Purpose: N⁶-methyladenosine (m⁶A) is the most abundant internal modification on eukaryotic mRNA and gained increasing attention recently. More and more evidence suggest that m⁶A methylation plays crucial role in tumor genesis and development. However, its role in prostate cancer remains largely unknown. Methods: METTL3 expression status in prostate cancer was analyzed by using TCGA database and Western blotting. m⁶A content was analyzed by using RNA Methylation Quantification Kit. The role of METTL3 in prostate cancer cells was determined by proliferation, survival, colony formation, and invasion assays. The m⁶A level of GLI1 RNA was detected by methylated RNA immunoprecipitation (MeRIP) assay. In vivo role of METTL3 was studied on xenograft models. Results: We found that m⁶A methyltransferase METTL3 was overexpressed in prostate cancer cell lines, together with increased m⁶A content. Functionally, silencing of METTL3 by shRNA in prostate cancer cell lines resulted in decreased m⁶A content, cell proliferation, survival, colony formation, and invasion. Interestingly, overexpression of wild-type METTL3 abrogated the repression effect of METTL3 depletion on m⁶A content, cell proliferation, survival, colony formation, and invasion, while the overexpression of m⁶A catalytic site mutant METTL3 was unable to rescue the inhibitory effect caused by METTL3 depletion. Further mechanism analysis demonstrated that METTL3 silence decreased the m⁶A modification and expression of GLI1, an important component of hedgehog pathway, which led to cell apoptosis. Moreover, depletion of METTL3 inhibited tumor growth in vivo. Conclusion: Our results suggested that the m⁶A methyltransferase METTL3 promotes the growth and motility of prostate cancer cells by regulating hedgehog pathway.

Objectives: Although evidence supports a role for inflammation in interstitial cystitis/bladder pain syndrome (IC/BPS), the mechanism remains unknown. We determined whether inflammation causes an elevated expression of nerve growth factor (NGF) and transient receptor potential vanilloid receptor subtype 1 (TRPV1) and correlated them with the symptoms. Methods: Bladder biopsies were obtained from 53 IC/BPS patients and 27 controls, and hematoxylin and eosin staining, immunostaining and Western blotting were performed to detect inflammation, TRPV1-immunoreactive and PGP9.5-immunoreactive nerve fibers, and NGF, respectively. Symptoms were assessed using the Pelvic Pain/Urgency/Frequency (PUF) questionnaire and pain visual analogue scale scores. Suburothelial nerve fiber density was quantified and correlated with PUF scores. Results: Increased severity of inflammation was correlated with a higher TRPV1-immunoreactive nerve fiber density (r = 0.4113, p = 0.0024) and higher NGF levels (r = 0.3775, p = 0.0052). Suburothelial TRPV1-immunoreactive nerve fiber density was significantly correlated with pain scores and urgency scores (r = 0.3320, p = 0.0145 and r = 0.3823, p = 0.0039, respectively). PGP9.5-immunoreactive nerve fibers were significantly increased in IC/BPS (p = 0.0193) and had a positive relationship with inflammation severity (r = 0.6138, p < 0.0001). Conclusions: Our study revealed increased severity of inflammation correlated with a higher expression of TRPV1-immunoreactive nerve fibers and NGF in IC/BPS and correlated with clinical symptoms.

Nanoscopic therapeutic systems that incorporate therapeutic agents, molecular targeting, and imaging capabilities have gained momentum and exhibited significant therapeutic potential. In this study, multifunctional polymeric nanoparticles with controlled drug delivery, cancer‐targeted capability, and efficient magnetic resonance imaging (MRI) contrast characteristics were formulated and applied in the treatment of castration‐resistant prostate cancer (CRPC). The “core‐shell” targeted nanoparticles (NPs) were synthesized by the self‐assembly of a prefunctionalized amphiphilic triblock copolymer composed of poly(lactic‐co‐glycolic‐acid) (PLGA), polyethylene glycol (PEG), and the Wy5a aptamer (Apt), which have been screened for targeting the CRPC cell line PC‐3 by cell‐SELEX technique as described in our previous study. Docetaxel (Dtxl) and a cluster of hydrophobic superparamagnetic iron oxide (SPIO) nanoparticles were simultaneously encapsulated into the targeted nanoparticles. The targeted NPs showed a controlled drug release and an increased contrast‐enhanced MRI capability. The presence of Wy5a on the nanoparticle surface resulted in the cancer‐targeted delivery to PC‐3 cells in vitro and in vivo. In vitro MRI and cytotoxicity studies demonstrated the ultrasensitive MRI and increased cytotoxicity of these targeted NPs. In vivo studies revealed that the targeted NPs exhibited a more efficacious antitumor capability without significant systemic toxicity. Our data suggested that these targeted NPs may be a promising drug delivery system for the efficacious treatment of CRPC.

Background: Previous studies have demonstrated that glial cells play an important role in the generation and maintenance of neuropathic pain. Activated glial cells produce numerous mediators such as proinflammatory cytokines that facilitate neuronal activity and synaptic plasticity. Similarly, bladder pain syndrome/interstitial cystitis shares many characteristics of neuropathic pain. However, related report on the involvement of spinal glia in bladder pain syndrome/interstitial cystitis-associated pathological pain and the underlying mechanisms are still lacking. The present study investigated spinal glial activation and underlying molecular mechanisms in a rat model of bladder pain syndrome/interstitial cystitis. Results: A rat model of bladder pain syndrome/interstitial cystitis was established via systemic injection with cyclophosphamide. Mechanical allodynia was tested with von Frey monofilaments and up-down method. Moreover, Western blots and double immunofluorescence were used to detect the expression and location of glial fibrillary acidic protein, OX42/Iba1, P-P38, NeuN, interleukin (IL)-1β, phosphorylation of N-methyl-D-aspartate receptor 1 (P-NR1), and IL-1 receptor I (IL-1RI) in the L6-S1 spinal cord. We found that glial fibrillary acidic protein rather than OX42/Iba1 or P-P38 was significantly increased in the spinal cord of cyclophosphamide-induced cystitis. L-alpha-aminoadipate but not minocycline markedly attenuated the allodynia. Furthermore, we found that spinal IL-1β was dramatically increased in cyclophosphamide-induced cystitis, and activated astrocytes were the only source of IL-1β release, which contributed to allodynia in cystitis rats. Besides, spinal P-NR1 was statistically increased in cyclophosphamide-induced cystitis and only localized in IL-1RI positive neurons in spinal dorsal horn. Additionally, NR antagonist significantly attenuated the cystitis-induced pain. Interestingly, the time course of the P-NR1 expression paralleled to that of IL-1β or glial fibrillary acidic protein. Conclusions: Our results demonstrated that astrocytic activation but not microglial activation contributed to the allodynia in cyclophosphamide-induced cystitis and IL-1β released from astrocytes might bind to its endogenous receptor on the neurons inducing the phosphorylation of NR1 subunit, leading to sensory neuronal hyperexcitability and pathological pain.