Formazan Granules Research Articles

Study of cellular immune response in mice Balb/c teated with LPS of Klebsiella pneumonia antigen and Glycyrrhiza glabra extract.

In this study, lipopolysaccharides of Klebsiella pneumonia and concentrate of Glycyrrhiza glabra were utilized to examine the cellular immunity in mice Balb/c (in vivo). Conversely, little is thought about cell invulnerability prompted by lipopolysaccharides and Glycyrrhiza glabra extract. A few parameters were utilized to accomplish this investigation, are neutrophil WBC (PMNs) rate, phagocytosis coefficient of PMNs at various eras 30, 60, 90 and 120 minutes, Formazan granules arrangement in them, and movement restraint factor (MIF).

Evaluation of influence several drugs with local antimicrobial activity against local immunity cells

Currently in clinical practice widely used drugs local antimicrobial drugs such as Tantum Verde (benzydamine) Tantum Rosa (benzydamine) Miramistin (benzyldimethyl-miristoilamino-propylammonium) Hexoral (hexetidin), chlorhexidine (chlorhexidine), Septolete total (benzydamine + cetylpyridinium chloride). The mechanism of action of these very similar. We evaluated the effect of these drugs on the viability of lymphoid tissue cells and their effect on the neutrophilic part of the immune system, which are the most important factors of local immunity and, at the same time, part of the immune system which is responsible for innate immunity. We used peripheral blood from 6 healthy donors and 6 patients with inflammatory diseases (abscess of the abdominal cavity). Evaluation of the viability of lymphocytes was performed in a test using trypan blue. The functional state of neutrophils was performed in a nitro-blue tetrazolium test. The final concentration of the studied drugs in all experiments was 10% of the initial recommended for local use. The results of the study showed that Miramistin, Hexoral, Chlorhexidine, Septolete Total cause the death of lymphocytes isolated from healthy donors and patients with severe inflammation process. Tantum Verde and Tantum Rose do not cause the death of lymphocytes. All studied drugs (except Tantum Verde and Tantum Rose in healthy donors) reduce the number of neutrophils containing formazan granules, which indicates the suppression of the activity of the NADPH oxidase system. Patients’ neutrophils witch were activated by inflammatory process under the influence of miramistin experiencing short-term excessive activation of the NADPH-oxidase system, which can lead to tissue damage in severe inflammation.

Results of cytochemical investigation of neutrophil granulocytes in patients with purulent-inflammatory diseases of fine tissues on the type 2 diabetes mellitus

Patients with purulent-inflammatory diseases of soft tissues on the background of diabetes type 2 are resistant changes in various parts of the immune system. Hyperglycemia, hyperlipidemia, insulin resistance and adaptive hyperinsulinemia affect the cells of the immune system, promote the development of metabolic immunosuppression with the formation of stable immune dysfunction. The purpose of the study to investigate and analyze the metabolic state of immunocompetent cells in patients with purulent-inflammatory diseases of soft tissues on the background of DMD type 2. The study was conducted in 47 patients (the main group) with purulent-inflammatory diseases of soft-tissue on the background of diabetes mellitus, and 20 healthy volunteers (the comparison group) using the cytohistochemical restoration method of NST based on the percentage composition of neutrophils that have cytoplasm of formazan granules. The spontaneous neutrophil response was studied, as well as the parameters of the NST-test in stimulation of neutrophil granulocytes in vitro to assess the bactericidal, resource potential and their ability to complete phagocytosis. Results of the study showed that the decrease in the activity of myeloperoxidase and the NST-test in the conditions of stimulation of neutrophil granulocytes is a sign of unsatisfactory state of the bactericidal system of cells, which may be the cause of acute inflammatory processes in the body that accompany type 2 diabetes, which leads to functional exhaustion of the macrophage level of the immune body protection. At GZZMT against the background of DM 2 type compared with the norm, the “spontaneous” NST-test of neutrophils is high, which is due to the presence of inflammatory process. The index of stimulation of neutrophils is lower in the main group (1.6 times: 5.3±0.2% in the main group versus 8.4±0.4% in the control, p≤0.05), which is an indicator of a relatively weaker stimulation of those populations of neutrophils that were responsible for high indices in the “spontaneous” NST test, and the impaired state of cellular metabolism. So, in patients with type 2 diabetes, a violation of the bactericidal system of organism protection, which is one of the causes of the development of infectious and inflammatory processes.

Effects of Lipopolysaccharides of Pseudomonas Aeruginosa and Aqueous Extract of Ginkgo Biloba, Ginkgoaceae, on Cellular Immune Response in Mice Balb/c

In this study, lipopolysaccharides (LPS) of Pseudomonas aeruginosa IAN5 isolate and aqueous extract of Ginkgo biloba leaves were used to investigate the cellular immunity in mice Balb/c (in vivo). Some parameters were used to achieve this study, are percentages of polymorphonuclear neutrophils (PMNs), phagocytosis coefficient of PMNs at different time periods 30, 60, 90 and 120 minutes, Formazan granules formation in them, and migration inhibition factor (MIF). The immunization of mice with the LPS antigen affected delay-type hypersensitivity and increased activity of PMNs in Candida sp. and reduced NBT, while inhibited migration of PMNs. The immunization did not affect the macrophages of PMNs. The injected treatment with LPS and Ginkgo biloba extract showed the best results significantly (p<0.05) compared to LPS and control treatments, whereas the Ginkgo biloba extract alone showed no significant differences (p<0.05).

Microscopic analysis of MTT stained boar sperm cells

The ability of sperm cells to develop colored formazan by reduction of MTT was used earlier to develop a spectrophotometric assay to determine the viability of sperm cells for several mammalian species. It was the objective of the present study to visualize microscopically the location of the formazan in boar sperm cells. The MTT staining process of boar sperm cells can be divided into a series of morphological events. Incubation of the sperm cells in the presence of MTT resulted after a few min in a diffuse staining of the midpiece of the sperm cells. Upon further incubation the staining of the midpiece became more intense, and gradually the formation of packed formazan granules became more visible. At the same time, a small formazan stained granule appeared medially on the sperm head, which increased in size during further incubation. After incubation for about 1 h the midpiece granules were intensely stained and more clearly distinct as granules, while aggregation of sperm cells occurred. Around 90% of the sperm cells showed these staining events. At the end of the staining the formazan granules have disappeared from both the sperm cells and medium, whereas formazan crystals appeared as thin crystal threads, that became heavily aggregated in the incubation medium. It was concluded that formazan is taken up by lipid droplets in the cytoplasm. Further, the use of the MTT assay to test for sperm viability should be regarded as a qualitative assay, whereas its practical use at artificial insemination (AI) Stations is limited.

Vital Dye Reaction and Granule Localization in Periplasm of Escherichia coli

BackgroundTetrazolium salts are widely used in biology as indicators of metabolic activity – hence termed vital dyes – but their reduction site is still debated despite decades of intensive research. The prototype, 2,3,5- triphenyl tetrazolium chloride, which was first synthesized a century ago, often generates a single formazan granule at the old pole of Escherichia coli cells after reduction. So far, no explanation for their pole localization has been proposed.Method/Principal FindingsHere we provide evidence that the granules form in the periplasm of bacterial cells. A source of reducing power is deduced to be thiol groups destined to become disulfides, since deletion of dsbA, coding for thiol-oxidase, enhances the formation of reduced formazan. However, pervasive reduction did not result in a random distribution of formazan aggregates. In filamentous cells, large granules appear at regular intervals of about four normal cell-lengths, consistent with a diffusion-to-capture model. Computer simulations of a minimal biophysical model showed that the pole localization of granules is a spontaneous process, i.e. small granules in a normal size bacterium have lower energy at the poles. This biased their diffusion to the poles. They kept growing there and eventually became fixed.ConclusionsWe observed that formazan granules formed in the periplasm after reduction of tetrazolium, which calls for re-evaluation of previous studies using cell-free systems that liberate inaccessible intracellular reductant and potentially generate artifacts. The localization of formazan granules in E. coli cells can now be understood. In living bacteria, the seeds formed at or migrated to the new pole would become visible only when that new pole already became an old pole, because of the relatively slow growth rate of granules relative to cell division.

MTT assay for cell viability: Intracellular localization of the formazan product is in lipid droplets

Although MTT is widely used to assess cytotoxicity and cell viability, the precise localization of its reduced formazan product is still unclear. In the present study the localization of MTT formazan was studied by direct microscopic observation of living HeLa cells and by colocalization analysis with organelle-selective fluorescent probes. MTT formazan granules did not colocalize with mitochondria as revealed by rhodamine 123 labeling or autofluorescence. Likewise, no colocalization was observed between MTT formazan granules and lysosomes labeled by neutral red. Taking into account the lipophilic character and lipid solubility of MTT formazan, an evaluation of the MTT reaction was performed after treatment of cells with sunflower oil emulsions to induce a massive occurrence of lipid droplets. Under this condition, lipid droplets revealed a large amount of MTT formazan deposits. Kinetic studies on the viability of MTT-treated cells showed no harmful effects at short times. Quantitative structure–activity relations (QSAR) models were used to predict and explain the localization of both the MTT tetrazolium salt and its formazan product. These predictions were in agreement with experimental observations on the accumulation of MTT formazan product in lipid droplets.

Proteomic Identification of Betaig-h3 as a Lysophosphatidic Acid-Induced Secreted Protein of Human Mesenchymal Stem Cells: Paracrine Activation of A549 Lung Adenocarcinoma Cells by Betaig-h3

Lysophosphatidic acid (LPA) is enriched in the serum and malignant effusion of cancer patients and plays a key role in tumorigenesis and metastasis. LPA-activated mesenchymal stem cells promote tumorigenic potentials of cancer cells through a paracrine mechanism. LPA-conditioned medium (LPA CM) from human adipose tissue-derived mesenchymal stem cells (hASCs) elicited adhesion and proliferation of A549 human lung adenocarcinoma cells. To identify proteins involved in the LPA-stimulated paracrine functions of hASCs, we analyzed the LPA CM using liquid-chromatography tandem mass spectrometry-based shotgun proteomics. We identified βig-h3, an extracellular matrix protein that is implicated in tumorigenesis and metastasis, as an LPA-induced secreted protein in hASCs. LPA-induced βig-h3 expression was abrogated by pretreating hASCs with the LPA receptor(1/3) inhibitor Ki16425 or small interfering RNA-mediated silencing of endogenous LPA(1). LPA-induced βig-h3 expression was blocked by treating the cells with the Rho kinase inhibitor Y27632, implying that LPA-induced βig-h3 expression is mediated by the LPA(1)- Rho kinase pathway. Immunodepletion or siRNA-mediated silencing of βig-h3 abrogated LPA CM-stimulated adhesion and proliferation of A549 cells, whereas retroviral overexpression of βig-h3 in hASCs potentiated it. Furthermore, recombinant βig-h3 protein stimulated the proliferation and adhesion of A549 human lung adenocarcinoma cells. These results suggest that hASC-derived βig-h3 plays a key role in tumorigenesis by stimulating the adhesion and proliferation of cancer cells and it can be applicable as a biomarker and therapeutic target for lung cancer.

Localization of MTT formazan in lipid droplets. An alternative hypothesis about the nature of formazan granules and aggregates.

MTT (3-(4, 5-dimethyl-2-thiazolyl)-2, 5-dihphenyltetrazolium bromide) assay is a widely used method to assess cell viability and proliferation. MTT is readily taken up by cells and enzymatically reduced to formazan, a dark compound which accumulates in cytoplasmic granules. Formazan is later eliminated by the cell by a mechanisms often indicated as exocytosis, that produces characteristic needle-like aggregates on the cell surface. The shape of formazan aggregates and the rate of exocytosis change in the presence of bioactive amyloid beta peptides (Abeta) and cholesterol. Though the cellular mechanisms involved in MTT reduction have been extensively investigated, the exact nature of formazan granules and the process of exocytosis are still obscure. Using Nile Red, which stains differentially neutral and polar lipids, and a fluorescent analog of cholesterol (NBD-cholesterol), we found that formazan localized in lipid droplets, consistent with the lipophilic nature of formazan. However, formazan granules and aggregates were also found to form after killing cells with paraformaldehyde fixation. Moreover, formazan aggregates were also obtained in cell-free media, using ascorbic acid to reduce MTT. The density and shape of formazan aggregates obtained in cell-free media was sensitive to cholesterol and Abeta. In cells, electron microscopy failed to detect the presence of secretory vesicles, but revealed unusual fibers of 50 nm of diameter extending throughout the cytoplasm. Taken together, these findings suggest that formazan efflux is driven by physico-chemical interactions at molecular level without involving higher cytological mechanisms.

Cholesterol Interferes with the MTT Assay in Human Epithelial-Like (A549) and Endothelial (HLMVE and HCAE) Cells

Metabolically active cells are able to convert the MTT [3-(4,5-dimethythiazol-2-yl)-2,5-diphenyltetrazolium bromide] dye to blue formazan. This is the basis of the MTT assay, which is among the most widely used screening methods to evaluate cell viability and proliferation. When testing the effects of cholesterol products on the viability of human pulmonary epithelial-like A549 cells using trypan blue staining (cell numbers) and the MTT assay, results were inconsistent. The MTT assay indicated greater than 50% loss of viability with exposure of cells to cholesterol, whereas there was no decrease in viability indicated by trypan blue exclusion and propidium iodide uptake. A similar decrease in MTT reduction was obtained upon cholesterol treatment in human lung microvascular endothelial cells (HLMVECs) and human coronary artery endothelial cells (HCAECs) without loss of viability. This suggested a direct interference of cholesterol with the assay. However, using a cell-free system, there was no decrease in the reduction of MTT by ascorbic acid during incubation with a similar concentration of cholesterol. Light microscopy revealed enhanced exocytosis of formazan granules in presence of cholesterol. Incubation with apolipoprotein A-1 decreased cholesterol-mediated inhibition of MTT assay. These studies indicate decreased MTT reduction as a result of enhanced exocytosis of formazan due to cholesterol. A careful validation of viability assay procedures is therefore suggested in experiments where cholesterol is a constituent, to avoid a potential bias in concluding results of cytotoxicity studies.

Separation of active and inactive fractions from starved culture of Vibrio parahaemolyticus by density dependent cell sorting

The co-existence of physiologically different cells in bacterial cultures is a general phenomenon. We have examined the applicability of the density dependent cell sorting (DDCS) method to separate subpopulations from a long-term starvation culture of Vibrio parahaemolyticus. The cells were subjected to Percoll density gradient and separated into 12 fractions of different buoyant densities, followed by measuring the cell numbers, culturability, respiratory activity and leucine incorporation activity. While more than 78% of cells were in lighter fractions, about 95% of culturable cells were present in heavier fractions. The high-density subpopulations also had high proportion of cells capable of forming formazan granules. Although this was accompanied by the cell specific INT-reduction rate, both leucine incorporation rates and INT-reduction rates per cell had a peak at mid-density fraction. The present results indicated that DDCS could be used to separate subpopulations of different physiological conditions.

Enhancement of MTT, a tetrazolium salt, exocytosis by amyloid β-protein and chloroquine in cultured rat astrocytes

The effect of amyloid β-protein (A β) on the cellular reducing activity has been a controversial issue. We determined the cellular reducing activity in cultured astrocytes using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium (WST-8) reduction assays following A β treatment. MTT reduction was inhibited whereas WST-8 reduction was unaffected by the A β treatment. Furthermore, the early extracellular appearance of MTT formazan, a reduced product of MTT, was observed in association with the rapid disappearance of intracellular formazan granules. Notably, similar results were obtained in cultures treated with chloroquine, a perturbant of membrane trafficking. Our results suggest that MTT formazan exocytosis is enhanced in a similar manner by A β and chloroquine and that this biological activity of A β may underlie the pathogenesis of Alzheimer's disease.

Effects of parent Shamo cocks on the histochemical properties of M. iliotibialis lateralis and M. supracoracoideus on their crossbred broilers

1. Four Shamo (a Japanese game bird) cocks showing different characteristics in the histochemical properties of M. iliotibialis lateralis (ITL) were crossed with White Rock hens to produce male and female crossbred broilers of the 4 lines (90 d of age). Normal broilers (56 d) were used, for comparison. 2. Histochemical properties of ITL and M. supracoracoideus (SC) were compared among the crossbred lines and normal broilers. Myofibres were divided into Types II R, II I and II W showing high, moderate and low reduced nicotinamide adenine dinucleotide dehydrogenase (NADH-DH) activities, respectively. 3. In the ITL of the crossbred cockerels, the percentage of Type II R and II I fibres decreased and conversely Type II W increased in comparison to those in the Shamo. 4. Sex differences of the histochemical properties were recognised only in the ITL of the crossbred, in which the percentage of Type II R fibres was greater in the male. 5. The different characteristics of the parent Shamo cocks were reproduced only in the different fibre type composition of the ITL muscle in the crossbred cockerels. 6. The histochemical features of fibre type seemed to develop with bird age, particularly subsarcolemmal accumulation of formazan granules (indicating high NADH-DH activity) in Type IIR fibres. 7. Breed, line, sex and age differences in the histochemical properties were demonstrated clearly in ITL but not in SC.

Comparative studies on the histochemical properties of M. iliotibialis lateralis from Kumamoto Cochin crossbred roaster and broiler chickens

1. Histochemical properties of M. iliotibialis lateralis were compared between Kumamoto Cochin (a Japanese native breed) crossbred roasters (KC roasters, 112 d of age) and normal broilers (56 d of age). 2. Myofibres were divided into Types II‐R, II‐I and II‐W showing high, moderate and low reduced nicotinamide adenine dinucleotide dehydrogenase (NADH‐DH) activities, respectively. 3. Subsarcolemmal formazan granules indicating very strong NADH‐DH activity in Type II‐R fibres were observed only in KC roasters. In both sexes the percentage of Type II‐R fibres present was greater in KC roasters than in broilers. 4. These results indicated marked differences in histochemical properties and meat quality between KC roasters and broilers.

Neutrophil NADPH oxidase activity in chronic myeloproliferative and myelodysplastic diseases by microscopic and photometric assays.

Neutrophil NADPH oxidase is a membrane-bound enzyme complex responsible for the reduction of oxygen to superoxide anion in the respiratory burst. Impaired neutrophil function has often been reported in myeloproliferative diseases and myelodysplastic syndromes (MDSs), but its laboratory features have not been well characterized. Traditionally, NADPH oxidase activity has been evaluated with a microscopic method by nitroblue tetrazolium salt reduction to blue-black insoluble formazan granules identified in positive neutrophils by microscopy. We investigated neutrophil NADPH oxidase in 22 patients with chronic myeloproliferative disorders (cMPDs) and 15 patients with MDSs, using the microscopic method as well as a photometric microplate assay, which monitored the cytochemical reaction for 30 min. The relationship between cMPD and patient susceptibility to infections was also investigated. In the photometric assay, the mean enzyme activity in MDSs and cMPDs was lower than in normal subjects. NADPH oxidase activity was greater in cMPDs (except myelofibrosis) than in MDSs (except chronic myelomonocytic leukemia), and these differences were less evident with microscopy. Moreover, for cMPDs, patients with susceptibility to infections showed a lower NADPH oxidase activity than patients without infections.

Survival of Denitrifiers in Nitrate-Free, Anaerobic Environments

Experiments were undertaken to explain the occurrence of a high denitrification capacity in anaerobic, NO(3)-free habitats. Deep layers of freshwater sediments that were buried more than 40 years ago and digested sludge were the habitats studied. The denitrifier populations were 3.1 x 10 and 3.1 x 10 cells cm in deep sediments from a river and lake, respectively, and 5.3 x 10 cells cm in digested sludge. The denitrification capacities of the samples reflected the population densities. Strict anaerobic procedures were used to obtain the predominant isolates that would grow on anaerobic medium with NO(3). All strict anaerobes isolated failed to denitrify. All isolates that denitrified were aerobic, gram-negative bacteria, particularly species of Pseudomonas and Alcaligenes. No detectable growth was observed when these strains were incubated with electron acceptors other than NO(3) or O(2). When representative isolates were added to sterile, O(2)- and NO(3)-free porewater from their original locations at their natural densities (10 cells cm), no change in viable population was noted over 3 months of incubation. Metabolic activity was demonstrated in these cells by slow formation of formazan granules when exposed to tetrazolium and by observation of motile cells. When [C]glucose was added to cell suspensions of the pseudomonads that had been starved for 3 months without electron acceptors (O(2) or NO(3)), C-labeled products, including cell biomass, CO(2), and fermentation products, were produced. The high denitrification capacity of these anaerobic environments appears to be due to conventional respiratory denitrifiers. These organisms have the capacity for long-term survival without O(2) or NO(3) and appear to be capable of providing for their maintenance by carrying on a low level of fermentation.

Oxygen derived free radicals in osteoclasts: The specificity and location of the nitroblue tetrazolium reaction

Oxygen derived free radicals are generated by osteoclasts. In a novel culture system, isolated rat osteoclasts were stained when nitroblue tetrazolium (NBT) was reduced by cellular oxidants to formazan, an insoluble precipitate. Superoxide dismutase (SOD) inhibited the accumulation of formazan by the isolated osteoclasts. Osteoclasts in mouse calvarial organ cultures also reduced NBT to formazan. The reaction products were localized to the area of the osteoclast-bone interface. At the light microscopic level, the formazan granules appeared to be concentrated within the cytoplasm. Formazan accumulation was significantly inhibited by calcitonin (hCT). The inhibition of NBT reduction by SOD indicates that the isolated osteoclasts were capable of producing superoxide. The localization of the formazan granules between the external osteoclastic membrane and the bone, and the inhibition of this reaction during hCT exposure suggests that oxygen derived free radicals may contribute to bone resorption.

Early morphological changes in the striated muscles in normal and dystrophic chickens

Histological and histochemical analyses were performed on the anterior latissimus dorsi muscle (ALD, red muscle) and the posterior latissimus dorsi muscle (PLD, white muscle) in normal (line 412) and dystrophic chickens (line 413) from 19 day embryos to 6 weeks of age. PLD, the white muscle, in dystrophic chickens showed higher percentages of red and intermediate fibres than those of normal chickens during the early development of muscles. Increases of the oxidative enzyme activities and the numbers of NADH-TR formazan granules in the white fibres of PLD were already found at 1 week of age in dystrophic chicken. Fibre types, oxidative enzyme activities and NADH-TR formazan granules showed no differences in ALD between normal and dystrophic chickens. These results suggest that increases of oxidative enzyme activities and formazan granule numbers and incomplete fibre type differentiation in PLD of dystrophic chickens are early pathological processes in such birds.

Electron-cytochemical localization of succinic semialdehyde dehydrogenase activity in Purkinje neurons and hepatocytes of the rat

A method is presented for the ultrastructural demonstration of succinate semialdehyde dehydrogenase (SSADH) activity in cerebellar Purkinje neurons and liver hepatocytes; SSADH is an enzyme involved in the degradation of γ-aminobutyric acid (GABA). Incubation media originally used for light microscopy were considered. Reaction products were mainly detected when fresh tissue was used. In Purkinje cells, grains ascribable to SSADH activity were localized on the mitochondria (especially on the outer membrane); some extramitochondrial formazan deposits were also found. After brief fixation by immersion or perfusion, only a few formazan granules were detected in the cytoplasm. A similar distribution pattern was observed in hepatocytes, in which extramitochondrial grains and grains on the nuclear membrane were frequent. The actual existence and the possible meaning of extramitochondrial SSADH activity is critically discussed on the basis of the data in the literature.

Nitrofurantoin-stimulated oxidant production in pulmonary endothelial cells.

Nitrofurantoin, a urinary antiseptic, is associated with significant pulmonary toxicity. Our study indicates that nitrofurantoin may produce lung injury by directly stimulating lung parenchymal cells to generate toxic oxygen species such as superoxide and hydrogen peroxide, which can overwhelm cellular antioxidant defenses and result in permanent injury to the cells. Nitrofurantoin (10(-3) mol/L) stimulates bovine pulmonary artery endothelial cells to release 3.7 +/- 0.4 mumol/L superoxide per 10(5) cells and 4.4 +/- 0.5 mumol/L hydrogen peroxide per 10(5) cells (p less than 0.001 compared with endothelial cells without nitrofurantoin). Endothelial cells treated with nitroblue tetrazolium, a yellow dye reduced by superoxide to insoluble blue formazan, can be monitored both spectrophotometrically and morphologically. Nitrofurantoin (10(-5), 10(-4), and 10(-3) mol/L) stimulated pulmonary endothelial cells to reduce nitroblue tetrazolium monitored spectrophotometrically as 0.022 +/- 0.001, 0.032 +/- 0.002, and 0.071 +/- 0.004 delta A515 per 10(5) cells per hour, respectively (p less than 0.001, comparison of cells with 10(-4) and 10(-3) mol/L nitrofurantoin vs. control cells. From the same dose-response curve, endothelial cells incubated with nitrofurantoin morphologically demonstrated formazan granules in the cytoplasm in 17% +/- 6%, 71% +/- 9%, and 92% +/- 5% of cells. Nitrofurantoin (10(-5), 10(-4), and 10(-3) mol/L) directly injures 51Cr-labeled pulmonary endothelial cells, with injury expressed as a cytotoxic index of 1 +/- 1, 20 +/- 4, and 51 +/- 3, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)