Use of X-Ray Fluorescence Microscopy for Studies on Research Models of Hepatocellular Carcinoma.

Introduction: TheraSphere® microspheres containing yttrium 90Y are among many radioembolization agents used clinically to reduce liver tumor burden, and their effects on cancer volume reduction are well-established. At the same time, concerns about off target tissue injury often limit their use. Deeper investigation into tissue distribution and long-term impact of these microspheres could inform us about additional ways to use them in practice. Methods: Healthy rat liver and rabbit liver tumor samples from animals treated with TheraSpheres were sectioned and their elemental maps were generated by X-ray fluorescence microscopy (XFM) at the Advanced Photon Source (APS) synchrotron at Argonne National Laboratory (ANL). Results: Elemental imaging allowed us to identify the presence and distribution of TheraSpheres in animal tissues without the need for additional sample manipulation or staining. Ionizing radiation produced by 90Y radioactive contaminants present in these microspheres makes processing TheraSphere treated samples complex. Accumulation of microspheres in macrophages was observed. Conclusions: This is the first study that used XFM to evaluate the location of microspheres and radionuclides in animal liver and tumor samples introduced through radioembolization. XFM has shown promise in expanding our understanding of radioembolization and could be used for investigation of human patient samples in the future.

Paramagnetic Mn8Fe4-co-Polystyrene Nanobeads as a Potential T1-T2 Multimodal Magnetic Resonance Imaging Contrast Agent with In Vivo Studies.

In developing a cluster-nanocarrier design, as a magnetic resonance imaging contrast agent, we have investigated the enhanced relaxivity of a manganese and iron-oxo cluster grafted within a porous polystyrene nanobead with increased relaxivity due to a higher surface area. The synthesis of the cluster-nanocarrier for the cluster Mn8Fe4O12(O2CC6H4CH═CH2)16(H2O)4, cross-linked with polystyrene (the nanocarrier), under miniemulsion conditions is described. By including a branched hydrophobe, iso-octane, the resulting nanobeads are porous and ∼70 nm in diameter. The increased surface area of the nanobeads compared to nonporous nanobeads leads to an enhancement in relaxivity; r1 increases from 3.8 to 5.2 ± 0.1 mM-1 s-1, and r2 increases from 11.9 to 50.1 ± 4.8 mM-1 s-1, at 9.4 teslas, strengthening the potential for T1 and T2 imaging. Several metrics were used to assess stability, and the porosity produced no reduction in metal stability. Synchrotron X-ray fluorescence microscopy was used to demonstrate that the nanobeads remain intact in vivo. In depth, physicochemical characteristics were determined, including extensive pharmacokinetics, in vivo imaging, and systemic biodistribution analysis.

Elemental Zn and its Binding Protein Zinc-α2-Glycoprotein are Elevated in HPV-Positive Oropharyngeal Squamous Cell Carcinoma.

Human papillomavirus (HPV)-positive head and neck squamous cell carcinoma (HNSCC) is biologically distinct from HPV-negative HNSCC. Outside of HPV-status, few tumor-intrinsic variables have been identified that correlate to improved survival. As part of exploratory analysis into the trace elemental composition of oropharyngeal squamous cell carcinoma (OPSCC), we performed elemental quanitification by X-ray fluorescence microscopy (XFM) on a small cohort (n = 32) of patients with HPV-positive and -negative OPSCC and identified in HPV-positive cases increased zinc (Zn) concentrations in tumor tissue relative to normal tissue. Subsequent immunohistochemistry of six Zn-binding proteins-zinc-α2-glycoprotein (AZGP1), Lipocalin-1, Albumin, S100A7, S100A8 and S100A9-revealed that only AZGP1 expression significantly correlated to HPV-status (p < 0.001) and was also increased in tumor relative to normal tissue from HPV-positive OPSCC tumor samples. AZGP1 protein expression in our cohort significantly correlated to a prolonged recurrence-free survival (p = 0.029), similar to HNSCC cases from the TCGA (n = 499), where highest AZGP1 mRNA levels correlated to improved overall survival (p = 0.023). By showing for the first time that HPV-positive OPSCC patients have increased intratumoral Zn levels and AZGP1 expression, we identify possible positive prognostic biomarkers in HNSCC as well as possible mechanisms of increased sensitivity to chemoradiation in HPV-positive OPSCC.

Gas-Induced Segregation in Pt-Rh Alloy Nanoparticles Observed by In Situ Bragg Coherent Diffraction Imaging.

Bimetallic catalysts can undergo segregation or redistribution of the metals driven by oxidizing and reducing environments. Bragg coherent diffraction imaging (BCDI) was used to relate displacement fields to compositional distributions in crystalline Pt-Rh alloy nanoparticles. Three-dimensional images of internal composition showed that the radial distribution of compositions reverses partially between the surface shell and the core when gas flow changes between O_{2} and H_{2}. Our observation suggests that the elemental segregation of nanoparticle catalysts should be highly active during heterogeneous catalysis and can be a controlling factor in synthesis of electrocatalysts. In addition, our study exemplifies applications of BCDI for in situ 3D imaging of internal equilibrium compositions in other bimetallic alloy nanoparticles.

Encapsulation, controlled release, and antitumor efficacy of cisplatin delivered in liposomes composed of sterol-modified phospholipids.

We employed a recently introduced class of sterol-modified lipids (SML) to produce m-PEG-DSPE containing liposome compositions with a range of cis-platinum content release rates. SML have a cholesterol succinate attached to the phosphatidylglycerol head group and a fatty acid at the 2 position. These compositions were compared to the well-studied liposome phospholipid compositions: mPEG-DSPE/Hydrogenated Soy PC/cholesterol or mPEG-DSPE/POPC/cholesterol to determine the effect of the cis-platinum release extent on C26 tumor proliferation in the BALB/c colon carcinoma mouse model. The release rates of cis-platinum from liposomes composed of SML are a function of the acyl chain length. SML-liposomes with shorter acyl chain lengths C-8 provided more rapid cisplatin release, lower in vitro IC50, and were easier to formulate compared to liposomes using traditional phospholipid compositions. Similar to other liposome cis-platinum formulations, the half-life of m-PEG-DSPE SML liposome cisplatin is substantially longer than the free drug. This resulted in a higher tumor cisplatin concentration at 48h post-dosing compared to the free drug and higher Pt-DNA adducts in the tumor. Moreover, the maximum tolerated dose of the liposome formulations where up to four fold greater than the free drug. Using X-ray fluorescence spectroscopy on tumor sections, we compared the location of platinum, to the location of a fluorescence lipid incorporated in the liposomes. The liposome platinum co-localized with the fluorescent lipid and both were non-uniformly distributed in the tumor. Non-encapsulated Cis-platinum, albeit at a low concentration, was more uniformly distributed thorough the tumor. Three liposome formulations, including the well-studied hydrogenated HSPC composition, had better antitumor activity in the murine colon 26 carcinoma model as compared to the free drug at the same dose but the SML liposome platinum formulations did not perform better than the HSPC formulation.

Microdistribution of lead in human teeth using microbeam synchrotron radiation X-ray fluorescence (µ-SRXRF).

Lead (Pb) exposure is known to be associated with adverse effects on human health, especially during the prenatal period and early childhood. The Pb content in teeth has been suggested as a useful biomarker for the evaluation of cumulative Pb exposure. This study was designed to employ the microbeam synchrotron radiation X-ray fluorescence technique to determine the microdistribution of Pb within the tooth to evaluate the reliability of the technique and the effectiveness of tooth Pb as a biomarker of Pb exposure. The results showed that in the incisor sample, Pb primarily deposited in secondary dentine region close to the pulp and secondarily at enamel exterior. In addition, Pb colocalised with Zn, indicating a positive correlation between Pb and Zn. By contrast,in the two molar samples, Pb accumulated principally in the pulp, and secondarily in the enamel. At the same time, Pb in these two molar samples colocalised with Ca instead of Zn as was observed in the incisor sample. Several batches of line scans further confirmed the conclusions. The feasibility of using microbeam synchrotron radiation X-ray fluorescence to determine the microdistribution of Pb in teeth and of using the tooth Pb, especially in dentine, as a biomarker was discussed.

Distribution of Iron Oxide Core-Titanium Dioxide Shell Nanoparticles in VX2 Tumor Bearing Rabbits Introduced by Two Different Delivery Modalities.

This work compares intravenous (IV) versus fluoroscopy-guided transarterial intra-catheter (IC) delivery of iron oxide core-titanium dioxide shell nanoparticles (NPs) in vivo in VX2 model of liver cancer in rabbits. NPs coated with glucose and decorated with a peptide sequence from cortactin were administered to animals with developed VX2 liver cancer. Two hours after NPs delivery tumors, normal liver, kidney, lung and spleen tissues were harvested and used for a series on histological and elemental analysis tests. Quantification of NPs in tissues was done both by bulk inductively coupled plasma mass spectrometry (ICP-MS) analysis and by hard X-ray fluorescence microscopy. Both IV and IC NPs injection are feasible modalities for delivering NPs to VX2 liver tumors with comparable tumor accumulation. It is possible that this is an outcome of the fact that VX2 tumors are highly vascularized and hemorrhagic, and therefore enhanced permeability and retention (EPR) plays the most significant role in accumulation of nanoparticles in tumor tissue. It is, however, interesting to note that IV delivery led to increased sequestration of NPs by spleen and normal liver tissue, while IC delivery lead to more NP positive Kupffer cells. This difference is most likely a direct outcome of blood flow dynamics. Armed with this knowledge about nanoparticle delivery, we plan to test them as radiosensitizers in the future.