Sabien van Neerven

RAR/RXR and PPAR/RXR signaling in neurological and psychiatric diseases.

Retinoids are important signals in brain development. They regulate gene transcription by binding to retinoic acid receptors (RAR) and, as was discovered recently, a peroxisome proliferator-activated receptor (PPAR). Traditional ligands of PPAR are best known for their functions in lipid metabolism and inflammation. RAR and PPAR are ligand-activated transcription factors, which share members of the retinoid X receptor (RXR) family as heterodimeric partners. Both signal transduction pathways have recently been implicated in the progression of neurodegenerative and psychiatric diseases. Since inflammatory processes contribute to various neurodegenerative diseases, the anti-inflammatory activity of retinoids and PPARgamma agonists recommends them as potential therapeutic targets. In addition, genetic linkage studies, transgenic mouse models and experiments with vitamin A deprivation provide evidence that retinoic acid signaling is directly involved in physiology and pathology of motoneurons, of the basal ganglia and of cognitive functions. The activation of PPAR/RXR and RAR/RXR transcription factors has therefore been proposed as a therapeutic strategy in disorders of the central nervous system.

Inflammatory cytokine release of astrocytes in vitro is reduced by all-trans retinoic acid

In the central nervous system inflammation is mediated by microglia and astrocytes. To investigate its regulation, murine astrocyte cultures were treated with bacterial lipopolysaccharides (LPS) and analyzed with Affymetrix gene array, qRT-PCR and ELISA. Cells responded to LPS with a strong upregulation of pro-inflammatory cytokines and chemokines. Treatment with the transcriptional activator retinoic acid (RA) suppressed mRNA expression and protein release of several important cytokines (IL-1β 4%, IL-6 21%, TNFα 30%, IL-12p40 42%, and IL-12p35/p40 27%; p<0.01). The data are consistent with the hypothesis that all-trans RA takes part in endogenous anti-inflammatory feedback loops in the CNS.

Inflammatory chemokine release of astrocytes in vitro is reduced by all-trans retinoic acid.

J. Neurochem. (2010) 114, 1511–1526.

RAR/RXR and PPAR/RXR Signaling in Spinal Cord Injury

The retinoid acid receptors (RAR) and peroxisome proliferator-activated receptors (PPAR) have been implicated in the regulation of inflammatory reactions. Both receptor families contain ligand-activated transcription factors which form heterodimers with retinoid X receptors (RXR). We review data that imply RAR/RXR and PPAR/RXR pathways in physiological reactions after spinal cord injury. Experiments show how RAR signaling may improve axonal regeneration and modulate reactions of glia cells. While anti-inflammatory properties of PPAR are well documented in the periphery, their possible roles in the central nervous system have only recently become evident. Due to its anti-inflammatory function this transcription factor family promises to be a useful target after spinal cord or brain lesions.

Anti-inflammatory effect of retinoic acid on prostaglandin synthesis in cultured cortical astrocytes

Prostanoids are important mediators of inflammation and pain signaling. Although it is now well accepted that astrocytes participate in inflammatory reactions in the CNS, the molecular regulation of this activity is still largely unknown. Specifically, the regulation of prostanoid synthesis by this type of glia remains to be resolved. Recent evidence suggests that the transcriptional regulator retinoic acid (RA) is involved in regulation of the immune response. We have investigated the expression pattern of the enzymes that catalyze prostanoid and leukotriene synthesis in cultured cortical astrocytes, their stimulation by lipopolysaccharides (LPS) and their regulation by RA. The data indicate that astrocytes are an important source of prostaglandins (PGs) and that RA reduces their inflammatory biosynthesis. LPS treatment induced the expression of enzymes for the production of arachidonic acid and PGs but caused down‐regulation of a PG degrading enzyme and of leukotriene synthesizing enzymes that compete with PG synthesis. Consequently, the secretion of the PGE2 was highly increased after LPS exposure. RA counteracted the inflammatory regulation of cyclooxygenase (COX)‐2 mRNA and protein in astrocytes and thereby reduced the synthesis of PGE2 by approximately 60%. In the absence of LPS, RA enhanced the expression of COX‐1 mRNA. In conclusion, RA might be effective in suppressing inflammatory processes in the brain by inhibiting PG synthesis.

Pre-differentiation of mesenchymal stromal cells in combination with a microstructured nerve guide supports peripheral nerve regeneration in the rat sciatic nerve model.

Many bioartificial nerve guides have been investigated pre‐clinically for their nerve regeneration‐supporting function, often in comparison to autologous nerve transplantation, which is still regarded as the current clinical gold standard. Enrichment of these scaffolds with cells intended to support axonal regeneration has been explored as a strategy to boost axonal regeneration across these nerve guides Ansselin et al. (1998). In the present study, 20 mm rat sciatic nerve defects were implanted with a cell‐seeded microstructured collagen nerve guide (Perimaix) or an autologous nerve graft. Under the influence of seeded, pre‐differentiated mesenchymal stromal cells, axons regenerated well into the Perimaix nerve guide. Myelination‐related parameters, like myelin sheath thickness, benefitted from an additional seeding with pre‐differentiated mesenchymal stromal cells. Furthermore, both the number of retrogradely labelled sensory neurons and the axon density within the implant were elevated in the cell‐seeded scaffold group with pre‐differentiated mesenchymal stromal cells. However, a pre‐differentiation had no influence on functional recovery. An additional cell seeding of the Perimaix nerve guide with mesenchymal stromal cells led to an extent of functional recovery, independent of the differentiation status, similar to autologous nerve transplantation. These findings encourage further investigations on pre‐differentiated mesenchymal stromal cells as a cellular support for peripheral nerve regeneration.

A flexible, sterile, and cost-effective retractor system for microsurgery

We read with interest the publications by Di Summa (2009) and by Kuhbier (2011) and propose some short additional notes. We also introduce our simple and cost-effective approach for a microretractor system, which is different from the others, but which from our point of view combines their advantages and avoids certain disadvantages. Di Summa et al. presented the use of paper clips as microretractors. These paper clips were shaped, kept in place, and stabilized using a needle. They proposed optimal surgical exposure without device bulkiness, combined with advantages such as simplicity, costeffectiveness, avoidance of sharp tips, or stress on the wounds. This is similar to the procedure recommended by Braunstein in the 1960s, of using safety pins as self-retaining microretractors. The method by Di Summa et al., as outlined by Kuhbier et al. (2011), had disadvantages of the lack of stiffness, possible oxidation after sterilizing processes, and restrictions due to the short length of the clips. Kuhbier et al. themselves described a simple and inexpensive microretractor system using wires of 0.7 mm diameter, which were fixed to a metal frame. They claimed that their sterilizable retractors were firm, preventing from deformation and, more importantly, could be used in a more ‘‘universal manner’’ due to the lengths and shapes of the tips. With this letter, we introduce a microretractor system, which is a different approach from those mentioned above, but which meets requirements thought to be advantageous. Initially, the sharp tip of a commercially available sterile 20-gauge needle (Sterican, B. Braun Melsungen AG) is bent to obtain a blunt tip (Fig. 1A). The needle can then be divided into three segments by bending it to an angle of nearly 908 at three points: 1) at the base (i.e., transition from plastic to metal), 2) at the end of the first third, and 3) at the end of the second third—thus forming a hook-like shape. The base of the needle is bound with an elasticated silicon loop (Fig. 1A), which can be prepared and sterilized or bought in the form of prefabricated loops (ETHILOOP 1, 3 mm, Ethicon, Nordstedt). We prefer the latter option. The second end of the silicon loop is fixed into position with a second needle (Figs. 1B and 1C). Wound retraction may be not the most important scientific topic, but it is of great practical interest. As mentioned by Braunstein et al. (1960) and Halsted, ‘‘exposure is the key to surgery.’’ In microsurgical experiments, an effective retraction is essential and is necessarily required for an optimal overview over the surgical field as well as for convenient access to the desired anatomical structures. On the basis of our own experience with a couple of 100 microsurgical experiments, and in agreement with abovementioned publications, we summarize the proposed requirements for retractor systems and differentiate between obligatory from optional requirements: *Correspondence to: Ahmet Bozkurt, MD, PhD, Department of Plastic Surgery, Reconstructive and Hand Surgery, Burn Center, Medical Faculty, RWTH Aachen University, Aachen, Pauwelsstrasse 30, Aachen 52074, Germany. E-mail: abozkurt@ukaachen.de Received 18 May 2011; Accepted 2 June 2011 Published online 18 October 2011 in Wiley Online Library (wileyonlinelibrary. com). DOI 10.1002/micr.20934

Functional recovery not correlated with axon regeneration through olfactory ensheathing cell-seeded scaffolds in a model of acute spinal cord injury

The implantation of bioengineered scaffolds into lesion-induced gaps of the spinal cord is a promising strategy for promoting functional tissue repair because it can be combined with other intervention strategies. Our previous investigations showed that functional improvement following the implantation of a longitudinally microstructured collagen scaffold into unilateral mid-cervical spinal cord resection injuries of adult Lewis rats was associated with only poor axon regeneration within the scaffold. In an attempt to improve graft-host integration as well as functional recovery, scaffolds were seeded with highly enriched populations of syngeneic, olfactory bulb-derived ensheathing cells (OECs) prior to implantation into the same lesion model. Regenerating neurofilament-positive axons closely followed the trajectory of the donor OECs, as well as that of the migrating host cells within the scaffold. However, there was only a trend for increased numbers of regenerating axons above that supported by non-seeded scaffolds or in the untreated lesions. Nonetheless, significant functional recovery in skilled forelimb motor function was observed following the implantation of both seeded and non-seeded scaffolds which could not be correlated to the extent of axon regeneration within the scaffold. Mechanisms other than simple bridging of axon regeneration across the lesion must be responsible for the improved motor function.

Cell-enrichment with olfactory ensheathing cells has limited local extra beneficial effects on nerve regeneration supported by the nerve guide Perimaix

The reconstruction of peripheral nerve injuries is clinically challenging, and today, the autologous nerve transplantation is still considered as the only gold standard remedy for nerve lesions where a direct nerve coaptation is not possible. Nevertheless, the functional merits of many biomaterials have been tested as potential substitutes for the autologous nerve transplant. One of the strategies that have been pursued is the combination of bioengineered nerve guides with cellular enrichment. In this present study, we combined the previously evaluated collagen‐based and microstructured nerve guide Perimaix with olfactory ensheathing cell enrichment. Rat sciatic nerve defects of 20 mm were either bridged by a cell‐seeded or nonseeded nerve guide or an autologous nerve transplant. Animals were monitored for 12 weeks for structural and functional parameters. Seeded cells survived on Perimaix, and following implantation aligned along the microstructured Perimaix framework. Axonal densities within the cell‐seeded nerve guides were higher than in the nonseeded nerve guides and were comparable to the autograft. Additionally, cell‐seeding had local beneficial effects on myelination within the nerve guide, as myelin sheath thickness was enhanced when compared with the empty scaffold. Nevertheless, for bridging the nerve gap of 20 mm, both the cell‐seeded as well as nonseeded scaffolds were equally efficient regarding the functional outcome, which did not differ between the autograft, seeded or nonseeded groups. Our data demonstrate that olfactory ensheathing cell enrichment has local effects on nerve regeneration in combination with the Perimaix nerve guide. Surprisingly, for traversing the lesion gap, additional cell‐seeding is not crucial.