Epigenetic Processes in Cancer Research

Catalyst Volume 9

Abstract

Epigenetics is the study of the phenotypic variation caused by external factors (e.g. diet, nicotine use, and carcinogenic chemical exposure) that influence the mechanisms through which cells read and interpret genes. Epigenetic modifications are independent of genetic mutations. Currently, epigenetics offers significant promise in novel noninvasive cancer therapies and early diagnostic tools. Key epigenetic processes including DNA methylation and histone modification are reversible, unlike most genetic mutations. Reversing these processes in tumor suppressor genes can restore normal behavior in tumor cells. This review discusses the biological basis and treatment potential of these processes and provides a brief analysis of their potential application in cancer treatment.

Introduction

DNA is a double-helical structure packed into the nuclei of eukaryotic cells in the form of chromatin. The basic unit of DNA packing in chromatin is the nucleosome, a complex of eight histone proteins and approximately 140 DNA base pairs. Further coiling of repeating nucleosome fibers eventually yields a chromatid, a key component of a chromosome.

Gene expression is epigenetically regulated through alterations in chromatin structure and the double helix of DNA by addition of functional groups. DNA methylation and histone modification are the most understood mechanisms of such epigenetic modulation. Errant modification of functional group patterns in DNA or histone tails could result in unintended silencing of critical regulatory genes, a process which contributes to the development of several diseases, including cancer. Understanding these complex mechanisms is essential for the development of cancer therapies that reverse the inhibition of tumor-suppressor genes. This review seeks to outline the biochemical basis of the gene silencing effected by DNA methylation and histone modification, as well as the current developments in reversing these mechanisms for the prevention of cancer.

Histone Modification & Its Implications on Cancer

Histone modifications are reversible modifications at the N-terminals of histones in a nucleosome. These include, but are not limited to, acetylation, methylation, phosphorylation, and ubiquitination. Depending on the group added or removed from a given histone, local gene transcription can be upregulated or downregulated. Histone acetylation, for instance, modifies chromatin into a less-condensed conformation that is more transcriptionally active. Conversely, histone deacetylases (HDACs) remove acetyl groups, increasing the ionic attractions between positively charged histones and negatively charged DNA and tightening chromatin structure. In order to further condense chromatin, HDACs can also recruit K9 histone methyltransferases (HMTs) to methylate H3K9 histone residues, providing the condensing agent heterochromatin protein 1 (HP1) with a binding site. As a result of the condensed heterochromatin conformation, the cell has limited transcriptional access to the genome.

Recent research has established histone modifications as useful biomarkers in cancer diagnosis. HDACs, specifically HDAC1, can often be identified in elevated quantities in prostate and gastric cancer patients. Prostate cancer tumors have been characterized by general hypomethylation of histone residues H4K20me1 and H4K20me2, as well as hypermethylation of residue H3K27 and increased activity of its specific methyltransferase. These conditions are now associated with progression and metastasis of prostate cancer. Histone modifications play a multifaceted role in cancer diagnosis and treatment. In addition to serving as diagnostic biomarkers, they are also potential therapeutic agents. Drugs that specifically inhibit HDACs are currently being manufactured and assessed for clinical application after recent FDA approval.⁵

DNA Methylation & Its Implications on Cancer

In DNA methylation, a methyl group is covalently added to a cytosine ring in the DNA sequence, forming a 5-methylcytosine. Molecules of 5-methylcytosine are found primarily at cytosine-guanine dinucleotides (CpGs). Some areas of high CpG density, such as CpG islands, are characteristically unmethylated and are located in or near the promoter regions of genes, allowing them to play a role in gene expression. Enzymes termed DNA methyltransferases (DNMTs) facilitate the methylation of CpG residues. Three key members of this family include Dnmt1, Dnmt3a, and Dnmt3b. Dnmt1 is labeled as the “maintenance” DNMT due to its methylation of DNA near replication forks, which preserves the epigenetic inheritance of methylation patterns across cellular generations. The other two DNMTs serve to directly methylate other CpG residues. These enzymes can either be recruited by a transcription factor bound to the promoter region of a given gene to methylate a specific CpG island or simply methylate all CpG sites across a genome not protected by a transcription factor. Several families of proteins, such as the UHRF proteins, the zinc-finger proteins, and most notably, the MBD proteins, are responsible for the interpretation of these methylation patterns. For example, MBD proteins possess transcriptional repression domains that allow them to bind to methylated DNA and silence nearby genes.

For a malignant cancer to develop, critical tumor-suppressor genes such as p53 must be knocked out. This process can be accomplished through hypermethylation of CpG islands in certain promoter regions and genes. For example, when the DNA repair gene BRCA1 becomes hypermethylated, it is rendered inactive, which can lead to the development of breast cancer. The reversibility of methylation implies that these tumor-suppressor genes could be reactivated to effectively treat tumors. Infiltration of a cell by pharmaceutical agents that inhibit methylation-mediated suppression could restore the function of a silenced tumor-suppressor gene.⁶

One recent study demonstrated that azacitidine, an agent that can be incorporated into DNA, is capable of inducing hypomethylation. The chemical modification of this agent’s diphosphate form by a ribonucleotide reductase and subsequent phosphorylation creates a triphosphate form that displaces cytosine bases in DNA. As a result, DNMTs are limited in methylation functionality since they are isolated on a substituted DNA strand.⁷ Such manipulation of naturally occurring DNA methylation processes in cancer cells appears to be a promising method of restoring the normal function of tumor-suppressor genes.

Co-Application of DNA Methylation & Histone Modification

The discovery of the protein MeCP2 has validated the hypothesis that DNA methylation regulates histone acetylation patterns. MeCP2 recruits histone HDACs to the promoter regions of methylated CpG islands. The resulting hypoacetylated histones yield a highly condensed heterochromatin structure that represses transcription of nearby genes. Clinical studies have thus emphasized the utilization of both demethylating agents and histone deacetylase inhibitors for transcriptional activation of tumor-suppressor genes.⁸

Conclusion

The field of epigenetics has opened a door to novel, promising cancer therapies unimaginable just a decade ago. Manipulating DNA methylation and histone modification can reverse tumor-suppressor gene silencing. Reactivating tumor-suppressor genes through epigenetic modifications can halt and reverse cancer proliferation.

References

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Developments in Bone Regenerative Medicine Using Stem Cell Treatment

Catalyst Volume 9

Abstract

There is an acute need for alternatives to modern bone regeneration techniques, which have in vivo morbidity and high cost. Dental pulp stem cells (DPSCs) constitute an immunocompatible and easily accessible cell source that is capable of osteogenic differentiation. In this study, we engineered economical hard-soft intercalated substrates using various thicknesses of graphene/polybutadiene composites and polystyrene/polybutadiene blends. We investigated the ability of these scaffolds to increase proliferation and induce osteogenic differentiation in DPSCs without chemical inducers such as dexamethasone, which may accelerate cancer metastasis.

For each concentration, samples were prepared with dexamethasone as a positive control. Proliferation studies demonstrated the scaffolds’ effects on DPSC clonogenic potential: doubling times were shown to be statistically lower than controls for all substrates. Confocal microscopy and scanning electron microscopy/energy dispersive X-ray spectroscopy indicated widespread osteogenic differentiation of DPSCs cultured on graphene/polybutadiene substrates without dexamethasone. Further investigation of the interaction between hard-soft intercalated substrates and cells can yield promising results for regenerative therapy.

Introduction

Current mainstream bone regeneration techniques, such as autologous bone grafts, have many limitations, including donor site morbidity, graft resorption, and high cost.^1,2 An estimated 1.5 million individuals suffer from bone-disease related fractures each year, and about 54 million individuals in the United States have osteoporosis and low bone mass, placing them at increased risk for fracture.^2,3,4,5 Bone tissue scaffold implants have been explored in the past decade as an alternative option for bone regeneration treatments. In order to successfully regenerate bone tissue, scaffolds typically require the use of biochemical growth factors that are associated with side effects, such as the acceleration of cancer metastasis.^6,7 In addition, administering these factors in vivo is a challenge.⁶ The purpose of this project was to engineer and characterize a scaffold that would overcome these obstacles and induce osteogenesis by controlling the mechanical environment of the implanted cells.

First isolated in 2001 from the dental pulp chamber, dental pulp stem cells (DPSCs) are multipotent ecto-mesenchymal stem cells.^8,9 Previous studies have shown that these cells are capable of osteogenic, odontogenic, chondrogenic, and adipogenic differentiation.^10,11,12 Due to their highly proliferative nature and various osteogenic markers, DPSCs provide a promising source of stem cells for bone regeneration.¹¹

An ideal scaffold should be able to assist cellular attachment, proliferation and differentiation.¹³ While several types of substrates suitable for these purposes have been identified, such as polydimethylsiloxane¹⁴ and polymethyl methacrylate¹⁵, almost all of them require multiple administrations of growth factors to promote osteogenic differentiation.⁶ In recent years, the mechanical cues of the extracellular matrix (ECM) have been shown to play a key role in cell differentiation, and are a promising alternative to chemical inducers.^16,17

Recent studies demonstrate that hydrophobic materials show higher protein adsorption and cellular activity when compared to hydrophilic surfaces; therefore, we employed hydrophobic materials in our experimental scaffold.^18,19,20,21 Polybutadiene (PB) is a hydrophobic, biocompatible elastomer with low rigidity. Altering the thickness of PB films can vary the mechanical cues to cells, inducing the desired differentiation. DPSCs placed onto spin-casted PB films of different thicknesses have been observed to biomineralize calcium phosphate, supporting the idea that mechanical stimuli can initiate differentiation.^6,16,17 Atactic polystyrene (PS) is a rigid, inexpensive hydrophobic polymer.²² As PB is flexible and PS is hard, a polymer blend of PS-PB creates a rigid yet elastic surface that could mimic the mechanical properties of the ECM.

Recently, certain carbon compounds have been recognized as biomimetic.²³ The remarkable rigidity and elasticity of graphene, a one-atom thick nanomaterial, make it a compelling biocompatible scaffold material candidate.²⁴ Studies have also shown that using a thin sheet of graphene as a substrate enhances the growth and osteogenic differentiation of cells.²³

We hypothesized that DPSCs plated on hard-soft intercalated substrates—specifically, graphene-polybutadiene (G-PB) substrates and polystyrene-polybutadiene (PS-PB) substrates of varying thicknesses—would mimic the elasticity and rigidity of the bone ECM and thus induce osteogenesis without the use of chemical inducers, such as dexamethasone (DEX).

Materials and Methods

G-PB and PS-PB solutions were prepared through dissolution of varying amounts of graphene and PS in PB-toluene solutions of varying concentrations. Graphene was added to a thin PB solution (3 mg PB/mL toluene) to create a 1:1 G-PB ratio by mass. Graphene was added to a thick PB solution (20 mg PB/mL toluene) to create 1:1 and 1:5 G-PB ratios by mass. PS was added to a thick PB solution to create 1:1, 1:2, and 1:4 PS-PB blend ratios by mass. Spincasting was used to apply G-PB and PS-PB onto silicon wafers as layers of varying thicknesses (thin PB: 20.5nm, thick PB: 202.0nm).²⁵ Subsequently, DPSCs were plated onto the coated wafers either with or without dexamethasone (DEX). Following a culture period of eight days, the cells were counted with a hemacytometer to determine proliferation, and then stained with xylenol orange for qualitative analysis of calcification. Cell morphology and calcification of stained cells were determined through confocal microscopy and phase contrast fluorescent microscopy. Cell modulus and scaffold surface character were determined using atomic force microscopy. Finally, cell biomineralization was analyzed using scanning electron microscopy.

Results

Cell Proliferation and Morphology

To ensure the biocompatibility of graphene, cell proliferation studies were conducted on all G-PB substrates. Results showed that G-PB did not inhibit DPSC proliferation. The doubling time was lowest for 1:1 G-thick PB, while doubling time was observed to be greatest for 1:1 G-thin PB. Multiple two-sample t-tests showed that the graphene substrates had significantly lower doubling times than standard plastic monolayer (p < 0.001).

After days 3, 5, and 8, cell morphology of the DPSCs cultured on the G-PB and PS-PB films was analyzed using phase contrast fluorescent microscopy. Images showed normal cell morphology and growth, based on comparison with the control thin PB and thick PB samples. After day 16 and 21 of cell incubation, morphology of the DPSCs cultured on all substrates was analyzed using confocal microscopy. There was no distinctive difference among the morphology of the DPSC colonies. DPSCs appeared to be fibroblast-like and were confluent in culture by day 15 of incubation.

Modulus Studies

In order to establish a relation between rigidity of the cells and rigidity of the substrate the cells were growing on, modulus measurements were taken using atomic force microscopy. Modulus results are included in Figure 3.

Differentiation Studies

After day 16 of incubation, calcification of DPSCs on all substrates was analyzed using confocal microscopy. Imaging indicated preliminary calcification on all substrates with and without DEX. Qualitatively, the DEX samples exhibited much higher levels of calcification than their non-DEX counterparts, as evident in thin PB, 1:1 G-thin PB, and 1:4 PS-PB samples.

After days 16 and 21 of incubation, biomineralization by the DPSCs cultured on the substrates was analyzed by scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDX). The presence of white, granular deposits in SEM images indicates the formation of hydroxyapatite, which signifies differentiation. This differentiation was confirmed by the presence of calcium and phosphate peaks in EDX analysis. Other crystals (not biomineralized) were determined to be calcium carbonates by EDX analysis and were not indicative of DPSC differentiation.

On day 16, only thin PB induced with DEX was shown to have biomineralized with sporadic crystal deposits. By day 21, all samples were shown to have biomineralized to some degree, except for 1:1 PS-PB (non-DEX) and 1:4 PS-PB (non-DEX). Heavy biomineralization in crystal and dotted structures was apparent in DPSCs cultured on 1:1 G-thin PB (both with and without DEX). Furthermore, samples containing graphene appeared to have greater amounts of hydroxyapatite than the control groups. All PS-PB substrates biomineralized in the presence of DEX, while only 1:2 PS-PB was shown to biomineralize without DEX. While results indicate that while PS-PB copolymers generally require DEX for biomineralization and differentiation, this is not the case for G-PB substrates. Biomineralization occurred on DPSCs cultured on G-PB substrates without DEX, demonstrating the differentiating ability of the G-PB mechanical environment and its interactions with DPSCs.

Discussion

This study investigated the effect of hard-soft intercalated scaffolds on the proliferation and differentiation of DPSCs in vitro. As cells have been shown to respond to substrate mechanical cues, we monitored the effect of ECM-mimicking hard-soft intercalated substrates on the behavior of DPSCs. We chose graphene and polystyrene as the hard components, and used polybutadiene as a soft matrix.

By using AFM for characterization of the G-PB composite and PS-PB blend substrates, we demonstrated that all surfaces had proper phase separation and uniform dispersion. This ensured that DPSCs would be exposed to both the hard peaks and soft surfaces during culture, allowing us to draw valid conclusions regarding the effect of substrate mechanics.

Modulus studies on substrates indicated that 1:1 G-thin PB was the most rigid substrate and control thin PB was the second most rigid. In general, G-PB substrates were 8-20 times stiffer than PS-PB substrates. The high relative modulus of graphene-based substrates can be attributed to the stiffness of graphene itself. The cell modulus appeared highly correlated to the substrate modulus, both indicating that the greater the stiffness of the substrate, the greater the stiffness of DPSCs cultured on it and supporting the finding that substrate stiffness affects the cell ECM.^27,28 For example, 1:1 G-thin PB had the highest surface modulus, and DPSCs grown on 1:1 G-thin PB had the highest cell modulus. Conversely, DPSCs grown on 1:4 PS-PB had the lowest cell modulus, and 1:4 PS-PB had one of the lowest relative surface moduli. Another notable trend involves DEX; cells cultured with DEX had greater moduli than cells cultured without, suggesting a possible mechanism used by DEX to enhance stiffness and thereby osteogenesis of DPSCs.

Cell morphology studies indicated normal growth and normal cell shape on all substrates. Cell proliferation studies indicated that all samples had significantly lower cell doubling times than standard plastic monolayer (p < 0.001). Results confirm that graphene is not cytotoxic to DPSCs, which supports previous research.²⁷

SEM/EDX indicated that DPSCs grown on thick PB soft substrates appeared to have increased proliferation but limited biomineralization. In contrast, cells on the hardest substrates, 1:1 G-thin PB and thin PB, exhibited slower proliferation, but formed more calcium phosphate crystals, indicating greater biomineralization and osteogenic differentiation. The success of G-PB substrates in inducing osteogenic differentiation may be explained by the behavior of graphene itself. Graphene can influence cytoskeletal proteins, thus altering the differentiation of DPSCs through chemical and electrochemical means, such as hydrogen bonding with RGD peptides.^29,30 In addition, G-PB substrate stiffness may upregulate levels of alkaline phosphatase and osteocalcin, creating isometric tension in the DPSC actin network and resulting in greater crystal formation.³⁰ Overall, the proliferation results indicate that cells that undergo higher proliferation will undergo less crystal formation and osteogenic differentiation (and vice-versa).

The data presented here indicate that hard-soft intercalated substrates have the potential to enhance both proliferation and differentiation of DPSCs. G-PB substrates possess greater differentiation capabilities, whereas PS-PB substrates possess greater proliferative capabilities. Within graphene-based substrates, 1:1 G-thin PB induced the greatest biomineralization, performing better than various other substrates induced with DEX. This indicates that substrate stiffness is a potent stimulus that can serve as a promising alternative to biochemical factors like DEX.

Conclusion

The development of an ideal scaffold has been the focus of significant research in regenerative medicine. Altering the mechanical environment of the cell offers several advantages over current strategies, which are largely reliant on growth factors that can lead to acceleration of cancer metastasis. Within this study, the optimal scaffold for growth and differentiation of DPSCs was determined to be the 1:1 G-thin PB sample, which exhibited the greatest cell modulus, crystal deposition, and biomineralization. In addition, our study indicates two key relationships: one, the correlation between substrate and cell rigidity, and two, the tradeoff between scaffold-induced proliferation and scaffold-induced differentiation of cells, which depends on substrate characteristics. Further investigation of hard-soft intercalated substrates holds potential for developing safer and more cost-effective bone regeneration scaffolds.

References

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12. Chen, S. et al. Arch Oral Biol. 2005, 50(2), 227–236.
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16. Engler, A. J. et al. Cell. 2006, 126(4), 677–689.
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27. Jana, B. et al. Chem Commun. 2014, 50(78), 11595–11598.
28. Nayak, T. R. et al. ACS Nano. 2010, 4(12), 7717–7725.
29. Banks, J. M. et al. Biomaterials. 2014, 35(32), 8951–8959.
30. Arnsdorf, E. J. et al. J Cell Sci. 2009, 122(4), 546–553.

Developments in Gold Nanoparticles and Cancer Therapy

Catalyst Volume 9

Abstract

Nanotechnology has recently produced several breakthroughs in localized cancer therapy. Specifically, directing the accumulation of gold nanoparticles (GNPs) in cancerous tissue enables the targeted release of cytotoxic drugs and enhances the efficacy of established cancer therapy methods. This article will give a basic overview of the structure and design of GNPs, the role of GNPs in drug delivery and localized cancer therapy, and the challenges in developing and using GNPs for cancer treatment.

Introduction

Chemotherapy is currently the most broadly utilized method of treatment for most subtypes of cancer. However, cytotoxic chemotherapy drugs are limited by their lack of specificity; chemotherapeutic agents target all of the body’s most actively dividing cells, giving rise to a number of dangerous side effects.¹ GNPs have recently attracted interest due to their ability to act as localized cancer treatments—they offer a non-cytotoxic, versatile, specific targeting mechanism for cancer treatment and a high binding affinity for a wide variety of organic molecules.² Researchers have demonstrated the ability to chemically modify the surfaces of GNPs to induce binding to specific pharmaceutical agents, biomacromolecules, and malignant cell tissues. This allows GNPs to deliver therapeutic agents at tumor sites more precisely than standard intravenous chemotherapy can. GNPs also increase the efficiency of established cancer therapy methods, such as hyperthermia.³ This article will briefly cover the design and characteristics of GNPs, and then outline both the roles of GNPs in cancer therapy and the challenges in implementing GNP-based treatment options.

Design and Characteristics of Gold Nanoparticles

Nanoparticle Structure

To date, gold nanoparticles have been developed in several shapes and sizes.⁴ Although GNPs have also successfully been synthesized as rods, triangles, and hexagons, spherical GNPs have been demonstrated to be one of the most biocompatible nanoparticle models. GNP shape affects accumulation behavior in cells.⁴ A study by Tian et. al. found that hexagonal GNPs produced a greater rate of vesicular aggregation than both spherical and triangular GNPs.

Differences in GNP shape also cause variation in surface area and volume, which affects cellular uptake, biocompatibility, and therapeutic efficiency.⁴ For example, GNPs with greater surface area or more vertices possess enhanced cell binding capabilities but also heightened cell toxicity. Clearly, GNP nanoparticles must be designed with respect to their intended function.

Nanoparticle Surface Modification

In order to target specific cells or tissues, GNPs must undergo a ligand attachment process known as surface modification. The types of ligand particles attached to a GNP affect its overall behavior. For example, ligand particles consisting of inert molecular chains can stabilize nanoparticles against inefficient aggregation.⁵ Polyethylene glycol (PEG) is a hydrocarbon chain that stabilizes GNPs by repelling other molecules using steric effects; incoming molecules are unable to penetrate the PEG-modified surface of the GNPs.⁵ Certain ligand sequences can enable a GNP to strongly bind to a target molecule by molecular recognition, which is determined by geometric matching of the surfaces of the two molecules.⁵

Tumor cells often express more cell surface receptors than normal cells; targeting these receptors for drug delivery increases drug accumulation and therapeutic efficacy.⁶ However, the receptors on the surface of tumor cells must be exclusive to cancerous cells in order to optimize nanoparticle and drug targeting. For example, most tumor cells have integrin receptors.⁷ To target these residues, the surfaces of the therapeutic GNPs can be functionalized with the arginine-glycine-aspartic acid (RGD) sequence, which binds to key members within the integrin family.⁸ Successful targeting can lead to endocytosis and intracellular release of the therapeutic elements that the GNPs carry.

An important factor of GNP therapy is the efficient targeting and release of remedial agents at the designated cancerous site. There are two types of GNP targeting: passive and active. In passive targeting, nanoparticles accumulate at a specific site by physicochemical factors (e.g. size, molecular weight, and shape), extravasation, or pharmacological factors. Release can be triggered by internal factors such as pH changes or external stimuli such as application of light.² In active targeting, ligand molecules attached to the surface of a GNP render it capable of effectively delivering pharmaceutical agents and large biomacromolecules to specific cells in the body.

Gold Nanoparticles in Localized Cancer Therapy

Hyperthermia

Hyperthermia is a localized cancer therapy in which cancerous tissue is exposed to high temperatures to induce cell death. Placing gold nanoparticles at the site of therapy can improve the efficiency and effectiveness of hyperthermia, leading to lower levels of tumor growth. GNPs aggregated at cancerous tissues allow intense, localized increases in temperature that better induce cell death. In one study on mice, breast tumor tissue containing aggregated GNPs experienced a temperature increase 28°C higher than control breast tumor tissue when subjected to laser excitation.⁹ While the control tissue had recurring cancerous growth, the introduction of GNPs significantly increased the therapeutic temperature of the tumors and permanently damaged the cancerous tissue.

Organelle Targeting

GNPs are also capable of specifically targeting malfunctioning organelles in tumor cells, such as nuclei or mitochondria. The nucleus is an important target in localized cancer therapy since it controls the processes of cell growth, proliferation, and apoptosis, which are commonly defective in tumor cells. Accumulation of GNPs inside nuclei can disrupt faulty nuclear processes and eventually induce cell apoptosis. The structure of the GNP used to target the nucleus determines the final effect. For example, small spherical and “nanoflower”-shaped GNPs compromise nuclear functioning, but large GNPs do not.¹⁰

Dysfunctional mitochondria are also valuable targets in localized therapy as they control the energy supply of tumor cells and are key regulators of their apoptotic pathways.¹⁰ Specific organelle targeting causes internal cell damage to cancerous tissue only, sparing normal tissue from the damaging effects of therapeutic agents. This makes nuclear and mitochondrial targeting a desirable treatment option that merits further investigation.

Challenges of Gold Nanoparticles in Localized Cancer Therapy

Cellular Uptake

Significant difficulties have been encountered in engineering a viable method of cellular GNP uptake. Notably, GNPs must not only bind to a given cancer cell’s surface and undergo endocytosis into the cell, but they must also evade endosomes and lysosomes.¹⁰ These obstacles are present regardless of whether the GNPs are engineered to target specific organelles or release therapeutic agents inside cancerous cells. Recent research has demonstrated that GNPs can avoid digestion by being functionalized with certain surface groups, such as polyethylenimine, that allow them to escape endosomes and lysosomes.¹⁰

Toxic Effects on Local Tissue

The cytotoxic effects of GNPs on local cells and tissues remain poorly understood.¹¹ However, recent research developments have revealed a relationship between the shapes and sizes of GNPs and their cell toxicities. Larger GNPs have been found to be more cytotoxic than smaller ones.¹² Gold nanospheres were lethal at lower concentrations, while gold nanostars were less toxic.¹³ While different shapes and sizes of GNPs can be beneficial in various localized cancer therapies, GNPs must be optimized on an application-by-application basis with regard to their toxicity level.

Conclusion

Gold nanoparticles have emerged as viable agents for cancer therapy. GNPs are effective in targeting malignant cells specifically, making them less toxic to normal cells than traditional cancer therapies. By modifying their surfaces with different chemical groups, scientists can engineer GNPs to accumulate at specific tumor sites. The shape and size of a GNP also affect its behavior during targeting, accumulation, and cellular endocytosis. After accumulation, GNPs may be used to enhance the efficacy of established cancer therapies such as hyperthermia. Alternatively, GNPs can deliver chemotherapy drugs to tumor cells internally or target specific organelles inside the cell, such as the nucleus and the mitochondria.

Although some research has shown that GNPs themselves do not produce acute cytotoxicity in cells, other research has indicated that nanoparticle concentration, shape, and size may all affect cytotoxicity. Therefore, nanoparticle design should be optimized to increase cancerous cell death but limit cytotoxicity in nearby normal cells.

References

1. Estanquiero, M. et al. Colloid Surface B. 2015, 126, 631-648.
2. Pissuwan, D. et al. J Control Release. 2009, 149, 65-71.
3. Chatterjee, D. V. et al. Ther Deliv. 2011, 2, 1001-1014.
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13. Favi, P. M. et al. J Biomed Mater Red A. 2015, 103, 3449-3462. 

Mastering Mega Minds: Our Quest for Cognitive Development

Catalyst Volume 9

Humans are continuously pursuing perfection. This drive is what motivates scientific researchers and comic book authors to dream about the invention of bionic men. It seems inevitable that this quest has expanded to target humankind’s most prized possession: our brain. Cognitive enhancements are various technologies created in order to elevate human mental capacities. However, as scientists and entrepreneurs attempt to research and develop cognitive enhancements, society faces an ethical dilemma. Policy must help create a balance, maximizing the benefits of augmented mental processing while minimizing potential risks.

Cognitive enhancements are becoming increasingly prevalent and exist in numerous forms, from genetic engineering to brain stimulation devices to cognition-enhancing drugs. The vast differences between these categories make it difficult to generalize a single proposition that can effectively regulate enhancements as a whole. Overall, out of these types, prescription pills and stimulation devices currently have the largest potential for widespread usage.

Prescription pills exemplify the many benefits and drawbacks of using cognitive enhancements. ADHD medications like Ritalin and Adderall, which stimulate dopamine and norepinephrine activity in the brain, may be the most ubiquitous example of available cognitive enhancements. These drugs are especially abused among college students, who use these pills to stay awake for longer periods of time and enhance their attention while studying. In a collection of studies, 4.1 to 10.8% of American college students reported recreationally using a prescription stimulant in the past year, while the College Life Study determined that up to a quarter of undergraduates used stimulants at least once during college.^1,2 Students may not know or may disregard the fact that prolonged abuse has resulted in serious health concerns, including cardiopulmonary issues and addiction. When these medications are taken incorrectly, especially in conjunction with alcohol, users risk seizures and death.³

In addition to stimulants, there are a variety of other prescriptions that have been shown to improve cognitive function. Amphetamines affect neurotransmitters in the brain to increase consciousness and adjust sleep patterns. They achieve this by preventing dopamine reuptake and disrupting normal vesicular packaging, which also increases dopamine concentration in the synaptic cleft through reverse transport from the cytosol.⁴ These drugs are currently used by the armed forces to mitigate pilots’ fatigue in high-intensity situations. While usage of these drugs may help regulate pilots’ energy levels, this unfortunately means that pilots face heavy pressure to take amphetamines despite the possibility of addiction and the lack of approval from the U.S. Food and Drug Administration.⁵

Besides prescription medications, various technological devices exist or are being created that affect cognition. For instance, transcranial direct current stimulation (tDCS) and transcranial magnetic stimulation (TMS) are devices currently marketed to enhance cognitive functioning through online websites and non-medical clinics, even though they have not yet received comprehensive clinical evaluations for this purpose.⁶ tDCS works by placing electrodes on the scalp to target specific brain areas. The machine sends a small direct current through electrodes to stimulate or inhibit neuronal activity. Similarly, TMS uses magnetic fields to alter neural activity. These methods have been shown to improve cognitive abilities including working memory, attention, language, and decision-making. Though these improvements are generally short-term, one University of Oxford study used tDCS to produce long-term improvements in mathematical abilities. Researchers taught subjects a new numerical system and then tested their ability to process and map the numbers into space. Subjects who received tDCS stimulation to the posterior parietal cortex displayed increased performance and consistency up to six to seven months after the treatment. This evidence indicates that tDCS can be used for the development of mathematical abilities as well as the treatment of degenerative neurological disorders such as Alzheimer’s.⁷

Regulation of cognitive enhancements is a multifaceted issue for which the risks and benefits of widespread usage must be intensively examined. According to one perspective, enhancements possess the ability to maximize human efficiency. If an enhancement can enable the acceleration of technological development and enable individuals to solve issues that affect society, it could improve the quality of life for users and non-users alike. This is why bans on anabolic steroids are not directly comparable to those on cognitive enhancements. While both medications share the goal of helping humans accomplish tasks beyond their natural capabilities, cognitive enhancements could accelerate technological and societal advancement. This would be more beneficial to society than one individual’s enhanced physical prowess.

While discussing this, it should be noted that such enhancements will not instantaneously bestow the user with Einsteinian intellectual capabilities. In a recent meta-analysis of 48 academic studies with 1,409 participants, prescription stimulants were found to improve delayed working memory but only had modest effects on inhibitory control and short-term episodic memory. The report also discussed how in some situations, other methods, including getting adequate sleep and using cognitive techniques like mnemonics, are far more beneficial than taking drugs such as methylphenidate and amphetamines. Biomedical enhancements, however, have broad effects that are applicable to many situations, while traditional cognitive techniques that don’t directly change the biology behind neural processes are task-specific and only rarely produce significant improvements.⁸

However, if we allow enhancement use to grow unchecked, an extreme possibility is the creation of a dystopian society led by only those wealthy enough to afford cognitive enhancements. Speculation about other negative societal effects is endless; for example, widespread use of cognitive enhancements could create a cutthroat work environment with constant pressure to use prescription pills or cranial stimulation, despite side effects and cost, in order to compete in the job market.

The possibility of addiction to cognitive enhancements and issues of social stratification based on access or cost should not be disregarded. However, there are many proposed solutions to these issues. Possible governmental regulation proposed by neuroethics researchers includes ensuring that cognitive enhancements are not readily available and are only given to those who demonstrate knowledge of the risks and responsible use of such enhancements. Additionally, the creation of a national database, similar to the current system used to regulate addictive pain relievers, would also help control the amount of medication prescribed to individuals. This database could be an integrated system that allows doctors to view patients’ other prescriptions, ensuring that those who attempt to deceive s pharmacies to obtain medications for personal abuse or illegal resale could not easily abuse the system. Finally, to address the issue of potential social inequality, researchers at Oxford University’s Future of Humanity Institute proposed a system in which the government could support broad development, competition, public understanding, a price ceiling, and even subsidized access for disadvantaged groups, leading to greater equalized access to cognitive enhancements.⁹

Advancements have made it possible to alter our minds using medical technology. Society requires balance to regulate these enhancements, an environment that will promote safe use while preventing abuse. The regulation of cognitive enhancement technologies should occur at several levels to be effective, from market approval to individual use. When creating these laws, research should not be limited because that could inhibit the discovery of possible cures to cognitive disorders. Instead, the neuroethics community should focus on safety and public usage regulations with the mission of preventing abuse and social stratification. Cognitive enhancements have the potential to affect the ways we learn, work, and live. However, specific regulations to address the risks and implications of this growing technology are required; otherwise the results could be devastating.

References

1. McCabe, S.E. et al. J. Psychoactive Drugs 2006, 38, 43-56.
2. Arria, A.M. et al. Subst. Abus. 2008, 29(4), 19-38.
3. Morton, W.A.; Stockton, G. J. Clin. Psychiatry 2000, 2(5), 159-164.
4. España, R.; Scammel, T. SLEEP 2011, 34(7), 845-858.
5. Rasmussen, N. Am. J. Public Health 2008, 98(6), 974-985.
6. Maslen, H. et al. J. of Law and Biosci. 2014, 1, 68-93.
7. Kadosh, R.C. et al. Curr. Biol. 2010, 20, 2016-2020.
8. Ilieva, I.P. et al. J. Cogn. Neurosci. 2015, 1069-1089.
9. Bostrom, N.; Sandberg, A. Sci. Eng. Ethics 2009, 15, 311-341.

Defect Patch: The Band-Aid for the Heart

Catalyst Volume 9

Imagine hearing that your newborn, only a few minutes out of womb, has a heart defect and will only live a couple more days. Shockingly, 1 in every 125 babies is born with some type of con-genital heart defect, drastically reducing his or her lifespan.¹ However, research institutes and hospitals nationwide are testing solutions and advanced devices to treat this condition. The most promising approach is the defect patch, in which scaffolds of tissue are engineered to mimic a healthy heart. The heart is enormously complex; mimicking is easier said than done. These patches require a tensile strength (for the heart’s pulses and variances) that is greater than that of the left ventricle of the human.¹ To add to the difficulty of creating such a device, layers of the patch have to be not only tense and strong, but also soft and supple, as cardiac cells prefer mal-leable tissue environments.

Researchers have taken on this challenge and, through testing various biomaterials, have de-termined the compatibility of each material within the patch. The materials are judged on the ba-sis of their biocompatibility, biodegradable nature, reabsorption, strength, and shapeability.² Natural possibilities include gelatin, chitosan, fibrin, and submucosa.¹ Though gelatin is easily biode-gradable, it has poor strength and lacks cell surface adhesion properties. Similarly, fibrin binds to different receptors, but with weak compression.³ On the artificial side, the polyglycolic acid (PGA) polymer, is strong and porous, while the poly lactic co-glycolic acid (PLGA) polymer has regulated biological properties, but poor cell attachment. This trade-off between different components of a good patch is what makes the building and modification of these systems so difficult. Nevertheless, the future of defect patches is extremely promising.

An unnatural polymer that is often used in creating patches is polycaprolactone, or PCL. This material is covered with gelatin-chitosan hydrogel to form a hydrophilic (water-conducive) patch.¹ In the process of making the patch, many different solutions of PCL matrices are pre-pared. The tension of the patch is measured to make sure that it will not rip or become damaged due to increased heart rate as the child develops. The force of the patch must always be greater than that of the left ventricle to ensure that the patch and the heart muscles do not rupture.¹ Although many considerations must be accounted for in making this artificial patch, the malleability and adhesive strength of the device are the most important.¹ Imagine a 12-year-old child with a defect patch implanted in the heart. Suppose this child attempts to do a cardio workout, including 100 jumping jacks, a few laps around a track, and some pushups. The heart patch must be able to reach the ultimate tensile strain under stress without detaching or bursting. The PCL core of the patch must also be able to handle large bursts of activity. Finally, the patch must be able to grow with the child and the heart must be able to grow new cells around the patch. In summary, the PCL patch must be biodegradable, have sufficient mechanical strength, and remain viable under harsh conditions.

While artificial materials like PCL are effective, in some situations, the aforementioned design criteria are best fulfilled by patches made from natural biomaterials. For instance, chitosan serves as a good template for the outside portion of the patch.⁴ This material is biocompatible, bioabsorbable, and shapeable. Using natural materials can reduce the risk of vascularization, or the abnormal formation of blood vessels. They can also adapt to gradual changes of the heart. Natural patches being developed and tested in Dr. Jeffrey Jacot’s lab at Rice University include a core of stem cells, which can differentiate into more specialized cells as the heart grows. They currently contain amniotic fluid-derived stem cells (AFSC) which must be isolated from hu-mans.⁵ Researchers prepare a layer of chitosan (or fibrin in some cases) and polyethylene gly-col hydrogels to compose the outside part of the patch.⁴ They then inject AFSC into this matrix to form the final patch. The efficiency of the patch is measured by recording the stem cells’ ability to transform into new cells. In experiments, AFSC are able to differentiate into virtually any cell type, and are particularly promising in regenerative medicine.⁵ These initial prototypes are still being developed and thoroughly tested on rodents.⁶ A major limitation of this approach is the ina-bility of patches to adapt in rapidly developing hearts, such as those of human infants and patch testing on humans or even larger mammals has yet to be done. The most important challenges for the future of defect patches are flexibility and adaptability.⁶ After all, this patch is essentially a transformed and repaired body part. Through the work of labs like Dr. Jacot’s, cardiac defects in infants and children may be completely treatable with a patch. Hopefully, in the future, babies with this “Band-Aid” may have more than a few weeks to live, if not an entire lifetime.

References

1. Pok, S., et al., ACS Nano. 2014, 9822–9832.
2. Pok, S., et al., Acta Biomaterialia. 5630–5642.
3. Pok, S., et al., Journal of Cardiovascular Translational Research J. of Cardiovasc. Trans. Res. 2011, 646–654.
4. Tottey, S., Johnson, et al. Biomaterials. 2011, 32(1), 128– 136.
5. Benavides, O. M., et al., Tissue Engineering Part A. 1185–1194.
6. Pok, S., et al., Tissue Engineering Part A. 1877–1887.

How Bionic Eyes Are Changing the Way We See the World

Catalyst Volume 9

Most blind people wear sunglasses, but what if their glasses could actually restore their vision? Such a feat seems miraculous, but the development of new bionic prostheses may make such miracles a reality. These devices work in two ways: by replacing non-functional parts of the visual pathway or by creating alternative neural avenues to provide vision.

When attempting to repair or restore lost vision, it is important to understand how we normally receive and process visual information. Light enters the eye and is refracted by the cornea to the lens, which focuses the light onto the retina. The cells of the retina, namely photoreceptors, convert the light into electrical impulses, which are transmitted to the primary visual cortex by the optic nerve. In short, this process serves to translate light energy into electrical energy that our brain can interpret. For patients suffering from impaired or lost vision, one of the steps in this process is either malfunctioning or not functioning at all.^1,2

Many patients with non-functional vision can be treated with current surgical techniques. For example, many elderly individuals develop cataracts, in which the lens of the eye becomes increasingly opaque, resulting in blurred vision. This condition can be rectified fairly simply with a surgical replacement of the lens. However, loss of vision resulting from a problem with the retina or optic nerve can very rarely be corrected surgically due to the sensitive nature of these tissues. Such pathologies include retinitis pigmentosa, an inherited degenerative disease affecting retinal photoreceptors, and head trauma, which can damage the optic nerve. In these cases, a visual prosthesis may be the solution. These devices, often called “bionic eyes,” are designed to repair or replace damaged ocular functions. Such prostheses restore vision by targeting damaged components in the retina, optic nerve, or the brain itself.

One set of visual prostheses works by correcting impaired retinal function via electrode arrays implanted between the retinal layers. The electrodes serve as substitutes for lost or damaged photoreceptors, translating light energy to electrical impulses. The Boston Retinal Implant Project has developed a device involving an eyeglass-mounted camera and an antenna implanted in the skin near the eye.³ The camera transmits visual data to the antenna in a manner reminiscent of a radio broadcast. Then, the antenna decodes the signal and then sends it through a wire to an implanted subretinal electrode array, which relays it to the brain. The problem with this system is that the camera is fully external and unrelated to the eye’s position, meaning the patient must move his or her entire head to survey a scene. Germany’s Retinal Implant AG team seeks to rectify this problem with the Alpha IMS implant system. In this system, the camera itself is subretinal, and “converts light in each pixel into electrical currents.”²

The Alpha IMS system is still undergoing experimental clinical trials in Europe, but it is facing some complications. Firstly, the visual clarity of tested patients is around 20/1000, which is well below the standard for legal blindness. Secondly, the system’s power supply is implanted in a very high-risk surgical procedure, which can endanger patients. In an attempt to overcome the problems faced by both The Boston Retinal Implant Project and Retinal Implant AG, Dr. Daniel Palanker at Stanford and his colleagues are currently developing a subretinal prosthesis involving a goggle-mounted video camera and an implanted photodiode array. The camera receives incoming light and projects the image onto the photodiode array, which then converts the light into pulsed electrical currents. These currents stimulate nearby neurons to relay the signal to the brain. As Dr. Palanker says, “This method for delivering information is completely wireless, and it preserves the natural link between ocular movement and image perception.”² Human clinical trials are slated to begin in 2016, but Palanker and his team are confident that the device will be able to produce 20/250 visual acuity or better in affected patients.

A potentially safer set of visual prostheses includes suprachoroidal implants. Very similar to the aforementioned subretinal implants, these devices also replace damaged components of the retina. The only difference is that suprachoroidal implants are placed between the choroid layer and the sclera, rather than between the retinal layers. This difference in location allows these devices to be surgically implanted with less risk, as they do not breach the retina itself. Furthermore, these devices are larger compared to subretinal implants, “allowing them to cover a wider visual field, ideal for navigation purposes.” Development of suprachoroidal devices began in the 1990s at both Osaka University in Japan and Seoul National University in South Korea. Dr. Lauren Ayton and Dr. David Nayagam of the Bionic Vision Australia (BVA) research partnership are heading more current research. BVA has tested a prototype of a suprachoroidal device in patients with retinitis pigmentosa, and results have been promising. Patients were able to “better localize light, recognize basic shapes, orient in a room, and walk through mobility mazes with reduced collisions.” More testing is planned for the future, along with improvements to the device’s design.²

Both subretinal and suprachoroidal implants work by replacing damaged photoreceptors, but they rely on a functional neural network between the retina and the optic nerve. Replacing damaged photoreceptors will not help a patient if he or she lacks the neural network that can transmit the signal to the brain. This neural network is composed of ganglion cells at the back of the retina that connect to the optic nerve; these ganglion cells can be viewed as the “output neurons of the eye.” A third type of visual prosthesis targets these ganglion cells. So-called epiretinal implants are placed in the final cell layer of the retina, with electrodes directly stimulating the optic nerve. Because these devices are implanted in the last retinal layer, they work “regardless of the state of the upstream neurons”.² So the main advantage of an epiretinal implant is that, in cases of widespread retinal damage due to severe retinitis pigmentosa, the device provides a shortcut directly to the optic nerve.

The most promising example of an epiretinal device is the Argus II Visual Prosthesis System, developed by Second Sight. The device, composed of a glasses-mounted camera that wirelessly transmits visual data to an implanted microelectrode array, received FDA marketing approval in 2012. Clinical trials have shown a substantial increase in visual perception and acuity in patients with severe retinitis pigmentosa, and the system has been implanted in more than 50 patients to date.

The common limitation of all these visual prostheses (subretinal, suprachoroidal, and epiretinal) is that they rely on an intact and functional optic nerve. But some blind patients have damaged optic nerves due to head trauma. The optic nerve connects the eye to the brain, so for patients with damage in this region, bionics researchers must find a way to target the brain itself. Experiments in the early 20th century showed that, by stimulating certain parts of the brain, blind patients could perceive light flashes known as phosphenes. Building from these experiments, modern scientists are working to develop cortical prostheses implanted in either the visual cortex of the cerebrum or the lateral geniculate nucleus (LGN), both of which are key in the brain’s ability to interpret visual information. Such a device would not truly restore natural vision, but produce artificial vision through the elicitation of phosphene patterns.

One group working to develop a cortical implant is the Monash Vision Group (MVG) in Melbourne, Australia, coordinated by Dr. Collette Mann and co. MVG’s Gennaris bionic-vision system consists of a glasses-mounted camera, a small computerized vision processor, and a series of multi-electrode tiles implanted in the visual cortex. The camera transmits images to the vision processor, which converts the picture into a waveform pattern and wirelessly transmits it to the multi-electrode tiles. Each electrode on each tile can generate a phosphene; all the electrodes working in unison can generate phosphene patterns. As Dr. Mann says, “The patterns of phosphenes will create 2-D outlines of relevant shapes in the central visual field.”² The Illinois Institute of Technology is developing a similar device called an intracortical visual prosthesis, termed the IIT ICVP. The device’s developers seek to address the substantial number of blind patients in underdeveloped countries by making the device more affordable. The institute says that “one potential advantage of the IIT ICVP system is its modularity,” and that by using fewer parts, they “could make the ICVP economically viable, worldwide.”⁴

These visual prostheses represent the culmination of decades of work by hundreds of researchers across the globe. They portray a remarkable level of collaboration between scientists, engineers, clinicians, and more, all for the purpose of restoring vision to those who live without it. And with an estimated 40 million individuals worldwide suffering from some form of blindness, these devices are making miracles reality.

References

1. The Scientist Staff. The Eye. The Scientist, 2014, 28.
2. Various Researchers. The Bionic Eye. The Scientist, 2014, 28.
3. Boston Retinal Implant Project. http://biomed.brown.edu/Courses/BI108/2006-108websites/group03retinalimplants/mit.htm (accessed Oct. 9, 2015)
4. Intracortical Visual Prosthesis. http://neural.iit.edu/research/icvp/ (accessed Oct. 10, 2015)

Nano-Materials with Giga Impact

Catalyst Volume 9

What material is so diverse that it has applications in everything from improving human lives to protecting the earth? Few materials are capable of both treating prolific diseases like diabetes and creating batteries that last orders of magnitude longer than industry standards. None are as thin, lightweight, and inexpensive as carbon nanotubes.

Carbon nanotubes are molecular cylinders made entirely of carbon atoms, which form a hollow tube just a few nanometers thick, as illustrated in Figure 1. For perspective, a nanometer is one ten-thousandth the width of a human hair.¹ The first multi-walled nanotubes (MWNTs) were discovered by L. V. Radushkevich and V. M. Lukyanovich of Russia in 1951.² Morinobu Endo first discovered single-walled nanotubes (SWNTs) in 1976, although the discovery is commonly attributed to Sumio Iijima at NEC of Japan in 1991.^3,4

Since their discovery, nanotubes have been the subject of extensive research by universities and national labs for the variety of applications in which they can be used. Carbon nanotubes have proven to be an amazing material, with properties that surpass those of existing alternatives such as platinum, stainless steel, and lithium-ion cathodes. Because of their unique structure, carbon nanotubes are revolutionizing the fields of energy, healthcare, and the environment.

Energy

One of the foremost applications of carbon nanotubes is in energy. Researchers at the Los Alamos National Laboratory have demonstrated that carbon nanotubes doped with nitrogen can be used to create a chemical catalyst. The process of doping involves substitution of one type of atom for another; in this case, carbon atoms were substituted with nitrogen. The synthesized catalyst can be used in lithium-air batteries which can hold a charge 10 times greater than that of a lithium-ion battery. A key parameter in the battery’s operation is the Oxygen Reduction Reaction (ORR) activity, which is a measure of a chemical species’ ability to gain electrons. The ORR activity of the nitrogen-doped material complex is not only the highest of any non-precious metal catalyst in alkaline media, but also higher than that of precious metals such as platinum.⁵

In another major development, Dr. James Tour of Rice University has created a graphene-carbon nanotube complex upon which a “forest” of vertical nanotubes can be grown. This base of graphene is a single, flat sheet of carbon atoms ‒ essentially a carbon nanotube “unrolled.” The ratio of height-to-base in this complex is equivalent to that of a house on a standard-sized plot of land extending into space.⁶ The graphene and nanotubes are joined at their interface by heptagonal carbon rings, allowing the structure to have an enormous surface area of 2000 m² per gram and serve as a high potential storage mechanism in fast supercapacitors.⁷

Healthcare

Carbon nanotubes also show immense promise in the field of healthcare. Take Michael Strano of MIT, who has developed a sensor composed of nanotubes embedded in an injectable gel that can detect several molecules. Notably, it can detect nitrous oxide, an indicator of inflammation, and blood glucose levels, which diabetics must continuously monitor. The sensors take advantage of carbon nanotubes’ natural fluorescent properties; when these tubes are complexed with a molecule that then binds to a specific target, their fluorescence will increase or decrease.⁸

Perhaps the most important potential application for carbon nanotubes in healthcare lies in their cancer-fighting applications. In the human cell, there is a family of genes called HER2 that is responsible for the regulation of growth and proliferation of cells. Normal cells have two copies of this family, but 20-25% of breast cancer cells have three or more copies, resulting in quickly-growing tumor cells. Approximately 40,000 U.S. women are diagnosed every year with this type of breast cancer. Fortunately, Huixin He of Rutgers University and Yan Xiao of the National Institute of Standards and Technology have found that they can attach an anti-HER2 antibody to carbon nanotubes to kill these cells, as shown in Figure 2. Once inserted into the body, a near-infrared light at a wavelength of 785 nm can be reflected off the antibody-nanotube complex, indicating where tumor cells are present. The wavelength then increases to 808 nm, at which point the nanotubes absorb the light and vibrate to release enough heat to kill any attached HER2 tumor cells. This process has shown a near 100% success rate and leaves normal cells unharmed.⁹

Environment

Carbon nanotube technology also has environmental applications. Hui Ying Yang from Singapore has developed a water-purification membrane made of plasma-treated carbon nanotubes which can be integrated into portable, rechargeable, and inexpensive purification devices the size of a teapot. These new purifications devices are ideal for developing countries and remote locations, where large industrial purification plants would be too energy- and labor-intensive. Unlike other portable devices, this rechargeable device utilizes a membrane system that does not require a continuous power source, does not rely on thermal processes or reverse osmosis, and can filter for organic contaminants found in brine water - the most common water supply in these developing and rural areas.¹⁰

Oil spills may no longer be such devastating natural disasters either. Bobby Sumpter of the Oak Ridge National Laboratory demonstrated that doping carbon nanotubes with boron atoms alters the curvature of the tubes. Forty-five degree angles form, leading to a sponge-like structure of nanotubes. As these tubes are made of carbon, they attract hydrocarbons and repel water due to their hydrophobic properties, allowing the tubes to absorb up to 100 times their weight in oil. Additionally, these tubes can be reused, as burning or squeezing them was shown to cause no damage. Sumpter and his team used an iron catalyst in the growth process of the carbon nanotubes, enabling a magnet to easily control or remove the tubes from an oil cleanup scenario.¹¹

Carbon nanotubes provide an incredible opportunity to impact areas of great importance to human life - energy, healthcare, and environmental protection. The results of carbon nanotube research in these areas demonstrate the remarkable properties of this versatile and effective material. Further studies may soon lead to their everyday appearance in our lives, whether in purifying water, fighting cancer, or even making the earth a better, cleaner place for everyone. Big impacts can certainly come in small packages.

References

1. Nanocyl. Carbon Nanotubes. http://www.nanocyl.com/CNT-Expertise-Centre/Carbon-Nanotubes (accessed Sep 12, 2015).
2. Monthioux, M.; Kuznetsov, V. Guest Editorial: Who should be given the credit for the discovery of carbon nanotubes? Carbon 44. [Online] 2006. 1621. http://nanotube.msu.edu/HSS/2006/1/2006-1.pdf (accessed Nov 15, 2015)
3. Ecklund, P.; et al. Ugliengo, P. In International Assessment of Carbon Nanotube Manufacturing and Applications, Proceedings of the World Technology Evaluation Center, Inc. Baltimore, MD, June, 2007.
4. Nanogloss. The History of Carbon Nanotubes – Who Invented the Nanotube? http://nanogloss.com/nanotubes/the-history-of-carbon-nanotubes-who-invented-the-nanotube/#axzz3mtharE9D (accessed Sep 14, 2015).
5. Understanding Nano. Economical non-precious-metal catalyst capitalizes on carbon nanotubes. http://www.understandingnano.com/catalyst-nitrogen-carbon-nanotubes.html (accessed Sep 17, 2015).
6. Understanding Nano. James’ bond: A graphene/nanotube hybrid. http://www.understandingnano.com/graphene-nanotube-electrode.html (accessed Sep 19, 2015).
7. Yan, Z. et al. ACS Nano. Toward the Synthesis of Wafer-Scale Single-Crystal Graphene on Copper Foils 2012, 6 (10), 9110–9117.
8. Understanding Nano. New implantable sensor paves way to long-term monitoring. http://www.understandingnano.com/carbon-nanotubes-implant-sensor.html (accessed Sep 20, 2015).
9. Understanding Nano. Combining Nanotubes and Antibodies for Breast Cancer 'Search and Destroy' Missions. http://www.understandingnano.com/nanomedicine-nanotubes-breast-cancer.html (accessed Sep 22, 2015).
10. Understanding Nano. Plasma-treated nano filters help purify world water supply. http://www.understandingnano.com/nanotube-membranes-water-purification.html (accessed Sep 24, 2015).
11. Sumpter, B. et al. Covalently bonded three-dimensional carbon nanotube solids via boron induced nanojunctions. Nature [Online] 2012, doi: 10.1038/srep00363. http://www.nature.com/articles/srep00363 (accessed Mar 06, 2016).
12.  Huixin, H. et al. Anti-HER2 IgY antibody-functionalized single-walled carbon nanotubes for detection and selective destruction of breast cancer cells. BMC Cancer 2009, 9, 351.   

3D Organ Printing: A Way to Liver a Little Longer

Catalyst Volume 9

On average, 22 people in America die each day because a vital organ is unavailable to them.¹ However, recent advances in 3D printing have made manufacturing organs feasible for combating the growing problem of organ donor shortages.

3D printing utilizes additive manufacturing, a process in which successive layers of material are laid down in order to make objects of various shapes and geometries.² It was first described in 1986, when Charles W. Hull introduced his method of ‘stereolithography,’ in which thin layers of materials were added by curing ultraviolet light lasers. In the past few decades, 3D printing has driven innovations in many areas, including engineering and art by allowing rapid prototyping of various structures.² Over time, scientists have further developed 3D printing to employ biological materials as a modeling medium. Early iterations of this process utilized a spotting system to deposit cells into organized 3D matrices, allowing the engineering of human tissues and organs. This method, known as 3D bioprinting, required layer-by-layer precision and the exact placement of 3D components. The ultimate goal of 3D biological modeling is to assemble human tissue and organs that have the correct biological and mechanical properties for proper functioning to be used for clinical transplantation. In order to achieve this goal, modern 3D organ printing is usually accomplished using either biomimicry, autonomous self-assembly, and mini-tissues. Typically, a combination of all three techniques is utilized to achieve bioprinting with multiple structural and functional properties.

The first approach, biomimicry, involves the manufacture of identical components of cells and tissues. The goal of this process is to use the cells and tissues of the organ recipient to duplicate the structure of organs and the environment in which they reside. Ongoing research in engineering, biophysics, cell biology, imaging, biomaterials, and medicine is very important for this approach to prosper, as a thorough understanding of the microenvironment of functional and supporting cell types is needed to assemble organs that can survive.³

3D bioprinting can also be accomplished through autonomous self-assembly, a technique that uses the same mechanisms as embryonic organ development. Developing tissues have cellular components that produce their own extracellular matrix in order to create the structures of the cell. Through this approach, researchers hope to utilize cells themselves to create fully functional organs. Cells are the driving force of this process, as they ultimately determine the functional and structural properties of the tissues.³

The final approach used in 3D bioprinting involves mini-tissues and combines the processes of both biomimicry and self-assembly. Mini-tissues are the smallest structural units of organs and tissues. They are replicated and assembled into macro-tissue through self-assembly. Using these smaller, potentially undamaged portions of the organs, fully functional organs can be made. This approach is similar to autonomous self-assembly in that the organs are created by the cells and tissues themselves.

As modern technology makes it possible, techniques for organ printing continue to advance. Although successful clinical implementation of printed organs is currently limited to flat organs such as skin and blood vessels and hollow organs such as the bladder,³ current research is ongoing for more complex organs such as the heart, pancreas, or kidneys.

Despite the recent advances in bioprinting, issues still remain. Since cell growth occurs in an artificial environment, it is hard to supply the oxygen and nutrients needed to keep larger organs alive. Additionally, moral and ethical debates surround the science of cloning and printing organs.³ Some camps assert that organ printing manipulates and interferes with nature. Others feel that, when done morally, 3D bioprinting of organs will benefit mankind and improve the lives of millions. In addition to these debates, there is also concern about who will control the production and quality of bioprinted organs. There must be some regulation of the production of organs, and it may be difficult to decide how to distribute this power. Finally, the potential expense of 3D printed organs may limit access to lower socioeconomic classes. 3D printed organs, at least in their early years, will more likely than be expensive to produce and to buy.

Nevertheless, there is widespread excitement surrounding the current uses of 3D bioprinting. While clinical trials may be in the distant future, organ printing can currently act as an in vitro model for drug toxicity, drug discovery, and human disease modeling.⁴ Additionally, organ printing has applications in surgery, as doctors may plan surgical procedures with a replica of a patient’s organ made with information from MRI and CT images. Future implementation of 3D printed organs can help train medical students and explain complicated procedures to patients. Additionally, 3D printed tissue of organs can be utilized to repair livers and other damaged organs. Bioprinting is still young, but its widespread application is quickly becoming a possibility. With further research, 3D printing has the potential to save the lives of millions in need of organ transplants.

References

1. U.S. Department of Health and Human Services. Health Resources and Services Information. http://www.organdonor.gov/about/data.html (accessed Sept. 15, 2015)
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