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The sesquiterpene α-humulene is an important plant natural product, which has been used in the pharmaceutical industry due to its anti-inflammatory and anticancer activities. Although phytoextraction and chemical synthesis have previously been applied in α-humulene production, the low efficiency and high costs limit the development. In this study, Yarrowia lipolytica was engineered as the robust cell factory for sustainable α-humulene production. First, a chassis with high α-humulene output in the cytoplasm was constructed by integrating α-humulene synthases with high catalytic activity, optimizing the flux of mevalonate and acetyl-CoA pathways. Subsequently, the strategy of dual cytoplasmic-peroxisomal engineering was adopted in Y. lipolytica; the best strain GQ3006 generated by introducing 31 copies of 12 different genes could produce 2280.3± 38.2 mg/l (98.7 ± 4.2 mg/g dry cell weight) α-humulene, a 100-fold improvement relative to the baseline strain. To further improve the titer, a novel strategy for downregulation of squalene biosynthesis based on Cu

Whiteleg shrimp is a widely cultured crustacean, but frequent disease outbreaks have decreased production and caused significant losses. Toll-like receptors (TLRs) comprise a large innate immune family that is involved in the innate immune response. However, understanding of their regulatory mechanism is limited. In this study, PacBio sequencing and Illumina sequencing were applied to the gill and hepatopancreas tissues of whiteleg shrimp and an integrated transcript gene set was established. The upregulation of Toll1, Toll2 and Toll3 transcripts in the hepatopancreas tissue of whiteleg shrimp after iridescent virus infection implies that these proteins are involved in the immune response to the virus; simultaneously, the TRAF6 and relish transcripts in the Toll pathway were also upregulated, implying that the Toll pathway was activated. We predicted the three-dimensional structure of the five Toll proteins in whiteleg shrimp and humans and constructed a phylogenetic tree of the Toll protein family. In addition, there was a large discrepancy of Toll1 between invertebrates and vertebrates, presumably because of the loss of Toll1 protein sequence during the evolution process from invertebrates to vertebrates. Our research will improve the cognition of Toll protein family in invertebrates in terms of evolution, structure and function and provide theoretical guidance for researchers in this field.

It has been reported that chemokine CX3 CL1 can regulate various tumours by binding to its unique receptor CX3 CR1. However, the effect of CX3 CL1-CX3 CR1 on the lung adenocarcinoma and lung squamous cell carcinoma is still unclear. Here, we showed that CX3 CL1 can further invasion and migration of lung adenocarcinoma A549 and lung squamous cell carcinoma H520. In addition, Western blot and immunofluorescence test indicated CX3 CL1 up-regulated the phosphorylation level of cortactin, which is a marker of cell pseudopodium. Meanwhile, the phosphorylation levels of c-Src and c-Abl, which are closely related to the regulation of cortactin phosphorylation, are elevated. Nevertheless, the src/abl inhibitor bosutinib and mutations of cortactin phosphorylation site could inhibit the promotion effect of CX3 CL1 on invasion and migration of A549 and H520. Moreover, these results of MTT, Hoechst staining and Western blot suggested that CX3 CL1 had no effect on the proliferation and apoptosis of A549 and H520 in vitro. The effects of CX3 CL1 were also verified by the subcutaneous tumour formation in nude mice, which showed that it could promote proliferation and invasion of A549 in vivo. In summary, our results indicated that CX3 CL1 furthered invasion and migration in lung cancer cells partly via activating cortactin, and CX3 CL1 may be a potential molecule in regulating the migration and invasion of lung cancer.

Backgroud Pesticides are widely used to control insect infestation and weeds in agriculture. However, concerns about the pesticide residues in agricultural products have been raised in recent years because of public interest in health and food quality and safety. Thus, rapid, convenient, and accurate analytical methods for the detection and quantification of pesticides are urgently required. Results A nanohybrid system composed of gold nanoparticles (AuNPs) and tetrakis(N-methyl-4-pyridiniumyl) porphyrin (TMPyP) was used as an optical probe for the detection and quantification of five pesticides (Paraquat, Dipterex, Dursban, methyl thiophanate and Cartap). The method is based on the aggregation effect of pesticides on the carboxyl group modified by AuNPs. Subsequently, with the help of particle swarm optimization-optimized sample weighted least squares-support vector machine (PSO-OSWLS-SVM), all the pesticides could be successfully quantified. In addition, partial least squares discriminant analysis (PLS-DA) was applied and the five pesticides were satisfactorily recognized based on data array obtained from the ultraviolet visible (UV-visible) spectra of AuNP-TMPyP complex. Furthermore, the quantitative and qualitative analysis of the five pesticides could be also achieved in the complex real samples, in which all the relative standard deviations (RSDs) were less than 0.3‰ and all the linear absolute correlation coefficients were more than 0.9990. Furthermore, recognition rate of the training set and the prediction set based on multiplicative scatter correction (MSC), or second-order derivative (2nd derivative) UV-visible spectra in PLS-DA model could reach 100%. Conclusion This method was successfully applied for the rapid and accurate determination of multicomponent pesticide residues in real food samples. © 2020 Society of Chemical Industry.

This paper investigated the feasibility of data fusion of near-infrared (NIR) and fluorescence spectroscopy for rapid analysis of cheap vegetable oils in Chinese Camellia oleifera Abel. (COA) oil. Because practical frauds usually involve adulterations of multiple known and unknown cheap oils, traditional analytical methods aimed at detecting one or more known adulterants are insufficient to identify adulterated COA oil. Therefore, untargeted analysis was performed by developing class models of pure COA oil using robust one-class partial least squares (OCPLS). RESULTS The most accurate OCPLS model was obtained with fusion of standard normal variate (SNV)-NIR and SNV?fluorescence spectra with sensitivity of 0.954 and specificity of 0.91. Robust OCPLS could detect adulterations with 2% (w/w) or more cheap oils, including rapeseed oil, sunflower seed oil, corn oil and peanut oil. CONCLUSION Fusion of NIR and fluorescence data and chemometrics provided enhanced capacity for rapid and untargeted analysis of multiple adulterations in Chinese COA oils. ? 2018 Society of Chemical Industry

Metzincins are key molecules in the degradation of the extracellular matrix and play an important role in cellular processes such as cell migration, adhesion, and cell fusion of malignant tumors, including cutaneous melanoma (CM). We hypothesized that genetic variants of the metzincin metallopeptidase family genes would be associated with CM-specific survival (CMSS). To test this hypothesis, we first performed Cox proportional hazards regression analysis to evaluate the associations between genetic variants of 75 metzincin metallopeptidase family genes and CMSS using the dataset from the genome-wide association study (GWAS) from The University of Texas MD Anderson cancer Center (MDACC) which included 858 non-Hispanic white patients with CM, and then validated in the dataset from the Harvard GWAS study which had 409 non-Hispanic white patients with invasive CM. Four independent SNPs (MMP16 rs10090371 C > A, ADAMTS3 rs788935 T > C, TLL2 rs10882807 T > C and MMP9 rs3918251 A > G) were identified as predictors of CMSS, with a variant-allele attributed hazards ratio (HR) of 1.73 (1.32-2.29, 9.68E-05), 1.46 (1.15-1.85, 0.002), 1.68 (1.31-2.14, 3.32E-05) and 0.67 (0.51-0.87, 0.003), respectively, in the meta-analysis of these two GWAS studies. Combined analysis of risk genotypes of these four SNPs revealed a decreased CMSS in a dose-response manner as the number of risk genotypes increased (Ptrend

Rho GTPases control cell division, motility, adhesion, vesicular trafficking and phagocytosis, which may affect progression and/or prognosis of cancers. Here, we investigated associations between genetic variants of Rho GTPases-related genes and cutaneous melanoma-specific survival (CMSS) by re-analyzing a published melanoma genome-wide association study (GWAS) and validating the results in another melanoma GWAS. In the single-locus analysis of 36,018 SNPs in 129 Rho-related genes, 427 SNPs were significantly associated with CMSS (P C, ARHGAP22 rs3851552 T>C, ARHGAP44 rs72635537 C>T and ARHGEF10 rs7826362 A>T) were independently predictive of CMSS (a meta-analysis derived P=9.04 × 10−4, 9.58 × 10−4, 1.21 × 10−4 and 8.47 × 10−4, respectively). Additionally, patients with an increasing number of unfavorable genotypes (NUGs) of these loci had markedly reduced CMSS in both discovery dataset and validation dataset (Ptrend=1.47 × 10−7 and 3.12 × 10−5). The model including the NUGs and clinical variables demonstrated a significant improvement in predicting the five-year CMSS. Moreover, rs10916352C and rs3851552C alleles were significantly associated with an increased mRNA expression levels of RHOU (P=1.8 × 10−6) and ARHGAP22 (P=5.0 × 10−6), respectively. These results may provide promising prognostic biomarkers for CM personalized management and treatment. This article is protected by copyright. All rights reserved.

To identify genetic variants involved in prognosis of cutaneous melanoma (CM), we investigated associations of single nucleotide polymorphisms (SNPs) of genes in the integrin signaling pathway with CM survival by re-analyzing a published genome-wide association study (GWAS) from The University of Texas M.D. Anderson Cancer Center (MDACC), and then validated significant SNPs in another GWAS from Harvard University. In the MDACC study, 1,148 SNPs were significantly associated with CM-specific survival (CMSS) (P ≤ 0.050 and false-positive report probability ≤ 0.20), and nine SNPs were validated in the Harvard study (P ≤ 0.050). Among these, three independent SNPs (i.e., DOCK1 rs11018104 T>A, rs35748949 C>T and PAK2 rs1718404 C>T) showed a predictive role in CMSS, with an effect-allele attributed adjusted hazards ratio [adjHR of 1.50 (95% confidence interval (CI) = 1.18-1.90, P = 7.46E-04), 1.53 (1.18-1.97, 1.18E-03) and 0.58 (0.45-0.76, 5.60E-05), respectively]. Haplotype analysis revealed that a haplotype carrying two risk alleles A-T in DOCK1 was associated with the poorest survival in both MDACC (adjHR=1.73, 95% CI = 1.19-2.50, P = 0.004) and Harvard (adjHR = 1.95, 95% CI=1.14-3.33, P = 0.010) studies. In addition, patients with an increasing number of unfavorable genotypes (NUGs) for these three SNPs had a poorer survival. Incorporating NUGs with clinical variables showed a significantly improved ability to classify CMSS (AUC increased from 86.8% to 88.6%, P = 0.031). Genetic variants in the integrin signaling pathway may independently or jointly modulate the survival of CM patients. Further large, prospective studies are needed to validate these findings. This article is protected by copyright. All rights reserved.

Two new prenylated indole diterpenoids, tolypocladins K and L (1 and 2), together with a known analog terpendole L (3), were isolated from the solid fermentation culture of a mine soil-derived fungus Tolypocladium sp. XL115. Their structures and relative configurations were determined by comprehensive spectroscopic data analysis, as well as by comparison of their NMR data with those related known compounds. Compound 3 exhibited remarkable antibacterial activity against Micrococcus luteus with an MIC value of 6.25 μg/mL, and compounds 1 and 3 displayed moderate antifungal activity selectively against tested strains with MIC values of 25-50 μg/mL.

Geranylgeranyltransferase I (GGT), a protein prenyltransferase, is responsible for the posttranslational lipidation of Rho GTPases, such as Rac, Rho and Cdc42, all of which play an important role in neuronal synaptogenesis. We previously demonstrated that GGT promotes dendritic morphogenesis in cultured hippocampal neurons and cerebellar slices. We report here that inhibiting GGT activity decreases basal- and activity-dependent changes in spine density as well as in learning and memory ability of mice in vivo. We found that KCl- or bicuculline-induced dendritic spine density increases was abolished by specific GGT inhibitor GGTi-2147 treatment in cultured hippocampal neurons. GGTi-2147 lateral ventricular injection reduced GGT activity and membrane association of Rac and decreased the density of dendritic spines in the mouse hippocampus, frontal cortex and cerebellum. GGTi-2147 administration also impaired learning and memory ability of mice. More importantly, mice exposed to environmental enrichment (EE) showed increased spine density and learning and memory ability, which were significantly reversed by GGTi-2147 administration. These data demonstrate that inhibiting GGT activity prevents both basal- and activity-dependent changes in spine density in central nervous system both in vitro and in vivo. Manipulating GGT activity may be a promising strategy for the therapies of neurodevelopmental disorders, such as autism, depression, and schizophrenia.

Bmi-1 is involved in the development of several human cancers; however, its significance in glioma progression remains largely unknown. We report that downregulation of Bmi-1 clearly reduces glioma cell migration and invasion. Downregulation of Bmi-1 promotes the expression of the tumor suppressor p16, which is important in glioma cell motility. Reduction in glioma cell invasion due to downregulation of Bmi-1 could be rescued by p16 downregulation. These results show that Bmi-1 contributes to the motility of glioma cells by regulating the expression of p16.

The identification of genes involved in carcinogenesis and tumor progression is of great interest, since these genes might be possible as candidates for new tumor targeted therapy strategies. Our previous study shows that Golgi phosphoprotein 3 (GOLPH3) is involved in glioma cell migration and invasion, the critical characteristics of malignant gliomas. In this study, we explored the mechanism of GOLPH3 affecting cell migration and invasion and found that GOLPH3 promotes glioblastoma (GBM) cell migration and invasion via the mammalian target of rapamycin(mTOR)-Y-box binding protein-1 (YB1) pathway in vitro. Both the protein levels of GOLPH3 and YB1 were up-regulated in human glioma tissues and they exhibited direct correlation with each other. In addition, down-regulation of GOLPH3 inhibited glioma cell migration and invasion, while over-expression of GOLPH3 enhanced them. Meanwhile, GOLPH3 down-regulation led to a significant decrease of YB1 level as well as mTOR activity, both required for glioma cell migration and invasion. On the contrary, YB1 level and mTOR activity increased after GOLPH3 over-expression. YB1 down-regulation or mTOR ATP site inhibitor INK128 treatment inhibited cell migration and invasion, similar to the effect of GOLPH3 down-regulation. Furthermore, over-expression of GOLPH3 induced glioma cell migration and invasion was blocked by INK128 and YB1 down-regulation. Taken together, these results show that GOLPH3 promotes glioblastoma cell migration and invasion via the mTOR-YB1pathway, indicating that GOLPH3-mTOR-YB1 pathway might be a new therapeutic target for glioma treatment.

BACKGROUND This study attempts to elucidate the complex effect of Ca2+, an important divalent cation widely present in aqueous media, on perchlorate removal by autotrophic perchlorate reducing bacteria (PRB) associated with a zero-valent iron (ZVI) system. RESULTS Batch flask test results showed that, in the presence of larger amounts of Ca2+, perchlorate removal initiated earlier and completed faster. According to the increase of differential chloride concentration, the respective contribution of biodegradation to perchlorate removal was evaluated and found to be improved with increasing amount of Ca2+. It appears that Ca2+ can delay pH increase by combining with OH− and can consequently extend biodegradation time. Despite the major contribution of biodegradation, when ZVI and Ca2+ co-existed, physiochemical perchlorate removal occurred earlier by both adsorption via surface charge neutralization and co-precipitation via formation of larger particles. The positive effect of Ca2+ was further evidenced in subsequent continuous column tests. As empty bed residence time (EBRT) was decreased, more reliable and effective perchlorate removal performance was observed with additional Ca2+ concentration of 240 mg L−1. CONCLUSION Perchlorate removal by a PRB-ZVI system can be significantly improved with increasing amounts of Ca2+, which can be reasonably understood by the enhanced biodegradation and physiochemical removal process. © 2014 Society of Chemical Industry

Recently, calcyclin-binding protein or Siah-1-interacting protein (CacyBP/SIP), a component of a novel ubiquitinylation pathway, could regulate the β-catenin degradation (Fukushima et al., Immunity 2006, 24, 29 – 39). However, the potential role of CacyBP/SIP itself in human glioma cells has not been clarified. Here, we found that CacyBP/SIP was expressed highly in human glioma tissues. Silencing of CacyBP/SIP by short-hairpin RNA severely suppressed the proliferation of human glioma cell U251, which was at least partly mediated by downregulation of phospho-Akt (p-Akt) and phospho-β-catenin (p-β-catenin) as well as upregulation of p53 and p21. Furthermore, overexpression of CacyBP/SIP obviously promoted the proliferation of human glioma U251, which exhibited the exactly contrary trend in the expression of p-Akt, p-β-catenin, p53, and p21. Taken together, these findings suggest that CacyBP/SIP plays important roles in the proliferation of human glioma cell which might be involved in the development of human glioma. © 2014 IUBMB Life, 66(4):286–291, 2014

Ras-oncogenic pathway contributes to the pathogenesis of various tumours in humans, in which mutations of three canonical genes including H-Ras, N-Ras and K-Ras are the most common events. Dysregulation of Ras signalling can be tumourigenic, especially gliomas of the central nervous system. Rap proteins are members of the small GTPase superfamily that involved in many biological processes. However, it remains largely unclear as to whether and how Rap proteins are involved in the development of multiple gliomas. We found that the levels of the protein Rap2a and the activity of Rap2a (GTP-Rap2a) were weakly expressed in glioma tissues. Overexpressed Rap2a significantly inhibited the migration and invasion of glioma cells with an increase of GTP-Rap2a. Overexpression of the dominant-active (DA-Rap2a), but not the dominant-negative (DN-Rap2a) form of Rap2a, also similarly inhibited the migration and invasion of glioma cells by reducing the phosphorylation level of AKT. In contrast, downregulation of Rap2a promoted glioma migration and invasion, and raised the phosphorylation level of AKT, whereas these effects were inhibited by PI3K-specific inhibitor, LY294002. Thus unlike the other family members of Ras, Rab2a probably serves as a tumour suppressor in the pathogenesis of glioma.

Geranylgeranyltransferase I (GGT) is a prenyltransferase that mediates lipid modification of Rho small GTPases, such as Rho, Rac, and Cdc42, which are important for neuronal synaptogenesis. Although GGT is expressed in brain extensively, the function of GGT in central nerves system is largely unknown so far. We have previously demonstrated that GGT promotes the basal and neuronal activity and brain-derived neurotrophic factor (BDNF)-induced dendritic morphogenesis of cultured hippocampal neurons and cerebellar slices. This study is to explore the function and mechanism of GGT in neuronal synaptogenesis. We found that the protein level and activity of GGT gradually increased in rat hippocampus from P7 to P28 and subcellular located at synapse of neurons. The linear density of Synapsin 1 and post-synaptic density protein 95 increased by over-expression of GGT β, while reduced by inhibition or down-regulation of GGT. In addition, GGT and its known substrate Rac was activated by BDNF, which promotes synaptogenesis in cultured hippocampal neurons. Furthermore, BDNF-induced synaptogenesis was eliminated by GGT inhibition or down-regulation, as well as by non-prenylated Rac1 over-expression. Together, our data suggested that GGT mediates BDNF-induced neuronal synaptogenesis through Rac1 activation.

Background Vitiligo is an acquired pigment disorder characterized by areas of depigmented skin resulting from the loss of epidermal melanocytes. Recently, several investigations have documented the benefits of excimer phototherapy (e.g., using the 308-nm excimer laser or the 308-nm excimer lamp) for the treatment of vitiligo. Aim To compare the effectiveness of the 308-nm excimer laser with the 308-nm excimer lamp in the treatment of vitiligo patients. Methods This intervention study was designed as a randomized self-control trial. Fourteen subjects with 48 symmetrical vitiligo lesions were enrolled in this study. One lesion was treated with the 308-nm excimer laser, and its counterpart was treated with the 308-nm excimer lamp. Lesions were treated three times a week with the same dose on both sides for a total of 20 sessions. Results All of the patients completed the study, and 48 lesions were treated. The two treatments exhibited similar results in terms of repigmentation. Conclusions The 308-nm excimer lamp and the 308-nm excimer laser exhibited similar efficacies in treating vitiligo.

Growth hormone (GH) is usually released by somatotrophs in the anterior pituitary in response to the GH-releasing hormone and plays an important role in skeleton development and postnatal growth. However, it is unclear if extrapituitary GH exerts any effect on murine multilineage cells (MMCs). MMCs are multipotent progenitors that give rise to several lineages, including bone, cartilage, and fat. We have identified bone morphogenic protein 9 (BMP9) as one of the most osteogenic BMPs in MMCs by regulating a distinct set of downstream mediators. In this study, we find that GH is one of the most significantly upregulated genes by BMP9 in mouse MMCs through expression-profiling analysis. We confirm that GH is a direct early target of and upregulated by BMP9 signaling. Exogenous GH synergizes with BMP9 on inducing early and late osteogenic markers in MMCs. Furthermore, BMP9 and GH costimulation leads to a significant expansion of growth plate in cultured limb explants. Although GH alone does not induce de novo bone formation in an ectopic bone formation model, BMP9 and GH costimulated MMCs form more mature bone, which can be inhibited by silencing GH expression. The synergistic osteogenic activity between BMP9 and GH can be significantly blunted by JAK/STAT inhibitors, leading to a decrease in GH-regulated insulin-like growth factor 1 (IGF1) expression in MMCs. Our results strongly suggest that BMP9 may effectively regulate extrapituitary GH expression in MMCs. Thus, it is conceivable that the BMP9-GH-IGF axis may be exploited as an innovative strategy to enhance osteogenesis in regenerative medicine.

In this paper, we propose an optimization of MAC protocol design for wireless sensor networks, that accounts for cross-layering information, in terms of location accuracy for nodes and residual energy levels. In our proposed solution we encode this cross-layer information within a decreasing backoff function in the MAC. The protocol is optimized by appropriately selecting priority window lengths, and we have shown that accurate cross-layer information plays a crucial role in achieving an optimal performance at the MAC layer level. The estimation accuracy can be characterized spatially using a location reliability probability distribution function. We show that this distribution function greatly influences the design of the optimal backoff window parameters, and the overall throughput performance of the MAC protocol. Copyright © 2010 John Wiley & Sons, Ltd.

Efficient osteogenic differentiation and bone formation from mesenchymal stem cells (MSCs) should have clinical applications in treating nonunion fracture healing. MSCs are adherent bone marrow stromal cells that can self-renew and differentiate into osteogenic, chondrogenic, adipogenic, and myogenic lineages. We have identified bone morphogenetic protein 9 (BMP-9) as one of the most osteogenic BMPs. Here we investigate the effect of insulin-like growth factor 2 (IGF-2) on BMP-9-induced bone formation. We have found that endogenous IGF-2 expression is low in MSCs. Expression of IGF-2 can potentiate BMP-9-induced early osteogenic marker alkaline phosphatase (ALP) activity and the expression of later markers. IGF-2 has been shown to augment BMP-9-induced ectopic bone formation in the stem cell implantation assay. In perinatal limb explant culture assay, IGF-2 enhances BMP-9-induced endochondral ossification, whereas IGF-2 itself can promote the expansion of the hypertropic chondrocyte zone of the cultured limb explants. Expression of the IGF antagonists IGFBP3 and IGFBP4 leads to inhibition of the IGF-2 effect on BMP-9-induced ALP activity and matrix mineralization. Mechanistically, IGF-2 is further shown to enhance the BMP-9-induced BMPR-Smad reporter activity and Smad1/5/8 nuclear translocation. PI3-kinase (PI3K) inhibitor LY294002 abolishes the IGF-2 potentiation effect on BMP-9-mediated osteogenic signaling and can directly inhibit BMP-9 activity. These results demonstrate that BMP-9 crosstalks with IGF-2 through PI3K/AKT signaling pathway during osteogenic differentiation of MSCs. Taken together, our findings suggest that a combination of BMP-9 and IGF-2 may be explored as an effective bone-regeneration agent to treat large segmental bony defects, nonunion fracture, and/or osteoporotic fracture. © 2010 American Society for Bone and Mineral Research.

Background: Hepatic progenitor cells (HPCs) can be isolated from fetal liver and extrahepatic tissues. Retinoic acid (RA) signalling plays an important role in development, although the role of RA signalling in liver-specific progenitors is poorly understood. Aims: We sought to determine the role of RA in regulating hepatic differentiation. Methods: RNA was isolated from liver tissues of various developmental stages. Liver marker expression was assessed by reverse transcriptase-polymerase chain reaction and immunofluorescence staining. Reversibly immortalized HPCs derived from mouse embryonic day 14.5 (E14.5) liver (aka, HP14.5) were established. Albumin promoter-driven reporter (Alb-GLuc) was used to monitor hepatic differentiation. Glycogen synthesis was assayed as a marker for terminal hepatic differentiation. Results: Retinoic acid receptor (RAR)-a, retinoid X receptor (RXR)-a and RXR-g expressed in E12.5 to postnatal day 28 liver samples. Expression of RAR-b and RXR-b was low perinatally, whereas RAR-g was undetectable in prenatal tissues and increased postnatally. Retinal dehydrogenase 1 and 2 (Raldh1 and Raldh2) were expressed in all tissues, while Raldh3 was weakly expressed in prenatal samples but was readily detected postnatally. Nuclear receptor corepressors were highly expressed in all tissues, while expression of nuclear co-activators decreased in perinatal tissues and increased after birth. HP14.5 cells expressed high levels of early liver stem cell markers. Expression of RA signalling components and coregulators was readily detected in HP14.5. RA was shown to induce Alb-GLuc activity and late hepatocyte markers. RA was further shown to induce glycogen synthesis in HP14.5 cells, an important function of mature hepatocytes. Conclusions: Our results strongly suggest that RA signalling may play an important role in regulating hepatic differentiation.

Wnt/beta-catenin pathway plays an important role in regulating embryonic development. Hepatocytes differentiate from endoderm during development. Hepatic progenitor cells (HPCs) have been isolated from fetal liver and extrahepatic tissues. Most current studies in liver development and hepatic differentiation have been focused on Wnts, beta-catenin, and their receptors. Here, we sought to determine the role of Wnt antagonists in regulating hepatic differentiation of fetal liver-derived HPCs. Using mouse liver tissues derived from embryonic day E12.5 to postnatal day (PD) 28, we found that 13 of the 19 Wnt genes and almost all of Wnt receptors/co-receptors were expressed in most stages. However, Wnt antagonists SFRP2, SFRP3, and Dkk2 were only detected in the early stages. We established and characterized the reversible stable HPCs derived from E14.5 mouse fetal liver (HP14.5). HP14.5 cells were shown to express high levels of early liver progenitor cell markers, but low levels or none of late liver markers. HP14.5 cells were shown to differentiate into mature hepatocytes upon dexamethasone (Dex) stimulation. Dex-induced late marker expression and albumin promoter activity in HP14.5 cells were inhibited by exogenous expression of SFRP3. Furthermore, Dex-induced glycogen synthesis of PAS-positive HP14.5 cells was significantly inhibited by SFRP3. Therefore, our results have demonstrated that the expression of Wnt antagonists decreases as hepatic differentiation progresses, suggesting that a balanced Wnt signaling may be critical during mouse liver development and hepatic differentiation.

NeuroD1/BETA2, a member of the bHLH transcription factor family, is known to influence the fate of specific neuronal, endocrine and retinal cells. We report here that NeuroD1 mRNA is highly abundant in the developing and adult rat pineal gland. Pineal expression begins in the 17-day embryo at which time it is also detectable in other brain regions. Expression in the pineal gland increases during the embryonic period and is maintained thereafter at levels equivalent to those found in the cerebellum and retina. In contrast, NeuroD1 mRNA decreases markedly in non-cerebellar brain regions during development. Pineal NeuroD1 levels are similar during the day and night, and do not appear to be influenced by sympathetic neural input. Gene expression analysis of the pineal glands from neonatal NeuroD1 knockout mice identifies 127 transcripts that are down-regulated (>twofold, p twofold, p < 0.05). According to quantitative RT-PCR, the most dramatically down-regulated gene is kinesin family member 5C (100-fold) and the most dramatically up-regulated gene is glutamic acid decarboxylase 1( fourfold). Other impacted transcripts encode proteins involved in differentiation, development, signal transduction and trafficking. These findings represent the first step toward elucidating the role of NeuroD1 in the rodent pinealocyte.

Recent studies suggest that a common theme links the diverse elements of pineal photoneuroendocrine transduction – regulation via binding to 14-3-3 proteins. The elements include photoreception, neurotransmission, signal transduction and the synthesis of melatonin from tryptophan. We review general aspects of 14-3-3 proteins and their biological function as binding partners, and also focus on their roles in pineal photoneuroendocrine transduction. The function of the pineal gland as a photoneuroendocrine transducer involves multiple processes, including photoreception, signal transduction and the synthesis of melatonin from tryptophan. In addition, in birds and mammals, adrenergic neurotransmission plays a regulatory role. Recent studies suggest that a common theme links these diverse processes, one that has been generally ignored: regulation via binding to 14-3-3 proteins. We review general aspects of 14-3-3 proteins, their biological role as binding partners and their roles in pineal photoneuroendocrine transduction.