This summer-friendly textile has so many uses in medicine!

Mulberry silk has a wide range of application potentials in the medical field. Its natural origin, biocompatibility and good physical properties make it an ideal choice for various medical and biotechnology applications.

Written by | MY

The word “silk” is translated into English as silk. In fact, this word refers to the protein polymer produced by various insect larvae (such as silkworms, flies, scorpions, spiders and mites, etc.), that is, the material that forms cocoons. In daily life, smooth and beautiful silk usually refers to textiles made from mulberry silk produced by silkworms (Bombyx mori, Bombyx mori) [1]**. Silkworms have been domesticated by humans and have a history of thousands of years of cultivation. Mulberry silk occupies a dominant position in the sericulture industry, and most of the natural silk commonly seen on the market belongs to this type. **In addition, undomesticated silkworms also provide us with raw materials for silk. Common wild species include Ellis (Philosamia ricini/Samia ricini), Muga (Anthrraea asana/assamensis) and Dusal (Antheraea mylitta), which do not feed on mulberry trees [2].

Beautiful silk has always been considered a gorgeous fabric, but the uses of mulberry silk go far beyond that. In recent years, scientists have discovered many medical uses for mulberry silk, and its outstanding physical and biological properties are changing bioengineering.

Structure and properties of mulberry silk

Mulberry silk is mainly composed of fibroin and sericin. Fibroin is present inside the silk, accounting for about 70% of the weight of silk, and has a diameter of about 10–25 μm. Fibroin is composed of a 1:1 ratio of amino acid light chain (about 26 kDa, Da is the unit of molecular weight) and heavy chain (about 390 kDa), which are connected by disulfide bonds. Wrapped outside the fibroin is sericin, which accounts for 30% of the total weight (20–310 kDa) [3]. In addition, mulberry silk also contains about 0.8-1% fat/wax and 1-1.4% pigment/ash. Fibroin can be extracted and purified from sericin by boiling the cocoons in an alkaline solution [4].

When we talk about the excellent biological and mechanical properties of mulberry silk, we are actually discussing the properties of silk fibroin. Silk fibroin has a unique β-folded structure that allows the hydrogen bonds of the protein to form a tight stack between antiparallel chains; the larger hydrophobic regions are interlaced with the smaller hydrophilic regions, which promotes the assembly of silk and the strength and elasticity of silk fibers. It is this unique β-folded structure that makes silk considered one of the strongest biomaterials (with a pressure resistance greater than 1 GPa) [5].

In tissue engineering for in vitro culturing of human organ tissues, the materials used must match the hardness of the target tissue. Generally, a common method to improve the mechanical strength of biopolymers is "cross-linking", that is, cross-linking between different macromolecules or introducing macromolecular materials and tissues. However, the macromolecular compounds present in this process may cause adverse consequences such as cytotoxicity and immune response. The β-folded structure of silk fibroin allows it to obtain good mechanical strength without any cross-linking steps [6] . In the actual production process, by adjusting the content of the β-folded structure, silk fibroin materials with different morphologies and mechanical strengths can be obtained, thereby providing a stable and supportive environment for cell attachment, expansion and growth [3]. Therefore, mulberry silk has great potential for medical applications.

Figure 1. Schematic diagram of the structure of silkworm cocoons | Source: DOI 10.31031/RDMS.2019.10.000740

Biomedical applications of silk

1. Surgical sutures

The most common and oldest medical application of mulberry silk is surgical sutures, which are commonly found in absorbable and non-absorbable forms.

Around 200 BC, Hippocrates and his disciples gradually spread surgical medicine after reviving medical thought and teaching. Influenced by this, the Roman medical journalist and teacher Aurelius Cornelius Celsus (De Re Medicina) wrote De Re Medicina around 50 AD, in which he described how to use braided sutures. Galen of the same period first described the use of catgut as a suture material to suture the severed tendons of gladiators, and suggested that if possible, after flushing the wound with a large amount of diluted wine, one could try to suture the wound with silk thread. Although he did not put this theory into practice, his theory of using mulberry silk as a suture material influenced later people's exploration of sutures. In the 16th to 18th centuries, there were records of surgeons using silk to suture blood vessels. In 1986, non-absorbable sterile silk thread soaked in carbolic acid was promoted for use [7].

As modern scientists explore the structure of silk, absorbable sutures based on silk fibroin have been created and popularized. Absorbable sutures do not trigger immune rejection in the human body and will gradually degrade and absorb in the body, without the need to be removed again after surgery. They are often used for internal sutures, such as tissue repair or visceral surgery.

2. Wound Dressing

Another major use of silk fibroin is wound dressings. Its synthetic dressings in different forms help promote skin regeneration and healing. Wound healing is a complex process that involves many interactions between cells and cell matrices. The initial stage of wound healing is the generation of an inflammatory response, which can last for two days. At this stage, the coagulation cascade, inflammatory pathways, and the immune system are activated to prevent further loss of blood and fluid. The second stage is the formation of new tissue, which involves processes such as collagen/matrix deposition, epithelial regeneration, angiogenesis, and wound contraction [8].

An ideal wound dressing should have the following characteristics: 1) keep the wound area moist while avoiding excessive drying and absorbing wound exudate; 2) have good air permeability; 3) be able to prevent bacterial infection; 4) have good waterproof properties[9]. To meet the above characteristics, silk fibroin is often processed into wound dressings in different forms such as films and hydrogels. Common hydrogel dressings are usually made of naturally occurring macromolecular polymers such as chitosan, alginate, collagen and hyaluronic acid[10] . Researchers have found that hydrogel dressings made of silk fibroin can not only meet the above conditions, but also induce cell growth, migration, proliferation and production of extracellular matrix [11].

In addition, mixing silk fibroin with other natural hydrogel dressings can also improve their mechanical strength[12] . Films and porous sponges are two other common dressing forms. They have a large surface area and a network of interconnected pores, which facilitates the ingrowth and adhesion of tissue cells [3] . Experimenters have found that using silk fibroin sponges mixed with collagen can be used as a dermal substitute [13].

The reason why silk fibroin can accelerate wound healing is that it activates different wound healing cell signaling pathways during contact with the wound. First, silk fibroin promotes the expression of common proteins in the cell proliferation stage, such as epidermal growth factor (EGF), fibronectin, vascular endothelial growth factor (VEGF), IL-10 (interleukin 10), IL-1β, transforming growth factor (TGF) and cell cycle proteins through the NF-κB signaling pathway, thereby promoting cell growth and proliferation and the disappearance of inflammation. Secondly, silk fibroin activates the mitogen-activated protein kinase (MAPK) signaling pathway, which plays a key role in wound healing. Silk fibroin can also affect a series of cell anti-apoptosis signaling pathways, promote cell survival, and thus promote wound healing [11, 14].

Figure 2. Common structures of silk fibroin: A) silkworm cocoon B) sponge structure C) cushion-like macrostructure D) 3D printed porous structure E cushion-like microstructure F) film-like G) hydrogel. Image source: Reference [15]

3. Drug delivery vehicle

Drug delivery carriers have been a hot research area in recent years, and silk fibroin also has a place in this field. Compared with the common nanoparticle delivery systems on the market, in addition to the good biodegradability mentioned above, the biggest advantage of silk fibroin is its low immunogenicity and will not induce immune rejection in the body. The mild processing conditions of silk fibroin make it more suitable for encapsulating drugs that are sensitive to processing conditions and easy to fail.

Silk fibroin drug carriers can be processed into structures such as hydrogels, films, micro- and nanoparticles, nanofibers, and porous sponges, which are suitable for different drug delivery routes and treatment needs and have a wider range of application potentials[16] . By optimizing the design of silk fibroin structure, the drug release rate can be slowed down and the carrier stability can be enhanced, thus prolonging its circulation time and effect in the blood, and achieving the purpose of sustained drug delivery [17].

Currently, microparticles and nanoparticles have been used to deliver different types of drugs (such as curcumin, doxorubicin, and ibuprofen) to various types of cells in a time- or site-specific manner. Silk fibroin films have been used for the controlled release of drugs (such as dextran, epirubicin) and biologics (such as IgG and HIV inhibitors). In addition, they have been used to stabilize biologics such as horseradish peroxidase (HRP), glucose oxidase, vaccines, and monoclonal antibodies to extend their shelf life [16].

Researchers have also tried to use silk as a vaccine carrier. The silk protein microneedle patch for influenza vaccine developed by Vaxess, a Boston-based company, has completed phase I clinical trials. At the same time, silk protein vaccine carriers for COVID-19 are also under development. Although great progress has been made in the study of silk protein as a vaccine delivery carrier, this process is still in the exploration and development stage. There is still a lack of sufficient research on its targeting and specificity, and there is still a long way to go before the real product comes out.

4. Repairing the tissue scaffold

The goal of modern tissue engineering is to regenerate and replace damaged tissues and organs. The ideal material should not only successfully replace tissues or organs, but also have a supporting effect. This means that it should be fully integrated into the surrounding tissues and not trigger an immune response or produce adverse effects. Scaffolds made of silk fibroin meet the above two requirements very well. While not triggering an immune response, they provide support and protection for cell tissues, promote the deposition and growth of related cells on the scaffold, and thus restore damaged tissues[18].

Silk fibroin scaffolds have been widely explored for use in ligaments, tendons, and bone tissue. Bone is a special type of connective tissue, with collagen and hyaluronic acid being the main components of bone tissue. An ideal bone tissue scaffold should allow inorganic components to deposit on the scaffold to enhance bone hardness and strength while ensuring the toughness of bone tissue. Just as we use silk thread to enhance the strength and durability of fabrics when we weave cloth, silk fibroin scaffolds can provide a stable, solid, and suitable growth environment for bone cells, promoting the regeneration and repair of bone tissue. Experiments have shown that silk fibroin scaffolds can promote the osteogenic differentiation of human mesenchymal stem cells. In addition, researchers have found that the combination of silk fibroin scaffolds with other biomaterials (such as collagen or calcium phosphate inorganic components) can enhance the expression of bone morphogenetic proteins, thereby enhancing osteogenic properties [19].

Ligament and tendon tissues are composed of collagen and fibroblasts. They are dense fibrous connective tissues that lack natural regeneration capabilities. The high toughness and elasticity of ligaments and tendons make silk fibroin scaffolds the preferred biopolymer for ligament and tendon tissue engineering [20] . These scaffolds are obtained by mixing silk fibroin with natural biomaterials (such as type I collagen, hyaluronic acid and gelatin) and synthetic materials (such as polyelectrolytes and PLGA). In 2002, the first silk fibroin matrix was successfully applied to the design of the anterior cruciate ligament (ACL) [21].

In addition, silk fibroin can also be used as an ideal material for repairing perforated eardrums. The eardrum is a transparent structure located between the outer ear and the middle ear that receives sound while protecting the middle ear [22]. If the eardrum does not repair itself within 3 months, a chronic perforation will form, leading to hearing loss and repeated infections. The eardrum is mainly composed of keratinocytes, fibroblasts, and collagen (type II and type III). Scaffolds made of silk fibroin allow human eardrum keratinocytes to grow and proliferate, while accelerating the regeneration of the eardrum, thereby significantly accelerating the speed of hearing recovery. Experiments have shown that when silk fibroin films were transplanted into the ears of rats and guinea pigs, the perforations in the ears of rats and guinea pigs were closed after 7 days compared with the control group. The researchers also found that silk fibroin films have good sound transmission ability and excellent cartilage tensile strength, indicating that these membranes have great potential for regenerating chronic eardrum perforations in vivo [21].

summary

In addition to the important medical engineering applications mentioned above, silk fibroin can also be used to make various types of sensors. For example, by performing specific optical modifications on silk fibroin, changes in light absorption, fluorescence or refractive index can be detected, which can be applied to biosensing, environmental monitoring and medical diagnosis. By linking silk fibroin with specific biorecognition molecules, such as antibodies, oligonucleotides or enzymes, specific detection of biological molecules or pathogens can be achieved, which plays an important role in disease diagnosis, food safety monitoring and biodefense [23].

In the future, as the structure and properties of silk protein are further studied, people will further explore its compatibility with organisms and its potential applications in tissue engineering and regenerative medicine. Silk protein scaffolds may be designed for more complex tissue repair, such as the regeneration of organs such as the heart, liver and lungs; researchers will design silk protein drug carriers with targeted and controlled release functions for the treatment of cancer, inflammation and infectious diseases, etc.

I believe that in the future there will be more types and forms of silk protein products, which will bring more convenience to human life. Mulberry silk will become an important force in promoting human health and quality of life.

References

[1] Nguyen, TP, et al., Silk Fibroin-Based Biomaterials for Biomedical Applications: A Review. Polymers (Basel), 2019. 11(12).

[2] Holland, C., et al., The Biomedical Use of Silk: Past, Present, Future. Advanced Healthcare Materials, 2019. 8(1): p. 1800465.

[3] Qi, Y., et al., A Review of Structure Construction of Silk Fibroin Biomaterials from Single Structures to Multi-Level Structures. Int J Mol Sci, 2017. 18(3).

[4] Rockwood, DN, et al., Materials fabrication from Bombyx mori silk fibroin. Nat Protoc, 2011. 6(10): p. 1612-31.

[5] Cheng, Y., et al., On the strength of β-sheet crystallites of Bombyx mori silk fibroin. Journal of The Royal Society Interface, 2014. 11(96): p. 20140305.

[6] Lee, J., et al., Development of Silk Fibroin-Based Non-Crosslinking Thermosensitive Bioinks for 3D Bioprinting. Polymers, 2023. 15(17): p. 3567.

[7] Muffly, TM, AP Tizzano, and MD Walters, The history and evolution of sutures in pelvic surgery. JR Soc Med, 2011. 104(3): p. 107-12.

[8] Almadani, YH, et al., Wound Healing: A Comprehensive Review. Semin Plast Surg, 2021. 35(3): p. 141-144.

[9] Dhivya, S., VV Padma, and E. Santhini, Wound dressings - a review. Biomedicine (Taipei), 2015. 5(4): p. 22.

[10] Nguyen, HM, et al., Biomedical materials for wound dressing: recent advances and applications. RSC Adv, 2023. 13(8): p. 5509-5528.

[11] Mazurek, Ł., et al., Silk Fibroin Biomaterials and Their Beneficial Role in Skin Wound Healing. Biomolecules, 2022. 12(12).

[12] Zhang, H., et al., Silk fibroin hydrogels for biomedical applications. Smart Medicine, 2022. 1(1): p. e20220011.

[13] Naomi, R., J. Ratanavaraporn, and MB Fauzi, Comprehensive Review of Hybrid Collagen and Silk Fibroin for Cutaneous Wound Healing. Materials (Basel), 2020. 13(14).

[14] Park, YR, et al., NF-κB signaling is key in the wound healing processes of silk fibroin. Acta Biomaterialia, 2018. 67: p. 183-195.

[15] Belda Marín, C., et al., Silk Polymers and Nanoparticles: A Powerful Combination for the Design of Versatile Biomaterials. Frontiers in Chemistry, 2020. 8.

[16] Wani, SUD, et al., Silk Fibroin as an Efficient Biomaterial for Drug Delivery, Gene Therapy, and Wound Healing. Int J Mol Sci, 2022. 23(22).

[17] Tiwari, G., et al., Drug delivery systems: An updated review. Int J Pharm Investig, 2012. 2(1): p. 2-11.

[18] Lee, EJ, FK Kasper, and AG Mikos, Biomaterials for tissue engineering. Ann Biomed Eng, 2014. 42(2): p. 323-37.

[19] Li, M., et al., A Comprehensive Review on Silk Fibroin as a Persuasive Biomaterial for Bone Tissue Engineering. International Journal of Molecular Sciences, 2023. 24(3): p. 2660.

[20] Tang, Y., et al., Functional biomaterials for tendon/ligament repair and regeneration. Regen Biomater, 2022. 9: p. rbac062.

[21] Sun, W., et al., Silk Fibroin as a Functional Biomaterial for Tissue Engineering. International Journal of Molecular Sciences, 2021. 22(3): p. 1499.

[22] Sainsbury, E., et al., Tissue engineering and regenerative medicine strategies for the repair of tympanic membrane perforations. Biomater Biosyst, 2022. 6: p. 100046.

[23] Ru, M., et al., Recent progress in silk-based biosensors. International Journal of Biological Macromolecules, 2023. 224: p. 422-436.

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