the Future Prospects

the Future Prospects

(Parte 1 de 5)

The Future Prospects of Microbial Cellulose in Biomedical Applications

Wojciech K. Czaja,†,‡ David J. Young,† Marek Kawecki,§ and R. Malcolm Brown, Jr.*,†

Section of Molecular Genetics and Microbiology, University of Texas at Austin, Austin, Texas 78713, Institute of Technical Biochemistry, Technical University of Lodz, Stefanowskiego 4/10, Lodz 90-924, Poland, and Center of Burn Healing, Jana Pawla I 2, Siemianowice SÄlaskie, Poland

Received June 28, 2006; Revised Manuscript Received September 8, 2006

Microbial cellulose has proven to be a remarkably versatile biomaterial and can be used in wide variety of applied scientific endeavors, such as paper products, electronics, acoustics, and biomedical devices. In fact, biomedical devices recently have gained a significant amount of attention because of an increased interest in tissue-engineered products for both wound care and the regeneration of damaged or diseased organs. Due to its unique nanostructure and properties, microbial cellulose is a natural candidate for numerous medical and tissue-engineered applications. For example, a microbial cellulose membrane has been successfully used as a wound-healing device for severely damaged skin and as a small-diameter blood vessel replacement. The nonwoven ribbons of microbial cellulose microfibrils closely resemble the structure of native extracellullar matrices, suggesting that it could function as a scaffold for the production of many tissue-engineered constructs. In addition, microbial cellulose membranes, having a unique nanostructure, could have many other uses in wound healing and regenerative medicine, such as guided tissue regeneration (GTR), periodontal treatments, or as a replacement for dura mater (a membrane that surrounds brain tissue). In effect, microbial cellulose could function as a scaffold material for the regeneration of a wide variety of tissues, showing that it could eventually become an excellent platform technology for medicine. If microbial cellulose can be successfully mass produced, it will eventually become a vital biomaterial and will be used in the creation of a wide variety of medical devices and consumer products.

Introduction

Rapid progress has been made in recent years in the field of biomedical materials, which utilize both natural and synthetic polymers and which can be used in a variety of applications, including wound closure, drug delivery systems, novel vascular grafts, or scaffolds for in vitro or in vivo tissue engineering. Several microbially derived polysaccharides (i.e., hyaluronic acid, dextran, alginate, scleroglucan) have interesting physical and biological properties and are particularly useful in various biomedical applications. Microbial cellulose (MC), a polysaccharide synthesized in abundance by Acetobacter xylinum, has already been used quite successfully in wound-healing applications, proving that it could become a high-value product in the field of biotechnology.1-3

Traditional plant-originated cellulose and cellulose-based materials, usually in the form of woven cotton gauze dressings, have been used in medical applications for many years and are mainly utilized to stop bleeding. Even though this conventional dressing is not ideal, its use continues to be widespread. These cotton gauzes, consisting of an oxidized form of regenerated plant cellulose, were developed by Frantz during World War I, and have been successfully used as a hemostatic agent as well as an adhesion barrier.4-8 Another product, a plant cellulose sponge, has an established clinical application in wound-healing research as a component which stimulates granulation tissue in the wound bed after injury.9 In addition, several studies described the implantation of regenerated cellulose hydrogels and revealed their biocompatibility with connective tissue formation and long-term stability.9,10 Other in vitro studies showed that regenerated cellulose hydrogels promote bone cell attachment and proliferation and are very promising materials for orthopedic applications.10-13

Although chemically identical to plant cellulose, the cellulose synthesized by Acetobacter is characterized by a unique fibrillar nanostructure which determines its extraordinary physical and mechanical properties, characteristics which are quite promising for modern medicine and biomedical research. In this review, the structural features of microbial cellulose and its properties are discussed in relation to the current and future status of its application in medicine.

The Significant Biomedical Potential of Microbial Cellulose Stems from Its Unique Structure and Properties

Cellulose synthesis by Acetobacter is a complex process and involves (A) the polymerization of single glucose residues into linear !-1,4-glucan chains, (B) the extracellullar secretion of these linear chains, and (C) the assembly and crystallization of the glucan chains into hierarchically composed ribbons.14 As a result of these processes, a three-dimensional, gelatinous structure is formed on the surface of a liquid medium. The physical and mechanical properties of microbial cellulose membranes arise from their unique structure, which differs significantly from the structure of plant cellulose. Basically, well-separated nano- and microfibrils of microbial cellulose create an extensive surface area which allows it to hold a large amount of water while maintaining a high degree of conformability. The hydrogen bonds between these fibrillar units stabilize the whole structure and give it a great deal of

* Corresponding author. † University of Texas at Austin. ‡ Technical University of Lodz. § Center of Burn Healing.

10.1021/bm060620d C: $3.50 © x American Chemical Society PAGE EST: 1.2Published on Web 12/01/2006

mechanical strength.15-17 Even though plant cellulose is composed of microfibrils which are similar to those found within microbial cellulose, the plant cellulose microfibrils are part of a larger aggregation of the cell wall. Thus, microbial cellulose can absorb much higher volumes of liquids than plant-derived cellulose materials. On the basis of its recent clinical performance and according to the results of other research on the properties of this particular biomaterial, MC can be considered an ideal material for high-quality wound dressings. Table 1 summarizes most of the physical and mechanical properties of microbial cellulose which characterize it as an ideal wound dressing material. Interestingly, many Acetobacterstrains display significant differences in the cellulose production process (i.e., the rate of cellulose ribbon extrusion from a single cell may significantly vary between strains), as well as in the structure of the synthesized polymer. Figure 1 presents SEM images of cellulose structures synthesized by two different strains of Acetobacter. The differences in the size of the cellulose ribbons can be clearly seen. From a bioengineering point of view, these structural differences are of great importance since they can be used to create hybrid materials with desired properties consisting of cellulose products synthesized by different Acetobacter strains.

The given medical application should dictate the choice of the particular cellulose structure (specific Acetobacter strain). For example, implantable cellulose for artificial skin should ideally display high porosity, with interconnected pores of 50- 150 µm, in order to facilitate skin cell integration into the cellulose scaffold, whereas temporary wound dressings should have a nanoporous structure and should keep the wound moist during the healing process.18,19

One of the main requirements of any biomedical material is that it must be biocompatible, which is the ability to remain in contact with living tissue without causing any toxic or allergic side effects. A material composed of porous plant cellulose has been shown to be biocompatible with bone tissue and hepatocytes.9,20 Research conducted on an implanted cellulose sponge showed that it can be regarded as a slowly degradable material.9

As mentioned by the same authors, this material can be considered nondegradable if used as a temporary wound coverage for a short period of time.9 Unlike plant-originated cellulose, microbial cellulose is free of lignin and hemicelluloses. However, microbial cellulose is treated with strong bases in order to completely remove bacterial cells embedded in the polymer net.3,21 There are several in vivo biocompatibility studies that used MC on animal models. For example, Kolodziejczyk and Pomorski implanted pieces of microbial cellulose (1 cm in diameter) into subcutaneous pockets on rabbits and

Table 1. Properties of Microbial Cellulose Membranes and How They Relate to the Properties of an Ideal Wound Dressing Materiala properties of ideal wound care dressing properties of microbial cellulose maintain a moist environment at the wound/dressing surface high water holding capacity (typical membrane can hold up to 200 g of its dry mass in water); high water vapor transmission rate provide physical barrier against bacterialinfections nanoporous structure does not allow any external bacteria to penetrate into the wound bed highly absorbable partially dehydrated membrane is able to absorb fluid up to its original capacity.

Physical processing of the membrane (i.e., squeezing) can remove part of the initial water and allow the membrane to be more absorbable sterile, easy to use, and inexpensive membranes are easy to sterilize (by steam or γ-radiation) and package. The estimated cost of production of 1 cm2 is $0.02 available in various shapes and sizes ability to be molded in situ provide easy and close wound coverage, but allow easy and painless removal high elasticity and conformability significantly reduce pain during treatment the unique MC nanomorphology of never-dried membrane promotes specific interaction with nerve endings provide porosity for gaseous and fluid exchange highly porous material with pore sizes ranging from several nanometers to micrometers nontoxic, nonpyrogenic, and biocompatible biocompatible, nonpyrogenic, nontoxic provide high conformability and elasticity high elasticity and conformability provide mechanical stability high mechanical strength [Young’s modulus value of several GPa]Refs 90-97.

Figure 1. Structure of cellulose produced by two different Acetobacter strains clearly indicate differences. (A) NQ5, (B) E25; much larger cellulose ribbons of NQ5 are clearly distinguishable. Whereas the NQ5 strain creates a highly compact and rigid membrane, the E25 strain produces a more gelatinous, yet still rigid form of cellulose, which is highly translucent (images captured by Dwight Romanovicz, University of Texas at Austin).

B Czaja et al. Biomacromolecules periodically examined them after 1 and 3 weeks.2 The implants did not cause any macroscopic inflammatory responses, and histological observations showed only a small number of giant cells and a thin layer of fibroblasts at the interface between the cellulose and the tissue.2 Positive results were also obtained by Oster et al. in an in vitro study using mouse fibroblasts cells.23 A specific in vivo biocompatibility study of microbial cellulose has also been conducted by Klemm et al., who implanted cellulose in the form of a hollow tube as an interposition segment of the carotid arteries of rats.24 In a recent, very systematic study by Helenius et al., pieces of microbial cellulose were implanted into rats.25 Those implants evaluated after 1, 4, and 12 weeks showed no macroscopic or histologic signs of inflammation and no presence of giant cells. Also, according to the authors, no chronic inflammatory responses were observed throughout the course of the studies.25 Instead, they observed the formation of new blood vessels around and inside the implanted cellulose.25 Interestingly, the authors also noticed that cells, mostly fibroblasts, were able to significantly penetrate the more porous bottom side of a microbial cellulose membrane. The newly formed tissue, integrated with MC, contained fibroblasts and newly synthesized collagen.

Microbial Cellulose as a Wound-Healing System: Temporary Wound Coverage

Microbial Cellulose in the Treatment of Chronic Wounds and Burns. Wound healing is a dynamic process that involves the complex interaction of various cell types, extracellular matrix (ECM) molecules, and soluble compounds.26 Typically, normal wound healing progresses through a series of processes including homeostasis, inflammation, granulation tissue formation, and remodeling.26 Chronic wounds, such as ulcers, do not heal because one or more of these processes fail to function properly. Thus, successful wound treatments improve the tissue repair process by counteracting the inherent abnormalities of the chronic wound. Once the barriers to normal tissue repair are removed, the healing process can begin, which involves autolytic debridement, granulation tissue formation, and re-epithelization.26

In order to eliminate the hostile environment within the chronic wound and to facilitate proper healing, wound dressings of various types have been developed and administered. For example, ulcers are typically treated with dressings such as hydrogels, hydrocolloids, synthetic and biological membranes, and alginate.27 In 1962, George Winter discovered that healing, and specifically re-epithialization, was accelerated if the wound was kept moist.28 Since then, almost all effective wound dressings are designed to maintain a moist environment within the affected region. In fact, proteolytic activity may be elevated in a moist environment, resulting in the stimulation and accumulation of growth factors.29 Moist dressings are permeable to water, and this property has advantages for wound healing. For example, high water vapor permeable dressings show enhanced healing, probably due to an increased concentration of growth-promoting factors within the exudate and to the creation of a more extensive ECM of fibrin(ogen) and fibronectin.30

Burns are very complex injuries, causing extensive damage to skin tissues. The healing process involves the regeneration of the epidermis and the repair of the dermis, both of which result in the formation of scar tissue.31 One of the major goals of burn therapy is to quickly accomplish effective wound closure so as to increase the rate of healing and to provide immediate pain relief.32-34 In addition, proper wound management must prohibit the wound from becoming infected and dehydrated.35,36 Despite the fact that many different biological and synthetic wound dressings have already been developed, the search for an ideal wound dressing is still in progress. According to the modern approaches in the field of wound healing, an ideal wound dressing system must be structurally and functionally similar to autograft skin.31,37

Because of its unique properties, microbial cellulose (MC) has been shown to be an highly effective wound dressing material. In fact, the results of various studies indicate that topical applications of MC membranes improve the healing process of burns and chronic wounds. The progress in this field has been discussed in a recent publication.1 In addition, a recent study conducted in Poland used never-dried MC membranes in order to treat patients with severe second-degree burns.38 This study showed that the skin of the patients whose burns were covered with never-dried MC membranes healed faster (faster re-epithelialization) than the wounds of patients who received a conventional wound dressing (such as wet gauze and ointments).38 The Polish study also found that MC membranes actually performed better than conventional wound dressings in (1) conforming to the wound surface (excellent molding to all facial contours and a high degree of adherence even to the contoured parts such as nose, mouth, etc. were observed), (2) maintaining a moist environment within the wound, (3) significantly reducing pain, (4) accelerating re-epithelialization and the formation of granulation tissue, and (5) reducing scar formation.38,39 These MC membranes can be created in any shape and size, which is beneficial for the treatment of large and difficult to cover areas of the body (Figure 2).

(Parte 1 de 5)

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