Erythropoietin Ameliorates Rat Experimental Autoimmune Neuritis by Inducing Transforming Growth Factor-Beta in Macrophages

Erythropoietin (EPO) is a pleiotropic cytokine originally identified for its role in erythropoiesis. In addition, in various preclinical models EPO exhibited protective activity against tissue injury. There is an urgent need for potent treatments of autoimmune driven disorders of the peripheral nervous system (PNS), such as the Guillain-Barré syndrome (GBS), a disabling autoimmune disease associated with relevant morbidity and mortality. To test the therapeutic potential of EPO in experimental autoimmune neuritis (EAN) - an animal model of human GBS – immunological and clinical effects were investigated in a preventive and a therapeutic paradigm. Treatment with EPO reduced clinical disease severity and if given therapeutically also shortened the recovery phase of EAN. Clinical findings were mirrored by decreased inflammation within the peripheral nerve, and myelin was well maintained in treated animals. In contrast, EPO increased the number of macrophages especially in later stages of the experimental disease phase. Furthermore, the anti-inflammatory cytokine transforming growth factor (TGF)-beta was upregulated in the treated cohorts. In vitro experiments revealed less proliferation of T cells in the presence of EPO and TGF-beta was moderately induced, while the secretion of other cytokines was almost not altered by EPO. Our data suggest that EPO revealed its beneficial properties by the induction of beneficial macrophages and the modulation of the immune system towards anti-inflammatory responses in the PNS. Further studies are warranted to elaborate the clinical usefulness of EPO for treating immune-mediated neuropathies in affected patients.     

Erythropoietin ameliorates rat experimental autoimmune neuritis by inducing transforming growth factor-β in macrophages

Erythropoietin (EPO) is a pleiotropic cytokine originally identified for its role in erythropoiesis. In addition, in various preclinical models EPO exhibited protective activity against tissue injury. There is an urgent need for potent treatments of autoimmune driven disorders of the peripheral nervous system (PNS), such as the Guillain-Barré syndrome (GBS), a disabling autoimmune disease associated with relevant morbidity and mortality. To test the therapeutic potential of EPO in experimental autoimmune neuritis (EAN) - an animal model of human GBS--immunological and clinical effects were investigated in a preventive and a therapeutic paradigm. Treatment with EPO reduced clinical disease severity and if given therapeutically also shortened the recovery phase of EAN. Clinical findings were mirrored by decreased inflammation within the peripheral nerve, and myelin was well maintained in treated animals. In contrast, EPO increased the number of macrophages especially in later stages of the experimental disease phase. Furthermore, the anti-inflammatory cytokine transforming growth factor (TGF)-beta was upregulated in the treated cohorts. In vitro experiments revealed less proliferation of T cells in the presence of EPO and TGF-beta was moderately induced, while the secretion of other cytokines was almost not altered by EPO. Our data suggest that EPO revealed its beneficial properties by the induction of beneficial macrophages and the modulation of the immune system towards anti-inflammatory responses in the PNS. Further studies are warranted to elaborate the clinical usefulness of EPO for treating immune-mediated neuropathies in affected patients.     

Small-molecule-induced Rho-inhibition: NSAIDs after spinal cord injury

Limited axonal plasticity within the central nervous system (CNS) is a major restriction for functional recovery after CNS injury. The small GTPase RhoA is a key molecule of the converging downstream cascade that leads to the inhibition of axonal re-growth. The Rho-pathway integrates growth inhibitory signals derived from extracellular cues, such as chondroitin sulfate proteoglycans, Nogo-A, myelin-associated glycoprotein, oligodendrocyte-myelin glycoprotein, Ephrins and repulsive guidance molecule-A, into the damaged axon. Consequently, the activation of RhoA results in growth cone collapse and finally outgrowth failure. In turn, the inhibition of RhoA-activation blinds the injured axon to its growth inhibitory environment resulting in enhanced axonal sprouting and plasticity. This has been demonstrated in various CNS-injury models for direct RhoA-inhibition and for downstream/upstream blockade of the RhoA-associated pathway. In addition, RhoA-inhibition reduces apoptotic cell death and secondary damage and improves locomotor recovery in clinically relevant models after experimental spinal cord injury (SCI). Unexpectedly, a subset of "small molecules" from the group of non-steroid anti-inflammatory drugs, particularly the FDA-approved ibuprofen, has recently been identified as (1) inhibiting RhoA-activation, (2) enhancing axonal sprouting/regeneration, (3) protecting "tissue at risk" (neuroprotection) and (4) improving motor recovery confined to realistic therapeutical time-frames in clinically relevant SCI models. Here, we survey the effect of small-molecule-induced RhoA-inhibition on axonal plasticity and neurofunctional outcome in CNS injury paradigms. Furthermore, we discuss the body of preclinical evidence for a possible clinical translation with a focus on ibuprofen and illustrate putative risks and benefits for the treatment of acute SCI.     

AMIGO3 Is an NgR1/p75 Co-Receptor Signalling Axon Growth Inhibition in the Acute Phase of Adult Central Nervous System Injury

Axon regeneration in the injured adult CNS is reportedly inhibited by myelin-derived inhibitory molecules, after binding to a receptor complex comprised of the Nogo-66 receptor (NgR1) and two transmembrane co-receptors p75/TROY and LINGO-1. However, the post-injury expression pattern for LINGO-1 is inconsistent with its proposed function. We demonstrated that AMIGO3 levels were significantly higher acutely than those of LINGO-1 in dorsal column lesions and reduced in models of dorsal root ganglion neuron (DRGN) axon regeneration. Similarly, AMIGO3 levels were raised in the retina immediately after optic nerve crush, whilst levels were suppressed in regenerating optic nerves, induced by intravitreal peripheral nerve implantation. AMIGO3 interacted functionally with NgR1-p75/TROY in non-neuronal cells and in brain lysates, mediating RhoA activation in response to CNS myelin. Knockdown of AMIGO3 in myelin-inhibited adult primary DRG and retinal cultures promoted disinhibited neurite growth when cells were stimulated with appropriate neurotrophic factors. These findings demonstrate that AMIGO3 substitutes for LINGO-1 in the NgR1-p75/TROY inhibitory signalling complex and suggests that the NgR1-p75/TROY-AMIGO3 receptor complex mediates myelin-induced inhibition of axon growth acutely in the CNS. Thus, antagonizing AMIGO3 rather than LINGO-1 immediately after CNS injury is likely to be a more effective therapeutic strategy for promoting CNS axon regeneration when combined with neurotrophic factor administration.     

精氨酸酶Ⅰ参与环腺苷酸促进脑缺血再灌注大鼠的轴突再生

环腺苷酸（cAMP）作为细胞内的重要第二信使之一,主要通过激活下游cAMP依赖性蛋白激酶A（PKA）,进一步激活转录因子-cAMP效应元件结合蛋（CREB）,达到促进损伤轴突再生的作用.精氨酸酶Ⅰ主要是通过促进多胺的表达,从而克服髓鞘相关抑制因子对轴突再生的抑制作用,达到促进轴突再生的效果.在脑缺血中,cAMP促进轴突再生的过程是否有精氨酸酶Ⅰ的参与及其与RhoA信号通路的关系尚不清楚.本研究采用线栓法制备脑缺血再灌注模型（MACO）,采用Longa 5评分法对大鼠运动功能进行评分,利用逆转录聚合酶链反应（RT-PCR）和Western蛋白印迹方法分别检测缺血灶周边脑组织生长相关蛋白43（GAP-43）和RhoA的mRNA和蛋白表达,免疫组化法进行GAP-43的形态学检测,作为轴突再生的标志.通过尾静脉注射cAMP类似物db-cAMP增加脑缺血后大鼠脑组织内cAMP的浓度后发现：db-cAMP处理可明显降低MACO大鼠的运动功能评分,且可促进GAP-43 mRNA及蛋白的表达,抑制RhoA mRNA及蛋白的表达,由此可见db-cAMP处理可促进脑缺血后大鼠运动功能的恢复,且这一过程与抑制RhoA通路,进而促进轴突再生有关;通过在db-cAMP的基础上给予精氨酸酶Ⅰ拮抗剂NOHA来降低精氨酸酶Ⅰ的活性发现：给予NOHA的大鼠运动功能评分明显增加,这一变化趋势与RhoA mRNA及蛋白表达的变化趋势相一致,而与GAP-43 mRNA及蛋白表达的变化趋势相反. 因此可推断：精氨酸酶Ⅰ参与了db-cAMP促进轴突再生、改善脑缺血后大鼠运动功能的过程,且与钝化RhoA通路有关.     

硫酸软骨素蛋白多糖与神经系统的发育和再生

硫酸软骨素蛋白多糖（chondroitin sulfate proteoglycans,CSPGs）是中枢神经系统（CNS）细胞外基质中的重要组成成分,在CNS的发育、成熟后正常功能的维持中发挥重要功能,如发育中影响神经细胞的迁移和轴突生长,成年后参与神经可塑性的控制等;而病理条件下,如CNS受损后又可做为胶质瘢痕的重要组分抑制受损神经的再生。研究发现,用酶降解CSPGs的糖氨多糖链或阻断其合成可以有效地削弱CSPGs对受损神经的抑制作用,促进轴突再生。然而,精确调控CSPGs特定时空表达模式的分子机制,以及功能发挥所涉及的完整信号转导通路还有待进一步研究。     

Antibodies against gangliosides and ganglioside complexes in Guillain-Barré syndrome: new aspects of research

Guillain-Barré syndrome (GBS) is an acute autoimmune neuropathy, often preceded by an infection. Serum anti-ganglioside antibodies are frequently elevated in titer. Those antibodies are useful for diagnosis. Some of them also may be directly involved in the pathogenetic mechanisms by binding to the regions where the respective target ganglioside is specifically localized. We have recently found the presence of the antibody that specifically recognizes a new conformational epitope formed by two gangliosides (ganglioside complex) in the acute-phase sera of some GBS patients. In particular, the antibodies against GD1a/GD1b and/or GD1b/GT1b complexes are associated with severe GBS requiring artificial ventilation. Some patients with Miller Fisher syndrome also have antibodies against ganglioside complexes including GQ1b; such as GQ1b/GM1 and GQ1b/GD1a. Gangliosides along with other components as cholesterol are known to form lipid rafts, in which the carbohydrate portions of two different gangliosides may form a new conformational epitope. Within the rafts, gangliosides are considered to interact with important receptors or signal transducers. The antibodies against ganglioside complexes may therefore directly cause nerve conduction failure and severe disability in GBS. More study is needed to elucidate the roles of the antibodies against ganglioside complexes.     

Antibodies against gangliosides and ganglioside complexes in Guillain–Barré syndrome: New aspects of research

Guillain–Barré syndrome (GBS) is an acute autoimmune neuropathy, often preceded by an infection. Serum anti-ganglioside antibodies are frequently elevated in titer. Those antibodies are useful for diagnosis. Some of them also may be directly involved in the pathogenetic mechanisms by binding to the regions where the respective target ganglioside is specifically localized. We have recently found the presence of the antibody that specifically recognizes a new conformational epitope formed by two gangliosides (ganglioside complex) in the acute-phase sera of some GBS patients. In particular, the antibodies against GD1a/GD1b and/or GD1b/GT1b complexes are associated with severe GBS requiring artificial ventilation. Some patients with Miller Fisher syndrome also have antibodies against ganglioside complexes including GQ1b; such as GQ1b/GM1 and GQ1b/GD1a. Gangliosides along with other components as cholesterol are known to form lipid rafts, in which the carbohydrate portions of two different gangliosides may form a new conformational epitope. Within the rafts, gangliosides are considered to interact with important receptors or signal transducers. The antibodies against ganglioside complexes may therefore directly cause nerve conduction failure and severe disability in GBS. More study is needed to elucidate the roles of the antibodies against ganglioside complexes.     

Synergistic Effects of Bone Mesenchymal Stem Cells and Chondroitinase ABC on Nerve Regeneration After Acellular Nerve Allograft in Rats

This study aimed to evaluate whether combination therapy of bone marrow stromal cells (BMSCs) transplantation and chondroitinase ABC (ChABC) treatment further enhances axonal regeneration and functional recovery after acellular nerve allograft repair of the sciatic nerve gap in rats. Eight Sprague–Dawley rats were used as nerve donors, and 32 Wistar rats were randomly divided into four groups: Group I: acellular rat sciatic nerve (ARSN) group; Group II: ChABC treatment; Group III: BMSCs transplantation; and Group IV: ChABC treatment and BMSCs transplantation. The results showed that compared with ARSN control group, BMSC transplantation promoted axonal regeneration, the secretion of neural trophic factors NGF, BDNF and axon angiogenesis in nerve graft. ChABC treatment degraded chondroitin sulfate proteoglycans in ARSN in vitro and in vivo and improved BMSCs survival in ARSN. The combination therapy caused much better beneficial effects evidenced by increasing sciatic function index, nerve conduction velocity, restoration rate of tibialis anterior wet muscle weight, and myelinated nerve number, but did not further boost the therapeutic effects on neurotrophic factor production, axon angiogenesis, and sensory functional recovery by BMSC transplantation. Taken together, for the first time, we demonstrate the synergistic effects of BMSC transplantation and BMSCs treatment on peripheral nerve regeneration, and our findings may help establish novel strategies for cell transplantation therapy for peripheral nerve injury.     

Functional motor recovery is improved due to local placement of GDNF microspheres after delayed nerve repair

The majority of bioengineering strategies to promote peripheral nerve regeneration after injury have focused on therapies to bridge large nerve defects while fewer therapies are being developed to treat other nerve injuries, such as nerve transection. We constructed delivery systems using fibrin gels containing either free GDNF or polylactide–glycolic acid (PLGA) microspheres with GDNF to treat delayed nerve repair, where ELISA verified GDNF release. We determined the formulation of microspheres containing GDNF that optimized nerve regeneration and functional recovery in a rat model of delayed nerve repair. Experimental groups underwent delayed nerve repair and treatment with GDNF microspheres in fibrin glue at the repair site or control treatments (empty microspheres or free GDNF without microspheres). Contractile muscle force, muscle mass, and MUNE were measured 12 weeks following treatment, where GDNF microspheres (2 weeks formulation) were superior compared to either no GDNF or short‐term release of free GDNF to nerve. Nerve histology distal to the repair site demonstrated increased axon counts and fiber diameters due to GDNF microspheres (2 weeks formulation). GDNF microspheres partially reversed the deleterious effects of chronic nerve injury, and recovery was slightly favored with the 2 weeks formulation compared to the 4 weeks formulation. Biotechnol. Bioeng. 2013; 110: 1272–1281. © 2012 Wiley Periodicals, Inc.     

Curiosity and cure: translational research strategies for neural repair-mediated rehabilitation

Clinicians who seek interventions for neural repair in patients with paralysis and other impairments may extrapolate the results of cell culture and rodent experiments into the framework of a preclinical study. These experiments, however, must be interpreted within the context of the model and the highly constrained hypothesis and manipulation being tested. Rodent models of repair for stroke and spinal cord injury offer examples of potential pitfalls in the interpretation of results from developmental gene activation, transgenic mice, endogeneous neurogenesis, cellular transplantation, axon regeneration and remyelination, dendritic proliferation, activity-dependent adaptations, skills learning, and behavioral testing. Preclinical experiments that inform the design of human trials ideally include a lesion of etiology, volume and location that reflects the human disease; examine changes induced by injury and by repair procedures both near and remote from the lesion; distinguish between reactive molecular and histologic changes versus changes critical to repair cascades; employ explicit training paradigms for the reacquisition of testable skills; correlate morphologic and physiologic measures of repair with behavioral measures of task reacquisition; reproduce key results in more than one laboratory, in different strains or species of rodent, and in a larger mammal; and generalize the results across several disease models, such as axonal regeneration in a stroke and spinal cord injury platform. Collaborations between basic and clinical scientists in the development of translational animal models of injury and repair can propel experiments for ethical bench-to-bedside therapies to augment the rehabilitation of disabled patients.     

Axonal Regeneration and Remyelination Evaluation of Chitosan/Gelatin-Based Nerve Guide Combined with Transforming Growth Factor-β1 and Schwann Cells

Despite efforts in peripheral nerve injury and regeneration, it is difficult to achieve a functional recovery following extended peripheral nerve lesions. Even if artificial nerve conduit, cell components and growth factors can enhance nerve regeneration, integration in peripheral nerve repair and regeneration remains yet to be explored. For this study, we used chitosan/gelatin nerve graft constructed with collagenous matrices as a vehicle for Schwann cells and transforming growth factor-β1 to bridge a 10-mm gap of the sciatic nerve and explored the feasibility of improving regeneration and reinnervation in rats. The nerve regeneration was assessed with functional recovery, electrophysiological test, retrograde labeling, and immunohistochemistry analysis during the post-operative period of 16 weeks. The results showed that the internal sides of the conduits were compact enough to prevent the connective tissues from ingrowth. Nerve conduction velocity, average regenerated myelin area, and myelinated axon count were similar to those treated with autograft (p > 0.05) but significantly higher than those bridged with chitosan/gelatin nerve graft alone (p < 0.05). Evidences from retrograde labeling and immunohistochemistry analysis are further provided in support of improving axonal regeneration and remyelination. A designed graft incorporating all of the tissue-engineering strategies for peripheral nerve regeneration may provide great progress in tissue engineering for nerve repair.     

Th1/Th2/Th17/Treg cytokines in Guillain–Barré syndrome and experimental autoimmune neuritis

Guillain–Barré syndrome (GBS) is an immune-mediated acute inflammatory disorder in the peripheral nervous system (PNS) of humans characterized by inflammatory infiltration and damage to myelin and axon. Experimental autoimmune neuritis (EAN) is a useful animal model for GBS. Although GBS and EAN have been widely studied, the pathophysiological basis of GBS/EAN remains largely unknown. Immunocompetent cells together with cytokines produced by various cells contribute to the inflammatory process of EAN by acting as mediators or effectors. Both GBS and EAN have hitherto been attributed to T helper (Th)1 cells-mediated disorders, however, some changes in GBS and EAN could not be explained by the pathogenic role of Th1 cells and a disturbance of the Th1/Th2 balance, which has previously been considered to be important for the homeostatic maintenance of the immune responses and to explain the adaptive immunity and autoimmune diseases. The Th1/Th2 paradigm in autoimmune diseases has been greatly challenged in recent years, with the identification of a particular T cell subset Th17 cells. Studies on the associations between Th17 cells/cytokines and GBS/EAN are reviewed. But some of them occasionally yield conflicting results, indicating an intricate network of cytokines in immune response.     

Role of Non-coding RNAs in Axon Regeneration after Peripheral Nerve Injury

Peripheral nerve injury (PNI) may lead to disability and neuropathic pain, which constitutes a substantial economic burden to patients and society. It was found that the peripheral nervous system (PNS) has the ability to regenerate after injury due to a permissive microenvironment mainly provided by Schwann cells (SCs) and the intrinsic growth capacity of neurons; however, the results of injury repair are not always satisfactory. Effective, long-distance axon regeneration after PNI is achieved by precise regulation of gene expression. Numerous studies have shown that in the process of peripheral nerve damage and repair, differential expression of non-coding RNAs (ncRNAs) significantly affects axon regeneration, especially expression of microRNAs (miRNAs), long non-coding RNAs (lncRNAs) and circular RNAs (circRNAs). In the present article, we review the cellular and molecular mechanisms of axon regeneration after PNI, and analyze the roles of these ncRNAs in nerve repair. In addition, we discuss the characteristics and functions of these ncRNAs. Finally, we provide a thorough perspective on the functional mechanisms of ncRNAs in nervous injury repair, and explore the potential these ncRNAs offer as targets of nerve injury treatment.     

Urokinase plasminogen receptor and the fibrinolytic complex play a role in nerve repair after nerve crush in mice, and in human neuropathies

Remodeling of extracellular matrix (ECM) is a critical step in peripheral nerve regeneration. In fact, in human neuropathies, endoneurial ECM enriched in fibrin and vitronectin associates with poor regeneration and worse clinical prognosis. Accordingly in animal models, modification of the fibrinolytic complex activity has profound effects on nerve regeneration: high fibrinolytic activity and low levels of fibrin correlate with better nerve regeneration. The urokinase plasminogen receptor (uPAR) is a major component of the fibrinolytic complex, and binding to urokinase plasminogen activator (uPA) promotes fibrinolysis and cell movement. uPAR is expressed in peripheral nerves, however, little is known on its potential function on nerve development and regeneration. Thus, we investigated uPAR null mice and observed that uPAR is dispensable for nerve development, whereas, loss of uPAR affects nerve regeneration. uPAR null mice showed reduced nerve repair after sciatic nerve crush. This was a consequence of reduced fibrinolytic activity and increased deposition of endoneurial fibrin and vitronectin. Exogenous fibrinolysis in uPAR null mice rescued nerve repair after sciatic nerve crush. Finally, we measured the fibrinolytic activity in sural nerve biopsies from patients with peripheral neuropathies. We showed that neuropathies with defective regeneration had reduced fibrinolytic activity. On the contrary, neuropathies with signs of active regeneration displayed higher fibrinolytic activity. Overall, our results suggest that enforced fibrinolysis may facilitate regeneration and outcome of peripheral neuropathies.     

Rho activation patterns after spinal cord injury and the role of activated Rho in apoptosis in the central nervous system

Growth inhibitory proteins in the central nervous system (CNS) block axon growth and regeneration by signaling to Rho, an intracellular GTPase. It is not known how CNS trauma affects the expression and activation of RhoA. Here we detect GTP-bound RhoA in spinal cord homogenates and report that spinal cord injury (SCI) in both rats and mice activates RhoA over 10-fold in the absence of changes in RhoA expression. In situ Rho-GTP detection revealed that both neurons and glial cells showed Rho activation at SCI lesion sites. Application of a Rho antagonist (C3-05) reversed Rho activation and reduced the number of TUNEL-labeled cells by approximately 50% in both injured mouse and rat, showing a role for activated Rho in cell death after CNS injury. Next, we examined the role of the p75 neurotrophin receptor (p75NTR) in Rho signaling. After SCI, an up-regulation of p75NTR was detected by Western blot and observed in both neurons and glia. Treatment with C3-05 blocked the increase in p75NTR expression. Experiments with p75NTR-null mutant mice showed that immediate Rho activation after SCI is p75NTR dependent. Our results indicate that blocking overactivation of Rho after SCI protects cells from p75NTR-dependent apoptosis.     

Rho inhibition recruits DCC to the neuronal plasma membrane and enhances axon chemoattraction to netrin 1

Molecular cues, such as netrin 1, guide axons by influencing growth cone motility. Rho GTPases are a family of intracellular proteins that regulate the cytoskeleton, substrate adhesion and vesicle trafficking. Activation of the RhoA subfamily of Rho GTPases is essential for chemorepellent axon guidance; however, their role during axonal chemoattraction is unclear. Here, we show that netrin 1, through its receptor DCC, inhibits RhoA in embryonic spinal commissural neurons. To determine whether netrin 1-mediated chemoattraction requires Rho function, we inhibited Rho signaling and assayed axon outgrowth and turning towards netrin 1. Additionally, we examined two important mechanisms that influence the guidance of axons to netrin 1: substrate adhesion and transport of the netrin receptor DCC to the plasma membrane. We found that inhibiting Rho signaling increased plasma membrane DCC and adhesion to substrate-bound netrin 1, and also enhanced netrin 1-mediated axon outgrowth and chemoattractive axon turning. Conversely, overexpression of RhoA or constitutively active RhoA inhibited axonal responses to netrin 1. These findings provide evidence that Rho signaling reduces axonal chemoattraction to netrin 1 by limiting the amount of plasma membrane DCC at the growth cone, and suggest that netrin 1-mediated inhibition of RhoA activates a positive-feedback mechanism that facilitates chemoattraction to netrin 1. Notably, these findings also have relevance for CNS regeneration research. Inhibiting RhoA promotes axon regeneration by disrupting inhibitory responses to myelin and the glial scar. By contrast, we demonstrate that axon chemoattraction to netrin 1 is not only maintained but enhanced, suggesting that this might facilitate directing regenerating axons to appropriate targets.     

Enhanced Neurite Outgrowth of Human Model (NT2) Neurons by Small-Molecule Inhibitors of Rho/ROCK Signaling

Axonal injury in the adult human central nervous system often results in loss of sensation and motor functions. Promoting regeneration of severed axons requires the inactivation of growth inhibitory influences from the tissue environment and stimulation of the neuron intrinsic growth potential. Especially glial cell derived factors, such as chondroitin sulfate proteoglycans, Nogo-A, myelin-associated glycoprotein, and myelin in general, prevent axon regeneration. Most of the glial growth inhibiting factors converge onto the Rho/ROCK signaling pathway in neurons. Although conditions in the injured nervous system are clearly different from those during neurite outgrowth in vitro, here we use a chemical approach to manipulate Rho/ROCK signalling with small-molecule agents to encourage neurite outgrowth in cell culture. The development of therapeutic treatments requires drug testing not only on neurons of experimental animals, but also on human neurons. Using human NT2 model neurons, we demonstrate that the pain reliever Ibuprofen decreases RhoA (Ras homolog gene family, member A GTPase) activation and promotes neurite growth. Inhibition of the downstream effector Rho kinase by the drug Y-27632 results in a strong increase in neurite outgrowth. Conversely, activation of the Rho pathway by lysophosphatidic acid results in growth cone collapse and eventually to neurite retraction. Finally, we show that blocking of Rho kinase, but not RhoA results in an increase in neurons bearing neurites. Due to its anti-inflammatory and neurite growth promoting action, the use of a pharmacological treatment of damaged neural tissue with Ibuprofen should be explored.