Time-lapse imaging of the dynamics of CNS glial-axonal interactions in vitro and ex vivo.

PLoS One. 2012;7(1):e30775

Authors: Ioannidou K, Anderson KI, Strachan D, Edgar JM, Barnett SC

Abstract
BACKGROUND: Myelination is an exquisite and dynamic example of heterologous cell-cell interaction, which consists of the concentric wrapping of multiple layers of oligodendrocyte membrane around neuronal axons. Understanding the mechanism by which oligodendrocytes ensheath axons may bring us closer to designing strategies to promote remyelination in demyelinating diseases. The main aim of this study was to follow glial-axonal interactions over time both in vitro and ex vivo to visualize the various stages of myelination.
METHODOLOGY/PRINCIPAL FINDINGS: WE TOOK TWO APPROACHES TO FOLLOW MYELINATION OVER TIME: i) time-lapse imaging of mixed CNS myelinating cultures generated from mouse spinal cord to which exogenous GFP-labelled murine cells were added, and ii) ex vivo imaging of the spinal cord of shiverer (Mbp mutant) mice, transplanted with GFP-labelled murine neurospheres. We demonstrate that oligodendrocyte-axonal interactions are dynamic events with continuous retraction and extension of oligodendroglial processes. Using cytoplasmic and membrane-GFP labelled cells to examine different components of the myelin-like sheath, we provide evidence from time-lapse fluorescence microscopy and confocal microscopy that the oligodendrocytes' cytoplasm-filled processes initially spiral around the axon in a corkscrew-like manner. This is followed subsequently by focal expansion of the corkscrew process to form short cuffs, which then extend longitudinally along the axons. We predict from this model that these spiral cuffs must extend over each other first before extending to form internodes of myelin.
CONCLUSION: These experiments show the feasibility of visualizing the dynamics of glial-axonal interaction during myelination over time. Moreover, these approaches complement each other with the in vitro approach allowing visualization of an entire internodal length of myelin and the ex vivo approach validating the in vitro data.

PMID: 22303455 [PubMed - in process]

Establishment and characterization of a transgenic mouse model for in vivo imaging of Bmp4 expression in the pancreas.

PLoS One. 2011;6(9):e24956

Authors: Yasunaga M, Oumi N, Osaki M, Kazuki Y, Nakanishi T, Oshimura M, Sato K

Abstract
Type-2 diabetes results from the development of insulin resistance and a concomitant impairment of insulin secretion. Bone morphogenetic protein 4 (Bmp4)-Bmp receptor 1A signaling in β cells has recently been reported to be required for insulin production and secretion. In addition, Bmp4 blocks the differentiation and promotes the expansion of endocrine progenitor cells. Bmp4 therefore regulates the maintenance of homeostasis in the pancreas. In this study, we constructed a reporter plasmid carrying 7-kb enhancer and promoter region of the Bmp4 gene upstream of the firefly luciferase gene. We used this construct to produce transgenic mice by pro-nuclear microinjection, for subsequent in vivo monitoring of Bmp4 expression. The bioluminescent signal was detected mainly in the pancreas in three independent lines of transgenic mice. Furthermore, the bioluminescent signal was enhanced in association with the autophagy response to 24-h fasting. These results suggest that pancreatic expression of Bmp4 is involved in responding to the physiological environment, including through autophagy. These mouse models represent useful tools for toxicological screening, and for investigating the mechanisms responsible for pancreatic Bmp4 functions in vivo, with relevance to improving our understanding of pancreatic diseases.

PMID: 21949805 [PubMed - indexed for MEDLINE]

Monitoring the glioma tropism of bone marrow-derived progenitor cells by 2-photon laser scanning microscopy and positron emission tomography.

Neuro Oncol. 2012 Feb 1;

Authors: Hasenbach K, Wiehr S, Herrmann C, Mannheim J, Cay F, von Kürthy G, Bolmont T, Grathwohl SA, Weller M, Lengerke C, Pichler BJ, Tabatabai G

Abstract
Intracerebral experimental gliomas attract intravenously injected murine or human bone marrow-derived hematopoietic progenitor and stem cells (HPC) in vitro, ex vivo, and in vivo, indicating that these progenitor cells might be suitable vehicles for a cell-based delivery of therapeutic molecules to malignant gliomas. With regard to therapeutic application, it is important to investigate cell fates in vivo (i.e., the time-dependent intratumoral and systemic distribution after intravenously injection). Conventional histological analysis has limitations in this regard because longitudinal monitoring is precluded. Here, we used 2-photon laser scanning microscopy (2PLSM), positron emission tomography (PET), and MRI to study the fate of intravenously injected HPC carrying fluorescence, bioluminescence, and PET reporter genes in glioma-bearing mice. Our 2PLSM-based monitoring studies revealed that HPC homing to intracerebral experimental gliomas occurred already within the first 6 h and was most efficient within the first 24 h after intravenous injection. The highest PET signals were detected in intracerebral gliomas, whereas the tracer uptake in other organs, notably spleen, lung, liver, and muscle, remained at background levels. The results have important implications for designing schedules for therapeutic cell-based anti-glioma approaches. Moreover, the PET reporter-based imaging technique will allow noninvasive monitoring of cell fate in future cell-based therapeutic antiglioma approaches.

PMID: 22298526 [PubMed - as supplied by publisher]

Integrated multimodal endomicroscopy platform for simultaneous en face optical coherence and two-photon fluorescence imaging.

Opt Lett. 2012 Feb 1;37(3):362-4

Authors: Xi J, Chen Y, Zhang Y, Murari K, Li MJ, Li X

Abstract
We report an all-fiber-optic scanning, multimodal endomicroscope capable of simultaneous optical coherence tomography (OCT) and two-photon fluorescence (TPF) imaging. Both imaging modalities share the same miniature fiber-optic scanning endomicroscope, which consists of a double-clad fiber with a core operating in single mode at both the OCT (1310 nm) and two-photon excitation (1550 nm) wavelengths, a piezoelectric two-dimensional fiber-optic beam scanner, and a miniature aspherical compound lens suitable for simultaneous acquisition of en face OCT and TPF images. A fiber-optic wavelength division multiplexer was employed in the integrated platform to combine the low coherence OCT light source and the femtosecond two-photon excitation laser into the same optical path. Preliminary imaging results of cell cultures and mouse tissue ex vivo demonstrate the feasibility of simultaneous real-time OCT and TPF imaging in a scanning endomicroscopy setting for the first time.

PMID: 22297353 [PubMed - in process]

The energy demand of fast neuronal network oscillations: insights from brain slice preparations.

Front Pharmacol. 2011;2:90

Authors: Kann O

Abstract
Fast neuronal network oscillations in the gamma range (30-100 Hz) in the cerebral cortex have been implicated in higher cognitive functions such as sensual perception, working memory, and, perhaps, consciousness. However, little is known about the energy demand of gamma oscillations. This is mainly caused by technical limitations that are associated with simultaneous recordings of neuronal activity and energy metabolism in small neuronal networks and at the level of mitochondria in vivo. Thus recent studies have focused on brain slice preparations to address the energy demand of gamma oscillations in vitro. Here, reports will be summarized and discussed that combined electrophysiological recordings, oxygen sensor microelectrodes, and live-cell fluorescence imaging in acutely prepared slices and organotypic slice cultures of the hippocampus from both, mouse and rat. These reports consistently show that gamma oscillations can be reliably induced in hippocampal slice preparations by different pharmacological tools. They suggest that gamma oscillations are associated with high energy demand, requiring both rapid adaptation of oxidative energy metabolism and sufficient supply with oxygen and nutrients. These findings might help to explain the exceptional vulnerability of higher cognitive functions during pathological processes of the brain, such as circulatory disturbances, genetic mitochondrial diseases, and neurodegeneration.

PMID: 22291647 [PubMed - in process]

Fractionated photothermal antitumor therapy with multidye nanoparticles.

Int J Nanomedicine. 2012;7:351-7

Authors: Gutwein LG, Singh AK, Hahn MA, Rule MC, Knapik JA, Moudgil BM, Brown SC, Grobmyer SR

Abstract
PURPOSE: Photothermal therapy is an emerging cancer treatment paradigm which involves highly localized heating and killing of tumor cells, due to the presence of nanomaterials that can strongly absorb near-infrared (NIR) light. In addition to having deep penetration depths in tissue, NIR light is innocuous to normal cells. Little is known currently about the fate of nanomaterials post photothermal ablation and the implications thereof. The purpose of this investigation was to define the intratumoral fate of nanoparticles (NPs) after photothermal therapy in vivo and characterize the use of novel multidye theranostic NPs (MDT-NPs) for fractionated photothermal antitumor therapy.
METHODS: The photothermal and fluorescent properties of MDT-NPs were first characterized. To investigate the fate of nanomaterials following photothermal ablation in vivo, novel MDT-NPs and a murine mammary tumor model were used. Intratumoral injection of MDT-NPs and real-time fluorescence imaging before and after fractionated photothermal therapy was performed to study the intratumoral fate of MDT-NPs. Gross tumor and histological changes were made comparing MDT-NP treated and control tumor-bearing mice.
RESULTS: The dual dye-loaded mesoporous NPs (ie, MDT-NPs; circa 100 nm) retained both their NIR absorbing and NIR fluorescent capabilities after photoactivation. In vivo MDT-NPs remained localized in the intratumoral position after photothermal ablation. With fractionated photothermal therapy, there was significant treatment effect observed macroscopically (P = 0.026) in experimental tumor-bearing mice compared to control treated tumor-bearing mice.
CONCLUSION: Fractionated photothermal therapy for cancer represents a new therapeutic paradigm enabled by the application of novel functional nanomaterials. MDT-NPs may advance clinical treatment of cancer by enabling fractionated real-time image guided photothermal therapy.

PMID: 22287844 [PubMed - in process]

Cancer cell-selective in vivo near infrared photoimmunotherapy targeting specific membrane molecules.

Nat Med. 2011 Dec;17(12):1685-91

Authors: Mitsunaga M, Ogawa M, Kosaka N, Rosenblum LT, Choyke PL, Kobayashi H

Abstract
Three major modes of cancer therapy (surgery, radiation and chemotherapy) are the mainstay of modern oncologic therapy. To minimize the side effects of these therapies, molecular-targeted cancer therapies, including armed antibody therapy, have been developed with limited success. In this study, we have developed a new type of molecular-targeted cancer therapy, photoimmunotherapy (PIT), that uses a target-specific photosensitizer based on a near-infrared (NIR) phthalocyanine dye, IR700, conjugated to monoclonal antibodies (mAbs) targeting epidermal growth factor receptors. Cell death was induced immediately after irradiating mAb-IR700-bound target cells with NIR light. We observed in vivo tumor shrinkage after irradiation with NIR light in target cells expressing the epidermal growth factor receptor. The mAb-IR700 conjugates were most effective when bound to the cell membrane and produced no phototoxicity when not bound, suggesting a different mechanism for PIT as compared to conventional photodynamic therapies. Target-selective PIT enables treatment of cancer based on mAb binding to the cell membrane.

PMID: 22057348 [PubMed - indexed for MEDLINE]

High mobility group box 1 promotes endothelial cell angiogenic behavior in vitro and improves muscle perfusion in vivo in response to ischemic injury.

J Vasc Surg. 2012 Jan;55(1):180-91

Authors: Sachdev U, Cui X, Hong G, Namkoong S, Karlsson JM, Baty CJ, Tzeng E

Abstract
OBJECTIVES: The angiogenic drive in skeletal muscle ischemia remains poorly understood. Innate inflammatory pathways are activated during tissue injury and repair, suggesting that this highly conserved pathway may be involved in ischemia-induced angiogenesis. We hypothesize that one of the endogenous ligands for innate immune signaling, high mobility group box 1 (HMGB1), in combination with autophagic responses to hypoxia or nutrient deprivation, plays an important role in angiogenesis.
METHODS: Human dermal microvascular endothelial cells (ECs) were cultured in normoxia or hypoxia (1% oxygen). Immunocytochemical analysis of HMGB1 subcellular localization, evaluation of tube formation, and Western blot analysis of myotubule light-chain 3I (LC3I) conversion to LC3II, as a marker of autophagy, were conducted. 3-Methyladenine (3MA), chloroquine, or rapamycin were administered to inhibit or promote autophagy, respectively. In vivo, a murine hind limb ischemia model was performed. Muscle samples were collected at 4 hours to evaluate for nuclear HMGB1 and at 14 days to examine endothelial density. Perfusion recovery in the hind limbs was calculated by laser Doppler perfusion imaging (LDPI).
RESULTS: Hypoxic ECs exhibited reduced nuclear HMGB1 staining compared with normoxic cells (mean fluorescence intensity, 186.9 ± 17.1 vs 236.0 ± 1.6, P = .01) with a concomitant increase in cytosolic staining. HMGB1 treatment of ECs enhanced tube formation, an angiogenic phenotype of ECs. Neutralization of endogenous HMGB1 markedly impaired tube formation and inhibited LC3II formation. Inhibition of autophagy with 3MA or chloroquine abrogated tube formation, whereas its induction with rapamycin enhanced tubing and promoted HMGB1 translocation. In vivo, ischemic skeletal muscle showed reduced numbers of HMGB1-positive myocyte nuclei compared with nonischemic muscle (34.9% ± 1.9% vs 51.7% ± 2.0%, P < .001). Injection of HMGB1 into ischemic hind limbs increased perfusion recovery by 21% and increased EC density (49.2 ± 4.1 vs 34.2 ± 3.4 ECs/high-powered field, respectively; P = .02) at 14 days compared with control hind limbs.
CONCLUSIONS: Nuclear release of HMGB1 and autophagy occur in ECs in response to hypoxia or serum depletion. HMGB1 and autophagy are necessary and likely play an interdependent role in promoting the angiogenic behavior of ECs. In vivo, HMGB1 promotes perfusion recovery and increased EC density after ischemic injury. These findings suggest a possible mechanistic link between autophagy and HMGB1 in EC angiogenic behavior and support the importance of innate immune pathways in angiogenesis.

PMID: 21944908 [PubMed - indexed for MEDLINE]

A fluorescent chromophore TOTO-3 as a 'smart probe' for the assessment of ultrasound-mediated local drug delivery in vivo.

Contrast Media Mol Imaging. 2011 Jul-Aug;6(4):267-74

Authors: Deckers R, Yudina A, Cardoit LC, Moonen CT

Abstract
Many potent anti-cancer drugs have an intracellular mode of action, but are limited in crossing the cell membrane, resulting in a reduced clinical efficacy. Ultrasound (US) is known to facilitate the penetration of drugs into tumors cells. However (molecular) imaging techniques that monitor in vivo the underlying processes of US-triggered drug delivery are lacking. The objective of this study was to demonstrate the feasibility of using a fluorescent nuclear acid stain (TOTO-3) as a model drug to monitor in real-time US-mediated delivery by in vivo fluorescence imaging. Following co-injection of TOTO-3 and microbubbles US was applied to the tumor. The time course of the drug delivery process was monitored by fluorescence imaging. Immunohistological analysis and in vitro experiments were performed to investigate the results in more detail. A significant signal intensity enhancement of the US-treated tumor was observed that indicates intracellular delivery of the dye. In the control tumor TOTO-3 signal was strongly associated with macrophages, which was not the case for the sonicated tumor. The capability of macrophages to uptake TOTO-3 was confirmed in vitro. This study demonstrates that an optical contrast agent with similar characteristics to an anti-cancer drug may be used for continuous in vivo monitoring of the drug delivery process.

PMID: 21861287 [PubMed - indexed for MEDLINE]

Pharmacological characterization of a recombinant, fluorescent somatostatin receptor agonist.

Bioconjug Chem. 2011 Sep 21;22(9):1768-75

Authors: Sreenivasan VK, Stremovskiy OA, Kelf TA, Heblinski M, Goodchild AK, Connor M, Deyev SM, Zvyagin AV

Abstract
Somatostatin (SST) is a peptide neurotransmitter/hormone found in several mammalian tissue types. Apart from its natural importance, labeled SST/analogues are utilized in clinical applications such as targeting/diagnosis of neuroendocrine tumors. We report on the development and characterization of a novel, recombinant, fluorescent somatostatin analogue that has potential to elucidate somatostatin-activated cell signaling. SST was genetically fused with a monomeric-red fluorescent protein (mRFP) as the fluorescent label. The attachment of SST to mRFP had no detectable effect on its fluorescent properties. This analogue's potency to activate the endogenous and transfected somatostatin receptors was characterized using assays of membrane potential and Ca(2+) mobilization and immunocytochemistry. SST-mRFP was found to be an effective somatostatin receptor agonist, able to trigger the membrane hyperpolarization, mobilization of the intracellular Ca(2+) and receptor-ligand internalization in cells expressing somatostatin receptors. This complex represents a novel optical reporter due to its red emission spectral band suitable for in vivo imaging and tracking of the somatostatin receptor signaling pathways, affording higher resolution and sensitivity than those of the state-of-the-art radiolabeling bioassays.

PMID: 21823634 [PubMed - indexed for MEDLINE]

On the use of micron-sized iron oxide particles (MPIOS) to label resting monocytes in bone marrow.

Mol Imaging Biol. 2011 Oct;13(5):819-24

Authors: Tang KS, Hann B, Shapiro EM

Abstract
PURPOSE: The use of MRI to monitor immune cell infiltration into various pathologies is well established. In an effort to boost the magnetic material within immune cells, this work attempted to label resting monocytes within bone marrow, in mice, by intravenous administration of micron-sized iron oxide particles (MPIOs), similar in fashion to the administration of (U)SPIO.
PROCEDURES: MPIOs were incubated with various immune cells both in culture, and in whole blood. Flow cytometry and histology were used to analyze magnetic cell labeling. Also, MPIOs were injected intravenously into mice. In vivo, high-resolution 3-D MRI was performed on mouse legs, and signal changes were quantified. Flow cytometry and histology were used to analyze magnetic cell labeling of bone marrow resident cells.
RESULTS: It is demonstrated here that monocytes and neutrophils can indeed endocytose MPIOs both in cell culture and ex vivo in whole blood. However, despite rapid accumulation of MPIOs within the bone marrow following injection, MPIOs did not label monocytes or any other hematopoietic cell type in the marrow. Hypotheses are drawn to explain these results in light of recent usage of MPIOs for immune cell tracking.
CONCLUSIONS: Systemic administration of various MPIO formulations showed that MPIOs arrive in bone marrow rapidly following injection and remain there for at least 7 days. Data also shows slow clearance of some particles from the tissue over this period. While MPIOs can efficiently label monocytes in culture and in whole blood ex vivo, they were not found to label bone marrow resident monocytes.

PMID: 20936363 [PubMed - indexed for MEDLINE]

Imaging the bio-distribution of fluorescent probes using multispectral epi-illumination cryoslicing imaging.

Mol Imaging Biol. 2011 Oct;13(5):874-85

Authors: Sarantopoulos A, Themelis G, Ntziachristos V

Abstract
PURPOSE: The increasing availability of fluorescent probes for in vivo optical imaging enables the interrogation of complex biological processes in small animals serving as models for human-like tissue function and disease. However, the validation of probe bio-distribution during their development or the study of different disease models, in support of in vivo imaging studies, is not straightforward.
PROCEDURES: The imaging system developed consists of a customized multispectral planar imager that has been adapted on a commercial cryomicrotome and provides a powerful modality for ex vivo imaging of small animals.
RESULTS: The ability to capture 3D anatomical (color) and fluorescence volumetric distributions of multiple fluorescent markers in high resolution is showcased.
CONCLUSIONS: Serving both as a method for accurately imaging the bio-distribution of multiple fluorescent agents inside organisms and as a modality for the validation of non-invasive methods, multispectral cryoslicing imaging offers useful insights to ex vivo optical imaging of molecular probes.

PMID: 20838910 [PubMed - indexed for MEDLINE]

Multifunctional unimolecular micelles for cancer-targeted drug delivery and positron emission tomography imaging.

Biomaterials. 2012 Jan 24;

Authors: Xiao Y, Hong H, Javadi A, Engle JW, Xu W, Yang Y, Zhang Y, Barnhart TE, Cai W, Gong S

Abstract
A multifunctional unimolecular micelle made of a hyperbranched amphiphilic block copolymer was designed, synthesized, and characterized for cancer-targeted drug delivery and non-invasive positron emission tomography (PET) imaging in tumor-bearing mice. The hyperbranched amphiphilic block copolymer, Boltorn(®) H40-poly(L-glutamate-hydrazone-doxorubicin)-b-poly(ethylene glycol) (i.e., H40-P(LG-Hyd-DOX)-b-PEG), was conjugated with cyclo(Arg-Gly-Asp-D-Phe-Cys) peptides (cRGD, for integrin α(v)β(3) targeting) and macrocyclic chelators (1,4,7-triazacyclononane-N, N', N''-triacetic acid [NOTA], for (64)Cu-labeling and PET imaging) (i.e., H40-P(LG-Hyd-DOX)-b-PEG-OCH(3)/cRGD/NOTA, also referred to as H40-DOX-cRGD). The anti-cancer drug, doxorubicin (DOX) was covalently conjugated onto the hydrophobic segments of the amphiphilic block copolymer arms (i.e., PLG) via a pH-labile hydrazone linkage to enable pH-controlled drug release. The unimolecular micelles exhibited a uniform size distribution and pH-sensitive drug release behavior. cRGD-conjugated unimolecular micelles (i.e., H40-DOX-cRGD) exhibited a much higher cellular uptake in U87MG human glioblastoma cells due to integrin α(v)β(3)-mediated endocytosis than non-targeted unimolecular micelles (i.e., H40-DOX), thereby leading to a significantly higher cytotoxicity. In U87MG tumor-bearing mice, H40-DOX-cRGD-(64)Cu also exhibited a much higher level of tumor accumulation than H40-DOX-(64)Cu, measured by non-invasive PET imaging and confirmed by biodistribution studies and ex vivo fluorescence imaging. We believe that unimolecular micelles formed by hyperbranched amphiphilic block copolymers that synergistically integrate passive and active tumor-targeting abilities with pH-controlled drug release and PET imaging capabilities provide the basis for future cancer theranostics.

PMID: 22281424 [PubMed - as supplied by publisher]

The targeted delivery of anticancer drugs to brain glioma by PEGylated oxidized multi-walled carbon nanotubes modified with angiopep-2.

Biomaterials. 2012 Jan 24;

Authors: Ren J, Shen S, Wang D, Xi Z, Guo L, Pang Z, Qian Y, Sun X, Jiang X

Abstract
In this study, a dual-targeting drug delivery system based on PEGylated oxidized multi-walled carbon nanotubes (O-MWNTs) modified with angiopep-2 (O-MWNTs-PEG-ANG) was successfully developed for treatment of brain glioma. O-MWNTs can not only distribute in brains but also accumulate in tumors, and have ultrahigh surface area with remarkably high loading anticancer drug of doxorubicin (DOX), which was selected as drug carrier. Angiopep-2 can specifically combine to the low-density lipoprotein receptor-related protein (LRP) receptor overexpressed on the blood-brain barrier (BBB) and glioma cells, which was selected as targeting ligand. The cooperative dual-targeting to brain glioma by O-MWNTs-PEG-ANG was evaluated by intracellular tracking in vitro and fluorescence imaging in vivo, which demonstrated that the combination of O-MWNTs-PEG and angiopep-2 constituted an ideal dual-targeting drug delivery system. The anti-glioma effect of DOX-loaded O-MWNTs-PEG-ANG (DOX-O-MWNTs-PEG-ANG) was assessed by C6 cytotoxicity and median survival time of glioma bearing mice, which showed a better anti-glioma effect than DOX. The biological safety of O-MWNTs-PEG-ANG was evaluated by BCEC and C6 cytotoxicity, hematology analysis and CD68 immunohistochemical analysis, which proved O-MWNTs-PEG-ANG was good biocompatibility and low toxicity. The biological safety of DOX-O-MWNTs-PEG-ANG was evaluated by histopathological analysis, which suggested a lower cardiac toxicity than DOX. In conclusion, O-MWNTs-PEG-ANG is a promising dual-targeting carrier to deliver DOX for the treatment of brain tumor.

PMID: 22281423 [PubMed - as supplied by publisher]

Real-Time Monitoring of Glutathione-Triggered Thiopurine Anticancer Drug Release in Live Cells Investigated by Surface-Enhanced Raman Scattering.

Anal Chem. 2012 Jan 26;

Authors: Ock K, Jeon WI, Ganbold EO, Kim M, Park J, Seo JH, Cho K, Joo SW, Lee SY

Abstract
We investigated in vitro and in vivo glutathione (GSH)-induced intracellular thiopurine anticancer drug release on gold nanoparticle (Au NP) surfaces by means of label-free confocal Raman spectroscopy. Direct monitoring of GSH-triggered release of 6-mercaptopurine (6MP) and 6-thioguanine (6TG) was achieved in real time. Live cell imaging technique provides a nanomolar range release of 6MP and 6TG from Au NP surfaces after the injection of external GSH. In vivo SERS spectra of 6TG were obtained from the subcutaneous sites in living mice after GSH treatment. GSH-triggered releases of Cy5-dye assembled on 6TG-capped Au NPs were also compared using fluorescence measurements. Our work demonstrates that the time-lapse Raman spectroscopic tools are useful for monitoring of the controlled release of thiopurine drug molecules in vitro and in vivo.

PMID: 22280519 [PubMed - as supplied by publisher]

[Development of a new photosensitizer on the basis of ytterbium porphyrazine complex].

Biofizika. 2011 Nov-Dec;56(6):1117-24

Authors:

Abstract
The tetraphenyltetracyanoporphyrazine complex of ytterbium has been studied as a potential photosensitizer for fluorescence diagnostics and photodynamic therapy (PDT) of cancer. It has been shown that the new compound has an intensive absorption and fluorescence in the "tissue optical window". In particular, the absorption maximum of the complex is at the wavelength of 590 nm, and the fluorescence emission maximum is at 640 nm. A strong fluorescence enhancement with a 50-fold increase in the quantum yield has been revealed in blood serum. The experiments on human cancer cells line have demonstrated that the complex penetrates the cells in vitro and is located around the nuclei. The biodistribution and pharmacokinetics of the complex in animals have been investigated in vivo by a new method of transillumination fluorescence imaging using a peculiar setup. It has been found that the period of maximum uptake of the complex in mouse cervical carcinoma is from 3 to 6 h after i.v. injection, with the half-life in the tumor being 24 h. However, the selectivity of the complex in the tumor is not high enough. The time of clearance from the body is about 48 h. The area of the strongest fluorescence in the abdominal cavity in in vivo images is anatomically recognized as the intestine. This indicates that the new compounds undergo mainly the hepatic clearance mainly. The conventional methods ex vivo (confocal microscopy and point spectroscopic measurements) have detected the largest content of the complex in the intestine, liver, skin and tumor tissue. In general, the optical characteristics of the ytterbium porphyrazine complex as well as the features of its interaction with biological objects make it promising drug candidate for the photodynamic therapy and/or fluorescence diagnostics of cancer. However, a search for other novel formulations possessing a higher tumor selectivity remains an urgent problem.

PMID: 22279757 [PubMed - in process]

Pharmacokinetics and biodistribution of human serum albumin-TIMP-2 fusion protein using near-infrared optical imaging.

J Pharm Pharm Sci. 2011 Jul;14(3):368-77

Authors: Lee MS, Kim YH, Kim YJ, Kwon SH, Bang JK, Lee SM, Song YS, Hahm DH, Shim I, Han D, Her S

Abstract
PURPOSE: TIMP-2 has been studied as an attractive cancer therapeutic candidate, and a TIMP-2 fusion protein (HSA/TIMP-2) displayed effective anticancer activity, despite a lack of information about its pharmacokinetics (PK) and biodistribution. The purpose of this work was to assess the PK and biodistribution of HSA/TIMP-2 as well as to quantify accumulated HSA/TIMP-2 in tumors.
METHODS: Cy5.5 near-infrared (NIR) fluorescence was conjugated to the HSA/TIMP-2 protein (Cy5.5-HSA/TIMP-2) for monitoring spatio-temporal changes in vivo. For PK and biodistribution analysis, 0.2 μg/g body weight of Cy5.5-HSA/TIMP-2 was injected into MAT-LyLu prostate tumor xenografts, which were then imaged using an IVIS-200 optical imaging system. To quantify the accumulated HSA/TIMP-2 in tumors, we introduced a standard curve with depth-corrected fluorescence measurement.
RESULTS: In the vascular tube formation assay with human umbilical vein endothelial cells (HUVECs), Cy5.5-HSA/TIMP-2 showed an antiangiogenic effect. In prostate cancer xenografts, Cy5.5-HSA/TIMP-2 exhibited a prolongation of blood half-life to 19.6 h and relatively preferential distribution to the tumor. The amount of tumor-accumulated Cy5.5-HSA/TIMP-2 was calculated to be 4.5 ± 0.5 ng/g body weight at 2 days, representing 2.25 ± 0.25% of the initial dose.
CONCLUSIONS: We evaluated the pharmacokinetic profile and biodistribution of HSA/TIMP-2 with favorable results, providing new information for more effective approaches to cancer therapeutics using HSA/TIMP-2. Additionally, real-time in vivo fluorescence imaging analysis using a depth-corrected standard curve may serve as a platform to quantify biodistributed drug in anticancer therapeutic studies.

PMID: 21962154 [PubMed - indexed for MEDLINE]

Investigation of glucose-modified liposomes using polyethylene glycols with different chain lengths as the linkers for brain targeting.

Int J Nanomedicine. 2012;7:163-75

Authors: Xie F, Yao N, Qin Y, Zhang Q, Chen H, Yuan M, Tang J, Li X, Fan W, Zhang Q, Wu Y, Hai L, He Q

Abstract
BACKGROUND: An intimidating challenge to transporting drugs into the brain parenchyma is the presence of the blood-brain barrier (BBB). Glucose is an essential nutritional substance for brain function sustenance, which cannot be synthesized by the brain. Its transport primarily depends on the glucose transporters on the brain capillary endothelial cells. In this paper, the brain-targeted properties of glucose-modified liposomes using polyethylene glycols with different chain lengths as the linkers were compared and evaluated to establish an optimized drug-delivery system.
METHODS: Coumarin 6-loaded liposomes (GLU200-LIP, GLU400-LIP, GLU1000-LIP, and GLU2000-LIP) composed of phospholipids and glucose-derived cholesterols were prepared by thin-film dispersion-ultrasound method. The BBB model in vitro was developed to evaluate the transendothelial ability of the different liposomes crossing the BBB. The biodistribution of liposomes in the mice brains was identified by in vivo and ex vivo nearinfrared fluorescence imaging and confocal laser scanning microscopy and further analyzed quantitatively by high-performance liquid chromatography.
RESULTS: Glucose-derived cholesterols were synthesized and identified, and coumarin 6-loaded liposomes were prepared successfully. The particle sizes of the four types of glucose-modified liposomes were around or smaller than 100 nm with a polydispersity index less than 0.300. GLU400-LIP, GLU1000-LIP, and GLU2000-LIP achieved higher cumulative cleared volumes on BBB model in vitro after 6 hours compared with GLU200-LIP (P < 0.05) and were significantly higher than that of the conventional liposome (P < 0.001). The qualitative and quantitative biodistribution results in the mice showed that the accumulation of GLU1000-LIP in the brain was the highest among all the groups (P < 0.01 versus LIP).
CONCLUSION: The data indicated that GLU400-LIP, GLU1000-LIP, and GLU2000-LIP all possess the potential of brain targeting, among which GLU1000-LIP, as a promising drug-delivery system, exhibited the strongest brain delivery capacity.

PMID: 22275832 [PubMed - in process]

In vivo NIR imaging with PbS quantum dots entrapped in biodegradable micelles.

J Biomed Mater Res A. 2012 Jan 24;

Authors: Cao J, Zhu H, Deng D, Xue B, Tang L, Mahounga D, Qian Z, Gu Y

Abstract
In this article, we firstly synthesized oil-soluble PbS quantum dots (QDs) emitting in the near-infrared (NIR) spectral range through a two-phase method, which exhibit a conveniently tunable photoluminescence (PL) emission (from ∼750 to 872 nm) with a narrow PL bandwidth, as well as a high (up to 40%) PL quantum yield (QY). Next, the as-prepared oil-soluble NIR PbS QDs were applied to the in vivo imaging of tumors by entrapping in biodegradable micelles (N-succinyl-N'-octyl nanomicelles, SOC) which had hydrophobic inner cores. Transmission electron microscope results show well dispersed spherical shaped QDs-loaded SOC micelles with 100 nm diameter. The QY of PbS QDs entrapped into SOC has no significant change compared to free QDs. Besides, both in vitro and in vivo acute toxicity results demonstrated that the QDs-loaded micelles have low cytotoxicity. Furthermore, in vivo imaging of PbS QDs-loaded SOC injected intravenously into tumor-bearing nude mice showed the NIR QDs-loaded micelles can accumulate in the tumor tissue due to the enhanced permeability and retention effect of SOC micelles. These results confirm that the as-prepared PbS QDs could be used as fluorescence probes to study the biodistribution of nanocarriers and their intracellular pathways, as well as their passive targeted behavior to tumors in preclinical research. © 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part A:, 2012.

PMID: 22275223 [PubMed - as supplied by publisher]

Prediction of proapoptotic anticancer therapeutic response in vivo based on cell death visualization and TRAIL death ligand-receptor interaction.

Cancer Biol Ther. 2011 Aug 15;12(4):335-48

Authors: Zhou L, Wang W, Dicker DT, Humphreys RC, El-Deiry WS

Abstract
Tumor growth is often associated with insufficient apoptosis. The Tumor Necrosis Factor (TNF)-Related Apoptosis-Inducing Ligand (TRAIL) and its proapoptotic receptors death receptor 4 (DR4) and DR5 agonistic monoclonal antibodies are being developed as targeted therapeutics because they kill cancer cells while sparing normal cells. A challenge to targeted therapeutics is the selection of patients who are most likely to benefit from targeted drugs because of the heterogeneity of cancer. Molecular imaging may be useful in targeted drug development by assessing the target expression and drug-target interaction, and for predicting therapeutic response. We hypothesized that the cell surface expression level of DR4/5 may predict the proapoptotic targeted therapeutic response if the signaling pathway downstream is intact. The goal of this proof-of-concept study was to develop a molecular imaging strategy to predict proapoptotic anti-cancer therapy response at an early stage of treatment. TRAIL and the DR5 agonistic monoclonal antibody HGS-ETR2 (Lexatumumab, TRM-2) were labeled with a near-infrared dye and these were used to image the TRAIL receptors on cultured TRAIL sensitive and TRAIL resistant human tumor cells as well as tumor xenografts. Imaging of cells and tumor-bearing animals was conducted with near infrared fluorescence imagers and apoptosis in cells was assessed by western blots of PARP-cleavage and flow cytometry of sub-G1 content. Apoptosis in tumors was evaluated by imaging near-infrared dye-labeled Annexin V and tumor tissue activated caspase-3 staining. Both in vitro and in vivo studies showed that imaging of death inducing ligand-receptor interaction was consistent with the apoptosis readout. Thus TRAIL sensitive tumors that express TRAIL receptors underwent cell death following treatment whereas tumors lacking TRAIL receptor expression were shown to be TRAIL resistant. In vivo molecular imaging of TRAIL receptor expression correlated with response to TRAIL therapy and an apoptotic response in vivo.

PMID: 21785270 [PubMed - indexed for MEDLINE]