Introduction
There are an estimated 300,000 anterior cruciate ligament (ACL) repairs each year in the United States. The aim of surgery is to restore the functional stability to the ACL-deficient knee. ACL reconstruction involves the replacement of the ruptured or damaged tendon, with a graft either procured intraoperatively from the patient (autograft) or recovered from human cadavers (allograft).
“The use of allograft tissue for anterior cruciate ligament reconstruction has increased steadily over the past decade.” Allograft options for ACL reconstruction include bone–patellar tendon–bone (BPTB), Achilles tendon, anterior tibialis tendon, posterior tibialis tendon, peroneus longus tendon, semitendinosus tendon, gracilis tendon, and fascia lata grafts. The use of allograft tissue has the advantages of reduced donor site morbidity, selectable graft sizes and types, decreased operative time, and decreased postoperative pain and stiffness. Additionally in cases of revision surgery where autograft options have already been exhausted, allografts provide an ideal tissue source.
The Allografts for ACL Reconstruction Survey Report by the American Academy of Orthopaedic Surgeons revealed that 47.8% of the respondents would be less likely to use allografts because of disease transmission concerns. Others in the survey cited risk of disease transmission as a reason they have decreased the use of allografts in their practice.
Despite concerns associated with allograft safety, a 2004 questionnaire of 340 hospitals showed that surgeons were involved in the selection of the source of allografts in only 15% of hospitals. “Further, a survey of 236 hospitals revealed that 34% of orthopedic surgeons utilizing allografts are unaware of the processing methods used for the graft.” It is vital for orthopaedic surgeons utilizing allograft tissue to be involved in the tissue selection process and understand how the tissue is processed.
This chapter will focus on the chemical sterilization techniques various allograft suppliers utilize to eliminate the possibility of disease transmission.
Allograft-Associated Disease Transmission
“A survey performed by the AATB reported that, during the period of 2003 to 2004, there were 192 reports of possible allograft-related infections to tissue banks and 1.35 million grafts were distributed during the same period. This is an overall reporting incidence of 0.014%.” Since surgeons have little incentive to self-report postoperative infections, the actual incidence may be underreported. However rare disease transmission associated with allografts may be, the impact is quite significant when it does occur.
Allograft-Associated Disease Transmission
“A survey performed by the AATB reported that, during the period of 2003 to 2004, there were 192 reports of possible allograft-related infections to tissue banks and 1.35 million grafts were distributed during the same period. This is an overall reporting incidence of 0.014%.” Since surgeons have little incentive to self-report postoperative infections, the actual incidence may be underreported. However rare disease transmission associated with allografts may be, the impact is quite significant when it does occur.
Aseptic Versus Sterile Allografts
It is imperative for surgeons using allografts to be knowledgeable about tissue processing methods. While the majority of tissue banks accredited by the American Association of Tissue Banks procure tissue, screen donors, and perform cultures to detect diseases using similar methods, subsequent processing of the tissue after procurement can be vastly different. It is critical to draw a clear distinction between aseptically processed and sterilized allografts.
Aseptic Tissue Processing
Aseptic tissue processing minimizes the introduction of new microorganisms onto cadaveric tissue during tissue procurement by means of a controlled environment. Aseptically processed tissue does not address the microorganisms that originated from the cadaver.
In order to reduce the chance of disease transmission, some aseptic tissue processors use antimicrobial solutions. “These solutions may consist of antiseptics or antibiotics, or both. These solutions, however, may not penetrate the tissue and thus may only be effective against surface contaminants. Infectious agents deep within the tissue matrices (for example, from contamination after postmortem tissue invasion by bowel flora) may remain.”
In a 2005 study of 331 patients at a surgical center in California, 11 patients (3.3%) met the case definition of an infection. The infection rate for patients who received aseptically processed allografts was 4.4% (11 of 250 patients), compared with 0% (0 of 81) for patients who received autografts or sterile allografts. The use of sterile allograft tissue appears to be associated with a significant reduction in the risk of postoperative infection.
Sterilization
In order to eliminate the chance of disease transmission, some tissue processors choose to sterilize their tissue. “Sterilization has been defined as a validated process used to render a product free from viable microorganisms.”
“According to the Association for the Advancement of Medical Instrumentation, a standards-setting organization for the medical industry, sterility assurance level (SAL) is defined as the probability of an item being non-sterile after it has been exposed to a validated sterilization process.” When a product is sterilized to achieve an SAL of 10 –6 , it means that there is less than a 1 in 1,000,000 possibility of a contaminating organism surviving the treatment.
Sterilization can be achieved in a variety of ways: heat and pressure, irradiation, the use of chemical sterilants, or a combination thereof. Sterilization of musculoskeletal tissue must be performed with care, as biomechanical properties of the tissue should not be adversely affected posttreatment.
Current Allografts
Each allograft tissue supplier uses different proprietary methods for aseptic processing or sterilization of allografts. The majority of ACL allografts are supplied by Musculoskeletal Transplant Foundation (MTF), RTI Surgical, LifeNet Health, and AlloSource. Despite the body of literature that exists comparing allografts to autografts, there are no studies to date that compare different allograft tissue processing techniques and their effects on clinical outcomes.
Musculoskeletal Transplant Foundation
MTF aseptically processes ACL allograft tissue. Once tissue is procured, culture results determine the pathway in which the tissue will be processed. Tissue is disinfected through an antibiotic soak with agitation, followed by a purified water rinse. Donor tissue with results that comply with aseptic parameters are not irradiated; all other tissue (approximately 65%) is exposed to 1.2–1.8 Mrad of gamma irradiation in a frozen state during processing or as a terminal sterilization step.
A prospective clinical study by Poehling et al. showed patients undergoing ACL reconstruction utilizing MTF. Cases involving the Achilles tendon achieved similar long-term outcomes to cases of ACL reconstruction with BPTB autografts. The study evaluated patients postoperatively at 1–2 weeks, 6 weeks, 3 months, 6 months, and then annually for 5 years. It is worth noting that of the top four ACL tissue processors, MTF is the only one that provides aseptically processed tissue.
RTI Surgical
RTI Surgical utilizes the BioCleanse tissue sterilization process to eliminate all viruses, spores, bacteria, and fungi. This process first involves sequential ultrasonic baths of detergent, hydrogen peroxide, alcohol, and water for fixed periods of time at low temperatures. Within each bath sequence, a series of pressure/vacuum cycles are used to improve matrix penetration.
The pressure variations drive the sterilants in and out of the tissue. Soft tissue grafts treated by this method achieve an SAL of 10 –6 .
The biomechanical properties of BioCleanse-processed ACL allografts have been examined by several investigators. Schimizzi et al. reported anterior tibialis allografts exposed to the BioCleanse process did not adversely affect the biomechanical properties in terms of stiffness or failure load. Jones et al. and Reid at al. found the same to be true with BioCleanse-treated BPTB allografts.
A 2-year clinical study by Indelicato et al. indicates that BioCleanse-processed BPTB allografts did not demonstrate a statistical difference in clinical outcomes when compared with aseptically processed BPTB allografts.
LifeNet Health
“LifeNet Health introduced Allowash in 1995, when it was a revolutionary process in cleaning and disinfection. Allowash solution is a combination of three detergents. These detergents include Brij 35, Nonoxynol 9, and Nonidet P40 for solubilizing lipids.” Additionally, the process uses 3% hydrogen peroxide for disinfection and 70% isopropanol for disinfection and as a drying agent. The Allowash process concludes with a controlled, low-level dose of gamma irradiation below 1.80 Mrad, which is administered at low temperatures after packaging and is able to achieve an SAL of 10 –6 .
AlloSource
“AlloSource uses the AlloTrue technology—a unique, patented process designed to penetrate deep within donor tissue to remove blood and lipids and reduce bioburden. AlloTrue utilizes a combination of treatments of antibiotics, alcohol, peroxide (cut tissue only), and multiple water rinses. The deep penetrating cleansing, with minimal tissue manipulation, results in microbial inactivation.” AlloTrue and terminal sterilization with 1.09–1.25 Mrad irradiation achieves an SAL of 10 –6 .
Chemicals Used in Anterior Cruciate Ligament Tissue Sterilization
Detergents
Detergents are used by RTI Surgical and LifeNet Health in their proprietary processing of ACL allograft. Ionic, non-ionic, and zwitterionic (a neutral molecule with a positive and negative charge) detergents solubilize cell membranes, lipids, and dissociate DNA from proteins, and they are therefore effective in removing fat and other cellular material from tissue.
Alcohol
Alcohol is used by RTI Surgical, LifeNet Health, and AlloSource in the processing of ACL allografts.
The Centers for Disease Control and Prevention Guideline for Disinfection and Sterilization in Healthcare Facilities covers the effectiveness of alcohol as a bactericidal agent. Pseudomonas aeruginosa, Serratia marcescens , Escherichia coli, Salmonella typhosa, Staphylococcus aureus , and Streptococcus pyogenes are all killed in 10 seconds by concentrations of ethanol ranging from 60% to 95% (v/v). Isopropanol was found to be more effective for killing E. coli and S. aureus .
Ethanol is also a potent virucidal agent. At 60%–80% it can inactivate all of the lipophilic or enveloped viruses like human immunodeficiency virus (HIV), hepatitis B virus, herpes, vaccinia, and influenza virus, and many hydrophilic viruses like adenovirus, enterovirus, rhinovirus, and rotaviruses. Isopropanol is not effective against hydrophilic viruses but is fully active against lipophilic viruses.
Alcohols further aid in tissue decellularization by dehydrating and lysing cells. Alcohols such as isopropanol, ethanol, and methanol are more effective than lipase in removing lipids from tissue and are capable of rendering adipose tissue lipid-free in a relatively brief period.
Hydrogen Peroxide
Hydrogen peroxide is used by RTI Surgical, LifeNet Health, and AlloSource in the processing of allografts. Hydrogen peroxide inactivates microorganisms (bacteria, yeasts, fungi, viruses, and spores) through oxidization. It can be converted into highly reactive hydroxyl radicals (OH), which attack membrane lipids, DNA, and other essential cell components to inactivate bacteria. It can be readily decomposed to water and oxygen spontaneously or by enzymes known as peroxidases possessed by living beings.
Supercritical Carbon Dioxide
NovaSterilis has developed a technique of sterilization using supercritical carbon dioxide at low temperatures and relatively low pressures. This results in transient acidification, which is lethal to viruses and bacteria.
In a study by Baldini et al., mechanical testing of anterior or posterior tibialis tendons treated with supercritical carbon dioxide (CO 2 ) yielded no significant difference in failure load when compared with unprocessed tendons. “The supercritical CO 2 sterilization method resulted in significantly lower stiffness than unprocessed allografts. However, the stiffness and strength of all groups tested were greater than that of published values of the native intact ACL.” As this is a relatively new process, further research will be required to ensure success in vivo.
Other Chemical Sterilants
Several other chemical sterilants such as ethylene oxide and dimethyl sulfoxide have been used for tissue sterilization in the past but are no longer commonly used today.
Related posts:
Risk of Anterior Cruciate Ligament Injury as a Function of Type of Playing Surface
Hamstring Harvest Technique for Anterior Cruciate Ligament Reconstruction
Intraoperative Fluoroscopy for Anterior Cruciate Ligament Tunnel Placement
Arthrex TightRope Fixation of a Soft Tissue Graft
Sonographically Guided Anterior Cruciate Ligament Injection: Technique and Potential Use for the Treatment of Partial Anterior Cruciate Ligament Tear
Factors Associated with Increased Allograft Failure Rate in Anterior Cruciate Ligament Reconstruction
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