Salicylic acid directly binds to ribosomal protein S3 and suppresses CDK4 expression in colorectal cancer cells.

Colorectal cancer is a significant cause of morbidity and represents a serious public health issue in many countries. The development of a breakthrough preventive method for colorectal cancer is urgently needed. Aspirin has recently been attracting attention as a cancer preventive drug, and its inhibitory effects on the development of various cancers have been reported in several large prospective studies. However, the underlying molecular mechanisms have not yet been elucidated in detail. In the present study, we attempted to identify the target proteins of aspirin using a chemical biology technique with salicylic acid, the main metabolite of aspirin. We generated salicylic acid-presenting FG beads and purified salicylic acid-binding proteins from human colorectal cancer HT-29 cells. The results obtained showed the potential of ribosomal protein S3 (RPS3) as one of the target proteins of salicylic acid. The depletion of RPS3 by siRNA reduced CDK4 expression and induced G1 phase arrest in human colorectal cancer cells. These results were consistent with the effects induced by the treatment with sodium salicylate, suggesting that salicylic acid negatively regulates the function of RPS3. Collectively, the present results show the potential of RPS3 as a novel target for salicylic acid in the protective effects of aspirin against colorectal cancer, thereby supporting RPS3 as a target molecule for cancer prevention.

Abemaciclib is a potent inhibitor of DYRK1A and HIP kinases involved in transcriptional regulation.

Homeodomain-interacting protein kinases (HIPKs) belong to the CMGC kinase family and are closely related to dual-specificity tyrosine phosphorylation-regulated kinases (DYRKs). HIPKs are regulators of various signaling pathways and involved in the pathology of cancer, chronic fibrosis, diabetes, and multiple neurodegenerative diseases. Here, we report the crystal structure of HIPK3 in its apo form at 2.5 Å resolution. Recombinant HIPKs and DYRK1A are auto-activated and phosphorylate the negative elongation factor SPT5, the transcription factor c-Myc, and the C-terminal domain of RNA polymerase II, suggesting a direct function in transcriptional regulation. Based on a database search, we identified abemaciclib, an FDA-approved Cdk4/Cdk6 inhibitor used for the treatment of metastatic breast cancer, as potent inhibitor of HIPK2, HIPK3, and DYRK1A. We determined the crystal structures of HIPK3 and DYRK1A bound to abemaciclib, showing a similar binding mode to the hinge region of the kinase as observed for Cdk6. Remarkably, DYRK1A is inhibited by abemaciclib to the same extent as Cdk4/Cdk6 in vitro, raising the question of whether targeting of DYRK1A contributes to the transcriptional inhibition and therapeutic activity of abemaciclib.

Downregulation of RPS14 inhibits the proliferation and metastasis of estrogen receptor-positive breast cancer cells.

Ribosomal protein S14 (RPS14) is a component of the 40S ribosomal subunit and is considered to be indispensable for ribosomal biogenesis. Previously, we found that RPS14 was significantly downregulated in estrogen receptor-positive (ER+) breast cancer cells following treatment with 4-hydroxytamoxifen (4-OH-TAM). However, its role in breast cancer remains poorly understood. In the present study, we sought to demonstrate, for the first time, that RPS14 is highly expressed in ER+ breast cancer tissues and its downregulation can significantly inhibit the proliferation, cycle, and metastasis of ER+ breast cancer cells, as well as induce cell apoptosis. Quantitative RT-PCR and western blotting were used to determine the expression of target genes. Herein, lentivirus-mediated small hairpin RNA (shRNA) targeting RPS14 was designed to determine the impact of RPS14 knockdown on ER+ breast cancer cells. Further, bioinformatics analysis was used to reveal the significance of differentially expressed genes in RPS14 knockdown breast cancer cells. RPS14 was highly expressed in ER+ breast cancer tissues compared to ER- tissues. The downregulation of RPS14 in two ER+ breast cancer cell lines suppressed cell proliferation, cell cycle and metastasis, and induced apoptosis. Based on bioinformatics analysis, the expression level of several significant genes, such as ASNS, Ret, and S100A4, was altered in breast cancer cells after RPS14 downregulation. Furthermore, the BAG2 and interferon signaling pathways were identified to be significantly activated. The downregulation of RPS14 in ER+ breast cancer cells can inhibit their proliferation and metastasis.

Ribosome Protection Proteins-"New" Players in the Global Arms Race with Antibiotic-Resistant Pathogens.

Bacteria have evolved an array of mechanisms enabling them to resist the inhibitory effect of antibiotics, a significant proportion of which target the ribosome. Indeed, resistance mechanisms have been identified for nearly every antibiotic that is currently used in clinical practice. With the ever-increasing list of multi-drug-resistant pathogens and very few novel antibiotics in the pharmaceutical pipeline, treatable infections are likely to become life-threatening once again. Most of the prevalent resistance mechanisms are well understood and their clinical significance is recognized. In contrast, ribosome protection protein-mediated resistance has flown under the radar for a long time and has been considered a minor factor in the clinical setting. Not until the recent discovery of the ATP-binding cassette family F protein-mediated resistance in an extensive list of human pathogens has the significance of ribosome protection proteins been truly appreciated. Understanding the underlying resistance mechanism has the potential to guide the development of novel therapeutic approaches to evade or overcome the resistance. In this review, we discuss the latest developments regarding ribosome protection proteins focusing on the current antimicrobial arsenal and pharmaceutical pipeline as well as potential implications for the future of fighting bacterial infections in the time of "superbugs."

Quantitative proteomics reveal the protective effects of EDS against osteoarthritis via attenuating inflammation and modulating immune response.

ETHNOPHARMACOLOGICAL RELEVANCE: Epimedium brevicornu Maxim, Dioscorea nipponica Makino, and Salvia miltiorrhiza Bunge formula (EDS) are three traditional Chinese medicines commonly combined and used to treat osteoarthritis (OA). However, the mechanism of its therapeutic effect on OA is still unclear. AIM OF THE STUDY: The aim of this study was to investigate the potential anti osteoarthritis mechanism of EDS in the treatment of OA rats' model by quantitative proteomics. MATERIALS AND METHODS: A papain-induced rat OA model was established, and then EDS was intragastrically administered for 28 days. A label-free quantification proteomics was performed to evaluate the holistic efficacy of EDS against OA and identify the possible protein profiles mechanisms. The expression levels of critical changed proteins were validated by RT-qPCR and Western blotting. The effects of EDS were then assessed by evaluating pathologic changes in the affected knee joint and measuring pressure pain threshold, acoustic reflex threshold, angle of joint curvature. RESULTS: Proteomics analysis showed that 62 proteins were significantly upregulated and 208 proteins were downregulated in OA group compared to control group. The changed proteins were involved in activation of humoral immunity response, complement cascade activation, leukocyte mediated immunity, acute inflammatory response, endocytosis regulation, and proteolysis regulation. The EDS treatment partially restored the protein profile changes. The protective effects of EDS on pathologic changes in OA rats' knee joint and pain threshold assessment were consisted with the proteomics results. CONCLUSIONS: The results suggest that EDS exerted synergistic therapeutic efficacies to against OA through suppressing inflammation, modulating the immune system, relieving joint pain, and attenuating cartilage degradation.

Peptide-Conjugated Phosphorodiamidate Morpholino Oligomers Retain Activity against Multidrug-Resistant Pseudomonas aeruginosa In Vitro and In Vivo.

Most antimicrobials currently in the clinical pipeline are modifications of existing classes of antibiotics and are considered short-term solutions due to the emergence of resistance. Pseudomonas aeruginosa represents a major challenge for new antimicrobial drug discovery due to its versatile lifestyle, ability to develop resistance to most antibiotic classes, and capacity to form robust biofilms on surfaces and in certain hosts such as those living with cystic fibrosis (CF). A precision antibiotic approach to treating Pseudomonas could be achieved with an antisense method, specifically by using peptide-conjugated phosphorodiamidate morpholino oligomers (PPMOs). Here, we demonstrate that PPMOs targeting acpP (acyl carrier protein), lpxC (UDP-(3-O-acyl)-N-acetylglucosamine deacetylase), and rpsJ (30S ribosomal protein S10) inhibited the in vitro growth of several multidrug-resistant clinical P. aeruginosa isolates at levels equivalent to those that were effective against sensitive strains. Lead PPMOs reduced established pseudomonal biofilms alone or in combination with tobramycin or piperacillin-tazobactam. Lead PPMO dosing alone or combined with tobramycin in an acute pneumonia model reduced lung bacterial burden in treated mice at 24 h and reduced morbidity up to 5 days postinfection. PPMOs reduced bacterial burden of extensively drug-resistant P. aeruginosa in the same model and resulted in superior survival compared to conventional antibiotics. These data suggest that lead PPMOs alone or in combination with clinically relevant antibiotics represent a promising therapeutic approach for combating P. aeruginosa infections.IMPORTANCE Numerous Gram-negative bacteria are becoming increasingly resistant to multiple, if not all, classes of existing antibiotics. Multidrug-resistant Pseudomonas aeruginosa bacteria are a major cause of health care-associated infections in a variety of clinical settings, endangering patients who are immunocompromised or those who suffer from chronic infections, such as people with cystic fibrosis (CF). Herein, we utilize antisense molecules that target mRNA of genes essential to bacterial growth, preventing the formation of the target proteins, including acpP, rpsJ, and lpxC We demonstrate here that antisense molecules targeted to essential genes, alone or in combination with clinically relevant antibiotics, were effective in reducing biofilms and protected mice in a lethal model of acute pneumonia.

SLFN11 inhibits hepatocellular carcinoma tumorigenesis and metastasis by targeting RPS4X via mTOR pathway.

Hepatocellular carcinoma (HCC) remains one of the most refractory malignancies worldwide. Schlafen family member 11 (SLFN11) has been reported to play an important role in inhibiting the production of human immunodeficiency virus 1 (HIV-1). However, whether SLFN11 also inhibits hepatitis B virus (HBV), and affects HBV-induced HCC remain to be systematically investigated. Methods: qRT-PCR, western blot and immunohistochemical (IHC) staining were conducted to investigate the potential role and prognostic value of SLFN11 in HCC. Then SLFN11 was stably overexpressed or knocked down in HCC cell lines. To further explore the potential biological function of SLFN11 in HCC, cell counting kit-8 (CCK-8) assays, colony formation assays, wound healing assays and transwell cell migration and invasion assays were performed in vitro. Meanwhile, HCC subcutaneous xenograft tumor models were established for in vivo assays. Subsequently, immunoprecipitation (IP) and liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) analyses were applied to understand the molecular mechanisms of SLFN11 in HCC. Co-IP, immunofluorescence and IHC staining were used to analyze the relationship between ribosomal protein S4 X-linked (RPS4X) and SLFN11. Finally, the therapeutic potential of SLFN11 with mTOR pathway inhibitor INK128 on inhibiting HCC growth and metastasis was evaluated in vitro and in vivo orthotopic xenograft mouse models. Results: We demonstrate that SLFN11 expression is decreased in HCC, which is associated with shorter overall survival and higher recurrence rates in patients. In addition, we show that low SLFN11 expression is associated with aggressive clinicopathologic characteristics. Moreover, overexpression of SLFN11 inhibits HCC cell proliferation, migration, and invasion, facilitates apoptosis in vitro, and impedes HCC growth and metastasis in vivo, all of which are attenuated by SLFN11 knockdown. Mechanistically, SLFN11 physically associates with RPS4X and blocks the mTOR signaling pathway. In orthotopic mouse models, overexpression of SLFN11 or inhibition of mTOR pathway inhibitor by INK128 reverses HCC progression and metastasis. Conclusions: SLFN11 may serve as a powerful prognostic biomarker and putative tumor suppressor by suppressing the mTOR signaling pathway via RPS4X in HCC. Our study may therefore offer a novel therapeutic strategy for treating HCC patients with the mTOR pathway inhibitor INK128.

Translational control of antibiotic resistance.

Many antibiotics available in the clinic today directly inhibit bacterial translation. Despite the past success of such drugs, their efficacy is diminishing with the spread of antibiotic resistance. Through the use of ribosomal modifications, ribosomal protection proteins, translation elongation factors and mistranslation, many pathogens are able to establish resistance to common therapeutics. However, current efforts in drug discovery are focused on overcoming these obstacles through the modification or discovery of new treatment options. Here, we provide an overview for common mechanisms of resistance to translation-targeting drugs and summarize several important breakthroughs in recent drug development.

Identification of the targets of hematoporphyrin derivative in lung adenocarcinoma using integrated network analysis.

BACKGROUND: Hematoporphyrin derivative (HPD) has a sensibilization effect in lung adenocarcinoma. This study was conducted to identify the target genes of HPD in lung adenocarcinoma. METHODS: RNA sequencing was performed using the lung adenocarcinoma cell line A549 after no treatment or treatment with X-ray or X-ray + HPD. The differentially expressed genes (DEGs) were screened using Mfuzz package by noise-robust soft clustering analysis. Enrichment analysis was carried out using "BioCloud" online tool. Protein-protein interaction (PPI) network and module analyses were performed using Cytoscape software. Using WebGestalt tool and integrated transcription factor platform (ITFP), microRNA target and transcription factor (TF) target pairs were separately predicted. An integrated regulatory network was visualized with Cytoscape software. RESULTS: A total of 815 DEGs in the gene set G1 (continuously dysregulated genes along with changes in processing conditions [untreated-treated with X-ray-X-ray + treated with HPD]) and 464 DEGs in the gene set G2 (significantly dysregulated between X-ray + HPD-treated group and untreated/X-ray-treated group) were screened. The significant module identified from the PPI network for gene set G1 showed that ribosomal protein L3 (RPL3) gene could interact with heat shock protein 90 kDa alpha, class A member 1 (HSP90AA1). TFs AAA domain containing 2 (ATAD2) and protein inhibitor of activated STAT 1 (PIAS1) were separately predicted for the genes in gene set G1 and G2, respectively. In the integrated network for gene set G2, ubiquitin-specific peptidase 25 (USP25) was targeted by miR-200b, miR-200c, and miR-429. CONCLUSION: RPL3, HSP90AA1, ATAD2, and PIAS1 as well as USP25, which is targeted by miR-200b, miR-200c, and miR-429, may be the potential targets of HPD in lung adenocarcinoma.

Identification of the targets of hematoporphyrin derivative in lung adenocarcinoma using integrated network analysis

BACKGROUND: Hematoporphyrin derivative (HPD) has a sensibilization effect in lung adenocarcinoma. This study was conducted to identify the target genes of HPD in lung adenocarcinoma. METHODS: RNA sequencing was performed using the lung adenocarcinoma cell line A549 after no treatment or treatment with X-ray or X-ray + HPD. The differentially expressed genes (DEGs) were screened using Mfuzz package by noise-robust soft clustering analysis. Enrichment analysis was carried out using "BioCloud" online tool. Protein-protein interaction (PPI) network and module analyses were performed using Cytoscape software. Using WebGestalt tool and integrated transcription factor platform (ITFP), microRNA target and transcription factor (TF) target pairs were separately predicted. An integrated regulatory network was visualized with Cytoscape software. RESULTS: A total of 815 DEGs in the gene set G1 (continuously dysregulated genes along with changes in processing conditions [untreated-treated with X-ray-X-ray + treated with HPD]) and 464 DEGs in the gene set G2 (significantly dysregulated between X-ray + HPD-treated group and untreated/X-ray-treated group) were screened. The significant module identified from the PPI network for gene set G1 showed that ribosomal protein L3 (RPL3) gene could interact with heat shock protein 90 kDa alpha, class A member 1 (HSP90AA1). TFs AAA domain containing 2 (ATAD2) and protein inhibitor of activated STAT 1 (PIAS1) were separately predicted for the genes in gene set G1 and G2, respectively. In the integrated network for gene set G2, ubiquitin-specific peptidase 25 (USP25) was targeted by miR-200b, miR-200c, and miR-429. CONCLUSION: RPL3, HSP90AA1, ATAD2, and PIAS1 as well as USP25, which is targeted by miR-200b, miR-200c, and miR-429, may be the potential targets of HPD in lung adenocarcinoma.

Liposome mediated double-stranded RNA delivery to silence ribosomal protein P0 in the tick Rhipicephalus haemaphysaloides.

Control of ticks has been achieved primarily by the application of acaricides, which has drawbacks such as environmental contamination leading to the selection of pesticide-resistant ticks. The potential of dsRNA to suppress genes critical for tick survival due to its sequence specificity suggests that dsRNAs could be developed as tailor-made pesticides. In this study, the dsRNA of P0 gene from the tick, Rhipicephalus haemaphysaloides, was evaluated as a potential anti-tick agent. Effects of using different dsRNA delivery methods were tested by quantitative RT-PCR and tick bioassays to determine survival, feeding and reproduction. The results showed that P0 dsRNAs could be effectively delivered into ticks and silenced by incubating with liposomes. Incubation time was found to be the most important factor in dsRNA delivery and gene silencing compared with liposome types and dsRNA concentration. The effects of P0 dsRNA treatment on ticks were found to be significant on blood feeding, molting or reproduction. These data show that anti-tick agents based on dsRNAs could have potential use in tick control.

The plant sesquiterpene lactone parthenolide inhibits Wnt/ß-catenin signaling by blocking synthesis of the transcriptional regulators TCF4/LEF1.

The Wnt/ß-catenin pathway is essential for embryonic development and homeostasis, but excessive activation of this pathway is frequently observed in various human diseases, including cancer. Current therapeutic drugs targeting the Wnt pathway often lack sufficient efficacy, and new compounds targeting this pathway are therefore greatly needed. Here we report that the plant-derived natural product parthenolide (PTL), a sesquiterpene lactone, inhibits Wnt signaling. We found that PTL dose-dependently inhibits Wnt3a- and CHIR99021-induced transcriptional activity assessed with the T-cell factor (TCF)/lymphoid enhancer factor (LEF) firefly luciferase (TOPFlash) assay in HEK293 cells. Further investigations revealed that PTL decreases the levels of the transcription factors TCF4/LEF1 without affecting ß-catenin stability or subcellular distribution. Moreover, this effect of PTL on TCF4/LEF1 was related to protein synthesis rather than to proteasome-mediated degradation. Of note, siRNA-mediated knockdown of RPL10, a ribosome protein PTL binds, substantially decreased TCF4/LEF1 protein levels and also Wnt3a-induced TOPFlash activities, suggesting a potential mechanism by which PTL may repress Wnt/ß-catenin signaling. In summary, PTL binds RPL10 and thereby potently inhibits the Wnt/ß-catenin pathway.

Molecular resistance mechanisms of Mycoplasma agalactiae to macrolides and lincomycin.

The extensive use of antimicrobials for disease control has caused a remarkable decrease in antimicrobial susceptibility of different animal mycoplasma species, including Mycoplasma agalactiae (M. agalactiae), the main causative agent of contagious agalactia. However, the molecular mechanisms behind M. agalactiae resistance to macrolides and lincomycin have not yet been elucidated. The aim of the present study was to investigate the association between minimum inhibitory concentration (MIC) values of different antimicrobials and mutations in the 23S rRNA gene and ribosomal proteins L4 and L22, analysing both field isolates (n=50) and in vitro selected resistant mutants of M. agalactiae. The obtained MIC results of the studied field isolates demonstrate an increasing development of tylosin resistance in this bacterium, in comparison to previous studies. Interestingly, predicted amino acid changes in L22 (Ser89Leu and Gln90Lys/His) were the first variations observed when MICs of M. agalactiae started to increase (tylosin MIC ≥0.8µg/ml), whereas mutations at positions 2058 or 2059 of domain V of the 23S rRNA gene appeared from MIC values of 1.6µg/ml. These results were consistent in both field isolates and in vitro selected mutants of M. agalactiae. Thus, although in other mycoplasma species resistance to macrolides and lincosamides had been mainly related to mutations in the 23S rRNA gene, this work demonstrates the role of alterations in ribosomal protein L22 in decreased susceptibility of M. agalactiae. Moreover, these mutations can be used as molecular markers to set an interpretative breakpoint of antimicrobial resistance for M. agalactiae.

Avilamycin and evernimicin induce structural changes in rProteins uL16 and CTC that enhance the inhibition of A-site tRNA binding.

Two structurally unique ribosomal antibiotics belonging to the orthosomycin family, avilamycin and evernimicin, possess activity against Enterococci, Staphylococci, and Streptococci, and other Gram-positive bacteria. Here, we describe the high-resolution crystal structures of the eubacterial large ribosomal subunit in complex with them. Their extended binding sites span the A-tRNA entrance corridor, thus inhibiting protein biosynthesis by blocking the binding site of the A-tRNA elbow, a mechanism not shared with other known antibiotics. Along with using the ribosomal components that bind and discriminate the A-tRNA-namely, ribosomal RNA (rRNA) helices H89, H91, and ribosomal proteins (rProtein) uL16-these structures revealed novel interactions with domain 2 of the CTC protein, a feature typical to various Gram-positive bacteria. Furthermore, analysis of these structures explained how single nucleotide mutations and methylations in helices H89 and H91 confer resistance to orthosomycins and revealed the sequence variations in 23S rRNA nucleotides alongside the difference in the lengths of the eukaryotic and prokaryotic α1 helix of protein uL16 that play a key role in the selectivity of those drugs. The accurate interpretation of the crystal structures that could be performed beyond that recently reported in cryo-EM models provide structural insights that may be useful for the design of novel pathogen-specific antibiotics, and for improving the potency of orthosomycins. Because both drugs are extensively metabolized in vivo, their environmental toxicity is very low, thus placing them at the frontline of drugs with reduced ecological hazards.

Pulsatile delivery of a leucine supplement during long-term continuous enteral feeding enhances lean growth in term neonatal pigs.

Neonatal pigs are used as a model to study and optimize the clinical treatment of infants who are unable to maintain oral feeding. Using this model, we have shown previously that pulsatile administration of leucine during continuous feeding over 24 h via orogastric tube enhanced protein synthesis in skeletal muscle compared with continuous feeding alone. To determine the long-term effects of leucine pulses, neonatal piglets (n = 11-12/group) were continuously fed formula via orogastric tube for 21 days, with an additional parenteral infusion of either leucine (CON + LEU; 800 µmol·kg-1·h-1) or alanine (CON + ALA) for 1 h every 4 h. The results show that body and muscle weights and lean gain were ∼25% greater, and fat gain was 48% lower in CON + LEU than CON + ALA; weights of other tissues were unaffected by treatment. Fractional protein synthesis rates in longissimus dorsi, gastrocnemius, and soleus muscles were ∼30% higher in CON + LEU compared with CON + ALA and were associated with decreased Deptor abundance and increased mTORC1, mTORC2, 4E-BP1, and S6K1 phosphorylation, SNAT2 abundance, and association of eIF4E with eIF4G and RagC with mTOR. There were no treatment effects on PKB, eIF2α, eEF2, or PRAS40 phosphorylation, Rheb, SLC38A9, v-ATPase, LAMTOR1, LAMTOR2, RagA, RagC, and LAT1 abundance, the proportion of polysomes to nonpolysomes, or the proportion of mRNAs encoding rpS4 or rpS8 associated with polysomes. Our results demonstrate that pulsatile delivery of a leucine supplement during 21 days of continuous enteral feeding enhances lean growth by stimulating the mTORC1-dependent translation initiation pathway, leading to protein synthesis in skeletal muscle of neonates.

Novel patented therapeutic approaches targeting the 37/67 kDa laminin receptor for treatment of cancer and Alzheimer's disease.

INTRODUCTION: The 37/67 kDa high-affinity laminin receptor (laminin receptor precursor/laminin receptor, LRP/LR) is a multi-faceted cellular receptor. It plays a vital role in the malignancy of various cancer types where it is seen to contribute to invasion, adhesion, apoptosis evasion and angiogenesis. Furthermore, it has been found to play an important role in facilitating the processes leading to neurotoxicity in Alzheimer's disease (AD). Various therapeutic options targeting this receptor have been patented with the outlook on application for the treatment/prevention of these diseases. AREAS COVERED: The various roles that LRP/LR plays in cancer, AD and infectious diseases caused by viruses and bacteria have been examined in detail and an overview of the current patented therapeutic strategies targeting this receptor is given. EXPERT OPINION: Molecular tools directed against LRP/LR, such as antibodies and small interfering RNA, could prove to be effective in the prevention of metastasis and angiogenesis while inducing apoptosis in cancers. Moreover, these strategies could also be applied to AD where LRP/LR is seen to facilitate the production and internalization of the neurotoxic Aß peptide. This review provides a comprehensive overview of the mechanisms by which LRP/LR is involved in eliciting pathogenic events, while showing how the use of patented approaches targeting this receptor could be used to treat them.

67kDa laminin receptor regulates the activation of MAPKs through DUSPs in glioma cell line U251.

PURPOSE: All-trans-retinoic-acid (ATRA), the active derivative of vitamin A, is critical in regulating cell cycle as well as inhibiting tumor growth and angiogenesis. It has been used in the clinical treatment of leukemia. 67kDa laminin receptor (67LR), as one of the receptor of laminin, plays an important role in tumor cells invasion, proliferation and metastasis. Current research indicates that 67LR is highly expressed in glioma and is associated with tumor progression. However, the underlying molecular mechanisms, especially the signaling pathways involved, have not been reported yet. Therefore it is of great importance to clarify its downstream targets. METHODS: The U251 glioma cell line was used in this study. Cell Counting Kit-8 was used in cell proliferation assay. Quantitative real-time PCR (qRT-PCR) was used to determine the transcription level of dual specificity phosphatases (DUSPs). Western blot analysis was used to detect the expression of mitogen activated protein kinases (MAPKs) and phosphorylated MAPKs. RESULTS: 67LR could influence the transcription of DUSPs and expression of MAPKs. ATRA could enhance the expression of 67LR in U251 cells and this enhancement was dose-dependent. ATRA was able to inhibit the growth of U251 cells. CONCLUSIONS: ATRA expressed significant therapeutic effect on glioma cells, and 67LR is not the only factor that can influence the proliferation of U251 cells.

Analysis of rpsL and rrs genes mutations related to streptomycin resistance in Mdr and Xdr clinical isolates of Mycobacterium tuberculosis.

INTRODUCTION: Streptomycin is a bactericidal and aminoglycoside antibiotic. It is one of the most effective drugs for treatment of multi-drug Tuberculosis disease. Incidence of resistance is increasingly reported. Its action mechanism is by inhibition of binding aminoacyl tRNA to position "A" in elongation phase, which finally it causes to stop bacterial protein synthesis. In this study, resistance rapid investigation to streptomycin was conducted in clinical strains of Mycobacterium tuberculosis. MATERIALS AND METHODS: In this study, among 105 strains of phlegm-positive and culture-positive Mycobacterium tuberculosis, 45 strains of resistant and sensitive to streptomycin were selected for possible mutations examination in genes rrs and rpsL. Specific primers that used for PCR were named rpsL 1, rpsL 2 and rrsR, Frrs. PCR products were sequenced. RESULT: PCR Products represents 504 bp band for gene rpsL and 1027 bp for gene rrs that shows proper selection of primers and determining an amplification appropriate program. From 26 resistant strains to streptomycin 26 strain have mutation in rpsL gene and 1 strain have alteration in rrs gene. In this study 19 strains were sensitive to streptomycin that have no mutation in these gene. CONCLUSIONS: Streptomycin resistance is mainly related to mutation at codons 43 and 88 "rpsL" gene and to a lesser extent "rrs" that are the greatest cause of drug resistance to streptomycin.

Ellagic acid modulates the expression of oral innate immune mediators: potential role in mucosal protection.

BACKGROUND: Ellagic acid (EA) found in various fruits such as pomegranates, blackberries, raspberries, strawberries, and walnuts has different pharmacological functions including antioxidant, antitumor, antiallergic, anti-inflammatory, antibacterial, and antiviral activities. It is not known, however, if EA could enhance mucosal innate immunity. Our goal was to determine the effects of EA on the expression of innate immune mediators produced by oral epithelial cells. METHODS: Culture of primary human gingival epithelial cells (HGEs) was performed in duplicate, and after the primary HGEs had been treated with EA at a concentration ranging from 12.5 to 100 µM for 18 h the cells and supernatants were harvested. The expression of innate immune mediators including human ß-defensin 2 (hBD2), secretory leukocyte protease inhibitor (SLPI), and various cytokines and chemokines was measured at both transcriptional and translational levels by using quantitative real-time PCR, ELISA, and Luminex assay. RESULTS: In the presence of EA, the expression of hBD2-and SLPI mRNA was 3.7-folds and 2.6-folds greater than untreated controls, respectively, and consistent with their secreted protein levels. For cytokines and chemokines, increased expression of RANTES, IL-2, and IL-1ß was found in response to EA. In contrast, EA decreased the expression of IL-6, IL-8, and TNF-α. CONCLUSIONS: This study demonstrated that oral innate immunity is affected by EA found in fruits. Thus, it may play some roles in mucosal innate immunity. The potential of EA for modulating the innate immune mediators may lead to developing a new topical agent to treat and/or prevent immune-mediated oral diseases.

Genome-wide screen identifies signaling pathways that regulate autophagy during Caenorhabditis elegans development.

The mechanisms that coordinate the regulation of autophagy with developmental signaling during multicellular organism development remain largely unknown. Here, we show that impaired function of ribosomal protein RPL-43 causes an accumulation of SQST-1 aggregates in the larval intestine, which are removed upon autophagy induction. Using this model to screen for autophagy regulators, we identify 139 genes that promote autophagy activity upon inactivation. Various signaling pathways, including Sma/Mab TGF-ß signaling, lin-35/Rb signaling, the XBP-1-mediated ER stress response, and the ATFS-1-mediated mitochondrial stress response, regulate the expression of autophagy genes independently of the TFEB homolog HLH-30. Our study thus provides a framework for understanding the role of signaling pathways in regulating autophagy under physiological conditions.