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81.
Fernandes Talma R.L. Suzuki-Kemmelmeier Fumie Constantin Jorgete Bracht Adelar 《Molecular and cellular biochemistry》1999,195(1-2):207-217
The heterogeneity of the liver parenchyma in relation to uric acid production from adenosine was investigated using the bivascularly perfused rat liver in the anterograde and retrograde modes. Adenosine was infused in livers from fed rats during 20 min at four different concentrations (20, 50, 100 and 200 M) according to four experimental protocols as follows: (A) anterograde perfusion, with adenosine infusion into the portal vein; (B) anterograde perfusion, with adenosine in the hepatic artery, (C) retrograde perfusion, with adenosine in the hepatic vein; (D) retrograde perfusion, with adenosine in the hepatic artery. With protocols A, B, and D uric acid production from adenosine was always characterized by initial bursts followed by progressive decreases toward smaller steady-states. With protocol C the initial burst was present only when 200 M adenosine was infused. The initial bursts in uric acid production were accompanied by simultaneous increases in the ratio of uric acid production/adenosine uptake rate. These initial bursts are thus representing increments in the production of uric acid that are not corresponded by similar increments in the metabolic uptake rates of adenosine. Global analysis of uric acid production revealed that the final steady-state rates were approximately equal for all infusion rates with protocols A, B and C, but smaller with protocol D. This difference, however, can be explained in terms of the differences in accessible cellular spaces, which are much smaller when protocol D is employed. When the analysis was performed in terms of the extra amounts of uric acid produced during the infusion of adenosine, where the initial bursts are also taken into account, different dose-response curves were found for each experimental protocol. These differences cannot be explained in terms of the accessible cell spaces and they are likely to reflect regional heterogeneities. From the various dose-response curves and from the known characteristics of the microcirculation of the rat liver it can be concluded that the initial bursts in uric acid production are generated in periportal hepatocytes. The reason for this heterogeneity could be related to the metabolic effects of adenosine, especially to oxygen uptake inhibition, which is likely to produce changes in the ATP/AMP ratios. 相似文献
82.
Park YC Kim JB Heo Y Park DC Lee IS Chung HW Han JH Chung WG Vendeland SC Whanger PD 《Biological trace element research》2004,98(2):143-157
Intestinal metabolism of the subtoxic level of selenite in rats was investigated using a double-perfusion system, which is
an in situ, in vitro preparation in which the intestinal lumen and its vasculature are perfused simultaneously. The toxicity of sodium
selenite was determined by inhibition of 3-O-methyl glucose (3MG) absorption and by histological examination. Levels of 1.2 mM selenite were required to significantly (p<0.05) reduce 3MG intestinal absorption (58±11%, mean±SD). Cation-exchange chromatography was used to determine the chemical
forms of Se from selenite after using luminal concentrations of 1–200 μM in vascular perfusates. The chemical forms were selenite, selenodiglutathione (GS-Se-SG), mixed selenoglutathione plus cysteine
(GS-Se-CYS), selenodicysteine (CYS-Se-CYS), protein-bound Se, and unidentified selenocompounds. Selenite was the predominant
selenocompound found in vascular perfusate, but protein-bound Se was the predominant metabolite from selenite present in the
vascular effuents. There was a corresponding increase of all metabolites with increased levels of selenite with time of absorption,
but not with increased concentration of luminal selenite. 相似文献
83.
Somatostatin suppresses ghrelin secretion from the rat stomach 总被引:6,自引:0,他引:6
Shimada M Date Y Mondal MS Toshinai K Shimbara T Fukunaga K Murakami N Miyazato M Kangawa K Yoshimatsu H Matsuo H Nakazato M 《Biochemical and biophysical research communications》2003,302(3):520-525
Ghrelin is an acylated peptide that stimulates food intake and the secretion of growth hormone. While ghrelin is predominantly synthesized in a subset of endocrine cells in the oxyntic gland of the human and rat stomach, the mechanism regulating ghrelin secretion remains unknown. Somatostatin, a peptide produced in the gastric oxyntic mucosa, is known to suppress secretion of several gastrointestinal peptides in a paracrine fashion. By double immunohistochemistry, we demonstrated that somatostatin-immunoreactive cells contact ghrelin-immunoreactive cells. A single intravenous injection of somatostatin reduced the systemic plasma concentration of ghrelin in rats. Continuous infusion of somatostatin into the gastric artery of the vascularly perfused rat stomach suppressed ghrelin secretion in both dose- and time-dependent manner. These findings indicate that ghrelin secretion from the stomach is regulated by gastric somatostatin. 相似文献
84.
Effects of oxygen on engineered cardiac muscle 总被引:6,自引:0,他引:6
Carrier RL Rupnick M Langer R Schoen FJ Freed LE Vunjak-Novakovic G 《Biotechnology and bioengineering》2002,78(6):617-625
Concentration gradients associated with the in vitro cultivation of engineered tissues that are vascularized in vivo result in the formation of only a thin peripheral tissue-like region (e.g., approximately 100 microm for engineered cardiac muscle) around a relatively cell-free interior. We previously demonstrated that diffusional gradients within engineered cardiac constructs can be minimized by direct perfusion of culture medium through the construct. In the present study, we measured the effects of medium perfusion rate and local oxygen concentration (p(O2)) on the in vitro reconstruction of engineered cardiac muscle. Neonatal rat cardiomyocytes were seeded onto biodegradable polymer scaffolds (fibrous discs, 1.1 cm diameter x 2 mm thick, made of polyglycolic acid, 24 x 10(6) cells per scaffold). The resulting cell-polymer constructs were cultured for a total of 12 days in serially connected cartridges (n = 1-8), each containing one construct directly perfused with culture medium at a flow rate of 0.2-3.0 mL/min. In all groups, oxygen concentration decreased due to cell respiration, and depended on construct position in the series and medium flow rate. Higher perfusion rates and higher p(O2) correlated with more aerobic cell metabolism, and higher DNA and protein contents. Constructs cultured at p(O2) of 160 mm Hg had 50% higher DNA and protein contents, markedly higher expression of sarcomeric alpha-actin, better organized sarcomeres and cell junctions, and 4.5-fold higher rate of cell respiration as compared to constructs cultured at p(O2) of 60 mm Hg. Contraction rates of the corresponding cardiac cell monolayers were 40% higher at p(O2) of 160 than 60 mm Hg. The control of oxygen concentration in cell microenvironment can thus improve the structure and function of engineered cardiac muscle. Experiments of this kind can form a basis for controlled studies of the effects of oxygen on the in vitro development of engineered tissues. 相似文献
85.
Transport characteristics of certain inorganic elements such as copper, magnesium, selenium and iron have been studied in maternal-fetal direction in normal pregnancies, using in vitro perfusion of isolated placental lobules. Copper, selenium, magnesium and iron salts corresponding to twice physiological concentrations were injected as a 100 l bolus, into the maternal arterial perfusate. Serial perfusate samples were collected from venous outflows for a study period of 5 min. Concentrations of various inorganic elements and their transport kinetics were determined. Transport fractions of copper, selenium, magnesium and iron averaged 0.14, 0.19, 0.06 and 0.23% of maternal load respectively. The pharmacokinetic parameters such as area under the curve, clearance, elimination constant, and time for maximum response showed some significant differences between the various elements. We speculate that copper and selenium share the same transport pathway along a concentration gradient in maternal-fetal direction, while for iron and magnesium, active transport plays a predominant role for element transfer across the human placental membrane. 相似文献
86.
Kai Iding Dirk Lütkemeyer Elisabeth Fraune Kurt Gerlach Jürgen Lehmann 《Cytotechnology》2000,34(1-2):141-150
Since 1969 much attention has been devoted to the useof spinfilter systems for retention of mammalian cellsin continuous perfusion cultivations. Previousinvestigations dealt with hydrodynamic conditions,fouling processes and upscaling. But hydrodynamicconditions and fouling processes seem to have asecondary importance in spinfilter performance duringauthentic perfusion cultivations. Obviously,alterations in culture condition are more relevantespecially during long-term processes. Therefore, ourpratical approach focussed on the performance qualityof a commercially available 20 m spinfilterduring a perfusion cultivation of a recombinant CHOcell line in pilot scale regarding the followingissues: 1) retention of viable cells in thebioreactor; 2) removal of dead cells and cell debrisfrom the bioreactor; 3) alterations in culturecondition; and 4) changes in perfusion mode.Furthermore, we tested the performance of 20 mspinfilters in 2 and 100 l pilot scale using solidmodel particles instead of cells. Our investigationsshowed that retention of viable cells in pilot scalewas independent of spinfilter rotation velocity andperfusion rate; the retention increased from 75 to 95%corresponding to operation time, enlarging celldiameter and enhanced formation of aggregates in theculture during the perfusion cultivation. By means ofthe Cell Counter and Analyzer System (CASY) anoperation cut off of 13 m was determined forthis spinfilter. Using solid model particles in 2 lscale, optimal retention was achieved at a tip speedof 0.43 m s-1 (141 rpm) – furtherenhancement of spinfilter rotation velocity up to0.56 m s-1 (185 rpm) decreased the retentionrapidly. In pilot scale best retention performance wasobtained with tip speeds of 0.37 m s-1(35 rpm) and 1.26 m s-1 (120 rpm). Hence,significant retention in pilot scale could already beachieved with low agitation. Therefore, the additionof shear force protectives could be avoided so thatthe purification of the target protein from thesupernatant would be facilitated. 相似文献
87.
Monocyte-colony inhibition factor (M-CIF) was produced in microcarrier perfusion cultures from engineered Chinese hamster
ovary (CHO) cells. Three and fifteen liter microcarrier perfusion bioreactors equipped with internal spin filters were operated
for over two months. Approximately 60 L and 300 L of culture filtrate were harvested from the 3L and 15L microcarrier perfusion
bioreactors respectively. During the perfusion operation, cell density reached 2–6 × 106 cells/ml. Importantly, stable expression of M-CIF from the CHO cells under non-selection condition was maintained at a level
of 4–10 mg/L. Specific productivity was maintained at 1.8–3.4 mg/billion cells/day. The ability of the recombinant CHO cells
to migrate from microcarrier to microcarrier under our proprietary HGS-CHO-3 medium greatly facilitated microcarrier culture
scale-up and microcarrier replenishment. Future directions for microcarrier perfusion system scale-up and process development
are highlighted.
This revised version was published online in August 2006 with corrections to the Cover Date. 相似文献
88.
An alternative culture system has been developed based on a conventional tissue culture plate (3.5 cm diameter) which is changed into a closed perfusion chamber. The system can easily be scaled up from one to several chambers. The shape and the size of the area of cell growth may be designed to individual experimental demands. The whole culture chamber is optically accessible, so cell growth and morphology can be evaluated by light microscopy. Furthermore the cellular physiology can be characterised by any fluorimetric assay using a bottom type fluorescence reader. A peristaltic pump sustains a constant medium flow through the chamber thus creating true homeostasis. The use of HPLC-valves and connectors allows the switching between different media or assay solutions. Thus it is possible to perform in situ assays also measuring transient effects. A protocol for vitality tests using calcein-AM is worked out for an adherent cell line and for a suspension cell line. The lower detection limits are 7 × 102 cells cm-2 for the adherent cells and 5 × 104 cells mL-1 for the suspension cells. The upper limits are 1–2 × 105 cells cm-2 respectively 8 × 106 cells mL-1. 相似文献
89.
Bote G. Bruinsma James H. Avruch Pepijn D. Weeder Gautham V. Sridharan Basak E. Uygun Negin G. Karimian Robert J. Porte James F. Markmann Heidi Yeh Korkut Uygun 《Journal of visualized experiments : JoVE》2015,(98)
There is currently a severe shortage of liver grafts available for transplantation. Novel organ preservation techniques are needed to expand the pool of donor livers. Machine perfusion of donor liver grafts is an alternative to traditional cold storage of livers and holds much promise as a modality to expand the donor organ pool. We have recently described the potential benefit of subnormothermic machine perfusion of human livers. Machine perfused livers showed improving function and restoration of tissue ATP levels. Additionally, machine perfusion of liver grafts at subnormothermic temperatures allows for objective assessment of the functionality and suitability of a liver for transplantation. In these ways a great many livers that were previously discarded due to their suboptimal quality can be rescued via the restorative effects of machine perfusion and utilized for transplantation. Here we describe this technique of subnormothermic machine perfusion in detail. Human liver grafts allocated for research are perfused via the hepatic artery and portal vein with an acellular oxygenated perfusate at 21 °C. 相似文献
90.