Lysosomes and the "toxicity" of Rickettsiales. IV. Ultrastructural studies of macrophages infected with a cytopathic L cell-grown C. psittaci 6BC strain

1973 ◽  
Vol 19 (3) ◽  
pp. 315-320 ◽  
Author(s):  
Nonna Kordová ◽  
Jan Hoogstraten ◽  
John C. Wilt
Keyword(s):  
Acid Phosphatase ◽  
Neutral Red ◽  
Contributory Factor ◽  
Enzyme Release ◽  
High Concentration ◽  
Ultrastructural Studies ◽  
Cytoplasmic Organelles ◽  
L Cell ◽  
Mouse Macrophages

Mouse macrophages in vitro inoculated with a low dose of a cytopathic L cell-grown C. psittaci 6BC strain were harvested at times when a delayed cytopathic effect (CPE) was observed by light microscopy and when a cytochemical test for acid phosphatase revealed lysosomal enzyme release. An array of different lysosomal types was revealed by ultrastructural studies such as digestive vesicles with single or double outer membranes and whorls of membranes, dense residual bodies, aggregated and single scattered ferritin-like granules, and fragments of membranes. Enlargement of cysternae of endoplasmic reticulum, disintegration of cytoplasmic organelles, and thinning of ground cytoplasm were also observed. Cytoplasmic degeneration has been observed both within membrane-limited structures and in wider areas not limited by recognizable membranes.A high concentration of the agent resulted in an early toxic effect; this was accompanied by a rapid vacuolization of the cytoplasm of the macrophages which showed a poorly developed ultrastructure, and absence of mitochondria and cytoplasmic organelles. Cytochemical tests indicated that these alterations correlated with release of lysosomal acid phosphatase and disintegration of neutral red stained granules. Ultrastructural studies support the view that disintegration of lysosomes may be considered an important contributory factor in the pathology of chlamydial infection.

Lysosomes and the "toxicity" of Rickettsiales. I. Cytochemical studies of macrophages inoculated in vitro with C. psittaci 6BC

1972 ◽  
Vol 18 (4) ◽  
pp. 457-464 ◽  
Author(s):  
Nonna Kordová ◽  
John C. Wilt
Keyword(s):  
Acid Phosphatase ◽  
Neutral Red ◽  
Immune Serum ◽  
Cytotoxic Action ◽  
Irreversible Damage ◽  
Lysosomal Acid ◽  
Control Serum ◽  
Mouse Macrophages ◽  
Rapid Destruction

Toxic doses of C. psittaci 6BC strain produced within 3–6 h after inoculation a rapidly progressive irreversible damage of mouse macrophages in vitro, an early leakage of a lysosomal enzyme, acid phosphatase, as well as a rapid disintegration of neutral red-stained cytoplasmic granules i.e. lysosomes. Pretreatment of the agent with immune serum neutralized the cytotoxic action of the agent and a reversible activation of lysosomes occurred. Pretreatment of the agent with undiluted control serum protected the macrophages from the early "toxic" injury; however, a cytopathic effect (CPE) occurred 48 h after inoculation and was accompanied by a release of lysosomal acid phosphatase. Heat-inactivated Chlamydiae produced an early agglutination of macrophages with leakage of lysosomal acid phosphatase, disintegration of neutral red-stained lysosomes, as well as a progressive irreversible damage of the macrophage. Concentrated, heat-inactivated agent-free, leuco-agglutinating material from 6BC infected L cells produced a similar injury of macrophages and their lysosomes. The findings are related to an earlier report of a rapid destruction of macrophages by C. psittaci in the intact animal (15). The possible mechanisms and factors of the "toxicity" of Rickettsiales are discussed.

scholarly journals THE PARTICULATE HYDROLASES OF MACROPHAGES

1963 ◽  
Vol 118 (6) ◽  
pp. 1009-1020 ◽  
Author(s):  
Zanvil A. Cohn ◽  
Edith Wiener
Keyword(s):  
Acid Phosphatase ◽  
Yeast Cell ◽  
Cell Walls ◽  
Hydrolytic Enzymes ◽  
Neutral Red ◽  
Biochemical Properties ◽  
Extracellular Medium ◽  
Cell Content ◽  
Prolonged Incubation

The influence of phagocytosis on the morphological and biochemical properties of macrophage hydrolase-containing granules has been studied in vitro. Following the uptake of large numbers of heat-killed bacteria, an intracellular rearrangement of hydrolytic enzymes occurred. This was associated with the solubilization of 50 to 60 per cent of the total cell content of acid phosphatase, cathepsin, lysozyme, beta glucuronidase, acid ribonuclease, and acid desoxyribonuclease and with a corresponding decrease in granule-bound enzyme. With more prolonged incubation the majority of the soluble intracellular pool of acid ribonuclease and lysozyme was lost to the extracellular medium. No change in the total content of any of the hydrolases was noted during 180 minutes of incubation in vitro. The morphological fate of the granules was studied by a histochemical method for acid phosphatase. After the phagocytosis of yeast cell walls there was a disappearance of acid phosphatase-positive granules and an accumulation of reaction product about the ingested particle. Experiments employing macrophages which were supravitally stained with neutral red also demonstrated the loss of neutral red-positive granules and the accumulation of the dye about the yeast cell walls. These results strongly suggest that lysis of macrophage granules occurs following phagocytosis and that a portion of the granule contents are then resegregated within the newly formed phagocytic vacuole.

Lysosomes and the "toxicity" of Rickettsials. II. Non-cytocidal interactions of egg-grown C. psittaci 6BC and in vitro macrophages

1972 ◽  
Vol 18 (6) ◽  
pp. 869-873 ◽  
Author(s):  
Nonna Kordová ◽  
Linda Poffenroth ◽  
John C. Wilt
Keyword(s):  
Acid Phosphatase ◽  
Cell Damage ◽  
Giant Cells ◽  
Host Cells ◽  
Marked Contrast ◽  
Lysosomal Acid ◽  
Cytoplasmic Vacuoles ◽  
L Cell

During the infection of cultured mouse peritoneal phagocytes with egg-grown C. psittaci 6BC strain, lysosomes retained their integrity and host cells were not damaged. Infected monocytes showed greater ability than uninfected monocytes to spread and transform into giant cells containing enlarged nuclei and masses of cytoplasm with clear cytoplasmic vacuoles. Chlamydial particles were released from the cytoplasm of infected phagocytes by pseudopodia-like extrusions. These events were in marked contrast to the effect of L cell grown C. psittaci 6BC strain which caused early leakage of lysosomal acid phosphatase into the cytoplasm of macrophages and induced a rapidly progressive irreversible cell damage (14).

scholarly journals THE DIFFERENTIATION OF MONONUCLEAR PHAGOCYTES

1965 ◽  
Vol 121 (1) ◽  
pp. 153-170 ◽  
Author(s):  
Zanvil A. Cohn ◽  
Belinda Benson
Keyword(s):  
Acid Phosphatase ◽  
Specific Activity ◽  
Total Content ◽  
Progressive Increase ◽  
Neutral Red ◽  
Mononuclear Phagocytes ◽  
In Vitro Differentiation ◽  
Dense Granules

The in vitro differentiation of homogeneous populations of monocyte-like cells from the unstimulated mouse peritoneal cavity is described. Under the conditions employed, a progressive increase in cell size occurs without significant cell division. This process is characterized morphologically by the accumulation of phase-dense and neutral red-positive granules, mitochondria, and lipid droplets. The phase-dense granules react strongly for acid phosphatase. Biochemical determinations indicate marked increases in the total content and specific activity of acid phosphatase, cathepsin, and ß-glucuronidase. The production of acid phosphatase is more rapid and extensive than that of the other two hydrolases. From these data it appears that the conversion of a monocyte-like cell to a mature macrophage is accompanied by the formation of increased numbers of lysosome-like cytoplasmic organelles. Mouse peritoneal phagocytes stimulated in vivo with a bacterial lipopolysaccharide undergo a similar series of morphological and biochemical events.

Probing the ultrastructure of the mitotic and meiotic spindle in eggs: An expeditious approach based on semi-thin (0.25 μm) serial sections

1983 ◽  
Vol 41 ◽  
pp. 552-555
Author(s):  
Conly L. Rieder ◽  
S. Bowser ◽  
R. Nowogrodzki ◽  
K. Ross ◽  
G. Sluder
Keyword(s):  
Small Sample Size ◽  
Small Sample ◽  
Meiotic Spindle ◽  
Serial Sections ◽  
Mitotic Apparatus ◽  
Thin Sections ◽  
Cell Cycles ◽  
Ultrastructural Studies

Eggs have long been a favorite material for studying the mechanism of karyokinesis in-vivo and in-vitro. They can be obtained in great numbers and, when fertilized, divide synchronously over many cell cycles. However, they are not considered to be a practical system for ultrastructural studies on the mitotic apparatus (MA) for several reasons, the most obvious of which is that sectioning them is a formidable task: over 1000 ultra-thin sections need to be cut from a single 80-100 μm diameter egg and of these sections only a small percentage will contain the area or structure of interest. Thus it is difficult and time consuming to obtain reliable ultrastructural data concerning the MA of eggs; and when it is obtained it is necessarily based on a small sample size.We have recently developed a procedure which will facilitate many studies concerned with the ultrastructure of the MA in eggs. It is based on the availability of biological HVEM's and on the observation that 0.25 μm thick serial sections can be screened at high resolution for content (after mounting on slot grids and staining with uranyl and lead) by phase contrast light microscopy (LM; Figs 1-2).

Ultrastructural studies on the interaction of haemophilus influenzae with human endothelial cells in vitro

1990 ◽  
Vol 48 (3) ◽  
pp. 636-637
Author(s):  
D.J.P. Ferguson ◽  
M. Virji ◽  
H. Kayhty ◽  
E.R. Moxon
Keyword(s):  
Endothelial Cells ◽  
Haemophilus Influenzae ◽  
Intercellular Junctions ◽  
Blood Stream ◽  
Ultrastructural Studies ◽  
Endothelial Barriers ◽  
Serotype B ◽  
Meningitis In Children ◽  
Respiratory Epithelial

Haemophilus influenzae is a human pathogen which causes meningitis in children. Systemic H. influenzae infection is largely confined to encapsulated serotype b organisms and is a major cause of meningitis in the U.K. and elsewhere. However, the pathogenesis of the disease is still poorly understood. Studies in the infant rat model, in which intranasal challenge results in bacteraemia, have shown that H. influenzae enters submucosal tissues and disseminates to the blood stream within minutes. The rapidity of these events suggests that H. influenzae penetrates both respiratory epithelial and endothelial barriers with great efficiency. It is not known whether the bacteria penetrate via the intercellular junctions, are translocated within the cells or carried across the cellular barrier in 'trojan horse' fashion within phagocytes. In the present studies, we have challenged cultured human umbilical cord_vein endothelial cells (HUVECs) with both capsulated (b+) and capsule-deficient (b-) isogenic variants of one strain of H. influenzae in order to investigate the interaction between the bacteria and HUVEC and the effect of the capsule.

Histochemical and Ultrastructural Studies of the Digestive Epithelium in a Gastropod Gametolytic Organ

1980 ◽  
Vol 38 ◽  
pp. 568-569
Author(s):  
R. L. Reeder ◽  
S. H. Rogers ◽  
W. A. Shannon
Keyword(s):  
Acid Phosphatase ◽  
Genital Tract ◽  
Reproductive Tract ◽  
Ultrastructural Level ◽  
Conclusive Evidence ◽  
Digestive Organ ◽  
Limited Information ◽  
Ultrastructural Studies ◽  
Morphological Studies

Numerous morphological studies have dealt with the spermatheca of pulmonate gastropods. This globular organ, which is attached to the female portion of the reproductive tract by a long duct in these monoecious animals, has had various functions ascribed to it. Recent histochemical demonstrations of deoxyribonuclease, ribonuclease, protease, and acid phosphatase have provided, however, conclusive evidence that it is a digestive organ for the degradation of superfluous sperm and genital tract secretions. Only limited information concerning the spermatheca is available at the ultrastructural level, a fact providing the stimulus for the present study of this organ in Sonorella santaritana, a desert mountain snail from Arizona.

The Effects of Shock Vancomycin Concentrations on the Formation of Heteroresistance in Staphylococcus aureus

2020 ◽  
Vol 65 (9-10) ◽  
pp. 3-7
Author(s):  
V. V. Gostev ◽  
Yu. V. Sopova ◽  
O. S. Kalinogorskaya ◽  
M. E. Velizhanina ◽  
I. V. Lazareva ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
In Vitro Selection ◽  
Methicillin Resistant Staphylococcus Aureus ◽  
High Concentration ◽  
Short Term ◽  
High Concentrations ◽  
Selection Of ◽  
Selection Of Resistance ◽  
Test Cultures

Glycopeptides are the basis of the treatment of infections caused by MRSA (Methicillin-Resistant Staphylococcus aureus). Previously, it was demonstrated that antibiotic tolerant phenotypes are formed during selection of resistance under the influence of high concentrations of antibiotics. The present study uses a similar in vitro selection model with vancomycin. Clinical isolates of MRSA belonging to genetic lines ST8 and ST239, as well as the MSSA (ATCC29213) strain, were included in the experiment. Test isolates were incubated for five hours in a medium with a high concentration of vancomycin (50 μg/ml). Test cultures were grown on the medium without antibiotic for 18 hours after each exposure. A total of ten exposure cycles were performed. Vancomycin was characterized by bacteriostatic action; the proportion of surviving cells after exposure was 70–100%. After selection, there was a slight increase in the MIC to vancomycin (MIC 2 μg/ml), teicoplanin (MIC 1.5–3 μg/ml) and daptomycin (MIC 0.25–2 μg/ml). According to the results of PAP analysis, all strains showed an increase in the area under curve depending on the concentration of vancomycin after selection, while a heteroresistant phenotype (with PAP/AUC 0.9) was detected in three isolates. All isolates showed walK mutations (T188S, D235N, E261V, V380I, and G223D). Exposure to short-term shock concentrations of vancomycin promotes the formation of heteroresistance in both MRSA and MSSA. Formation of VISA phenotypes is possible during therapy with vancomycin.

Vancomycin Adsorption During in vitro Model of Hemoperfusion with HA380 Cartridge

Nephron ◽  
2021 ◽  
pp. 1-7
Author(s):  
Ilaria Godi ◽  
Anna Lorenzin ◽  
Silvia De Rosa ◽  
Gianlorenzo Golino ◽  
Maira Knust ◽  
...  
Keyword(s):  
Complete Removal ◽  
Potential Interaction ◽  
Blood Purification ◽  
Therapeutic Monitoring ◽  
High Concentration ◽  
Langmuir Isotherm Model ◽  
Vitro Model ◽  
Kinetics Of ◽  
Balanced Solution

<b><i>Introduction:</i></b> A critical point for using blood purification during sepsis may be the potential interaction with antimicrobial therapy, the mainstay of sepsis treatment. The aim of our study was to investigate the vancomycin removal during hemoperfusion (HP) using HA380 cartridge. <b><i>Methods:</i></b> This is an experimental study, in which 500 mL of solution was circulated in a closed-circuit (blood flow of 250 mL/min) simulating HP ran using HA380. Vancomycin was added to reach a through concentration or a very high concentration to evaluate the removal ratio (RR) during 120 min of HP. Comparison between blood-crystalloid solution and balanced solution was performed by using Kruskal-Wallis test. The kinetics of vancomycin removal and the adsorption isotherm were evaluated. <b><i>Results:</i></b> We found a complete removal of vancomycin at baseline through concentration of 23.0 ± 7.4 mg/L. Using extremely high concentration (baseline 777.0 ± 62.2 mg/L), RR was 90.1 ± 0.6% at 5 min and 99.2 ± 0.6% at 120 min. No difference in terms of RR was found between blood-crystalloid mixture and balanced solution. The kinetics of the vancomycin reduction followed an exponential decay. Repeated boluses (total amount of 2,000 mg) resulted in cumulative adsorption of 1,919.4 mg with RR of 96.6 ± 1.4%, regardless of the amount injected (100 vs. 500 mg). Vancomycin adsorption onto HA380 followed the Langmuir isotherm model. <b><i>Conclusions:</i></b> A considerable amount of vancomycin was rapidly removed during in vitro HP with HA380. Clinical studies are needed to determine whether this may lead to underdosing. Drug therapeutic monitoring is highly recommended when using HA380 for blood purification in patients receiving vancomycin.

scholarly journals Replacement of Albumin by Preovulatory Oviductal Fluid in Swim-Up Sperm Preparation Method Modifies Boar Sperm Parameters and Improves In Vitro Penetration of Oocytes

Animals ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 1202
Author(s):  
Sergio Navarro-Serna ◽  
Evelyne París-Oller ◽  
Ondrej Simonik ◽  
Raquel Romar ◽  
Joaquín Gadea
Keyword(s):  
Calcium Concentration ◽  
Preparation Method ◽  
Culture Media ◽  
Penetration Rate ◽  
Sperm Parameters ◽  
High Concentration ◽  
Sperm Preparation ◽  
Boar Sperm ◽  
Oviductal Fluid

More suitable and efficient methods to protect gametes from external harmful effects during in vitro handling can be achieved by adding preovulatory porcine oviductal fluid (pOF) to in vitro culture media. The objective of this study was to assess the swim-up procedure’s suitability as a sperm selection method using a medium supplemented with 1mg/mL BSA, 1% preovulatory pOF (v/v), 1% v/v pOF plus 1mg/mL BSA, and 5mg/mL BSA. After selection, various sperm parameters were studied, such as sperm recovery rate, sperm morphology, motility (by CASA), vitality, acrosome status and intracellular calcium (by flow cytometry) and ability to penetrate oocytes in vitro. Around 2% of sperm were recovered after swim-up, and the replacement of BSA by pOF showed a beneficial reduction of motility parameters calcium concentration, resulting in an increased penetration rate. The combination of albumin and oviductal fluid in the medium did not improve the sperm parameters results, whereas a high concentration of BSA increased sperm morphological abnormalities, motility, and acrosome damage, with a reduction of calcium concentration and penetration rate. In conclusion, the replacement of albumin by preovulatory oviductal fluid in the swim-up sperm preparation method modifies boar sperm parameters and improves the in vitro penetration of oocytes.

Sign in / Sign up

Export Citation Format

Share Document