Background free in vivo29Si MR imaging with hyperpolarized PEGylated silicon nanoparticles.

This study demonstrated the potential of 50 nm PEGylated Si NPs for high-resolution in vivo29Si MR imaging, emphasizing their biocompatibility and water dispersibility. The acquisition of in vivo Si MR images using the lowest reported dose after subcutaneous and intraperitoneal administration opens new avenues for future 29Si MR studies.

Simultaneous analytical method for 296 pesticide multiresidues in root and rhizome based herbal medicines with GC-MS/MS.

A method for the simultaneous analysis of pesticide multiresidues in three root/rhizome-based herbal medicines (Cnidium officinale, Rehmannia glutinosa, and Paeonia lactiflora) was developed with GC-MS/MS. To determine the concentrations of pesticide residues, 5 g of dried samples were saturated with distilled water, extracted with 10 mL of 0.1% formic acid in acetonitrile/ethyl acetate (7:3, v/v), and then partitioned using magnesium sulfate and sodium chloride. The organic layer was purified with Oasis PRiME HLB plus light, followed by a cleanup with dispersive solid-phase extraction containing alumina. The sample was then injected into GC-MS/MS (2 µL) using a pulsed injection mode at 15 psi and analyzed using multiple reaction monitoring (MRM) modes. The limit of quantitation for the 296 target pesticides was within 0.002-0.05 mg/kg. Among them, 77.7-88.5% showed recoveries between 70% and 120% with relative standard deviations (RSDs) ≤20% at fortified levels of 0.01, and 0.05 mg/kg. The analytical method was successfully applied to real herbal samples obtained from commercial markets, and 10 pesticides were quantitatively determined from these samples.

Dynamic Nuclear Polarization of Selectively 29Si-Enriched Core@shell Silica Nanoparticles.

29Si silica nanoparticles (SiO2 NPs) are promising magnetic resonance imaging (MRI) probes that possess advantageous properties for in vivo applications, including suitable biocompatibility, tailorable properties, and high water dispersibility. Dynamic nuclear polarization (DNP) is used to enhance 29Si MR signals via enhanced nuclear spin alignment; to date, there has been limited success employing DNP for SiO2 NPs due to the lack of endogenous electronic defects that are required for the process. To create opportunities for SiO2-based 29Si MRI probes, we synthesized variously featured SiO2 NPs with selective 29Si isotope enrichment on homogeneous and core@shell structures (shell thickness: 10 nm, core size: 40 nm), and identified the critical factors for optimal DNP signal enhancement as well as the effective hyperpolarization depth when using an exogenous radical. Based on the synthetic design, this critical factor is the proportion of 29Si in the shell layer regardless of core enrichment. Furthermore, the effective depth of hyperpolarization is less than 10 nm between the surface and core, which demonstrates an approximately 40% elongated diffusion length for the shell-enriched NPs compared to the natural abundance NPs. This improved regulation of surface properties facilitates the development of isotopically enriched SiO2 NPs as hyperpolarized contrast agents for in vivo MRI.

Simultaneous Determination of Pyrethroid Insecticides in Foods of Animal Origins Using the Modified QuEChERS Method and Gas Chromatography-Mass Spectrometry.

Pyrethroid insecticides are used in agriculture to treat parasites in livestock. This study developed a simultaneous residue analysis method to measure seventeen pyrethroid insecticides in foods of animal origin, including beef, pork, chicken, milk, and eggs. The method, which comprises instrumental analysis using gas chromatography-tandem mass spectrometry and a modified QuEChERS (quick, easy, cheap, effective, rugged, and safe) method for pretreatment, was optimized to verify the applicability of the method. A mixture of acetonitrile, ethyl acetate, and original salt (MgSO4 4 g, NaCl 1 g) was used as the extraction solvent and salt. MgSO4 (150 mg) primary secondary amine (25 mg) and graphitized carbon black (25 mg) were selected for dispersive solid phase extraction (d-SPE). The method limit of quantitation was 0.01 mg/L, and the linearity of the matrix-matched calibration curves was reasonable (R2 > 0.99). Recovery tests were performed at three concentrations (LOQ, 10 LOQ, and 50 LOQ). Good recoveries (75.2109.8%) and reproducibility (coefficient of variation <10%) were obtained. The matrix effects were in the range of −35.8 to 56.0%. The established method was fully validated and can be used as an official analytical method for quantifying pyrethroid insecticides in animal commodities.

A cowpea severe mosaic virus-based vector simplifies virus-induced gene silencing and foreign protein expression in soybean.

BACKGROUND: Soybean gene functions cannot be easily interrogated through transgenic disruption (knock-out) of genes-of-interest, or transgenic overexpression of proteins-of-interest, because soybean transformation is time-consuming and technically challenging. An attractive alternative is to administer transient gene silencing or overexpression with a plant virus-based vector. However, existing virus-induced gene silencing (VIGS) and/or overexpression vectors suitable for soybean have various drawbacks that hinder their widespread adoption. RESULTS: We describe the development of a new vector based on cowpea severe mosaic virus (CPSMV), a plus-strand RNA virus with its genome divided into two RNA segments, RNA1 and RNA2. This vector, designated FZ, incorporates a cloning site in the RNA2 cDNA, permitting insertion of nonviral sequences. When paired with an optimized RNA1 construct, FZ readily infects both Nicotiana benthamiana and soybean. As a result, FZ constructs destined for soybean can be first delivered to N. benthamiana in order to propagate the modified viruses to high titers. FZ-based silencing constructs induced robust silencing of phytoene desaturase genes in N. benthamiana, multiple soybean accessions, and cowpea. Meanwhile, FZ supported systemic expression of fluorescent proteins mNeonGreen and mCherry in N. benthamiana and soybean. Finally, FZ-mediated expression of the Arabidopsis transcription factor MYB75 caused N. benthamiana to bear brown leaves and purple, twisted flowers, indicating that MYB75 retained the function of activating anthocyanin synthesis pathways in a different plant. CONCLUSIONS: The new CPSMV-derived FZ vector provides a convenient and versatile soybean functional genomics tool that is expected to accelerate the characterization of soybean genes controlling crucial productivity traits.

MRI measurement of alanine uptake in a mouse xenograft model of U-87 MG glioblastoma.

The potential use of alanine as an MRI contrast agent was investigated. The relaxation properties of alanine solutions were measured at 9.4 T. The T2 relaxivity caused by the chemical exchange (R2ex) between amine protons and water protons was 0.10 mM-1 s-1 at 37 °C. As a demonstration, alanine uptake in a mouse xenograft model of U-87 MG glioblastoma was measured using MRI, and was compared with immunohistochemistry staining of ASCT2, a transporter that imports amino acids into cancer cells. Statistically significant (p = 0.0079) differences in ASCT2 distribution were found between regions that show strong and weak alanine uptake in MRI. To better understand the influence of perfusion, the effect of ASCT2 inhibition on the alanine uptake in MRI was investigated, and dynamic contrast enhanced MRI was compared with alanine MRI.

Simultaneous determination of nereistoxin insecticides in foods of animal origins by combining pH-dependent reversible partitioning with hydrophilic interaction chromatography-mass spectrometry.

Although nereistoxin insecticides (NIs) are banned for animal husbandry operations, they are still used because of their high insecticidal activities. Therefore, a reliable residue analysis method for the simultaneous detection of cartap, bensultap, thiocyclam, and nereistoxin in foods of animal origins, including beef, pork, chicken, milk, and eggs, was developed using hydrophilic interaction liquid chromatography-mass spectrometry (HILIC-LC-MS/MS). The NIs were extracted with an acidic cysteine and formate buffer solution and hydrolyzed to nereistoxin. The molarity and pH of the buffer were optimized at 20 mM and 3, respectively, to keep the pH of the extracts at 4-5. pH-dependent acid-base partitioning coupled with salting-out-assisted liquid-liquid extraction using acetonitrile was performed for purification and for the direct introduction of the extracts to LC. The optimal pH values were 5 and 9 for the acid-base partitioning. Nereistoxin quantitation was achieved with consistent column retention (RSD < 0.6%) and a high degree of separation (N > 106). The matrix-dependent method limit of quantitation was 2 µg nereistoxin/kg, and the calibration curve showed good linearity (R2 > 0.998). The recovery efficiencies were in the range of 89.2-109.9% with relative standard deviations less than 10%, and matrix effects did not exceed ± 10%, which satisfied the criteria outlined in the European SANTE/12682/2019 guidelines.

29Si Isotope-Enriched Silicon Nanoparticles for an Efficient Hyperpolarized Magnetic Resonance Imaging Probe.

Silicon particles have garnered attention as promising biomedical probes for hyperpolarized 29Si magnetic resonance imaging and spectroscopy. However, due to the limited levels of hyperpolarization for nanosized silicon particles, microscale silicon particles have primarily been the focus of dynamic nuclear polarization (DNP) applications, including in vivo magnetic resonance imaging (MRI). To address these current challenges, we developed a facile synthetic method for partially 29Si-enriched porous silicon nanoparticles (NPs) (160 nm) and examined their usability in hyperpolarized 29Si MRI agents with enhanced signals in spectroscopy and imaging. Hyperpolarization characteristics, such as the build-up constant, the depolarization time (T1), and the overall enhancement of the 29Si-enriched silicon NPs (10 and 15%), were thoroughly investigated and compared with those of a naturally abundant NP (4.7%). During optimal DNP conditions, the 15% enriched silicon NPs showed more than 16-fold higher enhancements─far beyond the enrichment ratio─than the naturally abundant sample, further improving the signal-to-noise ratio in in vivo 29Si MRI. The 29Si-enriched porous silicon NPs used in this work are potentially capable to serve as drug-delivery vehicles in addition to hyperpolarized 29Si in vivo, further enabling their potential future applicability as a theragnostic platform.

Genetic changes and growth promotion of glioblastoma by magnetic nanoparticles and a magnetic field.

Aim: To confirm the biological effects of manganese ferrite magnetic nanoparticles (MFMNPs) and an external magnetic field on glioblastoma cells. Methods: U-87MG glioblastoma cells were prepared, into which the uptake of MFMNPs was high. The cells were then exposed to an external magnetic field using a neodymium magnet in vitro and in vivo. Results:LRP6 and TCF7 mRNA levels involved in the Wnt/ß-catenin signaling pathway were elevated by the influence of MFMNPs and the external magnetic field. MFMNPs and the external magnetic field also accelerated tumor growth by approximately 7 days and decreased survival rates in animal experiments. Conclusion: When MFMNPs and an external magnetic field are applied for a long time on glioblastoma cells, mRNA expression related to Wnt/ß-catenin signaling is increased and tumor growth is promoted.

L-glutamine as a T2 exchange contrast agent.

PURPOSE: The potential of L-glutamine as a T2 exchange contrast agent in MRI was investigated. METHODS: The T2 relaxation rate of L-glutamine solutions prepared in various concentrations was measured at 9.4 T. A series of T2 -weighted images in a mouse cancer model was acquired with an L-glutamine solution infusion. RESULTS: The T2 relaxivity caused by the exchange (R2ex ) at 37°C was 0.069 s-1 mM-1 and 0.102 s-1 mM-1 for glutamine and glutamate solutions at pH = 7.2, respectively. The R2ex of glutamine at pH = 6.1-6.7 was in the 0.097-0.1 s-1 mM-1 range. No significant dependence of T1 on the concentration of glutamine was observed. The dynamic measurement of T2 -weighted images in vivo showed that the glutamine uptake was primarily observed at the localized part of the tumor CONCLUSION: L-glutamine can be used as a T2 exchange contrast agent and images of glutamine uptake in vivo can be acquired.

Correction: Monitoring metal-amyloid-ß complexation by a FRET-based probe: design, detection, and inhibitor screening.

[This corrects the article DOI: 10.1039/C8SC04943B.].

Monitoring metal-amyloid-ß complexation by a FRET-based probe: design, detection, and inhibitor screening.

Aggregation of amyloidogenic peptides could cause the onset and progression of neurodegenerative diseases, such as Alzheimer's disease and Parkinson's disease. These amyloidogenic peptides can coordinate to metal ions, including Zn(ii), which can subsequently affect the peptides' aggregation and toxicity, leading to neurodegeneration. Unfortunately, the detection of metal-amyloidogenic peptide complexation has been very challenging. Herein, we report the development and utilization of a probe (A-1) capable of monitoring metal-amyloid-ß (Aß) complexation based on Förster resonance energy transfer (FRET). Our probe, A-1, is composed of Aß1-21 grafted with a pair of FRET donor and acceptor capable of providing a FRET signal upon Zn(ii) binding even at nanomolar concentrations. The FRET intensity of A-1 increases upon Zn(ii) binding and decreases when Zn(ii)-bound A-1 aggregates. Moreover, as the FRET intensity of Zn(ii)-added A-1 is drastically changed when their interaction is disrupted, A-1 can be used for screening a chemical library to determine effective inhibitors against metal-Aß interaction. Eight natural products (out of 145 compounds; >80% inhibition) were identified as such inhibitors in vitro, and six of them could reduce Zn(ii)-Aß-induced toxicity in living cells, suggesting structural moieties useful for inhibitor design. Overall, we demonstrate the design of a FRET-based probe for investigating metal-amyloidogenic peptide complexation as well as the feasibility of screening inhibitors against metal-bound amyloidogenic peptides, providing effective and efficient methods for understanding their pathology and finding therapeutic candidates against neurodegenerative disorders.

A prospective Phase II study for the efficacy of radiotherapy in combination with zoledronic acid in treating painful bone metastases from gastrointestinal cancers.

We investigated the efficacy of combined radiotherapy (RT) and zoledronic acid in treating painful bone metastases from gastrointestinal cancers. Sixty patients were prospectively enrolled between November 2014 and July 2016. The most common primary cancer type was hepatocellular carcinoma (HCC, n = 25), followed by colorectal cancer (n = 6). Patients received external beam RT of 30-54 Gy in 10-17 fractions or 20 Gy in 5 fractions for symptomatic bone metastases. On the first day of RT, patients received 4 mg intravenous zoledronic acid, which was repeated monthly for a total of six cycles. The mean pain score before treatment was 6.7, and it decreased to 2.8 at 1 month and 2.1 at 3 months (P < 0.001).The overall pain response rates at 1 and 3 months were 95% and 96%, respectively. Among the 24 patients who underwent magnetic resonance imaging, 71% were responders, with a complete response in 1 patient and partial in 16 patients. Combined treatment significantly decreased levels of macrophage inflammatory protein-1α and matrix metalloproteinase (MMP)-2 and -3 compared with baseline (all P < 0.05). In HCC patients, IL-6 and MMP-9 levels were significantly lower 1 month after treatment (P < 0.05). The mean quality of life (QOL) score improved from 66 to 56 at 1 month (P < 0.001) and 55 at 3 months (P = 0.016). The median survival was 7 months. In conclusion, RT with zoledronic acid decreased bone pain and improved QOL in patients with painful bone metastases from gastrointestinal cancers. Radiographic findings and serum biomarker measurements were closely correlated with therapeutic responses.

Spectrum of mitochondrial genome instability and implication of mitochondrial haplogroups in Korean patients with acute myeloid leukemia.

BACKGROUND: Mitochondrial DNA (mtDNA) mutations may regulate the progression and chemosensitivity of leukemia. Few studies regarding mitochondrial aberrations and haplogroups in acute myeloid leukemia (AML) and their clinical impacts have been reported. Therefore, we focused on the mtDNA length heteroplasmies minisatellite instability (MSI), copy number alterations, and distribution of mitochondrial haplogroups in Korean patients with AML. METHODS: This study investigated 74 adult patients with AML and 70 controls to evaluate mtDNA sequence alterations, MSI, mtDNA copy number, haplogroups, and their clinical implications. The hypervariable (HV) control regions (HV1 and HV2), tRNAleu1 gene, and cytochrome b gene of mtDNA were analyzed. Two mtDNA minisatellite markers, 16189 poly-C (16184CCCCCTCCCC16193, 5CT4C) and 303 poly-C (303CCCCCCCTCCCCC315, 7CT5C), were used to examine the mtDNA MSI. RESULTS: In AML, most mtDNA sequence variants were single nucleotide substitutions, but there were no significant differences compared to those in controls. The number of mtMSI patterns increased in AML. The mean mtDNA copy number of AML patients increased approximately 9-fold compared to that of controls (P<0.0001). Haplogroup D4 was found in AML with a higher frequency compared to that in controls (31.0% vs. 15.7%, P=0.046). None of the aforementioned factors showed significant impacts on the outcomes. CONCLUSION: AML cells disclosed more heterogeneous patterns with the mtMSI markers and had increased mtDNA copy numbers. These findings implicate mitochondrial genome instability in primary AML cells. Therefore, mtDNA haplogroup D4 might be associated with AML risk among Koreans.

Inter-alpha Inhibitor H4 as a Potential Biomarker Predicting the Treatment Outcomes in Patients with Hepatocellular Carcinoma.

PURPOSE: Early prediction of treatment outcomes represents an essential step towards increased treatment efficacy and survival in patients with hepatocellular carcinoma (HCC). In this study, we performed two-dimensional electrophoresis (2-DE) followed by protein profiling to identify biomarkers predictive of therapeutic outcomes in patients with HCC who received liver-directed therapy (LDTx) involving local radiotherapy (RT), and studied the underlying mechanisms of the identified proteins. MATERIALS AND METHODS: 2-DE analysis was conducted by pooling sera from patients with a good or poor prognosis; serum proteomic profiles of the two groups were compared and analyzed using matrixassisted laser desorption/ionization time-of-flight mass spectrometry. Identified proteins were confirmed via enzyme-linked immunosorbent assay. An invasion assay was performed after overexpression and knockdown of target protein in Huh7 cells. RESULTS: Levels of inter-alpha inhibitor H4 (ITIH4), fibrinogen gamma chain, keratin 9/1 complex, carbonic anhydrase I, and carbonmonoxyhemoglobin S were changed by more than 4-fold in response to LDTx. In particular, pre-LDTx ITIH4 expression was more than 5-fold higher in patients with a good prognosis, compared to patients with a poor prognosis. The migration ability of Huh7 cells was significantly suppressed and enhanced by ITIH4 overexpression and knockdown, respectively. The tumors of patients with HCC and a good prognosis expressed high levels of ITIH4, compared to those of patients with a poor prognosis. CONCLUSION: Taken together, ITIH4 may be a potential therapeutic target that could inhibit cancer metastasis, as well as a prognostic marker for patients with HCC who are receiving LDTx.

Molecular Imaging of CD44-Overexpressing Gastric Cancer in Mice Using T2 MR Imaging.

Novel diagnostic techniques have been developed in many research area using targetable contrast agents with magnetic resonance imaging (MRI) for cancer diagnosis. For cancer diagnosis, the use of MRI with biocompatible targeting moieties and manganese ferrite nanoparticles (MFNPs) is preferred. Thus, we synthesized MFNPs using a thermal decomposition method which enables sensitive T2 or T2 Turbo Spin Echo (TSE) MRI and coated them with hyaluronic acid (HA). The high targeting ability of HA-MFNPs was observed at MKN-45 cells (gastric cancer cell line) which high-expressing CD44 in contrast with MKN-28 cells which low-expressing CD44. We also prepared the gastric cancer mice model using MKN-45 cells which has the stem-like property was implanted into BALB/c nude mice. And then HA-MFNPs of the T2 contrast enhancement effects and targeting ability were investigated by in vivo MR imaging. As a result of these studies, we conclude that HA coated MFNPs can be effectively used as a novel probes for visualizing gastric cancer stem cells.

Clinical and oncologic outcomes of totally robotic total mesorectal excision for rectal cancer: initial results in a center for minimally invasive surgery.

PURPOSE: A robotic system was mainly designed to allow precise dissection in deep and narrow spaces. We report the clinical and oncologic outcomes of totally robotic total mesorectal excision for rectal cancer. METHODS: Between July 2009 and January 2012, 60 consecutive patients undergoing robotic surgery for rectal cancer at the Eulji University Hospital were included. RESULTS: The mean total operation time, docking time, and surgeon console time were 466.8 ± 115.6, 7.5 ± 6.7, and 261 ± 87.5 min, respectively. Oral intake of diet was started at 3.3 ± 0.9 days and the mean hospital stay was 8.6 ± 2.4 days. All 60 procedures were technically successful without the need for conversion to open or laparoscopic surgery. Complications included anastomotic leakage, anastomotic stricture, postoperative bleeding, ileus, and perineal wound infection in 3 (5 %), 1 (1.7 %), 2 (3.3 %), 2 (3.3 %), and 1 (1.7 %) patient, respectively. The mean distal resection margin and total number of lymph nodes harvested was 3.1 ± 1.7 cm and 20.1 ± 11.5, respectively. During the mean follow-up period of 48.5 months (range, 7-75), the 4-year overall and disease-free survival rates were 87.7 and 72.8 %, respectively. CONCLUSIONS: A totally robotic approach for rectal cancer operations was a time-consuming procedure, although we already had a lot experience in laparoscopic colorectal surgery. However, the dexterity of the robotic surgery could enable the surgeon to expand the choice of surgical methods according to the condition of the rectal cancer without the need for conversion.

Radiation Inhibits Interleukin-12 Production via Inhibition of C-Rel through the Interleukin-6/ Signal Transducer and Activator of Transcription 3 Signaling Pathway in Dendritic Cells.

Radiotherapy (RT) is a potent anti-tumor modality. However, unwanted effects including increased recurrence and metastasis that involve factors such as cytokines, which induce complex molecular mechanisms, have also been reported. In a previous study, we showed that interleukin (IL)-12 and radiotherapy combination treatment suppressed tumor growth and metastasis in a hepatoma mouse model. In this study, we investigated the mechanism underlying the IL-12 anti-tumor effect during radiotherapy. In tumor-bearing mice, irradiation decreased IL-12 expression in the tumors and spleens. However, a number of dendritic cells infiltrated into the tumors in which IL-12 expression did not decrease. To further study the underlying detailed mechanism for this decrease in IL-12, LPS-stimulated bone marrow-derived dendritic cells (BMDCs) were irradiated, and then IL-12- and IL-6-associated molecules were examined in irradiated tumors and BMDCs. Irradiation resulted in IL-12 suppression and IL-6 increase. IL-6 and signal transducer and activator of transcription 3 (STAT3) inhibitors restored the irradiation-induced IL-12 decrease via suppression of C-Rel activation. Taken together, our study suggests that irradiation-induced IL-6 can decrease IL-12 production through the inhibition of C-Rel phosphorylation by the IL-6/STAT3 signaling pathway.

Irradiation-induced localization of IL-12-expressing mesenchymal stem cells to enhance the curative effect in murine metastatic hepatoma.

Irradiation in conjunction with gene therapy is considered for efficient cancer treatment. Mesenchymal stem cells (MSCs), due to their irradiation-promotable tumor tropism, are ideal delivery vehicles for gene therapy. In this study, we investigated whether treatment with radiation and interleukin (IL)-12-expressing MSCs (MSCs/IL-12) exerts improved antitumor effects on murine metastatic hepatoma. HCa-I and Hepa 1-6 cells were utilized to generate heterotopic murine hepatoma models. Tumor-bearing mice were treated with irradiation or MSCs/IL-12 alone, or a combination. Monocyte chemoattractant protein-1 (MCP-1/CCL2) expression was assessed in irradiated hepatoma tissues to confirm a chemotactic effect. Combination treatment strategies were established and their therapeutic efficacies were evaluated by monitoring tumor growth, metastasis and survival rate. IL-12 expression was assessed and the apoptotic activity and immunological alterations in the tumor microenvironment were examined. MCP-1/CCL2 expression and localization of MSCs/IL-12 increased in the irradiated murine hepatoma cells. The antitumor effects, including suppression of pulmonary metastasis and survival rate improvements, were increased by the combination treatment with irradiation and MSCs/IL-12. IL-12 expression was increased in tumor cells, causing proliferation of cluster of differentiation 8(+) T-lymphocytes and natural killer cells. The apoptotic activity increased, indicating that the cytotoxicity of immune cells was involved in the antitumor effect of the combined treatment. Treatment with irradiation and MSCs/IL-12 showed effectiveness in treating murine metastatic hepatoma. IL-12-induced proliferation of immune cells played an important role in apoptosis of tumor cells. Our results suggest that treatment with irradiation and MSCs/IL-12 may be a useful strategy for enhancing antitumor activity in metastatic hepatoma.