Abstract

Articular cartilage defects are not common in the glenohumeral joint and are mostly found in patients after shoulder trauma, in patients with recurrent instability, or in patients who underwent previous surgical treatment. Articular cartilage defects lead to pain and loss of motion, consequently causing shoulder function impairment and reducing quality of life. In young patients, the use of osteochondral allografts for the treatment of humeral head defects may avoid well-known complications of shoulder arthroplasty. The goal of this Technical Note is to describe a step-by-step protocol for the harvesting, transport, and preservation of fresh humeral head osteochondral tissue for use in allograft transplantation.

Articular cartilage defects are relatively common in the glenohumeral joint, with rates of around 5% to 17% according to some authors, and are mostly found in patients after shoulder trauma, in patients with recurrent instability, or in patients who underwent previous surgical treatment.1Bhatia S. Hsu A. Lin E.C. et al.Surgical treatment options for the young and active middle-aged patient with glenohumeral arthritis., 2Capito N.M. Owens B.D. Sherman S.L. Smith M.J. Osteochondral allografts in shoulder surgical procedures., 3Riff A.J. Yanke A.B. Shin J.J. Romeo A.A. Cole B.J. Midterm results of osteochondral allograft transplantation to the humeral head. Hence, diagnosis can be difficult, and defects are frequently found during arthroscopic or open surgical treatment of other diseases.4Sedil A.J. Kraeutler M.J. Management of articular cartilage defects in the glenohumeral joint.Focal cartilage defects lead to pain and loss of motion, consequently causing shoulder function impairment and reducing quality of life. Although arthroplasty and humeral head resurfacing techniques are associated with improvement in pain and function, these techniques are more suitable for older and low-demand patients. On the other hand, in younger patients, autologous chondrocyte implantation, osteochondral autograft, and osteochondral allograft are described as treatment options owing to the poorer outcomes of arthroplasty described in this population.2Capito N.M. Owens B.D. Sherman S.L. Smith M.J. Osteochondral allografts in shoulder surgical procedures.,3Riff A.J. Yanke A.B. Shin J.J. Romeo A.A. Cole B.J. Midterm results of osteochondral allograft transplantation to the humeral head.,5Provencher M.T. Sanchez G. Schantz K. et al.Anatomic humeral head reconstruction with fresh osteochondral talus allograft for recurrent glenohumeral instability with reverse Hill-Sachs lesion. Besides, humeral head reconstruction with osteochondral allografts can improve shoulder range of movement, leading to better functional outcome scores and resulting in lower subsequent dislocation rates.6Wang K.C. Waterman B.R. Cotter E.J. Frank R.M. Cole B.J. Fresh osteochondral transplantation for focal chondral defect of the humerus associated with anchor arthropathy and failed SLAP repair.,7Saltzman B.M. Riboh J.C. Cole B.J. Yanke A.B. Humeral head reconstruction with osteochondral allograft transplantation.Most literature regarding allografts involves the use of articular cartilage specimens from the knee joint. However, this type of allograft does not seem suitable for reconstruction of the articular surface of the humeral head because of differences in both cartilage thickness and curvature.4Sedil A.J. Kraeutler M.J. Management of articular cartilage defects in the glenohumeral joint.,5Provencher M.T. Sanchez G. Schantz K. et al.Anatomic humeral head reconstruction with fresh osteochondral talus allograft for recurrent glenohumeral instability with reverse Hill-Sachs lesion. Moreover, there is no consensus in the literature regarding the standardization of a protocol for the harvest, transport, and preservation of humeral osteochondral allografts in tissue banks.8Beer A.J. Tauro T.M. Redondo M.L. Christian D.R. Cole B.J. Frank R.M. Use of allografts in orthopaedic surgery: Safety, procurement, storage, and outcomes. So, the goal of this Technical Note is to describe a step-by-step protocol for the harvesting, transport, and preservation of fresh humeral head osteochondral tissue for use in allograft transplantation (Video 1).Surgical Technique Tissue Harvesting and TransportAfter positive notification by the state’s transplant central registry of a viable corpse donor (Table 1), a team (5 doctors and 3 nurses) is dispatched to the hospital where the donor’s death occurred. On arrival, the team prepares the body for the removal of all bone tissue (Table 2), in the supine position, and collects blood for serologic testing (between 72 hours before and 6 hours after circulation arrest at room temperature; if the corpse is refrigerated, between 2°C and 8°C, up to 24 hours after circulation arrest). Detergent chlorhexidine is used to perform asepsis, and alcoholic chlorhexidine is used for antisepsis. The surgical drapes are then placed.

Table 1Exclusion Criteria for Organ Donation

NOTE. The exclusion criteria are divided into 5 groups according to national regulations. If the corpse donor does not fulfill any of the criteria, serologic testing is performed and then harvest begins.

BCG, bacillus Calmette-Guérin vaccine; HIV, human immunodeficiency virus.

Table 2Routine Harvesting Protocol According to Donor Age

NOTE. The harvesting protocol depends on the donor age. Bone is collected in patients aged 10 to 70 years. In patients aged between 18 and 55 years, bone and tendons are collected. Moreover, in patients aged between 15 and 45 years, bone, tendons, and osteochondral tissue are collected. Other tissues can be harvested depending on requests made by the orthopaedic division to the tissue bank.

The surgical technique for harvesting osteochondral tissue from the proximal humerus consists of performing the deltopectoral approach, using the coracoid process and the lateral surface of the humerus as anatomic references (Fig 1A ). After skin incision, the deltoid and pectoralis major muscles are shifted laterally and medially, respectively. To improve visualization, a pectoralis major tenotomy is performed, and the deltoid muscle is detached from the lateral and posterior surfaces of the humerus with the aid of a surgical remover; 24-cm Hohmann surgical retractors may also be used to facilitate exposure.

Fig 1Surgical technique for harvesting osteochondral tissue from proximal humerus. (A) For harvesting, patient is placed supine, and the deltopectoral approach is performed (in the example, in the left shoulder), using the coracoid process and the lateral surface of the humerus as anatomic references (dashed line). (B) After completion of the osteotomy with the aid of a clamp, proximal humerus piece elevation and opening of the whole posterior and inferior capsule are performed, with removal of remaining muscle until complete release of the piece.

Tenotomy of the subscapularis is performed at its insertion on the humeral lesser tuberosity. It is extremely important to avoid damaging the articular cartilage in this step. External rotation of the upper limb helps decrease this risk and increases surgical exposure. In this step, the axillary vessels and nerve, which pass inferior to the subscapularis muscle, are sectioned. The teres major tendon is also detached from the crest of the humeral lesser tuberosity. The rotator interval is opened, with tenotomy of the long head of the biceps, in the region of the supraglenoid tubercle. To facilitate exposure, tenotomy of the supraspinatus, infraspinatus, and teres minor tendons is performed under internal rotation of the upper limb, with extreme care taken not to damage the humeral articular cartilage. At the posterior humeral surface, the origin of the triceps muscle lateral head is sectioned.

A reciprocating saw and a bone clamp are used in the resection of the proximal humeral extremity, which is performed 10 to 15 cm below the humeral greater tuberosity (Fig 1B). After osteotomy with the aid of the clamp, proximal humeral piece elevation and opening of the whole posterior and inferior capsule are performed, with removal of remaining muscle until complete release of the piece. Two tissue samples are taken at this time and stored in a container at a controlled temperature (4°C) for microbiological analysis (fungi and aerobic bacteria) in the tissue bank.The humeral proximal extremity is placed in 0.9% saline solution and stored in triple sterile plastic packing, which is appropriately sealed and contains the corpse donor identification number and initials, tissue identification with laterality, and harvesting date. Packages are stored in a container at a controlled temperature (4°C) and sent for processing at the tissue bank (Fig 2). The container inner temperature must be registered during tissue placement and removal for storage in the tissue bank refrigerator at 4°C. After removal of the humeral proximal extremity, the corpse donor body is recomposed, resembling natural anatomy, using polyvinyl chloride pipes or broomsticks, which are fixed to the glenoid with crossed Kirschner wires and covered with compresses before wound closure.

Fig 2Graft accommodation for transport to tissue bank. (A, B) Packages containing the tissue are stored in a container at a controlled temperature (4°C) and sent for processing at the tissue bank.

 Tissue Reception and StorageOn arrival in the tissue bank facility, corpse donor data (name, harvesting date, and amount and type of tissue harvested) are registered. All legal documentation must be checked at this moment. When tissue is removed from the transport container, the temperature and weight are measured and visual analysis of the packages is performed; these data are registered on the identification tag, which is sealed with the package. The tissue is then stored in the “tissue-reception refrigerator” at 4°C until processing (Fig 3). The samples collected during harvest are sent for microbiological analysis.

Fig 3Tissue storage for processing. On arrival in the tissue bank facility, corpse donor data (name, harvesting date, and amount and type of tissue harvested) are registered and checked. In the meantime, tissue is stored in the tissue-reception refrigerator at 4°C until processing.

In the next step, the proximal humeral extremity undergoes radiologic screening. The tissue package is transported to the radiology unit in the same container at a controlled temperature. After radiographs are obtained, the tissue is transported back to the tissue bank and is stored in the tissue-reception refrigerator. The radiographic report is evaluated by the physician responsible for the tissue bank, who defines whether the tissue is adequate to continue processing.

 Osteochondral Tissue ProcessingTissue processing is performed in an ISO class 5 room (Fig 4) located within the tissue bank by a member of the staff as soon as the team arrives in the facility. Processing is performed in a sterile room with negative pressure, organized using 3 tables. On the first table, 2 sterile flasks are prepared for before and after lavage (Fig 5A ). All surgical instruments are positioned on the second table, such as clamps, scalpels, sterile drapes, syringes, oscillatory saws, batteries, and a lathe (Fig 5B). Finally, the third table is prepared with the sealer, scale, culture medium, and packages (Fig 5C).

Fig 4Tissue bank facility. (A-C) Tissue processing is performed in an ISO class 5 room, in a sterile manner and with negative pressure, located within the tissue bank.

Fig 5Tissue processing organization. The room for tissue processing is organized using 3 tables to facilitate the procedure. (A) On the first table, 2 sterile flasks are prepared for before lavage (black arrow) and after lavage (white arrow). (B) All surgical instruments are positioned on the second table, such as clamps, scalpels (black arrow), sterile drapes, syringes, oscillatory saws, batteries (black circle), and a lathe (white arrow). (C) The third table is prepared with the sealer (white arrow), scale (black arrow), culture medium, and packages (black circle).

The next step consists of cleaning plus removal of muscle and bone tissue not necessary for the transplant procedure and visual evaluation, with exclusion in the case of any articular cartilage injury. The first step of processing begins with the opening of 2 of the 3 sterile packages containing the tissue on the first table (Fig 6A ). The physician responsible for processing catches the inner package, which is the most sterile, with a surgical clamp. The package is opened in a sterile (pre-lavage) flask. In this step, samples of the solution in which the tissue was immersed since harvesting are collected for microbiological analysis (fungi and anaerobic and aerobic bacteria) in culture flasks.

Fig 6Tissue processing step-by-step: part 1. (A) Processing begins with the opening of 2 of the 3 sterile packages containing the tissue. (B, C) The tissue is fixed in a conventional lathe; then, skeletonization begins with removal of all remaining muscular and tendinous insertions. (D) The proximal third of the humerus is left with only cartilage and bone. (E) A tissue fragment is collected for histopathologic evaluation.

On the second table, tissue is fixed in a conventional lathe. Skeletonization begins with the removal of all remaining muscular and tendinous insertions (Fig 6 B and C), using scalpels and mechanical saws, leaving the proximal third of the humerus with only cartilage and bone (Fig 6D). During this step, it is important to make a few pauses to irrigation of the tissue with the pre-lavage solution to minimize damage to the humeral cartilage due to the heat caused by the mechanical saw. At the end of this step, a tissue fragment is collected for histopathologic evaluation (Fig 6E).In the sequence, the tissue returns to the first table for continuous lavage with 0.9% saline solution in a second (post-lavage) flask. Three samples of this solution are collected for microbiological analysis (fungi and anaerobic and aerobic bacteria) in culture flasks (Fig 7A ). Then, the tissue is dried with sterile compresses and transported to the third table, on which osteochondral tissue is measured with a graduated scale (Fig 7B). Tissue weighing is performed on a digital scale. After measurements, osteochondral tissue is stored in a sterile plastic package (internal package) containing the preservation medium, composed of Iscove’s cell culture medium without serum (Thermo Fisher Scientific, Waltham, MA) supplemented with 10% human albumin and vancomycin, 100 μg/mL, to reduce the risk of chondrocyte death (Fig 7C). Double and triple sterile package sealing is then performed. Double package sealing, in this step, is not easy because of the tissue size and owing to the preservation medium, which sometimes leaks, making adequate sealing more difficult.

Fig 7Tissue processing step-by-step: part 2. (A) The tissue undergoes continuous lavage with 0.9% saline solution in a second (post-lavage) flask, and samples of this solution are collected for microbiological analysis in culture flasks. (B) Osteochondral tissue is measured with a graduated scale (dashed line). (C) After measurements, osteochondral tissue is stored in a sterile plastic package containing the preservation medium (oval).

Processed tissue is stored in the “quarantine refrigerator” until the results of all the microbiological and histopathologic tests are known (Fig 8). In the case of positive culture findings, the tissue is discarded. If findings are negative, tissue is ready for transplant and the patient’s surgery can be scheduled. All data are recorded in the corpse donor files and in the host patient files. Table 3 presents pearls and pitfalls regarding humeral head graft harvesting and processing.

Fig 8Tissue quarantine after processing. Processed tissue is stored in the quarantine refrigerator until the results of all the microbiological and histopathologic tests are known.

Table 3Pearls and Pitfalls

 Osteochondral Tissue Transplantation PlanningHost osteochondral lesions are diagnosed by standard radiographs of the shoulder and computed tomography (CT). The goal of CT is to measure the size of the humeral head cartilage defect, besides identifying the absence of a posterior glenoid border fracture or bone avulsion of the infraspinatus or supraspinatus tendon. Axial CT images are obtained immediately below the coracoid process. A circle is positioned over the humeral head; a line is then drawn from the lesser tuberosity to the posterior margin of the cartilage adjacent to the infraspinatus insertion, and a second line is drawn tangent to the defect region toward the anterior line. The angle between these lines is denominated the defect angle (Fig 9). On the basis of the defect size, the size of the humeral head allograft can be determined.

Fig 9Calculation of host defect size. The size of the humeral head cartilage defect is measured on axial computed tomography images obtained immediately below the coracoid process. A circle is positioned over the humeral head (white arrow); a line (solid white line) is then drawn from the lesser tuberosity to the posterior margin of cartilage adjacent to the infraspinatus insertion, and a second line (dashed white line) is drawn tangent to the defect region toward the anterior line. The angle between these lines (A) is denominated the defect angle, which is used to calculate the size of the allograft to be transplanted.

On the surgery date, osteochondral allograft is transported in a cooled container at 4°C and removed from the package only at the moment of transplant (Fig 10A ). At the time of surgery, an osteotomy is performed in the anatomic humeral neck of the graft for isolation of the fresh humeral head osteochondral allograft (Fig 10B), which is adapted for transplantation and filling of the cartilage defect, restoring the spherical humeral head anatomy (Fig 10C).

Fig 10Preparation of osteochondral graft for transplantation. (A) The proximal third of the humerus is removed from the package only at the moment of transplant. An osteotomy is performed in the anatomic humeral neck of the graft for isolation of the fresh humeral head osteochondral allograft (dashed line) (B) and adapted for transplantation and filling of the cartilage defect, restoring the spherical humeral head anatomy (C).

DiscussionOwing to poorer long-term results of arthroplasty in young patients, alternative techniques that can be applied to patients younger than 50 years who present with major humeral head chondral lesions have been studied to restore the articular surface anatomy in a biological way using restorative techniques such as the fresh osteochondral allograft transplantation.9Harb A. Von Horn A. Gocalek K. et al.Lactated Ringer-based storage solutions are equally well suited for the storage of fresh osteochondral allografts as cell culture medium-based storage solutions. Osteochondral allografts are similar to autografts and present the advantages of a shorter surgical time, lower harvest-associated morbidity, and the possibility of treating lesions larger than 2 cm.2Capito N.M. Owens B.D. Sherman S.L. Smith M.J. Osteochondral allografts in shoulder surgical procedures.. The harvesting procedure requires a corpse donor and fitting of the articular geometry of the graft to the patient.10Torrie A.M. Kesler W.W. Elkin J. Gallo R.A. Osteochondral allografts can be fresh, when stored at 4°C, or frozen, when stored at –80°C. Frozen grafts at –80°C lead to a considerable reduction of the host immune response while preserving the biomechanical properties, although chondrocyte viability significantly decreases over time.11Ball S.T. Amiel D. Williams S.K. et al.The effects of storage on fresh human osteochondral allografts. Fresh osteochondral allografts refer to tissue harvested 24 hours after donor death and stored, almost aseptically, in a refrigerator at 4°C, until microbiological tests are performed to allow the transplant. This type of graft presents, as its main advantage, the fact that it contains hyaline cartilage with subchondral bone support, as well as viable chondrocytes at higher numbers.12Tschon M. Veronesi F. Giannini S. Fini M. Fresh osteochondral allotransplants: Outcomes, failures, and future developments., 13Stoker A.M. Stannard J.P. Kuroki K. Bozynski C.C. Pfeiffer F.M. Cook J.L. Validation of the Missouri Osteochondral Allograft Preservation System for the maintenance of osteochondral allograft quality during prolonged storage., 14Bruns J. Werner M. Habermann C. Osteochondritis dissecans: Etiology, pathology, and imaging with a special focus on the knee joint. However, fresh osteochondral allograft viability decreases over time14Bruns J. Werner M. Habermann C. Osteochondritis dissecans: Etiology, pathology, and imaging with a special focus on the knee joint.,15Cinats D. Miller S. Abusara Z. et al.Evolution of a novel tissue preservation protocol to optimize osteochondral transplantation outcomes. and with the medium used for storage.11Ball S.T. Amiel D. Williams S.K. et al.The effects of storage on fresh human osteochondral allografts.Preservation of human osteochondral allografts has previously been studied for the treatment of knee lesions.16De Sousa E.B. Aguiar D.P. Barcelos J.F. Duarte M.E.L. Olej B. Approaches to preserve human osteochondral allografts.,17Vivacqua T.A. Prinz R.D. Cavanellas N. Barreto J.M. de Sousa E.B. Aguiar D.P. Protocol for harvest, transport, and storage of human osteochondral tissue. Regarding the shoulder, few studies have evaluated the use of osteochondral allografts for treating humeral head articular cartilage lesions.7Saltzman B.M. Riboh J.C. Cole B.J. Yanke A.B. Humeral head reconstruction with osteochondral allograft transplantation. The few existing reports have mainly described the use of frozen allografts,7Saltzman B.M. Riboh J.C. Cole B.J. Yanke A.B. Humeral head reconstruction with osteochondral allograft transplantation.,18Allograft reconstruction of segmental defects of the humeral head for the treatment of chronic locked posterior dislocation of the shoulder.,19Snir N. Wolfson T.S. Hamula M.J. Gyftopoulos S. Meislin R.J. Arthroscopic anatomic humeral head reconstruction with osteochondral allograft transplantation for large Hill-Sachs lesions. whereas studies involving the use of fresh allografts are scarce.20Black L.O. Ko J.W.K. Quilici S.M. Crawford D.C. Fresh osteochondral allograft to the humeral head for treatment of an engaging reverse Hill-Sachs lesion: Technical case report and literature review.,21Proencher M.T. Leclere L.E. Ghodadra N. Solomon D.J. Postsurgical glenohumeral anchor arthropathy treated with a fresh distal tibia allograft to the glenoid and a fresh allograft to the humeral head. Those studies, however, although rare, have presented promising results regarding the use of this type of graft, showing good functional results, pain improvement, and no complications. When chondrocytes remain viable during storage, they maintain matrix integrity and, so, the graft properties.8Beer A.J. Tauro T.M. Redondo M.L. Christian D.R. Cole B.J. Frank R.M. Use of allografts in orthopaedic surgery: Safety, procurement, storage, and outcomes. Besides, long-term allograft transplant survival depends on graft chondrocyte viability, on matrix maintenance, and on the graft incorporation to host bone.8Beer A.J. Tauro T.M. Redondo M.L. Christian D.R. Cole B.J. Frank R.M. Use of allografts in orthopaedic surgery: Safety, procurement, storage, and outcomes.,22Krych A.J. Saris D.B.F. Stuart M.J. Hacken B. Cartilage injury in the knee: Assessment and treatment options., 23De Caro F. Bisicchia S. Amendola A. Ding L. Large fresh osteochondral allografts of the knee: A systematic clinical and basic science review of the literature., 24Schreiner A.J. Stocker A.M. Bozynski C.C. Kuroki K. Stannard J.P. Cook J.L. Clinical application of the basic science of articular cartilage pathology and treatment. Table 4 presents the advantages, disadvantages, and risks of the described technique. On the basis of this report, as described in this protocol, after adequate harvesting, transport, and storage, fresh humeral head osteochondral allograft is an adequate option for the treatment of patients with massive proximal humeral chondral lesions.

Table 4Advantages, Disadvantages, and Risks

Acknowledgment

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