Archival single-cell genomics reveals persistent subclones during DCIS progression.

Ductal carcinoma in situ (DCIS) is a common precursor of invasive breast cancer. Our understanding of its genomic progression to recurrent disease remains poor, partly due to challenges associated with the genomic profiling of formalin-fixed paraffin-embedded (FFPE) materials. Here, we developed Arc-well, a high-throughput single-cell DNA-sequencing method that is compatible with FFPE materials. We validated our method by profiling 40,330 single cells from cell lines, a frozen tissue, and 27 FFPE samples from breast, lung, and prostate tumors stored for 3-31 years. Analysis of 10 patients with matched DCIS and cancers that recurred 2-16 years later show that many primary DCIS had already undergone whole-genome doubling and clonal diversification and that they shared genomic lineages with persistent subclones in the recurrences. Evolutionary analysis suggests that most DCIS cases in our cohort underwent an evolutionary bottleneck, and further identified chromosome aberrations in the persistent subclones that were associated with recurrence.

A spatially resolved single-cell genomic atlas of the adult human breast.

The adult human breast is comprised of an intricate network of epithelial ducts and lobules that are embedded in connective and adipose tissue1-3. Although most previous studies have focused on the breast epithelial system4-6, many of the non-epithelial cell types remain understudied. Here we constructed the comprehensive Human Breast Cell Atlas (HBCA) at single-cell and spatial resolution. Our single-cell transcriptomics study profiled 714,331 cells from 126 women, and 117,346 nuclei from 20 women, identifying 12 major cell types and 58 biological cell states. These data reveal abundant perivascular, endothelial and immune cell populations, and highly diverse luminal epithelial cell states. Spatial mapping using four different technologies revealed an unexpectedly rich ecosystem of tissue-resident immune cells, as well as distinct molecular differences between ductal and lobular regions. Collectively, these data provide a reference of the adult normal breast tissue for studying mammary biology and diseases such as breast cancer.

Simple oligonucleotide-based multiplexing of single-cell chromatin accessibility.

Microdroplet single-cell ATAC-seq is widely used to measure chromatin accessibility, however, highly scalable and simple sample multiplexing procedures are not available. Here, we present a transposome-assisted single nucleus barcoding approach for ATAC-seq (SNuBar-ATAC) that utilizes a single oligonucleotide adaptor for multiplexing samples during the existing tagmentation step and does not require a pre-labeling procedure. The accuracy and scalability of SNuBar-ATAC was evaluated using cell line mixture experiments. We applied SNuBar-ATAC to investigate treatment-induced chromatin accessibility dynamics by multiplexing 28 mice with lung tumors that received different combinations of chemo, radiation, and targeted immunotherapy. We also applied SNuBar-ATAC to study spatial epigenetic heterogeneity by multiplexing 32 regions from a human breast tissue. Additionally, we show that SNuBar can multiplex single cell ATAC and RNA multiomic assays in cell lines and human breast tissue samples. Our data show that SNuBar is a highly accurate, easy-to-use, and scalable system for multiplexing scATAC-seq and scATAC and RNA co-assay experiments.

Breast tumours maintain a reservoir of subclonal diversity during expansion.

Our knowledge of copy number evolution during the expansion of primary breast tumours is limited1,2. Here, to investigate this process, we developed a single-cell, single-molecule DNA-sequencing method and performed copy number analysis of 16,178 single cells from 8 human triple-negative breast cancers and 4 cell lines. The results show that breast tumours and cell lines comprise a large milieu of subclones (7-22) that are organized into a few (3-5) major superclones. Evolutionary analysis suggests that after clonal TP53 mutations, multiple loss-of-heterozygosity events and genome doubling, there was a period of transient genomic instability followed by ongoing copy number evolution during the primary tumour expansion. By subcloning single daughter cells in culture, we show that tumour cells rediversify their genomes and do not retain isogenic properties. These data show that triple-negative breast cancers continue to evolve chromosome aberrations and maintain a reservoir of subclonal diversity during primary tumour growth.

Exploration of plant metabolomics variation and absorption characteristics of water-extracted Rheum tanguticum and ethanol-extracted Rheum tanguticum by UHPLC-Q-TOF-MS/MS.

BACKGROUND AND OBJECTIVE: The herb Rheum tanguticum (RT), a member of the Polygonaceae family, is listed in the Chinese Pharmacopoeia and has been widely used to treat cardiovascular and gastrointestinal disease. The research aimed to identify the different substances from two kinds of RT extraction methods and the in vivo biotransformation of RT components. METHODS: In this study, by using ultrahigh-performance liquid chromatography coupled with quadrupole-time-of-flight tandem mass spectrometry (UHPLC-Q-TOF-MS/MS), we have investigated the metabolomic variation and the in vivo metabolism of RT. A post-acquisition data processing software, PeakView, was applied to an accurate qualitative analysis of the chemical components in RT. RESULTS: Through plant metabolomics analysis, 24 related, differentially expressed metabolites of RT water extract and alcohol extract were obtained. Combined with novel identification strategies and systematic in vivo metabolism analysis, a total of 101 compounds were discovered or tentatively identified in rat serum (including 15 prototype compounds and 86 metabolites). CONCLUSION: In this study, a combination of extraction methods, liquid chromatography-mass spectrometry (LC-MS) technology, and in vivo animal metabolism studies have been established for the screening, identification, and research of chemical active components of natural medicines. LC-MS analysis combined with plant metabolomics was used to study the differential metabolites between different extraction methods of RT. Based on UHPLC-Q-TOF-MS/MS technology, the composition and metabolism of rat plasma before and after RT administration were analysed in vivo, and 15 prototype components and 86 metabolites were detected.

Long Non-Coding RNA Malat1 Increases the Rescuing Effect of Quercetin on TNFα-Impaired Bone Marrow Stem Cell Osteogenesis and Ovariectomy-Induced Osteoporosis.

Osteoporosis, a common systematic bone homeostasis disorder related disease, still urgently needs innovative treatment methods. Several natural small molecules were found to be effective therapeutics in osteoporosis. In the present study, quercetin was screened out from a library of natural small molecular compounds by a dual luciferase reporter system. Quercetin was found to upregulate Wnt/ß-catenin while inhibiting NF-κB signaling activities, and thereby rescuing osteoporosis-induced tumor necrosis factor alpha (TNFα) impaired BMSCs osteogenesis. Furthermore, a putative functional lncRNA, Malat1, was shown to be a key mediator in quercetin regulated signaling activities and TNFα-impaired BMSCs osteogenesis, as mentioned above. In an ovariectomy (OVX)-induced osteoporosis mouse model, quercetin administration could significantly rescue OVX-induced bone loss and structure deterioration. Serum levels of Malat1 were also obviously rescued in the OVX model after quercetin treatment. In conclusion, our study demonstrated that quercetin could rescue TNFα-impaired BMSCs osteogenesis in vitro and osteoporosis-induced bone loss in vivo, in a Malat1-dependent manner, suggesting that quercetin may serve as a therapeutic candidate for osteoporosis treatment.

Cranial Bone Transport Promotes Angiogenesis, Neurogenesis, and Modulates Meningeal Lymphatic Function in Middle Cerebral Artery Occlusion Rats.

BACKGROUND: Ischemic stroke is a leading cause of death and disability worldwide. However, the time window for quickly dissolving clots and restoring cerebral blood flow, using tissue-type plasminogen activator treatment is rather limited, resulting in many patients experiencing long-term functional impairments if not death. This study aims to determine the roles of cranial bone transport (CBT), a novel, effective, and simple surgical technique, in the recovery of ischemic stroke using middle cerebral artery occlusion (MCAO) rat model. METHODS: CBT was performed by slowly sliding a bone segment in skull with a special frame and a speed of 0.25 mm/12 hours for 10 days following MCAO. Morris water maze, rotarod test, and catwalk gait analysis were used to study the neurological behaviors, and infarct area and cerebral flow were evaluated during CBT process. Immunofluorescence staining of CD31 and Nestin/Sox2 (sex determining region Y box 2) was performed to study the angiogenesis and neurogenesis. OVA-A647 (ovalbumin-Alexa Fluor 647) was intracisterna magna injected to evaluate the meningeal lymphatic drainage function. RESULTS: CBT treatment has significantly reduced the ischemic lesions areas and improved the neurological deficits in MCAO rats compared with the rats in the control groups. CBT treatment significantly promoted angiogenesis and neurogenesis in the brain of MCAO rats. The drainage function of meningeal lymphatic vessels in MCAO rats was significantly impaired compared with normal rats. Ablation of meningeal lymphatic drainage led to increased neuroinflammation and aggravated neurological deficits and ischemic injury in MCAO rats. CBT treatment significantly improved the meningeal lymphatic drainage function and alleviated T-cell infiltration in MCAO rats. CONCLUSIONS: This study provided evidence for the possible mechanisms on how CBT attenuates ischemic stroke injury and facilitates rapid neuronal function recovery, suggesting that CBT may be an alternative treatment strategy for managing ischemic stroke.

Water-based Fe3O4 magnetic fluid-coated Aspergillus niger spores for treating liquid contaminated with Cr(VI).

The use of magnetic biosorbents for the remediation of heavy metals has attracted increasing attention due to their ease of separation and reusability. We developed a method for preparing superparamagnetic biosorbent materials using water-based magnetic fluids. Water-based magnetic fluid-spores (WMFSs) were obtained by combining water-based magnetic fluid (WMF) with Aspergillus niger spores at ratios of 0.6:1 (WMFS1), 0.8:1 (WMFS2), 1:1 (WMFS3), 1.2:1 (WMFS4), and 1.4:1 (WMFS5). A magnetic composite material was prepared from magnetic nanoparticles and spores in a ratio of 1:1 as a control. The adsorption efficiency and separation effect of WMFS3 were significantly better than those of the magnetic composite material. The morphology and structure of WMFS3 were characterized by performing transmission electron microscopy. The results showed that Fe3O4 magnetic particles were uniformly coated on the spore surface. The superparamagnetism of WMFS3 was tested using a vibrating sample magnetometer. At pH 2.0, the maximum adsorption capacity of WMFS3 for Cr(VI) was 105 mg/g; in the pH range of 2.0-3.0, the adsorption equilibrium time of WMFS3 was 60 min. Thus, the adsorption process conformed to the pseudo-second-order kinetic model and Freundlich isotherm. Thermodynamic studies showed that the process was spontaneous and endothermic. The adsorption mechanisms of WMF3 for Cr(VI) included electrostatic, reduction, and complexation adsorption. This biosorbent material showed excellent adsorption performance for Cr(VI) and is promising for wastewater resource applications.

Astrocytes in the rostral ventromedial medulla contribute to the maintenance of oro-facial hyperalgesia induced by late removal of dental occlusal interference.

BACKGROUND: Astrocytes in the rostral ventromedial medulla (RVM) contribute to descending pain modulation, but their role in oro-facial pain induced by persistent experimental dental occlusal interference (PEOI) or following EOI removal (REOI) is unknown. OBJECTIVE: To explore the involvement of RVM astrocytes in PEOI-induced oro-facial hyperalgesia or its maintenance following REOI. METHODS: Male rats were randomly assigned into five groups: sham-EOI, postoperative day 6 and 14 of PEOI (PEOI 6 d and PEOI 14 d), postoperative day 6 following REOI on day 3 (REOI 3 d) and postoperative day 14 following REOI on day 8 (REOI 8 d). The nociceptive head withdrawal threshold (HWT) and activities of RVM ON- or OFF-cells were recorded before and after intra-RVM astrocyte gap junction blocker carbenoxolone (CBX) microinjection. RVM astrocytes were labelled immunohistochemically with glial fibrillary acidic protein (GFAP) and analysed semi-quantitatively. RESULTS: Persistent experimental dental occlusal interference-induced oro-facial hyperalgesia, as reflected in decreased HWTs, was partially inhibited by REOI at day 3 but not at day 8 after EOI placement. Increased GFAP-staining area occurred only in REOI 8 d group in which CBX could inhibit the maintained hyperalgesia; CBX was ineffective in inhibiting hyperalgesia in PEOI 14 d group. OFF-cell activities showed no change, but the spontaneous activity and responses of ON-cells were significantly enhanced that could be suppressed by CBX in REOI 8 d group. CONCLUSION: Rostral ventromedial medulla astrocytes may not participate in PEOI-induced oro-facial hyperalgesia or hyperalgesia inhibition by early REOI but are involved in the maintenance of oro-facial hyperalgesia by late REOI.

Local administration of allogeneic or autologous bone marrow-derived mesenchymal stromal cells enhances bone formation similarly in distraction osteogenesis.

BACKGROUND AIMS: Distraction osteogenesis (DO) is a surgical technique to promote bone regeneration that requires a long time for bone healing. Bone marrow-derived mesenchymal stromal cells (MSCs) have been applied to accelerate bone formation in DO. Allogeneic MSCs are attractive, as they could be ready to use in clinics. Whether allogeneic MSCs would have an effect similar to autologous MSCs with regard to promoting bone formation in DO is still unknown. This study compares the effect of autologous MSCs versus allogeneic MSCs on bone formation in a rat DO model. METHODS: Rat bone marrow-derived MSCs were isolated, characterized and expanded in vitro. Adult rats were subjected to right tibia transverse osteotomy. On the third day of distraction, each rat received one injection of phosphate-buffered saline (PBS), autologous MSCs or allogeneic MSCs at the distraction site. Tibiae were harvested after 28 days of consolidation for micro-computed tomography examination, mechanical test and histological analysis. RESULTS: Results showed that treatment with both allogeneic and autologous MSCs promoted bone formation, with significantly higher bone mass, mechanical properties and mineral apposition rate as well as expression of angiogenic and bone formation markers at the regeneration sites compared with the PBS-treated group. No statistical difference in bone formation was found between the allogeneic and autologous MSC treatment groups. CONCLUSIONS: This study indicates that allogeneic and autologous MSCs have a similar effect on promoting bone consolidation in DO. MSCs from an allogeneic source could be used off-the-shelf with DO to achieve early bone healing.

A novel p.Pro871Leu missense mutation in SPECC1L gene causing craniosynostosis in a patient.

INTRODUCTION: Craniosynostosis is one of the most common craniofacial abnormalities. It involves premature closure of one or more cranial sutures. Mutations in many genes have been and continue to be identified in patients. SETTINGS AND SAMPLE POPULATION: Whole blood samples were collected from the patient and family members. MATERIAL AND METHODS: Whole exome sequencing was performed to identify potential mutations in the patient. The results were verified by Sanger sequencing by comparing SPECC1L gene sequence of blood samples from 100 unrelated population-matched controls. RESULTS: The patient presented with craniosynostosis with fusion of the bicoronal and sagittal sutures. A novel missense mutation (c.2612C>T, p.Pro871Leu) in the SPECC1L gene was identified. Gene analysis showed a missense mutation in exon1 of SPECC1L that led to an amino acid substitution in the region between coiled-coil domain 3 and calponin homology domain. CONCLUSION: Our observations expand the molecular spectrum of gene mutations in craniosynostosis and emphasize the importance of gene testing in the diagnosis of craniosynostosis. The observations also reinforce the characteristics of SPECC1L-related cranial disorders.

17ß-Estradiol Exacerbated Experimental Occlusal Interference-Induced Chronic Masseter Hyperalgesia by Increasing the Neuronal Excitability and TRPV1 Function of Trigeminal Ganglion in Ovariectomized Rats.

Pain symptoms in temporomandibular disorders (TMD) predominantly affect reproductive women, suggesting that estrogen regulates pain perception. However, how estrogen contributes to chronic TMD pain remains largely unclear. In the present study, we performed behavioral tests, electrophysiology, Western blot and immunofluorescence to investigate the role and underlying mechanisms of estrogen in dental experimental occlusal interference (EOI)-induced chronic masseter mechanical hyperalgesia in rats. We found that long-term 17ß-estradiol (E2) replacement exacerbated EOI-induced masseter hyperalgesia in a dose-dependent manner in ovariectomized (OVX) rats. Whole-cell patch-clamp recordings demonstrated that E2 (100 nM) treatment enhanced the excitability of isolated trigeminal ganglion (TG) neurons in OVX and OVX EOI rats, and EOI increased the functional expression of transient receptor potential vanilloid-1 (TRPV1). In addition, E2 replacement upregulated the protein expression of TRPV1 in EOI-treated OVX rats. Importantly, intraganglionic administration of the TRPV1 antagonist AMG-9810 strongly attenuated the facilitatory effect of E2 on EOI-induced masseter mechanical sensitivity. These results demonstrate that E2 exacerbated EOI-induced chronic masseter mechanical hyperalgesia by increasing TG neuronal excitability and TRPV1 function. Our study helps to elucidate the E2 actions in chronic myogenic TMD pain and may provide new therapeutic targets for relieving estrogen-sensitive pain.

Comparative Transcriptomic and Proteomic Analyses Prove that IFN-λ1 is a More Potent Inducer of ISGs than IFN-α against Porcine Epidemic Diarrhea Virus in Porcine Intestinal Epithelial Cells.

Type III interferon (IFN-λ) is currently considered to be largely nonredundant to type I interferon (IFN-α) in antivirus infection, especially in epithelial cells. Previous studies reported that, compared with IFN-α, IFN-λ exhibited stronger induction of interferon-stimulated genes (ISGs) at the transcriptional level in intestinal epithelial cells and stronger inhibition of porcine epidemic diarrhea virus (PEDV). In this study, the different mechanisms of ISG upregulation induced by IFN-α and IFN-λ1 were compared at the mRNA and protein levels in the porcine intestinal epithelial cell model (IPEC-J2). It was proved that IFN-λ1 consistently exhibited stronger stimulation effects at both levels. At the mRNA level, 132 genes were significantly upregulated upon IFN-λ1 stimulation, while 42 genes upon IFN-α stimulation. At the protein level, 47 proteins were significantly upregulated upon IFN-λ1 stimulation, but only 8 proteins were upregulated upon IFN-α stimulation. The shared upregulated genes/proteins by IFN-λ1 in both transcriptional and translational omics, especially the regulation factors of ISG15, were involved in the JAK-STAT signaling pathway. Compared to IFN-α, IFN-λ1 could induce more consistent upregulation of the key ISGs (ISG15, USP18, OASL, and RSAD2) at 3-24 h postinduction as measured by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) validation. It was further confirmed through functional analysis that ISG15 and RSAD2 could inhibit PEDV infection in dose-dependent manners. This study provided solid evidence that IFN-λ1 could induce a more unique and higher ISG expression level, which exhibited anti-PEDV effects on porcine intestinal epithelial cells.

CUDC-907, a novel dual PI3K and HDAC inhibitor, in prostate cancer: Antitumour activity and molecular mechanism of action.

Targeting the androgen receptor (AR) signalling pathway remains the main therapeutic option for advanced prostate cancer. However, resistance to AR-targeting inhibitors represents a great challenge, highlighting the need for new therapies. Activation of the PI3K/AKT pathway and increased expression of histone deacetylases (HDACs) are common aberrations in prostate cancer, suggesting that inhibition of such targets may be a viable therapeutic strategy for this patient population. Previous reports demonstrated that combination of PI3K inhibitors (PI3KIs) with histone deacetylase inhibitors (HDACIs) resulted in synergistic antitumour activities against preclinical models of prostate cancer. In this study, we demonstrate that the novel dual PI3K and HDAC inhibitor CUDC-907 has promising antitumour activity against prostate cancer cell lines in vitro and castration-resistant LuCaP 35CR patient-derived xenograft (PDX) mouse model in vivo. CUDC-907-induced apoptosis was partially dependent on Mcl-1, Bcl-xL, Bim and c-Myc. Further, down-regulation of Wee1, CHK1, RRM1 and RRM2 contributed to CUDC-907-induced DNA damage and apoptosis. In the LuCaP 35CR PDX model, treatment with CUDC-907 resulted in significant inhibition of tumour growth. These findings support the clinical development of CUDC-907 for the treatment of prostate cancer.

Comparative Analysis of Gammaherpesvirus Circular RNA Repertoires: Conserved and Unique Viral Circular RNAs.

Recent studies have identified circular RNAs (circRNAs) expressed from the Epstein-Barr virus (EBV) and Kaposi's sarcoma herpesvirus (KSHV) human DNA tumor viruses. To gain initial insights into the potential relevance of EBV circRNAs in virus biology and disease, we assessed the circRNAome of the interspecies homologue rhesus macaque lymphocryptovirus (rLCV) in a naturally occurring lymphoma from a simian immunodeficiency virus (SIV)-infected rhesus macaque. This analysis revealed rLCV orthologues of the latency-associated EBV circular RNAs circRPMS1_E4_E3a and circEBNA_U. Also identified in two samples displaying unusually high lytic gene expression was a novel rLCV circRNA that contains both conserved and rLCV-specific RPMS1 exons and whose backsplice junctions flank an rLCV lytic origin of replication (OriLyt). Analysis of a lytic infection model for the murid herpesvirus 68 (MHV68) rhadinovirus identified a cluster of circRNAs near an MHV68 lytic origin of replication, with the most abundant of these, circM11_ORF69, spanning the OriLyt. Lastly, analysis of KSHV latency and reactivation models revealed the latency associated circRNA originating from the vIRF4 gene as the predominant viral circRNA. Together, the results of this study broaden our appreciation for circRNA repertoires in the Lymphocryptovirus and Rhadinovirus genera of gammaherpesviruses and provide evolutionary support for viral circRNA functions in latency and viral replication.IMPORTANCE Infection with oncogenic gammaherpesviruses leads to long-term viral persistence through a dynamic interplay between the virus and the host immune system. Critical for remodeling of the host cell environment after the immune responses are viral noncoding RNAs that modulate host signaling pathways without attracting adaptive immune recognition. Despite the importance of noncoding RNAs in persistent infection, the circRNA class of noncoding RNAs has only recently been identified in gammaherpesviruses. Accordingly, their roles in virus infection and associated oncogenesis are unknown. Here we report evolutionary conservation of EBV-encoded circRNAs determined by assessing the circRNAome in rLCV-infected lymphomas from an SIV-infected rhesus macaque, and we report latent and lytic circRNAs from KSHV and MHV68. These experiments demonstrate utilization of the circular RNA class of RNAs across 4 members of the gammaherpesvirus subfamily, and they identify orthologues and potential homoplastic circRNAs, implying conserved circRNA functions in virus biology and associated malignancies.

Raddeanin A down-regulates androgen receptor and its splice variants in prostate cancer.

Castration-resistant progression of prostate cancer is a major cause of prostate cancer mortality, and increased expression and activity of the full-length and the splice variants of androgen receptor (AR) have been indicated to drive castration resistance. Consequently, there is an urgent need to develop agents that can target both the full-length and the splice variants of AR for more effective treatment of prostate cancer. In the present study, we showed that raddeanin A (RA), an oleanane-type triterpenoid saponin, suppresses the transcriptional activities of both the full-length and the splice variants of AR. This is attributable to their decreased expression as a result of RA induction of proteasome-mediated degradation and inhibition of the transcription of the AR gene. We further showed the potential of using RA to enhance the growth inhibitory efficacy of docetaxel, the first-line chemotherapy for prostate cancer. This study identifies RA as a new agent to target both the full-length and the splice variants of AR and provides a rationale for further developing RA for prostate cancer treatment.

Novel self-immobilized biomass mixture based on mycelium pellets for wastewater treatment: A review.

Mycelial pellets, as a novel biomass material, can adsorb pollutants as a biosorbent, or combine other substances and organisms to form self-immobilized biomixture (SIB) to remove pollutants from wastewater. The pellets are eco-friendly, have a good self-immobilization capacity, and are easy to filter. In addition, some mycelial fungi can remove the pollutants in water through biodegradation. This study reviewed biomixture based on mycelial pellets and the two ways, through which SIB remove pollutants in water: pure pellets and the pellets with other materials. The characteristics and functions of each part of SIB were discussed. The study also highlighted the shortcomings of the technology and provided recommendations for further development of this technology.

Recombinant mouse periostin ameliorates coronal sutures fusion in Twist1+/- mice.

BACKGROUND: Saethre-Chotzen syndrome is an autosomal dominantly inherited disorder caused by mutations in the twist family basic helix-loop-helix transcription factor 1 (TWIST1) gene. Surgical procedures are frequently required to reduce morphological and functional defects in patients with Saethre-Chotzen syndrome. Therefore, the development of noninvasive procedures to treat Saethre-Chotzen syndrome is critical. We identified that periostin, which is an extracellular matrix protein that plays an important role in both bone and connective tissues, is downregulated in craniosynostosis patients. METHODS: We aimed to verify the effects of different concentrations (0, 50, 100, and 200 µg/l) of recombinant mouse periostin in Twist1+/- mice (a mouse model of Saethre-Chotzen syndrome) coronal suture cells in vitro and in vivo. Cell proliferation, migration, and osteogenic differentiation were observed and detected. Twist1+/- mice were also injected with recombinant mouse periostin to verify the treatment effects. RESULTS: Cell Counting Kit-8 results showed that recombinant mouse periostin inhibited the proliferation of suture-derived cells in a time- and concentration-dependent manner. Cell migration was also suppressed when treated with recombinant mouse periostin. Real-time quantitative PCR and Western blotting results suggested that messenger ribonucleic acid and protein expression of alkaline phosphatase, bone sialoprotein, collagen type I, and osteocalcin were all downregulated after treatment with recombinant mouse periostin. However, the expression of Wnt-3a, Wnt-1, and ß-catenin were upregulated. The in vivo results demonstrated that periostin-treated Twist1+/- mice showed patent coronal sutures in comparison with non-treated Twist1+/- mice which have coronal craniosynostosis. CONCLUSION: Our results suggest that recombinant mouse periostin can inhibit coronal suture cell proliferation and migration and suppress osteogenic differentiation of suture-derived cells via Wnt canonical signaling, as well as ameliorate coronal suture fusion in Twist1+/- mice.

AKT isoform-specific expression and activation across cancer lineages.

BACKGROUND: Aberrant AKT activation is prevalent across human cancer lineages, providing an important therapeutic target. AKT comprises three isoforms that mediate critical non-redundant, even opposing functions in cancer pathophysiology. Therefore, targeting specific AKT isoforms in particular cancers may be more effective than pan-AKT inhibition while avoiding disadvantages of pan-AKT inhibition. Currently, AKT isoform-specific expression and activation in cancer are not clearly characterized. METHODS: We systematically characterized AKT isoform-specific expression and activation in 211 cancer cell lines derived from different lineages and genetic backgrounds using a reverse-phase protein array platform. RESULTS: We found that phosphorylation, but not expression, of AKT1 and AKT2 was coordinated in most but not all cells. Different cancer lineages displayed differential AKT1 and AKT2 expression and phosphorylation. A PIK3CA hotspot mutation H1047R but not E545K was associated with selective activation of AKT2 but not AKT1. CONCLUSIONS: Our study identified and validated AKT isoform-specific expression and phosphorylation in certain cell lines and demonstrated that genetic changes can affect AKT isoform-specific activation. These results provide a more precise understanding of AKT isoform-specific signaling and, in addition, facilitate AKT isoform targeting for personalized cancer therapies.

Impact of tumor architecture on disease recurrence and cancer-specific mortality of upper tract urothelial carcinoma treated with radical nephroureterectomy.

Upper tract urinary carcinoma (UTUC) is a relatively uncommon but aggressive disease. Recent publications have assessed the prognostic significance of tumor architecture in UTUC, but there is still controversy regarding the significance and importance of tumor architecture on disease recurrence. We retrospectively reviewed the medical records of 101 patients with clinical UTUC who had undergone surgery. Univariate and multivariate analyses were conducted to identify factors associated with disease recurrence and cancer-specific mortality. As our single center study and the limited sample size may influence the clinical significance, we further quantitatively combined the results with those of existing published literature through a meta-analysis compiled from searching several databases. At a median follow-up of 41.3 months, 25 patients experienced disease recurrence. Spearman's correlation analysis showed that tumor architecture was found to be positively correlated with the tumor location and the histological grade. Kaplan-Meier curves showed that patients with sessile tumor architecture had significantly poor recurrence free survival (RFS) and cancer specific survival (CSS). Furthermore, multivariate analysis suggested that tumor architecture was independent prognostic factors for RFS (Hazard ratio, HR = 2.648) and CSS (HR = 2.072) in UTUC patients. A meta-analysis of investigating tumor architecture and its effects on UTUC prognosis was conducted. After searching PubMed, Medline, Embase, Cochrane Library and Scopus databases, 17 articles met the eligibility criteria for this analysis. The eligible studies included a total of 14,368 patients and combined results showed that sessile tumor architecture was associated with both disease recurrence with a pooled HR estimate of 1.454 and cancer-specific mortality with a pooled HR estimate of 1.416. Tumor architecture is an independent predictor for disease recurrence after radical nephroureterectomy for UTUC. Therefore, closer surveillance is necessary, especially in patients with sessile tumor architecture.