Holding solutions for grafts are essential for optimal hair transplantation results. Very little research, however, examines holding solutions ideal for follicular grafts. There are, in actuality, there are numerous facets we must consider both about holding solutions and their temperature. These include:
- How does chilling effect follicular grafts?
- If chilling is beneficial, then what are the best temperatures?
- What is the best holding solution and at what temperature?
- Are any holding solutions bad for our grafts?
Also significant, more and more transplant surgeons are considering additives to their holding solutions. These include antioxidants, adenosine triphosphate (ATP), and oxygen. One product of note, and still undergoing research, is Vitasol. This unique holding solution has ATP-rich liposomes that could be of major benefit.
Unfortunately, near all research into transplant preservation looks at major organs such as the kidney, liver, myocardium, and pancreas. This paper, in turn, will specifically focus on holding solutions and temperatures ideal for hair follicle grafts.
Below is a review of literature pertaining to follicular survival rates and health. My hope is to help direct and encourage greater study in this area of interest. Further, and thus far, no effort has been made to reproduce these studies. Each contributes to our greater knowledge, yes. While important, none have generated specific protocols. Hair specialists choose their holding solution and temperature according to their own judgment and knowledge. Useful for direction for further research, the do not withstand statistical scrutiny.
- Kim finds normal saline at 4 °C does not improve graft health unless more than 6 hours out of body. That mentioned, 6 hours out-of-body and always decreases the survivability of follicles regardless of holding solution temperature 4 °C. (1)
- A study by Beehner found that cold storage in normal saline improves the survival rate of grafts exposed to crush injury.(2)
- Uebel finds that 15% more follicles survive the transplantation process with platelet rich plasma (PRP). However, this has not been reproduced or confirmed.(3)
- Krugluger demonstrates that transient effluvium was stopped in 6 patients through PBS-buffered tissue culture medium (TCM) with nitric oxide added as an inhibitor of apoptosis.(8)
- Krugluger finds in a separate study that PBS containing B12 encourages better elongation of hair follicles that those stored in PBS alone.(9) Interesting, we also remain unclear if elongation is a good indicator of graft vitality.
- Beehner finds that chilling grafts in Hypothermosol and ATP improves the survival rate of grafts over normal saline. This considers a number of storage times, from 2 hours up to 96 hours, along with the usual 6-hour mark.(10)
- Forhair's Dr.Cole shows that pH declines in normal saline no matter temperature. This is a result of time itself. However, pH remained constant in PBS due to cellular buffers.(11)
Specific solutions seem to offer advantages to different types of transplant organs.(13) Many regular solutions are excellent at preserving tissue. Additives for solutions can also help improve tissue survivability.(14) No protocolz, solution, or temperature, however has yet been defined for hair grafts. With this in mind, we will look at the basic science behind biopreservation.
Basic Tenets and Theories
Most tissues have a very high oxygen requirement. Unfortunately, the tissue's oxygen solubility is quite low. Interrupting bloodflow rapidly inhibits aerobic energy. Loss of circulation also deprives cells of necessary metabolites and eliminates removal of waste products.
Excision of grafts for transplantation results in ischemia, or inadequate blood supply. Such events contribute to cell death. Ischemia in hair transplant surgery can last many hours and often exceeds 4-6 hours. Advances in holding solutions and temperature reduction may offer ways to improve hair transplant survival
Different cell types have different tolerances for maintaining optimal health. (19) For example, the ischemic tolerance of the brain is only 6 minutes at 37 degrees. This, however, extends to nearly 60 minutes when the body temperature reduces to 17 degrees. (16) Kidneys are able to tolerate much longer periods of "warm ischemia" (i.e., nonhypothermic ischemia). Total necrosis of the majority of tubules occurs after 60 minutes. As indicated by Kim, hair follicles at room temperature tolerate even longer periods of ischemia. (1)
Therefore, it makes sense to look for a preservation solution and temperature that is ideal for the hair follicle. We must approach storage solutions with a molecular-based logic to design the ideal holding solution for specific cell types of the hair follicle. Solutions should support the cellular proteome, genome, and fragmentome in addition to cellular structures such as the mitochondria, cell membrane, and nucleus.
The proteome is all the proteins expressed by a genome and the fragmentome refers to the peptide fragments.
Cost of various products extant today:
Normal Saline: $44.20 USD case of 12 1 OOOcc bottles BSS: $132.18 USD case of8 500cc bottles Ringer's Lactate: $146.53 USD case of 16 lOOOcc bottles Hypothermosol: $1740.00 USD case of30 lOOcc bottles Vitasol: (liposomal ATP) $1354.00 USD per liter
As you can see, the cost varies greatly between the different storage solutions, so it is important to examine the differences in the various storage solutions.
As mentioned, many storage solutions are available. The following two sections further detail intracellular and extracellular options.
Intracellular storage solution
These are hypertonic solutions with elevated K+ levels and reduced Na levels more similar to the intracellular space that is unable to cross the cell membrane. This provides osmotic support. Important, as reduction in temperature and water-controlling cell pumps that regulate the osmotic gradient are inactivated. The higher osmotic pressure found in these storage solutions inhibit passive influx of water and thereby reduce the risk of cell swelling. Examples of intracellular storage solutions include Hypothermosol, Collins, Euro Collins, Viaspan (University of Wisconsin Solution), CryoStor, Celsior, HTK-Custodial, Unisol, and KPS I.
Tissue stored in intracellular storage solutions should be chilled, but at what temperature? According to Dr. Abey Matthew, grafts stored in Hypothermosol should be kept below l2°C and above freezing with a recognized usage range of 2°-8°C.(20) The composition of Hypothermosol is noted below: (19)
Extracellular Storage Solutions
These are isotonic solutions with a plasma-like complement of ions that mimics the normal extracellular environment of cells. Examples include normal saline, Ringer's Lactate, BSS, and tissue culture media. These solutions offer lackluster preservation at reduced temperatures. They also lead to cell swelling at lower temperatures due to reasons explained above. For this theoretical reason, specialists should never chill extracellular storage solutions. Nevertheless, specialists commonly use these solutions in combination with hypothermic temperatures. To summarize, there are strong hypothetical reasons based upon decades of large organ transplant research that conclude the following:
- 1. Synthetic preservation solutions that specifically inhibit detrimental cellular changes like ischemia and hypoxia may be better than culture medium or saline, which specialists commonly use.
- 2. Chilling of grafts should only be done when the grafts are stored in an intracellular medium.
- 3. If using an extracellular medium to store grafts then they should avoid chilling tissues.
- 4. However, not chilling the tissue presents its own problems.The metabolic rate and associated ATP demands do not decrease while the supply of ATP dramatically diminishes with the loss of normal perfusion of the tissue.
Every organ has an optimal temperature for storage based on the interaction of hypothermia, the nature of the cell, and its chemical composition. For cardiac muscle, the optimal temperature is 10°-20° Celsius.(23) For the kidney, some studies show that 10° Celsius is superior to 5° and 5° is superior to 0.5° (23,25). We are yet to study the optimal temperature for hair grafts. Parsley, for example, feels the optimal storage temperature for hair follicle grafts is between 8°-14° Celsius, but he bases this on optimal temperatures for other organs rather than for hair follicles. (26)
Unfortunately, ice-based systems can lead to the temperature abruptly falling below 4°C and then staying there for a prolonged period of time. We do not know if the temperature below 4°C has a negative impact on the survival of hair follicles. Furthermore, we do not know if the abrupt drop in temperature is as good as a gradual decline in temperature. Some theorize that a gradual decline in temperature is better for hair follicle grafts. Dr. Parsley first moves his hair follicle tissue to a room temperature holding reservoir and then transfers it to an ice-based system to achieve a more gradual decline in temperature from body temperature.(25)
We always traumatize tissue when we take it from the body. Then we further traumatize it upon chilling. Finally, it is traumatized once again when warmed. Based on this theory of traumatization, it may be harmful to our grafts if we allow our grafts to rise above the low temperature in our holding solutions to a temperature closer to room temperature and then suddenly shock them again by re-chilling them. Striving to maintain a constant temperature throughout the chilling process is ideal based on this rationalization.
Consequently, I have created a chilling pump that circulates a cool mixture of propylene glycol through my storage dishes. I've found that this system allows me to maintain a constant temperature and may benefit someone interested in doing studies into the optimal temperature for graft storage. More recently, I created the Graft Chilling Plate (GCP). The GCP is a compact chiller designed to hold one or two petri dishes and is capable of storing grafts at a stable temperature throughout the duration of the hair transplant procedure. It will hold multiple stainless steel cones (developed by Dr. Bill Parsley) that allow the transfer of multiple grafts at one time.
Temperature Depends on Holding Solution
We know that certain tissues respond better to specific temperatures. Furthermore, some tissues respond better to a specific or customized holding solution. It remains for us to define these in terms of hair follicles. We remain uncertain whether there is any benefit from chilling grafts below room temperature. We do know, however, that, for reasons discussed, there is strong evidence that only hypertonic solutions should be used when chilling grafts. Furthermore, that data also suggest to only use isotonic solutions when maintaining grafts at room temperature.
Despite this knowledge, hair transplant data at this time suggest that chilled grafts in normal saline or Ringer's Lactate seem to thrive just as well as when they are placed at room temperature in these solutions. It is uncertain whether follicle survival is better in hypertonic solutions at a cool temperature as is the case with other organs. There are a variety of different holding solutions of both the intracellular and extracellular composition. For reasons stated in this article, extracellular holding solutions should not be chilled. Conversely, as discussed, do not store grafts at room temperature in intracellular holding solutions.
This is the primary reason Cole Instruments is designing chilling devices. This puts absolute control of temperature in the hands of specialists. Ideal for research and everyday procedures, our chilling devices include the ability to control temperature by each degree celsius. Learn more about this and other Cole Instrument devices by contacting with a representative today.
The original article was posted at forhair.com: The Optimal Holding Solution and Temperature for Hair Follicle