Cryopreservation of Stem Cells
With stress-filled modern lifestyles causing a sharp uptick in long term and chronic health problems, there is increasing interest in the field of cryopreservation of stem cells as a contingency plan against future ailments. The great utility of preserving these cells boils down to one simple fact: stem cell are a guaranteed match with the patient-turned-donor due to their autologous nature, that is, there is no chance of rejection since the cells are from the patient’s own body. Some potential areas of application are:
1. Infertility:
·Frozen pre-pubertal testicular tissue can be utilized to restore fertility in cancer patients who lost their spermatogonial stem cells. There are also indications that human amniotic epithelial cells can be used to improve ovarian function in mice, which could be extended to human models as a management strategy for premature ovarian failure or insufficiency caused by cancer.
2. Neurodegenerative diseases:
With the discovery of neural stem cells, It is possible to delay the progression of diseases such as Parkinson’s or Alzheimer’s. Haematopoietic stem cells can be extracted by relatively non-invasive means (such as from umbilical cord blood or dental pulp), but a need arises to preserve them in a viable condition for long periods of time. In order to do so, the most commonly used cryopreservant is Dimethyl Sulfoxide, or DMSO. This molecule prevents the formation of water crystals within the cell and greatly reduces cell damage during the freezing phase. With current technology, cell cultures under long term preservation are generally estimated to retain viability for 7-15 years, depending on stability of storage conditions.
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Parameters to be monitored:
Due to the sensitive nature of the cells, various parameters must be monitored closely in order to ensure minimal damage and contamination, as highlighted below:
1. Temperature: In order to prevent pathogenic contamination or natural degradation, stem cells are frozen at very low temperatures of -196℃ in liquid nitrogen, -156℃ in vapor-phase nitrogen, or at -80℃ in mechanical freezers.
2. Freezing rate: Controlled freezing, wherein the temperature of the cells is decreased by small amounts over set periods of time (such as 1℃/min) up to a certain temperature, and then by larger amounts over similarly set time periods (such as 4℃/min), is agreed to be the safest approach to cell freezing. This is mainly due to the significant damage observed in samples where heat was liberated during uncontrolled freezing, resulting in intra-cellular crystalline ice formation. Recent studies support this method’s desirability, as cells frozen under these protocols showed high viability percentages post-thawing.
3. Cell concentration: In the past, a high cell concentration (beyond 2 × 10−7/ml) was thought to be detrimental to the viability of the cells. However, recent studies have disputed this idea and have shown that higher cell concentrations are well-tolerated and viable.
4. Thawing: The standard method to preserve viability while minimising the toxicity of the cryopreservative is to warm the cells in a water bath at 37℃ until all ice crystals disappear.
5. Washing: DMSO, although a widely used cryopreservant, is known to be clinically toxic. In order to reduce its detrimental effects in the patient, a standard two-step dilution process is followed. However, this procedure is labour-intensive and susceptible to cell loss.
Risks to be Considered:
Even if all parameters are well under control, certain risks must be taken into consideration with respect to the final graft of stem cells such as:
1. Toxicity of DMSO: Although it was established to be a non-stem cell toxic preservant, DMSO has significant side effects in patients who receive the stem cells such as nausea, vomiting, abdominal cramps, cardiovascular, respiratory, renal, Neurotoxic and hepatotoxic symptoms. Trials with lower concentrations of DMSO resulted in proving that a 10% cryopreservant solution of DMSO was superior to other concentrations.
2. Contamination: Microbial contamination can prove severely detrimental to immunosuppressed and immunocompromised patients. Some bacteria are stable in liquid nitrogen, which might cause fatal contamination in the preserved cells. Moreover, bone marrow derived stem cells are more likely to be infected due to the nature of the harvesting procedure.
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Currently, cryopreservation facilities remain expensive and inaccessible to many due to the intricate nature of the process. Being a nascent field, it can be expected to grow greatly in the near future in terms of affordability and efficiency.
Ethical Implications of Stem Cell Technology
Stem cells have increased the expectations among doctors, scientists, patients, and the general public due to their ability of differentiating into various types of cells. In addition to developing novel human tissues and biomaterials created from stem cells for use in pharmaceutical genomics and regenerative medicine, stem cell researchers are working to find treatments for genetic abnormalities. Results observed from completed and ongoing clinical research studies show huge therapeutic potential for stem cell-based therapeutics in the treatment of degenerative, autoimmune, and genetic disorders.
However, the clinical applications of stem cell-based therapy has raised ethical and safety concerns. We shall see some of the ethical challenges faced regarding human embryonic stem cell (hESC), induced pluripotent stem cell (iPSC), and mesenchymal stem cell (MSC)-based therapies in the upcoming section.
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