bluuebunny/crossref_metadata_2025 huggingface dataset:
Lycopene phytocomplex, but not pure lycopene, is able to trigger apoptosis and improve the efficacy of photodynamic therapy in HL60 human leukemia cells
Authors: Ettorre Anna , Frosali Simona , Andreassi Marco , Di Stefano Anna
Published: April 2011

We compared the ability of pure lycopene (Lyco) versus lycopene phytocomplex (LycoC) to induce apoptosis in vitro. We found that LycoC, but not Lyco, was able to trigger apoptosis in HL60 cells, as documented by subdiploid DNA content and phosphatidylserine exposure. LycoC-induced apoptosis was associated with reactive oxygen species (ROS) generation and loss of mitochondrial transmembrane potential, suggesting that LycoC triggered apoptosis via a mitochondrial pathway. We also verified the redox state of cells by measuring glutathione (GSH) content, but only a small percentage of cells showed GSH depletion, suggesting that the loss of GSH may be a secondary consequence of ROS generation. Moreover, LycoC pretreatment effectively increased apoptosis induced by photodynamic therapy (PDT), a mode of cancer treatment using a photosensitizer and visible light. LycoC pretreatment was even more potent in improving PDT than pretreatment with ascorbic acid or alpha-tocopherol (or the two combined). Our results demonstrate that LycoC has a stronger cytotoxic effect than Lyco and is a better source of agents able to trigger apoptosis in HL60 cells and improve the efficacy of PDT in vitro.
Investigation of toxicological effects of Shuanghuanglian injection in Beagle dogs by metabonomic and traditional approaches
Authors: Yan Guangyan , Zhao Yinglan , Deng Pengchi , Lv Lei , Wang Yanli , Bu Qian , Liu Bin , Hu Chunyan , Zhuo Yanqiang , Yang Xunning , Wang Li , Cen Xiaobo
Published: April 2011

In this study, clinical biochemistry, hematology, histopathology and 1H nuclear magnetic resonance spectroscopy-based metabonomic approaches were applied to investigate the toxicological effects of Shuanghuanglian (SHL) injection after intravenous administration (dosed at 4, 12 and 36 mL stock/kg) in Beagle dogs for 30 d. Decreases in red blood cells, hemoglobin, mean cell volume, mean corpuscular hemoglobin and mean corpuscular hemoglobin concentration were observed in the high-dose group. Elevated reticulocytes, total bilirubin and direct bilirubin were also observed in this group. Moreover, significant hemosiderosis and Prussian blue positivity were detected in the liver, spleen and kidney from high-dose group animals, and transmission electron microscopy examination revealed an appreciable number of acanthrocytes in the liver. These results collectively indicate that SHL injection has the potential to cause hemolytic anemia. Metabonomic analysis showed increases in serum lactate, choline and phosphocholine but a decrease in taurine in treated groups and these findings may underlie the toxicological mechanism of SHL injection. In summary, SHL injection shows hemolytic effects in Beagle dogs; moreover, serum choline and phosphocholine as well as lactate and taurine may be the biomarkers for hemolytic anemia induced by SHL injection.
Acute intermittent hypoxia exposures enhance arterial oxygen delivery
Authors: Zhang Peizhen , Downey H Fred , Shi Xiangrong
Published: April 2011

Physiological adaptations to intermittent hypoxia (IH) conditioning are based on the cumulative effect of repeated IH exposures. The present study sought to test the hypothesis that acute IH exposures would promote arterial O2 delivery and regional tissue oxygenation. Changes in arterial O2 saturation (SaO2, oximeter), forearm muscle and cerebral tissue oxygenations (SmO2 and ScO2, near-infrared spectroscopy) were compared during five repeated hypoxia exposures (10 ± 0.2% O2 for 5-min each) interposed with four-minute inhalation of room air in 11 healthy subjects (24 ± 0.9 y). Baseline, prehypoxia partial pressure of end-tidal O2 (PETO2, mass spectrometer) and SaO2 (107 ± 2 mmHg and 97.3 ± 0.3%) were decreased ( P < 0.05) after the first bout as compared with those during normoxia prior to the second (94 ± 2 mmHg and 96.2 ± 0.4%) and the fifth (92 ± 3 mmHg and 95.7 ± 0.7%) episodes of IH exposures, whereas partial pressure of end-tidal CO2, tidal volume and breathing frequency were similar. Arterial O2 dissociation in terms of per unit decrease in PETO2 during hypoxia, i.e. the slope of SaO2/PETO2, was augmented ( P = 0.0025) from 0.71 ± 0.09%/mmHg during the first hypoxia bout to 1.39 ± 0.15%/mmHg and 1.47 ± 0.16%/mmHg during the second and the fifth bouts, respectively. Fractional muscle tissue O2 extraction rate (SmO2D, i.e. normalized difference between SaO2 and SmO2) progressively decreased ( P < 0.01) during IH; however, fractional cerebral tissue O2 extraction rate (ScO2D, i.e. normalized difference between SaO2 and ScO2) did not decrease during hypoxia ( P = 0.94). ScO2D during normoxia tended to increase ( P = 0.089) following repeated IH exposures. We conclude that enhanced arterial O2 delivery with repeated IH exposures serves as a compensatory mechanism to potentiate O2 availability during hypoxia.
Differential involvement of insulin-like growth factor-I and estrogen on prolactin cells in the mouse anterior pituitary
Authors: Hikake Tamiki , Hayashi Shinji , Chambon Pierre , Watanabe Hajime , Iguchi Taisen , Sato Tomomi
Published: April 2011

Estrogen and insulin-like growth factor-I (IGF-I) stimulate prolactin (PRL) production, release and proliferation of PRL-producing cells (PRL cells) in the anterior pituitary. PRL cells in adult estrogen receptor α (ER α) knockout ( αERKO) mice and IGF-I knockout (IGF-IKO) mice are decreased considerably in number. To investigate a correlation between 17 β-estradiol (E2) and IGF-I on PRL production, IGF-I wild-type (WT) or IGF-IKO mice were ovariectomized at day 8 and the number of PRL cells was examined at days 20 and 60. Although PRL cell number at day 20 and WT or IGF-IKO mice ovariectomized at day 8 was similar to that in intact WT or IGF-IKO mice, PRL cells in adult WT or IGF-IKO mice ovariectomized at day 8 were significantly decreased as compared with those in intact WT or IGF-IKO mice. Therefore, estrogen is essential for PRL cell differentiation between days 20 and 60, regardless of IGF-I. While PRL cells in WT ovariectomized mice increased from days 20 to 60, those in IGF-IKO ovariectomized mice did not increase, suggesting that IGF-I modified PRL cell differentiation after day 20. ICI 182,780 (anti-estrogen) treatment canceled an increase of PRL cells in 30-day-old ovariectomized WT mice, indicating that the presence of ER α is important. The number of PRL cells in αERKO mice was similar to that in WT mice at day 20; however, PRL cells in αERKO mice at day 60 were not increased in number from day 20, supporting the idea that estrogen is essential for PRL cell differentiation after day 20. Finally, the percentage of PRL cells in IGF-IKO mice was decreased as compared with that in WT mice at day 20; therefore, IGF-I affects PRL cells before day 20. In conclusion, PRL cell differentiation is differently regulated by E2 and IGF-I depending on the age.
A combination of the metabolic enzyme inhibitor APO866 and the immune adjuvant L-1-methyl tryptophan induces additive antitumor activity
Authors: Yang Huei-Jiun , Yen Meng-Chi , Lin Chi-Chen , Lin Chiu-Mei , Chen Yi-Ling , Weng Tzu-Yang , Huang Tzu-Ting , Wu Chao-Liang , Lai Ming-Derg
Published: April 2011

Many types of malignant cells have a higher nicotinamide adenine dinucleotide (NAD) turnover rate than normal cells, as well as the ability to escape immune responses. Indoleamine 2,3-dioxygenase (IDO) is reported to be a negative immune regulator. Overexpression of IDO in dendritic cells is observed in tumor-draining lymph nodes. IDO-expressing dendritic cells suppress T-cell activation and promote immune tolerance. The nicotinamide phosphoribosyl transferase (NAMPT) inhibitor APO866 (also called FK866 or WK175) selectively inhibits tumor growth through intracellular NAD depletion. The IDO-specific inhibitor L-1-methyl-tryptophan (L-1MT) activates immune responses and reduces tumor volume in murine tumor models. We combined L-1MT and APO866 treatments and tested their antitumor effects in the murine gastric and bladder tumor models. In immune-competent mice, a combination of APO866 and L-1MT had a better therapeutic effect than did either L-1MT or APO866 alone. The intracellular level of NAD was suppressed by APO866 but not L-1MT. However, an additive inhibitory effect on tumor growth was not observed in tumor-bearing immune-deficient mice. The new strategy of combining a metabolic inhibitor and an immune adjuvant induced a potent therapeutic effect.
Human cardiac explant-conditioned medium: soluble factors and cardiomyogenic effect on mesenchymal stem cells
Authors: Schittini Andressa V , Celedon Paola F , Stimamiglio Marco A , Krieger Marco , Hansen Paula , da Costa Francisco Diniz Affonso , Goldenberg Samuel , Dallagiovanna Bruno , Correa Alejandro
Published: April 2011

The use of conditioned medium (CM) from human cardiac explants (HCEs) as a potential source of paracrine factors for adult stem cell signaling has never been evaluated. We hypothesized that HCEs might provide a source of soluble factors triggering the differentiation of mesenchymal stem cells (MSCs) into cardiomyocyte-like cells. By using two-dimensional electrophoresis (2-DE) gels/mass spectrometry and antibody macroarray assays, we found that HCEs release macromolecules, including cytokines, growth factors and myocardial and metabolism-related proteins into the culture medium. We identified a total of 20 proteins in the HCE-CM. However, as shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and 2-DE, these 20 proteins account for only a fraction of the total number of proteins present in the HCE-CM. We also found that CM increased the proliferation of bone marrow-derived-MSCs (BM-MSCs) in vitro. Unlike the other effects, this effect was most evident after 48 h of culture. Moreover, we examined the effect of HCE-CM on levels of mRNA and protein for specific cardiac markers. We showed that a surprisingly big fraction of BM-MSCs (3.4–5.0%) treated in vitro with HCE-CM became elongated and began to express cardiac markers, consistent with their possible differentiation into cardiomyocyte-like cells. Our in vitro model may be useful not only per se, but also for studies of the mechanisms of action of soluble factors involved in cell differentiation, paving the way for possible new protein-based treatments in the future.
Vitamin D nutritional policy needs a vision for the future
Authors: Norman Anthony W , Bouillon Roger
Published: April 2011

Historically vitamin D is known to be essential for normal bone growth and quality, and thus appropriate dietary vitamin D supplementation can eliminate vitamin D deficiency childhood rickets and adult osteomalacia. In spite of many government and medical associations' worldwide guidelines for the reference daily intake (RDI) of vitamin D, scientists and nutritionists from many countries agree that at present about half of elderly North Americans and Western Europeans and probably also of the rest of the world are not receiving enough vitamin D to maintain healthy bone. In addition, over the past decade there has been a dramatic increase in our understanding of the many biological actions that result from vitamin D acting through its daughter steroid hormone, 1 α,25-dihydroxyvitamin D3[1 α,25(OH)2D3] in collaboration with its cognate vitamin D receptor (VDR). Consequently, evidence has accumulated that beside intestine and bone, there are five additional physiological systems where the VDR with 1 α,25(OH)2D generates biological responses. These include the immune system (both the innate and adaptive), pancreas and metabolic homeostasis, heart-cardiovascular, muscle and brain systems as well as the control of the cell cycle, and thus of the disease process of cancer. Acting through the VDR, 1 α,25(OH)2D3can produce a wide array of favorable biological effects that collectively are projected to contribute to the improvement of human health. Responsible medicine demands that worldwide vitamin D nutritional guidelines reflect current scientific knowledge about vitamin D's spectrum of activities. Thus, worldwide vitamin D nutritional policy is now at a crossroads. This paper presents several proposed policy changes with regard to the amount of vitamin D daily intake that if implemented will maximize vitamin D's contribution to reducing the frequency of many diseases, which would then increase the quality and longevity of life and significantly reduce the cost of medical care worldwide.
Basal and induced granulopoiesis in outbred, F1 hybrid and inbred mice: can inbreeding depression influence the experimental practice?
Authors: Hofer Michal , Pospíšil Milan , Dušek Ladislav , Holá Jiřina , Hoferová Zuzana , Weiterová Lenka
Published: April 2011

In this study we examined differences in selected indices of granulopoiesis in outbred, F1 hybrid and inbred mouse strains. Specifically, serum granulocyte colony-stimulating factor (G-CSF) levels, numbers of marrow granulocyte–macrophage progenitor cells and morphologically recognizable proliferative marrow granulocytic precursor cells were evaluated. These parameters were determined in untreated controls, and in mice exposed either to a non-specific stimulus (injection of saline) or to a granulopoiesis-enhancing stimulus (administration of a cyclooxygenase-2 inhibitor, meloxicam). Lower levels of G-CSF were detectable in the outbred ICR mice, which also demonstrated an enhanced response to both types of the stimuli. Considering the fact that outbred mice are closer to natural mammalian populations, including human ones, the possibility of using outbred mice, instead of the often used inbred strains, for experiments evaluating the effects of pharmacological interventions on hematopoiesis should be investigated.
3,3′-Dioctadecylindocarbocyanine-low-density lipoprotein uptake and flow patterns in the rabbit aorta–iliac bifurcation under three perfusion flow conditions
Authors: Ding Zufeng , Fan Yubo , Deng Xiaoyan , Sun Anqiang , Kang Hongyan
Published: April 2011

The aim of this study was to elucidate which of the following two factors plays a more important role in the localization of atherogenesis: the barrier function of the arterial endothelium modulated by wall shear stress or flow-dependent low-density lipoprotein (LDL) concentration at the blood/wall interface. To determine this, the rabbit aorto-iliac bifurcation was perfused with 3,3′-dioctadecylindocarbocyanine (DiI)-LDL solution under three different flow conditions: (i) forward flow (perfused in the in vivo flow direction); (ii) backward flow (perfused in a reversed flow direction); and (iii) static group (no flow). The results showed that there was a peak in the curve of DiI-LDL uptake distribution along the lateral wall of the bifurcation for all three groups, which was located in the branching areas where the endothelial cells were round and polygonal with no preferred orientation. Nevertheless, the peak of the forward flow group was much sharper than those of the other two groups. The overall DiI-LDL uptake was the highest for the static group. The present experimental study supports the concept that both the barrier function of the endothelium modulated by wall shear stress and the mass transport phenomenon of LDL concentration polarization are involved in the infiltration/accumulation of atherogenic lipids within the arterial wall. Nevertheless, the latter might play a larger role in the localization of atherogenesis.
Increases in energy intake, insulin resistance and stress in rats before Wenchuan earthquake far from the epicenter
Authors: Chen Lu-Lu , Hu Xiang , Zheng Juan , Zhang Hao-Hao , Kong Wen , Yang Wei-Hong , Zeng Tian-Shu , Zhang Jiao-Yue , Yue Ling
Published: April 2011

The study of pre-earthquake (PE) behavior in animals has always been shrouded by controversy. There is very little scientific evidence showing that animals can sense the coming of an earthquake and that their organisms undergo physiological changes during the PE period. On the day of the Wenchuan earthquake, prior to the time of its actual occurrence, we were coincidentally able to measure the insulin sensitivity and stress level in rats that were originally part of another study. We detected defects in insulin signaling and a decrease in glucose uptake in skeletal muscle (SkM) and adipose tissue (AT), indicating impaired insulin sensitivity. These changes were associated with significantly increased plasma corticosterone concentration and elevated HSD11B1 mRNA expression in SkM and AT. The increase in insulin resistance (IR) could be attributed to elevated local (SkM and AT) and systemic stress. Interestingly, we also noticed that the food intake in rats showed a sudden increase two days before the earthquake and reached a peak on the day of the earthquake itself. Our observations suggest the possibility that the rats underwent PE physiological changes consisting of an increase in the stress level and consequently leading to an increase in food intake and IR.
Delivery of TFPI-2 using SonoVue and adenovirus results in the suppression of thrombosis and arterial re-stenosis
Authors: Wang Yuxue , Zhou Jie , Zhang Yanrong , Wang Ximing , Chen Juan
Published: April 2011

Genes could be used to treat atherosclerosis. The key problem is how to target a gene through the walls of arteries in free-flowing blood. TFPI-2 has been shown to suppress thrombosis and arterial re-stenosis, which indicates its potential function in gene therapy for atherosclerosis. The microbubble ultrasound contrast agent is widely applied in diagnostic imaging, and could be used for transferring genes into arteries. By transfecting TFPI-2 into arteries using SonoVue (a kind of microbubble ultrasound contrast agent), we identified TFPI-2 as an available factor for inhibiting the proliferation of vascular endothelial cells in vivo. Compared with adenovirus, SonoVue showed similar gene transfection efficiency, but the latter showed stronger inhibition of thrombosis and arterial re-stenosis with a high expression of TFPI-2 protein in vitro and in vivo. SonoVue was less damaging when transfecting genes into the arterial wall. These data indicate that transfecting human TFPI-2 into the arterial wall may suppress thrombosis and arterial re-stenosis, and reduce atherosclerosis.
Renal and metabolic effects of three months of decarbonated cola beverages in rats
Authors: Celec Peter , Pálffy Roland , Gardlík Roman , Behuliak Michal , Hodosy Július , Jáni Peter , Božek Peter , Šebeková Katarína
Published: April 2011

Epidemiological studies have shown an association between the intake of cola beverages and chronic kidney diseases. Experimental evidence for the negative effects of cola intake on kidneys is lacking. Male Wistar rats had ad libitum access to water (control group) or three different sugar-sweetened cola beverages for three months. Despite very high cola intake (daily cca 140 mL), no differences were found in body weight, kidney weight, glomerular morphology, oxidative and carbonyl stress or expression of selected marker genes in the renal cortex. Interestingly, all groups consuming cola beverages had lower blood glucose levels during an oral glucose tolerance test, suggesting improved insulin sensitivity. Despite hyperfiltration (5–6-fold increase in diuresis), cola beverages had no effect on assessed parameters of renal function, histology, gene expression or oxidative stress. Moreover, cola intake seems to increase creatinine clearance and to decrease plasma levels of urea. In our study increased insulin sensitivity and altered renal functional parameters were observed in rats receiving cola beverages for three months. Whether the findings are due to the short duration of the study or interspecies metabolic differences should be uncovered in further studies. Even more interesting might be the analysis of effects of cola intake in animal models of diabetes.
Alterations of bone marrow-derived endothelial progenitor cells following acute pulmonary embolism in mice
Authors: Wan Jun , Lu Li-Jin , Miao Ran , Liu Jie , Xu Xiao-Xue , Yang Ting , Hu Qing-Hua , Wang Jun , Wang Chen
Published: April 2011

Pulmonary embolism (PE) is a common, lethal, ischemic disease. PE-induced endothelium injury plays a critical role in the pathophysiological consequences of PE. Endothelial progenitor cells (EPCs) can be mobilized from the bone marrow to enter circulation and play important roles in repair of damaged endothelium. However, it is not yet known if EPC mobilization results from PE. The alterations of the quantity and function of bone marrow-derived EPCs were detected in acute pulmonary embolism (APE) events in mice, and the possible role of the endothelial nitric oxide synthase (eNOS)/nitric oxide (NO) pathway in those alterations was explored. APE models were established by injection of autologous thrombi into the right jugular vein of C57BL/6 mice. Mice were divided into sham and experimental groups including one hour (1H), one day (1D) and two day (2D) groups after injection. The results showed that in the APE 1D group, the thrombi were easily found in the large or medium pulmonary vessel. And CD133+ or CD34+ cells in bone marrow increased significantly, while CD133+/vascular endothelial growth factor receptor 2+ EPCs decreased. After seven days in culture, the abilities of incorporation into a vascular network, adhesion to fibronectin, migration and proliferation of bone marrow-derived EPCs in the APE 1D group increased significantly. The mRNA and protein expression levels of eNOS in EPCs increased in the APE 1D group. Treatment of EPCs with NG-nitro-L-arginine methyl ester inhibited functional alterations induced by APE. The results suggested that APE events stimulate the mobilization of EPCs from bone marrow, and enhance their functions. The eNOS/NO pathway may be involved in this process.
Study on ASTC-a-1 cells labeled with superparamagnetic iron oxide and its magnetic resonance imaging
Authors: Yu Ming-Xi , Chen Wen-Li , Zhou Quan , Gao Peng
Published: April 2011

The aim of this preliminary study is to explore the feasibility of incorporating superparamagnetic iron oxide (SPIO) with poly-l-lysine (PLL) for labeling and magnetic resonance imaging (MRI) of human lung adenocarcinoma cells. Endorem (5–200 μg/mL) was incubated with ASTC-a-1 cells in the presence of PLL (1.5 μg/mL) for 0.5–24 h. The presence of SPIO in labeled cells was examined by Prussian blue stain. The effects of SPIO on cell proliferation, reactive oxygen species (ROS) generation and the mitochondria were also examined. In vitro MRI of SPIO-PLL-labeled cells was performed using a clinical 1.5 T MRI system. The labeling efficiency of >99% could be achieved after incubating for 0.5 h with 25 μg/mL of SPIO in the presence of PLL (1.5 μg/mL). Higher concentrations of SPIO (e.g. >50 μg/mL) could induce significant cell death, which might be mediated by changes in intracellular ROS level and mitochondrial membrane potential. In vitro MRI showed the decrease in MRI signal intensity on T1WI, T2WI and T2*WI sequences. In conclusion, MRI can trace SPIO-labeled cancer cells according to changes in T1WI, T2WI and T2*WI sequences. It should be noted that at higher concentrations, SPIO can cause cell damage.
Essential role of HDL on endothelial progenitor cell proliferation with PI3K/Akt/cyclin D1 as the signal pathway
Authors: Zhang Qiuhua , Yin Hongchao , Liu Peimao , Zhang Hua , She Mingpeng
Published: April 2011

High-density lipoprotein (HDL) is known as an important factor in vascular wall remodeling that also affects gene expression in cell proliferation and differentiation. In this article, the role of HDL on endothelial progenitor cell (EPC) proliferation, angiogenesis and the signal pathway involved was studied, particularly the influence of HDL in strengthening the promoting effect of EPCs on wound healing of the arterial wall in hypercholesterolemic rats. Mononuclear cells isolated from rat bone marrow displayed characteristics of EPCs after cultivation. The role of HDL on EPC function and the signal pathway involved were studied by Western blotting, in vitro migration and ‘tube’ formation. Re-endothelialization and the number of circulating EPCs were compared between normal rats, hypercholesterolemic rats and hypercholesterolemic rats with HDL treatment. Results showed that HDL participated in the healing process by promoting EPC proliferation, migration and ‘tube’ formation. HDL activates cyclin D1 via phosphatidylinositol 3-kinase (PI3K)/Akt stimulation. Inhibition of PI3K/Akt via pharmacological or small interfering RNA approaches significantly attenuated HDL-induced EPC migration, proliferation and ‘tube’ formation. Results of experiments in vivo showed that HDL increased the number of circulating EPCs and promoted re-endothelialization in wound healing. These findings demonstrate for the first time that PI3K/Akt-dependent cyclin D1 activation plays an essential role in HDL-induced EPC proliferation, migration and angiogenesis.
Enhanced Gadd45 expression and delayed G2/M progression are p53-dependent in zinc-supplemented human bronchial epithelial cells
Authors: Shih Rita S M , Wong Stephen H K , Schoene Norberta W , Zhang Jun Jun , Lei Kai Y
Published: April 2011

Zinc is an essential nutrient for humans; however, this study demonstrated for the first time that an elevated zinc status, created by culturing cells at optimal plasma zinc concentration attainable by oral zinc supplementation, is cytotoxic for normal human bronchial epithelial (NHBE) cells. p53 plays a central role in the modulation of cell signal transduction in response to the stress from DNA damage, hypoxia and oncogene activation. The present study was designed to determine whether the previously reported increased Gadd45 expression and delayed G2/M cell cycle progression in zinc-supplemented NHBE cells is p53-dependent, and to decipher the mechanisms responsible for the regulation of Gadd45 expressions by p53, and elucidate the Gadd45 functions in impaired cell growth and cell cycle progression in NHBE cells. Cells were cultured for one passage in different concentrations of zinc: <0.4 μmol/L (ZD) as severe zinc-deficient; 4 μmol/L (ZN) as normal zinc level in culture medium; 16 μmol/L (ZA) as normal human plasma zinc level; and 32 μmol/L (ZS) as the high end of plasma zinc attainable by oral supplementation. Inhibition of cell growth and upregulation of p53 mRNA and protein expression were observed in ZS cells. Most importantly, ZS treatment also enhanced Gadd45 nuclear protein level and promoter activity, decreased CDK1-Cyclin B1 level and delayed G2/M cell cycle progression. These changes were normalized to those observed in ZN by treating ZS cells with Pifitherin, an inhibitor of p53 transactivation activity. Thus, our findings support the p53 dependency of the Gadd45-CDK1/Cyclin B1-G2/M cell cycle progression pathway in ZS NHBE cells.
Wistar-Kyoto and spontaneously hypertensive rat blood pressure after embryo transfer into different wombs and cross-suckling
Authors: Lee Jong Y , Azar Silvia H
Published: April 2011

Blood pressure (BP) varies based on genetic and environmental factors. To test genetic and environmental influences on body weight (BW) and BP, one-cell homozygous embryos were transferred into spontaneously hypertensive (SHR, pup:shr) or (Wistar-Kyoto normotensive [WKY], pup:wky) normotensive rats' oviducts (embryos: s,w; oviduct-uterine: S,W), cross-suckled at birth (nurses S,W) and weaned to normal diets at day-21. BP at day-120 was measured by radiotelemetry and analyzed by methods of linear least square rhythmometry and analysis of variance. Genetics dominantly affected shr BP, causing it to be significantly higher at birth (24.6 ± 1.8 in sS versus 21.8 ± 1.7 mmHg in wW, P < 0.005), and at day-120 (198 ± 0.5 in sSS versus 127 ± 0.2 mmHg in wWW, P < 0.001), with lower BW than those of wky (5.3 ± 0.2 versus 5.7 ± 0.2 g at birth, 332 ± 5 versus 404 ± 6 g at day-120, both P < 0.001). Surprisingly, uterine-suckling milieus lowered shr BP significantly at day-120 (198 ± 0.5 in sSS versus 178 ± 0.5, 147 ± 0.6, 179 ± 0.5 mmHg in sSW, sWS, sWW, respectively, all P < 0.01). BP was slightly elevated when wky-genetics were implanted into the S-uterine by 4 mmHg (wSW, P < 0.05), whereas implanting shr embryos into the W-uterine environment (sWS) lowered BP by 51 mmHg ( P < 0.001). In summary, the hypertensive shr-strain showed significantly lower BP when provided with an WKY-uterine environment and/or by WKY-nursing mothers, indicating that environment can modify genetic influences; yet the shr MESORs (rhythm-adjusted 24-h mean: midline estimating statistic of rhythm) lowered by WKY environments remained above MESORs encountered in wky-donors.
Matrix metalloproteinases are possible mediators for the development of alimentary tract mucositis in the dark agouti rat
Authors: Al-Dasooqi Noor , Gibson Rachel J , Bowen Joanne M , Logan Richard M , Stringer Andrea M , Keefe Dorothy M
Published: April 2011

Alimentary tract (AT) mucositis is a serious and debilitating side-effect of cancer therapy primarily characterized by damage of the mucous membranes throughout the AT. It is well established that this damage is a result of up-regulation of stress response genes and pro-inflammatory cytokines. Matrix metalloproteinases (MMPs) have been shown to function in several of the pathways known to be up-regulated in mucositis and play a key role in tissue injury and inflammation in many gastrointestinal disorders. This study aims to characterize the expression of multiple MMPs including MMP-1, -2, -3, -9 and -12 and their inhibitors, tissue inhibitor of metalloproteinase (TIMP)-1 and -2, in a rat model of irinotecan-induced mucositis. Dark agouti rats were administered a single 200 mg/kg intraperitoneal dose of irinotecan and killed at 1, 6, 24, 48, 72, 96 and 144 h following treatment. Hematoxylin and eosin staining, immunohistochemistry and realtime polymerase chain reaction were used to assess histopathological damage and MMP expression in the jejunum and colon. Marked histopathological evidence of mucositis was observed in the jejunum and colon as early as six hours following irinotecan treatment. A significant alteration in both gene expression and tissue levels of MMPs and TIMPs was noted following irinotecan. The increase in MMP-2, -3, -9 and -12 levels was associated with inflammatory infiltrate and maximum tissue damage. In contrast, MMP-1 expression correlated with tissue restitution. TIMP-1 and -2 levels were significantly altered in the jejunum following irinotecan. The augmentation in the expression profiles of MMPs and their inhibitors correlated with histopathological alterations observed in the tissue following irinotecan. This prompts the consideration of MMPs as possible mediators of chemotherapy-induced mucositis.
Differential modulation of angiogenesis by advanced glycation end products
Authors: Devi Manju S , Sudhakaran Perumana R
Published: April 2011

Divergent angiogenic responses occur in different organs in a diabetic state. Many of the pathological effects were mediated by the advanced glycation end products (AGEs) of non-enzymatically glycated molecules. Investigations were carried out using different angiogenic model systems to examine whether the angiogenic response to AGEs is influenced by the cellular microenvironment. AGE-albumin increased angiogenesis in chick chorioallantoic membrane (CAM). It also increased sprouting in rat aortic rings and the expression of angiogenic markers CD31 and E-selectin and the angiogenic growth factor, vascular endothelial growth factor (VEGF) in human umbilical vein endothelial cells (HUVECs) in culture, suggesting a proangiogenic effect. But in a serum-supplemented condition, AGE-albumin inhibited aortic sprouting and expression of angiogenic markers and VEGF production by HUVECs, suggesting an antiangiogenic effect in the presence of serum. Blocking of the AGE effect by the antioxidants, N-acetyl cysteine and ascorbic acid, suggested that the AGE effect involved oxidant stress. Reversal of the AGE effect by LY 294 002, an inhibitor of the Akt pathway and increased phosphorylation of Akt in cells maintained in serum-free medium, suggested the involvement of the Akt pathway in mediating the AGE effect; such an effect was absent in a serum-supplemented condition. These opposing effects of AGE-albumin on angiogenesis in the presence and absence of serum suggested that the AGE accumulated in a hyperglycemic condition can affect angiogenesis depending on the microenvironment of the cells.
Digital analysis of hepatic sections in mice accurately quantitates triglycerides and selected properties of lipid droplets
Authors: Ge Fengxia , Lobdell Harrison , Zhou Shengli , Hu Chunguang , Berk Paul D
Published: April 2011

We describe a method for the histologic evaluation of lipid accumulation in the livers of various mouse models of hepatic steatosis based on quantitative digital analysis of Oil Red O (ORO) accumulation in fresh-frozen hepatic sections. The process involves two principal steps: identification and digital photographic imaging of areas appropriate for analysis, followed by digital determination of the fraction of the identified area (Area Fraction) exhibiting ORO staining. The Area Fraction, designated the Digital Steatosis Score, is a valuable aspect of the histologic assessment of the liver, especially in various forms of alcoholic and non-alcoholic liver diseases. The method is rapid, requiring ∼3 min per specimen, and highly reproducible, avoiding the inevitably subjective, semi-quantitative evaluation of lipid content inherent in visual steatosis scoring systems. In normal mice and in six different mouse models of fatty liver, the Area Fraction was highly correlated with hepatic triglyceride content ( P < 0.01). The coefficient of variation of repeated determinations of the Area Fraction by two different observers was ±6.4%. If made available in clinical settings, rapid, accurate quantitation of liver triglycerides by this method could be very useful in specific conditions such as assessment of donor livers for transplantation.
Association of vascular endothelial growth factor −634C/G polymorphism and diabetic retinopathy in type 2 diabetic Han Chinese
Authors: Yang Ying , Andresen Bradley T , Yang Ke , Zhang Ying , Li Xiaojin , Li Xianli , Wang Hui
Published: April 2011

Genetic factors are important in the pathogenesis of diabetic retinopathy (DR); there is a clear association of increased expression of vascular endothelial growth factor (VEGF) with DR as well as numerous VEGF polymorphisms that are linked to increased VEGF levels and DR. In this study, the relationships between the VEGF promoter polymorphism −634C/G, plasma VEGF levels and DR were examined in the Han Chinese. Ninety-six healthy subjects and 285 subjects with type 2 diabetes were enrolled in this study. The diabetic subjects were divided into three groups depending on the degree of DR as determined by fundus photography and fluorescent angiography. Along with standard clinical characteristics, the −634C/G polymorphism was examined using TaqMan allelic discrimination, and plasma VEGF levels were analyzed by enzyme-linked immunosorbent assay. The distribution of the polymorphism differed significantly between patients with and without retinopathy; this was most pronounced between the no DR and proliferative DR groups. Significantly greater plasma VEGF levels were present in those with the −634CC genotype, and only the proliferative DR group had elevated plasma VEGF levels. Logistic regression revealed that the −634C/G polymorphism is strongly associated with DR. This study suggests that diabetic Han Chinese carrying the −634CC VEGF promoter polymorphism have a genetic risk of DR, and this polymorphism may be a major factor influencing plasma VEGF levels. Therefore, this polymorphism may be used as a biomarker at the onset of diabetes in the Han Chinese to predict the risk of DR, allowing for clinicians to treat these patients more aggressively.
Contact by melanoma cells causes malignant transformation of human epithelial-like stem cells via alpha V integrin activation of transforming growth factor β1 signaling
Authors: Sun Hongyu , Hu Kaimeng , Wu Minjuan , Xiong Jun , Yuan Li , Tang Ying , Yang Yongji , Liu Houqi
Published: April 2011

The embryonic microenvironment is known to suppress the tumorigenic phenotype of aggressive cancer cells; however, the effects of tumorigenic microenvironments on stem cells have not been sufficiently explored due to the lack of suitable model systems. In order to study the tumorigenic microenviornment, we developed a novel in vitro model system for induction of malignant transformation of human epithelial-like stem cells (hEpSCs), involving co-cultivation and close contact of hEpSCs with the A375 melanoma cell line, together with mutagen treatment of hEpSCs with dimethylbenzanthracene (DMBA). Both factors (close contact and mutagen treatment) were required to transform hEpSCs in vitro and cause phenotypic changes characteristic of epithelial to mesenchymal transition (EMT), including colony formation, decreased E-cadherin and increased N-cadherin and vimentin expression. Direct contact between tumor cells and hEpSCs treated with DMBA increased integrin alpha V (ITGAV gene) expression and caused local activation of the transforming growth factor (TGF)-β1/Smad signaling pathways in hEpSCs. The novel model system described here is being used to elucidate the microenvironmental factors and biological mechanisms involved in the induction of neoplastic progression in hEpSCs in vitro by A375 melanoma cells. A better understanding of the molecular mechanisms by which melanoma cells exert these effects on hEpSCs may open up new avenues for therapeutic and preventive cancer interventions.
Molecular biomarker panels for assessment of selenium status in rats
Authors: Sunde Roger A
Published: April 2011

Molecular biomarkers are mRNA transcripts that indicate the (nutrient) status of an organism or tissue. Molecular biomarker panels have the potential to readily and more accurately determine nutrient status than individual traditional biomarkers. To study the efficacy of molecular biomarker panels for predicting selenium (Se) status, we examined 30 biomarkers from rats fed graded levels of Se from deficient to eight times the minimum Se requirement, including four liver and four kidney traditional biomarkers, and 13 liver and nine kidney selenoprotein mRNA levels. Multiple regression analysis against liver and kidney Se and glutathione peroxidase-1 (Gpx1) activity, with stepwise single elimination of biomarkers that did not significantly contribute, was used to identify biomarker panels with significant ( P < 0.05) regression coefficients. Resulting regression equations were then used to predict Se status, and compared with traditional Se biomarkers panels. Over the full spectrum of Se status from 0 to 0.8 μg Se/g diet, the resulting 4-selenoprotein mRNA biomarker panel predicted liver Se concentration with a correlation of 0.948, which was nominally higher and statistically the same as the correlation of 0.909 for the panel based on Gpx1 activity. The molecular biomarker panels for predicting kidney Se and liver and kidney Gpx1 activity were all comparable to predictions based on traditional biomarkers. These analyses show that molecular biomarker panels can be used to predict accurately two traditional biomarkers of Se status. The resulting analyses also illustrate that additional orthogonal biomarkers reflecting higher Se intakes are needed to better predict supernutritional Se status and further strengthen this approach.
The influence of a 6 mT static magnetic field on apoptotic cell phagocytosis depends on monocyte/macrophage differentiation
Authors: Dini Luciana , Panzarini Elisa
Published: April 2011

In a previous work we showed that a 6 mT static magnetic field (SMF) interferes with monocyte/macrophage 12- O-tetradecanoylphorbol-13-acetate (TPA)-induced differentiation of promonocytes (U937 cells) and monocytes (THP-1 cells). In this study we investigated whether in the same cells and under the same conditions, phagocytosis of apoptotic cells is influenced by 6 mT SMF exposure. Fluid phase endocytosis and phagocytosis of latex particles were also analyzed for comparison. The results indicate that SMF exposure has effects on phagocytosis but not on fluid phase endocytosis, and that these effects are greater at the late stages of macrophage differentiation (THP-1 > U937 cells). The phagocytosis index and rate of phagocytosis decreased under SMF exposure while the number of latex particles bound to the plasma membrane of TPA-differentiated U937 and THP-1 cells increased. Conversely, the rate of phagocytosis of apoptotic cells increased under SMF exposure, while the number of apoptotic cells bound to the plasma membrane of isolated human Kupffer cells, Raw 264.7 macrophages and TPA-differentiated THP-1 and U937 cells decreased. In non-differentiated U937 and THP-1 cells, the SMF exposure enhanced the number of cell-surface bound apoptotic cells and latex beads.
Isolation and characterization of the antibreast carcinoma cell growth components of Vernonia amygdalina extracts
Authors: Luo Xuan , Oyugi Daniel A , Lin Cuiwu , Izevbigie Ernest B , Lee Ken S
Published: April 2011

Vernonia amygdalina (VA) is widely used for medicinal and food purposes in tropical Africa. Many health benefits (antioxidant, antimicrobial, anticancer activities and more) of VA extracts have been reported. The mechanisms of actions have also been described. We have previously reported that VA extracts elicited growth inhibitory activities in human estrogen receptor-positive (ER+) cells (MCF-7 cells) and ductal carcinoma cells (BT-549) in vitro. The active components in the organic solvent (chloroform)-extracted VA have been previously determined. However, the active components in the ethanolic extracts of VA have not been previously studied. Hence, the objectives of this study are to isolate and characterize the active components of the ethanolic extracts of VA using liquid–liquid extraction, thin layer chromatography and column techniques. Fractionation of the ethanolic extracts of VA yielded three fractions named A1, A2 and A3, and A2 retained the DNA synthesis-inhibitory activity of the extracts. Subsequent fractionation of A2 yielded fraction A2B whose activity was 16 and three times more potent than the ethanolic fraction and fraction A2, respectively. The treatment of cells with 100 μg/mL of either the ethanolic VA extracts, fraction A2 or fraction A2B resulted in a 23% ( P < 0.01), 86% ( P < 0.0001) and 97% ( P < 0.0001) inhibition of DNA synthesis compared with vehicle-treated controls, respectively. Further purification of A2B by high-speed countercurrent chromatography and confirmed by spectroscopic analysis revealed that the major active components of A2B (65% by weight) were steroid glucosides.
Identification of fetal hemoglobin-inducing agents using the human leukemia KU812 cell line
Authors: Zein Sima , Li Wei , Ramakrishnan Valya , Lou Tzu-Fang , Sivanand Sharanya , Mackie Ashley , Pace Betty
Published: April 2011

Fetal hemoglobin (HbF) ameliorates the clinical severity of sickle cell disease; therefore continued research to identify efficacious HbF-inducing agents is desirable. In this study, we investigated KU812 leukemia cells that express the fetal γ-globin and adult β-globin genes, as a system for screening and discovery of novel HbF inducers. KU812 cells were analyzed in the presence or absence of fetal bovine serum and then expression levels of the globin genes, cell surface markers and transcription factors were quantified by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). For comparison, primary erythroid cells were grown in a two-phase liquid culture system. After drug inductions for 48–72 h, globin mRNA and HbF levels were quantified by RT-qPCR and enzyme-linked immunosorbent assay, respectively. Erythroid markers and transcription factors expression levels in KU812 cells were comparable to days 7–14 erythroid cells. We also tested several drugs including butyrate, trichostatin A, scriptaid, suberoylanilide hydroxamic acid and hydroxyurea, which induced γ-globin in KU812 cells; however, some agents also induced β-globin. A novel agent STI-571 was studied in the system, which non-selectively induced the globin genes. Additional studies showed comparable globin gene response patterns in KU812 and primary erythroid cells after treatments with the various drug inducers. Mechanisms of drug-mediated γ-globin induction in KU812 cells require signaling through the p38 mitogen-activated protein kinase pathway similar to that previously demonstrated in primary erythroid cells. These data suggest that KU812 cells serve as a good screening system to identify potential HbF inducers for the treatment of β-hemoglobinopathies.
Tracking deposition of a 14C-radiolabeled kudzu hairy root-derived isoflavone-rich fraction into bone
Authors: Mun Jonathan G , Grannan Michael D , Lachcik Pamela J , Rogers Randy B , Yousef Gad G , Grace Mary H , Janle Elsa M , Wu Qing Li , Simon James E , Weaver Connie M , Lila Mary Ann
Published: April 2011

Hairy roots were induced in four genotypes from three kudzu species ( Pueraria montana var. lobata, P. lobata and P. phaseoloides) in vitro using Agrobacterium rhizogenes to stimulate rapid secondary metabolite synthesis. Hairy roots from P. montana var. lobata (United States Department of Agriculture no. PI 434246) yielded the highest puerarin and total isoflavone content and the greatest new biomass per growth cycle among the genotypes evaluated. Hairy roots from this genotype were selected for radiolabeling using 14C-sucrose as a carbon source. Isoflavones from radiolabeled kudzu hairy root cultures were extracted with 80% methanol, partitioned by solvent extraction, and then subfractionated by Sephadex LH-20 gel filtration. Radiolabeled isoflavones were isolated in a highly enriched fraction, which contained predominantly puerarin, daidzin and malonyl-daidzin and had an average radioactivity of 8.614 MBq/g (232.8 μCi/g) dry fraction. The 14C-radiolabeled, isoflavone-rich fraction was orally administered at a dose of 60 mg/kg body weight to male Sprague–Dawley rats implanted with a jugular catheter, a subcutaneous ultrafiltrate probe and a brain microdialysate probe. Serum, interstitial fluid, brain microdialysate, urine and feces were collected using a Culex® Automated Blood Collection System for 24 h. At the end of this period, rats were sacrificed and major tissues were collected. Analysis by a scintillation counter confirmed that a bolus dose of 14C-radiolabeled, isoflavone-rich kudzu fraction reached bone tissues, which accumulated 0.011%, 0.09% and 0.003% of the administered dose in femur, tibia and vertebrae, respectively. Femurs extracted with 80% methanol were analyzed by high-performance liquid chromatography with electrospray ionization–mass spectrometry and were found to contain trace quantities of puerarin, daidzein and puerarin glucuronide. This study demonstrates that kudzu isoflavones and metabolites are capable of reaching bone tissues, where they may contribute to the prevention of osteoporosis and the promotion of bone health.