Chitosan-Surfactant Composite Nanocoatings on Glass and Zinc Surfaces Prepared from Aqueous Solutions.

Hydrophobic coatings from chitosan-surfactant composites (ca. 400 nm thick by UV-Vis spectroscopy) for possible corrosion protection were developed on glass and zinc substrates. The surfactants (sodium dodecyl sulfate, SDS or sodium dodecylbenzenesulfonate, and SDBS) were added to the chitosan by two methods: mixing the surfactants with the aqueous chitosan solutions before film deposition or impregnating the deposited chitosan films with surfactants from their aqueous solutions. For the mixed coatings, it was found that the lower surface tension of solutions (40-45 mN/m) corresponded to more hydrophobic (80-90°) coatings in every case. The hydrophobicity of the impregnated coatings was especially significant (88° for SDS and 100° for SDBS). Atomic force microscopy studies revealed a slight increase in roughness (max 1.005) for the most hydrophobic coatings. The accumulation of surfactants in the layer was only significant (0.8-1.0 sulfur atomic %) in the impregnated samples according to X-ray photoelectron spectroscopy. Polarization and electron impedance spectroscopy tests confirmed better barrier properties for these samples (40-50% pseudo-porosity instead of 94%). The degree of swelling in a water vapor atmosphere was significantly lower in the case of the impregnated coatings (ca. 25%) than that of the native ones (ca. 75%), measured by spectroscopic ellipsometry. Accordingly, good barrier layer properties require advantageous bulk properties in addition to surface hydrophobicity.

Laboratory characterization of leukemic cell procoagulants.

BACKGROUND: In acute myeloid leukemias, there is an increased chance to develop thrombotic disorders. We hypothesized that in addition to leukemic promyelocytes, monocytic leukemia cells may also have a higher procoagulant activity. METHODS: Fibrin formation was assessed by a one-stage clotting assay using a magnetic coagulometer. The thrombin generation test (TGT) of magnetically isolated normal human monocytes, intact leukemic cells and their isolated microparticles was performed by a fluorimetric assay. Phosphatidylserine (PS) expression of leukemic cells and microparticle number determinations were carried out by flow cytometry. RESULTS: All cell lines displayed a significant procoagulant potential compared to isolated normal human monocytes. In the TGT test, the mean of lagtime and the time to peak parameters were significantly shorter in leukemic cells (3.9-4.7 and 9.9-10.3 min) compared to monocytes (14.9 and 26.5 min). The mean of peak thrombin in various monocytic leukemia cell lines was 112.1-132.9 nM vs. 75.1 nM in monocytes; however, no significant difference was observed in the ETP parameter. Factor VII-deficient plasma abolished all procoagulant activity, whereas factor XII-deficient plasma did not affect the speed of fibrin formation and thrombin generation but modulated the amount of thrombin. Factor XI-deficient plasma affected the time to peak values in one leukemic cell line and also attenuated peak thrombin. Leukemia cell-derived microparticles from all three cell lines exerted a procoagulant effect by significantly shortening the lagtime in TGT; there was a nonsignificant difference in case of ETP parameter. CONCLUSIONS: All investigated monocytic leukemia cell lines exhibited significant thrombin generation. This phenomenon was achieved by the procoagulants on the surface of leukemic cells as well as by their microparticles.

[Local infiltration analgesia in total joint replacement]. / Helyi infiltrációs érzéstelenítés alkalmazása nagyízületi endoprotézisek beültetése során.

INTRODUCTION: Total hip and knee replacment surgeries are characterized by severe postoperative pain. Local infiltration analgesia is proved to be very effective. However this method has not been widely used in Hungary. AIM: To evaluate the efficacy of the local infiltration analgesia with modified components in patients underwent total hip or knee replacement surgery. METHOD: Data of 99 consecutive patients underwent primary total hip or knee replacement surgery were evaluated prospectively. In all the 99 surgeries modified local infiltration analgesia was applied. Postoperative pain reported on a visual analog scale was recorded as well as the need for further analgetics during the first 18 hours after surgery. The cost of the analgetic drugs was calculated. The control group comprised 97 consecutive patients underwent total hip or knee replacement, where local infiltration analgesia was not applied. Statistical analysis was done. RESULTS: Patients received local infiltration analgesia reported significantly less pain (p<0.001). The need for postoperatively given analgetics was almost 50% less, and the cost of all postoperative analgetics was 47% less than in the control group. CONCLUSION: In total hip and knee replacement surgeries the modified local infiltration analgesia decreases postoperative pain effectively and contribute to the early mobilization of the patients. Orv. Hetil., 2017, 158(9), 352-357.

7DHC-induced changes of Kv1.3 operation contributes to modified T cell function in Smith-Lemli-Opitz syndrome.

In vitro manipulation of membrane sterol level affects the regulation of ion channels and consequently certain cellular functions; however, a comprehensive study that confirms the pathophysiological significance of these results is missing. The malfunction of 7-dehydrocholesterol (7DHC) reductase in Smith-Lemli-Opitz syndrome (SLOS) leads to the elevation of the 7-dehydrocholesterol level in the plasma membrane. T lymphocytes were isolated from SLOS patients to assess the effect of the in vivo altered membrane sterol composition on the operation of the voltage-gated Kv1.3 channel and the ion channel-dependent mitogenic responses. We found that the kinetic and equilibrium parameters of Kv1.3 activation changed in SLOS cells. Identical changes in Kv1.3 operation were observed when control/healthy T cells were loaded with 7DHC. Removal of the putative sterol binding sites on Kv1.3 resulted in a phenotype that was not influenced by the elevation in membrane sterol level. Functional assays exhibited impaired activation and proliferation rate of T cells probably partially due to the modified Kv1.3 operation. We concluded that the altered membrane sterol composition hindered the operation of Kv1.3 as well as the ion channel-controlled T cell functions.

Altered lipid subfraction profile and impaired antioxidant defense of high-density lipoprotein in Smith-Lemli-Opitz syndrome.

BACKGROUND: Smith-Lemli-Opitz syndrome (SLOS) is a rare disease caused by biallelic mutation in the 7-dehydrocholesterol (7DHC) reductase gene. High oxidizability of 7DHC and the appearance of small-sized low-density lipoprotein (LDL) subfractions indicate increased endogenous oxidative stress that is counterbalanced by natural antioxidant defense mechanisms including the high-density lipoprotein (HDL)-associated paraoxonase-1 (PON1) enzyme. PON1 prevents lipoproteins from oxidative modifications; however, PON1 activity and the distribution of lipoprotein subfractions have not been studied in SLOS. METHODS: 7DHC levels and PON1 arylesterase activities were measured spectrophotometrically in 11 SLOS patients and 10 healthy children. Lipoprotein subfractions were detected by polyacrylamide gel electrophoresis. RESULTS: Compared to controls, there was a shift towards the small-dense LDL subfraction and the large HDL subfraction in SLOS. PON1 arylesterase activity was significantly decreased in SLOS patients and correlated negatively with the proportion of small-dense LDL subfraction and the proportion of large HDL subfraction. Significant positive correlations were detected between PON1 arylesterase activity and the ratios of intermediate and small HDL subfractions. CONCLUSIONS: Decreased PON1 activity and the deleterious shift in the distribution of lipoprotein subfractions may contribute to the impaired antioxidant status observed in SLOS. Monitoring of serum PON1 arylesterase activity may be a complementary biomarker in SLOS.

[Inborn error of cholesterol biosynthesis: Smith-Lemli-Opitz syndrome]. / A koleszterin-bioszintézis veleszületett zavara: a Smith-Lemli-Opitz-szindróma.

Smith-Lemli-Opitz syndrome is an autosomal recessive mental retardation and multiple malformation syndrome caused by deficiency of the 7-dehydrocholesterol reductase, the enzyme catalyzing the last step in cholesterol biosynthesis. The authors summarize the pathophysiology, epidemiology, clinical picture, diagnostics and therapy of the disease based on a review of the international literature. Since 2004, fourteen patients have been diagnosed with Smith-Lemli-Opitz syndrome in Hungary, which suggests an underdiagnosis of the disease based upon estimated incidence data. Due to deficiency of the 7-dehydrocholesterol reductase, serum cholesterol concentration is low and 7-dehydrocholesterol concentration is elevated in blood and tissues; the latter being highly specific for the syndrome. Detection of disease-causing mutations makes the prenatal diagnosis possible. The clinical spectrum is wide, the most common symptom is syndactyly of the second and third toes. Standard therapy is cholesterol supplementation. Recent publications suggest that oxidative compounds of 7-dehydrocholesterol may play a role in the pathophysiology of the disease as well.

[Long-term follow-up after tympanostomy tube insertion in children with serous otitis media]. / Gyermekkori otitis media serosa miatt tubussal ellátott betegek hosszú távú nyomon követése.

INTRODUCTION: The authors report about the efficacy of inserted tympanostomy tube in children with serous otitis media. AIM: The aim of the authors was to assess the status of eardrum, the function of Eustachian tube and hearing level 10 years after the use of tympanostomy tube. METHOD: Patients filled out a questionnaire and microscopic examination of tympanic membrane, tympanometry, Eustachian tube function examination, and audiometry tests were performed. RESULTS: In the period of 2003-2004, ventilation tube insertion was performed in 711 patients in the ENT Department of Pediatric Health Center of University of Szeged. In 349 patients adenotomy and tympanostomy tube insertion, in 18 cases tonsillectomy and grommet insertion and in 344 patients only typmanostomy tube insertion were performed. Due to objective difficulties (address change, no phone number) 453 patients were asked for control test and 312 persons accepted the invitation. Normal hearing level was found in 84.6% of patients and normal tympanometry result occurred in 82%. Tympanic ventilation disorder, perforation of tympanic membrane, sensorineural hearing loss and sensorineural hearing loss due to noise exposure were diagnosed. CONCLUSIONS: Application of tympanostomy tube is effective in the treatment of serous otitis media resulting from ventilation disorder. The authors draw attention to the importance of tympanometry examination to prevent the adhesive processes and cholesteatoma in chronic ventilation disorder of the middle ear.

[Results of clinical and genetic diagnosis of rare diseases in the Eastern region of Hungary (2007-2013)]. / Ritka genetikai betegségek klinikai és genetikai diagnosztikájában szerzett tapasztalataink a kelet-magyarországi régióban (2007-2013).

INTRODUCTION: 80% of rare diseases have a genetic origin, and 50% manifest themselves as congenital anomalies. Their adequate health care includes early recognition of genetic anomalies and prevention of recurrence. AIM: The aims of the authors were to provide correct diagnoses to patients with multiple congenital anomalies with or without mental retardation attending to the outpatient clinic of the Clinical Genetics Center at the University of Debrecen in the time interval between August 1, 2007 and March 31, 2013, establish the possibility of prenatal diagnosis, assess the distribution of different genetic mechanisms in the background of rare genetic diseases, compare them with international data, and develop an algorithm for the diagnostic approach of rare genetic diseases applicable in Hungary. METHOD: Clinical data and genetic results of patients were evaluated, and patients were categorized into one of the ten proposed etiological groups, based on which the distribution of genetic causes was defined. RESULTS: Clinical diagnosis was achieved in 64.3% of patients, confirmed genetic diagnosis in 37.8%, while 35.7% of patients remained undiagnosed. Several dysmorphic syndromes and metabolic disorders were first diagnosed in Hungary, two of which unique in the literature. CONCLUSIONS: In the centre of the authors the diagnostic effectiveness of chromosome aberrations exceeds the international standards, that of known microdeletions and dysmorphic syndromes meets international data, and the genetic diagnosis of mendelian disorders and submicroscopic copy number changes remain below international figures.

Correlations between the anti-corrosion properties and the photocatalytic behavior of epoxy coatings incorporating modified graphene oxide deposited on a zinc substrate.

This research aimed to create a substrate-coating system based on zinc and an epoxy resin incorporating modified graphene oxide, which possesses two key characteristics: effective resistance against corrosion and the ability to harness photocatalytic properties. Furthermore, correlations between the anti-corrosion properties and the photocatalytic behaviour of the coatings were made. Thin epoxy (EP) layers embedding 0.1 wt% graphene oxide (GO), reduced graphene oxide (rGO), and modified graphene oxide with (3-aminopropyl)-triethoxysilane (APTES) or poly(amidoamine) (PAMAM) dendrimer were applied on a zinc (Zn) substrate using the dip-coating method. Anti-corrosion properties of coated Zn samples were investigated through electrochemical impedance spectroscopy (EIS) measurements. They showed that the corrosion protection effect is more prominent for EP containing functionalized GO, the highest in the case of GO-PAMAM. The results of the EIS measurements indicated also that the corrosion protection provided by EP-rGO is smaller than that of EP. The photocatalytic properties of the coatings were studied by exposure of the samples to Methylene Blue (MB) solution followed by monitoring the model dye degradation through UV-Vis measurements. To determine the changes in the anti-corrosion properties due to photocatalysis, the coated Zn samples were put through additional EIS measurements. The same coatings applied to a glass substrate lacked photocatalytic properties, indicating that the Zn substrate is accountable for the degradation of MB. Furthermore, the incorporation of GO or functionalized GO into the coating amplifies this effect. From EIS spectra, it was determined that the protective properties loss observed after 3 days is due to coating delamination during exposure to MB solution, the EP-GO-APTES retaining the best adhesion of the coating, 98% remaining on Zn after a cross-hatch test. The corrosion measurements were complemented by examining the morphology and structure of the coatings and the modified GO particles. All things considered, the Zn/EP-GO-APTES system shows the best ability to break down organic pollutants, keeping a good anti-corrosive property and adhesion.

Wetting and swelling behaviour of N-acetylated thin chitosan coatings in aqueous media.

Chitosan nanocoatings (thickness range of 120-540 nm) were produced on glass, zinc and silicon substrates with dip-coating and spin coating techniques to study their pH-dependent wetting and swelling behaviour. The coatings were N-acetylated with the methanolic solution of acetic anhydride to increase the degree of acetylation from 36 % to 100 % (according to ATR-FTIR studies). The measured contact angles of Britton-Robinson (BR) buffer solutions (pH 6.0, 7.4 and 9.0) were lower on the acetylated surfaces (ca. 50°), than that of their native counterparts (ca. 70°) and does not depend on the pH. Contrary, contact angles on the native coating deteriorated 10°-15° with increasing the pH. In addition, for native coatings, the decrease of the contact angles over time also showed a pH dependence: at pH 9.0 the contact angle decreased by 7° in 10 min, while at pH 6.0 it decreased by 13° and at a much faster rate. The constraint swelling of the coatings in BR puffer solutions was studied in situ by scanning angle reflectometry. The swelling degree of the native coatings increased significantly with decreasing pH (from 250 % to 500 %) due to the increased number of protonated amino groups, while the swelling degree of acetylated coatings was ca. 160 % regardless of the pH. The barrier properties of the coatings were studied by electrochemical tests on zinc substrates. The analysis of polarization curves showed the more permeable character of the acetylated coatings despite the non-polar character of the bulk coating matrix. It can be concluded that in the case of native coatings, 49 % of the absorbed water is in bound form, which does not assist ion transport, while in the case of acetylated coatings, this value is only 33 %.

Lactoferricin mediates anti-inflammatory and anti-catabolic effects via inhibition of IL-1 and LPS activity in the intervertebral disc.

The catabolic cytokine interleukin-1 (IL-1) and endotoxin lipopolysaccharide (LPS) are well-known inflammatory mediators involved in degenerative disc disease, and inhibitors of IL-1 and LPS may potentially be used to slow or prevent disc degeneration in vivo. Here, we elucidate the striking anti-catabolic and anti-inflammatory effects of bovine lactoferricin (LfcinB) in the intervertebral disc (IVD) via antagonism of both IL-1 and LPS-mediated catabolic activity using in vitro and ex vivo analyses. Specifically, we demonstrate the biological counteraction of LfcinB against IL-1 and LPS-mediated proteoglycan (PG) depletion, matrix-degrading enzyme production, and enzyme activity in long-term (alginate beads) and short-term (monolayer) culture models using bovine and human nucleus pulposus (NP) cells. LfcinB significantly attenuates the IL-1 and LPS-mediated suppression of PG production and synthesis, and thus restores PG accumulation and pericellular matrix formation. Simultaneously, LfcinB antagonizes catabolic factor mediated induction of multiple cartilage-degrading enzymes, including MMP-1, MMP-3, MMP-13, ADAMTS-4, and ADAMTS-5, in bovine NP cells at both mRNA and protein levels. LfcinB also suppresses the catabolic factor-induced stimulation of oxidative and inflammatory factors such as iNOS, IL-6, and toll-like receptor-2 (TLR-2) and TLR-4. Finally, the ability of LfcinB to antagonize IL-1 and LPS-mediated suppression of PG is upheld in an en bloc intradiscal microinjection model followed by ex vivo organ culture using both mouse and rabbit IVD tissue, suggesting a potential therapeutic benefit of LfcinB on degenerative disc disease in the future.

Relation between biomarkers and clinical severity in patients with Smith-Lemli-Opitz syndrome.

UNLABELLED: Smith-Lemli-Opitz syndrome (SLOS), a multiple congenital anomaly with severe mental retardation, is caused by decreased activity of 7-dehydrocholesterol reductase. Fifteen Hungarian patients were diagnosed with SLOS on the basis of clinical symptoms, serum cholesterol, 7-dehydrocholesterol, and molecular genetic testing. Their age at the time of diagnosis in mild SLOS (n = 4, clinical score <20) was 0.5-18 years, cholesterol was 2.37 ± 0.8 mmol/L, and 7DHC was 0.38 ± 0.14 mmol/L. In the group of typical SLOS (n = 7, score 20-50), the diagnosis was set up earlier (age of 0.1-7 years); t-cholesterol was 1.47 ± 0.7 mmol/L, and 7DHC was 0.53 ± 0.20 mmol/L. Patients with severe SLOS (n = 4, clinical score > 50) died as newborns and had the lowest t-cholesterol (0.66 ± 0.27 mmol/L), and 7DHC was 0.47 ± 0.14 mmol/L. Correlation coefficient with clinical severity was 0.74 for initial t-cholesterol and 0.669 for Cho/7DHC. Statistically significant difference was between the initial t-cholesterol of mild and severe SLOS (p = 0.01), and between the Cho/7DHC ratios of groups (p = 0.004). In severe SLOS, the percentage of α-lipoprotein was significantly lower than in typical (p = 0.003) and mild SLOS (p = 0.004). Although serum albumin, total bilirubin, and hemostasis parameters remained in the reference range during cholesterol supplementation (n = 10) combined with statin therapy (n = 9), increase of aspartate aminotransferase and alanine aminotransferase in 50 % of the patients probably refers to a reversible alteration of liver function; therefore, statin therapy was suspended. CONCLUSION: life expectancy is fundamentally determined by the initial t-cholesterol, but dehydrocholesterol and α-lipoprotein have prognostic value. Accumulation of hepatotoxic DHC may inhibit the synthesis of α-lipoproteins, decreasing the reverse cholesterol transport. During statin therapy, we suggest monitoring of lipid parameters and liver function.

Bisphenol A Diglycidyl Ether-Primary Amine Cooligomer-poly(ε-caprolactone) Networks: Synthesis and Characterization.

In this work, the preparation and systematic investigation of cross-linked polyurethane-epoxy (PU-EP) polymer systems are reported. The PU-EP polymers were prepared using a reaction of isocyanate (NCO)-terminated PU-prepolymer with diglycidyl ether of bisphenol A (DGEBA)-amine cooligomer. The oligomerization of DGEBA was carried out by adding furfurylamine (FA) or ethanolamine (EA), resulting in DGEBA-amine cooligomers. For the synthesis of NCO-terminated PU-prepolymer, poly(ε-caprolactone)diol (PCD) (Mn = 2 kg/mol) and 1,6-hexamethylene diisocyanate (HDI) were used. The cross-linking was achieved by adding DGEBA-amine cooligomer to PU-prepolymer, in which the obtained urethane bonds, due to the presence of free hydroxil groups in the activated DGEBA, served as netpoints. During cross-linking, ethanolamine provides an additional free hydroxyl group for the formation of a new urethane bond, while furfurylamine can serve as a thermoreversible coupling element (e.g., Diels-Alder adduct). The PU-EP networks were characterized using attenuated total reflectance Fourier-transform infrared spectroscopy (ATR-FTIR), differential scanning calorimetry (DSC), dynamical mechanical analysis (DMA) and scanning electron microscopy (SEM). The DMA curves of some PU-EPs (depending on the compositions and the synthetic method) revealed a plateau-like region above the melting temperature (Tm) of PCD, confirming the presence of a cross-linked structure. This property resulted in a shape memory (SM) behavior for these samples, which can be fine-tuned in the presence of furfurylamine through the formation of additional thermoreversible bonds (e.g., Diels-Alder adduct).

Temporary Anti-Corrosive Double Layer on Zinc Substrate Based on Chitosan Hydrogel and Epoxy Resin.

In practice, metal structures are frequently transported or stored before being used. Even in such circumstances, the corrosion process caused by environmental factors (moisture, salty air, etc.) can occur quite easily. To avoid this, metal surfaces can be protected with temporary coatings. The objective of this research was to develop coatings that exhibit effective protective characteristics while also allowing for easy removal, if required. Novel, chitosan/epoxy double layers were prepared on zinc by dip-coating to obtain temporary tailor-made and peelable-on-demand, anti-corrosive coatings. Chitosan hydrogel fulfills the role of a primer that acts as an intermediary between the zinc substrate and the epoxy film to obtain better adhesion and specialization. The resulting coatings were characterized using electrochemical impedance spectroscopy, contact angle measurements, Raman spectroscopy, and scanning electron microscopy. The impedance of the bare zinc was increased by three orders of magnitude when the protective coatings were applied, proving efficient anti-corrosive protection. The chitosan sublayer improved the adhesion of the protective epoxy coating. The structural integrity and absolute impedance of the protective layers were conserved in both basic and neutral environments. However, after fulfilling its lifespan, the chitosan/epoxy double-layered coating could be removed after treatment with a mild acid without damaging the substrate. This was because of the hydrophilic properties of the epoxy layer, as well as the tendency of chitosan to swell in acidic conditions.

Effect of the Preparation Method on the Properties of Eugenol-Doped Titanium Dioxide (TiO2) Sol-Gel Coating on Titanium (Ti) Substrates.

The focus of this study was the preparation of sol-gel titanium dioxide (TiO2) coatings, by the dip-coating technique, on Ti6Al4V (TiGr5) and specific Ti implant substrates. In order to confer antibacterial properties to the layers, Eugenol was introduced in the coatings in two separate ways: firstly by introducing the Eugenol in the sol (Eug-TiO2), and secondly by impregnating into the already deposed TiO2 coating (TiO2/Eug). Optimization of Eugenol concentration as well as long term were performed in orderboth short- and long-term Eugenol concentration was performed to investigate the prepared samples thoroughly. The samples were investigated by electrochemical impedance spectroscopy (EIS) and potentiodynamic polarization curves (PDP). To investigate their resistance against Gram-negative Escherichia coli bacteria, microbiological analysis was performed on coatings prepared on glass substrates. Structural studies (FT-IR analysis, Raman spectroscopy) were performed to confirm Eugenol-TiO2 interactions. Coating thicknesses and adhesion were also determined for all samples. The results show that Eug-TiO2 presented with improved anticorrosive effects and significant antibacterial properties, compared to the other investigated samples.

Barrier behaviour of partially N-acetylated chitosan layers in aqueous media.

Chitosan coatings of 353 ± 12 nm thickness were prepared on glass and zinc substrates by dip-coating method to study their barrier-behaviour. The coatings were chemically modified to increase their degree of acetylation (DA) from ca. 44 % up to ca. 98 % resulting a quasi-chitin coating. The effect of the acetylation reaction was studied by infrared spectroscopy, and the structural changes of the native and acetylated coatings were investigated by UV-Vis spectrophotometry and X-ray diffraction. The surface properties of the coated samples were characterized by wettability measurements - advancing water contact angle decreased from ca. 80° (native) to ca. 43° (fully acetylated) - and microscopic (SEM, AFM) studies. The barrier behaviour of the chitosan layer depending on the DA was evaluated by electrochemical impedance spectroscopy studies and with a special mesoporous silica - chitosan bilayer system by measuring the amount of dye (Rhodamine 6G) accumulated in the silica through the chitosan coating during an impregnation step. These methods showed significant decrease in the barrier-effect of the coatings with increasing DA (accumulation of approximately six times more dye and a reduction of charge transfer resistance by an order of magnitude), due to the structural and ionization changes in the coatings.

Biological effects of the plant-derived polyphenol resveratrol in human articular cartilage and chondrosarcoma cells.

The natural phytoestrogen resveratrol (RSV) may have therapeutic potential for arthritic conditions. RSV is chondroprotective for articular cartilage in rabbit models for arthritis, but its biological effects on human articular cartilage and chondrosarcoma cells are unknown. Effects of RSV on human articular cartilage homeostasis were studied by assessing production of matrix-degrading enzymes (MMP-13, ADAMTS4, and ADAMTS5), as well as proteoglycan production and synthesis. The counteractions of RSV against catabolic factors (e.g., FGF-2 or IL-1ß) were examined by in vitro and ex vivo using monolayer, three-dimensional alginate beads and cartilage explants cultures, respectively. RSV improves cell viability of articular chondrocytes and effectively antagonizes cartilage-degrading protease production that was initiated by catabolic and/or anti-anabolic cytokines in human articular chondrocytes. RSV significantly also enhances BMP7-promoted proteoglycan synthesis as assessed by (35) S-sulfate incorporation. Protein-DNA interaction arrays suggest that RSV inhibits the activation of transcription factors involved in inflammation and cartilage catabolic signaling pathways, including direct downstream regulators of MAPK (e.g., AP-1, PEA3) and NFκB. RSV selectively compromises survival of human chondrosarcoma cells, but not primary articular chondrocytes, revealing cell-specific activity of RSV on non-tumorigenic versus tumor-derived cells. We propose that RSV exerts its chondroprotective functions, in part, by deactivating p53-induced apoptosis in human primary chondrocytes, but not human chondrosarcoma. Our findings suggest that RSV has potential as a unique biologic treatment for both prevention and treatment of cartilage degenerative diseases.

Subtelomeric 6.7 Mb trisomy 10p and 5.6 Mb monosomy 21q detected by FISH and array-CGH in three related patients.

Cryptic subtelomeric chromosomal aberrations are responsible for 5-10% of moderate/severe and 1% of mild intellectual disability. Unbalanced subtelomeric chromosomal rearrangements result in variable phenotypes which seem to be highly influenced by both the size of the duplication/deletion and the chromosomes involved in the translocation. We report on three related patients with moderate intellectual disability, language delay, hypotonia, facial dysmorphism, cardiac anomalies, scoliosis, and kyphosis in whom a familial (maternal) unbalanced submicroscopic translocation was found by subtelomeric fluorescence in situ hybridization (FISH). This rearrangement resulted in a partial trisomy 10pter and partial monosomy 21qter. The karyotype was 46,XY.ish der(21)t(10;21)(p14;q22.2). Confirmation of a 6.7 Mb size distal duplication of the p15.3-14 region of chromosome 10 and a 5.6 Mb distal deletion of the q22.2-22.3 region of chromosome 21 was obtained by array-CGH. To our best knowledge, such a composition of subtelomeric unbalanced translocations has not yet been published. Detection of this aberration in successive pregnancies of carrier members of the family by prenatal FISH could prevent the recurrence of the disease. Furthermore, detection of the rearrangements and identification of genes located in the chromosomal regions involved might be of interest.

Detection of internal tandem duplications in the FLT3 gene by different electrophoretic methods.

BACKGROUND: In acute myeloid leukemia (AML), the internal tandem duplication (ITD) in the juxtamembrane domain of the FLT3 (Fms-like tyrosine kinase 3) gene is one of the most frequent genetic alterations associated with poor prognosis. METHODS: A complex evaluation of the analytical properties of the three most frequently used detection methods--PCR followed by agarose (AGE), polyacrylamide (PAGE) or capillary electrophoresis (CE)--was performed on 95 DNA samples obtained from 73 AML patients. RESULTS: All the three methods verified the presence of a mutant allele in 20 samples from 18 patients. AGE and PAGE could detect the presence of 1%-2% mutant allele, while the detection limit of CE was 0.28%. However, acceptable reproducibility (inter-assay CV <25%) of the mutant allele rate determination was only achievable above 1.5% mutant/total allele rate. The reproducibility of the ITD size determination by CE was much better, but the ITD size calculated by PeakScanner or GeneScan analysis was 7% lower as compared to values obtained by DNA sequencing. The presence of multiple ITD was over-estimated by PAGE and AGE due to the formation of heteroduplexes. CONCLUSIONS: This study suggests the use of PCR+CE in the diagnostics and the follow-up of AML patients. The data further supports the importance of proper analytical evaluation of home-made molecular biological diagnostic tests.

Biochemical and Clinical Effects of Vitamin E Supplementation in Hungarian Smith-Lemli-Opitz Syndrome Patients.

Smith-Lemli-Opitz syndrome (SLOS) is a severe monogenic disorder resulting in low cholesterol and high 7-dehydrocholesterol (7-DHC) levels. 7-DHC-derived oxysterols likely contribute to disease pathophysiology, and thus antioxidant treatment might be beneficial because of high oxidative stress. In a three-year prospective study, we investigated the effects of vitamin E supplementation in six SLOS patients already receiving dietary cholesterol treatment. Plasma vitamin A and E concentrations were determined by the high-performance liquid chromatography (HPLC) method. At baseline, plasma 7-DHC, 8-dehydrocholesterol (8-DHC) and cholesterol levels were determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS) method. The clinical effect of the supplementation was assessed by performing structured parental interviews. At baseline, patients were characterized by low or low-normal plasma vitamin E concentrations (7.19-15.68 µmol/L), while vitamin A concentrations were found to be normal or high (1.26-2.68 µmol/L). Vitamin E supplementation resulted in correction or significant elevation of plasma vitamin E concentration in all patients. We observed reduced aggression, self-injury, irritability, hyperactivity, attention deficit, repetitive behavior, sleep disturbance, skin photosensitivity and/or eczema in 3/6 patients, with notable individual variability. Clinical response to therapy was associated with a low baseline 7-DHC + 8-DHC/cholesterol ratio (0.2-0.4). We suggest that determination of vitamin E status is important in SLOS patients. Supplementation of vitamin E should be considered and might be beneficial.