Reprogramming by drug-like molecules leads to regeneration of cochlear hair cell-like cells in adult mice.

Strategies to overcome irreversible cochlear hair cell (HC) damage and loss in mammals are of vital importance to hearing recovery in patients with permanent hearing loss. In mature mammalian cochlea, co-activation of Myc and Notch1 reprograms supporting cells (SC) and promotes HC regeneration. Understanding of the underlying mechanisms may aid the development of a clinically relevant approach to achieve HC regeneration in the nontransgenic mature cochlea. By single-cell RNAseq, we show that MYC/NICD "rejuvenates" the adult mouse cochlea by activating multiple pathways including Wnt and cyclase activator of cyclic AMP (cAMP), whose blockade suppresses HC-like cell regeneration despite Myc/Notch activation. We screened and identified a combination (the cocktail) of drug-like molecules composing of small molecules and small interfering RNAs to activate the pathways of Myc, Notch1, Wnt and cAMP. We show that the cocktail effectively replaces Myc and Notch1 transgenes and reprograms fully mature wild-type (WT) SCs for HC-like cells regeneration in vitro. Finally, we demonstrate the cocktail is capable of reprogramming adult cochlea for HC-like cells regeneration in WT mice with HC loss in vivo. Our study identifies a strategy by a clinically relevant approach to reprogram mature inner ear for HC-like cells regeneration, laying the foundation for hearing restoration by HC regeneration.

SIRT3 inhibits prostate cancer metastasis through regulation of FOXO3A by suppressing Wnt/ß-catenin pathway.

SIRT3, a mitochondrial NAD+-dependent deacetylase, has been reported to restrain prostate cancer growth both in vitro and in vivo, however, its role in metastatic prostate cancer has not been revealed. In this study, we reported that SIRT3 inhibited the epithelial-mesenchymal transition (EMT) and migration of prostatic cancer cells in vitro and their metastasis in vivo. Consistently, based on analyses of tissue microarray and microarray datasets, lower SIRT3 expression level was correlated with higher prostate cancer Gleason scores, and SIRT3 expression were significantly decreased in metastatic tissues compared with prostate tumor tissues. Mechanistically, SIRT3 promoted FOXO3A expression by attenuating Wnt/ß-catenin pathway, thereby inhibiting EMT and migration of prostate cancer cells. Indeed, SIRT3's inhibitory effect on EMT and migration of prostate cancer cells can be rescued after applying Wnt/ß-catenin pathway activator LiCl, or boosted by wnt inhibitor XAV939. Together, this study revealed a novel mechanism for prostate cancer metastasis that involves SIRT3/ Wnt/ß-catenin/ FOXO3A signaling to modulate EMT and cell migration.

Demethylzeylasteral exerts potent efficacy against non-small-cell lung cancer via the P53 signaling pathway.

Lung cancer has one of the highest mortality rates worldwide, with non-small-cell lung cancer (NSCLC) constituting approximately 85% of all cases. Demethylzeylasteral (DEM), extracted from Tripterygium wilfordii Hook F, exhibits notable anti-tumor properties. In this study, we revealed that DEM could effectively induce NSCLC cell apoptosis. Specifically, DEM can dose-dependently suppress the viability and migration of human NSCLC cells. RNA-seq analysis revealed that DEM regulates the P53-signaling pathway, which was further validated by assessing crucial proteins involved in this pathway. Biacore analysis indicated that DEM has high affinity with the P53 protein. The CDX model demonstrated DEM's anti-tumor actions. This work provided evidence that DEM-P53 interaction stabilizes P53 protein and triggers downstream anti-tumor activities. These findings indicate that DEM treatment holds promise as a potential therapeutic approach for NSCLC, which warrants further clinical assessment in patients with NSCLC.

Non-flammable solvent-free liquid polymer electrolyte for lithium metal batteries.

As a replacement for highly flammable and volatile organic liquid electrolyte, solid polymer electrolyte shows attractive practical prospect in high-energy lithium metal batteries. However, unsatisfied interface performance and ionic conductivities are two critical challenges. A common strategy involves introducing organic solvents or plasticizers, but this violates the original intention of security design. Here, an electrolyte concept called liquid polymer electrolyte without any small molecular solvents is proposed for safe and high-performance batteries, based on the design of a room-temperature liquid-state brush-like polymer as the sole solvent of lithium salts. This liquid polymer electrolyte is non-flammable and exhibits high ionic conductivity (1.09 [Formula: see text] 10-4 S cm-1 at 25 °C), significant lithium dendrite suppression, and stable long-term cycling over a wide operating temperature range ( ≥ 1000 cycles at 60 °C and 90 °C). Moreover, the pouch cell can resist thermal abuse, vacuum environment, and mechanical abuse. This electrolyte and design strategy are expected to provide enlightening ideas for the development of safe and high-performance polymer electrolytes.

A Novel in vitro Model Delineating Hair Cell Regeneration and Neural Reinnervation in Adult Mouse Cochlea.

The study of an adult mammalian auditory system, such as regeneration, has been hampered by the lack of an in vitro system in which hypotheses can be tested efficiently. This is primarily due to the fact that the adult inner ear is encased in the toughest bone of the body, whereas its removal leads to the death of the sensory epithelium in culture. We hypothesized that we could take advantage of the integral cochlear structure to maintain the overall inner ear architecture and improve sensory epithelium survival in culture. We showed that by culturing adult mouse cochlea with the (surrounding) bone intact, the supporting cells (SCs) survived and almost all hair cells (HCs) degenerated. To evaluate the utility of the explant culture system, we demonstrated that the overexpression of Atoh1, an HC fate-determining factor, is sufficient to induce transdifferentiation of adult SCs to HC-like cells (HCLCs). Transdifferentiation-derived HCLCs resemble developmentally young HCs and are able to attract adult ganglion neurites. Furthermore, using a damage model, we showed that degenerated adult ganglions respond to regenerated HCLCs by directional neurite outgrowth that leads to HCLC-neuron contacts, strongly supporting the intrinsic properties of the HCLCs in establishing HCLC-neuron connections. The adult whole cochlear explant culture is suitable for diverse studies of the adult inner ear including regeneration, HC-neuron pathways, and inner ear drug screening.

Stepwise Induction of Inner Ear Hair Cells From Mouse Embryonic Fibroblasts via Mesenchymal- to-Epithelial Transition and Formation of Otic Epithelial Cells.

Although embryonic stem cells or induced pluripotent stem cells are able to differentiate into inner ear hair cells (HCs), they have drawbacks limiting their clinical application, including a potential risk of tumourigenicity. Direct reprogramming of fibroblasts to inner ear HCs could offer an alternative solution to this problem. Here, we present a stepwise guidance protocol to induce mouse embryonic fibroblasts to differentiate into inner ear HC-like cells (HCLs) via mesenchymal-to-epithelial transition and then acquisition of otic sensory epithelial cell traits by overexpression of three key transcription factors. These induced HCLs express multiple HC-specific proteins, display protrusions reminiscent of ciliary bundle structures, respond to voltage stimulation, form functional mechanotransduction channels, and exhibit a transcriptional profile of HC signature. Together, our work provides a new method to produce functional HCLs in vitro, which may have important implications for studies of HC development, drug discovery, and cell replacement therapy for hearing loss.

Novel sulfonated poly(ether ether ketone)/triphenylamine hybrid membrane for vanadium redox flow battery applications.

A novel sulfonated poly(ether ether ketone)/triphenylamine hybrid membrane with various triphenylamine loadings (1%, 2% and 5%) has been successfully fabricated. Optimum triphenylamine loading was confirmed by exploring the physicochemical properties and morphology of different membranes. The hybrid membrane exhibited lower vanadium permeability than pristine SPEEK membranes due to the acid-base interaction between amine groups and sulfonated groups. Introduction of triphenylamine also improved the proton conductivity because the nitrogen atom of triphenylamine can be protonated and contribute to the proton transfer. As the result, the hybrid membrane demonstrated higher ion selectivity compared with SPEEK and Nafion115 membranes. The VRFB single cell with SPEEK/TPAM-1% membrane showed better performance compared to a Nafion115 membrane at the current density of 60 mA cm-2. The SPEEK/TPAM hybrid membrane has great potential for VRFB application.

Renewed proliferation in adult mouse cochlea and regeneration of hair cells.

The adult mammalian inner ear lacks the capacity to divide or regenerate. Damage to inner ear generally leads to permanent hearing loss in humans. Here, we present that reprogramming of the adult inner ear induces renewed proliferation and regeneration of inner ear cell types. Co-activation of cell cycle activator Myc and inner ear progenitor gene Notch1 induces robust proliferation of diverse adult cochlear sensory epithelial cell types. Transient MYC and NOTCH activities enable adult supporting cells to respond to transcription factor Atoh1 and efficiently transdifferentiate into hair cell-like cells. Furthermore, we uncover that mTOR pathway participates in MYC/NOTCH-mediated proliferation and regeneration. These regenerated hair cell-like cells take up the styryl dye FM1-43 and are likely to form connections with adult spiral ganglion neurons, supporting that Myc and Notch1 co-activation is sufficient to reprogram fully mature supporting cells to proliferate and regenerate hair cell-like cells in adult mammalian auditory organs.

Inactivation of STAT3 Signaling Impairs Hair Cell Differentiation in the Developing Mouse Cochlea.

Although STAT3 signaling is demonstrated to regulate sensory cell differentiation and regeneration in the zebrafish, its exact role is still unclear in mammalian cochleae. Here, we report that STAT3 and its activated form are specifically expressed in hair cells during mouse cochlear development. Importantly, conditional cochlear deletion of Stat3 leads to an inhibition on hair cell differentiation in mice in vivo and in vitro. By cell fate analysis, inactivation of STAT3 signaling shifts the cell division modes from asymmetric to symmetric divisions from supporting cells. Moreover, inhibition of Notch signaling stimulates STAT3 phosphorylation, and inactivation of STAT3 signaling attenuates production of supernumerary hair cells induced by a Notch pathway inhibitor. Our findings highlight an important role of the STAT3 signaling during mouse cochlear hair cell differentiation and may have clinical implications for the recovery of hair cell loss-induced hearing impairment.

Regulation of Prostate Development and Benign Prostatic Hyperplasia by Autocrine Cholinergic Signaling via Maintaining the Epithelial Progenitor Cells in Proliferating Status.

Regulation of prostate epithelial progenitor cells is important in prostate development and prostate diseases. Our previous study demonstrated a function of autocrine cholinergic signaling (ACS) in promoting prostate cancer growth and castration resistance. However, whether or not such ACS also plays a role in prostate development is unknown. Here, we report that ACS promoted the proliferation and inhibited the differentiation of prostate epithelial progenitor cells in organotypic cultures. These results were confirmed by ex vivo lineage tracing assays and in vivo renal capsule recombination assays. Moreover, we found that M3 cholinergic receptor (CHRM3) was upregulated in a large subset of benign prostatic hyperplasia (BPH) tissues compared with normal tissues. Activation of CHRM3 also promoted the proliferation of BPH cells. Together, our findings identify a role of ACS in maintaining prostate epithelial progenitor cells in the proliferating state, and blockade of ACS may have clinical implications for the management of BPH.

Autocrine Activation of CHRM3 Promotes Prostate Cancer Growth and Castration Resistance via CaM/CaMKK-Mediated Phosphorylation of Akt.

PURPOSE: Although a previous study reported nerve ending-derived acetylcholine promoted prostate cancer invasion and metastasis by regulating the microenvironment of cancer cells, the present study aims to determine whether there is autocrine cholinergic signaling in prostate epithelial cells that promotes prostate cancer growth and castration resistance. EXPERIMENTAL DESIGN: In this study, IHC was performed to detect protein expression in mouse prostate tissue sections and human prostate cancer tissue sections. Subcutaneously and orthotopically xenografted tumor models were established to evaluate the functions of autocrine cholinergic signaling in regulating prostate cancer growth and castration resistance. Western blotting analysis was performed to assess the autocrine cholinergic signaling-induced signaling pathway. RESULTS: We found the expression of choline acetyltransferase (ChAT), the secretion of acetylcholine and the expression of CHRM3 in prostate epithelial cells, supporting the presence of autocrine cholinergic signaling in the prostate epithelium. In addition, we found that CHRM3 was upregulated in clinical prostate cancer tissues compared with adjacent non-cancer tissues. Overexpression of CHRM3 or activation of CHRM3 by carbachol promoted cell proliferation, migration, and castration resistance. On the contrary, blockading CHRM3 by shRNA or treatment with darifenacin inhibited prostate cancer growth and castration resistance both in vitro and in vivo. Furthermore, we found that autocrine cholinergic signaling caused calmodulin/calmodulin-dependent protein kinase kinase (CaM/CaMKK)-mediated phosphorylation of Akt. CONCLUSIONS: These findings suggest that blockade of CHRM3 may represent a novel adjuvant therapy for castration-resistant prostate cancer.

Adjudin protects rodent cochlear hair cells against gentamicin ototoxicity via the SIRT3-ROS pathway.

Hearing loss resulting from hair cell degeneration is a common disease that affects millions of people worldwide. Strategies to overcome the apparent irreversible hair cell loss in mammals become paramount for hearing protection. Here we reported that, by using a well-established gentamicin-induced hair cell loss model in vitro, adjudin, a multi-functional small molecule drug, protected cochlear hair cells from gentamicin damage. Immunohistochemistry, Western blotting and quantitative RT-PCR analyses revealed that adjudin exerted its otoprotective effects by up-regulating the level of Sirt3, a member of Sirtuin family protein located in mitochondria, which regulates reactive oxygen species (ROS) production in cochlear cells and inhibits the production of ROS and apoptotic cells induced by gentamicin. Sirt3 silencing experiments confirmed that Sirt3-ROS signaling axis mediated hair cell protection against gentamicin by adjudin, at least in part. Furthermore, adjudin's otoprotection effects were also observed in an in vivo gentamicin-injured animal model. Taken together, these findings identify adjudin as a novel otoprotective small molecule via elevating Sirt3 levels and Sirt3 may be of therapeutic value in hair cell protection from ototoxic insults.

SIRT3 inhibits prostate cancer by destabilizing oncoprotein c-MYC through regulation of the PI3K/Akt pathway.

SIRT3 is involved in aging-related diseases including cancer, but its role in prostate cancer and detailed regulatory function are not known. We found that SIRT3 was moderately down-regulated in prostate carcinomas. Overexpression of SIRT3 by lentiviral transfection inhibited prostate cancer growth both in vitro and in vivo, whereas knockdown of SIRT3 increased prostate tumor growth. Mechanistically, the tumor suppression effect of SIRT3 was achieved via its inhibition of the PI3K/Akt pathway. Notably, upregulation of SIRT3 suppressed the phosphorylation of Akt, leading to the ubiquitination and degradation of oncoprotein c-MYC; this could be attenuated by constitutive activation of PI3K/Akt signaling. Collectively, our results unveiled SIRT3's tumor suppressive function and the underlying mechanism in prostate cancer, which might provide therapeutic implications for the disease.

Adjudin--A Male Contraceptive with Other Biological Activities.

BACKGROUND: Adjudin has been explored as a male contraceptive for the last 15 years since its initial synthesis in the late 1990s. More than 50 papers have been published and listed in PubMed in which its mechanism that induces exfoliation of germ cells from the seminiferous epithelium, such as its effects on actin microfilaments at the apical ES (ectoplasmic specialization, a testis-specific actin-rich anchoring junction) has been delineated. OBJECTIVE: Recent studies have demonstrated that, besides its activity to induce germ cell exfoliation from the seminiferous epithelium to cause reversible infertility in male rodents, adjudin possesses other biological activities, which include anti-cancer, anti-inflammation in the brain, and anti-ototoxicity induced by gentamicin in rodents. Results of these findings likely spark the interest of investigators to explore other medical use of this and other indazole-based compounds, possibly mediated by the signaling pathway(s) in the mitochondria of mammalian cells following treatment with adjudin. In this review, we carefully evaluate these recent findings. METHODS: Papers published and listed at www.pubmed.org and patents pertinent to adjudin and its related compounds were searched. Findings were reviewed and critically evaluated, and summarized herein. RESULTS: Adjudin is a novel compound that possesses anti-spermatogenetic activity. Furthermore, it possesses anti-cancer, anti-inflammation, anti-neurodegeneration, and anti-ototoxicity activities based on studies using different in vitro and in vivo models. CONCLUSION: Studies on adjudin should be expanded to better understand its biological activities so that it can become a useful drug for treatment of other ailments besides serving as a male contraceptive.