Profibrotic Subsets of SPP1+ Macrophages and POSTN+ Fibroblasts Contribute to Fibrotic Scarring in Acne Keloidalis.

Acne keloidalis is a primary scarring alopecia characterized by longstanding inflammation in the scalp causing keloid-like scar formation and hair loss. Histologically, acne keloidalis is characterized by mixed leukocytic infiltrates in the acute stage followed by a granulomatous reaction and extensive fibrosis in the later stages. To further explore its pathogenesis, bulk RNA sequencing, single-cell RNA sequencing, and spatial transcriptomics were applied to occipital scalp biopsy specimens of lesional and adjacent no-lesional skin in patients with clinically active disease. Unbiased clustering revealed 19 distinct cell populations, including 2 notable populations: POSTN+ fibroblasts with enriched extracellular matrix signatures and SPP1+ myeloid cells with an M2 macrophage phenotype. Cell communication analyses indicated that fibroblasts and myeloid cells communicated by SPP1 signaling networks in lesional skin. A reverse transcriptomics in silico approach identified corticosteroids as possessing the capability to reverse the gene expression signatures of SPP1+ myeloid cells and POSTN+ fibroblasts. Intralesional corticosteroid injection greatly reduced SPP1 and POSTN gene expression as well as acne keloidalis disease activity. Spatial transcriptomics and immunofluorescence staining verified microanatomic specificity of SPP1+ myeloid cells and POSTN+ fibroblasts with disease activity. In summary, the communication between POSTN+ fibroblasts and SPP1+ myeloid cells by SPP1 axis may contribute to the pathogenesis of acne keloidalis.

Amelioration of the brain structural connectivity is accompanied with changes of gut microbiota in a tuberous sclerosis complex mouse model.

Tuberous sclerosis complex (TSC) is a genetic disease that causes benign tumors and dysfunctions in many organs, including the brain. Aside from the brain malformations, many individuals with TSC exhibit neuropsychiatric symptoms. Among these symptoms, autism spectrum disorder (ASD) is one of the most common co-morbidities, affecting up to 60% of the population. Past neuroimaging studies strongly suggested that the impairments in brain connectivity contribute to ASD, whether or not TSC-related. Specifically, the tract-based diffusion tensor imaging (DTI) analysis provides information on the fiber integrity and has been used to study the neuropathological changes in the white matter of TSC patients with ASD symptoms. In our previous study, curcumin, a diet-derived mTOR inhibitor has been shown to effectively mitigate learning and memory deficits and anxiety-like behavior in Tsc2+/- mice via inhibiting astroglial proliferation. Recently, gut microbiota, which is greatly influenced by the diet, has been considered to play an important role in regulating several components of the central nervous system, including glial functions. In this study, we showed that the abnormal social behavior in the Tsc2+/- mice can be ameliorated by the dietary curcumin treatment. Second, using tract-based DTI analysis, we found that the Tsc2+/- mice exhibited altered fractional anisotropy, axial and radial diffusivities of axonal bundles connecting the prefrontal cortex, nucleus accumbens, hypothalamus, and amygdala, indicating a decreased brain network. Third, the dietary curcumin treatment improved the DTI metrics, in accordance with changes in the gut microbiota composition. At the bacterial phylum level, we showed that the abundances of Actinobacteria, Verrucomicrobia, and Tenericutes were significantly correlated with the DTI metrics FA, AD, and RD, respectively. Finally, we revealed that the expression of myelin-associated proteins, myelin bassic protein (MBP) and proteolipid protein (PLP) was increased after the treatment. Overall, we showed a strong correlation between structural connectivity alterations and social behavioral deficits, as well as the diet-dependent changes in gut microbiota composition.

Recombinant Klotho attenuates IFNγ receptor signaling and SAMHD1 expression through blocking NF-κB translocation in glomerular mesangial cells.

Interferon gamma (IFNγ) is a cytokine implicated in the pathogenesis of autoimmune diseases. SAM and HD domain-containing protein 1 (SAMHD1) is an IFNγ-inducible protein that modulates cellular dNTP levels. Mutations in the human SAMHD1 gene cause Aicardi-Goutières (AG) syndrome, an autoimmune disease sharing similar clinical features with systemic lupus erythematosus (SLE). Klotho is an anti-inflammatory protein which suppresses aging through multiple mechanisms. Implication of Klotho in autoimmune response is identified in rheumatologic diseases such as SLE. Little information exists regarding the effect of Klotho in lupus nephritis, one of the prevalent symptoms of SLE. The present study verified the effect of IFNγ on SAMHD1 and Klotho expression in MES-13 glomerular mesangial cells, a special cell type in glomerulus that is critically involved in lupus nephritis. IFNγ upregulated SAMHD1 expression in MES-13 cells through the Janus kinase-signal transducer and activator of transcription 1 (JAK-STAT1) and the nuclear factor kappa B (NFκB) signaling pathways. IFNγ decreased Klotho protein expression in MES-13 cells. Treatment of MES-13 cells with recombinant Klotho protein inhibited SAMHD1 expression by blocking IFNγ-induced NFκB nuclear translocation, but showed no effect on JAK-STAT1 signaling. Collectively, our findings support the protective role of Klotho in attenuating lupus nephritis through the inhibition of IFNγ-induced SAMHD1 expression and IFNγ downstream signaling in MES-13 cells.

Detection of endophenotypes associated with neuropsychiatric deficiencies in a mouse model of tuberous sclerosis complex using diffusion tensor imaging.

Tuberous sclerosis complex (TSC) is a rare hereditary disease, which results from the mutation of either TSC1 or TSC2, and its clinical features include benign tumors and dysfunctions in numerous organs, including the brain. Many individuals with TSC manifest neuropsychiatric symptoms, such as learning impairments, cognitive deficits and anxiety. Current pharmacological treatment for TSC is the use of mTOR inhibitors. However, they are not effective in treating neuropsychiatric symptoms. We previously used curcumin, a diet-derived mTOR inhibitor, which possesses both anti-inflammatory and antiproliferative properties, to improve learning and memory deficits in Tsc2+/- mice. Diffusion tensor imaging (DTI) provides microstructural information in brain tissue and has been used to study the neuropathological changes in TSC. In this study, we confirmed that the impaired recognition memory and increased anxiety-like behavior in Tsc2+/- mice can be reversed by curcumin treatment. Second, we found altered fractional anisotropy and mean diffusivity in the anterior cingulate cortex and the hippocampus of the Tsc2+/- mice, which may indicate altered circuitry. Finally, the mTOR complex 1 hyperactivity was found in the cortex and hippocampus, coinciding with abnormal cortical myelination and increased glial fibrillary acidic protein expression in the hippocampal CA1 of Tsc2+/- mice, both of which can be rescued with curcumin treatment. Overall, DTI is sensitive to the subtle alterations that cannot be detected by conventional imaging, suggesting that noninvasive DTI may be suitable for longitudinally monitoring the in vivo neuropathology associated with the neuropsychiatric symptoms in TSC, thereby facilitating future clinical trials of pharmacological treatments.

Purα regulates the induction of Znf179 transcription during neuronal differentiation.

Development of the mammalian central nervous system is an important process, which is accomplished through precise regulations of many different genes. Zinc finger protein 179 (Znf179) is one of the essential genes that plays a critical role in neuronal differentiation. In our previous study, Znf179 knockout mice displayed brain malformation and impaired brain functions. We have also previously shown that Znf179 involves in cell cycle regulation, but the regulatory mechanism of Znf179 expression is not yet fully characterized. Herein, we identified that Purα is an essential factor for the promotor activity of Znf179. We also showed concurrent expression of Znf179 and Purα during neuronal differentiation. We also found that overexpression of Purα increased Znf179 expression in neuronal differentiated P19 cells. Through its direct binding to Znf179, as shown using DAPA, Purα upregulates Znf179 expression, suggesting that Purα is important for the regulation of Znf179 expression during neuronal differentiation. Our data indicated that Purα is involved in the transcriptional regulation of Znf179 gene during neuronal differentiation, and is indispensable during the brain development.

Promyelocytic leukemia zinc finger is involved in the formation of deep layer cortical neurons.

BACKGROUND: Promyelocytic leukemia zinc finger (Plzf), a transcriptional regulator involved in a lot of important biological processes during development, has been implied to maintain neural stem cells and inhibit their differentiation into neurons. However, the effects of Plzf on brain structures and functions are still not clarified. RESULTS: We showed that Plzf expression was detected as early as embryonic day (E) 9.5 in Pax6+ cells in the mouse brain, and was completely disappeared in telencephalon before the initiation of cortical neurogenesis. Loss of Plzf resulted in a smaller cerebral cortex with a decrease in the number of Tbr1+ deep layer neurons due to a decrease of mitotic cell number in the ventricular zone of forebrain at early developmental stage. Microarray, qRT-PCR, and flow cytometry analysis identified dysregulation of Mash1 proneural gene expression. We also observed an impairment of recognition memory in Plzf-deficient mice. CONCLUSIONS: Plzf is expressed at early stages of brain development and involved in the formation of deep layer cortical neurons. Loss of Plzf results in dysregulation of Mash1, microcephaly with reduced numbers of early-born neurons, and impairment of recognition memory.

Morphological control of mitochondria as the novel mechanism of Gastrodia elata in attenuating mutant huntingtin-induced protein aggregations.

BACKGROUND: According to Compendium of Materia Medica, Gastrodia elata (GE) Blume as a top grade and frequently prescribed herbal medicine has been used in treating dizziness, headaches, and epilepsy, indicating a neuroprotective effect. Because GE is capable of suppressing a hyperactive liver and thus calming endogenous wind, and because Huntington's disease (HD) can be classified as a phenomenon of disturbed liver wind, it is suggested that GE might be beneficial in treating HD. However, although current studies support GE for the prevention of diverse neurodegenerations such as HD, its detailed mechanisms remain elusive. PURPOSE: To investigate the molecular mechanism of GE in preventing HD by focusing on mitochondrial morphology, which is highly associated with HD etiology and thus proposed as a therapeutic target of neurodegenerations. STUDY DESIGN/METHODS: The overexpression of the mutant huntingtin (mHTT) gene in rat pheochromocytoma (PC12) cells was used as an in vitro cell model of HD. A filter retardation assay was applied to measure protein aggregations during HTT expression. Cotransfection with mitochondrial fusion and fission genes was used to test their relationships with HTT aggregates by monitoring with a confocal laser scanning microscope and filter retardation assay. Western blot analysis was used to estimate protein expression under different drug treatments or cotransfections with other related genes. RESULTS: The overexpression of mutant but not normal HTT genes significantly resulted in protein aggregations in PC12 cells. GE dose-dependently attenuated mHTT-induced protein aggregations and free radical formations. GE significantly reversed mHTT-induced mitochondrial fragmentation and dysregulation of mitochondrial fusion and fission molecules. The overexpression of mitochondrial fusion genes attenuated mHTT-induced protein aggregations. Further, Mdivi-1, a DRP1 fission molecule inhibitor, significantly reversed mHTT-induced protein aggregations and mitochondrial fragmentation. CONCLUSION: GE attenuated mHTT aggregations through the control of mitochondrial fusion and the fission pathway.

Ketamine ameliorates severe traumatic event-induced antidepressant-resistant depression in a rat model through ERK activation.

Treatment-resistant depression (TRD) is a major public health issue, as it is common for patients with depression to fail to respond to adequate trials of antidepressants. However, a well-established animal model of TRD is still warranted. The present study focused on selective serotonin reuptake inhibitor (SSRI) resistance, and aimed to investigate whether higher levels of traumatic stress caused by greater numbers of foot-shocks may lead to severe depression and to examine the feasibility of this as an animal model of SSRI-resistant depression. To reveal the correlation between traumatic stress and severe depression, rats received 3, 6 and 10 tone (conditioned stimulus, CS)-shock (unconditioned stimulus, US) pairings to mimic mild, moderate, and severe traumatic events, and subsequent depressive-like behaviors and protein immunocontents were analyzed. The antidepressant efficacy was assessed for ketamine and SSRI (i.e., fluoxetine) treatment. We found that only the severe stress group presented depressive-like behaviors. Phosphorylation of extracellular signal-regulated kinases (ERKs) was decreased in the amygdala and prefrontal cortex (PFC). The immunocontents of GluA1 and PSD 95 were increased in the amygdala and decreased in the PFC. Moreover, the glutamate-related abnormalities in the amygdala and PFC were normalized by single-dose (10 mg/kg, i.p.) ketamine treatment. In contrast, the depressive-like behaviors were not reversed by 28 days of fluoxetine treatment (10 mg/kg, i.p.) in the severe stress group. Our data demonstrated that high levels of traumatic stress could lead to SSRI-resistant depressive symptoms through impacts on the glutamatergic system, and that this rat model has the potential to be a feasible animal model of SSRI-resistant depression.

Leukemia Inhibitory Factor Promotes Castration-resistant Prostate Cancer and Neuroendocrine Differentiation by Activated ZBTB46.

PURPOSE: The molecular targets for castration-resistant prostate cancer (CRPC) are unknown because the disease inevitably recurs, and therapeutic approaches for patients with CRPC remain less well understood. We sought to investigate regulatory mechanisms that result in increased therapeutic resistance, which is associated with neuroendocrine differentiation of prostate cancer and linked to dysregulation of the androgen-responsive pathway. EXPERIMENTAL DESIGN: The underlying intracellular mechanism that sustains the oncogenic network involved in neuroendocrine differentiation and therapeutic resistance of prostate cancer was evaluated to investigate and identify effectors. Multiple sets of samples with prostate adenocarcinomas and CRPC were assessed via IHC and other assays. RESULTS: We demonstrated that leukemia inhibitory factor (LIF) was induced by androgen deprivation therapy (ADT) and was upregulated by ZBTB46 in prostate cancer to promote CRPC and neuroendocrine differentiation. LIF was found to be induced in patients with prostate cancer after ADT and was associated with enriched nuclear ZBTB46 staining in high-grade prostate tumors. In prostate cancer cells, high ZBTB46 output was responsible for the activation of LIF-STAT3 signaling and neuroendocrine-like features. The abundance of LIF was mediated by ADT-induced ZBTB46 through a physical interaction with the regulatory sequence of LIF. Analysis of serum from patients showed that cases of higher tumor grade and metastatic prostate cancer exhibited higher LIF titers. CONCLUSIONS: Our findings suggest that LIF is a potent serum biomarker for diagnosing advanced prostate cancer and that targeting the ZBTB46-LIF axis may therefore inhibit CRPC development and neuroendocrine differentiation after ADT.

E2f1 regulates the induction of promyelocytic leukemia zinc finger transcription in neuronal differentiation of pluripotent P19 embryonal carcinoma cells.

During brain development, the expression of promyelocytic leukemia zinc finger (Plzf) in neural stem cells is precisely controlled to maintain the balance between neural stem cell self-renewal and differentiation. However, the mechanism underlying transcriptional regulation of Plzf in neural stem cell is still unclear. Herein, using P19 embryonal carcinoma cells as a model, we observed that Plzf expression was induced in the P19-derived embryonic bodies, which enrich neural stem-like cell populations, as demonstrated by the expression of neural stem cell markers, Nestin and Sox2. We then characterized the Plzf promoter and identified two E2f1 binding sites (-755/-751 and -53/-49, the transcription start site was designated as +1) are important for the activation of Plzf promoter. Finally, we found that the induction of Plzf in the neural stem-like cells derived from pluripotent P19 cells is decrease by E2f1 knockdown. Taken together, we conclude that E2f1 is an important transcription factor that regulates Plzf transcription and may involve in maintaining the self-renewal ability of neural stem cells.

Gene-Embedded Nanostructural Biotic-Abiotic Optoelectrode Arrays Applied for Synchronous Brain Optogenetics and Neural Signal Recording.

Optogenetics is a recently established neuromodulation technique in which photostimulation is used to manipulate neurons with high temporal and spatial precision. However, sequential genetic and optical insertion with double brain implantation tends to cause excessive tissue damage. In addition, the incorporation of light-sensitive genes requires the utilization of viral vectors, which remains a safety concern. Here, by combining device fabrication design, nanotechnology, and cell targeting technology, we developed a new gene-embedded optoelectrode array for neural implantation to enable spatiotemporal electroporation (EP) for gene delivery/transfection, photomodulation, and synchronous electrical monitoring of neural signals in the brain via one-time implantation. A biotic-abiotic neural interface (called PG) composed of reduced graphene oxide and conductive polyelectrolyte 3,4-ethylenedioxythiophene-modified amphiphilic chitosan was developed to form a nanostructural hydrogel with assembled nanodomains for encapsulating nonviral gene vectors (called PEI-NT-pDNA) formulated by neurotensin (NT) and polyethylenimine (PEI)-coupled plasmid DNA (pDNA). The PG can maintain high charge storage ability to respond to a minimal current of 125 µA for controllable gene delivery. The in vitro analysis of PG-PEI-NT-pDNA on the microelectrode array chip showed that the microelectrodes provided electrically inductive electropermeabilization, which permitted gene transfection into localized rat adrenal pheochromocytoma cells with a strong green fluorescent protein expression that was up to 8-fold higher than that in nontreated cells. Furthermore, the in vivo implantation enabled on-demand spatiotemporal gene transfection to neurons with 10-fold enhancement of targeting ability compared with astrocytes. Finally, using the real optogenetic opsin channelrhodopsin-2, the flexible neural probe incorporated with an optical waveguide fiber displayed photoevoked extracellular spikes in the thalamic ventrobasal region after focal EP for only 7 days, which provided a proof of concept for the use of photomodulation to facilitate neural therapies.

Potential therapeutic effect of curcumin, a natural mTOR inhibitor, in tuberous sclerosis complex.

BACKGROUND: Curcumin is a polyphenol natural product of the plant Curcuma longa. Recent studies suggest that curcumin inhibit mTOR activity in vitro, which prompts us to investigate curcumin function as a new class of mTOR inhibitor suitable for tuberous sclerosis complex (TSC) treatment. PURPOSE: We aim to investigate the efficacy of curcumin in the treatment of TSC related manifestations in animal model. STUDY DESIGN: Solid lipid curcumin particle (SLCP), a novel curcumin formulation, was used to treat TSC related manifestations in Tsc2 knockout mice. METHODS: The novel object recognition test was used to analyze the recognition memory function. The long-term potentiation was studied using electrophysiological analysis. Western blotting was used to assess the protein expression and activation status. RESULTS: Recognition memory deficit began as early as 4 weeks of age in both male and female Tsc2+/- mice. Oral administration with SLCP activates AMPK activity and inhibits mTOR activity in the brain tissue of Tsc2+/- mice, and can rescue the electrophysiological abnormality and object recognition memory loss in the mice. CONCLUSIONS: Our results suggest that SLCP could be an effective treatment for TSC patients.

Znf179 E3 ligase-mediated TDP-43 polyubiquitination is involved in TDP-43- ubiquitinated inclusions (UBI) (+)-related neurodegenerative pathology.

BACKGROUND: The brain predominantly expressed RING finger protein, Znf179, is known to be important for embryonic neuronal differentiation during brain development. Downregulation of Znf179 has been observed in motor neurons of adult mouse models for amyotrophic lateral sclerosis (ALS), yet the molecular function of Znf179 in neurodegeneration has never been previously described. Znf179 contains the classical C3HC4 RING finger domain, and numerous proteins containing C3HC4 RING finger domain act as E3 ubiquitin ligases. Hence, we are interested to identify whether Znf179 possesses E3 ligase activity and its role in ALS neuropathy. METHODS: We used in vivo and in vitro ubiquitination assay to examine the E3 ligase autoubiquitination activity of Znf179 and its effect on 26S proteasome activity. To search for the candidate substrates of Znf179, we immunoprecipitated Znf179 and subjected to mass spectrometry (MS) analysis to identify its interacting proteins. We found that ALS/ FTLD-U (frontotemporal lobar degeneration (FTLD) with ubiquitin inclusions)-related neurodegenerative TDP-43 protein is the E3 ligase substrate of Znf179. To further clarify the role of E3 ubiquitin ligase Znf179 in neurodegenerative TDP-43-UBI (ubiquitinated inclusions) (+) proteinopathy, the effect of Znf179-mediated TDP-43 polyubiquitination on TDP-43 protein stability, aggregate formation and nucleus/cytoplasm mislocalization were evaluated in vitro cell culture system and in vivo animal model. RESULTS: Here we report that Znf179 is a RING E3 ubiquitin ligase which possesses autoubiquitination feature and regulates 26S proteasome activity through modulating the protein expression levels of 19S/20S proteasome subunits. Our immunoprecipitation assay and MS analysis results revealed that the neuropathological TDP-43 protein is one of its E3 ligase substrate. Znf179 interactes with TDP-43 protein and mediates polyubiquitination of TDP-43 in vitro and in vivo. In neurodegenerative TDP-43 proteinopathy, we found that Znf179-mediated polyubiquitination of TDP-43 accelerates its protein turnover rate and attenuates insoluble pathologic TDP-43 aggregates, while knockout of Znf179 in mouse brain results in accumulation of insoluble TDP-43 and cytosolic TDP-43 inclusions in cortex, hippocampus and midbrain regions. CONCLUSIONS: Here we unveil the important role for the novel E3 ligase Znf179 in TDP-43-mediated neuropathy, and provide a potential therapeutic strategy for combating ALS/ FTLD-U neurodegenerative pathologies.

Author Correction: Znf179 induces differentiation and growth arrest of human primary glioblastoma multiforme in a p53-dependent cell cycle pathway.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.

LGK974, a PORCUPINE inhibitor, mitigates cytotoxicity in an in vitro model of Parkinson's disease by interfering with the WNT/ß-CATENIN pathway.

Paraquat (PQ) as an herbicide has been demonstrated to impair dopaminergic (DAergic) neurons and highly correlate with the etiology of Parkinson's disease (PD). WNT/ß-CATENIN signaling is known for the specification and neurogenesis of midbrain DAergic neurons and implicated as a therapeutic target in treating many diseases, such as cancer and degenerative diseases. LGK974, a WNT pathway inhibitor, is currently under clinical trial for patients with malignancies. Since the exact role of WNT/ß-CATENIN signaling in mediating PD is undetermined, LGK974 was used to examine its effect on the PQ-induced cell model of PD. LGK974 attenuated PQ-induced apoptosis and released mitochondrial pro-poptotic molecules in human neuroblastoma SH-SY5Y cell. PQ increased the levels of ß-CATENIN, non-phosphorylated (Ser33/37/Thr41) ß-CATENIN, and phosphorylated glycogen synthase kinase (GSK)-3α/ß. PQ also increased the nuclear translocation of ß-CATENIN, which can be attenuated by LKG974. Furthermore, LGK974 attenuated the PQ-induced release of mitochondrial proapoptotic factors and WNT agonist 1-induced cell death. Taken together, we have shown for the first time that LGK974 mediated through the WNT/ß-CATENIN pathway to prevent PQ-induced cell death.

Highly sensitive/selective 3D nanostructured immunoparticle-based interface on a multichannel sensor array for detecting amyloid-beta in Alzheimer's disease.

Accumulation of ß-amyloid (Aß) peptides is highly associated with Alzheimer's disease (AD) progression in prevailing studies. The successful development of an ultrasensitive detection assay for Aß is a challenging task, especially from blood-based samples. Methods: We have developed a one-step electrophoresis/electropolymerization strategy for preparing a CSIP hierarchical immunoelectrochemical interface that is easily integrated into a PoCT device. The interface includes conductive silk fibroin-based immunoparticles (CSIPs) via electropolymerized Poly(3,4-ethylenedioxythiophene) (PEDOT) bridging to enable on-site electrochemical detection of serum amyloid-ß42 (Aß42) and -ß40 (Aß40) peptides from an AD blood test. In addition, micro-positron emission tomography (microPET) neuroimaging and behavioral tests were simultaneously performed. Results: This nanostructured conductive interface favors penetration of water-soluble biomolecules and catalyzes a redox reaction, providing limits of detection (LOD) of 6.63 pg/mL for Aß40 and 3.74 pg/mL for Aß42. Our proof-of-concept study confirms that the multi-sensing electrochemical immunosensor array (MEIA) platform enables simultaneous measurement of serum Aß42 and Aß40 peptide levels and is more informative in early stage AD animals than amyloid-labeling Aß plaque PET imaging and behavioral tests. Conclusion: We believe this study greatly expands the applications of silk fibroin-based materials, is an important contribution to the advancement of biomaterials, and would also be valuable in the design of new types of multichannel electrochemical immunosensor arrays for the detection of other diseases.

Upregulation of Znf179 acetylation by SAHA protects cells against oxidative stress.

The accumulation of reactive oxygen species (ROS) commonly occurs during normal aging and during some acute/chronic progressive disorders. In order to avoid oxidative damage, scavenging of these radicals is important. Previously, we identified zinc finger protein 179 (Znf179) as a neuroprotector that increases antioxidant enzymes against superoxide radicals. However, the molecular mechanisms involved in the activation and regulation of Znf179 remain unresolved. Here, by performing sequence alignment, bioinformatics analysis, immunoprecipitation using two specific acetyl-lysine antibodies, and treatment with the histone deacetylase (HDAC) inhibitor SAHA, we determined the lysine-specific acetylation of Znf179. Furthermore, we investigated Znf179 interaction with HDACs and revealed that peroxide insult induced a dissociation of Znf179-HDAC1/HDAC6, causing an increase in Znf179 acetylation. Importantly, HDAC inhibition by SAHA further prompted Znf179 hyperacetylation, which promoted Znf179 to form a transcriptional complex with Sp1 and increased antioxidant gene expression against oxidative attack. In summary, the results obtained in this study showed that Znf179 was regulated by HDACs and that Znf179 acetylation was a critical mechanism in the induction of antioxidant defense systems. Additionally, HDAC inhibitors may have therapeutic potential for induction of Znf179 acetylation, strengthening the Znf179 protective functions against neurodegenerative processes.

Nck2 is essential for limb trajectory selection by spinal motor axons.

BACKGROUND: The development of a functioning nervous system requires precise assembly of neuronal connections, which can be achieved by the guidance of axonal growth cones to their proper targets. How axons are guided by signals transmitted to the cytoskeleton through cell surface-expressed guidance receptors remains unclear. We investigated the function of Nck2 adaptor protein as an essential guidance intermediary in the context of spinal lateral motor column (LMC) motor axon trajectory into the limb. RESULTS: Nck2 mRNA and protein are preferentially expressed in the medial subgroups of chick LMC neurons during axon trajectory into the limb. Nck2 loss- and gain-of-function in LMC neurons using in ovo electroporation perturb LMC axon trajectory selection demonstrating an essential role of Nck2 in motor axon guidance. We also showed that Nck2 knockdown and overexpression perturb the growth preference of LMC neurites against ephrins in vitro and Eph-mediated redirection of LMC axons in vivo. Finally, the significant changes of LMC neurite growth preference against ephrins in the context of Nck2 and α2-chimaerin loss- and gain-of-function implicated Nck2 function to modulate α2-chimaerin activity. CONCLUSIONS: Here, we showed that Nck2 is required for Eph-mediated axon trajectory selection from spinal motor neurons through possible interaction with α2-chimaerin. Developmental Dynamics 247:1043-1056, 2018. © 2018 Wiley Periodicals, Inc.

Androgen deprivation therapy-induced epithelial-mesenchymal transition of prostate cancer through downregulating SPDEF and activating CCL2.

The chemokine CC motif ligand 2 (CCL2) is important in recruiting tumor-associated macrophages and is involved in the development of castration-resistance prostate cancer (CRPC) after androgen-deprivation therapy (ADT); however, the underlying mechanism remains unclear. We found that inactivation of the androgen receptor (AR) reduces a transcriptional repressor (SAM pointed domain-containing ETS transcription factor, SPDEF) of CCL2, which mediates epithelial-to-mesenchymal transition (EMT) of prostate tumor cells. Cell lines derived from a prostate-specific Pten/Trp53-null mouse and capable of a spontaneous EMT were utilized for identification of CCL2, and showed that reduced SPDEF expression was associated with an elevated CCL2-activated EMT. AR signaling inhibits CCL2 through a SPDEF-mediated mechanism in that the SPDEF recognizes the CCL2 promoter and transcriptionally represses its activity. Ectopically expressed SPDEF reduced the EMT and rescued expression of CCL2 in SPDEF-expressing cells, which induced the EMT and promotes malignant functions of prostate cancer cells. In tissues from prostate cancer patients with ADT, low SPDEF levels were correlated with high CCL2 expression compared to patients without ADT. We present a novel mechanism that contributes to the EMT and metastatic phenotype observed in a subset of ADT-resistant prostate cancer, where the CCL2 is stimulated through the inactivated of AR-mediated SPDEF.

Disruption of ETV6 leads to TWIST1-dependent progression and resistance to epidermal growth factor receptor tyrosine kinase inhibitors in prostate cancer.

BACKGROUND: ETS variant gene 6 (ETV6) is a putative tumor suppressor and repressed by epidermal growth factor receptor (EGFR) signaling in prostate cancer. Since EGFR antagonists seem ineffective in castration-resistant prostate cancer (CRPC), we aim to study the role of ETV6 in the development of drug resistance. METHODS: Etv6 target gene was validated by ChIP and promoter reporter assays. Correlation of ETV6 and TWIST1 was analyzed in human clinical datasets and tissue samples. Migration, invasion, and metastasis assays were used to measure the cellular responses after perturbation of ETV6 -TWIST1 axis. Proliferation and tumor growth in xenograft model were performed to evaluate the drug sensitivities of EGFR-tyrosine kinase inhibitors (TKIs). RESULTS: ETV6 inhibits TWIST1 expression and disruption of ETV6 promotes TWIST1-dependent malignant phenotypes. Importantly, ETV6 is required to the anti-proliferation effects of EGFR-TKIs, partly due to the inhibitory function of ETV6 on TWIST1. We also found that EGFR-RAS signaling is tightly controlled by ETV6, supporting its role in TKI sensitivity. CONCLUSIONS: Our study demonstrates that disruption of ETV6 contributes to EGFR-TKI resistance, which is likely due to derepression of TWIST1 and activation of EGFR-RAS signaling. Our results implicate ETV6 as a potential marker for predicting efficacy of an EGFR-targeted anticancer approach. Combination treatment of TWIST1 inhibitors could sensitize the anti-proliferation effects of EGFR-TKIs.