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Invasive aspergillosis (IA) represents a leading cause of mortality in immunocompromised patients. Although adoptive immunotherapy with Aspergillus-specific T cells (Asp-STs) represents a promising therapeutic approach against IA, the complex and costly production limits its broader application. We generated Asp-STs from a single blood draw of healthy individuals or IA patients in only 10 days, by either Aspergillus fumigatus (AF) lysate or peptide stimulation of mononuclear cells. The cells were phenotypically and functionally characterized, and safety was assessed in xenografts. Healthy donor-derived and lysate- or peptide-pulsed Asp-STs presented comparable fold expansion, immunophenotype, and Th1 responses. Upon cross-stimulation, only the lysate-pulsed Asp-STs were empowered to respond to peptide stimulation, although both cell products induced hyphal damage. Importantly, Asp-STs cross-reacted with other fungal species and did not induce alloreactivity in vivo. IA patient-derived T cells displayed an anergic phenotype that prohibited sufficient expansion and yield of meaningful doses of Asp-STs for autologous immunotherapy. Using a rapid and simple process, we generated, from healthy donors but not IA patients, functionally active Asp-STs of broad specificity and at clinically relevant numbers. Such an approach may form the basis for the effective management of IA in the context of allogeneic hematopoietic cell transplantation.

Endobronchial ultrasound endoscopy is a state of the art diagnostic endoscopic procedure for the thorax. Firstly it was designed mainly for the staging of lung cancer and of course for the diagnosis of suspicious findings in large central airways. The main limitation of the equipment is the diameter of the instrument and therefore it can only be guided through large airways. However; the diameter of the working channel also provides a large tissue sample nowadays with the 19G biopsy needle. We will provide our experience with the 22G needle of the endobronchial convex-probe in several medical situations of the thorax.

The integrity of buildings and the safety of their occupants can be catastrophically affected by the action of contact or near- eld explosions. Numerical simulation studies should be carried out in order to better understand the response of reinforced concrete (RC) structures to such dynamic impulsive actions. When concrete structural elements are subjected to near- eld blast loads, large deformations, rate effects and overloading render the modelling of concrete extremely complex. Significant research has been done since September 11, 2001, in order to improve the computational abilities, to understand the complex non-linear behaviour of concrete under extreme loading conditions and to obtain reliable numerical results. Simulations can be done using ‘user-developed’ codes or commercial hydrocodes. One of the most common hydrocode is LS-DYNA, where concrete can be treated as a homogeneous material. In this particular project LS-DYNA is used along with the Winfrith Concrete Model to simulate the response of a two-layer RC slab to several contact detonations (using C4, Composition-4). The aim is to achieve a contact explosion between the target (slab) and the explosive since literature can provide us with only limited information regarding this. The slab and the high explosive are explicitly modelled using the multi-material arbitrary Lagrangian–Eulerian approach (MM-ALE). The obtained numerical results are validated using experimental data. The aforementioned numerical results of the damage are in favourable agreement with the experimental data, with average errors lying under 10%. In addition, it was observed that the increase in the explosive mass led to a change of the damage modefrom penetration to perforation. Concrete’s crack pattern was also investigated.

We present a case of a 56-year-old female, with a familial history of breast, lung and brain cancer, which revealed a breast tumor, located in the upper outer quadrant of the left breast. During the routinely staging examinations, a 15 cm intrathoracic tumor was found in the upper left mediastinum, penetrating the pericardium and a smaller tumor, in the left side of parietal pleura. Core biopsies from both lesions, revealed a lobular carcinoma of the breast classic type, grade II (e-cadherin-, ER+, PR+, Her-2-, Ki-67 10%) and a B3 thymoma (CK19+, CD5+) penetrating the pericardium and the left lung. A synchronous removal of both tumors was scheduled, including median sternotomy and left intrapericardial pneumonectomy, followed by a modified radical left mastectomy and a sentinel lymph node biopsy. The postoperative course was uneventful. This case advocates that thymoma patients appear to have a predisposition towards developing additional neoplasms, as breast carcinoma. Clinicians should be aware of the increased incidence of extrathymic cancers, occurring in thymoma patients.

Mice, in contrast to other species, have the ability to recover after severe spinal cord damage without any therapeutic intervention. We studied the development of Wallerian degeneration of the long spinal tracts and also the contribution of proliferative neuronal and glial cells after complete spinal cord transaction including the dura matter in C57bl/6 mice. In addition, we monitored the functional recovery in association with histological and immunohistochemical observations. Critical point of time after SCT was the second week, when the lesion reached its maximum size, was well-vascularized and begun to self-limit drastically, thereafter. However, slight degree of functional recovery began from 48 hours after spinal cord transection. Immunophenotype analysis of the proliferative cells revealed that endogenous neurogenesis was of the glial, and not of the neuronal cell-type proliferation. Glial cells seemed to almost completely replace cellular loss, to remyelinate spared axons, to restore blood barrier and to support the rest, undamaged, neurons. Intense proliferation of glial cells occurred in the first 48 hours after injury and began to decline thereafter. Consequently, therapeutic interventions aiming at stimulation of endogenous neurogenesis should be initiated within the first hours after injury, while new approaches should target at stimulating not only the glial but also the endogenous neuronal cell-type proliferation.

Granulocyte colony-stimulating factor (G-CSF)-mobilized blood stem cells may become the preferable source of hematopoietic stem cells (HSCs) for gene therapy because of the higher yield of cells compared with conventional bone marrow harvesting. A G-CSF-associated risk of splenic rupture has been recognized in normal donors of HSCs, but limited information is available about the G-CSF effect in the presence of splenomegaly and extramedullary hematopoiesis. We investigated the G-CSF effect in a thalassemic mouse model (HBB(th-3)) as compared with a normal strain (C57BL/6), in terms of safety, mobilization efficacy, and distribution of stem cells among hematopoietic compartments. There was no death or clinical sequelae of splenic rupture in G-CSF-treated animals of either strain; however, hemorrhagic infarcts in the spleen were detected with low frequency in G-CSF-treated HBB(th-3) mice (12.5%). HBB(th-3) mice mobilized less effectively than C57BL/6 mice (Lin(-)Sca-1(+)c-Kit(+) cells/microl of peripheral blood mononuclear cells [PBMCs]: 90 +/- 55 vs. 255 +/- 174, respectively, p = 0.01; CFU-GM/ml PBMCs: 390 +/- 262 vs. 1131 +/- 875, p = 0.01) because of increased splenic trapping of hematopoietic stem and progenitor cells (Lin(-)Sca-1(+)c-Kit(+) cells per spleen (x10(5)): 487 +/- 35 vs. 109 +/- 19.6, p = 0.01; CFU-GM per spleen (x10(2)): 1470 +/- 347 vs. 530 +/- 425, p = 0.0006). Splenectomy restored the mobilization proficiency of thalassemic mice at comparable levels to normal mice and resulted in the development of a hematopoietic compensatory mechanism in the thalassemic liver that protected splenectomized mice from severe anemia. Our data imply that, in view of human gene therapy for thalassemia, either multiple cycles or alternative ways of mobilization may be required for a sufficient yield of transplantable HSCs. In addition, strategies to minimize the risk of G-CSF-induced splenic infarcts should be explored in a clinical setting.

Objective On the basis of the recently recognized potential of bone marrow (BM) cells to give rise to hepatocytes, we investigated the possibility that granulocyte colony-stimulating factor (G-CSF)–mobilized BM cells could home to the injured liver and promote tissue repair. We also examined the origin of cells (endogenous or BM) reconstituting liver after damage. Methods Acute and chronic liver injury models were generated by injecting CCl4 in C57Bl6 mice and G-CSF was administered in hematopoietic stem cell (HSC) mobilization doses. After sex-mismatched BM transplantation into lethally irradiated recipients and treatment with CCl4 ± G-CSF, sry (sex-determining region for Y chromosome) protein was detected by immunohistochemistry in liver sections. Double immunohistochemistry for sry and ki-67 protein was used to define the origin of proliferating cells reconstituting liver after injury. Results In both acute and chronic liver injury model, G-CSF administration ameliorated the histological damage and accelerated the regeneration process. This was accompanied by a strong survival benefit in G-CSF–treated group vs CCl4 group. Quantitative analysis showed higher percentage of BM-origin hepatocytes in the CCl4 + G-CSF group compared with the CCl4 group, although the liver engraftment rate still remained rather low. Double staining for ki-67 and sry demonstrated that the recovery acceleration after chemical injury and G-CSF treatment was mainly mediated by increased proliferation of host hepatocytes (ki-67+/sry−) with less support from BM-origin cells (ki-67+/sry+). Conclusion G-CSF treatment significantly improved survival and liver histology in chemically injured mice, predominantly by promoting endogenous repair mechanisms. Therefore, mobilization with G-CSF might offer a novel therapeutic approach for the treatment of acute and chronic liver diseases in humans.

High numbers of genetically modified hematopoietic stem cells (HSCs) equipped with enhanced engrafting potential are required for successful stem cell gene therapy. By using thalassemia as a model, we investigated the functional properties of hematopoietic stem and progenitor cells (HSPCs) from Hbb(th3)/45.2(+) mice after mobilization with G-CSF, plerixafor, or G-CSF+plerixafor and the engraftment kinetics of primed cells after competitive primary and noncompetitive secondary transplantation. G-CSF+plerixafor yielded the highest numbers of HSPCs, while G-CSF+plerixafor-mobilized Hbb(th3)/45.2(+) cells, either unmanipulated or transduced with a reporter vector, achieved faster hematologic reconstitution and higher levels of donor chimerism over all other types of mobilized cells, after competitive transplantation to B6.BoyJ/45.1(+) recipients. The engraftment benefit observed in the G-CSF+plerixafor group was attributed to the more primitive stem cell phenotype of G-CSF+plerixafor-LSK cells, characterized by higher CD150(+)/CD48 expression. Moreover, secondary G-CSF+plerixafor recipients displayed stable or even higher chimerism levels as compared with primary engrafted mice, thus maintaining or further improving engraftment levels over G-CSF- or plerixafor-secondary recipients. Plerixafor-primed cells displayed the lowest competiveness over all other mobilized cells after primary or secondary transplantation, probably because of the higher frequency of more actively proliferating LK cells. Overall, the higher HSC yields, the faster hematological recovery, and the superiority in long-term engraftment indicate G-CSF+plerixafor-mobilized blood as an optimal graft source, not only for thalassemia gene therapy, but also for stem cell gene therapy applications in general.

Bone marrow (BM) could serve as a source of cells facilitating liver repopulation in case of hepatic damage. Currently available hematopoietic stem cell (HSC) mobilizing agents, were comparatively tested for healing potential in liver fibrosis. Carbon tetrachloride (CCl4)-injured mice previously reconstituted with Green Fluorescent Protein BM were mobilized with Granulocyte-Colony Stimulating Factor (G-CSF), Plerixafor or G-CSF+Plerixafor. Hepatic fibrosis, stellate cell activation and oval stem cell frequency were measured by Gomori and by immunohistochemistry for a-Smooth Muscle Actin and Cytokeratin-19, respectively. Angiogenesis was evaluated by ELISA and immunohistochemistry. Quantitative real-time PCR was used to determine the mRNA levels of liver Peroxisome Proliferator-Activated Receptor gamma (PPAR-γ), Interleukin-6 (IL-6) and Tumor Necrosis-alpha (TNFα). BM-derived cells were tracked by double immunofluorescence. The spontaneous migration of mobilized HSCs towards injured liver and its cytokine secretion profile was determined in transwell culture systems. Either single-agent mobilization or the combination of agents significantly ameliorated hepatic damage by decreasing fibrosis and restoring the abnormal vascular network in the liver of mobilized mice compared to CCl4-only mice. The degree of fibrosis reduction was similar among all mobilized mice despite that G-CSF+Plerixafor yielded significantly higher numbers of circulating HSCs over other agents. The liver homing potential of variously mobilized HSCs differed among the agents. An extended G-CSF treatment provided the highest anti-fibrotic effect over all tested modalities, induced by the proliferation of hepatic stem cells and decreased hepatic inflammation. Plerixafor-mobilized HSCs, despite their reduced liver homing potential, reversed fibrosis mainly by increasing hepatic PPAR-γ and VEGF expression. In all groups, BM-derived mature hepatocytes as well as liver-committed BM stem cells were detected only at low frequencies, further supporting the concept that alternative mechanisms rather than direct HSC effects regulate liver recovery. Overall, our data suggest that G-CSF, Plerixafor and G-CSF+Plerixafor act differentially during the wound healing process, ultimately providing a potent anti-fibrotic effect.

Objective The role of mesenchymal stem cells (MSC) in experimental arthritis is undoubtedly conflicting. This study explored the effect of bone marrow-derived MSC in previously untested and pathogenetically different models of rheumatoid arthritis (RA). Methods MSC were tested both in an induced (adjuvant-induced) and a spontaneous (K/BxN) arthritis model. Arthritis was assessed clinically and histologically. The proliferation of splenocytes and fibroblast-like synoviocytes (FLS) in the presence of MSC was measured by radioactivity incorporation. Toll-like receptor (TLR) expression was measured by real-time PCR. T-regulatory cell (Treg) frequency, T-cell apoptosis and cytokine secretion were monitored by flow cytometry. Results MSC, in vitro, strongly inhibited critical cell populations; splenocytes and FLS. In contrast, MSC proved ineffective in vivo, unless they were administered before disease onset, an effect implying that the inflammatory arthritic milieu potentially abrogates MSC immunomodulatory properties. In order to alleviate inflammation before MSC infusion, the authors administered, at arthritis onset, a short course with a proteasome inhibitor, bortezomib, whereas MSC were infused when established disease was expected. The bortezomib plus MSC group demonstrated a significantly decreased arthritis score over arthritic, MSC-only, bortezomib-only groups, also confirmed by histology and immunohistochemistry. The bortezomib plus MSC combination restored TLR expression and Treg frequency in blood and normalised FLS and splenocyte proliferation, apoptosis and cytokine secretion. Conclusion MSC lose their immunomodulatory properties when infused in the inflammatory micromilieu of autoimmune arthritis. Conditioning of the recipient with bortezomib alters the disease microenvironment enabling MSC to modulate arthritis. Should milieu limitations also operate in human disease, this approach could serve as a strategy to treat RA by MSC.

Objective. To explore the effect of bortezomib in splenocytes and fibroblast-like synoviocytes (FLS) and its in vivo potency in a rat model of adjuvant-induced arthritis (AIA), which resembles human rheumatoid arthritis (RA). Methods. AIA was induced with Freund’s complete adjuvant. Splenocyte and FLS proliferation and apoptosis were measured by radioactivity incorporation and flow cytometry, respectively. The invasiveness of FLS from rats with AIA was tested in a Transwell system. The pattern of cytokine secretion was evaluated by cytometric bead array in splenocyte supernatants. Bortezomib was administered prophylactically or therapeutically, and arthritis was assessed clinically and histologically. Immunohistochemistry was performed for markers of inflammation and angiogenesis in joints. Hematologic and biochemical parameters were tested in peripheral blood (PB). Representative animals were examined by computed tomography (CT) scanning before and after bortezomib administration. The expression of Toll-like receptor 2 (TLR-2), TLR-3, and TLR-4 in PB and FLS was measured by real-time polymerase chain reaction, and alterations in specific cell populations in PB and spleen were determined by flow cytometry. Results. In vitro, bortezomib exhibited significant inhibitory and proapoptotic activity in splenocytes and FLS from rats with AIA, altered the inflammatory cytokine pattern, and reduced the invasiveness of FLS from rats with AIA. In vivo, bortezomib significantly ameliorated disease severity. Remission was associated with improved histology and decreased expression of CD3, CD79a, CD11b, cyclooxygenase 1, and factor VIII in target tissues as well as down-regulation of TLR expression in PB and cultured FLS. CT scanning demonstrated a bone healing effect after treatment. Conclusion. Our findings suggest that bortezomib affects AIA in a pleiotropic manner and that this drug may be effective in RA.

For end-stage liver disease, liver transplantation provides the only definite cure; however, many patients die while in the waiting list for donation. Various stem cell populations have been described to contribute to liver regeneration and there is accumulating evidence for the participation of hematopoietic stem cells (HSCs) in this process. We here report two cases treated with boost infusions of autologous mobilized HSCs to regenerate cirrhotic liver. The procedure was safe and well tolerated. Both patients showed a lasting amelioration in the clinical course of the disease during the 30 months of follow-up. These results suggest that the procedure may be considered in a clinical study setting as a bridging therapy until organ transplantation becomes available or to reverse a decompensated cirrhosis to a compensated one. Additional studies in this area will be required to document efficacy and evaluate toxicity.

Cyclin D1 expression was evaluated by immunohistochemical analysis and biotin-labeled in situ hybridization (ISH) in a series of 71 decalcified, paraffin-embedded bone marrow biopsy specimens from patients with multiple myeloma (MM). Cyclin D1 messenger RNA (mRNA) overexpression was detected by ISH in 23 (32%) of 71 cases, whereas cyclin D1 protein was identified by immunohistochemical analysis in 17 (24%) of 71 specimens. All cases that were positive by immunohistochemical analysis also were positive by ISH. Statistically significant associations were found between cyclin D1 overexpression and grade of plasma cell differentiation and between cyclin D1 overexpression and extent of bone marrow infiltration. Our findings demonstrate the following: (1) ISH for cyclin D1 mRNA is a sensitive method for the evaluation of cyclin D1 overexpression in paraffin-embedded bone marrow biopsy specimens with MM. (2) ISH is more sensitive than immunohistochemical analysis in the assessment of cyclin D1 expression. (3) Cyclin D1 overexpression in MM is correlated positively with higher histologic grade and stage.

Mantle cell lymphoma (MCL) is characterized by the chromosomal translocation t(11;14), which involves rearrangement of the bcl-1 proto-oncogene to the immunoglobulin heavy chain gene and results in overexpression of cyclin D1 mRNA. In this study, we evaluated the diagnostic relevance of three methods that may be helpful in the diagnosis of MCL: in situ hybridization (ISH) and a stringent reverse transcriptase-polymerase chain reaction (RT-PCR) protocol for cyclin D1 mRNA, and immunohistochemistry for cyclin D1 protein. The study group included 37 paraffin-embedded specimens (25 from lymph nodes and 12 from extranodal tissues) from 30 patients. MCL diagnosis was performed according to the Revised European-American Classification of Lymphoid Neoplasms. Twenty-nine patients with non-MCL lymphoproliferative disorders comprised the control group. Biotin-labeled ISH was performed in 28 cases of MCL, 24 (86%) of which were found to be positive. As shown by ISH in extranodal tissues, cyclin D1 mRNA was present not only in neoplastic lymphoid cells, but in other cell types as well. For this reason, RT-PCR results were considered reliable for MCL diagnosis only on informative material (from tissues that do not normally express cyclin D1); this method was evaluated as positive in 16 of 18 (89%) MCL cases. Cyclin D1 immunopositivity was present in 20 of 29 (69%) MCL cases. No members of the control group were found to express cyclin D1 mRNA by either ISH or RT-PCR under the stringent conditions used. In conclusion, stringent RT-PCR for cyclin D1 expression can be helpful in MCL diagnosis in paraffin-embedded material from lymph nodes. ISH is a sensitive method for cyclin D1 mRNA detection; its sensitivity is superior to that of cyclin D1 immunohistochemistry and similar to that of the stringent RT-PCR used. ISH is very specific as well, clearly more specific than RT-PCR, because it allows the correlation of molecular findings with morphology. This method can be applied on all types of paraffin-embedded tissues and provides an accurate tool for MCL diagnosis.

Abstract 3234 Poster Board III-171 Gene therapy has been recently postulated as a realistic therapeutic potential for thalassemia and the mobilized autologous hematopoietic stem cells (HSCs) may represent the preferable source of stem cells for genetic modification due to the higher yield of HSCs compared to conventional bone marrow (bm) harvest. We have previously shown (manuscript under revision) that G-CSF mobilization in the HBBth-3 thalassemic mouse model is less efficient compared to normal C57Bl6 strain, mainly due to increased trapping of hematopoietic stem (Lin-sca-1+ckit+–LSK) and progenitor cells (CFU-GM) in the enlarged thalassemic spleen. The novel mobilizer, AMD3100 (plerixafor, mozobil), has been shown to reversibly bind to CXCR4 and inhibit the interaction between SDF-1 and CXCR4 within the bm microenvironment, resulting in the egress of CD34+ cells into the circulation of healthy donors and cancer patients. The addition of AMD to G-CSF results in even greater increases in circulating CD34+cells. We explored in the current study whether AMD alone or in combination with G-CSF improves the mobilization efficiency of thalassemic mice. C57 and HBBth-3 mice received G-CSF-alone at 250microgr/kgX7 days, AMD-alone at 5mg/kgX3 days or the combination of two with AMD administered in the evening of days 5-7 of G-CSF administration. Hematopoietic tissues (blood, bm, spleen) were collected and the absolute LSK and CFU-GM numbers were calculated based on their frequency within tissues (by FCM and clonogenic assays) in relation to the individual cell count per tissue. AMD-alone didn't significantly affect the HSC yield as compared to G-CSF mobilization in thal mice (LSK/μl blood: 103±85 vs 69±26 p=ns), although it significantly increased the circulating Colony Forming Cells (CFU-GM/ml blood: 1205±533 vs 330±87, p=0,05). In contrast, the AMD+G-CSF combination significantly improved the mobilization efficiency of HBBth-3 mice over the G-CSF-treated group (LSK cells/μl blood: 224±104 vs 69±26 p=0,04, CFU-GM/ml blood: 1671±984 vs 330±87 p=0,05, respectively) at levels comparable to normal mice treated with G-CSF (LSK cells/ μl blood: 241±167, CFU-GM/ml blood: 1235±1140, respectively). AMD induced a “detachment” of stem cells from the bm because reduced numbers of bm LSK cells were counted in the AMD-alone group as compared to the untreated group (LSK/2 femurs×103: 692±429 vs 1687±1016, respectively, p=0,05). This was in contrast to the marrow hyperplasia caused by G-CSF over the steady-state condition (LSK/2 femurs×103: 2684±1743 vs 1687±1016 p=0,02 / CFU-GM/2femurs:111.841±15.391 vs 76.774±31.728 p=0,01). Consequently, the combination of AMD+G-CSF resulted in increased numbers of circulating stem and progenitor cells without inducing marrow hyperplasia as compared to steady-state condition (LSK/2femurs×103: 1681±862 vs 1686±1017, p=ns / CFU-GM/2femurs: 76.774±31.728 vs 82.905±26.277, p=ns). AMD, also in contrast to G-CSF, did not cause increased trapping of stem and progenitor cells in the spleen compared to the untreated condition (LSK cells/spleen×103: 4738±2970 vs 8303±4166 p=ns / CFU-GM/spleen:146.269±93.174 vs 98.518±25.549, p=ns). However, the combination of AMD+G-CSF still resulted in splenic sequestration of progenitor cells (CFU-GM/spleen: 412.176±157.417 vs 98.518±25.549, p=0,0003) but not of LSK cells (LSK cells/spleen×103: 10.200±7.260 vs 8.300±4.166 p=ns). Overall, the combination of AMD3100+G-CSF seems to restore the less efficient mobilization in a thalassemic mouse model. This combination may prove beneficial in a GT setting for obtaining the high numbers of HSCs needed for genetic correction. In addition, the combination of AMD3100+G-CSF, by avoiding the marrow hyperplasia induced by G-CSF alone, indicates a better safety profile because it will not further burden the hyperplastic –due to the increased erythroid demand and the intramarrow destruction of erythroblasts-thalassemic bone marrow. Disclosures No relevant conflicts of interest to declare.

The recently recognized potential of mesenchymal stem cells (MSCs) to differentiate into a broad spectrum of tissues and to act as immune regulators beyond the barriers of embryonic germ layers and major histocombatibility comlex (MHC) restriction, has emerged intense research interest on their possible use in a broad spectrum of clinical entities. Although the immunoregulatory potential of MSCs has been shown to effectively control GvHD in several preclinical and clinical studies, their role in autoimmune diseases has not been extensively explored in animal models. The goal of this study was to investigate the in vitro effect of rat bone marrow-derived MSCs on cultured fibrobIast-like synoviocytes (FLS) and T-cells from the spleen after induction of adjuvant arthritis (AA) by FCA as well as their in vivo effect in a rat model of AA resembling human rheumatoid arthritis. MSCs were isolated from bone marrow and were characterized by CD45 negativity and CD54, CD29 positivity in FCM analysis. Differentiation assays were performed to confirm their adipogenic, osteogenic and chondrogenic potential. Culture of AA-FLS in the presence of supernatant from syngeneic (syng) or allogeneic (allo) MSCs at passage 2–3, reduced the AA-FLS (p

Mice, unlike rats and humans, have a self recovery mechanism of spinal cord injury. Whether the hematopoietic system is involved in this mechanism is under investigation. In this study we tested whether bone marrow cells transplanted or mobilized by a growth factor in mice with spinal cord injury, can accelerate the recovery. C57bl/6 female mice 10 to 12 weeks of age underwent spinal cord incision in an open operation. The injury was performed as a complete transection including the dura mater and the whole circumference of the cord at the T10-T11 intervertebral space with a micro scalpel (No 11). Group A mice received 200μg/kg/day G-CSF subcutaneously for 7 days, starting 24 hours after operation. Group B mice received 106 light density bone marrow cells from C576bl/6 donor mice intravenously 24 hours after operation. Control group mice received no treatment. Histological evaluation was performed at 48 hours, 1 week, 3 weeks and 5 weeks postoperatively. Paraffin embedded longitudinal samples of spinal cord were cut as serial sections. Spinal cord damage was estimated by measuring the maximum diameter of the area of axonal damage and disruption of astrocytic network using immunostaining for neurofilaments and GFAP. Antibodies against CD68 were applied to identify macrophage aggregations. All measurements were performed by morphometric photo analysis. The volume of fibroblastic infiltration was estimated using a grading system (0–7), based on Van Gieson stain for connective tissue. Functional deficits and recovery over time were evaluated by testing hind limb reflex and coordinated motor function (Kuhn and Wrathal functional tests, modified by Seki et al, 2002). All tests have been videotaped. Outcome scores at 48 hours, 1 week, 3 weeks and 5 weeks postoperatively for the control group, group A and group B mice were analyzed with the Mann-Whitney U test. 48 hours post operatively all mice in all groups were paralyzed in both hind limbs. Gradual improvement was observed in all groups. At week 3 there was a significant difference between the mean scores of functional tests for both treated groups (A and B) compared with the mean scores of the control group. Statistically significant difference (p