Neural stem cell-like cells derived from autologous bone mesenchymal stem cells for the treatment of patients with cerebral palsy

Abstract

Background: Stem cell therapy is a promising treatment for cerebral palsy, which refers to a category of brain diseases that are associated with chronic motor disability in children. Autologous MSCs may be a better cell source and have been studied for the treatment of cerebral palsy because of their functions in tissue repair and the regulation of immunological processes.

Methods: To assess neural stem cell-like (NSC-like) cells derived from autologous marrow mesenchymal stem cells as a novel treatment for patients with moderate-to-severe cerebral palsy, a total of 60 cerebral palsy patients were enrolled in this open-label, non-randomised, observer-blinded controlled clinical study with a 6-months follow-up. For the transplantation group, a total of 30 cerebral palsy patients received an autologous NSC-like cells transplantation (1-2 × 107 cells into the subarachnoid cavity) and rehabilitation treatments whereas 30 patients in the control group only received rehabilitation treatment.

Results: We recorded the gross motor function measurement scores, language quotients, and adverse events up to 6 months post-treatment. The gross motor function measurement scores in the transplantation group were significantly higher at month 3 (the score increase was 42.6, 95% CI: 9.8-75.3, P=.011) and month 6 (the score increase was 58.6, 95% CI: 25.8-91.4, P=.001) post-treatment compared with the baseline scores. The increase in the Gross Motor Function Measurement scores in the control group was not significant. The increases in the language quotients at months 1, 3, and 6 post-treatment were not statistically significant when compared with the baseline quotients in both groups. All the 60 patients survived, and none of the patients experienced serious adverse events or complications.

Conclusion: Our results indicated that NSC-like cells are safe and effective for the treatment of motor deficits related to cerebral palsy. Further randomised clinical trials are necessary to establish the efficacy of this procedure.

Figure 1

Study protocol. A total of 60 patients with cerebral palsy with Gross Motor Function Classification System (GMFCS) levels III-V were enrolled in this study. For the transplantation group, a total of 30 cerebral palsy patients received the autologous NSC-like cell transplantation and rehabilitation treatments whereas 30 patients in the control group only received rehabilitation treatment. The study protocol is shown. Motor ability was assessed using the Gross Motor Function Measure questionnaire-88. Language ability was measured using the Gesell questionnaire. All of the assessments were conducted by a specifically assigned person, and the assessment results were recorded in the case report. A safety evaluation was performed during the 6-month follow-up to detect instances of death, any serious clinical events, abnormal electroencephalogram results, and neuroimaging complications.

Figure 2

MSCs can be induced to differentiate into NSC-like cells and exhibit multipotency by generating progenies of different neural lineages in vitro. (A) During differentiation, MSCs develop the properties of neural stem cells (NSCs). The induced cells were observed in the bright fields after 0.5 days and after 14 days. The growth curves of the MSCs and the NSC-like cells, which were obtained by enumerating the cells at each time point under a haemocytometer, are shown (n=3; the calculated doubling times were 20 hours and 40 hours, respectively). (B) After induction, the NSC-like cells were fixed, and immunofluorescence was used to detect the expression of Nestin and Tuj-1. The cells were counterstained using DAPI. The scale bar is 10 μm. To assess the induction efficiency, the cells were co-labelled with antibodies against Nestin and Tuj-1, Sox-2 or Sox1, and the proportions of double-positive cells were 66.83%, 90.92% and 64.71%, respectively. The tests were repeated three times. (C) To determine whether the induced NSC-like cells could differentiate into both neurons and glia cells, MSCs were induced for 14 days under the same conditions and stained using antibodies against neuron- and glial cell-specific proteins. The scale bar is 10 μm.

Figure 3

A comparison of the baseline characteristics of the patients in the two groups. (A) A comparison of the number of patients according to the Gross Motor Function Classification System (GMFCS) levels in the transplantation (Trans) and control groups. Statistically significant differences were detected using the chi-squared test. (B-D): A comparison of the mean ages (B), mean baseline Gross Motor Function Measure (GMFM) scores (C) and mean baseline language developmental quotients (LDQs) (D) of the patients in the transplantation (Trans) and control groups is shown. Statistically significant differences were detected between the groups using a Student t-test.

Figure 4

A comparison of the kinetic Gross Motor Function Measure (GMFM) scores of the patients in the two groups. (A) A comparison of the mean GMFM scores before treatment (0 months) and at 1, 3, and 6 months post-treatment for the patients in the transplantation (Trans) and control groups. (B-D) A comparison of the mean GMFM scores for the patients with GMFCS levels III (B), IV(C) and V(D) in the transplantation (Trans) and control groups. The data were expressed as the mean ± S.E.M. Statistically significant differences were detected between the groups using a two-tailed Student’s t-test, * P<0.05, ** P<0.01, *** P<0.001. (E-F) Changes in the GMFM scores of the patients in the transplantation (Trans) group (E) and in the control group (F) before and after treatment. Each line indicates the kinetic scores of an individual patient.

Figure 5

A comparison of the kinetic language developmental quotients of the patients in the two groups. (A) A comparison of the mean language developmental quotients before (0 months) and after treatment for the patients in the transplantation (Trans) and control groups. (B-D) A comparison of the mean language developmental quotients for the patients with GMFCS levels III (B), IV(C) and V (D) in the transplantation (Trans) and control groups. No statistically significant differences were detected between the groups using a two-tailed Student’s t-test.