Short hairpin RNA expression for enhancing the resistance of Bombyx mori (Bm) to nucleopolyhedrovirus in vitro and in vivo

A new paradigm of RNAi technology has been studied for enhancing the resistance to virus in plants and animals. Previous studies have shown that the Bombyx mori (Bm) U6 promoter based shRNA is an effective tool for inducing RNAi in Bombyx mori cell line. However, widespread knockdown and induction of phenotypes in Bm larvae have not been fully demonstrated. In this study, we examined Bm U6 promoter based shRNA expression for suppressing Bm nucleopolyhedrovirus (NPV) in the Bm cell line and silkworm larvae. We measured the relative expression level of replication genes of BmNPV in hemolymph of silkworm larvae and BmN cells transfected with recombinant targeting shRNA by quantitative real time polymerase chain reaction (PCR). These results indicated that the recombinant shRNA expression system was a useful tool for resistance to BmNPV in vivo and in vitro. The approach opens the door of RNAi technology as a wide range of strategies that offer a technically simpler, cheaper, and quicker gene-knockdown by recombinant shRNA for future genetics in silkworm Bm and other related species.Key words: RNA interference (RNAi), Silkworm Bombyx mori (Bm) cell line, short hairpin RNA (shRNA), Bm nucleopolyhedrovirus (BmNPV), quantitative real time polymerase chain reaction, Bm U6 promoter

Genes related to the very early stage of ConA-induced fulminant hepatitis: a gene-chip-based study in a mouse model

<p>Abstract</p> <p>Background</p> <p>Due to the high morbidity and mortality of fulminant hepatitis, early diagnosis followed by early effective treatment is the key for prognosis improvement. So far, little is known about the gene expression changes in the early stage of this serious illness. Identification of the genes related to the very early stage of fulminant hepatitis development may provide precise clues for early diagnosis.</p> <p>Results</p> <p>Balb/C mice were used for ConA injection to induce fulminant hepatitis that was confirmed by pathological and biochemical examination. After a gene chip-based screening, the data of gene expression in the liver, was further dissected by ANOVA analysis, gene expression profiles, gene network construction and real-time RT-PCR.</p> <p>At the very early stage of ConA-triggered fulminant hepatitis, totally 1,473 genes with different expression variations were identified. Among these, 26 genes were finally selected for further investigation. The data from gene network analysis demonstrate that two genes, MPDZ and Acsl1, localized in the core of the network.</p> <p>Conclusions</p> <p>At the early stages of fulminant hepatitis, expression of twenty-six genes involved in protein transport, transcription regulation and cell metabolism altered significantly. These genes form a network and have shown strong correlation with fulminant hepatitis development. Our study provides several potential targets for the early diagnosis of fulminant hepatitis.</p

Sulforaphane induces adipocyte browning and promotes glucose and lipid utilization

Scope: Obesity is closely related to the imbalance of white adipose tissue storing excess calories, and brown adipose tissue dissipating energy to produce heat in mammals. Recent studies revealed that acquisition of brown characteristics by white adipocytes, termed “browning,” may positively contribute to cellular bioenergetics and metabolism homeostasis. The goal was to investigate the putative effects of natural antioxidant sulforaphane (1-isothiocyanate-4-methyl-sulfonyl butane; SFN) on browning of white adipocytes. Methods and Results: 3T3-L1 mature white adipocytes were treated with SFN for 48 h, and then the mitochondrial content, function, and energy utilization were assessed. SFN was found to induce 3T3-L1 adipocytes browning based on the increased mitochondrial content and activity of respiratory chain enzymes, whereas the mechanism involved the upregulation of nuclear factor E2-related factor 2/ sirtuin1/ peroxisome proliferator-activated receptor gamma coactivator 1 alpha signaling. SFN enhanced uncoupling protein 1 expression, a marker for brown adipocyte, leading to the decrease in cellular ATP. SFN also enhanced glucose uptake and oxidative utilization, lipolysis and fatty acid oxidation in 3T3-L1 adipocytes. Conclusion: SFN-induced browning of white adipocytes enhanced the utilization of cellular fuel, and the application of SFN is a promising strategy to combat obesity and obesity-related metabolic disorder

Neurospheres from rat adipose-derived stem cells could be induced into functional Schwann cell-like cells in vitro

<p>Abstract</p> <p>Background</p> <p>Schwann cells (SC) which are myelin-forming cells in peripheral nervous system are very useful for the treatment of diseases of peripheral nervous system and central nervous system. However, it is difficult to obtain sufficient large number of SC for clinical use, so alternative cell systems are desired.</p> <p>Results</p> <p>Using a procedure similar to the one used for propagation of neural stem cells, we could induce rat adipose-derived stem cells (ADSC) into floating neurospheres. In addition to being able to differentiate into neuronal- and glial-like cells, neurospheres could be induced to differentiate into SC-like cells. SC-like cells were bi- or tri-polar in shape and immunopositive for nestin and SC markers p75, GFAP and S-100, identical to genuine SC. We also found that SC-like cells could induce the differentiation of SH-SY5Y neuroblastoma cells efficiently, perhaps through secretion of soluble substances. We showed further that SC-like cells could form myelin structures with PC12 cell neurites in vitro.</p> <p>Conclusion</p> <p>These findings indicated that ADSC could differentiate into SC-like cells in terms of morphology, phenotype and functional capacities. SC-like cells induced from ADSC may be useful for the treatment of neurological diseases.</p

Suppression of <i>TREX1</i> deficiency-induced cellular senescence and interferonopathies by inhibition of DNA damage response

TREX1 encodes a major DNA exonuclease and mutations of this gene are associated with type I interferonopathies in human. Mice with Trex1 deletion or mutation have shortened life spans accompanied by a senescence-associated secretory phenotype. However, the contribution of cellular senescence in TREX1 deficiency-induced type I interferonopathies remains unknown. We found that features of cellular senescence present in Trex1−/− mice are induced by multiple factors, particularly DNA damage. The cGAS-STING and DNA damage response pathways are required for maintaining TREX1 deletion-induced cellular senescence. Inhibition of the DNA damage response, such as with Checkpoint kinase 2 (CHK2) inhibitor, partially alleviated progression of type I interferonopathies and lupus-like features in the mice. These data provide insights into the initiation and development of type I interferonopathies and lupus-like diseases, and may help inform the development of targeted therapeutics

Comparative study of mesenchymal stem cells from C57BL/10 and mdx mice

<p>Abstract</p> <p>Background</p> <p>Human mesenchymal stem cells (MSCs) have been studied and applied extensively because of their ability to self-renew and differentiate into various cell types. Since most human diseases models are murine, mouse MSCs should have been studied in detail. The mdx mouse – a Duchenne muscular dystrophy model – was produced by introducing a point mutation in the dystrophin gene. To understand the role of dystrophin in MSCs, we compared MSCs from mdx and C57BL/10 mice, focusing particularly on the aspects of light and electron microscopic morphology, immunophenotyping, and differentiation potential.</p> <p>Results</p> <p>Our study showed that at passage 10, mdx-MSCs exhibited increased heterochromatin, larger vacuoles, and more lysosomes under electron microscopy compared to C57BL/10-MSCs. C57BL/10-MSCs formed a few myotubes, while mdx-MSCs did not at the same passages. By passage 21, mdx-MSCs but not C57BL/10-MSCs had gradually lost their proliferative ability. In addition, a significant difference in the expression of CD34, not Sca-1 and CD11b, was observed between the MSCs from the 2 mice.</p> <p>Conclusion</p> <p>Our current study reveals that the MSCs from the 2 mice, namely, C57BL/10 and mdx, exhibit differences in proliferative and myogenic abilities. The results suggest that the changes in mouse MSC behavior may be influenced by lack of dystrophin protein in mdx mouse.</p