Miguel A. Rico

Allogeneic bone marrow stromal cell transplantation after cerebral hemorrhage achieves cell transdifferentiation and modulates endogenous neurogenesis

BACKGROUND AIMS When a severe neurologic lesion occurs as a consequence of intracerebral hemorrhage (ICH), there is no effective treatment available for improving the outcome. However, cell therapy has opened new perspectives on reducing neurologic sequels subsequent to this disease. METHODS In this study, ICH was induced by stereotactic injection of 0.5 U collagenase type IV in the striatum of adult Wistar rats, and 2 h later a group of animals (n = 48) was subjected to intracerebral injection of 2 × 10(6) allogeneic bone marrow stromal cells (BMSC), while a control group (n = 48) received saline only. Eight animals from each group were killed at 48 h, 72 h, 7 days, 14 days, 21 days and 28 days. At these time-points, endogenous neurogenesis and survival of transplanted BMSC were studied. RESULTS Our findings show that after allogeneic BMSC transplantation, donor cells can survive in the brain tissue expressing neuronal and astroglial markers. Furthermore, BMSC transplantation enhances endogenous neurogenesis and inhibits apoptosis of newborn neural cells. CONCLUSIONS Although these results should be extrapolated to human disease with caution, it is obvious that cell therapy using allogeneic BMSC transplantation offers great promise for developing novel and efficacious strategies in patients suffering ICH.

The pig model of chronic paraplegia: A challenge for experimental studies in spinal cord injury

The regenerative medicine techniques that are beginning to be applied to the nervous system have led to increased hope in the treatment of diseases that have been considered incurable and that require experimental models on which to test new therapeutic strategies. We present our experience with adult pigs (minipigs) that have undergone a traumatic spinal cord injury (SCI) experimental model, and that have been followed for 1 year. We describe the surgical aspects of our SCI model by acute compression and also describe protocols for daily care and rehabilitation that are necessary to maintain the paraplegic pigs in good health during the months following the injury. Furthermore, we provide in detail the main complications that arise with this experimental model and the treatments used to address these complications. Suitable housing conditions, daily rehabilitation and prevention of complications (i.e., taking the same care applied to patients following SCI) are essential for achieving the absence of mortality and long-term maintenance of the animals. We consider the model that is described here to be feasible and useful for preliminary testing of novel therapeutic strategies aimed at regeneration of the injured spinal cord in paraplegic patients.

Late transplantation of allogeneic bone marrow stromal cells improves neurologic deficits subsequent to intracerebral hemorrhage

BACKGROUND AIMS Stem cell therapy seems to be a promising therapeutic tool for treating central nervous system (CNS) injuries. Bone marrow stromal cell (BMSC) transplantation influences functional outcome subsequent to intracerebral hemorrhage (ICH), and enhances endogenous neurogenesis in acute condition studies. We investigated whether late administration of BMSC improves functional deficits subsequent to ICH. METHODS Experimental ICH was induced by stereotactic injection of 0.5 IU collagenase type IV in the striatum of adult female Wistar rats, and 2 months later intralesional administration of 5 × 10(6) allogeneic BMSC from male donors rats in saline (n = 10), or saline only (n = 10), was performed. In the following 6 months, functional outcome was evaluated in each animal by rotarod, modified neurologic severity score (mNSS) and video-tracking box (VTB) tests. To study the behavior of BMSC after transplantation, in situ hybridization studies were performed, with double labeling of the chromosome Y-linked SrY-gene, and neuronal nuclei (NeuN) protein or gliofibrillary acidic protein (GFAP). RESULTS The assessment test revealed significant improvements in functional outcome for the BMSC-treated animals after 2 months of follow-up. Histologic results showed that functional outcome was associated with strong reactivation of endogenous neurogenesis. Furthermore, intralesional BMSC not only integrated in the injured tissue but also showed phenotypic expression of GFAP and NeuN. CONCLUSIONS Late intracerebral transplantation of allogeneic BMSC induces functional recovery after ICH. The possibility of using this type of cell therapy to reverse the consequences of hemorrhagic stroke in humans should be considered.

An approach to personalized cell therapy in chronic complete paraplegia: The Puerta de Hierro phase I/II clinical trial.

BACKGROUND AIMS Cell transplantation in patients suffering spinal cord injury (SCI) is in its initial stages, but currently there is confusion about the results because of the disparity in the techniques used, the route of administration, and the criteria for selecting patients. METHODS We conducted a clinical trial involving 12 patients with complete and chronic paraplegia (average time of chronicity, 13.86 years; SD, 9.36). The characteristics of SCI in magnetic resonance imaging (MRI) were evaluated for a personalized local administration of expanded autologous bone marrow mesenchymal stromal cells (MSCs) supported in autologous plasma, with the number of MSCs ranging from 100 × 10(6) to 230 × 10(6). An additional 30 × 10(6) MSCs were administered 3 months later by lumbar puncture into the subarachnoid space. Outcomes were evaluated at 3, 6, 9 and 12 months after surgery through clinical, urodynamic, neurophysiological and neuroimaging studies. RESULTS Cell transplantation is a safe procedure. All patients experienced improvement, primarily in sensitivity and sphincter control. Infralesional motor activity, according to clinical and neurophysiological studies, was obtained by more than 50% of the patients. Decreases in spasms and spasticity, and improved sexual function were also common findings. Clinical improvement seems to be dose-dependent but was not influenced by the chronicity of the SCI. CONCLUSION Personalized cell therapy with MSCs is safe and leads to clear improvements in clinical aspects and quality of life for patients with complete and chronically established paraplegia.

Failure of Delayed Intravenous Administration of Bone Marrow Stromal Cells after Traumatic Brain Injury

Traumatic brain injury (TBI) is a leading cause of mortality and morbidity worldwide. Currently, there is no effective strategy to treat the functional sequelae associated with TBI. Experimental evidence shows that the intravenous administration of bone marrow stromal cells (BMSC) during the first week after TBI prevents neurological deficits, but no experimental studies have shown evidence of the effect of intravenous BMSC on chronic brain injury sequelae. Here we studied the effect of intravenous administration of BMSC on functional outcomes 2 months after experimental TBI in rats. Adult Wistar rats were subjected to weight-drop impact causing severe brain injury, and 2 months later BMSC in saline, or saline alone, was intravenously injected. All experimental groups were evaluated by means of the modified Neurological Severity Score (mNSS), and internal zone Permanence Time (izPT) after video-tracking box (VTB) analysis, over the following 2 months to test the efficacy of BMSC therapy. At the end of the study period the animals were sacrificed and their brains were studied to evaluate possible differences between groups. Two months after BMSC administration no significant differences were detected in the motor and sensory evaluation between animals treated with BMSC and controls, and no differences were detected after histological study of the brains. Our present results suggest that intravenous administration of BMSC after TBI, when the neurological deficits are well established, has no beneficial effect in neurological outcomes or on brain tissue.

Repeated subarachnoid administrations of autologous mesenchymal stromal cells supported in autologous plasma improve quality of life in patients suffering incomplete spinal cord injury

BACKGROUND AIMS Cell therapy with mesenchymal stromal cells (MSCs) offers new hope for patients suffering from spinal cord injury (SCI). METHODS Ten patients with established incomplete SCI received four subarachnoid administrations of 30 × 106 autologous bone marrow MSCs, supported in autologous plasma, at months 1, 4, 7 and 10 of the study, and were followed until the month 12. Urodynamic, neurophysiological and neuroimaging studies were performed at months 6 and 12, and compared with basal studies. RESULTS Variable improvement was found in the patients of the series. All of them showed some degree of improvement in sensitivity and motor function. Sexual function improved in two of the eight male patients. Neuropathic pain was present in four patients before treatment; it disappeared in two of them and decreased in another. Clear improvement in bladder and bowel control were found in all patients suffering previous dysfunction. Before treatment, seven patients suffered spasms, and two improved. Before cell therapy, nine patients suffered variable degree of spasticity, and 3 of them showed clear decrease at the end of follow-up. At this time, nine patients showed infra-lesional electromyographic recordings suggesting active muscle reinnervation, and eight patients showed improvement in bladder compliance. After three administrations of MSCs, mean values of brain-derived neurotrophic factor, glial-derived neurotrophic factor, ciliary neurotrophic factor, and neurotrophin 3 and 4 showed slight increases compared with basal levels, but without statistically significant difference. CONCLUSIONS Administration of repeated doses of MSCs by subarachnoid route is a well-tolerated procedure that is able to achieve progressive and significant improvement in the quality of life of patients suffering incomplete SCI.

The severity of brain damage determines bone marrow stromal cell therapy efficacy in a traumatic brain injury model.

BACKGROUND: Patients who survive traumatic brain injury (TBI) can undergo serious sensorial and motor function deficits. Once damage occurs, there is no effective treatment to bring patients to full recovery. Recent studies, however, show bone marrow stromal cells (BMSC) as a potential therapy for TBI. METHODS: This study was designed to determine whether the degree of neurologic deficits influences the efficacy of cell therapy using intracerebral transplantation of BMSC in an experimental model of chronically established TBI. Adult Wistar rats were subjected to weight-drop impact causing TBI. Two months later, the animals were classified according to levels of neurologic deficits. To achieve this, we used two different functional tests: the modified Neurologic Severity Score test and internal zone Permanence Time in Video-Tracking-Box analysis. Saline only or saline containing BMSC was injected into injured brain tissue of the animals that were classified having moderate or severe neurologic damage depending on the level of established functional deficits. All experimental groups were evaluated in the course of the following 2 months to study the efficacy of BMSC administration. The animals were then killed and their brains were studied. RESULTS: Our results showed that significant functional improvement was seen when BMSC was injected into animals with moderate brain damage, but no significant improvement was found in animals with severe functional deficits when compared with controls. CONCLUSION: These findings suggest that the severity of neurologic damage may determine the potential effect of cell therapy when applied to chronically established TBI.

Cell therapy with autologous mesenchymal stromal cells in post-traumatic syringomyelia

BACKGROUND AIMS Recently, clinical studies show that cell therapy with mesenchymal stromal cells (MSCs) improves the sequelae chronically established in paraplegic patients, being necessary to know which of them can obtain better benefit. METHODS We present here a phase 2 clinical trial that includes six paraplegic patients with post-traumatic syringomyelia who received 300 million MSCs inside the syrinx and who were followed up for 6 months. Clinical scales, urodynamic, neurophysiological, magnetic resonance (MR) and studies of ano-rectal manometry were performed to assess possible improvements. RESULTS In all the cases, MR at the end of the study showed a clear reduction of the syrinx, and, at this time, signs of improvement in the urodynamic studies were found. Moreover, four patients improved in ano-rectal manometry. Four patients improved in neurophysiological studies, with signs of improvement in evoked potentials in three patients. In the American Spinal Injury Association (ASIA) assessment, only two patients improved in sensitivity, but clinical improvement in neurogenic bowel dysfunction was observed in four patients and three patients described improvement in bladder dysfunction. Spasms reduced in two of the five patients who had them previous to cell therapy, and spasticity was improved in the other two patients. Three patients had neuropathic pain before treatment, and it was reduced or disappeared completely during the study. Only two adverse events ocurred, without relation to the cell therapy. CONCLUSIONS Cell therapy can be considered as a new alternative to the treatment of post-traumatic syringomyelia, achieving reduction of syrinx and clinical improvements in individual patients.

Intrathecal administration of autologous mesenchymal stromal cells for spinal cord injury: Safety and efficacy of the 100/3 guideline

BACKGROUND AIMS Cell therapy with autologous mesenchymal stromal cells (MSCs) in patients with spinal cord injury (SCI) is beginning, and the search for its better clinical application is an urgent need. METHODS We present a phase 2 clinical trial in patients with chronic SCI who received three intrathecal administrations of 100 x 106 MSCs and were followed for 10 months from the first administration. Efficacy analysis was performed on nine patients, and safety analysis was performed on 11 patients. Clinical scales, urodynamic, neurophysiological and neuroimaging studies were performed previous to treatment and at the end of the follow-up. RESULTS The treatment was well-tolerated, without any adverse event related to MSC administration. Patients showed variable clinical improvement in sensitivity, motor power, spasms, spasticity, neuropathic pain, sexual function or sphincter dysfunction, regardless of the level or degree of injury, age or time elapsed from the SCI. In the course of follow-up three patients, initially classified as ASIA A, B and C, changed to ASIA B, C and D, respectively. In urodynamic studies, at the end of follow-up, 66.6% of the patients showed decrease in postmicturition residue and improvement in bladder compliance. At this time, neurophysiological studies showed that 55.5% of patients improved in somatosensory or motor-evoked potentials, and that 44.4% of patients improved in voluntary muscle contraction together with infralesional active muscle reinnervation. CONCLUSIONS The present guideline for cell therapy is safe and shows efficacy in patients with SCI, mainly in recovery of sphincter dysfunction, neuropathic pain and sensitivity.