Search PubMed⌕ Search

Biomedical subjects

H Falk

Publications and source records attributed to H Falk.

At least 19 recordsLinked to original sources
PubMed2006

Relationship between retroviral vector membrane and vector stability.

The present work studies the physico-chemical properties of retroviral vector membrane, in order to provide some explanation for the inactivation kinetics of these vectors and to devise new ways of improving transduction efficiency. For this purpose, vectors with an amphotropic envelope produced by TE Fly A7 cells at two culture temperatures (37 and 32 degrees C) were characterized by different techniques. Electron paramagnetic resonance (EPR) results showed that vectors produced at 32 degrees C are more rigid than those produced at 37 degrees C. Further characterization of vector membrane composition allowed us to conclude that the vector inactivation rate increases with elevated cholesterol to phospholipid ratio. Differential scanning calorimetry (DSC) showed that production temperature also affects the conformation of the membrane proteins. Transduction studies using HCT116 cells and tri-dimensional organ cultures of mouse skin showed that vectors produced at 37 degrees C have higher stability and thus higher transduction efficiency in gene therapy relevant cells as compared with vectors produced at 32 degrees C. Overall, vectors produced at 37 degrees C show an increased stability at temperatures below 4 degrees C. Since vector membrane physico-chemical properties are affected in response to changes in culture temperature, such changes, along with alterations in medium composition, can be used prospectively to improve the stability and the transduction efficiency of retroviral vectors for therapeutic purposes.

Animals↗
PubMed2005

Solid tissues can be manipulated ex vivo and used as vehicles for gene therapy.

BACKGROUND: Organ fragments can be cultured for weeks in vitro if they are prepared of microscopic thickness and if the basic organ structure is preserved. Such organ fragments, which we termed micro-organs (MOs), express in culture endogenous tissue-specific gene products. We have exploited this methodology to engineer MOs ex vivo by gene transfer. METHODS: MOs prepared from spleen, lung, colon and skin were infected using: herpes simplex type-1, adeno virus, vaccinia virus and murine leukemia virus (MuLV), carrying the reporter gene beta-galactosidase. RESULTS: All four viral vectors infected MOs in culture, with adeno infection giving significantly higher values. After optimization, high levels of expression (> 15% positive cells), comparable to those obtained with the adeno construct, were also obtained using the MuLV construct both in vitro and after implantation into syngeneic hosts. After implantation, the engineered tissue was found to remain localized, become vascularized, and to express the transduced gene for several months. CONCLUSIONS: The system can be used to study interactions between viruses and tissues both ex vivo and in vivo. Furthermore, the approach proposes a novel platform for ex vivo gene therapy. Such engineered structures could be used as autologous biological pumps for continuous secretion in vivo of gene products of clinical importance.

Animals↗
PubMed2003

Continuous measurement of metals in flue gas using ICP-OES.

A system capable of continuously measuring a range of metallic elements in the effluent gas from incinerators and other similar industrial processes, and providing on-line results has been developed. With a state-of-the-art mobile laboratory measurements were taken from a UK municipal solid waste incinerator. The detection system used was an ICP-OES, with a modified torch to allow the introduction of flue gas directly into the plasma. Metals that were investigated were Ni, Hg, V, Al, Na, Ca, Cu, Sn, Pb, Sb, As, Cd and Tl, with limits of detection in the range 0.0004 mg m(-3) to 0.1 mg m(-3) being calculated. Emission measurements produced data that showed that the MSWI plants emission were significantly lower than the emission limits specified in EC 2000/76/EC.

Journal Article↗
PubMed2001

Gene transfer by viral vectors into blood vessels in a rat model of retinopathy of prematurity.

AIMS: To test the feasibility of gene transfer into hyaloid blood vessels and into preretinal neovascularisation in a rat model of retinopathy of prematurity (ROP), using different viral vectors. METHODS: Newborn rats were exposed to alternating hypoxic and hyperoxic conditions in order to induce ocular neovascularisation (ROP rats). Adenovirus, herpes simplex, vaccinia, and retroviral (MuLV based) vectors, all carrying the beta galactosidase (beta-gal) gene, were injected intravitreally on postnatal day 18 (P18). Two sets of controls were also examined: P18 ROP rats injected with saline and P18 rats that were raised in room air before the viral vectors or saline were injected. Two days after injection, the rats were killed, eyes enucleated, and beta-gal expression was examined by X-gal staining in whole mounts and in histological sections. RESULTS: Intravitreal injection of the adenovirus and vaccinia vectors yielded marked beta-gal expression in hyaloid blood vessels in the rat ROP model. Retinal expression of beta-gal with these vectors was limited almost exclusively to the vicinity of the injection site. Injection of herpes simplex yielded a punctuate pattern of beta-gal expression in the retina but not in blood vessels. No significant beta-gal expression occurred in rat eyes injected with the retroviral vector. CONCLUSIONS: Adenovirus is an efficient vector for gene transfer into blood vessels in an animal model of ROP. This may be a first step towards utilising gene transfer as a tool for modulating ocular neovascularisation for experimental and therapeutic purposes.

Animals↗
PubMed2001

In vitro study of the photocytotoxicity of some hypericin analogs on different cell lines.

In the present study, hypericin analogs with an increased hydrophilic character were synthesized. As chemical modifications alter the lipophilicity/hydrophilicity balance together with the photophysical/chemical background of the molecule the influence of these structural changes on the cellular uptake, retention and subcellular localization in HeLa cells was investigated. Besides, their photocytotoxic effects using three cell lines (HeLa, MCF-7, A431), as well as their plasma protein binding were also assessed. To assess the relative hydrophilic/lipophilic character of hypericin and analogs their retention times were determined on a reversed phase high performance liquid chromatography (C-18) column. The retention time of all the hypericin analogs was < 46 min, except for dibenzyltetramethylhypericin (118 min), while the retention time of hypericin was > 200 min (solvent system: methanol/citrate buffer 30 mM pH 7; 70/30). Hypericin, hexa-, penta- and dibenzyltetramethylhypericin displayed a potent antiproliferative effect at the nanomolar range after photosensitization (3.6 J/cm2). On the contrary, photoactivated tetrasulfonhypericin and fringelite D had no antiproliferative effect on the three cell lines, whereas hypericin polyethylene glycol showed only an intermediate cytotoxic effect on A431 cells. In dark conditions no antiproliferative effect was observed for any photosensitizer. The antiproliferative photo-effect correlated well with the intracellular accumulation as measured using HeLa cells. In general, the photocytotoxic hypericin analogs concentrated to a large extent, while the noncytotoxic compounds were not taken up by the HeLa cells. Furthermore, confocal laser microscopy revealed that all photosensitizers mainly concentrated in the perinuclear region, probably corresponding with Golgi apparatus and the endoplasmic reticulum, except for tetrasulfonhypericin which located at the plasma membrane. In addition, the plasma protein binding studies illustrated that hypericin bind extensively to the low-density lipoproteins, while the other hypericin analogs were mainly bound to heavy proteins (mostly albumin) and to a small extent to low-density lipoproteins.

Anthracenes↗
PubMed2001

Toward hypericin-derived potential photodynamic therapy agents.

To optimize a hypericin derivative as a potential photodynamic therapy agent its light-induced singlet oxygen/superoxide radical formation capability should be enhanced and its long-wavelength absorption band should be bathochromically shifted to better match medicinal lasers. A heavy-atom-substituted derivative was realized by electrophilic iodination of hypericin to yield 2,5-diiodo-hypericin. Using photodestruction of bilirubin IX alpha this derivative was demonstrated to exhibit an enhanced light-induced singlet oxygen/superoxide radical formation capability as compared to hypericin. With respect to a bathochromically shifted derivative styryl residues were attached to the methyl groups of hypericin by de novo ring synthesis. Although the long-wavelength absorption band of this derivative displayed a bathochromic shift of nearly 40 nm it unfortunately immediately underwent an intramolecular [2 + 2] cycloaddition to yield the corresponding cyclobutane derivative in which the added conjugation system became interrupted.

Anthracenes↗
PubMed2000

Characterisation of the high-molecular weight fructan isolated from garlic (Allium sativum L.).

A high molecular weight fructan was isolated from garlic and the structure determined by enzymatic, chemical and spectroscopic (NMR) methods. It was found that the garlic fructan belongs to the neokestose family. It has a (2 --> 1)-linked beta-D-Fruf backbone with (2 --> 6)-linked beta-D-Fruf side chains. A structural model was postulated for a degree of polymerisation of about 58. This model was substantiated using an endo-inulinase purified from Aspergillus ficuum and by 1H and 13C NMR spectroscopy.

Aspergillus↗
PubMed2000

A retrospective radiographic outcome assessment study of intra-bony defects treated by osseous surgery or by bone graft procedures.

BACKGROUND: Intra-bony defects remain a significant therapeutic problem in periodontal therapy. Various non-surgical and surgical treatment modalities are being used. The long-term stability following treatment of intra-bony defects is poorly documented. OBJECTIVES: To assess changes in intra-bony defects after either osseous surgery or open flap debridement in combination with grafting procedures with demineralized freeze-dried bone allografts (DFDBA). METHOD: Pre- and post-surgical computer digitized images of intra-oral radiographs from 60 patients who had received periodontal surgery to manage intrabony defects were analyzed by linear measurements. RESULTS: 36 patients were treated with osseous surgery and 24 had received flap procedures and grafting with DFDBA. Post-surgical radiographs were obtained on average after 4.8 years (SD+/-2.8) and after 9.6 years (SD+/-3.6). A minor mean bone fill of 0.0 mm (SD+/-0.8) for osseous surgery sites and 0.5 mm (SD+/-0.9) for DFDBA sites, was noticed, but this gain was within the margin of measurement errors. Osseous surgery and modified Widman flap procedures with DFDBA resulted in crestal resorption, on average 1.7 mm (SD+/-1.5) and 1.5 mm (SD+/-1.5) and remaining mean defect depth of 2.0 mm (SD+/-1.4) and 2.5 mm (SD+/-1.6), respectively. CONCLUSIONS: Bone changes following bone graft procedures with DFDBA did not differ from those following osseous surgery, and neither procedure resulted in defect resolution with bone fill. It was also concluded that over the study period, stable treatment results were obtained as a result of both osseous surgery and modified Widman flap procedures with adjunct DFDBA.

Analysis of Variance↗
PubMed2000

Healing following GTR treatment of intrabony defects distal to mandibular 2nd molars using resorbable and non-resorbable barriers.

AIMS: The objectives of the present, randomised clinical trial were (i) to evaluate the healing of periodontal intrabony defects at the distal aspect of mandibular 2nd molars using a resorbable polylactic acid (PLA) barrier and a non-resorbable polytetrafluoroethylene (e-PTFE) barrier and (ii) to compare the therapeutic effect of the bioresorbable versus the non-resorbable barrier. METHOD: 19 patients with intrabony defects distal to mandibular 2nd molars > or = 4 mm (on radiographs) were included in the study. The defects all remained 5 years after surgical removal of impacted 3rd molars. Following flap elevation and defect debridement, the defects were randomly covered with, either a resorbable PLA or a non-resorbable e-PTFE barrier. Flaps were repositioned and sutured to completely cover the barriers. Treatment was evaluated clinically after 1 year by measurements of probing depth (PD), probing attachment level (PAL), and probing bone level (PBL) and radiographically by measurements of bone levels on computer digitised images of radiographs taken immediately before and 1 year postsurgery. RESULTS: Both treatments resulted in significant PD reduction, PAL gain, and bone fill. The total PD reduction was 5.3 +/- 1.9 mm for the PLA treated sites and 3.7 +/- 1.7 mm for the e-PTFE treated sites (p<0.05). The corresponding values for PAL gain were 4.7 +/- 0.7 mm and 3.6 +/- 1.7 mm (p<0.05) and for PBL gain 5.1 +/- 1.2 and 3.3 +/- 2.0 mm (p<0.05). Radiographic bone fill averaged 3.4 +/- 1.2 for the PLA and 2.0 +/- 1.6 mm for the e-PTFE barriers (p<0.05). Radiographic bone level measurements were significantly smaller than the corresponding clinical measurements, indicating that radiographs tend to underestimate bone fill. CONCLUSIONS: GTR treatment of deep intrabony defects distal to mandibular second molars using resorbable PLA barriers resulted in significant PD reduction, PAL gain and bone fill at least equivalent to the results obtained using non-resorbable e-PTFE barriers.

Adult↗
PubMed1999

Rapid estimation of avidin and streptavidin by fluorescence quenching or fluorescence polarization.

A new biotin-carboxyfluorescein conjugate has been presented in the accompanying study (G. Kada et al., Biochim. Biophys. Acta 000 (1999) 000-000) which contains ethylene diamine as a 4-atom spacer. This so-called biotin-4-fluorescein showed exceptionally fast and tight binding to avidin and streptavidin, and binding was accompanied by strong quenching. In the present study the specific quenching of 'biotin-4-fluorescein' was utilized to measure (strept)avidin concentrations (0.2-2 nM) by the extent of fluorescence quenching at 8 nM ligand concentration. Adsorption of (strept)avidin to the assay tubes was suppressed by inclusion of bovine serum albumin (0.1 mg/ml). Virtually the same specific response to avidin and streptavidin was also observed with commercial 'fluorescein-biotin', except that >10 h incubation times were required. The slow association of 'fluorescein-biotin' was attributed to the anti-cooperative binding which is due to the much longer spacer as compared to 'biotin-4-fluorescein'. The third ligand tested in this study was 'biotin-4-FITC' which was analogous to 'biotin-4-fluorescein' except that carboxyfluorescein was replaced by the fluorescein isothiocyanate residue. Surprisingly, this probe was much less quenched by avidin but this was compensated by an exceptionally high fluorescence polarization in the avidin-bound state. In conclusion, the new ligand 'biotin-4-fluorescein' appeared to be the most general and convenient probe: quenching was most pronounced and linearly dependent on (strept)avidin concentrations, the dose response for streptavidin was almost the same as for avidin, and the association kinetics were fast enough to reach equilibrium within 30 min incubation time.

Avidin↗
PubMed1999

Accurate measurement of avidin and streptavidin in crude biofluids with a new, optimized biotin-fluorescein conjugate.

A new biotin-fluorescein conjugate with an ethylene diamine spacer was found to be the first fluorescent biotin derivative which truly mimicked d-biotin in terms of high affinity, fast association, and non-cooperative binding to avidin and streptavidin tetramers. These exceptional properties were attributed to the small size/length of the new ligand since all larger/longer biotin derivatives are known for their mutual steric hindrance and anti-cooperative binding in 4:1 complexes with avidin and streptavidin tetramers. Specific binding of the new biotin-fluorescein conjugate towards avidin and streptavidin was accompanied by 84-88% quenching of ligand fluorescence. In the accompanying study this effect was used for rapid estimation of avidin and streptavidin in a new 'single tube assay'. In the present study the strong quenching effect was utilized to accurately monitor stoichiometric titration of biotin-binding sites in samples with >/=200 pM avidin or streptavidin. The concentration was calculated from the consumption of fluorescent ligand up to the distinct breakpoint in the fluorescence titration profile which was marked by the abrupt appearance of strongly fluorescent ligands which were in excess. Due to this protocol the assay was not perturbed by background fluorescence or coloration in the unknown samples. The new fluorescence titration assay is particularly suited for quick checks on short notice because getting started only means to thaw an aliquot of a standardized stock solution of fluorescent ligand. No calibration is required for the individual assay and the ligand stock solution needs to be restandardized once per week (or once per year) when stored at -25 degrees C (or at -70 degrees C, respectively).

Avidin↗
PubMed1999

Treatment of class II furcation defects using resorbable and non-resorbable GTR barriers.

The present study was performed to evaluate the therapeutic effect of a resorbable matrix barrier (test) compared to a non-resorbable barrier (control) on the healing of periodontal tissues in class II furcation defects. Eleven patients, each with two class II furcation defects, underwent guided tissue regeneration surgery. Random selection was used to decide in which of the two sites in each patient test barrier and in which control barrier would be placed. Clinical measurements of the Plaque Index, the Bleeding Index, the Position of the Gingival Margin, as well as Vertical and Horizontal Probing Attachment Levels were made. Whether the barriers were exposed and adverse events were also recorded. It was demonstrated that both procedures led to clinical improvement at the furcation sites. Nine out of eleven defects treated with the test barrier showed improvement while 7 out of 11 achieved the same goal with control barrier. None of the improved sites, though, was completely closed. The conclusion was that within the limits of this study, resorbable test barrier can be used for guided tissue regeneration (GTR) in class II furcation defects with at least as good results as those achieved with non-resorbable control barrier but with the additional benefit of only one operation.

Absorbable Implants↗
PubMed1997

Variant mouse lymphoma cells with modified response to interferon demonstrate enhanced immunogenicity.

We have previously developed an experimental model for the xenogenization of malignant lymphoma. From highly tumorigenic S49 mouse lymphoma cells that proliferate in suspension culture (designated T-25), we selected variant clones that grew as an adherent monolayer (designated T-25-Adh) and were non-tumorigenic in syngeneic mice. Furthermore, priming of syngeneic hosts with T-25-Adh cells protected them against subsequent challenges with the tumorigenic T-25 cells. Several lines of evidence have indicated that antigens of an endogenous mouse mammary tumor virus (MMTV) are involved in the immunogenicity of T-25-Adh cells. Since interferon (IFN) is known to affect retroviral assembly and maturation on the cell membrane, we have studied the effects of IFN on endogenous MMTV-related structures, as well as on the immunogenicity of T-25-Adh cells. We observed that mouse alpha and beta interferons affect the morphogenesis of intracellular MMTV-related precursors in the immunogenic T-25-Adh cells, but not in tumorigenic T-25 cells. From T-25-Adh cells we selected variants that were either high responders or low responders to the above-mentioned interferon effect. The high-response variants were significantly more protective against tumorigenic T-25 cells than the low-response variants. Involvement of MMTV-related antigens in the immune response of the host to T-25-Adh cells was further suggested by immunoelectron-microscopical analysis, demonstrating that antisera from mice, immunized with T-25-Adh cells, interacted specifically with cell-surface MMTV budding particles. These findings indicate a novel method for xenogenization of lymphoma cells by IFN. Since endogenous retroviruses are present in all tissues of the mouse, this approach might be applicable to a wide variety of tumors.

Animals↗
PubMed1997

Guided tissue regeneration therapy of 203 consecutively treated intrabony defects using a bioabsorbable matrix barrier. Clinical and radiographic findings.

THE AIM OF THIS RETROSPECTIVE three-center study was to evaluate guided tissue regeneration (GTR) therapy in a clinical periodontal setting. The material consisted of 203 consecutively treated intrabony defects > or = 4 mm in 143 patients using a bioabsorbable matrix barrier. Each center followed the same protocol for presurgical, intrasurgical, and follow up examinations. Initial therapy, surgical, and follow-up treatments followed the routine of each center. Treatment was evaluated after 1 year by clinical assessments for probing depth (PD) reduction and clinical attachment level (CAL) gain and by bone fill from computer digitized radiographs. Initial intrabony defect depth averaged 6.3 +/- 1.0 mm clinically and 5.7 +/- 1.8 mm radiographically. Mean PD was reduced from 9.0 +/- 1.0 mm to 3.3 +/- 1.0 mm. Mean CAL gain amounted to 4.8 +/- 1.5 mm corresponding to 79 +/- 13% of the initial intrabony defect depth; 78% of the defects exhibited CAL gain > or = 4 mm. Bone fill averaged 3.2 +2- 1.8 mm. Together with a crestal resorption of 1.1 +/- 1.4 mm this resulted in a defect resolution of 4.3 +/- 1.9 mm or 72%. Forty-seven percent (47%) of the variability in CAL gain could be explained by defect depth, defect width, early barrier exposure, and presence of plaque in the treated area. CAL gain and bone fill were positively correlated to the intrabony defect depth; i.e., the deeper the defect the more the CAL gain and bone fill. Sites with barrier exposure during the first 2 weeks of healing showed significantly less CAL gain than sites at which exposure occurred at a later stage or not at all. Presence of plaque in the treated area had a significant negative impact on both CAL gain and bone fill. It was concluded that GTR-treatment of intrabony defects > or = 4 mm in a periodontal specialist practice will result in clinical attachment level gain and bone fill comparable to what has been demonstrated in case studies and controlled clinical trials. The predictability to obtain CAL gain > or = 4 mm in defects > or = 4 mm was 78%.

Alveolar Bone Loss↗