BHLHE22 Expression Is Associated with a Proinflammatory Immune Microenvironment and Confers a Favorable Prognosis in Endometrial Cancer.

Endometrial cancer (EC) rates are rising annually. Additional prediction markers need to be evaluated because only 10-20% of EC cases show an objective response to immune-checkpoint inhibitors (ICIs). Our previous methylomic study found that BHLHE22 is hypermethylated in EC tissues and can be detected using a Pap-smear sample. BHLHE22, a basic helix loop helix transcription factor family member, is known as a transcriptional repressor and is involved in cell differentiation. However, the role of BHLHE22 in EC remains poorly understood. Herein, we analyzed BHLHE22 expression in 54 paired cancer and normal endometrial tissue samples, and confirmed with databases (TCGA, GTEx, and human protein atlas). We found that BHLHE22 protein expression was significantly downregulated in EC compared with normal endometrium. High BHLHE22 expression was associated with microsatellite-instable subtype, endometrioid type, grade, and age. It showed a significant favorable survival. BHLHE22 overexpression inhibited the proliferation and migration of EC cells. Functional enrichment analysis showed that BHLHE22 was significantly associated with immune-related pathways. Furthermore, BHLHE22 was positively correlated with proinflammatory leukocyte infiltration and expression of chemokine genes in EC. In conclusion, BHLHE22 regulates immune-related pathways and modulates the immune microenvironment of EC.

Hepatocyte Nuclear Factor 4-α (HNF4α) controls the insulin resistance-induced pancreatic ß-cell mass expansion.

BACKGROUND: Regardless of the etiology, any type of DM presents a reduction of insulin-secreting cell mass, so it is important to investigate pathways that induce the increase of this cell mass. AIM: Based on the fact that (1) HNF4α is crucial for ß-cell proliferation, (2) DEX-induced IR promotes ß-cell mass expansion, and (3) the stimulation of ß-cell mass expansion may be an important target for DM therapies, we aimed to investigate whether DEX-induced proliferation of ß pancreatic cells is dependent on HNF4α. METHODS: We used WildType (WT) and Knockout (KO) mice for HNF4-α, treated or not with 100 mg/Kg/day of DEX, for 5 consecutive days. One day after the last injection of DEX the IR was confirmed by ipITT and the mice were euthanized for pancreas removal. RESULTS: In comparison to WT, KO mice presented increased glucose tolerance, lower fasting glucose and increased glucose-stimulates insulin secretion (GSIS). DEX induced IR in both KO and WT mice. In addition, DEX-induced ß-cell mass expansion and an increase in the Ki67 immunostaining were observed only in WT mice, evidencing that IR-induced ß-cell mass expansion is dependent on HNF4α. Also, we observed that DEX-treatment, in an HNF4α-dependent way, promoted an increase in PDX1, PAX4 and NGN3 gene expression. CONCLUSIONS: Our results strongly suggest that DEX-induced IR promotes ß-cell mass expansion through processes of proliferation and neogenesis that depend on the HNF4α activity, pointing to HNF4α as a possible therapeutic target in DM treatment.

The Potential Roles of Dec1 and Dec2 in Periodontal Inflammation.

Periodontal inflammation is a common inflammatory disease associated with chronic inflammation that can ultimately lead to alveolar attachment loss and bone destruction. Understanding autophagy and pyroptosis has suggested their significant roles in inflammation. In recent years, studies of differentiated embryo-chondrocyte expressed genes 1 and 2 (Dec1 and Dec2) have shown that they play important functions in autophagy and in pyroptosis, which contribute to the onset of periodontal inflammation. In this review, we summarize recent studies on the roles of clock genes, including Dec1 and Dec2, that are related to periodontal inflammation and other diseases.

Genome-wide Analysis of Basic Helix-Loop-Helix Family Genes and Expression Analysis in Response to Drought and Salt Stresses in Hibiscus hamabo Sieb. et Zucc.

The basic helix-loop-helix (bHLH) family of transcription factors is one of the most significant and biggest in plants. It is involved in the regulation of both growth and development, as well as stress response. Numerous members of the bHLH family have been found and characterized in woody plants in recent years. However, no systematic study of the bHLH gene family has been published for Hibiscus hamabo Sieb. et Zucc. In this research, we identified 162 bHLH proteins (HhbHLHs) from the genomic and transcriptomic datasets of H. hamabo, which were phylogenetically divided into 19 subfamilies. According to a gene structural study, the number of exon-introns in HhbHLHs varied between zero and seventeen. MEME research revealed that the majority of HhbHLH proteins contained three conserved motifs, 1, 4, and 5. The examination of promoter cis-elements revealed that the majority of HhbHLH genes had several cis-elements involved in plant growth and development and abiotic stress responses. In addition, the overexpression of HhbHLH2 increased salt and drought stress tolerance in Arabidopsis.

HIF-1α and HIF-2α redundantly promote retinal neovascularization in patients with ischemic retinal disease.

Therapies targeting VEGF have proven only modestly effective for the treatment of proliferative sickle cell retinopathy (PSR), the leading cause of blindness in patients with sickle cell disease. Here, we shift our attention upstream from the genes that promote retinal neovascularization (NV) to the transcription factors that regulate their expression. We demonstrated increased expression of HIF-1α and HIF-2α in the ischemic inner retina of PSR eyes. Although both HIFs participated in promoting VEGF expression by hypoxic retinal Müller cells, HIF-1 alone was sufficient to promote retinal NV in mice, suggesting that therapies targeting only HIF-2 would not be adequate to prevent PSR. Nonetheless, administration of a HIF-2-specific inhibitor currently in clinical trials (PT2385) inhibited NV in the oxygen-induced retinopathy (OIR) mouse model. To unravel these discordant observations, we examined the expression of HIFs in OIR mice and demonstrated rapid but transient accumulation of HIF-1α but delayed and sustained accumulation of HIF-2α; simultaneous expression of HIF-1α and HIF-2α was not observed. Staggered HIF expression was corroborated in hypoxic adult mouse retinal explants but not in human retinal organoids, suggesting that this phenomenon may be unique to mice. Using pharmacological inhibition or an in vivo nanoparticle-mediated RNAi approach, we demonstrated that inhibiting either HIF was effective for preventing NV in OIR mice. Collectively, these results explain why inhibition of either HIF-1α or HIF-2α is equally effective for preventing retinal NV in mice but suggest that therapies targeting both HIFs will be necessary to prevent NV in patients with PSR.

atoh8 expression pattern in early zebrafish embryonic development.

Atonal homologue 8 (atoh8) is a basic helix-loop-helix transcription factor expressed in a variety of embryonic tissues. While several studies have implicated atoh8 in various developmental pathways in other species, its role in zebrafish development remains uncertain. So far, no studies have dealt with an in-depth in situ analysis of the tissue distribution of atoh8 in embryonic zebrafish. We set out to pinpoint the exact location of atoh8 expression in a detailed spatio-temporal analysis in zebrafish during the first 24 h of development (hpf). To our surprise, we observed transcription from pre-segmentation stages in the paraxial mesoderm and during the segmentation stages in the somitic sclerotome and not-as previously reported-in the myotome. With progressing maturation of the somites, the restriction of atoh8 to the sclerotomal compartment became evident. Double in situ hybridisation with atoh8 and myoD revealed that both genes are expressed in the somites at coinciding developmental stages; however, their domains do not spatially overlap. A second domain of atoh8 expression emerged in the embryonic brain in the developing cerebellum and hindbrain. Here, we observed a specific expression pattern which was again in contrast to the previously published suggestion of atoh8 transcription in neural crest cells. Our findings point towards a possible role of atoh8 in sclerotome, cerebellum and hindbrain development. More importantly, the results of this expression analysis provide new insights into early sclerotome development in zebrafish-a field of research in developmental biology which has not received much attention so far.

Nuclear expression of VDR and AHR is mutually exclusive in glandular cells in endometriosis.

The vitamin D receptor (VDR) and aryl hydrocarbon receptor (AHR) are two nuclear receptors that exert their effects by binding with ligands and forming a molecular complex. These complexes translocate to the nucleus and activate the expression of a series of genes which have a response element to VDR or AHR. Both receptors have been identified in the pathogenesis of endometriosis, a common disease characterized by the formation of endometrium-like tissue in ectopic zones. Despite numerous therapies, there is no definitive cure for endometriosis at the pharmacological level. Our study aims to describe the location and the expression of VDR and AHR at the protein level. For this purpose, an evaluation was performed using tissue from the three normal phases of the endometrium (proliferative, early, and late secretory) and in endometriosis by immunohistochemistry, using anti-VDR and anti-AHR antibodies. We demonstrate that in the nuclei of glandular cells in endometriosis, the expression of VDR and AHR is mutually exclusive-when the expression of one receptor is high, the other one is low-suggesting a possible target in the treatment of endometriosis. We also identify a significant change in the expression of glandular cytoplasmic AHR between the proliferative and late secretory endometrium.

Wutou decoction attenuates rheumatoid arthritis by modulating the Ahr/LOC101928120/SHC1 pathway.

CONTEXT: Wutou decoction (WTD) is a Chinese herbal formula alleviating rheumatoid arthritis (RA). SHC adaptor protein 1 (SHC1) regulates apoptosis, inflammation, and the production of reactive oxygen species (ROS). The LOC101928120 gene is located near the SHC1 gene. Bioinformatics analysis showed that the long non-coding RNA LOC101928120 binds to histone deacetylase HDAC1 that might regulate SHC1 expression. The LOC101928120 gene might be targeted by the transcriptional factor Aryl hydrocarbon receptor (Ahr). OBJECTIVE: This study determines the involvement of the Ahr/LOC101928120/SHC1 pathway in WTD alleviation of RA. MATERIALS AND METHODS: Wistar rats were injected with complete Freund's adjuvant in the hind footpad to construe the RA model. WTD (9.8 g/kg/day) was administered intragastrically for 15 days. The CHON-001 chondrocyte cells were treated with IL-1ß (10 ng/mL) alone or in combination with WTD (1 µg/mL). A RNA pull-down assay was performed to determine the interaction between LOC101928120 and HDAC1. Ahr targeting the LOC101928120 gene was detected using luciferase reporter and chromatin immunoprecipitation assays. RESULTS: WTD alleviated the swelling of the hind paw in rats with RA and suppressed the chondrocyte apoptosis and ROS production caused by IL-1ß. WTD decreased SHC1 but increased LOC101928120 in IL-1ß-treated chondrocytes. SHC1 knockdown and LOC101928120 overexpression also showed the protection. However, LOC101928120 knockdown attenuated the protective effects of WTD. WTD stimulated Ahr, which promoted LOC101928120 transcription. LOC101928120 recruited HDAC1 to the promoter region of the SHC1 gene, thereby decreasing SHC1. DISCUSSION AND CONCLUSION: This study revealed a new mechanism by which WTD alleviates RA by modulating the Ahr/LOC101928120/SHC1 pathway.

CircMEMO1 modulates the promoter methylation and expression of TCF21 to regulate hepatocellular carcinoma progression and sorafenib treatment sensitivity.

BACKGROUND: Cirrhosis is a recognized risk factor for developing hepatocellular carcinoma (HCC). Few studies have reported the expression profile of circRNAs in HCC samples compared to paratumour dysplastic nodule (DN) samples. METHODS: The Arraystar Human circRNA Array combined with laser capture microdissection (LCM) was used to analyse the expression profile of circRNAs in HCC samples compared to paratumour DN samples. Then, both in vitro and in vivo HCC models were used to determine the role and mechanism of key circRNA in HCC progression and treatment sensitivity. RESULTS: We found that circMEMO1 was significantly downregulated in HCC samples and that the level of circMEMO1 was closely related to the OS and disease-free survival (DFS) of HCC patients. Mechanistic analysis revealed that circMEMO1 can modulate the promoter methylation and gene expression of TCF21 to regulate HCC progression by acting as a sponge for miR-106b-5p, which targets the TET family of genes and increases the 5hmC level. More importantly, circMEMO1 can increase the sensitivity of HCC cells to sorafenib treatment. CONCLUSION: Our study determined that circMEMO1 can promote the demethylation and expression of TCF21 and can be considered a crucial epigenetic modifier in HCC progression.

HIF-2α activation potentiates oxidative cell death in colorectal cancers by increasing cellular iron.

Hypoxia is a hallmark of solid tumors that promotes cell growth, survival, and metastasis and confers resistance to chemo and radiotherapies. Hypoxic responses are largely mediated by the transcription factors hypoxia-inducible factor 1α (HIF-1α) and HIF-2α. Our work demonstrates that HIF-2α is essential for colorectal cancer (CRC) progression. However, targeting hypoxic cells is difficult, and tumors rapidly acquire resistance to inhibitors of HIF-2α. To overcome this limitation, we performed a small molecule screen to identify HIF-2α-dependent vulnerabilities. Several known ferroptosis activators and dimethyl fumarate (DMF), a cell-permeable mitochondrial metabolite derivative, led to selective synthetic lethality in HIF-2α-expressing tumor enteroids. Our work demonstrated that HIF-2α integrated 2 independent forms of cell death via regulation of cellular iron and oxidation. First, activation of HIF-2α upregulated lipid and iron regulatory genes in CRC cells and colon tumors in mice and led to a ferroptosis-susceptible cell state. Second, via an iron-dependent, lipid peroxidation-independent pathway, HIF-2α activation potentiated ROS via irreversible cysteine oxidation and enhanced cell death. Inhibition or knockdown of HIF-2α decreased ROS and resistance to oxidative cell death in vitro and in vivo. Our results demonstrated a mechanistic vulnerability in cancer cells that were dependent on HIF-2α that can be leveraged for CRC treatment.

Genetic and epigenetic changes in the eutopic endometrium of women with endometriosis: association with decreased endometrial αvß3 integrin expression.

About 40% of women with infertility and 70% of women with pelvic pain suffer from endometriosis. The pregnancy rate in women undergoing IVF with low endometrial integrin αvß3 (LEI) expression is significantly lower compared to the women with high endometrial integrin αvß3 (HEI). Mid-secretory eutopic endometrial biopsies were obtained from healthy controls (C; n=3), and women with HEI (n=4) and LEI (n=4) and endometriosis. Changes in gene expression were assessed using human gene arrays and DNA methylation data were derived using 385 K Two-Array Promoter Arrays. Transcriptional analysis revealed that LEI and C groups clustered separately with 396 differentially expressed genes (DEGs) (P<0.01: 275 up and 121 down) demonstrating that transcriptional and epigenetic changes are distinct in the LEI eutopic endometrium compared to the C and HEI group. In contrast, HEI vs C and HEI vs LEI comparisons only identified 83 and 45 DEGs, respectively. The methylation promoter array identified 1304 differentially methylated regions in the LEI vs C comparison. The overlap of gene and methylation array data identified 14 epigenetically dysregulated genes and quantitative RT-PCR analysis validated the transcriptomic findings. The analysis also revealed that aryl hydrocarbon receptor (AHR) was hypomethylated and significantly overexpressed in LEI samples compared to C. Further analysis validated that AHR transcript and protein expression are significantly (P<0.05) increased in LEI women compared to C. The increase in AHR, together with the altered methylation status of the 14 additional genes, may provide a diagnostic tool to identify the subset of women who have endometriosis-associated infertility.

Circular RNA circ_0003420 mediates inflammation in sepsis-induced liver damage by downregulating neuronal PAS domain protein 4.

OBJECTIVE: Our aim was to investigate whether circular RNA (circRNA) circ_0003420 mediates inflammation in sepsis-induced liver damage and to determine the mechanism involved. MATERIALS AND METHODS: Liver tissue samples from patients with sepsis and healthy subjects were used to identify differentially expressed circRNAs. Additionally, Kupffer cells were treated with lipopolysaccharide (LPS) to establish an in vitro model of sepsis-induced liver damage. Cell viability and proliferation were measured with a cell counting kit-8 and 5-ethynyl-2'-deoxyuridine (EdU) labeling, respectively. Relative mRNA and protein levels of IL-6, IL-1ß, tumor necrosis factor (TNF)-α, and neuronal PAS domain protein 4 (NPAS4) were determined via reverse-transcription quantitative PCR and western blotting, respectively. RESULTS: We observed circ_0003420 upregulation accompanied by NPAS4 downregulation in liver samples from patients with sepsis-associated damage and in Kupffer cells treated with LPS. Results of in vitro experiments indicated that LPS treatment reduced cell viability and induced well-pronounced apoptosis and inflammatory signs. Circ_0003420 silencing counteracted LPS's influence on cell proliferation, apoptosis, and inflammation signs. Bioinformatics and a dual-luciferase reporter assay revealed that circ_0003420 targets NPAS4 mRNA and negatively correlates with NPAS4 expression. Moreover, NPAS4 knockdown recovered the apoptosis rate and expression levels of inflammatory cytokines in the LPS-treated circ_0003420 knockdown cells, whereas NPAS4 overexpression had similar effects on Kupffer cell properties as circ_0003420 silencing. CONCLUSION: We demonstrate that circ_0003420 targets NPAS4 mRNA thereby mediating the cell damage and inflammation caused by LPS. This study provides a possible target for treatment of liver damage induced by sepsis.

NUPR1- CHOP experssion, autophagosome formation and apoptosis in the postmortem striatum of chronic methamphetamine user.

Methamphetamine (Meth) is a neuro-stimulator substrate which might lead to neural cell death and the activation of several interconnected cellular pathways as well. However, the precise molecular mechanisms underlying Meth-induced neural cell death remained unclear yet. The current study aimed to assess the specific relationship between long-term Meth exposure and several endoplasmic reticulum stress, autophagy, and apoptosis associated markers including C/EBP homologous protein (CHOP), Tribbles homolog 3(Trib3), Nuclear protein 1(NUPR1), and Beclin-1 expression in postmortem human striatum. Therefore, the effects of long-term Meth exposure on autophagy and apoptosis in the striatum of postmortem users were evaluated and molecular, immunehistochemical, and histological examinations were performed on 10 control and 10 Meth-addicted brains. The level of CHOP, Trib3, NUPR1, and Beclin-1, Microtubule-associated proteins 1A/1B light chain 3B(LC3), Caspase 3, and Autophagy protein 5 (ATG5) were measured by using qPCR and immunohistochemistry. Stereological neural cell counting, Hematoxylin and Eosin, Nissl and Tunel staining were also performed. Based on our findings, the expression level of CHOP, Trib3, NUPR1, and Beclin-1 in the striatum of Meth group were significantly higher than the control group. Besides, the neuronal cell death was substantially increased in the striatum based on data obtained from the Tunel assay and the stereological analysis. Long-term presence of Meth in the brain can induce ER stress and overexpression of NUPR1 which is associated with the upregulation of CHOP, a pro-apoptotic transcription factor. Moreover, an increase in Trib3 expression is implicated in CHOP-dependent autophagic cell death during Meth-induced ER stress accompanied by an increase in neuronal cell death in the striatum of the postmortem human brains. Beclin 1 expression was also upregulated which may due to the activation of autophagic mechanisms upon prolonged Meth exposure.

Photo-dependent membrane-less organelles formed from plant phyB and PIF6 proteins in mammalian cells.

Plant photobodies are the membrane-less organelles (MLOs) that can be generated by protein-protein interactions between active form of phytochrome B (phyB) and phytochrome-interacting factors (PIFs). These organelles regulate plant photomorphogenesis. In this study, we developed two chimeric proteins with fluorescent proteins, phyB fused to EGFP and PIF6 fused to mCherry, and investigated their exogenous expression in mammalian cells by confocal fluorescence microscopy. Results showed that irradiation with diffused 630-nm light induced formation and subsequent increase in sizes of the MLOs. The assembly and disassembly of the photo-inducible MLOs in the mammalian cell cytoplasm obeyed the laws inherent in the concentration-dependent phase separation of biopolymers. The sizes of MLOs formed from phyB and PIF6 in mammalian cells corresponded to the sizes of the so-called "early" photobodies in plant cells. These results suggested that the first step for the formation of plant photobodies might be based on the light-dependent liquid-liquid phase separation of PIFs and other proteins that can specifically interact with the active form of phyB. The developed chimeric proteins in principle can be used to control the assembly and disassembly of photo-inducible MLOs, and thereby to regulate various intracellular processes in mammalian cells.

Hypoxia and its possible relationship with endometrial receptivity in adenomyosis: a preliminary study.

BACKGROUND: Adenomyosis (AM) is an important cause of female infertility. However, the underlying mechanism remains unclear. This report describes a preliminary study of hypoxia and its possible association with endometrial receptivity in AM. METHODS: The study was divided into in vitro and in vivo experiments. In vitro, expression levels of the endometrial receptivity markers HOXA10 and HOXA11 in the implantation period were examined using real-time PCR and western blotting. Endometrial expression of hypoxia-inducible factor (HIF)-1α, HIF-2α, and HIF-3α was determined using immunohistochemistry. In vivo, using an AM mouse model established by oral administration of tamoxifen, we inhibited expression of HIF-2α using an HIF-2α antagonist (PT2399; 30 mg/kg body weight, twice daily by oral gavage for 2 days) and then examined expression levels of Hoxa10 and Hoxa11 using real-time PCR and western blotting. RESULTS: Endometrial mRNA and protein expression levels of HOXA10 and HOXA11 were significantly lower in patients with AM than in control patients. Expression of HIF-2α was significantly higher in the AM group than in the control group, whereas that of HIF-1α and HIF-3α was equivalent in both groups. In vivo analysis showed that administration of the HIF-2α antagonist resulted in increased expression of Hoxa10 and Hoxa11 at both the mRNA and protein levels in AM model mice. CONCLUSIONS: HIF-2α overexpression may be one reason for decreased endometrial receptivity in AM. The current findings provide insight into HIF-2α-mediated AM-related infertility and suggest that PT2399 has potential as a treatment for AM. TRIAL REGISTRATION: This trial was retrospectively registered.

Identifying age-specific gene signatures of the human cerebral cortex with joint analysis of transcriptomes and functional connectomes.

The human cerebral cortex undergoes profound structural and functional dynamic variations across the lifespan, whereas the underlying molecular mechanisms remain unclear. Here, with a novel method transcriptome-connectome correlation analysis (TCA), which integrates the brain functional magnetic resonance images and region-specific transcriptomes, we identify age-specific cortex (ASC) gene signatures for adolescence, early adulthood and late adulthood. The ASC gene signatures are significantly correlated with the cortical thickness (P-value <2.00e-3) and myelination (P-value <1.00e-3), two key brain structural features that vary in accordance with brain development. In addition to the molecular underpinning of age-related brain functions, the ASC gene signatures allow delineation of the molecular mechanisms of neuropsychiatric disorders, such as the regulation between ARNT2 and its target gene ETF1 involved in Schizophrenia. We further validate the ASC gene signatures with published gene sets associated with the adult cortex, and confirm the robustness of TCA on other brain image datasets. Availability: All scripts are written in R. Scripts for the TCA method and related statistics result can be freely accessed at https://github.com/Soulnature/TCA. Additional data related to this paper may be requested from the authors.

Effects of Neurod1 Expression on Mouse and Human Schwannoma Cells.

OBJECTIVES: The objective was to explore the effect of the proneuronal transcription factor neurogenic differentiation 1 (Neurod1, ND1) on Schwann cells (SC) and schwannoma cell proliferation. METHODS: Using a variety of transgenic mouse lines, we investigated how expression of Neurod1 effects medulloblastoma (MB) growth, schwannoma tumor progression, vestibular function, and SC cell proliferation. Primary human vestibular schwannoma (VS) cell cultures were transduced with adenoviral vectors expressing Neurod1. Cell proliferation was assessed by 5-ethynyl-2'-deoxyuridine (EdU) uptake. STUDY DESIGN: Basic science investigation. RESULTS: Expression of Neurod1 reduced the growth of slow-growing but not fast-growing MB models. Gene transfer of Neurod1 in human schwannoma cultures significantly reduced cell proliferation in dose-dependent way. Deletion of the neurofibromatosis type 2 (Nf2) tumor-suppressor gene via Cre expression in SCs led to increased intraganglionic SC proliferation and mildly reduced vestibular sensory-evoked potentials (VsEP) responses compared to age-matched wild-type littermates. The effect of Neurod1-induced expression on intraganglionic SC proliferation in animals lacking Nf2 was mild and highly variable. Sciatic nerve axotomy significantly increased SC proliferation in wild-type and Nf2-null animals, and expression of Neurod1 reduced the proliferative capacity of both wild-type and Nf2-null SCs following nerve injury. CONCLUSION: Expression of Neurod1 reduces slow-growing MB progression and reduces human SC proliferation in primary VS cultures. In a genetic mouse model of schwannomas, we find some effects of Neurod1 expression; however, the high variability indicates that more tightly regulated Neurod1 expression levels that mimic our in vitro data are needed to fully validate Neurod1 effects on schwannoma progression. LEVEL OF EVIDENCE: NA Laryngoscope, 131:E259-E270, 2021.

Long-term self-renewing stem cells in the adult mouse hippocampus identified by intravital imaging.

Neural stem cells (NSCs) generate neurons throughout life in the mammalian hippocampus. However, the potential for long-term self-renewal of individual NSCs within the adult brain remains unclear. We used two-photon microscopy and followed NSCs that were genetically labeled through conditional recombination driven by the regulatory elements of the stem cell-expressed genes GLI family zinc finger 1 (Gli1) or achaete-scute homolog 1 (Ascl1). Through intravital imaging of NSCs and their progeny, we identify a population of Gli1-targeted NSCs showing long-term self-renewal in the adult hippocampus. In contrast, once activated, Ascl1-targeted NSCs undergo limited proliferative activity before they become exhausted. Using single-cell RNA sequencing, we show that Gli1- and Ascl1-targeted cells have highly similar yet distinct transcriptional profiles, supporting the existence of heterogeneous NSC populations with diverse behavioral properties. Thus, we here identify long-term self-renewing NSCs that contribute to the generation of new neurons in the adult hippocampus.

R-sulforaphane modulates the expression profile of AhR, ERα, Nrf2, NQO1, and GSTP in human breast cell lines.

Our previous study showed remarkable differences in the effect of R-sulforaphane (R-SFN) on the expression of CYPs 19, 1A1, 1A2, and 1B1 in ER(+) MCF7, ER( -) MDA-MB-231, and non-tumorigenic immortalized MCF10A (8). This study aimed to evaluate the effect of R-SFN on phase II enzymes induction and expression of AhR, Nrf2, and ERα in the same breast cell lines. The results showed increased expression of GSTP as a result of treatment with R-SFN in breast cancer cells. An increased NQO1 transcript and protein levels were found in all breast cells, with the most significant increase in MCF7 cells. Similarly, the enhancement of Nrf2 expression was noticed in all tested cells. AhR gene transcript and protein were decreased in MCF7 cells. In MDA-MB-231, increased AhR mRNA was not confirmed at the protein level. No differences were found in the expression of ERα. Overall, the results of the present study extended our earlier suggestions on the possible interference of R-SFN with estrogens homeostasis in breast cancer cells differing in ERα status, as well as in non-tumorigenic immortalized breast epithelial cells. While some of R-SFN effects might be beneficial and useful in breast cancer prevention, the others, particularly GSTP induction, may lead to adverse effects.

Decreased prolyl hydroxylase 3 mRNA expression in oncocytomas compared with clear cell renal cell carcinoma.

INTRODUCTION: Hypoxia inducible factors (HIF) and prolyl hydroxylase domain (PHD) enzymes play a central role in tumor progression in clear cell renal cell carcinoma (ccRCC). However, there are currently no data regarding the behavior of this pathway (HIF/PHD) in a large number of benign renal tumors, the oncocytomas. The aim of the present study was to compare the expression levels of these factors between ccRCC and oncocytoma tumors. MATERIAL AND METHODS: A total of 56 fresh frozen specimens from patients with ccRCC and 14 oncocytoma specimens were analyzed via reverse transcription-quantitative polymerase chain reaction in order to assess the expression levels of HIF-1α, HIF-2α, PHD1, PHD2, and PHD3. The analysis involved both fresh frozen tumor samples as well as adjacent normal kidney tissues. RESULTS: In ccRCC, HIF-1α and HIF-2α levels were upregulated in 65.5% and 71.4% of cases, respectively. PHD3 was downregulated only in 15.4% of the ccRCC cases, in contrast with oncocytoma cases, which exhibited low expression levels in the majority. The upregulation of PHD3 messenger RNA (mRNA) levels in ccRCC when compared with oncocytoma was statistically significant (P<0.001). No other comparisons (HIF-1α, HIF-2α, PHD1, and PHD2) were significantly different. HIF-2α and PHD3 mRNA expression levels were negatively correlated with Fuhrman Grade (P=0.029 and P=0.026, respectively) in ccRCC. CONCLUSION: To the best of our knowledge, this is the first time that the HIF/PHD pathway was compared between ccRCC and a common benign tumor, identifying the upregulation of PHD3 as the possible underlying factor guiding the difference in the behavior of ccRCC.