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When people are born no spare parts are readily available for them. Soft tissue regeneration occurs quite fast but what about bone recovery? It was proved that implants — artificial bone replacements — were in use as long ago as some time BC. A piece of a mandibula of an Inca (6th century BC) was found in modern Honduras, around the De Los Muertos plateau, bearing three dental implants made of sea mussel shells. A female skull belonging to the 1st century BC, with a metal implant of the upper jaw canine was discovered in the province of Chantambre, France.
Packing of Gamaplant-paste-forte used to fill in bone defects
A human organism perceives any foreign body as hostile. Currently, up to 35 per cent of implants used in traumatic surgery result in rejection. That means that patients experience significant pain and face recurring operations. Human bone tissue consists mainly of hydroxyapatite nanocrystals (65 per cent) and collagen (25 per cent). It also includes specialised cells and proteins — growth factors. So the question is whether there is a way to increase the biocompatibility of implants and to make them as similar as possible to human bone tissues. Scientists and doctors have been attempting to address this task for decades now, which involved multiple approaches. A principally new technology was offered at the Gamaleya Research Institute for Epidemiology and Microbiology, Russian Academy of Medical Sciences.
The Gamaleya Research Institute is famous for its advanced developments. It was here that Lev Zilber, a world renowned virologist and immunologist who discovered the tick-borne encephalitis virus, created his viral genetic theory of cancer origin which served as a foundation for modern cancer immunology. Alexander Fridenstein, leading immunohematology expert, corresponding member of Russian Academy of Medical Sciences, discovered stromal bone marrow stem cells while working at the Institute. His research of bone tissue recovery and transplantation helped to create Russian technologies for osteoplastic materials.
The Gamaleya Research Institute served as a lead agency in creating new-generation implants according to the project titled “The Development of Experimental-Industrial Technologies for the Generation of Hydroxyapatite/Collagen Composite Preparations/Coatings for Implant Materials” which was initiated by the Ministry of Education and Science within the Research and Development on Priority Directions for Russian Sci.... According to the contract stipulations the researchers acquired RUR140 million of funding and had three years to perform the work
The fundamental novelty of the implantation technology developed at the Gamaleya Research Institute is related to the introduction of proteins — growth factors and bone tissue regeneration factors — into the composite implant material, as well as to the creation of metal implants whose surface facilitates the retention of biocoating that is as close to bone tissue composition as possible and also contains growth factors. Due to the bone tissue growth factors the implant not only provides the foundation for the bone to grow on but also becomes an active agent initiating bone tissue formation.
Spine pedicle fixation screw with a composite preparation/coating
Until today, no medicines of such kind were manufactured in this country. Bone tissue growth factors were generated based on genetic engineering approach in the laboratory of biologically active nanostructures at the Gamaleya Research Institute led by Vladimir Lunin. In the US, medicines containing bone tissue growth factors and regeneration factors were approved for use since 2002. Up until recently, Russia lagged behind significantly in this area as morphogenetic bone proteins hold the leading positions in the list of medical innovations of primary importance.
The laboratory led by Lunin used the assistance of others. The basis of the implants — nanostructured titanium — was produced at Ufa State Aviation and Technical University. Today, pure titanium is the most promising and widely used substance for transplantation material. Theoretically, it is possible to increase its durability due to nickel alloy and other alloys but those, however, are toxic for the body. Therefore, extra durability can be achieved using nanotechnologies that allow to change the titanium structure by crumbling the metal grain to bring it to nanosize, and ensure the Damascus steel effect. The difference is that nanostructured titanium is created not via forging but via drawing and rolling. As a result, the titanium product can be made very thin while preserving its durability. It is important for dental implants, for example — teeth experience significant loads, and large screws are impossible to install in the jaw due to its thinness. Titanium is not only durable but also rather light which is extremely important when an implant is intended to stay in the body for the whole life span of the patient.
Then, the nanostructured titanium is used to make precision titanium bars with checked geometry used by Moscow-based Konmet, CJSC, to produce dental implants. The implant surface is grit-blasted to create relief.
Experts of Iskra Innovation Research Centre, Ufa, working closely with local surgeons specialising in trauma and spine illnesses developed an original model of titanium implants for traumatology and orthopedics — a device intended to correct and fixate the spine. The device consists of a set of screws and pins that can be connected together into various constructions depending on the specifics of the trauma or the illness of a particular patient; the appliance is manufactured in Pushchino-on-Oka by Deost Scientific Production Association. Belgorod State University is responsible for modifying the surface of one of the screws — the one for the pediclefixation — to impart certain special properties to it. After that, a biocoating — the key technology aspect — is applied to the implant.
“When we commenced our research we knew from the available literature that we had to address three tasks at once, — said Vladimir Lunin. — We needed to create the surface optimal for the human body. First, the implant surface is hydrophobic — that is, its wettability must be ensured. Second, it has a negative charge which is not good either as the tissue cells are charged negatively as well. Thus, double repulsion occurs between the human cells and the implant surface due to the lack of wettability and similar charge. Third, a certain relief is required — that is, the surface should not be smooth.”
Gamaplant-paste-Forte composite medicine close to human bone tissue in properties is intended for filling bone defects and coating implants
Switzerland-based Straumann, the leading global manufacturer of dental implants, demonstrates the hydrophilism of the implant surface in its promotional brochures. Before the screw is processed using a special substance it “displaces” water when emerged in it, while after the processing a hydrophilic screw pulls the water upwards similar to a sheet of paper that absorbs water when one side of it is submerged into water. Swiss developers managed to create an implant showing a 10 millimetre “ascent” of liquid. The presentation offered to the Ministry of Education and Science commission shows a spine fixation screw with the surface processed using the microarc oxidation technology, or MAO, developed by the experts at Belgorod State University. Russian researchers managed to create an implant ensuring 30 mm liquid “ascent.” That is a significant achievement quite able to meet competition with global implant manufacturers.
Modern technologies make a broad use of changes in surface properties of materials. For example, similar surface wetting effect is implemented by Toyota to make car windows. Due to the creation of a hydrophilic film water spreads upon the glass without forming drops on the surface of the window. But here the researchers face a slightly different task of creating a large smooth surface whereas the implant, unlike car windows, should be as uneven as possible in order for the cells to “engage” to it. A principle of “sending a ball into a pit” applies in this case. A cell is about 20 micrometer in size; therefore, pits on the titanium surface should have slightly larger diameter for the cell to be able to “crawl” inside and “rest” there.
The technology used to create hydrophilic and uneven surface of titanium implants is based on the surface electrolytic processing in solutions containing hydroxyapatite, one of the main bone components. The method was adapted for the implants developed by the scientists at Belgorod State University in the course of the project; it is titled microarc oxidation, or MAO. It allows the hydroxyapatite atoms to penetrate the titanium structure. A gradient is created: the implant surface consists virtually of hydroxyapatite only with titanium atoms prevailing in the deeper layers. Due to the gradient structure, the surface becomes very durable which ensures that no desquamation of the surface layer occurs during screwdriving into the bone. This technological solution ensures not only high level of hydrophily (wettiness) of the material but also addresses the charge issue as this surface bears no charge — as well as the biocompatibility problem because hydroxyapatite is a generic substance for bone tissue. The task of attaching body cells “building” the bone to the implant surface gets addressed too — as well as the objective of the union of the newly generated bone and the implant. Electrolysis results in the formation of oxygen bubbles on the surface of the implant which facilitates the creation of porous structure presenting an ideal relief for the interaction between the implant and the body cells and bone.
Human bone tissue contains 20 bone morphogenetic proteins responsible for skeleton formation. “But we are unable to completely reproduce the nature, — said Anna Karyagina, leading scientific associate of the laboratory. — It is impossible to create 20 recombinant proteins similar to those synthesised in the human body, mix them up, put them in the right areas of the bone and properly “seal” them. Scientists model the main mechanism allowing the body to launch the regeneration processes fast and delivering the pre-manufactured building material based on one recombinant protein similar to the human one to the trauma spot.”
Chromatographic columns used to rectify protein samples
“When a bone brakes, it results in redistributing the bone tissue density around the spot of the break. Building material is delivered from the surrounding bones and after some time the spot of the break can become harder than the adjacent bone areas. For example, an arm brake leads to slower nail growth. And we here bring quality building material externally to the trauma area, that can be used by the organism immediately.”
Many medicines based on recombinant interferon are produced in this country. Unlike genetically modified food products that still face negative perception on the part of the consumers these medicines had more luck as they are broadly used in medical practice. To make bone tissue growth factors as well as interferons, bacteria are used. Due to a built-in human gene, the microorganisms begin the synthesis of a protein similar to that generated in the human body. When such protein gets into a human organism it is recognised as “friendly” and gets implemented in the bone along with the body own proteins. Growth factors facilitate the differentiation of mesenchymal stem cells discovered by А. Ya. Fridenstein. They help growth, mobilisation of calcium and phosphorus from other cells, and accelerate blood vessel invasion. Thus, a layer of composite coating applied to the implant is used as a bait for the cells of the organism and as a building material to grow its own bone.
Medical and biological study of the developed medicines and implants with the calcium-phosphate coating was conducted on rats and other laboratory animals. In particular, a hole was drilled in a rat shin bone to insert a coating-covered implant. For the reference experiment, coating-free implant was used. In all experiments involving the calcium-phosphate coating, the implant invasion in the bone was much faster with the implants sitting tighter in the bone.
The laboratory animal keeping conditions should be also mentioned. The Gamaleya Research Institute has one of the best vivaria in the country which is even certified according to the European standard. Each animal has a cage of its own; there is an automatic system intended to feed the animals and to maintain all the optimal conditions. All that means that their living standards are nearly as high as those of the laboratory personnel that also receives extraordinary care. A cozy dining room has a refrigerator personally filled with various delicacies by the laboratory head.
The laboratory currently has 46 employees. Each of them is a specialist in his or her narrow field. The laboratory personnel all together is “capable of solving any problems within any time frames,” said the head of the laboratory.
Reports concerning the Ministry of Education and Science project. Reports for each project stage are provided in three copies weighing from five to eleven kilograms; reports also include documents supplied by seven co-executors involved in the project.
Researchers are the deficient aspect of Russian science today. “In the 1990s we had a redundancy of uncalled scientific personnel combined with the lack of reagents, equipment or space; today the situation we face is quite the opposite, — explained Vladimir Lunin. — Reagents and equipment are relatively cheap but we have an acute shortage of people capable of organising the process, building technological chains and creating an end-to-end technology process aimed at exploring a certain problem.”
The laboratory led by Vladimir Lunin has special people responsible only for developing and reconciling various paperwork. There are experts in economics, accounting, agreement drafting. The laboratory performed huge chunk of work in the stage of state certification of the developed medicines. The institute administration was stunned when the laboratory managed to walk the complete way from the start of medical trial to the medicine registration in just six months. All actions were performed according to a detailed daily schedule. It took as much effort to close the project too. All reports concerning just the final stage of the project weighed approximately 11 kilograms, and the complete stack of reports and technology documentation comprises a tightly packed filing cabinet.
The Institute receives patents for the technical developments together with the Ministry of Education and Science. However, many developments are impossible to cover with patents. According to Vladimir Lunin, “the simplest” technical solutions are preserved in the form of know-how.
The contract stipulated that the Institute built the production area to manufacture the composite substance intended for implant coating. The pilot batch production and quality control comply with the Good Manufacturing Practice. This international system of standards and rules was established specially for manufacturing medicines and medical products. According to the system, absolutely every production and medicine parameters are controlled — instead of checking random batches.
In November 2010, the developers received the registration certificate and the certificate of conformance for Gamalant-paste Forte composite medicine for bone transplantation and use jointly with metal implants. The medicine is expected to ship in February 2011.
A natural and important issue arises: who will organise training for doctors targeted at using this new generation substance? “We need to attract serious specialists understanding the necessity of such training, — said Vladimir Lunin. — This medicine cannot be introduced in medical practice through promotional leaflets. It must be applied strictly according the instructions, and that should be done by professional surgeons. Careless application of the medicine might result in outgrowths on the bone. If the medicine gets into a muscle, a bone can grow there as well. Fortunately, when the project work was nearing its ending there already were many surgeons who “understood” the medicine. Currently, we work with many departments of the N.N. Priorov Central Research Institute of Traumatology and Orthopedics, and the Institute in question is already planning to start using the medicine.
The laboratory led by Lunin continues its work on new osteoplastic materials. The researchers are currently busy developing low-invasive forms — that is, the medicines that can be introduced directly to the spot of the break with a syringe or a catheter. If the break requires no metal constructions, and mere external fixation is enough but partial bone destruction is present in the picture such medicines can be injected there without further trauma for the organism. This research is conducted as a separate project funded by the Ministry of Industry and Trade. According to the contract, manufacturing facilities for the medicines are to be implemented at the Gamaleya Research Institute in 2011.