Mechanisms of peripheral neurotoxicity associated with four chemotherapy drugs using human induced pluripotent stem cell-derived peripheral neurons.

The awareness of the long-term toxicities of cancer survivors after chemotherapy treatment has been gradually strengthened as the population of cancer survivors grows. Generally, chemotherapy-induced peripheral neurotoxicity (CIPN) is studied by animal models which are not only expensive and time-consuming, but also species-specific differences. The generation of human induced pluripotent stem cells (hiPSCs) and differentiation of peripheral neurons have provided an in vitro model to elucidate the risk of CIPN. Here, we developed a drug-induced peripheral neurotoxicity model using hiPSC-derived peripheral neurons (hiPSC-PNs) to study the mechanisms of different chemotherapeutic agents on neuronal viability using LDH assay, a cell apoptosis assay determined by caspase 3/7 activation, neurite outgrowth, ion channel expression and neurotransmitter release following treatment of cisplatin, bortezomib, ixabepilone, or pomalidomide. Our data showed that the multiple endpoints of the hiPSC-PNs model had different sensitivity to various chemotherapeutic agents. Furthermore, the chemotherapeutics separated cell viability from the decrease in neurite lengthand changed levels of ion channels and neurotransmitters to a certain extent. Thus, we study the mechanisms of peripheral neurotoxicity induced by chemotherapeutic agents through changes in these indicators.

MicroRNA-29a activates a multi-component growth and invasion program in glioblastoma.

BACKGROUND: Glioblastoma is a malignant brain tumor characterized by rapid growth, diffuse invasion and therapeutic resistance. We recently used microRNA expression profiles to subclassify glioblastoma into five genetically and clinically distinct subclasses, and showed that microRNAs both define and contribute to the phenotypes of these subclasses. Here we show that miR-29a activates a multi-faceted growth and invasion program that promotes glioblastoma aggressiveness. METHODS: microRNA expression profiles from 197 glioblastomas were analyzed to identify the candidate miRNAs that are correlated to glioblastoma aggressiveness. The candidate miRNA, miR-29a, was further studied in vitro and in vivo. RESULTS: Members of the miR-29 subfamily display increased expression in the two glioblastoma subclasses with the worst prognoses (astrocytic and neural). We observed that miR-29a is among the microRNAs that are most positively-correlated with PTEN copy number in glioblastoma, and that miR-29a promotes glioblastoma growth and invasion in part by targeting PTEN. In PTEN-deficient glioblastoma cells, however, miR-29a nevertheless activates AKT by downregulating the metastasis suppressor, EphB3. In addition, miR-29a robustly promotes invasion in PTEN-deficient glioblastoma cells by repressing translation of the Sox4 transcription factor, and this upregulates the invasion-promoting protein, HIC5. Indeed, we identified Sox4 as the most anti-correlated predicted target of miR-29a in glioblastoma. Importantly, inhibition of endogenous miR-29a decreases glioblastoma growth and invasion in vitro and in vivo, and increased miR-29a expression in glioblastoma specimens correlates with decreased patient survival. CONCLUSIONS: Taken together, these data identify miR-29a as a master regulator of glioblastoma growth and invasion.

Protective efficacy of intravenous transplantation of adipose-derived stem cells for the prevention of radiation-induced salivary gland damage.

OBJECTIVE: High-dose radiation therapy in the head and neck area can lead to irreversible damage to salivary glands (SGs) with consequent xerostomia. Adipose-derived stem cells (ADSCs) have been shown to repair or rescue damaged SGs. Thus, we investigated the protective efficacy of ADSCs in the prevention of SG damage induced by high dose radiation. METHODS: Third-passage ADSCs (1×10(6)) were transplanted by intravenous infusion into the tail-vein of 8-week-old C57BL/6 mice, immediately after local irritation at a dose of 18Gy. The process was repeated twice a week during a period of six consecutive weeks. Eight weeks after radiation, functional evaluations were conducted by measuring salivary flow rate (SFR). Histological, immunohistochemical and transmission electron microscopic (TEM) examinations were performed to analyze microstructural and ultrastructural changes, microvessel density, amylase production, apoptosis, and proliferation activity. RESULTS: Intravenously administrated ADSCs could home to irradiated SGs within 24h after infusion, significantly increasing SG weights, improving SFR, and preserving the microscopic morphologies of SGs eight weeks post-radiation. More functional acini, higher amylase production levels, and higher microvessel densities were observed in ADSC-treated SGs than in irradiated SGs. Additionally, enhanced cell proliferation activity and reduced radiation-induced SG apoptosis was observed in the ADSC-treated group when compared with the irradiated group. CONCLUSION: Systemic administration of ADSCs immediately after radiation at a dose of 18Gy can protect both the morphology and function of SGs eight weeks after radiation in mice, and can be used as a protective measure for the prevention of SG damage induced by high-dose radiation.

A major role for microRNAs in glioblastoma cancer stem-like cells.

Studies have demonstrated that miRNAs contribute to the maintenance and phenotype of in several cancer types. This review will focus on the roles of a few well studied miRNAs in cancer stem-like cells of glioblastoma.

Numb regulates glioma stem cell fate and growth by altering epidermal growth factor receptor and Skp1-Cullin-F-box ubiquitin ligase activity.

Glioblastoma contains a hierarchy of stem-like cancer cells, but how this hierarchy is established is unclear. Here, we show that asymmetric Numb localization specifies glioblastoma stem-like cell (GSC) fate in a manner that does not require Notch inhibition. Numb is asymmetrically localized to CD133-hi GSCs. The predominant Numb isoform, Numb4, decreases Notch and promotes a CD133-hi, radial glial-like phenotype. However, upregulation of a novel Numb isoform, Numb4 delta 7 (Numb4d7), increases Notch and AKT activation while nevertheless maintaining CD133-hi fate specification. Numb knockdown increases Notch and promotes growth while favoring a CD133-lo, glial progenitor-like phenotype. We report the novel finding that Numb4 (but not Numb4d7) promotes SCF(Fbw7) ubiquitin ligase assembly and activation to increase Notch degradation. However, both Numb isoforms decrease epidermal growth factor receptor (EGFR) expression, thereby regulating GSC fate. Small molecule inhibition of EGFR activity phenocopies the effect of Numb on CD133 and Pax6. Clinically, homozygous NUMB deletions and low Numb mRNA expression occur primarily in a subgroup of proneural glioblastomas. Higher Numb expression is found in classical and mesenchymal glioblastomas and correlates with decreased survival. Thus, decreased Numb promotes glioblastoma growth, but the remaining Numb establishes a phenotypically diverse stem-like cell hierarchy that increases tumor aggressiveness and therapeutic resistance.

The TBP-PP2A mitotic complex bookmarks genes by preventing condensin action.

To maintain phenotypes of cell lineages, cells must 'remember' which genes were active before mitosis entry and transmit this information to their daughter cells so that expression patterns can be faithfully re-established in G1. This phenomenon is called gene bookmarking. However, during mitosis transcription ceases, most sequence-specific proteins dissociate from DNA and the chromatin is tightly compacted, making it difficult to understand how gene activity 'memory' is maintained through this stage of the cell cycle. A feature of gene bookmarking is that in mitotic cells, the promoters of formerly active genes lack compaction, but how compaction of these regions is inhibited is unknown. Here we show that during mitosis, TATA-binding protein (TBP), which remains bound to DNA during mitosis, recruits PP2A. TBP also interacts with condensin to allow efficient dephosphorylation and inactivation of condensin near these promoters to inhibit their compaction. Further, ChIP-on-chip data show that TBP is bound to many chromosomal sites during mitosis, and is higher in transcribed regions but low in regions containing pseudogenes and genes whose expression is tissue-restricted. These results suggest that TBP is involved not only in gene transcription during interphase but also in preserving the memory of gene activity through mitosis to daughter cells.

HSF1-TPR interaction facilitates export of stress-induced HSP70 mRNA.

Stress conditions inhibit mRNA export, but mRNAs encoding heat shock proteins continue to be efficiently exported from the nucleus during stress. How HSP mRNAs bypass this stress-associated export inhibition was not known. Here, we show that HSF1, the transcription factor that binds HSP promoters after stress to induce their transcription, interacts with the nuclear pore-associating TPR protein in a stress-responsive manner. TPR is brought into proximity of the HSP70 promoter after stress and preferentially associates with mRNAs transcribed from this promoter. Disruption of the HSF1-TPR interaction inhibits the export of mRNAs expressed from the HSP70 promoter, both endogenous HSP70 mRNA and a luciferase reporter mRNA. These results suggest that HSP mRNA export escapes stress inhibition via HSF1-mediated recruitment of the nuclear pore-associating protein TPR to HSP genes, thereby functionally connecting the first and last nuclear steps of the gene expression pathway, transcription and mRNA export.

Increased pathogen resistance and yield in transgenic plants expressing combinations of the modified antimicrobial peptides based on indolicidin and magainin.

Reverse peptide of indolicidin (Rev4), a 13-residue peptide based on the sequence of indolicidin, has been shown to possess both strong antimicrobial and protease inhibitory activities in vitro. To evaluate its efficacy in vivo, we produced and evaluated transgenic tobacco (Nicotiana tabacum L.) and Arabidopsis thaliana [(L.) Heynh.] plants expressing Rev4 with different signal peptide sequences for pathogen resistance. All transgenic plants showed normal growth and development, an indication of no or low cytotoxicity of the peptide. Furthermore, the transgenic plants exhibited elevated resistance to three bacterial and two oomycete pathogens. Interestingly, tobacco plants expressing Rev4 displayed enhanced yield compared to the control as indicated by an increased biomass production by as much as 34% in two field trials. When Rev4 was coexpressed with another antimicrobial peptide, Myp30, the disease resistance levels in the transgenic Arabidopsis were enhanced. These findings suggest the potential of using these peptides to protect plants from microbial pathogens and to enhance yield.

Mechanism of hsp70i gene bookmarking.

In contrast to most genomic DNA in mitotic cells, the promoter regions of some genes, such as the stress-inducible hsp70i gene that codes for a heat shock protein, remain uncompacted, a phenomenon called bookmarking. Here we show that hsp70i bookmarking is mediated by a transcription factor called HSF2, which binds this promoter in mitotic cells, recruits protein phosphatase 2A, and interacts with the CAP-G subunit of the condensin enzyme to promote efficient dephosphorylation and inactivation of condensin complexes in the vicinity, thereby preventing compaction at this site. Blocking HSF2-mediated bookmarking by HSF2 RNA interference decreases hsp70i induction and survival of stressed cells in the G1 phase, which demonstrates the biological importance of gene bookmarking.