Ansha Qian

Evidence That Osteogenic Progenitor Cells in the Human Tunica Albuginea May Originate from Stem Cells: Implications for Peyronie Disease

Abstract Tissue ossification in Peyronie disease (commonly known as Peyronies disease [PD]), a localized fibrotic lesion within the tunica albuginea (TA) of the penis, may result from osteogenic differentiation of fibroblasts, myofibroblasts, and/or adult stem cells in the TA, and may be triggered by chronic inflammation, oxidative stress, and profibrotic factors like transforming growth factor beta 1 (TGFB1). In this study, we have investigated whether cultures of cells from normal TA and PD plaques undergo osteogenesis, express markers for stem cells, and originate other cell lineages via processes modulated by TGFB1. We found that TA and PD cells in osteogenic medium (OM) expressed osteogenic markers, alkaline phosphatase, and osteopontin and underwent calcification. PD cells, but not TA cells, formed foci in soft agar that were positive for alkaline phosphatase and calcification and expressed the mRNAs for osteoblast-specific factors pleiotrophin and periostin and bone morphogenic protein 2. Both cultures expressed stem cell marker CD34 antigen but not protein tyrosine phosphatase, receptor type c. TA and PD cells expressed smooth-muscle cell markers smoothelin and transgelin. None of the cultures underwent adipogenesis in adipogenic medium. Incubation with TGFB1 increased osteogenesis and myofibroblast differentiation and reduced CD34 antigen expression in both cultures. TA and PD cells modulated the differentiation of the multipotent C3H 10T(1/2) cells in dual cultures, into osteoblasts and myofibroblasts. In conclusion, both TA and PD cultures contain cells, presumably stem cells, that undergo osteogenic and myofibroblast differentiation, and may induce these processes by paracrine interactions. This may explain progression of fibrosis in the PD plaque and its eventual calcification.

Gene expression profiles in the Peyronie’s disease plaque

OBJECTIVES To provide molecular insight into the pathophysiology of Peyronies disease (PD), a preliminary profile of differential gene expression between the PD plaque and control tunica albuginea was obtained with DNA microarrays. METHODS Seven PD plaques and five control tunica albugineas were studied. cDNA specimens were prepared from RNA isolated from one calcified PD plaque and one control tissue and hybridized with the Clontech Atlas 1.2 Array. Another set of plaque and control RNA samples was hybridized with the Affymetrix GeneChip. Relative changes of greater than 2.0 defined up-regulation and down-regulation, respectively. RNA from the remaining tissues was used to determine, by reverse transcriptase-polymerase chain reaction and Western blot analysis, the expression of selected individual genes. RESULTS Some of up-regulated genes in the PD plaque detected by the Clontech assay were pleiotrophin, monocyte chemotactic protein 1, and early growth response protein, which are involved in osteoblast recruitment, inflammation, and fibroblast proliferation, respectively. Ubiquitin and Id-2, which are involved in tissue remodeling, were down-regulated. The Affymetrix DNA chips identified the up-regulation of elastase (involved in elastic fiber degradation) and the myofibroblast markers alpha and gamma-smooth muscle actin, desmin, and others, as well as the down-regulation of collagenase IV and transforming growth factor-beta modulators. Four of the five genes selected for reverse transcriptase-polymerase chain reaction and Western blotting confirmed the DNA microarray results. CONCLUSIONS In the PD tissue, the genes involved in collagen synthesis, myofibroblast differentiation, tissue remodeling, inflammation, ossification, and proteolysis are up-regulated, and the genes that inhibit some of these processes and collagenase are down-regulated.

Gene expression in Peyronie's disease

Currently, surgical intervention is the only efficacious treatment for Peyronies disease (PD), a fibromatosis of the tunica albuginea of the penis. Therapies based on the molecular pathways for this disease could provide alternatives to surgical treatment but only recently has the pathophysiology of the Peyronies disease plaque been investigated at the molecular level. In this review, we examine the current knowledge of gene expression in the PD plaque and the relationship of PD with other fibrotic conditions such as Dupytrens disease. TGFβ1, along with other growth factors, pro-fibrotic genes, and collagen, are expressed in fibroblasts and myofibroblasts. Myofibroblasts are normally involved in wound contracture and largely eliminated via apoptosis during the late stages of wound remodeling. In the PD plaque, however, these cells persist and may play an important role in the PD plaque fibrosis. The expression levels of TGFβ1 and pro- and anti-fibrotic gene products, along with the nitric oxide/reactive oxygen species (NO/ROS) ratio in the tunica albuginea, appear to be essential for the formation and progression of the PD plaque and effect the expression of multiple genes. This can be assessed with the recently developed DNA-based chip arrays and results with the PD plaque have been encouraging. OSF-1 (osteoblast recruitment), MCP-1 (macrophage recruitment), procollagenase IV (collagenase degradation), and other fibrotic genes have been identified as being possible candidate regulatory genes. Finally, possible therapeutic avenues for gene-based therapy in the treatment of PD are discussed that may eventually reduce the need for surgical intervention.

Profibrotic Role of Myostatin in Peyronie's Disease

INTRODUCTION The primary histologic finding in many urologic disorders, including Peyronies disease (PD), is fibrosis, mainly mediated by the transforming growth factor beta1 (TGFbeta1). AIM To determine whether another member of the TGFbeta family, myostatin, (i) is expressed in the human PD plaque and normal tunica albuginea (TA), their cell cultures, and the TGFbeta1-induced PD lesion in the rat model; (ii) is responsible for myofibroblast generation, collagen deposition, and plaque formation; and (iii) mediates the profibrotic effects of TGFbeta1 in PD. METHODS Human TA and PD tissue sections, and cell cultures from both tissues incubated with myostatin and TGFbeta1 were subjected to immunocytochemistry for myostatin and alpha-smooth muscle actin (ASMA). The cells were assayed by western blot, Real time-Polymerase chain reaction (RT-PCR), and ribonuclease protection. Myostatin cDNA and shRNA were injected, with or without TGFbeta1, in the rat penile TA, and plaque size was estimated by Masson. MAIN OUTCOME MEASURES Myostatin expression in the human TA, the PD plaque, and their cell cultures, and myostatin effects on the PD-like plaque in the rat. RESULTS A threefold overexpression of myostatin was found in the PD plaque as compared with the TA. In PD cells, myostatin expression was mainly in the myofibroblasts, and in the TA cells, it increased upon passage paralleling myofibroblast differentiation and was up-regulated by TGFbeta1. Myostatin or its cDNA construct increased the myofibroblast number and collagen in TA cells. Myostatin was detected in the TGFbeta1-induced PD-like plaque of the rat partly in the myofibroblasts, and in the TA. Myostatin cDNA injected in the TA induced a plaque and intensified the TGFbeta1 lesion, which was not reduced by myostatin shRNA. CONCLUSIONS Myostatin is overexpressed in the PD plaque, partly because of myofibroblast generation. Although myostatin induces a plaque in the rat TA, it does not appear to mediate the one triggered by TGFbeta1, thus suggesting that both proteins act concurrently and that therapy should target their common downstream effectors.