Olfactory ensheathing cell transplantation improves sympathetic skin responses in chronic spinal cord injury

Forty-three patients with chronic spinal cord injury for over 6 months were transplanted with bryonic olfactory ensheathing cells, 2-4 × 106, into multiple sites in the injured area under the sur-gical microscope. The sympathetic skin response in patients was measured with an electromyo-graphy/evoked potential instrument 1 day before transplantation and 3-8 weeks after trans-tion. Spinal nerve function of patients was assessed using the American Spinal Injury Association impairment scale. The sympathetic skin response was elicited in 32 cases before olfactory en-sheathing celltransplantation, while it was observed in 34 cases after transplantation. tantly, sympathetic skin response latency decreased significantly and amplitude increased cantly after transplantation. Transplantation of olfactory ensheathing cells also improved American Spinal Injury Association scores for movement, pain and light touch. Our findings indicate that factory ensheathing celltransplantation improves motor, sensory and autonomic nerve functions in patients with chronic spinal cord injury.

Meta analysis of olfactory ensheathing cell transplantation promoting functional recovery of motor nerves in rats with complete spinal cord transection

OBJECTIVE： To evaluate the effects of olfactory ensheathing cell transplantation on functional recovery of rats with complete spinal cord transection. DATA SOURCES： A computer-based online search of Medline （1989-2013）, Embase （1989- 2013）, Cochrane library （1989-2013）, Chinese Biomedical Literature Database （1989-2013）, China National Knowledge Infrastructure （1989-2013）, VIP （1989-2013）, Wanfang databases （1989-2013） and Chinese Clinical Trial Register was conducted to collect randomized controlled trial data regarding olfactory ensheathing cell transplantation for the treatment of complete spinal cord transection in rats. SELECTION CRITERIA： Randomized controlled trials investigating olfactory ensheathing cell transplantation and other transplantation methods for promoting neurological functional recov- ery of rats with complete spinal cord transection were included in the analysis. Meta analysis was conducted using RevMan 4.2.2 software. MAIN OUTCOME MEASURES： Basso, Beattie and Bresnahan scores of rats with complete spinal cord transection were evaluated in this study. RESULTS： Six randomized controlled trials with high quality methodology were included. Meta analysis showed that Basso, Beattie and Bresnahan scores were significantly higher in the olfacto- ry ensheathing cell transplantation group compared with the control group （WMD = 3.16, 95% （21 （1.68, 4.65）; P 〈 0.00001）. CONCLUSION： Experimental studies have shown that olfactory ensheathing cell transplantation can promote the functional recovery of motor nerves in rats with complete spinal cord transection.

Transplantation of low-power laser-irradiated olfactory ensheathing cells to promote repair of spinal cord injury in rats

BACKGROUND： Previous studies have demonstrated that low-power laser （LPL） irradiation can promote the regeneration of peripheral nerves and central nerves, as well as influence cellular proliferation. Therefore, it is thought to be a potential treatment for spinal cord injury. OBJECTIVE： Utilizing histological observations and behavioral evaluations, the aim of this study was to investigate the influence of transplanted olfactory ensheathing cells （OECs）, irradiated by LPL, on functional repair of rats following transversal spinal cord injury. DESIGN, TIME AND SETTING： A randomized, controlled, animal experiment was performed at the animal experimental center in the First Affiliated Hospital of Xinjiang Medical University between January 2007 and February 2008. MATERIALS： A total of 52 Sprague Dawley rats were included in this experiment. Twelve rats were used to harvest OECs, some of which were irradiated by LPL on days 3, 5, and 7 in culture. The remaining 40 rats were used to establish T12 complete spinal cord transection injury. DMEM/F12 medium was purchased from Sigma, USA, Fluorogold was provided by Chemicon, USA, and the LY/JG650-D500-16 low-power laser was produced by Xi＇an Lingyue Electromechanical Science And Technology Co., Ltd., China. METHODS： The successful rat models were randomly divided into three groups： OEC transplantation, LPL-irradiated OEC transplantation, and control. These animals were microinjected with OEC suspension, LPL-irradiated OEC suspension, and DMEM/F12 medium （10μL） respectively 4 weeks after spinal cord was completely transected at the T12 level. MAIN OUTCOME MEASURES： Spinal cord injury was observed using hematoxylin-eosin staining Expression of nerve growth factor receptor p75 and glial fibrillary acidic protein were determined using immunohistochemical staining. Regeneration of spinal nerve fibers in rats was assayed by Fluorogold retrograde labeling method. Basso, Beattie and Bresnahan （BBB） scores were used to evaluate motor functions of rat lower limbs. RESULTS： Structural disturbances were observed following spinal cord injury in each group, and a large amount of scar tissue covered the broken ends, accompanied by porosis and inflammatory cell infiltration. Following OEC transplantation, the distal end connected to the proximal end. nerve growth factor receptor p75 and glial fibrillary acidic protein immunohistochemistry revealed positive OECs in the cephalad and caudal area of rats that received LPL-irradiated OEC transplantation. In the OECs group, only glial fibrillary acidic protein staining was observed. No staining was found in the control group. Neural fibers labeled with Fluorogold extended across the lesion area and into the cephalad and caudal area in the OECs and LPL-irradiated OECs groups, but were not present in the control group. BBB scores revealed statistically significant differences among the three groups （P 〈 0.05）： OECs irradiated by LPL group 〉 OECs group 〉 control group. CONCLUSION： Transplantation of OECs and LPL-irradiated OECs promoted functional repair in the injured spinal cord of rats, although LPL-irradiated OECs resulted in greater beneficial effects.

Transplantation of neural stem cells, Schwann cells and olfactory ensheathing cells for spinal cord injury:A Web of Science-based literature analysis

OBJECTIVE： To identify global research trends in transplantation of neural stem cells, Schwann cells and olfactory ensheathing cells for spinal cord injury. DATA RETRIEVAL： We performed a bibliometric analysis of studies on transplantation of neural stem cells, Schwann cells and olfactory ensheathing cells for spinal cord injury published from 2002 to 2011 and retrieved from the Web of Science, using the key words spinal cord injury along with either neural stem cell, Schwann cell or olfactory ensheathing cell. SELECTION CRITERIA： Inclusion criteria： （a） peer-reviewed published articles on neural stem cells, Schwann cells or olfactory ensheathing cells for spinal cord injury indexed in the Web of Science; （b） original research articles, reviews, meeting abstracts, proceedings papers, book chapters, editorial materials and news items; and （c） published between 2002 and 2011. Exclusion criteria： （a） articles that required manual searching or telephone access; （b） documents that were not published in the public domain; and （c） corrected papers. MAIN OUTCOME MEASURES： （1）Annual publication output, distribution by journal, distribution by institution and top-cited articles on neural stem cells; （2） annual publication output, distribution by journal, distribution by institution and top-cited articles on Schwann cells; （3） annual publication output, distribution by journal, distribution by institution and top-cited articles on olfactory ensheathing cells. RESULTS： This analysis, based on articles indexed in the Web of Science, identified several research trends among studies published over the past 10 years in transplantation of neural stem cells, Schwann cells and olfactory ensheathing cells for spinal cord injury. The number of publications increased over the 10-year period examined. Most papers appeared in journals with a focus on neurology, such as Journal of Neurotrauma, Experimental Neurology and Gila. Research institutes publishing on the use of neural stem cells to repair spinal cord injury were mostly in the USA and Canada. Those publishing on the use of Schwann cells were mostly in the USA and Canada as well. Those publishing on the use of olfactory ensheathing cells were mostly in the UK, the USA and Canada. CONCLUSION： On the basis of the large number of studies around the world, cell transplantation has proven to be the most promising therapeutic approach for spinal cord injury.

Olfactory ensheathing cell transplantation for spinal cord injury An 18-year bibliometric analysis based on the Web of Science

OBJECTIVE： Olfactory ensheathing cell （OEC） transplantation is a promising new approach for the treatment of spinal cord injury （SCI）, and an increasing number of scientific publications are devoted to this treatment strategy. This bibliometric analysis was conducted to assess global research trends in OEC transplantation for SCI. DATA SOURCE： All of the data in this study originate from the Web of Science maintained by the Institute for Scientific Information, USA, and includes SCI-EXPANDED, SSCI, A＆HCI, CPCI-S, CPCI-SSH, BKCI-S, BKCI-SSH, CCR-EXPANDED and IC. The Institute for Scientific Information＇s Web of Science was searched using the keywords ＂olfactory ensheathing cells＂ or ＂OECs＂ or ＂olfactory ensheathing gila＂ or ＂OEG＂ or ＂olfactory ensheathing glial cells＂ or ＂OEGs＂ and ＂spinal cord injury＂ or ＂SCI＂ or ＂spinal injury＂ or ＂spinal transection＂ for literature published from January 1898 to May 2012. DATA SELECTION： Original articles, reviews, proceedings papers and meeting abstracts, book chapters and editorial materials on OEC transplantation for SCI were included. Simultaneously, unpublished literature and literature for which manual information retrieval was required were excluded. MAIN OUTCOME MEASURES： All selected literatures addressing OEC transplantation for SCI were evaluated in the following aspects： publication year, document type, language, author, institution, times cited, Web of Science category, core source title, countries/territories and funding agency.RESULTS： In the Web of Science published by the Institute for Scientific Information, the earliest literature record was in April, 1995. Four hundred and fourteen publications addressing OEC transplantation for SCI were added to the data library in the past 18 years, with an annually increasing trend. Of 415 records, 405 publications were in English. Two hundred and fifty-nine articles ranked first in the distribution of document type, followed by 141 reviews. Thirty articles and 20 reviews, cited more than 55 times by the date the publication data were downloaded by us, can be regarded as the most classical references. The journal Experimental Neurology published the most literature （32 records）, followed by Glia. The United States had the most literature, followed by China. In addition, Yale University was the most productive institution in the world, while The Second Military Medical University contributed the most in China. The journal Experimental Neurology published the most OEC transplantation literature in the United States, while Neural Regeneration Research published the most in China. CONCLUSION： This analysis provides insight into the current state and trends in OEC transplantation for SCI research. Furthermore, we anticipate that this analysis will help encourage international cooperation and teamwork on OEC transplantation for SCI to facilitate the development of more effective treatments for SCI.

Olfactory ensheathing cell transplantation and inhibition of NogoA,NgR and RhoA expression in the damaged zone to ameliorate spinal cord injury

BACKGROUND： Olfactory ensheathing cell （OEC） transplantation promotes repair of spinal cord injury. Neural regeneration inhibits binding of the myelin protein Nogo to its receptor （NgR）, activates downstream inhibitory signal RhoA, and leads to axonal degeneration. OBJECTIVE： To determine the relationship between OECs transplantation for spinal cord injury and NogoA, NgR, and RhoA protein expression in the damaged zone. DESIGN, TIME AND SETTING： A randomized, controlled, animal experiment was performed from September 2006 to May 2007 at the Key Laboratory of Environment and Genes in Xi＇an Jiaotong University School of Medicine, China. MATERIALS： OECs were harvested from healthy, adult, male, Sprague Dawley rats aged 6 months. Mouse anti-rat NogoA, NgR, and RheA monoclonal antibodies were utilized for detection. METHODS： A total of 40 adult Sprague Dawley rats were randomly assigned to four groups： normal, model, OECs, and DF12, with 10 animals in each group. Transverse section spinal cord injury was established in the OECs and DF12 groups, followed by injection of 1μL OECs suspension （1×10^8/mL） or equivalent DF12 medium at 1 mm above and below the injury site. MAIN OUTCOME MEASURES： Immunohistochemistry and Western blot were utilized to detect NogoA, NgR, and RhoA expression in the spinal cord injury lesions. Morphological changes were observed by argyrophilia staining, and lower extremity function of the animals was assessed using Basso, Beattie, and Bresnahan scores. RESULTS： Eight weeks following OECs transplantation, a significant increase in new axons was observed in the OECs group, and nerve fibers crossed the injury site to repair spinal cord injury. Qualitative and quantitative results from the OECs group were superior to the model and DF12 groups. At 8 weeks after transplantation, Basso, Beattie, and Bresnahan scores were significantly greater in the OECs group compared with the model and DF12 groups （P 〈 0.01）, but expression of NogoA, NgR, and RhoA protein was significantly decreased compared with the model and DF12 groups （P〈 0.05）. CONCLUSION： OEC transplantation could inhibit NogoA, NgR, and RhoA expression in spinal cord injury lesions, thereby promoting repair of spinal cord injury.

Morphological properties and proliferation analysis of olfactory ensheathing cells seeded onto three-dimensional collagen-heparan sulfate biological scaffolds

This study aimed to examine the differences in the morphological properties and proliferation of olfactory ensheathing cells in three-dimensional culture on collagen-heparan sulfate biological scaffolds and in two-dimensional culture on common flat culture plates. The proliferation rate of olfactory ensheathing cells in three-dimensional culture was higher than that in two-dimensional culture, as detected by an M-I-r assay. In addition, more than half of the olfactory ensheathing cells subcultured using the trypsinization method in three-dimensional culture displayed a spindly Schwann cell-like morphology with extremely long processes, while they showed a flat astrocyte-like morphology in two-dimensional culture. Moreover, spindle-shaped olfactory ensheathing cells tended to adopt an elongated bipolar morphology under both culture conditions. Experimental findings indicate that the morphological properties and proliferation of olfactory ensheathing cells in three-dimensional culture on collagen-heparan sulfate biological scaffolds are better than those in two-dimensional culture.

Olfactory ensheathing cells for spinal cord repair: crucial differences between subpopulations of the glia

OECs for spinal cord repair： Is repairing the iniured spinal cord by olfactory ensheathing cell （OEC） transplantation pos- sible？ A recent human trial in which a paralysed man regained some function after transplantation of partially purified OECs suggests that this therapy may be a successful approach （Ta- bakow et al., 2014）. In another human trial in which olfactory mucosa lamina propria was transplanted, patients recovered some motor and sensory function （Wang et al., 2015）. While these results show promise, it is clear that improvements are needed to provide patients with increased functional output. Strategies to improve the therapeutic use of OECs may include improving the purification of the OECs used for transplantation, using them in combination with growth factors to combat the inhibitory environment and improve anon growth, the use of nerve bridges, advanced physiotherapy and the use of exo- skeleton robotics to reinforce functional connections. Of all these approaches, it is probably is primarily addressed to ensure crucial that the purity of OECs consistency in outcomes.

Simplified methods to isolate,culture and purify olfactory ensheathing cells

Conventional methods for harvesting, culturing and purifying olfactory ensheathing cells are complicated, time-consuming, and poorly reproducible. Olfactory bulbs were detached from adult Sprague Dawley rats and olfactory ensheathing cells were isolated using shearing, dispersion processes. After the primary cultures reached confluence, the cells were purified using a three-step process. The olfactory ensheathing cells attached and grew rapidly. The purity of the olfactory ensheathing cells increased following the three purification steps, eventually exceeding 95%. These cells could be maintained for an extended period time in culture. This simple, inexpensive, reproducible method of harvesting, culturing and purifying olfactory ensheathing cells shortens the culture cycle and provides sufficient olfactory ensheathing cells of controllable purity.

Gene and protein expression profiles of olfactory ensheathing cells from olfactory bulb versus olfactory mucosa

Olfactory ensheathing cells(OECs) from the olfactory bulb(OB) and the olfactory mucosa(OM) have the capacity to repair nerve injury. However, the difference in the therapeutic effect between OB-derived OECs and OM-derived OECs remains unclear. In this study, we extracted OECs from OB and OM and compared the gene and protein expression profiles of the cells using transcriptomics and non-quantitative proteomics techniques. The results revealed that both OB-derived OECs and OM-derived OECs highly expressed genes and proteins that regulate cell growth, proliferation, apoptosis and vascular endothelial cell regeneration. The differentially expressed genes and proteins of OB-derived OECs play a key role in regulation of nerve regeneration and axon regeneration and extension, transmission of nerve impulses and response to axon injury. The differentially expressed genes and proteins of OM-derived OECs mainly participate in the positive regulation of inflammatory response, defense response, cytokine binding, cell migration and wound healing. These findings suggest that differentially expressed genes and proteins may explain why OB-derived OECs and OM-derived OECs exhibit different therapeutic roles. This study was approved by the Animal Ethics Committee of the General Hospital of Ningxia Medical University(approval No. 2017-073) on February 13, 2017.

Effects of olfactory ensheathing cells on the proliferation and differentiation of neural stem cells

BACKGROUND： Olfactory ensheathing cells can promote oriented differentiation and proliferation of neural stem cells by cell-secreted neural factors. OBJECTIVE： To observe the effect of olfactory ensheathing cells on the differentiation and proliferation of neural stem cells. DESIGN, TIME AND SETTING： Cytology was performed at the Department of Neurology, Tongji Medical College, Huazhong University of Science and Technology, China, from September 2007 to October 2008. MATERIALS： Mouse anti-nestin polyclonal antibody （Chemicon, USA）, mouse anti-glial fibrillary acidic protein （GFAP） IgG1, mouse anti-2＇, 3＇-cyclic nucleotide 3＇-phosphodiesterase （CNPase） IgG1, mouse anti-Tubulin Class-Ill IgG1 （Neo Markers, USA）, Avidin-labeled Cy3 （KPL, USA）, and goat anti-mouse IgGl： fluorescein isothiocyanate （FITC） （Serotec, UK） were used in this study. METHODS：Tissues were isolated from the embryonic olfactory bulb and subependymal region of Wistar rats. Serum-free DMEM/F12 culture media was used for co-culture experiments. Neural stem cells were incubated in serum-free or 5% fetal bovine serum-containing DMEM/F12 as controls. MAIN OUTCOME MEASURES： After 7 days of co-culture, neural stem cells and olfactory ensheathing cells underwent immunofluorescent staining for nestin, tubulin, glial fibrillary acidic protein, and CNPase. RESULTS： Olfactory ensheathing cells promoted proliferation and differentiation of neural stem cells into neuron-like cells, astrocytes and oligodendrocytes. The proportion of neuron-like cells was 78.2%, but the proportion of neurons in 5% fetal bovine serum DMEM/F12 was 48.3%. In the serum-free DMEM/F12, neural stem cells contracted, unevenly adhered to the glassware wall, or underwent apoptosis at 7 days. CONCLUSION： Olfactory ensheathing cells promote differentiation of neural stem cells mainly into neuron-like cells, and accelerate proliferation of neural stem cells. The outcome is better compared with serum-free medium or medium containing 5% fetal bovine serum.

Effect of olfactory ensheathing cell transplantation on myelin repair and motor function in a rat model of experimental allergic encephalomyelitis

BACKGROUND： Olfactory ensheathing cell transplantation can activate axonal regeneration and enhance myelin repair, which are beneficial for treating demyelinating diseases. OBJECTIVE： To explore the effects of olfactory ensheathing cell transplantation on myelin repair, synaptophysin expression, and motor function in a rat model of experimental allergic encephalomyelitis. DESIGN, TIME AND SETTING： A randomized, controlled experiment was performed at the Laboratory of Provincial Hospital affiliated to Shandong University between August 2006 and September 2007. MATERIALS： Dibenzylamine （Hoechst 33342）, luxol fast blue, and rabbit anti-rat synaptophysin antibody were provided by Sigma, USA. METHODS： Olfactory ensheatbing cells extracted from neonatal Wistar rats were cultured for 10-14 days and labeled with dibenzylamine. Spinal cord extracted from a healthy guinea pig was homogenized and equally mixed with complete Freund＇s adjuvant; thereafter, the mixture was intracutaneously injected into two posterior voix pedis of healthy male Wistar rats to establish models of experimental allergic encephalomyelitis. Rats were randomly divided into a control encephalomyelitis group and an olfactory ensheathing cell transplantation group, 36 rats in each group. Physiological saline （2 μ L） or an olfactory ensheathing cell suspension （2 μ L） was separately injected along lateral cerebral ventricle at day 7 post-model induction. MAIN OUTCOME MEASURES： The migration and distribution of olfactory ensheathing cells were observed under fluorescence microscopy; myelin repair was detected using hematoxylin-eosin staining and luxol fast blue staining; synaptophysin expression was measured using immunohistochemical staining; motor function was evaluated using a motor function scale. RESULTS： Olfactory ensheatbing cells could survive in vivo and migrate to the distal end of the transplant focus and spinal cord, and survived 21 days. Hematoxylin-eosin staining and luxol fast blue staining indicated that myelin in the transplantation group was intact, and the inflammatory focus gradually disappeared. Transplantation increased synaptophysin expression （P 〈 0.05 versus control） and motor function （P 〈 0.05）. CONCLUSION： Olfactory ensheathing cell transplantation can promote myelin repair, increase synaptophysin protein expression, and ameliorate motor function in a rat model of experimental allergic encephalomyelitis.

Nogo-A expression in injured spinal cord following human olfactory mucosa-derived olfactory ensheathing cells transplantation

Transplantation of olfactory bulb-derived olfactory ensheathing cells （OECs） promotes motor functional recovery in rats with acute spinal cord injury, possibly by Nogo-A expression changes at the injury site. The present study transplanted OECs derived from the olfactory mucosa （OM） of rats OM-derived OEC （OM-OEC） transplantation significantly reduced the increase of Nogo-A protein and mRNA expression caused by spinal cord injury, supporting the hypothesis that OM-OECs improve spinal cord regeneration by reducing Nogo-A expression.

Immunity phenomena following olfactory ensheathing cell transplantation into experimental allergic encephalomyelitis rat brain

Olfactory ensheathing cells （OECs） can promote axonal regeneration and remyelination for the treatment of spinal cord injury. OECs can also treat experimental allergic encephalomyelitis （EAE）, but it remains unclear whether OECs might be rejected by the immune system in the brain including the destruction of the blood-brain barrier under inflammation, the release of inflammatory factors, the activation of local antigen-presenting cells （e.g., microglia cells） and antigen drainage. We found that OECs expressed major histocompatibility complex （MHC）-I molecules on the cell surface, barely expressed MHC-II, but MHC-II could be induced by interferon-v, suggesting that OECs have certain immunogenicity. When OECs were transplanted into normal animal brains, no OECs were phagocytosed by dendritic cells in the cervical lymph node, and OECs did not induce lymphocyte proliferation, which indicates that OECs share some immune privilege under normal conditions. However, OECs in the rat EAE brain were phagocytosed by dendritic cells in the cervical lymph node and enhanced lymphocyte proliferation. These findings suggest that OECs are rejected because of increased immunogenicity in EAE brain, and that brain inflammation, in particular activated dendritic cells, may be a prerequisite for rejecting OECs.

Bilateral olfactory ensheathing cell transplantation promotes neurological function in a rat model of cerebral infarction

In the present study, olfactory ensheathing cells were transplanted into the cortices of infarcted （infarct transplantation group）, normal （normal transplantation group）, and bilateral hemispheres （bilateral transplantation group）. Olfactory ensheathing cells migrated to the infarct focus. The number of growth associated protein 43-positive cells and nerve fibers was slightly increased in the infarct area. These changes were more evident in the bilateral cortical transplantation group. Results demonstrated that transplanted olfactory ensheathing cells can migrate in rats with cerebra infarction. The olfactory ensheathing cells on the normal side can also promote neurological function. Bilateral cortical transplantation exhibited superior effects over unilateral transplantation.

Microencapsulation improves inhibitory effects of transplanted olfactory ensheathing cells on pain after sciatic nerve injury

Olfactory bulb tissue transplantation inhibits P2X2/3 receptor-mediated neuropathic pain. However, the olfactory bulb has a complex cellular composition, and the mechanism underlying the action of purified transplanted olfactory ensheathing cells（OECs） remains unclear. In the present study, we microencapsulated OECs in alginic acid, and transplanted free and microencapsulated OECs into the region surrounding the injured sciatic nerve in rat models of chronic constriction injury. We assessed mechanical nociception in the rat models 7 and 14 days after surgery by measuring paw withdrawal threshold, and examined P2X2/3 receptor expression in L4–5 dorsal root ganglia using immunohistochemistry. Rats that received free and microencapsulated OEC transplants showed greater withdrawal thresholds than untreated model rats, and weaker P2X2/3 receptor immunoreactivity in dorsal root ganglia. At 14 days, paw withdrawal threshold was much higher in the microencapsulated OEC-treated animals. Our results confirm that microencapsulated OEC transplantation suppresses P2X2/3 receptor expression in L4–5 dorsal root ganglia in rat models of neuropathic pain and reduces allodynia, and also suggest that transplantation of microencapsulated OECs is more effective than transplantation of free OECs for the treatment of neuropathic pain.

Animal experiments and clinical application of olfactory ensheathing cell transplantation for treatment of spinal cord injury

BACKGROUND： The olfactory epithelium can still generate new neurons after arresting its growth and development in the human body. Axons can still be generated and pass through peripheral tissue to reach the olfactory bulb. Thus, olfactory cells have been widely used in the repair of spinal cord injury. OBJECTIVE： Using animal experiments in conjunction with a clinical study of olfactory ensheathing cells, this paper was designed to clarify the function and application prospects of olfactory ensheathing cells, as well as the existing problems with their application. RETRIEVAL STRATEGY： Using the terms ＂olfactory ensheathing cells, spinal cord injury＂, we retrieved manuscripts published from January 1990 to June 2007. The languages were limited to English and Chinese. Inclusion criteria： studies addressing the characteristics, basic study, clinical application and prospects of olfactory ensheathing cells; studies that were recently published or were published in high-impact journals. Exclusion criteria： repetitive studies. LITERATURE EVALUATION： The included 29 manuscripts were primarily clinical or basic experimental studies. DATA SYNTHESIS： Following spinal cord injury, spinal neurons die, neurotrophic factors are lacking, and the existing glial scar and cavities hinder axonal growth. One method to repair spinal cord injury is to interfere with the above-mentioned factors based on animal experiments. Myelination and axonal regeneration are the keys to spinal cord injury therapy. Olfactory ensheathing cells can secrete several neurotrophic factors, inhibit horizontal cell reactions, have noticeable neuroprotective effects, and possess a very strong reproductive activity, so they have many advantages in the fields of cell transplantation and gene therapy. However, there still exist many questions and uncertainties, such as the best time window and dose, as well as complications of olfactory ensheathing cell transplantation; precise mechanism of action after olfactory ensheathing cell transplantation; the number and length of growing axons as well as whether axonal growth is consistent with spinal cord recovery after olfactory ensheathing cell transplantation; and the long-term curative effect of olfactory ensheathing cells transplanted into spinal cord injury patients. CONCLUSION： Both animal experiments and clinical application have demonstrated that olfactory ensheathing cell transplantation helps spinal neurofunctional recovery, but the mechanism of action requires further investigation.

Culture and purification of human fetal olfactory ensheathing cells using different attachment rates combined with intermittent NT3 nutrition

Objective：To explore a simple and pragmatic method to obtain sufficient olfactory ensheathing cells from human fetus by selective attachment of harvested cells combined with intermittent NT3 nutrition. Methods：DMEM/F12 culture solution including 10% fetal bovine serum or NT3 was used to culture olfactory ensheathing cells intermittently every 48 h. The cell state and growth rates of OECs were observed, and P75 staining was used to estimate the purity of the cells. Results：Human fetal OECs were positive with P75 immunocytochemical staining. OECs in dipolar or tripolar shape formed networks by their processes in vitro. The purity of OECs in ＂good state＂ was about 95% at 9 d and 83% on 12 d, respectively. Conclusion：The method of using different attachment rates combined with intermittent NT3 addition is a simple and effective way to culture and purify OECs.

Semaphorin 3A expression in spinal cord injured rats after olfactory ensheathing cell transplantation

Semaphorin 3A expression is thought to increase following spinal cord injury.The impact of olfactory ensheathing cell transplantation remains unclear.The current study demonstrated that spinal cord hemorrhage,edema,degeneration,necrosis,cyst formation,proliferation of glial cells,regeneration of nerve fibers and various pathological reactions occurred following a simple cross-section of spinal cord injury.Transplantation of olfactory ensheathing cells was found to significantly relieve the pathological reactions in the spinal cord described above,decrease the extent of necrosis in damaged neurons and nerve fibers,and downregulate semaphorin 3A expression in the injured zone.The results confirmed that olfactory ensheathing cell transplantation plays a protective role on the injured spinal cord by reducing the expression of semaphorin 3A.

Expression and biological activity of double replica retrovirus carrier-mediated neurotrophin-3 in olfactory ensheathing cells

BACKGROUND： Previous studies have demonstrated that the combination of olfactory ensheathing cells （OECs） and neurotrophic factor-3 （NT-3） in the rat lateral ventricle can promote nerve axonal regeneration and myelin sheath repair. However, this effect remains very short-lived. OBJECTIVE： To transfect NT-3 into OECs and to observe the biological activity of OEC-expressing NT-3. DESIGN, TIME AND SETTING： This genetic engineering, in vitro experiment was performed in the Provincial Hospital Affiliated to Shandong University between January 2007 and October 2008. MATERIALS： Trizol Reagent kit was purchased from Gibco, USA; reverse transcription kit, NT-3 Emax ImmunoAssay System reagent was purchased from Promega, USA. METHODS： Neonatal Wistar rat OECs were established as primary cultures and were transfected with pN2A-NT-3 viral vector. The OECs with the highest virus titer and stable cellular growth served as the transfection group; OECs transfected with NT-3-free retrovirus carrier pN2A served as the empty vector group; un-transfected OECs served as the control group. After adherence, the logarithmically cultured PC12-TrkC cells were plated in OECs supernatant from the transfection and empty vector groups, as well as 20 μL PBS, and cultured for 4 days. MAIN OUTCOME MEASURES： NT-3 mRNA expression in OECs, fluorescence of NT-3-positive cells in the transfection group and control group; influence of OECs secreting NT-3 on the differentiation ratio of PC12-TrkC cells. RESULTS： NT-3 mRNA expression was observed 24 hours after transfection and lasted for 28 days which was greater than the control and empty vector groups （P 〈 0.01）. A large number of NT-3-positive cells were observed in the transfection group, and immunofluorescence was greater than the control and empty vector groups. PC12-TrkC cells co-cultured with OECs from the transfection group exhibited a thick and long cell process, increased cell density, and the differentiation ratio was increased （P 〈 0.01）. CONCLUSION： Recombinant double replica retrovirus NT-3 gene was stably and effectively expressed in OECs, and the expressed NT-3 possessed biological activity that promoted neuronal survival.