An Arabidopsis Retention and Splicing complex regulates root and embryo development through pre-mRNA splicing.

Pre-mRNA splicing is an important step in the posttranscriptional processing of transcripts and a key regulator of development. The heterotrimeric retention and splicing (RES) complex plays vital roles in the growth and development of yeast, zebrafish, and humans by mediating pre-mRNA splicing of multiple genes. However, whether the RES complex is conserved in plants and what specific functions it has remain unknown. In this study, we identified Arabidopsis (Arabidopsis thaliana) BUD13 (AtBUD13), GROWTH, DEVELOPMENT AND SPLICING 1 (GDS1), and DAWDLE (DDL) as the counterparts of the yeast RES complex subunits Bud site selection protein 13 (Bud13), U2 snRNP component Snu17 (Snu17), and Pre-mRNA leakage protein 1, respectively. Moreover, we showed that RES is an ancient complex evolutionarily conserved in eukaryotes. GDS1 directly interacts with both AtBUD13 and DDL in nuclear speckles. The BUD13 domain of AtBUD13 and the RNA recognition motif domain of GDS1 are necessary and sufficient for AtBUD13-GDS1 interaction. Mutants of AtBUD13, GDS1, and DDL failed to properly splice multiple genes involved in cell proliferation and showed defects in early embryogenesis and root development. In addition, we found that GDS1 and DDL interact, respectively, with the U2 small nuclear ribonucleoproteins auxiliary factor AtU2AF65B and the NineTeen Complex-related splicing factor SKIP, which are essential for early steps of spliceosome assembly and recognition of splice sites. Altogether, our work reveals that the Arabidopsis RES complex is important for root and early embryo development by modulating pre-mRNA splicing.

AtBUD13 affects pre-mRNA splicing and is essential for embryo development in Arabidopsis.

Pre-mRNA splicing is an important step for gene expression regulation. Yeast Bud13p (bud-site selection protein 13) regulates the budding pattern and pre-mRNA splicing in yeast cells; however, no Bud13p homologs have been identified in plants. Here, we isolated two mutants that carry T-DNA insertions at the At1g31870 locus and shows early embryo lethality and seed abortion. At1g31870 encodes an Arabidopsis homolog of yeast Bud13p, AtBUD13. Although AtBUD13 homologs are widely distributed in eukaryotic organisms, phylogenetic analysis revealed that their protein domain organization is more complex in multicellular species. AtBUD13 is expressed throughout plant development including embryogenesis and AtBUD13 proteins is localized in the nucleus in Arabidopsis. RNA-seq analysis revealed that AtBUD13 mutation predominantly results in the intron retention, especially for shorter introns (≤100 bases). Within this group of genes, we identified 52 genes involved in embryogenesis, out of which 22 are involved in nucleic acid metabolism. Our results demonstrate that AtBUD13 plays critical roles in early embryo development by effecting pre-mRNA splicing.

Regulation of flowering transition by alternative splicing: the role of the U2 auxiliary factor.

Flowering transition is regulated by complex genetic networks in response to endogenous and environmental signals. Pre-mRNA splicing is an essential step for the post-transcriptional regulation of gene expression. Alternative splicing of key flowering genes has been investigated in detail over the past decade. However, few splicing factors have been identified as being involved in flowering transition. Human heterodimeric splicing factor U2 snRNP auxiliary factor (U2AF) consists of two subunits, U2AF35 and U2AF65, and functions in 3' splice site recognition in mRNA splicing. Recent studies reveal that Arabidopsis U2AF65a/b and U2AF35a/b play important roles in the splicing of key flowering genes. We summarize recent advances in research on splicing-regulated flowering transition by focusing on the role of Arabidopsis U2AF in the splicing of key flowering-related genes at ambient temperature and in the abscisic acid signaling pathways.

AtU2AF65b functions in abscisic acid mediated flowering via regulating the precursor messenger RNA splicing of ABI5 and FLC in Arabidopsis.

In mammalians and yeast, the splicing factor U2AF65/Mud2p functions in precursor messenger RNA (pre-mRNA) processing. Arabidopsis AtU2AF65b encodes a putative U2AF65 but its specific functions in plants are unknown. This paper examines the function of AtU2AF65b as a negative regulator of flowering time in Arabidopsis. We investigated the expression and function of AtU2AF65b in abscisic acid (ABA)-regulated flowering as well as the transcript abundance and pre-mRNA splicing of flowering-related genes in the knock-out mutants of AtU2AF65b. The atu2af65b mutants show early-flowering phenotype under both long-day and short-day conditions. The transcript accumulation of the flowering repressor gene FLOWERING LOCUS C (FLC) is reduced in the shoot apex of atu2af65b, due to both increased intron retention and reduced transcription activation. Reduced transcription of FLC results, at least partially, from the abnormal splicing and reduced transcript abundance of ABSCISIC ACID-INSENSITIVE 5 (ABI5), which encodes an activator of FLC in ABA-regulated flowering signaling. Additionally, the expression of AtU2AF65b is promoted by ABA. Transition to flowering and splicing of FLC and ABI5 in the atu2af65b mutants are compromised during ABA-induced flowering. ABA-responsive AtU2AF65b functions in the pre-mRNA splicing of FLC and ABI5 in shoot apex, whereby AtU2AF65b is involved in ABA-mediated flowering transition in Arabidopsis.

14-3-3zeta is involved in the anticancer effect of metformin in colorectal carcinoma.

Metformin is a promising drug for cancer prevention and treatment, especially in the diabetic population. We aimed to test whether 14-3-3zeta affects the anticancer effect of metformin on colorectal carcinoma (CRC). In this study, we confirmed that higher 14-3-3zeta expression was found in CRC tissues than in pericarcinoma tissues, and in CRC tissue of patients with diabetes than in those without diabetes. A knockdown of 14-3-3zeta inhibited CRC proliferation and promoted apoptosis in vitro and in vivo. Then, we created stable cell lines with under-expressed 14-3-3zeta from SW480 and HCT15 cells after infection by a lentiviral vector carrying short hairpin RNA targeting 14-3-3zeta (named LV-sh14-3-3zeta). Of note, metformin induced apoptosis and retarded tumor growth in the CRCs with overexpressed 14-3-3zeta, whereas this action was attenuated when 14-3-3zeta was knocked down. Moreover, either metformin or downregulation of 14-3-3zeta noticeably activated AMP-dependent protein kinase (AMPK) signaling, whereas the effect of metformin was attenuated when the 14-3-3zeta expression was decreased. Taken together, our results suggest that 14-3-3zeta may be associated with carcinogenesis and poor prognosis of CRCs associated with diabetes, and metformin may reverse the AMPK inhibition caused by 14-3-3zeta in CRCs in the background of diabetes. Our study should lead to a better understanding of the anticancer activity of metformin and points to possible application of metformin to the treatment of cancers overexpressing 14-3-3zeta.

[Bone Morphogenetic Proteins from CT26 Regulate the Expression of PD-L1 in Murine Dendritic Cells and Macrophages].

OBJECTIVES: To analyze the influence of bone morphogenetic proteins (BMPs) from CT26 on PD-L1 of dendritic cells and macrophages. METHODS: In vivo, we respectively inoculated CT26 colon cancer cells subcutaneously and intraperitoneally to BALB/c mice.The mice were randomly assigned to three groups and treated with normal saline; BMPs inhibitor LDN193189; BMPs inhibitor LDN193189 combined with paclitaxel, respectively. The treatments started on the eighth day after inoculating, when the tumor volume reached 150 mm3 or the abdominal circumference was greater than 6 cm. After 2 weeks of treatments, the mice were sacrificed.The counts of dendritic cells and macrophages and the expression of PD-L1 in tumors or ascites were detected by flow cytometry (FCM).In vivo, the dendritic cells and macrophages from normal BALB/c mice bone marrow were exposed to: no treatment; CT26 supernatant; CT26; CT26 supernatant and LDN193189; CT26 and LDN193189; CT26 supernatant, LDN193189 and paclitaxel; CT26, LDN193189 and paclitaxel. BMPs from CT26 was detected by ELISA.The counts of dendritic cells and macrophages and their PD-L1 expressions were detected by FCM. IRF-1 expression was detected by real-time (RT)-PCR and Western blot. RESULTS: In vivo, LDN193189 treated mice had the greatest tumor size or abdominal circumference, with least dendritic cells andCM(155.3mm]macrophages and expressions of PD-L1.In vivo, ELISA test results showed that the concentration of BMPs in CT26 supernatant was (0.59±0.09) ng/mL. FCM, RT-PCR and Western blot showed that dendritic cells and macrophages exposed to CT26 supernatant and LDN193189 or CT26 and LDN193189 expressed the least PD-L1 and IRF-1, which was close to those without treatment. While added the PTX to the above treatment, the expressions of PD-L1 and IRF-1 increased in the test results. CONCLUSIONS: BMPs from CT26 up-regulate the expression of PD-L1 in murine dendritic cells and macrophages.

Chinese herbal medicine Guizhi Fuling Formula for treatment of uterine fibroids: a systematic review of randomised clinical trials.

BACKGROUND: Guizhi Fuling Formula is widely applied for uterine fibroids in China. Many clinical trials are reported. This study assessed the efficacy and safety of Guizhi Fuling Formula for the treatment of uterine fibroids. METHODS: PubMed, Cochrane CENTRAL, EMBASE, and four Chinese databases were searched through May 2013. We included randomised controlled trials (RCTs) that tested Guizhi Fuling Formula for uterine fibroids, compared with no intervention, placebo, pharmaceutical medication, or other Chinese patent medicines approved by the State Food and Drug Administration of China. Authors extracted data and assessed the quality independently. We applied RevMan 5.2.0 software to analyse data of included randomised trials. RESULTS: A total of 38 RCTs involving 3816 participants were identified. The methodological quality of the included trials was generally poor. Meta-analyses demonstrated that Guizhi Fuling Formula plus mifepristone were more effective than mifepristone alone in reducing the volume of fibroids (in total volume of multiple fibroids, MD -19.41 cm(3), 95% CI -28.68 to -10.14; in average volume of multiple fibroids, MD -1.00 cm(3), 95% CI -1.23 to -0.76; in average volume of maximum fibroids, MD -3.35 cm(3), 95% CI -4.84 to -1.87, I(2) = 93%, random effects model). Guizhi Fuling Formula significantly improved symptoms of dysmenorrhea either when it was used alone (RR 2.27, 95% CI 1.04 to 4.97) or in combination with mifepristone (RR 2.35, 95% CI 1.15 to 4.82). No serious adverse events were reported. CONCLUSIONS: Guizhi Fuling Formula appears to have additional benefit based on mifepristone treatment in reducing volume of fibroids. However, due to high risk of bias of the trials, we could not draw confirmative conclusions on its benefit. Future clinical trials should be well-designed and avoid the issues that are identified in this study.

[Chinese medicine clinical trial protocol design and report specifications].

Clinical trial protocol is the document that illustrates the background of a clinical trial, theoretic basis, objective, design, methods, and organization, as well as statistical calculating, implement, and conditions for completion. Clinical trial protocol is the basic measure for ensuring the validity of scientific results and reducing bias. In order to optimize the design of clinical trial protocol, we generalize main problems in Chinese medicine clinical trials, key points of clinical trial protocol, as well as report standards.

Molecular mechanism of endophytic bacteria DX120E regulating polyamine metabolism and promoting plant growth in sugarcane.

Introduction: Sugarcane endophytic nitrogen-fixing bacterium Klebsiella variícola DX120E displayed broad impact on growth, but the exact biological mechanism, especially polyamines (PAs) role, is still meager. Methods: To reveal this relationship, the content of polyamine oxidase (PAO), PAs, reactive oxygen species (ROS)-scavenging antioxidative enzymes, phytohormones, 1-aminocyclopropane-1-carboxylic synthase (ACS), chlorophyll content, and biomass were determined in sugarcane incubated with the DX120E strain. In addition, expression levels of the genes associated with polyamine metabolism were measured by transcriptomic analysis. Results: Genomic analysis of Klebsiella variícola DX120E revealed that 39 genes were involved in polyamine metabolism, transport, and the strain secrete PAs in vitro. Following a 7-day inoculation period, DX120E stimulated an increase in the polyamine oxidase (PAO) enzyme in sugarcane leaves, however, the overall PAs content was reduced. At 15 days, the levels of PAs, ROS-scavenging antioxidative enzymes, and phytohormones showed an upward trend, especially spermidine (Spd), putrescine (Put), catalase (CAT), auxin (IAA), gibberellin (GA), and ACS showed a significant up-regulation. The GO and KEGG enrichment analysis found a total of 73 differentially expressed genes, involving in the cell wall (9), stimulus response (13), peroxidase activity (33), hormone (14) and polyamine metabolism (4). Discussion: This study demonstrated that endophytic nitrogen-fixing bacteria stimulated polyamine metabolism and phytohormones production in sugarcane plant tissues, resulting in enhanced growth. Dual RNA-seq analyses provided insight into the early-stage interaction between sugarcane seedlings and endophytic bacteria at the transcriptional level. It showed how diverse metabolic processes selectively use distinct molecules to complete the cell functions under present circumstances.

[Reporting specification of systematic reviews on Chinese medicine and methodological evaluation].

Currently, the number of systematic reviews on Chinese medicine (CM) increases gradually. However, the quality of the reviews varied, which resulted in great limitations in guiding clinical practice. This article refers to the Cochrane Handbook and PRISMA Statement to introduce the reporting specification of the systematic review, including asking a research question, review methods, presentation of the results, discussion and conclusion. We analyzed the methodology issues in the published systematic reviews on CM in order to improve the quality of future reviews.

[Systematic review on methodology of randomized controlled trials of post-marketing Chinese patent drugs for treatment of type 2 diabetes].

BACKGROUND: Randomized controlled trial (RCT) is considered as the gold standard for the efficacy assessment of medicines. With the increasing number of Chinese patent drugs for treatment of type 2 diabetes, the methodology of post-marketing RCTs evaluating the efficacy and specific effect has become more important. OBJECTIVE: To investigate post-marketing Chinese patent drugs for treatment of type 2 diabetes, as well as the methodological quality of post-marketing RCTs. SEARCH STRATEGY: Literature was searched from the books of Newly Compiled Traditional Chinese Patent Medicine and Chinese Pharmacopeia, the websites of the State Food and Drug Administration and the Ministry of Human Resources and Social Security of the People's Republic of China, China National Knowledge Infrastructure Database, Chongqing VIP Chinese Science and Technology Periodical Database, Chinese Biomedical Database (SinoMed) and Wanfang Data. The time period for searching ran from the commencement of each database to August 2011. INCLUSION CRITERIA: RCTs of post-marketing Chinese patent drugs for treatment of type 2 diabetes with intervention course no less than 3 months. DATA EXTRACTION AND ANALYSIS: Two authors independently evaluated the research quality of the RCTs by the checklist of risk bias assessment and the data collection forms based on the CONSORT Statement. Independent double data-extraction was performed. RESULTS: The authors identified a total of 149 Chinese patent drugs for treatment of type 2 diabetes. According to different indicative syndromes, the Chinese patent drugs can be divided into the following types, namely, yin deficiency and interior heat (n=48, 32%), dual deficiency of qi and yin (n=58, 39%) and dual deficiency of qi and yin combined with blood stasis (n=22, 15%). A total of 41 RCTs meeting the inclusion criteria were included. Neither multicenter RCTs nor endpoint outcome reports were found. Risk bias analysis showed that 81% of the included studies reported randomization for grouping without sequence generation, 98% of these studies did not report concealment of random numbers, 5% used placebo, 10% reported outcome attrition bias and no study employed the analysis of intention-to-treat and 98% reported the diagnostic criteria for type 2 diabetes. The participants mainly consisted of outpatients without complications (76%). The minimum and maximum sample size was 40 and 300 (106 ± 60), respectively. CONCLUSION: The inclusion and exclusion criteria and outcome measures did not match the purposes and contents of post-marketing research in the included studies. They also failed to reflect the basic principles of traditional Chinese medicine in the process of diagnosis and treatment. The demographic characteristics of the patients, the indications for medicine and the syndrome differentiation process were not reported sufficiently and transparently. In order to improve the post-marketing research and promote the rational use of Chinese patent drugs, it is recommended that phase IV clinical trials should establish clear research purpose as well as hypothesis first, and choose scientific and evidence-based study design and outcome measures. In addition, guidelines for implementation of post-marketing research should be developed.

[Design, manufacture and evaluation of a multi-parameter controllable automatic fire-acupuncture instrument].

A multi-parameter controllable automatic fire-acupuncture instrument was developed by integrating traditional fire needling with modern medical device technology. A gun-like appearance was designed for easy hand-held operation, the electromagnetic induction was for heating needle body, a scale knob was for controlling the needle insertion depth, the combination of electromagnetic ejection and spring return was for the precise control of the needle retention time; and the changeable single ste-rile needle or multiple needles were adopted to meet individual demand, obtain high efficiency and prevent infection. All of these designs are associated with the overall process control system to ensure the exact controllability of needle body temperature, needling density, insertion depth and needle retention time. Besides, this device is advantageous at handy and aseptic operation with high efficiency, conformability and visualization. In this research, this instrument was tested in animals for the impacts of automatic fire needling on skin damage and fur growth. It is found that the accurate control of each parameter is of the significant advantage in the safety and effectiveness of treatment, which lays a solid foundation for the subsequent systematic review on safety and effectiveness.

Plant-Specific AtS40.4 Acts as a Negative Regulator in Abscisic Acid Signaling During Seed Germination and Seedling Growth in Arabidopsis.

Abscisic acid (ABA) is an important phytohormone regulating plant growth, development and stress responses. A multitude of key factors implicated in ABA signaling have been identified; however, the regulation network of these factors needs for further information. AtS40.4, a plant-specific DUF584 domain-containing protein, was identified previously as a senescence regulator in Arabidopsis. In this study, our finding showed that AtS40.4 was negatively involved in ABA signaling during seed germination and early seedling growth. AtS40.4 was highly expressed in seeds and seedlings, and the expression level was promoted by ABA. AtS40.4 was localized both in the nucleus and the cytoplasm. Moreover, the subcellular localization pattern of AtS40.4 was affected by ABA. The knockdown mutants of AtS40.4 exhibited an increased sensitivity to ABA, whereas the overexpression of AtS40.4 decreased the ABA response during seed germination and seedling growth of Arabidopsis. Furthermore, AtS40.4 was involved in ABRE-dependent ABA signaling and influenced the expression levels of ABA INSENTIVE (ABI)1-5 and SnRK2.6. Further genetic evidence demonstrated that AtS40.4 functioned upstream of ABI4. These findings support the notion that AtS40.4 is a novel negative regulator of the ABA response network during seed germination and early seedling growth.

RNA Splicing of FLC Modulates the Transition to Flowering.

Flowering is a critical stage of plant development and is closely correlated with seed production and crop yield. Flowering transition is regulated by complex genetic networks in response to endogenous and environmental signals. FLOWERING LOCUS C (FLC) is a central repressor in the flowering transition of Arabidopsis thaliana. The regulation of FLC expression is well studied at transcriptional and post-transcriptional levels. A subset of antisense transcripts from FLC locus, collectively termed cold-induced long antisense intragenic RNAs (COOLAIR), repress FLC expression under cold exposure. Recent studies have provided important insights into the alternative splicing of COOLAIR and FLC sense transcripts in response to developmental and environmental cues. Herein, at the 20th anniversary of FLC functional identification, we summarise new research advances in the alternative splicing of FLC sense and antisense transcripts that regulates flowering.

Assessment of Concurrent Chemoradiotherapy plus Induction Chemotherapy in Advanced Nasopharyngeal Carcinoma: Cisplatin, Fluorouracil, and Docetaxel versus Gemcitabine and Cisplatin.

Induction chemotherapy treatment for nasopharyngeal carcinoma (NPC) is controversial. The aim of this study was to evaluate the treatment outcomes and toxicities between two induction chemotherapy regimens, with both followed by concurrent chemoradiotherapy. The first strategy used docetaxel, cisplatin and fluorouracil for induction chemotherapy (TPF), and the second utilised gemcitabine and cisplatin (GP). A retrospective analysis was performed on eligible NPC patients attending our hospital between May 2009 and Dec 2014. A total of 113 patients were enrolled with 58 patients receiving TPF and 55 receiving GP induction chemotherapy. Ninety-four patients (83.2%) were alive after 36-months follow-up. The median overall survival (OS) and progression-free survival (PFS) time were 48.3 and 39.7 months, respectively. The 3-year OS for the TPF regimen was 87.9% and 87.4% with GP chemotherapy (P = 0.928). The 3-year PFS of the TPF treatment was 84.5%, while it was 83.5% for the GP group (P = 0.551). Univariate analysis showed that lymph node metastasis was a significant PFS prognostic factor, while N3 stage was an independent predictor of PFS and distant failure-free survival (DMFS) in multivariate analysis. There were no significant differences in adverse toxicities or treatment efficacy between the chemotherapy regimens in the treatment of locoregionally advanced NPC.

Antiangiogenic therapy using endostatin increases the number of ALDH+ lung cancer stem cells by generating intratumor hypoxia.

Antiangiogenic therapy is becoming a promising option for cancer treatment. However, many investigations have recently indicated that these therapies may have limited efficacy, and the cancers in most patients eventually develop resistance to these therapies. There is considerable recently acquired evidence for an association of such resistance with cancer stem-like cells (CSLCs). Here, we used xenograft tumor murine models to further suggest that antiangiogenic agents actually increase the invasive and metastatic properties of lung cancer cells. In our experiments with murine lung cancer xenografts, we found that the antiangiogenic agent endostatin increased the population of ALDH+ cells, and did so by generating intratumoral hypoxia in the xenografts. We further showed endostatin to cause an increase in the CSLC population by accelerating the generation of tumor hypoxia and by recruiting TAMs, MDSCs and Treg cells, which are inflammatory and immunosuppressive cells and which can secrete cytokines and growth factors such as IL-6, EGF, and TGF-ß into the tumor microenvironment. All these factors are related with increased CSLC population in tumors. These results imply that improving the clinical efficacy of antiangiogenic treatments will require the concurrent use of CSLC-targeting agents.

Endostatin improves cancer-associated systemic syndrome in a lung cancer model.

Cancer-associated systemic syndrome (CASS) is characterized by a constellation of symptoms, including progressive weight loss, anemia, endocrine disorders, gastrointestinal dysfunction, muscle and adipose atrophy, hepatic peliosis and kidney failure. The present study assesses the effects of endostatin on CASS and any possible underlying mechanism in tumor-bearing mice. The results showed that the inoculation of Lewis lung carcinoma cells into mice led to CASS that was characterized by a notable decrease in body weight, severe anemia phenotype, disordered biochemistry, hepatosplenomegaly, and a marked increase in serum vascular endothelial growth factor (VEGF), tumor necrosis factor α and interleukin-6 (IL-6). The continuous injection of 10 mg/kg/day endostatin suppressed tumor growth and alleviated CASS in the tumor-bearing mice, as shown by weight gain, improvement in biochemistry and anemia, and the preservation of organ function. The effects of endostatin on CASS in the tumor-bearing mice were accompanied by the downregulation of serum VEGF and IL-6. Collectively, these findings indicate that endostatin improves CASS in tumor-bearing mice by decreasing the serum levels of VEGF and IL-6.

Traditional Chinese medicine for chronic fatigue syndrome: a systematic review of randomized clinical trials.

BACKGROUND: There is no curative treatment for chronic fatigue syndrome (CFS). Traditional Chinese medicine (TCM) is widely used in the treatment of CFS in China. OBJECTIVE: To evaluate the effectiveness and safety of TCM for CFS. METHODS: The protocol of this review is registered at PROSPERO. We searched six main databases for randomized clinical trials (RCTs) on TCM for CFS from their inception to September 2013. The Cochrane risk of bias tool was used to assess the methodological quality. We used RevMan 5.1 to synthesize the results. RESULTS: 23 RCTs involving 1776 participants were identified. The risk of bias of the included studies was high. The types of TCM interventions varied, including Chinese herbal medicine, acupuncture, qigong, moxibustion, and acupoint application. The results of meta-analyses and several individual studies showed that TCM alone or in combination with other interventions significantly alleviated fatigue symptoms as measured by Chalder's fatigue scale, fatigue severity scale, fatigue assessment instrument by Joseph E. Schwartz, Bell's fatigue scale, and guiding principle of clinical research on new drugs of TCM for fatigue symptom. There was no enough evidence that TCM could improve the quality of life for CFS patients. The included studies did not report serious adverse events. CONCLUSIONS: TCM appears to be effective to alleviate the fatigue symptom for people with CFS. However, due to the high risk of bias of the included studies, larger, well-designed studies are needed to confirm the potential benefit in the future.

Anti-tumor effects of penfluridol through dysregulation of cholesterol homeostasis.

BACKGROUND: Psychiatric patients appear to be at lower risk of cancer. Some antipsychotic drugs might have inhibitory effects on tumor growth, including penfluridol, a strong agent. To test this, we conducted a study to determine whether penfluridol exerts cytotoxic effects on tumor cells and, if so, to explore its anti-tumor mechanisms. METHODS: Growth inhibition of mouse cancer cell lines by penfluridol was determined using the 3- (4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. Cytotoxic activity was determined by clonogenic cell survival and trypan blue assays. Animal tumor models of these cancer cells were established and to evaluate penfluridol for its anti-tumor efficacy in vivo. Unesterified cholesterol in cancer cells was examined by filipin staining. Serum total cholesterol and tumor total cholesterol were detected using the cholesterol oxidase/p- aminophenazone (CHOD-PAP) method. RESULTS: Penfluridol inhibited the proliferation of B16 melanoma (B16/ F10), LL/2 lung carcinoma (LL/2), CT26 colon carcinoma (CT26) and 4T1 breast cancer (4T1) cells in vitro. In vivo penfluridol was particularly effective at inhibiting LL/2 lung tumor growth, and obviously prolonged the survival time of mice bearing LL/2 lung tumors implanted subcutaneously. Accumulated unesterified cholesterol was found in all of the cancer cells treated with penfluridol, and this effect was most evident in LL/2, 4T1 and CT26 cells. No significant difference in serum cholesterol levels was found between the normal saline-treated mice and the penfluridol-treated mice. However, a dose-dependent decrease of total cholesterol in tumor tissues was observed in penfluridol-treated mice, which was most evident in B16/F10-, LL/2-, and 4T1-tumor-bearing mice. CONCLUSION: Our results suggested that penfluridol is not only cytotoxic to cancer cells in vitro but can also inhibit tumor growth in vivo. Dysregulation of cholesterol homeostasis by penfluridol may be involved in its anti-tumor mechanisms.

Chinese herbal medicine for gout: a systematic review of randomized clinical trials.

Patients with gout referring to Chinese herbal medicine are not rare in China, and a great number of clinical trials on herbal medicine have been published. However, there has not been a systematic review to summarize the evidence of Chinese herbal medicine for gout. The aim of this study is to evaluate the evidence for the effectiveness and safety of Chinese herbal medicine for gout. We searched for randomized clinical trials on Chinese herbal medicine for gout till December 2012. Cochrane risk of bias tool was used to assess the methodological quality. RevMan 5.2 was used to synthesize the results. We included 57 trials involving 4,527 gout patients. The quality of trials was generally poor. No trial reported health-related quality of life in patients. There is not enough evidence showing that herbal medicine was statistically more effective than conventional medications in pain relief [mean difference (MD), -0.03; 95% confidence interval (CI), -0.06, 0.00], but herbal medicine combined with conventional medicines may have better effectiveness (MD, -0.33; 95% CI, -0.59, -0.07). Trials that reported function limitation relief found herbal medicine more effective than conventional medications (MD, -0.23; 95% CI, -0.32, -0.15). There was no evidence showing that herbal medicine prevents gout recurrence better. Twenty-five out of 41 trials, involving 23 different herbal prescriptions, found statistical significance in lowering serum uric acid level, and the overall effect from Chinese herbal medicine in inflammation relief is better than conventional therapies in 19 trials with 17 different prescriptions. The current data show that herbal medicine leads to fewer side reactions compared to conventional therapies [risk ratio (RR), 0.11; 95% CI, 0.08 to 0.15]. Chinese herbal medicine may have clinical effectiveness for functional recovery in patients with gout, and lead to a safe control of serum uric acid level and inflammation severity. Due to low quality of trials, trials with higher methodological quality and less heterogeneity are needed in the future.