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Magnetic gold nanoparticles (mGNPs) with uniform size and morphology synthesized by our sonication treatment method were covalently bound with fluorescein isothiocyanate (FITC) molecules. Driven by an external magnetic field, FITC-labelled nanoparticles were delivered into plant cells with and without cell walls, evident from sectional transmission electron microscopy images. Confocal images further indicate that the green fluorescence in canola protoplasts and walled cells indeed came from the FITC molecules, instead of the chloroplasts' autofluorescence. FITC- labelled nanoparticles had a delivery efficiency of 95% based on confocal images. In further study, plasmids were covalently bound with mGNPs, and delivered into canola cells with and without cell walls. After culturing for 48 h followed by staining with 5-bromo-4-chloro-3-indolyl-β-D-glucuronic acid (X-Gluc), blue colour appeared in the protoplasts, while the walled canola cells showed a green colour that can be interpreted as the combination of blue and yellow from the suspension cells themselves. The presence of the blue colour indicates the expression of the GUS gene; therefore, the plasmids were successfully delivered into the canola cells. Furthermore, on examination, mGNPs were considered to be noncytotoxic by fluorescein diacetate staining. Key words: magnetic gold nanoparticles, FITC-labelled nanoparticle delivery, plasmid delivery, canola protoplast, walled canola cells, confocal image, sectional transmission electron microscopy image. A l'aide de leur traitement de sonication, les auteurs ont synthetise des nanoparticules d'or magnetiques (mNPOs) de morphologies et dimensions uniformes, et les ont attachees par lien covalent a des molecules d'isothiocyanate de fluoresceine (ITCF). Sous l'effet d'un champ magnetique externe, ils ont insere ces nanoparticules marquees au ITCF dans des cellules vegetales, avec ou sans paroi cellulaire, et mises en evidence par imageries en sections a l'aide de la microscopie electronique par transmission. Les images confocales demontrent de plus que la fluorescence verte dans les protoplastes de canola et les cellules avec paroi provient bien des molecules ITCF, plutot que de l'autofluorescence des chloroplastes. Les nanoparticules marquees au ITCF montrent une efficacite d'implantation de 95 %, basee sur les images confocales. Pour la poursuite de l'etude, les auteurs ont lie des plasmides en covalence avec des mNOPs avant de les implanter dans des cellules de canola avec ou sans paroi. Apres 48 heures de culture suivi d'une coloration avec X-Gluc, on observe une couleur bleue dans les protoplastes, alors que les cellules avec paroi montrent une couleur verte que l'on peut interpreter comme une combinaison du bleue et de la couleur verte de la suspension des cellules elles-memes. La presence de la couleur bleue indique l'expression du gene GUS, ce qui confirme l'implantation reussie des plasmides dans les cellules de canola. De plus, on a constate que les mNOPs ne sont pas toxiques lors de la coloration a l'diacetate de fluoresceine. [Traduit par la Redaction] Mots-cles : nanoparticules d'or magnetique, implantation de nanoparticules marquees a la ITCF, implantation de plasmides, protoplastes de canola, image confocale, imagerie en sections a l'aide de la microscopie electronique par transmission., Introduction In recent years, nanomaterials have been increasingly used in biomedical applications, such as gold nanoparticles for drug and gene delivery into mammalian cells (Levy et al. 2010; Ghosh et [...]

Objective: To explore the effect of myeloid leukemia-1 (Mcl-1) gene on the proliferation and apoptosis of Hela cells and the sensitivity of cervical cancer chemotherapy by antisense technology., Methods: Mcl-1 antisense oligonucleotide(AS-ODN)was transfected into Hela cells with lipofectamine 2000. The expression of Mcl-1 was analyzed by Western blot, the cell viability was detected by MTT assay, and apoptosis was evaluated by flow cytometry., Results: Mcl-1 AS-ODN arrested the cell cycle at G1/S,greatly inhibited the cell growth and induced apoptosis. The sensitivity of Hela cells on chemotherapy was low. There was obvious increase in the apoptosis rate by chemotherapy drugs and growth inhibition rate after inhibiting the expression of Mc1-1., Conclusion: Mcl-1 AS-ODN can not only inhibit Hela cell proliferation and induce apoptosis, but also increase the sensitivity of chemotherapy. Mcl-1 may be a potential target gene for cervical cancer chemotherapy.

Introduction: It was recently suggested that heat shock protein (HSP)70, an intracellular protein, is a potential mediator of inflammatory disease when it is released into the extracellular compartment. Although elevated HSP70 levels have been identified in rheumatoid arthritis (RA) synovial tissues and RA synovial fluid compared with patients with osteoarthritis and healthy individuals, it remains unclear what role extracellular HSP70 plays in the pathogenesis of RA. This study was conducted to investigate the effects of extracellular HSP70 on the production of RA-associated cytokines in fibroblast-like synoviocytes from patients with RA and to elucidate the mechanisms involved., Methods: IL-6, IL-8 and monocyte chemoattractant protein (MCP)-1 levels in culture supernatants were measured using enzyme-linked immunosorbent assays. Activation of mitogen-activated protein kinases (MAPKs), such as extracellular signal-regulated protein kinases (ERKs), c-Jun amino-terminal kinase (JNK) and p38 MAPK, was detected using Western blotting. Nuclear translocation of nuclear factor-kappaB (NF-kappaB) and degradation of the inhibitory protein IkappaBalpha were examined using immunohistochemistry and Western blotting., Results: Human HSP70 downregulated IL-6, IL-8 and MCP-1 production in RA fibroblast-like synoviocytes induced by tumour necrosis factor (TNF)-alpha in a concentration dependent manner. HSP70 inhibited the activation of ERK, JNK and p38 MAPK in fibroblast-like synoviocytes stimulated by TNF-alpha. Furthermore, HSP70 also significantly inhibited nuclear translocation of nuclear factor-kappaB and degradation of IkappaBalpha induced by TNF-alpha., Conclusion: Extracellular HSP70 has an anti-inflammatory effect on RA by downregulating production of IL-6, IL-8 and MCP-1 in fibroblast-like synoviocytes, which is mediated through inhibited activation of the MAPKs and NF-kappaB signal pathways.

The objective of this study was to evaluate the negative regulatory role of heat shock protein 70 (HSP70) on endotoxin-induced activation of inflammatory cytokine signaling pathways in a macrophage cell line. Our studies show that elevation of HSP70 either by activation of the heat shock response (HSR) or through forced expression of the hsp70.1 gene downregulates cytokine expression. Our experiments showed that activation of the HSR and HSP70 overexpression could inhibit LPS-mediated expression of the proinflammatory cytokines TNF-alpha and IL-1 at the mRNA and protein levels. We also investigated the effects of HSP70 elevation on signaling pathways downstream of LPS and its receptors, including the NF-kappaB and mitogen-activated protein kinase (MAPK) pathways. The effects of HSP70 on cytokine expression were correlated with its effects on activation of NF-kappaB, a known activator of the tnfalpha and Il-1 genes. Overexpression of HSP70 inhibited the nuclear translocation of p65, the transcriptionally active component of the NF-kappaB complex, and prevented the degradation of IkappaBalpha, the regulator of NF-kappaB activity. However, HSP70 elevation did not markedly inhibit signaling through the MAPK arm of the LPS-induced pathway, suggesting that the effects of HSP70 are mediated primarily through the NF-kappaB cascade. Our experiments therefore suggested that elevated levels of HSP70 inhibit LPS-induced production of inflammatory cytokines by a mechanisms involving inactivation of NF-kappaB but cast doubt on significant role for the MAPK pathway in these effects.