TIDILAP: Treatment of iron deficiency in lipoprotein apheresis patients --A prospective observational multi-center cohort study comparing efficacy, safety and tolerability of ferric gluconate with ferric carboxymaltose.

OBJECTIVES: Iron deficiency (ID) and iron deficiency anemia (IDA) are common findings in patients undergoing lipoprotein apheresis (LA). Different intravenous (iv) formulations are used to treat ID in LA patients, however guidelines and data on ID/IDA management in LA patients are lacking. We therefore performed a prospective observational multi-center cohort study of ID/IDA in LA patients, comparing two approved i.v. iron formulations, ferric gluconate (FG) and ferric carboxymaltose (FCM). METHODS: Inclusion criteria were a) serum ferritin <100 µg/L or b) serum ferritin <300 µg/L and transferrin saturation <20%. Patients received either FG (62.5 mg weekly) or FCM (500 mg once in ID or up to 1000 mg if IDA was present) i.v. until iron deficiency was resolved. Efficacy and safety were determined by repeated laboratory and clinical assessment. Iron parameters pre and post apheresis were measured to better understand the pathogenesis of ID/IDA in LA patients. RESULTS: 80% of LA patients treated at the three participating centers presented with ID/IDA; 129 patients were included in the study. Serum ferritin and transferrin levels were reduced following apheresis (by 18% (p < 0.0001) and by 13% (p < 0.0001) respectively). Both FG and FCM were effective and well tolerated in the treatment of ID/IDA in LA patients. FCM led to a quicker repletion of iron stores (p < 0.05), while improvement of ID/IDA symptoms was not different. Number and severity of adverse events did not differ between FG and FCM, no severe adverse events occurred. CONCLUSIONS: Our results suggest that FG and FCM are equally safe, well-tolerated and effective in treating ID/IDA in LA patients. These data form the basis for follow-up randomized controlled trials to establish clinical guidelines.

An active atmospheric methane sink in high Arctic mineral cryosols.

Methane (CH4) emission by carbon-rich cryosols at the high latitudes in Northern Hemisphere has been studied extensively. In contrast, data on the CH4 emission potential of carbon-poor cryosols is limited, despite their spatial predominance. This work employs CH4 flux measurements in the field and under laboratory conditions to show that the mineral cryosols at Axel Heiberg Island in the Canadian high Arctic consistently consume atmospheric CH4. Omics analyses present the first molecular evidence of active atmospheric CH4-oxidizing bacteria (atmMOB) in permafrost-affected cryosols, with the prevalent atmMOB genotype in our acidic mineral cryosols being closely related to Upland Soil Cluster α. The atmospheric (atm) CH4 uptake at the study site increases with ground temperature between 0 °C and 18 °C. Consequently, the atm CH4 sink strength is predicted to increase by a factor of 5-30 as the Arctic warms by 5-15 °C over a century. We demonstrate that acidic mineral cryosols are a previously unrecognized potential of CH4 sink that requires further investigation to determine its potential impact on larger scales. This study also calls attention to the poleward distribution of atmMOB, as well as to the potential influence of microbial atm CH4 oxidation, in the context of regional CH4 flux models and global warming.

Profile of secreted hydrolases, associated proteins, and SlpA in Thermoanaerobacterium saccharolyticum during the degradation of hemicellulose.

Thermoanaerobacterium saccharolyticum, a Gram-positive thermophilic anaerobic bacterium, grows robustly on insoluble hemicellulose, which requires a specialized suite of secreted and transmembrane proteins. We report here the characterization of proteins secreted by this organism. Cultures were grown on hemicellulose, glucose, xylose, starch, and xylan in pH-controlled bioreactors, and samples were analyzed via spotted microarrays and liquid chromatography-mass spectrometry. Key hydrolases and transporters employed by T. saccharolyticum for growth on hemicellulose were, for the most part, hitherto uncharacterized and existed in two clusters (Tsac_1445 through Tsac_1464 for xylan/xylose and Tsac_1344 through Tsac_1349 for starch). A phosphotransferase system subunit, Tsac_0032, also appeared to be exclusive to growth on glucose. Previously identified hydrolases that showed strong conditional expression changes included XynA (Tsac_1459), XynC (Tsac_0897), and a pullulanase, Apu (Tsac_1342). An omnipresent transcript and protein making up a large percentage of the overall secretome, Tsac_0361, was tentatively identified as the primary S-layer component in T. saccharolyticum, and deletion of the Tsac_0361 gene resulted in gross morphological changes to the cells. The view of hemicellulose degradation revealed here will be enabling for metabolic engineering efforts in biofuel-producing organisms that degrade cellulose well but lack the ability to catabolize C5 sugars.

[Immunoadsorption for steroid-unresponsive multiple sclerosis-relapses: clinical data of 14 patients]. / Immunadsorption bei steroidrefraktärem Schub der Multiplen Sklerose : Klinische Erfahrungen bei 14 Patienten.

BACKGROUND: Multiple sclerosis (MS) is a chronic inflammatory demyelinating immune-mediated disease of the central nervous system, often associated with relapses. Plasma exchange (PE) has become established as an escalation therapy for steroid-unresponsive relapses in national and international guidelines. PE is a non-selective apheresis method with elimination of the entire plasma with subsequent substitution. Selective extracorporeal elimination of autoantibodies and immune complexes with immunoadsorption (IA) is increasingly replacing PE for the treatment of autoimmune neurological diseases due to its equivalent efficacy and advantageous safety profile. The use of IA for MS still remains to become established. The aim of this retrospective investigation was to evaluate efficacy and safety of IA in patients with steroid-unresponsive relapses. PATIENTS AND METHODS: Fourteen patients with steroid-unresponsive MS relapses were retrospectively analysed. Patients received six IA treatments within 2 weeks using the single-use tryptophan adsorber. Peripheral venous access was used in 11 patients, and 3 patients needed a central line. The plasma volume treated was 2 l per IA. Efficacy criteria were improvement in symptoms of MS relapses which were measured with the Kurtzke scale (EDSS, FS) and visual acuity measurements for patients with optic neuritis. RESULTS: In 12 of 14 patients the major symptom of MS relapse improved to a clinically relevant extent after tryptophan IA; no patient got worse, corresponding to a response rate of 86%. Mean EDSS and FS in patients with spastic paresis (n=4) and dizziness (n=2) as well as mean visual acuity in patients with optic neuritis (n=8) significantly improved after IA. IA treatments were safe, with good tolerability, and no severe adverse events occurred. CONCLUSION: Immunoadsorption for the treatment of steroid-unresponsive MS relapses was safe and effective. The response rate was comparable to published results with PE. With IA, in contrast to unselective PE, administration of human plasma products is not necessary, avoiding associated risks.

[Fibrinogen/LDL apheresis for treatment of sudden hearing loss: an observational study on 152 patients]. / H.E.L.P.-Apherese bei der Behandlung des Hörsturzes: Eine Anwendungsbeobachtung an 152 Patienten.

Disturbances of cochlear microcirculation are among the most discussed causes of sudden sensorineural hearing loss. Increased levels of cholesterol and fibrinogen seem to act as risk factors for inner ear disorders. Fibrinogen/LDL apheresis greatly reduces the concentration of plasma fibrinogen thus leading to improved cochlear blood flow. In a retrospective case series remission rates of 152 patients suffering from sudden sensorineural hearing loss and resistant to former treatment were investigated after treatment with a single apheresis. Complete remission was reported in 11% of patients, partial remission in 43%. 37% had no change of hearing threshold and 2% reported a decrease in hearing. Rates of complete remissions decreased from 22% within the first 2 weeks after onset of hearing loss to 14% after 6 weeks. In the same period of time rates of partial remissions decreased from 33% to 13%. The present study shows that apheresis achieved complete or partial remission in 54% of patients even after unsuccessful treatment with another therapy and the therapeutic window lies by approximately 6 weeks.

Formation and characterization of iron-oligonucleotide complexes with matrix-assisted laser desorption/ionization fourier transform ion cyclotron resonance mass spectrometry.

Iron-containing oligonucleotide negative ions can be generated by matrix-assisted laser desorption/ionization from a stainless steel target disk (by either defocusing the laser beam or by mixing iron salts such as FeCl3 with the matrix compound during the sample preparation). High resolution mass measurements reveal the presence of both Fe2+ (as M + Fe - 3H)- and Fe3+ (as M + Fe - 4H)- in the metal-oligonucleotide ions. The presence of Fe3+ is unexpected, and must involve replacement of protons from the nucleic bases or ribose groups as well as the phosphate groups of the oligonucleotides. Inspection of a range of small oligonucleotides and mononucleotides reveals that the presence of both Fe2+ and Fe3+ in the iron-biomolecule complexes is dependent on the number of acidic hydrogens that can be replaced in the oligonucleotide or nucleotide. Collisional dissociation of several metal-tetranucleotide ions revealed that the presence of the iron ion alters the fragmentation observed. The iron atom was observed to be present in all of the fragment ions, and, whenever possible, seemed to enhance the abundance of fragment ions containing both iron and a guanine nucleic base. These results suggest that iron may serve as a useful probe for characterizing phosphorylated biomolecules.

Long-term measurements of energy expenditure in severe burn injury.

The objective of this study was to evaluate resting energy expenditure (REE) in spontaneously breathing and artificially ventilated burn patients during the entire intensive care period. In 27 patients with 51 +/- 20% body surface area burned (BSAB) the REE was determined via indirect calorimetry. Three groups were formed according to the mortality prognosis index of Zawacki et al. In groups A, B, and C the predicted mortality rates were <20%, 20% to 80%, and >80%, respectively. The frequency of acute respiratory distress syndrome (ARDS), sepsis, renal failure, and mortality increased from group A toward group C. The REE test revealed wide individual variation and was usually overestimated by all tested formulas. The mean REE was comparable in groups A, B, and C during the first 20 days (49 +/- 16% vs. 59 +/- 21% vs. 57 +/- 18% above the REE calculated by the Harris-Benedict equation, or HBEE). The REE of patients in groups A and B declined after this period, whereas the long-term ventilated patients in the prognostically unfavorable group C showed a high REE up to the 45th day, usually accompanied by severe organ dysfunction and major metabolic disorders. During this time a nutritional regimen meeting the actual REE could not be achieved. In the clinical situation when indirect calorimetry is not available, REE can be stated to be 50% to 60% above HBEE in patients with >20% BSAB for at least 20 days. Expecting a stable clinical course in patients with a predicted mortality of <20% (group A), oral nutrition usually seems sufficient after a short period of artificial nutritional support (1 week). Patients with a predicted mortality of more than 20% have a complication-burdened clinical course and a prolonged period of ventilation (groups B and C). These patients need parenteral and enteral nutrition for at least 20 days after trauma to prevent severe malnutrition.

Influence of severe burn injury on the expression of RM 3/1 and HLA-DR antigens in human blood monocytes.

The effect of severe burns on the expression of the glucocorticoid-inducible RM 3/1 and HLA-DR antigens in blood monocytes was studied in patients with less than or more than 50% total body surface area (TBSA) burned. All patients showed a strong increase in the portion of RM 3/1+ monocytes within 1 day after injury. In patients with more than 50% TBSA, RM 3/1+ cells decreased after 2 days; in those with less than 50% TBSA, cells decreased after 3 days HLA-DR+ monocytes decreased within 4 days in both groups. In patients with less than 50% TBSA, HLA-DR+ monocytes slowly increased thereafter to basic levels. In patients with more than 50% TBSA, HLA-DR+ monocytes further decreased, then slowly increased, however, did not reach basic levels. This long-lasting decrease was evidence in the nonsurvivors. These results show that severe burns differently affect monocyte antigens. The induction of the anti-inflammatory subtype RM 3/1 and the decrease of the immunoregulatory HLA-DR antigens may contribute to the immunosuppression observed after burn injury.

Investigation of oligonucleotide fragmentation with matrix-assisted laser desorption/ionization Fourier-transform mass spectrometry and sustained off-resonance irradiation.

Matrix-assisted laser desorption/ionization (MALDI) can be combined with Fourier-transform ion cyclotron resonance mass spectrometry (FTMS) for the detailed structural examination of biomolecules such as peptides and oligonucleotides. We have been able to detect molecular ions for bovine heart cytochrome c (MW = 12,327) by MALDI-FTMS (355 nm laser desorption, 2,5-dihydroxybenzoic acid matrix). Although the mass resolution of these molecular ions is poor, the experiments verify that the MALDI-FTMS mass range for our 3-tesla instrument is in excess of m/z 12,000. Accurate mass measurements and selective dissociation experiments were used to examine the fragmentation pathways of small oligonucleotides in detail. Sustained off-resonance irradiation (SORI) was found to be superior to conventional on-resonance collisionally activated dissociation (CAD) for the efficient dissociation and detection of fragment ions for oligonucleotides. These experiments indicated that oligonucleotide fragmentation is a complex process and results not only from simple elimination of nucleic bases and cleavages of phosphate ester bonds, but also by rearrangement processes in which a terminal phosphate moiety can be transferred to an internal phosphate group.

Assessment of microcirculation of an axial skin flap using indocyanine green fluorescence angiography.

In many cases the complexities of skin-flap microcirculation are difficult to assess despite all the subjective and objective examination techniques available today. Adequate microcirculation is essential for tissue viability, so any method employed for studying microcirculation should provide as accurate an assessment of the prevailing conditions as possible. Of all the clinical methods, the fluorescence technique using the dye sodium fluorescein has so far provided the most reliable results. However, the pharmacokinetic properties of this tracer have prevented the technique from becoming established in clinical practice. The fluorescent dye indocyanine green (Cardio Green), on the other hand, has far more favorable pharmacokinetics. In an experimental animal model, the fluorescence technique using indocyanine green (indocyanine green angiography, ICGA) was used to study postoperative changes in the microcirculation of a skin flap. On the day of operation, indocyanine green angiography revealed a state of hemodynamic imbalance for which the organism was able to compensate in the postoperative phase with the aid of humoral, physical, and metabolic factors. With indocyanine green angiography it was possible to quantify objectively the new hemodynamic equilibrium. Basically, microcirculation may be quantified in temporal and spatial terms. The significant objectivity of indocyanine green angiography and short intervals between each examination favor its possible and meaningful use in clinical practice and give cause for continuing studies.

Analysis of modified oligonucleotides by matrix-assisted laser desorption/ionization Fourier transform mass spectrometry.

Matrix-assisted laser desorption/ionization (MALDI) Fourier transform ion cyclotron resonance mass spectrometry (FTMS) has been applied to the structural characterization of modified oligodeoxyribonucleotide 4-, 6-, and 11-mers. Each oligonucleotide contained one modified base, either an O6-methyl-substituted guanine, an N6-(10R)-trans-opened benzo[a]pyrenediol epoxide adduct of adenine, or an N2-(R)-styrene oxide adduct of guanine. 3-Hydroxypicolinic acid was used as the MALDI matrix for molecular weight and purity determinations, while either 2,5-dihydroxybenzoic acid (DHBA) or an anthranilic/nicotinic acid (AA/NA) mixture was used to induce fragmentation for the production of structurally significant fragment ions. For the 4- and 6-mers, the oligonucleotide sequence could be obtained from the direct AA/NA or DHBA spectra. Sequence information was also obtained by inserting a time delay between the laser desorption event and ion detection to permit metastable decomposition. For the 11-mers, high-mass sequence ions were not detected. Although similar sequence ions were observed in both the positive and the negative ion mass spectra, more fragmentation was generally observed in the positive ion mode. In the positive ion mode, modified base fragment ions were observed when DHBA was used, and these fragments were examined using accurate mass measurements, collisionally induced dissociations, and ion-molecule reactions to characterize the modified base. MALDI-FTMS signals from one sample application can be used for the measurement of hundreds of spectra. The direct MALDI-FT mass spectra show matrix-dependent, structurally informative fragments, and CID experiments can be implemented using low-picomole sample quantities.

Countinuous Laser-Doppler free-flap monitoring via the telephone line. Preliminary experiences.

It is possible to link a Laser-Doppler to a DOS-compatible personal computer in order to monitor microcirculation of the free flap after plastic surgery. We connected such a system to a second computer by telephone modems, in order to control the Laser-Doppler via the telephone line from a distant location using only commercially available hardware and software. The aim was to develop a low-cost system which would not require the purchase or construction of any specialised equipment. Using this system, it was possible to show the Laser-Doppler waveforms to colleagues who were not at the patient site, for their (more experienced) opinions without any further delays. Preliminary tests on five cases showed that it was indeed possible to transfer recorded information about the blood flow rate of free skin flaps from one computer to a remote one, without any loss of information. In these cases, the measured blood flow and clinical surveillance of the blood supply were adequate, thus making revision of the tissues not necessary. This preliminary study suggests that standard 'off-the-shelf' personal computer tools may be useful in improving the monitoring of microcirculation after plastic surgery.

Characterization of radiation-induced products of thymidine 3'-monophosphate and thymidylyl (3'-->5') thymidine by high-performance liquid chromatography and laser-desorption Fourier-transform mass spectrometry.

High-performance liquid chromatography (HPLC) and laser-desorption Fourier-transform mass spectrometry (LD FTMS) have been applied for direct measurements of radiation-induced products of nucleic acid constituents containing thymidine. Laser desorption FTMS could be used for the direct detection (neither hydrolyzed nor derivatized) of X ray-induced decomposition products of aqueous thymidine monophosphate. After these initial experiments, a variety of hydrogenated and hydroxylated thymine standards were acquired and examined by FTMS to assist in the identification of unknown radiation-induced decomposition products of thymine-containing nucleotides and dinucleotides. To extend these studies to dinucleotides, the radiation-induced products generated by the gamma radiolysis of thymidylyl (3'-->5') thymidine (TpT) were isolated by reverse-phase HPLC and identified by LD FTMS. Thymine and thymidine 3'-monophosphate were observed as the major products in this case. Several of the minor products of the HPLC profile were pooled in a single fraction and characterized simultaneously by LD FTMS. The resulting mass spectra indicated the presence of hydroxy-5,6-dihydrothymidine monophosphate, 5,6-dihydrothymidine monophosphate and thymidine monophosphate, thymine glycol, hydroxy-5,6-dihydrothymine, 5-hydroxy-methyluracil and 5,6-dihydrothymine. The combination of HPLC purification and LD FTMS structural characterization provides a useful tool for the direct measurement of radiation-induced products of nucleotides and dinucleotides.

[In-house radio-assisted emergency call system in continuous monitoring of microcirculation using laser doppler flowmetry]. / Dienstfunkergestütztes Notrufsystem bei kontinuierlicher Mikrozirkulationsüberwachung mit Hilfe der Laser-Doppler-Flowmetry.

Instrumental monitoring is being increasingly employed in the postoperative phase after microsurgery. For this purpose, laser Doppler flowmetry has proven to be very useful. Monitoring of the perfusion of tissue with blood is mainly by clinical examination, which, however, requires the physical presence of the observer, and this is generally not possible on a continuous basis. By connecting up a laser Doppler unit with a computer and telephone modem and making use of the in-house radio system, it proved possible to produce an automatic emergency call system that responds to a worsening of perfusion and alerts the responsible doctor. For this purpose, the software program "call-doc" was developed, which provides the necessary monitoring and control functions. A general perfusion-independent emergency call can also be triggered at the touch of a button.

Treatment of skin defects using suspensions of in vitro cultured keratinocytes.

The in vitro cultivation of keratinocytes and their application in the form of confluent sheets to cover various kinds of skin defects involves a number of problematical steps which could be improved by using single cell suspensions instead. Therefore we developed a method to apply keratinocytes suspended as single cells in a fibrin gel. By testing the feasibility of this method in different experimental animal models we found that it facilitates cultivation as well as application of the cells, moreover, this method allows a much more flexible use of the cells, i.e. it is easier to consider the clinical condition of the patient than by the conventional method.

Structural characterization of polycyclic aromatic hydrocarbon dihydrodiol epoxide DNA adducts using matrix-assisted laser desorption/ionization Fourier transform mass spectrometry.

Matrix-assisted laser desorption/ionization (MALDI) Fourier transform ion cyclotron resonance mass spectrometry (FTMS) has been applied for the structural characterization of four polycyclic aromatic hydrocarbon dihydrodiol epoxide (PAHDE) adducts, including the 5,6-dimethylchrysene DE adduct of 2'-deoxyadenosine, the 5-methyl- and 5,6-dimethylchrysene DE adducts of 2'-deoxyguanosine, and the benzo[a]pyrene-DE adduct of 2'-deoxyguanosyl 3'-phosphate. Measurement of positive and negative ion mass spectra, accurate mass determinations, and CID experiments were carried out using 10-40 ng (20-70 pmol) of sample. An evaluation of five MALDI matrices showed that matrix selection can be used to control the degree of analyte fragmentation. Three MALDI matrices commonly used for the analysis of proteins (sinapinic acid, ferulic acid, 2,5-dihydroxybenzoic acid) gave positive ion adduct mass spectra showing protonated or sodiated molecular ions accompanied by abundant, structurally informative fragment ions. Fragmentation was significantly reduced when working with two matrices used for oligonucleotide analysis (an anthranilic-nicotinic acid mixture and 3-hydroxypicolinic acid). Using the CID capabilities of FTMS, isolation and activation of the MALDI-produced ions was used to provide additional structural information. While characteristic negative ions were not detected for the adenosyladduct, the guanosyl and guanosyl 3'-phosphate adducts gave [M-H]- ions when the anthranilic-nicotinic acid matrix mixture was used. The guanosyl adducts also showed [M-H-2H2O]- fragments. Compared with FAB or FAB-MS/MS for the analysis of underivatized PAH-DE adducts, MALDI-FTMS signals are long-lived, the direct MALDI-FT mass spectra show more structurally informative fragments, and accurate mass and CID experiments require lower sample quantities.

[Ergospirometry in sarcoidosis patients]. / Spiroergometrie bei Sarkoidosepatienten.

Our investigation should clarify the question of whether cardiopulmonary exercise testing (CPX) offers additional information of clinical relevance in patients with pulmonary sarcoidosis. We investigated the relationship between changes of lung function including the diffusing capacity and the changes of gas exchange. Furthermore, we determined the correlation of functional and morphological lung changes on the basis of radiological parameters and on the influence of a steroid therapy over the gas exchange. Forty-seven patients with histologically diagnosed pulmonary sarcoidosis were examined. The lung function test includes the measurement of lung volumes with spirometry and the diffusing capacity of carbon monoxide in the single breath technique. Minute ventilation, oxygen uptake at maximum, oxygen pulse, functional dead space/tidal volume ratio, anaerobic threshold, and alveolar-arterial oxygen difference were measured continuously during exercise. We found significant correlations between the lung volumes and the CPX data. An increased relative minute ventilation and an increased alveolar-arterial oxygen difference with exercise allows a more objective differentiation in radiological stages I and II than does the lung function at rest. A steroid therapy improves the effectivity of alveolar ventilation and an improvement of the ventilation-perfusion-match. The CPX detects early changes of pathological gas-exchange, before the lung function at rest is evident pathologically. It offers additional information on the pathophysiology of sarcoidosis, and is useful in reviewing steroid therapy.

Synthesis and Characterization of Molybdenum Carbide Clusters MonC4n, (n = 1 to 4).

Laser radiation (XeCl laser, 308-nanometer wavelength) focused into a cell containing Mo(CO)(6) vapor produced ultrafine particles in the extended waist of the laser beam. Negative ion mass spectrometry revealed molybdenum carbide cluster ions with a stoichiometry MonC4n (n = 1 to 4). The MonC4n(-) (n = 2 to 4) ions are completely unreactive with NH(3), H(2)O, and O(2), suggesting structures in which the molybdenum atoms are unavailable for coordination to additional ligands. Collision-induced dissociation studies of these anions show the loss of MoC(4) units as the main fragmentation pathway. This observation, together with the lack of addition reactions, provides a basis for structures in which a planar cluster of two, three, or four molybdenum atoms is surrounded by, and bonded to, carbon dimers.