The use of chemokine-releasing tissue engineering scaffolds in a model of inflammatory response-mediated melanoma cancer metastasis.

Inflammatory responses and associated products have been implicated in cancer metastasis. However, the relationship between these two processes is uncertain due to the lack of a suitable model. Taking advantage of localized and controllable inflammatory responses induced by biomaterial implantation and the capability of tissue scaffolds to release a wide variety of chemokines, we report a novel system for studying the molecular mechanisms of inflammation-mediated cancer metastasis. The animal model is comprised of an initial subcutaneous implantation of biomaterial microspheres which prompt localized inflammatory responses, followed by the transplantation of metastatic cancer cells into the peritoneal cavity or blood circulation. Histological results demonstrated that substantial numbers of B16F10 cells were recruited to the site nearby biomaterial implants. There was a strong correlation between the degree of biomaterial-mediated inflammatory responses and number of recruited cancer cells. Inflammation-mediated cancer cell migration was inhibited by small molecule inhibitors of CXCR4 but not by neutralizing antibody against CCL21. Using chemokine-releasing scaffolds, further studies were carried out to explore the possibility of enhancing cancer cell recruitment. Interestingly, erythropoietin (EPO) releasing scaffolds, but not stromal cell-derived factor-1α-releasing scaffolds, were found to accumulate substantially more melanoma cells than controls. Rather unexpectedly, perhaps by indirectly reducing circulating cancer cells, mice implanted with EPO-releasing scaffolds had ~30% longer life span than other groups. These results suggest that chemokine-releasing scaffolds may potentially function as implantable cancer traps and serve as powerful tools for studying cancer distraction and even selective annihilation of circulating metastatic cancer cells.

Extracellular matrix expression of human prolapsed vaginal wall.

OBJECTIVE: To compare the mRNA expression of extracellular matrix (ECM) proteins in postmenopausal prolapsed versus non-prolapsed anterior vaginal wall (AVW) tissue. We hypothesized that the weakening of the tissue leading to prolapse was due to decreased collagen production from a downregulation at the transcriptional level. METHODS: Following IRB approval, full thickness samples of redundant AVW were excised from consecutive age-equivalent, postmenopausal, women undergoing cystocele repair (prolapse, stage III or IV), or radical cystectomy (control, no clinical findings of prolapse). Total RNA was isolated, cDNA was synthesized, and quantitative real-time polymerase chain reaction (PCR) was conducted to assess the mRNA expression of collagens type I and III, pro-elastin, MMP3, MMP10, and MMP11. The significance of the difference of mRNA expression between prolapse and control tissues was tested using Student's t-test followed by Mann-Whitney Rank Sum Test. RESULTS: A 5.3-fold increase in collagen type I mRNA was found in prolapse (n = 47) over control (n = 7) tissues (P = 0.009). Type III collagen mRNA was also significantly increased to a 3.3 times higher level (P = 0.017). The ratio of type III to type I was decreased from 15.6 in controls to 9.7 in prolapse. An increasing trend in pro-elastin and MMP mRNA expression was found in prolapse, but this was not statistically significant. CONCLUSION: In this controlled study, the increase found in collagen mRNA expression disproved our hypothesis. To the contrary, this defective prolapsed tissue can signal its need for ECM replenishment. The message, however, is not being effectively translated to assist in tissue remodeling.

A comprehensive approach toward novel serum biomarkers for benign prostatic hyperplasia: the MPSA Consortium.

PURPOSE: Clinical benign prostatic hyperplasia is primarily diagnosed based on a diverse array of progressive lower urinary tract symptoms and is likely distinct from histological benign prostatic hyperplasia, which is detected by the presence of nonmalignant proliferation of prostate cells but may or may not be associated with symptoms. Pharmacological management of lower urinary tract symptoms has emerged as an effective initial treatment for clinical benign prostatic hyperplasia due to the introduction of new drug therapies shown to be effective in recent large clinical trials. Despite advances in symptom management and research into disease pathology, diagnostic strategies for the prediction of benign prostatic hyperplasia progression and response to drug modalities are lacking, and questions remain as to the molecular differences underlying clinical (symptomatic) vs histological (nonsymptomatic) benign prostatic hyperplasia. MATERIALS AND METHODS: As part of the Medical Therapy of Prostatic Symptoms (MTOPS) clinical trial, which demonstrated the effectiveness of combination drug therapy in slowing benign prostatic hyperplasia progression, an archive of biological specimens linked to clinical data was collected for future profiling of disease pathology and changes associated with response to drug therapy. The MTOPS Prostatic Samples Analysis (MPSA) Consortium was established to identify and validate molecular markers that may better define benign prostatic hyperplasia related pathologies, identify risk of progression of lower urinary tract symptoms, and predict response to drug therapy using the MTOPS archive. The cooperating MPSA Biomarker Discovery Sites and Pathology Coordinating Center use diverse methodologies and scientific approaches as well as unique expertise to address the goals of the Consortium. RESULTS: To date the MPSA has identified a number of promising biomarkers as well as other molecular and cellular changes associated with benign prostatic hyperplasia. CONCLUSIONS: These findings and ongoing Consortium discovery efforts have the potential to provide a greater understanding of the defects underlying disease pathology, and may lead to the development of early and more effective pharmacological treatment strategies for benign prostatic hyperplasia.

Prostatic stromal cells derived from benign prostatic hyperplasia specimens possess stem cell like property.

INTRODUCTION: The hyper-proliferative activity of stromal smooth muscle (SM) cells is believed to be responsible for the pathogenesis of benign prostatic hyperplasia (BPH). We have observed that those stromal cells can differentiate into unrelated specialized cells. We thus hypothesize that stromal cells derived from adults prostate specimens may contain adult stem cells. To test this hypothesis, human prostate stromal primary cultures were established and used for characterization of their stem cell properties. METHODS: Immunoblotting, immunohistochemistry, RT-PCR, and tissue culture techniques were used to characterize the primary cultured human prostate-derived stromal cells for their stem cell and differentiation properties. The plasticity of these stromal cells was analyzed using cell culture and histology techniques. RESULTS: Primary cultured prostate stromal cells from BPH patient possess polygonal and elongated fibroblast/myofibroblast cellular morphology. They are positive in CD30, CD34, CD44, NSE, CD133, Flt-1, stem cell factor (SCF), and neuron-specific enolase (NSE), but negative in C-Kit, stem cell antigen (SCA), SH2, CD11b. Expression of SM myogenic markers in these cells may be induced by sodium butyrate (NaBu) treatment. Induction to osteogenic and adipogenic differentiation in these cells is also evident. CONCLUSIONS: Our study on primary stromal cells from BPH patients have yielded many interesting findings that these prostate stroma cells possess: (1) mesenchymal stem cell (MSC) markers; (2) strong proliferative potential; and (3) ability to differentiate or transdifferentiate to myogenic, adipogenic, and osteogenic lineages. These cell preparations may serve as a potential tool for studies in prostate adult stem cell research and the regulation of benign prostatic hyperplasia.

Elastin expression and elastic fibre width in the anterior vaginal wall of postmenopausal women with and without prolapse.

OBJECTIVES: To compare elastin expression and elastic fibre width in the anterior vaginal wall of postmenopausal women with and with no bladder prolapse. PATIENTS AND METHODS: Full-thickness specimens were obtained from the upper lateral anterior vaginal wall of women having a large cystocele repaired (stage III or IV; prolapse group, 33) and the same location in patients with no prolapse having radical cystectomy (control group, 10). The percentage of elastin-positive tissue and elastic fibre width were measured by immunohistochemistry on 6 microm thick tissue sections from 10 random field readings per sample using image analysis software. The examiner was unaware of sample identity and the patients' clinical history. RESULTS: The age was comparable between the control and prolapse groups (median 70.5 years), and the parity, vaginal deliveries, hormone replacement use, cigarette smokers and body mass index were no different between the groups. Immunohistochemical staining and morphometric analysis indicated that elastin expression in the prolapse group was 10.6%, vs 14.4% in the control group (P = 0.049). The median width of elastic fibres was 0.9 microm in the prolapse and 1.8 microm in the control groups (P < 0.001). Elastin expression and elastic fibre width appeared to be stable with increasing age in the prolapse group. CONCLUSIONS: In this case-control study investigating elastin changes in postmenopausal women with prolapse, the elastin expression and fibre width were significantly lower in the vaginal wall of patients with a large cystocele than in controls of a similar age.

Changes in detrusor smooth muscle myosin heavy chain mRNA expression following spinal cord injury in the mouse.

AIMS: Smooth muscle myosin heavy chain (SMMHC) isoform composition has been shown to be developmentally regulated and to be associated with functional changes in smooth muscle activity. In this study, we sought to determine expression patterns of SMMHC isoforms in a murine model of spinal cord injury (SCI) and to compare these expression patterns to neurologic, cytometric, and morphometric findings. MATERIALS AND METHODS: Baseline cystometry was performed on adult, female mice followed by either thoracic spinal cord transection (SCI) or sham operation (Sham). At 1, 3, or 6 weeks postoperatively neurologic evaluation and cystometry were performed, bladders were harvested, and expression patterns of SMMHC isoforms (SM1 vs. SM2 and SMA vs. SMB) were assessed by RT-PCR. Morphometrics utilizing computer-assisted color image analysis was also performed on all bladders. RESULTS: There was a significant increase in bladder weight and capacity 1 week following SCI which normalized over time, however, morphometric analysis did not reveal an alteration in tissue composition amongst the three groups. One week following SCI, SM1 was predominantly expressed over SM2 and began to normalize at 3 weeks. This coincided with the emergence of reflex voiding and detrusor overactivity. SMA was expressed following SCI only, and the number of bladders found to express SMA decreased with increasing duration since SCI. CONCLUSIONS: Smooth muscle myosin heavy chain mRNA expression patterns appear to be affected by SCI. We believe the induction of SMA may be a factor in altered bladder function following injury.

Alpha(2) macroglobulin, a PSA binding protein, is expressed in human prostate stroma.

BACKGROUND: Benign prostatic hyperplasia (BPH) is characterized as a stromal process. The stroma smooth muscle (SM) may alter its phenotype during the progression of BPH. We have identified gene transcripts that may be differentially expressed in BPH using a differential display method. Among the fragments isolated, alpha(2) macroglobulin (alpha(2)-M) is one of the most interesting. alpha(2)-M is a binding protein of a variety of proteinases, including prostatic specific antigen (PSA). It also plays roles in molecular trapping and targeting. In this study, we characterized alpha(2)-M expression in the human prostate. METHODS: Differential display was used to identify and isolate the differentially expressed transcripts between normal prostate and BPH tissues. RT-PCR, Western blot, in situ hybridization, and immunohistochemistry were utilized to confirm and characterize alpha(2)-M expression in the prostate. RESULTS: Real-time RT-PCR results revealed that a 3.2-fold increase in alpha(2)-M mRNA expression is observed in BPH compared with normal prostate tissue. A 1.9-fold increase at protein level was also observed. In situ hybridization and immunohistochemistry showed that alpha(2)-M expression is primarily localized to the stromal compartment. Cultured primary stroma cells maintained alpha(2)-M expression, while prostate epithelial cells had a significantly lower level of alpha(2)-M expression. Furthermore, stromal cells in culture produce and secrete alpha(2)-M in the medium. CONCLUSIONS: We identified alpha(2)-M expression in the human prostate. An increased alpha(2)-M expression appears to be associated with BPH. Considering the unique features of its protein binding and targeting properties, alpha(2)-M expressed in the prostate may play an important role in regulating benign and malignant prostatic growth.