For the study of certain biological processes it is interesting to be able to grow cells in the laboratory ( in vitro ), instead of resorting to tissue samples ( in vivo ). This is especially relevant when studying how certain infectious agents develop, and the responses of cells to these agents. In this article we will explain the main techniques used for laboratory cell cultures, and some of their applications.
Cell culture overview
Laboratory tissue culture has its origins in the 19th century, when the English physician Sydney Ringer developed a saline solution capable of keeping an animal heart beating outside the body. The components of this solution, called Ringer’s Solution, were sodium, potassium, calcium, and magnesium chloride.
The technique later improved, and in the middle of the 20th century it made it possible to advance in the study of viruses, by being able to cultivate animal cells in the laboratory to be infected by them. Thus, the vaccine of Jonas Salk polio was the first to benefit from this technique in the 1950s.
Growth of cells in culture
For their growth, cells are kept in conditions of temperature and pH similar to those of the human body. The atmosphere in which cells grow is also important, as it must recreate the conditions in which they are found within the body, that is, 95% oxygen and 5% CO 2 that must be continuously eliminated.
Types of cell cultures
Cells can grow in isolation in a liquid medium (such as cells in the bloodstream) or on a surface (usually cells that are part of tissues). Cell surface cultivation is frequently carried out in Petri dishes, made up of two circular plastic parts that allow air to pass through. There are other techniques, although less widespread, that allow cell culture in 3D , for example by magnetic levitation.
A problem with culturing eukaryotic cells is that they can only divide a limited number of times (known as the Hayflick limit), due to restrictions in DNA replication related to the ends of chromosomes, called telomeres. Cells require a particular enzyme, telomerase, to prevent the degradation of telomeres in successive replications.
Since you want to work with specific tissue cells, the use of stem cells (which can divide continuously but are not differentiated) is not common. Instead, immortal tumor cells are used, which are differentiated and can divide unlimitedly.
Cell lines for culture and their applications
A group of similar cells that can be continuously grown in the laboratory is called a cell line. Of all the existing cell lines from tumor cells, the most widely used and known are HeLa cells . These are cells derived from a tissue sample affected by cervical cancer from an African-American woman named Henrietta Lacks, who died of this disease in 1951.
Its cells remain alive and have been used in countless cynical experiments since that year. The cell line name is an acronym for the patient’s name. These cells can divide indefinitely thanks to the fact that they have the enzyme telomerase .
A striking feature of HeLa cells is that they have 82 chromosomes instead of the 46 that are common in human cells. Due to the rapid division of this cell line, which causes errors during cell division, and the gene transfer effect of the human papillomavirus that had caused cancer in Henrietta Lacks, the chromosomes are tripled (3n) instead of duplicates (2n), as is usual in human cells, in addition to having additional copies of other chromosomes.
HeLa cell cell culture applications
HeLa cells have been used in numerous medical applications. In the 1950s they were used to test the efficacy of the polio vaccine developed by Jonas Salk. Cells infected by the virus died, while the vaccine prevented this from happening. Without the enormous number of HeLa cells that could be produced in the laboratory, the tests to obtain the vaccine would have been much more difficult to perform. In 1954 the vaccine was ready for extensive use in humans, which allowed the eradication of polio in many countries.
In addition, the study of HeLa cells identified the human papillomavirus and linked it to cervical cancer , a discovery that earned Harald zur Hausen the Nobel Prize in medicine in 2008 and led to the development of the HPV vaccine.
HeLa cells also allowed, almost accidentally by contaminating a different cell culture, the discovery of a staining technique that allows chromosomes to be observed with precision . This led to the first estimation of the number of chromosomes in HeLa cells, and also to the exact determination of the number of chromosomes in human cells, which until then were believed to be 48. This has had important applications in the study of genetic diseases. such as Down syndrome, caused by an alteration in the number of chromosomes (extra copy of chromosome 13).
On the other hand, these cells have been used extensively for the study of cancer and different treatments that specifically affect tumor cells. Thus, they have been used to study techniques that cause programmed cell death. In 1965, HeLa cells were hybridized with other animal cells, a milestone in genetic studies that allowed the development of genetic analysis techniques that led to the Human Genome Project, started in 1990, which culminated in the complete sequencing of the human genome in 2003. .
Other cell lines
Other human cell lines that are used in laboratories, to cite just a few examples, are BT-20 (breast cancer), Calu-3 (lung adenocarcinoma), DAOY (brain medulloblastoma), Jurkat (T lymphocytes, leukemia) or WM39 (melanoma). There are also cell lines from fetuses that did not develop, such as HEK293 from kidney epithelium.
Likewise, various cell lines from other animals are used, both tumor cells and embryonic cells. The most common are mice, but there are also established cell lines from dogs, hamsters, zebrafish, or some insects. A widely used cell type is Vero cells, derived from the kidney of the African green monkey ( Cercopithecus aethiops ). These are immortal cells with an abnormal number of chromosomes that are used in the study of various diseases.
Usefulness of cell cultures in the study of viruses
Since viruses require living cells to reproduce , their in vitro research requires cell cultures of the appropriate tissue. The presence of virus in cell cultures can be determined by fluorescence detection of antigens or by the appearance of dead cells in the cell culture, forming characteristic circular “plaques” in the culture medium. Some viruses that can be identified in a sample using these techniques are adenoviruses, enteroviruses, herpes, influenza, rhinovirus, chickenpox, or mumps.
Techniques are currently being developed for the study of SARS-CoV-2 in the laboratory using cell cultures. There is still no standardized pattern for these techniques , so different clinical trials have used different cell lines. The most widely used has been the Vero cell line (specifically Vero E6), but also others such as Caco-2 (human colorectal adenocarcinoma) or ATTC MK2 (Rhesus monkey kidney epithelium, Macaca mulatta ).
The culture of viruses in the laboratory is essential for the development of any vaccine, as it allows its efficiency to be tested in the early stages of clinical trials, before proceeding to human trials.