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At least 271 records · Page 15Linked to original sources
PubMed1985

Efficacy of intermittent versus continuous administration of netilmicin in a two-compartment in vitro model.

Several aminoglycoside dosage regimens were studied in a kinetic in vitro model. Pseudomonas aeruginosa, Escherichia coli, Klebsiella pneumoniae, and Staphylococcus aureus were exposed in serially placed artificial capillary units to netilmicin concentrations that changed based on human two-compartment pharmacokinetics. The same total dose per 24 h was administered as a continuous infusion (3.7 micrograms/ml) or in 1-h infusions given every 24 (24 micrograms/ml) or 8 h (8 micrograms/ml). The once daily administration showed the best response in terms of either faster killing of E. coli, K. pneumoniae, and S. aureus or greater reduction of the inocula of P. aeruginosa. After 28 h of treatment, however, all regimens reduced the nonpseudomonads by more than 99.99%, whereas all three P. aeruginosa strains regrew to greater than 10(8) CFU/ml due to selection of resistant subpopulations. In contrast to the bactericidal effect of the first dose, no killing occurred after subsequent doses if the ratio of peak drug concentration to MIC was low (less than or equal to 6). These results support the concept of administering high doses of aminoglycosides once every 24 h.

Bacteria↗
PubMed1993

In vitro models of human colorectal cancer.

Epithelial cell lines that differentiate in vitro have been isolated from hereditary and sporadic colorectal adenomas representing different stages in tumour progression, from small adenomas with a low malignant potential to large adenomas with a relatively high malignant potential. The majority of cell cultures derived from small adenomas senesced, whereas the larger adenomas were more likely to give rise to an immortal cell line. Karyotypic analysis has shown that specific abnormalities of chromosomes 1, 6, 7, 13, 14, 17, 18 and 22 occur in these adenoma cell lines. Abnormalities of chromosome 1 have been implicated in tumour progression and the in vitro immortalization of colorectal adenomas. Molecular and cellular changes involving abnormalities of chromosomes 1 and 18, TP53 and ras gene mutations and reduced response to the growth inhibitory effects of TGFB and sodium butyrate, which occur during tumour progression, suggest that the in vitro model has relevance to in vivo carcinogenesis.

Adenoma↗
PubMed1997

An in vitro model for toxin-mediated vascular leak syndrome: ricin toxin A chain increases the permeability of human endothelial cell monolayers.

Vascular leak syndrome (VLS) is the dose-limiting toxicity observed in clinical trials of immunotoxins containing ricin toxin A chain (RTA). RTA itself is thought to cause VLS by damaging vascular endothelial cells, but the exact mechanism remains unclear. This is partially due to the paucity of appropriate models. To study VLS, we developed an in vitro model in which human umbilical vein-derived endothelial cells were first grown to confluence on microporous supports and then cultured under low pressure in the presence or absence of RTA. Endothelial cell barrier function was assessed by measuring the volume of fluid that passed through each monolayer per unit time. We found that RTA significantly increased monolayer permeability at times and concentrations consistent with the onset of VLS in patients treated with RTA-based immunotoxins. Scanning electron microscopy showed that intercellular gaps formed in endothelial monolayers exposed to RTA. Intercellular gap formation followed endothelial cell death caused by the enzymatic activity of RTA. We conclude that RTA is directly toxic to endothelial cells in vitro and speculate that this contributes to VLS in vivo.

Capillary Permeability↗
PubMed2000

Investigation of an in vitro model for predicting the effect of food components on calcium availability from meals.

The availability of calcium from semi-synthetic meals and model food systems was studied by an in vitro method using equilibrium dialysis after simulating gastric digestion. Twelve different food components (phytate, oxalate, wheat fibre- and barley fibre-extract, D-sorbitol, xylitol, galactitol, casein, three different casein phosphopeptide preparations, and lactose) were added to the semi-synthetic meal and their effect on the relative index of availability of 45Ca was studied. Phytate, oxalate, wheat fibre-extract and barley fibre-extract, and, surprisingly, casein had a negative effect on availability of calcium. The most pronounced effect was observed for the addition of phytate. The other dietary factors had no effect on calcium availability. Therefore, this rapid and inexpensive in vitro model could be a useful screening method for factors affecting calcium absorption, especially inhibitory factors, in future human calcium bioavailability studies.

Biological Availability↗
PubMed2004

Molecular phenotyping of HCS-2/8 cells as an in vitro model of human chondrocytes.

OBJECTIVE: Cultures of primary articular chondrocytes for studying chondrocyte biology are notoriously difficult to handle. One alternative is the use of chondrocytic cell lines. Because the HCS-2/8 cells are the most widely used cell line in cartilage research, we investigated the molecular phenotype of these cells by mRNA-expression profiling. DESIGN: Monolayers of HCS-2/8 cells were cultured to sub-confluence, confluence and over-confluence; primary human chondrocytes were grown in monolayer culture and alginate-bead cultures and several other chondrocytic cell lines were cultured as monolayers. RNA was isolated and analyzed by cDNA array profiling using Affymetrix GeneChips (U95A/U95Av2) and quantitative PCR. RESULTS: Important similarities, but also remarkable differences between the HCS-2/8 cells and adult human articular chondrocytes were detected: Aggrecan and several cartilage typical collagens as well as SOX9 transcripts were strongly expressed in HCS-2/8 cells, whereas HCS-2/8 cells expressed hardly any chondrocyte-typical cartilage matrix degrading enzymes. Of all culturing conditions, clustering analysis showed that HCS-2/8 cultured at confluence are most closely related to primary chondrocytes. CONCLUSION: Our study confirms how careful one needs to be in choosing in vitro model systems for investigating effects of interest. The major issue of chondrocyte cell lines appears to be that they mainly proliferate and show less expression of genes of matrix synthesis and turnover. A successful approach will have to select suitable chondrocyte cell lines and to validate findings obtained using primary chondrocytes. This allows to establish a reproducible in vitro model showing the property of interest and subsequently to relate back the obtained results to the physiologic situation.

Adult↗
PubMed1998

Axonal transport of herpes simplex virus-1 in an in vitro model based on the isolated sciatic nerve of the frog Rana ridibunda.

An in vitro model for the study of the axonal transport of herpes simplex virus-1 (HSV-1) in the nerve fibres of the sciatic nerve of the frog Rana ridibunda, has been developed. The nerve was placed along a three-chambered bath consisting of three isolated chambers arranged in series: the stimulating, perfusion and recording chambers. The HSV-1 inoculum was placed in the stimulating chamber, where the proximal part of the isolated sciatic nerve was immersed. HSV-1 was detected after 24-36 h in the recording chamber, where the distal part of the nerve was immersed in Dulbecco's Modified Eagle Medium (DMEM), indicating an axonal transport speed of 46-60 mm/day. The evoked maximum compound action potentials generated in the stimulating chamber was monitored continuously in the recording chamber as an indication of the viability of the nerve during axonal transport. The in vitro method presented here is a useful tool for the pharmacological study of various parameters, e.g. drugs diluted in the perfusion chamber, ionising radiation and temperature, which may affect the axonal transport or other properties of HSV-1.

Animals↗
PubMed2000

p53, p21(WAF1/CIP1), and MDM2 involvement in proliferation and apoptosis in an in vitro model of conditionally immortalized human vascular smooth muscle cells.

Using an in vitro model of a conditionally immortalized cell line, we investigated how human vascular smooth muscle cells (VSMCs) are affected by the expression of simian virus 40 (SV40) large T antigen (LT antigen), which binds to cell cycle regulators, such as the tumor suppressor protein p53. Cells were obtained after infection of saphenous vein-derived VSMCs with a nonreplicative retroviral vector containing a temperature-sensitive (ts) mutant of SV40 LT antigen and were shown to have maintained some characteristics and responses of VSMCs. Under permissive temperature conditions (36 degrees C), the increased rate of cell proliferation was shown to be associated with expression of LT antigen and with LT-antigen binding to and inactivation of p53. p53 inactivation failed to block apoptosis induced by serum withdrawal or by UV irradiation. Downregulation of LT-antigen expression at the nonpermissive temperature (39 degrees C) was shown to be associated with growth arrest, increased expression of the cell cycle inhibitor p21(WAF1/CIP1), increased MDM2-promoter activity, and differential expression of MDM2 gene products, suggesting that p53-induced transcription/transactivation may be involved in VSMC cell cycle control but not necessarily apoptosis. The established SMC line HVTs-SM1 may be a useful model for the study of processes involved in myointimal hyperplasia and cellular aging, as well as for the study of cell cycle control in general.

Actins↗
PubMed1998

Follicular bisection in hair transplantation surgery: an in vitro model.

The aim of this study was to evaluate, in an in vitro model, the survival and growth rates of transversely sectioned human hair follicles to assess experimentally the soundness of this approach as a future possible method for "duplicating" available donor hair grafts in hair transplantation procedures. A total of 300 human anagen hair follicles was obtained from 10 healthy male patients. Follicles were thus randomly assigned to one of the following groups: group A (control; n = 100 follicles), cultured intact as dissected, and group B (experimental; n = 200 follicles), transversely transected, parallel to the epidermal surface and immediately below the bulge area, to obtain 200 lower-half follicles and 200 upper-half follicles. Isolated hair follicles from both groups were maintained in culture for 10 days. The length of each follicle was measured immediately following isolation and at the end of the 10-day culture period. No statistically significant differences were found between the growth rate of intact follicles (mean 10-day growth rate = 2.71 mm) and of lower-half follicles (mean 10-day growth-rate = 2.64 mm), whereas a statistically significant difference was found between the growth rate of follicles from the two above-mentioned groups and the growth rate of the "upper-half" follicles (mean 10-day growth rate = 1.07 mm). Histologic analysis demonstrated that both intact and lower-half follicles maintained a normal histologic appearance, whereas in upper-half follicle sections we invariably detected a region of intense cell proliferation, reminiscent of a regenerated follicular papilla, surrounding the lowermost part of the follicle. In our opinion, the reported in vitro survival rate of transected human hair follicles might represent an interesting starting point in striving to augment the number of donor hairs available during a hair transplantation procedure.

Adult↗
PubMed2000

Anti-interleukin-1 and anti-CD44 interventions producing significant inhibition of cartilage destruction in an in vitro model of cartilage invasion by rheumatoid arthritis synovial fibroblasts.

OBJECTIVE: To establish an in vitro model for the investigation of destructive processes in rheumatoid arthritis (RA), to study the interaction between fibroblasts, macrophages, and chondrocytes, and to evaluate strategies to inhibit joint destruction in RA. METHODS: Human and bovine chondrocytes cultured in sponges pretreated with native bovine embryonic extracellular matrix produced a cartilaginous matrix reflected by the incorporation of 35S into proteoglycans. The 3-dimensional culture system was optimized for the number of chondrocytes (10(5) cells/sponge), the timing of 35S incorporation (day 21 after chondrocyte isolation), and the medium (20% fetal calf serum). RA synovial fibroblasts (RASF; 10(5)) were added, and the matrix destruction mediated by these RASF was monitored by the release of 35S. The system was modulated by the addition of monocytes (U937 cells), cytokines (interleukin-1beta [IL-1beta] and tumor necrosis factor alpha [TNFalpha]), interleukin-1 receptor antagonist (IL-1Ra), and monoclonal antibodies against IL-1beta and CD44. RESULTS: RASF destroyed the bovine cartilaginous matrix within 2 weeks (days 5-12) and the human cartilaginous matrix within 3 weeks (days 10-18). Compared with the effect of RASF alone (mean +/- SD 948 +/-180 counts per minute/week), the addition of U937 cells (a monocytic cell line), IL-1beta, or TNFalpha to the incubation medium increased the destruction of human cartilaginous matrix by at least 71% up to 90% (ranging from 1,618+/-204 cpm/week to 1,802+/-307 cpm/week). IL-1Ra and anti-IL-1beta monoclonal antibodies reduced the destruction of human matrix by 45% and 35%, respectively; this was partially reversed by the addition of U937 cells. The pretreatment of RASF with anti-CD44 monoclonal antibody (an adhesion molecule and receptor for hyaluronic acid) inhibited the destruction of the cartilaginous matrix by an average of 41% over 3 weeks. CONCLUSION: This model is envisioned to study distinct aspects of human destructive joint diseases under in vitro conditions and to replace and/or supplement animal experiments in basic research and drug testing. Based on the fact that proinflammatory cytokines enhance destruction whereas IL-1Ra and antibodies against IL-1beta and CD44 inhibit the process, it is concluded that anti-IL-1- and anti-CD44-directed therapies may help prevent cartilage destruction in RA.

Animals↗
PubMed1998

The effect of fibrin glue patch in an in vitro model of postdural puncture leakage.

UNLABELLED: We studied the possibility of stopping a continuing transdural leakage with fibrin glue, a biologic adhesive, in an in vitro model. The model was made by sealing the bottom of a tube filled with saline to a height of 50 cm with a human lyophilized dural specimen. Dural punctures were performed with a 17-gauge Tuohy needle. The needle was then withdrawn, and 0.8 mL of fibrin glue was injected through the same needle to seal the defect. The column was refilled 3 min after sealing. The pressure in the intrathecal chamber was measured during the procedure. Macroscopic and microscopic histological studies of the dura and the fibrin plug were performed. In the five cases studied, the leak was sealed by the fibrin plug at closing pressures of 25-35 cm H2O, and no further leakage was detected after refilling. The dural specimens showed a fibrin glue plug stuck at the edges of the hole. We conclude that fibrin glue stops leakage of fluid from dural holes created by a 17-gauge Tuohy needle in an in vitro pressurized model. IMPLICATIONS: We explored the possibility of repairing a cerebrospinal fluid leak produced by an accidental dural puncture during epidural anesthesia by percutaneously injecting tissue adhesive in vitro. This technique seems promising for the prophylaxis and treatment of the headache associated with this leakage but requires further study in vivo.

Dura Mater↗
PubMed1995

In vitro models of the blood-brain barrier to study bacterial meningitis.

In vitro models of the blood-brain barrier involving culturing cerebral microvascular endothelial cells may provide information critical to understanding diseases of the central nervous system, such as bacterial meningitis, that would be difficult to obtain from clinical studies or from in vivo models. These models may also identify targets for therapeutic intervention.

Animals↗
PubMed1996

The effects of copper deficiency on human lymphoid and myeloid cells: an in vitro model.

Cu has long been known to influence immune responses. An in vitro model system was established in which human myeloid (HL-60), B-lymphoid (Raji) and T-lymphoid (Molt-3) cell lines could be grown in culture media of varying Cu levels. Initially Cu was removed from the medium by dialysis of fetal calf serum against a metal-ion chelator, minor depletion of other trace metals being obviated by repletion with appropriate metal salts. The growth rate of HL-60 was significantly (P < 0.05) inhibited by 72 h Cu depletion. Molt-3 cells required a longer period, up to 144 h, in Cu-depleted medium before growth was impaired. Raji-cell growth was not affected. These results confirmed clinical observations that T-cell functions were more sensitive to Cu deprivation than B cells. Analysis of intracellular metal levels in Molt-3 cells showed that Cu levels had been significantly lowered (P < 0.05) although Ca2+ levels were raised. Intracellular activity of the antioxidant enzyme superoxide dismutase (EC 1.15.1.1) was significantly impaired (P < 0.05) in Molt-3 cells grown in Cu-depleted medium. Activity of the mitochondrial enzyme cytochrome c oxidase (EC 1.9.3.1) was also significantly impaired (P < 0.05) by Cu depletion. Each of these findings indicates an increase in the potential for cellular damage by reduced antioxidant activity, impairment of normal mitochondrial activity and excessive Ca2+ influx. A major consequence of the type of damage occurring under these circumstances is membrane disruption. This was confirmed by scanning electron microscopy of Molt-3 cells grown under varying Cu levels.

B-Lymphocytes↗
PubMed2000

p53, p21(WAF1/CIP1), and MDM2 involvement in the proliferation and apoptosis in an in vitro model of conditionally immortalized human vascular smooth muscle cells.

Using an in vitro model of a conditionally immortalized cell line, we have investigated how human vascular smooth muscle cells (VSMCs) are affected by the expression of simian virus 40 (SV40) large T antigen (LT antigen), which binds to cell cycle regulators such as the tumor suppressor protein p53. Cells were obtained after infection of saphenous vein-derived VSMCs with a nonreplicative retroviral vector containing a temperature-sensitive mutant of SV40 LT antigen and were shown to have maintained some characteristics and responses of VSMCs. Under permissive-temperature conditions (36 degrees C), the increased rate of cell proliferation was shown to be associated with expression of LT antigen and with LT antigen binding to and inactivation of p53. p53 inactivation failed to block apoptosis induced by serum withdrawal or UV irradiation. Downregulation of LT antigen expression at a nonpermissive temperature (39 degrees C) was shown to be associated with growth arrest, increased expression of the cell cycle inhibitor p21(WAF1/CIP1), increased murine double minute-2 promoter activity, and differential expression of murine double minute-2 gene products, suggesting that p53-induced transcription/transactivation may be involved in VSMC cycle control but not necessarily in apoptosis. The established SMC line HVTs-SM1 may be a useful model for study of the processes involved in myointimal hyperplasia and cellular aging, as well as for the study of cell cycle control in general.

Antigens, Polyomavirus Transforming↗
PubMed1994

Chemostat flow cell system: an in vitro model for the evaluation of antiplaque agents.

We developed an experimental in vitro model of dental plaque to assess the potential efficacy of antiplaque agents. The model used a chemostat, which provided a continuous source of 5 species of oral bacteria grown in an artificial "saliva-like" medium. This mixture was pumped through six flow cells, each containing two types of surfaces on which plaque formed and was subsequently measured. Formation of bacterial plaque on hydroxyapatite surfaces was assessed by measurement of the DNA and protein content of the plaque film. The amount of bacterial plaque formed on germanium surfaces was measured by attenuated total reflectance (ATR/FT-IR) spectroscopy. Plaque viability was also assessed by a fluorescent staining technique. The quantity of plaque formed on both types of surfaces gradually increased with the duration of flow (from 24 to 72 h) through the cells during a 72-hour experimental period. The flow cells were then pulsed with experimental treatment solutions for 30 s, twice daily. Parallel to results of human clinical studies, the model was capable of discriminating among water, a placebo mouthrinse, and an active antimicrobial mouthrinse formulation containing 0.03% triclosan. It therefore offers a valuable alternative to animal model testing and allows for more rapid evaluations under well-controlled experimental conditions.

Actinomyces viscosus↗
PubMed2006

[A study of human fecal microbiocenosis by experimental in vitro modeling of constipation].

A study of human fecal microbiocenosis using experimental in vitro modeling of constipation was carried out. The study revealed certain dynamics of the incidence, proportion and ratio of microorganisms isolated from the feces at various stages of cultivation. The character of the changes of these parameters depended on the type of the microorganism. According to the results of the experiment, transitory flora (staphylococci; candidas) begins to disappear within the first day of cultivation. If intestinal evacuation is retarded for more than 2 or 3 days, distortion of the microbiological characteristics of the residential microflora, bifidobacteria and escherichiae, is possible. These processes may lead to a false positive result of dysbiosis analysis. The authors conclude that dysbiotic changes in constipation are caused by fecal retention. That is why correct diagnostics of dysbiosis is possible only after regular everyday stool is reestablished.

Adult↗
PubMedDate not supplied

An in vitro model of dermatophyte invasion of the human hair follicle.

A novel in vitro model for the study of hair invasion by Trichophyton mentagrophytes was developed. Hair was obtained by microdissection and plucking. Following inoculation of the hair follicle with arthroconidia growth of the fungus was seen on the hair and within the follicle. Growth was observed to begin at the shaft end and to extend along the hair shaft towards the bulb area. In follicles maintained in organ culture the inner root sheath in particular was invaded by T. mentagrophytes and provided a good substrate for fungal growth. Initially, the cuticle formed a barrier to fungal penetration of the hair. After incubation for 4 days, germlings of T. mentagrophytes were seen to penetrate under the cuticle and in between the layers of cuticular cells to invade the cortex. There was no evidence of intracellular growth; fungal elements were seen intercellularly. There were similarities between the findings in this study of the process of hair invasion by dermatophyte fungi and that in the natural disease.

Adult↗
PubMed1992

Interactions of lung cancer cells with the human mesothelial cell monolayer: an in vitro model for cancer invasion.

An in vitro monolayer model for cancer invasion was developed, comprising cultured layers of human pleura-derived mesothelial cells (PM cells). The cells were isolated from normal human pleura at autopsy and cultured in RPMI-1640 supplemented with 10% fetal calf serum; they were polygonal in shape, forming a pavement-like structure, and preserved the morphologic characteristics of mesothelial cells. When small cell lung cancer cells (OC-10 cells) were seeded on to the monolayer of PM cells, they attached themselves to the monolayer, invaded it and formed flattened cancer cell nests beneath it. By counting the number of cancer cells penetrated, the in vitro invasiveness (invasive capacity) of the cancer cells was assayed. The invasive capacity of poorly invasive cells (10N), which were cloned from OC-10 cells, was five times less than that of the parental OC-10 cells. These results indicate our system to provide a useful model for studying human cancer cell invasion.

Carcinoma, Small Cell↗
PubMed2000

A novel in vitro model of Brunner's gland secretion in the guinea pig duodenum.

A novel in vitro model that combined functional and morphological techniques was employed to directly examine pathways regulating Brunner's gland secretion in isolation from epithelium. In vitro submucosal preparations were dissected from guinea pig duodenum. A videomicroscopy technique was used to measure changes in luminal diameter of glandular acini as an index of activation of secretion. Carbachol elicited concentration-dependent dilations of the lumen (EC(50) = 2 microM) by activating muscarinic receptors on acinar cells. Ultrastructural and histological analyses demonstrated that dilation was accompanied by single and compound exocytosis of mucin-containing granules and the accumulation of mucoid material within the lumen. Inflammatory mediators (histamine, PGE(1), PGE(2)) and intestinal hormones (CCK, gastrin, vasoactive intestinal polypeptide, secretin) also stimulated glandular secretion, whereas activation of submucosal secretomotor neurons by 5-hydroxytryptamine did not. This study directly demonstrates that multiple hormonal, inflammatory, and neurocrine agents activate Brunner's glands, whereas many have dissimilar effects on the epithelium. This suggests that Brunner's glands are regulated by pathways that act both in parallel to and in isolation from those controlling epithelial secretion.

Animals↗