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Biomedical subjects

O Keil

Publications and source records attributed to O Keil.

14 recordsLinked to original sources
PubMed2006

A novel siRNA-lipoplex technology for RNA interference in the mouse vascular endothelium.

For the application of RNA interference (RNAi) in vivo the functional delivery of short interfering RNAs (siRNAs) is still the major obstacle. Therefore, delivery technologies need to be established for the systemic application of RNAi in vivo. Here we report uptake, biodistribution and in vivo efficacy of siRNA molecules formulated into siRNA-lipoplexes. The applied formulation is based on complex formation of positively charged liposomes, a mixture of cationic and fusogenic lipids complexed with the negatively charged siRNA. We determined by fluorescence microscopy the temporal and spatial distribution of fluorescently labeled siRNA-lipoplexes, the body clearance and endothelial cell type specific uptake after single intravenous injection. Furthermore, by using siRNA molecules for targeting endothelia-specifically expressed genes, such as CD31 and Tie2, we were able to demonstrate downregulation of the corresponding mRNA and protein in vivo. Taken together, we show the applicability of this non-viral delivery technology for inducing RNAi in the vasculature of mice after systemic application.

Animals↗
PubMed2006

RNA interference in the mouse vascular endothelium by systemic administration of siRNA-lipoplexes for cancer therapy.

RNA interference (RNAi) entails the potential for novel therapeutic strategies through the silencing of disease-causing genes in vivo. However, recent studies have raised an issue regarding applicable routes of administration for small interfering RNA (siRNA) molecules as therapeutics. In this study, we demonstrate that liposomally formulated siRNA molecules, the so-called siRNA-lipoplexes, but not naked siRNAs, are delivered to the tumor endothelial cells in vivo by microscopy. In addition, functional intracellular delivery of formulated siRNA targeting the tumor suppressor PTEN is shown in endothelial cells of the liver and tumor. Finally, the therapeutic potential of systemically administered siRNA(CD31)-lipoplexes is established by inhibition of tumor growth in two different xenograft mouse models. Our findings corroborate the applicability of this liposomal siRNA delivery technology for inducing RNAi to modulate gene expression levels in angiogenesis-dependent processes. In addition, our results advocate CD31 as a promising therapeutic target for antiangiogenic intervention. Therefore, our study provides a basis for the development of antiangiogenic cancer therapies based on RNAi.

3T3 Cells↗
PubMed2001

Novel lipophilic chloroquine analogues for a highly efficient gene transfer into gynecological tumors.

Liposomal vectors based on cationic lipids have been proven to be an attractive alternative to viral vectors in gene therapy protocols with regard to safety and manufacturing concerns. In order to improve the transfection efficiency we have synthesized two novel carboxycholesteryl-modified chloroquine analogues. Due to their potential endosomal buffering capacity these compounds enable the efficient transfection of various gynecological tumors and therefore are promising reagents in gene therapy applications.

Cell Survival↗
PubMed2001

Efficiency and toxicity of liposome-mediated gene transfer to corneal endothelial cells.

Gene transfer to corneal endothelial cells could be an important advance to modulate functions of these critical cells and is a field of current investigations. The development of gene transfer methods is a prerequisite for gene therapy to realize its full potential. We attempted to investigate and optimize the efficacy and safety of cationic liposome mediated gene transfer into corneal endothelial cells using different lipid formulations. Mono- and polycationic lipids and the neutral helper lipid dioleolphosphotidyl-ethanolamine (DOPE) were used for preparation of cationic liposomes. Six liposomal formulations containing DAC/DOPE 30/70 (DAC 30), DOSGA/DOPE 30/70 (DOSGA 30), DOSGA 100, DMRIE/DOPE 50/50 (DMRIE 50) and SP/DOPE 20/80 (SP 20) were complexed with the pUT 651-plasmid, encoding the E. coli beta-galactosidase gene. Subconfluent primary and passaged bovine corneal endothelial cells (BCEC) were transfected with different amounts of liposomes and DNA or uncomplexed free DNA as control. Quantitative expression of beta-galactosidase was measured using a colorimetric assay. In order to assess the effects on cell viability and growth, a modified acidic phosphatase assay was employed. Differences were detected using these various liposome preparations. Transfection experiments demonstrated the highest gene expression using SP 20> DMRIE 50 ranging at approximately 3 mU per beta-gal per well. Low expression of beta-galactosidase was achieved using DAC 30, DOSGA 30 and DOSGA 100. No beta-galactosidase expression was found in control dishes. There was no difference seen following transfection of primary or subsequent passages of BCEC. As indicated by the acid phosphatase assay, no significant toxicity was detected for the most efficient lipids used. Of the preparations studied, SP 20 appeared as the optimal vehicle for plasmid-mediated transfection of BCEC. The ability to deliver genes to BCEC via liposomes could be valuable, since the use of other vectors for transfection may be limited by undesired effects.

Animals↗
PubMed2000

Optimization of nonviral gene transfer of vascular smooth muscle cells in vitro and in vivo.

Gene therapy strategies for the prevention of restenosis postangioplasty are promising. Nonviral gene transfer to the arterial wall in vivo has so far been limited by poor efficiency. This study aimed to optimize transfection of primary vascular smooth muscle cells using cationic nonviral formulations based on cholesterol derivates (DC-, DAC-, DCQ-, and Sp-Chol), double-chained amphiphils (LipofectAMINE, DOTMA, DOSGA, DOSPER, and DOCSPER), or heterogeneous reagents (Superfect, Effectene, and Tfx-50). Estimation of transfection efficiencies was performed using galactosidase assays at different ratios of transfection reagent to plasmid DNA with reporter gene. Toxicity was monitored by analyzing cell metabolism. Transfer efficiency and safety were determined in a porcine restenosis model for local gene therapy using morphometry, histology, galactosidase assays, and reverse-transcriptase polymerase chain reaction. The highest in vitro transfection efficiency was achieved using the recently developed DOCSPER liposomes, with transfer rates of at least 20% in vascular smooth muscle cells. Transfer efficiency was further enhanced up to 20% by complexing with poly-L-lysine. Transfection efficiency in vivo in a porcine restenosis model was up to 15% of adventitial cells using DOCSPER versus 0.1% using LipofectAMINE. Toxicity in vivo and in vitro was lowest using DOCSPER. Increased biological effects were demonstrated following optimization of transfer conditions.

Angioplasty↗
PubMed1984

Assessment of the impact of medical devices on the quality of care.

Medical technology and medical devices play major roles in the diagnosis and treatment of patients in health care facilities. A facility should use a rational process to reach the decision to acquire a new piece of technology; this process should recognize the importance of both the machine and the user in modern technology. Moreover, each health care facility should assure that a newly acquired technological advance does not pose safety hazards to patients; that the device, when used in the clinical situation, produces the desired results reliably and consistently with its intended purpose; that there are appropriate means of identifying performance defects or opportunities for greater use of the existing device; and that the end of the device's useful life is anticipated so that quality does not decrease and dangers to patients do not increase due to equipment obsolescence. Successfully applying QA principles, consistent with each phase in the life of medical technological devices, should ensure equipment of high quality and thus benefit a health care facility and its patients.

Equipment Safety↗