Cellular senescence is a barrier to unlimited cellular proliferation. Senescent cells are alive and metabolically active but are unable to further divide. Cellular senescence may be the consequence of a number of stressing stimuli such as telomere shortening, exposure to DNA damaging agents and the expression of activated oncogenes. Its study impinges on a number of important biological mechanisms related to organismal aging and tumor development. Therefore, the study of the pathways involved in its establishment, maintenance or escape is of interest for many scientists.
Unfortunately, senescent cells are not amenable to the usual cellular techniques commonly used in molecular and cellular biology to modulate cellular factors: plasmids cannot be efficiently transfected by standard calcium precipitation or liposomes-based techniques and microinjection is hard due to the flattened morphology of senescent cells. Furthermore, their general little availability make them precious and suggest the use of approaches that minimize their use yet retain informative value. An approach can be the use of lentiviral vectors, recombinant viruses that can integrate and express also in non-dividing cells. The generation of lentiviruses expressing RNA interference transcripts against cellular genes has opened the possibility to knockdown individual genes in senescent cells.
Nevertheless, not all molecular biology laboratories are equipped or have experience with lentivirus production.
An easier and equally efficient alternative is the transfection of short interefering RNAs (siRNAs) by lipofection. Differently from plasmid DNA, siRNAs can efficiently enter the cells and their transfection efficacy can be directly monitored by the use of fluorescent labeled RNA oligonucleotides.
We present here a brief method that allows the transfection of human senescent cells (regardless of the cause of cellular senescence) with an efficiency of around 70%.