Effect of muscle-derived stem cells on the restoration of corpora cavernosa smooth muscle and erectile function in the aged rat.
ABSTRACT To determine whether skeletal muscle-derived stem cells (MDSCs) convert into smooth muscle cells (SMCs) both in vitro and in vivo, and in so doing ameliorate the erectile dysfunction (ED) of aged rats, and whether endogenous stem cells are present in the rat corpora cavernosa.
MDSCs were obtained from mouse muscle, and shown by immunocytochemistry for alpha-smooth muscle actin (alpha SMA) to originate in vitro in myofibroblasts and SMCs, discriminating SMCs by calponin 1 expression. In vivo these MDSCs, labelled with 4',6-diamidino-2-phenylindole, were implanted into the corpora cavernosa of young adult (5-month old) and aged (20-month old) rats for 2 and 4 weeks. Histological changes were assessed by immunohistochemistry and quantitative Western blot. Functional changes were determined by electrical field stimulation (EFS) of the cavernosal nerve.
The exogenous cells replicated and converted into SMCs, as shown in corporal tissue sections by confocal immunofluorescence microscopy for proliferating cell nuclear antigen (PCNA), alpha SMA, and smoothelin, and also by Western blot for alpha SMA and PCNA. MDSC differentiation was confirmed by the activation of the alpha SMA promoter-linked beta-galactosidase in transfected cells, both in vitro and after implantation in the corpora. Putative endogenous stem cells were shown in corporal tissue sections and Western blots by detecting CD34 and a possible Sca1 variant. EFS showed that implanted MDSCs raised in aged rats the maximal intracavernosal pressure/mean arterial pressure levels above (2 weeks) or up to (4 weeks) those of young adult rats.
MDSCs implanted into the corpora cavernosa of aged rats converted into SMCs and corrected ED, and endogenous cells expressing stem cell markers were also found in untreated tissue. This suggests that exogenous stem cell implantation and/or endogenous stem cell modulation might be viable therapeutic approaches for ageing-related ED.
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ABSTRACT: The intracellular mechanisms that regulate neurogenesis remain unclear. Using neurospheres isolated from the subventricular zone (SVZ) of the adult rat, we investigated the effect of cyclic guanosine monophosphate (cGMP) and its signaling pathway on the induction of neurogenesis. Neurospheres expressed phosphodiesterase 5 (PDE5) and treatment of neurospheres with Sildenafil, a specific inhibitor of PDE5, significantly increased cGMP levels and neurogenesis. In addition, incubation of neurospheres with Sildenafil significantly phosphorylated Akt, which was associated with an increase of phosphorylation of glycogen synthase kinase 3 (GSK-3), a downstream target of Akt. Coincubation of neurospheres with Sildenafil and LY 294002, a pharmacological inhibitor of PI3-K/Akt, abolished Sildenafil-induced phosphorylated Akt and GSK-3. Furthermore, LY 294002 blocked Sildenafil-increased SVZ cell proliferation. These data suggest that Sildenafil-enhanced neurogenesis likely occurs through activation of the PI3-K/Akt/GSK-3 pathway.Journal of Cerebral Blood Flow & Metabolism 10/2005; 25(9):1150-8. · 5.40 Impact Factor
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ABSTRACT: Three populations of myogenic cells were isolated from normal mouse skeletal muscle based on their adhesion characteristics and proliferation behaviors. Although two of these populations displayed satellite cell characteristics, a third population of long-time proliferating cells expressing hematopoietic stem cell markers was also identified. This third population comprises cells that retain their phenotype for more than 30 passages with normal karyotype and can differentiate into muscle, neural, and endothelial lineages both in vitro and in vivo. In contrast to the other two populations of myogenic cells, the transplantation of the long-time proliferating cells improved the efficiency of muscle regeneration and dystrophin delivery to dystrophic muscle. The long-time proliferating cells' ability to proliferate in vivo for an extended period of time, combined with their strong capacity for self-renewal, their multipotent differentiation, and their immune-privileged behavior, reveals, at least in part, the basis for the improvement of cell transplantation. Our results suggest that this novel population of muscle-derived stem cells will significantly improve muscle cell-mediated therapies.The Journal of Cell Biology 06/2002; 157(5):851-64. · 10.82 Impact Factor
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ABSTRACT: Nitric oxide (NO) is a short lived diatomic free radical species synthesized by nitric oxide synthases (NOS). The physiological roles of NO depend on its local concentrations as well as availability and the nature of downstream target molecules. At low nanomolar concentrations, activation of soluble guanylyl cyclase (sGC) is the major event initiated by NO. The resulting elevation in the intracellular cyclic GMP (cGMP) levels serves as signals for regulating diverse cellular and physiological processes. The participation of NO and cGMP in diverse physiological processes is made possible through cell type specific spatio-temporal regulation of NO and cGMP synthesis and signal diversity downstream of cGMP achieved through specific target selection. Thus cyclic GMP directly regulates the activities of its downstream effectors such as Protein Kinase G (PKG), Cyclic Nucleotide Gated channels (CNG) and Cyclic nucleotide phosphodiesterases, which in turn regulate the activities of a number of proteins that are involved in regulating diverse cellular and physiological processes. Localization and activity of the NO-cGMP signaling pathway components are regulated by G-protein coupled receptors, receptor and non receptor tyrosine kinases, phosphatases and other signaling molecules. NO also serves as a powerful paracrine factor. At micromolar concentrations, NO reacts with superoxide anion to form reactive peroxinitrite, thereby leading to the oxidation of important cellular proteins. Extensive research efforts over the past two decades have shown that NO is an important modulator of axon outgrowth and guidance, synaptic plasticity, neural precursor proliferation as well as neuronal survival. Excessive NO production as that evoked by inflammatory signals has been identified as one of the major causative reasons for the pathogenesis of a number of neurodegenerative diseases such as ALS, Alzheimers and Parkinson diseases. Regenerative therapies involving transplantation of embryonic stem cells (ES cells) and ES cell derived lineage committed neural precursor cells have recently shown promising results in animal models of Parkinson disease (PD). Recent studies from our laboratory have shown that a functional NO-cGMP signaling system is operative early during the differentiation of embryonic stem cells. The cell type specific, spatio-temporally regulated NO-cGMP signaling pathways are well suited for inductive signals to use them for important cell fate decision making and lineage commitment processes. We believe that manipulating the NO-cGMP signaling system will be an important tool for large scale generation of lineage committed precursor cells to be used for regenerative therapies.Neurochemical Research 01/2007; 32(4-5):681-94. · 2.13 Impact Factor