Electrical Vestibular Stimuli to Enhance Vestibulo-Motor Output and Improve Subject Comfort

Electrical vestibular stimulation is often used to assess vestibulo-motor and postural responses in both clinical and research settings. Stochastic vestibular stimulation (SVS) is a recently established technique with many advantages over its square-wave counterpart; however, the evoked muscle responses remain relatively small. Although the vestibular-evoked responses can be enhanced by increasing the stimulus amplitude, subjects often perceive these higher intensity electrical stimuli as noxious or painful. Here, we developed multisine vestibular stimulation (MVS) signals that include precise frequency contributions to increase signal-to-noise ratios (SNR) of stimulus-evoked muscle and motor responses. Subjects were exposed to three different MVS stimuli to establish that: 1) MVS signals evoke equivalent vestibulo-motor responses compared to SVS while improving subject comfort and reducing experimentation time, 2) stimulus-evoked vestibulo-motor responses are reliably estimated as a linear system and 3) specific components of the cumulant density time domain vestibulo-motor responses can be targeted by controlling the frequency content of the input stimulus. Our results revealed that in comparison to SVS, MVS signals increased the SNR 3–6 times, reduced the minimum experimentation time by 85% and improved subjective measures of comfort by 20–80%. Vestibulo-motor responses measured using both EMG and force were not substantially affected by nonlinear distortions. In addition, by limiting the contribution of high frequencies within the MVS input stimulus, the magnitude of the medium latency time domain motor output response was increased by 58%. These results demonstrate that MVS stimuli can be designed to target and enhance vestibulo-motor output responses while simultaneously improving subject comfort, which should prove beneficial for both research and clinical applications.     

Blood Born miRNAs Signatures that Can Serve as Disease Specific Biomarkers Are Not Significantly Affected by Overall Fitness and Exercise

Blood born micro(mi)RNA expression pattern have been reported for various human diseases with signatures specific for diseases. To evaluate these biomarkers, it is mandatory to know possible changes of miRNA signatures in healthy individuals under different physiological conditions. We analyzed the miRNA expression in peripheral blood of elite endurance athletes and moderatly active controls. Blood drawing was done before and after exhaustive exercise in each group. After Benjamini-Hochberg adjustment we did not find any miRNA with significant p-values when comparing miRNA expression between the different groups. We found, however, 24 different miRNAs with an expression fold change of minimum 1.5 in at least one of the comparisons (athletes before vs after exercise, athletes before exercise vs controls and athletes after exercise vs controls). The observed changes are not significant in contrast to the expression changes of the blood born miRNA expression reported for many human diseases. These data support the idea of disease associated miRNA patterns useful as biomarkers that are not readily altered by physiological conditions.     

CD70-restricted specific activation of TRAILR1 or TRAILR2 using scFv-targeted TRAIL mutants

To combine the CD27 stimulation inhibitory effect of blocking CD70 antibodies with an antibody-dependent cellular cytotoxicity (ADCC)-independent, cell death-inducing activity for targeting of CD70-expressing tumors, we evaluated here fusion proteins of the apoptosis-inducing TNF family member TRAIL and a single-chain variable fragment (scFv) derived from a high-affinity llama-derived anti-human CD70 antibody (lαhCD70). A fusion protein of scFv:lαhCD70 with TNC-TRAIL, a stabilized form of TRAIL, showed strongly enhanced apoptosis induction upon CD70 binding and furthermore efficiently interfered with CD70-CD27 interaction. Noteworthy, introduction of recently identified mutations that discriminate between TRAILR1 and TRAILR2 binding into the TRAIL part of scFv:lαhCD70-TNC-TRAIL resulted in TRAIL death receptor-specific fusion proteins with CD70-restricted activity.     

Circadian and Ultradian Time Patterns in Human Behaviour: Part 1: Activity Monitoring of Families from Prepartum to Postpartum

Time patterns of activity-rest rhythms during and after pregnancy are increasingly recognised as important factors for the well-being and health of young families. This longitudinal study examined activity-rest patterns of couples during late pregnancy and subsequently the alterations in the periodic structure of parental and neonatal time patterns during the first four months after birth. Part I concentrates on the effects of late pregnancy and birth to the mother's rest-activity patterns and those of the father and, after birth, what time pattern the infant developed. Part II attempts to clarify how activity patterns of the entire family agree or disagree with each other and investigates how the infant synchronises with the environment that includes the process of parent-infant interaction. Activity data of, so far, seven families (father, mother and child) were continuously recorded using non-invasive Actiwatch units. Recordings of parental activity started at the beginning of the 37th week of gestation, and were continued in parallel with the infants' recordings in three series of three weeks each until four months after birth: 1st to 3rd week, 7th to 9th week and 13th to 15th week of life. In a standardised diary, record was kept of household routines, parental activities, type of feeding, initiation of sleep or waking up. Activity data of seven non-pregnant women were collected and used as a control. Irregular nocturnal activity epochs occurred frequently in pregnant women and were absent in non-pregnant women. Period lengthenings and shortenings of the circadian rhythms appeared in both parents from prepartum to postpartum. Activity at night increased from prepartum to postpartum in mothers and fathers. Three infants showed a marked circadian rhythm between day 3 and 14 after birth. All seven infants showed a predominant circadian rhythm between day 8 and 19 after birth. The onset of daytime activity of mothers and their infants corresponded well to each other. Postpartum frequency spectra of parents and child always had some ultradian components in common. Time patterns of activity-rest rhythms of couples and parents are shown to be altered during and after pregnancy and we suggest that the infants' adaptation to the environment begins during the first week that includes the process of mother-infant interaction.     

Treatment-independent miRNA signature in blood of wilms tumor patients

ABSTRACT: BACKGROUND: Blood-born miRNA signatures have recently been reported for various tumor diseases. Here, we compared themiRNA signature in Wilms tumor patients prior and after preoperative chemotherapy according to SIOPprotocol 2001. RESULTS: We did not find a significant difference between miRNA signature of both groups. However both, Wilmstumor patients prior and after chemotherapy showed a miRNA signature different from healthy controls. Thesignature of Wilms tumor patients prior to chemotherapy showed an accuracy of 97.5% and of patients afterchemotherapy an accuracy of 97.0%, each as compared to healthy controls. CONCLUSION: Our results provide evidence for a blood-born Wilms tumor miRNA signature largely independent of fourweeks preoperative chemotherapy treatment.     

Selective inhibition of FLICE-like inhibitory protein expression with small interfering RNA oligonucleotides is sufficient to sensitize tumor cells for TRAIL-induced apoptosis

BACKGROUND: Most tumors express death receptors and their activation represents a potential selective approach in cancer treatment. The most promising candidate for tumor selective death receptor-activation is tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)/Apo2L, which activates the death receptors TRAIL-R1 and TRAIL-R2, and induces apoptosis preferentially in tumor cells but not in normal tissues. However, many cancer cells are not or only moderately sensitive towards TRAIL and require cotreatment with irradiation or chemotherapy to yield a therapeutically reasonable apoptotic response. Because chemotherapy can have a broad range of unwanted side effects, more specific means for sensitizing tumor cells for TRAIL are desirable. The expression of the cellular FLICE-like inhibitory protein (cFLIP) is regarded as a major cause of TRAIL resistance. We therefore analyzed the usefulness of targeting FLIP to sensitize tumor cells for TRAIL-induced apoptosis. MATERIALS AND METHODS: To selectively interfere with expression of cFLIP short double-stranded RNA oligonucleotides (small interfering RNAs [siRNAs]) were introduced in the human cell lines SV80 and KB by electroporation. Effects of siRNA on FLIP expression were analyzed by Western blotting and RNase protection assay and correlated with TRAIL sensitivity upon stimulation with recombinant soluble TRAIL and TRAIL-R1- and TRAIL-R2-specific agonistic antibodies. RESULTS: FLIP expression can be inhibited by RNA interference using siRNAs, evident from reduced levels of FLIP-mRNA and FLIP protein. Inhibition of cFLIP expression sensitizes cells for apoptosis induction by TRAIL and other death ligands. In accordance with the presumed function of FLIP as an inhibitor of death receptor-induced caspase-8 activation, down-regulation of FLIP by siRNAs enhanced TRAIL-induced caspase-8 activation. CONCLUSION: Inhibition of FLIP expression was sufficient to sensitize tumor cells for TRAIL-induced apoptosis. The combination of TRAIL and FLIP-targeting siRNA could therefore be a useful strategy to attack cancer cells, which are resistant to TRAIL alone.     

Dye affinity chromatography for fast and simple purification of fucoidan from marine brown algae

Fucoidan is a sulfated polysaccharide with promising pharmacological applications. Due to its medicinal properties, there is a demand for a separation technique that yields a high purification grade. Here, we present a novel purification tool for recovering fucoidan from the marine brown macroalgae Fucus vesiculosus. The developed method is based on amino‐derivatized Sepabeads^® EC‐EA. The beads were modified with toluidine blue (TB), a thiazine derivative, to exploit the strong donor acceptor interactions between the cationic dye and the anionic polysaccharide. The adsorption kinetics and the binding capacity of the resin were analyzed. A Sips model was used to approximate the adsorption isotherm, resulting in a maximum capacity of 127.7 mg fucoidan per g adsorbent. Investigation of the effect of adsorption step's pH on purity and chemical structure was performed by TB and Fourier transform infra‐red spectroscopy assays. Results showed that adsorption at pH 1 and 6 had negligible effects on fucoidan's chemical structure. However, purity was actually improved by 1.55‐ and 1.69‐fold at pH 1 and 6, respectively, with an average yield of 5 g/100 g dried algae powder. In contrast, only a 1.46‐fold increment was observed in fucoidan purified by the traditional method at pH 2, with a yield of 7.5 g/100 g dried algae powder. Furthermore, fucoidan purified by this method at pH 6 complies with, or even exceeds the quality of the commercially available (≥95% pure) fucoidan (Sigma‐Aldrich^®) with respect to molecular weight and sulfur content. Therefore, dye affinity chromatography provides more advantages than the classically used techniques for fucoidan purification.     

Nicotinic acetylcholine receptors of Drosophila: three subunits encoded by genomically linked genes can co-assemble into the same receptor complex

The second beta-like subunit (SBD) is a putative structural subunit of Drosophila melanogaster nicotinic acetylcholine receptors (nAChRs). Here we have produced specific antibodies against SBD to study, which other nAChR subunits can co-assemble with SBD in receptor complexes of the Drosophila nervous system. Immunohistochemical studies in the adult optic lobe revealed that SBD has a distribution similar to that of the alpha-subunit ALS in the synaptic neuropil. The subunits ALS, D(alpha)2 and SBD can be co-purified by alpha-bungarotoxin affinity chromatography. Moreover, anti-SBD antibodies co-precipitate ALS and D(alpha)2 and, vice versa, ALS and D(alpha)2 antibodies co-immunoprecipitate SBD protein. Two-step immunoaffinity chromatography with immobilized antibodies against ALS and D(alpha)2 revealed the existence of nAChR complexes that include ALS, D(alpha)2 and SBD as integral components. Interestingly, the genes encoding these three subunits appear to be directly linked in the Drosophila genome at region 96 A of the third chromosome. In addition, SBD appears to be a component of a different receptor complex, which includes the ARD protein as an additional beta-subunit, but neither ALS nor D(alpha)2 nor the third alpha-subunit D(alpha)3. These findings suggest a considerable complexity of the Drosophila nicotinic receptor system.     

ChipcheckII—Predicting Binding Curves for Multiple Analyte Strands on Small DNA Microarrays

Incomplete binding, saturation, and cross-hybridization between partially complementary strands complicate the parallel detection of nucleic acids via DNA microarrays. Treating the competing equilibria governing binding to microarrays requires computational tools. We have developed the web-based program ChipCheckII that calculates total hybridization matrices for target strands interacting with probes on small DNA microarrays. The program can be used to compute the extent of cross-hybridization and other phenomena affecting fidelity of detection based on sequences, quantities of strands, and hybridization conditions as inputs. Enthalpy and entropy of duplex formation are generated locally with UNAfold, including those for complexes that are partially matched. Simulated binding versus temperature curves for portions of a commercial genome chip demonstrate the extent to which cross-hybridization can complicate DNA detection. ChipCheckII is expected to aid nucleic acid chemists in developing high fidelity DNA microarrays.