Lipopolysaccharide-pretreated mesenchymal stem cell-conditioned medium optimized with 10 kDa filter attenuates the injury of H9c2 cardiomyocytes in a model of hypoxia/reoxygenation.

Mesenchymal stem cell-derived conditioned medium (MSC-CM) improves cardiac function, which is partly attributed to the released paracrine factors. Since such cardioprotection is moderate and transient, it is essential that MSC-CM's effective components are optimized to alleviate myocardial injury. To optimize MSC-CM, MSCs were treated with or without lipopolysaccharides (LPSs) for 48 h (serum-free), and the supernatant was collected. Then, LPS-CM (MSC stimulated by LPS) was further treated with LPS remover (LPS Re-CM) or was concentrated with a 10 kDa cutoff filter (10 kDa-CM). Enzyme-linked immunosorbent assay showed that all the pretreatments increased the levels of vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF), and insulin growth factor (IGF) except LPS Re-CM; 10 kDa-CM was superior to the other CMs. Cell Counting Kit-8 displayed that the viability of injured H9c2 cells was enhanced with the increase in the MSC-CM concentration. We also found that the 10 kDa-CM significantly alleviated H9c2 hypoxia/reoxygenation (H/R) injury, as evidenced by the increased Bcl-2/Bax ratio, and decreased the levels of lactate dehydrogenase and cardiac troponin. Transmission electron microscopy (TEM), TdT-mediated dUTP nick-end labelling (TUNEL), and hematoxylin and eosin staining (H&E) confirmed that 10 kDa-CM inhibited H/R-induced H9c2 morphological changes. Proteomic analysis identified 41 differentially expressed proteins in 10 kDa-CM, among which anti-inflammation, proangiogenesis, and antiapoptosis were related to cardiac protection. This study indicates that 10 kDa-CM protects H9c2 cardiomyocytes from H/R injury by preserving most of the protective factors, such as VEGF, HGF, and IGF, in MSC-CM.

Intra-individual comparison of therapeutic responses to vascular disrupting agent CA4P between rodent primary and secondary liver cancers.

AIM: To compare therapeutic responses of a vascular-disrupting-agent, combretastatin-A4-phosphate (CA4P), among hepatocellular carcinomas (HCCs) and implanted rhabdomyosarcoma (R1) in the same rats by magnetic-resonance-imaging (MRI), microangiography and histopathology. METHODS: Thirty-six HCCs were created by diethylnitrosamine gavage in 14 rats that were also intrahepatically implanted with one R1 per rat as monitored by T2-/T1-weighted images (T2WI/T1WI) on a 3.0T clinical MRI-scanner. Vascular response and tumoral necrosis were detected by dynamic contrast-enhanced (DCE-) and CE-MRI before, 1 h after and 12 h after CA4P iv at 10 mg/kg (treatment group n = 7) or phosphate-buffered saline at 1.0 mL/kg (control group n = 7). Tumor blood supply was calculated by a semiquantitative DCE parameter of area under the time signal intensity curve (AUC30). In vivo MRI findings were verified by postmortem techniques. RESULTS: On CE-T1WIs, unlike the negative response in all tumors of control animals, in treatment group CA4P caused rapid extensive vascular shutdown in all R1-tumors, but mildly or spottily in HCCs at 1 h. Consequently, tumor necrosis occurred massively in R1-tumors but patchily in HCCs at 12 h. AUC30 revealed vascular closure (66%) in R1-tumors at 1 h (P < 0.05), followed by further perfusion decrease at 12 h (P < 0.01), while less significant vascular clogging occurred in HCCs. Histomorphologically, CA4P induced more extensive necrosis in R1-tumors (92.6%) than in HCCs (50.2%) (P < 0.01); tumor vascularity heterogeneously scored +~+++ in HCCs but homogeneously scored ++ in R1-tumors. CONCLUSION: This study suggests superior performance of CA4P in metastatic over primary liver cancers, which could guide future clinical applications of vascular-disrupting-agents.​.

Differential diagnosis of gallstones by using hypericin as a fluorescent optical imaging agent.

AIM: To explore the feasibility of using hypericin as an optical imaging probe with affinity for cholesterol for differential fluorescent detection of human gallstones. METHODS: Cholesterol, mixed and pigment stones from cholecystectomy patients were incubated with hypericin or solvent. After 72 h, the stones were analysed for fluorescence (365 nm) and treated with 2-propanol/dimethyl sulfoxide for high performance liquid chromatography (HPLC) analysis. Rats with virtual gallbladder containing human cholesterol, mixed or pigment gallstones (VGHG) received 5 mg/kg hypericin or solvent and VGHG rats with cholesterol stones were given different hypericin doses (5-15 mg/kg). Twelve hours later, the stones were analysed at 365 nm. Biliary excretion and metabolites of hypericin were assessed in common bile duct (CBD) cannulated rats for 9 h using fluorospectrometry, HPLC and matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS). RESULTS: Homogeneous high fluorescence was seen on cholesterol stones either pre-incubated with hypericin or extracted from VGHG rats receiving hypericin. Mixed stones showed a dotted fluorescent pattern, whereas pigment and solvent-treated ones lacked fluorescence. HPLC showed 7.68, 6.65 and 0.08 × 10(-3) M of cholesterol in extracts from cholesterol, mixed, and pigment gallstones, respectively. Hypericin accounted for 2.0, 0.5 and 0.2 × 10(-6) M in that order. On cholesterol stones from VGHG rats receiving different hypericin doses, a positive correlation was observed between dose and fluorescence. In the bile from CBD-cannulated rats, fluorescence represented 20% of the injected dose with two peaks in 9 h. HPLC analysis revealed that hypericin conjugates reached 60% of the peak area. By MALDI-TOF MS, hypericin-glucuronide was detected. CONCLUSION: This study proves the potential use of hypericin for differential fluorescent detection of human gallstones regarding their chemical composition.

[Spatial-temporal pattern and factor diagnoses of agroecosystem health in major grain producing areas of Northeast China: A case study in Jilin Province]. / ä¸œåŒ—ç²®é£Ÿä¸»äº§åŒºå†œä¸šç”Ÿæ€ç³»ç»Ÿå¥åº·æ ¼å±€ä¸Žå› å­è¯Šæ–­­­ä»¥å‰æž—省为例.

It is of great significance to study agroecosystem health in major grain producing areas based on the theory and method of ecosystem health. This paper selected Jilin Province as the study area, and an evaluation index system of agroecosystem health was built based on the SSI-VOR conceptual framework model. Using the optimal combined weights method, comprehensive evaluation assessment, GIS spatial analysis and grey slope similarity incidence models, the spatial-temporal pattern of agroecosystem health and influence factors were analyzed from 2000 to 2011 in Jilin Pro-vince. The results indicated that, temporally, the composite index of agroecosystem health showed a rising trend in Jilin Province from 1995 to 2011, and the agroecosystem health level changed from not healthy to relatively healthy; spatially, the spatial discrepancy of agroecosystem health level tended to become larger, which remained unchanged in central area, while was gradually improved in southeast and west. The main contributors which improved the agroecosystem health level were economic driving force, environmental management and social development, while the main 'dragging' factors were ecological pressure, organization structure and input capacity. Finally, relevant measures were put forward to improve the agroecosystem health condition.

Separate calculation of DW-MRI in assessing therapeutic effect in liver tumors in rats.

AIM: To explore whether the antitumor effect of a vascular disrupting agent (VDA) would be enhanced by combining with an antiangiogenic agent, and whether such synergistic effects can be effectively evaluated with separate calculation of diffusion weighted magnetic resonance imaging (DW-MRI). METHODS: Thirty-seven rats with implanted liver tumors were randomized into the following three groups: (1) ZD6126, a kind of VDA; (2) ZDTHA, ZD6126 in combination with an antiangiogenic, thalidomide; and (3) control. Morphological DW-MRI were performed and quantified before, 4 h and 2 d after treatment. The apparent diffusion coefficient (ADC) values were calculated separately for low b values (ADC(low)), high b values (ADC(high)) and all b values (ADC(all)). The tissue perfusion contribution, ADC(perf), was calculated as ADC(low)-ADC(high). Imaging findings were finally verified by histopathology. RESULTS: The combination therapy with ZDTHA significantly delayed tumor growth due to synergistic effects by inducing cumulative tumor necrosis. In addition to delaying tumor growth, ZDTHA caused tumor necrosis in an additive manner, which was verified by HE staining. Although both ADC(high) and ADC(all) in the ZD6126 and ZDTHA groups were significantly higher compared to those in the control group on day 2, the entire tumor ADC(high) of ZDTHA was even higher than that of ZD6126, but the significant difference was not observed for ADC(all) between ZDTHA and ZD6126. This indicated that the perfusion insensitive ADC(high) values calculated from high b value images performed significantly better than ADC(all) for the monitoring of tumor necrosis on day 2. The perfusion sensitive ADC(perf) derived from ADC(low) by excluding high b value effects could better reflect the reduction of blood flow due to the vessel shutdown induced by ZD6126, compared to the ADC(low) at 4 h. The ADC(perf) could provide valuable perfusion information from DW-MRI data. CONCLUSION: The separate calculation of ADC is more useful than conventional averaged ADC in evaluating the efficacy of combination therapy with ZD6126 and thalidomide for solid tumors.

A single-dose toxicity study on non-radioactive iodinated hypericin for a targeted anticancer therapy in mice.

AIM: Hypericin (Hyp) and its radio-derivatives have been investigated in animal models with ischemic heart diseases and malignancies for diagnostic and therapeutic purposes. Before radioiodinated Hyp ((123)I-Hyp or (131)I-Hyp) can be considered as a clinically useful drug, vigorous evaluations on its chemotoxicity are necessary. In the present study, we examined the toxicity of a single dose of non-radioactive (127)I-Hyp in normal mice for 24 h and 14 d. METHODS: Studies were performed on 132 normal mice. (127)I -Hyp at a clinically relevant dose of 0.1 mg/kg body weight and a 100-times higher dose of 10 mg/kg was intravenously injected into 40 mice. The safety aspects of clinical manifestations, serological biochemistry, and histopathology were assessed. In another 72 mice, (127)I-Hyp was administered intravenously at assumed values to bracket the value of LD(50). The rest 20 mice were used in the control groups. RESULTS: At 24 h and 14 d following the injection of (127)I -Hyp at either 0.1 or 10 mg/kg, all mice tolerated well without mortality or any observable treatment-related symptoms. No significant differences were found in blood biochemical parameters between the test and control groups. All organs presented normal appearances upon histopathological inspection. The value of LD(50) of (127)I-Hyp in mice through intravenous injection was 20.26 mg/kg, with the 95% confidence interval between 18.90 and 21.55 mg/kg. CONCLUSION: The current study reveals a broad safety range of (127)I-Hyp, which not only supports the use of (123)I-Hyp or (131)I-Hyp in the necrosis targeting theragnostic strategy, but also serves as a valuable reference for exploring other possible applications for iodinated Hyp.