HepG2 is among the most studied and used human liver cancer cell lines in biomedical research studies. HepG2 was originally derived in 1975 from the liver tissue of a 15-year-old Caucasian male with hepatocellular carcinoma and has become an important research tool of scientists and researchers studying liver functions, drug metabolism, toxicity, and liver cancer processes. The cell line has preserved most of the features of normal hepatocytes, but due to being immortalised, offers experimental benefits due to continual cell culture.
HepG2 plays an important role in the current research that could not be overemphasized. These cells present an intermediate step between in vitro and excruciating in vivo models as a source of liver-specific processes to study under controlled conditions using the researchers. As compared to primary hepatocytes, in which the length of cell viability and availability is limited, the HepG2 cells can be propagated and therefore provide the right cells to do long term studies and in prestandardized experiments.
HepG2 Development and Origin
The hepatology research field changed with the advent of the HepG2 cell lines. This line of cells first was developed by Dr. Barbara Knowles and coworkers at the Wistar Institute who were able to immortalize carcinoma cells of the liver that preserved most liver cell properties. The name “HepG2” shows the liver origin (Hep> Hepatocyte and G2> Second generation of the given cell line).
Of special interest is the fact that HepG2 has kept many other features of hepatocyte cells even in spite of being a cancerous cell. Such cells still retain liver-specific proteins and have some metabolic activity levels and react to different stimuli in the same way as normal liver cells do. This paring of immortalization and amenability to study using a functional cell has made HepG2 an invaluable asset with regard to the study of normal liver physiology and pathology.
Features and HepG2 Properties
HepG2 cells have a number of unique properties which qualify them to be used in different kinds of research. Morphologically these have an epithelial like look; the cells are polygonal which is characteristic of the hepatocytes. They are monolayers and will grow in adherence attaining confluence in a relatively short period of time in favorable culture conditions.
The HepG2 cells have most of the hepatocyte functions as a functional point of view. They make albumin, transferrin and other plasma proteins typical of normal liver cells. The cells also convey cytochrome P450 enzymes but in lesser amount compared to primary hepatocytes hence finding usefulness in drug metabolism analysis. Also, HepG2 cells have a tendency to slightly retain the ability toward glucose metabolism and may react to hormonal changes, insulin, and glucagon.
HepG2 genetic profile shows similarities and differences in relation to the normal hepatocyte. They maintain the expression of numerous liver specific genes, but they have mutations typical of hepatocellular carcinoma. Such duality renders them especially useful in the investigation of phenomena related to the transformation of normal livercyles to malignant ones.
Research and Drug Development Procedures
HepG2 has gained massive use in different areas of research due to its ease of use. Pharmacologically, the cells are used as the main hepatotoxicity screen. The HepG2 cell line is regularly used by the pharmaceutical industry in screening the potential drug candidates to the liver toxicity prior to moving the drug candidates to animal trials. In spite of being partial, the effect of the cells in metabolizing drugs using the cytochrome P450 pathways gives useful initial information with regard to drug safety.
HepG2 cell lines have come in handy in cancer studies especially. Scientists employ these cells in investigating the progression of hepatocellular carcinoma, evaluating new forms of therapy, and analyzing resistance. The cells also respond to other chemotherapy drugs, and targeted therapy, which render them practical in preclinical drug screening and combination therapy tests.
Another important field to use the HepG2 would be in environmental toxicology. The cells assist scientists in evaluating the hepatotoxic risks of environmental pollutants, industry, and food additives. They are standardized, and this explains why they could be reproduced in various studies and laboratories.
Culture Conditions and Caring
HepG2 cells can be cultured successfully only under given conditions which keep them viable and functional. They can grow in normal media which include DMEM or MEM containing 10 percent fetal bovine serum, antibiotics, and other supplements necessary. Most of the mammalian cell lines grow well at 37 o C at 5% CO2 atmosphere.
To ensure healthy growth of HepG2, subculturing should be often performed. When cells are at 8090 confluence (depending on density of the cells these passages may be after 3-4 days) cells should be passaged. The preservation of cell integrity and functioning can be provided through appropriate trypsinization tech and proper handling. Regular tests on what contaminates the samples and a morphological evaluation are some of the quality control you can use to be sure of quality results in the experiments.
The major protocols in maintaining HepG2 stocks are storage and cryopreservation. The cells are successfully frozen in media of 10 % DMSO at -80 o C in short-term or in liquid nitrogen at long-term storage. Successful recovery procedures and thawing protocol keep the cell alive even after cryopreservation.
Pros and Cons
HepG2 has many benefits that have encouraged its application in researches. The advantages to immortalized cells are the unlimited supply of experiment material with no variability over primary cell isolation. Their reproducible characteristics of growth and standardised protocols allow similar results both between the laboratories and at different points in time.
HepG2 is of special value in studies involving the hepatocyte, as a result of retaining liver-specific functions. In contrast with other cancer cell lines which can be deprived of the properties characteristic only to an organ, many features of hepatocytes can be preserved by HepG2 cells (such as protein production, some metabolic processes, and reaction to liver-specific stimuli).
Nonetheless, HepG2 also features some significant shortcomings that have to be taken into account by researchers. The major weakness is that they have less metabolic activity than the primary hepatocytes. HepG2 has low expression of the cytochrome P450 enzyme which may interfere with drug metabolism researches. Also they themselves are cancerous cells and hence may not be very true in normal liver physiology in every sense.
The advantages brought about by the immortalized nature brings about potential artifacts. HepG2 cells are not normal hepatocytes and they might respond to treatment in a different way and their transformed state may affect the results of an experiment. When designing studies and interpreting results these are some of the limitations that should be considered by researchers.
Future Ideas and Trends
The development of HepG2 research is promising and a number of interesting advancements are coming. The development of genetic engineering, such as CRISPR-Cas9 technology, were making it possible to produce modified lines of HepG2 cells with desired or improved features. Such designed cell lines may overcome some of the existing shortcomings, and they will continue having the benefits of the original HepG2.
The use of HepG2 is being transformed by three-dimensional culture systems and by organ-on-chip technologies. These superior culture techniques more closely reconstitute the liver microenvironment, and thus could augur better predictive inferences of HepG2-based studies. More comprehensive models are also shown when a co-culture system of HepG2 cells with other cells of the liver is utilized.
The coming together of omics technologies and HepG2 studies is allowing a fresh understanding of the liver biology and disease. HepG2 studies to address issues in genomics as well as proteomics and metabolomics are providing novel therapeutic targets and biomarkers in liver diseases. All of these measures will improve translational value of HepG2 research.
Conclusion
The HepG2 cell lines have already become a valuable instrumental component of liver studies, drug development, and toxicology researchers. Their immortalized cell nature and preservation of hepatocyte properties are their special features that became the gold standard of many studies of liver-related nature. Despite there being limitation to their use, they are still being developed technologically, making them more useful and accurate in their prediction.
HepG2 will probably continue to play a pertinent role in liver research as other more advanced models complement it as part of the changing research methods and advances in technology. It is important to know about the capabilities and limitations of HepG2 so that the researcher can know how to maximize the use of this cell in his study. Future research using HepG2 will discover more insights into liver research and disease that will help our future generations.
FAQs About HepG2
Q1 Why is HepG2 different to other liver cancer cell lines?
HepG2 cells are distinct in the fact that despite being born out of liver cancer, they maintained several properties of its normal counterparts, that is, hepatocytes. They retain the ability to act as sites of production of liver-specific proteins such as albumin, and to possess some metabolic functions, unlike most other cancer cell lines, which fail. Drug metabolism and liver functioning may be specifically studied using them.
Q2 What is the maximum length of the passage of the HepG2 cells and do they retain their traits during a long period?
The nature of HepG2 cells is being immortalized and, thus, they can be cultured indefinitely. Nevertheless, their characteristics can alter in long passages (usually after 20-30 passages). One should keep the use of cells to a period of 10-15 passages beyond the initial stock and it is regularly checked with respect to the expression of proteins and functional tests to maintain consistency.
Q3 Can HepG2 cells be used to understand drugs metabolism and toxicity?
Yes, to be restricted. HepG2 cells have low levels of cytochrome P450, which is used in the metabolism of drugs when compared to primary hepatocytes. They are good to do preliminary screening of toxicity and mechanistic experiments but cannot be used to make judgments of overall drug metabolism and this needs validation with primary hepatocytes or in vivo model.
Q4 What is the best culture environment of HepG2 cells?
The HepG2 cells thrive well in the DMEM or MEM with 10 percent fetal bovine serum antibiotics and other necessary precarious paraphernalia. Culture them in the presence of 5% CO2 gases and 37 o C temperature. Divide the subculture every 3-4 days when the cells have attained 80-90% growth, but using gentle techniques of trypsinization, to keep the cells undamaged.
Q5 Is it possible to study the issue of hepatitis virus infection in HepG2 cells?
HepG2 cells are less susceptible to hepatitis viruses, especially hepatitis B and C than primary hepatocytes. Although they may be infected in certain circumstances and are applied in certain viral research, they might not represent all elements of natural viral infection. Modified HepG2 lines or enhancement of other cell models is common in the case of complete research on the hepatitis virus.
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