Challenges in Maintaining the Genetic and Phenotypic Properties of Cell Culture

Challenges in Maintenance of Genotype and Phenotype of Cell Culture

Discuss The Challenges You Might Expect In Maintaining The Genetic And Phenotype Of Your Cells In Culture. Need Turnitin Report Plagiarism Free Work. In Text Citation For All Of The Paragraph. Provide Answers In Your Own Words.

The cells which are present in the bone marrow region and the blood flowing through the circulatory system are known as a hematopoietic stem cell (HSC) or blood stem cell. The HCS can develop into a different type of blood cells based on the trigger response received by the surface signaling system (Bruns, et al., 2014). The present report will discuss the challenges faced in the preservation of genetic and phenotypic properties of cell culture.

The genotype of a cell is defined as the genetic characteristics of a cell which differentiate from other cells. The phenotypic properties of cell culture are defined as the observable characteristic of cells in terms of morphology and biochemical properties. The recent developments in biomedical and biotechnological techniques have helped researches to develop an artificial micro-environment for the observation and cultivation of stem cell which is known as in vitro studies. The HSC’s properties to be multi-potent in nature along with their self-renewal capability were considered to have an indeterminate lifespan. However, the study conducted by (Sieburg, Cattarossi, & Muller-Sieburg, 2013), indicated that genetically identical HCS have a variable lifespan.  The genetic markups for the development of the stem cell either again in stem cell or final differentiated form are stored in the deoxyribonucleic acid (DNA), however, their expression is highly dependent on extracellular matrix (ECM) (Choi, Mahadik, & Harley, 2015). The maintenance of ECM is a highly challenging factor, as it hosts the cell culture environment and the signals direct the phenotypic development of HSC and hence the maintenance of the genotypic integrity of HSC. In order to maintain potential value stem cells in terms of its regenerative properties and its potency, a high level of precision is required in terms of the composition of ECM.

The identification of HSC is still one of the major challenges as the non-availability of reliable in situ assays along with the incompletely defined surface markers (Yokota, Oritani, Butz, Ewers, Vestweber, & Kanakura, 2012).

The monitoring of oxygen level in the in-vitro condition is a challenging aspect to maintain HSC cell culture, as the concentration of in the proximity of bone marrow region is 1.3 % to 4.2 % relative low than the rest of the body corresponding to the partial pressure of oxygen at 10 to 32 mm Hg (Spencer, et al., 2014). Therefore in order to maintain the genotypic and phenotypic nature of HSC culture oxygen monitoring should be carried out throughout the cell culture process (Zhang & Sadek, 2014).

The Importance of ECM in Preserving Genotypic Integrity of HSC

The HSC culture is generally grown in media, which are without any antibacterial or antifungal substance, therefore they are highly prone to contamination of fungal or bacterial infection, and therefore it is prime importance to maintain aseptic condition throughout the process of growth of animal cell culture. In a study conducted by (Fountain, et al., 1997), indicated that 1.2 % of the culture which was earlier stored in liquid nitrogen, when thawed and observed were contaminated with different kinds of waterborne and environmental organisms.

The kind of culture condition selection is also very important, in one of the studies reported by (Schmal, Seifert, Schäffer, Walter, Aicher, & Klein, 2016), indicated that 2D culture process was more favorable for the growth of hematopoietic stem and progenitor cells (HSPCs) in comparison to 3D growth condition in a mixed culture study of mesenchyme stromal cells (MSCs) with HSPCs. Therefore to maintain the genotypic and phenotypic activity of HSCs it is of prime importance that suitable culture conditions should be provided.

The dynamic cell culture condition faces stress due to the buildup of secondary metabolites along with the hydrodynamic stress, which makes the growth process difficult. The phenotype of cell culture is highly dependent on the pH; therefore continuous pH maintenance is very important for the prevention of cell culture (Kowalczyk, Waldron, Kresnowati, & Danquah, 2011).   

Conclusion

The properties of HSC to differentiate into the number of different type of cells and their self-renewal property make them a perfect candidate for many animal tissue culture studies along with the applicant in medical treatment. However, the genetic and phenotypic preservation of HSC has presented a difficult task as there are number of factors such as ECM, non-availability of the identification marker along with the controlled environment in terms of oxygen concentration.  In terms of culture condition, 2D growth conditions should be preferred in comparison to 3D growth conditions.

Reference

Bruns, I., Lucas, D., Pinho, S., Ahmed, J., Lambert, M. P., Kunisaki, Y., et al. (2014). Megakaryocytes regulate hematopoietic stem cell quiescence through CXCL4 secretion. Nature medicine, 1315.

Choi, J. S., Mahadik, B. P., & Harley, B. A. (2015). Engineering the hematopoietic stem cell niche: Frontiers in biomaterial science. Biotechnology journal, 1529-1545.

Fountain, D., Ralston, M., Higgins, N., Gorlin, J. B., Uhl, L., Wheeler, C., et al. (1997). Liquid nitrogen freezers: a potential source of microbial contamination of hematopoietic stem cell components. Transfusion, 585-591.

Kowalczyk, M., Waldron, K., Kresnowati, P., & Danquah, M. K. (2011). Process challenges relating to hematopoietic stem cell cultivation in bioreactors. Journal of industrial microbiology & biotechnology, 761-767.

Schmal, O., Seifert, J., Schäffer, T. E., Walter, C. B., Aicher, W. K., & Klein, G. (2016). Hematopoietic stem and progenitor cell expansion in contact with mesenchymal stromal cells in a hanging drop model uncovers disadvantages of 3D culture. Stem cells international.

Sieburg, H. B., Cattarossi, G., & Muller-Sieburg, C. E. (2013). Lifespan differences in hematopoietic stem cells are due to imperfect repair and unstable mean-reversion. PLoS computational biology, 1-15.

Spencer, J. A., Ferraro, F., Roussakis, E., Klein, A., Wu, J., Runnels, J. M., et al. (2014). Direct measurement of local oxygen concentration in the bone marrow of live animals. Nature, 269-273.

Yokota, T., Oritani, K., Butz, S., Ewers, S., Vestweber, D., & Kanakura, Y. (2012). Markers for hematopoietic stem cells: histories and recent achievements. Advances in Hematopoietic Stem Cell Research, 77-78.

Zhang, C. C., & Sadek, H. A. (2014). Hypoxia and metabolic properties of hematopoietic stem cells. Antioxidants & redox signaling, 1891-1901.