Sonodynamic therapy in combination with photodynamic therapy shows enhanced long-term cure of brain tumor.

This article presents the construction of a multimodality platform that can be used for efficient destruction of brain tumor by a combination of photodynamic and sonodynamic therapy. For in vivo studies, U87 patient-derived xenograft tumors were implanted subcutaneously in SCID mice. For the first time, it has been shown that the cell-death mechanism by both treatment modalities follows two different pathways. For example, exposing the U87 cells after 24 h incubation with HPPH [3-(1'-hexyloxy)ethyl-3-devinyl-pyropheophorbide-a) by ultrasound participate in an electron-transfer process with the surrounding biological substrates to form radicals and radical ions (Type I reaction); whereas in photodynamic therapy, the tumor destruction is mainly caused by highly reactive singlet oxygen (Type II reaction). The combination of photodynamic therapy and sonodynamic therapy both in vitro and in vivo have shown an improved cell kill/tumor response, that could be attributed to an additive and/or synergetic effect(s). Our results also indicate that the delivery of the HPPH to tumors can further be enhanced by using cationic polyacrylamide nanoparticles as a delivery vehicle. Exposing the nano-formulation with ultrasound also triggered the release of photosensitizer. The combination of photodynamic therapy and sonodynamic therapy strongly affects tumor vasculature as determined by dynamic contrast enhanced imaging using HSA-Gd(III)DTPA.

Targeted Nanoparticles for Fluorescence Imaging of Folate Receptor Positive Tumors.

This report presents the synthesis and folate receptor target-specificity of amino-functionalized polyacrylamide nanoparticles (AFPAA NPs) for near-infrared (NIR) fluorescence imaging of cancer. For the synthesis of desired nano-constructs, the AFPAA NPs (hereafter referred to as NPs) were reacted with a NIR cyanine dye (CD) bearing carboxylic acid functionality by following our previously reported approach, and the resulting conjugate (NP-CD) on further reaction with folic acid (FA) resulted in a new nano-construct, FA-NP-CD, which demonstrated significantly higher uptake in folate receptor-positive breast cancer cells (KB+) and in folate receptor over-expressed tumors in vivo. The target-specificity of these nanoparticles was further confirmed by inhibition assay in folate receptor-positive (KB+) and -negative (HT-1080) cell lines. To show the advantages of polyacrylamide (PAA)-based NPs in folate receptor target-specificity, the CD used in preparing the FA-NP-CD construct was also reacted with folic acid alone and the synthetic conjugate (CD-FA) was also investigated for its target-specificity. Interestingly, in contrast to NPs (FA-NP-CD), the CD-FA conjugate did not show any significant in vitro or in vivo specificity toward folate receptors, showing the advantages of PAA-based nanotechnology in delivering the desired agent to tumor cells.

Pluronic F-127: An Efficient Delivery Vehicle for 3-(1'-hexyloxy)ethyl-3-devinylpyropheophorbide-a (HPPH or Photochlor).

To determine the impact of delivery vehicles in photosensitizing efficacy of HPPH, a hydrophobic photosensitizer was dissolved in various formulations: 1% Tween 80/5% dextrose, Pluronic P-123 and Pluronic F-127 in 0.5%, 1% and 2% phosphate buffer solutions (PBS). HPPH was also conjugated to Pluronic F-127, and the resulting conjugate (PL-20) was formulated in PBS. Among the different delivery vehicles, only Pluronic P-123 displayed significant vehicle cytotoxicity, whereas Pluronic F127 was nontoxic. Compared to PL-20, HPPH formulated in Tween80 and Pluronic F-127 showed higher cell-uptake, but lower long-term retention in Colon26 cell compared to PL-20. The higher retention of PL-20 was similarly observed during in vivo uptake with BALB/c mice baring Ct26 tumors. In contrast to the in vitro uptake experiments, PL-20 showed slightly higher uptake compared to HPPH formulated in Tween or Pluronic-F127. A significant difference in pharmacokinetic profile was also observed between the HPPH-Pluronic formulation and PL-20. Under similar in vivo treatment parameters (drug dose 0.47 µmol kg-1 , light dose: 135 J cm-2 at 24 h post-injection of PS), HPPH formulated either in Tween or Pluronic F-127 formulation showed similar in vivo PDT efficacy (20-30% tumor cure on day 60), whereas PL-20 showed 40% tumor cure (day 60).

Phototriggered Release of Tumor-Imaging and Therapy Agents from Lyophilized Multifunctional Polyacrylamide Nanoparticles.

The success of polyacrylamide nanoparticles in drug delivery spurred the creation of variations in surface functional groups. We report herein a simple, reproducible, and efficient approach for the creation of modifiable nanoparticles that are characterized by their long-term stability and high loading efficiency. In our experiments, a hydrophobic photosensitizer, such as 2-[1-hexyloxyethyl]-2-devinyl pyropheophorbide-a (HPPH), was able to be postloaded at more than 90% efficiency across all types of nanoparticles (NPs). Moreover, the NPs have tunable release kinetics, ranging from 9% to 23%, released by 96 h in 1% serum albumin, depending on the surface modification used. Additionally, it was observed that the NPs had a photorelease mechanism where >60% of the payload was released when exposed to at least 2 J of light. This held true with the photosensitizer and hydrophobic chemotherapeutics like curcumin. To test the impact these modifications have in vitro, two different bladder cancer cell lines were chosen (UMUC3 and T24). These nanoparticles increase the efficacy of the photosensitizer by 4-fold in UMUC3, with the cationic and amino-functionalized particles having the highest efficacy. This increase in efficacy, high uptake, and favorable subcellular localization makes the cationic modification of the nanoparticle extremely attractive for future studies.

Potential trade-offs between biomineralization and immunity revealed by shell properties and gene expression profiles of two closely related Crassostrea species.

Species of the Ostreidae family are key ecosystem engineers and many of them - including Crassostrea gigas and Crassostreavirginica - are commercially important aquaculture species. Despite similarities in their morphology and ecology, these two species differ in their ability to defend against pathogens, potentially reflecting species-specific differential specialization of hemocytes on immune defense versus biomineralization. To test this hypothesis, we investigated the expression levels of immune- and biomineralization-related genes as well as mineralogical and mechanical properties of the shells and the calcium sequestration ability of the hemocytes of C. gigas and C. virginica The expression of biomineralization-related genes was higher in C. virginica than in C. gigas in multiple tissues including the mantle edge and hemocytes, while the expression of immune genes was higher in the hemocytes of C. gigas Hemocytes of C. virginica contained more calcium (stored intracellularly as calcium carbonate mineral) compared with those of C. gigas Analysis of the adult shells showed that the crystallinity of calcite was higher and the laths of the foliated layer of the shell were thicker in C. virginica than in C. gigas Mechanically, the shells of C. virginica were stiffer, harder and stronger than those of C. gigas Taken together, our results show that the species-specific differences in physiology (such as disease resistance and exoskeleton properties) are reflected at the cellular and molecular levels in the differential specialization of hemocytes on potentially competing functions (immunity and biomineralization) as well as different expression profiles of other tissues involved in biomineralization (such as the mantle edge).

Aggregation of Calcium Phosphate and Oxalate Phases in the Formation of Renal Stones.

The majority of human kidney stones are comprised of multiple calcium oxalate monohydrate (COM) crystals encasing a calcium phosphate nucleus. The physiochemical mechanism of nephrolithiasis has not been well determined on the molecular level; this is crucial to the control and prevention of renal stone formation. This work investigates the role of phosphate ions on the formation of calcium oxalate stones; recent work has identified amorphous calcium phosphate (ACP) as a rapidly forming initial precursor to the formation of calcium phosphate minerals in vivo. The effect of phosphate on the nucleation of COM has been investigated using the constant composition (CC) method in combination with scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Our findings indicate COM nucleation is strongly promoted by the presence of phosphate; this occurs at relatively low phosphate concentrations, undersaturated with respect to brushite (dicalcium phosphate dehydrate, DCPD) formation. The results show that ACP plays a crucial role in the nucleation of calcium oxalate stones by promoting the aggregation of amorphous calcium oxalate (ACO) precursors at early induction times. The coaggregations of ACP and ACO precursors induce the multiple-point nucleation of COM. These novel findings expand our knowledge of urinary stone development, providing potential targets for treating the condition at the molecular level.

Tracking Amorphous Precursor Formation and Transformation during Induction Stages of Nucleation.

Hydroxyapatite (HAP) participates in vertebral bone and tooth formation by a nonclassical hitherto unknown nucleation mechanism, in which amorphous precursors form and transform during long induction periods. Elucidation of the mechanism by which amorphous precursors assemble and transform is essential to understanding how hard tissues form in vivo and will advance the design and fabrication of new biomaterials. The combination of conductance and potentiometric techniques to monitor Ca-P mineral formation has given new insight into the mechanism of nucleation. Differences detected in the dehydration rates of calcium and phosphate ions indicate the formation of nonequilibrium calcium-deficient clusters. The aggregation of these clusters forms a calcium-deficient amorphous phase I [Ca-(HPO4)1+x ·nH2O]2x-) early in the induction period, which slowly transforms to amorphous phase II [Ca-(HPO4)·mH2O] by dehydration. Precritical nuclei form within amorphous phase II later in the induction period, leading to mineral formation.

Kinetics of canine dental calculus crystallization: an in vitro study on the influence of inorganic components of canine saliva.

This work identifies carbonated hydroxyapatite (CAP) as the primary component of canine dental calculus, and corrects the long held belief that canine dental calculus is primarily CaCO3 (calcite). CAP is known to be the principal crystalline component of human dental calculus, suggesting that there are previously unknown similarities in the calcification that occurs in these two unique oral environments. In vitro kinetic experiments mimicking the inorganic components of canine saliva have examined the mechanisms of dental calculus formation. The solutions were prepared so as to mimic the inorganic components of canine saliva; phosphate, carbonate, and magnesium ion concentrations were varied individually to investigate the roll of these ions in controlling the nature of the phases that is nucleated. To date, the inorganic components of the canine oral systems have not been investigated at concentrations that mimic those in vivo. The mineral composition of the synthetic calculi grown under these conditions closely resembled samples excised from canines. This finding adds new information about calculus formation in humans and canines, and their sensitivity to chemicals used to treat these conditions.

Surface-modification-directed controlled adsorption of serum albumin onto magnetite nanocuboids synthesized in a gel-diffusion technique.

Magnetite nanocuboids have been synthesized via gel-diffusion technique in agarose gel. Here, the agarose gel matrix has been used as an organic template for formation and growth modification of magnetite. Gel mineralization mimics the membrane-based biomineralization, controls the diffusion process and gives the micro/nano environment for the crystal growth. We also attempt to understand the influence of different surface modifications of synthesized magnetite nanocuboids on protein interaction. For this purpose, magnetite particles were coated with trimesic acid (benzene-1,3,5-tricarboxylic acid) and stearic acid, which generates a hydrophilic and a hydrophobic modified surface, respectively. We report controlled adsorption behavior of bovine serum albumin (BSA) by surface modification of magnetite nanocuboids with different functional groups. The adsorption capacity of BSA increases on trimesic acid-coated surfaces compared to bare magnetite surfaces, while it decreases on stearic acid-coated surfaces. In situ fluorescence spectroscopy has been used to analyze the tertiary protein structure in the adsorbed state on these three surfaces. Partial unfolding in the tertiary structure of BSA was observed upon adsorption onto bare magnetite surfaces. On trimesic acid-coated surfaces, tertiary unfolding of BSA was greater than on bare magnetite surfaces, while BSA undergoes minor tertiary structural change on stearic acid-coated surfaces.