Glucagon-like peptides (GLP-1/2) are co-produced and highlighted as key modulators to improve glucose homeostasis and insulin sensitivity after bariatric surgery. However, it is unknown if CNS GLP-2 plays any physiological role in the control of glucose homeostasis and insulin sensitivity. We show that mice lacking GLP-2 receptor (GLP-2R) in POMC neurons display glucose intolerance and hepatic insulin resistance. GLP-2R activation in POMC neurons is required for GLP-2 to enhance insulin-mediated suppression of hepatic glucose production (HGP) and gluconeogenesis. GLP-2 directly modulates excitability of POMC neurons in GLP-2R- and PI3K-dependent manners. GLP-2 initiates GLP-2R-p85α interaction and facilitates PI3K-Akt-dependent FoxO1 nuclear exclusion in POMC neurons. Central GLP-2 suppresses basal HGP and enhances insulin sensitivity, which are abolished in POMC-p110α KO mice. Thus, CNS GLP-2 plays a key physiological role in the control of hepatic glucose production through activating PI3K-dependent modulation of membrane excitability and nuclear transcription of POMC neurons in the brain.
Glucagon-like peptides and receptors; Glucose homeostasis; Hypothalamus; Insulin sensitivity
Neonatal engraftment by oligodendrocyte progenitor cells (OPCs) permits the myelination of congenitally dysmyelinated brain. To establish a potential autologous source of these cells, we developed a strategy by which to differentiate human induced pluripotential stem cells (hiPSCs) into OPCs. From 3 hiPSC lines, as well as from human embryonic stem cells (hESCs), we generated highly enriched OLIG2+/PDGFRα+/NKX2.2+/SOX10+ hOPCs, which could be further purified using fluorescence-activated cell sorting. hiPSC OPCs efficiently differentiated into both myelinogenic oligodendrocytes and astrocytes, in vitro and in vivo. Neonatally engrafted hiPSC OPCs robustly myelinated the brains of myelin-deficient shiverer mice, and substantially increased the survival of these mice. The speed and efficiency of myelination by hiPSC OPCs was higher than that previously observed using fetal tissue-derived OPCs, and no tumors from these grafts were noted as long as 9 months after transplant. These results suggest the utility of hiPSC-derived OPCs in treating disorders of myelin loss.
glial progenitor cell; iPS cell; embryonic stem cell; neural stem cell; remyelination
The harpin protein Hpa1 produced by the bacterial blight pathogen of rice induces several growth-promoting responses in plants, activating the ethylene signaling pathway, increasing photosynthesis rates and EXPANSIN (EXP) gene expression levels, and thereby enhancing the vegetative growth. This study was attempted to analyze any mechanistic connections among the above and the role of gibberellin in these responses. Hpa1-induced growth enhancement was evaluated in Arabidopsis, tomato, and rice. And growth-promoting responses were determined mainly as an increase of chlorophyll a/b ratio, which indicates a potential elevation of photosynthesis rates, and enhancements of photosynthesis and EXP expression in the three plant species. In Arabidopsis, Hpa1-induced growth-promoting responses were partially compromised by a defect in ethylene perception or gibberellin biosynthesis. In tomato and rice, compromises of Hpa1-induced growth-promoting responses were caused by a pharmacological treatment with an ethylene perception inhibitor or a gibberellin biosynthesis inhibitor. In the three plant species, moreover, Hpa1-induced growth-promoting responses were significantly impaired, but not totally eliminated, by abolishing ethylene perception or gibberellin synthesis. However, simultaneous nullifications in both ethylene perception and gibberellin biosynthesis almost canceled the full effects of Hpa1 on plant growth, photosynthesis, and EXP2 expression. Theses results suggest that ethylene and gibberellin coregulate Hpa1-induced plant growth enhancement and associated physiological and molecular responses.
Electronic supplementary material
The online version of this article (doi:10.1007/s00425-013-2013-y) contains supplementary material, which is available to authorized users.
Ethylene; Gibberellin; Hpa1; Plant growth enhancement; Signal transduction
Active DNA demethylation is an important part of epigenetic regulation in plants and animals. How active DNA demethylation is regulated and its relationship with histone modification patterns are unclear. Here, we report the discovery of IDM1, a regulator of DNA demethylation in Arabidopsis. IDM1 is required for preventing DNA hypermethylation of highly homologous multicopy genes and other repetitive sequences that are normally targeted for active DNA demethylation by Repressor of Silencing 1 and related 5-methylcytosine DNA glycosylases. IDM1 binds methylated DNA at chromatin sites lacking histone H3K4 di- or trimethylation and acetylates H3 to create a chromatin environment permissible for 5-methylcytosine DNA glycosylases to function. Our study reveals how some genes are indicated by multiple epigenetic marks for active DNA demethylation and protection from silencing.
Skeletal muscle mitochondrial dysfunction is associated with aging and diabetes, which decreases respiratory capacity and increases reactive oxygen species. Lipoic acid (LA) possesses antioxidative and antidiabetic properties. Metabolic action of LA ismediated by activation of AMP-activated protein kinase (AMPK), a cellular energy sensor that can regulate PGC-1α, a master regulator of mitochondrial biogenesis. We hypothesized that LA improves energy metabolism and mitochondrial biogenesis by enhancing AMPK-PGC-1α signalling in the skeletal muscle of aged mice.
C57BL/6 mice (24-month old, male) were supplemented with or without α-LA (0.75% in drinking water) for one month. In addition, metabolic action and cellular signalling of LA were studied in cultured mouse myoblastoma C2C12 cells.
LA supplementation improved body composition, glucose tolerance, and energy expenditure in the aged mice. LA increased skeletal muscle mitochondrial biogenesis with increased phosphorylation of AMPK and mRNA expression of PGC-1α and glucose transporter-4 (GLUT-4). Besides body fat mass, LA decreased lean mass and attenuated phosphorylation of mammalian target of rapamycin (mTOR) signalling in the skeletal muscle. In cultured C2C12 cells, LA increased glucose uptake and palmitate β-oxidation, but decreased protein synthesis, which was associated with increased phosphorylation of AMPK and expression of PGC-1α and GLUT-4, and attenuated phosphorylation of mTOR and p70S6 kinase.
We conclude that LA improves skeletal muscle energy metabolism in the aged mouse possibly through enhancing AMPK-PGC-1α-mediated mitochondrial biogenesis and function. Moreover, LA increases lean mass loss possibly by suppressing protein synthesis in the skeletal muscle by down-regulating the mTOR signalling pathway. Thus, LA may be a promising supplement for treatment of obesity and/or insulin resistance in older patients.
Lipoic acid; Aging; Mitochondrial biogenesis; Protein synthesis; Energy metabolism
Impact and friction model of nanofluid for molecular dynamics simulation was built which consists of two Cu plates and Cu-Ar nanofluid. The Cu-Ar nanofluid model consisted of eight spherical copper nanoparticles with each particle diameter of 4 nm and argon atoms as base liquid. The Lennard-Jones potential function was adopted to deal with the interactions between atoms. Thus motion states and interaction of nanoparticles at different time through impact and friction process could be obtained and friction mechanism of nanofluids could be analyzed. In the friction process, nanoparticles showed motions of rotation and translation, but effected by the interactions of nanoparticles, the rotation of nanoparticles was trapped during the compression process. In this process, agglomeration of nanoparticles was very apparent, with the pressure increasing, the phenomenon became more prominent. The reunited nanoparticles would provide supporting efforts for the whole channel, and in the meantime reduced the contact between two friction surfaces, therefore, strengthened lubrication and decreased friction. In the condition of overlarge positive pressure, the nanoparticles would be crashed and formed particles on atomic level and strayed in base liquid.
The Yunnan province is the epicenter of HIV-1 epidemics in China and a center for drug trafficking to the other parts of the world. In six prefectures of this province, a total of 132 IDUs were recruited to determine the seroprevalence of HCV and HIV-1 and the positive rates were 93.94% and 68.18%, respectively (P <0.001). Co-infection with HCV and HIV-1 was found among 89 IDUs, of whom several HCV fragments were amplified and sequenced. Sequences of the HCV 5′NCR-C and NS5B region were determined from 82 IDUs. Phylogenetic analyses showed consistent genotyping among 80 IDUs. Among them HCV genotypes 1a, 1b, 3a, 3b, 6a, 6n, and a tentatively assigned novel 6u subtype were found in 1 (1.25%), 16 (20%), 19 (23.75%), 24 (30%), 4 (5%), 9 (11.25%) and 7 (8.75%) individuals, respectively. In two IDUs, genotyping results were discordant, suggesting mixed HCV infections or recombination. The proportion of patients with HCV 1b tended to decrease from the north to south and from the east to west in this province. Genotype 3 and 6 strains were more frequent in the southern prefectures. The novel subtype 6u strains were only detected in Dehong which borders Myanmar. Our findings showed a unique pattern of HCV genotype distribution, which is similar to that in the southeastern Asian countries but distinct from that among the general population in China. Routes of drug trafficking and the resulting high prevalence of HIV-1 infection may have contributed to this pattern of HCV genotype distribution.
HCV; HIV-1; co-infection IDUs; genotypes
It is known that periodontal ligament stem cells (PDLSCs) can differentiate into cementoblast-like cells, adipocytes and collagen-forming cells. However, whether PDLSCs are able to differentiate into Schwann cells and which method is best for their neural induction remain unknown. We attempted to determine whether PDLSCs possessed the potential for neural differentiation in vitro.
Materials and methods
We isolated and multiplied PDLSCs from periodontal ligaments obtained from the teeth (n = 24) of 8-month-old beagle dogs. Four protocols with different chemicals and growth factors were adopted to induce the PDLSCs to differentiate into Schwann cells. Immunochemistry, RT-PCR and qRT-PCR were performed to investigate the in vitro neural differentiation potential of PDLSCs.
We compared the 4 different protocols and showed that all 4 protocols could successfully induce PDLSCs to express nestin, GFAP and S100, markers for Schwann cells. Further, qRT-PCR revealed relative differences in the expression levels of these 3 genes in differentiated PDLSCs obtained by different protocols.
We conclude that PDLSCs have neural/glial differentiation potential in vitro and that neural/glial differentiation can be induced in PDLSCs if suitable protocols are followed. We also found that supplementing the growth medium with suitable growth factors is more effective than applying chemicals alone. While nerve growth factor is more effective than platelet-derived growth factor for inducing neural/glial differentiation in PDLSCs, pre-induction of PDLSCs with dimethyl sulphoxide yields better results than those obtained with all-trans-retinoic acid.
periodontal ligament; stem cells; neural/glial differentiation; Schwann cells
Mutations in leucine-rich repeat kinase-2 (LRRK2) are the most common known cause of Parkinson disease (PD), but how this protein results in the pathobiology of PD is unknown. Moreover, there is variability in pathology among cases and α-synuclein (α-syn) neuronal inclusions are often present but whether LRRK2 is present in these pathological inclusions is controversial. This study characterizes novel LRRK2 antibodies, some of which preferentially recognize an aggregated form of LRRK2, as observed in cell culture models. Large perinuclear aggregates containing LRRK2 were promoted by proteasome inhibition and prevented by microtubule polymerization inhibition. Further, they were vimentin- and γ-tubulin- but not lamp1-immunoreactive, suggesting that these structures fit the definition of aggresomes. Inhibition of Hsp90 led to degradation of only the soluble/cytosolic pool of LRRK2, suggesting that the aggresomes formed independent of the stability provided by Hsp90. Although these novel anti-LRRK2 antibodies identified aggregates in model cell systems, they did not immunostain pathological inclusions in human brains. Further, co-expression of LRRK2 and α-syn did not recruit α-syn into aggresomes in cultured cells, even in the presence of proteasome inhibition. Thus, although LRRK2 is a model system for aggresome formation, LRRK2 is not present in α-syn pathological inclusions.
α-synuclein; Aggregation; Aggresome; Antibody; LRRK2; Parkinson disease
Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are the most common known cause of Parkinson's disease (PD). Whether loss of LRRK2 function accounts for neurodegeneration of dopamine neurons in PD is not known, nor is it known whether LRRK2 kinase activity modulates the susceptibility of dopamine (DA) neurons to the selective dopaminergic toxin, 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine (MPTP). To better understand the role of LRRK2 in DA neuronal survival and its role in the susceptibility of DA neurons to MPTP, we generated LRRK2 knockout (KO) mice lacking the kinase domain of LRRK2. Here we show that LRRK2 KO mice are viable and have no major abnormalities and live to adulthood. The dopaminergic system is normal in LRRK2 KO mice as assessed via HPLC for DA and its metabolites and via stereologic assessment of DA neuron number in young and aged mice. Importantly, there is no significant difference in the susceptibility of LRRK2 KO and wild type (WT) mice to MPTP. These results suggest that LRRK2 plays little if any role in the development and survival of DA neurons under physiologic conditions. Thus, PD due to LRRK2 mutations are likely not due to a loss of function. Moreover, LRRK2 is not required for the susceptibility of DA neurons to MPTP.
Parkinson's disease; LRRK2; MPTP; knockouts; mitochondria; Dopamine
Adipose-derived stem cells (ADSCs) could accomplish neural transdifferentiation with the presence of certain growth factors in vitro. It has been proved that bone marrow stromal cells (BMSCs) can realize neural transdifferentiation only by being co-cultured with Schwann cells (SCs), and in our former studies we have confirmed that ADSCs could do so too. This paper aims to investigate whether the neural induction efficiency of co-culture is as high as that of other strategies using chemicals or chemicals combined with some growth factors.
Material and methods
We isolated and multiplied ADSCs from adult Sprague-Dawley rats, and SCs from sciatic nerves of 1-to-2-day-old Sprague-Dawley rat pups, then induced ADSCs neural transdifferentiation through 2% dimethyl sulphoxide (DMSO) and DMSO combined with growth factors. Meanwhile we co-cultured ADSCs and SCs in Transwell culture dishes without intercellular contacts. Immunostaining and RT-PCR were adopted to investigate the neural transdifferentiation of ADSCs. Then we compared the expression differences for genes S100, nestin and GFAP of the above three protocols by real-time PCR.
Both immunostaining and RT-PCR proved that ADSCs could accomplish neural transdifferentiation through each of the above three protocols. And real-time PCR further shows that the gene expression relative quantities for the above three genes are not statistically different between co-culture and induction through DMSO combined with growth factors (p > 0.05), but both of them are statistically different from induction only by DMSO (p < 0.05).
Co-culturing ADSCs and SCs may be a simple, effective and practical way for ADSCs neural transdifferentiation.
stem cells; Schwann cells; neural transdifferentiation; co-culture
Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are associated with late-onset, autosomal-dominant, familial Parkinson's disease (PD) and also contribute to sporadic disease. The LRRK2 gene encodes a large protein with multiple domains, including functional Roc GTPase and protein kinase domains. Mutations in LRRK2 most likely cause disease through a toxic gain-of-function mechanism. The expression of human LRRK2 variants in cultured primary neurons induces toxicity that is dependent on intact GTP binding or kinase activities. However, the mechanism(s) underlying LRRK2-induced neuronal toxicity is poorly understood, and the contribution of GTPase and/or kinase activity to LRRK2 pathobiology is not well defined. To explore the pathobiology of LRRK2, we have developed a model of LRRK2 cytotoxicity in the baker's yeast Saccharomyces cerevisiae. Protein domain analysis in this model reveals that expression of GTPase domain-containing fragments of human LRRK2 are toxic. LRRK2 toxicity in yeast can be modulated by altering GTPase activity and is closely associated with defects in endocytic vesicular trafficking and autophagy. These truncated LRRK2 variants induce similar toxicity in both yeast and primary neuronal models and cause similar vesicular defects in yeast as full-length LRRK2 causes in primary neurons. The toxicity induced by truncated LRRK2 variants in yeast acts through a mechanism distinct from toxicity induced by human α-synuclein. A genome-wide genetic screen identified modifiers of LRRK2-induced toxicity in yeast including components of vesicular trafficking pathways, which can also modulate the trafficking defects caused by expression of truncated LRRK2 variants. Our results provide insight into the basic pathobiology of LRRK2 and suggest that the GTPase domain may contribute to the toxicity of LRRK2. These findings may guide future therapeutic strategies aimed at attenuating LRRK2-mediated neurodegeneration.
Parkinson's disease (PD) is the second most common neurodegenerative disorder. PD is considered to be caused by a combination of risk factors including environmental exposure, age, and a positive family history for disease. Several genes have been unambiguously implicated in PD. However, our knowledge is still limited about these genes and the disease mechanisms involved. Mutations in the LRRK2 gene account for up to 40% of PD in certain populations. Since a single-cell model, baker's yeast, has been employed successfully to study the function of genes related to PD and other neurodegenerative disorders, we developed a yeast model of LRRK2 cytotoxicity in this study to investigate the function of LRRK2. We dissected the LRRK2 protein into different fragments including the various functional domains and found that fragments including the GTPase domain of LRRK2 are toxic. This toxicity can be modulated by alterations in GTPase activity and correlates with defects in cellular trafficking. These truncated LRRK2 variants induce similar toxicity and trafficking defects in both yeast and primary neuronal models. This yeast model reveals an important role of GTPase activity in the basic pathobiology of LRRK2 and may guide future therapeutic strategies for PD.
Background and Aims
Delayed self-pollination is a mechanism that allows animal-pollinated plants to outcross while ensuring seed production in the absence of pollinators. This study aims to explore a new mechanism of delayed self-pollination facilitated by wind-driven corolla abscission in Incarvillea sinensis var. sinensis.
Floral morphology and development, and the process of delayed self-pollination were surveyed. Experiments dealing with pollinator and wind exclusion, pollination manipulations, and pollinator observations were conducted in the field.
Delayed self-pollination occurs when the abscising corolla driven by wind drags the adherent epipetalous stamens, thus leading to contact of anthers with stigma in late anthesis. There is no dichogamy and self-incompatibility in this species. The significantly higher proportion of abscised corolla under natural conditions as compared with that in wind-excluding tents indicates the importance of wind in corolla abscission. When pollinators were excluded, corolla abscission significantly increased the number of pollen grains deposited on the stigma and, as a result, the fruit and seed set. Only half of the flowers in plots were visited by pollinators, and the fruit set of emasculated flowers was significantly lower than that of untreated flowers in open pollination. This species has a sensitive stigma, and its two open stigmatic lobes closed soon after being touched by a pollinator, but always reopened if no or only little pollen was deposited.
This delayed self-pollination, which involved the movement of floral parts, the active participation of the wind and sensitive stigma, is quite different from that reported previously. This mechanism provides reproductive assurance for this species. The sensitive stigma contributes to ensuring seed production and reducing the interference of selfing with outcrossing. The pollination pattern, which combines actions by bees with indirect participation by wind, is also a new addition to ambophily.
Ambophily; anther movement; Bignoniaceae; corolla abscission; delayed self-pollination; Incarvillea sinensis var. sinensis; reproductive assurance; stigma closure
The oral health status of rural residents in the People's Republic of China has not been extensively studied and the relationship between poor oral health and esophageal cancer (EC) is unclear. We aim to report the oral health status of adults participating in an EC screening study conducted in a rural high-risk EC area of China and to explore the relationship between oral health and esophageal dysplasia.
National Health and Nutrition Examination Survey (NHANES) oral health examination procedures and the Modified Gingival Index (MGI) were used in a clinical study designed to examine risk factors for esophageal cancer and to test a new esophageal cytology sampling device. This study was conducted in three rural villages in China with high rates of EC in 2002 and was a collaborative effort involving investigators from the National Institutes of Health and the Cancer Institute of the Chinese Academy of Medical Sciences.
Nearly 17% of the study participants aged 40–67 years old were edentulous. Overall, the mean number of adjusted missing teeth (including third molars and retained dental roots) was 13.8 and 35% had 7 contacts or less. Women were more likely to experience greater tooth loss than men. The average age at the time of first tooth loss for those with no posterior functional contacts was approximately 41 years for men and 36 years for women. The mean DMFT (decayed, missing, and filled teeth) score for the study population was 8.5. Older persons, females, and individuals having lower educational attainment had higher DMFT scores. The prevalence of periodontal disease (defined as at least one site with 3 mm of attachment loss and 4 mm of pocket depth) was 44.7%, and 36.7% of the study participants had at least one site with 6 mm or more of attachment loss. Results from a parsimonious multivariate model indicate that participants with poor oral health wemore likely to have esophageal dysplasia (OR = 1.59; 95% CI 1.06, 2.39).
This report describes the first use of NHANES oral health protocols employed in a clinical study conducted outside of the United States. The extent and severity of poor oral health in this Chinese study group may be an important health problem and contributing factor to the prevalence of EC.
Direct cultivation of the first filial generation of gametophyte clones from different Laminaria species is a highly effective way of utilizing kelp heterozygous vigor (heterosis). A male gametophyte clone of L. longissima Miyabe and a female one of L. japonica Areschoug were hybridized, generating Dongfang No. 2 hybrid kelp. This hybrid kelp was used directly in trial cultivation, and its agronomical traits were evaluated. L. longissima and L. japonica are obviously different and complement each other in their morphological characteristics and ecological performances. The hybrid of their gametophyte clones, Dongfang No. 2, showed 56.8% heterozygous vigor in yield. It also showed increased yields of 41.0 and 76.4% compared to the widely used commercial kelps Variety 1 and Variety 2, respectively. In large-scale cultivation trials at different locations and in different years, Dongfang No. 2 attained significantly higher yields than Varieties 1 and 2, increasing yield by 26.4% on average over Variety 1 and by 65.0% over the other. Dongfang No. 2 has a robust holdfast and a wide, long and deep-brown uniform blade, which shows a distinct middle groove. In addition to yield, Dongfang No. 2 also demonstrates obvious heterozygous vigor in other agronomic traits. It is resistant to strong irradiance, as the two commercial varieties are, has an appropriate vegetative maturation time, and adapts well to a range of different culture conditions. The parentage analysis using AFLP of total DNA and SNP of the ITS region of ribosomal RNA transcription unit showed that Dongfang No. 2 is the real hybrid of L. japonica and L. longissima.
Dongfang No. 2; hybrid kelp; gametophyte clone; heterosis; hybridization
Previous studies of mRNA for classical glutathione peroxidase 1
(GPx1) demonstrated that hepatocytes of rats fed a selenium-deficient
diet have less cytoplasmic GPx1 mRNA than hepatocytes of rats fed a
selenium-adequate diet. This is because GPx1 mRNA is degraded by the
surveillance pathway called nonsense-mediated mRNA decay (NMD) when the
selenocysteine codon is recognized as nonsense. Here, we examine the
mechanism by which the abundance of phospholipid hydroperoxide
glutathione peroxidase (PHGPx) mRNA, another selenocysteine-encoding
mRNA, fails to decrease in the hepatocytes and testicular cells of rats
fed a selenium-deficient diet. We demonstrate with cultured NIH3T3
fibroblasts or H35 hepatocytes transiently transfected with PHGPx gene
variants under selenium-supplemented or selenium-deficient conditions
that PHGPx mRNA is, in fact, a substrate for NMD when the
selenocysteine codon is recognized as nonsense. We also demonstrate
that the endogenous PHGPx mRNA of untransfected H35 cells is subject to
NMD. The failure of previous reports to detect the NMD of PHGPx mRNA in
cultured cells is likely attributable to the expression of PHGPx cDNA
rather than the PHGPx gene. We conclude that 1) the sequence of the
PHGPx gene is adequate to support the NMD of product mRNA, and 2) there
is a mechanism in liver and testis but not cultured fibroblasts and
hepatocytes that precludes or masks the NMD of PHGPx mRNA.