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PubMed1971

Differences in distribution of esterase between cell fractions of rat liver homogenates prepared in various media. Relevance to the lysosomal location of the enzyme in the intact cell.

The distribution of esterase in subcellular fractions of rat liver homogenates was compared with that of the lysosomal enzyme acid phosphatase and the microsomal enzyme glucose 6-phosphatase. Most of the esterase from sucrose homogenate sediments with glucose 6-phosphatase and about 8% is recovered in the supernatant. However, up to 53% of the esterase can be washed from microtome sections of unfixed liver, in which less cellular damage would be expected than that caused by homogenization. About 40% of both esterase and acid phosphatase are recovered in the soluble fraction after homogenization in aqueous glycerol or in a two-phase system (Arcton 113-0.25m-sucrose), although glucose 6-phosphatase is still recovered in the microsomal fraction of such homogenates. The esterase of the microsomal fraction prepared from a sucrose homogenate is much more readily released by treatment with 0.26% deoxycholate than are other constituents of this fraction. The release of esterase from the microsomal fraction by the detergent and its concomitant release with acid phosphatase after homogenization in glycerol or the two-phase system suggests that a greater proportion of esterase may be present in lysosomes of the intact cell than is indicated by the results of standard fractionation procedures.

Acid Phosphatase↗
PubMed1980

The differential release of basal ATPase, Ca2+-dependent ATPase, 5'-nucleotidase and cholesterol during homogenization of skeletal muscle.

The influence of homogenization times on the presence of constituents in the microsomal fraction of skeletal muscle was investigated. Membranes having Ca2+-activated ATPase activity have a fragmentation pattern distinct from that of membranes displaying Ca2+-independent or basal ATPase activity. These latter membranes were found in highest specific concentration in the microsomal fraction prepared from homogenates subjected to short periods of homogenization. 5'-Nucleotidase (EC 3.1.3.5) activity paralleled that of basal ATPase on short periods of homogenization, as also did the specific concentration of cholesterol. Longer periods of homogenization led to a decrease in the specific activity of basal atpase, which reached its lowest value at 120s of homogenization, whereas the specific activity of 5'-nucleotidase and the specific concentration of cholesterol decreased initially in a similar manner to basal ATPase, but both increased substantially after the longest period of homogenization.

Adenosine Triphosphatases↗
PubMed1980

Crohn disease lymph node homogenates produce murine lymphoma in athymic mice.

To study the putative agent(s) related to Crohn disease, we intraperitoneally in injected mesenteric lymph node homogenates from four patients with active Crohn disease into 10-week-old athymic (nu/nu) mice. Control mice (nu/nu) were injected with homogenates of mesenteric lymph nodes from two patients with ulcerative colitis and four patients undergoing elective cholecystectomy, and with a homogenate of a cervical lymph node containing sarcoid granuloma. Thirty-four mice received filtered or unfiltered homogenates from Crohn disease lymph nodes. Thirty-two mice received homogenates or filtrates from lymph nodes of control patients. Four mice from the group injected with Crohn disease homogenates from four different patients developed generalized lymphadenopathy due to lymphoma 10-28 weeks after th injection. Two additional mice developed lymphadenopathy due to plasma cell hyperplasia. None of the control mice developed lymphomas or lymphadenopathy. Two lymphomas were homogenized, filtered, and injected intraperitoneally into a second group of nu/nu mice, which also developed lymphoma within 8 weeks of injection. Two lymphomas were cultured in vitro and B cell sur?ACE MARKERS WERE IDENTIFIED. Indirect immunofluorescence studies in two lymphomas showed cytoplasmic staining of lymphoma cells with sera from 10 patients with active Crohn disease but not with sera from 13 control subjects, including 6 with ulcerative colitis and 7 with other gastrointestinal disorders. These results suggest that a transmissible factor present in Crohn disease lymph nodes produces lymphoma in nu/nu mice. Furthermore, sera of Crohn disease patients contain an antibody that recognizes an "antigen(s)" in the murine lymphoma.

Animals↗
PubMed2005

Proficiency testing in immunopathology: establishing the homogeneity of test material.

AIMS: To develop a technique for homogeneity testing of serum aliquot samples suitable for use in the Quality Assurance Program in Clinical Immunology (QAP Pty Ltd). METHODS: Albumin was selected as the surrogate protein marker for the product to be tested and the coefficient of dispersion (COD) calculated as the measure of homogeneity. To detect changes in the average level of homogeneity, cumulative sum control (cusum) charts were used. RESULTS: The COD(%) for each triplicate reading of albumin obtained from 34 specimens was normally distributed with a mean of 0.49% and a standard deviation of 0.25%. In industrial quality control schemes the action line is generally set at the upper 99% confidence limits, hence any triplicate sample would be considered to have acceptable homogeneity if the COD was < or = 1.08%. Cusum charts were created to monitor albumin homogeneity over time. CONCLUSIONS: The use of albumin measurement as the surrogate appears statistically suitable for homogeneity testing in QAP programs for immunodiagnostic testing. CUSUM charts are particularly useful to monitor such homogeneity testing.

Albumins↗
PubMed1977

Tests for homogeneity of effect in an epidemiologic investigation.

A test for homogeneity of the effect of a factor in inducing disease in the various strata of a population should relate to the scaling in which homogeneity is anticipated. Popularity, it is the multiplicative effect on the odds ratio in which homogeneity is anticipated. An asymptotic test by M. Zelen (Biometrika 58:129-137, 1971) for homogeneity in this scaling is indicated to be invalid. Further, a test by J. L. Fleiss (Statistical Methods for Rates and Proportions. NY, Wiley, 1973) for homogeneity of the standardized difference is shown to be the same as the Zelen test, to be invalid in any case, and not to correspond to any scaling. The examples given include one in which results of two studies are apparently inhomogeneous by the test even though they give effectively identical results. It is advised that tests for homogeneity be conducted with care, and that thought be given to how effects might be made homogeneous rather than to trying to demonstrate the existence of heterogeneity.

Epidemiologic Methods↗
PubMed2005

Generation of 8-hydroxydeoxyguanosine from DNA using rat liver homogenates.

In relation to carcinogenesis, aging and other pathologic conditions, urinary 8-hydroxydeoxyguanosine (8OHdG) is widely used as a marker for evaluating the effect of oxidative stress on DNA. Because no reports have described how 8OHdG is generated from DNA in vivo or by biological materials, and how it is excreted into urine, the authors investigated the generation of 8OHdG from DNA, using rat liver homogenate. Oxidatively damaged DNA samples containing different levels of 8OHdG were prepared using ultraviolet irradiation with three different concentrations of riboflavin. Following incubation of damaged DNA samples with rat liver homogenates, the generation of 8OHdG from the DNA was determined using high-performance liquid chromatography with electrochemical detection after ultrafiltration of the incubation mixtures. The generation of 8OHdG was also tested with an anti-8OHdG antibody. The quantity of 8OHdG generated from the DNA by rat liver homogenates was dependent on the 8OHdG levels in the DNA: almost all 8OHdG in the DNA was released as 8OHdG by rat liver homogenates. Generation of 8OHdG correlated with the degradation of DNA. Interestingly, the generated 8OHdG was stable in the presence of rat liver homogenates, whereas deoxyguanosine (dG) rapidly disappeared in the same conditions. Less than 1/10,000 of dG was converted to 8OHdG when dG was incubated with rat liver homogenate. Incubation of 8-hydroxyguanine with rat liver homogenates did not generate 8OHdG. These findings suggest that most of the 8OHdG in DNA is released as 8OHdG during DNA degradation and that, because of its stability, 8OHdG is excreted into urine, thus providing a convenient measure of oxidative damage to DNA.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1984

Glutamine oxidation by dissociated cells and homogenates of rat brain: kinetics and inhibitor studies.

The rates of [U-14C]glutamine oxidation to 14CO2 were determined under a variety of experimental conditions using whole homogenates and dissociated cells from rat brain. The pattern of glutamine oxidation by homogenates differed from that by dissociated brain cells in several respects. The rates of glutamine oxidation by dissociated brain cells showed saturation kinetics with an apparent Km of 0.30 mM. Lineweaver-Burk plots of glutamine oxidation by homogenates revealed two linear segments with two apparent Km values (0.58 mM and 3.0 mM). In the presence of aminooxyacetate, however, the Lineweaver-Burk plots for homogenates were linear with a single Km of 0.47 mM. The oxidation of glutamine by homogenates was inhibited by both rotenone and antimycin A (80-85%), as were glutamate and glucose oxidation, suggesting that a significant amount of glutamine is oxidized via the tricarboxylic acid cycle. In the presence of aminooxyacetate, glutamine oxidation was inhibited less than 40%, whereas the oxidation of glutamate was inhibited 75%; in contrast, glucose oxidation was enhanced 50%. The rates of glutamine oxidation by homogenates were highest in the presence of high levels of potassium (50 mM) and low levels of sodium (2.5 mM). Varying ionic composition, however, had little or no effect on the rates of glutamine oxidation by dissociated brain cells. Measurements of glutamine oxidation by homogenates prepared from 2-, 10-, 15-, 25-, and 90-day-old rats revealed little or no age-dependent difference. In contrast, the oxidation by dissociated brain cells from 2-day-old animals was significantly less than that obtained for animals 10 days or older (7.76 vs. 15.6 nmol/h/mg).(ABSTRACT TRUNCATED AT 250 WORDS)

Aminooxyacetic Acid↗
PubMed1995

Melatonin reduces H2O2-induced lipid peroxidation in homogenates of different rat brain regions.

The ability of melatonin to modify H2O2-induced lipid peroxidation in brain homogenates was determined. The concentrations of brain malonaldehyde (MDA) and 4-hydroxyalkenals (4-HDA) were assayed as an index of induced membrane oxidative damage. Homogenates from five different regions of the brain (cerebral cortex, cerebellum, hippocampus, hypothalamus, and corpus striatum) derived from two different strains of rats, Sprague-Dawley and Wistar, were incubated with either H2O2 (5 mM) alone or H2O2 together with melatonin at increasing concentrations ranging from 0.1 to 4 mM. The basal level of lipid peroxidation was strain-dependent and about 100% higher in homogenates from the brain of Wistar rats than those measured in Sprague-Dawley rats. MDA + 4-HDA levels increased after H2O2 treatment in homogenates obtained from each region of the brain in both rat strains but the sensitivity of the homogenates from Sprague-Dawley rats was greater than that for the homogenates from Wistar rats (increases after H2O2 from 45 to 165% compared 20 to 40% for Sprague-Dawley and Wistar rats, respectively). Melatonin co-treatment reduced H2O2-induced lipid peroxidation in brain homogenates in a concentration-dependent manner; the degree of protection against lipid peroxidation was similar in all brain regions.

Animals↗
PubMed2002

Floquet wave homogenization of periodic anisotropic media.

A dynamic homogenization method based on Floquet wave theory is developed. The theory is based on the equivalency within the homogenization domain of Floquet waves in a periodic anisotropic medium and plane waves in a dispersive homogeneous anisotropic medium. A simple procedure has been developed to estimate analytically critical angles and the upper frequency bound of this homogenization domain. Using this method, the frequency dependent effective elastic constants are obtained and examples for [0/90] and [0/45/90/-45] composites are given. By comparison with an exact theory, it is shown that the time domain signal propagation in a periodic laminate is well described by the Floquet wave homogenization theory in the homogenization domain. It is also shown that in the static limit the results are identical to those calculated by static homogenization theory (the generalized method of cells). The potential applications of the method are discussed.

Journal Article↗
PubMed1988

Effect of dietary carbohydrates on bacterial cholyltaurine hydrolase in poultry intestinal homogenates.

The bile salt hydrolase activity in intestinal homogenates reflects composite activities of the gastrointestinal microbial consortia. We have proposed that specific transformations of conjugated bile acids by the intestinal microflora result in the production of metabolites which depress the growth of poultry. The influence of dietary carbohydrates on the physical and kinetic properties of cholyltaurine hydrolase activity, one such bile acid-transforming enzyme in gastrointestinal homogenates of young chickens, was characterized by using a sensitive radiochemical assay. Cholyltaurine hydrolase activity in crude extracts of ileal homogenates was increased twofold by 0.25% Triton X-100 and a freeze-thaw cycle. The pH optimum for cholyltaurine hydrolase from ileal homogenates was very broad and reflected the pH range of poultry intestinal contents (i.e., 5.8 to 6.4). The carbohydrate component of the diet did not affect the apparent temperature optimum (41 degrees C) or stability profile, nor did it affect the apparent Km for taurocholic acid hydrolysis (approximately 0.43 mM). The enzymes in intestinal homogenates were active on all taurine-conjugated bile acids tested. The carbohydrate component of the diet did, however, affect the specific activity of cholyltaurine hydrolase in ileal homogenates from chickens. The levels of cholyltaurine hydrolase activity (rye greater than sucrose greater than corn) in homogenates from birds fed the different diets were directly related to the amount of growth depression (rye greater than sucrose greater than corn) associated with feeding these dietary carbohydrates. These data suggest that intestinal levels of cholyltaurine hydrolase are correlated with the amount of carbohydrate-induced growth depression in poultry.

Amidohydrolases↗
PubMed1985

Passive calcium-buffering capacity of a rabbit ventricular homogenate preparation.

This study characterizes the passive Ca2+-binding capacity of a crude homogenate fraction prepared from rabbit ventricular tissue. The soluble and particulate fractions of the homogenate were separated by centrifugation, and the Ca2+-binding capacity of each fraction was measured separately with Ca2+-selective electrodes and then summed to obtain the Ca2+-binding capacity of the total tissue homogenate. Qualitatively, the Ca2+ bound to the homogenate exhibited both relatively high- (Km less than 1 microM) and low-affinity (Km greater than 10 microM) sites over the physiological range of free Ca2+ concentrations. The total homogenate bound 72, 128, and 308 mumol Ca2+/kg wet wt tissue at 1, 10, and 50 microM free Ca2+ concentrations, respectively. The Ca2+ bound to the homogenate was mostly La3+ displaceable, which suggests that it represents a largely noncompartmentalized, rapidly exchangeable fraction of Ca2+. Myofibrillar ATPase activity was examined as a function of free Ca2+ concentration in an ionic medium similar to that used in the Ca2+-binding experiments. At free Ca2+ concentrations that stimulated myofibrillar ATPase activity half maximally, the measured rise in Ca2+ bound to the homogenate above resting values was greater than 100 mumol/kg wet wt. These data provide base-line values for the passive Ca2+-buffering capacity of the myocardium, although relevance to the physiological setting must be interpreted with caution.

Adenosine Triphosphatases↗
PubMed2003

Peripheral nerve magnetic stimulation: influence of tissue non-homogeneity.

BACKGROUND: Peripheral nerves are situated in a highly non-homogeneous environment, including muscles, bones, blood vessels, etc. Time-varying magnetic field stimulation of the median and ulnar nerves in the carpal region is studied, with special consideration of the influence of non-homogeneities. METHODS: A detailed three-dimensional finite element model (FEM) of the anatomy of the wrist region was built to assess the induced currents distribution by external magnetic stimulation. The electromagnetic field distribution in the non-homogeneous domain was defined as an internal Dirichlet problem using the finite element method. The boundary conditions were obtained by analysis of the vector potential field excited by external current-driven coils. RESULTS: The results include evaluation and graphical representation of the induced current field distribution at various stimulation coil positions. Comparative study for the real non-homogeneous structure with anisotropic conductivities of the tissues and a mock homogeneous media is also presented. The possibility of achieving selective stimulation of either of the two nerves is assessed. CONCLUSION: The model developed could be useful in theoretical prediction of the current distribution in the nerves during diagnostic stimulation and therapeutic procedures involving electromagnetic excitation. The errors in applying homogeneous domain modeling rather than real non-homogeneous biological structures are demonstrated. The practical implications of the applied approach are valid for any arbitrary weakly conductive medium.

Child↗
PubMed1995

Susceptibility of insulin to proteolysis in rat lung homogenate and its protection from proteolysis by various protease inhibitors.

The objective of this study was to determine the stability of insulin in rat lung homogenate and to confirm the effectiveness of various protease inhibitors for insulin delivery from the lung. The stability of insulin in rat lung homogenate was compared with that in the small intestine. Insulin was rapidly degraded in lung homogenate similarly to degradation in the small intestinal homogenate. The effects of various protease inhibitors on the degradation of insulin were studied in the lung. Protease inhibitors such as Na-glycocholate (Na-GC; 10 mM), aprotinin (10 mg/ml), soybean trypsin inhibitor (STI; 10 mg/ml) and bacitracin (20 mM) effectively reduced insulin degradation in lung homogenate. The rank order of effectiveness for decreasing the insulin hydrolysis in lung homogenate was bacitracin > aprotinin > STI > Na-GC. In addition, a slight correlation was observed between the in situ pulmonary absorption of insulin and its stability in vitro in the lung homogenate in the presence of various protease inhibitors. These findings suggest that the coadministration of protease inhibitors would be useful for improving the stability of insulin in the lung.

Absorption↗
PubMed1977

Specific binding of prolactin to seminal vesicle, prostate and testicular homogenates of immature, mature and aged rats.

Ovine prolactin was iodinated by the lactoperoxidase method and purified by gel filtration on Sephadex G-100. The binding ability of the labelled hormone was determined, by incubation with liver homogenate from rabbits in late pregnancy, to be 8-8% total binding/mg protein, of which 86% was specific. The fraction of 125I-labelled ovine prolactin which bound most strongly was subsequently used to study its binding to rat seminal vesicle, prostate and testicular homogenates. The total binding to the seminal vesicle homogenate taken from mature (80-day-old) rats was the highest (11-69%/mg protein), but the greatest degree of binding specificity (82-6%) was to immature (30-day-old) rat prostate. Both total and specific binding to rat testicular homogenate were consistently very low. The binding specificity was demonstrated by displacement studies: while ovine prolactin caused displacement of specific binding, human chorionic gonadotropin, rat thyrotropin and human follicle-stimulating hormone did not cause any significant displacement of bound 125I-labelled ovine prolactin. Affinity constants (Ka) and binding capacities for the seminal vesicle and prostate homogenates were determined by Scatchard analysis and the effect of age on these parameters was studied. There was no difference in Ka between the aged (220-day-old), immature and mature rat tissue homogenates; however, a significant fall in binding capacity was observed in the mature rat prostate, and a further fall in the aged rat prostate. No such change was observed in the binding capacity of the seminal vesicle, as estimated by Scatchard analysis, although total and specific binding to the mature homogenates was higher than that of the other age groups.

Age Factors↗
PubMed2005

[The proapoptotic effect of the homogenate of the tissue of different parts of pig's full thickness dermal wounds on cultured fibroblasts].

OBJECTIVE: To observe the proapoptotic effect of the homogenate of different parts of pig's full thickness dermal wounds on cultured fibroblasts. METHODS: The tissues were dissected from the wound center and sub-neoepithelium separately 15 days after homogenization and sterilization, the specimens stored at -70 degrees C. The forth passage of the fibroblasts were cultured for 16 hours in different culture solutions and were grouped into 7 groups: DMEM containing 5% fetal bovine serum as Group I, DMEM containing 5% homogenate of tissue from wound center as Group II, DMEM containing 5% homogenate of tissue from sub-neoepithelium as Group III, the culture solution of Group I mixed with 10 microg/ml GM6001 in Group IV, with the culturing medium of Group III plus 10 microg/ml GM6001 as Group V, the culture solution of Group II mixed with 10 ng/ml aFGF as Group VII, and the culture solution of Group III mixed with 10 ng/ml aFGF as Group VII. In all groups except Group I, the fibroblasts of the 6 pigs were treated with the homogenate derived from the same animal respectively. After being incubated in Annexin V-FITC and PI, cells were analyzed by Flow Cytometry and the rate of apoptotic cells was acquired. The data were analyzed by SPSS 11.0 using Least-significant Difference test (LSD). RESULTS: The apoptotic rate of the 7 groups were as follows: 4.39% +/- 0.41% in Group I, 10.98% +/- 1.42% in Group II, 13.47% +/- 1.44% in Group III, 7.2% +/- 0.46% in Group IV, 12.1% +/- 0.85% in Group V, 3.9% +/- 0.63% in Group VI, 9.8% +/- 0.50% in Group VII; there were significant differences between every two groups except Group I and Group VI. CONCLUSION: Homogenate of the tissue derived from the sub-neoepithelium has greater proapoptotic effect than that from the wound center; the proapoptotic effect of homogenate of the tissue both under neoepithelium and in wound center can be significantly alleviated by acid fibroblast growth factor, partly because of MMPs.

Animals↗
PubMed2000

Disturbances of mouse pregnancy after injection of Ascaris homogenate during early organogenesis.

Administration of the Ascaris tegumental homogenate (0.6-1.2 g of Ascaris proteins/kg/day) at a early stage of organogenesis (5-9 days of pregnancy) had a harmful effect upon the developing mouse fetuses. It has been found that injection of the homogenate did not delay or prevent implantation, but causes a high rate of intrauterine deaths. The Ascaris homogenate significantly decreased the number of live fetuses per litter, increased the frequency of litter resorption, produced a delay in bone formation and induced pathological changes of fetal organs and tissues. The congenital malformations were noted in fetuses after injection of higher doses of Ascaris homogenate (exencephaly, craniomeningocele and internal hydrocephalus). No malformations were noted in control groups and after injection of minimum dose of the homogenate. The symptoms that occurred after administration of the tegumental homogenate to pregnant mice included: decreased body weight gain (p<0.001) as compared to controls, vaginal hemorrhage, intrauterine resorption of litter and mortality. These signs suggest that the Ascaris homogenate causes maternal toxicity.

Abnormalities, Drug-Induced↗
PubMedDate not supplied

The effect of SH-groups of cysteine on the rate of oxygen uptake by some homogenates of the rabbit tissues.

The effect of SH-groups of cysteine on the rate of oxygen uptake by some homogenates of the rabbit tissues. Acta Physiol. Pol., 1977, 28 (6): 541-551. In the present experiments the effect of SH-groups of cysteine on the respiration rate of homogenates of kidney, liver, brain, myocardium and skeletal muscle was investigated in the rabbit. Using the Warburg's method of respirometry it was found that cysteine added to the incubation medium modifies oxygen uptake by the above-mentioned tissue homogenates and that this reaction depends both on the kind of tissue and cysteine concentration in the medium. Addition of cysteine to the incubation medium in the concentration of 0.1 mg/ml exerted only slight, insignificant influence on the tissue respiration but in the concentration of 0.4 mg/ml it increased the respiration rate in homogenates of kidney (by 30%), liver (by 55%) and skeletal muscle (by 59%). Cysteine added in the concentration of 0.8 mg/ml increased the respiration rate of all the examined tissue homogenates. The strongest effect of cysteine in this concentration was found in the liver and skeletal muscle homogenates (an increase in O2 uptake by 88% and 89% respectively) and the lowest in the myocardium (by 53%). Under control conditions (without cysteine) kidney homogenates had the highest oxygen consumption and skeletal muscle ones the lowest.

Animals↗
PubMed1982

Dose-dependent induction of embryonic abnormalities in vitro by tissue homogenates of placenta and decidua.

Homogenate preparations from normal rat placental and decidual tissue induced abnormalities when included in the culture medium of rat embryos between Day 9.5 and Day 11.5. Abnormal embryos were produced between doses of 2.5 and 4 mg/ml for the placental homogenate and between doses of 1.2 and 4 mg/ml for the decidual homogenate, but were not produced by a solution of bovine serum albumin or by a protein preparation of rat lung tissue at the same concentration. The degree to which the embryos were malformed depended on the dose and which of the two homogenates was used. The decidual homogenate preparation was more pathogenic than the placental homogenate, but both were able to produce neural-tube defects and a severe reduction in embryonic size. The possible association between these findings and some known proteins within such homogenates is discussed.

Abnormalities, Drug-Induced↗