Publications by authors named "Pierre Moens"

46 Publications

Evaluation of autofluorescence quenching techniques on formalin- fixed chicken tissues.

J Immunol Methods 2021 Jul 2;496:113097. Epub 2021 Jul 2.

Animal Science, School of Environmental and Rural Science, University of New England, Armidale, Australia. Electronic address:

Autofluorescence (AF) in formalin-fixed and paraffin-embedded tissues limit their use in immunofluorescence staining techniques. Various methods have been used to reduce AF in human and animal tissues but no protocol has been optimized for avian tissues. The present study was undertaken to evaluate different treatment methods including ammonium chloride, glycine, Trypan blue, sodium borohydride, Sudan Black B, potassium permanganate, LED light, cupric sulphate combined with glycine, ammonium chloride and cupric sulphate in reducing AF in FFPE chicken tissues for the detection of FITC labelled antibodies against immune cell markers. Chicken tissues including conjunctiva, trachea and Harderian gland presented intense non-homogenous AF in cells resembling erythrocytes, connective cells and melanocytes. Only Sudan Black B effectively reduced AF in FFPE tissues; however, no specific fluorescent signal was observed for six FITC labelled antibodies against immune cell markers. Specific fluorescent signal from the FITC-labelled antibodies was observed in frozen chicken tissue sections with minimal AF, suggesting that the AF in FFPE tissues is related to the use of formaldehyde fixatives. In conclusion, this study demonstrates for the first time that AF quenching methods commonly used for other animal species are not appropriate for use in avian tissues and that frozen tissue sections are recommended for immunofluorescence staining techniques in poultry.
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http://dx.doi.org/10.1016/j.jim.2021.113097DOI Listing
July 2021

Intra operative assessment of the coronal balance in spinal deformity surgery : a technical note and retrospective study.

Acta Orthop Belg 2021 Mar;87(1):175-179

Obtaining a spine that is well balanced after fusion for scoliotic deformity is primordial for the patients' quality of life. A simple T-shaped instrument combined with standard intraoperative fluoroscopy can be of great help to evaluate the coronal alignment quickly. The aim of this study was to evaluate if a T-shaped device could predict the postoperative coronal balance. Before finalization of the rod fixation, the balance was checked by verifying the relationship between the T-shaped instrument and the upper instrumented vertebra (UIV), and final adjustments were made to correct the coronal balance. A retrospective study was conducted on 48 patients who underwent surgery to correct scoliotic deformity. Intraoperative and postoperative coronal alignment was measured independently by two observers. The mean intraoperative horizontal offset measured between T-shaped instrument and the center of the UIV was 1,69mm to the right with a standard deviation (SD) of 12,43 mm. On postoperative full spine radiographs, the mean offset between the centra sacral vertical line and the center of the UIV was 2,44mm to the left with a SD of 13,10mm. There is no significant difference in coronal balance between both measurements (p=0,12). With this technique we were able to predict the postoperative coronal balance in all but one patient (97,92%). We conclude that the use of a simple T-shaped instrument can provide adequate intraoperative assessment of coronal balance in correcting scoliotic deformity. Level of evidence : IV - case series.
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March 2021

Effect of methionine feeding on oxidative stress, intracellular calcium and contractility in cardiomyocytes isolated from male and female rats.

Mol Cell Biochem 2021 May 30;476(5):2039-2045. Epub 2021 Jan 30.

School of Science and Technology, University of New England, Armidale, NSW, 2351, Australia.

Homocysteine (Hcy) is a breakdown product of methionine metabolism. The risk of cardiovascular disease (CVD) correlates with an increase in plasma Hcy levels. The aim of this study was to investigate whether 1% methionine supplementation of adult rats altered intracellular reactive oxygen species (ROS) generation, intracellular Ca content, and contractile activity in freshly isolated cardiomyocytes. This was measured under normal conditions and during oxidative stress in freshly isolated cardiomyocytes. Single rat cardiomyocytes from both sexes were isolated by enzymatic and mechanical dispersion techniques. Fluorescence microscopy was used to measure ROS production and intracellular Ca concentration. Cell contraction was measured using a video camera. During exposure to 200 μM, HO female cardiomyocytes produced significantly fewer ROS and had a higher intracellular Ca concentration compared to male cardiomyocytes in control and methionine-fed conditions. The contractility of cardiomyocytes isolated from male rats was insignificantly decreased after methionine feeding compared to control, while the contractility of cardiomyocytes from female rats insignificantly reduced after methionine feeding and acute exposure to oxidative stress. These findings provide evidence that during exposure to 200 μM HO, cardiomyocytes from female rats produce less ROS and have higher intracellular Ca levels. There were no significant effects on contractility in cardiomyocytes from either gender and under any of the different conditions.
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http://dx.doi.org/10.1007/s11010-020-04011-2DOI Listing
May 2021

The prevalence of scoliosis within Belgian myelomeningocele population and the correlation with ambulatory status and neurological comorbidities: a chart audit.

Spinal Cord 2021 Jan 25. Epub 2021 Jan 25.

Physical and Rehabilitation Medicine, University Hospitals Leuven, Leuven, Belgium.

Study Design: Retrospective chart audit.

Objectives: Firstly determining the prevalence of scoliosis in myelomeningocele (MMC) patients of the University Hospitals Leuven. Secondly analyzing whether there are differences concerning distribution of radiological level, ambulatory status, hydrocephalus, tethered cord, and syringomyelia in MMC patients with/without scoliosis.

Setting: University Hospitals Leuven, spina bifida convention.

Methods: The following data were collected: age, gender, radiograph type, age at the time of the radiograph, position during radiograph, presence of fusion, age at the time of fusion, diagnosis of hydrocephalus, tethered cord, or syringomyelia, radiological level of MMC, ambulatory status, main Cobb angle, main curve convexity, and main curve location. Correlation between prevalence of scoliosis and ambulatory status, neurological comorbidities, and radiological level were investigated.

Results: There were 116 patients remaining, after excluding patients without MMC or useful images. The scoliosis prevalence in MMC patients was 78.4% (95% CI, 71.0-85.8) for Cobb angle ≥10°; 60.3% (95% CI, 51.4-69.2) for ≥20°, 52.6% (95% CI, 43.5-61.7) for ≥30°, and 36.6% (95% CI, 27.7-45.5) for an angle ≥40°. Wheelchair users had 4 to 8 times more chance of having scoliosis than patients able to walk on all surfaces without aid. Thoracolumbar and lumbar radiological levels had a slightly higher prevalence of scoliosis than sacral levels.

Conclusions: The high prevalence of scoliosis warrants a thorough screening and follow-up for MMC. There was no statistically significant difference between hydrocephalus, tethered cord, or syringomyelia regarding scoliosis. Future studies should focus on the interactions of the neurological comorbidities associated with MMC and scoliosis.
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http://dx.doi.org/10.1038/s41393-020-00611-3DOI Listing
January 2021

Effect of ageing and hypertension on the expression and activity of PEPT2 in normal and hypertrophic hearts.

Amino Acids 2021 Feb 6;53(2):183-193. Epub 2021 Jan 6.

School of Science and Technology, University of New England, Armidale, NSW, 2351, Australia.

Some dipeptides have been implicated in myocardial protection, but little is known about their membrane transporter PEPT2. The aim of this study was to determine whether the expression and activity of the cardiac-type PEPT2 cotransporter could be affected by ageing and/or hypertension. Sarcolemmal vesicles (SV) were isolated from the hearts of all rat groups using a standard procedure to investigate the transport activity and protein abundance by fluorescence spectroscopy and Western blot, respectively. SLC15A2 "PEPT2" gene expression was relatively quantified by RT-qPCR. In the Wistar rat groups, the protein and gene expression of PEPT2 were upregulated with ageing. These changes were accompanied by corresponding increases in the competitive inhibition and the transport rate (V) of β-Ala-Lys (AMCA) into SV isolated from middle-aged hearts. Although, the transport rate of β-Ala-Lys (AMCA) into SV isolated from old hearts was significantly the lowest compared to middle-aged and young adult hearts, the inhibition percentage of β-Ala-Lys (AMCA) transport by Gly-Gln was the highest. In the WKY and SHR rat groups, Y-SHR hypertrophied hearts showed an increase in PEPT2 gene expression accompanied by a significant decrease in protein expression and activity. With advanced age, however, M-SHR hypertrophied hearts revealed significantly lower gene expression, but higher protein expression and activity than Y-SHR hearts. These findings suggest that increased expression of PEPT2 cotransporter in all types of middle-aged hearts could be exploited to facilitate di-and tripeptide transport by PEPT2 in these hearts, which subsequently could result in improved myocardial protection in these populations.
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http://dx.doi.org/10.1007/s00726-020-02936-yDOI Listing
February 2021

Profilin: many facets of a small protein.

Biophys Rev 2020 Aug 13;12(4):827-849. Epub 2020 Jul 13.

Centre for Bioactive Discovery in Health and Ageing, School of Science and Technology, University of New England, Armidale, Australia.

Profilin is a ubiquitously expressed protein well known as a key regulator of actin polymerisation. The actin cytoskeleton is involved in almost all cellular processes including motility, endocytosis, metabolism, signal transduction and gene transcription. Hence, profilin's role in the cell goes beyond its direct and essential function in regulating actin dynamics. This review will focus on the interactions of Profilin 1 and its ligands at the plasma membrane, in the cytoplasm and the nucleus of the cells and the regulation of profilin activity within those cell compartments. We will discuss the interactions of profilin in cell signalling pathways and highlight the importance of the cell context in the multiple functions that this small essential protein has in conjunction with its role in cytoskeletal organisation and dynamics. We will review some of the mechanisms that control profilin expression and the implications of changed expression of profilin in the light of cancer biology and other pathologies.
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http://dx.doi.org/10.1007/s12551-020-00723-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7429631PMC
August 2020

The Transverse Gravitational Deviation Index, a Novel Gravity Line-Related Spinal Parameter, Relates to Balance Control and Health-Related Quality of Life in Adults With Spinal Deformity.

Spine (Phila Pa 1976) 2020 Jan;45(1):E25-E36

Department of Development and Regeneration, Institute for Orthopaedic Research and Training (IORT), KU Leuven, Belgium.

Study Design: Prospective cross-sectional case-control study design.

Objective: This study aims to analyze the relation between balance control as well as health-related quality of life (HRQOL) in patients with adult spinal deformity (ASD), with a novel gravity line (GL)-related 3D spinal alignment parameter, the transverse gravitational deviation index (TGDI), defined to quantify the transverse plane position of any vertebra with respect to the GL.

Summary Of Background Data: Demographic data and balance control have both been identified as important determinants of HRQOL in ASD patients during a preoperative setting. Therefore, a better understanding of the relation between spinal alignment and balance is required.

Methods: After informed consent, 15 asymptomatic healthy volunteers (mean age 60.1 ± 11.6 years old) and 55 ASD patients (mean age 63.5 ± 10.1 years old) were included. Relation between performance on BESTest as well as core outcome measures index (COMI) with spinopelvic alignment was explored using General Linear Modeling (GLM). A P-value ≤0.05 was considered statistically significant.

Results: The L3 TGDI was identified to relate to balance control in the total ASD population after correction for confounding demographic factors (P = 0.001; adjusted R = 0.500) and explained 19% of the observed variance in balance performance. In addition, COMI is related to L3 TGDI in a subgroup of ASD patients with combined coronal and sagittal malalignment of L3 (P = 0.027; slope B = 0.047), despite significant influence of age (P = 0.020).

Conclusion: In ASD patients with a combined coronal and sagittal malalignment of the L3 vertebra, both the level of balance impairment as well as HRQOL are related to the distance component of the L3 TGDI, that is, the offset between the center of the L3 vertebral body and the GL in the transverse plane.

Level Of Evidence: 2.
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http://dx.doi.org/10.1097/BRS.0000000000003301DOI Listing
January 2020

Degenerative changes of the hip following fixation for slipped capital femoral epiphysis: a minimum 18-year follow-up study.

Hip Int 2021 Mar 4;31(2):264-271. Epub 2019 Aug 4.

Orthopaedic Department, University Hospitals Leuven - Pellenberg, Belgium.

Introduction: fixation (ISF) is currently still the 'gold standard' treatment for slipped capital femoral epiphysis (SCFE) and has shown acceptable results at mid-term follow-up. This study aims to evaluate functional, clinical and radiographic long-term outcomes after this procedure.

Methods: We reviewed 64 SCFE patients (76 SCFE hips) treated with ISF between 1983 and 1998. 82.9% were stable hips and 17.1% unstable according to Loder's definition. Initial radiographs demonstrated a mild slip in 50%, moderate in 41.3% and severe in 8.7% based on the Southwick angle. Long-term outcomes were assessed using the modified Harris Hip Score (mHHS), University of California at Los Angeles (UCLA) and Tegner activity scores, visual analogue scale (VAS) pain, VAS function, flexion-adduction-internal rotation (FADIR) test, extent of internal-rotation at 90° of hip flexion and Tönnis classification for hip osteoarthritis (OA).

Results: 10 (15.6%) SCFE hips were converted to a total hip replacement (THR) after a mean of 16 years. 38 (59.4%) patients underwent a clinical and radiographic examination after a mean follow-up of 23 (range 18-33) years. 12 (18.8%) patients were lost to follow-up. 74% of SCFE hips demonstrated degenerative change on radiography or were converted to THR (Tönnis 1: 33.3%, 2: 18.5%, 3 or THR: 22.2%). There were 3 cases of avascular necrosis (AVN) all in unstable hips. Mean mHHS was 86.8/100, UCLA activity score 7.5/10, Tegner activity score 3.8/10, VAS pain 1.7/10 and VAS function 1.5/10. 20% of SCFE hips were found to have a positive FADIR-test and a limited internal-rotation of 19.7° versus 36.1° ( < 0.001) in contralateral normal hips.

Discussion: This long-term follow-up study of ISF for SCFE shows that although complication rates in terms of AVN are low, a high number of patients become symptomatic and have a limited function. Degenerative changes are common with 22.2% of hips developing end-stage hip OA (Tönnis 3 or THR). It is important that patients and parents are informed about these risks.
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http://dx.doi.org/10.1177/1120700019867248DOI Listing
March 2021

Cofilin and profilin: partners in cancer aggressiveness.

Biophys Rev 2018 Oct 19;10(5):1323-1335. Epub 2018 Jul 19.

Centre for Bioactive Discovery in Health and Ageing, School of Science and Technology, University of New England, Armidale, Australia.

This review covers aspects of cofilin and profilin regulations and their influence on actin polymerisation responsible for cell motility and metastasis. The regulation of their activity by phosphorylation and nitration, miRs, PI(4,5)P binding, pH, oxidative stress and post-translational modification is described. In this review, we have highlighted selected similarities, complementarities and differences between the two proteins and how their interplay affects actin filament dynamics.
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http://dx.doi.org/10.1007/s12551-018-0445-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6233335PMC
October 2018

Molecular changes to the rat renal cotransporters PEPT1 and PEPT2 due to ageing.

Mol Cell Biochem 2019 Feb 17;452(1-2):71-82. Epub 2018 Jul 17.

School of Science and Technology, University of New England, Armidale, NSW, 2351, Australia.

Renal PEPT1 and PEPT2 cotransporters play an important role in the balance of circulating body oligopeptides and selected peptidomimetic drugs. We aim to comprehensively characterise age-related changes of the renal PEPT cotransporters at the gene, protein, and functional level. Brush border membrane vesicles (BBMV) and outer medulla membrane vesicles (OMMV) were isolated from the kidneys of young, middle-aged and old rats. The protein expression of PEPT1 was not only increased in BBMV from old rats, but PEPT1 also appeared in OMMV from middle-aged and old rats. SLC15A1 gene expression in the renal cortex increased in middle-aged group. PEPT2 protein expression was not only increased with ageing, but PEPT2 also was found in BBMV from middle-aged and old groups. SLC15A2 gene expression in the renal outer medulla increased in the old group. These changes in the expressions and localisations of PEPT1 and PEPT2 could explain the changes to transport activity in BBMV and OMMV. These findings provide novel insights that would be useful for maintaining protein nutrition and optimising the delivery of some peptidomimetic drugs in elderly individuals.
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http://dx.doi.org/10.1007/s11010-018-3413-xDOI Listing
February 2019

The Prevalence of Scoliosis in Spina Bifida Subpopulations: A Systematic Review.

Am J Phys Med Rehabil 2018 11;97(11):848-854

From Physical and Rehabilitation Medicine, University Hospitals Leuven, Leuven, Belgium (AH, KP, CK); Clinical and Experimental Sciences Department, University of Brescia, Brescia, Italy (SN); IRCCS Fondazione Don Carlo Gnocchi ONLUS, Milan, Italy (SN); Department of Development and Regeneration, University Hospitals Leuven, Leuven, Belgium (KJ); and Orthopaedic Surgery, University Hospitals Leuven, Leuven, Belgium (PM, SS).

Prevalence of scoliosis within spina bifida subpopulations is important for diagnostics and therapeutic purposes. This review determined the prevalence of scoliosis within spina bifida subpopulations by means of a systematic literature review by using the following databases: Medline PubMed, Embase, Cochrane, and Pedro. All Dutch- and English-written literature using the MESH terms "spinal dysraphism," "neural tube defects," and "scoliosis" was analyzed using the exclusion criteria: animal studies, case reports, studies regarding the prevalence of spina bifida among patients with scoliosis, studies with inclusion of patients with scoliosis of less than 11 degrees without possibility to identify subgroups with scoliosis of greater than 10 degrees, studies without an own study group, articles comprising the same patient group as another article, neural tube defects besides spina bifida, and articles without specification of spina bifida subtype. It resulted in six articles, two concerning diastematomyelia (103 patients, 82 females and 21 males) and four about myelomeningocele (479 patients, 283 females and 196 males) with an overall weighted prevalence of scoliosis (20-degree Cobb angle cutoff) of 44.4% and 52.5%, respectively. It can be concluded that most studies have a lot of methodological flaws, so there is a need for further research with standardization of data collection to allow comparison of different data.
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http://dx.doi.org/10.1097/PHM.0000000000000966DOI Listing
November 2018

1D Self-Assembly and Ice Recrystallization Inhibition Activity of Antifreeze Glycopeptide-Functionalized Perylene Bisimides.

Chemistry 2018 Jun 8;24(31):7834-7839. Epub 2018 May 8.

School of Science and Technology, University of New England, Armidale, 2351, Australia.

Antifreeze glycoproteins (AFGPs) are polymeric natural products that have drawn considerable interest in diverse research fields owing to their potent ice recrystallization inhibition (IRI) activity. Self-assembled materials have emerged as a promising class of biomimetic ice growth inhibitor, yet the development of AFGP-based supramolecular materials that emulate the aggregative behavior of AFGPs have not yet been reported. This work reports the first example of the 1D self-assembly and IRI activity of AFGP-functionalized perylene bisimides (AFGP-PBIs). Glycopeptide-functionalized PBIs underwent 1D self-assembly in water and showed modest IRI activity, which could be tuned through substitution of the PBI core. This work presents essential proof-of-principle for the development of novel IRIs as potential supramolecular cryoprotectants and glycoprotein mimics.
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http://dx.doi.org/10.1002/chem.201800857DOI Listing
June 2018

A new use of β-Ala-Lys (AMCA) as a transport reporter for PEPT1 and PEPT2 in renal brush border membrane vesicles from the outer cortex and outer medulla.

Biochim Biophys Acta Biomembr 2018 May 30;1860(5):960-964. Epub 2017 Dec 30.

School of Science and Technology, University of New England Armidale, NSW 2351, Australia.

Integral membrane proteins PEPT1 and PEPT2 are essential for reabsorbing almost all hydrolysed or filtered di- and tripeptides alongside a wide range of peptidomimetic drugs in the kidney. The aim of this study was to investigate the potential use of the fluorophore-conjugated dipeptide β-Ala-Lys (AMCA) as a biosensor for measuring peptide transport activity in brush border membrane vesicles isolated from the outer cortex (BBMV-OC) and outer medulla (BBMV-OM) (representing PEPT1 and PEPT2 respectively). The vesicles were isolated using a dual magnesium precipitation and centrifugation technique. Intravesicular fluorescence accumulation was measured after incubating extra-vesicular media at pH6.6 and different concentrations of β-Ala-Lys (AMCA) with vesicles pre-equilibrated at pH7.4. Both BBMV-OC and BMMV-OM showed accumulation of an intravesicular fluorescence signal after 20min incubation. Changing the extra-vesicular pH to 7.4 caused a significant reduction in the β-Ala-Lys (AMCA) uptake into BBMV-OC at concentrations >100μM. When different concentrations of dipeptide, Gly-Gln was added, there was a significant inhibition of 100μM β-Ala-Lys (AMCA) uptake into BBMV-OC and BMMV-OM, reaching 69% and 80%, respectively. Kinetic analysis of β-Ala-Lys (AMCA) at 20min showed that the K and V were 783.7±115.7μM and 2191.2±133.9ΔF/min/mg for BBMV-OC, while BMMV-OM showed significantly higher affinity, but lower capacity at K=93.6±21.9μM and V=935.8±50.2ΔF/min/mg. These findings demonstrate the applicability of β-Ala-Lys (AMCA) as a biosensor to measure the transport activity of the renal-type PEPT1 and PEPT2 in BBMV-OC and BMMV-OM respectively.
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http://dx.doi.org/10.1016/j.bbamem.2017.12.021DOI Listing
May 2018

Quantitative image mean squared displacement (iMSD) analysis of the dynamics of profilin 1 at the membrane of live cells.

Methods 2018 05 11;140-141:119-125. Epub 2017 Dec 11.

Centre for Bioactive Discovery in Health and Ageing, School of Science and Technology, University of New England, Armidale, Australia. Electronic address:

Image mean square displacement analysis (iMSD) is a method allowing the mapping of diffusion dynamics of molecules in living cells. However, it can also be used to obtain quantitative information on the diffusion processes of fluorescently labelled molecules and how their diffusion dynamics change when the cell environment is modified. In this paper, we describe the use of iMSD to obtain quantitative data of the diffusion dynamics of a small cytoskeletal protein, profilin 1 (pfn1), at the membrane of live cells and how its diffusion is perturbed when the cells are treated with Cytochalasin D and/or the interactions of pfn1 are modified when its actin and polyphosphoinositide binding sites are mutated (pfn1-R88A). Using total internal reflection fluorescence microscopy images, we obtained data on isotropic and confined diffusion coefficients, the proportion of cell areas where isotropic diffusion is the major diffusion mode compared to the confined diffusion mode, the size of the confinement zones and the size of the domains of dynamic partitioning of pfn1. Using these quantitative data, we could demonstrate a decreased isotropic diffusion coefficient for the cells treated with Cytochalasin D and for the pfn1-R88A mutant. We could also see changes in the modes of diffusion between the different conditions and changes in the size of the zones of pfn1 confinements for the pfn1 treated with Cytochalasin D. All of this information was acquired in only a few minutes of imaging per cell and without the need to record thousands of single molecule trajectories.
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http://dx.doi.org/10.1016/j.ymeth.2017.12.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5995621PMC
May 2018

Kinetic Measurements of Di- and Tripeptide and Peptidomimetic Drug Transport in Different Kidney Regions Using the Fluorescent Membrane Potential-Sensitive Dye, DiS-C-(3).

J Membr Biol 2017 Dec 7;250(6):641-649. Epub 2017 Oct 7.

School of Science and Technology, University of New England, Armidale, NSW, 2351, Australia.

Tri- and dipeptides are transported in the kidney by PEPT1 and PEPT2 isoforms. The aim of this study was to investigate differences in transport kinetics between renal brush border (BBMV) and outer medulla (OMMV) membrane vesicles (where PEPT1 and PEPT2 are sequentially available) for a range of di- and tripeptides and peptidomimetic drugs. This was accomplished through the use of the potential-sensitive fluorescent dye 3,3'-dipropylthiacarbocyanine iodide [DiS-C-(3)]. BBMV and OMMV were prepared from the rat kidney using standard techniques. The presence of PEPT1 in BBMV and PEPT2 in OMMV was confirmed using Western blotting. Fluorescence changes were measured when extravesicular medium at pH 6.6 containing 0-1 mM substrates was added to a cuvette containing vesicles pre-equilibrated at pH 7.4 and 2.71 μM DiS-C-(3). An increase in fluorescence intensity occurred upon substrate addition reflecting the expected positive change in membrane potential difference. Of the range of substrates studied, OMMV manifested the highest affinity to cefadroxil and valacyclovir (K 4.3 ± 1.2 and 11.7 ± 3.2 µM, respectively) compared to other substrates, whilst the BBMV showed a higher affinity to Gly-His (K 15.4 ± 3.1 µM) compared to other substrates. In addition, OMMV showed higher affinity and capacity to Gly-Gln (K 47.1 ± 9.8 µM, 55.5 ± 2.8 ΔF/s/mg protein) than BBMV (K 78.1 ± 13.3 µM and 35.5 ± 1.7 ΔF/s/mg protein, respectively). In conclusion, this study successfully separated the expression of PEPT1 and PEPT2 into different vesicle preparations inferring their activity in different regions of the renal proximal tubule.
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http://dx.doi.org/10.1007/s00232-017-9990-xDOI Listing
December 2017

Proteomic and Microscopic Strategies towards the Analysis of the Cytoskeletal Networks in Major Neuropsychiatric Disorders.

Int J Mol Sci 2016 Apr 20;17(4). Epub 2016 Apr 20.

Center for Bioactive Discovery in Health and Aging, School of Science and Technology, University of New England, Armidale, NSW 2351, Australia.

Mental health disorders have become worldwide health priorities. It is estimated that in the next 20 years they will account for a 16 trillion United State dollars (US$) loss. Up to now, the underlying pathophysiology of psychiatric disorders remains elusive. Altered cytoskeleton proteins expression that may influence the assembly, organization and maintenance of cytoskeletal integrity has been reported in major depressive disorders, schizophrenia and to some extent bipolar disorders. The use of quantitative proteomics, dynamic microscopy and super-resolution microscopy to investigate disease-specific protein signatures holds great promise to improve our understanding of these disorders. In this review, we present the currently available quantitative proteomic approaches use in neurology, gel-based, stable isotope-labelling and label-free methodologies and evaluate their strengths and limitations. We also reported on enrichment/subfractionation methods that target the cytoskeleton associated proteins and discuss the need of alternative methods for further characterization of the neurocytoskeletal proteome. Finally, we present live cell imaging approaches and emerging dynamic microscopy technology that will provide the tools necessary to investigate protein interactions and their dynamics in the whole cells. While these areas of research are still in their infancy, they offer huge potential towards the understanding of the neuronal network stability and its modification across neuropsychiatric disorders.
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http://dx.doi.org/10.3390/ijms17040581DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4849037PMC
April 2016

Profilin-1 mediated cell-cycle arrest: searching for drug targets.

Cell Cycle 2015 27;14(23):3669-70. Epub 2015 Aug 27.

b School of Rural Medicine ; University of New England ; Armidale , NSW Australia.

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http://dx.doi.org/10.1080/15384101.2015.1086204DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4825793PMC
August 2016

Modes of diffusion of cholera toxin bound to GM1 on live cell membrane by image mean square displacement analysis.

Biophys J 2015 Mar;108(6):1448-1458

Centre for Bioactive Discovery in Health and Ageing, School of Science and Technology, University of New England, Armidale, Australia; Laboratory for Fluorescence Dynamics, Department of Biomedical Engineering, University of California, Irvine, Irvine, California.

The image-mean square displacement technique applies the calculation of the mean square displacement commonly used in single-molecule tracking to images without resolving single particles. The image-mean square displacement plot obtained is similar to the mean square displacement plot obtained using the single-particle tracking technique. This plot is then used to reconstruct the protein diffusion law and to identify whether the labeled molecules are undergoing pure isotropic, restricted, corralled, transiently confined, or directed diffusion. In our study total internal reflection fluorescence microscopy images were taken of Cholera toxin subunit B (CtxB) membrane-labeled NIH 3T3 mouse fibroblasts and MDA 231 MB cells. We found a population of CTxB undergoing purely isotropic diffusion and one displaying restricted diffusion with corral sizes ranging from 150 to ∼1800 nm. We show that the diffusion rate of CTxB bound to GM1 is independent of the size of the confinement, suggesting that the mechanism of confinement is different from the mechanism controlling the diffusion rate of CtxB. We highlight the potential effect of continuous illumination on the diffusion mode of CTxB. We also show that aggregation of CTxB/GM1 in large complexes occurs and that these aggregates tend to have slower diffusion rates.
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http://dx.doi.org/10.1016/j.bpj.2015.02.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4375681PMC
March 2015

Profilin-1 overexpression in MDA-MB-231 breast cancer cells is associated with alterations in proteomics biomarkers of cell proliferation, survival, and motility as revealed by global proteomics analyses.

OMICS 2014 Dec;18(12):778-91

1 School of Science and Technology, University of New England , Armidale, NSW, Australia .

Despite early screening programs and new therapeutic strategies, metastatic breast cancer is still the leading cause of cancer death in women in industrialized countries and regions. There is a need for novel biomarkers of susceptibility, progression, and therapeutic response. Global analyses or systems science approaches with omics technologies offer concrete ways forward in biomarker discovery for breast cancer. Previous studies have shown that expression of profilin-1 (PFN1), a ubiquitously expressed actin-binding protein, is downregulated in invasive and metastatic breast cancer. It has also been reported that PFN1 overexpression can suppress tumorigenic ability and motility/invasiveness of breast cancer cells. To obtain insights into the underlying molecular mechanisms of how elevating PFN1 level induces these phenotypic changes in breast cancer cells, we investigated the alteration in global protein expression profiles of breast cancer cells upon stable overexpression of PFN1 by a combination of three different proteome analysis methods (2-DE, iTRAQ, label-free). Using MDA-MB-231 as a model breast cancer cell line, we provide evidence that PFN1 overexpression is associated with alterations in the expression of proteins that have been functionally linked to cell proliferation (FKPB1A, HDGF, MIF, PRDX1, TXNRD1, LGALS1, STMN1, LASP1, S100A11, S100A6), survival (HSPE1, HSPB1, HSPD1, HSPA5 and PPIA, YWHAZ, CFL1, NME1) and motility (CFL1, CORO1B, PFN2, PLS3, FLNA, FLNB, NME2, ARHGDIB). In view of the pleotropic effects of PFN1 overexpression in breast cancer cells as suggested by these new findings, we propose that PFN1-induced phenotypic changes in cancer cells involve multiple mechanisms. Our data reported here might also offer innovative strategies for identification and validation of novel therapeutic targets and companion diagnostics for persons with, or susceptibility to, breast cancer.
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http://dx.doi.org/10.1089/omi.2014.0075DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4253143PMC
December 2014

Conradi-Hünermann-Happle syndrome: a novel heterozygous missense mutation, c.204G>T (p.W68C).

Pediatr Dermatol 2014 Jul-Aug;31(4):493-6. Epub 2014 Jun 11.

Department of Dermatology, University Hospitals Leuven, Leuven, Belgium.

Conradi-Hünermann-Happle syndrome (X-linked dominant chondrodysplasia punctata, CDPX2 [Online Mendelian Inheritance in Man 302960]) is a rare genodermatosis that presents with blaschkolinear ichthyosis, cicatricial alopecia, chondrodysplasia punctata, asymmetric shortening of the bones, and cataracts. In this case report we describe a child presenting with a patterned alopecia in which supplementary signs and clinical examination of the mother led to the suspicion of Conradi-Hünermann-Happle syndrome. Mutation analysis revealed a heterozygous novel missense mutation, c.204G>T (p.W68C), in exon 2.
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http://dx.doi.org/10.1111/pde.12336DOI Listing
May 2015

Sublaminar devices for the correction of scoliosis: metal wire versus polyester tape.

Acta Orthop Belg 2013 Apr;79(2):216-21

Department of Orthopaedic Surgery, University Hospitals Leuven, Pellenberg, Belgium.

The authors conducted a retrospective study comparing the corrective effect of two sublaminar techniques on scoliosis: the classical one, based on metal wire, and a more recent one, based on polyester tape (thoracic Universal Clamp), known to be safer (less risk of neurological damage, less laminar breakthrough) and compatible with MRI. Lumbar screws were used in both groups. The authors composed two groups of 25 scoliosis patients, matched for gender, age, aetiology, anterior release, number of levels fused, number of infections, major curve and flexibility: there was no significant difference. Only the follow-up period was different: 55 months in the metal wire group, versus 17 months in the polyester tape group (p < 0.001), but this was immaterial because the curves were compared one year after surgery. After one year there was no significant difference between both groups, as to correction in the coronal or in the sagittal plane. This means that the polyester tape technique offers an interesting alternative, given that it yields supplementary advantages, as mentioned above.
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April 2013

The effect of Luque-Galveston fusion on curve, respiratory function and quality of life in Duchenne muscular dystrophy.

Acta Orthop Belg 2011 Oct;77(5):659-65

University Hospital, Leuven, Belgium.

The aim of this retrospective study was to evaluate the long-term effect of the Luque-Galveston spinal fusion in Duchenne muscular dystrophy (DMD) patients. Twenty patients had undergone this operation at a mean age of 153 years (surgical group, A). The correction of their scoliosis amounted to +/- 55.8%, after an average follow-up period of 3 years. This is in accordance with the literature. The authors would therefore advise to perform spinal fusion in an early stage of the disease, once a rapid evolution of the scoliosis is seen. The decline of respiratory function slightly diminished after surgery, but not significantly. This means that no expectations should be made to improve respiratory function, as respiratory function decline continues relentlessly. Most authors agree with this statement. Patient satisfaction after surgery was relatively high, mainly because of an improved sitting balance, but only 60% of the questionnaires were available. Twenty-five other patients were not operated upon (non-surgical group, B). They had better results at ages 153 and 183, but this was mainly due to the fact that group B contained more benign cases according to the Oda classification.
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October 2011

Spondylolisthesis in twins: multifactorial etiology: a case report and review of the literature.

Spine (Phila Pa 1976) 2011 May;36(11):E741-6

Division of Orthopaedics, Department of Musculoskeletal Sciences, University Hospital Pellenberg, Pellenberg, Belgium.

Study Design: Report of a high dysplastic developmental spondylolisthesis in two identical twins of two unrelated families.

Objective: To investigate the multifactorial etiology of developmental spondylolisthesis.

Summary Of Background Data: Multiple studies have suggested an association between a high pelvic incidence and the presence of isthmic spondylolisthesis. Other studies suggest a genetic background for spondylolysis and a pattern of inheritance of susceptibility to spondylolysis and spondylolisthesis. Heterozygous cartilage-derived morphogenetic protein-1 (CDMP-1) mutation has previously been associated with spondylolysis and severe spondylolisthesis.

Methods: Two identical female twins presented with a developmental spondylolisthesis. Pelvic parameters, lumbar lordosis and grade of spondylolisthesis were calculated on a lateral standing spine radiograph. MRI is performed to confirm a high dysplastic developmental spondylolisthesis. Blood sample of these four individuals were analyzed for the presence of a CDMP-1 mutation, a cartilage-specific member of the TGF-b superfamily of secreted signaling molecules that plays a key role in chondrogenesis, growth, and patterning of the developing vertebrate skeleton.

Results: PI, SS, PT, LL, and SI are significantly greater in all of these patients in comparison with the general population. Spinal MRI confirms a high dysplastic developmental spondylolisthesis in both twins. Mutation analysis of the two coding exons of CDMP-1 did not reveal any mutation in all four individuals.

Conclusion: To our knowledge, this is the first report of a high dysplastic developmental spondylolisthesis in identical twins. The presence of a high dysplastic developmental spondylolisthesis in two identical twins shows the convergence in etiology of different factors such as genetics, maturation, critical age, female sex, high pelvic incidence. Although we cannot confirm that CDMP-1 mutation plays a key role in the etiology of spondylolysis/spondylolisthesis, neither can we rule out that CDMP-1 problems may be an etiology for at least a subpopulation of patients. However, the presence of a developmental spondylolisthesis in two sets of identical twins still suggests a genetic susceptibility to spondylolysis and spondylolisthesis.
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http://dx.doi.org/10.1097/BRS.0b013e3181f9f575DOI Listing
May 2011

Fluorescence correlation spectroscopy, raster image correlation spectroscopy, and number and brightness on a commercial confocal laser scanning microscope with analog detectors (Nikon C1).

Microsc Res Tech 2011 Apr 23;74(4):377-88. Epub 2010 Aug 23.

Centre for Bioactive Discovery in Health and Ageing, School of Science and Technology, University of New England, Armidale, Australia.

Fluorescence correlation spectroscopy (FCS) was developed in 1972 by Magde, Elson and Webb. Photon counting detectors and avalanche photodiodes have become standards in FCS to the point that there is a widespread belief that these detectors are essential to perform FCS experiments, despite the fact that FCS was developed using analog detectors. Spatial and temporal intensity fluctuation correlations using analog detection on a commercial Olympus Fluoview 300 microscope have been reported by Brown et al. (2008). However, each analog instrument has its own idiosyncrasies that need to be understood before using the instrument for FCS. In this work, we explore the capabilities of the Nikon C1, a low-cost confocal microscope, to obtain single point FCS, Raster-scan image correlation spectroscopy (RICS), and Number and Brightness data both in solution and incorporated into the membrane of giant unilamellar vesicles. We show that it is possible to obtain dynamic information about fluorescent molecules from single point FCS, RICS, and Number and Brightness using the Nikon C1. We highlighted the fact that care should be taken in selecting the acquisition parameters to avoid possible artifacts due to the detector noise. However, due to relatively large errors in determining the distribution of digital levels for a given microscope setting, the system is probably only adequate for determining relative brightness within the same image.
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http://dx.doi.org/10.1002/jemt.20919DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3001136PMC
April 2011

Plant-extract-induced changes in the proteome of the soil-borne pathogenic fungus Thielaviopsis basicola.

Proteomics 2010 Apr;10(8):1573-91

Molecular and Cellular Biology, School of Science and Technology, University of New England, Armidale, NSW, Australia.

Thielaviopsis basicola is a hemibiotroph fungus that causes black root rot disease in diverse plants with significant impact on cotton production in Australia. To elucidate how T. basicola growth and proteome are influenced by interactions with natural sources, this fungus was cultured in the presence of root extracts from non-host (wheat, hairy vetch) and susceptible host (cotton, lupin) plants. We found that T. basicola growth was significantly favored in the presence of host extracts, while hierarchical clustering analysis of 2-DE protein profiles of T. basicola showed plant species had a larger effect on the proteome than host/non-host status. Analysis by LC-MS/MS of unique and differentially expressed spots and identification using cross-species similarity searching and de novo sequencing allowed successful identification of 41 spots. These proteins were principally involved in primary metabolism with smaller numbers implicated in other diverse functions. Identification of several "morpho" proteins suggested morphological differences that were further microscopically investigated. Identification of several highly expressed spots suggested that vitamin B(6) is important in the T. basicola response to components present in hairy vetch extract, and finally, three spots, induced in the presence of lupin extract, may correspond to malic enzyme and be involved in lipid accumulation.
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http://dx.doi.org/10.1002/pmic.200900301DOI Listing
April 2010

Thoracolumbar intervertebral disc calcifications in an 8-year-old boy: case report and review of the literature.

Eur J Pediatr 2010 May 8;169(5):577-80. Epub 2009 Oct 8.

Department of Musculoskeletal Science, Division of Orthopedics-Pediatric Orthopedics, University Hospital Pellenberg, Weligerveld 1, Pellenberg, Belgium.

An 8-year-old boy presented himself with a nontraumatic acute onset of thoracolumbar pain with an antalgic scoliosis. His past medical history did not reveal any infectious disorder. The back pain kept him from sleep and sports. Imaging revealed calcified intervertebral discs from thoracic intervertebral disc 10-11 to lumbar disc 2-3, with compression of the 11th thoracic root. Conservative treatment with analgesia, nonsteroidal anti-inflammatory drugs, and rest reduced the symptoms within a few days. The boy became asymptomatic within 2 weeks. Magnetic resonance imaging changes diminished in 2 months. Intervertebral disc calcification in children is a rare benign disorder which mostly affects the cervical spine. This is the first case, to our knowledge, of symptomatic intervertebral disc calcification involving the lumbar spine up to lumbar disc 2-3.
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http://dx.doi.org/10.1007/s00431-009-1076-zDOI Listing
May 2010

Structure and functions of profilins.

Biophys Rev 2009 Jul 4;1(2):71-81. Epub 2009 Jun 4.

Centre for Bioactive Discovery in Health and Ageing, School of Science and Technology, University of New England, McClymont Bldg, Armidale, NSW, 2351, Australia.

Profilins are small actin-binding proteins found in eukaryotes and certain viruses that are involved in cell development, cytokinesis, membrane trafficking, and cell motility. Originally identified as an actin sequestering/binding protein, profilin has been involved in actin polymerization dynamics. It catalyzes the exchange of ADP/ATP in actin and increases the rate of polymerization. Profilins also interact with polyphosphoinositides (PPI) and proline-rich domains containing proteins. Through its interaction with PPIs, profilin has been linked to signaling pathways between the cell membrane and the cytoskeleton, while its role in membrane trafficking has been associated with its interaction with proline-rich domain-containing proteins. Depending on the organism, profilin is present in a various number of isoforms. Four isoforms of profilin have been reported in higher organisms, while only one or two isoforms are expressed in single-cell organisms. The affinity of these isoforms for their ligands varies between isoforms and should therefore modulate their functions. However, the significance and the functions of the different isoforms are not yet fully understood. The structures of many profilin isoforms have been solved both in the presence and the absence of actin and poly-L-proline. These structural studies will greatly improve our understanding of the differences and similarities between the different profilins. Structural stability studies of different profilins are also shedding some light on our understanding of the profilin/ligand interactions. Profilin is a multifaceted protein for which a dramatic increase in potential functions has been found in recent years; as such, it has been implicated in a variety of physiological and pathological processes.
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http://dx.doi.org/10.1007/s12551-009-0010-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5425664PMC
July 2009

Profilin interaction with phosphatidylinositol (4,5)-bisphosphate destabilizes the membrane of giant unilamellar vesicles.

Biophys J 2009 Jun;96(12):5112-21

Centre for Bioactive Discovery in Health and Ageing, School of Science and Technology, University of New England, Armidale, Australia.

Profilin, a small cytoskeletal protein, and phosphatidylinositol (4,5)-bisphosphate [PI(4,5)P2] have been implicated in cellular events that alter the cell morphology, such as endocytosis, cell motility, and formation of the cleavage furrow during cytokinesis. Profilin has been shown to interact with PI(4,5)P2, but the role of this interaction is still poorly understood. Using giant unilamellar vesicles (GUVs) as a simple model of the cell membrane, we investigated the interaction between profilin and PI(4,5)P2. A number and brightness analysis demonstrated that in the absence of profilin, molar ratios of PI(4,5)P2 above 4% result in lipid demixing and cluster formations. Furthermore, adding profilin to GUVs made with 1% PI(4,5)P2 leads to the formation of clusters of both profilin and PI(4,5)P2. However, due to the self-quenching of the dipyrrometheneboron difluoride-labeled PI(4,5)P2, we were unable to determine the size of these clusters. Finally, we show that the formation of these clusters results in the destabilization and deformation of the GUV membrane.
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http://dx.doi.org/10.1016/j.bpj.2009.03.034DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2712054PMC
June 2009

Bilateral patellar Köhler's disease in an eleven-year-old child with growth retardation: a case report.

Acta Orthop Belg 2009 Apr;75(2):273-6

Catholic University Hospitals, Leuven, Belgium.

Osteochondrosis of the primary ossification centre of the patella (Köhler's disease) has been reported as a rare cause of anterior knee pain in children between 5 and 9 years of age. The aetiology remains unclear. This is the first case report presenting a late bilateral patellar osteochondrosis with unilateral involvement of the proximal pole in an 11-year old boy with documented growth retardation. This case highlights the importance to thoroughly investigate young patients with patellar osteochondrosis for growth retardation. This case might indicate that growth retardation rather than growth spurt is an important aetiological factor of osteochondrosis.
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April 2009

The McHale procedure in the treatment of the painful chronically dislocated hip in adolescents and adults with cerebral palsy.

Acta Orthop Belg 2009 Apr;75(2):181-8

Department of Children's Orthopaedics, University Hospital Pellenberg, Leuven, Belgium.

We retrospectively studied the results achieved in adolescent and adult patients with cerebral palsy, following management of chronically painful dislocated hips with the McHale procedure. Thirteen patients with seventeen procedures were included in the study. The indication for surgery was pain in all patients. Limitation in sitting was also reported in eight cases and nursing care was difficult in ten of thirteen patients. All patients underwent clinical and radiological examination of the hips, and a questionnaire was answered by both parents and caretakers. Improvement was noted in all patients with respect to pain. In six patients sitting was tolerated for longer intervals and 10 out of 13 patients were easier to nurse owing to improved mobility of the hip. Although these results appear promising, the procedure had a high complication rate, including heterotopic ossification, residual pain caused by hardware, fracture and pulmonary complications, for which a second procedure was necessary in several cases. There was also in several cases a prolonged period of pain postoperatively, for which an additional procedure was needed in eight patients. The McHale procedure is a technique that can provide pain relief and improvement in motion. There is however a high incidence of failure and complications.
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April 2009
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