Pullout characteristics of percutaneous pedicle screws with different cement augmentation methods in elderly spines: An in vitro biomechanical study.

BACKGROUND: Vertebroplasty prefilling or fenestrated pedicle screw augmentation can be used to enhance pullout resistance in elderly patients. It is not clear which method offers the most reliable fixation strength if axial pullout and a bending moment is applied. The purpose of this study is to validate a new in vitro model aimed to reproduce a cut out mechanism of lumbar pedicle screws, to compare fixation strength in elderly spines with different cement augmentation techniques and to analyze factors that might influence the failure pattern. MATERIALS AND METHODS: Six human specimens (82-100 years) were instrumented percutaneously at L2, L3 and L4 by non-augmented screws, vertebroplasty augmentation and fenestrated screws. Cement distribution (2 ml PMMA) was analyzed on CT. Vertebral endplates and the rod were oriented at 45° to the horizontal plane. The vertebral body was held by resin in a cylinder, linked to an unconstrained pivot, on which traction (10 N/s) was applied until rupture. Load-displacement curves were compared to simultaneous video recordings. RESULTS: Median pullout forces were 488.5 N (195-500) for non-augmented screws, 643.5 N (270-1050) for vertebroplasty augmentation and 943.5 N (750-1084) for fenestrated screws. Cement augmentation through fenestrated screws led to significantly higher rupture forces compared to non-augmented screws (P=0.0039). The pullout force after vertebroplasty was variable and linked to cement distribution. A cement bolus around the distal screw tip led to pullout forces similar to non-augmented screws. A proximal cement bolus, as it was observed in fenestrated screws, led to higher pullout resistance. This cement distribution led to vertebral body fractures prior to screw pullout. CONCLUSION: The experimental setup tended to reproduce a pullout mechanism observed on radiographs, combining axial pullout and a bending moment. Cement augmentation with fenestrated screws increased pullout resistance significantly, whereas the fixation strength with the vertebroplasty prefilling method was linked to the cement distribution.

Surface roughness and morphology of three nanocomposites after two different polishing treatments by a multitechnique approach.

OBJECTIVES: The purpose of this study was to assess the surface roughness and morphology of three nanocomposites polished with two different polishing systems. METHODS: Specimens made of hybrid composite (Tetric Ceram [TC] as control) and nanocomposites: nanofilled (Filtek Supreme [FS]), nanofilled hybrid (Grandio [Gr]), complex nanofilled hybrid (Synergy D6 [Syn]) were polished with CompoSystem [CS] or Sof-Lex [SL] polishing discs. The average surface roughness (Ra) before and after polishing was measured using optical profilometry. Both AFM and SEM techniques were additionally used to analyze the surface morphology after polishing with the aim of relating the surface morphology and the surface roughness. Statistical analysis was done by ANOVA using a general linear model (alpha=0.05) with an adjustment for multiple comparisons. RESULTS: Within the same polishing system, FS exhibited the smoothest surface, followed by Syn, TC and Gr (p<0.0001). Sof-Lex polishing discs produced the smoothest surface compared to CompoSystem (p<0.0001). AFM and SEM observations confirmed that the surface roughness was related to the surface morphology and to the average filler size. SIGNIFICANCE: Positive correlation between the average filler size and the surface roughness suggest that using nanoparticles in the formulation does not necessary improve the surface texture. The nanofilled composite FS, which contains only nanofillers, showed the best results when associated to Sof-Lex polishing discs.

Calcium phosphate thin film processing by pulsed laser deposition and in situ assisted ultraviolet pulsed laser deposition.

Calcium orthophosphates (CaP) and hydroxyapatite (HA) were intensively studied in order to design and develop a new generation of bioactive and osteoconductive bone prostheses. The main drawback now in the CaP and HA thin films processing persists in their poor mechanical characteristics, namely hardness, tensile and cohesive strength, and adherence to the metallic substrate. We report here a critical comparison between the microstructure and mechanical properties of HA and CaP thin films grown by two methods. The films were grown by KrF* pulsed laser deposition (PLD) or KrF* pulsed laser deposition assisted by in situ ultraviolet radiation emitted by a low pressure Hg lamp (UV-assisted PLD). The PLD films were deposited at room temperature, in vacuum on Ti-5Al-2.5Fe alloy substrate previously coated with a TiN buffer layer. After deposition the films were annealed in ambient air at 500-600 degrees C. The UV-assisted PLD films were grown in (10(-2)-10(-1) Pa) oxygen directly on Ti-5Al-2.5Fe substrates heated at 500-600 degrees C. The films grown by classical PLD are crystalline and stoichiometric. The films grown by UV-assisted PLD were crystalline and exhibit the best mechanical characteristics with values of hardness and Young modulus of 6-7 and 150-170 GPa, respectively, which are unusually high for the calcium phosphate ceramics. To the difference of PLD films, in the case of UV-assisted PLD, the GIXRD spectra show the decomposition of HA in Ca(2)P(2)O(7), Ca(2)P(2)O(9) and CaO. The UV lamp radiation enhanced the gas reactivity and atoms mobility during processing, increasing the tensile strength of the film, while the HA structure was destroyed.

Structural basis for interaction of FGF-1, FGF-2, and FGF-7 with different heparan sulfate motifs.

Stromal cell-derived FGF-7 binds and activates only the resident FGFR2IIIb in epithelial cells while FGF-1 and FGF-2 exhibit a broader interaction with multiple isoforms of FGFR. Here we report the structure of FGF-7 that has been solved to 3.1 A resolution by molecular replacement with the structure of a dual function chimera of FGF-7 and FGF-1 (FGF-7/1) which was resolved to 2.3 A. Comparison of the FGF-7 structure to that of FGF-1 and FGF-2 revealed the strongly conserved Calpha backbone among the three FGF polypeptides and the surface hydrophobic patch that forms the primary receptor-binding domain. In contrast, a decrease and dispersion of the positive surface charge density characterized the heparin-binding domain of FGF-7 defined by homology to that of FGF-1 and FGF-2 in complexes with heparin. A simple heparin hexasaccharide that cocrystallized with FGF-1 and FGF-2 and protected both against protease in solution failed to exhibit the same properties with FGF-7. In contrast to FGF-1 and FGF-2, protection of FGF-7 was enhanced by heparin oligosaccharides of increased length with those exhibiting a 3-O-sulfate being the most effective. Protection of FGF-7 required interaction with specifically the fraction of crude heparin retained on antithrombin affinity columns. Conversely, heparin enriched by affinity for immobilized FGF-7 exhibited anti-factor Xa activity similar to that purified on an antithrombin affinity matrix. In contrast, an FGF-1 affinity matrix enriched the fraction of crude heparin with low anti-factor Xa activity. The results provide a structural basis to suggest that the unique FGF-7 heparin-binding (HB) domain underlies a specific restriction in respect to composition and length of the heparan sulfate motif that may impact specificity of localization, stability, and trafficking of FGF-7 in the microenvironment, and formation and activation of the FGFR2IIIb kinase signaling complex in epithelial cells.

Crystal structures of human DNA polymerase beta complexed with gapped and nicked DNA: evidence for an induced fit mechanism.

DNA polymerase beta (pol beta) fills single nucleotide (nt) gaps in DNA produced by the base excision repair pathway of mammalian cells. Crystal structures have been determined representing intermediates in the 1 nt gap-filling reaction of pol beta: the binary complex with a gapped DNA substrate (2.4 A resolution), the ternary complex including ddCTP (2.2 A), and the binary product complex containing only nicked DNA (2.6 A). Upon binding ddCTP to the binary gap complex, the thumb subdomain rotates into the closed conformation to contact the otherwise solvent-exposed ddCTP-template base pair. Thumb movement triggers further conformational changes which poise catalytic residue Asp192, dNTP, and template for nucleotidyl transfer, effectively assembling the active site. In the product nicked DNA complex, the thumb returns to the open conformation as in the gapped binary DNA complex, facilitating dissociation of the product. These findings suggest that pol beta may enhance fidelity by an induced fit mechanism in which correct base pairing between template and incoming dNTP induces alignment of catalytic groups for catalysis (via thumb closure), but incorrect base pairing will not. The structures also reveal that pol beta binds both gapped and nicked DNA with a 90 degrees kink occurring precisely at the 5'-phosphodiester linkage of the templating residue. If the DNA were not kinked in this way, contact between the thumb and dNTP-template base pair, presumably important for the checking mechanism, would be impossible, especially when the gap is but a single nucleotide. Such a 90 degrees kink may be a mechanistic feature employed by any polymerase involved in filling gaps to completion.

A structural basis for metal ion mutagenicity and nucleotide selectivity in human DNA polymerase beta.

When crystals of human DNA polymerase beta (pol beta) complexed with DNA [Pelletier, H., Sawaya, M. R., Wolfle, W., Wilson, S. H., & Kraut, J. (1996) Biochemistry 35, 12742-12761] are soaked in the presence of dATP and Mn2+, X-ray structural analysis shows that nucleotidyl transfer to the primer 3'-OH takes place directly in the crystals, even though the DNA is blunt-ended at the active site. Under similar crystal-soaking conditions, there is no evidence for a reaction when Mn2+ is replaced by Mg2+, which is thought to be the divalent metal ion utilized by most polymerases in vivo. These results suggest that one way Mn2+ may manifest its mutagenic effect on polymerases is by promoting greater reactivity than Mg2+ at the catalytic site, thereby allowing the nucleotidyl transfer reaction to take place with little or no regard to instructions from a template. Non-template-directed nucleotidyl transfer is also observed when pol beta-DNA cocrystals are soaked in the presence of dATP and Zn2+, but the reaction products differ in that the sugar moiety of the incorporated nucleotide appears distorted or otherwise cleaved, in agreement with reports that Zn2+ may act as a polymerase inhibitor rather than as a mutagen [Sirover, M. A., & Loeb, L. A. (1976) Science 194, 1434-1436]. Although no reaction is observed when crystals are soaked in the presence of dATP and other metal ions such as Ca2+, Co2+, Cr3+, or Ni2+, X-ray structural analyses show that these metal ions coordinate the triphosphate moiety of the nucleotide in a manner that differs from that observed with Mg2+. In addition, all metal ions tested, with the exception of Mg2+, promote a change in the side-chain position of aspartic acid 192, which is one of three highly conserved active-site carboxylate residues. Soaking experiments with nucleotides other than dATP (namely, dCTP, dGTP, dTTP, ATP, ddATP, ddCTP, AZT-TP, and dATP alpha S) reveal a non-base-specific binding site on pol beta for the triphosphate and sugar moieties of a nucleotide, suggesting a possible mechanism for nucleotide selectivity whereby triphosphate-sugar binding precedes a check for correct base pairing with the template.

Crystal structures of human DNA polymerase beta complexed with DNA: implications for catalytic mechanism, processivity, and fidelity.

Mammalian DNA polymerase beta (pol beta) is a small (39 kDa) DNA gap-filling enzyme that comprises an amino-terminal 8-kDa domain and a carboxy-terminal 31-kDa domain. In the work reported here, crystal structures of human pol beta complexed with blunt-ended segments of DNA show that, although the crystals belong to a different space group, the DNA is nevertheless bound in the pol beta binding channel in the same way as the DNA in previously reported structures of rat pol beta complexed with a template-primer and ddCTP [Pelletier, H., Sawaya, M. R., Kumar, A., Wilson, S. H., & Kraut, J. (1994) Science 264, 1891-1903]. The 8-kDa domain is in one of three previously observed positions relative to the 31-kDa domain, suggesting that the 8-kDa domain may assume only a small number of stable conformations. The thumb subdomain is in a more open position in the human pol beta-DNA binary complex than it is in the rat pol beta-DNA-ddCTP ternary complex, and a closing thumb upon nucleotide binding could represent the rate-limiting conformational change that has been observed in pre-steady-state kinetic studies. Intermolecular contacts between the DNA and the 8-kDa domain of a symmetry-related pol beta molecule reveal a plausible binding site on the 8-kDa domain for the downstream oligonucleotide of a gapped-DNA substrate; in addition to a lysine-rich binding pocket that accommodates a 5'-PO4 end group, the 8-kDa domain also contains a newly discovered helix-hairpin-helix (HhH) motif that binds to DNA in the same way as does a structurally and sequentially homologous HhH motif in the 31-kDa domain. DNA binding by both HhH motifs is facilitated by a metal ion. In that HhH motifs have been identified in other DNA repair enzymes and DNA polymerases, the HhH-DNA interactions observed in pol beta may be applicable to a broad range of DNA binding proteins. The sequence similarity between the HhH motif of endonuclease III from Escherichia coli and the HhH motif of the 8-kDa domain of pol beta is particularly striking in that all of the conserved residues are clustered in one short sequence segment, LPGVGXK, where LPGV corresponds to a type II beta-turn (the hairpin turn), and GXK corresponds to a part of the HhH motif that is proposed to be critical for DNA binding and catalysis for both enzymes. These results suggest that endonuclease III and the 8-kDa domain of pol beta may employ a similar mode of DNA binding and may have similar catalytic mechanisms for their respective DNA lyase activities. A model for productive binding of pol beta to a gapped-DNA substrate requires a 90 degrees bend in the single-stranded template, which could enhance nucleotide selectivity during DNA repair or replication.

Characterization of the metal ion binding helix-hairpin-helix motifs in human DNA polymerase beta by X-ray structural analysis.

X-ray crystallographic studies have shown that DNA binding by human polymerase beta (pol beta) occurs primarily through two structurally and sequentially homologous helix-hairpin-helix (HhH) motifs, one in the fingers subdomain and the other in the 8-kDa domain [Pelletier, H., Sawaya, M. R., Wolfle, W., Wilson, S. H., & Kraut, J. (1996a) Biochemistry 35, 12742-12761]. In that DNA binding by each HhH motif is facilitated by a metal ion, we set out to determine the identity of the metal ion that most likely binds to the HhH motif in vivo. Crystal soaking experiments were performed on human pol beta-DNA cocrystals with Mg2+, Ca2+, Na+, and K+, the four most prevalent metal ions in the cell, and in each case a data set was collected and the resulting structure was refined. Under the conditions tested, the HhH motifs of pol beta have an affinity for these biologically prevalent metal ions in the order Mg2+ < Ca2+ < Na+ < K+, with K+ displaying the strongest binding. Crystals soaked in the presence of Tl+, a commonly used spectroscopic probe for K+, were too X-ray-sensitive to establish the binding behavior of Tl+, but soaking experiments with Ba2+ and Cs+ resulted in relatively stable crystals that gave evidence of metal ion binding in both HhH motifs, confirming that larger monovalent and divalent metal ions are capable of binding to the HhH metal sites. Although Mn2+, which has been categorized as a potent polymerase mutagen, binds to the HhH motifs with a greater affinity than Mg2+, Mn2+ does not bind to the HhH motifs in the presence of equimolar concentrations of Na+. These results suggest that in vivo, where Mn2+ is present only in trace amounts, Mn2+ probably does not have a large effect on DNA binding and may instead manifest a mutagenic effect on pol beta primarily by distorting nucleotide binding or by directly affecting the catalytic step [Pelletier, H., Sawaya, M. R., Wolfle, W., Wilson, S. H., & Kraut, J. (1996b) Biochemistry 35, 12762-12777]. Crystal soaking experiments with 31-kDa apoenzyme crystals show that, in the absence of DNA, the HhH motif in the fingers subdomain binds metal ions with either much lower occupancy or not at all, indicating that metal ion binding is dependent on the presence of the DNA substrate.

Tooth thickness at the furcation entrance of lower molars.

Better understanding of the furcation anatomy may serve to decrease the risk of pulpal injury during rotary odontoplasty, a procedure often used in conjunction with guided tissue regeneration. The purpose of this study was to determine (i) the tooth thickness about the furcation entrance of lower molars, and (ii) whether there is a relationship between tooth thickness and patient age. 40 mandibular 1st molars (M1) (mean age = 36.2; range 10-65 years) and 40 mandibular 2nd molars (M2) (mean age = 37.9; range 14-70 years) were collected. Age, gender and furcation involvement (if any) were noted for each tooth at the time of extraction. Teeth were sectioned in half, buccal-lingual, at the furcation entrance with a rotary diamond blade. A standardized linear reference scale was placed on each experimental section and an 8 x 10 in. photograph generated. The distance from the floor of the pulp chamber to 5 predetermined sites on the root surface was calculated. The data were expressed as (a) the mean of each site and (b) the mean of each tooth (the average of the 5 points of each tooth). Analysis of covariance failed to show a relationship between thickness measurements and gender or furcation involvement. Thus, the data was subjected to simple regression analysis to determine the relationship of age with tooth and cementum thickness. This study revealed that by site, the mean measurements ranged from 2.7-3.0 mm for both M1 and M2. The single least/greatest measurements of the 5 sites were for M1: 1.6/4.7 mm and for M2: 1.8/4.2 mm. By tooth, the average distance from the pulp to the root surface was 2.83 mm (+/- 0.49) for M1 and 2.88 mm (+/- 0.44) for M2. Regression analysis of tooth thickness with age was significant for M1 only. The maximum slope of the 5 sites was approximately 0.3 mm/10 years. No relationship was found between cementum thickness and age for either tooth group. The results of this study indicate that the majority of times the pulp is 1.6-4.2 mm from the root surface in the vicinity of the furcation entrance of lower 1st and 2nd molars. Although tooth thickness in this area may increase with age, the amount is not enough to forego judicious odontoplasty on older patients.

Crystallization of fragment D from human fibrinogen.

Fragment D from human fibrinogen has been crystallized. The fragment, which is composed of three disulfide-linked chains (alpha' beta' gamma' = 88,000), was generated with either plasmin or mild trypsin digestion. The crystals diffracted out to 3.5 A; the space group is P2(1), unit cell dimensions a = 108 A, b = 48 A, c = 167 A, beta = 106 degrees. Fragment D was also co-crystallized with the ligand GPRP-amide, in which case the space group is consistent with P212121, unit cell dimensions a = 476 A, b = 82 A, c = 432 A.

Structures of ternary complexes of rat DNA polymerase beta, a DNA template-primer, and ddCTP.

Two ternary complexes of rat DNA polymerase beta (pol beta), a DNA template-primer, and dideoxycytidine triphosphate (ddCTP) have been determined at 2.9 A and 3.6 A resolution, respectively. ddCTP is the triphosphate of dideoxycytidine (ddC), a nucleoside analog that targets the reverse transcriptase of human immunodeficiency virus (HIV) and is at present used to treat AIDS. Although crystals of the two complexes belong to different space groups, the structures are similar, suggesting that the polymerase-DNA-ddCTP interactions are not affected by crystal packing forces. In the pol beta active site, the attacking 3'-OH of the elongating primer, the ddCTP phosphates, and two Mg2+ ions are all clustered around Asp190, Asp192, and Asp256. Two of these residues, Asp190 and Asp256, are present in the amino acid sequences of all polymerases so far studied and are also spatially similar in the four polymerases--the Klenow fragment of Escherichia coli DNA polymerase I, HIV-1 reverse transcriptase, T7 RNA polymerase, and rat DNA pol beta--whose crystal structures are now known. A two-metal ion mechanism is described for the nucleotidyl transfer reaction and may apply to all polymerases. In the ternary complex structures analyzed, pol beta binds to the DNA template-primer in a different manner from that recently proposed for other polymerase-DNA models.

Crystal structure of rat DNA polymerase beta: evidence for a common polymerase mechanism.

Structures of the 31-kilodalton catalytic domain of rat DNA polymerase beta (pol beta) and the whole 39-kilodalton enzyme were determined at 2.3 and 3.6 angstrom resolution, respectively. The 31-kilodalton domain is composed of fingers, palm, and thumb subdomains arranged to form a DNA binding channel reminiscent of the polymerase domains of the Klenow fragment of Escherichia coli DNA polymerase I, HIV-1 reverse transcriptase, and bacteriophage T7 RNA polymerase. The amino-terminal 8-kilodalton domain is attached to the fingers subdomain by a flexible hinge. The two invariant aspartates found in all polymerase sequences and implicated in catalytic activity have the same geometric arrangement within structurally similar but topologically distinct palms, indicating that the polymerases have maintained, or possibly re-evolved, a common nucleotidyl transfer mechanism. The location of Mn2+ and deoxyadenosine triphosphate in pol beta confirms the role of the invariant aspartates in metal ion and deoxynucleoside triphosphate binding.

Cariogenic feeding habits and fluoride supplementation during infancy and early childhood.

Parents of 240 children between nine months and three years of age were interviewed using a questionnaire in order to determine cariogenic feeding habits and fluoride supplementation. Mean age of weaning from the bottle was 14.6 months. After 18 months of age, children from minority ethnic groups were more frequently bottle-fed than French-Canadian children (p < .005). Giving a bottle in bed (34.6% of cases) was more often practised by less educated mothers (p = .007) or by minority ethnic groups (p = .002), and was seen as a cariogenic factor by 31% of parents. Fluoride was given in half of cases, mainly by highly educated mothers (p = .001) and was mentioned as a preventive measure by 27% of parents. Physicians should be aware of poor parental knowledge and practices of preventive dentistry, and must discuss cariogenic feeding habits and fluoride supplementation during well-baby visits.

Crystal structure of a complex between electron transfer partners, cytochrome c peroxidase and cytochrome c.

The crystal structure of a 1:1 complex between yeast cytochrome c peroxidase and yeast iso-1-cytochrome c was determined at 2.3 A resolution. This structure reveals a possible electron transfer pathway unlike any previously proposed for this extensively studied redox pair. The shortest straight line between the two hemes closely follows the peroxidase backbone chain of residues Ala194, Ala193, Gly192, and finally Trp191, the indole ring of which is perpendicular to, and in van der Waals contact with, the peroxidase heme. The crystal structure at 2.8 A of a complex between yeast cytochrome c peroxidase and horse heart cytochrome c was also determined. Although crystals of the two complexes (one with cytochrome c from yeast and the other with cytochrome c from horse) grew under very different conditions and belong to different space groups, the two complex structures are closely similar, suggesting that cytochrome c interacts with its redox partners in a highly specific manner.

Confluence-dependent resistance in human colon cancer cells: role of reduced drug accumulation and low intrinsic chemosensitivity of resting cells.

In vitro sensitivity of HT29 human colon cancer cells to doxorubicin (DXR), vincristine (VCR), etoposide (VP16), cisplatin (CDDP), melphalan (L-PAM) and 5-fluorouracil (5FU) was markedly reduced when cell-culture density increased. For some drugs, confluence-dependent resistance (CDR) was partly due to decreased intracellular drug accumulation; the ratio of mean intracellular drug content of non confluent to confluent cells (NC/C) was 2.5 for DXR, 4.1 for VCR and 7.4 for VP16. Altered drug penetration with confluence could be related to decrease of plasma membrane fluidity as measured by the fluorescence polarization method. Reduction of drug intracellular accumulation was nil or weak for L-PAM (NC/C = 1.0), CDDP (NC/C = 1.2) and 5 FU (NC/C = 1.8). Even if drug concentration was adjusted in culture medium to produce similar intracellular drug content in confluent and non confluent cells, higher intrinsic resistance of confluent cells was still evidenced for DXR and VP16 but not for VCR, the only agent without direct interaction with DNA. DXR- and VP16-induced DNA breakage was also less important in confluent than in non-confluent cells. CDR appeared closely related to an increased proportion of non-cycling cells at confluence, as demonstrated by flow cytometry, expression of nuclear antigen recognized by Ki67 MAb and expression of topoisomerase II. CDR is probably a major factor in the poor sensitivity of colorectal adenocarcinomas to chemotherapy.

Cytotoxic activity of lymphocytes infiltrating progressive and regressive tumor variants from a rat colonic cancer.

Two tumor cell variants (PROb and REGb) isolated from a single chemically-induced rat colon adenocarcinoma were previously shown to differ in their tumorigenicity. When injected into syngeneic BDIX rats, PROb cells induce progressive tumors whereas REGb cells give rise to tumors which always regress. PROb and REGb variants also differ in their capacity to induce an immune response in the syngeneic host. Regression of REGb tumors could have been mediated by cytotoxic effector cells acting at the tumor site. To test this hypothesis, the cytolytic activity of non-adherent lymphoid cells isolated from PROb and REGb tumors and from the spleen of the same animals were compared. The same number of infiltrating lymphocytes was recovered per gram of PROb or REGb tumor. The cytolytic activity of tumor infiltrating lymphocytes, as that of spleen lymphocytes, was predominantly non specific, as demonstrated by their ability to kill YAC-1 cells, an NK-sensitive cell line. PROb cells were relatively resistant to the cytotoxic activity of spleen or tumor infiltrating lymphocytes. In the regressing REGb tumors, the cytotoxic activity of tumor infiltrating lymphocytes to homologous cells or to YAC-1 cells was low and significantly inferior to that of the corresponding spleen lymphocytes. These results suggest that the cytotoxic activity of lymphocytes was impaired at the local, intratumoral level, even in spontaneously regressing tumors.

[Parental knowledge about their asthmatic children]. / Connaissances des parents d'enfants asthmatiques.

A questionnaire was administered to parents of 171 asthmatic children, and their knowledge of asthma was evaluated using a quantitative score. The mean age of children was 5 yr and the mean duration of their asthma was 3 yr. Each sign of the classical triad "noisy breathing, cough, indrawing" was mentioned by two-thirds of parents. Inhaled agents perceived as triggers of asthma were: animal hair (73.7%), dust (69.6%), pollen (60.2%), tobacco (44.4%), molds (14.6%). Other triggering agents mentioned were: stress (51.5%), infections (38%), exercise (13.5%). Mothers with college or university education knew more clinical signs of an attack (P less than 0.01) and more triggering factors (P less than 0.005). Parents satisfied with previous teaching knew more threatening signs of an attack (P less than 0.01). About 80% of those that used theophylline and 49.4% of those that used inhaled beta-2-agonists knew the correct mode of administration. Parents satisfied with previous teaching had better knowledge of the side-effects of theophylline (P less than 0.005) and beta-2-agonists (P less than 0.02). 58.5% of those that used cromolyn sodium did not know the mean duration of a therapeutic trial and 15% did not use it as prophylaxis. 57.1% of those that used oral corticosteroids did not know any side-effects of the drug. About half of the parents indicated that they would like to receive more information about the causes and the appropriate treatment of asthma. It was concluded that parental teaching should focus more on environmental and therapeutic issues.

Mechanisms of resistance of confluent human and rat colon cancer cells to anthracyclines: alteration of drug passive diffusion.

Two colon cancer cell lines, HT-29 (human) and DHD/K12/TRb (rat), were grown as monolayer cultures to various confluence degrees. The cytotoxic efficacies of doxorubicin and 4'-deoxydoxorubicin, evaluated by a survival assay, and the nuclear drug concentrations, measured by microspectrofluorometry, were shown to progressively decrease with the augmentation of confluence. This confluence dependent resistance (CDR) to anthracyclines was demonstrated independent of the multidrug resistance drug efflux mechanism. The cellular uptake of three compounds (sodium [51Cr]chromate, D-[14C]alanine, L-[14C]glucose) known to passively diffuse across the cell membrane as anthracyclines do was also reduced in confluent cells. After trypsin cell detachment, the kinetics of reversion of the sodium [51Cr]chromate uptake decrease and that of CDR were similar. Therefore, CDR may be attributed to a reduction of anthracycline cell intake due to a general alteration of passive diffusion across the cell membrane. However, CDR is only partly explained by this phenomenon since a reduced sensitivity of confluent cells was observed compared with nonconfluent cells for a similar amount of drug in their nuclei. CDR could explain the high resistance to anthracyclines of some solid tumors, such as colon tumors, in which cancer cells are tightly aggregated.

Potential usefulness of quinine to circumvent the anthracycline resistance in clinical practice.

Quinine, the widely used antimalaria agent, was found to increase the cytotoxicity of epideoxorubicin (epiDXR) in resistant DHD/K12 rat colon cancer cells in vitro. Quinine appeared as slightly less effective than quinidine or verapamil for anthracycline potentiation but its weaker cardiotoxicity could counterbalance this disadvantage in vivo. Serum from six patients treated by conventional doses of quinine (25-30 mg kg-1 day-1) was demonstrated to enhance the accumulation of epiDXR in DHD/K12 cells as judged by fluorescence microscopy and HPLC assay (1.6 to 6-fold compared with control serum). In this patients quinine concentrations in serum ranged from 4.4 to 10.1 micrograms ml-1. Our results suggest that quinine could be safely used as anthracycline resistance modifier in clinical practice.