INTRODUCTION

Although the first keratoplasties used fresh corneas from living donors with blind eyes due to a non-corneal cause, the idea of using post-mortem corneas had emerged since the beginning of the 20th century. This was not applied regularly until the following decades, when Vladimir Filatov (Ukraine, 1930) performed almost a thousand corneal homografts of cadaverous origin^1,2. From then on, the need to establish ethical, legislative and technical criteria, in order to regularize the use of donated eyes is essential. These will be centralized in one entity, the Eye Bank (EB), established for the collection, conservation and distribution of ocular tissues for transplants.

In the first story of preservation of a human cornea, for 8 days before being used in a lamellar graft, hemolyzed blood was used as preservation medium and the tissue was maintained at 5 °C³. The first modern EB, was founded by Richard Paton, in 1944 in New York. The preservation techniques were very simple, using the "wet chamber" - small glass jars, where the enucleated eyeballs were kept in a humid environment at low temperature, until they were used (Figure 1). The enucleation was performed in the shortest possible period after death. This variable, and the macroscopic aspect of the cornea, were the only quality controls that were used¹. The importance of an adequate evaluation of the corneal endothelium was only introduced years later^4,5.

In Spain, more than 60,000 patients have benefited from a keratoplasty, since the first corneal transplant was performed in 1940, by Ignacio Barraquer⁶. Aguilar Muñoz, in 1952, created the first EB in the Provincial Hospital of Madrid, which worked for a short period of time⁷. In 1962, Ignacio and Joaquín Barraquer founded the Eye Bank for the Treatment of Blindness in Barcelona, the first in continental Europe that continues to function (Figure 2).

ORGANIZATION OF THE EYE BANK

The main objective of the EB, is to guarantee the availability of the tissue with the highest levels of quality and safety. This includes respect for the legal and ethical principles intrinsic to a product of human origin donated altruistically, as well as the ability to ensure the principle of equitable distribution, when demand exceeds availability.

The legal framework: who authorizes

The first reference to tissues in Spanish legislation dates from 1950^8,9. However, it was not until 1996¹⁰ that a specific regulation was proposed. The European Directive 2004/23¹¹ and the Technical Directives that develop it^12-14, were transposed in our legal system in the RD 1301/2006¹⁵ that established the quality and safety standards for the donation of human cells and tissues and approved the norms of coordination and operation for use in humans. This was recently replaced by RD-law 9/2014¹⁶, without substantial changes for the organization, structure and functioning of EBs. The authority to authorize an EB in Spain is transferred to the Autonomous Communities.

When the activity involves the substantial manipulation of cells, its use in a place other than its origin, or with a different function, such as the ex vivo expansion of sclero-corneal limbus cells for the treatment of limbal deficiency, it is the Spanish Agency for Medicines and Health Products (AEMPS) who must authorize it.

What is authorized

For each type of tissue or cell group, specific activities and processes are authorized. For this, a description of them is required in the application, for the process, storage and distribution of cornea and sclera, or the cell cultures of the sclero-corneal limbus. In addition to tissues of ocular origin, authorization may include other tissues such as amniotic membrane, arteries, valves, skin, skeletal muscle, etc.

For the authorization of processes, it must be described what the tissues are going to be subjected to, in order to preserve or enhance their properties or cells, and to minimize the risks of microbiological contamination or disease transmission. The most frequently authorized processes within an EB do not involve substantial manipulation of the tissues:

a) Cornea, which can be: fresh (refrigerated at 4-8 °C), preserved in organotypic culture medium (at 35-37 °C), cryopreserved (at -196 °C), or pre-cut for anterior lamellar keratoplasty.

b) Sclera, either by preservation at room temperature or lyophilized.

How is it authorized

The EB must submit a request for authorization, accompanied by a report that complies with the requirements detailed in RD-Law 9/2014. In addition to describing the activities and processes, it must detail the organization chart, responsibilities, facilities, quality system, logistics of distribution, traceability and bio monitoring. Once the report has been reviewed, the competent authority may request clarifications and carry out a pre-authorization inspection. This will have a validity of no less than two years, nor more than four years. The renewal is granted only after confirming that the conditions that gave rise to its concession continue to be met. Any substantial modification of the same (for example, changes in the processing and preservation, transfer of the facilities, change of the technical responsible) requires a notice be submitted to said authority.

STRUCTURE OF THE EYE BANK

The structure of the EB is based on seven pillars, as detailed below (Figure 3):

Quality system

The EB must implement and maintain a quality management system, updated to ensure that the tissues and cells meet the legal and technical specifications. These will depend on the activities that are carried out (for example: Guide to Standards of Correct Manufacturing¹⁷, RD-law 9/2014¹⁶, Norm ISO 9001¹⁸), as well as the specifications of the tissues and cells obtained. Quality is the responsibility of all the personnel involved, that should contribute to its development and maintenance, and should be applied at all stages, from the selection of the donor to the transplant.

To build an adequate quality system, there are technical guides, such as the Good Tissue Practices for European Tissue Banks¹⁹ or the Guide to the Quality and Safety of Tissues and Cells for Human Application²⁰, or of a legal nature, such as Directives 2004/23¹¹, 2006/17¹², 2006/86¹³, or the Good Manufacturing Standards Guide¹⁷, which can be very helpful. This involves writing standard operating procedures (SOPs) for all the critical processes of the EB, which must have at least the following: a) training and reference manuals; b) forms for transmitting the information; c) necessary information on the origin and destination of tissues and cells; d) traceability on cells and tissues; e) detection and communication system of adverse effects and reactions¹⁶.

Personnel

The EB will appoint a technical manager who must have a university degree in the field of medicine or biomedical sciences and have practical experience in the area of EB activity not less than three years. Their functions include being responsible for organizing the activities and carrying them out in accordance with the provisions of the legislation. The EB may have its own medical team or have external advisors to review, supervise, analyze and if necessary, promote changes in their medical activities.

Staff training is essential to ensure that all team members are competent and capable of performing the tasks assigned to them. The technical staff-training program must be documented and registered. In order to maintain an optimum technical level, it is recommended to establish a continuous training program for all personnel. Training in prevention of occupational hazards, is also necessary and should essentially cover the risks inherent in the handling of biological material.

Facilities and equipment

The EB must have the premises and infrastructure necessary to carry out the activities for which it is authorized. Before starting the design of the EB, it is advisable to consider the following aspects:

- The design and construction of the facilities should follow the European Tissue and Cell Directives, and the principles and guidelines of the Standards of Correct Manufacturing¹⁷.

- Define what activities will be carried out, e.g. if the separation of the sclero-corneal cap from the eyeball will be done in the EB, or if the extractor personnel will do so in the donor.

- Estimate the number and type of processes to be carried out, in order to measure it appropriately.

- Define the flow of people and materials between the different processing areas.

- Design the distribution, considering that not all areas have the same technical and legal requirements, including cleanliness and air quality²⁰.

All this will depend in turn, on: a) the type of cells and tissues that are obtained; b) the manner in which those materials are preserved and stored until processed; c) the degree of exposure of the same during the processing; and d) the decontamination or sterilization methods applied.

Production, processing and quality control

The processing area must be designed, qualified and monitored to ensure that the air quality is adapted to the processes to be carried out. If tissue/cells are exposed directly to ambient air during processing, the quality of this (number of nonviable particles/mm³ and number of colony-forming units/mL) must be equivalent to that specified as "grade A", with an environment that should at least be "grade D", according to Annex I of the European Guide for Good Manufacturing Standards¹⁷.

All the critical processing processes carried out with the tissues/cells must be validated, either internally in the EB or retrospectively, based on the clinical results obtained with the tissues/cells supplied, or on published scientific data. All production processes must be described in the corresponding SOPs, as well as the procedures and approaches that may affect their quality and safety. The quality controls of both the processed tissues and the installation and critical equipment will be defined and validated depending on the type of production process.

External activities

Any activity or process that EB does not perform, requires agreements or collaboration agreements with companies, laboratories, or external medical services in which the scope and responsibilities of each party are clearly established. These may include: a) the donation process (detection, selection, donor evaluation and tissue removal); b) the environmental controls of the EB facilities; c) cleaning and maintenance; d) microbiological control of culture media; and e) the transport of tissues to transplant centers, among others.

Claims and withdrawals

The EB must have specific procedures, for the management of incidents and removal of the tissues, compromised in their quality or safety, in the event of a claim by the recipients or transplant centers. If there are other tissues from the same donor distributed, the EB should contact the centers that have transplanted them to know the status of the recipients, apply the precise treatment and follow up. If the tissues have not yet been transplanted, they will be removed and kept in quarantine until the investigation of the incident is complete.

Tracing

For an EB, it is fundamental to know if the tissues and cells transplanted have reached and satisfied the expectations of the patient and the clinician. The information on results and evolution of the transplants is very valuable, not only to know its effectiveness as a treatment, but also to optimize tissue preservation methods and identify adverse effects or reactions that may be due to deficiencies in its quality and safety.

OPERATION OF THE EYE BANK

The purpose of the EB is to prepare, preserve and store tissues and cells, both of autologous and allogeneic origin, so that they are used in the maximum number of patients. If the availability of tissues is lower than the demand, the EB must ensure the principle of equity in its distribution^16,20.

Given that tissues and cells that reach the EB come from altruistic donations, their activity should not be lucrative in any case. The competent authority will establish the regime of compensation, and charge of costs that can be applied, which can only be imputed to the transplant center and never to the patients. The transplant of any material of human origin, carries a risk of disease transmission, which must be minimized. Therefore, the selection and evaluation of the donor, serological screening, extraction, processing, preservation and storage of tissues and cells should be aimed at reducing this risk and ensuring its quality and safety (Figure 4).

Selection and procurement of donor materials

The EB establishes the selection criteria for donors, tissues and cells. The aspects of obtaining ocular tissues are discussed in another chapter of this book.

Quarantine, processing and quality controls

When receiving the tissues or cells in the EB, it must be verified that the transport container is correctly identified and maintains the specified temperature conditions. The primary tissue container must be intact, properly labelled and accompanied by the appropriate documentation. In case the materials are not going to be processed immediately, they will be stored in quarantine in a cold room (4-8 °C). The EB can receive the whole enucleated eyeball, kept in a wet chamber, or the corneoscleral cap already separated from the eyeball. In the first case we will proceed to separate said cap in aseptic conditions (after external asepsis of the globe), before submerging it in the specific corneal preservation media.

Given the importance of the corneal endothelium in the evaluation of tissue quality and viability, the EB should establish the criteria for measuring cell density and minimum acceptable values. In general, it is accepted that said value must be higher than 2,000 cells/mm², for the performance of a transplant that includes the replacement of the endothelium of the recipient. One of the recent advances is the preparation of pre-cut corneas for lamellar keratoplasty, or Descemet-endothelial membranes or rolls^21-23. These processes involve a risk of perforation or rupture of the lamina with the endothelium, which can be attributed to the difficulty of the technique, the learning curve, or the characteristics of the processed cornea. The previous preparation in the EB of the tissue for these procedures prevents such accidents from affecting the logistics of the transplant centers.

Quality controls during evaluation and processing are of two main types: a) microbiological, to minimize the risk of transmission of infectious agents, and b) viability, such as the endothelial count or the thickness of the pre-cut sheet. The acceptance criteria for such controls must be clearly defined, validated and adequately documented. Once processed, the tissue is stored in quarantine pending the results of the quality controls, both of the tissue (microbiological, serological and viability) and of the facility. The release of tissue for clinical use occurs after checking that the processed tissue obeys the criteria of quality and viability in all controls performed.

Release of tissues or cells

The release of tissues/cells for clinical use implies that the viability of the donor and the tissue, as well as the processing conditions, have been verified. This should include any information or additional results pending after the previous phases. For example, during the extraction of tissues from a donor that had been considered suitable, the extraction team observed a mass in the colon and referred it for histopathological study. If the latter concludes that it is a colon adenocarcinoma, the corneal button can be accepted for clinical use, but not the sclera, the limbal cells or the rest of the ocular tissues.

Distribution

By tissue and cell distribution, we understand the transport and delivery of these for application in a human being. This is a critical process, that must be previously validated, and that should include the packaging material, to ensure transport conditions such as temperature, or its resistance and deformability against shocks, as well as the time for which said environmental conditions will be maintained. Each EB, in agreement with its competent authority²⁴, establishes the distribution criteria in a transparent manner, giving priority to medical emergencies.

Traceability

Having an effective traceability system, as defined by RD-Law 9/2014¹⁶, allows acting quickly and efficiently in the event of any deviation in any step, from the selection of the donor to the transplant, which may affect the quality or safety of the tissues or cells provided, and thus avoid or minimize potential damage. For this, it is necessary to: ​​a) have a unique identification code; b) transfer critical information securely, preferably electronically; if done manually, check in duplicate and two people; c) update the information in real time: if a risk is identified, any delay in the transmission may increase the number of affected patients; d) clearly define the traceability responsibilities of each organization involved; e) store and safeguard the traceability information (donation, evaluation, procurement, storage, distribution and final use) for at least 30 years from its coding¹⁶. With regards to coding, Directive 565/2015²⁵ amends Directive 2006/86¹³ as far as technical requirements on cell and tissue coding are concerned; the member states will apply the legislation as of April 29, 2017.

Bio-vigilance

Any adverse effect or serious reaction that may influence the quality or safety of the tissues and cells must be noted and investigated, as this may be due to the processes of obtaining, evaluating, processing, storing and distributing them. Likewise, the corrective actions that derive from the investigation must be implemented. All the organizations involved in the donation, the EB and the transplant professionals, must establish a procedure of bio monitoring in which the responsibilities and the algorithms of notification, investigation and follow-up of these events, are clearly stated ^16,20,26,27, and can be defined as:

- Serious Adverse Reaction (SAR): the unexpected response of the donor or recipient, including a transmittable disease, associated with the acquisition or application of tissues or cells, which is fatal, life-threatening, disabling or incapacitating, or result in hospitalization or illness or prolonging them.

- Serious Adverse Effect (SAE): any unfavorable event related to the obtaining, evaluation, processing, storage and distribution of cells and tissues that may lead to the transmission of a disease, the death of the patient, or to states that endanger their life, disability or incapability or that may lead to hospitalization or illness or may prolong it.

In summary, a SAR is an incident in which the donor or recipient has been damaged, while in an SAE such a risk arises, but the damage has not yet occurred. The EB will provide the transplant professionals with information or a document model on how to report possible SAR/SAE. To do this, one should not wait until the investigation is completed or the suspicion confirmed.

CONCLUSIONS AND FUTURE PERSPECTIVES

In the EB the logistical and technical optimization, the adequate training of the personnel, together with a modern infrastructure, all within the regulatory norm, allows to generate a tissue of the best quality and to respond adequately to the needs and expectations of the recipients and the medical community.

The field of EB is in constant evolution. Among current and future advances, we can count (a) the growing tendency to use hot organotypic culture methods, which allow longer preservation times and therefore facilitates quality control and a better organization of the surgical schedule of the transplant centers, (b) the preparation of pre-cut or dissected tissue for lamellar keratoplasties (anterior or endothelial), thus relieving the surgeon of devoting his time to this aspect of the technique, and (c) the development of cellular therapies in the field of regenerative medicine, such as the ex vivo expansion of limbal stem cells or the other types that may be implemented. All these developments require the incorporation of increasingly qualified technical personnel, as well as the joint work of the EB with the transplant surgeons.