Integrated Bioinformatics Analysis Identified ASNS and DDIT3 as the Therapeutic Target in Castrate-Resistant Prostate Cancer.

Many studies have demonstrated the mechanisms of progression to castration-resistant prostate cancer (CRPC) and novel strategies for its treatment. Despite these advances, the molecular mechanisms underlying the progression to CRPC remain unclear, and currently, no effective treatments for CRPC are available. Here, we characterized the key genes involved in CRPC progression to gain insight into potential therapeutic targets. Bicalutamide-resistant prostate cancer cells derived from LNCaP were generated and named Bical R. RNA sequencing was used to identify differentially expressed genes (DEGs) between LNCaP and Bical R. In total, 631 DEGs (302 upregulated genes and 329 downregulated genes) were identified. The Cytohubba plug-in in Cytoscape was used to identify seven hub genes (ASNS, AGT, ATF3, ATF4, DDIT3, EFNA5, and VEGFA) associated with CRPC progression. Among these hub genes, ASNS and DDIT3 were markedly upregulated in CRPC cell lines and CRPC patient samples. The patients with high expression of ASNS and DDIT3 showed worse disease-free survival in patients with The Cancer Genome Atlas (TCGA)-prostate adenocarcinoma (PRAD) datasets. Our study revealed a potential association between ASNS and DDIT3 and the progression to CRPC. These results may contribute to the development of potential therapeutic targets and mechanisms underlying CRPC progression, aiming to improve clinical efficacy in CRPC treatment.

Erectile Dysfunction Treatment Using Stem Cell Delivery Patch in a Cavernous Nerve Injury Rat Model.

Erectile dysfunction (ED) is a common and feared complication of radical prostatectomy (RP) for prostate cancer. Recently, tissue engineering for post-prostatectomy ED has been attempted in which controlled interactions between cells, growth factors, and the extracellular matrix (ECM) are important for the structural integrity if nerve regeneration. In this study, we evaluated the effects of a biomechanical ECM patch on the morphology and behavior of human bone marrow-derived mesenchymal stem cells (hBMSCs) in a bilateral cavernous nerve injury (BCNI) rat model. The ECM patch, made of decellularized human fibroblast-derived ECM (hFDM) and a biocompatible polyvinyl alcohol (PVA) hydrogel, was tested with human bone marrow-derived mesenchymal stem cells (hBMSCs) on a bilateral cavernous nerve injury (BCNI) rat model. In vitro analysis showed that the hFDM/PVA + hBMSCs patches significantly increased neural development markers. In vivo experiments demonstrated that the rats treated with the hFDM/PVA patch had higher ICP/MAP ratios, higher ratios of smooth muscle to collagen, increased nNOS content, higher levels of eNOS protein expression, and higher cGMP levels compared to the BCNI group. These results indicate that the hFDM/PVA patch is effective in promoting angiogenesis, smooth muscle regeneration, and nitrergic nerve regeneration, which could contribute to improved erectile function in post-prostatectomy ED.

Niclosamide exerts anticancer effects through inhibition of the FOXM1-mediated DNA damage response in prostate cancer.

Niclosamide, an established anti-helminthic drug, has anticancer activity against various cancers including prostate cancer, but the underlying mechanisms have not yet been defined. We demonstrated the anticancer effects of niclosamide in castration-resistant prostate cancer (CRPC) cells, and elucidated the mechanism of action of niclosamide in CRPC. Niclosamide reduced cell proliferation and induced apoptosis of CRPC cells in vitro, and also reduced xenograft tumor growth in vivo. Niclosamide significantly increased the number of γH2AX- and 53BP1-positive cells. In RNA-sequencing, niclosamide induced extensive changes in gene expression including cell division, DNA replication, and DNA repair. Bioinformatics analysis using TCGA data set revealed that FOXM1 is an important target of niclosamide. In microarray assays, FOXM1 knockdown significantly inhibited several genes involved in DNA repair, and homologous recombination, in particular. Finally, FOXM1 strongly bound to EXO1 in CRPC cells, and FOXM1 knockdown significantly reduced EXO1-driven luciferase activity. Taken together, our results suggest that niclosamide exerts anticancer activity through inhibition of the FOXM1-mediated DNA damage response in CRPC.

HMGB1 promotes tumor progression and invasion through HMGB1/TNFR1/NF-κB axis in castration-resistant prostate cancer.

Prostate cancer (PCa) is the most common male cancer. Most patients treated with androgen deprivation therapy progress to castration-resistant PCa. To overcome the limitations of this treatment, there is an urgent need to identify more effective treatment targets. High mobility group box 1 protein (HMGB1) is known to be associated with progression, metastasis, and poor prognosis of several solid tumors; however, its role in PCa remains unclear. Thus, we aimed to evaluate the clinical significance and biological roles and mechanism of HMGB1 in PCa. We showed that increased expression of HMGB1 correlated with increased risk of aggressive PCa, and high expression of HMGB1 was associated with poor biochemical recurrence-free survival in a Korean cohort. Additionally, the inhibition of HMGB1 expression significantly reduced cell proliferation, invasive capacity, and NF-κB signaling in vitro. Our results indicated that HMGB1 is a critical factor in the development and progression of PCa. Moreover, we found that HMGB1 directly interacts with TNFR1, and TNFR1 overexpression in HMGB1 knockdown cells reversed the effects of HMGB1 knockdown. Importantly, our results suggest that HMGB1 binding to TNFR1 promotes tumor progression by activating the NF-κB signaling pathway in PCa; therefore, the HMGB1/TNFR1/NF-κB signaling pathway could serve as a novel therapeutic target for improving PCa therapy.

Stem Cell/Oxygen-Releasing Microparticle Enhances Erectile Function in a Cavernous Nerve Injury Model.

Erectile dysfunction caused by damage to the cavernous nerve is a common complication of radical prostatectomy for patients with localized prostate cancer. Various studies have investigated repair of damaged tissue and prevention of fibrosis in the corpus cavernosum using stem cell therapy. However, stem cell therapy has limitations, including insufficient nutrient and oxygen supply to transplanted stem cells. This study investigated whether stem cell/oxygen-releasing hollow microparticles (HPs) had therapeutic effect on erectile dysfunction in a rat model of bilateral cavernous nerve injury (BCNI). Therapeutic effects were observed in the BCNI model at 1, 2, and 4 weeks postcavernous nerve injury. Erectile function further improved after treatment with stem cell/oxygen-releasing HP system compared with treatment with only stem cells at 4 weeks. Stem cell/oxygen-releasing HP system increased cyclic guanosine monophosphate (cGMP) level and neuronal nitric oxide synthase (nNOS), endothelial nitric oxide synthase (eNOS), α-smooth muscle actin (α-SMA), and muscarinic acetylcholine receptor 3 (M3) expression while decreasing fibrosis and apoptosis in the corpus cavernosum. Our results clearly show that stem cell survival increases around transplanted stem cell/oxygen-releasing hybrid system site. Taken together, an oxygen-releasing HP system supported prolonged stem cell survival, sustaining the paracrine effect of the stem cells, and consequently enhancing erectile function. These findings show promise with regard to prolonged stem cell survival in stem cell applications for various diseases and types of tissue damage. Impact statement In this study, we used an oxygen-releasing hollow microparticles (HPs) system with stem cells to attempt to overcome certain limitations of stem cell therapy, including insufficient nutrient and oxygen supplies for transplanted stem cells. Our results demonstrated that a stem cell/oxygen-releasing HP hybrid system could further improve erectile function, cyclic guanosine monophosphate (cGMP) level, and NOS level in a bilateral cavernous nerve injury rat model through prolonged stem cell survival. Our data suggest that a stem cell/oxygen-releasing HP system is a promising clinical treatment option for postprostatectomy erectile dysfunction. Furthermore, this system may be relevant in different disease therapies and regenerative medicine.

Direct differentiation of insulin-producing cells from human urine-derived stem cells.

The loss of pancreatic ß-cells is a cause of diabetes. Therefore, replacement of pancreatic ß-cells is a logical strategy for the treatment of diabetes, and the generation of insulin-producing cells (IPCs) from stem cells has been widely investigated as an alternative source for pancreatic ß-cells. Here, we isolated stem cells from human urine and investigated their differentiation potential into IPCs. We checked the expression of surface stem cell markers and stem cell transcription factors, and found that the isolated human urine-derived stem cells (hUDSCs) expressed the stem cell markers CD44, CD90, CD105 and stage-specific embryonic antigen (SSEA)-4. In addition, these cells expressed octamer binding transcription factor (Oct)4 and vimentin. hUDSCs could differentiate into adipocytes and osteocytes, as evidenced by Oil-red O staining and Alizarin Red S-staining of differentiated cells, respectively. When we directly differentiated hUDSCs into IPCs, the differentiated cells expressed mRNA for pancreatic transcription factors such as neurogenin (Ngn)3 and pancreatic and duodenal homeobox (Pdx)1. Differentiated IPCs expressed insulin and glucagon mRNA and protein, and these IPCs also secreted insulin in response to glucose stimulation. In conclusion, we found that hUDSCs can be directly differentiated into IPCs, which secrete insulin in response to glucose.

GV1001 inhibits cell viability and induces apoptosis in castration-resistant prostate cancer cells through the AKT/NF-κB/VEGF pathway.

Purpose: We examined the effect of GV1001 in castration castration-resistant prostate cancer (CRPC) cell growth and invasion and explored the potential molecular mechanisms of action. Materials and Methods: The in vitro anti-cancer effects of GV1001 in CRPC cells were examined using cell viability assay, TUNEL assay, and flow cytometry analysis. To evaluate the effects of GV1001 on different steps of angiogenesis, wound healing assay, transwell invasion assay, endothelial cell tube formation assay, and western blot analysis were performed. Finally, the anti-cancer effects of GV1001 on tumor growth in vivo were examined in a CRPC xenograft model. Results: GV1001 inhibited cell viability and induced apoptosis in CRPC cells in vitro, accompanied by down-regulation of Bcl-2 and caspase-3. GV1001 also inhibited different steps of angiogenesis, such as migration, invasion, and endothelial tube formation, along with decreased expression of MMP-2, MMP-9, and CD31 and increased expression of TIMP-1 and TIMP-2. Mechanistically, GV1001 significantly decreased the levels of phosphorylated AKT, phosphorylated p65, and VEGF in CRPC cells in a dose-dependent manner. GV1001 was effective in suppressing tumor growth and inducing apoptosis in a CRPC xenograft mouse model. Conclusions: Our data demonstrated that GV1001 inhibited cell viability, induced apoptosis, and inhibited angiogenesis in CRPC cells by inhibition of the AKT/NF-κB/VEGF signaling pathway.

Author Correction: Prostate-specific extracellular vesicles as a novel biomarker in human prostate cancer.

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Stem Cells Seeded on Multilayered Scaffolds Implanted into an Injured Bladder Rat Model Improves Bladder Function.

Background: To investigate whether human adipose-derived stem cells (hADSCs) seeded on multilayered poly (l-lactide-co-ɛ-caprolactone) (PLCL) sheets improve bladder function in a rat model of detrusor smooth muscle-removed bladder. Methods: Male rats were randomly divided into 4 groups: Normal, injury (detrusor smooth muscle-removed bladder), PLCL (detrusor smooth muscle-removed bladder implanted with PLCL sheets), and PLCL + ADSC (detrusor smooth muscle-removed bladder implanted with PLCL sheets seeded with hADSCs). Four weeks after the treatment, physiological, histological, immunohistochemical, and immunoblot analyses were performed. Results: hADSCs were compatible with PLCL sheets. Further, the physiological study of PLCL + ADSC group showed significant improvement in compliance and contractility suggesting the functional improvement of the bladder. Histological, immunohistochemical and immunoblot analyses revealed the uniform distribution of hADSCs in between PLCL sheets as well as differentiation of hADSCs into smooth muscle cells (SMC) which is illustrated by the expression of SMC markers. Conclusion: hADSCs seeded on the multilayered PLCL sheets has the potential to differentiate into SMC, thus facilitating the recovery of compliance and contractility of the injured bladder.

High FOXM1 expression is a prognostic marker for poor clinical outcomes in prostate cancer.

Purpose: We aimed to investigate the expression of FOXM1 and to determine the relationships between FOXM1 expression and clinicopathologic characteristics in patients with PCa. Furthermore, we reconfirmed the prognostic impact of FOXM1 in different cohorts using already published data. Patients and Methods: Formalin-fixed, paraffin-embedded tissues were collected from patients with low- (n=17), intermediate- (n=36), and high-risk (n=29) disease, from patients with CRPC (n=2) and from patients with BPH (n=28). To analyze FOXM1 expression, we performed IHC analyses. Also, we analyzed gene expression data from cBioPortal to evaluate the associations between FOXM1 alteration and prognosis of PCa. Results: FOXM1 expression measured using Allred score differed between patients with BPH, and low-, intermediate-, and high-risk PCa (0.3, 1.5, 4.8, and 6.2, respectively; p<0.001). Patients with high FOXM1 expression had higher preoperative PSA levels (p=0.023), more advanced tumor stages (p=0.047), and higher pathologic Gleason score (p<0.001) than those with low FOXM1 expression. ROC curve analysis indicated that FOXM1 expression was a useful marker for discriminating PCa from BPH (AUC 0.851, 95% CI 0.783-0.920) and for discriminating high-risk PCa from low- and intermediate-risk PCa (AUC 0.807, 95% CI 0.719-0.894). In multivariate analyses, high FOXM1 expression was an independent predictor of BCR. Finally, in the TCGA dataset, FOXM1 alteration was associated with poor overall (p=4.521e-4) and disease-free survival (p=0.0108). Conclusions: In patients with PCa, high FOXM1 expression was associated with advanced tumor stages, high Gleason score, and poor prognosis. These data suggest a role of FOXM1 in biologically and clinically aggressive PCa.

Therapeutic Effect of Controlled Release of Dual Growth Factor Using Heparin-Pluronic Hydrogel/Gelatin-Poly (Ethylene Glycol)-Tyramine Hydrogel System in a Rat Model of Cavernous Nerve Injury.

The number of cases of erectile dysfunction (ED) caused after radical prostatectomy (RP) prostate cancer treatment is increasing steadily. Although various studies have been conducted for treatment of post-RP ED, there is still a need for more effective methods. A dual growth factor incorporated heparin-pluronic/gelatin-poly(ethylene glycol)-tyramine (HP/GPT) hydrogel, which consists of a basic fibroblast growth factor (bFGF)-loaded HP hydrogel and nerve growth factor (NGF)-loaded GPT hydrogel, can control dose and rate of growth factor release. In this study, we demonstrated that dual growth factor incorporated HP/GPT hydrogel could further improve erectile function in a rat model of bilateral cavernous nerve injury (BCNI). We showed that erectile function was decreased after BCNI, but it was further improved by treatment with a dual growth factor incorporated HP/GPT hydrogel compared with groups treated with single growth factor in a rat model of cavernous nerve injury. Also, we observed an increase in cyclic guanosine monophosphate (cGMP) levels in the dual growth factor group when compared with the groups treated with single growth factor. This effect was associated with greater upregulation of nitric oxide synthase and endothelial nitric oxide synthase expression in the penile tissue of the group treated with dual growth factor incorporated HP/GPT than in the other experimental groups. Apoptosis in the penile tissue treated with the dual growth factor incorporated HP/GPT hydrogel was lower than those treated singly with either bFGF or NGF incorporated GPT hydrogel. Both α-smooth muscle actin and CD31 expression increased in the group treated with dual growth factor incorporated HP/GPT hydrogel when compared to in the other experimental groups. Altogether, our results proved that the sequential and continuous release of growth factors from dual growth factor incorporated HP/GPT hydrogel prevented fibrosis and nerve damage induced by BCNI in the corpus cavernosum, and promoted the recovery of erectile function. Dual growth factor incorporated HP/GPT hydrogel may be a potent clinical application for the treatment of post-RP ED and could potentially be used various biomedical application in tissue regnerative medicine.

GV1001 Induces Apoptosis by Reducing Angiogenesis in Renal Cell Carcinoma Cells Both In Vitro and In Vivo.

OBJECTIVE: To investigate the anticancer effects of GV1001 and its biological mechanism of action in renal cell carcinoma (RCC). METHODS: The effects of GV1001 on cell survival and apoptosis in RCC cells were examined in vitro using cell viability assay, fluorescence-activated cell sorting, and terminal deoxynucleotidyl transferase dUTP nick end labeling assay. To evaluate the effect of GV1001 on migration, invasion, and angiogenesis, we used wound healing, invasion, endothelial cell tube formation assay, and western blot analysis. Furthermore, we used an RCC xenograft model with either phosphate buffered saline or GV1001 to confirm the anticancer effect of GV1001 in vivo. Tumor volume was monitored during treatment, and tumor weight was measured after animals were killed. Apoptosis and angiogenesis of the tumor tissue were assessed using hematoxylin and eosin staining, immunohistochemistry, and western blot analysis. RESULTS: GV1001 reduced cell viability and induced apoptosis in RCC cells in vitro. Furthermore, GV1001 suppressed the migration and invasion of RCC cells through regulation of matrix metalloproteinases and tissue inhibitors of metalloproteinases. In addition, GV1001 reduced angiogenesis via regulation of hypoxia-inducible factor 1α. In xenograft mouse model experiment, GV1001 reduced tumor growth and induced apoptosis. As in the in vitro results, GV1001 significantly reduced angiogenesis through regulation of hypoxia-inducible factor 1α in vivo. CONCLUSION: Our data demonstrated that GV1001 induced apoptosis through suppression of angiogenesis in RCCs both in vitro and in vivo, which suggests that GV1001 may be a potential therapeutic target for RCC.

Prostate-specific extracellular vesicles as a novel biomarker in human prostate cancer.

Extracellular vesicles (EVs) may play an important role in cancer development and progression. We aimed to investigate the prognostic potential of prostate-specific EVs in prostate cancer (PCa) patients. Plasma and prostate tissue were collected from patients who underwent surgery for PCa (n = 82) or benign prostatic hyperplasia (BPH, n = 28). To analyze the quantity of EVs in prostate, we performed transmission electron microscopy (TEM), immuno-TEM with CD63 and prostate-specific membrane antigen (PSMA), and immunofluorescence staining. After EV isolation from plasma, CD63 and PSMA concentration was measured using ELISA kits. PSMA-positive areas in prostate differed in patients with BPH, and low-, intermediate-, and high-risk PCa (2.4, 8.2, 17.5, 26.5%, p < 0.001). Plasma PSMA-positive EV concentration differed in patients with BPH, and low-, intermediate-, and high-risk PCa (21.9, 43.4, 49.2, 59.9 ng/mL, p < 0.001), and ROC curve analysis indicated that plasma PSMA-positive EV concentration differentiated PCa from BPH (AUC 0.943). Patients with lower plasma PSMA-positive EV concentration had greater prostate volume (50.2 vs. 33.4 cc, p < 0.001) and lower pathologic Gleason score (p = 0.025). During the median follow-up of 18 months, patients with lower plasma PSMA-positive EV concentration tended to have a lower risk of biochemical failure than those with higher levels of prostate-specific EVs (p = 0.085).

Combination Therapy Using Human Adipose-derived Stem Cells on the Cavernous Nerve and Low-energy Shockwaves on the Corpus Cavernosum in a Rat Model of Post-prostatectomy Erectile Dysfunction.

OBJECTIVE: To investigate combined therapeutic efficacy of human adipose-derived stem cells (h-ADSCs) application on injured cavernous nerve and low-energy shockwave therapy (SWT) on the corpus cavernosum in a rat model of post-prostatectomy erectile dysfunction. MATERIALS AND METHODS: Rats were randomly divided into 5 groups: control, bilateral cavernous nerve injury (BCNI), adipose-derived stem cell (ADSC) (BCNI group with h-ADSCs on the cavernous nerve), SWT (BCNI group with low-energy SWT on the corpus cavernosum), and ADSC/SWT (BCNI group with a combination of h-ADSCs and low-energy SWT). After 4 weeks, erectile function was assessed using intracavernosal pressure. The cavernous nerves and penile tissue were evaluated through immunostaining, Western blotting, and a cyclic guanosine monophosphate assay. RESULTS: ADSC/SWT significantly improved intracavernosal pressure compared to the other experimental group. ADSC had significantly increased ß-III tubulin expression of the cavernous nerve, and SWT had a markedly enhanced vascular endothelial growth factor expression in corpus cavernosum. The ADSC/SWT group had a significantly increased in alpha smooth muscle actin content (P < .05), neural nitric oxide synthase (nNOS) of the dorsal penile nerve (P < .05), endothelial nitric oxide synthase (eNOS) protein expression (P < .05), and cyclic guanosine monophosphate level (P < .05) compared to the ADSC or SWT alone group. In addition, ADSC/SWT reduces the apoptotic index in the corpus cavernosum. CONCLUSION: In this study, h-ADSCs showed an effect on the recovery of injured cavernous nerve and low-energy SWT improved angiogenesis in the corpus cavernosum. The h-ADSCs combined with low-energy SWT showed beneficial effect on the recovery of erectile function in a rat model of postprostatectomy erectile dysfunction.

Nanoengineered Polystyrene Surfaces with Nanopore Array Pattern Alters Cytoskeleton Organization and Enhances Induction of Neural Differentiation of Human Adipose-Derived Stem Cells.

Human adipose-derived stem cells (hADSCs) can differentiate into various cell types depending on chemical and topographical cues. One topographical cue recently noted to be successful in inducing differentiation is the nanoengineered polystyrene surface containing nanopore array-patterned substrate (NP substrate), which is designed to mimic the nanoscale topographical features of the extracellular matrix. In this study, efficacies of NP and flat substrates in inducing neural differentiation of hADSCs were examined by comparing their substrate-cell adhesion rates, filopodia growth, nuclei elongation, and expression of neural-specific markers. The polystyrene nano Petri dishes containing NP substrates were fabricated by a nano injection molding process using a nickel electroformed nano-mold insert (Diameter: 200 nm. Depth of pore: 500 nm. Center-to-center distance: 500 nm). Cytoskeleton and filopodia structures were observed by scanning electron microscopy and F-actin staining, while cell adhesion was tested by vinculin staining after 24 and 48 h of seeding. Expression of neural specific markers was examined by real-time quantitative polymerase chain reaction and immunocytochemistry. Results showed that NP substrates lead to greater substrate-cell adhesion, filopodia growth, nuclei elongation, and expression of neural specific markers compared to flat substrates. These results not only show the advantages of NP substrates, but they also suggest that further study into cell-substrate interactions may yield great benefits for biomaterial engineering.

Comparison between subcutaneous injection of basic fibroblast growth factor-hydrogel and intracavernous injection of adipose-derived stem cells in a rat model of cavernous nerve injury.

OBJECTIVE: To compare the effects of subcutaneous penile injection of basic fibroblast growth factor (bFGF)-hydrogel and intracavernous injection of human adipose-derived stem cells (h-ADSCs) on improving erectile function in a rat model of cavernous nerve injury. MATERIALS AND METHODS: Adult male Sprague-Dawley rats were randomly divided into 5 groups (n = 10 per group): age-matched control (normal group), bilateral cavernous nerve injury (BCNI group), penile subcutaneous injection of hydrogel after BCNI (hydrogel group), penile subcutaneous injection of bFGF-hydrogel after BCNI (bFGF-hydrogel group) and intracavernous injection of h-ADSCs after BCNI (ADSC group). Four weeks after the treatment, all rats underwent an erectile function test. Then, penile tissue was harvested for immunohistological analysis of bFGF, phalloidin, and cluster of differentiation (CD) 31. The cyclic guanosine monophosphate (cGMP) level of the corpus cavernosum was quantified by cGMP assay. RESULTS: From the functional test and immunohistological result, we observed that bFGF-hydrogel and h-ADSCs injection significantly elevated intracavernous pressure. The evaluation of filamentous actin content, CD31 expression, and cGMP concentration in the corpus cavernosum were meaningfully increased in the bFGF-hydrogel and ADSC groups compared with BCNI group. The bFGF released from bFGF-hydrogel prevented smooth muscle atrophy. Moreover, bFGF expression was significantly increased in bFGF-hydrogel group. CONCLUSION: The subcutaneous injection of bFGF-hydrogel prevented smooth muscle atrophy, increased the intracavernous pressure, and improved erectile function like an intracavernous injection of h-ADSCs.

The effect of PnTx2-6 protein from Phoneutria nigriventer spider toxin on improvement of erectile dysfunction in a rat model of cavernous nerve injury.

OBJECTIVE: To explore the therapeutic potential of PnTx2-6 injected 3 times a week for 4 weeks into the intracavernosal tissue in a rat model of bilateral cavernous nerve crush injury (BCNI). METHODS: Eight-week-old male Sprague-Dawley rats were randomly divided into the following 6 groups (n = 5 per group): age-matched control (normal group), BCNI (injury group), post-BCNI phosphate-buffered saline injection (PBS group), post-BCNI Sf9 cell-lysate injection (N/C group), post-BCNI injection of cell lysate from S9 cells infected with wild-type recombinant baculovirus (W/T group), and post-BCNI injection of cell lysate from S9 cells infected with recombinant baculovirus containing PnTx2-6 (PnTx2-6 group). Injections were delivered 3 times a week for 4 weeks. After 4 weeks, intracavernosal pressure-to-mean arterial pressure ratio, smooth muscle and collagen content via the Masson trichrome staining, levels of neural nitric oxide synthase, phosphoendothelial nitric oxide synthase, and cyclic guanosine monophosphate were all measured. RESULTS: The PnTx2-6 group showed significantly higher intracavernosal pressure-to-mean arterial pressure ratio (P <.05), smooth muscle-to-collagen ratio (P <.01), expression levels of neural nitric oxide synthase, phosphoendothelial nitric oxide synthase (P <.05), and cyclic guanosine monophosphate (P <.05) than all other experimental groups. CONCLUSION: We conclude that PnTx2-6 improved erectile function and prevented muscle atrophy in a rat model of BCNI via increased synthesis of nitric oxide and cyclic guanosine monophosphate.

Combined effects of brain-derived neurotrophic factor immobilized poly-lactic-co-glycolic acid membrane with human adipose-derived stem cells and basic fibroblast growth factor hydrogel on recovery of erectile dysfunction.

Erectile dysfunction (ED) is the most frequent long-term problem after radical prostatectomy. We aimed to evaluate whether the use of combination therapy with basic fibroblast growth factor (bFGF)-hydrogel on corpus cavernosum and with adipose-derived stem cells (ADSCs) and brain-derived neurotrophic factor (BDNF)-immobilized poly-lactic-co-glycolic acid (PLGA) membrane on the cavernous nerve (CN) could improve erectile function in a rat model of bilateral cavernous nerve crush injury (BCNI). Rats were randomly divided into five groups (n=15 per group): a normal group (N group), a group receiving saline application after bilateral cavernous nerve crush injury (BCNI), a group undergoing bFGF-hydrogel injection in the corpus cavernosum after BCNI (bFGF), a group receiving ADSC application covered with BDNF-membrane after BCNI (ADSC/BDNF), and a group undergoing coadministration of bFGF-hydrogel injection and BDNF-membrane with ADSCs after BDNF (bFGF+ADSC/BDNF). Four weeks postoperatively, the erectile function was assessed by detecting the ratio of intracavernous pressure (ICP) to mean arterial pressure (MAP). Smooth muscle and collagen contents were measured using Masson's trichrome staining. Neuronal nitric oxide synthase (nNOS) expression in the dorsal penile nerve was detected by immunostaining. The protein expression of the α-smooth muscle actin (α-SMA) and the cyclic guanosine monophosphate (cGMP) level of the corpus cavernosum were quantified by western blot and cGMP assay, respectively. In the bFGF+ADSC/BDNF group, the erectile function was significantly elevated compared with the BCNI and other treated groups and showed a significantly increased smooth muscle/collagen ratio, nNOS content, α-SMA expression, and cGMP level. In particular, there were no statistical differences in the ICP/MAP ratio, smooth muscle/collagen ratio, and α-SMA and cGMP levels between the bFGF+ADSC/BDNF group and normal group. Application of the BDNF-immobilized PLGA membrane with human ADSC into the CN and bFGF-incorporated hydrogel into the corpus carvernosum improved nearly normal erectile function in a rat model of postprostatectomy ED. This result suggests that a combined application of bFGF+ADSC/BDNF might be a promising treatment for postprostatectomy ED.

Combination therapy of human adipose-derived stem cells and basic fibroblast growth factor hydrogel in muscle regeneration.

Skeletal muscle regeneration after sport injury is inconsistent, and complete healing without fibrosis is very important. In this study, we determined whether the combination therapy using human adipose-derived stem cells (h-ADSCs) and basic fibroblast growth factor (bFGF) incorporated into hydrogel could enhance muscle regeneration in a muscle laceration animal model. The h-ADSCs and/or bFGF hydrogels were applied to the lacerated gastrocnemius muscle. Fast twitch muscle contraction improved significantly and fibrosis decreased significantly in combined h-ADSC and bFGF-hydrogel group compared to other experimental groups. Skeletal muscle differentiation of h-ADSCs was determined by immunohistochemistry (PKH-26/MyHC co-staining) and Western blot. Our data suggested that combination therapy of h-ADSCs and bFGF hydrogel resulted in functional recovery, revascularization and reinnervation with minimal fibrosis in lacerated muscle. A combination of h-ADSCs and bFGF hydrogel can be used as a promising therapy for skeletal muscle regeneration.