Plasmalogen, a glycerophospholipid crucial for Streptococcus mutans acid tolerance and colonization.

Plasmalogen is a specific glycerophospholipid present in both animal and bacterial organisms. It plays a crucial function in eukaryotic cellular processes and is closely related to several human diseases, including neurological disorders and cancers. Nonetheless, the precise biological role of plasmalogen in bacteria is not well understood. In this study, we identified SMU_438c as the enzyme responsible for plasmalogen production in Streptococcus mutans under anaerobic conditions. The heterologous expression of SMU_438c in a plasmalogen-negative strain, Streptococcus sanguinis, resulted in the production of plasmalogen, indicating that this enzyme is sufficient for plasmalogen production. Additionally, the plasmalogen-deficient S. mutans exhibited significantly lower acid tolerance and diminished its colonization in Drosophila flies compared to the wild-type strain and complemented strain. In summary, our data suggest that plasmalogen plays a vital role in bacterial stress tolerance and in vivo colonization. IMPORTANCE: This study sheds light on the biological role of plasmalogen, a specific glycerophospholipid, in bacteria, particularly in Streptococcus mutans. Plasmalogens are known for their significant roles in eukaryotic cells and have been linked to human diseases like neurological disorders and cancers. The enzyme SMU_438c, identified as essential for plasmalogen production under anaerobic conditions, was crucial for acid tolerance and in vivo colonization in Drosophila by S. mutans, underscoring its importance in bacterial stress response and colonization. These findings bridge the knowledge gap in bacterial physiology, highlighting plasmalogen's role in microbial survival and offering potential insights into microbial pathogenesis and host-microbe interactions.

Cardiac-Specific Expression of Cre Recombinase Leads to Age-Related Cardiac Dysfunction Associated with Tumor-like Growth of Atrial Cardiomyocyte and Ventricular Fibrosis and Ferroptosis.

Transgenic expression of Cre recombinase driven by a specific promoter is normally used to conditionally knockout a gene in a tissue- or cell-type-specific manner. In αMHC-Cre transgenic mouse model, expression of Cre recombinase is controlled by the myocardial-specific α-myosin heavy chain (αMHC) promoter, which is commonly used to edit myocardial-specific genes. Toxic effects of Cre expression have been reported, including intro-chromosome rearrangements, micronuclei formation and other forms of DNA damage, and cardiomyopathy was observed in cardiac-specific Cre transgenic mice. However, mechanisms associated with Cardiotoxicity of Cre remain poorly understood. In our study, our data unveiled that αMHC-Cre mice developed arrhythmias and died after six months progressively, and none of them survived more than one year. Histopathological examination showed that αMHC-Cre mice had aberrant proliferation of tumor-like tissue in the atrial chamber extended from and vacuolation of ventricular myocytes. Furthermore, the αMHC-Cre mice developed severe cardiac interstitial and perivascular fibrosis, accompanied by significant increase of expression levels of MMP-2 and MMP-9 in the cardiac atrium and ventricular. Moreover, cardiac-specific expression of Cre led to disintegration of the intercalated disc, along with altered proteins expression of the disc and calcium-handling abnormality. Comprehensively, we identified that the ferroptosis signaling pathway is involved in heart failure caused by cardiac-specific expression of Cre, on which oxidative stress results in cytoplasmic vacuole accumulation of lipid peroxidation on the myocardial cell membrane. Taken together, these results revealed that cardiac-specific expression of Cre recombinase can lead to atrial mesenchymal tumor-like growth in the mice, which causes cardiac dysfunction, including cardiac fibrosis, reduction of the intercalated disc and cardiomyocytes ferroptosis at the age older than six months in mice. Our study suggests that αMHC-Cre mouse models are effective in young mice, but not in old mice. Researchers need to be particularly careful when using αMHC-Cre mouse model to interpret those phenotypic impacts of gene responses. As the Cre-associated cardiac pathology matched mostly to that of the patients, the model could also be employed for investigating age-related cardiac dysfunction.

The Prognostic Significance of FKBP1A and Its Related Immune Infiltration in Liver Hepatocellular Carcinoma.

Liver hepatocellular carcinoma (LIHC) remains a global health challenge with poor prognosis and high mortality. FKBP1A was first discovered as a receptor for the immunosuppressant drug FK506 in immune cells and is critical for various tumors and cancers. However, the relationships between FKBP1A expression, cellular distribution, tumor immunity, and prognosis in LIHC remain unclear. Here, we investigated the expression level of FKBP1A and its prognostic value in LIHC via multiple datasets including ONCOMINE, TIMER, GEPIA, UALCAN, HCCDB, Kaplan-Meier plotter, LinkedOmics, and STRING. Human liver tissue microarray was employed to analyze the characteristics of FKBP1A protein including the expression level and pathological alteration in cellular distribution. FKBP1A expression was significantly higher in LIHC and correlated with tumor stage, grade and metastasis. The expression level of the FKBP1A protein was also increased in LIHC patients along with its accumulation in endoplasmic reticulum (ER). High FKBP1A expression was correlated with a poor survival rate in LIHC patients. The analysis of gene co-expression and the regulatory pathway network suggested that FKBP1A is mainly involved in protein synthesis, metabolism and the immune-related pathway. FKBP1A expression had a significantly positive association with the infiltration of hematopoietic immune cells including B cells, CD8+ T cells, CD4+ T cells, macrophages, neutrophils, and dendritic cells. Moreover, M2 macrophage infiltration was especially associated with a poor survival prognosis in LIHC. Furthermore, FKBP1A expression was significantly positively correlated with the expression of markers of M2 macrophages and immune checkpoint proteins such as PD-L1, CTLA-4, LAG3 and HAVCR2. Our study demonstrated that FKBP1A could be a potential prognostic target involved in tumor immune cell infiltration in LIHC.

Intraspecies interactions of Streptococcus mutans impact biofilm architecture and virulence determinants in childhood dental caries.

Early childhood dental caries (ECC) is the most common chronic disease among children, especially among low socioeconomic populations. Streptococcus mutans is most frequently associated with initiation of ECC. Although many studies report children with multiple S. mutans strains (i.e., genotypes) have greater odds of developing ECC, studies investigating intraspecies interactions in dental caries are lacking. This study investigates the impact of intraspecies interactions on cariogenic and fitness traits of clinical S. mutans isolates using in vitro and in vivo approaches. Association analysis evaluated if presence of multiple S. mutans genotypes within the first year of colonization was associated with caries. Initially, clinical S. mutans isolates from 10 children were evaluated. S. mutans strains (G09 and G18, most prevalent) isolated from one child were used for subsequent analysis. Biofilm analysis was performed for single and mixed cultures to assess cariogenic traits, including biofilm biomass, intra-polysaccharide, pH, and glucan. Confocal laser scanning microscopy (CLSM) and time-lapse imaging were used to evaluate spatial and temporal biofilm dynamics, respectively. A Drosophila model was used to assess colonization in vivo. Results showed the mean biofilm pH was significantly lower in co-cultured biofilms versus monoculture. Doubling of S. mutans biofilms was observed by CLSM and in vivo colonization in Drosophila for co-cultured S. mutans. Individual strains occupied specific domains in co-culture and G09 contributed most to increased co-culture biofilm thickness and colonization in Drosophila. Biofilm formation and acid production displayed distinct signatures in time-lapsed experiments. This study illuminates that intraspecies interactions of S. mutans significantly impacts biofilm acidity, architecture, and colonization.IMPORTANCEThis study sheds light on the complex dynamics of a key contributor to early childhood dental caries (ECC) by exploring intraspecies interactions of different S. mutans strains and their impact on cariogenic traits. Utilizing clinical isolates from children with ECC, the research highlights significant differences in biofilm architecture and acid production in mixed versus single genotype cultures. The findings reveal that co-cultured S. mutans strains exhibit increased cell density and acidity, with individual strains occupying distinct domains. These insights, enhanced by use of time-lapsed confocal laser scanning microscopy and a Drosophila model, offer a deeper understanding of ECC pathogenesis and potential avenues for targeted interventions.

Potential Oral Health Benefits of Ginseng and Its Extracts.

This review discusses the effects of ginseng and its extracts in the treatment of dental caries, periodontal diseases, endodontic diseases, oral cancers, oral mucosal diseases, and some other dental associations. In the meantime, bioavailability and safety application of ginseng products are discussed. All of the articles reviewed were from PubMed, Embase, Cochrane Library, Web of Science, China National Knowledge Infrastructure, Wanfang Data, and VIP Chinese Science and Technology Periodicals Full-Text Database through November 2022, including full-text English or non-English publications. Ginseng and its extracts were shown to have beneficial effects on oral diseases, and further studies are needed to understand the mechanisms and confirm the effects in humans.

Drosophila melanogaster as a model to study polymicrobial synergy and dysbiosis.

The fruit fly Drosophila melanogaster has emerged as a valuable model for investigating human biology, including the role of the microbiome in health and disease. Historically, studies involving the infection of D. melanogaster with single microbial species have yielded critical insights into bacterial colonization and host innate immunity. However, recent evidence has underscored that multiple microbial species can interact in complex ways through physical connections, metabolic cross-feeding, or signaling exchanges, with significant implications for healthy homeostasis and the initiation, progression, and outcomes of disease. As a result, researchers have shifted their focus toward developing more robust and representative in vivo models of co-infection to probe the intricacies of polymicrobial synergy and dysbiosis. This review provides a comprehensive overview of the pioneering work and recent advances in the field, highlighting the utility of Drosophila as an alternative model for studying the multifaceted microbial interactions that occur within the oral cavity and other body sites. We will discuss the factors and mechanisms that drive microbial community dynamics, as well as their impacts on host physiology and immune responses. Furthermore, this review will delve into the emerging evidence that connects oral microbes to systemic conditions in both health and disease. As our understanding of the microbiome continues to evolve, Drosophila offers a powerful and tractable model for unraveling the complex interplay between host and microbes including oral microbes, which has far-reaching implications for human health and the development of targeted therapeutic interventions.

Intraspecies interactions of Streptococcus mutans impact biofilm architecture and virulence determinants in childhood dental caries.

Early childhood dental caries (ECC) is the most common chronic disease among children with a heavy disease burden among low socioeconomic populations. Streptococcus mutans is most frequently associated with initiation of ECC. Many studies report children with multiple S. mutans strains (i.e., genotypes) having greater odds of developing ECC, studies investigating intraspecies interactions in dental caries are lacking. In this study, the impact of intraspecies interactions on cariogenic and fitness traits of clinical S. mutans isolates are investigated using in-vitro and in-vivo approaches. Initially clinical S. mutans isolates of 10 children from a longitudinal epidemiological study were evaluated. S. mutans strains (G09 and G18, most prevalent) isolated from one child were used for subsequent analysis. Association analysis was used to determine if presence of multiple S. mutans genotypes within the first-year of colonization was associated with caries. Biofilm analysis was performed for single and mixed cultures to assess cariogenic traits, including biofilm biomass, intra-polysaccharide, pH, and glucan. Confocal Laser Scanning Microscopy (CLSM) and time-lapse imaging were used to evaluate spatial and temporal biofilm dynamics, respectively. A Drosophila model was used to assess colonization in-vivo. Mean biofilm pH was significantly lower in co-cultured biofilms as compared with monoculture biofilms. Doubling of S. mutans in-vitro biofilms was observed by CLSM and in-vivo colonization in Drosophila for co-cultured S. mutans. Individual strains occupied specific domains in co-culture and G09 contributed most to increased co-culture biofilm thickness and colonization in Drosophila. Biofilm formation and acid production displayed distinct signatures in time-lapsed experiments.

Mosaic loss of chromosome Y in peripheral blood cells is associated with age-related macular degeneration in men.

BACKGROUND: Age-related macular degeneration (AMD) is the leading cause of severe vision loss in patients over 55 years old in the industrialized world. In the past 20 years, approximately 288 million patents have been affected by this disease. Despite this high prevalence, the molecular mechanism for AMD remains unclear, and there remains no effective treatment for this disease. The mosaic loss of Y chromosome (mLOY) has been identified as a common phenomenon in multiple age-related disease (i.e., oncogenesis and cardiovascular disease) has recently been identified by genome-wide analysis to be linked to AMD as well. As the Y chromosome mainly possesses three genomic functions, sister chromatin cohesion, cell cycle mitosis, and apoptotic signaling, here we characterize the Y chromosome euchromatic genes and non-chromosome AMD genes in relevance to cellular proliferation and apoptotic signaling of leukocytes. RESULTS: Using STRING, a publically available database of all protein-protein interaction, Grassmann et al. found the genes on the Y chromosome is mainly believed to take part in three major cellular genomic functions- sister chromatin cohesion, cell cycle mitosis, and apoptotic signaling. Based on data from the Ensembl Genome database, we focus on our discussion on coding genes found in the euchromatins but not the PAR1 and PAR2 regions of the Y chromosomes. All 14 known euchromatic genes on the Y chromosome short arm and all 31 known euchromatic genes on the Y chromosome long arm (Yq) are directly or indirectly involved in the cell cycle (meiosis and mitosis) and proliferation. We sorted non-Y chromosome AMD associated genes into these three categories to identify signaling pathways that may compound with cellular dysregulation due to mLOY. Of the genes associated with AMD, complement pathway genes such as C2, C9 and CFH/ARMD4 are associated with proliferation, receptor-mediated endocytosis genes such as APOE, DAB2 and others associated with apoptotic signaling. Because nucleated cells found in peripheral circulation are mainly composed of leukocytes with reduced expression of CD99, a protein essential for leukocytes adhesion, translocation, and function, mLOY in these cells likely affect retinal degeneration through altered immunological surveillance. In fact, there is precedence that circulating macrophage can stabilize and modify the cardiac rhythm and contractility post ischemic damage. Therefore, the most likely mechanism through which peripheral mLOY affects AMD development in men is through the role affected leukocytes play in retinal proliferation and apoptosis. CONCLUSIONS: mLOY in peripheral blood is newly discovered in AMD by Grassmann et al. as it is a common phenomenon in oncogenesis and cardiac dysfunction. Here the recent data conclude the possible mechanism for the newly identified link between mLOY and AMD, and provide support that mLOY in circulating macrophage-monocyte of affected male patients promotes AMD by targeting the retina and causing macular degeneration.

Template-Free Synthesis of g-C3N4 Nanoball/BiOCl Nanotube Heterojunction with Enhanced Photocatalytic Activity.

There are many reports on g-C3N4 nanosheet and BiOCl nanosheet, but few studies on other morphologies of g-C3N4 and BiOCl. Herein, a g-C3N4 nanoball/BiOCl nanotube heterojunction prepared by a simple one-step acetonitrile solvothermal method is reported. The XRD results prove that the g-C3N4/BiOCl composites can be prepared in one step. SEM results revealed that the g-C3N4 was spherical and the BiOCl was tubular. The HRTEM results indicate that g-C3N4 has an amorphous structure and that the (100) crystal plane of BiOCl borders the g-C3N4. Spherical g-C3N4 has a narrow band gap (approximately 1.94 eV), and the band gap of g-C3N4/BiOCl after modification was also narrow. When the BiOCl accounted for 30% of the g-C3N4/BiOCl by mass, the quasi-primary reaction rate constant of RhB degradation was 45 times that of g-C3N4. This successful preparation method for optimizing g-C3N4 involving simple one-step template-free synthesis may be adopted for the preparation of diverse-shapes and high-performance nanomaterials in the future.

RACK1 promotes breast carcinoma migration/metastasis via activation of the RhoA/Rho kinase pathway.

We aimed to gain a mechanistic understanding of the role of RACK1 in breast carcinoma migration/metastasis. Migration assays were conducted in breast carcinoma cell lines. siRNA targeting RACK1 as well as the Rho kinase inhibitor were also applied. Immunoprecipitation and immunofluorescence were used to study the RACK1/RhoA interaction. GTP-Rho pull-down assays were performed to assess the activation of RhoA. We also conducted immunohistochemistry in 160 breast carcinoma samples. Experiments in vitro showed that RACK1 promotes migration via interaction with RhoA and activation of the RhoA/Rho kinase pathway. Immunohistochemistry in 160 samples revealed that RACK1 is strongly correlated with accepted tumor spread indicators and RhoA (all P < 0.05). Kaplan-Meier survival analysis indicated a correlation between higher RACK1 expression and shorter survival times (P < 0.001). RACK1 is a prognostic factor that promotes breast carcinoma migration/metastasis by interacting with RhoA and activating the RhoA/Rho kinase pathway.

Autism-Associated Variant in the SLC6A3 Gene Alters the Oral Microbiome and Metabolism in a Murine Model.

Background: Altered dopamine (DA) signaling has been associated with autism spectrum disorder (ASD), a neurodevelopmental condition estimated to impact 1 in 54 children in the United States. There is growing evidence for alterations in both gastrointestinal function and oral microbiome composition in ASD. Recent work suggests that rare variants of the SLC6A3 gene encoding the DA transporter (DAT) identified in individuals with ASD result in structural and functional changes to the DAT. One such recently identified de novo mutation is a threonine to methionine substitution at position 356 of the DAT (DAT T356M). The DAT T356M variant is associated with ASD-like phenotypes in mice homozygous for the mutation (DAT T356M+/+), including social deficits, hyperactivity, and impaired DA signaling. Here, we determine the impact of this altered DA signaling as it relates to altered oral microbiota, and metabolic and gastrointestinal dysfunction. Methods: In the DAT T356M+/+ mouse, we determine the oral microbiota composition, metabolic function, and gastrointestinal (GI) function. We examined oral microbiota by 16S RNA sequencing. We measured metabolic function by examining glucose tolerance and we probed gastrointestinal parameters by measuring fecal dimensions and weight. Results: In the DAT T356M+/+ mouse, we evaluate how altered DA signaling relates to metabolic dysfunction and altered oral microbiota. We demonstrate that male DAT T356M+/+ mice weigh less (Wild type (WT) = 26.48 ± 0.6405 g, DAT T356M+/+ = 24.14 ± 0.4083 g) and have decreased body fat (WT = 14.89 ± 0.6206%, DAT T356M+/+ = 12.72 ± 0.4160%). These mice display improved glucose handling (WT = 32.60 ± 0.3298 kcal/g, DAT T356M+/+ = 36.97 ± 0.4910 kcal/g), and an altered oral microbiota. We found a significant decrease in Fusobacterium abundance. The abundance of Fusobacterium was associated with improved glucose handling and decreased body fat. Conclusions: Our findings provide new insights into how DAT dysfunction may alter gastrointestinal function, composition of the oral microbiota, and metabolism. Our data suggest that impaired DA signaling in ASD is associated with a number of metabolic and gastrointestinal changes which are common in individuals with ASD.

Therapeutic Targeting of Nemo-like Kinase in Primary and Acquired Endocrine-resistant Breast Cancer.

PURPOSE: Endocrine resistance remains a major clinical challenge in estrogen receptor (ER)-positive breast cancer. Despite the encouraging results from clinical trials for the drugs targeting known survival signaling, relapse is still inevitable. There is an unmet need to discover new drug targets in the unknown escape pathways. Here, we report Nemo-like kinase (NLK) as a new actionable kinase target that endows previously uncharacterized survival signaling in endocrine-resistant breast cancer. EXPERIMENTAL DESIGN: The effects of NLK inhibition on the viability of endocrine-resistant breast cancer cell lines were examined by MTS assay. The effect of VX-702 on NLK activity was verified by kinase assay. The modulation of ER and its coactivator, SRC-3, by NLK was examined by immunoprecipitation, kinase assay, luciferase assay, and RNA sequencing. The therapeutic effects of VX-702 and everolimus were tested on cell line- and patient-derived xenograft (PDX) tumor models. RESULTS: NLK overexpression endows reduced endocrine responsiveness and is associated with worse outcome of patients treated with tamoxifen. Mechanistically, NLK may function, at least in part, via enhancing the phosphorylation of ERα and its key coactivator, SRC-3, to modulate ERα transcriptional activity. Through interrogation of a kinase profiling database, we uncovered and verified a highly selective dual p38/NLK inhibitor, VX-702. Coadministration of VX-702 with the mTOR inhibitor, everolimus, demonstrated a significant therapeutic effect in cell line-derived xenograft and PDX tumor models of acquired or de novo endocrine resistance. CONCLUSIONS: Together, this study reveals the potential of therapeutic modulation of NLK for the management of the endocrine-resistant breast cancers with active NLK signaling.

RACK1: A superior independent predictor for poor clinical outcome in breast cancer.

We aimed to investigate the expression of RACK1 in breast cancer, evaluate its role in predicting prognosis and compare with commonly used biomarkers: Ki67, ER, PR and HER-2 for patients with breast cancer. The RACK1 expression and its clinical significance were examined in 160 breast carcinoma patients using immunohistochemistry. Correlations of RACK1 expression with other commonly used biomarkers and survival analyses were assessed. Immunohistochemistry results showed that the number of RACK1 cases scoring 0, 1, and 2 were 66, 54, and 40, respectively. RACK1 staining was strongly related to clinical stage, histological grade, Ki67, ER, PR and HER-2 (all p < 0.05). Consistently, all of the cases exhibiting RACK1 staining score 0 were survivors, whereas the majority (55.0%) of those exhibiting RACK1 staining score 2 were deaths. Kaplan-Meier survival analysis of 160 cases revealed a correlation between higher RACK1 expression levels and shorter overall survival times (p < 0.001). Univariate and multivariate analyses revealed that RACK1, tumor size, lymph node metastasis, and HER-2 were independent prognostic factors (all p < 0.05). Interestingly, receiver operator characteristic (ROC) curves showed that the ROC areas for RACK1, Ki67, ER, PR and HER-2 were 0.833, 0.766, 0.446, 0.387, and 0.689, respectively, and the superiority of RACK1 in sensitivity and specificity as biomarker was demonstrated. To our knowledge, it is the first time to investigate the expression of RACK1, and identified that RACK1 is a superior independent biomarker for diagnosis and prognosis comparing with currently widely used diagnostic index in breast carcinoma.

Overexpression and involvement of special AT-rich sequence binding protein 1 in multidrug resistance in human breast carcinoma cells.

Special AT-rich sequence binding protein (SATB) 1 has been proposed to act as a determinant for the acquisition of metastatic activity by controlling expression of a specific set of genes that promote metastatic activity. Here we found that SATB1 expression is upregulated in multidrug-resistant breast cancer cells that exhibit higher invasive potential than the parental cells. Apart from accelerating metastasis and inducing epithelial-mesenchymal transition, SATB1 was demonstrated to confer resistance to both P-glycoprotein-related and P-glycoprotein-non-related drugs on MCF7 cells, which was accompanied by decreasing accumulation of adriamycin in SATB1-overexpressing transfectants. SATB1 depletion could partially reverse the multidrug resistance (MDR) phenotype of MCF7/ADR in vitro and in vivo. The SATB1-induced P-glycoprotein-mediated MDR could be reversed by treatment with anti-P-glycoprotein mAb. Moreover, SATB1 plays an important role in anti-apoptotic activity in MCF7/ADR cells in response to adriamycin treatment, which suggests another mechanism contributing to SATB1-related MDR of breast cancers. These data provide new insights into the mode by which breast tumors acquire the MDR phenotype and also imply a role for SATB1 in this process.

RACK1 promotes breast carcinoma proliferation and invasion/metastasis in vitro and in vivo.

A yeast two-hybrid system was utilized to identify novel PI3K p110alpha-interacting proteins, of which receptor of activated protein kinase C1 (RACK1) was chosen for successive detailed analyses. Our aim was to investigate the function(s) of RACK1 and its involvement in mechanisms of breast carcinoma proliferation and invasion/metastasis. Experiments in breast carcinoma cell lines stably transfected with RACK1, as well as nude mouse models, showed that RACK1 promotes breast carcinoma proliferation and invasion/metastasis in vitro and in vivo. Conversely, knockdown of RACK1 by siRNA in vitro inhibited proliferation, migration, and invasion. In cell lines stably transfected with RACK1, p-AKT, cyclin D1, cyclin D3, and CD147 expression, as well as MMP2 activity, were elevated. RACK1-induced migration could be inhibited by the addition of Rho-kinase inhibitor. In 160 breast carcinoma cases, survival analyses established that RACK1 is an independent prognostic factor for poor outcome (P < 0.001). In conclusion, RACK1 is an independent prognosis-related factor and promotes breast carcinoma proliferation and invasion/metastasis in vitro and in vivo.

Twist1-mediated adriamycin-induced epithelial-mesenchymal transition relates to multidrug resistance and invasive potential in breast cancer cells.

PURPOSE: Besides its therapeutic effects, chemotherapeutic agents also enhance the malignancy of treated cancers in clinical situations. Recently, epithelial-mesenchymal transition (EMT) has attracted attention in studies of tumor progression. We aimed to test whether transient Adriamycin treatment induces EMT and apoptosis simultaneously in cancer cells, clarify why the same type of cells responds differentially (i.e., apoptosis, EMT) to Adriamycin treatment, and elucidate the role of Twist1, the master regulator of EMT, in this process. EXPERIMENTAL DESIGN: In unsynchronized MCF7 cells or cells synchronized at different phases, apoptosis, EMT, and concurrent events [multidrug resistance (MDR) and tumor invasion] after Adriamycin or/and Twist1 small interfering RNA treatment were examined in vitro and in vivo. The Adriamycin-induced Twist1 expression and the interaction of Twist1 with p53-Mdm2 were examined by immunoblotting and immunoprecipitation, respectively. RESULTS: We showed in vitro that Adriamycin induced EMT and apoptosis simultaneously in a cell cycle-dependent manner. Only the cells undergoing EMT displayed enhanced invasion and MDR. Twist1 depletion completely blocked the mesenchymal transformation, partially reversed MDR, and greatly abolished invasion induced by Adriamycin. Also, we confirmed in vivo that Twist1 RNA interference improved the efficacy of Adriamycin for breast cancers. Further, Twist1 reduction in Adriamycin-treated cells promoted p53-dependent p21 induction and disrupted the association of p53 with Mdm2. CONCLUSIONS: Our studies show the diverse responses to Adriamycin treatment in cells at different phases, suggest an unrecognized role of EMT in regulating MDR and invasion, and show the efficacy of Twist1 RNA interference in Adriamycin-based chemotherapies for breast cancer.

DIXDC1 targets p21 and cyclin D1 via PI3K pathway activation to promote colon cancer cell proliferation.

DIXDC1 is the human homolog of Ccd1, a recently identified DIX domain containing protein in zebrafish. It is a positive regulator in the Wnt signaling pathway functioning downstream of Wnt and upstream of Axin. Since Wnt pathway activation is correlated with human colon cancer formation and progression, the biological role of DIXDC1 in human colon cancer was examined. In the current study, up-regulation of DIXDC1 protein was detected in human colorectal adenocarcinoma tissues and was found to be correlated well with high cell proliferation index. Ectopic over-expression of DIXDC1 resulted in increased cell proliferation in vitro and accelerated tumorigenesis on nude mice in vivo. We also showed that DIXDC1 promoted G0/G1 to S phase transition concomitantly with up-regulation of cyclin D1 and down-regulation of p21 protein. DIXDC1 over-expression cells showed activation of the PI3K/AKT pathway. Both siRNA knockdown of DIXDC1 and blocking the PI3K pathway using a specific inhibitor caused G1/S phase arrest, as well as down-regulation of cyclin D1 and up-regulation of p21 in DIXDC1 over-expression colon cancer cells. Collectively, this study demonstrates that over-expression of DIXDC1 might target p21 and cyclin D1 to promote colon cancer cell proliferation and tumorigenesis at least partially through activation of the PI3K/Akt pathway.

Over-expression of ubiquitin carboxy terminal hydrolase-L1 induces apoptosis in breast cancer cells.

Ubiquitin carboxy terminal hydrolase-L1 (UCH-L1) belongs to the UCH proteases family that deubiquitinates ubiquitin-protein conjugates in the ubiquitin-proteasome system. Previous research showed that UCH-L1 was expressed in mouse retinal cells and testicular germ cells, and its function was associated with apoptosis. But it is still unclear whether UCH-L1 is concerned with apoptosis in tumor cells. In order to clarify the role of UCH-L1 in tumor cells, multi-drug resistance (MDR) human breast carcinoma cell line MCF7/Adr, that expresses relatively high UCH-L1, and its parental cell line MCF7, that expresses relatively low UCH-L1, were chosen for this study. We transfected pcDNA3.1-UCH-L1 plasmid and UCH-L1 siRNA into MCF7 and MCF7/Adr cells, respectively. Using 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay, western blot, Hoechst 33258 staining assay and flow cytometry, we found that over-expression of UCH-L1 in MCF7 cells induced apoptosis. On the other hand, silencing of UCH-L1 in MCF7/Adr cells led to the opposite effect. Moreover, to explore the mechanism underling these observations, we further investigated the expression of phospho-Akt and its downstream signal phospho-IkB-alpha and other signal molecules including Fas, Fas-L, Trail, DR4, DR5, Bax, cytochrome C, active caspase-3, phospho-p53, phospho-Mdm-2, Bcl-2, Bcl-xL, p21 and p27. The results indicated that the process of apoptosis triggered by UCH-L1 is, at least in part, probably through Phosphoinositide 3-kinase (PI3K)/Akt signal pathway. Our findings suggest that modulating the ubiquitination and deubiquitination pathway could be a novel method for tumor therapy.

Cervical actinomycosis with spinal cord compression. Case report and literature review.

Cervical actinomycosis with spinal cord compression is extremely rare. The clinical presentation of spinal actinomycosis may be nonspecific and back pain is the most consistent early symptom. Here, we present such a case with fever, pain in the neck and upper back, progressive weakness and numbness in all 4 limbs with difficulty ambulating, constipation and uroschesis. Correct diagnosis is difficult because the clinical and radiological findings of actinomycosis closely resemble metastatic tumors and other infectious processes. Timely surgical debridement and decompression contributed to the prompt improvement of the patient's conditions, and histopathological demonstration of the inflammatory granulation tissue and Gram-positive sulfur-containing filamentous bacteria led to the correct diagnosis of actinomycosis. The diagnosis must be made promptly because delayed treatment can result in irreversible neurologic damage or death. Timely and long-term antibacterial therapy is essential for the complete recovery of the patient with actinomycosis.

Interaction between CD147 and P-glycoprotein and their regulation by ubiquitination in breast cancer cells.

BACKGROUND: Multidrug-resistant cancer cells overexpressing P-glycoprotein (P-gp) display variations in invasive and metastatic ability through the upregulation of the extracellular matrix metalloproteinase (MMP) inducer (CD147). However, the direct linkage between these two proteins is still unclear. METHODS: We used immunoprecipitation, immunofluorescence analysis, migration and invasion assays, drug sensitivity assay and Western blot to measure the physical and functional interaction between P-gp and CD147. Then we transfected vectors carrying ubiquitin C-terminal hydrolase L1 (UCH-L1) or UCH-L1 siRNA into MCF7 and MCF7/Adr cells, respectively, and investigated the role of UCH-L1 in the regulation of the expression and degradation of P-gp, CD147 and MMP-1, MMP-2, and MMP-9 by quantitative real-time polymerase chain reaction, Western blot and immunoprecipitation. RESULTS: In this paper, we showed that P-gp and CD147 interacted with each other, and that the ubiquitin-proteasome pathway played an important role in the turnover of them. In addition, we found that inhibition of N-glycosylation increased the ubiquitination and degradation of P-gp and CD147, and affected their function. UCH-L1 not only regulated the expression of P-gp, CD147 and MMP-1, MMP-2, and MMP-9, but also the ubiquitination and degradation of P-gp and CD147 in breast cancer cells. CONCLUSION: Our results demonstrate a mechanism underlying the linkage between multidrug resistance and tumor metastasis, and suggest for the first time that modulating the ubiquitination of P-gp and CD147 might be a novel method for tumor therapy.