Search

A gene encoding an archaeal transcription regulator (TON_1525) homologous to the Tfx family DNA-binding protein has been identified to affect gene expression of carbon monoxide metabolism in the hyperthermophilic archaeon Thermococcus onnurineus NA1. To broadly understand gene regulation by the TON_1525 gene, a gene deletion mutant (Δ1525) was constructed and transcriptome changes were investigated. A total of 351 genes were found to be differentially expressed in the Δ1525 mutant. The expression levels of the hydrogenase genes were significantly changed; mbh (membrane-bound hydrogenase) and mch (membrane-bound carbon monoxide-dependent hydrogenase) genes were increased, and mfh 2 (membrane-bound formate-dependent hydrogenase) and soluble hydrogenase genes were decreased. The Δ1525 mutant was cultured in a medium supplemented with maltodextrin or carbon monoxide to investigate whether changes in gene expression lead to physiological changes. The maximum H 2 production rates of the Δ1525 mutant were significantly enhanced on both substrates compared to the wild-type strain. These results indicate that the TON_1525 gene can affect the expression of various genes including mbh and mch , consequently regulating the metabolism of T. onnurineus NA1. This study expands our understanding of the functional role of the Tfx family DNA-binding proteins. • Functional studies on the role of the Tfx family DNA-binding protein are limited. • Deletion of the TON_1525 gene caused expression changes in various genes. • The Δ1525 mutant showed enhanced H 2 production by upregulation of H 2 evolving hydrogenase genes. [ABSTRACT FROM AUTHOR]

Simple Summary: It is generally reported that the prognostic factors for ampulla of Vater cancer include T stage, N stage, and lymphovascular invasion and subtype, and the meaning of MUC stain is controversial. This retrospective study by the authors evaluated the significance of various MUC stains in AoV cancer using single-center clinicopathological data. The results showed that MUC5AC was significant for lymph node metastasis and was furthermore valuable as a prognostic factor for predicting overall survival. If this is evaluated in addition to the hematoxylin and eosin stain, which is commonly performed in preoperative biopsy, it is expected to be a method to select groups that require more extensive LN dissection and further prevent locoregional recurrence. Introduction: Mucins play a pivotal role in epithelial carcinogenesis; however, their role remains elusive in ampulla of Vater (AoV) cancer, regardless of histological subtype. Therefore, we investigated the clinical significance of MUC1, MUC2, MUC5AC, and MUC6 expression in AoV cancer. Methods: Using samples from 68 patients with AoV cancer, we performed immunohistochemical staining for MUC1, MUC2, MUC5AC, and MUC6 using a tissue microarray. Subsequently, we analyzed their expression patterns in relation to clinicopathological parameters and patient outcomes. Results: Of the patients, 98.5% exhibited positive expression for MUC1, while MUC2, MUC5AC, and MUC6 were expressed in 44.1%, 47.1%, and 41.2% of the patients, respectively. Correlation analyses between mucin expression and clinicopathological factors revealed no significant associations, except between MUC5AC expression and N stage. Univariate analysis demonstrated significant associations between MUC5AC expression and overall survival (OS). Multivariate analysis further confirmed that MUC5AC expression was a significant predictor of OS, along with the N stage. However, MUC5AC expression was not meaningfully associated with recurrence-free survival (RFS). The patients positive for MUC5AC expression had a considerably shorter OS than those with negative expression. Conclusions: Our study provides insights into the clinical impact of mucins on AoV cancer, regardless of the histological subtype. Although MUC1 expression is universal, MUC5AC expression is a significant prognostic indicator that correlates with lymph node metastasis and poor OS. These results emphasize the possible utility of MUC5AC as a biomarker for extensive lymph node dissection and the prognostic evaluation of patients with AoV cancer. [ABSTRACT FROM AUTHOR]

Malignant pleural mesothelioma (MPM) is associated with asbestos exposure. Asbestos can induce chronic inflammation which in turn can lead to silencing of tumour suppressor genes. Wnt signaling pathway can be affected by chronic inflammation and is aberrantly activated in many cancers including colon and MPM. SFRP genes are antagonists of Wnt pathway, and SFRPs are potential tumour suppressors in colon, gastric, breast, ovarian, and lung cancers and mesothelioma. This study investigated the expression and DNA methylation of SFRP genes in MPM cells lines with and without demethylation treatment. Sixty-six patient FFPE samples were analysed and have showed methylation of SFRP2 (56%) and SFRP5 (70%) in MPM. SFRP2 and SFRP5 tumour-suppressive activity in eleven MPM lines was confirmed, and long-term asbestos exposure led to reduced expression of the SFRP1 and SFRP2 genes in the mesothelium (MeT-5A) via epigenetic alterations. Finally, DNA methylation of SFRPs is detectable in MPM patient plasma samples, with methylated SFRP2 and SFRP5 showing a tendency towards greater abundance in patients. These data suggested that SFRP genes have tumour-suppresive activity in MPM and that methylated DNA from SFRP gene promoters has the potential to serve as a biomarker for MPM patient plasma. [ABSTRACT FROM AUTHOR]

Background: Osteosarcoma (OSA) is a common malignant bone tumor of large breed dogs that occurs at predictable anatomic sites. At the time of initial diagnosis, most affected dogs have occult pulmonary metastases. Even with aggressive surgical treatment combined with chemotherapy, the majority of dogs diagnosed with OSA live less than 1 year from the time of diagnosis. The ability to identify canine OSA cases most responsive to treatment is needed. In humans, OSA is also an aggressive tumor that is histologically and molecularly similar to canine OSA. The expression of the tumor suppressor gene product P16 by human OSA tissue has been linked to a favorable response to chemotherapy. Results: We identified an antibody that binds canine P16 and developed a canine OSA tissue microarray in order to test the hypothesis that P16 expression by canine OSA tissue is predictive of clinical outcome following amputation and chemotherapy. Although statistical significance was not reached, a trend was identified between the lack of canine OSA P16 expression and a shorter disease free interval.Conclusions: The identification of a molecular marker for canine OSA is an important goal and the results reported here justify a larger study. [ABSTRACT FROM AUTHOR]

Non-alcoholic fatty liver disease (NAFLD), characterized by fat accumulation in liver, is closely associated with central obesity, over-nutrition and other features of metabolic syndrome, which elevate the risk of developing hepatocellular carcinoma (HCC). The Wnt/β-catenin signaling pathway plays a significant role in the physiology and pathology of liver. Up to half of HCC patients have activation of Wnt/β-catenin signaling. However, the mutation frequencies of CTNNB1 (encoding β-catenin protein) or other antagonists targeting Wnt/β-catenin signaling are low in HCC patients, suggesting that genetic mutations are not the major factor driving abnormal β-catenin activities in HCC. Emerging evidence has demonstrated that obesity-induced metabolic pathways can deregulate chromatin modifiers such as histone deacetylase 8 to trigger undesired global epigenetic changes, thereby modifying gene expression program which contributes to oncogenic signaling. This review focuses on the aberrant epigenetic activation of Wnt/ β-catenin in the development of NAFLD-associated HCC. A deeper understanding of the molecular mechanisms underlying such deregulation may shed light on the identification of novel druggable epigenetic targets for the prevention and/or treatment of HCC in obese and diabetic patients. [ABSTRACT FROM AUTHOR]

The article presents a case study of a 4-year old boy with an asymptomatic mass at the back of right thigh. The morphology and immunophenotype of the tumor corresponded to the characterized group of soft tissue spindle cell lesions defined by a relatively uniform cytomorphology, perivascular hyalinization, and S100 and CD34 coexpression. The article adds benefits of parallel sequencing as the types and number of gene fusions these tumors can render single-gene assay insensitive.

Patients with dementia are diagnosed with deficiency patterns and interior patterns in traditional Chinese medicine due to decreased physical strength, mental atrophy including cognitive function, and decreased motor function in the gastrointestinal tract. Since "greater yin symptom" in Shanghanlun has been interpreted as interior, deficiency, and cold pattern in traditional Chinese medicine, it is necessary to determine whether Geijigadaehwang-tang (GDT) has therapeutic effects on neurodegenerative diseases and the underlying mechanism if it has such effects. Aims of the study: Trimethyltin (TMT), a neurotoxic organotin compound, has been used to induce several neurodegenerative diseases, including epilepsy and Alzheimer's disease. This study aimed to evaluate the therapeutic efficacy of GDT for TMT-induced hippocampal neurodegeneration and seizures and to determine the mechanisms involved at the molecular level. The main components of GDT were analyzed using ultra-performance liquid chromatography. TMT was used to induce neurotoxicity in microglial BV-2 cells and C57BL6 mice. GDT was administered at various doses to determine its neuroprotective and seizure inhibition effects. The inhibitory effects of GDT on TMT-induced apoptosis, inflammatory pathways, and oxidative stress pathways were determined in the mouse hippocampal tissues. GDT contained emodin, chrysophanol, albiflorin, paeoniflorin, 6-gingerol, and liquiritin apioside. In microglial BV-2 cells treated with TMT, GDT showed dose-dependent neuroprotective effects. Oral administration of GDT five times for 2.5 days before and after TMT injection inhibited seizures at doses of 180 and 540 mg/kg and inhibited neuronal death in the hippocampus. In hippocampal tissues extracted from mice, GDT inhibited the protein expression of ionized calcium binding adaptor molecule 1, glial fibrillary acidic protein, nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing protein 3, and phosphorylated nuclear factor (NF)-κB/total-NFκB ratio. Additionally, GDT inhibited the messenger RNA levels of tumor necrosis factor-α, inducible nitric oxide synthase, apoptosis-associated speck-like protein containing a caspase recruitment domain, caspase-1, interleukin-1β, nuclear factor erythroid-2-related factor 2, and heme oxygenase-1. This study's results imply that GDT might have neuroprotective potential in neurodegenerative diseases through neuronal death inhibition and anti-inflammatory and antioxidant mechanisms. [Display omitted] [ABSTRACT FROM AUTHOR]

The expression of the myogenic regulatory factors (MRFs), Myf5, MyoD, myogenin ( Mgn) and MRF4 have been analysed during the development of chicken embryo somites and limbs. In somites, Myf5 is expressed first in somites and paraxial mesoderm at HH stage 9 followed by MyoD at HH stage 12, and Mgn and MRF4 at HH stage 14. In older somites, Myf5 and MyoD are also expressed in the ventrally extending myotome prior to Mgn and MRF4 expression. In limb muscles a similar temporal sequence is observed with Myf5 expression detected first in forelimbs at HH stage 22, MyoD at HH stage 23, Mgn at HH stage 24 and MRF4 at HH stage 30. This report describes the precise time of onset of expression of each MRF in somites and limbs during chicken embryo development, and provides a detailed comparative timeline of MRF expression in different embryonic muscle groups. [ABSTRACT FROM AUTHOR]

Background: During the fourth larval (L4) stage, vulval cells of C. elegans undergo extensive morphogenesis accompanied by changes in gene expression. This phase of vulval development, occurring after the well-studied induction of vulval cells, is not well understood but is potentially a useful context in which to study how a complex temporal sequence of events is regulated during development. However, a system for precisely describing different phases of vulval development in the L4 stage has been lacking. Results: We defined ten sub-stages of L4 based on morphological criteria as observed using Nomarski microscopy (L4.0 ~ L4.9). Precise timing of each sub-stage at 20 °C was determined. We also re-examined the timing of expression for several gene expression markers, and correlated the sub-stages with the timing of other developmental events in the vulva and the uterus. Conclusions: This scheme allows the developmental timing of an L4 individual to be determined at approximately one-hour resolution without the need to resort to time course experiments. These well-defined developmental stages will enable more precise description of gene expression and other developmental events. [ABSTRACT FROM AUTHOR]

Background: During the fourth larval (L4) stage, vulval cells of C. elegans undergo extensive morphogenesis accompanied by changes in gene expression. This phase of vulval development, occurring after the well-studied induction of vulval cells, is not well understood but is potentially a useful context in which to study how a complex temporal sequence of events is regulated during development. However, a system for precisely describing different phases of vulval development in the L4 stage has been lacking. Results: We defined ten sub-stages of L4 based on morphological criteria as observed using Nomarski microscopy (L4.0 ~ L4.9). Precise timing of each sub-stage at 20 °C was determined. We also re-examined the timing of expression for several gene expression markers, and correlated the sub-stages with the timing of other developmental events in the vulva and the uterus. Conclusions: This scheme allows the developmental timing of an L4 individual to be determined at approximately one-hour resolution without the need to resort to time course experiments. These well-defined developmental stages will enable more precise description of gene expression and other developmental events. [ABSTRACT FROM AUTHOR]

Background: Plasmodium knowlesi is one of five Plasmodium species known to cause malaria in humans and can result in severe illness and death. While a zoonosis in humans, this simian malaria parasite species infects macaque monkeys and serves as an experimental model for in vivo, ex vivo and in vitro studies. It has underpinned malaria discoveries relating to host-pathogen interactions, the immune response and immune evasion strategies. This study investigated differences in P. knowlesi gene expression in samples from ex vivo and in vitro cultures. Methods: Gene expression profiles were generated using microarrays to compare the stage-specific transcripts detected for a clone of P. knowlesi propagated in the blood of a rhesus macaque host and then grown in an ex-vivo culture, and the same clone adapted to long-term in vitro culture. Parasite samples covering one blood-stage cycle were analysed at four-hour intervals. cDNA was generated and hybridized to an oligoarray representing the P. knowlesi genome. Two replicate experiments were developed from in vitro cultures. Expression values were filtered, normalized, and analysed using R and Perl language and applied to a sine wave model to determine changes in equilibrium and amplitude. Differentially expressed genes from ex vivo and in vitro time points were detected using limma R/Bioconductor and gene set enrichment analysis (GSEA). Results: Major differences were noted between the ex vivo and in vitro time courses in overall gene expression and the length of the cycle (25.5 hours ex vivo; 33.5 hours in vitro). GSEA of genes up-regulated ex vivo showed an enrichment of various genes including SICAvar, ribosomal- associated and histone acetylation pathway genes. In contrast, certain genes involved in metabolism and cell growth, such as porphobilinogen deaminase and tyrosine phosphatase, and one SICAvar gene, were significantly up-regulated in vitro. Conclusions: This study demonstrates how gene expression in P. knowlesi blood-stage parasites can differ dramatically depending on whether the parasites are grown in vivo, with only one cycle of development ex vivo, or as an adapted isolate in long-term in vitro culture. These data bring emphasis to the importance of studying the parasite, its biology and disease manifestations in the context of the host. [ABSTRACT FROM AUTHOR]

Artemisinin resistance in Plasmodium falciparum threatens global efforts to control and eliminate malaria. Polymorphisms in the kelch domain-carrying protein K13 are associated with artemisinin resistance, but the underlying molecular mechanisms are unknown. We analyzed the in vivo transcriptomes of 1043 isolates from patients with P. falciparum acute malaria and found that artemisinin resistance is associated with increased expression of unfolded protein response (UPR) pathways involving the major PROSC and TRiC chaperone complexes. Artemisinin-resistant parasites also exhibit decelerated progression through the first part of the asexual intraerythrocytic development cycle. These findings suggest that artemisinin-resistant parasites remain in a state of decelerated development at the young ring stage, whereas their up-regulated UPR pathways mitigate protein damage caused by artemisinin. The expression profiles of UPR-related genes also associate with the geographical origin of parasite isolates, further suggesting their role in emerging artemisinin resistance in the Greater Mekong Subregion. [ABSTRACT FROM AUTHOR]

Rapid assembly of DNA damage response (DDR) proteins at nuclear “repair” foci is a hallmark response of ionizing radiation (IR)-treated cells. The ubiquitin E3 ligases RNF8 and RNF168 are critical for foci formation, and here we aim to determine their dynamic mobility and abundance at individual foci in living cells. To this end, YFP-tagged RNF8 and RNF168 were expressed at physiological levels in MCF-7 cells, then analyzed by fluorescence recovery after photobleaching (FRAP) assays, nuclear retention measurement, and virus-like particles (VLPs)-based quantification. The results showed that RNF8 and RNF168 were both highly dynamic at IR-induced foci. Intriguingly, RNF8 displayed remarkably faster in vivo association/dissociation rates than RNF168, and RNF8-positive IR-foci were less resistant to detergent extraction. In addition, copy number assay revealed that RNF168 was two-fold more abundant than RNF8 at foci. Collectively, we show for the first time that RNF8 moves on-and-off nuclear DNA repair foci more than six-fold as quickly as RNF168. The faster kinetics of RNF8 recruitment explains why RNF8 is generally observed at DNA-breaks prior to RNF168. Moreover, our finding that RNF8 is less abundant than RNF168 identifies RNF8 as a rate-limiting determinant of focal repair complex assembly. [ABSTRACT FROM AUTHOR]

The acetolactate synthase ( als)-deficient mutant of Klebsiella pneumoniae fails to produce 1,3-propanediol (1,3-PD) or 2,3-butanediol (2,3-BD), and is defective in glycerol metabolism. In an effort to recover production of the industrially valuable 1,3-PD, we introduced the Zymomonas mobilis pyruvate decarboxylase ( pdc) and aldehyde dehydrogenase ( aldB) genes into the als-deficient mutant to activate the conversion of pyruvate to ethanol. Heterologous expression of pdc and aldB efficiently recovered glycerol metabolism in the 2,3-BD synthesis-defective mutant, enhancing the production of 1,3-PD by preventing the accumulation of pyruvate. Production of 1,3-PD in the pdc- and aldB-expressing als-deficient mutant was further enhanced by increasing the aeration rate. This system uses metabolic engineering to produce 1,3-PD while minimizing the generation of 2,3-BD, offering a breakthrough for the industrial production of 1,3-PD from crude glycerol. [ABSTRACT FROM AUTHOR]

Pleiotrophin (PTN) is a growth factor with both pro-angiogenic and limited pro-tumorigenic activity. We evaluated the potential for PTN to be used for safe angiogenic gene therapy using the full length gene and a truncated gene variant lacking the domain implicated in tumorigenesis. Mouse myoblasts were transduced to express full length or truncated PTN (PTN or T-PTN), along with a LacZ reporter gene, and injected into mouse limb muscle and myocardium. In cultured myoblasts, PTN was expressed and secreted via the Golgi apparatus, but T-PTN was not properly secreted. Nonetheless, no evidence of uncontrolled growth was observed in cells expressing either form of PTN. PTN gene delivery to myocardium, and non-ischemic skeletal muscle, did not result in a detectable change in vascularity or function. In ischemic hindlimb at 14 days post-implantation, intramuscular injection with PTN-expressing myoblasts led to a significant increase in skin perfusion and muscle arteriole density. We conclude that (1) delivery of the full length PTN gene to muscle can be accomplished without tumorigenesis, (2) the truncated PTN gene may be difficult to use in a gene therapy context due to inefficient secretion, (3) PTN gene delivery leads to functional benefit in the mouse acute ischemic hindlimb model. [ABSTRACT FROM AUTHOR]

Simple Summary: Ubiquitin-specific Peptidase 13 (USP13) is highly amplified and promotes tumorigenesis and metastasis in ovarian cancer. However, post-translational modifications and their functions on USP13 are largely unknown. This study revealed that USP13 is phosphorylated at Thr122, which upregulates the stability of USP13 at a post-translation level. Notably, mutation of Thr122 diminished the effect of USP13 on the increased proliferation of ovarian cancer cells. Overall, we uncovered a new mechanism for the regulation of USP13 stability via a post-translational modification, suggesting novel therapeutics targeting USP13 in USP13-amplified cancers. Ubiquitin-specific Peptidase 13 (USP13) is a deubiquitinating enzyme that regulates the stability or function of its substrate. USP13 is highly amplified in human ovarian cancer, and elevated expression of USP13 promotes tumorigenesis and metastasis of ovarian cancer. However, there is little known about USP13 post-translational modifications and their role in ovarian cancer. Here, we found that USP13 is phosphorylated at Thr122 in ovarian cancer cells. Phosphorylated Thr122 (pT122) on endogenous USP13 was observed in most human ovarian cancer cells, and the abundance of this phosphorylation was correlated to the total level of USP13. We further demonstrated that Casein kinase 2 (CK2) directly interacts with and phosphorylates USP13 at Thr122, which promotes the stability of USP13 protein. Finally, we showed that Threonine 122 is important for cell proliferation of ovarian cancer cells. Our findings may reveal a novel regulatory mechanism for USP13, which may lead to novel therapeutic targeting of USP13 in ovarian cancer. [ABSTRACT FROM AUTHOR]

Abstract: MicroRNAs (miRNAs) generally pair with the 3′UTRs of their target mRNAs to repress gene expression. It has reported that miRNA targets (TGs) are longer and evolve more slowly than non-targets (NTGs). We confirmed the observation and also found novel structural and expressional characteristics of TGs. The length difference between TGs and NTGs was greatest for the 3′UTRs, although a difference was also observed for CDSs and introns. Widely expressed genes were shorter for both TGs and NTGs; however, TGs were significantly longer than NTGs in all ranges of expression. TGs were more likely than NTGs to be widely expressed, which might explain why TGs evolve more slowly than NTGs. Finally, we found that TG mRNAs have faster decay rates. In addition, the decay rate of a TG mRNA transcript was found to be positively correlated with the number or density of target sites located in that TG''s mRNA transcript. [Copyright &y& Elsevier]

Abstract: Simultaneous silencing of multiple up-regulated genes is an attractive and viable strategy to treat many incurable diseases including cancer. Herein we report that multimerized siRNA conjugate composed of two different siRNA sequences in the same backbone shows more efficient inhibition of the two corresponding target genes at one time than physically mixed multimerized siRNA conjugates. Two model siRNAs against VEGF and GFP gene were chemically crosslinked via cleavable and noncleavable linkages for the preparation of dual gene targeted multimeric siRNA conjugates (DGT multi-siRNA). Cleavable DGT multi-siRNA with reducible disulfide linkages exhibited significantly higher gene silencing efficiencies at mRNA and protein expression levels than noncleavable DGT multi-siRNA, the physical mixture of naked siRNA, and that of single gene targeted multimeric siRNA (SGT multi-siRNA) with eliciting negligible immune response. DGT multi-siRNAs against two therapeutic siRNAs, anti-survivin and anti-bcl-2 targeted siRNA, also showed greatly enhanced apoptotic effect. This approach for concurrent suppression of combinatorial therapeutic target genes using cleavable multimeric siRNA structure can be potentially used for improved therapeutic efficacy. [ABSTRACT FROM AUTHOR]

Abstract: A gene encoding a serine protease inhibitor of Clonorchis sinensis (CsSERPIN) was identified and characterized. CsSERPIN contained an open reading frame of 1158bp that encoded 385 amino acid residues. Sequence analysis of the primary structure of CsSERPIN revealed that it had essential structural motifs including a reactive central loop (RCL), which well conserved in the serine protease inhibitor (serpin) superfamily. CsSERPIN was classified as a member of the ovalbumin-type serpin family on the basis of phylogenetic analysis and the absence of a classical N-terminal signal peptide. Recombinant CsSERPIN showed an inhibitory effect on chymotrypsin in a dose-dependent manner, but did not effectively inhibit trypsin, thrombin, elastases or cathepsin G. Optimal pH values of CsSERPIN were between 7.0 and 9.0, as evidenced by the rapid loss of inhibitory activity under acidic conditions. CsSERPIN was expressed at various developmental stages of the parasite, from eggs to adult worms, but its expression level was higher in eggs and adult worms than in metacercariae and juvenile worms. CsSERPIN was identified in the soluble extract of the parasite, but not in the excretory and secretory products (ESP) or insoluble extract of the parasite. Immunolocalization analysis of CsSERPIN showed that it mainly localized to the eggs and vitelline glands of the adult worm. These results suggest that intracellular CsSERPIN may be possibly involved in maintaining the physiology of eggs as well as in egg production of C. sinensis by regulating endogenous serine proteases. [ABSTRACT FROM AUTHOR]

Background and Purpose: Icariin may be the active ingredient in Herba Epimedii, a Chinese herb commonly used for treatment of osteoporosis. The present study aims to delineate the mechanism(s) by which icariin prevents bone loss after ovariectomy (OVX) in vivo and stimulates osteoblastic functions in vitro.Experimental Approach: Ovariectomized or sham-operated C57BL/6 mice were treated with vehicle, 17beta-oestradiol or icariin for 6 weeks. Total and trabecular bome mineral density (BMD) as well as polar stress-strain index of distal femur were measured by peripheral computed tomography. The mRNA expressions of OPG and RANKL in tibia were studied by RT-PCR. Interactions between the oestrogen receptor (ER) antagonist ICI182,780 and icariin were studied in UMR 106 cells. The functional transactivation of ERalpha and ERbeta as well as ERalpha phosphorylation by icariin were also assessed.Key Results: Icariin suppressed the loss of bone mass and strength in distal femur and increased the mRNA expression ratio of OPG/RANKL in tibia, following OVX. Icariin increased ER-dependent cell proliferation, alkaline phosphatase (ALP) activity, gene expression of OPG and the OPG/RANKL ratio in UMR 106 cells. Icariin did not activate ERE-luciferase activity in UMR 106 cells, via the ERalpha or the ERbeta-mediated pathway, but it did increase ERalpha phosphorylation at Ser118.Conclusions and Implications: Our results indicate that icariin exerts anabolic effects in bone possibly by activating ER in a ligand-independent manner. Its ability to prevent OVX-induced bone loss without inducing uterotrophic effects supports its use as an alternative regimen for management of postmenopausal osteoporosis. [ABSTRACT FROM AUTHOR]

Summary: Advanced stage papillary serous tumors of the ovary are responsible for the majority of ovarian cancer deaths, yet the molecular determinants modulating patient survival are poorly characterized. Here, we identify and validate a prognostic gene expression signature correlating with survival in a series of microdissected serous ovarian tumors. Independent evaluation confirmed the association of a prognostic gene microfibril-associated glycoprotein 2 (MAGP2) with poor prognosis, whereas in vitro mechanistic analyses demonstrated its ability to prolong tumor cell survival and stimulate endothelial cell motility and survival via the αVβ3 integrin receptor. Increased MAGP2 expression correlated with microvessel density suggesting a proangiogenic role in vivo. Thus, MAGP2 may serve as a survival-associated target. [Copyright &y& Elsevier]

Abstract: Proline-rich membrane anchor (PRiMA) is a molecule to organize acetylcholinesterase (AChE) into tetrameric globular form (G4) that anchors onto the plasma membrane in brain and muscle. In mammal, PRiMA is encoded by a single gene with two splicing variants, PRiMA I and PRiMA II: PRiMA II is different to PRiMA I by its absence of a C-terminal cytoplasmic domain. The existence of these isoforms has not been revealed in avian specie. By using RT-PCR and bioinformatic analyses, two splicing variants of PRiMA were identified in chicken cerebrum. One variant contains very similar domains as compared to mammalian PRiMA I. The other variant, named as PRiMA II, has a very distinct cytoplasmic C-terminus of having 26 amino acids. Both forms of chicken PRiMA were able to organize the formation of G4 AChE when that was over expressed together with AChET subunit in cultured cells. The level of PRiMA mRNA, mainly PRiMA I, was higher in slow-twitch muscle than that of in fast-twitch muscle of chicken. This finding suggests that the muscle fiber type-specific expression of G4 AChE in chicken could be a result of the different expression pattern of PRiMA in fast- and slow-twitch muscles. [Copyright &y& Elsevier]

Abstract: A good understanding of telomeres and telomerase biology is crucial for unraveling mechanisms related to aging and cancer. However, in vivo vertebrate studies of telomere biogenesis and telomerase function have been limited by the development of appropriate animal model systems. The present study aims to demonstrate evolutionary conservation of telomerase in vertebrate species, supporting the potential application of fish as vertebrate model for studying telomeres and telomerase function. Comparison of genomic and protein information among vertebrate TERTs (TElomerase Reverse Transcriptase), the Japanese medaka Oryzias latipes shares the highest similarity to that of the human than the other small size fish species studied (including pufferfish and zebrafish). The ubiquitous expression of TERT mRNA, the high constitutive level of telomerase activity, and the humanized telomere lengths further substantiate that Japanese medaka is an ideal vertebrate model for the study of telomere and telomerase-related mechanisms in vivo. Moreover, medaka exhibits fast, invariable growth and is able to provide a variety of useful developmental and reproductive endpoints for lifelong and multi-generational experiments. Our earlier and present findings support the use of medaka for studying organismal aging, tissue regeneration and carcinogenesis. [Copyright &y& Elsevier]

Human immunodeficiency virus (HIV)-based vectors are being increasingly used in vitro for gene transfer and in vivo for gene therapy. The proportion of integrated retroviral vectors that are silenced or remain transcriptionally active, and the stability of gene expression in the latter remains poorly explored. To study this, T cells were infected with an HIV-1-based vector construct containing a long terminal repeat-driven reporter gene. Only a small percentage of detectable integrated vector expressed gene product. In clones derived from cells with transcriptionally active vector, gene expression was remarkably stable with more than 80% continuing to express for greater than 18 months. Failure to continue expressing the vector was associated with epigenetic changes. Our data suggest that there are two forms of vector silencing: one occurring immediately after integration affecting the majority of the vectors, and one occurring in the much longer term affecting a small minority of vectors which had previously established expression.Gene Therapy (2007) 14, 741–751. doi:10.1038/sj.gt.3302923; published online 1 March 2007 [ABSTRACT FROM AUTHOR]

We isolated a transposon-induced nonmotile mutant of the cyanobacterium Synechocystis sp. PCC 6803. The mutant was revealed to have a Tn5 insertion in the slr1443 gene that showed sequence similarity to a eukaryotic-type protein kinase. Thick pili were not observed on the mutant cell surface under the electron microscope. The slr1443 gene was not involved in transcription or translation of the pilA1 gene encoding pilin, the major component of thick pili. In the mutant, lower molecular mass pilin peptides were detected than in the wild-type. The pilin variant was not truncated at the N- or C-terminus of mature PilA1. The reduced molecular mass may have resulted from insufficient post-translational modification. The amounts of pilin variants were remarkably reduced in the periplasmic and surface fractions. The pilin variants were released into liquid media without being assembled into pili. Our finding suggests that Slr1443 plays an important role in pilus biogenesis at the level of the post-translational modification of pilin. [Copyright &y& Elsevier]

The molecular events that drive the initiation and progression of ovarian adenocarcinoma are not well defined. We have investigated changes in gene expression in ovarian cancer cell lines compared to an immortalized human ovarian surface epithelial cell line (HOSE) using a cDNA array. We identified 17 genes that were under-expressed and 10 genes that were over-expressed in the cell lines compared to the HOSE cells. One of the genes under-expressed in the ovarian cancer cell lines, Id3, a transcriptional inactivator, was selected for further investigation. Id3 mRNA was expressed at reduced levels in 6 out of 9 ovarian cancer cell lines compared to the HOSE cells while at the protein level, all 7 ovarian cancer cell lines examined expressed the Id3 protein at greatly reduced levels. Expression of Id3 mRNA was also examined in primary ovarian tumours and was found in only 12/38 (32%) cases. A search was conducted for mutations ofId3 in primary ovarian cancers using single stranded conformation polymorphism (SSCP) analysis. Only one nucleotide substitution, present also in the corresponding constitutional DNA, was found in 94 ovarian tumours. Furthermore no association was found between LOH at 1p36 and lack of expression of Id3. These data suggest that Id3 is not the target of LOH at 1p36. [ABSTRACT FROM AUTHOR]

The modification of synaptic and neural connections in adults, including the formation and removal of synapses, depends on activity‐dependent synaptic and structural plasticity. MicroRNAs (miRNAs) play crucial roles in regulating these changes by targeting specific genes and regulating their expression. The fact that somatic and dendritic activity in neurons often occurs asynchronously highlights the need for spatial and dynamic regulation of protein synthesis in specific milieu and cellular loci. MicroRNAs, which can show distinct patterns of enrichment, help to establish the localized distribution of plasticity‐related proteins. The recent study using atomic force microscopy (AFM)‐based nanoscale imaging reveals that the abundance of miRNA(miR)‐134 is inversely correlated with the functional activity of dendritic spine structures. However, the miRNAs that are selectively upregulated in potentiated synapses, and which can thereby support prospective changes in synaptic efficacy, remain largely unknown. Using AFM force imaging, significant increases in miR‐132 in the dendritic regions abutting functionally‐active spines is discovered. This study provides evidence for miR‐132 as a novel positive miRNA regulator residing in dendritic shafts, and also suggests that activity‐dependent miRNAs localized in distinct sub‐compartments of neurons play bi‐directional roles in controlling synaptic transmission and synaptic plasticity. [ABSTRACT FROM AUTHOR]

Although there have been several analyses for identifying cancer-associated pathways, based on gene expression data, most of these are based on single pathway analyses, and thus do not consider correlations between pathways. In this paper, we propose a hierarchical structural component model for pathway analysis of gene expression data (HisCoM-PAGE), which accounts for the hierarchical structure of genes and pathways, as well as the correlations among pathways. Specifically, HisCoM-PAGE focuses on the survival phenotype and identifies its associated pathways. Moreover, its application to real biological data analysis of pancreatic cancer data demonstrated that HisCoM-PAGE could successfully identify pathways associated with pancreatic cancer prognosis. Simulation studies comparing the performance of HisCoM-PAGE with other competing methods such as Gene Set Enrichment Analysis (GSEA), Global Test, and Wald-type Test showed HisCoM-PAGE to have the highest power to detect causal pathways in most simulation scenarios. [ABSTRACT FROM AUTHOR]

Selective herbicides are used to control undesirable vegetation or weeds in fields without harming crops. Herbicide use for weed management can directly impact the densities of insect pests in agricultural communities as a result of insect mortality during and immediately after application. In insects, the glutathione S‐transferase (GST) enzyme is involved in both the detoxification process and the defense of cellular membranes against oxidative damage. In this study, two TmGSTs (TmGST‐iso1 and TmGST‐iso2) were identified and characterized to elucidate the GST family in Tenebrio molitor. Among the developmental stages of T. molitor, eggs had the highest expression levels of TmGST‐iso1. TmGST‐iso2 expression was highest in the pre‐pupal stage. TmGST‐iso1 expression was high in the guts of early and late larvae, whereas TmGST‐iso2 expression was not observed in early larvae. Adults, both male and female, had the highest levels of TmGST‐iso1 mRNA expression in the gut and reproductive organs, and the highest levels of TmGST‐iso2 expression in the reproductive organs. Quantitative polymerase chain reaction was used to investigate the impact of treatment with butachlor on the mRNA expression of TmGST‐iso1 and TmGST‐iso2 in larvae. TmGST‐iso1 expression increased in the butachlor‐treated group after 3 and 24 h, whereas TmGST‐iso2 expression peaked at 24 h after treatment. This study provides vital information about the detoxifying activities of T. molitor. [ABSTRACT FROM AUTHOR]

Agricultural intensification has led to significant increases in production, but the overuse of pesticides and associated hazards pose threats to biodiversity and ecological functions. Catalase (CAT), a key antioxidant enzyme, plays a crucial role in alleviating oxidative stress by directly interacting with toxins. In this study, we identified three CAT isoforms in Tenebrio molitor (TmCAT‐iso1, TmCAT‐iso2 and TmCAT‐iso3). These CATs possess a CAT domain, tetramer interface sites and a heme‐binding pocket. We examined the expression of Tm catalases across all developmental stages and in specific tissues using quantitative real time polymerase chain reaction (qRT‐PCR) experiments. Our findings demonstrate that TmCAT‐iso1 and TmCAT‐iso3 exhibit peak expression in young and late larval stages, respectively, whereas TmCAT‐iso2 shows peak expression during the egg and pre‐pupal stages. Tissue distribution analysis revealed the high expression of TmCAT‐iso1 and TmCAT‐iso2 in larval hemocytes, whereas TmCAT‐iso3 is predominantly expressed in larval Malpighian tubules. Furthermore, injection with chlorantraniliprole significantly elevated mRNA expression levels of TmCAT‐iso1, TmCAT‐iso2, and TmCAT‐iso3 in larval groups, compared with control groups. Our study highlights the distinct developmental stages and tissues where TmCATs are expressed. We also elucidated the effects of pesticide application on the expression of each TmCAT, revealing the physiological characteristics of CATs in response to these pesticides, which are dose‐ and time‐dependent in T. molitor. [ABSTRACT FROM AUTHOR]

Structural maintenance of chromosomes flexible hinge domain-containing 1 (SMCHD1) is an epigenetic regulator that mediates gene expression silencing at targeted sites across the genome. Our current understanding of SMCHD1's molecular mechanism, and how substitutions within SMCHD1 lead to the diseases, facioscapulohumeral muscular dystrophy (FSHD) and Bosma arhinia microphthalmia syndrome (BAMS), are only emerging. Recent structural studies of its two component domains — the N-terminal ATPase and C-terminal SMC hinge — suggest that dimerization of each domain plays a central role in SMCHD1 function. Here, using biophysical techniques, we demonstrate that the SMCHD1 ATPase undergoes dimerization in a process that is dependent on both the N-terminal UBL (Ubiquitin-like) domain and ATP binding. We show that neither the dimerization event, nor the presence of a C-terminal extension past the transducer domain, affect SMCHD1's in vitro catalytic activity as the rate of ATP turnover remains comparable to the monomeric protein. We further examined the functional importance of the N-terminal UBL domain in cells, revealing that its targeted deletion disrupts the localization of full-length SMCHD1 to chromatin. These findings implicate UBL-mediated SMCHD1 dimerization as a crucial step for chromatin interaction, and thereby for promoting SMCHD1-mediated gene silencing. [ABSTRACT FROM AUTHOR]

The Kir4.1 channel, an inwardly rectifying potassium ion (K+) channel, is located in the hair cells of the organ of Corti as well as the intermediate cells of the stria vascularis. The Kir4.1 channel has a crucial role in the generation of endolymphatic potential and maintenance of the resting membrane potential. However, the role and functions of the Kir4.1 channel in the progenitor remain undescribed. To observe the role of Kir4.1 in the progenitor treated with the one-shot ototoxic drugs (kanamycin and furosemide), we set the proper condition in culturing Immortomouse-derived HEI-OC1 cells to express the potassium-related channels well. And also, that was reproduced in mice experiments to show the important role of Kir4.1 in the survival of hair cells after treating the ototoxicity drugs. In our results, when kanamycin and furosemide drugs were cotreated with HEI-OC1 cells, the Kir4.1 channel did not change, but the expression levels of the NKCC1 cotransporter and KCNQ4 channel are decreased. This shows that inward and outward channels were blocked by the two drugs (kanamycin and furosemide). However, noteworthy here is that the expression level of Kir4.1 channel increased when kanamycin was treated alone. This shows that Kir4.1, an inwardly rectifying potassium channel, acts as an outward channel in place of the corresponding channel when the KCNQ4 channel, an outward channel, is blocked. These results suggest that the Kir4.1 channel has a role in maintaining K+ homeostasis in supporting cells, with K+ concentration compensator when the NKCC1 cotransporter and Kv7.4 (KCNQ4) channels are deficient. [ABSTRACT FROM AUTHOR]

Objectives: Periodontitis is a highly prevalent chronic inflammatory disease that results in destruction of tooth‐supporting structures followed by tooth‐loss. Until now, periodontitis has been regarded to be initiated by bacterial infection followed by aberrant host response. Although increasing evidence suggests a strong association between oxidative stress and periodontitis, precise molecular mechanism has been left unanswered. In this study, we investigated roles of SOD2, the main antioxidant enzyme maintaining reactive oxygen species (ROS) homeostasis, under inflammatory conditions. Methods: We computationally analyzed SOD2 expression in periodontitis. To confirm this data, immunoblot assay was performed with samples from periodontitis patients. The cellular mechanism of change in SOD2 expression was identified through immunoblot assay and immunofluorescence. To evaluate the molecular function of SOD2, we generated SOD2‐deficient cells by utilizing the CRISPR/Cas9 system. Results: We first determined that SOD2 expression was significantly increased in periodontitis. We also confirmed that SOD2 expression was upregulated through the NF‐κB pathway when the inflammatory signal was stronger and extended. Gene manipulation against SOD2 through the CRISPR/Cas9 system showed that the absence of SOD2 increased production of NLRP3 inflammasome components. Conclusions: Our study demonstrates that intracellular SOD2 has a protective role by suppressing NLRP inflammasome‐caspase‐1‐IL‐1β axis under inflammatory conditions. [ABSTRACT FROM AUTHOR]

Backgroud: There were limitations existing in programmed cell‐death ligand 1 (PD‐L1) as predictive biomarkers for breast cancer (BC), hence exploring the correlation between PD‐L1 levels and other biomarkers in BC may become a very useful therapeutic clinical tool. Methods: A total of 301 Chinese patients with different BC subtypes including 47 HR+/HER2+, 185 HR+/HER2−, 38 HR−/HER2+, and 31 triple‐negative breast cancer (TNBC) were enrolled in our study. Next‐generation sequencing based Yuansu450 gene panel was used for genomic alteration identification and PD‐L1 expression was tested using immunohistochemistry. Results: The most prevalent BC‐related mutations were TP53 mutations, followed by mutations in PIK3CA, ERBB2, CDK12, and GATA3 in our Chinese cohort. We found that mutations DDR2 and MYCL were only mutated in HR−/HER2+ subtype, whereas H3‐3A and NRAS mutations were only occurred in HR−/HER2− subtype. The percentage of patients with PD‐L1‐positive expression was higher in patients with HR‐/HER2− mainly due to the percentage of PD‐L1‐high level. Mutational frequencies of TP53, MYC, FAT4, PBRM1, PREX2 were observed to have significant differences among patients with different BC subtypes based on PD‐L1 levels. Moreover, a positive correlation was observed between TMB and PD‐L1 level in HR+/HER2− subtype, and showed that the proportion of patients with high PD‐L1 expression was higher than that of patients with low PD‐L1 expression in the HR+/HER2− and HR+/HER2+ cohorts with high Ki67 expression. Conclusions: The genomic alterations based on PD‐L1 and other biomarkers of different cohorts may provide more possibilities for the treatment of BC with different subtypes. [ABSTRACT FROM AUTHOR]

Cannabidiol, a non-psychoactive phytocannabinoid, has antioxidant and anti-inflammatory activity in keratinocytes. However, the signaling pathway through which cannabidiol exerts its effect on keratinocytes or whether it can modulate keratinocyte differentiation has not been fully elucidated yet. We investigated whether cannabidiol modulates epidermal differentiation and scavenges reactive oxygen species through the aryl hydrocarbon receptor (AhR) in keratinocytes and epidermal equivalents. We investigated the cannabidiol-induced activation of AhR using AhR luciferase reporter assay, qRT-PCR, western blot, and immunofluorescence assays. We also analyzed whether keratinocyte differentiation and antioxidant activity are regulated by cannabidiol-induced AhR activation. In both keratinocytes and epidermal equivalents, cannabidiol increased both the mRNA and protein expression of filaggrin, involucrin, NRF2, and NQO1 and the mRNA expression of the AhR target genes, including CYP1A1 and aryl hydrocarbon receptor repressor. Additionally, cannabidiol showed antioxidant activity that was attenuated by AhR knockdown or co-administration with an AhR antagonist. Moreover, cannabidiol increased the ratio of OVOL1/OVOL2 mRNA expression, which is a downstream regulator of AhR that mediates epidermal differentiation. In addition to increased expression of barrier-related proteins, cannabidiol-treated epidermal equivalent showed a more prominent granular layer than the control epidermis. The increased granular layer by cannabidiol was suppressed by the AhR antagonist. Cannabidiol can be a modulator of the AhR-OVOL1-filaggrin axis and AhR-NRF2-NQO1 signaling, thus indicating a potential use of cannabidiol in skin barrier enhancement and reducing oxidative stress. • CBD regulates AhR in keratinocytes and epidermal equivalents. • CBD induces epidermal differentiation by modulating the AhR-OVOL1 axis. • AhR orchestrates ROS scavenging through NRF2 regulation. [ABSTRACT FROM AUTHOR]

Ethnopharmacological relevance Lonicera japonica Thunberg is a traditional herbal medicine widely used in East Asia as an anti-bacterial, anti-inflammatory, and antiviral agent. This study was designed to investigate the effects of HS-23, ethanol extract of the dried flower buds of Lonicera japonica, in experimental models of sepsis and elucidate the mechanisms of action of HS-23. Materials and methods Male ICR mice were intravenously administered HS-23 (20 and 40 mg/kg) for 0 (immediately) and 24 h after cecal ligation and puncture (CLP) for survival tests, and HS-23 (40 mg/kg) immediately after CLP for biochemical assays. Results HS-23 improved sepsis-induced mortality, enhanced bacterial clearance, and attenuated multiple organ failure. The mechanisms of action of HS-23 included attenuation of increased toll-like receptor (TLR)4 protein and mRNA expression. HS-23 suppressed sepsis-induced increases in protein expression of myeloid differentiation primary response protein 88, p38 and c-Jun N-terminal kinase in both liver and lung, as well as TIR-domain-containing adapter-inducing interferon-β and interferon regulatory transcription factor 3 protein expression in liver. Conclusion The results of this study revealed that HS-23 attenuated sepsis through suppression of TLR signaling pathways. Therefore, our findings suggest that HS-23 might be useful as a potential therapeutic agent for treatment of sepsis. [ABSTRACT FROM AUTHOR]

Background: Genetic studies reveal that single-nucleotide polymorphisms (SNPs) of SPI1 are associated with Alzheimer's disease (AD), while their effects in the Chinese population remain unclear.Objective: We aimed to examine the AD-association of SPI1 SNPs in the Chinese population and investigate the underlying mechanisms of these SNPs in modulating AD risk.Methods: We conducted a genetic analysis of three SPI1 SNPs (i.e., rs1057233, rs3740688, and rs78245530) in a Chinese cohort (n = 333 patients with AD, n = 721 normal controls). We also probed public European-descent AD cohorts and gene expression datasets to investigate the putative functions of those SNPs.Results: We showed that SPI1 SNP rs3740688 is significantly associated with AD in the Chinese population (odds ratio [OR] = 0.72 [0.58-0.89]) and identified AD-protective SPI1 haplotypes β (tagged by rs1057233 and rs3740688) and γ (tagged by rs3740688 and rs78245530). Specifically, haplotypes β and γ are associated with decreased SPI1 gene expression level in the blood and brain tissues, respectively. The regulatory roles of these haplotypes are potentially mediated by changes in miRNA binding and the epigenetic landscape. Our results suggest that the AD-protective SPI1 haplotypes regulate pathways involved in immune and neuronal functions.Conclusion: This study is the first to report a significant association of SPI1 with AD in the Chinese population. It also identifies SPI1 haplotypes that are associated with SPI1 gene expression and decreased AD risk. [ABSTRACT FROM AUTHOR]

Breast cancer is highly heterogenous with temporal and spatial heterogeneity making it necessary for rebiopsy. DS-8201a, a new potential therapy for human epidermal growth factor receptor 2 (HER2) low expression breast cancer, had been proved that it could overcome heterogenous HER2 expression in a preclinical setting. In January 2014, a 23-year-old woman was presented with a lump in the right breast with bone metastasis, diagnosed as infiltrating ductal carcinoma, estrogen receptor (ER)+, progesterone receptor (PR)+, HER2 immunohistochemistry (IHC) 2+, and fluorescence in situ hybridization negative. The patient received a series of therapies including surgery, radiotherapy, endocrine therapy, target therapy, and chemotherapy. The longest progression-free survival was 17 months after surgery. Biopsy of liver metastasis in February 2020 showed triple negative (HER2−, ER−, PR−), which was quite different from the initial diagnosis in 2014, so retesting was performed and the results showed ER−, PR+ by 10%, HER2 IHC score of 1+, indicating heterogeneity of HER2 expression. In May 2020, DS-8201a treatment was initiated and continued for 10 cycles until November 2020. Remarkable relief in symptoms was observed after the first dose. A reduction in the metastatic lesion size (liver and brain) and improved liver function was observed during the therapy. This case indicated the heterogeneity of breast cancer, and impressive efficacy of DS-8201a in a heavily treated patient with HER2-low and HER2 heterogeneity. [ABSTRACT FROM AUTHOR]

Gekko gecko is used as a traditional medicine for various diseases including respiratory disorders in northeast Asian countries, mainly Korea, Japan, and China. Allergic asthma is a chronic respiratory disease caused by an inappropriate immune response. Due to the recent spread of coronavirus disease 2019, interest in the treatment of pulmonary disorders has rapidly increased. In this study, we investigated the anti-asthmatic effects of G. gecko extract (GGE) using an established mouse model of ovalbumin-induced asthma. To evaluate the anti-asthmatic effects of GGE, we evaluated histological changes and the responses of inflammatory mediators related to allergic airway inflammation. Furthermore, we investigated the regulatory effects of GGE on type 2 helper T (Th2) cell activation. Administration of GGE attenuated asthmatic phenotypes, including inflammatory cell infiltration, mucus production, and expression of Th2 cytokines. Furthermore, GGE treatment reduced Th2 cell activation and differentiation. These results indicate that GGE alleviates allergic airway inflammation by regulating Th2 cell activation and differentiation. [Display omitted] • Gekko gecko is traditionally used as therapeutic agent to treat pulmonary diseases in Asian countries. • Gekko gecko extracts reduced inflammatory cell infiltration and goblet cell hyperplasia in airway. • Gekko gecko extracts decreased allergic inflammation by regulating Th2 cell activation and polarization. [ABSTRACT FROM AUTHOR]

Glucose-6-phosphatase-β (G6Pase-β or G6PC3) deficiency, also known as severe congenital neutropenia syndrome 4, is characterized not only by neutropenia but also by impaired neutrophil energy homeostasis and functionality. We now show the syndrome is also associated with macrophage dysfunction, with murine G6pc3-/- macrophages having impairments in their respiratory burst, Chemotaxis, calcium flux, and phagocytic activities. Consistent with a glucose-6-phosphate (G6P) metabolism deficiency, G6pc3-/- macrophages also have a lower glucose uptake and lower levels of G6P, lactate, and ATP than wild-type macrophages. Furthermore, the expression of NADPH oxidase subunits and membrane translocation of p47Phox are down-regulated, and G6pc3-/- macrophages exhibit repressed trafficking in vivo both during an inflammatory response and in pregnancy. During pregnancy, the absence of G6Pase-ß activity also leads to impaired energy homeostasis in the uterus and reduced fertility of G6pc3-/- mothers. Together these results show that immune deficiencies in this congenital neutropenia syndrome extend beyond neutrophil dysfunction. [ABSTRACT FROM AUTHOR]

Infections with variant Creutzfeldt-Jakob disease (vCJD) have almost exclusively occurred in young patients, but the reasons for this age distribution are uncertain. Our data suggest that the pathogenesis of many peripherally acquired transmissible spongiform encephalopathy (TSE) agents is less efficient in aged individuals. Four vCJD cases linked to transfusion of vCJD-contaminated blood or blood products have been described. Three cases occurred in elderly patients, implying that intravenous exposure is more efficient in aged individuals than other peripheral routes. To test this hypothesis, young (6 to 8 weeks old) and aged (600 days old) mice were injected intravenously with a TSE agent. In aged and young mice, the intravenous route was more efficient than other peripheral routes of TSE agent exposure. However, in aged mice, disease pathogenesis was significantly reduced. Although most aged mice failed to develop clinical disease during their life spans, many showed histopathological signs of TSE disease in their brains. Thus, the effects of age on intravenous TSE pathogenesis may lead to significant levels of subclinical disease in the population. After peripheral exposure, many TSE agents accumulate upon follicular dendritic cells (FDCs) in lymphoid tissues before they infect the brain. In aged spleens, PrPC expression and TSE agent accumulation upon FDCs were reduced. Furthermore, the splenic marginal zone microarchitecture was substantially disturbed, adversely affecting the delivery of immune complexes to FDCs. This study is the first to suggest that the effects of aging on the microarchitecture and the function of the splenic marginal zone significantly influence the pathogenesis of an important pathogen. [ABSTRACT FROM AUTHOR]

Identification of gene expression profiles of cancer stem cells may have significant implications in the understanding of tumor biology and for the design of novel treatments targeted toward these cells. Here we report a potential ovarian cancer stem cell gene expression profile from isolated side population of fresh ascites obtained from women with high-grade advanced stage papillary serous ovarian adenocarcinoma. Affymetrix U133 Plus 2.0 microarrays were used to interrogate the differentially expressed genes between side population (SP) and main population (MP), and the results were analyzed by paired T-test using BRB-ArrayTools. We identified 138 up-regulated and 302 down-regulated genes that were differentially expressed between all 10 SP/MP pairs. Microarray data was validated using qRT-PCR and17/19 (89.5%) genes showed robust correlations between microarray and qRT-PCR expression data. The Pathway Studio analysis identified several genes involved in cell survival, differentiation, proliferation, and apoptosis which are unique to SP cells and a mechanism for the activation of Notch signaling is identified. To validate these findings, we have identified and isolated SP cells enriched for cancer stem cells from human ovarian cancer cell lines. The SP populations were having a higher colony forming efficiency in comparison to its MP counterpart and also capable of sustained expansion and differentiation in to SP and MP phenotypes. 50,000 SP cells produced tumor in nude mice whereas the same number of MP cells failed to give any tumor at 8 weeks after injection. The SP cells demonstrated a dose dependent sensitivity to specific γ-secretase inhibitors implicating the role of Notch signaling pathway in SP cell survival. Further the generated SP gene list was found to be enriched in recurrent ovarian cancer tumors. [ABSTRACT FROM AUTHOR]

Targeting host factors is a complementary strategy for the development of new antiviral drugs. We screened a library of isoxazolidine and isoxazole sulfonamides and found four compounds that inhibited HIV-1 infection in human CD4+ lymphocytic T cells with no toxicity at IC90 concentrations. Structure-activity relationship showed that benzyl sulfonamides and a halo-substituted aromatic ring on the heterocycle scaffold were critical for antiretroviral activity. The size and position of the incorporated halogen had a marked effect on the antiretroviral activity. The sulfonamide derivatives had no significant effect on HIV-1 entry, reverse transcription and integration but impaired a step necessary for activation of viral gene expression. This step was Tat-independent, strongly suggesting that the target is a cell factor. A virus partially resistant to the least potent compounds could be selected but could not be propagated in the long term, consistent with the possibility that HIV-1 may be less likely to develop resistance against drugs targeting some host factors. Here, we provide evidence that novel synthetic methods can be applied to develop small molecules with antiretroviral activity that target host factors important for HIV-1 replication. [ABSTRACT FROM AUTHOR]

Mechanisms for differential regulation of gene expression may underlie much of the phenotypic variation and adaptability of malaria parasites. Here we describe transcriptional variation among culture-adapted field isolates of Plasmodium falciparum, the species responsible for most malarial disease. It was found that genes coding for parasite protein export into the red cell cytosol and onto its surface, and genes coding for sexual stage proteins involved in parasite transmission are upregulated in field isolates compared with long-term laboratory isolates. Much of this variability was associated with the loss of small or large chromosomal segments, or other forms of gene copy number variation that are prevalent in the P. falciparum genome (copy number variants, CNVs). Expression levels of genes inside these segments were correlated to that of genes outside and adjacent to the segment boundaries, and this association declined with distance from the CNV boundary. This observation could not be explained by copy number variation in these adjacent genes. This suggests a localacting regulatory role for CNVs in transcription of neighboring genes and helps explain the chromosomal clustering that we observed here. Transcriptional co-regulation of physical clusters of adaptive genes may provide a way for the parasite to readily adapt to its highly heterogeneous and strongly selective environment. [ABSTRACT FROM AUTHOR]

Abstract: Members of the claudin family play important roles in the formation of tight junctions in the kidneys, liver and intestine. Claudin-19 (Cldn19), a newly identified member of this family, is highly expressed in the kidney of the mouse. To have a better understanding on mouse claudin-19 gene expression, a 0.9-kb DNA fragment containing the 5′-flanking region of the Cldn19 gene was isolated. DNA sequence comparison between the mouse and human Cldn19 promoter regions exhibited little homology. One transcription initiation site was located at 104 nucleotides upstream of the start codon (ATG) of the Cldn19 gene. The mouse claudin-19 promoter lacked typical CAAT or GC-box. Deletion constructs of the 0.9-kb DNA fragment were generated and fused to a promoterless luciferase (Luc) reporter plasmid. Transfection studies using various kidney cell lines (MDCK, mIMCD3 and HEK293) revealed that the minimal promoter fragment resided in the −39 to −108 region, which contained a number of binding sites for transcription factors including Sp1. Site-directed mutagenesis using specific oligo probes confirmed that Sp1 was crucial for Cldn19 transactivation in the three cell lines studied. Electromobility shift assay confirmed that the nuclear extracts of these cells bound to the Sp1 oligo derived from Cldn19 promoter, but not to the mutated Sp1 oligo probe. Moreover, this DNA–protein complex would be recognized by Sp1 antibody, indicating specific Sp1 binding. Collectively, our data suggest that Sp1 binds to the claudin-19 promoter region and is responsible for its expression in the kidney cell lines in vitro. [Copyright &y& Elsevier]

Emphysema is an important feature of chronic obstructive pulmonary disease (COPD). Genetic factors likely affect emphysema pathogenesis, but this question has predominantly been studied in those of European ancestry. In this study, we sought to determine genetic components of emphysema severity and characterize the potential function of the associated loci in Korean population. We performed a genome-wide association study (GWAS) on quantitative emphysema in subjects with or without COPD from two Korean COPD cohorts. We investigated the functional consequences of the loci using epigenetic annotation and gene expression data. We also compared our GWAS results with an epigenome-wide association study and previous differential gene expression analysis. In total, 548 subjects (476 [86.9%] male) including 514 COPD patients were evaluated. We identified one genome-wide significant SNP (P < 5.0 × 10–8), rs117084279, near PIBF1. We identified an additional 57 SNPs (P < 5.0 × 10–6) associated with emphysema in all subjects, and 106 SNPs (P < 5.0 × 10–6) in COPD patients. Of these candidate SNPs, 2 (rs12459249, rs11667314) near CYP2A6 were expression quantitative trait loci in lung tissue and a SNP (rs11214944) near NNMT was an expression quantitative trait locus in whole blood. Of note, rs11214944 was in linkage disequilibrium with variants in enhancer histone marks in lung tissue. Several genes near additional SNPs were identified in our previous EWAS study with nominal level of significance. We identified a novel SNP associated with quantitative emphysema on CT. Including the novel SNP, several candidate SNPs in our study may provide clues to the genetic etiology of emphysema in Asian populations. Further research and validation of the loci will help determine the genetic factors for the development of emphysema. [ABSTRACT FROM AUTHOR]

Epidemiological data have implicated the pituitary gonadotropin follicle stimulating hormone (FSH) as both a risk factor for and a protective agent against epithelial ovarian cancer. Yet, little is known about how this hormone could play such opposing roles in ovarian carcinogenesis. Complementary DNA microarrays containing 2400 named genes were used to examine FSH-induced gene expression changes in ovarian cancer (OC) and immortalized normal human ovarian surface epithelial (HOSE) cell lines. Two-way t-statistics analyses of array data identified two distinct sets of FSH-regulated genes in HOSE and in established OC cell lines established from patients (OVCA cell lines). Among the HOSE cell lines, FSH increased expression of 57% of the 312 genes and downregulated 43%. In contrast, FSH diminished expression of 92% of the 177 genes in the OVCA cell lines. All but 18 of the genes affected by FSH in HOSE cell lines were different from those altered in OVCA cell lines. Among the 18 overlapping genes, nine genes exhibited the same direction of change following FSH challenge, while the other nine showed discordance in response between HOSE and OVCA cell lines. The FSH-induced differential expression of seven out of nine genes was confirmed by real-time RT-PCR. Gene-specific antisense oligonuleotides (ODNs) were used to inhibit the expression of genes encoding GTPase activating protein (rap1GAP), neogenin, and restin in HOSE and OVCA cells. Antisense ODNs to neogenin and restin, but not an antisense ODN to rap1GAP, were effective in inhibiting OVCA cell growth, diminishing proliferating cell nuclear antigen expression, and increasing caspase 3 activities. Furthermore, the ODN to rap1GAP was further shown to be ineffective in altering migration properties of OVCA cell lines. HOSE cell proliferation was not affected by treatment with any of the antisense ODNs. In summary, gene profiling data reveal for the first time that FSH may exert different biological actions on... [ABSTRACT FROM AUTHOR]

Background: COVID-19 is a respiratory viral infection with unique features including a more chronic course and systemic disease manifestations including multiple organ involvement; and there are differences in disease severity between ethnic groups. The immunological basis for disease has not been fully characterised. Analysis of whole-blood RNA expression may provide valuable information on disease pathogenesis. Methods: We studied 45 patients with confirmed COVID-19 infection within 10 days from onset of illness and a control group of 19 asymptomatic healthy volunteers with no known exposure to COVID-19 in the previous 14 days. Relevant demographic and clinical information was collected and a blood sample was drawn from all participants for whole-blood RNA sequencing. We evaluated differentially-expressed genes in COVID-19 patients (log2 fold change ≥ 1 versus healthy controls; false-discovery rate < 0.05) and associated protein pathways and compared these to published whole-blood signatures for respiratory syncytial virus (RSV) and influenza. We developed a disease score reflecting the overall magnitude of expression of internally-validated genes and assessed the relationship between the disease score and clinical disease parameters. Results: We found 135 differentially-expressed genes in the patients with COVID-19 (median age 35 years; 82% male; 36% Chinese, 53% South Asian ethnicity). Of the 117 induced genes, 14 were found in datasets from RSV and 40 from influenza; 95 genes were unique to COVID-19. Protein pathways were mostly generic responses to viral infections, including apoptosis by P53-associated pathway, but also included some unique pathways such as viral carcinogenesis. There were no major qualitative differences in pathways between ethnic groups. The composite gene-expression score was correlated with the time from onset of symptoms and nasal swab qPCR CT values (both p < 0.01) but was not related to participant age, gender, ethnicity or the presence or absence of chest X-ray abnormalities (all p > 0.05). Conclusions: The whole-blood transcriptome of COVID-19 has overall similarity with other respiratory infections but there are some unique pathways that merit further exploration to determine clinical relevance. The approach to a disease score may be of value, but needs further validation in a population with a greater range of disease severity. [ABSTRACT FROM AUTHOR]