Mean Cell Concentration Research Articles

Composite characterization of freeze-dried human amnion membrane and human adipose tissue derived stromal cells for soft tissue engineering

Background: Soft tissue engineering has widely explored freeze-dried human amniotic membrane (FdHAM) and human adipose-derived stromal cells (hADSC) as separate components. In this study, the composite of FdHAM and hADSC combination was investigated regarding cell morphology and density, constituent components, and cytotoxicity evaluation. Methods: In vitro experimental laboratory research was conducted to analysis of the FdHAM and hADSC composites. Evaluation of cell viability and CD105 for hADSC, evaluation of pore diameter for amniotic membrane and assessment of composite cell morphology, constituent components, evaluation of cytotoxicity, and cell density was conducted. Results: Our morphology data indicated that clusters of hADSC with homogenous distribution on FdHAM. The viability test showed that the hADSCs were still dominantly alive with an average percentage of 69.3% with the mean cell concentration being 1.53 x 106/mL. The average FdHAM pore diameter was 1.90±0.30 µM. Our data suggested that FdHAM was nontoxic with good cell survivability after exposure to FdHAM. Conclusions: The structure of FdHAM and hADSC has excellent basic properties and therefore suitable for use as carriers of hADSC or in soft tissue engineering. Compared to the available literature, the characteristic of the FdHAM and hADSC composite is ideal and may be used as an end-product for use in humans.

Dispersion of a gyrotactic micro-organism suspension in a vertical pipe: the buoyancy–flow coupling effect

Understanding the transport of micro-organisms in pipes is crucial to many fundamental problems, such as bioconvection and biodiesel production. In this work, we investigate the velocity profile and dispersion of a suspension of negatively buoyant, gyrotactic micro-organisms in a vertical pipe. With an imposed flow rate, the non-uniform radial cell concentration typical of gyrotaxis distorts the simple Poiseuille flow through inhomogeneous buoyancy, which in turn affects the cell concentration distribution. By solving the fundamental Smoluchowski equation and the Navier–Stokes equation simultaneously, we account for this bidirectional buoyancy–flow coupling effect. Asymptotic dispersion coefficients, namely, drift velocity and dispersivity, are further calculated with the obtained radial velocity and cell concentration profiles, which are assumed to be steady, symmetric and axially invariant. Using the gyrotactic micro-organism Chlamydomonas augustae as an example, detailed results are given to illustrate the effect of buoyancy–flow coupling. In downwelling flows, the buoyancy–flow coupling effect intensifies with the Richardson number $Ri$ quantifying the mean cell concentration, but is strongest at a moderate flow strength. The buoyancy–flow coupling effect significantly enhances the velocity and cell concentration in the central region, as well as the drift velocity and dispersivity. In contrast, the buoyancy–flow coupling effect is comparatively limited in upwelling flows, due to the dominant influence of the no-slip boundary condition imposed at the wall. Comparisons with predictions of existing approximate models are also presented.

A comparison between the FlowCam 8100, microscopy, and sandwich hybridization assay for quantifying abundances of the saxitoxin-producing dinoflagellate, Alexandrium catenella

Light microscopy, FlowCam, and sandwich hybridization assay (SHA) are three approaches that facilitate the monitoring of harmful algal bloom (HAB) forming phytoplankton. Yet, cross-comparisons among these techniques have not been conducted. This study addressed that gap using the saxitoxin-producing ‘red tide’ dinoflagellate Alexandrium catenella, a species responsible for blooms and paralytic shellfish poisoning worldwide. To achieve this goal, the dynamic ranges of each technique were compared using A. catenella cultures spanning low (pre-bloom), moderate (bloom), and high (dense bloom) levels. To assess field detection, water samples containing very low (<3 cells mL−1) A. catenella levels were collected from Long Island Sound, USA (Jun–Aug 2021) and evaluated using each method. Field samples were also spiked with A. catenella to high (160 cells mL−1) or low (40 cells mL−1) concentrations. In general, microscopy, FlowCam, and SHA returned comparable A. catenella cell concentrations for all tests. Mean cell concentrations from laboratory intercalibration experiments were not significantly different for any method or concentration (ANOVA, p > 0.05). However, relative to microscopy at times SHA produced non-detect signals <2 cells mL−1 in field samples and the FlowCam slightly underestimated cell concentrations when A. catenella abundances were high in laboratory and field samples. Mean cell concentrations of spike experiments were not significantly different for any test date, sampling location, or method, despite variability among methods within the high concentration treatment (ANOVA, p > 0.05 for all treatments). Findings are relevant to HAB researchers, managers, and public health officials because they help reconcile disparate cell abundance datasets that inform numerical models and enhance HAB monitoring and prediction. Results are also likely broadly applicable to several HAB species.

Longitudinal profile of circulating endothelial cells in post-acute coronary syndrome patients

Introduction Patients who have experienced an acute coronary syndrome (ACS) are at risk of a recurrent event, but their level of risk varies. Because of their close temporal relationship with vascular injury, longitudinal measurements of circulating endothelial cells (CECs) carry potential to improve individual risk assessment. Methods We conducted an explorative nested case-control study within our multicenter, prospective, observational biomarker study (BIOMArCS) of 844 ACS patients. Following an index ACS, high-frequency blood sampling was performed during 1-year follow-up. CECs were identified using flow cytometric analyses in 15 cases with recurrent event, and 30 matched controls. Results Cases and controls had a median (25th-75thpercentile) age of 64.1 (58.1-75.1) years and 80% were men. During the months preceding the endpoint, the mean (95%CI) CEC concentration in cases was persistently higher than in controls (12.8 [8.2-20.0] versus 10.0 [7.0-14.4] cells/ml), although this difference was non-significant (P = 0.339). In controls, the mean cell concentration was significantly (P = 0.030) lower in post 30-day samples compared to samples collected within one day after index ACS: 10.1 (7.5-13.6) versus 17.0 (10.8-26.6) cells/ml. Similar results were observed for CEC subsets co-expressing CD133 and CD309 (VEGFR-2) or CD106 (VCAM-1). Conclusion Despite their close relation to vascular damage, no increase in cell concentrations were found prior to the occurrence of a secondary adverse cardiac event.

The prevalence of leukocyte abnormalities among Sudanese patients with sickle cell disease

Sickle cell disease (SCD) is an inherited blood disorder that affects red blood cells. The study was conducted in Elobied town during the period May 2011 to September 2011. The aim of this study is to detect the abnormalities of leucocytes among sickle cell anemic patients. 40 sickle cell anemic patients; age range between 8 months to 23 years. Blood sample was taken for all patients and the laboratory investigation were performed using automated estimation for: hemoglobin (Hb), Packed cell volume (PCV), red cell count (RBCs), mean cell volume (MCV), mean cell hemoglobin (MCH), mean cell concentration (MCHC), and total white blood cells, comment on blood film using manual methods. The conclusion of this study there is increase in total white blood cells with shift to left in neutrophil precursor in sickle cell patients with complications ,the most immature cells are band form, myelocytes and metamyelocytes, and there also lymphocytosis and neutrophilia which has been increases in response to infections.

Representative monitoring of the calcifying alga &lt;em&gt;Phacotus lenticularis&lt;/em&gt; (Chlamydophyceae) in lentic ecosystems

The biogenic carbonate precipitation by the freshwater alga Phacotus lenticularis may play a role in long-term carbon (C) fixation that has not yet been quantified. This is partly due to the absence of a standardised methodology to representatively sample and assess the cell density and sedimentation of P. lenticularis in lakes. The objective of the present study was to define an adequate sampling methodology taking into account the spatiotemporal variation of P. lenticularis as well as the sedimentation and dissolution of calcite shells. Simultaneous measurements in three different lake sub-basins of Lake Grosser Ostersee, Germany, showed that the spatial cell density of P. lenticularis was similar in each sub-basin. At all sites, the vertical P. lenticularis cell density maxima corresponded with the slowly downshifting thermocline from depths of 2 to 6 m. During the entire growth period, composite samples from 0 to 7 m included 89% of the total P. lenticularis population. Lake bathymetry, as well as external factors like wind exposure, did not appear to affect the abundance of these calcifying algae. Sediment traps at a depth below the thermocline (at 7 m) and 1 m above the lake bottom recorded sedimentation rates of P. lenticularis shell halves (sh) between 1.1×106 and 1.1×108 sh m−2 d−1, while mean cell concentrations in the water column were between 1.1×108 and 1.7×109 shells per m². Sinking velocity ranged between 3 and 4 m d−1. Sediment from traps installed at a depth of 7 m did not reflect mean shell concentrations in the water column above. Dissolution of carbonates reduced the number of shells in sediment traps at the lake bottom and during the storage of samples. A laboratory experiment showed that even distilled water used for dilution during microscopic analysis led to dissolution of P. lenticularis shells. In conclusion, combined sampling of P. lenticularis from open water and sediment traps close to the lake bottom delivers a most representative assessment of biogenic carbonate precipitation. Due to dissolution effects, high temporal resolution along with appropriate sample preservation are crucial, whereas spatial representativeness was already achieved with low number of sampling sites per lake.

Isolation, Expansion, Differentiation and Growth Kinetics Essay in Mesenchymal Stem Cells Culture from the Bone Marrow of Collared Peccaries (Tayassu tajacu)

Background: There are few studies on stem cell isolation in wild animals that provide isolation and culture protocols of these cells in vitro. Among the wild species studied, we present the collared peccary (Tayassu tajacu) as a model with potential to obtain and use MSC in preclinical studies. These animals are phylogenetically close to the domestic pig, popularly known as peccaries and found naturally in South America, Central America and the South of the United States. The aim of the present study was to establish a protocol for the isolation, in vitro cell expansion, differentiation and assessment of the stromal MSC growth curve before and after thawing.Materials, Methods &amp; Results: Mesenchymal stem cells (MSC) from collared peccary bone marrow (Tayassu tajacu) were isolated and expanded by centrifuge in Ficoll® solution and cultured in DMEM® High Glucose medium. The culture was assessed by assays of colony forming units CFU-F and growth curve by saturation (GCS). Cultures in the third passage, with 70% confluence, were replicated at 105 cells/mL concentration in the culture media to induce osteogenic cell differentiation and adipogenic cell differentiation, respectively. The MSC were frozen in nitrogen for 40 days, thawed and re-assessed for cell viability and GCS.Discussion: The bone marrow collected presented high mononuclear cellularity, with a mean variability of 94.5% and 60.83 ± 4.27 UFC were identified in the samples and cells with fibroblast-like-cell morphology were observed. When they were expanded, the mean cell viability was 95%, the mean cell concentration obtained was 233.31 ± 20.04 cells per 25cm2 bottle and the culture reached the growth plateau in GCS between the 13th and 16th day. The osteoblastic cell differentiation assay showed after 18 days, morphology similar to osteoblasts, with irregular cytoplasm limits, cell prolongation formation and flattened appearance. After staining with Alizarin Red, the nucleus presented a wine red coloring and the cytoplasm, more basophilic and well-defined, with calcium deposits inside the cells. The cultures submitted to adipogenic differentiation were large, hexagonal, irregular and presented birrefringent cytoplasm granules after the third week of culture. When stained with Oil Red it was observed that the cytoplasm granules were scattered small fat vacuoles and stained maroon. The viability after thawing was 78% and the mean cell concentration obtained in GCS was 199.71 ± 14.72 cells per 25 cm2 bottle. The curves reached the saturation plateau early, on the eighth day of observation. From then onwards the cultures entered became exhausted and the cell concentration of the samples decreased progressively until minimum values. These results showed the presence of a well-defined MSC population in the collared peccary bone marrow with a high rate of replication in vitro and potential for differentiation confirmed by the adipogenic and osteogenic lines. The cryopreservation technique adopted presented satisfactory results, but indicated a significant cell stress after thawing that justifies investigation of the apoptosis rates induced post thawing in the species. Furthermore, the bone marrow collection did not harm the animals and the facility of stromal MSC isolation and culture qualifies the collared peccary as a viable alternative model to obtain MSC and for studies in the area of cell therapy.

Prokaryoplankton standing stocks in oligotrophic gyre and equatorial provinces of the Atlantic Ocean: Evaluation of inter-annual variability

Temporal changes of prokaryoplankton in three different provinces of the Atlantic Ocean were examined between 1996 and 2004. The abundance and integrated biomass of three prokaryote groups (Prochlorococcus spp., Synechococcus spp. and other prokaryoplankton) were used to detect standing stock changes in the northern and southern oligotrophic gyres and in the equatorial region. Mean cell concentrations (±standard error of the mean) of Prochlorococcus spp., Synechococcus spp. and other prokaryoplankton above the nitracline in the northern oligotrophic gyre were 1.2×105 (±0.08), 5.0×103 (±1.22) and 0.9×106 (±0.03) cells mL−1, respectively. Similar concentrations of 1.2×105 (±0.06) Prochlorococcus mL−1, 1.9×103 (±0.29) Synechococcus mL−1 and 0.7×106 (±0.03) other prokaryoplankton mL−1 were measured in the southern oligotrophic gyre, with higher concentrations of all prokaryote groups in equatorial waters. Integrated biomass (±standard error of the mean) of Prochlorococcus spp. above the nitracline was 173 (±21) mg C m−2 in the northern oligotrophic gyre, 190 (±14) mg C m−2 in the southern oligotrophic gyre and 141 (±15) mg C m−2 in the equatorial region. Synechococcus spp. biomass was lower in each of the three provinces (18 (±2), 17 (±4) and 32 (±5) mg C m−2, respectively). The data showed no statistically significant inter-annual variability in Prochlorococcus or Synechococcus abundance or integrated biomass above the nitracline in any of the provinces. The abundance and biomass of the remaining prokaryoplankton were variable, but these variations could not be ascribed to seasonal differences and did not follow a clear inter-annual trend. In light of results presented here, recommendations on the frequency and spatial resolution of sampling needed to characterise province-scale temporal variability of prokaryoplankton communities have been suggested.

Elemental composition of Microcystis aeruginosa under conditions of lake nutrient depletion

Electron probe X-ray microanalysis (XRMA) was used to determine the elemental composition of the blue-green alga, Microcystis aeruginosa, in a stratified freshwater lake during the late summer. Colonies of this alga were initially observed in mid-July, at a time when phosphorus concentration in the lake water had decreased to minimal levels (total P 0.04 mg l−1). The P quota of these cells was high (mean concentration 132 mmol kg−1 dry weight) with a cell P to lake water P concentration ratio of 105. The elemental concentrations of Microcystis remained relatively stable throughout the sampling period (July–September), with mean cell concentrations of Mg, P, S and Ca showing no significant changes. Mean elemental ratios and the ratio of monovalent/divalent cations were also relatively constant (SE <10% mean). The pattern of cell elemental associations, determined by Factor and Pearson correlation analysis, was consistent throughout – with Mg, P, K and S forming a core tetrad of inter-correlated elements. The relative constancy of cell composition seen in Microcystis would be expected of an alga with a K-selection strategy. The continued high P quota over a period of nutrient depletion in lake water is consistent with the ability of this alga to sink to nutrient-rich lower regions of the water column.

A Snow Algal Community on Tyndall Glacier in the Southern Patagonia Icefield, Chile

We quantitatively investigated a snow algal community on Tyndall Glacier of the Southern Patagonia Icefield, Chile, at an elevation from 300 to 1500 m a.s.l. We observed 7 species of snow and ice algae (Chlorophyta and cyanobacteria) on the glacier. These species were Mesotaenium (M.) berggrenii, Cylindrocystis (Cyl.) brébissonii, Ancylonema sp., Closterium sp., Chloromonas sp., Oscillatoriaceae cyanobacterium, and an unknown alga. The spatial distribution of these algae differed among the species. M. berggrenii, Cyl. brébissonii, Ancylonema sp., and Closterium sp. appeared mainly in the lower-elevation area (370–770 m a.s.l.), the unknown alga in the higher-elevation area (900–1500 m a.s.l.), and Chloromonas sp. and Oscillatoriaceae cyanobacterium in the middle part of the glacier. The mean cell concentration and total cell volume biomass ranged from 0 to 9.2 × 104 (mean: 1.8 × 104) cells mL−1, from 0 to 327 (mean: 63) μL m−2, respectively. The cell volume biomass generally decreased with altitude. The community structure showed that M. berggrenii was dominant in the ice area (65–100% of total cell volume) and lower snow area (50–70%) and that the unknown alga was dominant in the higher snow area (100%). The Simpson’s species diversity index was significantly different among the study sites but was generally low (less than 1.9) at all sites. The cell volume biomass and diversity index are relatively smaller on the Patagonian glacier than those in algal communities on Alaskan and Himalayan glaciers. Lower nutrient levels in precipitation are likely to cause the smaller algal biomass on the glacier.

The effect of agricultural fertilizer on growth rate of benthic diatoms

The aim of this work was to consider a non-conventional medium for benthic diatoms culture, formulated with liquid agricultural fertilizers and with the ‘f/2’ medium used as control. Three strains of benthic marine diatoms ( Nitzschia laevis, Nitzschia thermalis var. minor and Navicula incerta) were grown with these two media, in batch cultures in 18-l plastic containers. There were no significant differences in mean cell concentration and growth rate between three diatoms raised with agricultural fertilizers compared to the standard ‘f/2’ medium. The cost for non-conventional culture media is less than 1/8 the cost of the ‘f/2’ medium.

Changes in the elemental composition of Asterionella formosa during the diatom spring bloom

Changes in the elemental composition of the diatom Asterionella formosa within mixed phytoplankton samples were determined over the spring bloom period using energy dispersive X-ray microanalysis. Using a 10 kV electron microprobe, X-ray information from the top 1-2 µm of the cell revealed overall mean concentrations of: Si (4636 mmol kg -1 dry wt), P (82), S (54), Cl (71) K (94) and Ca (90). Concentrations of all elements showed wide variation within each date sample, with unimodal frequency distributions approximating to a Normal distribution. Correlation of the entire data set demonstrated clear statistical associations between elements, including Si, P and K. Uptake of Si during the course of the diatom bloom led to a major fall in lake water concentration (1.0 to 0.07 mg l -1 ), which correlated with a decrease in the mean cell Si concentration (6735 to 3107 mmol kg -1 dry wt) during the final phase of the bloom. Mean cell concentrations of P and K also showed marked decreases with time, in close parallel with cell Si. These changes in P and K were attributed to the high level of internal correlation with Si and not to any significant decrease in P and K availability in the environment.

Dynamics of picophytoplankton, ultraphytoplankton and bacteria in the central equatorial Pacific

Pico- and ultraplankton are known to contribute significantly to overall biomass and primary productivity in the “high nutrient-low chlorophyll” waters of the equatorial Pacific. In order to understand the dynamics of this community on ecologically relevant time-scales, we examined the abundance, distribution and cellular characteristics of Prochlorococcus, Synechococcus, eukaryotic ultraphytoplankton and heterotrophic bacteria during two 20-day time-series at 0°N, 140°W in the spring and fall of 1992 (JGOFS time-series cruises, TS-I and TS-II). Prochlorococcus was numerically dominant among the autotrophic groups considered, with mean cell concentrations in surface waters on the order of 1.4 × 10 5 cells ml −1. Synechococcus and ultraphytoplankton abundances were 17–30-fold lower than those of Prochlorococcus, and heterotrophic bacterial abundances were 5–7-fold higher (during TS-I and TS-II, respectively). Daily cell abundances for all groups varied by factors of 1.5-2 within each time-series. Depth-integrated Prochlorococcus abundance averaged over each time-series was 25% lower during TS-II relative to TS-I; ultraphytoplankton abundance was 42% higher during the same period. Prochlorococcus and ultraphytoplankton both contributed significantly to the estimated total autotrophic biomass; Synechococcus contributed relatively little. Estimated total photosynthetic pico- plus ultraplankton biomass was on average 30% higher than heterotrophic bacterial biomass. Changes in the fluorescence and light scatter properties of individual Prochlorococcus cells were observed during the passage of a tropical instability wave during TS-II, and are hypothesized to reflect a physiological response among these cells to that event. Examination of bulk properties alone (e.g. cell numbers or total red fluorescence) would not have revealed these physiological changes. Lower bounds for Prochlorococcus-specific growth rates were calculated based on the DNA distributions of these populations at dusk. These rates were maximal at 15 or 30 m depth, where they approached one doubling per day. Changes in Prochlorococcus forward angle light scatter (FALS) from dawn to dusk were well correlated with these estimates of specific growth rate, an observation that allowed us to relate measurements of FALS to cell volume for Prochlorococcus.

Phytoplankton distribution along the eastern coast of the U.S.A.—VII. Mean cell concentrations and standing crop

Based on a 10 year data set, the phytoplankters that were most abundant and those that represented the greatest contribution (biovolume) to the standing crop were identified for several regions of the northeastern continental shelf of the United States. Pico-nanoplankton components (<10 μm), Skeletonema costatum and a mixed centric diatom category (<20 μm in size) represented 75% of the cells over the shelf. In contrast, several diatoms (e.g. Rhizosolenia spp.) and a dinoflagellate assemblage composed the bulk (95%) of the standing crop. A greater degree of relationships for species composition, occurrence and abundance was found over the outer shelf than among coastal regions.

Limitations of grazing rate equations: the case for time-series measurements

The limitations of traditional zooplankton grazing rate equations were analysed, and the relative advantages of taking time-series measurements or single end-point measurements of grazing rate examined. For zooplankters with variable feeding rates, the time-series approach is the only acceptable method. Use of end-point measurements to calculate feeding rates results in significant error if clearance rate changes or feeding ceases during the experiment, i.e. when the grazing coefficient is not constant, as is assumed in the clearance rate equations. The use of timeseries measurements is particularly important above the critical concentration for saturated ingestion rate. The functional response plot of ingestion rate versus mean cell concentration is inappropriate statistically and should be modified to avoid compounding variables appearing on both axes of the plot.

The cellular reaction of kidney after different physical injuries.

Autoradiographic investigations on kidney cells were performed after focal cryolesions (-180 to -196 degrees C) and focal heat application (740 degrees C). Cells were studied 12 h to 30 days after the lesions had been produced. In the damage granulation tissue the percentage of radioactively labelled fibroblasts as well as the percentage distribution of leucocytes, monocytes, macrophages, lymphocytes, fibroblasts and fibrocytes and the mean cell concentration were determined. There were no significant differences in the leucocytic and monocytic cell reactions after the two types of physical injuries. However, the percentage of fibroblasts, fibrocytes and macrophages was higher and the percentage of lymphocytes lower after cryonecrosis when compared to heat application. The cell concentration increased during the last 2 weeks of the experimental time after a thermolesion. The labelling index of the wound fibroblasts was significantly higher after the 10th day after thermonecrosis than after in sinus freezing. The increased cellular activity 2 to 4 weeks after heat coagulation of the kidney was probably induced by the delayed resorption of the carbonised necrotic tissue. The reduced phagocytic activity of macrophages might have depended on alteration and modification of molecular cell structures which were different after heat application of freezing. The different lymphocytic reaction seems to be the consequence of different immune responses of the lymphatic system. It is suggested that focal thermolesions may have a stimulatory effect on the cell-mediated immune response and that focal cryonecrosis may induce an increase in humoral immune response.

Long‐term T‐cell‐mediated immunity to epstein‐barr virus in man. IV. Development of T‐cell memory in convalescent infectious mononucleosis patients

Under appropriate culture conditions, EB virus infection of lymphocytes from seropositive donors leads to regression of transformation, and this was shown previously to be due to activation in a secondary immune response to T lymphocytes inhibitory for the autologous lymphoblastoid cell line. Regression can be quantified by determining the number of cells required for its expression. To investigate the development of memory T cells with EB-virus specificity in the primary infection, a comparison was made of the capacity for regression of lymphocytes from 16 cases of infectious mononucleosis (IM) and 13 normal donors. With 9 normal seropositive donors a mean lymphocyte concentration of 4.6 X 10(5)/ml was required to achieve 50% regression. In contrast, with 8 cases of IM tested within 1 week of onset, a much higher mean lymphocyte concentration (3.7 X 10(6)/ml) was necessary. Six of these IM cases, and another not tested in the first week, were tested on several occasions between 5 and 23 weeks after onset, and showed a slight reduction in the mean cell concentration required for regression (1.5 X 10(6)/ml). Six additional were tested 23--83 weeks after onset by which time the cell concentration required for 50% regression (mean = 4.5 X 10(5)/ml) had reached the level shown by normal seropositive donors. Regression did not occur with lymphocytes from seronegative donors, even at the highest cell concentration. Recombination cultures of T-cell-depleted and T-cell-enriched lymphocyte populations from 3 IM cases in ratios of 1:7 to 7:1 showed that the failure of regression in acute IM was not due simply to lack of sufficient numbers of T cells. The results indicate that EB-virus-specific memory T-cell activity as detected by the regression test is absent in the acute phase of IM, becomes evident at low levels 5--23 weeks after onset, and reaches a maximum after about 6 months.

Cytochrome P-450 distribution in rat liver and the effect of sodium phenobarbitone administration

A microspectrophotometric method for assaying cytochrome P-450 in fresh 24 μm unfixed cryostat sections of rat liver has been developed. When used to assay this cytochrome in sections of microsomal preparations it has yielded results equivalent to those obtained by the conventional spectrophotometric assay of the same preparations. Random measurements made throughout sections of liver have given mean values for cytochrome P-450 concentrations which are twice those measured in microsomes prepared from the livers of the same animals (not corrected for the yield in the homogenate). Measurements of the cytochrome P-450 content of liver cells by the microspectrophotometric method show that in liver from male Wistar rats, cells nearer to the central veins contain up to twice as much cytochrome P-450 as those nearer to the portal tract (mean cell concentrations of 26.4 (±4.4) μmol/l and 17.5 (±3.0) μmol/l respectively). In the livers from similar rats, killed at the same time, but which had received 1 mg/ml sodium phenobarbitone in their drinking water for one week, the cells near the central vein contained up to five times as much cytochrome P-450 as those near the portal tract (mean cell concentrations of 77.3 (±25.0) μmol/l and 28.3 (±9.6) μmol/l respectively). The results show a selective increase in cytochrome P-450 content by the cells in the centrilobular region after treatment with sodium phenobarbitone and a smaller increase by some of the cells in the periportal region.

Microbial Detection Method Based on Sensing Molecular Hydrogen

A simple method for detecting bacteria, based on the time of hydrogen evolution, was developed and tested against various members of the Enterobacteriaceae group. The test system consisted of (i) two electrodes, platinum and a reference electrode, (ii) a buffer amplifier, and (iii) a strip-chart recorder. Hydrogen evolution was measured by an increase in voltage in the negative (cathodic) direction and recorded on a strip-chart recorder. Hydrogen response curves consisted of (i) a lag period, (ii) a period of rapid buildup in potential due to hydrogen, and (iii) a period of decline in potential. A linear relationship was established between inoculum size and the time hydrogen was detected (lag period). Lag times ranged from 1 h for 10(6) cells/ml to 7 h for 10(0) cells/ml. For each 10-fold decrease in inoculum, length of the lag period increased 60 to 70 min. Mean cell concentrations at the time of hydrogen evolution were 10(6)/ml. Based on the linear relationship between inoculum size and lag period, these results indicate the potential application of the hydrogen-sensing method for rapidly detecting coliforms and other gas-producing microorganisms in a variety of clinical, food, and other samples.