Immobilizing bacteriorhodopsin on a single electron transistor

As awareness of potential human and environmental impacts from toxins has increased, so has the development of innovative sensors. Bacteriorhodopsin (bR) is a light activated proton pump contained in the purple membrane (PM) of the bacteria Halobacterium salinarum. Bacteriorhodopsin is a robust protein which can function in both wet and dry states and can withstand extreme environmental conditions. A single electron transistor(SET) is a nano-scale device that exploits the quantum mechanical properties of electrons to switch on and off. SETs have tremendous potential in practical applications due to their size, ultra low power requirements, and electrometer-like sensitivity. The main goal of this research was to create a bionanohybrid device by integrating bR with a SET device. This was achieved by a multidisciplinary approach. The SET devices were created by a combination of sputtering, photolithography, and focused ion beam machining. The bionanomaterial bacteriorhodopsin was created through oxidative fermentation and a series of transmembrane purification processes. The bR was then integrated with the SET by electrophoretic deposition, creating a bionanohybrid device. The bionanohybrid device was then characterized using a semiconductor parametric analyzer. Characterization demonstrated that the bR modulated the operational characteristics of the SET when bR was activated with light within its absorbance spectrum. To effectively integrate bacteriorhodopsin with microelectromechanical systems (MEMS) and nanoelectromechanical systems (NEMS), it is critical to know the electrical properties of the material and to understand how it will affect the functionality of the device. Tests were performed on dried films of bR to determine if there is a relationship between inductance, capacitance, and resistance (LCR) measurements and orientation, light-on/off, frequency, and time. The results indicated that the LCR measurements of the bR depended on the thickness and area of the film, but not on the orientation, as with other biological materials such as muscle. However, there was a transient LCR response for both oriented and unoriented bR which depended on light intensity. From the impedance measurements an empirical model was suggested for the bionanohybrid device. The empirical model is based on the dominant electrical characteristics of the bR which were the parallel capacitance and resistance. The empirical model suggests that it is possible to integrate bR with a SET without influencing its functional characteristics.

Correction of erosion after suburethral sling insertion for stress incontinence: results and related sexual function

BACKGROUND: Suburethral slings are commonly used for the surgical treatment of female stress incontinence; occasionally they can cause erosion and dyspareunia. OBJECTIVES: The primary aim of this study is to determine the outcome after reclosure of the vaginal epithelium for suburethral sling erosion. Sexual function was assessed before and after intervention using the Female Sexual Function Index (FSFI) questionnaire. DESIGN, SETTING, AND PARTICIPANTS: This was a prospective case-controlled study in which, between December 2005 and December 2007, we included patients who were referred to the Department of Urogynaecology because of vaginal erosion after suburethral sling insertion for urinary stress incontinence. For evaluation of sexual function, all patients filled in an FSFI questionnaire before intervention and at follow-up. All patients underwent gynaecological examination including colposcopy, and the site and size of the defect were noted. INTERVENTION: The edge of the vaginal epithelium was trimmed, mobilized, and closed with interrupted vertical Vicryl mattress sutures in a single layer. MEASUREMENTS: FSFI questionnaire and clinical findings. RESULTS AND LIMITATIONS: Twenty-one patients were included in the study. Eighteen patients with larger defects were operated on, and three defects healed after topical application of estrogen cream. In 16 patients, the defect had healed at follow-up; two patients with persisting defects were brought back to surgery and the procedure was repeated, paying particular attention to tension-free adaptation of vaginal tissue. In one patient, partial sling removal was performed after the second failed intervention. The domains of desire (p<0.0001), arousal (p<0.0003), lubrication (p<0.0001), satisfaction (p<0.0130), and pain (p<0.0001) improved significantly. Orgasm remained unchanged (p=0.4130; all two-tailed t-test). CONCLUSION: Suburethral erosion can be treated effectively by resuturing. Sexual function is improved in regard to desire, arousal, lubrication, satisfaction, and pain, but not orgasm. In septic patients and patients with a history of radiation, grossly infected tissue, or severe pain, excision of the mesh needs to be considered.

Integration of room temperature single electron transistor with CMOS subsystem

The single electron transistor (SET) is a charge-based device that may complement the dominant metal-oxide-semiconductor field effect transistor (MOSFET) technology. As the cost of scaling MOSFET to smaller dimensions are rising and the the basic functionality of MOSFET is encountering numerous challenges at dimensions smaller than 10nm, the SET has shown the potential to become the next generation device which operates based on the tunneling of electrons. Since the electron transfer mechanism of a SET device is based on the non-dissipative electron tunneling effect, the power consumption of a SET device is extremely low, estimated to be on the order of 10^-18J. The objectives of this research are to demonstrate technologies that would enable the mass produce of SET devices that are operational at room temperature and to integrate these devices on top of an active complementary-MOSFET (CMOS) substrate. To achieve these goals, two fabrication techniques are considered in this work. The Focus Ion Beam (FIB) technique is used to fabricate the islands and the tunnel junctions of the SET device. A Ultra-Violet (UV) light based Nano-Imprint Lithography (NIL) call Step-and-Flash- Imprint Lithography (SFIL) is used to fabricate the interconnections of the SET devices. Combining these two techniques, a full array of SET devices are fabricated on a planar substrate. Test and characterization of the SET devices has shown consistent Coulomb blockade effect, an important single electron characteristic. To realize a room temperature operational SET device that function as a logic device to work along CMOS, it is important to know the device behavior at different temperatures. Based on the theory developed for a single island SET device, a thermal analysis is carried out on the multi-island SET device and the observation of changes in Coulomb blockade effect is presented. The results show that the multi-island SET device operation highly depends on temperature. The important parameters that determine the SET operation is the effective capacitance Ceff and tunneling resistance Rt . These two parameters lead to the tunneling rate of an electron in the SET device, Γ. To obtain an accurate model for SET operation, the effects of the deviation in dimensions, the trap states in the insulation, and the background charge effect have to be taken into consideration. The theoretical and experimental evidence for these non-ideal effects are presented in this work.

QUANTUM CORRELATIONS OF LIGHTS IN MACROSCOPIC ENVIRONMENTS

This dissertation presents a detailed study in exploring quantum correlations of lights in macroscopic environments. We have explored quantum correlations of single photons, weak coherent states, and polarization-correlated/polarization-entangled photons in macroscopic environments. These included macroscopic mirrors, macroscopic photon number, spatially separated observers, noisy photons source and propagation medium with loss or disturbances. We proposed a measurement scheme for observing quantum correlations and entanglement in the spatial properties of two macroscopic mirrors using single photons spatial compass state. We explored the phase space distribution features of spatial compass states, such as chessboard pattern by using the Wigner function. The displacement and tilt correlations of the two mirrors were manifested through the propensities of the compass states. This technique can be used to extract Einstein-Podolsky-Rosen correlations (EPR) of the two mirrors. We then formulated the discrete-like property of the propensity Pb(m,n), which can be used to explore environmental perturbed quantum jumps of the EPR correlations in phase space. With single photons spatial compass state, the variances in position and momentum are much smaller than standard quantum limit when using a Gaussian TEM00 beam. We observed intrinsic quantum correlations of weak coherent states between two parties through balanced homodyne detection. Our scheme can be used as a supplement to decoy-state BB84 protocol and differential phase-shift QKD protocol. We prepared four types of bipartite correlations ±cos2(θ12) that shared between two parties. We also demonstrated bits correlations between two parties separated by 10 km optical fiber. The bits information will be protected by the large quantum phase fluctuation of weak coherent states, adding another physical layer of security to these protocols for quantum key distribution. Using 10 m of highly nonlinear fiber (HNLF) at 77 K, we observed coincidence to accidental-coincidence ratio of 130±5 for correlated photon-pair and Two-Photon Interference visibility >98% entangled photon-pair. We also verified the non-local behavior of polarization-entangled photon pair by violating Clauser-Horne-Shimony-Holt Bell’s inequality by more than 12 standard deviations. With the HNLF at 300 K (77 K), photon-pair production rate about factor 3(2) higher than a 300 m dispersion-shifted fiber is observed. Then, we studied quantum correlation and interference of photon-pairs; with one photon of the photon-air experiencing multiple scattering in a random medium. We observed that depolarization noise photon in multiple scattering degrading the purity of photon-pair, and the existence of Raman noise photon in a photon-pair source will contribute to the depolarization affect. We found that quantum correlation of polarization-entangled photon-pair is better preserved than polarization-correlated photon-pair as one photon of the photon-pair scattered through a random medium. Our findings showed that high purity polarization-entangled photon-pair is better candidate for long distance quantum key distribution.

Failures and complications in patients with birth defects restored with fixed dental prostheses and single crowns on teeth and/or implants

OBJECTIVES: To assess retrospectively, over at least 5 years, the incidences of technical and biological complications and failures in young adult patients with birth defects affecting the formation of teeth. MATERIAL AND METHODS: All insurance cases with a birth defect that had crowns and fixed dental prostheses (FDPs) inserted more than 5 years ago were contacted and asked to participate in a reexamination. RESULTS: The median age of the patients was 19.3 years (range 16.6-24.7 years) when prosthetic treatment was initiated. Over the median observation period of 15.7 years (range 7.4-24.9 years) and considering the treatment needs at the reexamination, 19 out of 33 patients (58%) with reconstructions on teeth remained free from all failures or complications. From the patients with FDPs and single unit crowns (SCs) on implants followed over a median observation period of 8 years (range 4.6-15.3 years), eight out of 17% or 47% needed a retreatment or repair at some point due to a failure or a complication. From the three groups of patients, the cases with amelogenesis/dentinogenesis imperfecta demonstrated the highest failure and complication rates. In the cases with cleft lip, alveolus and palate (CLAP) or hypodontia/oligodontia, 71% of the SCs and 73% of the FDPs on teeth (FDP T) remained complication free over a median observation period of about 16 years. Sixty-two percent of the SCs and 64% of the FDPs on implants remained complication free over 8 years. Complications occurred earlier with implant-supported reconstructions. CONCLUSIONS: Because healthy, pristine teeth can be left unprepared, implant-supported SCs and FDPs are the treatment choice in young adults with birth defects resulting in tooth agenesis and in whom the edentulous spaces cannot be closed by means of orthodontic therapy. However, the trend for earlier and more frequent complications with implant-supported reconstructions in young adults, expecting many years of function with the reconstructions, has to be weighed against the benefits of keeping teeth unprepared. In cases with CLAP in which anatomical conditions render implant placement difficult and in which teeth adjacent to the cleft require esthetic corrections, the conventional FDP T still remains the treatment of choice.

Prognostic clinical and radiological parameters for immediate facial nerve function following vestibular schwannoma surgery

OBJECTIVE: The paper aims to define the parameters available before surgery which could predict immediate facial nerve function after excision of a vestibular schwannoma (VS). METHODS: Ninety-nine patients with VS operated consecutively by a single surgeon using an identical surgical technique have been evaluated retrospectively. Data were collected regarding patients' sex, age at onset of symptoms and at surgery, initial symptoms, neurological status at presentation, early post-operative neurological status and complications. The main radiological parameters included in the study were tumour extension pattern, diameters, shape, and volume, as well as extent of bony changes of the internal auditory canal. RESULTS: As the tumour stage and volume increase, facial nerve function is worse after surgery (p < 0.001 and p < 0.05, respectively). Concomitantly, larger extra-meatal tumour diameters in three dimensions (sagittal, coronal and axial) led to worse function (p < 0.01). Anterior and/or caudal tumour extension (p = 0.001 and p = 0.004, respectively) had more significant correlation than posterior and/or cranial extension (p = 0.022 and p = 0.353, respectively). Polycyclic VS had the worst prognosis, followed by the tumours with oval shape. The extent of intra-meatal tumour growth does not correlate with immediate facial nerve outcome. The different angles, lengths and diameters of the internal auditory channel showed no significant correlation with facial nerve outcome. Patients with headache as an initial symptom and those with gait instability and/or pre-operative poor facial nerve function had significantly worse immediate facial nerve outcome. CONCLUSION: Our data suggests that the analysis of the radiological and neurological patient data prior to surgery could give reliable clues regarding the immediate post-operative facial nerve function.

Effect of urinary tract infection on reservoir function in patients with ileal bladder substitute

PURPOSE: We determined the functional consequences of urinary tract infection in patients with an ileal bladder substitute in terms of urinary continence, post-void residual and urinary retention. MATERIALS AND METHODS: A total of 48 patients with culture documented urinary tract infection (single organism, 10(5) or greater cfu) were retrospectively evaluated before, during and after the infection for changes in continence, post-void residual and urinary retention as well as for resolution of symptomatology after appropriate antibiotic therapy. RESULTS: Of the 48 patients 40 had a single infection while the remaining 8 had multiple urinary tract infection episodes. During daytime 27 of the 44 patients with previously good daytime continence experienced deterioration in their baseline voiding status while infected. Of the 40 patients who were previously continent at night 20 had incontinence while infected. There were 15 patients with documented post-void residual and urinary retention developed in 4 during the urinary tract infection. All patients returned to baseline continence status and reservoir function after appropriate antibiotic treatment based on objective and subjective assessments. CONCLUSIONS: Urinary tract infection may cause urinary incontinence in patients with ileal bladder substitutes. Therefore, when there are complaints of de novo urinary incontinence, a finding of post-void residual or an acute presentation of urinary retention, a urinary tract infection should be excluded. When the urinary tract infection is appropriately treated urinary continence and reservoir function return to their baseline status.

The role of common single-nucleotide polymorphisms on exon 9 and exon 12 skipping in nonmutated CFTR alleles

Classic cystic fibrosis (CF) is caused by two loss-of-function mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene, whereas patients with nonclassic CF have at least one copy of a mutant gene that retains partial function of the CFTR protein. In addition, there are several other phenotypes associated with CFTR gene mutations, such as idiopathic chronic pancreatitis. In CFTR-associated disorders and in nonclassic CF, often only one CFTR mutation or no CFTR mutations can be detected. In this study, we screened 23 patients with CFTR-associated disorders for CFTR mutations by complete gene testing and quantitative transcript analysis. Mutations were found in 10 patients. In cells from respiratory epithelium, we detected aberrant splicing of CFTR mRNA in all investigated individuals. We observed a highly significant association between the presence of coding single-nucleotide polymorphisms (coding SNPs, or cSNPs) and increased skipping of exon 9 and 12. This association was found both in patients and in normal individuals carrying the same cSNPs. The cSNPs c.1540A>G, c.2694T>G, and c.4521G>A may have affected pre-mRNA splicing by changing regulatory sequence motifs of exonic splice enhancers, leading to lower amounts of normal transcripts. The analysis of CFTR exons indicated that less frequent and weak exonic splicing enhancer (ESE) motifs make exon 12 vulnerable to skipping. The number of splice variants in individuals with cSNPs was similar to previously reported values for the T5 allele, suggesting that cSNPs may enhance susceptibility to CFTR related diseases. In addition, cSNPs may be responsible for variation in the phenotypic expression of CFTR mutations. Quantitative approaches rather than conventional genomic analysis are required to interpret the role of cSNPs.

Probing the function of ionotropic and G protein-coupled receptors in surface-confined membranes

This article reports on recent electrical and optical techniques for investigating cellular signaling reactions in artificial and native membranes immobilized on solid supports. The first part describes the formation of planar artificial lipid bilayers on gold electrodes, which reveal giga-ohm electrical resistance and the insertion and characterization of ionotropic receptors therein. These membranes are suited to record a few or even single ion channels by impedance spectroscopy. Such tethered membranes on planar arrays of microelectrodes offer mechanically robust, long-lasting measuring devices to probe the influence of different chemistries on biologically important ionotropic receptors and therefore will have a future impact to probe the function of channel proteins in basic science and in biosensor applications. In a second part, we present complementary approaches to form inside-out native membrane sheets that are immobilized on micrometer-sized beads or across submicrometer-sized holes machined in a planar support. Because the native membrane sheets are plasma membranes detached from live cells, these approaches offer a unique possibility to investigate cellular signaling processes, such as those mediated by ionotropic or G protein-coupled receptors, with original composition of lipids and proteins.

Compound heterozygous mutations affect protein folding and function in patients with congenital sucrase-isomaltase deficiency

BACKGROUND & AIMS: Congenital sucrase-isomaltase (SI) deficiency is an autosomal-recessive intestinal disorder characterized by a drastic reduction or absence of sucrase and isomaltase activities. Previous studies have indicated that single mutations underlie individual phenotypes of the disease. We investigated whether compound heterozygous mutations, observed in some patients, have a role in disease pathogenesis. METHODS: We introduced mutations into the SI complementary DNA that resulted in the amino acid substitutions V577G and G1073D (heterozygous mutations found in one group of patients) or C1229Y and F1745C (heterozygous mutations found in another group). The mutant genes were expressed transiently, alone or in combination, in COS cells and the effects were assessed at the protein, structural, and subcellular levels. RESULTS: The mutants SI-V577G, SI-G1073D, and SI-F1745C were misfolded and could not exit the endoplasmic reticulum, whereas SI-C1229Y was transported only to the Golgi apparatus. Co-expression of mutants found on each SI allele in patients did not alter the protein's biosynthetic features or improve its enzymatic activity. Importantly, the mutations C1229Y and F1745C, which lie in the sucrase domains of SI, prevented its targeting to the cell's apical membrane but did not affect protein folding or isomaltase activity. CONCLUSIONS: Compound heterozygosity is a novel pathogenic mechanism of congenital SI deficiency. The effects of mutations in the sucrase domain of SIC1229Y and SIF1745C indicate the importance of a direct interaction between isomaltase and sucrose and the role of sucrose as an intermolecular chaperone in the intracellular transport of SI.

Color and Scent: How Single Genes Influence Pollinator Attraction

A major function of angiosperm flowers is the recruitment of animal pollinators that serve to transfer pollen among conspecific plants. Distinct sets of floral characteristics, called pollination syndromes, are correlated with visitation by specific groups of pollinators. Switches among pollination syndromes have occurred in many plant families. Such switches must have involved coordinated changes in multiple traits and multiple genes. Two well-studied floral traits affecting pollinator attraction are petal color and scent production. We review current knowledge about the biosynthetic pathways for floral color and scent production and their interaction at the genetic and biochemical levels. A key question in the field concerns the genes that underlie natural variation in color and scent and how such genes affect pollinator preference, reproductive isolation, and ultimately speciation.

Topographic time-frequency decomposition of the EEG

Frequency-transformed EEG resting data has been widely used to describe normal and abnormal brain functional states as function of the spectral power in different frequency bands. This has yielded a series of clinically relevant findings. However, by transforming the EEG into the frequency domain, the initially excellent time resolution of time-domain EEG is lost. The topographic time-frequency decomposition is a novel computerized EEG analysis method that combines previously available techniques from time-domain spatial EEG analysis and time-frequency decomposition of single-channel time series. It yields a new, physiologically and statistically plausible topographic time-frequency representation of human multichannel EEG. The original EEG is accounted by the coefficients of a large set of user defined EEG like time-series, which are optimized for maximal spatial smoothness and minimal norm. These coefficients are then reduced to a small number of model scalp field configurations, which vary in intensity as a function of time and frequency. The result is thus a small number of EEG field configurations, each with a corresponding time-frequency (Wigner) plot. The method has several advantages: It does not assume that the data is composed of orthogonal elements, it does not assume stationarity, it produces topographical maps and it allows to include user-defined, specific EEG elements, such as spike and wave patterns. After a formal introduction of the method, several examples are given, which include artificial data and multichannel EEG during different physiological and pathological conditions.

Effect of lifetime endurance training on left atrial mechanical function and on the risk of atrial fibrillation

Background Left atrium (LA) dilation and P-wave duration are linked to the amount of endurance training and are risk factors for atrial fibrillation (AF). The aim of this study was to evaluate the impact of LA anatomical and electrical remodeling on its conduit and pump function measured by two-dimensional speckle tracking echocardiography (STE). Method Amateur male runners > 30 years were recruited. Study participants (n = 95) were stratified in 3 groups according to lifetime training hours: low (< 1500 h, n = 33), intermediate (1500 to 4500 h, n = 32) and high training group (> 4500 h, n = 30). Results No differences were found, between the groups, in terms of age, blood pressure, and diastolic function. LA maximal volume (30 ± 5, 33 ± 5 vs. 37 ± 6 ml/m2, p < 0.001), and conduit volume index (9 ± 3, 11 ± 3 vs. 12 ± 3 ml/m2, p < 0.001) increased significantly from the low to the high training group, unlike the STE parameters: pump strain − 15.0 ± 2.8, − 14.7 ± 2.7 vs. − 14.9 ± 2.6%, p = 0.927; conduit strain 23.3 ± 3.9, 22.1 ± 5.3 vs. 23.7 ± 5.7%, p = 0.455. Independent predictors of LA strain conduit function were age, maximal early diastolic velocity of the mitral annulus, heart rate and peak early diastolic filling velocity. The signal-averaged P-wave (135 ± 11, 139 ± 10 vs. 148 ± 14 ms, p < 0.001) increased from the low to the high training group. Four episodes of non-sustained AF were recorded in one runner of the high training group. Conclusion The LA anatomical and electrical remodeling does not have a negative impact on atrial mechanical function. Hence, a possible link between these risk factors for AF and its actual, rare occurrence in this athlete population, could not be uncovered in the present study.

Comparison between pulsed-wave Doppler- and tissue Doppler-derived Tei indices in fetuses with and without congenital heart disease

OBJECTIVES The aim of this study was to compare the right (RV) and left (LV) ventricular Tei indices obtained by pulsed-wave Doppler (PD) and tissue Doppler (TD) methods in fetuses with structurally normal and abnormal hearts. METHODS This was a retrospective cross-sectional study of 147 fetuses that had a fetal echocardiogram and Tei index measured during a 2-year period. The RV and LV Tei indices were measured using both PD and TD methods. The difference between the two methods of Tei index measurement was tested using paired sample t-test, Pearson correlation coefficient was used to examine their relationship, and the agreement between the methods was tested using Bland-Altman analysis. RESULTS A total of 87 fetuses had normal hearts and 60 had a congenital heart defect. Both PD and TD Tei indices were measured successfully from at least one ventricle in 123 cases and from both ventricles in 110 cases. The mean TD Tei index was significantly higher than the mean PD Tei index for both ventricles (P < 0.0001). There was a weak but statistically significant correlation between the PD and TD Tei indices of the right ventricle (r = 0.20, P = 0.029), whereas the PD and TD Tei indices of the left ventricle did not correlate significantly (r = 0.04, P = 0.684). When pairs of Tei indices measured by two different methods (123 pairs for the right ventricle and 111 for the left ventricle) were tested with Bland-Altman analysis, the bias and precision were 0.147 and 0.254, respectively, for the right ventricle, and 0.299 and 0.276, respectively, for the left ventricle. CONCLUSIONS Correlation between Tei indices measured by PD and TD methods is weak and the agreement between individual measurements is poor. Therefore, they should not be used interchangeably in the assessment of fetal cardiac function.

Expression, purification, and projection structure by single particle electron microscopy of functional human TRPM4 heterologously expressed in Xenopus laevis oocytes

Despite efforts implicating the cationic channel transient receptor potential melastatin member 4 (TRPM4) to cardiac, nervous, and immunological pathologies, little is known about its structure and function. In this study, we optimized the requirements for purification and extraction of functional human TRPM4 protein and investigated its supra-molecular assembly. We selected the Xenopus laevis oocyte expression system because it lacks endogenous TRPM4 expression, it is known to overexpress functional human membrane channels, can be used for structure-function analysis within the same system, and is easily scaled to improve yield and develop moderate throughput capabilities through the use of robotics. Negative-stain electron microscopy (EM) revealed various sized low-resolution particles. Single particle analysis identified the majority of the projections represented the monomeric form with additional oligomeric structures potentially characterized as tetramers. Two-electrode voltage clamp electrophysiology demonstrated that human TRPM4 is functionally expressed at the oocyte plasma membrane. This study opens the door for medium-throughput screening and structure-function determination of this important therapeutically relevant target.