Integrated metabolism and epigenetic modifications in the macrophages of mice in responses to cold stress.

The negative effects of low temperature can readily induce a variety of diseases. We sought to understand the reasons why cold stress induces disease by studying the mechanisms of fine-tuning in macrophages following cold exposure. We found that cold stress triggers increased macrophage activation accompanied by metabolic reprogramming of aerobic glycolysis. The discovery, by genome-wide RNA sequencing, of defective mitochondria in mice macrophages following cold exposure indicated that mitochondrial defects may contribute to this process. In addition, changes in metabolism drive the differentiation of macrophages by affecting histone modifications. Finally, we showed that histone acetylation and lactylation are modulators of macrophage differentiation following cold exposure. Collectively, metabolism-related epigenetic modifications are essential for the differentiation of macrophages in cold-stressed mice, and the regulation of metabolism may be crucial for alleviating the harm induced by cold stress.

Procyanidin B2 alleviates liver injury caused by cold stimulation through Sonic hedgehog signalling and autophagy.

Procyanidin B2 (PB2), a naturally occurring flavonoid abundant in a wide range of fruits, has been shown to exert antioxidant, anti-inflammatory and anticancer properties. However, the role of PB2 in the prevention of cold stimulation (CS)-induced liver injury. The present study was undertaken to determine the effects of PB2 on liver injury induced by cold stimulation and its potential molecular mechanisms. The present study results showed that treatment with PB2 significantly reduced CS-induced liver injury by alleviating histopathological changes and serum levels of alanine transaminase and aspartate transaminase. Moreover, treatment with PB2 inhibited secretion of inflammatory cytokines and oxidative stress in cold-stimulated mice. PB2 reduced cold stimulation-induced inflammation by inhibiting TLR4/NF-κB and Txnip/NLRP3 signalling. Treatment with PB2 reduced oxidative stress by activating Nrf-2/Keap1, AMPK/GSK3ß signalling pathways and autophagy. Furthermore, simultaneous application of Shh pathway inhibitor cyclopamine proved that PB2 targets the Hh pathway. More importantly, co-treatment with PB2 and cyclopamine showed better efficacy than monotherapy. In conclusion, our findings provide new evidence that PB2 has protective potential against CS-induced liver injury, which might be closely linked to the inhibition of Shh signalling pathway.

O-GlcNAc / Akt pathway regulates glucose metabolism and reduces apoptosis in liver of piglets with acute cold stress.

Cold stress is one of the serious factors restricting the development of animal husbandry in cold areas. Cold exposure can easily lead to cold stress, slow growth and even death of newborn animals. O-GlcNAcylation modification can act as type of "stress receptor" and"nutrition sensor" in a variety of stress responses, however, it is not clear how O-GlcNAcylation can regulate glucose metabolism in the liver of piglets under cold stress. In this study, piglets 21 days of age were exposed to 4 °C for 4 h or 8 h in a phytotron. Serum cortisol and other stress hormones were used to assess body status to establish a cold stress piglet model. The changes of glycogen in liver were detected by PAS. FDP and PA were also measured to study the glycolysis level of liver. To characterize potential mechanisms of O-GlcNAcylation on the livers of cold stress piglets, AKT, GSK3ß, GS, PFKFB2, AS160 and their corresponding phosphorylation were determined by Western blotting. Results show O-GlcNAcylation increased and apoptosis levels increased in the liver following cold exposure during excessive CORT or metabolic dysfunction. It is suggested that the acute cold exposure of piglets induced a sequential change in the level of O-GlcNAcylation, which may be one of the factors mediating liver cell apoptosis and glucose metabolism regulation by the O-GlcNAc/AKT pathway. These findings provide new insight into the mechanisms of the cold stress response, which can facilitate the development of new strategies to combat the effects of hypothermia.

Down-regulation of miR-383-5p suppresses apoptosis in oxidative stress rat hepatocytes by targeting Bcl2.

miRNAs are a class of small non-coding RNAs that are involved in various biological processes. In the preliminary work of the laboratory, found that miR-383-5p was down-regulated in the liver tissue of acute cold stress rats and has been shown to be an important regulatory factor in tumour proliferation, but there are very few studies involving the mediation of cold stress in rat liver tissues. Therefore, the purpose of this study was to determine the effect of miR-383-5p on the livers of cold stress rats by simulating the cold stress state of rat liver tissues in vitro using H2 O2 to induce rat hepatocyte oxidative stress. The results showed that MDA content, Caspase 3 and Cyto C protein levels increased significantly; GPx activity and SOD1 protein levels decreased significantly and miR-383-5p expression was significantly down-regulated in rat liver tissues after cold stress. Different concentrations of H2 O2 was added to rat hepatocytes, and the results showed that the expression of miR-383-5p, the ROS level, and the apoptosis rate in rat hepatocytes was increased significantly in a concentration-dependent fashion. Transfection of miR-383-5p inhibitor revealed that the apoptosis rate of rat hepatocytes, and the protein level of apoptosis-related protein Caspase 3 were reduced; the results of the dual-luciferase reporter gene assay showed that miR-383-5p targeted regulation of Bcl2. The results suggested that the expression of miR-383-5p was up-regulated in oxidative stress rat hepatocytes and may aggravate the apoptosis of rat hepatocytes induced by targeting inhibition of Bcl2 translation.

Identification, functional prediction, and key lncRNA verification of cold stress-related lncRNAs in rats liver.

Cold stimulation reduces the quality of animal products and increases animal mortality, causing huge losses to the livestock industry in cold regions. Long non-coding RNAs (lncRNAs) take part in many biological processes through transcriptional regulation, intracellular material transport, and chromosome remodeling. Although cold stress-related lncRNAs have been reported in plants, no research is available on the characteristic and functional analysis of lncRNAs after cold stress in rats. Here, we built a cold stress animal model firstly. Six SPF male Wistar rats were randomly divided to the acute cold stress group (4 °C, 12 h) and the normal group (24 °C, 12 h). lncRNA libraries were constructed by high-throughput sequencing (HTS) using rat livers. 2,120 new lncRNAs and 273 differentially expressed (DE) lncRNAs were identified in low temperature environments. The target genes of DElncRNA were predicted by cis and trans, and then functional and pathway analysis were performed to them. GO and KEGG analysis revealed that lncRNA targets were mainly participated in the regulation of nucleic acid binding, cold stimulation reaction, metabolic process, immune system processes, PI3K-Akt signaling pathway and pathways in cancer. Next, a interaction network between lncRNA and its targets was constructed. To further reveal the mechanism of cold stress, DElncRNA and DEmRNA were extracted to reconstruct a co-expression sub-network. We found the key lncRNA MSTRG.80946.2 in sub-network. Functional analysis of key lncRNA targets showed that targets were significantly enriched in fatty acid metabolism, the PI3K-Akt signaling pathway and pathways in cancer under cold stress. qRT-PCR confirmed the sequencing results. Finally, hub lncRNA MSTRG.80946.2 was characterized, and verified its relationship with related mRNAs by antisense oligonucleotide (ASO) interference and qRT-PCR. Results confirmed the accuracy of our analysis. To sum up, our work was the first to perform detailed characterization and functional analysis of cold stress-related lncRNAs in rats liver. lncRNAs played crucial roles in energy metabolism, growth and development, immunity and reproductive performance in cold stressed rats. The MSTRG.80946.2 was verified by network and experiments to be a key functional lncRNA under cold stress, regulating ACP1, TSPY1 and Tsn.

Metabonomics analysis of Zi goose follicular granulosa cells using ENO1 gene expression interference.

The Zi goose is native to North-east China and is noted for its high egg production. Alpha enolase (ENO1) is a glycolytic enzyme which functions as a plasminogen receptor in follicular granulosa cells (FGCs), with several studies showing that FGCs can support follicular development. By transfecting the ENO1 interfering plasmid (shRNA) into FGCs, ENO1 expression in these cells was downregulated, suggesting the successful knock-down of ENO1 in these cells. In this knock-down model, we detected 13 metabolites from FGCs using LC/MS. When compared with the non-coding shRNA (NC) group, the lower level metabolites were (R)-(+)-citronellic acid, altretamine, 3-hydroxycaproic acid, heptadecanoic acid, cholecalciferol vitamin D3, indole, benzoic acid, capric acid, caffeic acid, azelaic acid, 3,4-dihydroxyhydrocinnamic acid and cholic acid, while oleic acid was detected at high levels. To further examine the results of metabolomics, six key metabolites were verified by gas chromatography-mass spectrometry (GC-MS). We found that vitamin D3, indole, benzoic acid, capric acid and cholic acid were significantly downregulated in the shRNA group, while oleic acid was significantly upregulated. This observation was consistent with the metabolomics data. Through these studies, we found that decreased ENO1 levels altered certain metabolite levels in FGCs.

Cortisol Excess-Mediated Mitochondrial Damage Induced Hippocampal Neuronal Apoptosis in Mice Following Cold Exposure.

Cold stress can induce neuronal apoptosis in the hippocampus, but the internal mechanism involving neuronal loss induced by cold stress is not clear. In vivo, male and female C57BL/6 mice were exposed to 4 °C, 3 h per day for 1 week. In vitro, HT22 cells were treated with different concentrations of cortisol (CORT) for 3 h. In vivo, CORT levels in the hippocampus were measured using ELISA, western blotting, and immunohistochemistry to assess the neuronal population and oxidation of the hippocampus. In vitro, western blotting, immunofluorescence, flow cytometry, transmission electron microscopy, and other methods were used to characterize the mechanism of mitochondrial damage induced by CORT. The phenomena of excessive CORT-mediated oxidation stress and neuronal apoptosis were shown in mouse hippocampus tissue following cold exposure, involving mitochondrial oxidative stress and endogenous apoptotic pathway activation. These processes were mediated by acetylation of lysine 9 of histone 3, resulting in upregulation involving Adenosine 5'-monophosphate (AMP)-activated protein kinase (APMK) phosphorylation and translocation of Nrf2 to the nucleus. In addition, oxidation in male mice was more severe. These findings provide a new understanding of the underlying mechanisms of the cold stress response and explain the apoptosis process induced by CORT, which may influence the selection of animal models in future stress-related studies.

Rno-miR-425-5p targets the DLST and SLC16A1 genes to reduce liver damage caused by excessive energy mobilization under cold stress.

MicroRNAs (miRNAs) are a class of single-stranded non-coding small RNA molecules, which participate in the regulation of many physiological processes, and play a crucial role in cancer, metabolism and other processes. Rno-miR-425-5p has been shown to play a role in the response to cold stress. To explore the mechanism by which rno-miR-425-5p regulates the response to cold stress, we analysed the candidate target genes of rno-miR-425-5p. After verification in rat hepatocyte BRL cells and in rat liver tissue, we identified several target genes that were altered in expression in response to cold stress. In rat liver tissue, the expression of rno-miR-425-5p was significantly increased and the expression levels of target genes DLST and SLC16A1 were decreased under cold stress. The miRNA and mRNA levels were analysed by quantitative real-time PCR and the protein levels were detected by Western blot analysis. Combined with the results of bioinformatic analysis, we concluded that rno-miR-425-5p reduced the expression of DLST and SLC16A1, inhibiting energy release from the tricarboxylic acid cycle and preventing the liver from being injured by excessive energy mobilization.

Effects of Cold-inducible RNA-binding Protein (CIRP) on Liver Glycolysis during Acute Cold Exposure in C57BL/6 Mice.

Cold-inducible RNA-binding protein (CIRP) is a stress-responsive protein involved in several signal transduction pathways required for cellular function, which are associated with apoptosis and proliferation. The present study aimed to investigate the possible effects of CIRP-mediated regulation of glucose metabolism in the liver following acute cold exposure. The livers and serum of male C57BL/6 mice were collected following cold exposure at 4 °C for 0 h, 2 h, 4 h, and 6 h. Glucose metabolic markers and the expression of glucose metabolic-related proteins were detected in the liver. Acute cold exposure was found to increase the consumption of glycogen in the liver. Fructose-1,6-diphosphate (FDP) and pyruvic acid (PA) were found to show a brief increase followed by a sharp decrease during cold exposure. Anti-apoptotic protein (Bcl-2) expression was upregulated. CIRP protein expression displayed a sequential increase with prolonged acute cold exposure time. Acute cold exposure also increased the level of protein kinase B (AKT) phosphorylation, and activated the AKT-signaling pathway. Taken together, these findings indicate that acute cold exposure increased the expression of CIRP protein, which regulates mouse hepatic glucose metabolism and maintains hepatocyte energy balance through the AKT signaling pathway, thereby slowing the liver cell apoptosis caused by cold exposure.

Regulating glycolysis, the TLR4 signal pathway and expression of RBM3 in mouse liver in response to acute cold exposure.

At low temperatures, the liver increases glucose utilization and expresses RNA-binding motif 3 (RBM3) to cope with cold exposure. In this study, the expression of heat shock protein 70 (HSP70), Toll-like receptor 4 (TLR4), bone marrow differentiation factor 88 (MYD88), and phosphorylated nuclear factor-κB (NF-κB) was consistent with fluctuations in insulin in fasted cold-exposed mice. We also found up-regulation of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) in acute cold exposure with a decrease in core body temperature. RBM3 transcription and translation were activated 2 h after cold exposure. The anti-apoptotic factor Bcl-2/Bax ratio also increased, while expression of apoptosis factors: cleaved caspase-3, cleaved poly(ADP-ribose)polymerase 1 (PARP-1) and cytochrome-c (Cyt-c) was unchanged. Liver glycogen was depleted after 2 h of cold exposure, and blood glucose decreased after 4 h. Glycogen synthase kinase 3ß (GSK3ß) phosphorylation continued to increase to promote hepatic glycogen synthesis. We found a high level of protein kinase B (AKT) phosphorylation after 6 h of cold exposure. In addition, we demonstrated that after cold exposure for 2 h, in the liver, continued phosphorylation of fructose-2,6-diphosphate (PFKFB2) and decreased accumulation of glycogen intermediates fructose-1,6-diphosphate (FDP) and pyruvic acid (PA). In summary, the liver responds to cold exposure through a number of different pathways, including activation of HSP70/TLR4 signaling pathways, up-regulation of RBM3 expression, and increased glycolysis and glycogen synthesis. We propose a possible signaling pathway in which regulation of RBM3 expression by the liver affects the AKT metabolic signaling pathway. Lay summary In response to changes in ambient temperature, mice regulate global metabolism and gene expression through hormones. This study focused on the effects of environmental hypothermia on molecular pathways of glucose metabolism in the liver, which is the important metabolic organ in mice. This provides a basis for further study of mice against cold exposure damage.

Lactobacillus bulgaricus or Lactobacillus rhamnosus Suppresses NF-κB Signaling Pathway and Protects against AFB1-Induced Hepatitis: A Novel Potential Preventive Strategy for Aflatoxicosis?

Aflatoxin B1 (AFB1), a mycotoxin found in food and feed, is immunotoxic to animals and poses significant threat to the food industry and animal production. The primary target of AFB1 is the liver. To overcome aflatoxin toxicity, probiotic-mediated detoxification has been proposed. In the present study, to investigate the protective effects and molecular mechanisms of Lactobacillus bulgaricus or Lactobacillus rhamnosus against liver inflammatory responses to AFB1, mice were administered with AFB1 (300 µg/kg) and/or Lactobacillus intragastrically for 8 weeks. AML12 cells were cultured and treated with AFB1, BAY 11-7082 (an NF-κB inhibitor), and different concentrations of L. bulgaricus or L. rhamnosus. The body weight, liver index, histopathological changes, biochemical indices, cytokines, cytotoxicity, and activation of the NF-κB signaling pathway were measured. AFB1 exposure caused changes in liver histopathology and biochemical functions, altered inflammatory response, and activated the NF-κB pathway. Supplementation of L. bulgaricus or L. rhamnosus significantly prevented AFB1-induced liver injury and alleviated histopathological changes and inflammatory response by decreasing NF-κB p65 expression. The results of in vitro experiments revealed that L.rhamnosus evidently protected against AFB1-induced inflammatory response and decreased NF-κB p65 expression when compared with L. bulgaricus. These findings indicated that AFB1 exposure can cause inflammatory response by inducing hepatic injury, and supplementation of L. bulgaricus or L. rhamnosus can produce significant protective effect against AFB1-induced liver damage and inflammatory response by regulating the activation of the NF-κB signaling pathway.

Effects of Acute Cold Stress on Liver O-GlcNAcylation and Glycometabolism in Mice.

Protein O-linked ß-N-acetylglucosamine glycosylation (O-GlcNAcylation) regulates many biological processes. Studies have shown that O-GlcNAc modification levels can increase during acute stress and suggested that this may contribute to the survival of the cell. This study investigated the possible effects of O-GlcNAcylation that regulate glucose metabolism, apoptosis, and autophagy in the liver after acute cold stress. Male C57BL/6 mice were exposed to cold conditions (4 °C) for 0, 2, 4, and 6 h, then their livers were extracted and the expression of proteins involved in glucose metabolism, apoptosis, and autophagy was determined. It was found that acute cold stress increased global O-GlcNAcylation and protein kinase B (AKT) phosphorylation levels. This was accompanied by significantly increased activation levels of the glucose metabolism regulators 160 kDa AKT substrate (AS160), 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 2 (PFKFB2), and glycogen synthase kinase-3ß (GSK3ß). The levels of glycolytic intermediates, fructose-1,6-diphosphate (FDP) and pyruvic acid (PA), were found to show a brief increase followed by a sharp decrease. Additionally, adenosine triphosphate (ATP), as the main cellular energy source, had a sharp increase. Furthermore, the B-cell lymphoma 2(Bcl-2)/Bcl-2-associated X (Bax) ratio was found to increase, whereas cysteine-aspartic acid protease 3 (caspase-3) and light chain 3-II (LC3-II) levels were reduced after acute cold stress. Therefore, acute cold stress was found to increase O-GlcNAc modification levels, which may have resulted in the decrease of the essential processes of apoptosis and autophagy, promoting cell survival, while altering glycose transport, glycogen synthesis, and glycolysis in the liver.

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.

Identification of cold-responsive miRNAs in rats by deep sequencing.

miRNA is an endogenously noncoding sRNA, which is involved in post-transcription gene expression regulation of growth, tumor development and stress survival. As a biological marker, miRNA has been used for the early diagnosis of diseases and the evaluation of some physiological state. We constructed two small RNA libraries with the serums of rats treated or not with cold conditions (4℃ for 12h) by deep sequencing, in order to understand the miRNAs' expressions of cold-exposed rats and find new cold-responsive biological markers. 485 conserved miRNAs and 287 novel miRNAs were identified in the two libraries by comparing to the known miRNAs of rat in miRBase 21.0 Differential expression analysis showed that 56 conserved miRNAs and 3 novel miRNAs were expressed differentially in low ambient temperature. The qRT-PCR results confirmed that rno-miR-151-3p, rno-miR-210-3p, rno-miR-425-5p, rno-miR-383-5p, rno-miR-92a-3p, rno-miR-98-5p and rno-miR-328a-3p decreased significantly in rats serums treated with cold exposure. The expressions of the 7 miRNAs changed significantly in cold-exposed rats' livers too. rno-miR-383-5p decreased significantly, but all the others increased significantly. Thus, the 7 miRNAs were considered as cold-responsive miRNAs of rat. 670 target genes of the 7 cold-responsive miRNAs were predicted. KEGG analysis showed that they were enriched in 28 pathways and most of them were enriched by metabolic pathway. Overall, the results of this study suggest an important role for selected miRNA's in the response to cold stress.

[The research of the progesterone and estrogen secretion level and the FSHR, LHR gene quantitative in granular cells cultured in vitro in different time of laying hens].

OBJECTIVE: To research the hormone secretion levels of progesterone and estrogen and the gene expression levels of two go-nadotropin receptors follicle stimulating hormone receptor (FSHR) and luteinizing hormone receptor (LHR) in granular cells of laying hen, and the effect of culture time on the levels of hormone secretion and expression of related receptor gene in granulosa cells was inferred. METHODS: The experiment using the method of cells culture in vitro, the granular cells supernatants of hens were collected at 0 h, 24 h, 48 h, 72 h, 96 h, the progesterone and estrogen concentrations in cell supernatants were determined by ELISA kits, and detected the expression of FSHR and LHR gene in granular cells by real-time fluorescent quantitative PCR. RESULTS: The results showed that the progesterone and estrogen secretion reduced in the early culture of 0 h~48 h(P < 0.05), with the culture time increases to 72 h, the secretion of two hormones began to in-crease, and reaching the level of the initial level of culture. When the cells were cultured to 96 h, the rogesterone and estrogen secretion was reduced again. The lower levels of FSHR and LHR mRNA expression in granular cells appeared with the increase of culture time, compared with the group of cell culture to 0 h, the mRNA expression levels of each groups reduced obviously(P < 0.05). CONCLUSIONS: The amount of progesterone and estrogen in the cultured follicular granulosa cells decreased with the increase of in vitro culture time, and then increased. This might be related to the growth state of cells cultured in vitro. But on the whole, with the extension of the training time, the secretion of proges-terone and estrogen in the cells decreased. This may be related to the decreased expression of the FSHR and LHR genes in the two go-nadotropin receptors.

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), 11ß-hydroxysteroid dehydrogenase 2 (11ß-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)-1ß. 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.

Hsp70 suppresses apoptosis of BRL cells by regulating the expression of Bcl-2, cytochrome C, and caspase 8/3.

During cold stress, liver cells undergo apoptotic injury as a result of oxidative stress. Heat shock 70 kDa protein (Hsp70) is a protein involved in modulating a variety of physiological processes, including stress responses, proliferation, and apoptosis. In addition, Hsp70 regulates apoptotic signaling pathways in different manners, promoting or suppressing apoptosis. In this study, we investigated the effects of Hsp70 overexpression on hydrogen peroxide (H2O2)-induced apoptosis of Buffalo rat liver (BRL) cells and the underlying mechanisms of these effects. Our results show that in comparison with the control group, Hsp70 overexpression displayed increased protein levels of Bcl-2, and decreased cytochrome c (Cyt c), cleaved caspase 3, and cleaved caspase 8, but no apparent differences were found in levels of Bax. Furthermore, Hsp70 overexpression significantly suppresses the amount of apoptotic cells. Such findings indicate that overexpression of Hsp70 inhibits H2O2-mediated activation of caspase 8 and caspase 3, upregulates the expression of Bcl-2 which is a known anti-apoptotic protein, and decreases the release of Cyt c from the mitochondria into the cytoplasm, collectively decreasing cell apoptosis.

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.

ß-Hydroxybutyric sodium salt inhibition of growth hormone and prolactin secretion via the cAMP/PKA/CREB and AMPK signaling pathways in dairy cow anterior pituitary cells.

ß-hydroxybutyric acid (BHBA) regulates the synthesis and secretion of growth hormone (GH) and prolactin (PRL), but its mechanism is unknown. In this study, we detected the effects of BHBA on the activities of G protein signaling pathways, AMPK-α activity, GH, and PRL gene transcription, and GH and PRL secretion in dairy cow anterior pituitary cells (DCAPCs). The results showed that BHBA decreased intracellular cAMP levels and a subsequent reduction in protein kinase A (PKA) activity. Inhibition of PKA activity reduced cAMP response element-binding protein (CREB) phosphorylation, thereby inhibiting GH and PRL transcription and secretion. The effects of BHBA were attenuated by a specific Gαi inhibitor, pertussis toxin (PTX). In addition, intracellular BHBA uptake mediated by monocarboxylate transporter 1 (MCT1) could trigger AMPK signaling and result in the decrease in GH and PRL mRNA translation in DCAPCs cultured under low-glucose and non-glucose condition when compared with the high-glucose group. This study identifies a biochemical mechanism for the regulatory action of BHBA on GH and PRL gene transcription, translation, and secretion in DCAPCs, which may be one of the factors that regulate pituitary function during the transition period in dairy cows.

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.