Roth-Visal Ung

Inflammatory Cytokines and Reactive Oxygen Species as Mediators of Chronic Kidney Disease-Related Vascular Calcification

BACKGROUNDnVascular calcification, a regulated process in chronic kidney disease (CKD), requires vascular smooth muscle cell (VSMC) differentiation into osteoblast-like cells. This phenomenon can be enhanced by inflammatory cytokines and production of reactive oxygen species (ROS). In CKD rats with vascular calcification, we investigated whether inflammatory cytokines, ROS generation, and downstream signaling events are associated with CKD-related vascular calcification.nnnMETHODSnCKD was induced in male Wistar rats by renal mass ablation and vascular calcification was induced with a high calcium-phosphate diet and vitamin D supplementation (Ca/P/VitD). At week 3-6, hemodynamic parameters were determined and thoracic aorta was harvested for assessment of vascular calcification, macrophage infiltration, cytokines expression, VSMC differentiation, ROS generation, and related signaling pathway activation.nnnRESULTSnCKD rats treated with Ca/P/VitD developed medial calcification of thoracic aorta and increased pulse pressure and aortic pulse wave velocity. VSMC differentiation was confirmed by increased bone morphogenetic protein-2 and osteocalcin expression and reduced α-smooth muscle actin expression. The expression of interleukin-1β, interleukin-6, and tumor necrosis factor were also increased. The expression of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunits p22(phox) and p47(phox) were increased, whereas the expression of antioxidant enzymes (SOD1, SOD2, Gpx1, and Prdx1) was reduced in CKD + Ca/P/VitD rats. Oxidized peroxiredoxin, a sensor of ROS generation, was significantly increased and ROS-sensitive signaling pathways were activated in the aorta from CKD + Ca/P/VitD rats.nnnCONCLUSIONnThis study demonstrates a relationship between inflammation/ROS and arterial calcification in CKD and contributes to understanding of the complex pathways that mediate arterial calcification in CKD patients.

Specific role of dopamine D1 receptors in spinal network activation and rhythmic movement induction in vertebrates

Dopamine (DA) is well‐recognized for its determinant role in the modulation of various brain functions. DA was also found in in vitro isolated invertebrate preparations to activate per se the central pattern generator for locomotion. However, it is less clear whether such a role as an activator of central neural circuitries exists in vertebrate species. Here, we studied in vivo the effects induced by selective DA receptor agonists and antagonists on hindlimb movement generation in mice completely spinal cord‐transected (Tx) at the low‐thoracic level (Th9/10). Administration of D1/D5 receptor agonists (0.5–2.5 mg kg−1, i.p.) was found to acutely elicit rhythmic locomotor‐like movements (LMs) and non‐locomotor movements (NLMs) in untrained and non‐sensory stimulated animals. Comparable effects were found in mice lacking the D5 receptor (D5KO) whereas D1/D5 receptor antagonist‐pretreated animals (wild‐type or D5KO) failed to display D1/D5 agonist‐induced LMs. In contrast, administration of broad spectrum or selective D2, D3 or D4 agonists consistently failed to elicit significant hindlimb movements. Overall, the results clearly show in mice the existence of a role for D1 receptors in spinal network activation and corresponding rhythmic movement generation.

Spontaneous recovery of hindlimb movement in completely spinal cord transected mice: a comparison of assessment methods and conditions

Study design:To compare results obtained with a variety of locomotor rating scales in Th9/10 spinal cord transected (Tx) mice.Objectives:To assess spontaneous recovery with a variety of rating scales to find the most sensitive methods for assessing recovery levels in Tx mice and differences associated with gender and condition.Setting:Laval University Medical Center, Neuroscience Unit & Laval University, Department of Anatomy and Physiology, Quebec City, Quebec, Canada.Methods:Scales including the Basso, Beattie and Bresnahan (BBB), the Basso Mouse Score (BMS), the Antri, Orsal and Barthe (AOB), the Motor Function Score (MFS) and the Averaged Combined Score (ACOS) were used to assess, in open-field and treadmill conditions, spontaneous locomotor recovery in male and female Tx mice.Results:The ACOS scale revealed a progressive increase of spontaneous recovery during 5-weeks post-Tx. The other methods detected a progressive increase for the first 2–3 weeks post-Tx without any significant progress in weeks 4 and 5. Generally, scores obtained with each method were nonsignificantly different between males and females or between open-field and treadmill conditions.Conclusion:These results further confirm the existence of a limited but significant increase of locomotor function recovery, occurring without intervention, in Tx animals. Although each method could detect small levels of recovery, the ACOS method was discriminative enough to detect progressive changes up to 5 weeks post-Tx. In conclusion, the ACOS rating scale was the most discriminative method for assessing the spontaneous return of hindlimb movements found in Tx mice, both in open-field and treadmill conditions.

Vascular remodeling and media calcification increases arterial stiffness in chronic kidney disease

Abstract Background: Cardiovascular disease is the most common cause of death in patients with chronic kidney disease (CKD). Arterial stiffness and calcification are non-traditional risk factors of cardiovascular disease in CKD. In CKD rats, we investigated the involvement of smooth muscle cells differentiation to osteoblast-like cells and blood vessel wall remodeling, associated with media calcification, in arterial stiffness. Method: CKD with vascular calcification was induced by subtotal nephrectomy followed by treatment with a high calcium and phosphate diet, and vitamin D supplementation (Ca/P/VitD). At week 3–6, hemodynamic parameters and pulse wave velocity (PWV) were assessed. Vascular media calcification and remodeling were determined by histological von Kossa staining and confocal immunofluorescence analysis of osteocalcin, elastin, α-smooth muscle actin (α-SMA) and collagen-1. Results: Treatment of CKD rats with Ca/P/VitD, but not normal animals, induced a significant increase in pulse pressure and PWV (pu2009<u20090.05) and marked calcification in the media. In calcification areas, de novo expression of osteocalcin was observed, whereas α-SMA immunofluorescence levels were reduced (pu2009<u20090.01). The immunofluorescence levels of elastin were also reduced, which was related to disruption of elastic lamella. In contrast, collagen-1 immunofluorescence levels in areas of calcification were increased (pu2009<u20090.01). Changes in both α-SMA and elastin inversely correlated with the PWV. Conclusion: This study indicate that smooth muscle cells differentiation to osteoblast-like cells and the associated media remodeling, which includes disruption of elastic lamellas and deposition of collagen are, at least in part, associated with the increased arterial stiffness observed in CKD rats with vascular calcification.

Strain-dependent recovery of spontaneous hindlimb movement in spinal cord transected mice (CD1, C57BL/6, BALB/c).

Reorganization and plasticity after spinal cord injury have been recently shown to take place in sublesional neuronal networks, but the possibility of strain-dependent changes at that level has never been explored. The authors studied the spontaneous return of hindlimb movement in low-thoracic spinal cord transected (Tx) mice from 3 commonly used strains. Without intervention, most CD1, C57BL/6, and BALB/c mice displayed some hindlimb movement recovery after Tx. Although all assessment methods unanimously reported that CD1 displayed higher recovery levels than did the C57BL/6 and BALB/c, higher scores were generally found with the Antri-Orsal-Barthe (M. Antri, D. Orsal, & J. Y. Barthe, 2002) and the Average Combined Score (P. A. Guertin, 2005a) methods. Such spontaneous recovery in low-thoracic Tx mice is likely the result of neuronal plasticity at the lumbosacral spinal cord level, suggesting that these sublesional changes are strain dependent.

Oral administration of a tri-therapy for central pattern generator activation in paraplegic mice: Proof-of-concept of efficacy

Spinal cord injury (SCI) is a neurological condition, for which no cure exists, typically leading to an immediate and irreversible loss of sensory and voluntary motor functions accompanied by significant health problems. We conducted proof‐of‐concept experiments aimed at assessing efficacy upon oral administration of a novel combination therapy for central pattern generator (CPG) activation and corresponding locomotor movement generation in completely paraplegic animals. Co‐administration orally (by gavage) of buspirone, levodopa and carbidopa was found to dose‐dependently induce episodes of steady weight‐bearing stepping in low‐thoracic (Th9/10) spinal cord‐transected (Tx) mice (with no other form of assistance or training). Robust hindlimb stepping with weight‐bearing capabilities was induced with the tri‐therapy but not with clinically relevant doses of these compounds administered separately. These results provide evidence suggesting that this drug combination may be ideally suited to constitute a first‐in‐class therapy (CPG activator) for locomotor activity induction in chronic SCI individuals, given that efficacy was shown using commercially available brain‐permeable small molecules, already known as safe for the treatment of various neurological indications.

Effects on Locomotion, Muscle, Bone, and Blood Induced by a Combination Therapy Eliciting Weight-Bearing Stepping in Nonassisted Spinal Cord–Transected Mice

Background. The health benefits associated with physical activity–based rehabilitation in patients with no lower-extremity motor function after a spinal cord injury (SCI) is uncertain. Methods. The authors assessed signs of efficacy, safety, and utility associated with a novel pharmacological combination therapy to activate central pattern generator (CPG) activity and corresponding locomotor activity in complete thoracic Th9/10-transected mice. Results. Subcutaneous administration 4 times per week for 1 month of 1.5 mg/kg buspirone, 1.5 mg/kg apomorphine, 12.5 mg/kg benserazide, and 50 mg/kg L-DOPA induced episodes of weight-bearing stepping on a treadmill in nonassisted paraplegic mice for 45-minute sessions. Hindlimb muscle cross-sectional area and fiber area values as well as several blood cell constituent levels assessed at 30 days postinjury were positively affected by the combination therapy, as compared with controls. Episodes of locomotion remained effective on each treatment. Femoral bone mineral density loss was not prevented by triple therapy. Conclusion. Although translation of these findings needs further experimentation, similar pharmacological activation of the CPG offers a novel therapeutic target to provide some health benefits in motor-complete SCI patients.

Early adaptive changes in chronic paraplegic mice: a model to study rapid health degradation after spinal cord injury

Study design:Literature review.Objective:To describe quantitatively some of most important anatomic, systemic, and metabolic changes occurring soon (one month) after spinal cord trauma in mice.Setting:University Laval Medical Center.Results:Significant changes in weight, mechanical and contractile muscle properties, bone histomorphometry and biomechanics, deep-vein morphology, complete blood count, immune cell count, lipid metabolism and anabolic hormone levels were found occurring within 1 month in completely spinal cord transected (Th9/10) mice.Conclusion:These data reveal that many changes in mice and humans are comparable suggesting, in turn, that this model may be a valuable tool for neuroscientists to investigate the specific mechanisms associated with rapid health degradation post-SCI.

Effects of Spinal α2-Adrenoceptor and I1-Imidazoline Receptor Activation on Hindlimb Movement Induction in Spinal Cord-Injured Mice

A partial recovery of locomotor functions has been shown in spinal cord-transected (Tx) cats after regular treadmill training and repeated administration of clonidine, an α2-adrenoreceptor agonist. However, clonidine has generally failed to show prolocomotor effects in other models (e.g., rat or mudpuppy in vitro-isolated spinal cord preparations). The reasons for this discrepancy remain unclear, but they may suggest condition- or species-specific effects induced by clonidine. This study is aimed at examining both the acute (at 6 or 41 days post-Tx) and chronic effects of repeated (once a week for one month) clonidine administration (0.25–5.0 mg/kg i.p.) on hindlimb movement generation in Tx mice (thoracic segment9/10). Locomotor-like (LM) and nonlocomotor movements (NLM) were assessed both in open-field and treadmill conditions. The results show that clonidine consistently failed, in both conditions, to induce LM and NLM at all time points even though control experiments revealed hindlimb movements steadily induced by 8-hydroxy-2-(di-N-propylamino)-tetralin (8-OH-DPAT), a serotonin receptor agonist. In turn, clonidine acutely suppressed (I1-imidazoline receptor-mediated) the frequency of spontaneously occurring LM and NLM but apparently increased spinal excitability over time, because the frequency of spontaneous LM and NLM was significantly greater in clonidine-treated (before an injection) than vehicle-treated animals after repeated administration for a few weeks. The results clearly show that clonidine can not acutely induce hindlimb movements in untrained and otherwise nonstimulated (e.g., no tail or perineal pinching) Tx mice, although repeated administration may progressively facilitate the expression of spontaneous hindlimb movements.

Endothelin type A receptor blockade reduces vascular calcification and inflammation in rats with chronic kidney disease.

Objective: Arterial stiffness and calcification are nontraditional cardiovascular risk factors in chronic kidney disease (CKD). Using a rat model of CKD with mineral imbalance, medial vascular calcification has been associated with inflammation and increased endothelin-1 (ET-1) production. We therefore hypothesized that ET-1, through the endothelin type A (ETA) receptor, induces vascular inflammation, calcification and stiffness in CKD. Methods: CKD was induced in Wistar rats by renal mass ablation. To induce medial vascular calcification, mineral imbalance was established with a identified as calcium-rich/phosphate-rich diet and vitamin D supplementation (Ca/P/VitD). One group of CKDu200a+u200aCa/P/VitD rats was given the ETA receptor antagonist atrasentan (10u200amg/kg/day) for 6 weeks. Hemodynamic parameters including SBP, pulse pressure (PP) and pulse wave velocity (PWV) were determined. Vascular calcification, smooth muscle cells osteoblastic differentiation and expression of inflammatory markers such as inflammatory cytokines and calgranulins S100A8 and S100A9 were assessed in the thoracic aorta. Results: As compared with CKD control rats, CKDu200a+u200aCa/P/VitD rats developed medal vascular calcification that was associated with increased SBP, PP and PWV. These changes were also associated with increased macrophage infiltration and expression of IL-6, calgranulins and osteoblastic markers. Treatment of CKDu200a+u200aCa/P/VitD rats with atrasentan reduced vascular calcification, SBP, PP and PWV, macrophage infiltration and expression of IL-1&bgr;, IL-6, tumor necrosis factor, calgranulins and osteoblastic markers. Conclusion: This study shows that ETA receptor blockade reduced vascular inflammation, smooth muscle cells differentiation, calcification and stiffness indicating a pivotal role for ET-1 in medial vascular calcification in this rat remnant kidney model of CKD with mineral imbalance. Therefore, the endothelin system may be a potential therapeutic target for improving cardiovascular morbidity in patients with CKD.