Glucocorticoid Receptor/HCN4 Channels Interaction in Spinal Dorsal Horn Participates in the Regulation of Neuropathic Pain after Peripheral Nerve Injury in Rats.

We studied the interaction between glucocorticoid receptor (GR) and HCN4 channels in the rat model of spared nerve injury (SNI) in Sprague-Dawley rats (n=124). The animals were randomly divided into 6 groups: sham-operated (SO; n=24), SNI (reference group; n=20), and 4 experimental SNI groups intrathecally treated with dexamethasone (DEX; GR agonist; n=20), RU38486 (GR antagonist; n=20), ZD7288 (HCN channels blocker; n=20), and ZD7288+DEX (n=20). The paw mechanical withdrawal threshold (PWT) was measured one day before surgery (SO group) and on days 1, 3, 7, 14, and 21 after surgery. Behavioral results showed that mechanical hyperalgesia appeared on day 1 after SNI, while PWT decreased gradually with time. The expression of GR and HCN4 channels in L4-L6 dorsal horn of the spinal cord was detected by Western blotting and immunohistochemistry. In the reference group, SNI significantly increased GR expression up to day 14 after surgery in comparison with the SO group. The expression of GR showed a tendency to increase in the DEX group (with the maximum expression on days 14 and 21), significantly increased in the RU38486 group (maximum on day 7). In the ZD7288 group, GR expression was lower than in the SNI group and did not change throughout the experiment, suggesting that ZD7288 could block the expression of GR. In the DEX group, the expression of HCN4 channels was significantly higher on day 1 after SNI, but there were no differences in this parameter between the RU38486 and ZD7288 groups. In the ZD7288+DEX group, the expression of HCN4 channels significantly increased on days 14 and 21 after SNI. Thus, GR and HCN4 have the same linkage in the formation of central sensitization after SNI, but antagonists have no significant effect on the improvement of pain behavior.

COX-2-derived prostaglandins as mediators of the deleterious effects of nicotine in chronic kidney disease.

Tobacco smoking has been identified as a risk factor in the progression of chronic kidney disease (CKD). In previous studies, we showed that nicotine induces cyclooxygenase (COX)-2 expression in vivo and in vitro and that the administration of nicotine in vivo worsens the severity of renal injury in a model of subtotal renal ablation. In the present study, we tested the role of COX-2-derived prostaglandins on the deleterious effects of nicotine in CKD. Sham and 5/6 nephrectomy (5/6Nx) rats received tap water or nicotine (100 µg/mL) in the drinking water for 12 wk. Additional groups also systemically received the COX-2 inhibitor NS-398 (1.5 mg·kg-1·day-1 via osmotic minipump). The administration of nicotine worsened renal injury and proteinuria in 5/6Nx rats and increased proteinuria in sham rats. 5/6Nx rats had increased cortical production of the prostaglandins PGE2, PGI2, PGD2, and PGF2α and of thromboxane A2. In these rats, nicotine reduced the production of all prostaglandins examined except thromboxane A2. Treatment with the COX-2 inhibitor NS-398 resulted in complete inhibition of all prostaglandins studied and ameliorated renal injury and proteinuria in 5/6Nx rats on nicotine but not in 5/6 Nx rats on tap water. Nicotine also reduced the expression of megalin in all groups examined, and this was partially prevented by COX-2 inhibition. In the present study, we showed that in CKD, nicotine worsens renal injury at least in part by producing an imbalance in the production of prostaglandins. This imbalance in the production of prostaglandins likely plays a role in the deleterious effects of smoking on the progression of CKD.

Deficiency of brain structural sub-network underlying post-ischaemic stroke apathy.

BACKGROUND AND PURPOSE: This study aimed to reveal the structural basis of post-ischaemic stroke apathy, especially in relation to disruptions in structural connectivity. METHODS: Eighty-eight participants were included. The Apathy Evaluation Scale, clinician version, was used to characterize the severity of apathy. Diffusion tensor imaging tractography was used to examine white matter integrity and to reconstruct white matter networks using 90 nodes based on the automated anatomical labeling atlas. The degree for each node was extracted to determine the relationship to the severity of apathy. RESULTS: Apathy was not significantly associated with damage to any single brain region. The degrees of 24 nodes (limbic system, three nodes; frontal lobe, six; basal ganglia, two; temporal lobe, three; parietal lobe, three; insula, two; occipital lobe, five) were significantly correlated to the Apathy Evaluation Scale scores. These 24 nodes constituted an apathy-related sub-network and its global and local efficiencies were negatively correlated with apathy levels (global, r = -0.54, P < 0.01; local, r = -0.64, P < 0.01). Multivariate logistic regression indicated that decreased global efficiency of this sub-network was an independent risk factor for apathy (odds ratio 0.03, 95% confidence interval 0.01-0.04, P = 0.007). Efficiencies of the non-apathy-related sub-network (the remaining 66 nodes) did not correlate or predict the presence of apathy. CONCLUSIONS: Post-stroke apathy is not due to the dysfunction of a single region or circuit. Rather, it results from disconnection of a complex sub-network of brain regions. This provides new insights into the neuroanatomical basis of post-stroke apathy.

The study of the tumor stem cell properties of CD133+CD44+ cells in the human lung adenocarcinoma cell line A549.

We studied the tumor stem cell properties of the CD133+CD44+ subpopulation in the human lung adenocarcinoma cell line A549. A549 cells were classified into subpopulations based on differential expression patterns for CD133 and CD44. Cells from different subpopulations were cultured and subcutaneously injected into 32 nude mice. Our results as following, (1) The majority of A549 cells died, whereas only about 4.11% of cells divided and proliferated to form cell clones. (2) The expression of CD133 and CD44 in proliferative cancer cells was statistically significantly different from that in normal A549 cells (p < 0.001). (3) Cell proliferation in group A (CD133+CD44+) was the fastest among all groups. Cell proliferation in A549 cells was slower than in group A but faster than in groups B (CD133-CD44-), C (CD133-CD44+), and D (CD133+CD44-). (4) The tumorigenic capacity in cells from group A was significantly higher than that in cells from groups B (p<0.001), C (p<0.001) and D (p<0.04). In conclusion, CD133+CD44+ cells in the adenocarcinoma cell line A549 have expressive significant cancer stem cell properties with continuous proliferative capacity and differentiation potential.

Non-equilibrium thermodynamic analysis of transport properties in the nanofiltration of ionic liquid-water solutions.

Thenanofiltration of aqueous solutions of the ionic liquids (ILs) 1-butyl-3-methylimidazolium tetrafluoroborate ([Bmim]BF(4)), and 1-butyl-3-methylimidazolium bromide ([Bmim]Br) with a polyamide nanofiltration membrane was investigated. The practical transport coefficients, including hydrodynamic permeability (L(p)), reflection (sigma) and solute permeability (omega) were calculated in terms of a non-equilibrium thermodynamics approach. It was found that L(p) and sigma diminished as the concentration of the IL solutions increased. These characteristics are similar to those observed in inorganic electrolyte-water systems. In addition, it was shown that the rejection and volume flux for both ionic liquid solutions rose with feed pressure, while it decreased with feed concentration. The maximum rejection efficiencies for [Bmim]Br and [Bmim]BF(4) are 67 % and 60 %, respectively, on our experimental scale. All the data suggests that a highly efficient process for IL separation could be developed when the operating conditions are optimized further.

Development and validation of a novel 13-plex PCR system for commonly used short tandem repeats in horses (Equus caballus).

BACKGROUND: Due to the thriving development of the modern horse industry and the occurrence of horse related crimes, the demand for methods of individual horse identification, parentage tests and other genetic analyses is increasing. Previous methods had disadvantages that decreased the accuracy of the results, lacked the inclusion of all commonly used short tandem repeats (STR) or increased the experimental cost and time. OBJECTIVES: We aimed to develop a novel 13-plex STR typing system to resolve the above issues. STUDY DESIGN: Experimental study. METHODS: Twelve autosomal and most commonly used di-nucleotide STRs (AHT4, AHT5, ASB2, ASB17, ASB23, HMS2, HMS3, HMS6, HMS7, HTG4, HTG10 and VHL20), and a Y-chromosomal STR (YJ10) were included. We redesigned the primers of eight STRs to establish a novel multiplex PCR system and tested this system for species specificity, sensitivity and repeatability. RESULTS: Full profiles were easily generated in one fast PCR reaction using a low-cost polymerase, as little as 1 ng of horse DNA template and 13 pairs of primers labelled with fluorescent dyes. No full profile was generated from DNA templates of humans or other commonly encountered animals. We also established an allelic ladder that contained 110 alleles based on 200 horses from 12 breeds and calculated standard population genetic parameters based on 150 Thoroughbreds. Stutter analysis showed that the averages of the stutter ratios were distinctly lower than those of lower allele ratios and the combined probability of paternity exclusion for this system were 0.994659935 (CPEduo ) and 0.999854032 (CPEtrio ). MAIN LIMITATIONS: A nonspecific and relatively low peak at 316 bp was frequently observed in locus HMS2, which is a nonexistent allele in all horses and should be ignored. CONCLUSIONS: Our results indicate that this 13-plex STR genotyping system is sensitive, species-specific, cost-effective and robust for applications in the horse industry and forensic investigation.

JMJD3 enhances invasiveness and migratory capacity of non-small cell lung cancer cell via activating EMT signaling pathway.

OBJECTIVE: This study aimed to explore the expression of JMJD3 in non-small cell lung cancer (NSCLC) and to further study its association with clinical features and prognosis of patients. PATIENTS AND METHODS: Quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) was performed to examine the level of JMJD3 in 46 pairs of NSCLC tissues and para-cancerous specimens. The relationship between JMJD3 level and clinical features of NSCLC and patients' prognosis was analyzed. And JMJD3 expression in NSCLC cells was further verified by qRT-PCR. In addition, JMJD3 knockdown model was constructed using siRNA in cell lines including A549 and SPC-A1, and the effect of JMJD3 on the biological function of NSCLC cells was analyzed by cell counting kit-8 (CCK-8) and transwell migration and invasive-ness assays. Lastly, Western blot was performed to explore the potential mechanism. RESULTS: In this investigation, qRT-PCR results indicated that JMJD3 expression in above-mentioned tumor tissues was conspicuously higher than that in normal tissues. In addition, compared with patients with low level of JMJD3, patients with high level of JMJD3 had a higher incidence of lymphatic metastasis and distant metastasis and a lower overall survival rate. Meanwhile, the proliferation, invasiveness and migratory capacity of cells in the sh-JMJD3 group was conspicuously decreased when compared with the cells in negative control group. Western Blot results indicated that the levels of key proteins in EMT signaling pathway such as E-cadherin, N-cadherin, Vimentin, TGF-ß, and MMP-9 were notably decreased in sh-JMJD3 group. Besides, the addition of TGF-ß cytokines synergistically promoted the malignant progression of NSCLC induced by JMJD3. CONCLUSIONS: JMJD3 expression was found conspicuously increased in NSCLC, which might be close relevant to NSCLC lymphatic or distant metastasis as well as patients' poor prognosis. Therefore, we speculated that JMJD3 could promote invasiveness and migratory capacity of non-small cell lung cancer cells by activating EMT process.

Are older drivers' driving patterns during an on-road driving task representative of their real-world driving patterns?

OBJECTIVE: The current study investigated whether older drivers' driving patterns during a customized on-road driving task were representative of their real-world driving patterns. METHODS: Two hundred and eight participants (male: 68.80%; mean age = 81.52 years, SD = 3.37 years, range = 76.00-96.00 years) completed a customized on-road driving task that commenced from their home and was conducted in their own vehicle. Participants' real-world driving patterns for the preceding 4-month period were also collected via an in-car recording device (ICRD) that was installed in each participant's vehicle. RESULTS: During the 4-month period prior to completing the on-road driving task, participants' median real-world driving trip distance was 2.66 km (interquartile range [IQR] = 1.14-5.79 km) and their median on-road driving task trip distance was 4.41 km (IQR = 2.83-6.35 km). Most participants' on-road driving task trip distances were classified as representative of their real-world driving trip distances (95.2%, n = 198). CONCLUSIONS: These findings suggest that most older drivers were able to devise a driving route that was representative of their real-world driving trip distance. Future research will examine whether additional aspects of the on-road driving task (e.g., average speed, proportion of trips in different speed zones) are representative of participants' real-world driving patterns.

Ontogeny, sex dimorphism, and neonatal sex hormone determination of synapse-associated messenger RNAs in rat brain.

Sex hormones influence neurite outgrowth and synaptogenesis in certain hormone-dependent areas of the rat brain during neonatal development. These alterations are thought to mediate changes in brain structure and function between the sexes. Growth-associated protein 43 kDa (GAP-43) gene expression is estrogen-regulated in the adult ventromedial hypothalamus (VMH) and sexually dimorphic (M:F = 1.8:1) in adult cortex (CTX). Such effects intimate hormonal regulation of synaptic plasticity. To investigate the nature of these dimorphisms, the present study examined the ontogeny of expression of mRNAs encoding 3 neural-specific proteins: GAP-43, SCG10, and synaptosomal-associated protein 25 kDa (SNAP-25); and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), in the VMH and CTX; and also the effects of altering the neonatal sex hormonal milieu on the development of these adult dimorphisms. Levels of specific mRNAs in VMH and CTX were quantitated by slot-blot hybridization in rats of both sexes at different postnatal ages. To determine the involvement of neonatal sex hormones on the levels of these mRNAs, male neonatal rat pups were treated with an estrogen receptor antagonist or an aromatase inhibitor, and neonatal female pups were treated with testosterone or estrogen prior to slot-blot evaluations in adulthood. In VMH, GAP-43 mRNA levels were high on days P1 and P4 with a 3-fold decrease by day P23; in CTX, GAP-43 mRNA first increased by day P11, then fell to baseline by day P23. In VMH, SCG10 mRNA showed only small increases with time; but in CTX, there was a 5-fold drop from days P4 to P23. In VMH, SNAP-25 mRNA was low and changed only slightly; but in CTX there was a 5-fold increase between days P4 and P60. At birth, there was no sex dimorphism in either VMH or CTX, but the levels of all 3 neural-specific mRNAs were sexually dimorphic in adult CTX (M:F = 1.76 for GAP-43, 1.46 for SCG10, 1.44 for SNAP-25). GAPDH mRNA levels were regulated developmentally in VMH and CTX, but there was no sex dimorphism in either area. In male rats who received either an estrogen antagonist or aromatase inhibitor at birth, the CTX GAP-43 and SNAP-25 mRNA levels fell by 30%, to levels similar to untreated females. Conversely, in female rats, neonatal treatment with either testosterone or estrogen increased GAP-43 and SNAP-25 mRNA levels by about 30%, to levels similar to the untreated adult male. SCG10 levels did not demonstrate neonatal hormonal dependence.(ABSTRACT TRUNCATED AT 400 WORDS)

Characterization of the main effects, epistatic effects and their environmental interactions of QTLs on the genetic basis of yield traits in rice.

Main effects, epistatic effects and their environmental interactions of QTLs are all important genetic components of quantitative traits. In this study, we analyzed the main effects, epistatic effects of the QTLs, and QTL by environment interactions (QEs) underlying four yield traits, using a population of 240 recombinant inbred lines from a cross between two rice varieties tested in replicated field trials. A genetic linkage map with 220 DNA marker loci was constructed. A mixed linear model approach was used to detect QTLs with main effects, QTLs involved in digenic interactions and QEs. In total, 29 QTLs of main effects, and 35 digenic interactions involving 58 loci were detected for the four traits. Thirteen QTLs with main effects showed QEs; no QE was detected for the QTLs involved in epistatic interactions. The amount of variations explained by the QTLs of main effect were larger than the QTLs involved in epistatic interactions, which in turn were larger than QEs for all four traits. This study illustrates the ability of the analysis to assess the genetic components underlying the quantitative traits, and demonstrates the relative importance of the various components as the genetic basis of yield traits in this population.

Improved methods to determine optimal currents in electrical impedance tomography.

An electrical impedance tomography (EIT) system that uses the optimal current method to inject currents and the regularized Newton-Raphson algorithm to reconstruct an image of resistivity distribution is discussed. Iterative methods to derive the optimal current patterns through iterative physical measurements are developed. Direct methods to first determine the resistance matrix of a resistivity distribution through a set of current bases is injected and the measured voltage responses are stored. This permits iterative reconstruction techniques to operate on the stored data without requiring lengthy data taking from the object and reduces the effects of motion artifacts. The direct methods have superior performance as compared to the iterative methods in both optimal current and voltage generation. The results obtained with three sets of current bases are studied.

A robust image reconstruction algorithm and its parallel implementation in electrical impedance tomography.

An efficient and robust image reconstruction algorithm for static impedance imaging using Hachtel's augmented matrix method was developed. This improved Newton-Raphson method produced more accurate images by reducing the undesirable effects of the ill-conditioned Hessian matrix. It is demonstrated that the electrical impedance tomography (EIT) system could produce two-dimensional static images from a physical phantom with 7% spatial resolution at the center and 5% at the periphery. Static EIT image reconstruction requires a large amount of computation. In order to overcome the limitations on reducing the computation time by algorithmic approaches, the improved Newton-Raphson algorithm was implemented on a parallel computer system. It is shown that the parallel computation could reduce the computation time from hours to minutes.

Iterative reconstruction methods using regularization and optimal current patterns in electrical impedance tomography.

An iterative reconstruction method which minimizes the effects of ill-conditioning is discussed. Based on the modified Newton-Raphson algorithm, a regularization method which integrates prior information into the image reconstruction was developed. This improves the conditioning of the information matrix in the modified Newton-Raphson algorithm. Optimal current patterns were used to obtain voltages with maximal signal-to-noise ratio (SNR). A complete finite element model (FEM) was used for both the internal and the boundary electric fields. Reconstructed images from phantom data show that the use of regularization optimal current patterns, and a complete FEM model improves image accuracy. The authors also investigated factors affecting the image quality of the iterative algorithm such as the initial guess, image iteration, and optimal current updating.

Walsh function current patterns and data synthesis for electrical impedance tomography.

A data collection method which uses Walsh functions as injection current patterns is presented. This method can satisfy two conditions: the optimality of current patterns in every iteration and the single-time data measurement condition. The use of Walsh functions simplifies the design of current sources since only two levels of current (+1 and -1) are required, whereas sinusoidal injection requires a digital-to-analog converter to produce many different values of currents. Compared to diagonal or neighboring type of pulses as injection current patterns, Walsh injection current patterns provide more information about the interior of the subject since Walsh function simulate low and high spatial frequency patterns. Therefore, Walsh function injection uses the simplicity of pulse type injection and yields the better distinguishability or SNR of sinusoidal injection.

Using compound electrodes in electrical impedance tomography.

In electrical impedance tomography, we inject currents and measure voltages to estimate an object's resistivity distribution. The electrode configuration affects measured voltage data because the electrode-skin contact impedance is high and varies with electrode location. We developed a compound electrode which is composed of two electrodes: a large outer electrode to inject current and a small inner electrode to sense voltage. We used these compound electrodes to measure voltages from a physical phantom. We showed that the measured voltages from the compound electrodes are smaller in amplitude than those from conventional electrodes. This demonstrates that the compound electrode can minimize contact impedance voltage drop from the measured data. We used a finite element model for the compound electrode and incorporated the model into the regularized Newton-Raphson reconstruction algorithm. We performed a sensitivity study and showed that the reconstructed resistivity distributions are less dependent on the unknown contact resistance values for a compound electrode than a conventional electrode and that the use of a compound electrode results in improved images for the reconstruction algorithm.

Finite element modeling of electrode-skin contact impedance in electrical impedance tomography.

In electrical impedance tomography (EIT), we inject currents through and measure voltages from an array of surface electrodes. The measured voltages are sensitive to electrode-skin contact impedance because the contact impedance and the current density through this contact impedance are both high. We used large electrodes to provide a more uniform current distribution and reduce the contact impedance. A large electrode differs from a point electrode in that it has shunting and edge effects which cannot be modeled by a single resistor. We used the finite element method (FEM) to study the electric field distributions underneath an electrode, and developed three models: a FEM model, a simplified FEM model and a weighted load model. We showed that the FEM models considered both shunting and edge effects and matched closely the experimental measurements. FEM models for electrodes can be used to improve the performance of an electrical impedance tomography reconstruction algorithm.

Measuring lung resistivity using electrical impedance tomography.

We propose the use of electrical impedance tomography (EIT) imaging techniques in the measurement of lung resistivity for detection and monitoring of apnea and edema. In EIT, we inject currents into a subject using multiple electrodes and measure boundary voltages to reconstruct a cross-sectional image of internal resistivity distribution. We found that a simplified, therefore fast, version of the impedance imaging method can be used for detection and monitoring of apnea and edema. We have showed the feasibility of this method through computer simulations and human experiments. We speculate that the EIT imaging technique will be more reliable than the current impedance apnea monitoring method, since we are monitoring the change of internal lung resistivity. However, more study is required to verify that this method performs better in the presence of motion artifact than the conventional two-electrode impedance apnea monitoring method. Future work should include experiments which carefully simulate different kinds of motion artifacts.

Attachment kinetics of Vero cells onto CT-3 microcarriers.

The effects of cell seeding density, microcarrier concentration, agitation speed and age of seeding cells on the rate of cell attachment to the surface of CT-3 microcarriers were investigated. It was shown that the attachment followed first-order kinetics. When either the cell seeding density or the microcarrier concentration was increased, the kinetic constant increased due to an increase in the probability of collision between cells and microcarriers. However, at higher microcarrier concentrations, the increase in the kinetic constant with increasing microcarrier concentration was not significant. Cell attachment was decelerated upon increasing the agitation speed because of the shorter cell-microcarrier contact time. In addition, it was also demonstrated that cell attachment occurred more efficiently when seeding cells from the middle or early exponential growth phase were used. The process of cell attachment onto CT-3 microcarriers was investigated, and our results showed that the attachment stage was the rate-limiting step. These results will facilitate the optimization of the Vero cell culture process.

Finite-element method in electrical impedance tomography.

In electrical impedance tomography (EIT), current patterns are injected into a subject and boundary voltages are measured to reconstruct a cross-sectional image of resistivity distribution. Static EIT image reconstruction requires a computer model of a subject, an efficient data-collection method and robust and fast reconstruction algorithms. The finite-element method is used as the computer model. The paper describes the finite-element analysis software package developed, including an interactive graphical mesh generator and fast algorithms for solving linear systems of equations using sparse-matrix and vector techniques. Various models of irregularly shaped subjects are developed using mesh-design tools, including automatic mesh generation and optimisation using the Delaunay algorithm. Even though the software package is customised for use in electrical impedance tomography, it can be used for other biomedical research areas, such as impedance cardiography, cardiac defibrillation and impedance pneumography.

Skin impedance measurements using simple and compound electrodes.

We have studied the effect of the electrode configuration on the measurement of body impedance and found that the electrode configuration greatly affects the impedance measurement using the four-electrode method. We studied the characteristics of the compound electrode and found that the compound electrode provides the four-electrode method in a compact form. A new method of measuring the skin impedance using simple electrodes at low frequencies was developed. At high frequencies where the effect of internal tissue impedance is not negligible, we used the compensation method using compound electrodes, because they measure the voltage right under the skin. At 50 kHz, we measured the real part of the skin impedance of less than 80 omega on the thorax. We propose a simple instrument which can measure accurate skin impedance at various frequencies.