Feifan Zhou

Cancer photothermal therapy in the near-infrared region by using single-walled carbon nanotubes.

Single-walled carbon nanotubes (SWNTs) have a high optical absorbance in the near-infrared (NIR) region. In this special optical window, biological systems are known to be highly transparent. The optical properties of SWNTs provide an opportunity for selective photothermal therapy for cancer treatment. Specifically, CoMoCAT nanotubes with a uniform size (about 0.81 nm) and a narrow absorption peak at 980 nm are ideal candidates for such a novel approach. Here, CoMoCAT SWNTs are conjugated to folate, which can bind specifically to the surface of the folate receptor tumor markers. Folate-SWNT (FA-SWNT) targeted tumor cells were irradiated by a 980-nm laser. In our in vitro and in vivo experiments, FA-SWNT effectively enhanced the photothermal destruction on tumor cells and noticeably spared the photothermal destruction for nontargeted normal cells. Thus, SWNTs, combined with suitable tumor markers, can be used as novel nanomaterials for selective photothermal therapy for cancer treatment.

Mitochondrial oxidative stress causes mitochondrial fragmentation via differential modulation of mitochondrial fission–fusion proteins

Mitochondria are dynamic organelles that undergo continual fusion and fission to maintain their morphology and functions, but the mechanism involved is still not clear. Here, we investigated the effect of mitochondrial oxidative stress triggered by high‐fluence low‐power laser irradiation (HF‐LPLI) on mitochondrial dynamics in human lung adenocarcinoma cells (ASTC‐a‐1) and African green monkey SV40‐transformed kidney fibroblast cells (COS‐7). Upon HF‐LPLI‐triggered oxidative stress, mitochondria displayed a fragmented structure, which was abolished by exposure to dehydroascorbic acid, a reactive oxygen species scavenger, indicating that oxidative stress can induce mitochondrial fragmentation. Further study revealed that HF‐LPLI caused mitochondrial fragmentation by inhibiting fusion and enhancing fission. Mitochondrial translocation of the profission protein dynamin‐related protein 1 (Drp1) was observed following HF‐LPLI, demonstrating apoptosis‐related activation of Drp1. Notably, overexpression of Drp1 increased mitochondrial fragmentation and promoted HF‐LPLI‐induced apoptosis through promoting cytochrome c release and caspase‐9 activation, whereas overexpression of mitofusin 2 (Mfn2), a profusion protein, caused the opposite effects. Also, neither Drp1 overexpression nor Mfn2 overexpression affected mitochondrial reactive oxygen species generation, mitochondrial depolarization, or Bax activation. We conclude that mitochondrial oxidative stress mediated through Drp1 and Mfn2 causes an imbalance in mitochondrial fission–fusion, resulting in mitochondrial fragmentation, which contributes to mitochondrial and cell dysfunction.

Indocyanine Green-Containing Nanostructure as Near Infrared Dual-Functional Targeting Probes for Optical Imaging and Photothermal Therapy

Indocyanine green (ICG) is a near-infrared (NIR) imaging agent and is also an ideal light absorber for laser-mediated photothermal therapy. This NIR dye could serve as a basis of a dual-functional probe with integrated optical imaging and photothermal therapy capabilities. However, applications of ICG remain limited by its concentration-dependent aggregation, poor aqueous stability, nonspecific binding to proteins and lack of target specificity. To overcome these limitations, a novel ICG-containing nanostructure is designed utilizing the noncovalent self-assembly chemistry between phospholipid-polyethylene glycol (PL-PEG) and ICG. The interactions between both amphiphilic ICG and PL-PEG were studied using absorption and fluorescence spectroscopy. The properties of ICG-PL-PEG nanoprobe, such as absorption and fluorescence spectra, stability, morphology and size distribution, were also investigated. Two representative targeting molecules, namely, a small molecule, folic acid (FA), and a large protein, integrin α(v)β₃ monoclonal antibody (mAb), were conjugated to the surface of ICG-PL-PEG nanoprobe, displaying the diversity of ligand conjugation. The target specificity was confirmed using three cell lines with different levels of available folate receptors (FRs) or integrin α(v)β₃ expression via laser scanning confocal microscope and flow cytometry. This targeting ICG-PL-PEG nanoprobe could be internalized into targeted cells via ligand-receptor mediated endocytosis pathway. Our in vitro experiments showed that internalized ICG-PL-PEG could be used for cell imaging and selective photothermal cell destruction. These results represent the first demonstration of the dual functionality of ICG-containing nanostructure for targeted optical imaging and photothermal therapy of cancerous cells. This novel ICG-PL-PEG nanostructure, when conjugated with other therapeutic and imaging agents, could become a multifunctional probe for cancer diagnosis and treatment.

Enhanced tumor treatment using biofunctional indocyanine green-containing nanostructure by intratumoral or intravenous injection

Indocyanine green (ICG) is a conventional dye that can be used in clinical near-infrared (NIR) imaging, and it is also an effective light absorber for laser-mediated photothermal therapy. However, applications of ICG were limited due to its fast degradation in aqueous media and quick clearance from the body. Herein, an ICG-containing nanostructure, ICG-PL-PEG, was developed for photothermal therapy, which was self-assembled by ICG and phospholipid-polyethylene glycol (PL-PEG). Our in vitro and in vivo experiments demonstrated that ICG-PL-PEG suspension was more efficient in producing a NIR-dependent temperature increase than ICG alone, due to the increase of ICG monomers from the addition of PL-PEG to match the central wavelength of the 808 nm laser. When conjugated with integrin α(v)β(3) monoclonal antibody (mAb), ICG-PL-PEG could be selectively internalized and retained in target tumor cells. Irradiation of an 808 nm laser after intravenous administration of ICG-PL-PEG-mAb resulted in tumor suppression in mice, while ICG alone had only limited effect. This is the first time an ICG-containing nanostructure has been used through systemic administration to achieve an efficient in vivo photothermal effect for cancer treatment. Therefore, ICG-PL-PEG could be used as a fluorescent marker as well as a light-absorber for imaging-guided photothermal therapy. All the components of ICG-PL-PEG have been approved for human use. Therefore, this unique ICG-containing nanostructure has great potential in clinical applications.

New insights of transmembranal mechanism and subcellular localization of noncovalently modified single-walled carbon nanotubes.

Translocation and localization of single-walled carbon nanotubes (SWNTs) in normal and cancerous cells have significant biomedical implications. In this study, SWNTs functionalized with different biomolecules in cells were observed with confocal laser scanning microscopy. Functionalized with PL-PEG, SWNTs were found to localize exclusively in mitochondria of both tumor and normal cells due to mitochondrial transmembrane potential, but they were found mainly in lysosomes of macrophages due to phagocytosis. However, when conjugated with different molecules, the subcellular localization of the surface-modified SWNT-PL-PEG depended on how SWNTs enter the cells: inside mitochondria if crossing cell membrane or inside lysosomes if being endocytosized. We also show that mitochondrial SWNT-PL-PEG, when irradiated with a near-infrared light, can induce cell apoptosis due to mitochondrial damages. These findings provide a better mechanistic understanding of cellular localization of SWNTs, which could lead to advanced biomedical applications such as the design of molecular transporters and development of SWNT-assisted cancer therapies.

Mitochondria-Targeting Single-Walled Carbon Nanotubes for Cancer Photothermal Therapy

Nanomaterials have recently attracted much attention as efficient transducers for cancer photothermal therapy, based on their intrinsic absorption properties in the near-infrared region. This study explores a novel therapy model with mitochondria-targeting single-walled carbon nanotubes (SWNTs), which act efficiently to convert 980-nm laser energy into heat and selectively destroy the target mitochondria, thereby inducing mitochondrial depolarization, cytochrome c release, and caspase 3 activation. The laser+SWNTs process affords remarkable efficacy in suppressing tumor growth in a breast cancer model, and results in complete tumor regression in some cases. Laser+SWNTs could prove to be a promising selective local treatment modality, while minimizing adverse side effects.

Antitumor immunologically modified carbon nanotubes for photothermal therapy

An immunologically modified nanotube system was developed using an immunoadjuvant, glycated chitosan (GC), as surfactant of single-walled carbon nanotube (SWNTs). This SWNT-GC system not only retained both optical properties of SWNTs and immunological functions of GC, but also could enter cells due to the carrier properties of SWNTs. Cellular SWNTs induced thermal destruction of tumor cells when irradiated by a near-infrared laser and, at the same time, cellular GC could serve both as damage associated molecular pattern molecules (DAMPs) and pathogen associated molecular pattern molecules (PAMPs) to enhance the tumor immunogenicity and enhance the uptake and presentation of tumor antigens, leading to special antitumor response. Using this system and a 980 nm laser, we treated tumors, both in vitro and in vivo, and investigated the induced thermal and immunological effects. Laser + SWNT-GC afford a remarkable efficacy in suppressing tumor growth in animal cancer models, in many cases resulting in complete tumor regression and long-term survival. Mice successfully treated by Laser + SWNT-GC could establish resistance to tumor rechallenge. This system forms a multifunctional temporal-spatial continuum, which can synergize photothermal and immunological effects. The Laser + SWNT-GC could represent a promising treatment modality to induce systemic antitumor response through a local intervention, while minimizing the adverse side effects.

Functional single-walled carbon nanotubes based on an integrin αvβ3 monoclonal antibody for highly efficient cancer cell targeting

The application of single-walled carbon nanotubes (SWNTs) in the field of biomedicine is becoming an entirely new and exciting topic. In this study, a novel functional SWNT based on an integrin alpha(v)beta(3) monoclonal antibody was developed and was used for cancer cell targeting in vitro. SWNTs were first modified by phospholipid-bearing polyethylene glycol (PL-PEG). The PL-PEG functionalized SWNTs were then conjugated with protein A. A SWNT-integrin alpha(v)beta(3) monoclonal antibody system (SWNT-PEG-mAb) was thus constructed by conjugating protein A with the fluorescein labeled integrin alpha(v)beta(3) monoclonal antibody. In vitro study revealed that SWNT-PEG-mAb presented a high targeting efficiency on integrin alpha(v)beta(3)-positive U87MG cells with low cellular toxicity, while for integrin alpha(v)beta(3)-negative MCF-7 cells, the system had a low targeting efficiency, indicating that the high targeting to U87MG cells was due to the specific integrin targeting of the monoclonal antibody. In conclusion, SWNT-PEG-mAb developed in this research is a potential candidate for cancer imaging and drug delivery in cancer targeting therapy.

Low-level laser therapy regulates microglial function through Src-mediated signaling pathways: implications for neurodegenerative diseases.

BackgroundActivated microglial cells are an important pathological component in brains of patients with neurodegenerative diseases. The purpose of this study was to investigate the effect of He-Ne (632.8 nm, 64.6 mW/cm2) low-level laser therapy (LLLT), a non-damaging physical therapy, on activated microglia, and the subsequent signaling events of LLLT-induced neuroprotective effects and phagocytic responses.MethodsTo model microglial activation, we treated the microglial BV2 cells with lipopolysaccharide (LPS). For the LLLT-induced neuroprotective study, neuronal cells with activated microglial cells in a Transwell™ cell-culture system were used. For the phagocytosis study, fluorescence-labeled microspheres were added into the treated microglial cells to confirm the role of LLLT.ResultsOur results showed that LLLT (20 J/cm2) could attenuate toll-like receptor (TLR)-mediated proinflammatory responses in microglia, characterized by down-regulation of proinflammatory cytokine expression and nitric oxide (NO) production. LLLT-triggered TLR signaling inhibition was achieved by activating tyrosine kinases Src and Syk, which led to MyD88 tyrosine phosphorylation, thus impairing MyD88-dependent proinflammatory signaling cascade. In addition, we found that Src activation could enhance Rac1 activity and F-actin accumulation that typify microglial phagocytic activity. We also found that Src/PI3K/Akt inhibitors prevented LLLT-stimulated Akt (Ser473 and Thr308) phosphorylation and blocked Rac1 activity and actin-based microglial phagocytosis, indicating the activation of Src/PI3K/Akt/Rac1 signaling pathway.ConclusionsThe present study underlines the importance of Src in suppressing inflammation and enhancing microglial phagocytic function in activated microglia during LLLT stimulation. We have identified a new and important neuroprotective signaling pathway that consists of regulation of microglial phagocytosis and inflammation under LLLT treatment. Our research may provide a feasible therapeutic approach to control the progression of neurodegenerative diseases.

Glycated chitosan as a new non-toxic immunological stimulant

Chitosan is capable of stimulating immune responses. However, because chitosan is not water soluble, it has limited biological applications. By attaching galactose molecules to the chitosan molecules, a new water-soluble compound, glycated chitosan (GC), was synthesized. GC was designed for immune stimulations in combination with phototherapies in the treatment of metastatic tumors. To investigate the possible toxicity of GC, cultures of normal and tumor cells were incubated with GC of different concentrations and the cell viabilities were determined. For in vivo studies, GC solution was fed or injected to animals and its toxicity was determined through observations of animals and histological examinations of vital organs. No toxic effects of GC were observed in cultured cells or in animal studies. In addition, the immunological effect of GC was investigated through its stimulation of TNFα secretion by macrophages in vitro. In vivo studies showed enhancement of the survival of laser immunotherapy-treated rats bearing metastatic mammary tumors. Our in vitro and in vivo results indicated that GC was a strong immunological stimulant. Its non-toxic nature and immunological activity make GC a potential immunoadjuvant for treatment of metastatic tumors.