CDX2 as a Predictive Biomarker Involved in Immunotherapy Response Suppresses Metastasis through EMT in Colorectal Cancer.

Background: Studies have confirmed that Caudal Type Homeobox 2 (CDX2) plays a tumor suppressor role in colorectal cancer (CRC) and as a prognostic and predictive marker for colorectal cancer. The epithelial to mesenchymal transition (EMT) is a transdifferentiation process, providing migratory and invasive properties to cancer cells during tumor progression. However, the role of CDX2 during the activation of EMT in CRC maintains controversial. Aim: To investigate whether CDX2 is associated with EMT in CRC. Methods: Forty-six CRC patients were included in the study. Expressions of CDX2, E-cadherin, and N-cadherin in all CRC patients were detected by IHC. ROC assays were applied to detect cut-off points for IHC scores to distinguish high and low expressions of CDX2 in 46 CRC samples. The prognostic value of CDX2 was statistically analyzed. MTT, Western blot, invasion, and migration assays in vitro were employed to explore the function of CDX2. Results: We observed that high expressions of CDX2 and E-cadherin as well as low expressions of N-cadherin were significantly correlated with favorable prognosis. The levels of CDX2 protein exhibited a positive associated with E-cadherin while negative correlation with N-cadherin. Then, the low expression of CDX2 and high expression of CA199 in combination are positively related with poor prognosis. Overexpression of CDX2 reduced expression of MMP-2 and diminished cell proliferation, invasion, and migration, while knockdown CDX2 enhanced MMP-2 expression and increased cell proliferation, invasion, and migration in HCT-116 cells. CDX2 was correlated with expression of EMT markers. Overexpression of CDX2 suppressed the EMT markers indicating that CDX2 suppresses CRC cell viability, invasion, and metastasis through inhibiting EMT. Finally, we found that the expression of CDX2 was negatively associated with Th1 cells, macrophages, Th2 cells, cytotoxic cells, T cells, and T helper cells. Conclusions: These results indicated CDX2 as prognostic biomarkers involved in immunotherapy response for CRC. CDX2 loss promotes metastasis in CRC through a CDX2-dependent mechanism.

[Evaluation of 93 cases of mandibular first molar root bifurcation lesions treated with autologous dentin Granules combined with platelet-rich fibrin membrane].

PURPOSE: To investigate the efficacy of autologous dentin particles combined with platelet-rich fibrin membrane (PRF) in the treatment of root bifurcation lesions of mandibular first molar. METHODS: Ninety-three patients (93 teeth) with mandibular first molar root bifurcation lesions were selected from our department from February 2016 to October 2017. They were randomly divided into 2 groups. Forty-six patients with 46 teeth in the experimental group underwent autologous dentin particles combined with platelet-rich fibrin membrane, while patents in the control group (47 patients with 47 teeth) were treated with Bio-Oss implanted in the bone defect area covered with collagen membrane. The patients were revisted at 1, 3, 6 and 12 months after operation. The success rate of the operation group, the depth of periodontal pocket (PD), the loss of attachment (AL), the depth of penetration of the root bifurcation (HPD), and the bone density of the root bifurcation area before and after treatment. The data were recorded and compared with SPSS25.0 software package. RESULTS: The success rate was 97.83%(45/46) in the experimental group, 85.11%(40/47) in the control group, the difference between the two groups was significant(P<0.05). After treatment, PD, AL and HPD decreased significantly (P<0.05), and MGVs increased gradually. There was no significant difference in MGVs before treatment and 1 month after treatment in the experimental group (P>0.05). MGVs at other time points were significantly higher than those of the control group (P<0.05). PD, AL and HPD of the experimental group were lower significantly than the control group at each time point after treatment (P<0.05), and MGVs value was significantly higher than the control group (P<0.05). There was no significant difference in the incidence of complications(4.35% vs 6.38%, χ2=0.189, P>0.05). CONCLUSIONS: Autologous dentin particles combined with platelet-rich fibrin membrane is effective for the treatment of root bifurcation lesions of mandibular first molar, which is worthy of wide application.

Antifungal Mechanism of Dipicolinic Acid and Its Efficacy for the Biocontrol of Pear Valsa Canker.

Valsa pyri is a fatal canker pathogen that causes significant reduction of crop yield in pear orchards. V. pyri invades the trunk phloem, and is difficult to control by chemical treatment. In this work, it was found for the first time that Bacillus subtilis-produced dipicolinic acid (DPA) exhibits antifungal activity against different canker pathogens, including Alteraria alternata, Botryosphaeria dothidea, Rhizoctonia solani, and V. pyri. Growth inhibition of V. pyri was observed at less than 5 mM concentration (pH = 5.6). DPA showed the highest antifungal activity at acidic pH values and in the presence of bivalent metals, such as zinc(II), cobalt(II), and copper(II). Measurement of mRNA expression levels and scanning electron microscope (SEM) observations revealed that DPA causes V. pyri apoptosis via inhibition of chitin biosynthesis and subsequent cell lysis. Interestingly, DPA showed high stability in the pear bark and was able to cross the pear tree bark into the phloem, protecting the internal phases of the pear trunk. In preventive applications, DPA reduced the canker symptoms of V. pyri on Cuigan pear trees by 90%. Taken together, an efficient strategy for the management of V. pyri-caused canker disease was developed using a novel antifungal agent, DPA, with strong antifungal activity and particular diffusion properties.

Overexpression of IQGAP1 promotes the angiogenesis of esophageal squamous cell carcinoma through the AKT and ERK­mediated VEGF­VEGFR2 signaling pathway.

Angiogenesis is crucial for the progression of esophageal squamous cell carcinoma (ESCC). Anti­angiogenesis by targeting important molecules has been considered as one of the most promising and efficient strategy for cancer therapy. Recent studies have demonstrated that the IQ­domain GTPase activating protein 1 (IQGAP1) plays critical roles in tumorigenesis and cancer progression. We previously reported that IQGAP1 is overexpressed in ESCC, and IQGAP1 knockdown can decrease cell proliferation and metastasis ability in vitro and in vivo. However, the effects of IQGAP1 on the angiogenesis of ESCC and its underlying mechanisms remain unknown. In the present study, we found that IQGAP1 overexpression promoted tumor angiogenesis confirmed by human umbilical vascular endothelial cell (HUVEC) tube formation in vitro and chicken embryo chorioallantoic membrane (CAM) assay in vivo. Moreover, IQGAP1 overexpression in ESCC cells increased expression of vascular endothelial growth factor (VEGF) and phosphorylation of vascular endothelial growth factor receptor 2 (VEGFR2). Meanwhile, we found that levels of AKT and ERK phosphorylation were upregulated in IQGAP1­overexpressing cells. Importantly, IQGAP1­knockdown cells showed the opposing results. Furthermore, AKT and ERK inhibitors not only significantly decreased VEGF expression and VEGFR2 phosphorylation in IQGAP1­overexpressing cells, but also abolished the pro­angiogenic effect of IQGAP1 overexpression on angiogenesis in the HUVEC tube formation and chicken embryo CAM assay. Taken together, this evidence confirms that IQGAP1 overexpression promotes tumor angiogenesis via the AKT and ERK­mediated VEGF­VEGFR2 signaling pathway in ESCC, and IQGAP1 may be an attractive therapeutic target for cancer anti­angiogenesis treatment.

[Study on clinical efficacy and patients' satisfaction of MTA apical barrier technique in treatment of young permanent teeth with periapical inflammation].

PURPOSE: To compare the clinical efficacy of MTA apical barrier technique and Vitapex apexification in treatment of young permanent teeth with periapical inflammation, and to evaluate the satisfaction of patients. METHODS: Seventy-five cases of young permanent teeth with periapical inflammation were randomly divided into control group (n=37) and experimental group (n=38). Patients in the control group were treated with Vitapex apexification, while patients in the experimental group were treated with MTA apical barrier technique. The clinical efficacy of the two groups was compared at 3, 6, 9 months and 1 year after treatment, and the average treatment time and average treatment period were compared between 2 groups. The difference of patients' satisfaction with medical environment, health care service, late health care guidance, treatment cost, treatment period and treatment effect were compared between 2 groups. The clinical efficacy, treatment times and period, satisfaction of 2 groups were recorded and analyzed by using SPSS 19.0 software package. RESULTS: At 3 month and 6 month of revisit, the clinical efficacy of the experimental group was better than the control group, but there was no significant difference between 2 groups (P>0.05). At 9 months and 1 year of revisit, the total efficiency of the experimental group was significantly better than the control group (78.38%:94.74%, P=0.037；75.68%∶97.37%, P=0.006). The treatment time and treatment period of the experimental group were significantly lower than the control group (P<0.05), the values were (3.24±0.39) times, (0.68±0.23) months and (7.78±0.65) times, (8.24±2.95) months. Patients' satisfaction with medical treatment environment, health care service, late health care guidance and treatment period was not significant different between 2 groups (P>0.05). However, patients' satisfaction with treatment cost and treatment effect in the experimental group was significantly higher than the control group (P<0.05). CONCLUSIONS: MTA apical barrier technique has better clinical efficacy, less treatment time, shorter treatment period and higher satisfaction than Vitapex apexification. It is suitable for clinical application.

Long-term propagation of tree shrew spermatogonial stem cells in culture and successful generation of transgenic offspring.

Tree shrews have a close relationship to primates and have many advantages over rodents in biomedical research. However, the lack of gene manipulation methods has hindered the wider use of this animal. Spermatogonial stem cells (SSCs) have been successfully expanded in culture to permit sophisticated gene editing in the mouse and rat. Here, we describe a culture system for the long-term expansion of tree shrew SSCs without the loss of stem cell properties. In our study, thymus cell antigen 1 was used to enrich tree shrew SSCs. RNA-sequencing analysis revealed that the Wnt/ß-catenin signaling pathway was active in undifferentiated SSCs, but was downregulated upon the initiation of SSC differentiation. Exposure of tree shrew primary SSCs to recombinant Wnt3a protein during the initial passages of culture enhanced the survival of SSCs. Use of tree shrew Sertoli cells, but not mouse embryonic fibroblasts, as feeder was found to be necessary for tree shrew SSC proliferation, leading to a robust cell expansion and long-term culture. The expanded tree shrew SSCs were transfected with enhanced green fluorescent protein (EGFP)-expressing lentiviral vectors. After transplantation into sterilized adult male tree shrew's testes, the EGFP-tagged SSCs were able to restore spermatogenesis and successfully generate transgenic offspring. Moreover, these SSCs were suitable for the CRISPR/Cas9-mediated gene modification. The development of a culture system to expand tree shrew SSCs in combination with a gene editing approach paves the way for precise genome manipulation using the tree shrew.

Germ stem cells are active in postnatal mouse ovary under physiological conditions.

STUDY HYPOTHESIS: Are active ovarian germ stem cells present in postnatal mouse ovaries under physiological conditions? STUDY FINDING: Active ovarian germ stem cells exist and function in adult mouse ovaries under physiological conditions. WHAT IS KNOWN ALREADY: In vitro studies suggested the existence of germ stem cells in postnatal ovaries of mouse, pig and human. However, in vivo studies provided evidence against the existence of active germ stem cells in postnatal mouse ovaries. Thus, it remains controversial whether such germ stem cells really exist and function in vivo in postnatal mammalian ovaries. STUDY DESIGN, SAMPLES/MATERIALS, METHODS: Octamer-binding transcription factor 4 (Oct4)-MerCreMer transgenic mice were crossed with R26R-enhanced yellow fluorescent protein (EYFP) mice to establish a tamoxifen-inducible tracing system so that Oct4-expressing potential ovarian germ stem cells in young adult mice (5-6 weeks old) can be labeled with EYFP. The germ cell activities of DNA replication, mitotic division, entry into meiosis and progression to primordial follicle stage were investigated by means of immunofluorescent staining of ovarian tissues collected at different time points post-tamoxifen injection (1 day, 3 days, 2 months and 4 months). Meiosis entry and primordial follicle formation were also measured by EYFP-labeled single-cell RT-PCR. Germ cell proliferation and mitotic division were examined through 5-bromodeoxyuridine triphosphate incorporation assay. At each time point, ovaries from two to three animals were used for each set of experiment. MAIN RESULTS AND THE ROLE OF CHANCE: By labeling the Oct4-expressing small germ cells and tracing their fates for up to 4 months, we observed persistent meiosis entry and primordial follicle replenishment. Furthermore, we captured the transient processes of mitotic DNA replication as well as mitotic division of the marked germ cells at various time periods after tracing. These lines of evidence unambiguously support the presence of active germ stem cells in postnatal ovaries and their function in replenishing primordial follicle pool under physiological conditions. Moreover, we pointed out that Oct4(+) deleted in azoospermia-like (Dazl)(-) but not Oct4(+)Dazl(+) or Oct4(+) DEAD (Asp-Glu-Ala-Asp) Box Polypeptide 4 (Ddx4)(+) cells contain a population of germ stem cells in mouse ovary. LIMITATIONS, REASONS FOR CAUTION: This study was conducted in mice. Whether or not the results are applicable to human remain unclear. The future work should aim at identifying the specific ovarian germ stem cell marker and evaluating the significance of these stem cells to normal ovarian function. WIDER IMPLICATIONS OF THE FINDINGS: Clarifying the existence of active germ stem cells and their functional significance in postnatal mammalian ovaries could provide new insights in understanding the mechanism of ovarian aging and failure. LARGE SCALE DATA: Not applicable. STUDY FUNDING/COMPETING INTERESTS: This work was supported by the National Key Basic Research Program of China (grant number 2012CBA01300) and the National Natural Science Foundation of China to P.Z. (31571484). No competing interests are reported.

Filia Is an ESC-Specific Regulator of DNA Damage Response and Safeguards Genomic Stability.

Pluripotent stem cells (PSCs) hold great promise in cell-based therapy, but the genomic instability seen in culture hampers their full application. A greater understanding of the factors that regulate genomic stability in PSCs could help address this issue. Here we describe the identification of Filia as a specific regulator of genomic stability in mouse embryonic stem cells (ESCs). Filia expression is induced by genotoxic stress. Filia promotes centrosome integrity and regulates the DNA damage response (DDR) through multiple pathways, including DDR signaling, cell-cycle checkpoints and damage repair, ESC differentiation, and apoptosis. Filia depletion causes ESC genomic instability, induces resistance to apoptosis, and promotes malignant transformation. As part of its role in DDR, Filia interacts with PARP1 and stimulates its enzymatic activity. Filia also constitutively resides on centrosomes and translocates to DNA damage sites and mitochondria, consistent with its multifaceted roles in regulating centrosome integrity, damage repair, and apoptosis.

Lateral flow immunoassay for quantitative detection of ractopamine in swine urine.

A strip reader based lateral flow immunoassay (LFIA) was established for the rapid and quantitative detection of ractopamine (RAC) in swine urine. The ratio of the optical densities (ODs) of the test line (AT) to that of the control line (AC) was used to effectively minimize interference among strips and sample variations. The linear range for the quantitative detection of RAC was 0.2 ng/mL to 3.5 ng/mL with a median inhibitory concentration (IC50) of 0.59 ± 0.06 ng/mL. The limit of detection (LOD) of the LFIA was 0.13 ng/mL. The intra-assay recovery rates were 92.97%, 97.25%, and 107.41%, whereas the inter-assay rates were 80.07%, 108.17%, and 93.7%, respectively.

[Current progress and future direction in the biology of ovarian germ stem cells in mammals].

Whether or not oogenesis continues after birth in mammalian ovaries remains controversial. Since the 1950's, it has been generally accepted that oogenesis takes place during embryogenesis in mammals and ceases at birth. At birth, germ cells in mammalian ovaries have progressed to the diplotene stage of meiotic prophase and have formed primordial follicles with surrounding somatic cells. These primordial follicles represent follicle reserves of the reproductive life. However, this view has been recently challenged by a growing body of evidence showing the isolation and propagation of germ stem cells from mouse and human ovaries. These ovarian germ stem cells are capable of regenerating functional oocytes when transplanted back into recipient ovaries. Despite the discovery of the potential germ stem cells in mammalian ovaries, it remains uncertain whether these cells exist and function in ovaries under physiological conditions. Herein we review the current progress and future direction in this infant area.

[Preconception weight and perinatal health].

The health condition of a women relates closely to the health of her child and family members. The health of the mother is critical to her being able to conceive and give birth to healthy children and constructing a healthy lifestyle environment. Recent research has found a relationship between preconception weight and perinatal health. Abnormal weight (either overweight or underweight) was found to be the most prevalent of pre-pregnancy medical conditions studied. Therefore, the Center of Disease Control and Prevention in the United States has suggested that every woman should receive a preconception health assessment. Weight gain during pregnancy should be compared against preconception weight and body mass index. Following such a regimen can help improve maternal-infant perinatal health.