TLE3, transducing-like enhancer of split 3, suppresses osteoblast differentiation of bone marrow stromal cells

In senile osteoporosis the balance of adipogenesis and osteoblastogenesis in bone marrow stromal cells (BMSCs) is disrupted so that adipogenesis is increased with respect to osteoblastogenesis, and as a result, bone mass is decreased. While the molecular mechanisms controlling the balance between osteoblastogenesis and adipogenesis are of great interest, the exact nature of the signals regulating this process remains to be determined.     

Chitosan stabilizes platelet growth factors and modulates stem cell differentiation toward tissue regeneration

The idea of using chitosan as a functional delivery aid to support simultaneously PRP, stem cells and growth factors (GF) is associated with the intention to use morphogenic biomaterials to modulate the natural healing sequence in bone and other tissues. For example, chitosan–chondroitin sulfate loaded with platelet lysate was included in a poly(d,l-lactate) foam that was then seeded with human adipose-derived stem cells and cultured in vitro under osteogenic stimulus: the platelet lysate provided to the bone tissue the most suitable assortment of GF which induces the osteogenic differentiation of the mesenchymal stem cells. PDGF, FGF, IGF and TGF-β were protagonists in the repair of callus fractures. The release of GF from the composites of chitosan–PRP and either nano-hydroxyapatite or tricalcium phosphate was highly beneficial for enhancing MSC proliferation and differentiation, thus qualifying chitosan as an excellent vehicle. A number of biochemical characteristics of chitosan exert synergism with stem cells in the regeneration of soft tissues.     

Anticancer mechanisms and clinical application of alkylphospholipids

Synthetic alkylphospholipids (ALPs), such as edelfosine, miltefosine, perifosine, erucylphosphocholine and erufosine, represent a relatively new class of structurally related antitumor agents that act on cell membranes rather than on DNA. They selectively target proliferating (tumor) cells, inducing growth arrest and apoptosis, and are potent sensitizers of conventional chemo- and radiotherapy. ALPs easily insert in the outer leaflet of the plasma membrane and cross the membrane via an ATP-dependent CDC50a-containing ‘flippase’ complex (in carcinoma cells), or are internalized by lipid raft-dependent endocytosis (in lymphoma/leukemic cells). ALPs resist catabolic degradation, therefore accumulate in the cell and interfere with lipid-dependent survival signaling pathways, notably PI3K-Akt and Raf-Erk1/2, and de novo phospholipid biosynthesis. At the same time, stress pathways (e.g. stress-activated protein kinase/JNK) are activated to promote apoptosis. In many preclinical and clinical studies, perifosine was the most effective ALP, mainly because it inhibits Akt activity potently and consistently, also in vivo. This property is successfully exploited clinically in highly malignant tumors, such as multiple myeloma and neuroblastoma, in which a tyrosine kinase receptor/Akt pathway is amplified. In such cases, perifosine therapy is most effective in combination with conventional anticancer regimens or with rapamycin-type mTOR inhibitors, and may overcome resistance to these agents. This article is part of a Special Issue entitled Phospholipids and Phospholipid Metabolism.     

Mechanotransduction activates α5β1 integrin and PI3K/Akt signaling pathways in mandibular osteoblasts

It is unclear how bone cells at different sites detect mechanical loading and how site-specific mechanotransduction affects bone homeostasis. To differentiate the anabolic mechanical responses of mandibular cells from those of calvarial and long bone cells, we isolated osteoblasts from C57B6J mouse bones, cultured them for 1 week, and subjected them to therapeutic low intensity pulsed ultrasound (LIPUS). While the expression of the marker proteins of osteoblasts and osteocytes such as alkaline phosphatase and FGF23, as well as Wnt1 and β-catenin, was equally upregulated, the expression of mandibular osteoblast messages related to bone remodeling and apoptosis differed from that of messages of other osteoblasts, in that the messages encoding the pro-remodeling protein RANKL and the anti-apoptotic protein Bcl-2 were markedly upregulated from the very low baseline levels. Blockage of the PI3K and α₅β₁ integrin pathways showed that the mandibular osteoblast required mechanotransduction downstream of α₅β₁ integrin to upregulate expression of the proteins β-catenin, p-Akt, Bcl-2, and RANKL. Mandibular osteoblasts thus must be mechanically loaded to preserve their capability to promote remodeling and to insure osteoblast survival, both of which maintain intact mandibular bone tissue. In contrast, calvarial Bcl-2 is fully expressed, together with ILK and phosphorylated mTOR, in the absence of LIPUS. The antibody blocking α₅β₁ integrin suppressed both the baseline expression of all calvarial proteins examined and the LIPUS-induced expression of all mandibular proteins examined. These findings indicate that the cellular environment, in addition to the tridermic origin, determines site-specific bone homeostasis through the remodeling and survival of osteoblastic cells. Differentiated cells of the osteoblastic lineage at different sites transmit signals through transmembrane integrins such as α₅β₁ integrin in mandibular osteoblasts, whose signaling may play a major role in controlling bone homeostasis.     

Rab11a and its binding partners regulate the recycling of the ß1-adrenergic receptor

ß1-adrenergic receptors (ß1-AR) are internalized in response to agonists and then recycle back for another round of signaling. The serine 312 to alanine mutant of the ß1-AR (S312A) is internalized but does not recycle. We determined that WT ß1-AR and S312A were internalized initially to an early sorting compartment because they colocalized by > 70% with the early endosomal markers rab5a and early endosomal antigen-1 (EEA1). Subsequently, the WT ß1-AR trafficked via rab4a-expressing sorting endosomes to recycling endosomes. In recycling endosomes WT ß1-AR were colocalized by > 70% with the rab11 GTPase. S312A did not colocalize with either rab4a or rab11, instead they exited from early endosomes to late endosomes/lysosomes in which they were degraded. Rab11a played a prominent role in recycling of the WT ß1-AR because dominant negative rab11a inhibited, while constitutively active rab11a accelerated the recycling of the ß1-AR. Next, we determined the effect of each of the rab11-interacting proteins on trafficking of the WT ß1-AR. The recycling of the ß1-AR was markedly inhibited when myosin Vb, FIP2, FIP3 and rabphillin were knocked down. These data indicate that rab11a and a select group of its binding partners play a prominent role in recycling of the human ß1-AR.     

Eicosapentaenoic acid decreases expression of anandamide synthesis enzyme and cannabinoid receptor 2 in osteoblast-like cells

Anandamide (AEA) is an endogenous agonist for the cannabinoid receptor 2 (CB2) which is expressed in osteoblasts. Arachidonic acid (AA) is the precursor for AEA and dietary n-3 polyunsaturated fatty acids (PUFA) are known to reduce the concentrations of AA in tissues and cells. Therefore, we hypothesized that n-3 PUFA, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), which reduce AA in cells, could lower AEA in osteoblasts by altering enzyme expression of the endocannabinoid (EC) system. MC3T3-E1 osteoblast-like cells were grown for 6, 10, 15, 20, 25 or 30 days in osteogenic medium. Osteoblasts were treated with 10 μM of AA, EPA, DHA, oleic acid (OA) or EPA+DHA (5 μM each) for 72 h prior to their collection for measurement of mRNA and alkaline phosphatase (ALP) activity. Compared to vehicle control, osteoblasts treated with AA had higher levels of AA and n-6 PUFA while those treated with EPA and DHA had lower n-6 but higher n-3 PUFA. Independent of the fatty acid treatments, osteoblasts matured normally as evidenced by ALP activity. N-acyl phosphatidylethanolamine-selective phospholipase D (NAPE-PLD), fatty acid amide hydrolase (FAAH) and CB2 mRNA expression were higher at 20 days compared to 10 days. NAPE-PLD and CB2 mRNA was lower in osteoblasts treated with EPA compared to all other groups. Thus, mRNA expression for NAPE-PLD, FAAH, and CB2 increased during osteoblast maturation and EPA reduced mRNA for NAPE-PLD and CB2 receptor. In conclusion, EPA lowered mRNA levels for proteins of the EC system and mRNA for AEA synthesis/degradation is reported in osteoblasts.     

丛枝菌根真菌在田间的分布特征、代谢活性及其对甘薯生长的影响

研究了大田条件下丛枝菌根(AM)真菌的分布特征、代谢活性及其对甘薯的生长效应.结果表明,接种Glomus intraradices 8周后,甘薯地上部干重,薯块鲜重和薯块个数均明显高于不接种对照;植株地上部和根系的吸磷量显著提高.与不接种对照相比,接种处理的甘薯菌根侵染率、甘薯根外菌丝密度以及甘薯根内菌丝的活性(根内菌丝碱性磷酸酶活性)显著提高.进一步分析甘薯根际不同方位上的菌丝分布,发现接种处理中平行于垄的方向的菌丝密度显著高于苗子下方的菌丝密度,而不接种处理的各个方向总菌丝密度无差异;活菌丝(具琥珀酸脱氢酶活性的菌丝)密度在各个方向的分布规律与总菌丝密度的分布规律一致.接种后根内菌丝活性的增强,根外活性菌丝密度的增加及其分布特征的改变,是甘薯产量增加的主要原因.     

Protein repertoire impact of Ubiquitin-Proteasome System impairment: insight into the protective role of beta-estradiol

The Ubiquitin-Proteasome System (UPS) and the Autophagy-Lysosome Pathways (ALP) are key mechanisms for cellular homeostasis sustenance and protein clearance. A wide number of Neurodegenerative Diseases (NDs) are tied with UPS impairment and have been also described as proteinopathies caused by aggregate-prone proteins, not efficiently removed by proteasome. Despite the large knowledge on proteasome biological role, molecular mechanisms associated with its impairment are still blur. We have pursued a comprehensive proteomic investigation to evaluate the phenotypic rearrangements in protein repertoires associated with a UPS blockage. Different functional proteomic approaches have been employed to tackle UPS impairment impact on human NeuroBlastoma (NB) cell lines responsive to proteasome inhibition by Epoxomicin. 2-Dimensional Electrophoresis (2-DE) separation combined with Mass Spectrometry and Shotgun Proteomics experiments have been employed to design a thorough picture of protein profile. Unsupervised meta-analysis of the collected proteomic data revealed that all the identified proteins relate each other in a functional network centered on beta-estradiol. Moreover we showed that treatment of cells with beta-estradiol resulted in aggregate removal and increased cell survival due to activation of the autophagic pathway. Our data may provide the molecular basis for the use of beta-estradiol in neurodegenerative disorders by induction of protein aggregate removal.     

Association of three SNPs in interleukin-28B with graft hepatic dysfunction after liver transplantation in Chinese Han population

Background

Recurrent graft infection limited the effect of LT, early recognition and prophylaxis of HBV recurrence are very important, and interleukin 28B (IL‐28B) gene was reported to be associated with HBV infection.

Aims

To explore the association between IL-28B single-nucleotide polymorphisms (SNPs) and graft re-infection after liver transplantation(LT).

Methods

21 recipients with hepatitis B virus(HBV) recurrence and 157 recipients without HBV recurrence were included. We studied three SNPs in the promoter region of IL-28B gene at the positions rs12979860, rs12980275 and rs8099917 by HRM analysis (high-resolution melting curve analysis).

Results

Hepatic allograft dysfunction was more likely to be associated with IL-28B SNPs. However, there was no significant difference in the frequencies of IL-28B gene distribution in recipients with or without HBV recurrence.

Conclusion

IL-28B gene polymorphism may be associated with the prognosis of LT recipients but it needs more experiments.     

A novel mutation of the SLC25A13 gene in a Chinese patient with citrin deficiency detected by target next-generation sequencing

Type II citrullinaemia, also known as citrin deficiency, is an autosomal recessive metabolic disorder, which is caused by pathogenic mutations in the SLC25A13 gene on chromosome 7q21.3. One of the clinical manifestations of type II citrullinaemia is neonatal intrahepatic cholestatic hepatitis caused by citrin deficiency (NICCD, OMIM# 605814). In this study, a 5-month-old female Chinese neonate diagnosed with type II citrullinaemia was examined. The diagnosis was based on biochemical and clinical findings, including organic acid profiling using a gas chromatography mass spectrometry (GC/MS), and the patient's parents were unaffected. Approximately 14 kb of the exon sequences of the SLC25A13 and two relative genes (ASS1 and FAH) from the proband and 100 case-unrelated controls were captured by array-based capture method followed by high-throughput next-generation sequencing. Two single-nucleotide mutations were detected in the proband, including the previous reported c.1177+1G>A mutation and a novel c.754G>A mutation in the SLC25A13 gene. Sanger sequence results showed that the patient was a compound heterozygote for the two mutations. The novel mutation (c.754G>A), which is predicted to affect the normal structure and function of citrin, is a candidate pathogenic mutation. Target sequence capture combined with high-throughput next-generation sequencing technologies is proven to be an effective method for molecular genetic testing of type II citrullinaemia.