Screening and Identification of the Main Metabolites of Schisantherin a In Vivo and In Vitro by Using UHPLC-Q-TOF-MS/MS.

Schisantherin A is an active ingredient originating from Schisandra chinensis (Turcz.) which has hepatoprotective and anti-oxidation activities. In this study, in vitro metabolisms investigated on rat liver microsomes (RLMs) and in vivo metabolisms explored on male Sprague Dawley rats of Schisantherin A were tested, respectively. The metabolites of Schisantherin A were identified using ultra-high-performance liquid chromatography coupled with hybrid triple quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS/MS). Based on the method, 60 metabolites were successfully identified and structurally characterized including 48 phase-I and 12 phase-II metabolites. Among the metabolites, 45 metabolites were reported for the first time. Moreover, 56 and eight metabolites were detected in urine and bile and 19 metabolites were identified in rats' plasma. It demonstrated that hepatic and extra-hepatic metabolic pathways were both involved in Schisantherin A biotransformation in rats. Five in vitro metabolites were structurally characterized for the first time. The results indicated that the metabolic pathways mainly include oxidation, reduction, methylation, and conjugation with glucuronide, taurine, glucose, and glutathione groups. This study provides a practical strategy for rapidly screening and identifying metabolites, and the results provide basic data for future pharmacological and toxicology studies of Schisantherin A and other lignin ingredients.

Circ-Foxo3 is positively associated with the Foxo3 gene and leads to better prognosis of acute myeloid leukemia patients.

BACKGROUND: The Foxo3 gene, belonging to the forkhead family, is one of the classes of transcription factors characterized by a forkhead DNA-binding domain, which usually considered being a cancer suppressor gene. Circ-Foxo3 is a circular structure which connects the 3'end to the 5'end. Scholars detected that circ-Foxo3 could compete with Foxo3 for binding to some miRNAs. METHODS: In this study, we will test the expression of Foxo3 and circ-Foxo3 in de novo acute myeloid leukemia (AML) patients to explore the relationship between Foxo3 gene and circ-Foxo3. All the de novo AML samples and normal control samples was measured by real-time quantitative PCR. A receiver operating characteristic curve was conducted to differentiate AML patients from control people. Association of Foxo3 expression and overall survival was conducted by Kaplan-Meier survival analysis. RESULTS: We found that the expression of Foxo3 gene in de novo patients was significantly lower than control samples (P = 0.009). Meanwhile, circ-Foxo3 also expressed lower in de novo AML patients than in control samples (P = 0.040). In different classifications, this trend could be observed more remarkably. In non-M3 patients, the Foxo3 high patients' survival time was longer than Foxo3 low patients (P = 0.002). Besides, in non-favorable risk groups, patients with low expression of Foxo3 had longer survival time than Foxo3 high patients (P = 0.004). Furthermore, in normal Karyotypic patients, the overall survival time of patients with high-expressed Foxo3 was significantly longer than those with low expression (P = 0.034). Besides, Pearson analysis was also conducted between these two genes in AML patients. Results revealed that they were positively correlated (R = 0.63, P < 0.001). CONCLUSION: In conclusion, we found that low expression of circ-Foxo3 and Foxo3 were frequent in AML patients, and patients with high expression of Foxo3 often had a trend of better prognosis.

Low Expression of Pseudogene POU5F1B Affects Diagnosis and Prognosis in Acute Myeloid Leukemia (AML).

BACKGROUND The transcription factor Oct-4 is necessary for maintaining pluripotency and self-renewal of embryonic stem cells, and POU5F1B is a processed pseudogene of Oct-4 with coding capacity. The purpose of this study is to evaluate the expression and clinical implication of POU5F1B in AML. MATERIAL AND METHODS The expression of the POU5F1B transcript was evaluated in 175 newly diagnosed AML patients and 39 healthy controls by use of real-time quantitative PCR (RQ-PCR). RESULTS POU5F1B was underexpressed in AML compared with controls (P<0.001). The receiver operating characteristic (ROC) curve revealed that the POU5F1B transcript level was able to differentiate AML patients from healthy individuals (AUC=0.682). In non-APL AML patients, the POU5F1Blow group had significantly higher WBC than the POU5F1Bhigh group (20.2×109 vs. 4.6×109 L⁻¹, P=0.021). Among whole-cohort AML, non-APL AML, and intermediate-risk AML, POU5F1Bhigh patients had obviously higher complete remission (CR) rates than POU5F1Blow patients (P=0.012, P=0.012 and P=0.027). In addition, Kaplan-Meier analysis demonstrated better overall survival (OS, P=0.019, P=0.007 and P=0.046, respectively) in POU5F1Bhigh patients compared with POU5F1Blow patients. Furthermore, in multivariate survival analysis, POU5F1B was independently associated with OS in non-APL AML patients and intermediate-risk AML as a favorable prognostic factor. CONCLUSIONS POU5F1B was frequently underexpressed in AML, and might contribute to the diagnosis and prognosis of AML.

Novel Cucurbitane Triterpenes from the Tubers of Hemsleya amabilis with Their Cytotoxic Acitivity.

Chemical research of the medicinal plant Hemsleya amabilis (Cucurbitaceae) yielded five new cucurbitane-type triterpenes hemslelis A⁻E (1⁻5) by silica gel column, ODS column, and semi-HPLC techniques. Their structures were determined by spectroscopic analysis and examined alongside existing data from prior studies. Compounds 1⁻5 were evaluated for their cytotoxic activities against three human tumor cell lines, Hela, HCT-8, and HepG-2, with the IC50 ranging from 5.9 to 33.9 µM compared to Cisplatin.

Low Expression of FUS1 Is Negatively Correlated with miR-378 and May Predict Adverse Prognoses in Acute Myeloid Leukemia.

FUS1 is a tumor suppressor gene that has been found to be frequently lost in a variety of solid tumors. In this study, we aimed to investigate the expression status of the FUS1 gene in acute myeloid leukemia (AML), as well as its clinical significance. We further explored the correlation between the expression of FUS1 and miR-378 in AML. We detected expression of the FUS1 transcript in bone marrow mononuclear cells from 23 controls and 158 newly diagnosed AML patients by real-time quantitative polymerase chain reaction. Downregulated FUS1 expression was found in 139 out of 158 (87.97%) AML cases; this rate was significantly lower than that in all 23 controls (p = 0.012). Receiver operating characteristic curve analysis revealed that the FUS1 transcript level could discriminate AML patients from controls effectively (area under the ROC curve = 0.663). Kaplan-Meier analysis demonstrated that non-M3-AML patients with a low FUS1 expression had a shorter overall survival (p = 0.049) and leukemia-free survival (p = 0.051) than those with a high FUS1 expression. Furthermore, we studied the correlation between the expression of FUS1 and miR-378 in 53 newly diagnosed AML patients. We found that the correlation coefficient was -0.346, which showed that FUS1 and miR-378 were negatively correlated in AML patients (p = 0.011). These results indicate that the low expression of FUS1 is a common molecular event in AML.

Effect of Group Density on the Physiology and Aggressive Behavior of Male Brandt's Voles (Lasiopodomys brandtii).

Xin Dai, Ling-Yu Zhou, Jie-Xia Cao, Yan-Qi Zhang, Feng-Ping Yang, Ai-Qin Wang, Wan-Hong Wei, and Sheng-Mei Yang (2018) Population density is well known to influence animal physiology and behavior. How population density affects the aggressive behavior of the Brandt's vole (Lasiopodomys brandtii) is, however, little known. The aim of this study was to investigate the effect of group density on physiologic responses and aggressive behavior of male Brandt's voles and their potential underlying neuro-mechanism. The results show that increasing group density led to elevated serum corticosterone levels and increased spleen weight; it also induced more male-male aggressive behavior. By contrast, it had a negative effect on body growth and the weight of testis and epididymis. Aging also increased male-male aggressive behavior. Higher density reduced mRNA levels of tryptophan hydroxylase 2 (TPH2), 5-hydroxytryptamine receptor 1A (5HT1A), and 5-hydroxytryptamine receptor 1B (5HT1B) in the amygdala and the dorsal raphe nucleus (DRN). Our results demonstrate that higher population density can intensify stress reactions and male-male aggressive behavior in Brandt's voles at the price of inhibiting body growth and reproduction. Serotonergic systems in the amygdala and the DRN may take part in the control of aggressive behavior among male voles. Our results provide novel insights into the neuro-mechanism underlying the influence of population density on aggressive behavior in Brandt's vole, and imply that aggressive behavior may play an important role in the population fluctuation of the animal.

Reduced expression of chemerin is associated with poor clinical outcome in acute myeloid leukemia.

Chemerin is dysregulation in numerous solid cancers. However, only little is known about the role of chemerin in acute myeloid leukemia (AML). In this study, we aimed to investigate the expression and clinical significance of recently described chemerin in acute myeloid leukemia (AML). The expression of chemerin in 149 patients with de novo AML and 35 normal controls was quantified by Real-time quantitative PCR (RQ-PCR). Chemerin was down-expressed in AML compared with controls (P=0.042). A receiver operating characteristic (ROC) curve revealed that chemerin expression could differentiate patients with AML from control subjects (AUC=0.611, 95% CI: 0.490-0.732; P=0.042) respectively. The cohort of AML patients was divided into two groups according to the cut-off value of 0.0826 (79% sensitivity and 54% specificity, respectively). In addition, the AML patients with low chemerin expression had significantly shorter overall survival (OS) than those with high chemerin expression (P=0.049). Moreover, multivariate survival analysis confirmed that chemerin was an independent prognostic factor for AML patients. In conclusion, downregulation of chemerin might be a useful diagnostic and prognostic factor for AML patients.

Scutellaria barbata attenuates diabetic retinopathy by preventing retinal inflammation and the decreased expression of tight junction protein.

AIM: To observe the attenuation of ethanol extract of Herba Scutellaria barbata (SE) against diabetic retinopathy (DR) and its engaged mechanism. METHODS: C57BL/6J mice were intraperitoneally injected with streptozotocin (STZ, 55 mg/kg) for 5 consecutive days to induce diabetes. The diabetic mice were orally given with SE (100, 200 mg/kg) for 1mo at 1mo after STZ injection. Blood-retinal barrier (BRB) breakdown was detected by using Evans blue permeation assay. Real-time polymerase chain reaction (RT-PCR), Western blot and immunofluorescence staining were used to detect mRNA and protein expression. Enzyme-linked immunosorbent assay (ELISA) was used to detect serum contents of tumor necrosis factor-α (TNF-α) and interleukin (IL)-1ß. RESULTS: SE (100, 200 mg/kg) reversed the breakdown of BRB in STZ-induced diabetic mice. The decreased expression of retinal claudin-1 and claudin-19, which are both tight junction (TJ) proteins, was reversed by SE. SE decreased the increased serum contents and retinal mRNA expression of TNF-α and IL-1ß. SE also decreased the increased retinal expression of intercellular cell adhesion molecule-1 (ICAM-1). SE reduced the increased phosphorylation of nuclear factor kappa B (NFκB) p65 and its subsequent nuclear translocation in retinas from STZ-induced diabetic mice. Results of Western blot and retinal immunofluorescence staining of ionized calcium-binding adapter molecule 1 (Iba1) demonstrated that SE abrogated the activation of microglia cells in STZ-induced diabetic mice. CONCLUSION: SE attenuates the development of DR by inhibiting retinal inflammation and restoring the decreased expression of TJ proteins including claudin-1 and claudin-19.

Down-regulation of pseudogene Vimentin 2p is associated with poor outcome in de novo acute myeloid leukemia.

OBJECTIVES: This study was intended to investigate the expression status of Vimentin 2p (VIM 2p), a pseudogene of Vimentin, and further analyze its clinical significance in AML patients. METHODS: Real-time quantitative PCR (RQ-PCR) was employed to explore the expression status of VIM 2p in 128 patients with de novo AML and 36 healthy controls. RESULTS: The expression level of VIM 2p was significantly decreased compared with healthy controls (P< 0.001). The patients with low VIM 2p expression were identified in 93 of 128 (73%) of AML patients. No significant differences could be observed in sex, age, blood parameters, FAB/WHO subtypes, karyotype risks and ten gene mutations (FLT3-ITD, NPM1, C-KIT, IDH1/IDH2, DNMT3 A, C/EBPA, N/K-RAS and U2AF1) between VIM 2p low-expressed and high-expressed patients (P> 0.05). Patients with low VIM 2p expression had significantly shorter overall survival (OS) than those with high VIM 2p expression in whole AML cases (median 7 vs. 13 months, respectively, P= 0.032), besides cytogenetically normal AML (CN-AML) and non-M3 AML cohort (P= 0.042 and 0.045, respectively). CONCLUSIONS: These findings indicate that VIM 2p down-regulation is a common event in AML and may be associated with poor clinical outcome.

Pseudogene BMI1P1 expression as a novel predictor for acute myeloid leukemia development and prognosis.

The BMI1P1 levels of 144 de novo AML patients and 36 healthy donors were detected by real-time quantitative PCR (RQ-PCR). BMI1P1 was significantly down-regulated in AML compared with control (P < 0.001). A receiver operating characteristic (ROC) curve revealed that BMI1P1 expression could differentiate patients with AML from control subjects (AUC = 0.895, 95% CI: 0.835-0.954, P < 0.001). The percentage of blasts in bone marrow (BM) was significantly lower in BMI1P1 high-expressed group versus low-expressed group (P = 0.008). BMI1P1 high-expressed cases had significantly higher complete remission (CR) than BMI1P1 low-expressed cases (P = 0.023). Furthermore, Kaplan-Meier demonstrated that both whole AML cohort and non-M3-AML patients with low BMI1P1 expression showed shorter leukemia free survival (LFS, P = 0.002 and P = 0.01, respectively) and overall survival (OS, P < 0.001 and P = 0.011, respectively) than those with high BMI1P1 expression. Multivariate analysis also showed that BMI1P1 over-expression was an independent favorable prognostic factor for OS in both whole and non-M3 cohort of AML patients (HR = 0.462, 95% CI = 0.243-0.879, P = 0.019 and HR = 0.483, 95% CI = 0.254-0.919, P = 0.027). To further investigate the significance of BMI1P1 expression in the follow-up of AML patients, we monitored the BMI1P1 level in 26 de novo AML patients and found that the BMI1P1 level increased significantly from the initial diagnosis to post-CR (P < 0.001). These results indicated that BMI1P1 might contribute to the diagnosis of AML and the assessment of therapeutic effect.

Over-expression of miR-675 in formalin-fixed paraffin-embedded (FFPE) tissues of breast cancer patients.

BACKGROUND: Dysregulation of miR-675 has been found in a variety of solid tumors. MiR-675 has been suggested as having both oncogenic and tumor suppression properties in cancer. However, there is no evidence whether miR-675 is involved in breast cancer. The objective of this study was to evaluate the expression status of miR-675 and its clinical relevance in breast cancer patients. METHODS: The expression level of miR-675 was detected in 100 breast cancer patients and 38 cancer-free controls using real-time quantitative PCR. The clinicopathological characteristics of miR-675 in breast cancer were also investigated. All statistical analyses were performed using SPSS 20.0. RESULTS: The study showed that miR-675 was significantly up-regulated in breast cancer patients compared with controls (P < 0.01). There was no significant difference in age, lymph nodes stage, ER status and PR status between patients with and without miR-675 over-expression (P > 0.05). The frequency of miR-675 over-expression was higher in the patients of histological grade I-II than in others (50% versus 9%, P = 0.011). The expression level of miR-675 had a high correlation with miR-24/93/98/378 in breast cancer patients. CONCLUSIONS: Taken together, our study demonstrated that miR-675 in formalin-fixed paraffin-embedded (FFPE) tissues might serve as a good source for biomarker discovery and breast cancer validation.

High expression of OCT4 is frequent and may cause undesirable treatment outcomes in patients with acute myeloid leukemia.

In recent years, many researches have shown that OCT4 is overexpressed in both germ cell tumors and somatic cancers. Meanwhile, OCT4 has relationship with poor prognosis in a lot of solid tumors, such as hepatocellular carcinoma, gastric cancer, and esophageal cancer. In our study, we investigated the expression status of OCT4 and its clinical significance in patients with acute myeloid leukemia (AML) using real-time quantitative PCR. The receiver operating characteristic (ROC) curve reveals that the level of OCT4 expression could be available for a potential diagnostic biomarker for differentiating AML from controls with an area under the ROC curve (AUC) of 0.915 (95 % confidence interval (CI) 0.837-0.992; P < 0.001). At the cutoff value of 0.56, the sensitivity and the specificity are 75.9 and 81.2 %, respectively. The amount of white blood cell (WBC) of patients with high OCT4 expression is higher than that of patients with low OCT4 expression (18.2 × 10(9) versus 2.7 × 10(9) L(-1), P = 0.001). Among those patients who are less than 70 years old, patients with OCT4 high expression have significantly shorter overall survival (OS) than those without OCT4 high expression (P = 0.048). These findings suggest that OCT4 high expression is a common event and may have an adverse impact on prognosis in AML.

High expression of dual-specificity phosphatase 5 pseudogene 1 (DUSP5P1) is associated with poor prognosis in acute myeloid leukemia.

The purpose of this study was to investigate the expression status of Dual-Specificity Phosphatase 5 Pseudogene 1 (DUSP5P1) and its clinical relevance in patients with acute myeloid leukemia (AML). Real-time quantitative PCR (RQ-PCR) was performed to detect the status of DUSP5P1 expression in 89 patients with de novo AML and 24 normal controls. The level of DUSP5P1 expression was significantly up-regulated in AML compared to controls (P=0.031). The patients with high expression of DUSP5P1 had higher percentage of blasts in bone marrow (BM) than those without high expression (P=0.027). The occurrence rate of DUSP5P1 high expression was significantly higher in M1 (2/8, 25%) and M2 subtypes (9/33, 27%) than in M3 subtype (0/17, 0%) (P=0.034). At the same time, the frequency of DUSP5P1 high expression in patients with intermediate (13/53, 24%) and poor karyotypes (5/11, 45%) was significantly higher than that in patients with favorable karyotype (0/21, 0%) (P=0.003). Meanwhile, DUSP5P1 high-expressed patients had significantly shorter overall survival (OS) than those with low expression (median 4.5 vs. 10.5 months, respectively, P=0.038). Our findings indicated that high expression of DUSP5P1 may identify high-risk AML patients and is associated with poor prognosis in AML.