Fig. 2.

Accumulation of structurally defective mitochondria in idiopathic and Parkin-defect hAD-MSCs. (A) (a) An electron micrograph showing the primary hAD-MSCs from non-PD patients. Primary cultures of non-PD were examined by electron microscopy and the mitochondria (M) are shown in the image. Note that the inner matrices of the mitochondria are generally evenly filled with cristae. Bar=200 nm. (b) An electron micrograph showing the immortalized hAD-MSCs from non-PD patients. Immortalized non-PD cells were examined by electron microscopy and the mitochondria (M) are shown in the image. Note that the inner mitochondrial matrices are not only evenly filled but also clearly show distinguishable cristae. Nu, nucleus. Bar=200 nm. (c) An electron micrograph showing the primary hAD-MSCs from idiopathic PD patients. Primary cultures of PD were examined by electron microscopy and the mitochondria (M) are shown in the image. Note that the inner mitochondrial matrices show signs of disintegration and hollow areas are visible. Bar=200 nm. (d) An electron micrograph showing the immortalized hAD-MSCs from idiopathic PD patients. Immortalized PD cells were examined by electron microscopy and the mitochondria (M) are shown in the image. Note that the inner mitochondrial matrices show clearly visible signs of disintegration, resulting in the many hollow areas. Bar=200 nm. (e) An electron micrograph showing the primary hAD-MSCs from PD patients with Parkin deficiency. Primary cultures of Parkin were examined by electron microscopy and the mitochondria (M) are shown in the image. Note that the overall shapes of the mitochondria are grossly distorted and the inner mitochondrial matrices are either severely damaged or almost absent. Bar=200 nm. (f) An electron micrograph showing the immortalized hAD-MSCs from PD patients with Parkin deficiency. Immortalized Parkin cells were examined by electron microscopy and the mitochondria (M) are shown in the image. Note that the inner mitochondrial matrices of many mitochondria are almost absent. Bar=200 nm. The area of the mitochondria from the electron micrograph data in the idiopathic- and Parkin-defect cells was significantly decreased by 90.3% and 90.8%, in the primary cells, and by 94.8% and 95.0% in the immortalized cells, respectively, compared to the wild type EM. The number of mitochondria from the electron micrograph data for the idiopathic- and Parkin-defect cells was significantly decreased by 94.5% and 94.7%, in the primary cells, and by 94.8% and 95.0% in the immortalized cells, respectively, compared to the wild type EM. All values are the means±SEM. ^****p <0.0001.

(A).

(B) Analysis of mitochondrial immunostaining in primary and immortalized hAD-MSCs. non-PD showing the hAD-MSCs from non-PD patients before (a) and after immortalization (b); PD showing the hAD-MSCs from idiopathic PD patients before (c) and after immortalization (d); Parkin showing the hAD-MSCs from Parkin-defect PD patients before (e) and after immortalization (f), Bar=100 µm for primary cells, 20 µm for immortalized cells, respectively. The degree of mitochondrial staining in idiopathic- and Parkin-defect cells was significantly decreased by 43.2% and 92.3%, in the primary cells, and by 19.8% and 36.3% in the immortalized cells, respectively, compared to the non-PD cells; their expressions were normalized to DAPI. All values are the means±SEM. ^**p<0.01, ^****p<0.0001.

(C) Analysis of Mitotracker staining in primary and immortalized hAD-MSCs. non-PD showing the hAD-MSCs from non-PD patients before (a) and after immortalization (b); PD showing the hAD-MSCs from idiopathic PD patients before (c) and after immortalization (d); Parkin showing the hAD-MSCs from Parkin-defect PD patients before (e) and after immortalization (f); Bar=50 µm for primary cells, 10 µm for immortalized cells, respectively. The degree of Mitotracker staining in the idiopathic- and Parkin-defect cells were significantly decreased by 65.2% and 76.1% in the primary cells, and by 33.1% and 53.5% in the immortalized cells, respectively, compared to the non-PD cells; their expressions were normalized to DAPI. All values are the means±SEM. ^*p<0.05, ^***p<0.001, ^****p<0.0001.

(D) Analysis of mitochondrial immunofluorescence staining in brain cortex tissues from idiopathic, early onset PD and non-PD patients. The degree of mitochondrial staining in early onset and idiopathic PD tissues was significantly decreased by 21.0% and 49.0%, respectively, compared to the non-PD tissues; their expressions were normalized to DAPI. All values are the means±SEM. ^**p<0.01. Bar=20 µm.