuclear factor (NF)-κB and phosphorylated AKT were upregulated; 4) pro-inflammatory cytokines such as interleukin-6 and tumor necrosis factor-α were also upregulated; and 5) microglia were activated, while glial fibrillary acid protein and ionized calcium-binding adapter molecule 1 protein expression increased. Taken together, these results indicate that cold stress induces pro-inflammatory cytokine upregulation that leads to neuroinflammation and neuronal apoptosis in the hippocampi of adolescent mice. We believe that these effects are influenced by a GABAB/Rap1B/AKT/NF-κB pathway. Finally, male mice were more sensitive to the effects of cold stress than were female mice.