by Caitlin Smith
Need more physiological relevance in your work? Cell lines giving you inexplicable results? Moving to primary cell cultures could help – and it’s not as hard as you think. In fact, commercial primary cultures may give better and more consistent results than their homegrown counterparts. And vendors are quickly expanding their cell type offerings, along with specialized media.
The most famous types of primary cells, of course, are stem cells. “A tremendous amount of work using embryonic, induced, and adult stem cells is rapidly changing the way researchers perceive primary cells,” says Ben Buehrer, VP and CSO at Zen-Bio. “We have found a much greater interest in using human primary cells over the past several years.” In addition, culture quality is improving, and tools for genetic manipulation are increasingly available. “All these technologies allow researchers to use primary cells in ways that they have never been able to before,” says Joydeep Goswami, VP for primary and stem cell systems at Life Technologies, which markets primary cells under the Invitrogen brand, “and reduce the reliance on immortalized lines that often do not adequately replicate the performance of primary cells.”
Improving culture quality
Such a change in culture reliance may have already begun. For example, “40% of all cell biologists who use continuous cell lines in their research also use primary cells to achieve their experimental objectives,” says Robin Rothrock, director of product line strategy at ATCC. “The use of primary cells is widespread within the scientific community.”
Increasing availability and advances in quality are key factors in the growing use of primary cultures. Primary cell quality is determined by many factors that must happen concurrently to produce highly pure cell populations for primary cultures. “It’s a combination of the tissue, the skill of the person doing the dissection in the art of cell isolation, and superior media,” says Robin Rothrock, business unit brand director for tissue biology at ATCC. “Because even doing that initial isolation in the presence of superior media, at the end of the day, is going to give you better cells.” ATCC’s recent offerings include highly pure populations of human renal epithelial cells.
Others agree that cell isolation and media play a large role. “The availability of high-quality cells is addressed by advances in tissue isolation and cell biology that allow the design of better media to enable cells to survive,” says Goswami. Indeed, research efforts into factors that make primary cells happy are paying off. “We are investing significant resources into research and development through our subsidiary Lifeline Cell Technology to isolate primary cells and create optimized media for the support of each of these primary cell types,” says Jeffrey Janus, senior VP of International Stem Cell and CEO of Lifeline Cell Technology.
StemCell Technologies also has designed optimized media for the maintenance or differentiation of particular types of primary cells. Clive Glover, senior product manager for pluripotent stem cell biology at StemCell Technologies, notes an example of this challenge in mesenchymal stem cells, which “hold the potential to be used as a cell therapy against many diseases such as graft versus host disease and Crohn's disease, and to repair the myocardium following acute myocardial infarction,” he says. “To use these cells in clinical trials, a specialized cell culture medium must be designed that allows their expansion with full retention of their clinical function. The culture environment must also be animal component-free to avoid the possibility of transmission of animal viruses to the patient. MesenCult-ACF is a novel serum-free, animal component-free medium designed specifically to expand mesenchymal stem cells.”
Rather than developing tools for many types of cells, Life Technologies concentrates on providing a complete array of products for a few types. “The focus on complete workflow solutions on a few focused cell types [means] we are able to serve customer needs ranging from basic research, drug screening and cell therapy,” says Goswami. Their workflow includes cells, media, and the characterization, isolation, expansion, cryopreservation and engineering of primary cells. “Epilife/ S7 are providing researchers and therapy providers a way to expand keratinocytes, while technologies such as BacMam and Neon are helping them insert DNA into these cells much more easily and efficiently,” Goswami says.
The Health Protection Agency (HPA) Culture Collections in the UK, in contrast, offers over 100 types of primary cells, for which specialized media is available from the European Collection of Cell Cultures, one of four collections that constitute the HPA Culture Collections. “Our aim is the provision of primary cultures as near-physiologically relevant cell lines,” says Peter Thraves, senior scientist at HPA Culture Collections. “The majority [of our primary cell types] are only 2 passages away from the original tissue. The low passage number maximizes the number of population doublings left in the cultures and the physiological relevance of the cells.”
The importance of being human
Any cell culture work that aims for therapeutic applications in humans may gain more biologically relevant results from using human primary cell cultures. “A shift in use from animal cell and cell lines to human primary cells is essential to more realistically and rigorously model expected in vivo responses and, therefore, to advance both discovery and therapeutic translation,” says Ron Jankowski, director of research and product development at Cook MyoSite, which provides products for research on human primary skeletal muscle cells. One difficulty with studying human primary cells is that availability of tissue, especially healthy tissue, is often limited. “Primary cells must be studied as true controls along with the diseased cell type to understand what is ‘normal,’” says Janus. “They are also quite important to determine if a drug is a safe treatment for normal versus immortal cancerous or diseased cells.”
Other vendors seek to supply tools to researchers in specific areas of disease research. For example, Zen-Bio, which offers many types of human primary cells, is developing “a purified population of primary human mammary luminal epithelial cells and their donor-matched myoepithelial cells,” says Ben Buehrer, VP and CSO at Zen-Bio. “This will provide researchers with two distinct populations for breast cancer research. Currently there are no other human primary mammary luminal cells commercially available, so we are trying to fill that niche.”
Tissue engineering
Tissue engineering, or culturing primary cells in an approximation of their tissue of origin, is becoming more accessible too. “Tissue engineering and organogenesis are for me the most exciting development of the use of primary cells,” says Samuel Constant, chief operating officer at Epithelix. Their long shelf-life, reconstituted tissues are particularly suited for studying human respiratory diseases as well as for the evaluation of long-term chronic effects of drugs and the toxicity of potentially hazardous chemicals. “Epithelix is specialized in tissue engineering and focused on the human respiratory tract,” says Constant. “We sell 3D Human Airway Epithelia reconstituted in vitro called MucilAir™. MucilAir™ is in a homeostatic state and these tissues remain fully differentiated and functional up to one year.”
Janus expects to see an increase in organogenesis-type model systems. “Three-dimensional culturing of cells will allow an extended lifespan as well as a better model of in vivo function,” he says. International Stem Cell, through Lifeline Cell Technology, offers a three-dimensional skin model with more models – including a human corneal model – in development. “It is also possible that as we learn more about stem cells, especially mesenchymal and parthenogenetic stem cells, we will learn to differentiate them into pure cultures of primary cells that will have all the same or similar characteristics as primary cells derived from human tissue.” Indeed, researchers are finding combinations of precursors and optimized media that give strong, dependable differentiation. For example, according to Paul Diehl, director of commercial operations at B-Bridge International, “[We] sell rat osteoclast precursors with optimized differentiation medium that generates a differentiation rate of over 80%.”
Primary Cell, a partner of sells osteoclast precursors cells that grow and differentiate well, according to Primary Cell scientist Kyoko Shimizu.
“I still think that primary cell use will continue to grow,” says Buehrer. “New isolation, propagation, and culture techniques have made it easier and more cost effective to use primary cells. Also, the innovations with stem cells will continue to keep primary cells at the forefront of research.”