Braxton H. F. Chan
Age 17 | Cranbrook, Britich Columbia
INSPOSCIENCE Honor of Distinguised Bioinformatics 2020 | INSPOSCIENCE Research and Innovation Honorable Mention Award 2020 | Youth Innovation Showcase 2020, Finalist, BC, Canada | Canada-Wide Science Fair 2020 Finalist | Innovate BC Young Innovator Scholarship | Sanofi Biogenius Award | Genome British Columbia Award | Best of Fair, East Kootenay Regional Science Fair
BACKGROUND
Osteochondral defects (OCD) refers to a focal area of damage over the surface of an articular joint resulting in the loss of cartilage and bone. This is the result of acute trauma or an underlying bone disorder. This creates a painful joint. Multiple surgical techniques have been attempted to deal with the pain. This includes microfracturing techniques, osteoarticular transfer systems (OATS procedure) and unloading osteotomies. Unfortunately the results have not been shown to be effective. This ultimately leads to a total joint replacement and possibly amputation. OCD tends to occur in younger patients and results in a lifetime of pain and dysfunction. Fibrocartilage is a dynamic tissue found at the insertions of ligaments and tendons. It contains cartilage ground substance and chondrocytes. Fibrocartilage is unique as it has a high propensity to reattach to its insertion site if it is surgically repaired.
PURPOSE
The purpose of this project is to help patients with an Osteochondral defect. An OCD causes patients a lifetime of pain and discomfort. Patients are never able to fully recover from this disorder. By taking advantage of its physical and biological properties, fibrocartilage may be an ideal autologous tissue to transplant into osteochondral defects. Biological ingrowth and the re-establishment of articular cartilage function would need to be examined to determine if fibrocartilage could be a viable option to treat OCD.
HYPOTHESIS
By transplanting autologous fibrocartilage tissue into osteochondral defects, it is hypothesized that histological analysis and materials testing can determine whether fibrocartilage can be a clinically applicable surgical procedure to treat OCD.
PROCEDURE
Donor Recruitment
Consent was acquired from 11 patients undergoing total joint replacements (hips or knees) to donate their old natural joints for this research project. Osteochondral defects usually occur in hip and knee joints. This is why it was ideal to use the discarded joints from total hip and knee surgeries.
Protocol for creating OCD lesions
An area of intact articular cartilage surface was first identified. A curette was used to create an osteochondral defect by removing all the cartilage and underlying superficial bone layer. Throughout the procedure, the joint was kept moist by irrigation with an isotonic phosphate-buffered saline (PBS) solution. This solution was used to maintain tissue viability.
Harvesting of fibrocartilage tissue
During the harvesting of the hip or knee joints at the time of surgery, the labrum or anterior cruciate ligament (ACL) was also excised. Both these are sources of fibrocartilage. This tissue was kept moist by irrigation with an isotonic PBS solution.
Transplantation of fibrocartilage
The excised labrum or anterior cruciate ligament was shaped using a surgical blade to match the size of the osteochondral defect. This tissue was then positioned into the osteochondral defect and secured onto the underlying bone using a biodegradable 2.0mm suture anchor with ultrabraid sutures (Smith and Nephew) (see Figure 1).
Storage of the transplanted joints
All joints were stored at a temperature of 4oC to 6oC in the biopreservative X-VIVO-10 (Lonza) as recommended by Lonza. This is a commercially available product used for tissue preservation in clinical tissue banks.
Gross pathology
The joints were examined by gross inspection after 7 weeks in their biopreservative. The samples were examined after 7 weeks because this is the duration when fibrocartilage tends to reintegrate into their bone anchor after being detached.
Materials testing
Beslands HF-500N Digital Force Gauge Push and Pull Dynamometer was used to perform compression load testing. The joint samples were load tested in a hydrated environment (with PBS) at room temperature. The fibrocartilage transplant was oriented perpendicular to the force gauge indenter (Figure 2). This allows for an axial compression measurement while limiting shear loading.
Force measurements were performed on the fibrocartilage transplant and on the adjacent intact articular cartilage. Repeat measurements were performed allowing the tissue to relax in between measurements.
Sectioning and staining protocols
After 7 weeks, the joints were then preserved in 10% buffered formalin solution. When ready for histological examination, the samples were decalcified, dehydrated and embedded in paraffin. The specimens were cut longitudinally and 5 micron sections mounted for staining with hematoxylin and eosin (H&E)[1].
Quantifying historical sections
The histologic sections were examined with a clinical pathologist. Four qualitative observations were analyzed:
1) osteointegration
2) osteoconduction
3) cellular viability
4) fibrovascular formation
Statistical tests
Peak compression force measurements for the fibrocartilage trans- plant was compared to the adjacent intact articular cartilage. A two-way ANOVA with post hoc Tukey HSD Test was used to analyze for differences[2].
Figure 1A
Figure 1B
Figure 2
Figure 3A
Figure 3B
Figure 3C: Poorly adhered transplant
Figure 4
RESULTS/OBSERVATIONS
Gross pathology
Gross pathological examination shows the fibrocartilage transplant situated in the osteochondral defect (see Figure 3). The fibrocartilage is secured and adhered to the underlying bone. If the osteochondral defect had any remaining articular cartilage or subchondral bone, this created a barrier preventing the fibrocartilage transplant from adhering (see Figure 3).
Histology
H&E staining shows the viability of chondrocytes within the fibrocartilage transplant (Figure 4), osteoapposition of the fibrocartilage transplant to the trabecular bone (Figure 5) and fibrovascular formation (Figure 5).
Materials testing
The peak compression force for the fibrocartilage was shown to be similar to the adjacent intact articular cartilage (two-way ANOVA, SS=549.19, df=31, MS=15.36, F=0.32, P=0.81, HSD=9.18) (see Figure 6). This demonstrates how the fibrocartilage transplant is able to absorb the same stress experienced during functional weight-bearing.
DISCUSSION
Osteochondral defects in articular joints can be the result of repetitive micro trauma, genetic factors, vascular disruption of subchon- dral bone and endocrine abnormalities[3,4,5]. Osteochondral defects in hip and knee joints are similar. They both are caused from sim- ilar traumas and causes the patient similar amounts of disability.
The overall age- and gender-adjusted annual incidence of knee OCD lesions has been reported as 6.09 per 100,000 person-years[6].
OCD tends to occur in younger patients with the highest in- cidence between the ages of 11 and 15 years[6,7]. This can lead to a life-time of joint pain and dysfunction.
Several treatment options for OCD have been tried. This includes osteoarticular transfer system (OATS procedure), osteochondral picking, and unloading osteotomies. These options have not shown good long-term benefits and eventually leads to the requirement of an artificial joint replacement.
This study shows fibrocartilage can be a viable alternative to treat osteochondral defects. Throughout this experiment, the samples were biopreserved to simulate the clinical transplantation of the fibrocartilage autograft. Fibrocartilage was chosen as an ideal tissue source for several reasons. Fibrocartilage is strong since it functionally takes the load of the normal joint. It already has the capacity to reattach to bone after a surgical repair. Fibrocartilage can also be harvested from multiple sites in the body (for example, the calcaneal tendon) without any detrimental or functional deficits.
This study provides gross pathological evidence of adherence of the fibrocartilage transplant onto the OCD. Histological evidence shows viability of the transplant tissue and its capacity to osteoappose with the trabecular bone of the OCD. Material testing supports the ability of the transplant to absorb the stresses required for weight-bearing.
Further studies will now need to be performed to reproduce these results in the long-term. Phase two of this project will examine the fibrocartilage transplants after one year in vitro. In vivo fibrocartilage transplantation will then follow and, eventually, human clinical trials will be undertaken.
The surgical procedure to transplant the fibrocartilage can be performed arthroscopically. This means the surgery is minimally invasive and can be performed as an outpatient procedure. The fibrocartilage transplant can be readily harvested with ease from the ipsilateral affected limb. It would not compromise the patient’s function or require allograft precautions. Importantly, tissue rejection as seen with allograft transplantation would not occur with autograft transplantation.
By providing a biological substitute for the articular cartilage loss in osteochondral defects, the fibrocartilage transplant can postpone the need for multiple artificial joint replacements. This is particularly important as most patients with OCD are young. Joint replacements in younger patients often require multiple revisions with less predictable outcomes.
Figure 5
Figure 6
CONCLUSION
This study confirms my hypothesis and establishes a novel treatment option for osteochondral defects (OCD). Histological and biomechanical evidence was shown to support the use of autologous fibrocartilage transplants as a viable solution to OCD.
FUTURE STUDIES
Further studies will focus on applying growth factors, mes- enchymal stem cells and platelet-rich plasma (PRP) to augment the healing of the fibrocartilage transplant.
In vivo studies will need to be undertaken to determine the efficacy of the fibrocartilage transplants in clinical studies.
BIBLIOGRAPHY
Fischer, Jacobson, Rose & Zeller (2008) Hematoxylin and Eosin Staining of Tissue and Cell Sections, Cold Spring Harbor Protocols. Retrieved from HYPERLINK “http://cshprotocols.cshlp.org/content/2008/5/pdb.prot4986”http://cshprotocols.cshlp.org/content/2008/5/pdb.prot4986
Vasavada N (2016) One-way ANOVA with post-hoc Tukey HSD Test. Retrieved from HYPERLINK “http://astatsa.com/KruskalWallisTest/”http:// astatsa.com/KruskalWallisTest/
Grimm NL, Weiss JM, Kessler JI, Aoki SK. (2014) Osteochondritis dissecans of the knee: pathoanatomy, epidemiology, and diagnosis. Clin Sports Med, 33(2):181–8.
Nepple JJ, Milewski MD, Shea KG. (2016) Research in Osteochondritis Dissecans of the Knee: 2016 Update. J Knee Surg, 29(7):533–8.
Gornitzky AL, Mistovich RJ, Atuahuene B, Storey EP, Ganley TJ. (2016) Osteochondritis
Dissecans Lesions in Family Members: Does a Positive Family History Impact Phenotypic Potency? Clin Orthop Relat Res. 475(6): 1751
Pareek A, Sanders TL, Wu IT, Larson DR, Saris DBF & Krych AJ (2017) Incidence of Symptomatic Osteochondritis Dissecans Lesions of the Knee: A Population-Based Study in Olmsted County: Osteoarthritis Cartilage, 25(10): 1663-1671
ACKNOWLEDGEMENTS
RBC Future Launch
Smith and Nephew
Canada Stryker Canada
Dr. Catalin Taraboanta
ABOUT THE AUTHOR
Braxton H. F. Chan
Braxton is attending Grade 12 at Mount Baker Secondary School in Cranbrook, BC. He has a passion for Sciences, the Arts and sports. Braxton plays for the pre-collegiate Whitecaps Football Academy. He is also an accomplished surfer and kiteboarder. He is known for his intricate Origami creations.