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PubMed1996

Cell, tissue and organ culture as in vitro models to study the biology of squamous cell carcinomas of the head and neck.

In vitro models are currently being used to study head and neck squamous cell carcinoma (HNSCC). Several hundred HNSCC cell lines have been established by various investigators and used to study a broad spectrum of questions related to head and neck cancer. The head and neck model with respect to multistage carcinogenesis is now complete. Several techniques exist for the culture of normal epithelial cells from the upper aerodigestive tract (UADT). The biology of these UADT cells (oral cavity, oropharynx, hypopharynx and larynx) is being studied. Successful culture of premalignant lesions (dysplastic mucosa, leukoplakia, erythroplakia) has resulted in establishment of a limited number of premalignant cell lines and cell cultures. HPV infection of normal oral epithelial cells for immortalization (approximately premalignant cells) coupled with transformation with carcinogens (malignant cells) has established an experimental model for progression. Two in vivo models for oral carcinogenesis, the 7,12 dimethylbenz(a)anthracene-induced hamster cheek pouch model and the 4-nitroquinoline-N-oxide rat oral model, have been established in culture. Thus, multistage carcinogenesis models have been established from both human tissues and animal models and include cultures of normal, premalignant and malignant cells. Culture techniques for growing dissociated primary tumor cells for short term experimental analysis are being used. The culture of normal or tumor tissue as organ/explant cultures allows for the maintenance of normal cell-cell and cell-matrix interaction, but limits experimentation since these cultures cannot be propagated. Several three dimensional model systems are being used to obtain this histological complexity but allow for experimentation. The ability to culture normal, premalignant and malignant cells coupled with the use of a variety of culture techniques, should allow for the continued growth and experimentation in head and neck cancer research.

Animals↗
PubMedDate not supplied

An in vitro model and its application for the study of carotid Doppler spectral broadening.

A new in vitro model has been developed for studying the changes in the ultrasound Doppler spectrum that occur in the region of a stenosis. Pulsatile flow in rigid acrylic tubes was produced by means of a modified hemodialysis pump. The Doppler spectral waveforms were measured using a continuous wave Doppler system, a probe of a known field pattern, a real-time high resolution frequency analyzer, and a video display and recording system. The flow velocity waveforms were found to be nearly identical to those seen in the human carotid. Measurements were made to determine the critical stenosis and the results are similar to those reported from in vivo studies. In a preliminary study, the extent of spectral broadening was found to be dependent on the recording site in relation to the stenosis, the severity of the stenosis, and the flow rate. Using qualitative methods it was not possible to determine either the influence of the shape of the stenosis or the phase of the cardiac cycle on spectral broadening.

Carotid Arteries↗
PubMed1991

Killing kinetics of cefuroxime axetil against Haemophilus influenzae in an in-vitro model simulating serum concentration profiles after oral administration.

We have used a dynamic in-vitro model to determine the killing kinetics of two formulations of cefuroxime axetil against 18 clinical strains of Haemophilus influenzae. The concentration of cefuroxime axetil was adjusted dynamically to simulate the mean serum profile in healthy volunteers following the administration of a single 250 mg dose in oral suspension or tablet form. Bacterial numbers were reduced by 3 log10 cfu/mL during the eight-hour experimental period with both dosage form simulations. Our results suggest that cefuroxime axetil, 250 mg in oral suspension or tablet form, may protect against the dissemination of H. influenzae during episodes of bacteraemia.

Administration, Oral↗
PubMed1996

In vitro models for the effects of sex hormones on neurons.

The results of these two in vitro models share some striking similarities. In both, estrogen was able to induce or promote the formation of either dendrites themselves in hippocampal neurons or dendritic specializations in PC12 neurites, and these specializations were then able to induce interneural interactions. In both models, androgen was able to promote the development of axons that branched frequently, while not directly fostering interneuronal contact. These findings recapitulate in part some of the effects of estrogen and androgen on neurons in vivo and suggest the inherent ability of cells of neural crest origin to respond to these hormones with specific neural morphogenetic programs designed to alter interneuronal communication. In these ways, it seems likely that both sex hormones are acting as neural growth factors in cells that express the appropriate receptor, leading to stereotyped changes in neural growth and pattern formation. Through the examination of such subcellular mechanisms, we hope to further understand the effects of sex hormones on brain development and the ontogeny of behavioral, cognitive, and reproductive differences between the sexes.

Animals↗
PubMed1995

Artificial tumor: a novel heterotypic, polymorphic, three-dimensional in vitro model of individual human solid tumors.

We established a new complex three-dimensional in vitro model called artificial tumor (ArT), which stimulates individual human solid tumors. In contrast to the generally used spheroid models consisting of tumor cells only, the model described in the present report is composed of both tumor cells and tumor stromal cells. We created ArTs as a replica of colon carcinomas, composed of colon carcinoma cells, colon fibroblasts, and as supportive skeleton fibrous tissue from human colon. All cellular components were established from the same surgically obtained colon carcinoma specimen. The ArTs studied came from 4 types of colon carcinomas; mucinous, well, moderately and undifferentiated colon carcinomas. Among all types of colon carcinomas ArTs could be established with a high rate of success, i.e. 81%. Cells growing for 5 days on the fibrous tissue under normal culture conditions formed tumors up to 2,000 microns in diameter. Morphological studies showed that tumor cells grown as ArTs maintained the individual characteristics of the original tumor; for example, a comparable heterogeneity in CEA expression and/or maintenance of the differentiation stage. Polarized tumor cells were linked with the tumor stroma-simulating structure by their basal cell membrane, and formed central lumina lined by the apical membranes of the tumor cells, a structure quite similar to the tissue architecture of colon carcinomas in vivo.

Cell Division↗
PubMed2003

Rheological behavior and parameters of the in vitro model of lung surfactant systems: the role of the main phospholipid component.

The proposed in vitro model for studying the alveolar surface layer of the lungs enables one to investigate the surface intermolecular forces which influence the stability of the alveolus. The general role for the stability of the alveolus belongs to the phospholipids in the alveolar surfactant and predominantly to their main component dipalmitoylphosphatidylcholine (DPPC). The aim of the study was to investigate the rheological behavior of DPPC and exogenous surfactant preparations used in neonatal clinical practice. Data for the rheological behavior of the solutions of the commercially available surfactants, Infasurf, Exosurf and Survanta, as well as of DPPC (their main phospholipid component) at shear rates from 0.024 to 94.5 s(-1) under steady and transient flow conditions at 23 degrees C were obtained. Infasurf and Exosurf showed Newtonian rheological behavior, while Survanta revealed the shear-thinning behavior of a non-Newtonian pseudoplastic fluid. The rheological properties of aqueous solutions of DPPC containing 0.14 M NaCl at concentrations from 100 and 630 microg/ml of phospholipid (chosen from the dependence of the probability for bilayer film formation) were studied. Differences observed in the rheological properties of the exogenous surfactants were interpreted on the basis of their composition, the presence of other phospholipid components, certain additives and surfactant proteins, as well as the bulk structures formed from them. The relevance of the results for the delivery of exogenous surfactants and their spreading in replacement therapy is discussed.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1988

Spinal cord cultured neurons: an in vitro model to study GABA synaptic pharmacology.

In summary, spinal cord cultured neurons provide an in vitro model for studying GABA synaptic pharmacology. GABAA agonists stimulate 36Cl-influx in these cells in a concentration-dependent manner. The enhancing effect of GABA is potentiated by BZ agonists, inhibited by GABA antagonists, and attenuated by the inverse agonist beta-carboline, DMCM. These cells also exhibit the specific binding of [3H]flunitrazepam with a pharmacological specificity of the central BZ receptors. Finally, BZ receptors are coupled to GABA, barbiturate, and picrotoxin sites in these cells, as indicated by allosteric interactions both by binding and 36Cl-influx assay.

Animals↗
PubMed1980

An improved method to study antibacterial activity of antibiotics in an in vitro model simulating serum levels.

The bacterial activity of cefoxitin was studied in an improved in vitro model simulating serum levels. The peak concentration of cefoxitin was 110 mcg/ml as achieved after 1 g i.v. Half-life time was adjusted to 50 minutes. The time at which the viable count reached the mark 1% of the original inoculum after the lowest count was determined (regrowth period) ranged from one to 10 hours from the beginning of the experiments.

Anti-Bacterial Agents↗
PubMed2000

[Toxicity of preserved and unpreserved beta-blocker eyedrops in an in vitro model of human conjunctival cells].

PURPOSE: To compare the toxicity of a short-time application of timolol with benzalkonium chloride (timolol-BAC+) and unpreserved timolol (timolol-BAC-) in an in vitro model of human conjunctival cells. METHODS: Chang's conjunctival cell line (ATCC CCL 20.2) was treated for 15min. with 0.1%, 0.25% or 0.4% timolol-BAC(+) or BAC(-) and then examined immediately or 24h later. Cell viability, chromatin condensation, mitochondrial mass and activity, free radicals production were studied by microplate cold light cytometry. Moreover, relative cell number was evaluated by crystal violet colorimetric test. In addition, cell size and the expression of an apoptotic marker Apo2.7 were studied by flow cytometry. RESULTS: Timolol-BAC(+) induced a rapid decrease in cell viability ranging from 40% immediately after treatment to 85% 24h later. A small, significantly less important decrease in cell viability was also observed with all tested concentrations of timolol-BAC(-). 24h after treatment with 0.25% timolol-BAC(+), the relative cell number was reduced by 55% whereas it did not vary after 0.25% timolol -BAC(-) treatment. Only timolol-BAC(+) induced chromatin condensation, decrease in mitochondrial membrane potential and cell size reduction. Moreover, cells treated with timolol-BAC(+) overexpressed the apoptotic marker Apo2.7. Also reactive oxygen species (ROS) production was significantly more important after cell exposure to timolol-BAC(+). CONCLUSION: In our model of conjunctival cells in vitro, timolol-BAC(+) induced irreversible cytotoxic damage with some characteristics of apoptosis. The active compound of timolol-BAC(-) could be responsible for ROS production and for cell viability variations. Oxidative stress could also play a role in timolol-BAC(+)-induced toxicity. In vitro toxic effects of antiglaucoma drugs could, in part, explain some ocular surface disorders in long-term treated patients.

Adrenergic beta-Antagonists↗
PubMed1998

Amniotic membrane as an in vitro model for endometrium-extracellular matrix interactions.

The aim of the study was to develop an in vitro model to study the interaction between endometrial cells and extracellular matrix (ECM) and to evaluate the expression of cell adhesion molecules in endometrial cells and tissue fragments under in vitro conditions. Endometrial biopsies were collected from 32 patients. Samples were either digested using collagenase type I, or dissected mechanically. Adhesion of isolated cells and tissue fragments to stripped amniotic membranes and to coverslips coated with ECM components was studied. The expression of beta1 integrins and cadherins was assessed using immunohistochemistry. Collagenase digestion of endometrial biopsies yielded viable single cells. These cells did not adhere to either side of stripped amniotic membranes, and did not show expression of the cell adhesion molecules. In contrast, mechanically fragmented endometrium samples adhered to both sides of stripped amniotic membranes and showed immunohistochemical expression of E- and P-cadherin and integrin subunits alpha2, alpha3, alpha4, alpha5 and alpha6. Amniotic membranes, after stripping of epithelial lining, are suitable to study interactions between endometrial tissue and ECM in functional and structural studies. Endometrial cells after collagenase type I digestion do not adhere to stripped amniotic membrane and have lost expression of beta1 integrins and E- and P-cadherin.

Amnion↗
PubMed2006

Effect of heliox on albuterol delivery by metered-dose inhaler in pediatric in vitro models of mechanical ventilation.

STUDY OBJECTIVE: To determine the effect of varying concentrations of heliox, a mixture of helium and oxygen, on albuterol delivery administered by metered-dose inhaler (MDI) in pediatric mechanically ventilated models. DESIGN: Prospective in vitro laboratory study. SETTING: University-affiliated research laboratory. MODELS: The lungs of a 10-kg infant and 30-kg child receiving humidified pressure-regulated volume-controlled ventilation were simulated. The infant settings were an endotracheal tube (ETT) of 4.0 mm, tidal volume of 150 ml, positive end-expiratory pressure of 2 cm H(2)O, rate of 20 breaths/minute, inspiratory time of 0.7 second; the child settings were an ETT of 6.0 mm, tidal volume of 450 ml, positive end-expiratory pressure of 2 cm H(2)O, rate of 16 breaths/minute, and inspiratory time of 0.8 second. MEASUREMENTS AND MAIN RESULTS: Ten albuterol MDI canisters with chlorofluorocarbon propellants were each actuated once sequentially (total dose 1000 mug) with a commercially available aerosol holding chamber. Albuterol was collected onto a filter proximal to a lung simulator. The filter was rinsed, and concentrations were determined by high-performance liquid chromatography. In the infant model, heliox mixtures of 70:30, 60:40, and 50:50 were compared with nitrogen:oxygen (N(2):O(2)) mixtures in the same ratios. The effect of the 70:30 mixtures was also explored in a child model. Each gas mixture was tested 5 times. At all three ratios, albuterol delivery to the end of the ETT was improved with heliox compared with N(2):O(2) (approximately 7% vs 3-4%, p<0.0001, one-way analysis of variance [ANOVA] with a Bonferroni correction for multiple comparisons). No significant difference was noted in mean percentage albuterol delivery among the varying ratios of heliox studied. By two-way ANOVA, significantly greater albuterol delivery was noted with 70:30 heliox compared with 70:30 N(2):O(2) (7-8% vs 3%, p<0.0001), with no significant difference between the infant and child model (p=0.21). The gas mixture, model, and interaction of the two explained 88% of the variability in mean percentage albuterol delivery. CONCLUSION: Heliox increased albuterol delivery administered by MDI to the end of the ETT in these in vitro pediatric models of mechanical ventilation. Further studies are needed to determine if the improved albuterol delivery with heliox enhances clinical response in infants and children needing mechanical ventilation.

Aerosols↗
PubMed1999

An improved low-permeability in vitro-model of the blood-brain barrier: transport studies on retinoids, sucrose, haloperidol, caffeine and mannitol.

Primary cultures of porcine brain capillary endothelial cells grown on collagen coated polycarbonate membranes were used to build up an in vitro-model for the blood-brain barrier. Improved cultivation techniques allowed cell-storage and experiments under serum-free conditions. We employed this model to perform permeability studies in vitro with the radioactively labelled marker substances sucrose, retinoic acid, retinol, haloperidol, caffeine, and mannitol. Permeability values obtained with this blood-brain barrier model (1. 0x10-6 cm/s for sucrose, 6.2x10-6 cm/s for retinoic acid, 4.8x10-6 cm/s for retinol, 49.5x10-6 cm/s for haloperidol, 62.4x10-6 cm/s for caffeine, and 1.8x10-6 cm/s for mannitol) show a good correlation to data which are already known from in vivo-experiments. As judged by the sucrose permeability our blood-brain barrier model is less permeable than numerous other models published so far. Therefore it represents a powerful tool for in vitro-prediction of blood-brain barrier permeability of drugs and offers the possibility to scan a large quantity of drugs for their potential to enter the brain.

Animals↗
PubMed1997

Flow cytometric analysis of coronary stent-induced alterations of platelet antigens in an in vitro model.

One of the limitations of coronary stenting is the subacute thrombotic occlusion. In an in vitro model, we examined the effects of tantalum wire stents (n = 12) on platelet antigens. Platelet-rich plasma (PRP) was circulated in PVC tubing systems. At fixed intervals over a 10-min time course, aliquots of PRP were drawn, stained with monoclonal antibodies (CD41a, CD42b, CD62p, and CD63), and analyzed by flow cytometry. Within 2 minutes of the onset of circulation, expression of the activation-dependent antigens CD62p and CD63 increased in all tubing systems with stents. This early increase was followed by a progressive rise in fluorescence intensity of these neoantigens over the course of 10 minutes (p < 0.05 vs.. control system without stent). Antigens CD41a and CD42b did not show significant changes in either system. The artificial surfaces and shear forces of stent meshes induce alterations in platelet antigens. Flow cytometry provides a sensitive technique for in vitro testing of the thrombogenicity of coronary stents, and may be useful in further improving stent biocompatibility.

Adult↗
PubMed2003

Malignant hematopoietic cell lines: in vitro models for the study of mast cell leukemia.

Mast cells are multifunctional hematopoietic cells producing various proinflammatory mediators. They arise in the bone marrow from CD34+ myeloid progenitors under the influence of stem cell factor but reside extravascular in the tissues. Mastocytosis which is a rare disorder encompasses a heterogeneous group of entities characterized by abnormal proliferation and accumulation of mast cells, including mast cell leukemia. Progress in understanding the (patho)biology of mast cells has been hindered by the lack of genuine model systems, such as continuous cell lines. Now the two in vitro models HMC-1 and LAD 1/2 are available. Cell line HMC-1 was published in 1988; the sister cell lines LAD 1 and LAD 2 have been established in 2000. These cell lines were all derived from patients with mast cell leukemia-sarcoma. The cell lines have been properly authenticated; their immunophenotypic, cytogenetic, molecular, and functional features have been described in detail. Taken together, these cell lines reproduce faithfully most or all of the characteristics of primary normal or malignant cells in so far as these are known. Interest in this rare but nevertheless remarkable cell type should gain momentum with the availability of model systems.

Cell Culture Techniques↗
PubMed1975

Response of ampicillin resistant Escherichia coli to cephalosporins in an in vitro model simulating conditions of bacterial growth in the urinary bladder.

Five ampicillin resistant strains of Escherichia coli were exposed to cephalosporins in an in vitro model which simulates the hydrokinetic features of the urinary bladder. Although the strains showed substantial zones of inhibition when tested against cephalosporins by the disc diffusion method, the results in the bladder model suggest that, in conditions where the antibiotic concentration is being reduced by dilution and micturition as well as enzymic hydrolysis by the organism, activity of this group of agents may be insufficient to eradicate infection. It is suggested that the results warrant a closer investigation into the efficacy of cephalosporins against ampicillin resistant Gram negative bacilli in vivo.

Ampicillin↗
PubMed1980

An in vitro model for monitoring bacterial responses to antibiotic agents under simulated in vivo conditions.

A new in vitro model for the study of bacterial responses to antibacterial agents at exponentially varying concentrations, i. e. at given elimination half-life values simulating in vivo conditions, is described. The initial concentrations and elimination rates of the agents, composition of bacterial milieu, and commencing density of the bacterial population may be altered. The principal responses of key gram-negative and gram-positive species to bactericidal and bacteriostatic agents and experimental variation are described.

Anti-Bacterial Agents↗
PubMed1997

In-line pressure-flow module for in vitro modelling of haemodynamics and biosensor validation.

An in-line pressure-flow module for in vitro modelling of haemodynamics and biosensor validation has been developed. Studies show that good accuracy can be achieved in the measurement of pressure and of flow, in steady and pulstile flow systems. The model can be used for development, testing and evaluation of cardiovascular-mechanical-electrical anlogue models, cardiovascular prosthetics (i.e. valves, vascular grafts) and pressure and flow biosensors.

Animals↗
PubMed1985

Organ culture of young rat vas deferens as an in vitro model for the study of denervation supersensitivity.

To investigate whether organ culture is a suitable in vitro model for studying the mechanisms of denervation-induced supersensitivity, we cultured 1-week-old rat vas deferens for 3 days with a basic applied tension of 20 mg. Cultured muscles showed supersensitivity to norepinephrine and methacholine with concomitant elevation of the maximal response. To compare these changes with those caused by denervation, young rats were chemically denervated by injecting 6-hydroxydopamine, and consequent sensitivity changes were investigated. Denervated muscles showed non-specific supersensitivity to norepinephrine and methacholine but the maximal response did not increase. When these denervated muscles were organ-cultured, they showed no or only a slight increase in sensitivity to norepinephrine and methacholine, but the maximal response increased greatly. These observations led to the suggestion that the increase in sensitivity may be mediated through the same mechanisms as those for denervation supersensitivity. The elevation of the maximal response was suggested to be produced by the improvement of cell-to-cell conduction as well as some other unknown factor(s) probably specific to organ culture. Thus, it was concluded that organ-cultured 1-week-old rat vas deferens is a useful model to study the mechanisms of denervation supersensitivity.

Animals↗