D‐ and L‐[123I]‐2‐I‐phenylalanine show a long tumour retention compared with D‐ and L‐[123I]‐2‐I‐tyrosine in R1M rhabdomyosarcoma tumour‐bearing Wag/Rij rats

The aim of this study was the comparison of the tumour uptake and the long‐term retention of [¹²³I]‐2‐I‐L ‐phenylalanine and [¹²³I]‐2‐I‐D ‐phenylalanine with those of [¹²³I]‐2‐I‐L ‐tyrosine and [¹²³I]‐2‐I‐D ‐tyrosine in R1M rhabdomyosarcoma tumour‐bearing rats. The biodistribution of the radioactivity as a function of time in R1M tumour‐bearing rats was measured by planar gamma camera imaging (dynamic and static). If dissection was applied, the activity in the tumours and tissues of interest was measured by gamma counting. [¹²³I]‐2‐iodo‐L ‐phenylalanine, [¹²³I]‐2‐iodo‐D ‐phenylalaine, [¹²³I]‐2‐I‐L ‐tyrosine showed a considerable tumour uptake reaching a maximum between 10 and 30 min. At 30 min p.i. the differential uptake ratio values of this uptake were, respectively, 2.1, 2.3, 2.5 and 1.7. The activity in the tumour was shown to be related to a tumour cell uptake and not to an increased blood pool activity. All the tracers showed a clearance from the blood to the bladder without renal retention. At longer times both L ‐ and D ‐ [¹²³I]‐2‐I‐tyrosine were cleared for a large part from the tumours and the body. [¹²³I]‐2‐I‐L ‐Phe and [¹²³I]‐2‐I‐D ‐Phe showed a considerable and equal retention in the tumours: as compared with 0.5 h, 91% at 24 h and 80% at 48 h. This was related to the longer retention of activity in the blood pool noticed for these compounds (81% at 24 h and 65% at 48 h). The tumour‐to‐background ratio increased with 25% at those longer times. At short times all the tracers were taken up to a considerable extent in the tumours. In the R1M‐bearing Wag/Rij rat model only [¹²³I]‐2‐I‐L ‐phenylalanine and [¹²³I]‐2‐I‐D ‐phenylalanine showed an especially high retention at long times without any significant difference between the enantiomers. Copyright © 2007 John Wiley & Sons, Ltd.     

Influence of right ventricular pre‐ and afterload on right ventricular ejection fraction and preload recruitable stroke work relation

When right ventricular (RV) afterload is abnormally increased, it correlates inversely with right ventricular ejection fraction (RVEF). We tested, whether this would be different with normal afterload. Additionally, we investigated whether previous studies on the slope of RV preload recruitable stroke work (SW) relation, which used rather non‐physiological measures to change RV preload, could be transferred to more physiological loading conditions. RV volumes were determined by thermodilution in 16 patients with stable coronary artery disease and normal pulmonary artery pressure (PAP) at rest. Pre‐ and afterload were varied by body posture, nitroglycerin (NTG) application and by exercise at different body positions. At rest, the change from recumbent to sitting position decreased PAP, cardiac index (C_i), RV diastolic and systolic volumes, and RVEF. Additionally, mean pulmonary artery pressure (MPAP) correlated positively with both RVEF and cardiac index. After correction for mathematical coupling, the RV preload recruitable SW relation was: right ventricular stroke work index (RVSW_i) (10³ erg m^–2)= 8·1 × (RV end‐diastolic volume index ?4·9), with n=96, r=0·57, P≤0·001. Exercise abolished this correlation and led to an inverse correlation between RV end‐systolic volume (ESV) and RVSW. In conclusion, (i) RVEF correlates positively with RV afterload when afterload varies within normal range; (ii) the slope of the RV preload recruitable SW relation, which is obtained at steady state under normal loading conditions, is substantially flatter than previously described for dynamic changes of RV preload. With increasing afterload, preload loses its determining effect on RV performance, while afterload becomes more important. This puts earlier assumptions of an afterload independent RV preload recruitable SW relation into question.     

The effect of glutamine on protein balance and amino acid flux across arm and leg tissues in healthy volunteers

Background. Glutamine is important in nitrogen transportation and the physiological control of acid–base regulation. In addition, it has been assumed that glutamine regulates protein balance in skeletal muscles based on findings in both experimental and clinical studies. However, little information on glutamine and its effect on protein dynamics in normal individuals is available. Therefore, the aim of this study was to evaluate whether glutamine improves protein balance and uptake of various indispensable amino acids across peripheral tissue in healthy individuals. Material and methods. Standard primed constant infusions of L ‐[ring‐²H₅]phenylalanine and [ring 3,3‐²H₂]tyrosine (2 μmol kg^?1 h^?1) were performed after overnight fast in five healthy male volunteers before and during infusions of a standard and a glutamine/tyrosine enriched amino acid solution. Flux measurements of amino acids (AA) including 3‐methylhistidine, glucose, lactate and free fatty acids (FFA) were performed across arm and leg tissues. Results. Infusion of the standard AA solution (0·2 g N kg^?1 day^?1) increased the net uptake of individual amino acids, but provision of the enriched solution (0·4 g N kg^?1 day^?1) with increased amounts of glutamine and tyrosine seemed to compete unfavourably with the net uptake of other key amino acids as methionine and phenylalanine, which are indispensable in muscles for protein synthesis. Increased flux of amino acids across peripheral tissues did not influence on flux of glucose, free fatty acid and lactate. Conclusions. Glutamine provision did neither stimulate protein synthesis nor attenuate breakdown of either globular or myofibrillar proteins in skeletal muscles of healthy volunteers.     

Sustained‐release of naproxen sodium from electrospun‐aligned PLLA–PCL scaffolds

Spontaneous tendon healing may result in reduced tissue functionality. In view of this, tissue engineering (TE) emerges as a promising approach in promoting proper tendon regeneration. However, unfavourable post‐surgical adhesion formations restrict adequate tendon healing through the TE approach. Naproxen sodium (NPS), a non‐steroidal anti‐inflammatory drug (NSAID), has been demonstrated to prevent adhesions by inhibiting the inflammatory response. Therefore, in this study, various factors, such as polymer composition, i.e. different poly‐l ‐lactic acid (PLLA):polycaprolactone (PCL) ratios, and percentage of water:hexafluoroisopropanol (HFIP; as co‐solvent) ratios, were investigated to understand how these can influence the release of NPS from electrospun scaffolds. By adjusting the amount of water as the co‐solvent, NPS could be released sustainably for as long as 2 weeks. Scaffold breaking strength was also enhanced with the addition of water as the co‐solvent. This NPS‐loaded scaffold showed no significant cytotoxicity, and L929 murine fibroblasts cultured on the scaffolds were able to proliferate and align along the fibre orientation. These scaffolds with desirable tendon TE characteristics would be promising candidates in achieving better tendon regeneration in vivo. Copyright © 2015 John Wiley & Sons, Ltd.     

Localization,mechanism and reduction of renal retention of technetium‐99m labeled epidermal growth factor receptor‐specific nanobody in mice

Background

Nanobodies are single‐domain antigen binding fragments derived from functional heavy‐chain antibodies elicited in Camelidae. They are powerful probes for radioimmunoimaging, but their renal uptake is relatively high. In this study we have evaluated the role of megalin on the renal uptake of anti‐EGFR ^99mTc‐7C12 nanobody and the potency of gelofusine and/or lysine to reduce renal uptake of ^99mTc‐7C12.

Methods

First we compared the renal uptake of ^99mTc‐7C12 in megalin‐deficient and megalin‐wild‐type mice using pinhole SPECT/microCT and ex vivo analysis. The effect of gelofusine and lysine administration on renal accumulation of ^99mTc‐7C12 was analyzed in CD‐1 mice divided into lysine preload at 30 min before tracer injection (LysPreload), LysPreload + gelofusine coadministration (LysPreload + GeloCoad), lysine coadministration (LysCoad), gelofusine coadministration (GeloCoad) and LysCoad + GeloCoad. The combined effect of gelofusine and lysine on tumor uptake was tested in mice xenografts.

Results

Renal uptake of ^99mTc‐7C12 was 44.22 ± 3.46% lower in megalin‐deficient compared with megalin‐wild‐type mice. In CD‐1 mice, lysine preload had no effect on the renal retention whereas coinjection of lysine or gelofusine with the tracer resulted in 25.12 ± 2.99 and 36.22 ± 3.07% reduction, respectively. The combined effect of gelofusine and lysine was the most effective, namely a reduction of renal retention of 45.24 ± 2.09%. Gelofusine and lysine coadministration improved tumor uptake.

Conclusion

Megalin contributes to the renal accumulation of ^99mTc‐7C12. Gelofusine and lysine coinjection with the tracer reduces the renal uptake while tumor uptake is improved. Although this methodology allows for optimization of imaging protocol using nanobodies, further improvements are needed before using these molecules for radionuclide therapy. Copyright © 2010 John Wiley & Sons, Ltd.     

Syntheses and preliminary evaluation of [18F]AlF‐NOTA‐G‐TMTP1 for PET imaging of high aggressive hepatocellular carcinoma

The goal of this study is to evaluate a new ¹⁸F‐labeled imaging agent for diagnosing high metastatic (aggressive) hepatocellular carcinoma using positron emission tomography (PET). The new ¹⁸F‐labeled imaging agent [¹⁸F]AlF‐NOTA‐G‐TMTP1 was synthesized and radiolabeled with ¹⁸F using NOTA‐AlF chelation method. The tumor‐targeting characteristics of [¹⁸F]AlF‐NOTA‐G‐TMTP1 was assessed in HepG2, SMCC‐7721, HCC97L and HCCLM3 xenografts. The total synthesis time was about 20 min with radiochemical yield of 25 ± 6%. The specific activity was about 11.1–14.8 GBq/µmol at the end of synthesis based on the amount of peptide used and the amount of radioactivity trapped on the C₁₈ column. The log P value of [¹⁸F]AlF‐NOTA‐G‐TMTP1 was ‐3.166 ± 0.022. [¹⁸F]AlF‐NOTA‐G‐TMTP1 accumulated in SMCC‐7721 and HCCLM3 tumors (high metastatic potential) in vivo and result in tumor/muscle (T/M) ratios of 4.5 ± 0.3 and 4.7 ± 0.2 (n = 4) as measured by PET at 40 min post‐injection (p.i.). Meanwhile, the tumor/muscle (T/M) ratios of HepG2 and HCC97L tumors (low metastatic potential) were1.6 ± 0.3 and 1.8 ± 0.4. The tumor uptake of [¹⁸F]AlF‐NOTA‐G‐TMTP1 could be inhibited 61.9% and 57.6% by unlabeled G‐TMTP1 in SMCC‐7721 and HCCLM3 xenografts at 40 min p.i., respectively. Furthermore, [¹⁸F]AlF‐NOTA‐G‐TMTP1 showed pretty low activity in the liver and intestines in all tumor bearing mice, such in vivo distribution pattern would be advantageous for the detection of hepatic carcinoma. Overall, [¹⁸F]AlF‐NOTA‐G‐TMTP1 may specifically target high metastatic or/and aggressive hepatocellular carcinoma with low background activity and, therefore, holds the potential to be used as an imaging agent for detecting tumor lesions within the liver area. Copyright © 2016 John Wiley & Sons, Ltd.     

ITAM receptor‐mediated generation of reactive oxygen species in human platelets occurs via Syk‐dependent and Syk‐independent pathways

Background: Ligation of the platelet‐specific collagen receptor, GPVI/FcRγ, causes rapid, transient disulfide‐dependent homodimerization, and the production of intracellular reactive oxygen species (ROS) generated by the NADPH oxidase, linked to GPVI via TRAF4. Objectives: The aim of this study was to evaluate the role of early signaling events in ROS generation following engagement of either GPVI/FcRγ or a second immunoreceptor tyrosine‐based activation motif (ITAM)‐containing receptor on platelets, FcγRIIa. Methods and Results: Using an H₂DCF‐DA‐based flow cytometric assay to measure intracellular ROS, we show that treatment of platelets with either the GPVI agonists, collagen‐related peptide (CRP) or convulxin (Cvx), or the FcγRIIa agonist 14A2, increased intraplatelet ROS; other platelet agonists such as ADP and TRAP did not. Basal ROS in platelet‐rich plasma from 14 healthy donors displayed little inter‐individual variability. CRP, Cvx or 14A2 induced an initial burst of ROS within 2 min followed by additional ROS reaching a plateau after 15–20 min. The Syk inhibitor BAY61‐3606, which blocks ITAM‐dependent signaling, had no effect on the initial ROS burst, but completely inhibited the second phase. Conclusions: Together, these results show for the first time that ROS generation downstream of GPVI or FcγRIIa consists of two distinct phases: an initial Syk‐independent burst followed by additional Syk‐dependent generation.     

The anticonvulsant remacemide and its metabolite AR‐R12495AA attenuate spinal synaptic transmission and carrageenan‐induced inflammation in the young rat

The effects of the anticonvulsants remacemide [(±)‐2‐amino‐N‐(1‐methyl‐1,2‐diphenylethyl)‐acetamide hydrochloride] and its des ‐glycinated metabolite AR‐R12495AA [(±)‐1‐methyl‐1,2‐diphenylethylamine‐ monohydrochloride] on primary afferent‐induced synaptic transmission and frequency‐dependent summation of synaptic potentials were assessed in the young rat spinal cord in vitro. Behavioural studies in the rat determined the effects of these anticonvulsant compounds in the carrageenan model of inflammation. Recordings of the extracellular dorsal root‐evoked ventral root potential (DR‐VRP) revealed a significant reduction of the duration and t1/2 decay of the long latency, slow DR‐VRP by remacemide (50 and 100 μM) and AR‐R12495AA (25, 50 and 100 mM). The short‐latency, fast monosynaptic DR‐VRP peak was reduced by only the highest concentration of AR‐R12495AA (100 μM). In intracellular dorsal root‐evoked excitatory postsynaptic potentials (DR‐EPSPs) of single ventral horn neurons, AR‐R12495AA (100 μM) attenuated the time course of the long‐latency (slow) EPSP. Frequency‐dependent (0.5–2.0 Hz) summation of dorsal root‐evoked synaptic events (recorded extracellularly as the cumulative ventral root depolarization (CVRD), and intracellularly as wind‐up) was attenuated by remacemide (100 μM) and AR‐R12495AA (50 and 100 μM). Pre‐treatment with intra‐peritoneal injection of 75 mg/kg of remacemide or AR‐R12495AA caused a significant reduction of carrageenan‐induced mechanical hyperalgesia and oedema. These electrophysiological and behavioural data provide evidence that remacemide and AR‐R12495AA may also possess analgesic and anti‐inflammatory activity. Copyright 2000 European Federation of Chapters of the International Association for the Study of Pain     

Solanum indicum ssp. distichum extract is effective against l‐NAME‐induced hypertension in rats

Solanum indicum ssp. distichum is used as a vegetable in some parts of Africa and claimed in folk medicine to guard against cardiovascular disorders. It was of interest to study the potential blood pressure lowering effects of a standardized extract of the fruit. An ethanolic extract of the fruit, standardized to contain > 0.15% chlorogenic acids, was tested orally in both normotensive rats and in those rendered hypertensive by twice daily intraperitoneal injection of N^W‐nitro‐l ‐arginine methylester (l ‐NAME) for 1 week. The extract was either given at the same time as l ‐NAME or after the establishment of hypertension. The systolic blood pressure (SBP) was measured non‐invasively using a tail cuff computer‐aided monitoring device. Treatment of normotensive rats with the extract (30–300 mg/kg) for 4 weeks showed no hypotensive effect. Giving the extract (100 and 300 mg/kg) orally once daily during the 1 week hypertension induction period with l ‐NAME prevented the development of hypertension. Administration of the extract orally for 1 week after the establishment of hypertension tended to normalize the blood pressure. Pharmacological evidence for the antihypertensive activity of S. distichum is hereby reported for the first time. The extract showed good prophylactic as well as curative effect against l ‐NAME‐induced hypertension, whereby its content of chlorogenic acids may play a minor role. Other constituents may be responsible for the antihypertensive action. The findings support further development of the extract as a potential therapeutically useful antihypertensive agent.     

Phosphoinositide 3‐kinase mediates CD40 ligand‐induced oxidative stress and endothelial dysfunction via Rac1 and NADPH oxidase 2

Summary. Objectives: CD40 ligand (CD40L) has been implicated as an inducer of reactive oxygen species (ROS) generation in endothelial cells, but definitive evidence for this and the in vivo relevance haves not been demonstrated fully. We thus investigated whether phosphoinositide 3‐kinase (PI3K) was linked to ROS generation and endothelial reactivity in response to CD40L. Methods and Results: CD40L treatment activated PI3K activity by regulating the association between PI3K p85 and the CD40 receptor. CD40L exposure also stimulated the GTPase Rac1, which is known to activate NADPH oxidases, and enhanced ROS formation, whereas PI3K inhibition or depletion by small interfering RNA (siRNA) prevented these responses. Subsequently, PI3K overexpression activated Rac1 and increased ROS generation. These responses were not observed in the presence of inactive Rac1 or siRNA against the NADPH oxidase subunit NOX2. Protein kinase Cζ mediates PI3K‐regulated NADPH oxidase activation by promoting cellular p47phox translocation. Importantly, PI3K inhibition prevented CD40L‐mediated ROS generation and endothelial dysfunction in a mouse model. In summary, PI3K mediates CD40L‐induced ROS production and subsequent endothelial dysfunction. Conclusions: Targeting PI3K may provide a new therapeutic approach in diseases associated with oxidative stress and endothelial dysfunction.     

Human platelet antigen polymorphisms (HPA‐1, ‐2, ‐3, ‐4, ‐5 and ‐15) in major ethnic groups of Pakistan

Objectives: Gene frequencies of human platelet antigens (HPA) determine the magnitude of platelet immunological disorders like neonatal alloimmune thrombocytopenia, platelet refractoriness and ease of availability of particular HPA‐typed platelet donors in a given community. Background: However, the pattern of HPA in Pakistani population is not known. Aim: The aim of present study was to determine the gene frequencies of HPA (HPA‐1 to ‐5 and ‐15) in individuals belonging to major ethnic groups and castes of Pakistani population. Materials and Methods: HPA genotyping was done in 593 individuals belonging to all ethnic groups of Pakistan, by polymerase chain reaction‐sequence specific primers with detection on polyacrylamide electrophoresis. Results: The gene frequencies of the ‘a’ and ‘b’ alleles of HPA‐1 to ‐5 and ‐15 in Pakistanis were as follows: HPA‐1a/b, 0·885/0·115; HPA‐2a/b, 0·92/0·08; HPA‐3a/b, 0·69/0·31; HPA‐4a/b, 1/0; HPA‐5a/b, 0·9/0·1; HPA‐15a/b, 0·59/0·41. Except for significant difference regarding gene frequency of HPA‐3 between Pathans and Sindhis, there was no significant difference of HPA‐1 to ‐5 and ‐15 between major ethnic groups of Pakistan. The estimated mismatch probability regarding platelet antigens 1–5 and 15 in Pakistanis, after transfusion of random donor platelets, is from 14 to 37%. The expected incidence of neonatal alloimmune thrombocytopenia due to anti‐HPA‐1a in Pakistani pregnant females is < 1 of 1000 pregnancies and 8–12 of 1000 in case of anti‐HPA‐5b. Homozygosity of HPA‐1b, ‐2b and ‐5b genotypes ranged from 1 to 2% in the Pakistani population, whereas homozygosity of HPA‐3b and ‐15b was 11 and 18%. Conclusions: There is a need to establish donor registries typed for HPA in the transfusion centres of the country.     

Biodistribution and evaluation of 131I‐labeled neuropilin‐binding peptide for targeted tumor imaging

Neuropilin‐1 (NRP‐1) is overexpressed in several kinds of cancer cell and contributes to tumor aggressiveness. Recently, the arginine/lysine‐rich peptide with C‐terminal motifs (R/K)XX(R/K) indicated promising penetrating and transporting capability into NRP‐1 positive cancer cells. In the present study, we describe a ¹³¹I‐labeled C‐end rule motif peptide conjugate, Tyr–tLyp‐1, for NRP‐1 positive tumor targeting and imaging properties. Briefly, a truncated Lyp‐1 peptide was designed to expose its C‐end motif and conjugated to tyrosine for radiolabeling after structural modification. The peptide indicated specific binding to A549 cancer cells at 2 μM concentration, and its binding was dependent on NRP‐1 expression and could be inhibited by other NRP‐1‐binding peptides. In vivo imaging of ¹³¹I‐labeled Tyr–tLyp‐1peptide showed that a subcutaneous A549 xenograft tumor could be visualized using a SPECT/CT scanner. The tumor uptake of ¹³¹I‐Tyr–tLyp‐1 was 4.77 times higher than the uptake in muscles by SPECT/CT software quantification at 6 h post injection. Together, this study indicated that truncated Lyp‐1 peptide could specifically localize in NRP‐1 positive tumors and successfully mediate the ¹³¹I radionuclide diagnosis, indicating promising targeted imaging capability for NRP‐1 positive tumors. Copyright © 2016 John Wiley & Sons, Ltd.     

Leukocytes as a reservoir of circulating oncogenic DNA and regulatory targets of tumor‐derived extracellular vesicles

Essentials

• Tumor‐bearing mice were employed to follow oncogenic HRAS sequences in plasma, and blood cells.
• Cancer DNA accumulated in leukocytes above levels detected in exosomes, platelets and plasma.
• Extracellular vesicles and nucleosomes are required for uptake of tumor DNA by leukocytes.
• Uptake of tumor‐derived extracellular vesicles by leukocytes triggers coagulant phenotype.

Summary

Background

Tumor‐derived extracellular vesicles (EVs) and free nucleosomes (NSs) carry into the circulation a wealth of cancer‐specific, bioactive and poorly understood molecular cargoes, including genomic DNA (gDNA).

Objective

Here we investigated the distribution of extracellular oncogenic gDNA sequences (HRAS and HER2) in the circulation of tumor‐bearing mice.

Methods and Results

Surprisingly, circulating leukocytes (WBCs), especially neutrophils, contained the highest levels of mutant gDNA, which exceeded the amount of this material recovered from soluble fractions of plasma, circulating EVs, platelets, red blood cells (RBCs) and peripheral organs, as quantified by digital droplet PCR (ddPCR). Tumor excision resulted in disappearance of the WBC‐associated gDNA signal within 2–9 days, which is in line with the expected half‐life of these cells. EVs and nucleosomes were essential for the uptake of tumor‐derived extracellular DNA by neutrophil‐like cells and impacted their phenotype. Indeed, the exposure of granulocytic HL‐60 cells to EVs from HRAS‐driven cancer cells resulted in a selective increase in tissue factor (TF) procoagulant activity and interleukin 8 (IL‐8) production. The levels of circulating thrombin‐antithrombin complexes (TAT) were markedly elevated in mice harboring HRAS‐driven xenografts.

Conclusions

Myeloid cells may represent a hitherto unrecognized reservoir of cancer‐derived, EV/NS‐associated oncogenic gDNA in the circulation, and a possible novel platform for liquid biopsy in cancer. In addition, uptake of this material alters the phenotype of myeloid cells, induces procoagulant and proinflammatory activity and may contribute to systemic effects associated with cancer.

     

MRI tumor characterization using Gd‐GlyMe‐DOTA‐perfluorooctyl‐mannose‐conjugate (Gadofluorine M™), a protein‐avid contrast agent

The rationale and objectives were to define the MRI tumor‐characterizing potential of a new protein‐avid contrast agent, Gd‐GlyMe‐DOTA‐perfluorooctyl‐mannose‐conjugate (Gadofluorine M?; Schering AG, Berlin, Germany) in a chemically induced tumor model of varying malignancy. Because of the tendency for this agent to form large micelles in water and to bind strongly to hydrophobic sites on proteins, it was hypothesized that patterns of dynamic tumor enhancement could be used to differentiate benign from malignant lesions, to grade the severity of malignancies and to define areas of tumor necrosis. Gadofluorine M, 0.05 mmol Gd kg^?1, was administered intravenously to 28 anesthetized rats that had developed over 10 months mammary tumors of varying degrees of malignancy as a consequence of intraperitoneal administration of N‐ethyl‐N‐nitrosourea (ENU), 45–250 mg kg^?1. These tumors ranged histologically from benign fibroadenomas to highly undifferentiated adenocarcinomas. Dynamic enhancement data were analyzed kinetically using a two‐compartment tumor model to generate estimates of fractional plasma volume (fPV), apparent fractional extracellular volume (fEV*) and an endothelial transfer coefficient (K^PS) for this contrast agent. Tumors were examined microscopically for tumor type, degree of malignancy (Scarff–Bloom–Richardson score) and location of necrosis. Eighteen tumor‐bearing rats were successfully imaged. MRI data showed an immediate strong and gradually increasing tumor enhancement. K^PS and fEV*, but not fPV obtained from tumors correlated significantly (p < 0.05) with the SBR tumor grade, r = 0.65 and 0.56, respectively. Estimates for K^PS and fEV* but not fPV were significantly lower in a group consisting of benign and low‐grade malignant tumors compared with the group of less‐differentiated high‐grade tumors (1.61 ± 0.64 vs 3.37 ± 1.49, p < 0.01; 0.45 ± 0.17 vs 0.78 ± 0.24, p < 0.01; and 0.076 ± 0.048 vs 0.121 ± 0.088, p = 0.24, respectively). It is concluded that the protein‐avid MRI contrast agent Gadofluorine M enhances tumors of varying malignancy depending on the tumor grade, higher contrast agent accumulation for more malignant lesions. The results show potential utility for differentiating benign and low‐grade malignant lesions from high‐grade cancers. Copyright © 2006 John Wiley & Sons, Ltd.     

Mesenchymal stem cells protect from acute liver injury by attenuating hepatotoxicity of liver natural killer T cells in an inducible nitric oxide synthase‐ and indoleamine 2,3‐dioxygenase‐dependent manner

The effects of mesenchymal stem cells (MSCs) on the phenotype and function of natural killer T (NKT) cells is not understood. We used concanavalin A (Con A) and α‐galactosylceramide (α‐GalCer)‐induced liver injury to evaluate the effects of MSCs on NKT‐dependent hepatotoxicity. Mouse MSCs (mMSCs) significantly reduced Con A‐ and α‐GalCer‐mediated hepatitis in C57Bl/6 mice, as demonstrated by histopathological and biochemical analysis, attenuated the influx of inflammatory [T‐bet⁺, tumour necrosis factor‐α (TNF‐α), interferon‐γ (IFN‐γ)‐producing and GATA3⁺, interleukin‐4 (IL‐4)‐producing] liver NKT cells and downregulated TNF‐α, IFN‐γ and IL‐4 levels in the sera. The liver NKT cells cultured in vitro with mMSCs produced lower amounts of inflammatory cytokines (TNF‐α, IFN‐γ, IL‐4) and higher amounts of immunosuppressive IL‐10 upon α‐GalCer stimulation. mMSC treatment attenuated expression of apoptosis‐inducing ligands on liver NKT cells and suppressed the expression of pro‐apoptotic genes in the livers of α‐GalCer‐treated mice. mMSCs reduced the cytotoxicity of liver NKT cells against hepatocytes in vitro. The presence of 1‐methyl‐dl ‐tryptophan, a specific inhibitor of indoleamine 2,3‐dioxygenase (IDO), or l ‐N^G‐monomethyl arginine citrate, a specific inhibitor of inducible nitric oxide synthase (iNOS), in mMSC‐conditioned medium injected into α‐GalCer‐treated mice, counteracted the hepatoprotective effect of mMSCs in vivo and restored pro‐inflammatory cytokine production and cytotoxicity of NKT cells in vitro. Human MSCs attenuated the production of inflammatory cytokines in α‐GalCer‐stimulated human peripheral blood mononuclear cells in an iNOS‐ and IDO‐dependent manner and reduced their cytotoxicity against HepG2 cells. In conclusion, MSCs protect from acute liver injury by attenuating the cytotoxicity and capacity of liver NKT cells to produce inflammatory cytokines in an iNOS‐ and IDO‐dependent manner.     

Further analysis of acute antinociceptive and anti‐inflammatory actions of 4‐phenylselenyl‐7‐chloroquinoline in mice

A new quinoline containing selenium, 4‐phenylselenyl‐7‐chloroquinoline (4‐PSQ), was described and synthetized by our research group. Recently, we demonstrated the potential antinociceptive and anti‐inflammatory of 4‐PSQ. For this reason, the first objective of this study was to expand our previous findings by investigating the contribution of glutamatergic, serotonergic, and nitrergic systems to the acute antinociceptive action of this compound. Pretreatment with 4‐PSQ (0.01–25 mg/kg, p.o.) reduced the nociception induced by glutamate. MK‐801 (an uncompetitive antagonist of the N‐Methyl‐d ‐aspartate (NMDA) receptor) blocked the antinociceptive effect exerted by 4‐PSQ (25 mg/kg, p.o.) in the acetic acid‐induced abdominal writhing test. The pretreatment with WAY100635 (a selective antagonist of 5‐HT_1A receptor), ketanserin (a selective antagonist of 5‐HT_2A/2C receptor), and pindolol (a nonselective antagonist of 5‐HT_1A/1B receptors) partially blocked the antinociceptive effect caused by 4‐PSQ (25 mg/kg, per oral, p.o.) in the acetic acid‐induced abdominal writhing test. Nitric oxide precursor, l ‐arginine hydrochloride, partially reversed antinociception caused by 4‐PSQ or ω‐nitro‐l ‐arginine (l ‐NOARG). Treatments did not modify the locomotor and exploratory activities of mice. Additionally, the acute anti‐inflammatory effect of 4‐PSQ in a model of pleurisy induced by carrageenan in mice was investigated. 4‐PSQ reduced the cellular migration, pleural exudate accumulation, and myeloperoxidase activity induced by carrageenan exposure. 4‐PSQ protected against the increase in reactive species levels and reduction of nonprotein thiol levels induced by carrageenan. Data presented here showed that the modulation of serotonergic, nitrergic, and glutamatergic systems contributed to the antinociceptive effect of 4‐PSQ and it reinforced the therapeutic potential of this quinolinic compound for acute inflammation.     

Antitumor Effect of Docetaxel‐Loaded Lipid Microbubbles Combined With Ultrasound‐Targeted Microbubble Activation on VX2 Rabbit Liver Tumors

Objective. The purpose of the study was to explore the antitumor effect of docetaxel‐loaded lipid microbubbles combined with ultrasound‐targeted microbubble activation (UTMA) on VX2 rabbit liver tumors. Methods. Docetaxel‐loaded lipid microbubbles were made by a mechanical vibration technique. VX2 liver tumor models were established in 90 rabbits, which were randomly divided into 6 groups, including control, docetaxal‐loaded lipid microbubbles alone, docetaxal alone, docetaxal combined with ultrasound, pure lipid microbubbles combined with ultrasound, and docetaxel‐loaded lipid microbubbles combined with ultrasound (DOC+MB/US). The tumor volume and inhibition rate (IR) of tumor growth were calculated and compared. Apoptosis was detected by terminal deoxyuridine nick end labeling. Proliferating cell nuclear antigen and matrix metalloproteinase 2 (MMP2) protein expression was detected by immunohistochemistry. Caspase 3 and MMP2 messenger RNA (mRNA) expression was detected by in situ hybridization histochemistry. The tumor metastasis rate and survival time of the animals were compared. Results. The IR and apoptotic index of the DOC+MB/US group were the highest among all groups, and the proliferating labeling index was the lowest. Matrix metalloproteinase 2 protein and mRNA expression in the DOC+MB/US group was the lowest among all groups, and caspase 3 mRNA expression in the DOC+MB/US group was the highest. The extensive metastasis rate in the DOC+MB/US group was the lowest, and the survival time of the animals in the DOC+MB/US group was the longest. Conclusions. Docetaxel‐loaded lipid microbubbles combined with UTMA could inhibit the growth of VX2 rabbit liver tumors by deferring proliferation and promoting apoptosis, which may provide a novel targeted strategy for chemotherapy of liver carcinoma.     

Assessment of the biodistribution of an [18F]FDG‐loaded perfluorocarbon double emulsion using dynamic micro‐PET in rats

Perfluorocarbon (PFC) double emulsions loaded with a water‐soluble, therapeutic agent can be triggered by ultrasound in a process known as acoustic droplet vaporization. Elucidating the stability and biodistribution of these sonosensitive vehicles and encapsulated agents is critical in developing targeted drug delivery strategies using ultrasound. [¹⁸F]fluorodeoxyglucose (FDG) was encapsulated in a PFC double emulsion and the in vitro diffusion of FDG was assessed using a Franz diffusion cell. Using dynamic micro‐positron emission tomography and direct tissue sampling, the biodistribution of FDG administered as a solution (i.e. non‐emulsified) or as an emulsion was studied in Fisher 344 rats (n = 6) bearing subcutaneous 9L gliosarcoma. Standardized uptake values (SUVs) and area under the curve of the SUV (AUC_SUV) of FDG were calculated for various tissues. The FDG flux from the emulsion decreased by up to a factor of 6.9 compared with the FDG solution. FDG uptake, calculated from the AUC_SUV, decreased by 36% and 44% for brain and tumor, respectively, when comparing FDG solution vs FDG emulsion (p < 0.01). Decreases in AUC_SUV in highly metabolic tissues such as brain and tumor demonstrated retention of FDG within the double emulsion. No statistically significant differences in lung AUC_SUV were observed, suggesting minimal accumulation of the emulsion in the pulmonary capillary bed. The liver AUC_SUV increased by 356% for the FDG emulsion, thus indicating significant hepatic retention of the emulsion. Copyright © 2013 John Wiley & Sons, Ltd.     

An experimental burn wound‐healing study of non‐thermal atmospheric pressure microplasma jet arrays

In contrast with a thermal plasma surgical instrument based on coagulative and ablative properties, low‐temperature (non‐thermal) non‐equilibrium plasmas are known for novel medicinal effects on exposed tissue while minimizing undesirable tissue damage. In this study we demonstrated that arrays of non‐thermal microplasma jet devices fabricated from a transparent polymer can efficiently inactivate fungi (Candida albicans) as well as bacteria (Escherichia coli), both in vitro and in vivo, and that this leads to a significant wound‐healing effect. Microplasma jet arrays offer several advantages over conventional single‐jet devices, including superior packing density, inherent scalability for larger treatment areas, unprecedented material flexibility in a plasma jet device, and the selective generation of medically relevant reactive species at higher plasma densities. The therapeutic effects of our multi‐jet device were verified on second‐degree burns in animal rat models. Reduction of the wound area and the histology of the wound after treatment have been investigated, and expression of interleukin (IL)‐1α, ‐6 and ‐10 was verified to evaluate the healing effects. The consistent effectiveness of non‐thermal plasma treatment has been observed especially in decreasing wound size and promoting re‐epithelialization through collagen arrangement and the regulation of expression of inflammatory genes. Copyright © 2015 John Wiley & Sons, Ltd.